Role of Stem Cell Transplantation in the Treatment of Ulcerative Colitis

Introduction and Aim of the Work

- 1 -

Introduction

Inflammatory bowel disease (IBD) represents

a group of idiopathic chronic inflammatory intestinal

conditions. The two main disease categories are

Crohn's disease (CD) and ulcerative colitis (UC),

with both overlapping and distinct clinical and

pathological features (Charles, et al., 2009).

Ulcerative colitis is a chronic disease leading to

inflammation of the colon and in more severe degrees even

causing painful ulcers in the colon which can bleed, cause

mucous production and infection. Symptoms can recur or

be minimal for months and years. Common symptoms

include bloody diarrhea, abdominal pain and weight loss

which may be mild to severe and affect individual's quality

of life (Lakatos PL, et al., 2007).

In a meta-analysis performed by Mayo Clinic,

incidence of ulcerative colitis was reported as 2 to 14 per

thousand person-years (Mahid, et al., 2006).

The disease pathogenesis is still incompletely

understood. The genetic and environmental factors

such as altered luminal bacteria and enhanced the

intestinal permeability play a role in the dysregulation

of intestinal immunity, leading to the gastrointestinal injury

(Ricart E, et al., 2010).


- 2 -

Standard medical therapy is directed against the

inflammatory and immune processes that are known to play

an important role in the disease process. Medical therapy is

of variable success in ameliorating cardinal symptoms of

the disease (diarrhea, abdominal pain), in treating extra

intestinal manifestations, and in preventing complications


Currently, therapy is most often implemented in

a stepwise fashion, progressing through amino salicylates

[sulfasalazine, mesalazine (mesalamine)], corticosteroids,

immunosuppressive medications including tioguanine

(thioguanine) compounds (mercaptopurine, azathioprine),

methotrexate, and ciclosporin, and finally anti-TNF drugs.

This common approach is predicated on the addition of

more potent medications to agents that are believed

to be safer but that may also be less effective


Primary and secondary failure to respond to

approved therapies and, in some cases, inability to provide

a surgical solution to a particular patient due to extension

and \ or location of lesions represents unmet needs in the

treatment of IBD (Ricart E, et al., 2010).

A novel and exciting approach could be offered

through the current development in the field of

stem cell biology (Masson, et al, 2004).

Consequently, bone marrow stem cells have been

sought of as a promising new approach capable of

addressing mostly unmet medical needs (Weissman, 2000).


- 3 -

The considerable excitement surrounding the stem

cell field is based on the unique biological properties of

these cells and their capacity to self-renew and regenerate

tissue and organ systems, a flurry of studies reported bone

marrow derived stroma to brain, bone marrow to

liver, skin to brain, brain to heart and other such

stem cells differentiation (Morrison, 2000).

Two streams of research, experimental and clinical,

are the origin of the increasing utilization of stem cell

therapies for severe immune-mediated diseases (IMIDs)

including IBD. The considerable excitement surrounding

the stem cell field was initially based on the unique

biological properties of these cells; later, the

immunomodulatory ability of stem cell therapy has become

also apparent (Ricart E, et al., 2010).


- 4 -

Aim of the Work

To investigate the role of autologous bone marrow

stem cells intravenous injection in treatment for cases of

ulcerative colitis disease.

Ulcerative Colitis Chapter 1

- 5 -

Ulcerative Colitis

Inflammatory bowel disease:

Inflammatory bowel disease (IBD) commonly refers

to ulcerative colitis (UC) and Crohn's disease (CD), which

are chronic inflammatory diseases of the GI tract of

unknown etiology (Hyams, 2002).

Ulcerative colitis is characterized by diffuse mucosal

inflammation limited to the colon. It is classified according

to the maximal extent of inflammation observed at

colonoscopy, while Crohn's disease is characterized by

patchy, trans mural inflammation, which may affect any

part of the gastrointestinal tract, it may be defined by: age

of onset, location, or behavior (Silverberg, et al., 2005).

In particular, the definitions of ulcerative colitis

and Crohn's disease acknowledge the revised

Montreal classification which attempts to more

accurately characterize the clinical patterns of IBD

(Satsangi, et al., 2006).

Unclassified (IBDU) is the term best suited for the

minority of cases where a definitive distinction between

UC, CD, or other cause of colitis cannot be made after

considering clinical, radiological, endoscopic and

pathological criteria, because they have some features of

both conditions. Indeterminate colitis (IC) is a term

reserved for pathologists to describe overlapping features in

IBDU (Satsangi, et al., 2006).


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Ulcerative colitis:

Ulcerative colitis is a lifelong disease arising from an

interaction between genetic and environmental factors,

observed predominantly in the developed countries of the

world. The precise etiology is unknown and therefore

medical therapy to cure the disease is not yet available

(Dignass, et al., 2012).

It is a chronic inflammatory condition causing

continuous mucosal inflammation of the colon without

granulomas on biopsy, affecting the rectum and a variable

extent of the colon in continuity, which is characterized by

a relapsing & remitting course (Silverberg, et al., 2005).

Clinical disease activity is grouped into remission,

mild, moderate and severe. This refers to biological

activity and not to treatment responsiveness

(Rice-Oxley and Truelove, 1950).

The term severe colitis (or „acute severe colitis‟) is

preferred to „fulminant‟ colitis, because the term

„fulminant‟ is ill-defined. Severe colitis as defined

according to Truelove and Witt's' criteria is easy to apply in

outpatients, mandates hospital admission for intensive

treatment and defines an outcome (only 70% respond to

intensive therapy) (Dignass, et al., 2012).

Response is defined as clinical and endoscopic

improvement, depending on the activity index used.

In general, this means a decrease in the activity index

of >30%, plus a decrease in the rectal bleeding and

endoscopy sub scores, but there are many permutations

(D'Haens, et al., 2007).


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The term relapse is used to define a flare of

symptoms in a patient with established UC who is in

clinical remission, either spontaneously or after medical

treatment. It is considered when a combination of rectal

bleeding with an increase in stool frequency and abnormal

mucosa at sigmoidoscopy are present. It may be infrequent

(≤1/year), frequent (≥2 relapses/year), or continuous

(persistent symptoms of active UC without a period of

remission) (D'Haens G et al., 2007).

The term „chronic active disease‟ has been used in

the past to define a patient who is dependent on, refractory

to, or intolerant of steroids, or who has disease activity

despite immunomodulators. Since this term is ambiguous it

is best avoided. Instead, arbitrary, but more precise

definitions are preferred, including steroid-refractory or

steroid-dependence (Van Assche, et al., 2010).

Steroid-refractory colitis if patients have active

disease despite prednisolone up to 0.75 mg/kg/day over a

period of 4 weeks. Steroid-dependent colitis patients who

are either unable to reduce steroids below the equivalent of

prednisolone 10 mg/day within 3 months of starting

steroids, without recurrent active disease, or who have

a relapse within 3 months of stopping steroids

(Van Assche, et al., 2010).

Immunomodulator-refractory colitis patients who

have active disease or relapse in spite of thiopurines at an

appropriate dose for at least 3 months (i.e. azathioprine 2–

2.5 mg/kg/day or mercaptopurine 1– 1.5 mg/kg/day in the

absence of leucopenia) (Dignass, et al., 2012).


- 8 -

Classifications:

A. Classification according to disease extent

The preferred classification is an endoscopic

classification as outlined in the Montréal classification

into ulcerative proctitis (limited to the rectum), left-sided

colitis (up to the splenic flexure) and extensive colitis, and

by maximal extent upon follow up (Dignass, et al., 2012).

There are several reasons why patients with UC

should be classified according to disease extent. The extent

of inflammation will influence the patient's management

and the choice of delivery system for a given therapy. For

instance, topical therapy in the form of suppositories (for

proctitis) or enemas (for left-sided colitis) is often the first

line choice, but oral therapy often combined with

topical therapy is appropriate for extensive colitis.

Also, it influences start and frequency of surveillance


Table (1): The Montreal classification of UC (Silverberg, et al., 2005).

E1 Proctitis

Involvement limited to the rectum (i.e.

proximal extent of inflammation is distal to

recto-sigmoid junction)

E2 Left-sided

Involvement limited to the proportion of the

colon distal to the splenic flexure

(analogous to „distal‟ colitis)

E3 Extensive Involvement extends proximal to the

splenic flexure, including pan colitis


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B. Classification according to disease severity

Classification of UC based on disease severity is

useful for clinical practice and dictates the patient's

management. Many disease activity indices or criteria have

been proposed, but none have been adequately validated.

Although modifications of the original Truelove and Witts'

criteria are used in daily practice, the modified Mayo score

is used more frequently in current clinical trials.

For clinical practice a combination of clinical

features, laboratory findings, imaging modalities and

endoscopic parameters including histopathology will

all assist physicians in their patients' management


A distinction should be made between disease

activity at a point in time (remission, mild, moderate,

severe) and the response of disease to treatment. Moderate

colitis has become necessary to distinguish from mildly

active disease, because the efficacy of some treatments may

differ. The simplest clinical measure to distinguish

moderate from mildly active colitis is the presence of

mucosal friability (bleeding on light contact with the rectal

mucosa at sigmoidoscopy) (D'Haens, et al., 2007).

There is no fully validated definition of remission.

The best way of defining remission is a combination of

clinical parameters (i.e. stool frequency ≤ 3/day with no

bleeding) and a normal mucosa at endoscopy. Absence of

an acute inflammatory infiltrate at histology is helpful



- 10 -

Table (2): Disease severity index of UC (Truelove and Witts, 1995).

Mild Moderate Severe Bloody stools/day < 4 4 - 6 ≥ 6

Pulse < 90 bpm ≤ 90 bpm > 90 bpm

Temperature < 37.5 °C ≤ 37.8 °C > 37.8 °C

Hemoglobin > 11.5 g/dL ≥ 10.5 g/dL < 10.5 g/dL

ESR < 20 mm/h ≤ 30 mm/h > 30 mm/h

CRP Normal ≤ 30 mg/L > 30 mg/L

Table (3): Mayo activity scoring index (D'Haens, et al., 2007).

0 1 2 3 Stool

frequency Normal

1-2/day

> normal

3-4/day

> normal

5/day

> normal

Rectal

bleeding None Streaks Obvious

Mostly

blood

Endoscopic

finding Normal

Mild

friability

Moderate

friability

Spontaneous

bleeding

Global

assessment Normal Mild Moderate Severe

The Mayo score ranges from

0 to 12, with higher scores

indicating more severe disease.

0 to 1: Remission

2 to 5: Mild disease

6 to 9: Moderate disease

10 to 12: Severe disease

Table (4): Endoscopic scores for UC (Dignass, et al., 2012).

Baron Score (Baron JH, et al., 1964)

0 Normal: matt mucosa, ramifying vascular pattern clearly

visible, no spontaneous bleeding, no bleeding to light touch

1 Abnormal, but non-hemorrhagic: appearances between 0 and 2

2 Moderately hemorrhagic: bleeding to light touch,

but no spontaneous bleeding seen on initial inspection

3 Severely hemorrhagic: spontaneous bleeding seen ahead of

instrument at initial inspection and bleeds to light touch

Schroeder Score (Schroeder KW, et al., 1987)

0 Normal or inactive disease

1 Mild (erythema, decreased vascular pattern, mild friability)

2 Moderate (marked erythema, absent vascular pattern, friability, erosions)

3 Severe (spontaneous bleeding, ulceration)

Feagan Score (Feagan BG, et al., 2005)

0 Normal, smooth, glistening mucosa, with vascular pattern visible; not friable

1 Granular mucosa; vascular pattern not visible; not friable; hyperemia

2 As 1, with a friable mucosa, but not spontaneously bleeding

3 As 2, but mucosa spontaneously bleeding


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Pathophysiology:

Increasing evidence suggests that there is a defect in

the function of the intestinal immune system.

As a consequence, there is a breakdown of the defense

barrier of the gut, which, in turn, results in exposure of the

mucosa to microorganisms or their products. The result is

a chronic inflammatory process mediated by T-cells.

Hence, therapy should be directed at improving the

intestinal immune system. It has been postulated that

genetic factors may predispose certain individuals to

developing a "leaky gut" (William Tremaine, et al., 2008).

In ulcerative colitis, inflammation always begins in

the rectum, extends proximally a certain distance, and then

abruptly stops. A clear demarcation exists between

involved and uninvolved mucosa and no "skip areas" are

present. It primarily involves the mucosa and submucosa,

with formation of crypt abscesses and mucosal ulceration.

The mucosa typically appears granular and friable. The

small intestine is never involved, except when the distal

terminal ileum is inflamed in a superficial manner, referred

to as backwash ileitis (William Tremaine, et al., 2008).

In severe cases, pseudo polyps form, consisting of

areas of hyperplastic growth with swollen mucosa

surrounded by inflamed mucosa with shallow ulcers.

Necrosis can extend below the lamina propria to involve

the submucosa and the circular and longitudinal muscles,

although this is unusual. As the disease becomes chronic,

the colon becomes a rigid foreshortened tube that

lacks its usual haustral markings, leading to the

lead pipe appearance observed on barium enema

(William Tremaine, et al., 2008).


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Etiology:

The etiology of IBD is unknown. Environmental,

infectious, genetic, autoimmune, and host factors have

been suspected. Interactions among these factors may be

more important (Buhner, et al., 2006).

A. Genetic Factors

IBD is seen two to four times greater in the Jewish

population as compared with other ethnic groups.

Ashkenazi Jews have the greatest risk within the Jewish

population. Other epidemiologic studies have shown higher

rates in whites, lower rates in African Americans, and the

lowest rates in Asians (Ahmad, et al., 2001).

The prevalence of IBD is also increased in relatives

of those who have CD and UC. For patients who have UC,

the occurrence of IBD in their offspring was 6.26%; for

patients who have CD, the occurrence was 9.2%

(Orholm, et al., 1999).

Epidemiologic studies demonstrate familial

similitude for disease type, extent and extra-intestinal

manifestations for siblings with UC, but the concordance

rates are smaller than for CD. All studies that included the

evaluation of concordance rates between monozygotic and

dizygotic twins indicate that the genetic contribution to

disease susceptibility is smaller for UC than for CD

(Halfvarson, et al., 2003).

The region of the major histocompatibility complex

(MHC) locus on chromosome 6p that contains the genes

encoding the HLA Class I and II histocompatibility

molecules has been implicated in susceptibility to UC by

both association and linkage studies; however, the linkage


- 13 -

studies do not discriminate between risk for UC and CD.

Specifically, UC has been most consistently associated with

HLA Class II alleles (Satsangi, et al. 2003).

For example, in some populations the HLA-

DRB1*1502 allele (representing HLA-DR2) is positively

associated with UC and the HLA-DR4 and DR6 alleles

negatively associated; the differences in association among

populations may be accounted for by racial and ethnic

variability. The infrequent HLA-DRB1*0103 allele is

associated with extensive and severe UC and often

associated with the requisite for colectomy. Although there

is conflicting data in differing populations, other potential

genetic associations for UC include the interleukin-1 family

of genes on chromosome 2q13. Another potential

'functional candidate gene' is the multidrug resistance gene

(MDR1) that is located in an area of linkage on

chromosome 7 (Pallone; Silverberg; Ahmad, et al. 2003).

In addition, there is a strong likelihood that genetics

also impact on the incidence of extra-intestinal

complications of UC. In particular, the association between

HLA-B27 and the development of ankylosing spondylitis

and sacroiliitis in patients with UC has been reproduced

and approaches 100%. Peripheral arthropathies (type I and

II) accompanying UC are also associated with HLA

polymorphisms that associate with erythema nodosum and

uveitis (Orchard, et al. 2002).

Of note, the association of UC with primary

sclerosing cholangitis is also related to the presence of

several HLA Class II alleles and is modified (prevented)

by the environmental factor of cigarette smoking

(Mitchell, et al. 2002).


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B. Environmental Factors

Cigarette smoking:

There are several environmental clues to

susceptibility and development of UC (Krishnan and

Korzenik, 2002).

The long-standing finding that cigarette smoking

protects against the development of ulcerative colitis has

withstood the test of time. Indeed, case series continue to

demonstrate a protective effect of smoking on both the

development and course of UC (Abraham, et al., 2003).

Although smokers are less likely to develop UC, however,

ex-smokers are more likely to develop extensive or severe

colitis. Others believe that ex-smokers account for the

preponderance of the second age peak for UC in

patients > 40 years (Halme, et al., 2002).

The protective effect of smoking also extends to

the extra-intestinal manifestations and the post-surgical

complications of UC. For example, smoking protects

against the development of PSC, smoking, or non-smoking,

accounts for the differing incidence of PSC associated

with UC (Mitchell, et al., 2002).

Appendectomy:

Another consistent epidemiologic clue to the

pathogenesis of UC is the observation that appendectomy,

particularly at a younger age, both reduces the likelihood of

developing and the severity of disease. It seems to be an

additive protective factor to cigarette smoking against the

development of UC. In contrast to UC, prior appendectomy

does not seem to be protective against development of PSC

(Feeney et al.; Cosnes, et al.; Mitchell, et al., 2002).


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Bacteria:

One ubiquitous factor in animal models of colitis and

in human disease is the relationship with bacteria. In

experimental models of IBD, colitis does not develop in

animals that are raised in germ-free environments

(Sartor, 2004).

Commensal bacteria, not pathogens, are sufficient to

induce colitis, but this is determined by both host and

bacterial specificities. Also, different phenotypic patterns of

colitis are seen with specific bacterial species. Commensal

bacteria can induce a protective effect that can be

transmitted by bacteria-responsive regulatory CD4+ T-cells

(Cong, et al. 2002).

Although it has not been possible to identify bacterial

strains that are specific to UC, there are increased numbers

of mucosa-associated (adherent) Bacteroides species and

Enterobacteriaceae species in patients with inflamed

segments (Swidsinski, et al. 2002).

Whether early exposure to common environmental

microbes is protective against UC as it is with other

autoimmune disorders, in line with the so-called hygiene

hypothesis, remains to be determined (Weiss, 2002).

Alternatively, functional activity of microbial strains

may also lead to 'DYSBIOSIS' and affect the metabolic

activity of colonocytes or enterocytes, leading to the

development of UC. The potential inductive or protective

role of bacteria has also led to considerable interest in

prebiotic or probiotic therapies for UC and its

complications (Sartor, 2004).


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C. Immunologic Factors

From an immunologic perspective, UC has less of a

Th1 response pattern than CD (Pallone, et al. 2003).

In IBD there are chronic inflammatory changes in the

GIT. These are mediated by different immunologic factors

for each disease, although they are both a consequence of

T-cell activation (Pallone and Monteleone, 1998).

The cytokine expression is different comparing

ulcerative colitis and Crohn's disease. In ulcerative colitis,

the inflammation is thought to be regulated by Th2-cells,

which mediate B cells and antibody responses; however

this has not been proven. It has been shown that there is

increased expression of IL-5, which is a Th2 cytokine,

but IL-4, another Th2 cytokine, is not increased

(Fuss, et al., 1996).

The Th2 contribution may be helping the antibody

response, because in UC, there is an increase in IgG plasma

cells presumably mediated by T-cells (Macdonald, 2000).

Recent evidence indicates that, in contrast to the Th1

cytokines that are associated with the pathogenesis of

Crohn's disease (interferon-γ, TNF-α and IL-12),

animal models of ulcerative colitis may be associated

with increased natural killer cell activity and IL-13

(Heller, et al. 2002).

In addition, attention is being directed at the down

regulatory role of transforming growth factor-ß in colitis

and the possibility that defective signaling of transforming

growth factor-ß may account for inadequate tissue repair

(Pallone, et al. 2003).


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Diagnosis:

A gold standard for the diagnosis of UC is not

available. The diagnosis is made on the basis of clinical

suspicion supported by appropriate macroscopic findings

on colonoscopy, typical histological findings on biopsy

and negative stool examinations for infectious agents


There is some evidence to suggest that patients with

UC stratified by age have different outcomes. Patients

diagnosed before the age of 16 had a more aggressive

initial course, while older age at diagnosis was found

to be associated with a lower risk of colectomy

(Barreiro-de-Acosta, et al., 2010).

There is also some evidence that UC diagnosed in

the very young has a different etiology and prognosis. This

is taken into consideration by the pediatric modification to

the Montréal classification (Levine, et al., 2011).

It is a disease that used to carry a high mortality and

major morbidity. With modern medical and surgical

management, the disease now has a slight excess of

mortality in the first 2 years after diagnosis, but little

subsequent difference from the normal population. The

clinical course is marked by exacerbation and remission.

About 50% of patients have a relapse in any year. An

appreciable minority has frequently relapsing or chronic,

continuous disease and overall, 20-30% of patients with

pancolitis come to colectomy. After the first year

approximately 90% of patients are fully capable of work

(defined by < 1 month off work/year), although significant

employment problems remain an issue for a minority

(Langholz, et al., 1994).


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Clinical picture

The onset may be gradual or sudden. The course is

variable, with periods of exacerbation, improvement, and

remission that may occur with or without specific medical

therapy (William Tremaine, 2008).

Symptoms of UC are dependent upon extent and

severity of disease. The cardinal symptom of ulcerative

colitis is bloody diarrhea. Diarrhea may vary from 1 to 20

or more loose or liquid stools a day, usually worse in the

morning and immediately after meals, and patients with

moderate or severe symptoms often have nocturnal stools.

Constipation with rectal bleeding is a presenting symptom

in about 25% of patients with disease limited to the rectum.

Abdominal pain is usually cramping, which is worse after

meals or bowel movements. Anorexia, weight loss, and

nausea in the absence of bowel obstruction are common

with severe and extensive disease but uncommon with mild

to moderate disease or disease limited to the left colon. In

children, urgency, incontinence, and upper gastrointestinal

tract symptoms are more frequent and growth failure is

common. It is associated with an equivalent increased risk

of colonic carcinoma (Friedman, et al., 2008).

A full medical history should include detailed

questioning about the onset of symptoms, particularly the

stool frequency and consistency, recurrent episodes of

rectal bleeding or bloody diarrhea, urgency, tenesmus,

abdominal pain, incontinence, nocturnal diarrhea, weight

loss, features of extra-intestinal manifestations, and

systemic symptoms of malaise, anorexia, or fever are

features of a severe attack (Dignass, et al., 2012).


- 19 -

History should include recent travel, food

intolerances, contact with enteric infectious illnesses,

medication (antibiotics and NSAIDs drugs), smoking habit,

sexual practice, vaccination, family history of IBD,

Colorectal cancer, and previous appendictomy should be

explored (Dignass, et al., 2012).

However, severe colitis is still a potentially

life-threatening illness. Immediate admission to

hospital is warranted for all patients fulfilling

Truelove and Witts' criteria for severe colitis to

prevent delayed decision making which may lead

to increased perioperative morbidity and mortality


In mildly active ulcerative colitis, physical

examination findings are often normal or there

may be abdominal tenderness, particularly with

palpation over the sigmoid colon. Patients with

more severe disease may have pallor, dehydration,

tachycardia, fever, diminished bowel sounds,

and diffuse abdominal tenderness with rebound.

Tenderness with rebound is ominous and suggests toxic

dilatation or perforation (Sands, 2004).

Also, it should include general well-being, pulse rate,

body temperature, blood pressure, measurement

body weight and height, calculation of BMI, abdominal

examination for distention and tenderness, palpable

masses, perianal inspection, digital rectal examination,

oral inspection, check for anemia, fluid depletion,

and check for eye, skin and/or joint involvement

(Sands, 2004).


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Ulcerative colitis is associated with a wide variety of

systemic complications which classically called extra-

intestinal manifestations, immune-mediated phenomena

that affect the joints, eye, skin, or hepatobiliary tract, but

they can be defined more broadly to include complications

in other organ systems and complications that arise as a

direct pathophysiologic consequence of extensive bowel

inflammation or resection. It occurs in up to 36% of

patients (Edward, 2008).

Arthritis affecting the axial skeleton can be classified

into the more common, frequently asymptomatic,

sacroiliitis and the less common, more progressive,

ankylosing spondylitis. Symptomatic sacroiliitis manifests

as low back pain and stiffness, typically worse in the

morning and with rest while spondylitis resulting in

progressive stiffness and lordosis of the spine. The

symptoms usually accompany exacerbations but may

appear before the disease and don't necessarily follow its

course (Johns Hopkins, 2013).

About 19% of patients with UC experience

dermatological changes. The two most common

dermatologic manifestations are pyoderma gangrenosum

and erythema nodosum. Other dermatological sequelae

include dermatitis, erythematous rash, psoriasis, carcinoma,

urticaria, pityriasis, lupus erythematosus, vitiligo and

ecchymosis (Edward, 2008).

Ocular complications occur in 1-13% of patients

with IBD. The most common forms are anterior uveitis

(also known as iritis) and scleritis. An inflammatory

retinopathy or keratitis (corneal inflammation) may occur

less frequently. Symptoms include headache, photophobia

and blurred vision (Johns Hopkins, 2013).


- 21 -

The most important hepatobiliary condition

associated with IBD is primary sclerosing cholangitis. This

idiopathic chronic cholestatic liver disease is characterized

by inflammation and fibrosis of the biliary tree.

Autoimmune hepatitis is associated rarely with IBD, but

when it is, it usually is associated with ulcerative colitis.

Some patients may have features of both autoimmune

hepatitis and primary sclerosing cholangitis (the so-called

overlap syndrome) (Edward, 2008).

In most situations, extra-intestinal manifestations

respond to standard medical therapy. On rare occasions,

a total proctocolectomy may be necessary to control

severe extra intestinal manifestations of this disease

(Edward, 2008).

Fig.(1): Extra-colonic manifestations of UC (Johns Hopkins, 2013)


- 22 -

Endoscopy

Flexible proctosigmoidoscopy or colonoscopy with

multiple biopsies (at least two biopsies from five sites

including the distal ileum and rectum) is the first line

procedure for diagnosing colitis. It allows classification of

disease based on endoscopic extent, severity of mucosal

disease and histological features. Active disease

confirmed sigmoidoscopy as a first line procedure


No endoscopic feature is specific for UC. The most

useful endoscopic features of UC are considered to be

continuous and confluent colonic involvement with clear

demarcation of inflammation and rectal involvement.

Endoscopic severity of UC may be best reflected by the

presence of mucosal friability, spontaneous bleeding and

deep ulcerations (Dignass, et al., 2012).

There are mucosal changes including loss of

the normal vascular markings, mucosal granularity,

mucosal friability, mucous exudate, and focal ulceration

(William Tremaine, 2008).

Fig.(2): Endoscopic image of ulcerative colitis (Samir, 2004)


- 23 -

With colonoscopy, the extent of disease can be

determined and the terminal ileum can be examined for

evidence of backwash ileitis in UC or ileal involvement in

CD. Patients with left-sided UC may have inflammatory

changes around the appendix, called a cecal patch,

as a manifestation of the disease; this finding should

not be confused with segmental colitis due to CD


In acute severe colitis, full colonoscopy is rarely

needed and may be contraindicated, because of the risk of

perforation or hemorrhage. Patients should have abdominal

radiography. Also, phosphate enema is considered safe,

except with colonic dilatation (Terheggen, et al., 2008).

Endoscopic findings are predictive of outcome at for

patients with UC in remission. Endoscopic reassessment is

appropriate at a relapse, or for steroid-dependent or -

refractory UC or when considering colectomy. Patients

with UC (extending proximal to the rectum), the risk of

malignancy is increased above that for the general

population after 8-10 years of disease. So, periodic

colonoscopy with biopsies for surveillance for dysplasia is

indicated after 8-10 years of disease. The risk for patients

with less extensive UC (with involvement of the colon

distal to the splenic flexure) also is increased, but the

magnitude of the risk is not defined. There doesn't appear

to be an increased risk of the rectal cancer for ulcerative

proctitis without colitis above the rectum. Patients with

left-sided UC of 8-10 years‟ duration, or longer, should

undergo periodic surveillance biopsies. The optimal

interval between surveillance examinations has not been

defined, and the examinations usually are performed

at 1-2 year intervals (William Tremaine, 2008).


- 24 -

Table (5): UC Endoscopic Index of Severity (UCEIS) (Travis, et al., 2012)

Vasc

ula

r p

att

ern

Normal (0)

Normal vascular pattern with

arborisation of capillaries clearly

defined, or with blurring or patchy

loss of capillary margins

Patchy obliteration

(1) Patchy obliteration of vascular pattern

Obliterated (2) Complete obliteration of vascular pattern

Ble

edin

g

None (0) No visible blood

Mucosal (1)

Some spots or streaks of coagulated blood on

the surface of the mucosa ahead of the scope,

which can be washed away

Mild (2) Some free liquid blood in the lumen

Moderate

or severe (3)

Frank blood in the lumen ahead of

endoscope or visible oozing from mucosa

after washing intra-luminal blood, or visible

oozing from a hemorrhagic mucosa

Ero

sio

ns

& U

lcer

s None (0) Normal mucosa, no visible erosions or

ulcers

Erosions (1) Tiny (≤5 mm) defects in the mucosa, of a

white or yellow color with a flat edge

Superficial

ulcer (2)

Larger (N5 mm) defects in the mucosa,

which are discrete fibrin-covered

ulcers when compared to erosions,

but remain superficial

Deep ulcer (3) Deeper excavated defects in the mucosa,

with a slightly raised edge

Fig. (3): Histology of

normal colon and UC

(Johns Hopkins, 2013)


- 25 -

Histology

Histopathological examination of biopsy specimens

should be carried out according to the BSG guideline, „A

Structured Approach to Colorectal Biopsy Assessment‟

(Jenkins, et al., 1997).

For a reliable diagnosis of ulcerative colitis multiple

biopsies from five sites around the colon (including the

rectum) and the ileum should be obtained. Multiple implies

a minimum of two samples. Repeat biopsies after an

interval may help to solve differential diagnostic problems

and establish a definitive diagnosis especially in adults, by

showing additional features (Dignass, et al., 2012).

There should be an attempt to define the type of IBD,

to mention other coexistent diagnoses or complications and

to mention the absence or presence of any dysplasia and its

grade. Medical and surgical therapy may modify the

histological appearances of IBD and these should be taken

into account when assessing IBD biopsy pathology

(Hyde, et al., 2002).

Also, mucosal biopsy specimens from involved areas

of the gastrointestinal tract are useful for excluding self-

limited colitis and other infections and non-infectious

colitis due to ischemia, collagenous and lymphocytic

colitis, drug effect, radiation injury, and solitary rectal ulcer

syndrome. Non-caseating granulomas are a feature of CD

and can be helpful for distinguishing it from UC


A hallmark of active UC is the presence of a

polymorphonuclear cell infiltration into the epithelial crypts

(cryptitis) and lamina propria (Beckmann, et al., 2007).


- 26 -

Radiologic Features

Imaging can be helpful in diagnosis, assessment of

disease extent and severity and for investigation of

suspected complications. Each modality has its own

advantages and drawbacks and the tests are often

complimentary (Hall and Brenner, 2008).

Barium fluoroscopy (Double contrast barium

enema) allows for exquisite detail of the colonic mucosa,

and also allows bowel proximal to strictures to be assessed.

It is however contraindicated if acute severe colitis is

present due to the risk of perforation. Mucosal

inflammation lends a granular appearance to the surface of

the bowel. Mucosal ulcers are undermined (button-shaped

ulcers). When most of the mucosa has been lost, islands of

mucosa remain giving it a pseudo-polyp appearance. In

chronic cases the bowel becomes featureless with loss of

normal haustral markings, luminal narrowing and bowel

shortening (lead pipe sign). Small islands of residual

mucosa can grow into thin worm like structures (so-called

filiform polyps) (Roggeveen, et al., 2005).

A. Pseudo-polyp-------- B. Target sign ------- C. Loss of haustration

Fig.(4): Radiological features in UC (Gore, et al., 1996)


- 27 -

Computed Tomography (CT) will reflect the same

changes that are seen with a barium enema, with the

additional advantage of being able to directly visualize the

colonic wall, the terminal ileum and identify extra-colonic

complications, such as perforations or abscess formation. It

is important to note however that CT is insensitive to early

mucosal disease (Gore, et al., 1996).

A cross section of the inflamed and thickened bowel

is having a target appearance, due concentric rings of

varying attenuation, also known as mural stratification. In

chronic cases, submucosal fat deposition is seen

particularly in the rectum (fat halo sign). Strictures are also

common, and are not all malignant. Colorectal carcinoma is

often sessile. Focal loss of mural stratification or

excessive mural thickness (1.5 cm) should prompt

endoscopic evaluation (Gore, et al., 1996).

The current status of Magnetic Resonance Imaging

(MRI) in UC is that of a promising, noninvasive technique

for imaging extent of more severe disease. The most

striking abnormalities in UC are wall thickening and

increased enhancement. The median wall thickness in UC

ranges from 4.7-9.8 mm. In general, the more is severe the

inflammation, the thicker the colonic wall. A colonic wall

thickness <3 mm is usually considered as normal, 3-4 mm

as a "gray zone," and >4 mm as pathological. Enhancement

of the mucosa with no or less enhancement of

the submucosa is producing a low SI strip the

so-called submucosal stripe. Other features are the loss

of haustral markings, backwash ileitis shows

mild enhancement and no wall thickening and

there is increased SI of the pericolonic fat noted

(Gore, et al., 1996).


- 28 -

Plain film is nonspecific but may show evidence of

mural thickening. Ultrasound cannot comprehensively

assess the gut when used in isolation. Doppler are useful in

the assessing the degree of disease activity. It has

reasonable sensitivity for documenting presence of

complicating abscess, particularly in thinner patients and is

a useful first line test in this context (Dietrich, 2009).

Virtual colonography is an evolving technology.

The limited data currently available do not demonstrate a

diagnostic value for assessing the disease extent in patients

with suspected or proven UC (Dignass, et al., 2012).

Recent studies assess the assessment of the severity

of ulcerative colitis using endorectal ultrasonography

(ERUS) corresponds with clinical severity of the disease.

ERUS is a valuable, relatively cost-effective diagnostic tool

of high overall accuracy, which may be helpful in clinical

evaluation and monitoring of ulcerative colitis


A variety of nuclear medical techniques can be used

in the assessment of IBD, but they have no role in primary

diagnosis. Technetium-99m labelling of WBCs remain a

widely acceptable scintigraphic method for the evaluation

of disease extension and severity. Positron emission

tomography alone or with CT using fluorine-18

fluorodeoxy glucose is a promising method of measuring

inflammation in IBD. These techniques considered when

colonoscopy is not completed successfully or other imaging

modalities are negative (Stathaki, et al., 2009).


- 29 -

Laboratory Findings :

Laboratory studies are of value in assisting with the

management of IBD but are of minimal help in establishing

the diagnosis. Laboratory values may be used as surrogate

markers for inflammation, nutritional status and to look for

deficiencies of necessary vitamins and minerals. Serologic

studies have been proposed to help diagnose IBD and to

differentiate CD from UC (Kefalides and Hanaeur, 2002).

With new-onset IBD or at relapse, infection should

be ruled out with stool studies, including cultures for

bacterial pathogens and examinations for ova and parasites

and Clostridium difficile toxin. Any patient hospitalized

with a flare of colitis should, at a minimum, have a C

difficile toxin assay performed because, commonly,

pseudomembranous colitis is super-imposed on UC

(Kefalides and Hanaeur, 2002).

Also, there is an exudation of inflammatory cells into

the colonic lumen that can be identified as the presence of

fecal leukocytes (William Tremaine, 2008).

Acute phase reactants, including the ESR, CRP,

and transferrin, usually correlate with disease activity but

may be normal in mildly active disease. Hypokalemia, and

metabolic acidosis can occur with severe disease because of

their wasting with diarrhea (William Tremaine, 2008).

Blood tests can also be used to help determine

nutritional status. The most commonly used marker is

serum albumin and prealbumin. Hypoalbuminemia may

reflect malnutrition; it can also develop because of the

protein-losing enteropathy that can occur with active IBD

(Kefalides and Hanaeur, 2002).


- 30 -

Complete blood cell count (CBC) can be useful

indicators of disease activity and iron or vitamin

deficiency. An elevated WBC count is common in patients

with active inflammatory disease or due to a complicating

abscess but it does not necessarily indicate infection.

Anemia is common and may be an anemia of chronic

disease, presumably due to cytokine effects on the bone

marrow or an iron deficiency anemia due to blood loss that

is confirmed by serum iron studies or as a result of

malabsorption of vitamin B12 or folate especially with CD

(Kefalides PT and Hanaeur SB, 2002)

Cytomegalovirus (CMV) should be considered in

severe or refractory colitis, as reactivation is common in

patients with IBD on immunosuppression. Additional tests

may need for patients who have travelled abroad


Several new fecal tests have become available that

assist, initially, with the diagnosis of IBD in general, but

not specifically UC. Perhaps in the future they will

contribute to measurements of disease activity. For

example, elevated concentrations of calprotectin,

a neutrophil granulocyte-derived Ca++

binding protein,

have been evaluated as both a diagnostic assay to

identify inflammatory diarrhea and as measurement of

inflammatory activity (Beckmann, et al., 2007)

Similarly, the concentration of lactoferrin, another

neutrophil granulocyte-derived protein, can be quantified,

most recently by using a polyclonal antibody-based enzyme

linked immunoassay that can discriminate between active

IBD and IBS. It can quantify disease activity but has yet to

be incorporated into clinical trials as a disease-activity

endpoint (Summerton, et al., 2002; Kane, et al., 2003).


- 31 -

The peri-nuclear antineutrophil cytoplasmic

antibody (pANCA) is positive in about 2/3 of patients with

UC and about 1/3 of patients with CD while the

anti-Saccharomyces cerevisiae antibody (ASCA) is

positive in about 2/3 of patients with CD and about 1/3 of

patients with UC. These tests used together to help

distinguish UC from CD. However, the positive predictive

value of the two tests together is 63.6% for UC and 80%

for CD; thus, distinguishing the two diseases with these

tests is less than ideal (William Tremaine, 2008).

There are several potential roles for serologic

markers in UC. The most desirable would be as a

pathognomonic marker of disease specificity or prognosis,

the second as a screening tool to discriminate IBD

from other digestive disorders, and the third to assist in

the understanding of disease mechanisms or

immune interactions (Dubinsky, et al., 2002).

In adult populations the ability to diagnose UC by

flexible sigmoidoscopy or colonoscopy is less of an issue

and offers immediate confirmation and access to histology.

So, serologic markers have, thus far, not been necessary to

screen or exclude UC compared with conventional

investigations. The potential for serologic studies to

discriminate between UC & CD, particularly for patients

with 'indeterminate colitis' has been evaluated by

(Joossens, et al., 2002).

They found that the presence of pANCA does not

help to discriminate between UC and CD in patients with

IC. By contrast, the presence of anti-Saccharomyces

cerevisiae antibodies is more helpful, but their sensitivity in

indeterminate colitis is so low that a positive or negative

predictive value will be low (Vasiliauskas, 2003).


- 32 -

Nutritional aspects in UC

The inflammation of the GIT with the associated

symptoms of pain, nausea, and diarrhea is leading to

reduced food intake and uptake leading to malnutrition.

The prevalence of nutritional deficiencies and malnutrition

is higher in patients with CD than in patients with UC

(Lochs, et al., 2006).

The etiology of malnutrition is multifactorial.

Medication, increased exudative losses of protein or small

bowel bacterial overgrowth is additional causes of

malabsorption (Han, et al., 1999).

Anemia and iron deficiencies are more prevalent in

patients with UC. In patients with severe diarrhea,

low levels of potassium, magnesium, calcium and

phosphate can be encountered. The levels of fat

soluble vitamins correlate with the severity of steatorhea

(Remy Meier, 2008).

Table (6): Nutritional deficiencies in IBD (Remy Meier, 2008).


- 33 -

The diagnosis of malnutrition is done by using

several anthropometric and biochemical parameters as

weight, height, skinfold thickness, body composition

analysis, and serum albumin. It is also important to record

the food intake in the last 1-2 weeks. Patients often

consume an unbalanced diet which may lead to nutritional

deficiencies (Vagianos, et al., 2007).

Resting energy expenditure varies depending on

inflammatory activity. The Energy requirements have been

calculated with the Harris-Benedict equation. However,

physical activity, inflammatory activity, malabsorption and

the degree of obesity should be taken into account. The

more obese the less energy/kg is required and vice versa.

Consequently requirements may be calculated on the basis

of ideal BW (or adjusted BW) and may amount to 25-30

Cal/kg ideal BW/24h (Lochs, et al., 2006).

Protein requirements in patients with IBD are

generally increased. Inflammation induces a catabolic

response with endogenous proteolysis and ensuing negative

nitrogen balance. To limit nitrogen losses in patients with

active IBD, 1.5 g/kg BW protein per day should be

provided. It is recommended to increase protein intake to

2g protein/kg BW/24 h in infectious or severely

malnourished patients (Han, et al., 1999).

The nutritional support required during an acute

exacerbation differs from the nutritional regimen during

remission. The aims of nutritional support in IBD are to

treat or to prevent nutritional deficits, to reduce disease

activity, to improve growth and development in children

and adolescents, to reduce the need for surgery or

aggressive medical treatment, and to maintain remission

(Remy Meier, 2008).


- 34 -

There are no specific data available for ulcerative

colitis to improve or to maintain nutritional status with oral

supplements. Specific nutritional support is more beneficial

in Crohn's disease than in ulcerative colitis. Enteral

nutrition is the preferred route for nutritional repletion

because of the potential trophic effects on the intestinal

mucosa, the preservation of gastrointestinal function, the

beneficial effects on the intestinal flora and mucosal barrier

integrity (Harries, et al., 1983).

In addition, complication rates and costs have been

reported to be lower than with parenteral nutrition. For a

long time, it was suggested that bowel rest with TPN may

reduce intestinal inflammation and decrease disease activity

in patients with IBD. Parenteral nutrition is of limited

benefit in ulcerative colitis. Sometimes parenteral nutrition

is instituted to decrease a debilitating defecation frequency,

when patients are hospitalized with acute toxic colitis

(Remy Meier, 2008).

Enteral tube feeding has shown to be ineffective in

patients with active UC. The remission rates are not higher

than the rate of spontaneous remission when patients

consume normal food. A comparison of TPN with EN in

acute UC showed similar effects on nutritional status,

disease activity and complications, but neither TPN

nor EN had a positive effect on inflammatory activity

(González-Huix, et al., 1993).

Pre- and probiotics have been shown to be beneficial

in gastrointestinal diseases. Prebiotics are soluble poli- or

oligo-saccharides and serve in the intestine as substrates for

fermentation (Johannsson, et al., 1997).


- 35 -

Probiotics are non-pathogenic bacteria which are

able to exert positive health benefits in the gastrointestinal

tract. They are able to adhere to the intestinal mucosa and

can stimulate the secretion of serum IgA and mucus

production. They may reduce the levels of pro-

inflammatory cytokines and increase levels of anti-

inflammatory cytokines. Also, they can produce defenses

and heat shock proteins (Johannsson, et al., 1997).

Pre- and probiotics can interact with the commensal

bacteria and may therefore influence the intestinal

ecosystem. This effect is eminent in the colon, where

anaerobic bacteria can ferment non-absorbable dietary

carbohydrates. Through fermentation, the intestinal pH

decreases, which is stimulates the growth of non-

pathogenic bacteria and liberate short chain fatty acids.

Butyrate is the main energy source for the colonic epithelial

cells. This prevents the expression of specific genes

encoding cytokines intensifying inflammatory response.

Also, it increases apoptosis of inflammatory cells. So far,

the use of pre-and probiotics was found to be more

beneficial in UC than in CD. The use of a fermentable

Plantago ovate (dietary fibre) supplementation

achieved similar relapse rates in UC as Mesalazine

(Buda, et al., 2003; Gionchetti, et al., 2000).

Most patients with IBD in remission have a normal

nutritional status. There are no specific diets recommended

if the patients are in remission and can eat normally. A

normal "healthy" diet rich in fruits, vegetables and fish can

be recommended. In patients with ileum resection or

sulfasalazine treatment, vitamin B12 levels have to be

monitored. Ca++

and vitamin D status should be controlled

in patients treated with steroids (Belluzzi, et al., 1996).


- 36 -

Traditional pharmacological treatment of UC:

Therapy for IBD is a rapidly evolving field, with

many new biological agents under investigation that are

likely to change therapeutic strategies radically in the next

decade (Mowat, et al., 2011). Details of the principal drugs

can only be summarized in this document.

Amino salicylates

5-Aminosalicylic acid (5-ASA) or mesalazine

(„mesalamine‟ in the USA) can be delivered in millimolar

concentrations to the gut lumen by a variety of oral tablets,

sachets or suspensions using pH-dependent release

mechanism, multi-matrix delivery systems, or conjugation

via a diazo bond to a variety of carrier molecules with

release of 5-ASA after splitting by bacterial enzymes in

the large intestine. They can also be used as topical agents

in the form of liquid or foam enemas, or suppositories

(Sandborn and Hanauer, 2003).

They act on epithelial cells by a variety of

mechanisms to moderate the release of lipid mediators,

inflammatory cells, cytokines and reactive oxygen species

(Sutherland and Macdonald, 2006).

The choice of 5-ASA is debated, but is influenced by

tolerability (mesalazine is tolerated by 80% of those

unable to tolerate sulfasalazine), dose schedule (single or

twice daily dosing with better compliance), route of

delivery, availability and cost are relevant factors in

choice (Dignass, et al., 2009).


- 37 -

For UC, greater clinical improvement is associated

with doses of ≥ 2.0 g/day are more effective than < 2.0

g/day. Clinical improvement characteristically occurs at

twice the remission rate (Bergman and Parke, 2006).

There is now robust evidence to suggest that single

daily dosing is as effective as multiple dosing, and may

even be superior. Maintenance with all 5-ASA drugs may

reduce the risk of colorectal cancer by up to 75%

(Eaden, et al., 2000).

Side effects of 5-Aminosalicylic acid occur in

10-45% of patients, depending on the dose. Headache,

nausea, epigastric pain, diarrhea, thrombocytopenia,

rash and oligospermia in men are most common. Serious

idiosyncratic reactions (including Stevense Johnson

syndrome, pancreatitis, agranulocytosis, or alveolitis)

are rare (Van Staa, et al., 2004).

Also, they associated with nephrotoxicity (including

interstitial nephritis and nephrotic syndrome), which

appears both to be idiosyncratic and, in part, dose related

(Muller, et al., 2005).

For patients on maintenance 5-ASA, many clinicians

believe that creatinine and full blood count should be

monitored every 3–6 months and it should be stopped if

renal function deteriorates (Muller, et al., 2005).


- 38 -

Corticosteroids

Corticosteroids are used in the form of oral

prednisolone, prednisone, budesonide, or intravenous

hydrocortisone and methylprednisolone. Topical

suppositories, foam or liquid enemas also used. Many

strategies attempt to maximize topical effects

while limiting the systemic side effects of steroids

(Seow, et al., 2009).

Corticosteroids are potent anti-inflammatory agents

for moderate to severe relapses of UC. They have no role in

maintenance therapy (Manguso and Balzano, 2007).

They act through inhibition of several inflammatory

pathways: suppressing interleukin transcription,

suppression of arachidonic acid metabolism and stimulation

of apoptosis of lymphocytes within the lamina propria of

the gut (Benchimol, et al., 2008).

A combination of oral and rectal steroids is better

than either alone. Adverse events are significantly more

frequent at a dose of 60 mg/day compared to 40 mg/day,

without added benefit, so 40 mg/day appears optimal for

outpatient management of acute UC (Lee, et al., 1996).

Budesonide (colonic release preparation) is a poorly

absorbed corticosteroid with limited bioavailability and

extensive first-pass metabolism that has therapeutic benefit

with reduced systemic toxicity in ileo-caecal CD, or UC.

Beclometasone dipropionate has been studied in oral and

enema forms in UC, and is no better than 5-ASA also, it

appears as effective as prednisolone for mild - moderate

left-sided and extensive colitis (Campieri, et al., 2003).


- 39 -

Regimens of steroid therapy are various. There is no

evidence to support any particular regimen. The commonly

used regimen is done by a starting dose of 40 mg

prednisolone per day, reducing by 5 mg/d at weekly

intervals. A standard weaning strategy helps identify

patients who relapse rapidly or do not respond and

need adjunctive therapy. Shorter courses (< 3 weeks)

are associated with early relapse and doses of

prednisolone ≤ 15 mg day are ineffective for active disease

(Kane, et al., 2002).

Steroid resistance or unresponsiveness should

lead to escalation of treatment, or consideration of

surgery. Medical therapies include an immunosuppressive

appropriate to the acuteness and type of the disease

(Kane, et al., 2002).

Side effects of corticosteroids due to supra-

physiological doses include cosmetic (acne, moon face,

edema), sleep and mood disturbance, dyspepsia or glucose

intolerance (Subramanian, et al., 2008). Prolonged use

(usually >12 weeks, but sometimes less) include cataracts,

osteoporosis, osteonecrosis of the femoral head, myopathy

and susceptibility to infection (Newby, et al., 2005).

Efficacy should be balanced against side effects, but

decisive treatment of active disease in conjunction with a

strategy for complete withdrawal of steroids is often

appreciated by a patient suffering miserable symptoms

during withdrawal acute adrenal insufficiency and

corticosteroid withdrawal syndrome may occur

(Lichtenstein, et al., 2006). Guidelines recommend

monitoring for eye, bone and other side effects particularly

in patients on steroids for more than 3 months

(Abreu, et al., 2006).


- 40 -

Thiopurines

Azathioprine (AZA) or mercaptopurine (MP) is

widely used in UC and CD as adjunctive and sparing

therapy. Purine antimetabolites inhibit ribonucleotide

synthesis, but mechanism of immunomodulation is by

inducing T-cell apoptosis by modulating cell (Rac1)

signaling (Tiede, et al., 2003).

AZA is non-enzymatically metabolized to MP,

which involves loss of a nitro-imidazole side chain; this is

thought to explain some of the side effects seen with AZA

and which may be less of a problem with MP. MP is

subsequently metabolized to 6-thioguanine nucleotides (6-

TGN). 6-TGN has been used for treatment of IBD with

caution because of potential hepatotoxicity

(McGovern, et al., 2002; Bowen and Selby, 2000).

AZA is more effective than mesalazine at induction

of clinical and endoscopic remission in steroid dependent

ulcerative colitis and should be first-choice therapy in this

situation providing other causes of persistent symptoms

such as CMV or cancer have been excluded. Thiopurines

are effective maintenance therapy for patients with UC who

has failed or who cannot tolerate mesalazine and for

patients who require repeated courses of steroids. The

evidence for using thiopurines in UC is weaker than in CD

(Ardizzone, et al., 2006; Timmer, et al., 2007).

Tailoring or optimization can occur prior to or during

treatment. The appropriate maintenance dose of AZA is 2-

2.5 mg/kg/day and of MP is 0.75-1.5 mg/kg/day. The

„maximum‟ dose will differ between individuals and

effectively means that level at which leucopenia develops

(Gilissen, et al., 2005).


- 41 -

Adverse events occur in up to 20%. The commonest

are allergic reactions (fever, arthralgia, rash) that

characteristically occur after 2-3 weeks and cease

rapidly when the drug is withdrawn. Hepatotoxicity and

pancreatitis are uncommon (Gisbert and Gomollon, 2008).

Bone marrow toxicity has been reported to occur up

to 11 years after starting AZA and blood monitoring should

continue throughout thiopurine therapy. Manufacturers

recommend monitoring thiopurine therapy weekly by full

blood counts (FBCs) for the first 8 weeks of therapy

followed by blood tests at least every 3 months

(Colombel, et al., 2000).

Although a significant proportion of patients

experience adverse effects with thiopurines when the drug

is tolerated for 3 weeks, long-term benefit can be expected

(Macdonald, et al., 2009). In absolute terms, the risk

remains very small (<1% risk after 10 years of thiopurine

use) and the benefits of AZA outweigh any risks

(Lewis, et al., 2000).

In IBD, large population-based studies have shown

no increased risk. Whereas a second suggested a fourfold

increased risk of lymphoma in patients with IBD treated

with AZA/MP compared with background population

(Kandiel, et al., 2005).

There is an increased risk of non-melanoma skin

cancer in patients treated with thiopurines. Patients should

be advised to avoid excessive sun exposure and use a high-

strength sun block (Fraser, et al., 2002).


- 42 -

Methotrexate

Polyglutamated metabolites of methotrexate (MTX)

inhibit dihydrofolate reductase, but this cytotoxic effect

does not explain its anti-inflammatory effect. Inhibition of

cytokine and eicosanoid synthesis probably plays a role. It

is positioned as a second-line immunosuppressive agent in

patients resistant or intolerant of AZA, although it is

currently unclear whether thiopurines are any more

efficacious than MTX for induction or maintenance of

remission in IBD (Oren, et al., 1996).

A low dose (12.5 mg once weekly) was not shown to

be efficacious at inducing or maintaining remission. Using

larger weekly doses show higher response or remission

rates resistant or intolerant of AZA or MP. Parenteral

administration (either subcutaneous or intramuscular) may

be more effective that oral therapy and is recommended.

Monitoring therapy by measurement of FBC and liver

function tests are advisable before and within 4 weeks of

starting therapy, then monthly (Ei-Matary; Wahed, et al.,

2009; Nathan, et al., 2008).

Side effects are reported by 27- 49% of patients.

Early toxicity from MTX is primarily gastrointestinal

(nausea, vomiting, diarrhea and stomatitis). Co-prescription

of folic acid 5 mg (once a week, taken 3 days after MTX)

limits side effects. Long-term concerns are hepatotoxicity,

pneumonitis and opportunistic infections (Fraser, 2003).

MTX is teratogenic and should not be used if

conception considered. It may persist in tissues for long

periods; therefore conception should be avoided for 3-6

months after withdrawal. Breastfeeding isn't recommended

(Mahadevan and Kane, 2006)


- 43 -

Calcineurin inhibitors

Ciclosporin (CsA) is an inhibitor of calcineurin,

which prevents clonal expansion of T cell subsets

(Shibolet, et al., 2005). It is rapidly effective as a salvage

therapy for patients with refractory UC, who would

otherwise face colectomy, but its use is controversial

because of the narrow therapeutic index of it (including

mortality rates of 3–4%), toxicity and long-term failure

rate. The drug should rarely be continued for more than 3-6

months and its main role is a bridge to thiopurine therapy

(Lichtiger, et al., 1994).

Measurement of blood pressure, FBC, renal

function and CsA concentration (aim for 100-200 ng/ml)

are advisable at 0, 1 and 2 weeks, then monthly. Blood

cholesterol and Mg++

should be checked before starting due

to risk of seizures (McDonald, et al., 2005).

Minor side effects occur in 31-51%, including

tremor, paraesthesiae, malaise, headache, abnormal liver

function, gingival hyperplasia and hirsutism. Major

complications are reported in 17%, including renal

impairment, pneumonia, infections and neurotoxicity (Van

Assche, et al., 2003). Toxicity can be reduced by using

lower doses (2 mg/kg/day IV), by oral micro emulsion

CsA, or by mono-therapy without corticosteroids

(D’Haens, et al., 2001).

Tacrolimus is another calcineurin inhibitor often

preferred in the transplant setting to CsA. Data from trials

show that it is effective in treatment of steroid refractory

thiopurine naïve UC. A dose is of 0.025 mg/kg twice a day

should achieve trough levels of 10-15 ng/ml (Ogata;

Herrlinger, et al., 2006).


- 44 -

Anti-TNF therapies

There are presently two biological agents licensed

for the treatment of IBD in UK; both are monoclonal

antibodies against TNF α (anti-TNF) (Mowat, et al., 2011).

Infliximab (IFX) is a chimeric anti-TNF antibody,

consisting of 75% human IgG and 25% murine component

that actively binds membrane-bound and soluble TNFα.

IFX is given by IV infusion only (Mowat, et al., 2011).

Adalimumab (ADA) is a humanized anti-TNF

antibody, given by sub-cutaneous injection only. At the

present time both agents are licensed for the treatment of

Crohn's disease that has failed to respond to standard

immunosuppression (Mowat, et al., 2011).

Three intravenous infusions of IFX at 0, 2 and 6

weeks were effective in inducing clinical remission;

inducing endoscopic remission and clinical response at 8

weeks. The efficacy of infliximab for treating patients with

moderate to severe UC refractory to corticosteroids and

immunomodulators concluded that it was effective for

inducing clinical remission, clinical response, promoting

mucosal healing, and reducing the need for colectomy in

the short term (Lawson, et al., 2006).

Most recently, the anti-TNF antibody golimumab

has been shown to induce clinical remission and mucosal

healing. Treatment at weeks 0 and 2 (400/200 mg, 200/100

mg) significantly induced clinical remission and mucosal

healing at week 6 suggesting that several anti-TNF

antibodies favor mucosal healing in ulcerative colitis

(Sandborn, et al., 2012).


- 45 -

Treatment with anti-TNF therapy is relatively safe if

used for appropriate indications. It should be balanced with

the potential curative option of surgery in UC. Due to the

nature of their effects on TNF, all anti-TNF therapies share

a similar profile of adverse events, including increased risk

of infections from intracellular pathogens, most notably,

TB, other opportunistic infections, autoimmunity,

infusion reactions, and other more rare side-effects

(Mowat, et al., 2011).

Pre-treatment screening for exposure to TB is

important via a history, chest x-ray and tuberculin skin test

if applicable in patients who are about to begin anti-TNF

therapy (Theis and Rhodes, 2008).

The combination of IFX and a thiopurine analogue

or corticosteroids is probably justified to decrease

immunogenicity, which is the source of infusion reactions

and loss of response (Tekkis, et al., 2010).

There is insufficient evidence at present to

recommend the use of interferon γ release assays. Re-

activation of chronic hepatitis B has been reported in

patients treated with IFX. There are no data to suggest it

has any effect on course of chronic hepatitis C.

Pre-treatment screening for exposure to hepatitis B is

important; vaccination should be considered in the non-

immune high-risk patient (Esteve, et al., 2004).

Antibodies formation to infliximab (ATI) can

trigger both acute infusion reactions and delayed serum-

sickness-like reactions. Minor acute reactions usually

respond to slowing the infusion rate or treatment with

antihistamines, paracetamol and sometimes corticosteroids.

Episodic therapy and consequent „drug holiday‟ is


- 46 -

associated with increased formation of ATIs, and should be

avoided. ATI formation is associated with increased

incidence of infusion reactions and loss of response (Baert,

et al., 2003). Although ADA is a fully humanized antibody,

it is also associated with the formation of antibodies to

adalimumab (ATA) which have been shown to reduce

efficacy in rheumatoid arthritis and CD (West, et al., 2008).

Prolonged medical therapy for a potentially

pre-malignant condition with anti-TNF therapy creates its

own anxieties. The Mayo Clinic practice and Edinburgh

series confirmed the relatively rare occurrence of

malignancy including basal & squamous cell cancers

(Lees, et al., 2009).

Anti-TNF therapy was associated with an increased

risk of NHL when compared to the general population, but

the risk remained small (6.1 per 10000 patient-years). Anti-

TNF therapy also led to an increased rate of NHL

compared to those treated with immunosuppressants

alone, although this did not reach significance

(Kandiel, et al., 2005).

Reports of optic neuritis, seizure, and new onset or

exacerbation of central nervous system demyelinating

disorders, including multiple sclerosis, have been reported

with the use of all anti-TNFs. Also, Anti-TNF agents are

contraindicated for patients with class III-IV congestive

heart failure due to evidence of increased risks of death

from several clinical trials (Mowat, et al., 2011).


- 47 -

UC Management Guidelines

Active ulcerative colitis

When deciding the appropriate treatment strategy for

active ulcerative colitis one should consider the activity,

distribution and pattern of disease. The disease pattern

includes relapse frequency, course of disease, response

to previous medications, side-effect profile of

medication and extra-intestinal manifestations. The age

at onset and disease duration may also be important factors

(Silverberg, et al.,2005).

It is most important to distinguish patients with

severe ulcerative colitis necessitating hospital admission

from those with mild or moderately active disease

who can generally be managed as outpatients

(Schroeder, et al., 1987).

Patients should be encouraged to participate actively

in therapeutic decisions which should be tailored to the

individual (Munkholm, et al., 2010).

The choice of therapeutic strategy should be

influenced by the balance between drug potency and

side-effect profile; previous response to treatment

(especially when considering treatment of a relapse,

treatment of steroid-dependent or refractory disease, or

immunomodulator refractory disease); and the presence of

extra intestinal manifestations which may require

systemic therapy (Su, et al., 2007).


- 48 -

Treatment according to site of disease and disease activity

PROCTITIS

Choice of topical formulation should be determined

by the proximal extent of the inflammation (suppositories

for disease to the recto-sigmoid junction, foam or liquid

enemas for more proximal disease) along with patient

preference, such as ease of insertion or retention of enemas

(Safdi, et al., 1997).

A mesalazine 1 g suppository once daily is the

preferred initial treatment for mild or moderately active

proctitis. There is no dose response for topical therapy

above a dose of 1 g daily. Mesalazine foam enemas are an

alternative. Suppositories may deliver drug more

effectively to the rectum and are better tolerated than

enemas. Topical mesalazine is more effective than topical

steroids; consequently topical steroids should be reserved

as second line therapy for patients who are intolerant of

topical mesalazine (Dignass, et al., 2012).

Combining topical with oral mesalazine or topical

steroid is more effective than either alone and should be

considered for escalation of treatment. Oral mesalazine

alone is less effective. Patients who fail to improve on

oral/topical mesalazine and topical corticosteroids should

be treated with the addition of oral prednisolone.

Refractory proctitis may require treatment with

immunosuppressants or biologics (Dignass, et al., 2012).

Fig. (5): Extent of bowel

involvement in UC

(Johns Hopkins, 2013)


- 49 -

LEFT SIDED COLITIS

Left-sided colitis of mild-moderate severity should

initially be treated with an amino-salicylate enema 1

g/day combined with oral mesalazine >2 g/day. Foam

enemas are not inferior to liquid enemas for inducing

remission. Also, low volume enemas are not inferior to

high volume enemas and may be better tolerated. Topical

therapy with steroids or 5ASA alone as well as mono-

therapy with oral 5ASA is less effective than oral plus

topical 5ASA therapy. Topical mesalazine is more effective

than topical steroid (Dignass, et al., 2012).

Systemic corticosteroids are appropriate if a patient's

symptoms deteriorate, rectal bleeding persists beyond 10-

14 days, or sustained relief from all symptoms has not been

achieved after 40 days of appropriate mesalazine therapy


EXTENSIVE COLITIS

Extensive UC of mild-moderate severity should

initially be treated with oral 5ASA >2 g/day, which should

be combined with topical mesalazine to increase remission

rates if tolerated. Systemic corticosteroids are appropriate if

symptoms of active colitis do not respond to mesalazine by

oral prednisolone combined with steroid enemas. An

appropriate regimen for active disease is prednisolone 40

mg/day for 1 week, reducing by 5 mg/day/week resulting in

an 8 week course, and many different regimes are used



- 50 -

SEVERE UC OF ANY EXTENT

Acute severe UC is a potentially life-threatening

condition. In 1933, 75% patients died within the first year

after acute presentation with UC and in 1950 a mortality of

22% was reported amongst cases in the first year after

diagnosis. The response rate to appropriately dosed

intravenous steroids has not changed over the last 30 years

(Turner, et al., 2007).

Patients with bloody diarrhea ≥6/day and any signs

of systemic toxicity (tachycardia >90 bpm, fever >37.8 °C,

Hb <10.5 g/dL, or an ESR >30 mm/h) have severe colitis

and should be admitted to hospital for intensive treatment.

All patients admitted with severe colitis require appropriate

investigations to confirm the diagnosis and exclude enteric

infection (Van Assche, et al., 2011).

Corticosteroids are generally given intravenously

using methyl-prednisolone 60 mg/24 h or hydrocortisone

100 mg four times daily. Higher doses are no more

effective, but lower doses are less effective. Bolus injection

is as effective as continuous infusion. Treatment should be

given for a defined period as extending therapy

beyond 7 to 10 days carries no additional benefit

(Rosenberg, et al., 1990).

Consideration of mono-therapy with CsA or IFX if

there is no improvement by day 3, there is subsequent

deterioration or when steroids are best avoided, but for

patients already on immunosuppressive therapy, colectomy

is the first option. Other measures that may considered as

subcutaneous prophylactic low molecular weight heparin,

IV fluid, electrolyte replacement and blood transfusion



- 51 -

Table (7): Algorithm for treatment of active UC (Carter et al., 2004).

Refractory UC

Treatment decisions are depending on the pattern of

relapse, timing of relapse (disease remains active in spite of

treatment or relapse occurs when it tapered 'drug

dependent'), concurrent therapy, previous response to

therapy, adherence to maintenance therapy and patient

opinion (adverse effects, speed of response, convenience)

(Sandborn, et al., 2009).

Opinion is divided whether to use the same induction

treatment as before to achieve remission or to use more

potent therapy. Also, maintenance therapy should also be

optimized. Patients who have an early (<3 months) relapse

require further induction therapy, but should also

commence AZA or MP to reduce the risk of a subsequent

relapse. (Dignass, et al., 2012).


- 52 -

‘Steroid-dependent’, active ulcerative colitis

Thiopurines is the first choice of therapy for patients

who flare when steroids are withdrawn. Patients with active

disease despite steroid therapy require appropriate

induction therapy, including consideration of anti-TNF

therapy. Azathioprine is significantly more effective than

mesalazine at achieving clinical and endoscopic

remission in the treatment of steroid-dependent UC

(Ardizzone, et al., 2006).

Oral steroid-refractory ulcerative colitis

For active UC that is refractory to steroids, other

causes of persistent symptoms should be considered. If

active steroid-refractory UC is confirmed, alternative

therapy to induce steroid-free remission is required. Anti-

TNF therapy has clear evidence of benefit in this patient

group (Dignass, et al., 2012).

Adalimumab is now licensed for treatment of

severely active UC in Europe. Moderately active disease

outpatients have the surgical options or admission for IV

steroid therapy (Dignass, et al., 2012).

It should be noted that none of the treatments

discussed above have achieved steroid-free remission at

any time point in the majority of patients. The sequence (or

hierarchy) of therapy has to depend on the individual

circumstances and acceptability to the patient

(Ogata, et al., 2006).


- 53 -

Intravenous-steroid refractory UC of any extent

The response to intravenous steroids is best assessed

objectively around the third day. Treatment options

including colectomy should be discussed with patients with

severely active UC not responding to IV steroids. Second

line therapy with ciclosporin, infliximab or tacrolimus may

be appropriate. If there is no improvement within 4-7 days

of salvage therapy, colectomy is recommended. The timing

of colectomy for severe colitis remains one of the most

difficult decisions that a gastroenterologist has to make.

Delaying will increase the morbidity and mortality

associated with subsequent surgery (Dignass, et al., 2012;

Randall, et al., 2010).

Immunomodulator-refractory ulcerative colitis

Patients should be treated with anti TNF therapy or

tacrolimus although colectomy should also be considered.

Continued medical therapy that does not achieve a

clear clinical benefit is not recommended. It is best

reassessed to confirm diagnosis and exclude complications


Infliximab at either dose used (5 mg/kg or 10 mg/kg)

achieved clinical remission in a significantly greater

proportion of patients. Using infliximab (three IV infusions

at 0, 2, and 6 weeks) was effective in inducing clinical

remission in patients with moderate to severe UC refractory

to corticosteroids and/or immunomodulators (Lawson, et

al., 2006).Adalimumab also induced clinical remission with

standard induction dose of 160 mg, followed by 80 mg

after 2 weeks. Maintenance doses are then scheduled at 40

mg every other week (Gies, et al., 2010).


- 54 -

Refractory proctitis and distal colitis

They present common clinical dilemmas. It is clearly

important to consider and identify the etiology of the

refractory course. One obvious explanation is that the

disease is refractory to medication being prescribed.

However, alternative explanations include poor adherence

to prescribed therapy, delivery of an inadequate

concentration of drug to inflamed mucosa, unrecognized

complications (constipation or infection), and Inappropriate

diagnosis. Therefore, the initial step is to review current

symptoms and treatment to date, with a careful discussion

about adherence followed by reassessment of the diagnosis.

Next step is to ensure that conventional therapy has been

used appropriately. Attention in particular should be paid to

the formulation of topical therapy and whether it was used

in conjunction with an adequate dose of oral therapy.

Endoscopically documented patients have therapeutic

options include admission for IV steroid therapy.

Alternatively, there is open label evidence supporting the

use of salvage medical therapies (Dignass, et al., 2012;

Hebden, et al., 2000; Sandborn, et al., 1994).

An abdominal X-ray can be useful to diagnose

proximal constipation, since abnormal intestinal motility

induces proximal colonic stasis in patients with distal

colitis which may affect drug delivery. If there is visible

faecal loading, a laxative should be considered

(Järnerot, et al., 1985).

There is evidence from studies that appendicectomy

may improve outcome in patients with refractory proctitis.

The outcome of colectomy and pouch formation for distal

colitis is usually good (Brunel, et al., 1999).


- 55 -

Maintenance of remission

The goal of maintenance therapy in UC is to

maintain steroid-free remission, clinically and

endoscopically defined. More than half of patients with

UC have a relapse in the year following a flare. The

endpoint is the absence of relapse after 6 or 12 months

(Edwards and Truelove, 1963).

Maintenance treatment is recommended for all

patients. Intermittent therapy is acceptable in a few patients

with disease of limited extent. Choice of maintenance

treatment in UC is determined by disease extent, disease

course (frequency of flares), failure of previous

maintenance treatment, severity of the most recent flare,

treatment used for inducing remission during the most

recent flare, safety of maintenance treatment, and cancer

prevention. Options for a stepwise escalation of

maintenance therapy include dose escalation of oral/rectal

amino salicylates, the addition of azathioprine/

mercaptopurine or Infliximab/anti TNF therapy. Short term

use of systemic or topical steroids may be required when a

rapid response is needed (Dignass, et al., 2012).

Oral 5-ASA containing compounds are the first line

maintenance treatment in patients responding to 5-ASA or

steroids (oral or rectal). Rectal 5-ASA is the first line

in maintenance in proctitis and an alternative in

left-sided colitis. A combination of oral and rectal 5-ASA

can be used as a second line maintenance treatment


The minimum effective dose of oral 5-ASA is 1.2 g

per day. For rectal treatment 3 g/week in divided doses is

sufficient to maintain remission (Dignass, et al., 2012).


- 56 -

The dose can be tailored individually according to

efficacy. In some cases higher doses ± topical 5-ASA is

useful. There is no robust evidence to support the choice of

any specific preparation for maintenance. Although

sulfasalazine is equally or slightly more effective, other 5-

ASA preparations are preferred for toxicity reasons


E. Coli Nissle is an effective alternative to 5-ASA

for maintenance. No evidence has yet been reported that

any other probiotic is effective for maintaining of UC

remission. Also, insufficient data were regarded to

recommend antibiotics for maintenance of remission in UC


Azathioprine/mercaptopurine is recommended for

patients with mild to moderate disease activity who have

experienced early or frequent relapse whilst taking 5-ASA

at optimal dose or who are intolerant to 5-ASA, patients

that are steroid-dependent and for patients responding to

ciclosporin (or tacrolimus) for induction of remission


In patients responding to anti-TNF agents, both

maintaining remission with AZA/ MP and continuing anti-

TNF therapy with or without thiopurines are appropriate. In

patients with severe colitis responding to IV steroids, IV

CsA or IFX, AZA / MP should be considered to maintain

remission. However, in patients responding to INX

continuing it is also appropriate. The prior failure of

thiopurines favors maintenance with anti-TNF therapy. Due

to limited evidence, no recommendation can be given for

the duration of treatment with AZA or IFX, although

prolonged use of these medications may be considered if

needed (Dignass, et al., 2012).


- 57 -

Surgical options in UC

Up to 30% of patients will ultimately require

colectomy for ulcerative colitis. The decision to operate is

best taken by the gastroenterologist and colorectal surgeon

in conjunction with the patient (Hoie, et al., 2007).

Delay in appropriate surgery is associated with an

increased risk of surgical complication. A staged procedure

(colectomy first) is recommended in the acute case when

patients do not respond to medical therapy, or if a patient

has been taking 20 mg daily or more of prednisolone for

more than 6 weeks. If the appropriate laparoscopic skills

are available, a minimally invasive approach is feasible and

may convey some advantages (Dignass, et al., 2012).

The operation of choice in patients with acute severe

colitis failing to respond to intensive medical treatment is a

subtotal colectomy, end ileostomy and preservation of a

long rectal stump. Ileo-anal pouch is anther procedure that

involves panproctocolectomy with permanent end

ileostomy or ileoanal pull through procedure (IAPP)

(Brown, et al., 2008).

To aid decision-making many factors that predict the

need for colectomy in acute severe colitis can broadly be

divided into clinical markers such as stool frequency

(>12/day), pyrexia (>38◦C) and steroid-dependent disease

course, biochemical markers include a high CRP, low

albumin, and pH and radiological/endoscopic criteria

include the presence of colonic dilatation (>5.5 cm), the

depth of colonic ulceration, or mucosal islands on a plain

abdominal radiograph (Roussomoustakaki, et al., 1997).

Stem Cells and Stem Cell Therapy for UC Chapter 2

- 58 -

Stem Cells and Stem Cell therapy for UC

Discovery of Stem Cells:

. Stem cells are how we all begin. They are

undifferentiated cells that go on to develop into any of

more than 200 types of cell an adult human body holds

(Becker AJ, et al., 1963).

The stem cell is the origin of life, as stated first by

the great pathologist Rudolph Virchow, “All cells come

from cells”, and “All the cells of the human body arise

from a preexisting stem cell, the fertilized egg”

(Sell, 2004).

Fig. (6): Stem cells (Bloomberg News, 2012)


- 59 -

Introduction to Stem Cells:

The zygote is the ultimate stem cell produced from

fertilization. It is totipotent with the ability to produce all

the cell types of the species including the trophoblast and

the embryonic membranes. The zygote undergoes a process

of cell divisions and cell migrations known as cleavage. It

begins when several successive cell divisions resulting in a

doubling of the cell number and a reduction in the cell size.

At the 32 - 64 cell stage each cell is called a blastomere

(Sell, 2004).

The blastomeres stick together to form a tight ball of

cells called a morula. Each of these cells retains

totipotential. The next stage is the blastocyst which

consists of a hollow ball of cells; trophoblast cells along

the periphery develop into the embryonic membranes and

placenta while the inner cell mass develops into the fetus.

Beyond the blastocyst stage, development is characterized

by cell migration in addition to cell division

(Kuehnle and Goodell, 2002).

The gastrula is composed of three germ layers: the

ectoderm, mesoderm and endoderm. As development

proceeds, there is a loss of potential and a gain of

specialization, a process called determination. The cells of

the germ layers are more specialized than the fertilized egg

or the blastomere. The germ layer SCs give rise to

progenitor cells (precursor cells) which can further divide

to produce the terminally differentiated cell. The ectoderm

(outer layer) gives rise to the future nervous system and the

epidermis. The mesoderm (middle layer) gives rise to the

connective tissue, muscles, bones and blood, and the

endoderm (inner layer) forms the gastrointestinal tract and

the respiratory system (Kuehnle and Goodell, 2002).


- 60 -

Fig. (7): Development of Preimplantation Blastocyst in Humans(Terese Winslow, 2001).

Early in embryogenesis, some cells migrate to the

primitive gonad or genital ridge. These are the precursors to

the gonad of the organism and are called germinal cells.

These cells are not derived from any of the three germ

layers but appear to be set aside earlier. In adults, ovum and

sperm are derived from these cells by special cell division

called meiosis (Sell, 2004).

Fig. (8):Pluripotent stems cells (Mike Jones, 2006).


- 61 -

Definitions and Characteristics of SCs:

Stem cells (SCs) are undifferentiated cells that have

the capability of both Self-regeneration (self-renewal) and

Potency (differentiation) into mature specialized cells

through replication (Ricart E, et al., 2010).

Self-regeneration is the ability of stem cells to

divide and produce more stem cells. During early

development, the cell division is symmetrical i.e. each cell

divides to give rise to daughter cells each with the same

potential. Later in development, the cell divides

asymmetrically with one of the daughter cells produced is

also a stem cell and the other is a more differentiated cell

(Piscaglia, et al., 2008).

Potency is the ability to differentiate or the potential

to develop into different cell types. A totipotent stem cell

(e.g. fertilized egg) can develop into all cell types including

the embryonic membranes. A pluripotent stem cell can

develop into cells from all three germinal layers. A

multipotent stem cells could produce only cells of a

closely related family of cells (e.g. hematopoietic SCs).

Other cells can be oligopotent, bipotent or unipotent

depending on their ability to develop into few, two or one

other cell type(s) (Sell, 2004).

There are differences in how progenitor cell division

is described. For instance, according to one source (Robey,

2000), when a SC divides at least one of the daughter cells

it produces is also a SC; when a progenitor cell undergoes

cell division it produces two specialized cells. A different

source (Sell, 2004), explains that a progenitor cell

undergoes asymmetrical cell division, while a SCs

undergoes symmetrical cell division.


- 62 -

The apparent inconsistency of these two versions

illustrates the diversity and complexity of progenitor cells

and their role in differentiation. This diversity is reflected

in the nomenclature as well; progenitor cells are also called

Transit-amplifying cells, Precursor cells, Progenitors,

Lineage stem cells, and Tissue - determined stem cells


Table (8):Stem cells during differentiation at each stage (Sell, 2004).

Table (9): Types of cell division (Lindblad, 2004)

Early in development

Late in development: type 1

Late in development: type 2


- 63 -

Types of SCs:

A. Embryonic Stem Cells:

Embryonic stem cells (ESCs) are defined by

its origin. They are generally isolated from the

inner cell masses (ICMs) of blastocysts, which consist

of pluripotent cell populations that are able to

generate primitive ectoderm during embryogenesis.

More specifically the primitive ectoderm gives rise

during the gastrulating process to the primary germ

layers, including ectoderm, mesoderm, and endoderm

(Yao, et al., 2006).

They possess the dual ability to undergo unlimited

self-renewal and to differentiate in all fetal and

adult stem cells and their more differentiated

progenitors. They are pluripotent cells, capable of

forming tissues from all three germ layers in vitro and vivo

(Swenson and Theise, 2010).

Therefore, they represent a useful source of stem

cells for investigating the molecular events that are

involved in normal embryogenesis and generating a large

number of specific progenitors for cellular therapies. It

could use in studies of congenital birth defects,

chromosomal abnormality effects and childhood tumors

development (Trounson; Karp et al., 2006).


- 64 -

Advantages & Disadvantages of Embryonic SCs:

Advantages: They are

Flexible: They have the potential to make

anybody cell.

Immortal: One cell line can potentially supply

endless amounts of cells with carefully defined

characteristics.

Easily available: human embryos can be obtained

from fertility clinics in developed countries.

Disadvantages: They could be

Difficult to control: ESCs may be tumorigenic

when transplanted in vivo. The method for

inducing the cell type needed to treat a particular

disease must be defined and optimized

(Fujikawa, et al., 2005).

At odds with a patient's immune system: It is

possible that transplanted cells would differ in

their immune profile from that of the recipient

and so would be rejected. But this could be

avoided through genetic engineering them to

express MHC antigen of the recipient.


Ethically controversial: as life begins at

conception, doing research on human embryos is

unethical.


- 65 -

B. Adult Stem Cells:

The term adult stem cells (ASCs) refers to any cell

which is found in a developed organism that has two

properties: the ability to divide and create another cell like

itself and also divide and create a cell more differentiated

than itself. Also known as somatic (from Greek "of the

body") stem cells and germ line (giving rise to gametes)

stem cells, they can be found in children, as well as adults

(Doyonnas and Blau, 2004).

ASCs allow specific tissues to regenerate throughout

life. They also have the ability for self-renewal and multi-

lineage differentiation. In fact, the list for

identifying ASCs and lineage specific progenitor cells is

growing. A great deal of ASCs research has

focused on clarifying their capacity to divide or

self-renew indefinitely and their differentiation potential

(Preston, et al., 2003).

Distinct stem cell types have been established in

specific niches in many adult mammalian tissues and

organs, such as brain, skin, eyes, heart, kidneys,

lungs, gastrointestinal tract, pancreas, liver, breast,

ovaries, prostate, and testis(Griffiths, et al., 2005;

Liu, et al., 2004).

The replenishment of epithelial cell lineages within

the gastrointestinal tract (GIT) is a frequent process,

occurring every 2-7 days under physiological conditions.

This process may contribute to the generation of

new cell progenitors, which repopulate the damaged tissues

during diverse pathological disorders, such as inflammation

and ulceration (Schier and Wright, 2005).

http://en.wikipedia.org/wiki/Somatic


- 66 -

This process is regulated by multipotent SCs, which

give rise to all gastrointestinal epithelial cell lineages and

can regenerate whole intestinal crypts and gastric glands.

The SCs of the GIT are localized within the

niches in the intestinal crypts and gastric glands

(Brittan and Wright, 2002).

These SCs can give rise to all cell types within the

crypt, including absorptive, goblet, entero-endocrine and

Paneth cells. Similarly, the SCs in the large intestine or

colon, which are localized at the bottom of crypts, may also

give rise to the proliferative progenitors that differentiate

toward all lineages during epithelium regeneration

(Reya and Clever, 2005).

Fig. (9): Embryonic stem cells (Kuehnle and Goodell, 2002).


- 67 -

Advantages & Disadvantages of Adult SCs:

Advantages: They are

Already somewhat specialized: the inducement

may be simpler.

Immune hardy: Recipients who receive products

of their own SCs will not experience immune

rejection.

Flexible: it may be used to form other tissue

types.

Mixed degree of availability: Some ASCs are

easy to harvest and others, such as neural (brain)

stem cells, can be dangerous to the donor.

Disadvantages: They could be

Minimal quantity: difficulty in obtaining it in

large quantities

Finite: They don't live as long in a culture as

ESCs.

Genetically unsuitable: The harvested SCs may

carry genetic mutations for disease or become

defective during experimentation (Kuehnle and

Goodell, 2002).


- 68 -

Progenitor cells:

Stem cells generate an intermediate cell type or types

before they achieve their fully differentiated state. The

intermediate cell is called precursor or progenitor cell.

Progenitor cells in fetal or adult tissues are partially

differentiated cells that divide and give rise to differentiated

cells. Such cells are usually regarded as “committed” to

differentiate along a particular cellular pathway

(Robey, 2000).

The concept of a progenitor cell is difficult to define.

Like SCs, progenitor cells have a capacity to differentiate

into a specific type of cells. However, in contrast to SCs,

they are already far more specific than SCs: they are

pushed to differentiate into their “target” cell. Despite the

difficulty of defining progenitors, the term is frequently

used in researches (Noctor, et al., 2007).

Most progenitors are described as unipotant or

multipotant. In this point of view, they may be compared to

ASCs, on the other hand, they are said to be in a farther

stage of cell differentiation. They are in the “center”

between stem cells and fully differentiated cell. The kind of

potency they have depends on the type of their “parent”

stem cell and also on their niche (Noctor, et al., 2007).

Progenitors exhibit slow growth and their main role

is to replace cells lost by normal attrition. Growth factors

and cytokines trigger the progenitors towards the damage

tissue. At the same time, they differentiate to the target cell

to recover the injured tissue (Noctor, et al., 2007).


- 69 -

Plasticity of SCs:

Recently, several examples have been reported

which demonstrate that these stem cells, under certain

conditions, can be induced to form other cell types. This

property is called plasticity. For example neural stem cells

(NSCs) can give rise to blood and skeletal muscle. Also

BM cells can give rise to muscle, liver cells, and astrocytes

(Kuehnle and Goodell, 2002)

Therefore, they are considered multipotent, since

they can produce mature cell types of one or more lineages.

What determines SC potency largely depends on intrinsic

properties of SCs provided by the niche, microenvironment

where SCs reside, (Baharvand, et al., 2007).

Plasticity is the ability of stem cells to expand their

potential beyond the tissue from which they are derived.

For example, Dental pulp stem cells develop into tissue of

the teeth but can also develop into neural tissue (Nosrat, et

al., 2004). There are at least 2 alternative pathways to be

kept in mind when discussing mechanisms of SC plasticity:

i- Tran's differentiation is the direct conversion of one

cell type to another completely different one(Shen, et

al., 2003), e.g. Tran's differentiation of pancreatic

cells into hepatic cells and vice versa (Priller, 2004).

ii- Cell fusion: ESCs can fuse in vitro with a very small

proportion of BM stem cells, the fusion product, seen

as a new cell type, expressed genes of both fusion

partners in various degrees. Here the chromosomes of

the fused cells mosaic, presenting as a

mixture of chromosomes of different origin

(Doyonnas and Blau, 2004).

http://www.csa.com/discoveryguides/stemcell/gloss.php#trd

http://www.csa.com/discoveryguides/stemcell/gloss.php#cef


- 70 -

Bone Marrow:

Bone marrow (BM) contains hematopoietic stem

cells (HSCs) and stromal stem cells (SSCs) that collaborate

in a reciprocal manner at all stages, leading to the

generation of different BM and bloodstream cell lineages

(Murphy, et al., 2005).Bone marrow contains at least two

kinds of stem cells, hematopoietic stem cells and stem cells

for non-hematopoietic tissues (Friedenstein, et al., 1974;

Chopp, et al., 2000).

Recently, multipotent adult progenitor cells

(MAPCs), was identified which differentiated in vitro into

cells of all three germ layers and contributed to most

somatic tissues when injected into an early murine

blastocyst (Jiang, et al., 2002). A phenotypically identical

cell was isolated from human BM (Reyes, et al., 2001). It is

unclear, however, whether such MAPCs decline with donor

age, a phenomenon that has been observed for the

hematopoietic (Geiger and Van Zant, 2002) and

the mesenchymal stem cell compartment from BM

(Mendes, et al., 2002).

Fig. (10): Hematopoietic & stromal SCs differentiation (Terese Winslow, 2001).


- 71 -

Types of ASCs:

A. Hematopoietic Stem Cells:

The ability of HSCs to self-renew continuously in the

marrow and to differentiate into the full complement of cell

types found in blood qualifies them as the premier ASCs.

HSCs are among the few SCs to be isolated in adult

humans. They reside in the BM localized near endosteal

bone surface and sinusoidal endothelium and under some

conditions migrate to other tissues through the blood. HSCs

are also normally found in the fetal liver, spleen, umbilical

cord and placenta blood (Kiel, et al., 2005).

HSCs can migrate from endosteal surface into

vascular niche under specific stimuli such as injury and

after treatment with myeloablative agents. Hence, the

localization of HSCs and their progenitors within BM

vascular niche may allow their rapid release into

circulation. In addition, HSCs were also localized at the

sinusoidal endothelium on the spleen tissue sections,

it has been proposed that this site could also

constitute an extra medullary niche for HSC homing

(Wilson and Trumpp, 2006).

There is a growing body of evidence that HSCs are

plastic that, at least under some circumstances, they are

able to participate in the generation of tissues other than

those of the blood system. The expectations have risen that

HSCs will be able to give rise to multiple cell types from

all three germ layers. If HSCs are truly multipotant, their

potential for life-saving regenerative therapies may be

considerably expanded in the future (Murphy, et al., 2005)

.


- 72 -

B. Mesenchymal Stem Cell:

The BM stroma is a highly vascularized, complex

structure containing mesenchymal stem cells and

extracellular matrix elements supporting for hematopoiesis

(Neiva et al., 2005). They are variously referred to as

mesenchymal stem cells or marrow stromal cells. The term

"mesenchymal stem cells" has been used only relatively

recently. MSCs provide the stromal cells constituting the

microenvironment within the marrow cavities.

More recent data indicate broader potential for MSCs

(Jiang, et al., 2002).

Mesenchymal stem cells are an entirely different

type of adult stem cell. MSCs can be derived from bone

marrow, fat, or other tissues. MSCs are perhaps more

restricted in their differentiation potential to mesodermal

tissues such as fat, cartilage and bone. Transplanted MSCs

do not reconstitute hematopoietic lineages, but can alter the

host immune response. (Swenson and Theise, 2010).

Depending upon the environment, MSCs give rise to

many cell lineages including epithelial cells, astrocytes,

osteoblasts, chondrocytes, adipocytes and muscle,

promoting regeneration of damaged tissue in vivo

(Uccelli, et al., 2008).

In vitro, MSCs have vast proliferative potential, can

clonally regenerate and can give rise to differentiated

progeny. They also exhibit anti proliferative and anti-

inflammatory properties in vitro/vivo, making them

candidates for treatment of Immunomodulatory drugs

(IMiDs) (Dazzi, et al., 2007).


- 73 -

Among the BM stromal stem cells, there are

primitive mesodermal progenitor cells (MPCs) and

multipotent adult progenitor cells, which have been co-

purified with other mesenchymal stem cells, also as known

as bone marrow stromal cells (BMSCs). MPCs might

differentiate into different mesenchymal cell types

including bone-forming cells, osteoblasts, chondrocytes, &

adipocytes (Reyes, et al., 2001), as well as neuron-like cells

& glial cells, as in well-defined culture mediums in vitro

(Sanchez-Ramos, et al., 2000).

Furthermore, MAPCs and MSCs also show the

ability to migrate extensively throughout the circulation.

For instance, it has been reported that the transplantation of

MAPCs leads to their engraftment and differentiation into

the hematopoietic lineages and the epithelium of the liver,

lungs, and gut in non irradiated mice, which were further

enhanced in an irradiated host (Jiang, et al., 2002).

The injection of stromal BM cells was also

accompanied by their migration throughout the forebrain

and cerebellum, where they differentiated into astrocytes

(Kopen, et al., 1999).

Regardless of whether or not MSCs are true stem

cells, clinical benefit from MSCs may not require sustained

engraftment of large numbers of cells or differentiation into

specific tissues. It is possible that a therapeutic benefit can

be obtained by local paracrine production of growth factors

and by the provision of temporary antiproliferative and

immune-modulatory properties (Panes and Salas, 2009).


- 74 -

Role of Stem Cells in tissue regeneration:

If body tissue is damaged, stem cells head for the

damaged area and advance the process of healing.

However, day-to-day processes in the human body also rely

on stem cells: our erythrocytes only live for about 120 to

130 days, by which time they have become too old, cannot

transport enough oxygen and have to be replaced. This task

is taken over by the HSCs that can be found in the bone

marrow. According to theoretical calculations, about 350

million new erythrocytes are formed every minute. Most of

the other somatic cells are also replaced regularly: liver

cells after 10 to 15 days, white blood cells after 1 to 3 days

(Tuch, 2006).

During development, stem cells divide and produce

more specialized cells. Stem cells are also present in the

adult in far lesser numbers. Adult stem cells are observed in

continually-replenished cells such as blood cells and skin

cells, stem cells have recently been found in other issues

such as neural tissues (Hans Schöler, 2007).

Organ regeneration has long been believed to be

through organ-specific and tissue-specific stem cells.

Hematopoietic stem cells were believed to replenish blood

cells, stem cells of the gut to replace cells of the gut and so

on. Recently, using cell lineage tracking, stem cells from

one organ have been discovered that divide to form cells of

another organ. Hematopoietic stem cells can give rise to

liver, brain and kidney cells. This plasticity of adult stem

cells has been observed not only under experimental

conditions, but also in people who have received bone

marrow transplants (Hans Schöler, 2007).

http://www.csa.com/discoveryguides/stemcell/gloss.php?SID=0ecbnc5rna9k1ill21vm5g0076#pla


- 75 -

If the goal of medicine has been to treat symptoms of

diseases and possibly remove their causes, scientists and

physicians point now to a more ambitious target. The

primary objective of regenerative medicine is the complete

structural and functional recovery of the damaged organ

(Koh and Atala, 2004).

Regeneration coincides with tissue homeostasis and

involves the replacement of cells lost by normal

weal and tear and, ideally, following injury

(Ferguson and O'Kane, 2004).

Tissue regeneration is achieved by two mechanisms:

(1) Circulating stem cells divide and differentiate under

appropriate signaling by cytokines and growth factors, e.g.

blood cells; and

(2) Differentiated cells which are capable of division can

also self-repair, e.g. hepatocytes, endothelial cells, smooth

muscle cells, keratinocytes and fibroblasts. These fully

differentiated cells are limited to local repair. For more

extensive repair, stem cells are maintained in the quiescent

state, and can then be activated and mobilized to the

required site (Forbes, et al., 2002).

For wound healing in the skin, epidermal stem cells

and bone-marrow progenitor cells both contribute. Thus it

is likely that organ-specific progenitors and hematopoietic

stem cells are involved in repair, even for other organ repair

(Asahara and Isner, 2004).

http://www.csa.com/discoveryguides/stemcell/gloss.php?SID=0ecbnc5rna9k1ill21vm5g0076#cyt


- 76 -

Stem Cells Therapies:

There is great interest in the biology of ASCs

because of their capacity to self-renew and their high

plasticity. These traits enable ASCs to produce diverse

mature cell progenitors that actively participate in the

maintenance of homeostatic processes by replenishing the

cells that repopulate the tissues/organs during a lifespan

and regenerate damaged tissues during injury

(Leri, et al., 2005).

In general, embryonic, fetal and adult stem cells

show several common functional properties. Common

properties include their high self-renewal capacity and

potential to generate differentiated cell progenitors of

different lineages under simplified culture conditions in

vitro and after transplantation in host in vivo

(Trounson, 2006).

Therefore, the use of stem cells and their progenitors

is a promising strategy in cellular and genetic therapies for

multiple degenerative disorders, as well as adjuvant

immunotherapy for diverse aggressive cancer types

(Lindvall, et al., 2004).

Stem cell-based therapies hold great promise for

many diseases (Swenson and Theise, 2010).

The main clinical application of stem cells is as a

source of donor cells to be used to replace cells in

transplantation therapy. Parkinson and Alzheimer diseases,

muscular degenerative disorders, chronic liver and heart

failures, type 1 and 2 diabetes, arthritis, as well as skin, eye,

kidney, and hematopoietic disorders, could be treated by

the stem cell-based therapies (Hesdorffer, et al., 1998).


- 77 -

Induced pluripotent stem cells (iPSCs) can be

derived from adult somatic cells forced to reprogram to an

ES-like state in culture. iPSCs can be redirected toward

other phenotypes under specific culture conditions. iPSCs

are not derived from embryos, avoiding ethical concerns

regarding the use of human embryos. iPSCs can

also be made from any patient, allowing the production

of patient-specific and disease-specific cells for

research treatment. Undifferentiated ESCs and iPSCs can

both give rise to teratomas after transplantation, so

complete eradication of undifferentiated cells is a critical

step toward the clinical use of cell therapy when

they are used to produce differentiated cells for therapy


Adult stem cells exist in all types of tissues

throughout the body and function as a reservoir to replace

damaged or ageing cells (Ricart E, et al., 2010). They are

derived from mature organs and are much more limited in

differentiation potential compared with ESCs or iPSCs.

Also, ASCs might be better considered as

multipotent “progenitor” cells rather than stem cells


Due to the inability to harvest or expand SCs from

most adult organs efficiently and safely (e.g. liver, GIT,

heart), a majority of human stem cell trials have focused on

clinical applications for HSCs, MSCs, or both, which can

be easily obtained in clinically sufficient numbers from

peripheral blood, bone marrow, or umbilical cord blood and

placenta (Chamberlain, et al., 2007).


- 78 -

Stem cell transplantation in treatment of IBD

Applications of stem cells into autoimmune diseases

(AID) are currently being actively sought. Autoimmune

diseases result in the patient's own immune system

attacking its own tissues. The symptoms of the particular

autoimmune disease match the type of tissue that is under

attack. For example, autoimmune antibodies to a nerve

receptor can result in progressive paralysis

(Tyndall and Saccardi, 2002).

For stem cell therapeutics, one approach is to try to

suppress the patient's native immune system, which is

attacking the patient, and then replacing bone marrow with

other grafted stem cells. Human clinical trials using high

dose immunosuppression and chemotherapy, combined

with auto graft hematopoietic cell transplants have

been used in several autoimmune diseases including:

systemic sclerosis, rheumatoid arthritis, myasthenia gravis,

systemic lupus erythematosus and multiple sclerosis

(Burt, et al., 2003).

Intensive myelosuppressive or myeloablative

chemotherapy followed by transplantation of stem cells

derived directly from the bone marrow or from peripheral

blood after suitable conditioning, has revolutionized the

management of haematological malignancy and

haemoglobinopathies. Because these maneuvers cause

significant and prolonged alterations in the body's immune

system and function, SCT has been used for severe cases of

diseases that are believed to have an autoimmune basis

(Marmont and Van Bekkum, 1995).


- 79 -

Some diseases seem to have longer remissions while

others have more frequent recurrences. Furthermore, for

those patients in whom there is a relapse of the autoimmune

disease, may responded to agents that were ineffective

before the stem cell therapy (Tyndall and Saccardi, 2002).

Stem cell transplantation can either be allogeneic

(from a donor, usually HLA matched sibling)

(Soutar and King, 1995), or autologous (harvested from

the individual undergoing treatment with later reinfusion)

(Duncombe, 1997), also syngeneic transplantation

(from an identical twin) is possible (Snowden, et al., 1998).

HSCs have been proposed as a novel form of cell

therapy for damaged organs (Anversa and Nadal-Ginard,

2002). Hematopoietic stem cell transplantation (HSCT) has

been used to treat patients with other autoimmune diseases

that include loss of immune tolerance or a Th1-

predominant immune response (Lowenthal, et al., 1993).

Most patients with AID have a near-normal life

expectancy. Nevertheless, some patients suffer severe,

therapy-resistant progressive autoimmunity. HSCT is a

potential therapy for people with such severe refractory

diseases. HCT involves the administration of HSC, which

are capable of giving rise to all mature haematopoietic

cell types and some non haematopoietic cell types

(Sykes, et al., 2005).

IBD is an acute or chronic inflammation of the

mucosal layers of the intestine. It may be considered

an autoimmune disease, resulting from inappropriate

immune reactivity against one‟s own tissues in the GIT



- 80 -

The epithelial lining of the intestinal tract is

constantly being shed and renewed, using a stem cell

lineage system similar to the bone marrow's hematopoietic

stem cell system. The stem cells lie at the bottom crypts

of villi that line the intestinal wall and daughter

cells then differentiate as the migrate up the villi

(Marshman, et al., 2002).

The mainstay of clinical treatment is a combination

of anti-inflammatory agents and immunosuppressive

medications. Newer agents, including monoclonal

antibodies directed against TNF, are available for patients

who are unable to achieve remission with standard

immunosuppression. Despite treatment, the morbidity and

cost of IBD remain high. Corticosteroids are not effective

for maintaining remission of IBD and induce well-known

adverse side effects. New treatment approaches are needed


Recent advances reveal a primary role of

dysregulation of the immune system in the pathogenesis of

IBD. Sophisticated genome-wide association studies

implicate maladaptive signaling of a number of cytokines

and their receptors (Abraham and Cho, 2009).

Stem cell therapy for inflammatory bowel disease

(IBD) is still in its very early stages. It is known that

isolated hematopoietic stem cells in vitro can reconstitute,

among other things, epithelial cells of the GIT

(Theise, 2005).

The advantage with IBD is that it is an inflammatory

process, and inflamed tissue is one of the triggers that

appear to recruit circulating SCs. From a therapeutic

standpoint for IBD, this raises the possibility of treating


- 81 -

patient with SCs that are able to secrete an IBD alleviating

medication such as antibodies that can neutralize tumor

necrosis factor (TNF). The SCs would then be recruited

into the intestinal wall, locally secrete the TNF antibodies,

soothe the inflammation of the bowel and thus control the

symptoms of IBD (Brittan and Wright, 2002).

Recent studies on the use of HSCT for refractory

Crohn's disease and evaluations of patients who have

undergone HSCT for other reasons show remission of CD

following the transplantation. HSCTs were either

allogeneic or autologous. Allogeneic transplants may help

by overcoming CD's genetic predisposition to circulate

leukocytes, while autologous transplants may clear the

body of committed lymphocyte clones and restore the

lymphocytic imbalance. Most patients responded well and

some were in complete clinical remission 10 years after

transplantation (Hawkey, 2004).

BM transplant therapy for refractory IBD remains

experimental. This approach should be limited to patients

with severe disease who have exhausted standard

treatment options, and is best performed in the

context of an appropriately designed clinical trial


Results from phase I and phase II clinical trials

indicate that the approach is feasible and may result in a

remission with regard to tumor formation. Phase III trials

currently underway in Europe and the USA should help

clarify these issues (Swenson and Theise, 2010).


- 82 -

Stem cell transplantation Methods:

The recipient is prepared for the transplant by potent

immunosuppressive treatment, usually by chemotherapy

and/or total body irradiation. This may then be followed by

the transfer of autologous or allogeneic haematopoietic

cells to restore the host immune system (Burt, et al., 1998).

This procedure can cure autoimmune disease in

experimental animal models and is now being explored in

human clinical trials. So far, auto-HCT has been generally

preferred over allo-HCT because of the increased toxicity

and potential for rejection in allo-HCT. The risk of

GVHD in allo-HCT, which arises from the attack

of donor allogeneic T cells on recipient alloantigens, is

associated with significant morbidity and mortality

(Burt, et al., 1998).

It is an intensive procedure that most probably will

not replace current pharmacological treatment for less

severe, conservatively treatable AID (Burt, et al., 1998).

Fig.(11): Autologous HCT as a therapy for AID (Burt, et al., 1998).


- 83 -

The steps in autologous HCT (Sykes, et al., 2005):

a. Stem-cell collection: HSCs are either harvested from the

bone marrow under anesthesia or collected after

mobilization to the peripheral blood by treating the

patient with growth factor (for example, G-CSF).

b. HSC harvest, with or without ex vivo selection using

antibody specific for CD34 (a marker of HSCs): efforts

may or may not be made to deplete T and B cells from

the marrow or mobilized peripheral blood cells. This is

usually achieved by HSC positive selection on the basis

of CD34 expression. The HSC preparation is

cryopreserved.

c. In vivo lymphocyte depletion: various conditioning

regimens with the goal of minimizing the burden of auto

reactive B and T cells are given to patients. These

include combinations of chemotherapeutic agents, total-

body irradiation (TBI) and in vivo lymphocyte depletion

by antibodies, such as anti-thymocyte globulin (ATG),

and these treatments often deplete host HSC as well.

d. Stem-cell transplantation: after conditioning, the

cryopreserved HSC preparation is thawed and returned

to the recipient to reconstitute hematopoiesis


- 84 -

The steps in allogeneic HCT (Sykes, et al., 2005):

a. Mobilization regimen: these are similar to the methods

employed for autologous HCT, except that

chemotherapies are not used. Mobilization is achieved

using the growth factor G-CSF.

b. HSC harvest: This step is similar to that used for auto-

HCT.

c. In vivo lymphocyte depletion: this step depletes host T

and B cells to prevent rejection and treat the AID; it

may also be aimed at depleting donor T cells (to prevent

GVHD).

d. Stem - cell transplantation: as for auto-HCT.

e. Post - transplantation immunosuppression: as

prophylaxis against both rejection mediated by residual

host T cells and GVHD mediated by contaminating

donor T cells, allo-HCT usually requires additional post-

transplantation immunosuppression with agents such as

cyclosporine A.

Fig. (12):SCs transplant

using patient’s own cells

(Preeti Gokal Kochar, 2004)


- 85 -

Mechanisms by which HCT treat autoimmunity:

Several mechanisms for correction of autoimmunity

may apply to HCT (Burt, et al., 1998).

a. Immunomodulation by immunosuppressive

conditioning: potent immunosuppressive treatments

such as total body irradiation (TBI), cyclophosphamide

(CY), anti-CD2 antibodies (anti-CD2), anti-CD52

antibodies (anti-CD52), fludarabine & anti-thymocyte

globulin (ATG) can eliminate the majority of B and T

cells. In addition to alloreactive and auto reactive

memory B and T cells, others are depleted.

b. Immune-mediated destruction of auto reactive cells

(graft-versus-autoimmune host T and B cells). Donor T

cells recognize alloantigens and destroy residual

memory B and T cells. This mechanism is applicable to

allo-HCT only.

c. Deletion of alloreactive and auto reactive T cells in

thymus. If both donor and host cells contribute to

hematopoiesis and to the antigen-presenting cell (APC)

pool in the thymus, the new T-cell repertoire generated

in the recipient thymus is deleted of T cells recognizing

both allo- and autoantigens expressed by HSC of both

origins.

d. Induction of energy and deletion of auto reactive and

alloreactive T cells in the periphery. Costimulatory

blockade of the CD40−CD154 and CD28−CD80−CD86

pathways in concert with allogeneic BMT can overcome

the T-cell barrier to allo-HSC engraftment, which in

turn quickly produces donor-specific tolerance and

could tolerize cross-reactive auto reactive lymphocytes

as well. This mechanism plays a role only in allo-HCT.


- 86 -

e. Tolerization of peripheral auto reactive/alloreactive

T cells by regulatory T cells. CD4+CD25

+ regulatory

T cells have been implicated in the maintenance of

peripheral tolerance to organ-specific self-antigens as

well as alloantigens. The secretion of TGF- and IL-10

by regulatory T cells has been suggested to mediate this

process. Both allo-HCT and auto-HCT could

incorporate this mechanism.

f. Tolerization of auto reactive and alloreactive B cells. In

addition to alloantigens, haematopoietic chimaerism

could potentially tolerize pre-existing recipient B cells

to autoantigens expressed by donor haematopoietic

cells. This mechanism plays a role only in allo-HCT.

Fig.(13): Mechanism by which HCT ameliorate AID (Burt, et al., 1998).

Donor-derived cells are shown in green and host-derived cells are shown in red.


- 87 -

Potential risks of stem cell therapies

The risks of autologous or allogeneic hematopoietic

stem cell transplantation are well known, and include

hemorrhage, sepsis and GVHD. Allogeneic bone marrow

donors are thoroughly screened to prevent transmission of

infectious diseases, though acute infections may be

missed and laboratory errors are always possible


As well as the intrinsic dangers of SCT, which

include mucositis (a painful inflammation and ulceration of

the mucous membranes lining the digestive tract), there are

also some data suggesting that SCT may either worsen or at

least not prevent IBD in some patients. Some patients have

been reported to develop UC or colonic ulceration after

allogeneic SCT (Murayama, et al., 1995).

It would appear that the risk of infectious disease or

malignancy with autologous MSC therapy should be low.

As with allogeneic hematopoietic or whole-organ

transplantation, there is a small risk of transmission of

malignancy from donor to recipient. MSC themselves may

give rise to tumors, though evidence for this is limited


Reports suggest that MSC-derived tumor formation

may be a concern, and long-term MSC culture is best

avoided. Suppression of GVHD by allogeneic MSC may

also be coincident with reduction in the beneficial graft-vs.-

tumor effect, therefore it is essential to follow the long-term

incidence of hematologic disease recurrence in patients

treated with MSC (Swenson and Theise, 2010).


- 88 -

A clinical trial in stem cell therapies for UC

In 2010, a study included 44 patients with ulcerative

colitis (UC), which was implemented MSC transplantation

to determine the efficacy and safety of mesenchymal

stromal cells of bone marrow in the treatment of patients

with UC. 40 patients with UC who received standard

therapy and 12 patients treated by infliximab therapy.

The patients were observed for 24 months after

transplantation and evaluated using the average values of

indices of Rahmilevich clinical activity, indices of

endoscopic and histological activity scales Mayo and Gebs.

BM cells were obtained from the donor's sternum or iliac

crest and culture of it injected in the drip IV single dose.

The results show that in 34 patients after the

induction of MSCs was statistically significant compared

with the group of patients treated with drugs only 5-

aminosalicylic acid and corticosteroids, reducing the

clinical and morphological indices of inflammatory

activity. In 12 patients with UC include MSCs in the

treatment program did not have a therapeutic effect.

Application of MSC allowed canceling corticosteroids in

most patients with hormone-dependent and steroid

resistance forms of UC, and in 7 to reduce the dose of

prednisolone to 5 mg/day.

Finally researchers said that using of MSCs can be

evaluated as a new strategic direction for therapy UC.

MSC, have powerful immunomodulatory effects, reduce

the activity and stimulate regeneration of intestinal mucosa,

thereby increasing the duration of remission, reduces risk

of recurrence of disease (Lazebnik, et al., 2010).

Patients and Methods

- 89 -


This pilot study was a phase II randomized add-on

clinical trial that was done for the first time in EGYPT and

in Ain Shams University to investigate the role of bone

marrow stem cell transplantation as a potential line of

treatment in ulcerative colitis patients.

It was conducted on 10 patients with confirmed

diagnose of ulcerative colitis. Cases were collected from

Internal Medicine Departments and Out Patient Clinics

from Ain Shams, Nasser Institute and Electricity Hospitals.

This study hypothesized that infusion of HSCs may

help to reverse the inflammatory process in patients with

ulcerative colitis. Thus, we conducted a human trial to

evaluate safety and feasibility of autologous bone marrow

HSC transplantation in Egyptian patients with ulcerative

colitis and to evaluate the HSC transplantation as a

therapeutic option.

All patients assessed initially at January 2013 and

three months later at April 2013 after autologous

hematopoietic stem cells transplantation and follow up

includes comparison the changes in the patient's clinical

assessment, nutritional status, biochemical profile,

endoscopic findings, medication requirement, and quality

of life of those patients before and after potential therapy.

The patients of this study continued on the same

medications they were on previously but changes in their

use and dose were done according to the clinical

assessment throughout the follow up period.


- 90 -

All patients will be selected according to:

Inclusion criteria:

Adult Middle Eastern patients below 60 years old.

Documented diagnosed Ulcerative Colitis.

Written consent from every patient in our study.

Exclusion criteria:

Patients with Crohn's disease.

Patients with advanced systemic disease.

Patients with any malignancies or blood diseases.

Patients with other autoimmune diseases.

Refusal to sign a written consent.

All patients will be subjected to :

Clinical assessment :

Complete history taking.

Thorough medical examination.

Body mass index.

Investigations :

Laboratory markers (HB% - ALB - ESR - CRP).

Endoscopy : Lower GI Endoscopy (Colonoscopy).

Staging:(Mayo Scoring for Assessment of UC Activity)

Treatment modifications : Any changes in the type, form

or dose of different traditional pharmacological lines of

treatment according to case status.


- 91 -

Stem Cells Transplantation:

Stem cells transplantation was done in Nozha

International Hospital. All steps were performed within

complete sterile field in the following sequence:

A- Bone marrow aspiration:

1. The patient is placed in prone position on the

operating table.

2. Visual access is created to iliac crest by making a

puncture in the right iliac crest.

3. Penetrate bone marrow of ilium using standard

BM aspiration needle with needle length of

about 4 cm.

4. Upon penetration approximately 1 cm into

marrow cortex, the aspiration needle is advanced

forward gently through the marrow until the

needle contacts the cortical bone surface.

5. Spike with heparin solution (2000 µ/mL).

Ensure heparin solution has coated the entire

marrow contacting surfaces.

6. Procedure is complete after 100 ml of BM has

been aspirated from the penetration in the

right ilium.

B- Bone marrow transferring:

1. Remove Processing Disposables (PDs) from

packaging and place on an appropriate

workspace.

2. Slowly dispense 50 ml of bone marrow aspirate

(BMA) into the marrow chamber of each PD

through the RED access site.


- 92 -

C- Bone marrow processing:

1. Load centrifuge by placing the fully PDs into the

Smart PReP2 System (the Harvest instrument).

2. Close lid machine and press ʻSTARTʼ button to

start the process which takes 14 min. to be

finished. Then remove PDs from the Harvest

when cycle completed.

3. The plasma volume used for cell resuspension is

about 50 ml in chamber of each PD. a syringe is

used to withdraw unwanted plasma from the

chamber by being careful not to disturb the lower

cell layer.

4. To resuspend the BM cells into the plasma we

withdraw remaining plasma via a 50 ml syringe

with blunt needle, and gently inject it back and

forth into the plasma chamber. We repeat this 2-3

times till cells visibly suspend in plasma then we

withdraw total volume into the syringe.

5. Transfer full plasma volume which contains the

SCs (100ml) back in sterile field to a 350 ml

plastic bag (JMS Single Blood Bag).

D- Bone marrow injection:

1. A puncture of peripheral vein was performed by

using wide pore cannula.

2. IV infusion is performed through an infusion set

connects cannula with plastic bag to transfer

100ml of autologous BM mononuclear cell layer

contains SCs.

3. After that, the cannula was flushed with 10 ml of

normal saline and the procedure was finished.

After the stem cell infusion the catheter

was removed.


- 93 -

All patients take 500 ml saline 0.9% contains 1gm

cefoperazone (cefobid®) before starting the injection. They

were kept under medical observation for their vital data for

6 hours after that they discharged and reassessment

3 months later.

Bone marrow aspiration

Bone marrow transferring

Bone marrow processing

Bone marrow injection

Fig. (14): Stem Cells Transplantation for Patient No. 2

at Nozha International Hospital


- 94 -

Statistical Method

Statistical presentation and analysis of the present

study was conducted, using the mean, standard deviation,

Paired t-test, Chi- by SPSS V17.

Mean = n

x

Where: = sum & n = number of observations.

Standard Deviation [SD]:

SD 1

x-x 2

n

Standard Error [SE]:

n

SD SE

Paired t-test:

SE

dX

d

t

Where:

dX = Mean's difference between pre and post

SEd = Standard error of difference between pre and post.

Unpaired Student T-test was used to compare

between related samples.

Chi-square:

The hypothesis that the row and column variables are

independent, without indicating strength or direction of the

relationship. Pearson chi-square and likelihood-ratio chi-

square. Fisher's exact test and Yates' corrected chi-square

are computed for 2x2 tables.

Results

- 95 -

Results

This study was conducted on 10 patients with

confirmed diagnose of ulcerative colitis. The cases

chosen of those had a resistant active disease with

variable degrees of severity.

All cases were evaluated according to the

"Mayo Scoring System for Assessment of UC Activity"

which ranges from 0 to 12, with higher scores indicating

more severe disease (D'Haens, et al., 2007). This score

was used for both initial evaluation and 3 months post

therapy re-evaluation.

They are diagnosed and managed according to

European Crohn's and Colitis Organization (ECCO). They

were selected randomly of various extents and severity

assessed according to the Montreal classification and

Truelove & Witts criteria (Dignass, et al., 2012).

The patients included in this study are males (40%)

and females (60%). Their ages ranged from 24-50 years

with mean of (39.2 ± 10.2) and their BMI score ranged

from 20-33 with mean of (25.0 ± 4.7). All the patients

undergo to the autologous stem cell transplantation, and all

the patients tolerated the treatment protocol well without

any complications or side effects related to the procedure.

Results

- 96 -

Table (10): Showing comparison between patients

before and after the SCT as regard the presence of diarrhea

Diarrhea Negative Positive Total

Before N 0 10 10

% 0.00 100.00 100.00

After N 9 1 10

% 90.00 10.00 100.00

Chi-square

X2 4.9

P-value 0.026*

There were statistically significant differences as

regard the presence of diarrhea as 10 patients complained

of diarrhea associated with loose, mucus and bloody stool

in the initial clinical assessment but only one of them

continue complaining of this complaint in the final clinical

assessment after the procedure during the follow up period.

Fig. (15): Assessment of the presence of diarrhea

in patients pre and post the SCT procedure

Results

- 97 -

0

10

20

30

40

50

60

70

80

Before After

Negative

Positive

Rectal bleeding


before and after the SCT as regard the rectal bleeding

Rectal bleeding Negative Positive Total

Before N 4 6 10

% 40.00 60.00 100.00

After N 8 2 10

% 80.00 20.00 100.00

Chi-square X2 2.37

P-value 0.124

There were statistically none significant differences

as regard the presence of rectal bleeding as 6 patients

complained of rectal bleeding in the initial clinical

assessment, 4 of them become free of rectal bleeding in the

final clinical assessment after the procedure during the

follow up period.

Fig. (16): Assessment of the rectal bleeding


Results

- 98 -

0

10

20

30

40

50

60

70

80

Before After

Negative

Positive

Abdominal pain


before and after the SCT as regard the abdominal pain

Abdominal pain Negative Positive Total

Before N 2 8 10

% 20.00 80.00 100.00

After N 8 2 10

% 80.00 20.00 100.00

Chi-square

X2 1.011

P-value 0.315


as regard the presence of abdominal pain as8 patients

complained of abdominal pain in the initial clinical

assessment, 6 of them become free in the final clinical

assessment after the procedure during the follow up period.

Fig. (17): Assessment of the abdominal pain


Results

- 99 -


before and after the SCT as regard the disease extent

Disease extent E1 E2 E3 Total

Before N 0 4 6 10

% 0.00 40.00 60.00 100.00

After N 4 6 0 10

% 40.00 60.00 0.00 100.00

Chi-square

X2 0.277

P-value 0.599


as regard the disease site and extent as 4 patients having

left sided colitis and 6 have pancolitis in the initial

endoscopic assessment but by the end of the study

6patientshave left sided colitis and 4 have proctitis as

recorded in the final endoscopic assessment after the

procedure during the follow up period.

Fig. (18): Assessment of the disease extent


Results

- 100 -


before and after the SCT as regard the disease severity

Disease severity

After Total cases (before) Mild

Mild to moderate

Moderate

Be

fore

Moderate N 2 2 0 4

% 20.00 20.00 0.00 40.00

Moderate to sever

N 2 0 2 4

% 20.00 0.00 20.00 40.00

Sever N 0 2 0 2

% 0.00 20.00 0.00 20.00

Total cases (after)

N 4 4 2 10

% 40.00 40.00 20.00 100.0

Chi-square X2 10.008 P-

value 0.040*


regard the disease degree and severity as patients were:

4 moderate, 4 moderate to severe and 2 severe cases

in the initial assessment but by the end of the study

we have only 2 moderate case and 8 cases

evaluated as a mild or mild to moderate cases in

the final assessment after the procedure during the

follow up.

Fig. (19): Assessment of the disease severity


Results

- 101 -


before and after the SCT as regard the medical treatment

Medical treatment

After Total cases (before)

Rectal 5-ASA

Oral 5-ASA

5-ASA + Steroids

5-ASA + AZA

Be

fore

5-ASA + Steroids

N 1 2 2 0 5

% 10.00 20.00 20.00 0.00 50.00

5-ASA + AZA

N 2 2 0 1 5

% 20.00 20.00 0.00 10.00 50.00

Total cases (after)

N 3 4 2 1 10

% 30.00 40.00 20.00 10.00 100.0

Chi-square X2 3.333 P-

value 0.343


as regard the medical treatment modifications as patients

were divided in two groups, one group treated by 5-ASA

with steroids and other treated by 5-ASA with AZA in the

initial assessment but by the end of the study we have 7

patients treated with 5-ASA alone and 3 patient still on

steroids and AZA but with lower doses in final assessment

during the follow up.

Fig. (20): Assessment of the medical treatment


Results

- 102 -

Table (16): Showing comparison between patients before and after the SCT as regard the disease outcome

There were statistically significant differences as regard the disease degree and severity as patients were

divided in 5 cases of moderate, moderate to severe and severe pancolitis; and 5 cases of moderate and moderate

to severe left sided colitis in the initial global assessment but we end the study with having only 2 cases of

moderate left sided colitis case while other 8 cases evaluated between mild proctitis and left sided colitis in the

final global assessment during the follow up period.

Disease outcome After Total

cases (before)

Mild proctitis

Mild left sided colitis

Mild to moderate proctitis

Mild to moderate left sided colitis

Moderate left sided colitis

Be

fore

Moderate active left sided colitis

N 1 1 0 0 0 2

% 10.00 10.00 0.00 0.00 0.00 20.00

Moderate active pancolitis

N 0 0 1 1 0 2

% 0.00 0.00 10.00 10.00 0.00 20.00

Moderate to severe active left sided colitis

N 1 1 0 0 1 3

% 10.00 10.00 0.00 0.00 10.00 30.00

moderate to severe active pancolitis

N 0 0 1 0 0 1

% 0.00 0.00 10.00 0.00 0.00 10.00

Sever active pancolitis N 0 0 0 1 1 2

% 0 0 0 10.00 10.00 20.00

Total cases (after) N 2 2 2 2 2 10

% 20.00 20.00 20.00 20.00 20.00 100.0

Wilcoxon Signed Ranks Test

Z 17.27 P-

value 0.038*

Results

- 103 -

Fig. (21): Assessment of the disease outcome in patients pre and post SCT procedure

Results

- 104 -

Fig. (22): Endoscopic picture of a patient before and after SCT procedure.

Before After

Results

- 105 -


before and after the SCT as regard the motion frequency

Frequency Paired t-test

Range Mean ± SD t P-value

Before 5.000 - 10.000 8.000 ± 2.000

9.731 0.000*

After 3.000 - 5.000 3.600 ± 0.843


regard the motion frequency of the patients with ulcerative

colitis before and after the SCT with P values <0.05.

Fig. (23): Assessment of the motion frequency


Results

- 106 -


before and after the SCT as regard the body temperature

Temperature Paired t-test


Before 37.000 - 37.900 37.440 ± 0.381

1.354 0.209

After 36.900 - 37.500 37.220 ± 0.262

There were statistically non significant differences

as regard body temperature of the patients with ulcerative

colitis before and after the SCT with P values >0.05.

Fig. (24): Assessment of the body temperature


Results

- 107 -


before and after the SCT as regard the heart rate

Heart rate Paired t-test

Range Mean ± SD T P-value

Before 73.000 - 100.000 86.600 ± 10.679

3.321 0.009*

After 65.000 - 90.000 80.400 ± 8.809


regard the heart rate of the patients with ulcerative colitis

before and after the SCT with P values <0.05.

Fig. (25): Assessment of the heart rate


Results

- 108 -


before and after the SCT as regard the body mass index

BMI Paired t-test


Before 19.800 - 33.200 25.040 ± 4.739

0.178 0.863

After 20.500 - 32.300 24.980 ± 4.468


as regard the BMI of the patients with ulcerative colitis

before and after the SCT with P values >0.05.

Fig. (26): Assessment of the body mass index


Results

- 109 -


before and after the SCT as regard the serum albumin

ALB Paired t-test


Before 3.100 - 5.100 4.000 ± 0.772

-0.309 0.764

After 3.100 - 4.700 4.040 ± 0.659


as regard the serum albumin of the patients with ulcerative

colitis before and after the SCT with P values >0.05.

Fig. (27): Assessment of the serum albumin


Results

- 110 -

Table (22): Showing comparison between patients before

and after the SCT as regard the hemoglobin concentration

HB Paired t-test


Before 7.300 - 14.000 10.840 ± 2.688

-0.739 0.479

After 9.500 - 16.500 11.340 ± 2.744


as regard the HB % of the patients with ulcerative colitis

before and after the SCT with P values >0.05.

Fig. (28): Assessment of the hemoglobin concentration


Results

- 111 -

Table (23): Showing comparison between patients before

and after the SCT as regard erythrocyte sedimentation rate

ESR Paired t-test

Range Mean ± SD T P-value

Before 20.000 - 140.000 75.400 ± 49.332

3.628 0.006*

After 13.000 - 100.000 49.200 ± 32.913


regard the ESR of the patients with ulcerative colitis before

and after the SCT with P values <0.05.

Fig. (29): Assessment of the erythrocyte sedimentation rate


Results

- 112 -


before and after the SCT as regard the C-reactive protein

CRP Paired t-test


Before 16.000 - 24.000 20.667 ± 3.724

3.273 0.012*

After 11.000 - 19.000 14.500 ± 12.124


regard the CRP of the patients with ulcerative colitis before

and after the SCT with P values <0.05.

Fig. (30): Assessment of the C-reactive protein


Results

- 113 -


before and after the SCT as regard the activity score

Score Paired t-test


Before 7.000 - 9.000 7.800 ± 0.789

6.708 0.000*

After 5.000 - 7.000 5.800 ± 0.789


regard the activity score of the patients with ulcerative

colitis before and after the SCT with P values <0.05.

Fig. (31): Assessment of the activity score


Discussion

- 114 -

Discussion

„„Whatever deserves to exist deserves to be known‟‟

(Francis Bacon)

So much has been written and rewritten on stem

cells, their potentials, their properties, their possible uses

and their risks. Protagonists of a new scientific era, SCs are

promising tools for treating diseases of any sort, from

degenerative disorders to inborn errors of metabolism, to

organ failures. SCs represent the key to tissue genesis,

regeneration and homeostasis. This has led to the concept

of regenerative medicine, which is based on their potential

for therapies aimed to facilitate the repair of degenerating

or injured tissues (Piscaglia et al., 2008).

SCs can be obtained from various sources, including

embryos, fetal tissues, umbilical cord blood, and also

terminally differentiated organs. Once isolated, these cells

may be forced - ex vivo or in vivo - to expand and

differentiate into functional progenies suitable

for the cell replacement and the tissue engineering

(Mimeault et al., 2007).

Recent progress in cell biology resulting in the

isolation and characterization of bone marrow SCs and

progenitor cells further increases the expectation for a new

approach to the treatment of genetic and chronic diseases

(Sakaida et al.;2005).

Discussion

- 115 -

It is well known that ESCs may give rise to tumors,

while cancers derived from ASC-therapies haven‟t been

reported. Nonetheless, the long-term safety of ASC

infusion has not been adequately tested. Preclinical

studies and clinical trials with longer follow up

periods should be recommended prior to large-scale

clinical applications of such cell-based therapies

(Piscaglia, 2008).

The manipulation of adult stem cells (ASCs) seems to

be particularly promising, as it could improve the

endogenous regenerative potential without risk of

rejection and overcome the ethical and political

issues related to embryonic stem cell research

(Mimeault et al., 2007).

There are at least two types of SCs in the human

BM; mesenchymal SCs, and hematopoietic SCs.

HSC are continuously moving between the

bone marrow and peripheral blood. This movement is

critical for hematopoietic homeostasis & is hypothesized to

contribute to homeostasis and tissue repair

(Schachinger et al., 2006).

SC-based therapies could be used to cure

inherited or genetic degenerative alterations associated

with the loss of adult SC functions, such as cancers,

immune system and hematopoietic disorders,

cardiovascular, muscular and neurological diseases,

gastrointestinal pathologies, diabetes mellitus and

chronic hepatopathies (Mimeault et al., 2007).

Discussion

- 116 -

This pilot study was a phase II randomized add-on

cross-over clinical trial that was done in Ain Shams

University to investigate the role of bone marrow stem cell

transplantation as a potential line of treatment in ulcerative

colitis patients.

It was conducted on 10 patients with confirmed

diagnose of ulcerative colitis. Cases were collected from

Internal Medicine Departments and Out Patient Clinics

from Ain Shams, Nasser Institute and Electricity Hospitals.







therapeutic option compared to the conventional treatment.

All patients assessed before and 3 months after

autologous hematopoietic stem cells transplantation and

follow up includes comparison the changes in the patient's

clinical assessment, nutritional status, biochemical profile,



This study goes with agreement with Lazebnik, et

al., (2010), who initiated a pilot study to assess the safety

and efficacy of mesenchymal stromal cells of bone marrow

in the treatment of patients with UC. The patients were

observed for 24 months after transplantation and evaluated

using the average values of indices of Rahmilevich clinical

activity, indices of endoscopic and histological activity

scales Mayo and Gebs.

Discussion

- 117 -

In this study, there was improvement in the quality of

life in most of the patients after the procedure. Such

improvement was recorded according to the clinical,

laboratory and endoscopic assessment. This goes with the

trial of Lazebnik, et al., (2010), in which there was

improvement in the quality of life of 34 patients of the 44

patients included in their study at the end of follow up.

As regard clinical symptomatology, there was

significant improvement in most of the 10 patients

regarding the presence of diarrhea but in rectal bleeding,

abdominal pain and other associated symptoms includes

weight loss, anorexia, and urgency the improvement was

partially and statistically insignificant. Generally, most of

the patients regain their normal bowl habits with well-

formed stool and shifted form positive records at initial

clinical evaluation before the SCT to negative records at

the final clinical evaluation as they stopped complaining of

diarrhea, pain, bleeding, etc., after 3 months the SCT. This

goes with results of clinical trial of Lazebnik, et al., (2010).

There were statistically significant differences

regarding the motion frequency of the patients before SCT

with mean of (8.000 ± 2.000) and after SCT with mean of

(3.600 ± 0.843), with (P value <0.05) with improvement of

the frequency to the normal ranges. However, individual

non statistical analysis of each patient showed sustained

decline in most of patients over follow up period. Patients

No.1 and 3 decline from 9-10 to 3-4 times daily while

patients' No. 7, 8, 9 and 10 decline from 6-7 to 4-5 times

daily. This goes with results of Lazebnik, et al., (2010).

Discussion

- 118 -

This reflects that SCT has made improvement in the

quality of life to patients of UC but the mechanism for

the effect isn't clear but may reflect activation of

genes corresponding to an enterocyte differentiation

program in the transplanted SCs upon exposure to

injured colon environment.

Also, it found that heart rate was significantly

different before and after SCT while body temperature

was statistically insignificant. Vital data are considered as

supporting factors in assessment especially because it is

recorded in the former evaluation scoring systems of UC

but we didn't entitle them alone as they might be affected

by other several factors.

The body mass index (BMI) and serum albumin

which is an important laboratory marker that reflect the

nutritional status of the patients with ulcerative colitis

showed statistically non-significant differences in their

levels before and after the SCT in patients over the follow

up period. Analyzing the results of each patient without

statistical analysis showed limited increase in the level of

the s. albumin by about 0.5gm/dl in patients' No. 1 and 6

while slight decrease in patient No. 2 detected. Other

patient s. albumin didn't remarkably change. On the other

hand, BMI wasn't changed though out the study in the all of

the patient except patient No. 9 who gain about two

kilograms over his weight that recorded in his final

assessment which may be due to improved anorexia or

from the effect of the corticosteroids therapy. Results

regarding the BMI and s. albumin weren't evaluated in

study of Lazebnik, et al., (2010).

Discussion

- 119 -

Regarding the hemoglobin concentration level of

patients, there were statistically non-significant differences

before and after the SCT. Patient No. 3 showed elevation in

her HB% levels from 7.3g/dL before procedure to 9.5g/dL

after it in later follow up visits without blood transfusion.

This may be related to disease activity improvement. This

goes with results of clinical trial of Lazebnik, et al., (2010).

As regard the C-reactive protein (CRP), there was

statistically significant improvement in its level in patients

with Mean of (20.667 ± 3.724) before the SCT and a Mean

of (14.50 ± 12.124) 3 month after SCT. This also goes with

the results of the study of Lazebnik, et al., (2010).When

analyzing the results of each patient alone from the Master

Sheet; we found that the decreased was remarkable in some

patients as in patients' No. 1, 3, 4, 5, 6, 7 and 9. Other

patients showed not more than slight decrease as in patient

No. 2 showed no decrease at all. This difference between

the CRP results couldn't be confirmed due to small number

of patients. We need to have larger groups of patients to be

reevaluated upon with long-term follow-up.

Erythrocyte sedimentation rate (ESR) showed

statistically significant decrease in its results in patients 3

months after the SCT as differences regarding the ESR of

the patients before SCT with mean of (75.400 ± 49.332)

and 3 months after SCT with mean of (49.200 ± 32.913),

with (P value <0.05). When analyzing the results of each

patient alone from the Master Sheet, we found that the

decreased was in parallel with that in the CRP values as

mentioned and goes with results of Lazebnik, et al., (2010).

Discussion

- 120 -

Although the medical treatment changes

were statistically non-significant, it was considerable in

the individual non statistical analysis which showed

changes over the follow up period in the prescription, form

and dose of the used drugs for treatment of the patients in

our study. This also goes with the results of the study of

Lazebnik, et al., (2010).

There were significant differences regarding the

medications used in all the patients before and after the

SCT, as seven of them stopped using of steroids and

immune suppurations through gradual withdrawal of them

with sustained clinical and endoscopic response after SCT

throughout the follow up period.

On the other hand, two patients still on steroids that

planned to be stopped but in much slower rate than the

planned period of our study. Only one case remained using

azathioprine (AZA) after 3 months after SCT but in

lower dose as it decreased to100mg instead

of 150mg before it.

One of the targets in this study was to assess the

effect of SCT on the disease extent and severity. The result

analysis showed that SCT had significant effect on both.

Colonoscopy showed significant differences in distribution

and degree of inflammation after 3 months after the SCT.

This may be due to the complex pathophysiological

mechanisms of SCT that involved in the treatment of

ulcerative colitis. This also goes with the results of the

study of Lazebnik, et al., (2010).

Discussion

- 121 -

When analyzing the results of each patient alone

from the Master Sheet, it found that the improvement was

significant in most of the patients as they were having

moderate or moderate to severe, left sided or extensive

colitis in the initial assessment before SCT and become

mild or mild to moderate, proctitis or left sided colitis

after 3 months after SCT.

As regard the activity score index, there is

statistically significant difference between the score before

the SCT with mean of (7.800 ± 0.789) and 3 month after

the SCT with mean of (5.800 ± 0.789).

Patients No. 2, 4, 6 and 7 showed sustained decrease

in the activity score index from 8 to 6 over the whole

follow up period. The same occurred with

Patient No. 3 and 5 whose score decreased from 9 to 6 for

more than 3 months. Rest of patients showed slight

decrease in their score after SCT. Lazebnik, et al., (2010),

used different scoring indices to evaluated patients.

Disease outcome was promising in this study as

patients pass in a degree of inactivity but the correlation

between the procedure and the patients' improvements still

completely unclear. It could be analyzed as stems cells

effect by their dual action through induction mucosa

regeneration and immune system remodeling. In the study

of Lazebnik, et al., (2010), patients also showed

improvement at end of follow up period.

SCT may be able to improve the general condition of

patients with ulcerative colitis disease, at least temporarily;

until a curative treatment is available or other SCT is done.

Conclusions

- 122 -

Conclusions

Hematopoietic Stem Cell transplantation to patients

of ulcerative colitis disease is a safe and feasible procedure

without recorded complications like bleeding, thrombosis

or any system failure.

It can improve the quality of life of the patients as

well as the clinical illness, laboratory markers,

inflammatory process extent, activity degrees, and multiple

usage of medical treatment.

This improvement may be transient needs for longer

period of follow up to assess if SCT can consider as an

induction therapy of remissions or acts only as adjuvant

one helps to deliver the patient to inactivity much easier.

Recommendations

- 123 -

Recommendations

One of the limitations of our work was the fact that we

didn't track the infused HSCs in the patients‟ bodies. It

is very important to understand the way SCs act to

improve UC disease.

Evaluating the effect of SCT using different clinical trial

protocol as randomized controlled clinical trial study as

it has the strongest evidence of the clinical efficacy of

preventive and therapeutic procedures in the

clinical setting.

Clarify its effect on the other medications and

vice versa. Also, put it in comparison with

anti-TNF therapies.

Compare the use of Hematopoietic and Mesenchymal

stem cell therapy as regard the efficacy and side effects.

Larger number of patients and longer period of follow

up are needed to assess long lasting effects

and complications.

Evaluating the SCT as a maintenance therapy and

compare it with other available maintenance therapeutic

options as regard efficacy, cost and side effects.

Try different routes for SCT as local injection of stem

cells and whether it is better than intravenous injection.

Evaluating the SCT in patients of IBD due to other

causes rather than ulcerative colitis to see if there are

better results with other diseases or not.

Summary

- 124 -

Summary

Ulcerative colitis is an inflammatory bowel disease

leading to inflammation of the colon and causing painful

ulcers which can bleed, cause mucous production and

infection. Common symptoms include bloody diarrhea,

abdominal pain and weight loss which may be mild to

severe and affect individual's quality of life.

Currently, therapy is most often implemented in

a stepwise fashion, progressing through amino salicylates,

corticosteroids, immunosuppressives, and anti-TNF drugs.

Primary and secondary failure to respond to

approved therapies represents unmet needs in the

treatment of IBD.

A novel and exciting approach could be offered

through the current development in the field of

stem cell biology. Two streams of research, experimental

and clinical, are the origin of the increasing utilization of

stem cell therapies for severe immune-mediated diseases

(IMIDs) including IBD.

Other medical researches reflect that SCT has made

improvement in the quality of life to patients of UC but the

mechanism for the effect isn't clear but it interpreted

through the powerful immunomodulatory effects of

SCs and its ability to stimulate regeneration of

intestinal mucosa.

Summary

- 125 -

The present pilot study is a clinical trial that was

done in Ain Shams University to investigate the role of

bone marrow stem cell transplantation as a potential line of

treatment in ulcerative colitis patients. It was conducted on

10 patients diagnosed and managed according to European

Crohn's and Colitis Organization. They were selected

randomly of various extents and severity, and they

were assessed according to The Montreal classification,

Truelove & Witts severity scoring system and

Mayo activity score index.







therapeutic option compared to the conventional treatment.

All patients assessed before and 3 months after

autologous hematopoietic stem cells transplantation and

follow up includes comparison the changes in the patient's

clinical assessment, nutritional status, biochemical profile,



The study results showed that there was

improvement in the severity of the disease and the quality

of life in most of the patients over the 3 months after the

procedure. Also, results showed that the procedure was

well tolerated with no observed procedure-related

complications.

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Arabic summary

- 154 -

:هقدهة الرسالة

ؤد أحذ أيشاع انزبثبد انمن انضيخ انزبة انمن انزمشح

أ رزظ إفشاصاد أ رضف رمشحبد يؤنخ ك ظبحجإن انزبة انمن

أػشاع يب غؼم شبؽ خل يزفبرخ يخبؽخ ، ش انشع ثفزشاد

سثب ساد ،زض زا انشع أنؼذح أشش رخزفانشع رزكشس أ

ثبإلػبفخ إن آالو ف انجط ، فمذا ثبنص ، يرشم اإلسبل انذثأػشاع

أػشاع أخش.

حز ا، لذ أظشد انؼذذ ي كهب يسججبد زا انشع غش يؼشف

هؼذ ك ن ف ظس ، سثب انجئخ هؼايم انساصخنا انذساسبد رأصش

انجكزشخ دس ف رنك أؼب ، إال أ أغهت انظشبد رشعح أ يشع يبػ

يسزشف يب ث لبيذ ثحش لذ شش ثبنذسعخ األن ، يشع بدس سجب ،

.سب زا انشع ث شخض ي كم أنف شخض 44 - 2ي إطبثخػح كهك

انسشش، ؼزذ رشخض زا انشع ػه ػذح ػايم رشم انفحض

انزحبنم انخجشخ ، إػبفخ إن انظبس انزشخظ ػ ؽشك انمن ، انز

زح أخز ػبد نفحظب يكشسكثب رأكذ انزشخض ،إػبفخ إن رنك زى

ثشايظ يزؼذدح يؼزذح دنب ،زلف رمى انحبالد رظفب رجؼب ألسبنت

يكب حذح انشع ، ف انؼزبد انز رج ػه زبئظ انزمى نشػا ػالط

، ػمبساد انسبنفب يضم ثبنؼمبساد انزمهذخ يؼظى األؽجبء ػالط يشػبى جذأ

بػخ فؼبنزب انيضجطبد ػمبساد صجذلذ أ انكسرضبد انؼبداد انحخ ،

طبق د ي اسزخذايب ػه لذ حذ زب س أؼب ، إال ا انشع زا ف ػالط

انز اصجزذ انؼمبساد انجنعخ األعسبو انؼبدح ظشد حذضب كباسغ ،

.نك اسرفبع ركبنف انؼالط ثب حذ ي ازشبسب كزنك ، أؼبػبنخ خ فؼبن

حممب زبة انمن انزمشح يب ضال شكم رحذب نػالط ا ف انالغ إ

نهؼالط رك أكضش فبػهخ سهخ عذذحزمذى ، يب غؼهى ف ثحش دائى ن نألؽجبء

االثحبس انطجخ زظش انزطم نؼالط عذذ ألم س خ ، يغ رمذو انؼهى رطس

بعح نزا انشع.

ذػ نالزبو ، انغزػخ س انحبطم ف يغبل ثنعب انخالبانزط إ

اػذح نهكضش ي ثذأ ؼط إشبساد كؼالط خكب أ ارسبع اسزخذاو ز انزم

انؼه نؼالط شبف نب انجحش ، انز نى زطم االيشاع انضيخ انسزؼظخ

انكجشح انحطخ ثغبل انؼالط ثبنخالب انغزػخ رسزذ انؼغخسثب حز ا.

ػه لذسرب انزارخ ػه انزغذد انضح انفشذح ثنعب ، ػه خظبئظب

ذساسبد انيعخ ي لذ ازششد، األػؼبء انخزهفخػه رغذذ األسغخ

يخزهف مم خبع انؼظى إن إيكبخ حلششد االثحبس انؼهخ ، انز

نؼالط األيشاع. األػؼبء

-أ-

Arabic summary

- 155 -

انخالب انغزػخ ف يغبل ثنعب ظشد ثادس انزطس انحذش انزم

با خؼؼ كانز يز ألم ي ػمذ ي انضي، حش نحع أ ثؼغ انشػ

انز كب ؼب أؼب ػخ ثسجت أيشاع سشؽب انذيزنهؼالط ثبنخالب انغ

الحظا رحسب ف حبنزى اخزفبء ي يشع انزبة انمن انزمشح انضي

نألػشاع انظبحجخ نزا انشع ، يب عؼم انجبحض فكش ف اسزخذاو ز

يفظم.انزمخ ف ػالط زا انشع ثشكم

انغزػخ بنخال أبلخ ػه لذ اصجزذ انؼذذ ي االثحبس انؼبنخ ثأدنخ

ػضنب ك عدح ف انؼذذ ي أعضح عسى انجبنغانيزؼذدح انم

بض زحش راالشخبص انجبنغ، يؼبنغزب خبسعب صسػب يشح أخش ف

، لذ رى رفز ز انطشمخ نؼالط انخزهفخ أسغخ األعضح نزؼم ثشكم يبست ف

انز ح ثظذد -ػذح أيشاع، يب يشع انزبة انمن انزمشح انضي

اظشد انزبئظ دالالد يجششح. لذ –انجحش ف ػالع

طرق وأسالب البحث:

كانزحم ذفب شس ػ عبيؼخف رزى انز انزغشجخ انذساسخ ز

انذيخ انخالب انغزػخ نخبع انؼظى داخم األػخحم انكشف ػ دس

انمن انزبة ي ؼب انز انشػ ف يحزم ػالط كخؾ انفشػخ

رأصش ػه ي خالل زا اإلعشاء ف انؼالط رمى يذ فبػهخ انضي انزمشح

انزحبنم ، انزغزخ انحبنخ ، انسشش انزمى رحس انحبالد ي بحخ

سفغ ، انسزخذيخ األدخ ع عشػخ ، نظبسا زبئظ ، انؼهخ

عدح حبح انشػ.

ؼب ي انزبة انمن انزمشحيشػ 41ز انذساسخ ػه أعشذ

رى رشخظى ػالعى فمب نهزطبد انشسح ي يظخ )إك( األسثخ ،

انحبالد ؼذع رمى حبالرى فمب نألظخ انؼبش انؼزذح دنب ،لذ

يؼذ ، شس ػ يسزشفبد ف انؼبداد انخبسعخ انجبؽخ وبلسأ ي ػشائب

. انكشثبء، بطش

:ى وفقا لوا ل تن اختار جوع الورض

اإلدراج هعار:

.سخ 61شخض ششق أسط ثبنغ ألم ي .4

.يشغ انزبة انمن انزمشح ثزشخض يصك .2

.يافمخ خطخ ي انشغ ػه انزطع ف دساسزب .3

-ب-

Arabic summary

- 156 -

هعار االستبعاد:

كشض.ع انشػ انز ؼب ي يش .4

.خيزمذي انشػ انز ؼب ي أيشاع أخش يضيخ .2

.انشػ انز ؼب ي األساو انخجضخ أ أيشاع انذو .3

.أخش انشػ انز ؼب ي أيشاع يبػخ رارخ .4

.افمخ انخطخ نهزطع ثبنذساسخانسفغ انزلغ ػه .5

هزمى ن أخش عغ انحبالد ف دساسزب نزمى أن كبيم إػبدح ذخؼؼ

أشش ي حم انخالب انغزػخ ي خالل يؤشش شبؽ انزبة ثؼذ يشس صالصخ

.ػه حبنخ انشػ انمن انزمشح نزسغم رأصش صسع انخالب انغزػخ

:خضع جوع الورضى إلى ها ل

:التقن السرري .1

شبيمانأخز انزبسخ انشػ انكبيم انفحض انطج

الفحوصات: .2

(- %ESR – CRP –ALB -HBرحبنم يخجش )

:التنظر .3

فحض االسغخ ػه انمن ( انزظش انسفه)يظبس

لتقن نشاط الورض( "Mayoالتدرج )نظام " .4

سزى اسزخذاو زا انظبو ف كم ي انزمى األن انزكه

.42إن 1نشطذ االسزغبثخ نهؼالط حسبة دسعخ انشع ي

:تعدالت العالج .5

ؽشمخ أ عشػخ انؼــــــــالط فأ رغشاد ف ع أ

.فمب نحبنخ انشغ يخزهف خطؾ انؼالط انذائ انزمهذخ

بنخالب انغزػخ انزارخ ث رهمذ انؼالطانحبالد اناسدح ف دساسزب عغ

ثؼذ انذيخ ، رنك انغش يزبضح نخبع انؼظى ػ ؽشك انحم داخم األػخ

يؼمخ ثبنكبيم ، رمبد خالل عبص خبصيؼبنغزب ثبنطشد انشكض ثاسطخ

ف أسثغ خطاد سئسخ : انغزػخ نهشػ انخالب صسع ػهخلذ رذ

عهسخ احذح ف ذ عؼبر ، االسزخالص ، انمم ، انؼبنغخ، انحم

.انذن انضخ سزشفث

-ج-

Arabic summary

- 157 -

االستنتاج :

انزبة شػن انغزػخ انخالب صسعي خالل زبئظ ب انجحش عذب أ

خطشح يؼبػفبد ذث يك ، آي إعشاء انضي انزمشح انمن

.يسغهخ

عدح سفغ أ رؤد إن ككب عذب أ اسزخذاو ز انطشمخ نهؼالط

انزبة حذح ،انؼهخ ؼاليبدان ، نهشع ، األػشاع انؼبيخ انشػ حبح

إػبفخ إن خفغ اسزخذاو انؼمبساد انذائخ ، انشع شبؽ دسعخ ، انشع

انزمهذخ.

زبثؼخان ي أؽل فزشحن لز أ ػبثش نزنك حزبط انزحس زا ك لذ

ػالط اػزجبس حمب كإرا كب انؼالط ثبنخالب انغزػخ يب نزم نهشػ

يب إن انشغ إظبل ػه أ فمؾ لذ سزخذو كؼالط يسبػذ يفشد نهشع ، أو

.سمثظسح أ انشع خلس ثفزشح

التوصات:

شعان زا رحس ػه انغزػخ انخالب كفخ ػم نفى ؼكثانجحش.

يخزهفخ ثشركالد ثحش ػه ثبسزخذاو انغزػخرأصش انؼالط ثبنخالب رمى.

انؼكس. األخش األدخ انؼالط ػهرأصش زا رػح

ف ػالط زا انشع. انغزػخ ثبنخالب اسزخذاو األاع انخزهفخ ي يمبسخ

نهزمى ثظسح أػح.ل أؽ فزشح انشػ ي أكجش ػذد إن حبعخان

كؼالط رحفظ ؽم انفؼل. انغزػخرمى انؼالط ثبنخالب

انغزخ ف ػالط انشع.اسزخذاو ؽشق أخش نحم انخالب

نألاع األخش ي ايشاع انزبة انمن. كؼالط رمى انخالب انغزػخ

……..

-د-

Role of Stem Cell

Transplantation in the

Treatment of Ulcerative Colitis

Thesis

Submitted for the Partial Fulfillment

of Master Degree in Internal Medicine

By

Mohammed Fathy Sayed Mohammed Zaky

M.B.B.Ch.

Faculty of Medicine - Ain Shams University

Supervised by

Prof. Dr . Mohsen Mostafa Maher

Professor of Internal Medicine


Ass. Prof. Dr . Wesam Ahmed Ibrahim

Assistant Professor of Internal Medicine


Dr . Shereen Abou Bakr Saleh

Lecturer of Internal Medicine



2014

بسم هللا الرحمن الرحيم

(111سورة طه )

صدق هللا العظيم

Acknowledgement

At first and foremost thanks to “Allah” the most

merciful who gave me the power to finish this work.

I wish to express my deepest appreciation and

sincere gratitude to Prof. Dr. Mohsen Mostafa Maher,

Professor of Internal Medicine, Ain Shams University,

for his precious help throughout all stages of this work,

outstanding kind support and valuable instructions.

My greatest respect, appreciation and thanks to

Prof. Dr. Wesam Ahmed Ibrahim, Assistant Professor

of Internal Medicine, Ain Shams University, for her

valuables help, planning and supervision of this study.

It was a great honor to me to work under her guidance.

It is a great honor to express my thanks to

Prof. Dr. Shereen Abou Bakr Saleh, Lecturer of

Internal Medicine, Ain Shams University, who was very

kind, supportive and helpful throughout all the stages of

this work. She had such a great influence on my character

and behavior.

I owe many thanks, respect and appreciation to

Prof. Dr. Alaa El Din Ismail, Professor of General

Surgery, Ain Shams University, for offering me much of

his time and experience throughout the practical part of

this work and for help me to overcome the obstacles and

difficulties that arose along the way until finally the

project was completed.

Also, I am great fully indebted to

Prof. Dr. Doaa Zakaria Zaki, Lecturer of Tropical

Medicine, Ain Shams University, for her help, guidance

and revision of this work that helped me to finish this

project.

I am deeply grateful to my Mother, Father and

Brother, for their great support and help throughout my

life not only this great study….

Mohammed Fathy Sayed Zaky

List of Contents

Title Page

Introduction and Aim of the Work 1

Review of Literature

Chapter I

Chapter II

5

58

Patients and Methods 89

Results 95

Discussion 114

Conclusions 122

Recommendations 123

Summary 124

References 126

Arabic Summary -أ-

List of Abbreviations

ADA Adalimumab

AID Autoimmune Diseases

ALT Alanine Aminotransferase

ALB Albumin

APC Antigen Presenting Cell

ASA Amino salicylic Acid

ASCA Anti-Saccharomyces Cerevisiae Antibodies

ASCs Adult Stem Cells

ATA Antibodies formation to Adalimumab

ATI Antibodies formation to Infliximab

ATG Anti-thymocyte Globulin

AZA Azathioprine

BM Bone Marrow

BMA Bone Marrow Aspirate

BMI Body mass index

BMSCs Bone Marrow Stromal Cells

bpm Beat per minute

BSG British Society of Gastroenterology

BW Bodyweight

Cal Calories

CBC Complete Blood Cell Count

CD Crohn's Disease

CK Cytokeratin

CMV Cytomegalovirus

CRC Colorectal Cancer

CNS Central Nervous System

CRP C Reactive Protein

CsA Ciclosporin

CT Computed Tomography

CY Cyclophosphamide

List of Abbreviations (cont.)

DNA Deoxyribonucleic Acid

E1 Proctitis

E2 Left-sided Colitis

E3 Extensive Ulcerative Colitis

E. coli Escherichia Coli

ECCO European Crohn's and Colitis Organization

EIS Endoscopic Index of Severity

EN Enteral Nutrition

ERUS Endorectal Ultrasonography

ESCs Embryonic Stem Cells

ESR Erythrocyte Sedimentation Rate

FBCs Full Blood Counts

G-CSF Granulocyte Colony Stimulating Factor

g/dL gram/deciliter

GIT Gastro Intestinal Tract

GVHD Graft Versus Host Disease

HB Hemoglobin

HCT Haematopoietic Cell Transplantation

HLA Human Leukocyte Antigen

HSCs Hematopoietic Stem Cells

HSCT Hematopoietic Stem Cell Transplantation

IC Indeterminate Colitis

ICMs Inner Cell Masses

IBD Inflammatory Bowel Disease

IBDU Unclassified Inflammatory Bowel Disease

IBS Irritable Bowel Syndrome

IFN Interferon

IFX Infliximab

IL Interleukin

IMIDs Immune-mediated Diseases

List of Abbreviations (cont.)

IMiDs Immunomodulatory Drugs

iPSCs Induced Pluripotent Stem Cells

IV Intravascular

MAPCs Multipotent Adult Progenitor Cells

MDR Multidrug Resistance Gene

MHC Histocompatibility Complex

MTX Methotrexate

MP Mercaptopurine

MPCs Mesodermal Progenitor Cells

MRI Magnetic Resonance Imaging

MSCs Mesenchymal Stem Cells

NSAID Non-Steroidal Anti-Inflammatory Drug

NSC Neural Stem Cells

NHL Non Hodgkin Lymphoma

PD Processing Disposable

PSC Primary Sclerosing Cholangitis

SCT Stem Cell Therapy

SCs Stem Cells

SLE Systemic Lupus Erythematosus

SSCs Stromal Stem Cells

TB Tuberculosis

TBI Total Body Irradiation

TGF Transforming Growth Factor

TGN Thioguanine Nucleotides

TNF Tumor Necrosis Factor

TPN Total Parenteral Nutrition

UC Ulcerative Colitis

UK United Kingdom

USA United State of America

WBCs White Blood Cells

List of Figures

Fig. Subjects Page

(1) Extra-colonic manifestations of UC. 21

(2) Endoscopic image of ulcerative colitis. 22

(3) Histology of normal colon and UC 24

(4) Radiological features of ulcerative colitis. 26

(5) Extent of bowel involvement in UC. 48

(6) Stem cells. 58

(7) Development of Preimplantation Blastocyst 60

(8) Pluripotent stems cells. 60

(9) Embryonic stem cells. 63

(10) Hematopoietic & stromal SCs differentiation. 69

(11) Autologous HCT as a therapy for AID. 81

(12) SCs transplant using patient‟s own cells. 84

(13) Mechanism by which HCT ameliorate AID. 86

(14) Stem Cells Transplantation for Patients. 93

(15) Assessment of the presence of diarrhea. 96

(16) Assessment of the rectal bleeding. 97

(17) Assessment of the abdominal pain. 98

(18) Assessment of the disease extent. 99

(19) Assessment of the disease severity. 100

(20) Assessment of the medical treatment. 101

(21) Assessment of the disease outcome. 103

(22) Endoscopic picture of patient before and after SCT. 104

(23) Assessment of the motion frequency. 105

(24) Assessment of the body temperature. 106

(25) Assessment of the heart rate. 107

(26) Assessment of the body mass index. 108

(27) Assessment of the serum albumin. 109

(28) Assessment of the hemoglobin level. 110

(29) Assessment of the erythrocyte sedimentation rate. 111

(30) Assessment of the C-reactive protein. 112

(31) Assessment of the activity score. 113

List of Tables

Table Subjects Page

(1) The Montreal classification of UC. 8

(2) Disease severity index of UC 10

(3) Mayo activity scoring index. 10

(4) Endoscopic scores for UC. 10

(5) UC Endoscopic Index of Severity. 24

(6) Nutritional deficiencies in IBD. 32

(7) Algorithm for the treatment of UC. 51

(8) SCs during differentiation at each stage. 62

(9) Types of cell division. 62

(10) Comparison between patients as regard the presence of diarrhea 96

(11) Comparison between patients as regard the rectal bleeding. 97

(12) Comparison between patients as regard the abdominal pain. 98

(13) Comparison between patients as regard the disease extent. 99

(14) Comparison between patients as regard the disease severity. 100

(15) Comparison between patients as regard the medical treatment 101

(16) Comparison between patients as regard the disease outcome. 102

(17) Comparison between patients as regard the motion frequency 105

(18) Comparison between patients as regard the body temperature 106

(19) Comparison between patients as regard the heart rate. 107

(20) Comparison between patients as regard the body mass index. 108

(21) Comparison between patients as regard the serum albumin. 109

(22) Comparison between patients as regard the hemoglobin level 110

(23) Comparison between patients as regard the ESR. 111

(24) Comparison between patients as regard the CRP. 112

(25) Comparison between patients as regard the activity score. 113

Introduction

Aim of the Work

Review of Literature

Chapter I

Chapter II

Results

Discussion

Conclusion and

Recommendations

Summary

References

Arabic Summary

Role of Stem Cell Transplantation in the Treatment of Ulcerative Colitis

Health & Medicine

Transcript of Role of Stem Cell Transplantation in the Treatment of Ulcerative Colitis