RED BLOOD CELLS

by

Mary Yvonnette C. Nerves, MD, FPSP

Erythropoiesis

A process by which early erythroid precursor cells differentiate to becomethe mature RBCs

Primary regulator: ERYTHROPOIETIN

- stimulates red cell precursors at all levels ofmaturation to hasten the maturation process

- responsible for stimulating the premature releaseof reticulocytes into the bloodstream.

Erythropoiesis Total erythropoiesis:

- total number of red blood cells (RBCs) - measured by the myeloid-erythroid

(M:E) ratio from aspirate smears plus the estimate of cellularity from biopsy

sections

Effective erythropoiesis: - number of viable and functional RBCs available for physiologic needs - reflects the balance between the number ofcells produced and their life span- measured by the reticulocyte count, which is normally 1% of the total RBC count

Stages of Maturation1. Pronormoblast (Rubriblast)2. Basophilic Normoblast (Prorubriblast)3. Polychromatophilic Normoblast

(Rubricyte)4. Orthochromatic Normoblast

(Metarubricyte)5. Reticulocyte6. Erythrocyte

Pronormoblast Earliest recognizable and largest cell of the

erythrocyte series Morphology: - Size: 12 – 20 um

- Nucleus: large round, oval, dark violet; finechromatin; 1 – 2 nucleoli

- Cytoplasm: deep blue spotty, basophilic w/ a perinuclear halo

- N/C Ratio: 8:1 - BM (%): 1

Basophilic Normoblast

Hemoglobin synthesis begins at this stage

Morphology:

- Size: 10 – 15 um

- Nucleus: large round to sl oval; condensed,coarse chromatin; 0 – 1 nucleoli

- Cytoplasm: deeply basophilic; clusters of free ribosomes

- N/C Ratio: 6:1

- BM (%): 1-4

Polychromatic Normoblast Increased production of hemoglobin pigmentation and decreasing amounts of RNA

Last stage in which the cell is capable of mitoses

Morphology:

- Size: 10 - 15 um

- Nucleus: round nucleus, deep staining, may be centrally or eccentrically located; coarse & clumped chromatin

- Nucleoli: 0

Morphology:

- Cytoplasm: abundant blue-gray (RNA) to pink-gray (hemoglobin)

- N/C Ratio: 4:1

- BM (%): 10-20

Orthochromatic Normoblast

The last nucleated stage

Cannot synthesize DNA and cannot undergo cellular division

The NRBC sometimes seen in the peripheralcirculation

Morphology:

- Size: 8 - 10 um

- Nucleus: small pyknotic nucleus; dense chromatin; 0 nucleoli

- Cytoplasm: abundant red-orange cytoplasmuniform in color

- N/C Ratio: 1:2

- BM (%): 5-10

Reticulocyte Slightly larger than the mature RBC with residual amts of RNA Reticulocyte count: an index of bone marrow

activity or effective erythropoiesi Morphology: - Size: 8 - 10 um - Nucleus: anucleate cell containing small

amt of basophilic reticulum (RNA) - Nucleoli: 0 - Cytoplasm: large amt of blue-pink staining

hemoglobin cytoplasm

Erythrocyte

A biconcave 6 – 8 um disc

Life span: 120 days

Main function: to transport hemoglobin, aprotein that delivers oxygen from the

lungs to tissues and cells

Contains 90% hemoglobin and 10% H2O

normal conc of RBCs varies w/ age, sex &geographic distribution

Morphology:

- Size: 7 - 8 um

- Nucleus: anucleated cell

- Nucleoli: 0

- Cytoplasm: pink staining, zone of centralpallor is 1/3 of cell diameter devoid of hemoglobin

- N/C Ratio: NA

Hemoglobin: Structure & Function

A conjugated protein that serves as the vehicle for the transportation of O2 and CO2

When fully saturated, each gram of Hgb can hold 1.34 mL of O2 A molecule of Hgb consists of 2 pairs of polypeptide chains (“globin”) and 4 prostheticheme grps each contg 1 atom of ferrous iron

DESCRIPTION of TERMS

SIZE DESCRIPTORS

Anisocytosis: variation in the sizeof the RBCs due to a pathologic condition

Normocytic: normal sized biconcave disc RBC- normal MCV

Microcytic: Smaller RBCs less than 6 um- MCV < 80 fl- Defect / Change: abn size due to failure

of hgb synthesis- Dse: IDA, Thalassemia, Chronic dse

Macrocytic: Larger RBCs greater than 9um- MCV > 90 fl- Defect / Change: impaired DNA synthesis

/ stress erythropoiesis - Dse: Megaloblastic anemia / liver dse /

MDS / Alcoholism / Malaria

Macrocytic Microcytic

CHROMICITY DESCRIPTORS Normochromic: normal in color; pale central

area occupies less than 1/3

- Defect / Change: normal amt of Hgb

- Normal indices

Hypochromic: an RBC that has a decreasedHgb complement

- central pallor exceeds 1/3 of diameter ofcell

- Defect / Change: reduced Hgb content ( MCHC)

- Assoc conditions: IDA / Thalassemia

“Hyperchromic”: no central pallor

- Defect / Change: greater than normal MCHC

- Assoc condition: Spherocytosis

“Hyperchromic” Hypochromic

Polychromasia: blue-gray coloration

- Defect / Change: presence of RNA

- Assoc condition: increased erythropoietic activity / hemorrhage / hemolysis

SHAPE DESCRIPTORS Poikilocytosis: variation in shape of the RBC

- Defect / Change: irreversible alteration of membrane

- Assoc conditions: Anemia / Hemolytic states

Discocyte: normal biconcave erythrocyte - 6 – 8 um diameter; 0 – 2 um thickness

- Aka: Normocyte

Normal Red Cells (SEM)

Acanthocyte: spheroid w/ 3 – 12 irreg spikesor spicules

- Aka: spur cell

- decreased cell volume

- Defect / Change: inc ratio of chole to lecithin

- Assoc conditions:

end-stage liver dse

Pyruvate kinase def

Hemolytic anemia

Abetalipoproteinemia

Blister cell: contains 1 or more vacuoles

- Aka: Bite cells

- thinned periphery

- Defect / Change: formed by removal of Heinz bodies

- Assoc conditions:

Hemolytic episodes

G6PD def

Hemoglobinopathies

Codocyte : peripheral rim of Hgb surr by clear area & central hemoglobinizedarea (bull’s eye)

- Aka: target cell

- Defect / Change: excess of surface to volume ratio

- Assoc conditions:

Hemoglobinopathies

Thalassemia

Liver dse

Postsplenectomy

Dacryocyte: teardrop or pear-shaped w/single elongated point or tail

- Aka: tear drop cell- Defect / Change: squeezing &

fragmentation during splenicpassage

- Assoc conditions: Myeloid metaplasiaThalassemiaMegaloblastic anemiaHypersplenism

Drepanocyte: crescent-shaped cell thatlacks zone of central pallor

- Aka: Sickle cell

- Defect / Change: polymerization ofdeoxygenated Hgb

- Assoc conditions:

Sickle cell anemia

SC disease

S-thalassemia

Echinocyte: regular 10-30 scalloped shortprojections evenly distributed / spiny-like

- Aka: Burr cell / crenated RBC- Defect / Change: Depletion of ATP Exposure to hypertonic soln Artifact in air drying- Assoc conditions:

UremiaCirrhosis / HepatitisChronic renal dse

Ovalocyte: egglike or oval-shaped cell

- Defect / Change:

Hgb has bipolar arrangement

Reduction in membrane chole

- Assoc conditions:

Megaloblastic BM

Myelodysplasia

Sickle cell anemia

Elliptocyte: rod or cigar shape, generallynarrower than ovalocytes

- Defect / Change: polarization of Hgb

- Assoc conditions:

Thalassemia

Iron def

Hereditary elliptocytosis

Schistocyte: Fragmented RBCs varying in size & shape

- Aka: Helmet cells- Defect / Change: extreme fragmentation

produced by damage of RBC by fibrin, altered vessel walls, prosthetic heart

valves- Assoc conditions:

DIC / TTP / BurnsMicroangiopathic hemolytic anemia

Spherocyte: smaller in diameter than normal RBC w/ concentrated Hgb content; no visible central pallor- Defect / Change:

lowest surface area to volume ratiodefect of loss of membrane

- Assoc conditions: Hereditary spherocytosisIso- & autoimmune hemolytic anemiaSevere burnsHemoglobinopathies

Stomatocyte: normal sized cell w/ slitlike area in center

- Defect / Change:

artifact of slow drying

known to have inc permeability to Na+

- Assoc conditions:

Hereditary stomatocytosis

Acute alcoholism

Liver dse

RED CELL INCLUSIONS

Basophilic Stippling - cytoplasmic remnants of RNA- Fine: thin round dark blue granules

uniformly distributed- Defect/Change: represents

polychromasia (reticulocyte)- Coarse: medium sized uniformly

distributed- Defect/Change: represents

impaired erythropoiesis

Basophilic Stippling - Assoc conditions:

Thalassemia

Lead Poisoning

Increased reticulocytosis

Cabot Ring - rings, loops, or figure eights; red to

purple- Defect / Change: remnants of

microtubules of mitotic spindle

- Assoc conditions:Megaloblastic anemiaDyserythropoiesis

Heinz bodies - deep purple irregularly shaped inclusions

found on RBC inner surface of membrane - Defect / Change: represent precipitated,

denatured Hgb due to oxidative injury - Assoc conditions:

Hereditary defects in HMSG6PD defUnstable HgbsSplenectomized ptsThalassemia

Howell-Jolly bodies: coarse round denselystained purple 1-2 um granules

eccentrically located on periphery of membrane

- Defect / Change: nuclear remnants; contain DNA

- Assoc conditions:Megaloblastic anemiaSevere hemolytic processThalassemiaAccelerated erythropoiesis

Pappenheimer bodies: small, 2-3 um irregular basophilic inclusions that aggregate in small clusters near periphery w/ Wright’s stain

- Defect / Change: unused iron (nonheme) deposits

- Assoc conditions:Sideroblastic anemiaDefective erythropoiesisMDSHemolytic anemiaThalassemia

Ringed Sideroblasts- Nucleated RBC that contains nonheme

iron particles (siderotic granules) arranged in ring form

- Defect / Change: excessive iron overload in mitochondria of normoblasts

- due to defective heme synthesis- Assoc conditions:

Sideroblastic anemiaMDS

Ringed SideroblastsPrussian blue iron stain showing excess accumulation of iron as ferritin in mitochondria ringing nucleus.

Siderocyte: non-nucleated cell containing iron granules

- Defect / Change: excessive iron overload in mitochondria of normoblasts- due to defective heme synthesis

- Assoc conditions:Sideroblastic anemiaMDS

Autoagglutination: clumping of RBCs- Defect / Change: presence of antibody- Assoc conditions:

Cold agglutininAHA

Rouleaux Formation: alignment of RBCslinear appearing as stacks of coins

- Defect / Change: concentration of fibrinogen &

immunoglobulin- Assoc conditions:

MM / Waldenstrom’s macroglobulinemia

Red Cell Studies

Hematologic tests used to measure several important parameters that reflect rbc

structure and function: 1) Hemoglobin determination2) Erythrocyte count3) Hematocrit4) Erythrocyte Indices: MCH, MCHC,

MCV 5) Reticulocyte Count6) Osmotic Fragility Test7) Erythrocyte Sedimentation Rate (ESR)

Measurement (units) Men Women

Hemoglobin (gm/dL) 13.6-17.2 12.0-15.0

Hematocrit (%) 39-49 33-43

Red cell count (106/μL) 4.3-5.9 3.5-5.0

Reticulocyte count (%) 0.5-1.5

Mean cell volume (μm3) 82-96

Mean corpuscular hemoglobin (pg) 27-33

Mean corpuscular hemoglobin concentration (gm/dL) 33-37

RBC distribution width 11.5-14.5

Adult Reference Ranges for Red Blood Cells

Hemoglobin

- involves lysing the erythrocytes, thus producingan evenly distributed solution of

hemoglobin in the sample

- Hemiglobincyanide Mtd: blood is diluted in a soln of K3Fe(CN6). The K3Fe(CN6) oxidizes Hgbs to hemiglobin (metHgb) and K cyanide provides cyanide ions to form HiCN, w/c has a broad absorption max at a wl of 540 nm

Erythrocyte Count- involves counting the number of rbcs per unitvolume of whole blood. - expressed as number of cells per unit volume,

specifically cells/µL- NV: Female = 4.2 - 5.4 x 106/µL

Males = 4.7 - 6.1 x  106/µL

Hematocrit- sometimes referred to as the Packed CellVolume (PCV) or volume of packed red cells - is the ratio of the volume of RBCs to that of the

whole blood- varies with age and sex- expressed as a percentage or as adecimal fraction

Plasma

Buffy coat

Red cells

Erythrocyte Indices1) Mean Cell Volume (MCV)

- average volume of red cells- calculated from the Hct and RBC count

MCV = Hct x 1000 RBC (in millions/uL)

- expressed in femtoliters (fl) or cubic micrometers

2) Mean Cell Hemoglobin (MCH)- content (weight) of Hgb of the average red cell- calculated from the Hgb and RBC

count

MCH = Hgb (in g/L) RBC (/L)

- value is expressed in picograms (pg)

3) Mean Cell Hemoglobin Concentration (MCHC)- the average conc of Hgb in a given

volume of packed red cells- calculated from the Hgb conc & the Hct

MCHC = Hgb (in g/dL)

Hct

- expressed in g/dL

Morphologic Classification of Anemias

Type of Anemia Blood Constants

MCV (mm3 or fl)

MCHC (g.Hb/dl.RBC or

mmol/l)

Microcytic hypochromic 60-87 20-30

Macrocytic normochromic 103-160 32-36

Normocytic normochromic 87-103 32-36

Microcytic normochromic 60-87 32-36

Reticulocyte Count- Principle: Reticulocytes are immature non-nucleated red cells that contain RNA andcontinue to synthesize Hgb after the loss ofthe nucleus- Supravital staining: blood is briefly incubated

in a soln of new MB or BCB, the RNA isprecipitated as a dye-ribonucleoproteincomplex dark blue network (reticulum or

filamentous strand) - NV: 0.5 – 1.5% or 24 – 84 x 109/L

Osmotic Fragility Test (OFT)- a measure of the ability of red cells to take up

fluid without lysing- Red cells are suspended in a series of tubes

contg hypotonic solns of NaCl solns varying from 0.9% to 0.0%, incubated at room temp for 30 mins and centrifuged - the percent hemolysis in the supernatant solns

is measured & plotted for each NaCl conc.

- The larger the amount of red cell membrane(surface area) in relation to the size of thecell, the more fluid the cell is capable ofabsorbing before rupturing- Cells that are more spherical, w/ a decreased

surface/volume ratio, have a limited capacity to expand in hypotonic solns & lyse at a higher conc of NaCl than do normal biconcave cells OFT

- Cells that are hypochromic & flatter have a greater capacity to expand in hypotonic solns, lyse at a lower conc than do normal cells, & are said to have decreased osmotic fragility

- Cells with increased surface/volume ratio are osmotic resistant IDA, thalassemia, liver dse, & reticulocytosis

Erythrocyte Sedimentation Rate (ESR)- detect and monitor an inflammatory response

to tissue injury (an acute phase response) in which there is a change in the plasma conc of several proteins

- Principle: When well-mixed venous blood is placed in a vertical tube, RBCs will tend to fall toward the bottom. The length of the

fall of the top of the column of RBCs in a given interval of time is called the ESR

- ESR is affected by (3) FACTORS:a) erythrocytesb) plasma compositionc) mechanical / technical factors

- Red Cell Factors: Anemia increases ESR (change in RBC plasma ratio

favors rouleaux fotn) ESR is directly proportional to the weight of thecell

aggregate & inversely proportional to the surface area

Microcytes sediment slower than macrocytes Rouleaux accelerate the ESR Red cells w/an abnormal or irregular shape hinder

rouleaux fotn & lower the ESR

- Plasma Factors: Elevated levels of fibrinogen accelerate ESR Albumin & lecithin retard ESR Cholesterol accelerate ESR

- Mechanical / Technical Factors: A tilt of 3o can cause errors up to 30% ESR ESR increases as the temp increases ESR tubes with a narrower than standard bore will

generally yield lower ESR ESR stands fro > 60 mins falsely elevated ESR Greater conc of EDTA falsely low ESR

- Methods: Westergren Mtd / Wintrobe Mtd

ERYTHROCYTE DISORDERS

Two main disorders affecting RBCs:

1. Polycythemia (Erythrocytosis)

- an elevated Hct level above the normal range

2. Anemia

- a reduction below normal limits of the total circulating red cell mass

Relative Reduced plasma volume (hemoconcentration)

Absolute

Primary Polycythemia vera, rare erythropoietin receptor mutations (low erythropoietin)

Secondary High erythropoietin

  Appropriate: lung disease, high-altitude living, cyanotic heart disease

  Inappropriate: erythropoietin-secreting tumors (e.g., renal cell carcinoma, hepatocellular carcinoma, cerebellar hemangioblastoma)

Pathophysiologic Classification of Polycythemia

POLYCYTHEMIAMay be classified into (2) major conditions:

1) Relative Polycythemia

- an increase in the Hct or red cell count as a result of decreased plasma volume

- total red cell mass is NOT increased

- Assoc conditions: acute dehydration or hemoconcentration / pts on diuretic

therapy / Gaisbock’s syndrome (psedopolycythemia or stress erythrocytosis)

- BM: Normal

2) Absolute (or Secondary) Polycythemia- an erythropoietin mediated increase in

RBCs and Hgb due primarily to ahypoxic situation

- increase in the total red cell mass in the body assoc w/ normal or sl increased plasma volume

- Assoc conditions: tumors / anabolic steroids / & renal dso such as cystic dse, hydronephrosis / & adrenal cortical hyperplasia

- BM: Erythroid hyperplasia

3) Polycythemia rubra vera (Primary Erythrocytosis)- an absolute increase in all cell types,

RBCs, WBCs and platelets- not dependent on erythropoietin levels- BM: all three cell lines increased

(panhyperplasia)

ANEMIA

Decreased oxygen carrying capacity of the blood

Anemia may also be "defined" in terms of the Hb content

Hb < 12 g/dL in an adult male

Hb < 11 g/dL in an adult female

AnemiaMales: Hb < 13.5 Hct < 41Female: Hb < 12 Hct < 36

MCV

MicrocyticMCV < 80

NormocyticMCV 80-100

MacrocyticMCV > 100

Reticulocyte Count

Low

High

Marrow FailureAplastic anemiaMyelofibrosisLeukemia /MetastasisRenal failureAnemia of Chronic disease

Iron deficiency anemiaThalassemiaAnemia of Chronic diseaseSideroblastic anemia

Megaloblastic anemiaAlcoholic liver disease

Sickle cell anemiaG6PD def anemiaHereditary spherocytosisAIHAPNH

ANEMIAS SECONDARY TO BLOOD LOSS

Acute: e.g., hemorrhage due to trauma, massive GI bleeding,

or child delivery. Usually the iron stores remain normal.

Chronic: e.g., bleeding peptic ulcer or excessive menstrual

bleeding.

HYPOPROLIFERATIVE ANEMIAS (Impaired Production)

reduced production of red cells can be subdivided into: deficiency of haematinics

iron deficiency B12 and folate deficiency

dyserythropoiesis (production of defective cells) anaemia of chronic disorders (AOCD) myelodysplasia sideroblastic anaemia

marrow infiltration (myelophthisic anemia) aplasia (failure of production of cells)

aplastic anaemia red cell aplasia

Normal forms of iron (Fe) and iron metabolism Functional iron is found in Hb, myoglobin, and

enzymes (catalase & cytochromes) Ferritin: physiological storage form Hemosiderin: degraded ferritin + lysosomal

debris (Prussian blue positive) Iron is transported by transferrin

Iron Deficiency Anemia

causes: Dietary deficiency: elderly, children and poor Increased demand: children & pregnant women Decreased absoprtion:

generalized malabsorption

after gastrectomy Chronic blood loss:

GI bleeding (e.g. peptic ulceration, carcinoma of stomach or colon)

menorrhagia

urinary tract bleeding

Hook worm (Ancylostoma duodenale adult worm sucks 0.2 ml blood/day)

Lab Findings: Microcytic, hypochromic anemia. Low serum iron BM: show absence of iron Ferritin: Low serum ferritin indicates low body stores

of iron Transferrin: These carrier proteins will be

unsaturated and available to bind iron, hence the Total Iron Binding Capacity (TIBC) is increased with anemia.

Char by iron being trapped in BM macrophages

Can be grouped in 3 categories:

- chronic microbial infections (eg. Osteomyelitis)

- chronic immune disorders (eg. RA)

- Neoplasms (eg. lymphoma, breast/lung CA)

Chronic inflamm dso inc IL-1, TNF, IF-Gamma - reduction in renal erythropoietin marrow erythroid

precursors do notproliferate

- hepcidin synthesis in liver inhibits release of iron

Anemia of Chronic Disease (AOCD)

Labs: low serum iron

increased serum ferritin

decreased TIBC

normochromic, normocytic anemia or hypochromic, microcytic anemia

A group of dso in which the blood and BM hematopoietic cells display changes

Pathogenesis: impaired DNA synthesis (delayed mitoses) while RNA is not impaired; this produces nuclear-cytoplasmic asynchrony

Megaloblastic anemias can be divided into groups:

- anemia caused by B12 deficiency

- anemia caused by folate deficiency

- anemias nonresponsive to either therapy

Megaloblastic Anemia

important background knowledge: B12:

vitamin B12 is required for DNA replication and inhibition of transcription of DNA to RNA

B12 is normally absorbed from gut by the following mechanism:

- secretion of intrinsic factor by parietal cells in stomach

- binding of intrinsic factor and vitamin B12 in lumen

- intrinsic factor- B12 complex is absorbed in terminal ileum through pinocytotic vesicles

Folate: - folate is required for DNA replication and inhibition of

transcription of DNA to RNA

lack of B12 or folate means that RNA builds up and the cells become too large

Causes: causes of vitamin B12 deficiency (pernicious

anaemia) lack of intrinsic factor

- atrophic gastritis - parietal cells are destroyed - gastrectomy

malabsorption of B12 not related to lack of intrinsic factor

- tropical sprue or bacterial overgrowth of terminal ileum - ileal disease (e.g. Crohn's disease affecting the terminal

ileum) - fish tape-worm (these attach to intestinal wall, and

therefore in large enough numbers, may prevent B12-intrinsic factor complex absorption in terminal ileum)

- poor diet - rare

causes of folate deficiency poor diet - especially in alcoholics malabsorption - coeliac disease increased cell turnover (e.g. leukaemia, chronic

haemolysis, pregnancy) antifolate drugs (e.g. phenytoin)

Peripheral Blood Findings:

WBCs Normal or decreased

RBCs Decreased

Hgb Decreased

MCV Increased (usualy > 100 fl)

MCH Increased

MCHC Normalor slightly decreased

Platelet count Normal or decreased

Reticulocyte count Decreased

Reticulocyte Count

Normal or decreased Increased

Macrocyte normal, no hyersegmented

neutrophils

Macro-ovalocytes,Hypersegmented

neutrophils

BM not megaloblastic

R/O Refractory anemia,Sideroblastic anemia,

Myelodysplasia, drug induction,

liver disease,Aplastic anemia

HemolyticHemorrhagic

deficiency

Morphologic Abnormalities: Large RBC's with nuclear-cytoplasmic dyssynchrony Ovalocytes: The large RBC's tend to have an oval-shape.

Hypersegmented Neutrophils: One of the earliest signs of disease. 5 or 6 lobes

Howell-Jolly Bodies: Nuclear fragments seen in Megaloblastic anemia.

pancytopenia associated w/ a severe reduction in the amt of hematopoietic tissue that results in deficient production of blood cells

Etiology: Acquired

idiopathic Chemical agents Physical agents Viral infections

Inherited Fanconi’s anemia

Aplastic Anemia

Pure red cell aplasia: erythrocyte stem cells are suppressed, but the other formed elements of blood are unaffected

Anemia due to isolated depletion of erythroid precursors in the marrow, and may be acute or chronic.

Lab Findings:

- Normochromic, normocytic or macrocytic anemia

- Reticulocytes are decreased or absent because it is hypoproliferative.

- BM: hypocellular or dry tap

reduction in all cell lines

HEMOLYTIC ANEMIAS (Increased Destruction)

Grp of dso that can be inherited, acquired,or drug-induced

Char by an increased red cell destruction or shortened survival of the RBC

Char by increased BM activity, polychromasia, nucleated RBCs and an increased reticulocyte count w/ stress reticulocytes

Hemolytic anemias share the ff. features:

1. shortened red cell life span, that is, premature destruction of red cells

2. elevated erythropoietin levels and increased erythropoiesis in the marrow & other sites

3. accumulation of products of Hgb catabolism,due to an increased rate of red celldestruction

HEMOLYTIC ANEMIAS Acquired

Immune-mediated- Autoimmune- Alloimmune (Transfusion)- Drug-induced

Microangiopathic Infection

Hereditary Enzymopathies Membranopathies Hemoglobinopathies

INTRINSIC DEFECTSHereditary Spherocytosis

abnormal cell membrane assoc cytoskeletoncausing red cells to be spherical and

fragile

principle defect is an abnormality of the membrane protein ankyrin

Lab findings: Normocytic, hyperchromic anemia (normal MCV and increased

MCHC)

- increased pigment catabolism, erythroid hyperplasia, & reticulocytosis

- red cells with increased OFT

Glucose-6-Phosphate Dehydrogenase Deficiency

Normal: G6PD metabolises glucose, and formssmall amounts of ATP (which

maintains the cell cytoskeleton and membrane) and NADPH (which mops up free radicals) G6PD def renders the cell susceptible to damage

by free radicals an X-linked recessive condition, in which haemolytic crises are precipitated by infections or certain drugs Lab findings: poikilocytes & spherocytes, & Heinz

bodies (stain w/ methyl violet)

HEMOGLOBINOPATHIES Normal Hgb:

HbA / HbF / HbA2 (adult)

Hb Gower-1 and 2 / Hb-Portland (embryonic)

Hemoglobin-A alpha2, beta2 Predominant hemoglobin in adults

Hemoglobin-A2

alpha2, delta2

Found in normal adults

Hemoglobin-F alpha2, gamma2

Fetal (cord) hemoglobin, with higher O2-binding affinity

Hemoglobin-S alpha2, beta2 (beta: Glu-6

---> Val)

Sickle-Cell Hemoglobin. Sickle crisis can result from low O2-tension.

Hemoglobin-C alpha2, beta2 (beta: Glu-6

---> Lys)

Hemoglobin-C Disease. Second most common hemoglobinopathy.

THALASSEMIACaused by impaired production of one of the polypeptide chains of the Hb moleculeEpidemiology: Mediterranean, African & Asian ancestryautosomal recessive disease Types according to clinical severity:thalassaemia major = homozygote;

thalassaemia minor = heterozygote Types according to molecular defect: beta thalassaemia alpha thalassaemia

Beta-Thalassemia Major (Homozygous state):

- severe hypochromic, microcytic anaemia, hepato- splenomegaly, marrow hyperplasia causing

skeletal deformities, haemochromatosis develops with repeated transfusions

Minor (Heterozygous states):

- reduction in HbA, but increase in HbA2; mild anaemia with hypochromia

Clinical Nomenclature

Genotype Disease Molecular Genetics

β-Thalassemias

Thalassemia major

Homozygous β0-thalassemia (β0/β0)

Severe; requires blood transfusions

Rare gene deletions in β0/β0

Defects in transcription, processing, or translation of β-globin mRNA

  Homozygous β+-thalassemia (β+/β+)

 

Thalassemia intermedia

β0/ββ+/β+

Severe, but does not require regular blood transfusions

Thalassemia minor

β0/ββ+/β

Asymptomatic with mild or absent anemia; red cell abnormalities seen

Alpha-Thalassemianote that there are 4 copies of the alpha globin gene (not 2), and therefore four possible degrees of alpha thalassaemia exist

3 good copies - silent carrier 2 good copies - mild anaemia with microcytosis 1 good copy - moderate haemolytic anaemia

with hypochromia and mycrocytosis; HbH (tetramer of beta)

0 good copies - lethal in utero (hydrops fetalis)

α-Thalassemias

Hydrops fetails -/--/- Lethal in utero without transfusions

Mainly gene deletions

HbH disease -/--/α Severe; resembles β-thalassemia intermedia

α-Thalassemia trait

-/-α/α (Asian)-/α-/α (black African)

Asymptomatic, like β-thalassemia minor

Silent carrier -/αα/α Asymptomatic; no red cell abnormality

Thalassemia MajorPatient with thalassemia major due to heterozygous hemoglobin E/B thalassemia. Note prominent target cells, anisopoikilocytosis, and three nucleated red cells (normoblasts) 

Sickle Cell Disease Endemic to Sub-saharan Africa, due to heterozygous advantage conferred against Falciparum Malaria (infected RBC's preferentially sickle and are thus taken to the spleen and sequestered, limiting the spread of infection) PATHOGENESIS: Point-mutation of Glu Val at 6th

position of beta-globin chain Pathophysio: abn Hgb polymerises at low O2

saturation causing abnormal rigidity and deformity of red cells and become

abnormality fragile (and undergo haemolysis and sludge in small vessels) autosomal recessive, with a point mutation in beta gene

forming an abnormal HbS; more common in Negroes

Lab. Findings

Smear: normochromic, normocytic anemia, increased polychromasia, normoblasts are present, numerous target cells, Howell-Jolly and Pappenheimer bodies are present, sickle cells

OFT decreased

BM: normoblastic hyperplasia w/ increased iron storage

Electrophoresis: no HbA, 80% HbS (SCD)

Sickle Cells (SEM)Scanning electron micrograph (SEM) showing sickle cells obstructing small vessel. 

EXTRINSIC DEFECTSImmune Hemolytic Anemias

Dso in w/c erythrocyte survival is reduced because of the deposition of Ig &/or `

complement on the red cell membrane

Classification:

1. Autoimmune Hemolytic Anemia

2. Isoimmune Hemolytic Anemia

3. Drug-induced Hemolytic Anemia

LAB: (+) direct & indirect antiglobulin tests

Agglutination of erythrocytes is seen on this peripheral blood smear

Coomb’s Test

Traumatic Hemolytic AnemiaChar by striking morphologic abn of the red cells,

w/c include fragments (schistocytes) & irregularly contracted cells (triangular cells, helmet cells)

MICROANGIOPATHIC HEMOLYSIS: RBC's being damaged by intravascular fibrin-clots, in small vessels. DIC, TTP, HUS.

MACROANGIOPATHIC HEMOLYSIS: Damage by artifical heart valves.

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