Philomène Toquet

Abnormalities of the LDL metabolism:physiology, pathophysiology and treatments

Leticia Szadai

2015-20163rd year of MedicineJPEMS Program - Pathophysiology module

Under the supervision of Zsófia Mezei-Leprán MD., Ph.D

Publically presented on:October 6th 2015 in Szeged

Erwan Williamson

C O N T E N T

1.INTRODUCTION

2.PHYSIOLOGY OF LDL METABOLISM2.1.Synthesis of LDL2.1.1.Exogenous pathway2.1.2.Endogenous pathway2.1.3.The reverse cholesterol pathway

2.2.Regulation of uptake of LDL by cells2.2.1.Hepatic uptake of LDL2.2.2.Extra-hepatic uptake of LDL

3.PATHOPHYSIOLOGY OF LDL METABOLISM3.1.Increased synthesis3.1.1.Metabolic syndrome3.1.2.Diabetes type 1 and 23.1.3.Hypothyroidism3.1.4.Hypercorticism3.1.5.Nephrotic syndrome3.1.6.Infection3.1.7.Ethanol intake3.1.8.Drug intake3.1.9.Menopause3.2.Reduce uptake by the cells 3.2.1.Familial hypercholesterolemia3.2.2.Familial defective ApoB-1003.2.3.PCSK9 mutation3.2.4.ARH mutation3.2.5.Cholesterol 7-hydroxylase deficiency3.3.Harmful LDL3.3.1.Sitosterolemia3.3.2.Oxidized LDL3.3.3.Small density LDL3.3.4.Uremia

4.Treatments5.Summary6.References7.Declaration

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ABBREVIATIONS:

ABCA1: ATP-binding cassette transporter A1 Apos: apo(lipo)proteinsARH: autosomal recessive hypercholesterolemia gene ASCVD: arteriosclerosis cardiovascular diseaseCAD: coronary artery diseasesCE: cholesteryl esterCETP: cholesteryl ester transfer proteinChol-E: cholesterol estersCM: chylomicronCVD: cardiovascular diseasesCYP7A1: Cholesterol 7 -hydroxylaseEGF: epidermal growth factorFDB: familial defective apoB-100FH: familial hypercholesterolemiaHDL : high-density lipoproteinheFH: heterozygous FHHL: hepatic lipaseHMG-CoA: 3-hydroxy-3-methyl-glutaryl-CoAhoFH: homozygous FHIDL: intermediate density lipoproteinLCAT: lecithin-cholesterol acyltranferaseLDL: low-density lipoproteinLDL-C: low-density lipoprotein cholesterolLDLR: LDL receptorsLP: lipoproteinLPL: lipoprotein lipaseLPS: lipopolysaccharidesLXR: liver X receptorMTP: microsomal triglyceride transfer proteinNPC1L1: Niemann-Pick C1-like 1ox-LDL: oxidized LDLPCSK9: proprotein convertase subtilisin/kexin type 9PPAR: peroxisome proliferator-activated receptorsdLDL: small dense LDLSRBI: scavenger receptor BISREBP: sterol regulatory element binding proteinTG: triglycerideTSH: thyroid - stimulating hormoneVLDL: very-low-density lipoproteinWHO: World Health Organization

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1.INTRODUCTION

The abnormalities of LDL metabolism are usually associated with the most severe

cardiovascular diseases (CVD), for example coronary artery diseases (CAD), peripheral

artery diseases and stroke including atherosclerosis. These disorders of fat metabolism

strongly correlate with plasma low-density lipoprotein cholesterol (LDL-C) level (Marais,

2015).

If the balance of LDL metabolism collapses, it causes a rise in the concentration of

serum LDL and the building-up of arterial plaque that leads to atherosclerosis. However,

nowadays the elevation plasma LDL level becomes a global proposition. As the pathological

amount of LDL and its inner content, cholesterol, are accumulating in vessel walls, it affects

a blockage in the bloodstream. Consequently, the raised level of cholesterol increases the risk

of ischemic heart disease and stroke.

Figure 1 - Raised BC: 2008 http://www.who.int/gho/ncd/risk_factors/cholesterol_text/en/

These diseases give an image about the significant consequences of the abnormalities

of LDL metabolism, like increased cholesterol, all over the world. Actually, according to

WHO statistic the incidence of elevated cholesterol level was the highest in Europe (54% for

both genders). This average is followed by America with 48% for both genders, and then

Africa with 22.6% and South East Asia with 29.0% in 2008 (Fig. 1.).

Overall, a big number of patients are concerned about the abnormalities of LDL

metabolism, therefore the first aim of the present report to demonstrate the importance of

LDL abnormalities and its therapy in a point of view.

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2.PHYSIOLOGY OF LDL METABOLISM

Dietary fat and endogenous lipid synthesized in the liver must be transported between

tissues and organs to be metabolized there. This plasma transportation is provided by water

soluble macromolecules: lipoproteins. Lipoprotein metabolism abnormalities are a major

cause of atherosclerosis.

2.1.Synthesis of LDL

The low-density lipoprotein (LDL) is a globular molecular complex. Besides

chylomicron, very-low-density lipoprotein (VLDL), intermediate density lipoprotein (IDL),

high-density lipoprotein (HDL), it belongs to the lipoprotein’s family, which are responsible

for transporting lipid molecules, especially cholesterol. The structure of LDL, as other

lipoproteins, might be separated to amphilic and nonpolar portions. The micro emulsion’s

surface has an encompassment of amphilic phospholipids and non-esterified cholesterol while

the inner part contains a large number of hydrophobic triacylglycerol and cholesterol esters

(Chol-E) with a function of storing and carrying of cholesterol. The different types of lipid-

protein complexes are differentiated by their densities, diameters and molecular weight. The

more the complex contains proteins, the higher density it has. And the more the complex

contains lipids, the lower density it has.(chylomicrons <VLDL <IDL <LDL <HDL)

(Silbernagl, 2009).

These complexes also are distinguished from other complexes by their proteins

composition (Figure 2).

Lipoprotein particle Apolipoprotein

VLDL (very low density lipoproteins) Apo-B100, Apo-CI, Apo-E

IDL (intermediary density lipoproteins) Apo-B100, Apo-E

LDL (low density proteins) Apo-B100

HDL (high density proteins) Apo-A1

Chylomicrons Apo-B48, Apo-E, Apo-CII

Figure 2 - Apoproteins of lipoproteins

The apo(lipo)proteins (Apos) that are needed for serving lipids to the targeted cells. For

instance, ApoAII and ApoB48 as structural elements, while ApoB100 and ApoE as ligands,

ApoAI and ApoCII as enzyme activators. The way, which leads to LDL synthesis, is an

endogenous pathway. It also exist an exogenous pathway, which can implicate into

endogenous pathway and interferes with the LDL metabolism (Silbernagl, 2009).

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2.1.1.Exogenous pathway

Dietary lipids are absorbed via the small intestine and packaged into large particles

known as chylomicrons, which contain the intestine-specific ApoB48. Chylomicrons

transport lipids from the gut to the periphery (skeletal, muscle and fat tissue), where their

ApoCII activates the endothelial lipoprotein lipase (LPL); thus free fatty acids are split off

and are taken up by muscle cells and fat tissue. Chylomicrons remnant particles eventually

return to the liver where they bind to receptor via ApoE. They are endocytosed and in this

way deliver their triglycerides as well as their cholesterol and cholesterol esters. Some,

relatively enriched in cholesterol, integrate endogenous hepatic lipid pathways (Silbernagl,

2009).

2.1.2.Endogenous pathway

VLDL transports endogenous triglycerides, phospholipids, cholesterol, and

cholesterol esters. Nascent VLDL released from the liver contains apoB100, cholesterol,

cholesterol esters and triglycerides. In the bloodstream, VLDL picks up ApoCII (essential for

LPL’s operation) and ApoE given by HDL. At this point, nascent VLDL becomes a mature

VLDL. VLDL will come in contact with LPL on endothelial cells. LPL will remove

triglycerides from VLDL for storage and energy production. The triglycerides are removed

from VLDL with LPL. The composition of the molecule changes and it becomes

intermediate-density lipoprotein, IDL. 50% of IDLs are recognized by receptors in the liver

cells because of the apoB100 and apoE, their contents are endocytosed. For the other 50% of

IDL, when their cholesterol content becomes greater than the content of triglyceride due to

action of hepatic lipase, they become LDL, with apoB100 as the primary apolipoprotein. In

the end of this procedure, the ApoB100 of LDL binds to the LDLR, on the surface of hepatic

and extrahepatic cells. The LDL-LDLR complex is internalized in an endocytic vesicle to

participate in a lysis subsequently the receptor will recirculate to the cell membrane to bind a

new LDL molecule. As a result of this process, the intracellular cholesterol level mighty

increase, for this reason the synthesis of LDL receptor and 3-hydroxy-3-methyl-glutaryl-CoA

reductase (HMG-CoA reductase) will be inhibited (Fig. 3.) (Silbernagl, 2009).

The cholesterol content of cell is stored, used for cell membrane structure, or converted into

other products such as steroid hormones or bile acids. If the cellular intake of LDL is

damaged, resulting elevation of serum cholesterol level that induces atherosclerosis.

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Figure 3 - Regulation of LDL receptor expression and cholesterol synthesis.

http://i0.wp.com/www.namrata.co/wp-content/uploads/2012/11/cholesterol-

metabolism.jpg?resize=628%2C484

2.1.3.The reverse cholesterol pathway

High-density lipoprotein (HDL) plays an essential role in plasma lipid transport. HDL

is also the major vehicle for the transport of cholesterol from peripheral cells to the liver for

excretion and catabolism. This process, known as reverse cholesterol transport (Fig. 4.),

occurs in three stages: extravascular, intravascular, and intrahepatic. In the extravascular

phase, non-esterified cholesterol is removed from cell membranes by interaction with the

ATP-binding cassette transporter A1 (ABCA1). Dietary cholesterols (animals and vegetable)

enter in the enterocyte via Niemann-Pick C1-like 1 (NPC1-L1) receptor exposed on the

luminal surface. Only dietary animal cholesterol is kept in the cell to be secreted in the blood

via receptors. After being absorbed, vegetable cholesterol is thought it away in digestive tract

via ABCG5 and ABCG8 receptors. Mutations of those receptors can cause a specific disease.

It leads to secretion of vegetable cholesterol in the blood stream and increases the circulating

cholesterol level.

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Figure 4 - The revers cholesterol transport http://www.hindawi.com/journals/ppar/2009/501739/fig3/

In the intravascular phase, a plasma enzyme called lecithin-cholesterol

acyltranferase (LCAT) converts the free cholesterol into cholesterol ester, which is then

sequestered into the core of lipoprotein particle. HDL particles increase in size as they

circulate through the bloodstream and incorporate more cholesterol and phospholipid

molecules from cells and other lipoproteins, for example by the interaction with ABCG1

transporter. HDL engulfs cholesterol in excess in cells. For example, cholesterol from

macrophages, which are coming from athermanous plaque. In the direct pathway, HDL

transports cholesterol mostly to the liver or steroidogenic organs such as adrenals, ovary and

testes. It is removed by HDL receptors such as scavenger receptor BI (SRBI), which mediate

the selective uptake of cholesterol from HDL. Mature HDL gets back to nascent HDL and is

able to capture cholesterol again. Indirect pathway is probably the relevant one. It is mediated

by CETP. This protein exchanges triglycerides of IDL or VLDL against cholesterol-esters of

HDL. CETP decreases the concentration of HDL cholesterol and increases the concentrations

of CE in VLDL, IDL and LDL. As the result, IDLs are processed to LDL, which is removed

from the circulation by LDL receptor pathway (Griffin, 2013).

2.2.Regulation of uptake of LDL by cells

Low-density lipoproteins are the major carrier of cholesterol in the blood, accounting

for more than 60% of total plasma cholesterol. Hepatic and extrahepatic tissues through LDL

receptor-mediated endocytosis where it is internalized and degraded take up LDL to free

cholesterol and amino acids. Uptake of native and oxidized LDL by monocyte-derived

macrophages in the vessel wall results accumulation of cholesterol and formation of

atherosclerotic plaques.

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2.2.1.Hepatic uptake of LDL

This receptor is regulated by lots of genes, proteins and transcription factors and when

they are mutated, it leads to many diseases like hypercholesterolemia, diabetes,

atherosclerosis, etc. LDL receptors complexes are present in clathrin-coated pits on the cell

surface, which when bounds to LDL-cholesterol via adaptin, are pinched off to form clathrin-

coated vesicles inside the cell. The modular adaptor protein, autosomal recessive

hypercholesterolemia gene (ARH) is required for LDL binding and internalization. Lacking

autosomal recessive hypercholesterolemia gene (ARH-/-) has severe hypercholesterolemia

due to impaired hepatic clearance of LDL (Michaely, 2004). This allows LDL-cholesterol to

be endocytosis and prevents the LDL diffusing around the membrane surface. This occurs in

all nucleated cell but mainly in the liver, which removes 70% of LDL, form the circulation.

Once the coated vesicle is internalized it will shed its clathrin-coat and will fuse with an

acidic late endosome. The receptors are then either destroyed or they can be recycled via

endocytic cycle back to the surface of the cell ready to receive another LDL. The level of free

intracellular cholesterol regulates synthesis of receptor. If it is in excess for the need of the

cell then the transcription of the receptor gene will be inhibited

(http://ndtvinfo.gq/LDL_receptor ). The number of LDL receptor isn’t only regulated by the

blood cholesterol level but also by some molecules. Indeed, it proved that proprotein

convertase subtilisin/kexin type 9 (PCSK9) accelerates degradation of LDL receptors

(LDLR). PCSK9 is secreted from liver and binds a repeat domain in the extracellular portion

of LDL receptor. PCSK9 locks the receptor in an open conformation that targets it to

lysosomes for degradations. Some research study prove that heterozygote person for PCSK9

null mutations in 2.6% of African-Americans was founds to be associated with 28%

reduction in LDL cholesterol and 88% reduction in cardiovascular risk (Seidah, 2014).

LDL receptor (LDLR) can recognize differently apolipoproteins:

- ApoB100 (major pathway): VLDL, IDL, LDL can be endocytosed.

- ApoE (minor pathway): VLDL, IDL and chylomicrons can be endocytosed.

ApoB and microsomal triglyceride transfer protein (MTP) are necessary for lipoprotein

assembly. MTP's lipid transfer activity is required for the assembly of lipoproteins. This

activity renders nascent ApoB secretion-competent and may be involved in the import of

triglycerides into the lumen of endoplasmic reticulum. In addition, MTP binds to apoB with

high affinity involving ionic interactions. Some drugs have been developed in order to

prevent the assembly of apo B-containing lipoproteins; thus inhibiting the synthesis of

chylomicrons and VLDL and leading to decrease in plasma levels of LDL-C (Hussain, 2003).

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2.2.2.Extra-hepatic uptake of LDL

The liver produces bile. It is a melange of 97% water, 0.7% bile salt, 0.2% bilirubin,

0.51% acids (cholesterol, fatty acids). Bile acids allow fragmentation of large globules of

dietary lipids leading to the formation of micro-droplets. This emulsion then facilitates the

digestion of lipids by pancreatic lipase. Bile acid synthesis plays a critical role in the

maintenance of mammalian cholesterol homeostasis. There are two pathways, which lead to

the formation of bile acids: classical and alternative. Cholesterol 7-hydroxylase is the rate-

limiting enzyme in the synthesis of bile acid form cholesterol in the classical pathway. This

enzyme is the subject of feedback control. It is downregulated by sterol regulatory element-

binding protein (SREBP) when the level of plasma cholesterol is low. On the opposite, it is

upregulated by the nuclear receptor LXR (liver X receptor) when cholesterol levels are high.

The effect of this upregulation increases the production of bile acids and reduce the level of

cholesterol in hepatocytes (Chawla, 2000).

3.PATHOPHYSIOLOGY OF LDL METABOLISM

The blood concentration of LDL has been proven to be causally linked to the prevalence

of atherosclerosis (Huszar, 2000) and cardiovascular diseases. An increase of this low-density

lipoprotein can be caused by several mechanisms; in this article we will try to review those

mechanisms by citing the most famous pathologies described in the literature.

3.1.Increased synthesis

An increased synthesis originates from deregulations often due to acquired poly-factorial

causes, both genetic and environmental and it not well known until this day.

3.1.1.Metabolic syndrome

The first etiology of high plasma level of LDL is the rich diet in saturated fatty acids

and cholesterol. It is often responsible for hypercholesterolemia and thus an increase of LDL

plasma level. But this type of diet does not always explain the appearance of clinical and

biological symptoms and is often linked to many other factors.

3.1.2.Diabetes type 1 and 2

Type 1 and type 2 diabetes are described as being risk factors for CVD and

atherosclerosis by their influence on LDL metabolism. It is proven that patients with diabetes

can have higher concentration of LDL and smaller LDL particles (Garvey, 2003), both risk

factors.

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3.1.3.Hypothyroidism

Thyroid hormones have been shown to have multiple effects on lipid metabolism and

it’s been decades that hypothyroidism is known to induce hyperlipidemia. Approximately 1

to 11% of dyslipidemic patients have subclinical hypothyroidism (Pearce, 2012) even though

the effect of thyroid hormones depletion is not clear. An increase of thyroid - stimulating

hormone (TSH) would have an impact on the increased synthesis of ApoB and the higher

level of LDL cholesterol. T3 hormone may have an impact on sterol regulatory element

binding protein-2 (SERBP-2) activity hence; a lower level of this hormone would mean a

decrease of LDLR expression and thus a lower LDL clearance. Hypothyroidism could also

affect intestinal cholesterol absorption due to thyroid hormone actions on Niemann-Pick C1-

like 1 protein in the gut. Hypothyroidism seems also to have an effect on the size of LDL

particle as it induces an increase of small dense LDL (Abbas, 2008).

3.1.4.Hypercorticism

It has been shown that in hypercortisolism diseases like Crushing’s syndrome,

patients has metabolic syndrome associating high level of LDL, oxidized LDL and enhance

the inflammatory process leading to the development of cardiovascular disease (Boero,

2015).

3.1.5.Nephrotic syndrome

Abnormal lipid metabolism is often described in patients with renal alterations,

especially in nephrotic syndrome where we can observe high level of LDL as well as other

dyslipidemia. The appearance of mechanism is still unclear but nephrotic syndrome has been

shown to increase apolipoprotein levels like ApoB. It is believe that ApoB transcription is

induced by low oncotic pressure caused by the renal failure (Vega, 1995)

3.1.6.Infection

Some bacteria and virus has been proven to be responsible for coronary heart disease

by significantly increasing plasma level of cholesterol, triglycerides and LDL (Al-Ghamdi,

2011). It is interesting to see that LDL plasma level also have a consequence on the immune

system. In fact, there is a significant relation between the plasma cholesterol level and the

risk on infection as showed a 15 year cohort study (Iribarren, 1998).

3.1.7.Ethanol intake

It seems that the alcohol intake has an effect on LDL cholesterol level even though

the studies conducted on the subject shows variable results. Those different results are

believed to be linked to polymorphism in the population genome. Thus, studies on Italian and

Turkish subject showed that alcohol intake increases the circulating LDL whereas studies on

Japoneses brought opposite results (Brinton, 2012). Other studies showed that a low alcohol

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intake on monkey does not significantly change the amount of plasma LDL while a high

intake tends to elevate it.

3.1.8.Drug intake

It is known that some drugs increase the plasma level of LDL. It is the case for the

beta-blockers, such as atenolol, researchers has found an increase amount of LDL in the

patient receiving such treatment (Fonseca, 2010).

3.1.9.Menopause

Some studies show that LDL levels increase when women reach their menopause

(Corr, 2000) linked to more frequent CHD. It is believed to be due to the sudden low sex

hormone concentration, which were positively influencing biliary secretions.

3.2.Decrease uptake by cell

3.2.1.Familial hypercholesterolemia

The genetic mutations leading to hyperlipidemia and hypercholesterolemia can affect

a lot of different protein implicated in the LDL metabolism leading to variable diseases that

differ by their clinical and biological diagnosis.

The most well known and most described of all is the familial hypercholesterolemia (FH).

This disease is characterized by a mutation in the LDLR gene. Patients mutated in one of

these two alleles have heterozygous FH (heFH) while a patient presenting two mutated alleles

has homozygous FH (hoFH). This monogenic disease has the highest incidence,

approximately one out of 500 people (Golstein, 2001) in 2001 for heFH, and is one of the

most severe forms especially for those with hoFH. Its inheritance has an autosomal

codominant pattern. Patients with hoFH have a homogenous distribution of their high LDL

levels whereas heFH patients has different amount of plasma-LDL depending on their diet,

environment and genetic predisposition. hoFH have several clinical symptoms. They develop

cutaneous xanthomas and coronary atherosclerosis at a young age. Aortic root and coronary

ostia are often initially damaged by LDL accumulation in the endothelial lumen. If the young

patients are not taken care of quickly, they die prematurely of atherosclerotic cardiovascular

complications. The longer the exposition to high LDL level is, worsen the symptoms and the

prognostic are. Researchers found out that LDL levels in hoFH are inversely related to the

remaining LDLR activity. This is why “receptor-negative” patients with less than 2% of

normal LDLR activity rarely survive the second decade while “receptor-positive” patients

with 2 to 25% of normal LDLR activity develop clinically observable atherosclerosis disease

by the age of 30.

Patient presenting heFH have significantly lower and variable LDL levels and thus, much

more dependent on other factors making their prognosis difficult.

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Familial hypercholesterolemia caused by an abnormal LDLR can be caused by over 900

mutations of its gene. Some of them result in lowering the number of receptors on the cell’s

surface while others disrupt the receptor's ability to remove low-density lipoproteins from the

blood. The majority of them occurred in the ligand binding and epidermal growth factor

(EGF) precursor homology regions in the 5' region. The majority of its mutations are

missense, small insertion, and deletion. Approximately 10% to 15% of LDLR mutations are

large rearrangements, such as exonic deletions and duplications, which cannot be detected by

full-gene sequencing.

The diagnosis of FH is based on the familial history and the appearance of premature

coronary atherosclerosis, abnormal plasma lipidic composition and xanthomas. Because of

the high number of mutations, the molecular diagnosis is difficult except if the patient comes

from a population where a limited number of mutations predominate.

3.2.2.Familial defective ApoB-100

A less common disease has been described as similar to FH, with a reduced rate of

LDL catabolism but with a normal LDLR activity. Familial defective apoB-100 (FDB) is

characterized in most cases by a missense mutation (Arg3500Gln) in the LDLR-binding

domain of this apolipoprotein B-100. Approximately 60 different mutations has been found

affecting this gene. Its incidence can go up to 1 in 1,000 in Central Europe but is usually far

less common world wild. Like in FH, high LDL cholesterol levels, normal triglycerides,

xanthomas and early atherosclerosis characterize FDB. Even though the mean concentration

of LDL is higher in FDB patient than in controls, studies often show a wide standard

deviation. The appearance of atherosclerosis disease, like coronary disease, is slower but has

been proved to be more dangerous than FH’s mainly because, in FDB, LDL itself is

atherogenic. LDL produced from FDB homozygotes binds the LDLR with ~10% of normal

affinity and its clearance reach one-third to one-quarter of the normal value so that we find in

the bloodstream more particles carrying the deficient ApoB protein than particle bearing the

normal one.

The clinical diagnosis is very similar to heFH; patients presenting high level of plasma LDL

and xanthomas. The most common mutation upon wish ApoB-100 is mutated can be detected

using molecular techniques. It is important to notice that FDB is not always due to the single

base pair substitution and can occur by other ApoB mutations.

3.2.3.PCSK9 mutation

Mutations altering the PCSK9 protein are relatively new. Because PCSK9 inhibits

endocytic recycling of LDLR, its gain-of-function missense mutation is responsible for a

greater LDLR lysosomal degradation. Through this mechanism, the plasma LDL levels are

much higher and the risk of CVD is increased. As different studies showed, depending of the

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cell type, PCSK9 has variable effects on LDLR metabolism; in kidney cells it seems to

reduce LDLR levels whereas, in fibroblast, it does not affect LDLR expression (Benjannet,

2004). We identified three missense gain-of-function mutations in PCSK9 gene (Abifadel,

2003). Even though PCSK9 mutations is a very rare cause of hypercholesterolemia compared

to mutation of LDLR and ApoB, its study is not without interest. Indeed, by studying this

mutation, we are able to say that PCSK9 has a very important role in the LDL uptake by

cells. An overexpression of this protein, mutated or wild-type, enhances the LDLR

degradation by a post-transcriptional mechanism. On the other hand, three loss-of-function

mutations (in-frame deletion and missenses) have also been described (Cameron, 2006). A

large prospective study held for 15 years showed that those mutations reduced LDL levels by

28% and thus the frequency of CHD by 88% (Cohen, 2006). In addition with statins, the

results were even greater.

3.2.4.ARH mutation

ARH is an adaptor protein needed for a normal LDLR endocytosis. When ARH is

mutated, the number of LDLR is greatly increased on the plasma membrane and LDL

binding is also increased but its degradation is reduced. This is explained by understanding

ARH role: by being an adapter between the LDL receptor and the complex clathrin/AP2, a

loss-of-function mutation is responsible for an altered endocytosis of the ligand-receptor

complex. It’s interesting to note that the binding between the lipoprotein ApoB-100 and its

receptor is perfectly normal.

Once again, the accumulation of LDL in the blood vessels leads to increase risk of

atherosclerosis and CVD. The diagnosis of ARH deficiency cannot be distinguished from

heFH or hoFH, the LDL plasma levels being between those two levels. One difference than

can be found is in the onset of vascular alteration that can be clinically diagnosed; coronary

atherosclerosis happen later than hoFH patients. ARH patients can also have large and bulky

xanthomas.

3.2.5.Cholesterol 7-hydroxylase deficiency

The genetic deficiency of cholesterol 7α-hydroxylase is responsible for a lower

classical bile acid biosynthesis. This disease is extremely rare and leads to

hypertriglyceridemia and hypercholesterolemia. A study made by Pullinger et al. on three

siblings presenting CYP7A1 showed a bile acid synthesis reduced by 94% (Pullinger, 2002).

The hypercholesterolemia could be explained by the reduced hepatic LDLR activity due to

7α-hydroxylase deficiencies. The discoveries of this mutation are very important and suggest

that the anabolism of cholesterol into bile acid is essential for the homeostasis of plasma

cholesterol levels.

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3.3.Harmful LDL

3.3.1.Sitosterolemia

In normal individual, cholesterol represents more than 99% of the sterols. In

sitosterolemia, the plasma levels of a different form of sterol is elevated by more than 30 to

100-fold; sistosterol (Escolà-Gil, 2014). Affected individuals can have plasma LDL levels as

elevated as in hoFH, especially in childhood. They develop planar xanthomas, aortic stenosis

and premature CHD. A specific sitosterolemia feature is the appearance of a low-level

hemolysis caused by the presence of plant sterol in erythrocyte membranes. Those patients

have an increase fractional absorption of exogenous sterols and a defect in the acid bile

biosynthesis leading to the accumulation of both cholesterol and sistosterol in the circulation

and tissues. This disease is caused by mutations in two genes encoding ABC half-transporters

(ABCG5 and ABCG8) (Berge, 2000). The diagnosis is based on the xanthomatosis visible in

affected patients, and on the fact that they are hypercholesterolemia while their parents are

normocholesterolemic. The biochemical diagnosis can be made by extraction of lipids from

the plasma and gas-liquid chromatography of the sample.

3.3.2.Ox-LDL

Oxidized LDL (ox-LDL) is now a good biochemical marker of the oxidative stress.

Ox-LDL activates monocytes increasing the risk of LDL to infiltrate the blood walls.

Metabolic syndrome is a complex disorder associating multiples dysregulation. We know that

it provokes low-chronic inflammation and high oxidative stress added to the atherosclerosis

and CHD risks (Holvoet, 2008). In case of biliary retention, it has been proven that the

amount of ox-LDL is greatly increased (Comert, 2006). Inflammation and complement

activation is also a source of oxidation as immune complex made of LDL seems to generate

an oxidative stress leading to the production of ox-LDL (Salisubry, 2014)

3.3.3.Sd-LDL

The size of the LDL particle has also been linked to the increase of CHD risk. The

higher percentage of small dense LDL (sdLDL) is associated to the CHD events. High levels

of sdLDL have been correlated with inflammatory markers, metabolic syndrome and altered

lipid profiles (Brunzell, 2012; Kobayashi, 2015; Santamarina-Fojo, 2004).

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Figure 5 - Formation of sdLDL (Kobayashi, 2015)

In hypertriglyceridemic condition, the CETP and the hepatic lipase is activated leading to a

cholesterol-rich VLDL, thus a cholesterol-rich LDL. The HDL particle is transporting few

concentration of cholesterol and is full of triglycerides. Therefore, the LDL is more harmful

and the HDL hepatic cholesterol clearance is lowered.

3.3.4.Uremia

Studies showed that patients suffering from uremia presented a vicious cycle leading

to harmful LDL. The decreased catabolism of IDL and LDL characteristic of uremia leads to

their increased half-life in the plasma and further alteration of the ApoB protein. Those

modifications can be oxidation, carbamylation and glycation leading to the lower binding

LDL to its receptor, therefore the even greater reduction of their clearance. To compensate

this decreased liver clearance, there is an increase clearance of these modified particles by the

scavenger pathway. LDL particles are then taken up by macrophages via those scavenger

receptors turning them into foam cells in the endothelial wall and enhancing atherogenesis.

4.TREATMENTS

The numbers of special therapies against LDL abnormalities has increased spectacularly

in the last few decades. These treatments’ efficiency is based on its innovation and incidence.

Nowadays, against atherogenic dyslipidemia, doctors and researchers could distinguish

therapies between acquired and genetic diseases. In this part of the composition, I

demonstrate a full range of treatments about the LDL debate.

At first, there are genetic disorders in the LDL abnormal metabolism which including

familial hypercholesterinemia, single-gene defect such as hyperlipidemia, familial

hyperlipidemia. The familial hypercholesterinemia, as in the pathophysiology part it is

written, causes very high level of low-density lipoprotein cholesterol by the mutation of the

LDLR gene. It could be inherited in an autosomal dominant way and appear in a homozygous

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or a heterozygous form (Vogt, 2015). Then, ApoB defect as familial defective ApoB-100,

PCSK9 mutation and ARH mutation are also classified to congenital disorders.

A therapy for all of the genetic caused dyslipidemia is difficult to find, but a treatment, as

early as possible after the final diagnosis, is mostly needed. The best touchstone in this case is

the usage of statins. Statin is a HMG Co-A reductase inhibitor, which stops the assembly of

cholesterol; leads to lowering LDL-C level and reduce the motility rate of post myocardial

infarctions. It has a negative effect on increased synthesis of cholesterol then LDL. However,

like every drug, statins also have side effects. Those who have muscle complaints, these are

the most significant symptoms that stop the statin use (Moriarty, 2014). Statins are usually

expended for acquired and genetic diseases, too. Besides statins, there are other acceptable

medications, which are used simultaneously with these cholesterol synthesis inhibitors. Next,

we place emphasis on the importance of PCSK9 inhibitors. Pro-protein convertase

subtilisin/kexin type 9 (PCSK9) is a serine protease that has a big role in LDLR circulation.

By binding to this molecule, the PCSK9 enhance the receptor’s internalization in the

lysosome and its degradation. That process leads to a decreased LDL receptor’ recirculation

back to the cell membrane. With PCSK9 inhibitors, the latter mechanism is blocked and these

retardants inhibit the LDLRs degradation then increase the expression of the LDLRs on the

cell surface. This activity conducts reducing LDL-C level in plasma.

Figure 6 - Fig.1.Therapeutic mechanism of pro-protein convertase subtilisin/kexin type 9 inhibition. (Ahn, 2015)

The PCSK9 inhibitors are mostly monoclonal antibodies including Alirocumab,

Evolocumab, Bococizmumab, LGT-209, RG7652, and LY3015014. The other PCSK9-

directed agents are small interfering RNA (as ALN-PCSO2), antisense oligonucleotide

mimetic peptides and fusion protein using adnectin technology (as BMS-962476). All of

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these inhibitors are humanized with successful data (Ahn, 2015). These drugs are commonly

used against congenital reduced hepatic uptake.

Figure 7 - PCSK9-directed agents in development (Ahn, 2015)

For reducing LDL level and avoid cardiovascular diseases, there are other targeted

mechanisms that are mediated by MTP. MTP are emerged in enterocytes, hepatocytes and

they are needed in the synthesis of ApoB-containing lipoproteins. These proteins have also a

big part of transferation phospholipids, triglyceride, cholesterol esters to the ApoB and

synthesis of VLDL and chylomicron. By using MTP inhibitors, the TG level decreases due to

blocked secretions of VLDL and synthesis of ApoB. Furthermore, these synthetic obstacles

reduce dietary fat absorption by chylomicron and LDL-C concentration, too. These days, one

type of MTP, so-called lomitapide, was approved and successful for treating homozygous

familial hypercholesterinemia. Normally, ApoB plays a central role in assembling and

secreting ApoB-containing lipoproteins and its concentration indicates the amount of LDL in

plasma. Mipomersen is a 20 nucleotide antisense oligonucleotide that is used against ApoB

mRNA. These inhibitors establish a bond with ApoB mRNA and generate a substrate against

RNase H1, which cause lower level of ApoB protein and reducing concentration of LDL.

Before, the mipormesen is becoming a humanized and efficacious nucleotide structure, it

should undergo in two chemical modifications. First, it needs to be chemically transformed as

a psosphorothioated ester to survive the hydrolysis and degradation by enzymes. Second, it

should be modified with methoxyl sugar residues to gain a more stable version of

mipormersen (Ahn, 2015). The lomitapide and the mipomersen have also successful results

on treatments against congenital reduce hepatic uptake.

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Figure 8 - Therapeutic mechanism of lomitapide and mipomersen. (Ahn, 2015)

Finally, the last therapy method against arteriosclerosis cardiovascular disease

(ASCVD) is apolipoprotein A1 mimetics. However, these agents belong to HDL-C by

imitating the effect of ApoA1 which leads to cholesterol intake from macrophages causally

inverse progression of atherosclerosis (Ahn, 2015). It is also used against congenital

disorders.

As in the physiology part was mentioned, CETP has a big role in transferring cholesteryl

ester and triglyceride among the circulating lipoproteins, especially HDL, LDL, VLDL. The

CETP modulator (JTT-705) inhibits the CE and TG transport HDL between ApoB containing

lipoprotein in blood. It has a good effect on HDL level elevation because in this case the TG

and CE will deliver from HDL3 to HDL2 then to the liver. Thus, the LDL concentration will

decrease. The CETP inhibitor ‘s effect could be seen against harmful LDL mechanism.

Figure 9 - Fig.4. The effects of dalcetrapib on CETP-mediated cholesterol transport. (Shinkai, 2012)

In addition, we would emphasis other lipid-lowering agents. For instance, 10% reduction of

LDL and 20% increase of HDL due to Vitamin B3 that blocks the VLDL synthesis. Then,

fibrates could decrease the LDL level with 15% and the triglyceride level with 35% by

19

stimulating lipoprotein lipase, fatty acid oxidation, and triglycerides elimination. The 2-

Azetidiones blocks the sterol transporter at brush border and intestinal absorption of

cholesterol, this process reduces the LDL concentration by 15%. The bile acid sequestrates

also decrease the cholesterol and the LDL level. The last agent, omega-3, which inhibit acyl

Co-A:1,2diacylglicerol acyltransferase and lead to reduction of non HDL cholesterol, ratio of

total cholesterol to HDL-C, VLDL, ApoC, phospholipase A2, arachidonic acid and LDL

(http://emedicine.medscape.com/article/2172172-overview; Farinde, 2015). All of these

agents have a positive effect on patients who are suffering from acquired form of LDL

abnormalities.

These diseases are also presented in the palette of LDL’ abnormal attitude. It is including

metabolic syndrome, diabetes, hypothyroidism, hypercorticism, nephritic syndrome,

infection, ethanol intake, types of drugs and menopause. The aim of treatment in acquired

hypercholesterinemia is reducing the total and coronary vascular diseases. For a prosperous

cure, the ailments should be parried by a lifestyle modification. The therapy is involving three

pilots, these are „dietary changes, exercise, behavioral therapy” [Najam, 2015]. Patients’

menu should not consist saturated fats, trans fats, especially cholesterol and include

vegetables, fruits, omega-3 (eskimo-dietary), that increases the HDL concentration in plasma.

Taking exercises are also needed two or three times per week to accelerate the speed of LDL

metabolism. Last but not least, for a successful therapy, the patients’ strong mentality is

indispensable. With a strength of will, most of the cases the strong mentality could vanish the

diseases.

Finally, for the next year, these treatments will be more and more developed to stop this

abnormal mechanism and reduce the number of patients, which is everyone’s interest.

5.SUMMARY

To sum up, there are many LDL regulatory mechanisms, which can be disturb and lead to

several diseases. On one hand, some of those dysfunctions can be unavoidable. Receptors’

mutations, enzymes’ mutations, lead to an overproduction of cholesterol or a reducing LDL

regulation. As the consequences, the excess of LDL accumulates in arteries and possibly

causes atherosclerosis. This excess can also interferes with some organs’ physiological

processes and leads to severe dysfunctions. On the other hand, some diseases can be

avoidable. In order to prevent atherosclerosis, recommended levels of LDL and HDL are

dependent of the number of cardiovascular risk factors. Those rates can be respected by

supporting healthy lifestyles (reducing rich food in animal cholesterol and triglyceride, stop

smoking and do exercises) and following a good compliance of treatment.

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http://emedicine.medscape.com/article/2172172-overview

http://ndtvinfo.gq/LDL_receptor

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Table of content

Figure 1 - Raised BC: 2008http://www.who.int/gho/ncd/risk_factors/cholesterol_text/en/

4

Figure 2 - Table 1. Apoproteins of lipoproteins................................................................5

Figure 3 - Regulation of LDL receptor expression and cholesterol synthesis..................7

Figure 4 - The revers cholesterol transport.......................................................................8

Figure 5 - Formation of sdLDL (Kobayashi, 2015).......................................................17

Figure 6 - Fig.1.Therapeutic mechanism of pro-protein convertase subtilisin/kexin type 9

inhibition. (Ahn, 2015).............................................................................................20

Figure 7 - PCSK9-directed agents in development (Ahn, 2015)....................................21

Figure 8 - Therapeutic mechanism of lomitapide and mipomersen. (Ahn, 2015)..........22

Figure 9 - Fig.4. The effects of dalcetrapib on CETP-mediated cholesterol transport. (Shinkai,

2012).........................................................................................................................23

23