**2.1 Changes of routinely measured lipid parameters in CRI and HD**

The changes of two basic lipid parameters – the concentration of triglycerides (TG) and total cholesterol (TC) in different forms of renal disease are in Table 3. Increased triglyceride concentration is a general feature of kidney disease whereas increased cholesterol is not. Assesment the risk of atherosclerosis related morbidity from these two parameters is of course not possible because they do not reflect the real metabolic situation sufficiently. Lipids are insoluble in blood plasma and therefore they can circulate only in the form of lipoproteins. On the other side increased TG alone is an important warning sign of the presence of highly

Lipid and Lipoprotein Abnormalities in Chronic Renal Insufficiency: Review 353

> 50 < 0,940 80 – 95 2-7 3-9

1,125 – 1,210 5 – 10 15 – 25 20 – 30

**TG CH PL** 

**Lipoprotein Size, nm Density, g/l Lipid content, %** 

0,950 – 1,005 1,000 – 1,019 1,019 – 1,063

1,063 – 1,125

Apoprotein, function **Gene Finding**  A I – activator of LCAT 11q23-qter HDL, CHY

Inhibitor of LCAT 11q21-q23 HDL, CHY A IV – activator of LCAT 11q23-qter HDL, CHY

B 48 - short form of B 100 The same as B100 CHY C I –Cofactor of LCAT 19q12-q13.2 CHY, VLDL C II – activator of lipoprotein lipase 19q12-q13.2 CHY, VLDL, HDL C III – inhibitor of VLDL transfer into liver 11q23-qter CHY, VLDL, IDL D – regulator of cholesterol ester transfer 3q14.2-qter HDL E – Transfer of CHY remnants into liver 19q12-q13.2 CHY, VLDL, IDL

After a meal rich in fat the peak value of chylomicrons occurs approximately in 3 hours and they disappear in 8 – 9 hours. Visible plasma turbidity is a clear sign of chylomicron presence (mostly caused when blood sampling is not realized in fasting state). Chylomicrons are not involved singificantly in atherogenesis but in the case of their decreased catabolism (which is a case in kidney disease) the accumulated remnants can be

Endogenous lipid carrier particles are formed in the liver as VLDL. They contain a bigger variant of apoB (apoB100) but the function of this protein is the same as in chylomicrons. Apoproteins B are of key importance during the assembly of lipoprotein particles and also serve as a docking structure when the lipoprotein binds to the LDL-receptor. The interaction of VLDL with HDL and with the tissues is similar to that of chylomicrons and during their circulation they became smaller – known as IDL and LDL. LDL particles during their relatively long life span (t1/2 = 2 – 4 hours and in CRI probably even more) can undergo oxidation, glycation and carbamylation. These random postsynthetic events alter the quality of liporoteins with a profound effect on their cellular metabolism – e.g. the uptake of damaged LDL particles through the scavenger receptor and not through the LDL-receptor. Another important topic important for the rate of atherosclerosis (not only) in CRI patients

binding 2p23-24 LDL, IDL, VLDL

Table 5. Basic characteristics of lipoproteins (According to Žák, 2002, modified; PL =

80 – 500

9 – 12 5 - 9

A II – activator of liver lipase,

B 100 - key protein for assembly and receptor

Table 6. Apoprotein function and their occurence in lipoproteins

**Chylomicrons Chylomicron remnants** 

> **VLDL IDL LDL**

**HDL2 HDL3**

atherogenic (Chan et al, 2009).

phospholipids)

atherogenic triglyceride rich lipoprotein particles (TRL). The concentration of total cholesterol on the other side does not provide a valuable information because its carriers can be less and more atherogenic or even antiatherogenic. In table 4 data on changes of HDL-cholesterol, non-HDL-cholesterol and the highly atherogenhic Lp(a) in CRI are shown. LDL-cholesterol is not involved in this table for reasons of its analytical uncertainity which is further analyzed in part 3 of this chapter. From data in Table 3 and 4 the shift towards accelerated atherosclerosis is evident – high amount of triglycerides in the lipoproteins and a low level of antiatherogenic HDL particles regardless on the form and stage of kidney disease (Vazari, 2006, Saland et Ginsberg, 2007, Tsimihodimos et al., 2008, Attman et al., 2010).


Table 3. Triglycerides and total cholesterol in different conditions associated with kidney disease


Table 4. Non-HDL cholesterol, HDL cholesterol and lipoprotein(a) in chronic renal insufficiency, hemodialysis and peritoneal dialysis

### **2.2 Lipoprotein metabolism in health and kidney disease**

Lipoproteins are complex particles consisting from lipids transferred from and to the tissues and a phospholipid envelope containing different proteins. They are not passive containers of TG and cholesterol but dynamic particles with variable composition. Both lipoprotein turnover and composition is altered in kidney diseases and therefore basic knowledge about it is essential for the proper assessment of cardiovascular risk in this special case. Lipoprotens are divided int three families: Chylomicrons and their remnants, the VLDL-IDL-LDL family and the HDL family. Their composition and apoprotein content is in Tables 5 and 6.

Chylomicrons produced in enterocytes are carriers of lipids from guts to liver. There are much bigger than the other lipoproteins and during ultracentrifugation they do not sediment at all. Chylomicrons contain one molecule of apoprotein is B48 and some apoprotein molecules A I, A II and A IV. During their life span they acquire apo E and apo C from HDL particles. In contact with muscle and fat tissue capillary endothelial cells they release fatty acids through triglyceride lipolysis catalyzed by endothelium-bound lipoprotein lipase. Remnants of chylomicrons are removed by the liver.

atherogenic triglyceride rich lipoprotein particles (TRL). The concentration of total cholesterol on the other side does not provide a valuable information because its carriers can be less and more atherogenic or even antiatherogenic. In table 4 data on changes of HDL-cholesterol, non-HDL-cholesterol and the highly atherogenhic Lp(a) in CRI are shown. LDL-cholesterol is not involved in this table for reasons of its analytical uncertainity which is further analyzed in part 3 of this chapter. From data in Table 3 and 4 the shift towards accelerated atherosclerosis is evident – high amount of triglycerides in the lipoproteins and a low level of antiatherogenic HDL particles regardless on the form and stage of kidney disease (Vazari, 2006, Saland et

**Condition Triglycerides Total cholesterol**  Minimal proteinuria Increased No change or decrease Heavy proteinuria Increased Highly increased in

Hemodialysis Increased Controversial data – mostly

Chronic renal insufficiency Increased No change

Peritoneal dialysis Increased Increased Table 3. Triglycerides and total cholesterol in different conditions associated with kidney

**Condition Non-HDL cholesterol HDL-cholesterol Lp(a)**

insufficiency Increased Decreased Increased Hemodialysis Increased Decreased Increased Peritoneal dialysis Increased Decreased Increased

Lipoproteins are complex particles consisting from lipids transferred from and to the tissues and a phospholipid envelope containing different proteins. They are not passive containers of TG and cholesterol but dynamic particles with variable composition. Both lipoprotein turnover and composition is altered in kidney diseases and therefore basic knowledge about it is essential for the proper assessment of cardiovascular risk in this special case. Lipoprotens are divided int three families: Chylomicrons and their remnants, the VLDL-IDL-LDL family and the HDL family. Their composition and apoprotein content is in Tables

Chylomicrons produced in enterocytes are carriers of lipids from guts to liver. There are much bigger than the other lipoproteins and during ultracentrifugation they do not sediment at all. Chylomicrons contain one molecule of apoprotein is B48 and some apoprotein molecules A I, A II and A IV. During their life span they acquire apo E and apo C from HDL particles. In contact with muscle and fat tissue capillary endothelial cells they release fatty acids through triglyceride lipolysis catalyzed by endothelium-bound

lipoprotein lipase. Remnants of chylomicrons are removed by the liver.

Table 4. Non-HDL cholesterol, HDL cholesterol and lipoprotein(a) in chronic renal

insufficiency, hemodialysis and peritoneal dialysis

**2.2 Lipoprotein metabolism in health and kidney disease** 

nephrotic syndrome

no change

Ginsberg, 2007, Tsimihodimos et al., 2008, Attman et al., 2010).

disease

5 and 6.

Chronic renal


Table 5. Basic characteristics of lipoproteins (According to Žák, 2002, modified; PL = phospholipids)


Table 6. Apoprotein function and their occurence in lipoproteins

After a meal rich in fat the peak value of chylomicrons occurs approximately in 3 hours and they disappear in 8 – 9 hours. Visible plasma turbidity is a clear sign of chylomicron presence (mostly caused when blood sampling is not realized in fasting state). Chylomicrons are not involved singificantly in atherogenesis but in the case of their decreased catabolism (which is a case in kidney disease) the accumulated remnants can be atherogenic (Chan et al, 2009).

Endogenous lipid carrier particles are formed in the liver as VLDL. They contain a bigger variant of apoB (apoB100) but the function of this protein is the same as in chylomicrons. Apoproteins B are of key importance during the assembly of lipoprotein particles and also serve as a docking structure when the lipoprotein binds to the LDL-receptor. The interaction of VLDL with HDL and with the tissues is similar to that of chylomicrons and during their circulation they became smaller – known as IDL and LDL. LDL particles during their relatively long life span (t1/2 = 2 – 4 hours and in CRI probably even more) can undergo oxidation, glycation and carbamylation. These random postsynthetic events alter the quality of liporoteins with a profound effect on their cellular metabolism – e.g. the uptake of damaged LDL particles through the scavenger receptor and not through the LDL-receptor. Another important topic important for the rate of atherosclerosis (not only) in CRI patients

Lipid and Lipoprotein Abnormalities in Chronic Renal Insufficiency: Review 355

Tiacylglycerols and cholesterol are insoluble in water. They circulate in the blood

Lipoproteins are complex particles; their cholesterol content (measured or calculated as

Lipoproteins are dynamic, changing their lipid and protein composition during their life

Lipoproteins are heterogeneous and their different density is not a simple biological variation but it has profound effect on their metabolism. Small dense lipoproteins are

In some people and in some pathological conditions there are also abnormal lipoproteins in the blood. One of them is lipoprotein(a) which is highly aterogenic and its

Some of the assay methods for lipids and lipoproteins are not bulletproof from analytical

**3. Methods for measurement and lipids, lipoproteins with special attention to** 

Triglyceride and total cholesterol assays are the starting points of lipid status assessment also today but their information value is different. Strictly speaking both are "artefacts" but whereas total triglycerides provide an important information about the presence of TRLs,

Lipoproteins are prone to differential precipitation in arteficial conditions (e.g. heparinmanganese or dextran-magnesium solutions) and this allowed to develop methods to separate them without ultracentrifugation or electrophoresis. Based on this procedure in 70's first the direct measurement of HDL-C was developed and introduced into clinical chemistry. On the other side direct measurement of LDL-C was not possible until the end of the century. In the 25 year long interim period there was only one possibility to estimate the

Table 9. Pitfalls in interpretation of basic lipid parameters from clinical point of view

**3.1 Basic lipid assays and the short history of lipid and lipoprotein measurement**  The first cumbersome but at that time reliable colorimetric methods for cholesterol and triglyceride assays were developed in the first half of the last century. From today's point of view obsolete methods were crucial for understanding the association between lipid metabolism and cardiovascular disease. Later there were replaced with enzymatic assays and adjusted for the use in automatic analyzers. Isolation of lipoproteins by ultracentrifugation giving basis for the today lipoprotein terminology was introduced in the 40's of the last century. Analytical and preparative ultracentrifugation is also today the gold standard method for quantification, separation and research on lipoproteins. About the same time paper electrophoresis (later replaced by agarose and polyacrylamide gel) gave the rise not only to an alternative nomenclature (alpha, pre-beta and beta lipoproteins) but also to the epoch-making phenotypic classification of dyslipoproteinemias by Fredrickson & Lees in 1965 and a better understanding of the pathobiochemistry of atherosclerosis. This methodology is currently reserved only for specialized applications. (For an excellent review

HDL-C and LDL-C) does not reflect their real composition and structure.

exclusively as lipoproteins.

span in the circulation.

more atherogenic than those with low density.

concentration is increased in renal disease.

point of view – see part 3 of this chapter

**LDL-cholesterol measurement** 

on this topic see Mcnamara et al., 2006).

**3.2 Estimation of LDL-cholesterol** 

total cholesterol itself has a limited diagnostic value.

is the presence of small dense LDL particles and (Table 7) and increased levels of abnormal particles as Lp(a) – Tables 4 and 8. All these features render the interpretation of basic laboratory parameters of lipid/lipoprotein metabolism in regard to the assessment of atherosclerosis difficult (Table 9).

Despite the complicated and sometimes contradictory data on LDL metabolism in CRI, ESDR and HD it is possible to summarize the underlying picture in a relatively simple way. According to Ikerwaka et al (2005) and Chan et al (2009) all lipid and lipoprotein alterations associated with LDL particles have a common soil – their decreased catabolism. Increased synthesis can play an additonal role in the case of massive proteinuria.


Table 7. Normal density distribution of LDL particles


Table 8. Structure of Lp(a)

The basic function of HDL is the reverse transport of surplus cholesterol from the tissues to liver. HDL particles are synthetized as discoid particles poor in lipids. Nascent HDL (or HDL3) particles contain a lot of different apoproteins (70 % of total protein is Apo A-I and 20 % Apo A-II) crucial in TG and cholesterol metabolism which they exchange with chylomicrons and VLDL. Cholesterol bound to the surface of HDL is esterified by the enzyme LCAT and the esters are internalized. The cholesterol-ester loaded HDL2 particles unload their content through a specific receptor into the liver. The low number and decreased function of LDL particles (manifest as "low HDL-C") is probably a consequence of the enzyme LCAT and the apoprotein A-I. Recently Rosenson et al (2011) highlighted the topic of HDL heterogeneity and its possible role in the pathophysiology of accelerated atherosclerosis in kidney disease. According to this concept not the low level of "good cholesterol" but rather the altered constellation of HDL structure and function is of importance in atherogenesis.

is the presence of small dense LDL particles and (Table 7) and increased levels of abnormal particles as Lp(a) – Tables 4 and 8. All these features render the interpretation of basic laboratory parameters of lipid/lipoprotein metabolism in regard to the assessment of

Despite the complicated and sometimes contradictory data on LDL metabolism in CRI, ESDR and HD it is possible to summarize the underlying picture in a relatively simple way. According to Ikerwaka et al (2005) and Chan et al (2009) all lipid and lipoprotein alterations associated with LDL particles have a common soil – their decreased catabolism. Increased

> **Class Density, g/ml Size, nm % of LDL**  I 1.026 27.5 – 26.0 3 II 1.028 26.0 – 25.5 16 III 1.034 25.5 – 24.2 50 IV 1.048 24.2 – 21.8 22 V > 1.048 < 21.8 9

synthesis can play an additonal role in the case of massive proteinuria.

Lipoprotein Lp(a) has a similar structure and composition as LDL particles

Proteins: ApoB 100 and an abnormal protein, Apo(a) attached to the B 100 Apo(a) has a variable molecular weight between 300 – 800 kDa, its structure is similar to the plasminogen (involved in fibrinolysis).

Concentration: 0 – 1200 mg/l with abnormal distribution of values. As atherogenic are

Lp(a) in CRI: Increased but the cause of increase is not clear. The basic level of Apo(a) is probably associated with gene polymorphisms and the kidney disease is a provocative factor for further increase (Danesh et al, 2000)

The basic function of HDL is the reverse transport of surplus cholesterol from the tissues to liver. HDL particles are synthetized as discoid particles poor in lipids. Nascent HDL (or HDL3) particles contain a lot of different apoproteins (70 % of total protein is Apo A-I and 20 % Apo A-II) crucial in TG and cholesterol metabolism which they exchange with chylomicrons and VLDL. Cholesterol bound to the surface of HDL is esterified by the enzyme LCAT and the esters are internalized. The cholesterol-ester loaded HDL2 particles unload their content through a specific receptor into the liver. The low number and decreased function of LDL particles (manifest as "low HDL-C") is probably a consequence of the enzyme LCAT and the apoprotein A-I. Recently Rosenson et al (2011) highlighted the topic of HDL heterogeneity and its possible role in the pathophysiology of accelerated atherosclerosis in kidney disease. According to this concept not the low level of "good cholesterol" but rather the altered constellation of HDL structure and function is of

Table 7. Normal density distribution of LDL particles

Subtypes: 6 different forms – F, B and S1 – S4

considered concentrations above 200 – 300 mg/l

atherosclerosis difficult (Table 9).

Density: 1,050 – 1,100 Size: 25 – 35 nm

Table 8. Structure of Lp(a)

importance in atherogenesis.

Tiacylglycerols and cholesterol are insoluble in water. They circulate in the blood exclusively as lipoproteins.

Lipoproteins are complex particles; their cholesterol content (measured or calculated as HDL-C and LDL-C) does not reflect their real composition and structure.

Lipoproteins are dynamic, changing their lipid and protein composition during their life span in the circulation.

Lipoproteins are heterogeneous and their different density is not a simple biological variation but it has profound effect on their metabolism. Small dense lipoproteins are more atherogenic than those with low density.

In some people and in some pathological conditions there are also abnormal lipoproteins in the blood. One of them is lipoprotein(a) which is highly aterogenic and its concentration is increased in renal disease.

Some of the assay methods for lipids and lipoproteins are not bulletproof from analytical point of view – see part 3 of this chapter

Table 9. Pitfalls in interpretation of basic lipid parameters from clinical point of view
