Increased VLDL in nephrotic patients results from a decreased catabolism while increased LDL results from increased synthesis

Abstract

Increased VLDL in nephrotic patients results from a decreased catabolism while increased LDL results from increased synthesis. Increased very low density lipoprotein (VLDL) in nephrotic patients results from a decreased catabolism while increased low density lipoprotein (LDL) results from increased synthesis. Hyperlipidemia is a hallmark of nephrotic syndrome that has been associated with increased risk for ischemic heart disease as well as a loss of renal function in these patients. The hyperlipidemia usually is characterized by increased cholesterol levels, although hypertriglyceridemia may be present as well. The factors that determine the phenotype of nephrotic dyslipidemia are not understood, nor has the primary stimulus for nephrotic hyperlipidemia been identified. One hypothesis is that nephrotic hyperlipidemia is the result of a coordinate increase in synthesis of proteins by the liver. To address these issues we simultaneously measured the in vivo rate of VLDL apolipoprotein B100 (apo B100) secretion, LDL apo B100 synthesis and albumin synthesis in patients with a nephrotic syndrome (N = 8) and compared them with a control group (N = 7) using a primed/continuous infusion of the stable isotope L-[1-13C] valine for six hours. Kinetic data were analyzed by multicompartmental analysis. Patients studied had combined hyperlipidemia as reflected by an significant increase in both VLDL and LDL apo B100 pool sizes. In contrast, the albumin pool size was significantly decreased. VLDL apo B100 levels were primarily increased as a consequence of a decrease in fractional catabolic rate (FCR) rather than from an increase in the absolute synthesis rate (ASR). Both VLDL apo B100 and triglycerides were inversely related to the fractional catabolism (FCR) of VLDL apo B100 (r2= 0.708; P = 0.0088) while neither had any relationship to the ASR of VLDL apo B100. In contrast to VLDL, increased LDL apo B100 was not a consequence of decreased catabolism. The LDL apo B100 ASR was significantly increased (P = 0.001) in the nephrotic patients compared to controls. Low density lipoprotein apo B100 ASR was greater than that of VLDL apo B100 in some patients, suggesting that LDL in these patients was not only derived from VLDL delipidation, but also by an alternative secretory pathway. There was no clear relationship between the ASR of VLDL apo B100 and the ASR of albumin within the current study population. Our data indicate that increased VLDL in nephrotic patients results from a decreased catabolism, while increased LDL results from increased synthesis. Increased VLDL in nephrotic patients results from a decreased catabolism while increased LDL results from increased synthesis. Increased very low density lipoprotein (VLDL) in nephrotic patients results from a decreased catabolism while increased low density lipoprotein (LDL) results from increased synthesis. Hyperlipidemia is a hallmark of nephrotic syndrome that has been associated with increased risk for ischemic heart disease as well as a loss of renal function in these patients. The hyperlipidemia usually is characterized by increased cholesterol levels, although hypertriglyceridemia may be present as well. The factors that determine the phenotype of nephrotic dyslipidemia are not understood, nor has the primary stimulus for nephrotic hyperlipidemia been identified. One hypothesis is that nephrotic hyperlipidemia is the result of a coordinate increase in synthesis of proteins by the liver. To address these issues we simultaneously measured the in vivo rate of VLDL apolipoprotein B100 (apo B100) secretion, LDL apo B100 synthesis and albumin synthesis in patients with a nephrotic syndrome (N = 8) and compared them with a control group (N = 7) using a primed/continuous infusion of the stable isotope L-[1-13C] valine for six hours. Kinetic data were analyzed by multicompartmental analysis. Patients studied had combined hyperlipidemia as reflected by an significant increase in both VLDL and LDL apo B100 pool sizes. In contrast, the albumin pool size was significantly decreased. VLDL apo B100 levels were primarily increased as a consequence of a decrease in fractional catabolic rate (FCR) rather than from an increase in the absolute synthesis rate (ASR). Both VLDL apo B100 and triglycerides were inversely related to the fractional catabolism (FCR) of VLDL apo B100 (r2= 0.708; P = 0.0088) while neither had any relationship to the ASR of VLDL apo B100. In contrast to VLDL, increased LDL apo B100 was not a consequence of decreased catabolism. The LDL apo B100 ASR was significantly increased (P = 0.001) in the nephrotic patients compared to controls. Low density lipoprotein apo B100 ASR was greater than that of VLDL apo B100 in some patients, suggesting that LDL in these patients was not only derived from VLDL delipidation, but also by an alternative secretory pathway. There was no clear relationship between the ASR of VLDL apo B100 and the ASR of albumin within the current study population. Our data indicate that increased VLDL in nephrotic patients results from a decreased catabolism, while increased LDL results from increased synthesis. apolipoprotein absolute synthesis rate cholesterol colloid oncotic pressure fractional catabolic rate fractional rate of synthesis high density lipoprotein intermediate density lipoprotein low density lipoprotein nephrotic syndrome triglyceride very low density lipoprotein Hyperlipidemia is one hallmark of the nephrotic syndrome (NS)1.Kaysen G.A. Hyperlipidemia of the nephrotic syndrome.Kidney Int. 1991; 31: S8-S15Google Scholar. Hyperlipidemia in the nephrotic syndrome has been proposed to result from increased synthesis and decreased catabolism of lipoproteins2.Warwick G.L. Packard C.J. Demant T. Bedford D.K. Boulton-Jones J.M. Shepherd J. Metabolism of apolipoprotein B-containing lipoproteins in subjects with nephrotic-range proteinuria.Kidney Int. 1991; 40: 129-138Abstract Full Text PDF PubMed Scopus (79) Google Scholar, 3.Vega G.L. Grundy S.M. Lovastatin therapy in nephrotic hyperlipidemia: Effects on lipoprotein metabolism.Kidney Int. 1988; 33: 1160-1168Abstract Full Text PDF PubMed Scopus (102) Google Scholar, 4.Warwick G.L. Caslake M.J. Boulton Jones J.M. Dagen M. Packard C.J. Shepherd J. Low-density lipoprotein metabolism in the nephrotic syndrome.Metabolism. 1990; 39: 187-192Abstract Full Text PDF PubMed Scopus (79) Google Scholar, 5.Aguilar Salinas C.A. Barrett P.H. Kelber J. Delmez J. Schonfeld G. Physiologic mechanisms of action of lovastatin in nephrotic syndrome.J Lipid Res. 1995; 36: 188-199PubMed Google Scholar, although the relative contribution of each has not been quantitated. The mechanism of hyperlipidemia is of clinical significance, since hyperlipidemia has been implicated in increased cardiovascular risk and in progression of renal injury in nephrotic patients6.Keane W.F. Mulcahy W.S. Kasiske B.L. Kim Y. O'DONNELL M.P. Hyperlipidemia and progressive renal disease.Kidney Int. 1991; 39: S41-S48Google Scholar, 7.Kasiske B.L. O'DONNELL M.P. Cowardin W. Keane W.F. Lipids and the kidney.Hypertension. 1990; 15: 443-450Crossref PubMed Scopus (72) Google Scholar, 8.Keane W.F. O'DONNELL M.P. Kasiske B.L. Schmitz P.G. Lipids and the progression of renal disease.J Am Soc Nephrol. 1990; 1: S69-S74Google Scholar. Although an increase in total plasma cholesterol is the most common abnormality, plasma triglyceride (TG) levels are also increased in patients with more severe proteinuria4.Warwick G.L. Caslake M.J. Boulton Jones J.M. Dagen M. Packard C.J. Shepherd J. Low-density lipoprotein metabolism in the nephrotic syndrome.Metabolism. 1990; 39: 187-192Abstract Full Text PDF PubMed Scopus (79) Google Scholar, 9.Ohta T. Matsuda I. Lipid and apolipoprotein levels in patients with nephrotic syndrome.Clin Chim Acta. 1981; 117: 133-143Crossref PubMed Scopus (54) Google Scholar, 10.Appel G.B. Blum C.B. Chien S. Kunis C.L. Appel A.S. The hyperlipidemia of the nepfrotic syndrome. Relation to plasma albumin concentration, oncotic pressure, and viscosity.N Engl J Med. 1985; 312: 1544-1548Crossref PubMed Scopus (195) Google Scholar, but this increase is not invariably present. It is not known why some patients with the nephrotic syndrome develop increased TG levels while others do not. While some investigators have reported an increased rate of very low density lipoprotein/low density lipoprotein (VLDL/LDL) secretion, others have not been able to confirm this finding. This could be due to differences in methodology or to differences in the patient population studied. Direct measurement of VLDL catabolism using 125I labeled lipoproteins is complicated by the fact that VLDL contains several apolipoproteins and more than one may be labeled. Of greater concern is that most studies employing iodination of proteins use oxidizing reagents that might modify lipids and may lead to different processing (such as scavenger receptor) so that they are processed differently than native lipoproteins. We avoided these potential artifacts by directly measuring synthesis and secretion of apolipoprotein B100 (apo B100) in nephrotic patients using stable isotopes. Although stable isotope measurements require extensive protein isolation procedures, complex multicompartmental modeling and specific equipment, no radiation is involved and this method has been proven to be a feasible approach to measure apo B metabolism5.Aguilar Salinas C.A. Barrett P.H. Kelber J. Delmez J. Schonfeld G. Physiologic mechanisms of action of lovastatin in nephrotic syndrome.J Lipid Res. 1995; 36: 188-199PubMed Google Scholar, 11.Parhofer K.G. Barrett P.H.R. Bier D.M. Schonfeld G. Determination of kinetic parameters of apolipoprotein B metabolism using amino acids labeled with stable isotopes.J Lipid Res. 1991; 32: 1311-1323PubMed Google Scholar, 12.Cohn J.S. Wagner D.A. Cohn S.D. Millar J.S. Schaefer E.J. Measurement of very low density and low density lipoprotein apolipoprotein (Apo) B-100 and high density lipoprotein Apo A-I production in human subjects using deuterated leucine. Effect of fasting and feeding.J Clin Invest. 1990; 85: 804-811Crossref PubMed Scopus (156) Google Scholar. Synthesis of a group of proteins, including albumin, is increased proportionally in the nephrotic syndrome in experimental animals13.Sun X. Kaysen G.A. Albumin and transferrin synthesis are increased in H4 cells by serum from analbuminemic or nephrotic rats.Kidney Int. 1994; 45: 1381-1387Abstract Full Text PDF PubMed Scopus (15) Google Scholar, 14.Lewandowski A.E. Liao W.S.L. Stinson-Fisher C.A. Kent J.D. Jefferson L.S. Effects of experimentally induced nephrosis on protein synthesis in rat liver.Am J Physiol. 1988; 254: C634-C642PubMed Google Scholar, 15.Kaysen G.A. Albumin metabolism in the nephrotic syndrome: the effect of dietary protein intake.Am J Kidney Dis. 1988; 12: 461-480Abstract Full Text PDF PubMed Scopus (35) Google Scholar and in humans16.Ballmer P.E. Weber B.K. Roy Chaudhurry P. Mcnurlan M.A. Watson H. Power D.A. Garlick P.J. Elevation of albumin synthesis rates in nephrotic patients measured with leucine.Kidney Int. 1992; 41: 132-138Abstract Full Text PDF PubMed Scopus (42) Google Scholar, 17.Kaysen G.A. Gambertoglio J. Jimenez I. Jones H. Hutchison F.N. Effect of dietary protein intake on albumin homeostasis in nephrotic patients.Kidney Int. 1986; 29: 572-577Abstract Full Text PDF PubMed Scopus (177) Google Scholar, 18.DE SAIN-VAN DER VELDEN, MGM, KAYSEN, GA, DE MEER, K, STELLAARD, F, VOORBIJ, HAM, REIJNGOUD, DJ, RABELINK, TJ, KOOMANS, HA: Proportionate increase of fibrinogen and albumin synthesis in nephrotic patients. Measurements with stable isotopes. Kidney Int (in press)Google Scholar, leading to the hypothesis by Marsh and Sparks19.Marsh J.B. Sparks C.E. Hepatic secretion of lipoproteins in the rat and the effect of experimental nephrosis.J Clin Invest. 1979; 64: 1229-1237Crossref PubMed Scopus (98) Google Scholar that hyperlipidemia might be a result of a coordinated increase in synthesis of albumin and other proteins by the liver, including lipoproteins. In addition, Davis et al20.Davis R.A. Engelhorn S.C. Weinstein D.B. Steinberg D. Very low density lipoprotein secretion by cultured rat hepatocytes. Inhibition by albumin and other macromolecules.J Biol Chem. 1980; 255: 2039-2045Abstract Full Text PDF PubMed Google Scholar suggested that low albumin states might exert an effect on hepatic synthesis of apo B100 by less direct means, specifically by increasing the availability of free fatty acids. In the present study, synthesis of apo B100 in VLDL and LDL was therefore measured directly and simultaneously with that of albumin synthesis in patients with nephrotic syndrome with a wide range of proteinuria, albumin and hyperlipidemia, using endogenous labeling with infused 13C valine as a precursor. Eight patients (6 males, 2 females) were recruited for the study from the renal division of the University Hospital Utrecht. The mean age was 48 ± 4 years (median 51; range 31 to 63 years). All had a stable nephrotic syndrome for at least three months duration. Six of them had membranous glomerulonephritis, one had steroid-resistant minimal change disease and one had focal glomerular sclerosis. Patients were prescribed a diet containing 0.8 g protein/kg body wt/day and 100 mmol sodium/day for at least two weeks before starting the infusion study. Besides diuretics, none of them received medication, or medication was stopped at least two weeks before the infusion study. Control studies were done in seven healthy subjects (3 males, 4 females) who were on a similar diet as the nephrotic syndrome patients. The mean age of the control group was 33 ± 2 years (median 33; range 25 to 40 years). One day before the infusion study, the subjects collected a 24-hour urine sample that was analyzed for urea, creatinine, protein and albumin. All patients and volunteers agreed to participate after signing an informed consent form, in accordance with the Helsinki Declaration of Human Rights. This study was approved by the Institutional Ethical Committee for studies in humans. L-[1-13C] valine (isotope mole fraction > 0.99; MassTrace, Woburn, MA, USA) was dissolved in sterile 0.9% saline and sterilized through a 0.22 μm filter. All chemicals were obtained from Riedel de Haën (Seelze, Germany) unless indicated otherwise. Density solutions were made with KBr in 0.9% NaCl, NaN3 (0.01%) and 1 mmol/liter EDTA. The infusion protocol has been described in detail previously18.DE SAIN-VAN DER VELDEN, MGM, KAYSEN, GA, DE MEER, K, STELLAARD, F, VOORBIJ, HAM, REIJNGOUD, DJ, RABELINK, TJ, KOOMANS, HA: Proportionate increase of fibrinogen and albumin synthesis in nephrotic patients. Measurements with stable isotopes. Kidney Int (in press)Google Scholar. In short, samples were taken to measure the Evans blue distribution volume, an index for plasma volume21.Freinkel N. Schreiner G.E. Athens J.W. Simultaneous distribution of T-1824 and I131-labelled human serum albumin in man.J Clin Invest. 1953; 32: 138-148Crossref PubMed Scopus (9) Google Scholar. At the baseline (time, t = 0), a priming dose of 15 μmol/kg L-[1-13C] valine was administered intravenously over two minutes, followed by a continuous infusion of 15 μmol/kg/hr L-[1-13C] valine during six hours. Blood samples (5 ml) were collected into heparin-containing tubes and into EDTA containing tubes (10 ml). Samples were taken from the contralateral arm at t = 0, 15, 30, 60, 120, 180, 240, 270, 300, 330 and 360 minutes. Samples were kept on ice (maximum 1 hr) until plasma was separated by centrifugation (20 min, 3000 rpm, 4°C). Plasma samples for isolation of albumin and lipoproteins were immediately stored at -80°C or used directly. The isolation of free amino acids from plasma and the isolation of albumin from heparin plasma, which was based on differential solubility in absolute ethanol from TCA-precipitated proteins, was performed as described in detail previously18.DE SAIN-VAN DER VELDEN, MGM, KAYSEN, GA, DE MEER, K, STELLAARD, F, VOORBIJ, HAM, REIJNGOUD, DJ, RABELINK, TJ, KOOMANS, HA: Proportionate increase of fibrinogen and albumin synthesis in nephrotic patients. Measurements with stable isotopes. Kidney Int (in press)Google Scholar. Lipoproteins were isolated from plasma by sequential ultracentrifugation. For the VLDL isolation, 3.0 ml EDTA plasma was over layered by 2.8 ml of a density solution 1.006 kg/liter and ultracentrifugated for 20 hours at 40,000 rpm in a 50.3 ti rotor (Beckman Instruments, Inc., Palo Alto, CA, USA). After tube slicing, the supernatant was washed by again adding 2.8 ml of the density solution 1.006 kg/liter to 3 ml supernatant. The VLDL fraction was obtained after centrifugation and tube slicing again. Low density lipoprotein was isolated as described elsewhere22.Pekelharing H.L.M. Kleinveld H.A. Duif P.F.C.C.M. Bouma B.N. Van Rijn H.J.M. Effect of lipoprotein(a) and LDL on plasminogen binding to extracellular matrix and on matrix-dependent plasminogen activation by tissue plasminogen activator.Thromb Haemost. 1996; 75: 497-502PubMed Google Scholar. In short, 2.5 ml infranatant of the first run was adjusted with KBr to a density of 1.24 kg/liter and overlayered by 5 ml of a density solution 1.12 kg/liter, 2.0 ml of a density solution 1.06 kg/liter and 2.5 ml of a density solution 1.006 kg/liter followed by ultracentrifugation using a swinging out bucket rotor (SW-40; Beckman). Density bands were aspirated in 0.5 ml fractions. Fraction 13 to 21 contained LDL that was free from intermediate density lipoprotein (IDL) and high density lipoprotein (HDL). Fractions with Lp(a) contents < 28 mg/liter (detection limit) were taken. Control experiments were performed and showed that enrichment in this fraction is a good representation of the total LDL pool. In two patients who had a detectable Lp(a) concentration in this fraction, the sample was pre-treated with lysine-sepharose to remove the Lp(a) (< 28 mg/liter). Apolipoprotein B100 was isolated from the VLDL and LDL fractions by precipitation with isopropanol as described by Egusa et al23.Egusa G. Brady D.W. Grundy S.M. Howard B.V. Isopropanol precipitation method for the determination of apolipoprotein B specific activity and plasma concentrations during metabolic studies of very low density lipoprotein and low density lipoprotein apolipoprotein B.J Lipid Res. 1983; 24: 1261-1267Abstract Full Text PDF PubMed Google Scholar. The precipitate was delipidated with ethanol/ether (3:1) and thereafter with ether. At each step of the extraction, the protein-solvent mixture was incubated at -20°C overnight and was centrifugated at 2000 rpm for 30 minutes. The solvent was removed by aspiration. After evaporation the remaining ether fraction, apo B100 was hydrolyzed with 6 N HCl for 24 hours at 110°C24.Hill R.L. Hydrolysis of proteins.Advantages of Prot Chem. 1965; 20: 37-107Crossref PubMed Scopus (264) Google Scholar. The hydrolysates were supplied to cation-exchange resin as described previously18.DE SAIN-VAN DER VELDEN, MGM, KAYSEN, GA, DE MEER, K, STELLAARD, F, VOORBIJ, HAM, REIJNGOUD, DJ, RABELINK, TJ, KOOMANS, HA: Proportionate increase of fibrinogen and albumin synthesis in nephrotic patients. Measurements with stable isotopes. Kidney Int (in press)Google Scholar. For quantification of lipids and lipoproteins, plasma samples were subjected to a single ultracentrifugation step according to Redgrave, Roberts and West25.Redgrave T.G. Roberts D.C.K. West C.E. Separation of plasma lipoproteins by density-gradient ultracentrifugation.Anal Biochem. 1975; 65: 42-49Crossref PubMed Scopus (870) Google Scholar. Plasma, VLDL, IDL and LDL fractions were assayed for total cholesterol and triglyceride on a Synchron CX4 (Beckman). The amount of HDL was measured by precipitating a small aliquot of the bottom fraction using dextran sulphate-Mg2+26.VÄISÄNEN S. GÄVERT J. Julkunen A. Voutilainen E. Penttila I. Contents of apolipoprotein A-I, A-II and B of the human serum fractions for high-density and low-density lipoproteins prepared by common precipitation methods.Scand J Clin Lab Invest. 1992; 52: 853-862Crossref PubMed Scopus (14) Google Scholar. Plasma was assayed for apo A1 and for apo B100 using a routine nephelometric assay (Behringwerke AG, Marburg, Germany). For the measurement of apo B100 in the VLDL and IDL fractions, a nephelometric assay was used in which the detection limit of the apo B100 assay was 12 mg/liter. Derivatization of the isolated amino acids and of the isolated apo B100 was done according to the method of Hušek27.Husek P. Rapid derivatization and gas chromatographic determination of amino acids.J Chromatogr. 1991; 552: 289-299Crossref Scopus (261) Google Scholar. The N(O,S)-methoxycarbonyl methyl ester derivatives of plasma free amino acids and of apo B100 from VLDL were analyzed by gas chromatography/mass spectrometry, and the derivatives of apo B100 LDL and of albumin were analyzed by gas chromatography combustion isotope ratio mass spectrometry as described previously18.DE SAIN-VAN DER VELDEN, MGM, KAYSEN, GA, DE MEER, K, STELLAARD, F, VOORBIJ, HAM, REIJNGOUD, DJ, RABELINK, TJ, KOOMANS, HA: Proportionate increase of fibrinogen and albumin synthesis in nephrotic patients. Measurements with stable isotopes. Kidney Int (in press)Google Scholar. The apo B100 valine tracer/tracee ratio in VLDL and LDL were analyzed by mathematical compartmental modeling Figure 1 using SAAM II software28.SAAM II User Guide. Seattle, University of WashingtonGoogle Scholar. The model consists of an plasma amino acid pool, a delay compartment that accounts for the time delay in appearance of VLDL apo B 100 into plasma, and a series of delipidation compartments representing VLDL, IDL and LDL pools. In this model k-values represent the fractional rate at which 13C valine appeared into various apo B100 lipoprotein pools. Simulations were done using the plasma valine tracer/tracee ratio as a forcing function to the model, which allowed an adjustment of the various k-value for an optimal fit to the VLDL apo B100 and LDL data simultaneously. The k-value (2,1) represents the fractional rate of synthesis (FSR) of apo B VLDL. The k-value (0,5) represents the fractional rate of catabolism (FCR) of LDL. Since the measurements were done while apo B100 metabolism was in steady state, the fractional clearance rate was equal to the fractional production rate (FCR = FSR). The tracer/tracee data of albumin were analyzed by fitting the data to a multicompartmental model consisting of a plasma amino acid pool a delay element and an albumin pool29.Foster D.M. Barrett P.H.R. Toffolo G. Beltz W.F. Cobelli C. Estimating the fractional synthetic rate of plasma apolipoproteins and lipids from stable isotope data.J Lipid Res. 1993; 34: 2193-2205PubMed Google Scholar. The absolute synthesis rate (ASR), which is the amount of protein synthesized per day, were calculated by multiplying pool sizes by the corresponding FSR. Pool sizes were calculated by multiplying the plasma concentrations of albumin, VLDL apo B100 and LDL apo B100 with the plasma volume measured by Evans blue. To adjust for different body wts, these quantities are expressed per kg body wt. Statistics were performed using t-test. If the normality test or equal variance test failed a Mann-Whitney Rank sum test was used. All data are presented as means ±SEM Correlations were performed by linear regression analysis. As expected, plasma albumin concentration and plasma colloid oncotic pressure (COP) were significant decreased compared to the control group Table 1. Creatinine clearance was also decreased significantly. Urine urea excretion, as index of protein intake, was not significantly different from that in the control subjects. Total triglyceride (TG), VLDL-TG, IDL-TG, total cholesterol (chol), VLDL-chol, IDL-chol and LDL-chol were significantly elevated in the patients compared to controls Table 2. The plasma apo B 100 pool was significantly (P < 0.001) increased in the nephrotic patients (2.39 ± 0.29 g/liter) compared to the control group (1.03 ± 0.09 g/liter), while HDL-chol Table 2 and its apolipoprotein A1 (1.45 ± 0.15 g/liter vs. 1.26 ± 0.08 g/liter) were not significantly different between nephrotic patients and controls. Cholesterol levels were increased above the normal range in all patients and ranged from 7.3 to 15.9 mmol/liter. In contrast, TG levels ranged from 1.0 to 7.7 mmol/liter. Very low density lipoprotein apo B100 pool size was increased above the maximum value found in control subjects in six nephrotic patients, but was within the normal range in two (No. 7 and 8). Pool sizes of LDL apo B100 were significantly increased while the pool sizes of albumin were significantly decreased Table 3 compared to normal subjects.Table 1Clinical characteristics of the patients and control subjectsTable 2Levels of plasma lipids, lipoproteins and apolipoproteins A1 and B100 of both groupsTable 2Levels of plasma lipids, lipoproteins and apolipoproteins A1 and B100 of both groupsTable 3Kinetics of apo B100 and albuminTable 3Kinetics of apo B100 and albumin Enrichment in plasma free valine reached a plateau after 60 minutes and remained stable during the study, for both patients and controls (data not shown). The mean coefficient of variation of plateau was 1.5%. The fraction rate of appearance of 13C valine in VLDL apo B100 (and thereby the FSR) tended to be lower in patients than controls (data not shown). Since in steady state FSR is equal to fractional catabolic rate (FCR), the FCR of VLDL apo B100 tended to be lower in the entire patient group, although this value did not reach significance (Table 3 and Figure 2a). Within the group of six patients with elevated levels of VLDL apo B100, the FCR was significantly decreased (13.6 ± 3.6%/hr in nephrotic vs. 26.5 ± 3.2%/hr in controls, P = 0.02). Since the VLDL apo B100 pool size was increased more than fivefold in nephrotic patients, the absolute rate of 13C valine incorporation was greater in the nephrotic group compared to controls (Table 3 and Figure 2b). In four nephrotic patients (No. 2, 3, 5 and 6), the calculated ASR of VLDL apo B100 was above the highest value found in the control group, while the remaining patients (No. 1, 4, 7 and 8) had a calculated ASR of VLDL apo B100 similar to those found in the control group Figure 4b. In the patient group, plasma VLDL apo B100 concentration correlated inversely with VLDL apo B 100 FCR (Figure 3a; r2= 0.708; P = 0.0088) and bore no relationship to VLDL apo B 100 ASR (Figure 3b; r2= 0.245; P = 0.21), suggesting that the expanded VLDL apo B 100 pool size was a consequence of a decrease in FCR, rather than an increase in ASR. Furthermore, the FCR of VLDL apo B100 correlated with plasma VLDL-chol (r2= 0.716; P = 0.0081), plasma VLDL-TG (r2= 0.784; P = 0.019) and total plasma triglyceride (r2= 0.776; P = 0.0038). Those nephrotic patients whose triglycerides were increased all had a decreased FCR of VLDL apo B100, while those whose triglycerides were within the normal range all had a FCR of VLDL apo B100 that was not reduced. The VLDL apo B100 concentration did not correlate with either albumin concentration or urinary albumin secretion (r2= 0.060; P = 0.559 and r2= 0.071; P = 0.522, respectively).Figure 4Box plots showing the fractional catabolic rate (FCR %/hr; A) and the absolute synthesis rate (ASR) of low density lipoprotein (LDL) apolipoprotein (apo) B100 (mg/kg/day; B) in both patients and controls. Shown are the mean (black line), median (dotted line) and range (vertical bars) (*P = 0.001).View Large Image Figure ViewerDownload (PPT)Figure 3Relationship between plasma very low density lipoprotein (VLDL) apolipoprotein (apo) B100 (mg/liter) and the fractional catabolic rate (FCR) of apo B100 (%/hr) in nephrotic patients (A; r2= 0.708, P = 0.0088). The 95% confidence limits of the whole group are shown on either side of the regression line (N = 8). Relationship between plasma VLDL apo B100 (mg/liter) and the absolute synthesis rate (ASR) of VLDL apo B100 (mg/kg/day) for nephrotic patients (B; r2= 0.25, P = 0.21; N = 8).View Large Image Figure ViewerDownload (PPT) Low density lipoprotein apolipoprotein B100 concentration and pool size was increased more than twofold in the patient group compared to controls. In contrast to VLDL apo B100, the ASR of LDL apo B100 was significantly (P < 0.001) increased from a mean of 6.0 ± 0.8 mg/kg/day in the control subjects to 28 ± 7 mg/kg/day in 7 nephrotic patients Figure 4b. The FCR of LDL could not be fitted by the model that we used in one patient (No. 4). This finding caused us to consider the possibility of a different pathway for synthesis of LDL from the usually accepted VLDL-IDL-LDL route in this patient. The increase in LDL apo B100 synthesis was responsible for the increase in LDL apo B100 concentration and pool size, since the FCR of LDL apo B100 actually tended to be higher in patients than controls, although this difference did not achieve significance, again due to the wide range in the individual values (Table 3 and Figure 4a). The absolute rate of LDL apo B100 synthesis was greater than the rate of synthesis of VLDL apo B100 in three of the nephrotic patients Table 3, suggesting that the relationship between VLDL and LDL synthesis is not a direct precursor product relationship, but instead that at least a component of the LDL pool arises either directly or by a pathway that does not include VLDL. In the patient group there was no relationship between the ASR of albumin and that of apo B100 in LDL or between plasma albumin concentration and the ASR of LDL apo B100 (r2= 0.017; P = 0.779 and r2= 0.026; P = 0.728, respectively). In contrast to VLDL apo B100, incorpo