lipoprotein-X Research Articles

Lipoprotein-X reduces LDL atherogenicity in primary biliary cirrhosis by preventing LDL oxidation

Hypercholesterolemic human LDL contains oxidized subfractions that have atherogenic properties. Paradoxically, atherosclerosis incidence is low in patients with primary biliary cirrhosis (PBC), a disease characterized by marked increases in plasma LDL, including the LDL subfraction lipoprotein-X (Lp-X). To investigate the mechanisms underlying this paradox, we first examined the propensity to oxidation of unfractionated LDL isolated from PBC patients. After prolonged incubation with copper, PBC-LDL failed to increase the oxidation index or electrophoretic mobility noted in control LDL. An admixture of PBC-LDL or Lp-X with control LDL prevented oxidation of the latter in a dose-dependent manner. PBC-LDL was also noncompetitive against copper-oxidized LDL (oxLDL) for binding with a murine monoclonal anti-oxLDL antibody in a competitive ELISA. OxLDL exerts its proapoptotic and antiangiogenic effects in part by inhibiting fibroblast growth factor 2 (FGF2) expression. Preincubation of oxLDL with PBC-LDL, but not control LDL, attenuated the inhibitory effects of oxLDL on FGF2 expression in cultured bovine aortic endothelial cells (ECs). The antioxidant and prosurvival properties of PBC-LDL diminished after the patients underwent orthotopic liver transplantation. These results suggest that Lp-X reduces LDL atherogenicity by preventing LDL oxidation to protect EC integrity in the presence of hypercholesterolemia. They also suggest that altering LDL composition may be as important as reducing LDL concentration in preventing or treating atherosclerosis.

A Novel in Vivo Lecithin-Cholesterol Acyltransferase (LCAT)-Deficient Mouse Expressing Predominantly LpX Is Associated with Spontaneous Glomerulopathy

Complete lecithin cholesterol acyltransferase (LCAT) deficiency is a rare genetic cause of extreme reduction in high density lipoproteins and there is a high prevalence of chronic renal dysfunction that may progress to renal failure. Previous in vitro studies suggest the vesicular lipoprotein X (LpX) particles commonly seen in LCAT-deficient plasmas may be causative. To test this hypothesis, we have generated a novel murine model that selectively accumulate LpX in the circulation by cross breeding the sterol regulatory element binding protein (SREBP) 1a transgenic mice (S+) with the LCAT knockout (lcat-/-) mice. Fast protein liquid chromatography fractionation of pooled plasma lipids revealed that virtually all cholesterol is concentrated in the very low density lipoprotein (VLDL)-sized fractions. These fractions are enriched in free cholesterol and phospholipid but extremely poor in triglyceride. Electron microscopy of the d <1.063 g/ml fraction of the S+lcat-/- mice revealed abnormal large vesicular particles, suggestive of LpX. The S+lcat-/- mice developed glomerular lesions spontaneously evident at 6 months with glomerular and tubulointerstitial lipid-deposits. Immunohistochemical staining with RhoA showed marked positive focal staining in glomeruli in the S+lcat-/- mice and undetectable in the S+/lcat+/+ control. By 10 months of age, the kidneys showed progressive glomerular injury including segmental foam cell infiltrates, mesangial expansion, and hyalinosis. Renal abnormalities are very similar to those seen in human LCAT deficiency. We conclude that the selective high-level accumulation of plasma LpX in the S+lcat-/- mice is strongly associated with a spontaneous glomerulopathy, providing in vivo evidence that LpX contributes to the LCAT deficiency-related nephropathy.

Lurgi develops new propylene process

Abnormal lipid metabolism manifests as hypercholesterolemia in patients with obstructive jaundice due to lipoprotein X (LpX). Our aim was to explore the clinical laboratory characteristics of patients with obstructive jaundice accompanied by dyslipidemia in a large number of samples.A total of 665 patients with obstructive jaundice were included and categorized into two groups (with/without dyslipidemia) based on the ratio of the sum of HDL-c and LDL-c to total cholesterol [(HDL-c + LDL-c)/TC] with a cut-off value of 0.695. Laboratory liver, kidney, and blood lipid parameters were determined. Cholesterol composition assessment was performed by ultracentrifugation and high-performance liquid chromatography (UC-HPLC), and serum protein profiles were analyzed by capillary electrophoresis.Liver function in patients with obstructive jaundice accompanied by dyslipidemia was more aggravated than that in patients with simple obstructive jaundice (P < 0.05). The (HDL-c + LDL-c)/TC ratio was negatively correlated with bilirubin levels (P < 0.05). In addition, the difference in ApoB/LDL-c ratios was statistically significant between the obstructive jaundice accompanied by dyslipidemia group and healthy control group (P < 0.05). The LDL-c concentration determined by the UC-HPLC method was more than five times that determined by the enzymatic method (P < 0.05). Bisalbuminemia was found in 43 of 60 patients with obstructive jaundice accompanied by hypercholesterolemia.In patients with obstructive jaundice, the decreased (HDL-c + LDL-c)/TC ratio may be a novel marker to identify dyslipidemia secondary to LpX. The decreased ratio was associated with poor liver function and indicated disease progression.

Lipoprotein-X stimulates monocyte chemoattractant protein-1 expression in mesangial cells via nuclear factor-κB

Lipoprotein-X (Lp-X) is an abnormal lipoprotein found in the plasma of patients with familial lecithin:cholesterol acyltransferase (LCAT) deficiency. The majority of patients with this disorder develop progressive glomerulosclerosis. One key event in the pathogenesis of glomerulosclerosis is the infiltration of monocytes into affected glomeruli. Mesangial cells can synthesize and secrete monocyte chemoattractant protein-1 (MCP-1), an important chemoattractant for monocytes. The objective of the present study was to examine the effect of Lp-X on MCP-1 expression in mesangial cells leading to an enhanced monocyte chemotaxis and to elucidate the mechanisms involved in this process. Lp-X was isolated from the plasma of a patient with familial LCAT deficiency. After rat mesangial cells were incubated with Lp-X for four or six hours, the expression of MCP-1 mRNA was determined by nuclease protection assay, and MCP-1 protein was measured by Western immunoblotting analysis. Monocyte chemotaxis was determined by using a Micro Chemotaxis Chamber. Lp-X (50 to 100 nmol/mL) stimulated mesangial cell MCP-1 mRNA expression (137 to 220%) and MCP-1 protein levels (233 to 375%). Conditioned media collected from Lp-X-treated mesangial cells stimulated human acute monocytic leukemia (THP-1) monocyte chemotaxis (165 to 200%). The increase in MCP-1 expression in mesangial cells was associated with an elevation of intracellular diacylglycerol levels, and activation of protein kinase C (PKC) as well as nuclear factor-kappa B (NF-kappa B). These results suggest that Lp-X participates in the pathogenesis of glomerulosclerosis and subsequent renal failure in familial LCAT deficient patients by stimulating monocyte infiltration via a mechanism involving mesangial MCP-1 expression.

Hyperlipidemia and atherosclerosis associated with liver disease in ferrochelatase-deficient mice

Erythropoietic protoporphyria (EPP) is an inherited disorder of heme synthesis caused by deficiency of the mitochondrial enzyme ferrochelatase. EPP in humans is associated with liver disease, hypertriglyceridemia, and a low level of high density lipoprotein (HDL) cholesterol. To explore consequences of ferrochelatase deficiency in lipid metabolism, we have analyzed hepatic lipid content and plasma lipoprotein levels in chow-fed BALB/c mice homozygous (fch/fch) or heterozygous (fch/+) for a point mutation in the ferrochelatase gene and in wild-type controls (+/+). Livers of fch/fch mice show bile duct proliferation and biliary fibrosis, but bile formation is not impaired. The free cholesterol content of fch/fch livers is significantly increased when compared with fch/+ and +/+ livers. Plasma cholesterol in fch/fch mice (9.9 ± 6.4 mM) is elevated when compared with fch/+ and +/+ mice (2.9 ± 0.2 and 2.5 ± 0.3 mM, respectively), because of an increased cholesterol content in the very low density lipoprotein-sized fractions, whereas HDL cholesterol is reduced. The ratio of cholesteryl ester to free cholesterol is 4.3 ± 0.6, 3.3 ± 0.3, and 0.3 ± 0.1 in the plasma of +/+, fch/+, and fch/fch mice, respectively. The latter is not due to reduced lecithin:cholesterol acyltransferase activity in plasma of fch/fch mice but to the presence of lipoprotein-X (Lp-X), a particle composed of bile-type lipids usually seen only in cholestatic conditions. Expression of mdr2, essential for biliary phospholipid/ cholesterol secretion, is increased in fch/fch livers. In spite of this, biliary phospholipid/cholesterol secretion is reduced relative to that of bile salts. It is postulated that an inability of bile salts to stimulate lipid secretion adequately leads to formation of Lp-X in this noncholestatic condition. Distinct atherosclerotic lesions were found in aged fch/fch mice. Thus, ferrochelatase deficiency in mice leads to liver disease associated with altered hepatic lipid metabolism, a characteristic hyperlipidemia, and development of atherosclerosis.—Bloks, V. W., T. Plösch, H. van Goor, H. Roelofsen, J. Baller, R. Havinga, H. J. Verkade, A. van Tol, P. L. M. Jansen, and F. Kuipers. Hyperlipidemia and atherosclerosis associated with liver disease in ferrochelatase-deficient mice. J. Lipid Res. 2001. 42: 41–50.

Evaluation of Two Different Homogeneous Assays for LDL-Cholesterol in Lipoprotein-X-positive Serum

The purpose of this study was to evaluate the performance of two homogeneous assays for LDL-cholesterol (LDL-C), a polyethylene/cyclodextrin (PC) assay and a detergent (D) assay, which are based on different principles, in cholestatic serum. We compared serum LDL-C concentrations determined by the two assays for healthy normolipidemic subjects (n = 42) and cholestatic patients (n = 51). LDL-C concentrations obtained with the homogeneous assays were also compared with those obtained by HPLC for patients' sera. In the interference study, conjugated bile acids were added to normal serum, and their effects on the two assays were examined. The effects of lipoprotein-X (LP-X), intermediate-density lipoprotein (IDL), and apolipoprotein (apo) E-rich HDL on the LDL-C assays were also investigated by adding these lipoproteins to normal serum. The LDL-C concentrations obtained with the D assay were higher than those obtained with the PC assay in the serum with high LP-X. The bias for LDL-C vs LP-X in cholestatic serum correlated with LP-X concentration (r = 0.582; P: <0.0001; n = 51). In the interference study, no effect of bile acids on the LDL-C assays was observed. However, the D assay measured 51.0% of the cholesterol in LP-X, whereas no reactivity was observed for LP-X in the PC assay. In addition, the D assay and the PC assay measured IDL-cholesterol at 31.2% and 52.4%, respectively, and measured apo E-rich HDL-C at 7.6% and 17.8%, respectively. Although both homogeneous LDL-C assays are suitable for most cases, the present study showed that each homogeneous assay has a different limitation for cholestatic serum with gross alterations in lipoproteins.

Class III P-glycoproteins mediate the formation of lipoprotein X in the mouse.

Cholestasis is associated with hypercholesterolemia and appearance of the abnormal lipoprotein X (LpX) in plasma. Using mice with a disrupted Mdr2 gene, we tested the hypothesis that LpX originates as a biliary lipid vesicle. Mdr2-deficient mice lack Mdr2 P-glycoprotein, the canalicular translocator for phosphatidylcholine, and secrete virtually no phospholipid and cholesterol in bile. Bile duct ligation of Mdr2(+)/+ mice induced a dramatic increase in the plasma cholesterol and phospholipid concentration. Agarose electrophoresis, density gradient ultracentrifugation, gel permeation, and electron microscopy revealed that the majority of phospholipid and cholesterol was present as LpX, a 40-100 nm vesicle with an aqueous lumen. In contrast, the plasma cholesterol and phospholipid concentration in Mdr2(-)/- mice decreased upon bile duct ligation, and plasma fractionation revealed a complete absence of LpX. In mice with various expression levels of Mdr2 or MDR3, the human homolog of Mdr2, we observed that the plasma level of cholesterol and phospholipid during cholestasis correlated very closely with the expression level of these canalicular P-glycoproteins. These data demonstrate that during cholestasis there is a quantitative shift of lipid secretion from bile to the plasma compartment in the form of LpX. The concentration of this lipoprotein is determined by the activity of the canalicular phospholipid translocator.

Catabolism of lipoprotein-X induced by infusion of 10% fat emulsion

The clinical significance of lipoprotein-X (Lp-X) induced by intravenous infusion of 10% fat emulsion was assessed, with special reference to atherogenesis, by in vitro experiment using purified Lp-X from the sera of patients receiving Intralipid 10%. Lp-X appeared after long-term intravenous infusion of 10% fat emulsion in the patients with intestinal fistula due to the anastomotic leakage. To clarify the role of Lp-X in terms of atherogenicity, the cholesterol metabolism of Lp-X in macrophages as scavenger cells and in hepatocytes as parenchymal cells was studied. When [ 3H] cholesterol-labeled Lp-X or oxidized low-density lipoprotein (o-LDL) was incubated with J-774 macrophages, the incorporation of Lp-X into macrophages was negligible compared with o-LDL. When Lp-X or high-density lipoprotein (HDL) was incubated with J-774 macrophages laden with [ 3H] cholesterol, the release of cholesterol from macrophages was enhanced by Lp-X as well as HDL. When [ 3H] cholesterol-labeled Lp-X LDL or HDL was incubated with the human hepatoma cell line of Hep G 2 cells, the incorporation of Lp-X into Hep G 2 cells was less than that of LDL, but similar to that of HDL. From these findings, it is suggested that the catabolism of Lp-X cholesterol generated with intravenous 10% fat emulsion was mediated by hepatocytes rather than by macrophages, indicating that the hyperlipidemia due to increased Lp-X may not be atherogenic.

Uptake and metabolism of lipoprotein-X in mesangial cells.

Progressive glomerulosclerosis is a major complication in patients with familial lecithin:cholesterol acyltransferase (LCAT) deficiency. The lack of LCAT activity results in the accumulation of an abnormal lipoprotein, lipoprotein-X (Lp-X), in the plasma of these patients. Lipoprotein-X contains high levels of unesterified cholesterol and phosphatidylcholine. Lp-X may play a role in the accumulation of lipids in the kidney, which in turn may lead to glomerulosclerosis. The objective of this study is to examine the uptake and metabolism of Lp-X by rat mesangial cells. Our results suggest that Lp-X is taken up by mesangial cells and that the lipids in Lp-X are metabolized. Lysosomes containing unesterified cholesterol and phosphatidylcholine, in a molar ratio similar to Lp-X, were synthesized to investigate the roles individual apolipoproteins (apo CI, II, III and E) play in the uptake of Lp-X. Both apo CI and CIII inhibited its uptake while apo CII (1.5 fold) and E (4 fold) stimulated the uptake of Lp-X. Very low density lipoprotein (VLDL) and low density lipoprotein (LDL) inhibited Lp-X uptake by mesangial cells. However, at higher concentrations of high density lipoprotein (HDL), the uptake of Lp-X was stimulated. Proteoglycans have an important role in regulating the uptake of Lp-X, while cytoskeleton-dependent phagocytosis and the scavenger receptor do not appear to be involved.

Effect of lipoprotein-X on lipid metabolism in rat kidney.

Lipoprotein-X (Lp-X) is found in the plasma of patients with familial lecithin: cholesterol acyltransferase (LCAT) deficiency syndromes. The majority of the patients with this disorder develop progressive glomerulosclerosis. In this study, the effect of Lp-X on lipid metabolism in perfused rat kidney was investigated. Lp-X was isolated from plasma of patients with familial LCAT deficiency by sequential ultracentrifugation and gel filtration column chromatography. Rat kidneys were perfused for 1-2 h with Krebs-Henseleit buffer containing 20 microM [1-(14)C]acetate or 20 microM [Me-3H]choline. In the presence of Lp-X, no significant difference in the incorporation of radioactivity into triglycerides, cholesterol, phosphocholine, CDP-choline and sphingomyelin was observed. However, incorporation of radioactivity into cholesteryl esters and phosphatidylcholine was significantly elevated in Lp-X perfused kidneys. The contents of cholesterol, cholesteryl esters and phosphatidylcholine were also significantly increased in Lp-X perfused kidneys. The increase in lipid content in the Lp-X perfused kidney is attributed to the direct deposition of Lp-X lipids into the organ. The increase in the labelling of cholesteryl esters was attributed to the increase of available substrate (cholesterol) for the acyl-CoA:cholesterol acyltransferase (ACAT) reaction. The increase in phosphatidylcholine labelling was caused by a reduced turnover of the newly synthesized labelled phosphatidylcholine during Lp-X perfusion.

Role of lecithin:cholesterol acyltransferase and apolipoprotein A-I in cholesterol esterification in lipoprotein-X in vitro

Lipoprotein-X (Lp-X) is an abnormal particle present in the plasma of patients with familial lecithin:cholesterol acyltransferase (LCAT) deficiency syndromes or cholestatic liver disease. Compared to other lipoproteins, Lp-X contains a high content of unesterified cholesterol (30%, w/w) to phosphatidylcholine (60%, w/w). The objective of this study was to evaluate the role of LCAT and apolipoprotein A-I (apoA-I) in Lp-X metabolism in vitro and to elucidate the regulation of cholesterol esterification in this unique lipoprotein. Lp-X isolated from sera of patients with obstructive jaundice had a high content of unesterified cholesterol and phosphatidylcholine and contained apolipoprotein E, apoCs, and albumin. Although human recombinant LCAT used as an enzyme source did bind to isolated Lp-X, no cholesterol esterification was detected. However, addition of human apoA-I in the presence of albumin resulted in significant cholesterol esterification in Lp-X (Vmax 0.25 +/- 0.04 nmol/h per microgram LCAT protein). Exogenous apoA-I did not change the size of Lp-X particle as determined by quasi-elastic light scattering analysis. A reduction in Lp-X size was observed when both apoA-I and LCAT were included in the reaction mixture (from 47 nm to 42 nm). Furthermore, addition of apoA-I (but not HDL) dramatically changed the electrophoretic mobility of Lp-X from cathodic to anodic migration. Such changes are not due to displacement of apoC or apoE proteins from Lp-X by apoA-I. While increasing apoA-I concentration (up to 35 micrograms/ml) in the reaction mixture stimulated cholesterol esterification in Lp-X, addition of apoA-I at the concentration of 8 micrograms/ml inhibited cholesterol esterification in VLDL, LDL, and HDL by more than 50%. Albumin was required for the LCAT reaction to Lp-X. Our results suggest that while LCAT binds to isolated Lp-X, apoA-I is needed for the LCAT reaction to proceed. The presence of apoA-I does not result in the displacement of apoCs and apoE from Lp-X and addition of apoA-I changes the electrophoretic mobility but not the size of Lp-X.

Lipoprotein-X fails to inhibit hydroxymethylglutaryl coenzyme A reductase in HepG2 cells

Abnormalities of lipid composition and metabolism are frequently observed in patients with cholestatic liver disease. Both elevated low-density lipoprotein (LDL) levels and the appearance of lipoprotein-X (LP-X) in plasma underlie the high incidence of hypercholesterolemia in this population. We tested the hypothesis that the hypercholesterolemia of cholestasis may reflect a failure of normal feedback regulation of hepatic cholesterolgenesis by determining the influence of LP-X on the rate-limiting enzyme of cholesterol synthesis, hydroxymethylglutaryl coenzyme A (HMG CoA) reductase. Cultured human hepatoma (HepG2) cells were incubated in purified lipoprotein for 24 hours, harvested, and then assayed for HMG CoA reductase activity and mass. LDL isolated from either normal controls or patients with cholestasis decreased reductase activity in a dose-dependent fashion (2 to 30 μg cholesterol/mL media) to a level approximately 50% of that measured in cells incubated in lipid-deficient serum. LP-X failed to downregulate enzyme activity compared with LDL, with little change in reductase activity at cholesterol concentrations (30 μg/mL media) that produced maximal reductase inhibition by LDL. Three distinct LP-X subspecies were purified from the plasma of a patient with primary biliary cirrhosis (PBC) and tested in an analogous manner. All LP-X subspecies were similar in their inability to decrease reductase activity as compared with LDL. HMG CoA reductase mass was increased approximately twofold in cells incubated with LP-X, as estimated by Western blot analysis. These results suggest that LP-X may contribute to hypercholesterolemia in the cholestatic patient by not effectively downregulating hepatic cholesterol synthesis.

Effect of Intensive Chemo- and Radiotherapy Preceding Bone Marrow Transplantation on Lipid and Lipoprotein Metabolism

Background: Organ toxicity related to the conditioning regimen preceding bone marrow transplantation is a source of transplant-related mortality and a common cause of morbidity. A frequent finding of toxicity analysis is the occurrence of regimen-related hepatic toxicity, which has been found in more than 80% of patients receiving conditioning therapy. In order to characterize the effect of regimen-related hepatic toxicity on lipid and lipoprotein metabolism we studied patients before, during and after conditioning therapy preceding bone marrow transplantation. Patients and Methods: Sera from 22 patients were examined for total cholesterol, free cholesterol, triglycerides, lipoprotein electrophoretic patterns, apolipoproteins A-I, A-II and B, phospholipids, lipoprotein-X, and fatty acid composition before, during and after bone marrow transplantation. In addition, erythrocyte membranes were analyzed for cholesterol content, phospholipid fractions, and fatty acid composition. Marrow ablation and immunosuppression for bone marrow transplantation conditioning were achieved by treatment with high-dose chemotherapy, mostly combined with total-body irradiation. Results: After conditioning therapy, the serum concentrations of total cholesterol, HDL cholesterol, LDL cholesterol, apolipoproteins A-I, A-II, and B were decreased, whereas concentrations of free cholesterol, phospholipids, and triglycerides were increased. In 13 of 14 patients examined, lipoprotein-X (LP-X) could be identified in serum after conditioning therapy. In erythrocyte membranes, free cholesterol and the ratio free cholesterol/phospholipid increased, whereas arachidonic acid decreased after conditioning therapy. Conclusions: The lipid and lipoprotein abnormalities observed in patients after conditioning therapy are similar to those in hepatocellular injury, indicating a ‘secondary’ LCAT deficiency. However, side effects of lipid emulsions infused during the observation period as part of total parenteral nutrition may have affected study results as well.

Management of lipoprotein-x accumulation in severe cholestasis by semi-selective ldl-apheresis

Liver disorders characterized by prolonged bile stasis are often associated with the accumulation of an abnormal lipoprotein, lipoprotein-X (LP-X), in plasma. LP-X is separated in the low-density lipoprotein (LDL) density range, but lacks apolipoprotein B and does not interact with the LDL receptor; LP-X can cause hyperlipidemia, cutaneous xanthomas, and worsening of arterial disease. We report the case of a patient with severe cholestasis, markedly elevated plasma cholesterol levels (26.8 to 31.5 mmol/L), mainly due to a massive accumulation of LP-X in plasma, and diffuse xanthomas. To reduce the elevated cholesterol levels, the patient was given extracorporeal treatment aimed at removing atherogenic lipoprotein (LDL-apheresis). LDL-apheresis was performed at weekly or bi-weekly intervals, either by a semi-selective technique using filters with a defined pore diameter (double filtration, DF) or by a more selective technique using dextran-sulfate-cellulose (DSC) columns able to bind LDL. The semi-selective DF technique proved more effective than DSC, removing 48% of total cholesterol (compared to 30% with DSC), and lowering cholesterol levels to 11.1 mmol/L in 6 weeks. DF removed both LDL and LP-X from plasma, whereas DSC selectively decreased the LDL content. The reduction of plasma cholesterol levels was associated with a complete regression of the xanthomas, supporting DF apheresis as a first-choice treatment for patients with massive LP-X accumulation due to cholestasis.

Impaired Efficacy of Selective Ldl-Apheresis in Primary Biliary Cirrhosis

Low-density lipoprotein apheresis (LDL-apheresis) was done with either cascade filtration (DF) or dextran sulfate cellulose adsorption (DSC) in a patient with primary biliary cirrhosis who developed severe dyslipidemia associated with cholestasis and accumulation of lipoprotein-X (LP-X). The extracorporeal treatment was initially performed weekly, and resulted in a sharp drop in total cholesterol from 1038 to 430 mg/dl. During the next four months the patient was treated every 10-15 days, and pre-apheresis cholesterol levels were maintained between 438 and 505 mg/dl, until an orthotopic liver transplantation was successfully performed. With semi-selective DF a mean 47.1% of total cholesterol was removed per procedure compared to 30.0% with DSC, although the volume of treated plasma was 38.0 vs 49.9 ml/kg body weight. The changes in plasma cholesterol levels during DSC and DF showed that the kinetics of cholesterol removal were similar with both techniques, but the efficacy differed; DF removed both LDL and LP-X from plasma, whereas DSC selectively lowered the LDL content. Cascade filtration may therefore be considered as a first-choice treatment for patients with LP-X accumulation due to cholestasis.

Management of lipoprotein-X accumulation in severe cholestasis by semi-selective LDL-apheresis

Liver disorders characterized by prolonged bile stasis are often associated with the accumulation of an abnormal lipoprotein, lipoprotein-X (LP-X), in plasma. LP-X is separated in the lowdensity lipoprotein (LDL) density range, but lacks apolipoprotein B and does not interact with the LDL receptor, LP-X can cause hyperlipidemia, cutaneous xanthomas, and worsening of arterial disease. We report the case of a patient with severe cholestasis, markedly elevated plasma cholesterol levels (26.8 to 31.5 mmol/L), mainly due to a massive accumulation of LP-X in plasma, and diffuse xanthomas. To reduce the elevated cholesterol levels, the patient was given extracorporeal treatment aimed at removing atherogenic lipoprotein (LDL-apheresis). LDLapheresis was performed at weekly or bi-weekly intervals, either by a semi-selective technique using filters with a defined pore diameter (double filtration, DF) or by a more selective technique using dextran-sulfate-cellulose (DSC) columns able to bind LDL. The semi-selective DF technique proved more effective than DSC, removing 48% of total cholesterol (compared to 30% with DSC), and lowering cholesterol levels to 11.1 mmol/L in 6 weeks. DF removed both LDL and LP-X from plasma, whereas DSC selectively decreased the LDL content. The reduction of plasma cholesterol levels was associated with a complete regression of the xanthomas, supporting DF apheresis as a first-choice treatment for patients with massive LP-X accumulation due to cholestasis.

Effect of fat-emulsion phospholipids on serum lipoprotein profile during 1 mo of cyclic total parenteral nutrition

Changes in serum lipoprotein determined by selective precipitation were investigated in 11 adult patients during 1 mo of parenteral nutrition. Patients were divided into two groups that received a similar nutrient regimen except for Intralipid (IL) phospholipid, which was higher in group A (10% IL, n = 5) than in group B (20% IL, n = 6), 139 +/- 15 vs 71 +/- 0.5 mg.kg-1.d-1 (P less than 0.01). Lipoprotein X (LPX) detected soon after IL infusions were started reached its highest concentrations in group A. LPX concentrations correlated with phospholipid intakes on days 7 and 15 but not on day 29. Significant increases in the cholesterol and phospholipid content of low-density-lipoprotein-very-low-density-lipoprotein fractions were observed only in group A. It is suggested that these changes were induced by the twofold-higher intake of phospholipids in group A. With regard to the possible involvement of LPX in lipid overloading of the reticuloendothelial system and hepatocytes, administration of 20% IL seems preferable to 10% IL.

Effects of intravenous infusions of commercial fat emulsions (Intralipid 10 or 20%) on rat plasma lipoproteins: Phospholipids in excess are the main precursors of lipoprotein-X-like particles

Like most commercial parenteral emulsions, Intralipid ® contains the same amount of phospholipids (12 mg/ml) to stabilize 100 or 200 mg of soybean oil (10 or 20% formula, respectively). By centrifugation, 10 or 20% Intralipid was separated into a supernatant, fat particles containing the bulk of triacylglycerols stabilized by a fraction of phospholipids and an infranatant — called mesophase — consisting mainly of phospholipids used in excess as emulsifier. We observed that the initial triacylglycerol phospholipid ratio of the emulsion ( 100 12 and 200 12 , respectively) determines the size of the triacylglycerol-rich particles (260 and 350 nm) as well as the phospholipid content of the mesophase (6.02 and 4.67 mg/ml). To understand the mechanism of the lipoprotein-X (LPX) accumulation generally reported after intravenous fat infusions, plasma lipid levels and lipoprotein profiles were first compared in the rats after infusion (at a constant rate of 0.5 or 1 ml/h for 43 h) of Intralipid 10 or 20%. For the same intravenous triacylglycerol load (100 mg/h), rats infused with Intralipid 10% at 1 ml/h displayed higher triacylglycerol levels than rats infused with the 20% emulsion at 0.5 ml/h, suggesting that the size of exogenous fat particles modulated the catabolic rate of their triacylglycerols. The plasma levels of LPX varied according to the infusion rate of phospholipids not associated with triacylglycerol-rich particles of the emulsion. Moreover, an apo E and apo B enrichment of plasma and an elevation of the apo B48/apo B100 ratio was always observed after Intralipid infusions. In order to confirm that phospholipids of the mesophase are the main LPX precursors, lipoprotein profiles were then compared in the rats after intravenous infusion, at a constant rate of 1 ml/h, of either the mesophase or a suspension of triacylglycerol-rich particles isolated from Intralipid 20%. As expected, significant LPX amounts were only detected in rats infused with the pure mesophase of the emulsion. It was concluded that products of the lipolysis of exogenous fat particles play only a minor role in the formation of LPX. In fact these abnormal lipoproteins are generated by phospholipids of the mesophase which, like infused liposomes, actively mobilize endogenous free cholesterol. Consequently, in order to be considered as true chylomicron models for safe fat delivery in parenteral nutrition and in order to prevent some detrimental effects on cholesterol metabolism, commercial emulsions should be cleared of phospholipid excess.

Lipoprotein pattern and plasma lipoprotein lipase activities in patients with primary biliary cirrhosis. Relationship with increase of HDL2 fraction in Lp-X-positive and Lp-X-negative subjects.

Plasma lipids, apoprotein A-I and B in serum and in lipoprotein fractions (VLDL + LDL, HDL2, and HDL3) obtained by preparative ultracentrifugation, as well as postheparin lipoprotein lipase activity (H-TGL and LPL) were evaluated in 17 subjects with primary biliary cirrhosis (stage II and III) subdivided into two groups according to the presence or absence of lipoprotein X (Lp-X). A reduction in total lipoprotein lipase activity was observed in both patient groups, compared to controls (P less than 0.01); the hepatic lipoprotein lipase was significantly reduced (P less than 0.01) only in the Lp-X-positive group. The lipid (477.8 +/- 154.3 vs 239.6 +/- 51.1; P less than 0.01) and protein (147.4 +/- 37.1 vs 83.3 +/- 19.7; P less than 0.01) masses in the VLDL + LDL fraction of the Lp-X-positive group were increased compared to controls. In the same group, the HDL2 fraction also showed an increase in lipid (186.6 +/- 80.0 vs 77.9 +/- 21.6; P less than 0.01) and protein (133.9 +/- 60.0 vs 67.9 +/- 16.5; P less than 0.01) masses; in addition, the HDL2 percent lipid composition was different in the two patient groups, showing a decrease in esterified cholesterol (20.4 +/- 3.6 vs 25.7 +/- 2.2; P less than 0.01) and an increase in phospholipids (59.2 +/- 2.9 vs 54.8 +/- 2.6; p less than 0.01) in the Lp-X-positive group.(ABSTRACT TRUNCATED AT 250 WORDS)

A simple method for quantitative determination of lipoprotein-X (LP-X) in human serum

A simple method for quantitative determination of lipoprotein-X (LP-X) in human serum