Changes in apolipoprotein A-I mRNA level in the liver of rats with experimental nephrotic syndrome

In previous studies we had shown that: one of the most specific feature of hyperlipoproteinemia found in rats with experimental nephrotic syndrome is the accumulation of apolipoprotein A-I-rich HDL in plasma and this disorder is associated with an overproduction of apolipoprotein A-I by the liver. The present study was designed to investigate whether the increased hepatic synthesis of apolipoprotein A-I was due to an accumulation of functionally active apolipoprotein A-I mRNA in liver of nephrotic rats. Hepatic mRNA was translated in vitro by rabbit reticulocyte lysate in the presence of [35S]methionine and in vitro synthesized apolipoprotein A-I, albumin and apolipoprotein E were immunoprecipitated by specific rabbit IgG. In nephrotic rats the amount of in vitro synthesized apolipoprotein A-I was almost twice that found in the controls, suggesting that functionally active apolipoprotein A-I mRNA was increased in liver of nephrotic rats. To confirm that this difference in apolipoprotein A-I mRNA activity was due to an actual increase of hepatic apolipoprotein A-I mRNA sequences, we performed nucleic acid hybridization experiments (northern blot) using several cloned cDNA probes (rat and human apolipoprotein A-I, rat apolipoprotein E and apolipoprotein A-II). The results indicate that in nephrotic rats the amount of hybridizable apolipoprotein A-I mRNA sequences was about 3-fold higher than that in controls. In contrast, there was no difference in the amount of hybridizable apolipoprotein A-II and apolipoprotein E mRNA sequences, indicating that the change in apolipoprotein A-I mRNA induced by the nephrotic state was specific for this mRNA.     

Differential effect of ultracentrifugation on apolipoprotein A-I-containing lipoprotein subpopulations

Two populations of apolipoprotein (apo) A-I-containing lipoprotein particles are found in high density lipoproteins (HDL): those that also contain apo A-II[Lp(A-I w A-II)] and those that do not [Lp(A-I w/o A-II)]. Lp(A-I w/o A-II) comprised two distinct particle sizes with mean hydrates Stokes diameter of 10.5 nm for Lp(A-I w/o A-II)1 and 8.5 nm for Lp(A-I w/o A-II)2. To study the effect of ultracentrifugation on these particles, Lp(A-I w/o A-II) and Lp(A-I w A-II) were isolated from the plasma and the ultracentrifugal HDL (d 1.063-1.21 g/ml fractions) of five normolipidemic and three hyperlipidemic subjects. The size subpopulations of these particles were studied by gradient polyacrylamide gel electrophoresis. Several consistent differences were detected between plasma Lp(A-I w/o A-II) and HDL Lp(A-I w/o A-II). First, in all subjects, the relative proportion of Lp(A-I w/o A-II)1 to Lp(A-I w/o A-II)2 isolated from HDL was reduced. Second, particles larger than Lp(A-I w/o A-II)1 and smaller than Lp(A-I w/o A-II)2 were considerably reduced in HDL. Third, a distinct population of particles with approximate Stokes diameter of 7.1 nm usually absent in plasma was detected in HDL Lp(A-I w/o A-II). Little difference in subpopulation distribution was detected between Lp(A-I w A-II) isolated from the plasma and HDL of the same subject. When plasma Lp(A-I w/o A-II) and Lp(A-I w A-II) were centrifuged, 14% and 4% of A-I were, respectively, recovered in the D greater than 1.21 g/ml fraction. Only 2% A-II was found in this density fraction. These studies show that the Lp(A-I w/o A-II) particles are less stable than Lp(A-I w A-II) particles upon ultracentrifugation. Among the various Lp(A-I w/o A-II) subpopulations, particles larger than Lp(A-I w/o A-II)1 and smaller than Lp(A-I w/o A-II)2 are most labile.     

Apolipoprotein A-II/A-I ratio is a key determinant in vivo of HDL concentration and formation of pre-beta HDL containing apolipoprotein A-II

Overexpression of human apolipoprotein A-II (apo A-II) in mice induced postprandial hypertriglyceridemia and marked reduction in plasma HDL concentration and particle size [Boisfer et al. (1999) J. Biol. Chem. 274, 11564-11572]. We presently compared lipoprotein metabolism in three transgenic lines displaying plasma concentrations of human apo A-II ranging from normal to 4 times higher, under ad libitum feeding and after an overnight fast. Fasting dramatically decreased VLDL and lowered circulating human apo A-II in transgenic mice; conversely, plasma HDL levels increased in all genotypes. The apo A-I content of HDL was inversely related to the expression of human apo A-II, probably reflecting displacement of apo A-I by an excess of apo A-II. Thus, the molar ratios of apo A-II/A-I in HDL were significantly higher in fed as compared with fasted animals of the same transgenic line, while endogenous LCAT activity concomitantly decreased. The number and size of HDL particles decreased in direct proportion to the level of human apo A-II expression. Apo A-II was abundantly present in all HDL particles, in contrast to apo A-I mainly present in large ones. Two novel findings were the presence of pre-beta migrating HDL transporting only human apo A-II in the higher-expressing mice and the increase of plasma HDL concentrations by fasting in control and transgenic mice. These findings highlight the reciprocal modifications of VLDL and HDL induced by the feeding-fasting transition and the key role of the molar ratio of apo A-II/A-I as a determinant of HDL particle metabolism and pre-beta HDL formation.     

Apolipoproteins A-I,A-II and E are independently distributed among intracellular and newly secreted HDL of human hepatoma cells

Whereas hepatocytes secrete the major human plasma high density lipoproteins (HDL)-protein, apo A-I, as lipid-free and lipidated species, the biogenic itineraries of apo A-II and apo E are unknown. Human plasma and HepG2 cell-derived apo A-II and apo E occur as monomers, homodimers and heterodimers. Dimerization of apo A-II, which is more lipophilic than apo A-I, is catalyzed by lipid surfaces. Thus, we hypothesized that lipidation of intracellular and secreted apo A-II exceeds that of apo A-I, and once lipidated, apo A-II dimerizes. Fractionation of HepG2 cell lysate and media by size exclusion chromatography showed that intracellular apo A-II and apo E are fully lipidated and occur on nascent HDL and VLDL respectively, while only 45% of intracellular apo A-I is lipidated. Secreted apo A-II and apo E occur on small HDL and on LDL and large HDL respectively. HDL particles containing both apo A-II and apo A-I form only after secretion from both HepG2 and Huh7 hepatoma cells. Apo A-II dimerizes intracellularly while intracellular apo E is monomeric but after secretion associates with HDL and subsequently dimerizes. Thus, HDL apolipoproteins A-I, A-II and E have distinct intracellular and post-secretory pathways of hepatic lipidation and dimerization in the process of HDL formation. These early forms of HDL are expected to follow different apolipoprotein-specific pathways through plasma remodeling and reverse cholesterol transport.     

Immunoaffinity fractionation of high-density lipoprotein subclasses 2 and 3 using anti-apolipoprotein A-I and A-II immunosorbent gels

High-density lipoprotein (HDL) subclasses 2 and 3 prepared by density gradient ultracentrifugation have been further fractionated by immunoaffinity chromatography using antibody affinity gels targetting the major HDL apolipoproteins, A-I and A-II. Fractions containing A-I without A-II (AI w/o AII) and A-I with A-II (AI w AII) were isolated from both density ranges. Whereas there were similar concentrations of the major subfraction (HDL3(AI w AII] in both males and females, the remaining subfractions were present in higher concentrations in females as compared to males, in the order HDL3 (AI w/o AII) less than HDL2(AI w AII) less than HDL2(AI w/o AII). The difference was most marked for HDL2 (AI w/o AII), where plasma concentrations in females were almost 3-fold greater than in males. Compositional analyses indicated that the plasma concentrations of the fractions, rather than their compositions, were the major determinants of male-female differences in HDL levels. In contrast, fractions defined by similar apolipoprotein criteria and isolated from different density subclasses (i.e., HDL2(AI w/o AII) vs. HDL3(AI w/o AII) and HDL2(AI w AII) vs. HDL3(AI w AII] showed major compositional differences. This is suggestive of distinct lipoprotein particles.     

Calcium transport in basolateral plasma membranes from kidney cortex of Milan hypertensive rats

Ca2+ transport was investigated in basolateral plasma membranes (BLM) isolated from kidney cortex of the Milan strain of genetically hypertensive rats (MHS) and their normotensive controls (MNS) during a pre-hypertensive stage (age 3-4 weeks). It was found that the Vmax of ATP-dependent Ca2+ transport (in the presence of calmodulin) was about 16% lower in MHS than in control rats. In membranes from MNS rats which had been isolated in the presence of EGTA, the ATP-dependent Ca2+ transport showed a hyperbolic Ca2+ concentration dependence, a high Km (Ca2+) and a low Vmax; upon addition of exogenous calmodulin, the kinetics became sigmoidal, the Km (Ca2+) was decreased and the Vmax was increased. In membranes from MHS rats, the Ca2+ concentration dependence of ATP-driven Ca2+ transport was sigmoidal and the Ca2+ affinity was high in the absence of added calmodulin. Addition of exogenous calmodulin to these membranes resulted in an increase in Vmax, but no change in other kinetic parameters. Low-affinity hyperbolic kinetics of Ca2+ transport could only be obtained in MHS rats if the membranes were extracted with hypotonic EDTA and hypertonic KCl. These data suggest that the plasma membrane Ca2+-ATPase, which catalyses the ATP-dependent Ca2+ transport, exists in BLM of pre-hypertensive MHS rats predominantly in an activated, high-affinity form.     

Interaction between high-density lipoprotein subpopulations in apo B-free and abetalipoproteinemic plasma.

Two populations of high-density lipoprotein (HDL) particles exist in human plasma. Both contain apolipoprotein (apo) A-I, but only one contains apo A-II: Lp(AI w AII) and Lp(AI w/o AII). To study the extent of interaction between these particles, apo B-free plasma prepared by the selective removal of apo B-containing lipoproteins (LpB) from the plasma of three normolipidemic (NL) subjects and whole plasma from two patients with abetalipoproteinemia (ABL) were incubated at 37 degrees C for 24 h. Apo B-free plasma samples were used to avoid lipid-exchange between HDL and LpB. Lp(AI w AII) and Lp(AI w/o AII) were isolated from each apo B-free plasma sample before and after incubation and their protein and lipid contents quantified. Before incubation, ABL plasma had reduced levels of Lp(AI w AII) and Lp(AI w/o AII), (40% and 70% of normals, respectively). Compared to the HDL of apo B-free NL plasma, ABL HDL had higher relative contents of free cholesterol, phospholipid and total lipid, and contained more particles with apparent hydrated Stokes diameter in the 9.2-17.0 nm region. These differences were particularly pronounced in particles without apo A-II. Despite their differences, the total cholesterol contents of Lp(AI w AII) increased, while that of Lp(AI w/o AII) decreased in all five plasma samples and the amount of apo A-I in Lp(AI w AII) increased by 6-8 mg/dl in four during the incubation. These compositional changes were accompanied by a relative reduction of particles in the 7.0-8.2 nm Stokes diameter size region and an increase of particles in the 9.2-11.2 nm region. These data are consistent with intravascular modulation between HDL particles with and without apo A-II. The observed increase in apo A-II-associated cholesterol and apo A-I, could involve either the transfer of cholesterol and apo A-I from particles without apo A-II to those with A-II, or the transfer of apo A-II from Lp(AI w AII) to Lp(AI w/o AII). The exact mechanism and direction of the transfer remain to be determined.     

原发性肾病综合征高脂血症发生机制的研究进展*

摘要：原发性肾病综合征高脂血症主要表现为血浆总胆固醇（Ch）和低密度脂蛋白胆固醇（LDL-Ch）明显升高,甘油三酯（TG）和极 低密度脂蛋白胆固醇（VLDL-Ch）升高。高密度脂蛋白胆固醇（HDL-Ch）浓度可降低或不变,但HDL的亚型分布异常,即HDL3 增 加而HDL2 减少。这提示我们HDL3 转变为富含CH的HDL2 成熟障碍。不过,关于高脂血症在肾病综合征的发生机制较为复杂, 还不十分清楚。但是目前有关该机制的观点主要集中在（1）低白蛋白血症刺激肝脏合成胆固醇、甘油三酯、脂蛋白增加,（2）外周 脂蛋白的清除障碍,（3）高密度脂蛋白（HDL）的成熟障碍,本文就此做一综述。     

Distribution of cholesterol and apolipoprotein A-I and A-II in human high density lipoprotein subfractions separated by CsCl equilibrium gradient centrifugation: evidence for HDL subpopulations with differing A-I/A-II molar ratios.

The purpose of this experiment was to characterize the high density lipoproteins (HDL) as a function of hydrated density. HDL was subfractionated on the basis of hydrated density by CsCl density gradient centrifugation of whole serum or the d 1.063-1.25 g/ml HDL fraction isolated from three men and three women. Apolipoprotein A-I and A-II quantitation by radial immunodiffusion showed that the A-I/A-II ratio varied with the lipoprotein hydrated density. The A-I/A-II molar ratio of HDL lipoproteins banding between d 1.106 and 1.150 g/ml was nearly constant at 2.2 +/- 0.2. In the density range 1.151-1.25 g/ml the A-I/A-II ratio increased as the density increased. On the other hand, in the density range between 1.077 and 1.105 the A-I/A-II ratio increased as the density decreased, ranging from 2.8 +/- 0.5 for the d 1.093-1.105 g/ml fraction to 5.6 +/- 1.3 for the d 1.077-1.082 g/ml fraction. The d 1.063-1.076 g/ml fraction and the d 1.077-1.082 g/ml fractions had comparable A-I/A-II ratios. Serum and the d 1.063-1.25 g/ml HDL fraction exhibited similar trends. The cholesterol/(A-I + A-II) ratio decreased as the density increased in all 12 samples (six serum and six HDL) examined. Gradient gel electrophoresis of the density gradient fractions showed that as the density increased from 1.063 to 1.200 g/ml the apparent molecular weight decreased from 3.9 x 10(5) to 1.1 x 10(5). HDL subfractions with the same hydrated densities had comparable molecular weights and A-I/A-II and cholesterol/(A-I + A-II) ratios when isolated from men or women. HDL contains subpopulations that differ in the A-I/A-II molar ratio.-Cheung, M. C., and J. J. Albers. Distribution of cholesterol and apolipoprotein A-I and A-II in human high density lipoprotein subfractions separated by CsCl equilibrium gradient centrifugation: evidence for HDL subpopulations with differing A-I/A-II molar ratios.     

Characterization of human high-density lipoprotein subclasses LP A-I and LP A-I/A-II and binding to HepG2 cells

Plasma HDL can be classified according to their apolipoprotein content into at least two types of lipoprotein particles: lipoproteins containing both apo A-I and apo A-II (LP A-I/A-II) and lipoproteins with apo A-I but without apo A-II (LP A-I). LP A-I and LP A-I/A-II were isolated by immuno-affinity chromatography. LP A-I has a higher cholesterol content and less protein compared to LP A-I/A-II. The average particle mass of LP A-I is higher (379 kDa) than the average particle weight of LP A-I/A-II (269 kDa). The binding of 125I-LP A-I to HepG2 cells at 4 degrees C, as well as the uptake of [3H]cholesteryl ether-labelled LP A-I by HepG2 cells at 37 degrees C, was significantly higher than the binding and uptake of LP A-I/A-II. It is likely that both binding and uptake are mediated by apo A-I. Our results do not provide evidence in favor of a specific role for apo A-II in the binding and uptake of HDL by HepG2 cells.     

原发性肾病综合征高脂血症发生机制的研究进展

原发性肾病综合征高脂血症主要表现为血浆总胆固醇（Ch）和低密度脂蛋白胆固醇（LDL—Ch）明显升高,甘油三酯（TG）和极低密度脂蛋白胆固醇（VLDL—Ch）升高。高密度脂蛋白胆固醇（HDL—Ch）浓度可降低或不变,但HDL的亚型分布异常,即HDL3增加而HDL2减少。这提示我们HDL3转变为富含CH的HDL2成熟障碍。不过,关于高脂血症在肾病综合征的发生机制较为复杂,还不十分清楚。但是目前有关该机制的观点主要集中在（1）低白蛋白血症刺激肝脏合成胆固醇、甘油三酯、脂蛋白增加,（2）外周脂蛋白的清除障碍,（3）高密度脂蛋白（HDL）的成熟障碍,本文就此做一综述。     

Stimulation of lecithin:cholesterol acyltransferase activity by apolipoprotein A-II in the presence of apolipoprotein A-I

Various combinations of incorporation and addition of apolipoprotein A-I (apo A-I) and apolipoprotein A-II (apo A-II) individually or together to a defined lecithin-cholesterol (250/12.5 molar ratio) liposome prepared by the cholate dialysis procedure were used to study the effect of apo A-II on lecithin:cholesterol acyltransferase (LCAT, EC 2.3.1.43) activity of both purified enzyme preparations and plasma. When apo A-I (0.1-3.0 nmol/assay) alone was incorporated or added to the liposome, apo A-I effectively activated the enzyme. By contrast, when apo A-II (0.1-3.0 nmol/assay) alone was incorporated into or added to the liposome, apo A-II exhibited minimal activation of LCAT activity, approximately 1% of the activity obtained by an equal amount of apo A-I. Addition of apo A-II (0.1-3.0 nmol/assay) together with apo A-I (0.8 nmol/assay) to the liposome reduced the LCAT activity to approximately 30% of the level obtained with addition of apo A-I alone. On the other hand, addition of apo A-II (0.1-3.0 nmol/assay) or addition of lecithin-cholesterol liposome containing apo A-II (0.1-3.0 nmol/assay) to lecithin-cholesterol liposome containing apo A-I (0.8 nmol/assay) did not significantly alter apo A-I activation of LCAT activity. However, when the same amounts (0.1-3.0 nmol/assay) of apo A-II were incorporated together with apo A-I (0.8 nmol/assay) into the liposome, apo A-II significantly stimulated LCAT activity as compared to activity obtained with incorporation of apo A-I alone. The maximal stimulation was obtained with 0.4 nmol apo A-II/assay for both purified and plasma enzyme. At this apo A-II concentration, approximately 4-fold and 1.8-fold stimulation was observed for purified enzyme and plasma enzyme, respectively. These results indicated that apo A-II must be incorporated together with apo A-I into lecithin-cholesterol liposomes to exert its stimulatory effect on LCAT activity and that apo A-II in high-density lipoprotein may play an important role in the regulation of LCAT activity.     

Inhibitory effect of resveratrol on proteinuria, hypoalbuminemia and hyperlipidemia in nephritic rats.

The effect of resveratrol, a polyphenolic compound present in grapes and other plants, on proteinuria, hypoalbuminemia and hyperlipidemia was studied in rats with glomerulonephritis. The nephritis was induced by an intravenous injection of anti-rat kidney glomerular basement membrane rabbit antiserum. Nephritic rats were given oral intubation of resveratrol (5 mg/day/100 g body weight) for 14 days, while control nephritic rats as well as normal ones were similarly given vehicle alone. By resveratrol treatment, enlargement in liver and kidney due to nephritis induction was significantly reduced, together with partial restoration of nephritis-induced reduction in body weight gain. Both proteinuria and hypoalbuminemia, characteristic symptoms to nephrotic syndrome, were significantly remedied, that is, urinary protein excretion was suppressed and serum albumin concentration was increased by resveratrol treatment. Resveratrol also suppressed significantly hyperlipidemia incident to nephritis, the hypotriglyceridemic action being more prominent than the hypocholesterolemic one. From these results, resveratrol is suggested to be a potent anti-glomerulonephritic food factor capable of suppressing proteinuria, hypoalbuminemia and hyperlipidemia at the same time.     

The origin and metabolism of a nascent pre-β high density lipoprotein involved in cellular cholesterol efflux

The pre-β HDL fraction constitutes a heterogeneous population of discoid nascent HDL particles. They transport from 1 to 25 % of total human plasma apo A-I. Pre-β HDL particles are generated de novo by interaction between ABCA1 transporters and monomolecular lipid-free apo A-I. Most probably, the binding of apo A-I to ABCA1 initiates the generation of the phospholipid-apo A-I complex which induces free cholesterol efflux. The lipid-poor nascent pre-β HDL particle associates with more lipids through exposure to the ABCG1 transporter and apo M. The maturation of pre-β HDL into the spherical α-HDL containing apo A-I is mediated by LCAT, which esterifies free cholesterol and thereby forms a hydrophobic core of the lipoprotein particle. LCAT is also a key factor in promoting the formation of the HDL particle containing apo A-I and apo A-II by fusion of the spherical α-HDL containing apo A-I and the nascent discoid HDL containing apo A-II. The plasma remodelling of mature HDL particles by lipid transfer proteins and hepatic lipase causes the dissociation of lipid-free/lipid-poor apo A-I, which can either interact with ABCA1 transporters and be incorporated back into pre-existing HDL particles, or eventually be catabolized in the kidney. The formation of pre-β HDL and the cycling of apo A-I between the pre-β and α-HDL particles are thought to be crucial mechanisms of reverse cholesterol transport and the expression of ABCA1 in macrophages may play a main role in the protection against atherosclerosis.     

Lipoprotein lipase and hepatic lipase: their relationship with HDL subspecies Lp(A-I) and Lp(A-I,A-II)

HDL subspecies Lp(A-I) and Lp(A-I,A-II) have different anti-atherogenic potentials. To determine the role of lipoprotein lipase (LPL) and hepatic lipase (HL) in regulating these particles, we measured these enzyme activities in 28 healthy subjects with well-controlled Type 1 diabetes, and studied their relationship with Lp(A-I) and Lp(A-I,A-II). LPL was positively correlated with the apolipoprotein A-I (apoA-I), cholesterol, and phospholipid mass in total Lp(A-I), and with the apoA-I in large Lp(A-I) (r >or= 0.58, P >or= 0.001). HL was negatively correlated with all the above Lp(A-I) parameters plus Lp(A-I) triglyceride (r >or= -0.53, P or= 0.50, P 相似文献   

Podocyte-secreted angiopoietin-like-4 mediates proteinuria in glucocorticoid-sensitive nephrotic syndrome

The main manifestations of nephrotic syndrome include proteinuria, hypoalbuminemia, edema, hyperlipidemia and lipiduria. Common causes of nephrotic syndrome are diabetic nephropathy, minimal change disease (MCD), focal and segmental glomerulosclerosis (FSGS) and membranous nephropathy. Among the primary glomerular diseases, MCD is usually sensitive to glucocorticoid treatment, whereas the other diseases show variable responses. Despite the identification of key structural proteins in the glomerular capillary loop which may contribute to defects in ultrafiltration, many of the disease mechanisms of nephrotic syndrome remain unresolved. In this study, we show that the glomerular expression of angiopoietin-like-4 (Angptl4), a secreted glycoprotein, is glucocorticoid sensitive and is highly upregulated in the serum and in podocytes in experimental models of MCD and in the human disease. Podocyte-specific transgenic overexpression of Angptl4 (NPHS2-Angptl4) in rats induced nephrotic-range, and selective, proteinuria (over 500-fold increase in albuminuria), loss of glomerular basement membrane (GBM) charge and foot process effacement, whereas transgenic expression specifically in the adipose tissue (aP2-Angptl4) resulted in increased circulating Angptl4, but no proteinuria. Angptl4(-/-) mice that were injected with lipopolysaccharide (LPS) or nephritogenic antisera developed markedly less proteinuria than did control mice. Angptl4 secreted from podocytes in some forms of nephrotic syndrome lacks normal sialylation. When we fed the sialic acid precursor N-acetyl-D-mannosamine (ManNAc) to NPHS2-Angptl4 transgenic rats it increased the sialylation of Angptl4 and decreased albuminuria by more than 40%. These results suggest that podocyte-secreted Angptl4 has a key role in nephrotic syndrome.     

Proteinuria increases oxylipid concentrations in VLDL and HDL but not LDL particles in the rat

We previously established that proteinuria alters the apolipoprotein content of lipoproteins. This study was conducted to establish whether proteinuria also alters the concentrations of oxidized lipids within lipoprotein density fractions. To this end, we induced passive Heymann nephritis in Sprague Dawley rats and measured an array of alkaline-stable oxylipids in VLDL, LDL, and HDL particles. Proteinuria increased the total oxylipid amounts in the HDL and VLDL fractions. More importantly, these levels were increased when expressed per unit lipoprotein protein, indicating that the oxidized lipid load per particle was increased. Epoxides and diols increased approximately 2-fold in HDL and approximately 5-fold in VLDL, whereas LDL showed approximately 2-fold decreases. The hydroxyeicosatetraenoic acids and hydroxyoctadecadienoic acids (HODEs) increased >4-fold in HDL and >20-fold in VLDL, whereas LDL showed approximately 2-fold decreases in the HODEs. Therefore, nephrotic syndrome alters the lipoprotein oxylipid composition independently of an increase in total lipoprotein levels. These proteinuria-induced changes may be associated with the cardiovascular risk of lipoprotein oxidation.     

Effect of copper deficiency on the composition of three high-density lipoprotein subclasses as separated by heparin-affinity chromatography

Copper deficiency in rats produces a hypercholesterolemia with a marked increase in HDL fraction. This study investigated changes in the plasma distribution and composition of HDL subclasses as affected by copper deficiency. Plasma HDL were separated into the following three subclasses by heparin-affinity chromatography: HDL containing no apo E but high in apo A-I (HDL-E0); HDL with an intermediate level of apo E (HDL-E1); and HDL highly enriched in apo E but low in apo A-I (HDL-E2). The compositional analysis showed that the hypercholesterolemia observed in copper-deficient rats was due specifically to an increase in plasma cholesterol carried by HDL-E0. Copper deficiency did not alter the percent distribution of apo A-I in HDL-E0, but lowered the apo A-I content in HDL-E1 and HDL-E2, with an increase in apo E in these subclasses. The total plasma concentration of apo A-I was, however, significantly elevated in Cu-deficient rats, which was attributable to an increase in the total number of circulating HDL particles. No difference was noted between Cu-deficient and control groups in the distribution of free cholesterol or the ratio of free cholesterol to esterified cholesterol in any of the HDL subclasses. The present results and earlier observations suggest that copper deficiency may produce a defect in the plasma clearance or tissue uptake of the HDL subclass high in apo A-I but devoid of apo E (HDL-E0), which may be mediated by the specific apo A-I receptor or non-endocytotic transfer of HDL-E0 cholesterol to the liver. Such metabolic defects may partly explain the simultaneous increases in both plasma HDL cholesterol and apo A-I and altered cholesterol homeostasis observed in copper deficiency.     

Increased arterial smooth muscle Ca2+ signaling, vasoconstriction, and myogenic reactivity in Milan hypertensive rats

The Milan hypertensive strain (MHS) rats are a genetic model of hypertension with adducin gene polymorphisms linked to enhanced renal tubular Na(+) reabsorption. Recently we demonstrated that Ca(2+) signaling is augmented in freshly isolated mesenteric artery myocytes from MHS rats. This is associated with greatly enhanced expression of Na(+)/Ca(2+) exchanger-1 (NCX1), C-type transient receptor potential (TRPC6) protein, and sarco(endo)plasmic reticulum Ca(2+)-ATPase (SERCA2) compared with arteries from Milan normotensive strain (MNS) rats. Here, we test the hypothesis that the enhanced Ca(2+) signaling in MHS arterial smooth muscle is directly reflected in augmented vasoconstriction [myogenic and phenylephrine (PE)-evoked responses] in isolated mesenteric small arteries. Systolic blood pressure was higher in MHS (145 ± 1 mmHg) than in MNS (112 ± 1 mmHg; P < 0.001; n = 16 each) rats. Pressurized mesenteric resistance arteries from MHS rats had significantly augmented myogenic tone and reactivity and enhanced constriction to low-dose (1-100 nM) PE. Isolated MHS arterial myocytes exhibited approximately twofold increased peak Ca(2+) signals in response to 5 μM PE or ATP in the absence and presence of extracellular Ca(2+). These augmented responses are consistent with increased vasoconstrictor-evoked sarcoplasmic reticulum (SR) Ca(2+) release and increased Ca(2+) entry, respectively. The increased SR Ca(2+) release correlates with a doubling of inositol 1,4,5-trisphosphate receptor type 1 and tripling of SERCA2 expression. Pressurized MHS arteries also exhibited a ～70% increase in 100 nM ouabain-induced vasoconstriction compared with MNS arteries. These functional alterations reveal that, in a genetic model of hypertension linked to renal dysfunction, multiple mechanisms within the arterial myocytes contribute to enhanced Ca(2+) signaling and myogenic and vasoconstrictor-induced arterial constriction. MHS rats have elevated plasma levels of endogenous ouabain, which may initiate the protein upregulation and enhanced Ca(2+) signaling. These molecular and functional changes provide a mechanism for the increased peripheral vascular resistance (whole body autoregulation) that underlies the sustained hypertension.