High density lipoprotein metabolism in low density lipoprotein receptor-deficient mice.

The LDL receptor (LDLR) and scavenger receptor class B type I (SR-BI) play physiological roles in LDL and HDL metabolism in vivo. In this study, we explored HDL metabolism in LDLR-deficient mice in comparison with WT littermates. Murine HDL was radiolabeled in the protein ((125)I) and in the cholesteryl ester (CE) moiety ([(3)H]). The metabolism of (125)I-/[(3)H]HDL was investigated in plasma and in tissues of mice and in murine hepatocytes. In WT mice, liver and adrenals selectively take up HDL-associated CE ([(3)H]). In contrast, in LDLR(-/-) mice, selective HDL CE uptake is significantly reduced in liver and adrenals. In hepatocytes isolated from LDLR(-/-) mice, selective HDL CE uptake is substantially diminished compared with WT liver cells. Hepatic and adrenal protein expression of lipoprotein receptors SR-BI, cluster of differentiation 36 (CD36), and LDL receptor-related protein 1 (LRP1) was analyzed by immunoblots. The respective protein levels were identical both in hepatic and adrenal membranes prepared from WT or from LDLR(-/-) mice. In summary, an LDLR deficiency substantially decreases selective HDL CE uptake by liver and adrenals. This decrease is independent from regulation of receptor proteins like SR-BI, CD36, and LRP1. Thus, LDLR expression has a substantial impact on both HDL and LDL metabolism in mice.

Scavenger receptor CD36 mediates uptake of high density lipoproteins in mice and by cultured cells.

The mechanisms of HDL-mediated cholesterol transport from peripheral tissues to the liver are incompletely defined. Here the function of scavenger receptor cluster of differentiation 36 (CD36) for HDL uptake by the liver was investigated. CD36 knockout (KO) mice, which were the model, have a 37% increase (P = 0.008) of plasma HDL cholesterol compared with wild-type (WT) littermates. To explore the mechanism of this increase, HDL metabolism was investigated with HDL radiolabeled in the apolipoprotein (¹²5I) and cholesteryl ester (CE, [³H]) moiety. Liver uptake of [³H] and ¹²5I from HDL decreased in CD36 KO mice and the difference, i. e. hepatic selective CE uptake ([³H]¹²5I), declined (-33%, P = 0.0003) in CD36 KO compared with WT mice. Hepatic HDL holo-particle uptake (¹²5I) decreased (-29%, P = 0.0038) in CD36 KO mice. In vitro, uptake of ¹²5I-/[³H]HDL by primary liver cells from WT or CD36 KO mice revealed a diminished HDL uptake in CD36-deficient hepatocytes. Adenovirus-mediated expression of CD36 in cells induced an increase in selective CE uptake from HDL and a stimulation of holo-particle internalization. In conclusion, CD36 plays a role in HDL uptake in mice and by cultured cells. A physiologic function of CD36 in HDL metabolism in vivo is suggested.

Hepatic ATP-binding cassette transporter A1 is a key molecule in high-density lipoprotein cholesteryl ester metabolism in mice.

OBJECTIVE: Mutations in ATP-binding cassette transporter A1 (ABCA1), the cellular lipid transport molecule mutated in Tangier disease, result in the rapid turnover of high-density lipoprotein (HDL)-associated apolipoproteins that presumably are cleared by the kidneys. However, the role of ABCA1 in the liver for HDL apolipoprotein and cholesteryl ester (CE) catabolism in vivo is unknown. METHODS AND RESULTS: Murine HDL was radiolabeled with 125I in its apolipoprotein and with [3H]cholesteryl oleyl ether in its CE moiety. HDL protein and lipid metabolism in plasma and HDL uptake by tissues were investigated in ABCA1-overexpressing bacterial artificial chromosome (BAC)-transgenic (BAC+) mice and in mice harboring deletions of total (ABCA1-/-) and liver-specific ABCA1 (ABCA1(-L/-L)). In BAC+ mice with elevated ABCA1 expression, fractional catabolic rates (FCRs) of both the protein and the lipid tracer were significantly decreased in plasma and in the liver, yielding a diminished hepatic selective CE uptake from HDL. In contrast, ABCA1-/- or ABCA1(-L/-L) mice had significantly increased plasma and liver FCRs for both HDL tracers. An ABCA1 deficiency was associated with increased selective HDL CE uptake by the liver under all experimental conditions. CONCLUSIONS: Hepatic ABCA1 has a critical role for HDL catabolism in plasma and for HDL selective CE uptake by the liver.

Scavenger receptor class B type I mediates the selective uptake of high-density lipoprotein-associated cholesteryl ester by the liver in mice.

OBJECTIVE: High-density lipoprotein (HDL) cholesteryl esters (CE) are taken up by liver and adrenals selectively, ie, independent from particle internalization. Class B type I scavenger receptor (SR-BI) mediates this uptake in vitro. The role of SR-BI in HDL metabolism was explored in mice. METHODS AND RESULTS: Mice with a mutation in the SR-BI gene (SR-BI KO) and wild-type (WT) littermates were used. Mutants had increased HDL cholesterol. HDL was labeled with 125I (protein) and [3H] (CE). After HDL injection, blood samples were drawn and finally the mice were euthanized. In WT, the plasma decay of HDL-associated [3H] is faster compared with 125I and this represents whole-body selective CE uptake. In SR-BI KO, the decay of both tracers is similar, yielding no selective CE removal. In WT liver and adrenals, uptake of [3H] is higher than 125I, showing selective uptake. In SR-BI KO, liver uptake of [3H] and 125I are similar, proposing no selective HDL CE uptake. In SR-BI KO adrenals, selective uptake is reduced; however, even in the absence of SR-BI, this uptake is detected using WT-HDL. CONCLUSIONS: SR-BI mediates selective uptake of HDL CE by the liver. In adrenals, an alternative mechanism or mechanisms can play a role in selective CE uptake.

Scavenger receptor BI (SR-BI) mediates a higher selective cholesteryl ester uptake from LpA-I compared with LpA-I:A-II lipoprotein particles.

Scavenger receptor class B, type I (SR-BI) mediates the selective uptake of high-density lipoprotein- (HDL-) associated cholesteryl esters (CE), i.e. lipid uptake independent from HDL holo-particle internalisation. This pathway contributes to the HDL-mediated CE delivery to the liver. From human plasma HDL, two major lipoprotein subfractions can be isolated: one contains apolipoprotein (apo) A-I and apo A-II (LpA-I:A-II) as dominant protein components, whereas in the other population apo A-II is absent (LpA-I). In this investigation the role of SR-BI in selective CE uptake from HDL, LpA-I and LpA-I:A-II was explored. HDL(3) (d=1.125-1.21 g/ml), LpA-I and LpA-I:A-II were isolated from human plasma and radiolabeled in the protein (125I) as well as in the CE moiety ([3H]). Baby hamster kidney (BHK) cells were stably transfected with the full-length human SR-BI cDNA and these cells demonstrate SR-BI expression in immunoblots. In contrast, no SR-BI protein was detectable in control BHK cells (vector). To investigate lipoprotein uptake, cells incubated (37 degrees C, 4 h) in medium containing radiolabeled HDL(3), LpA-I or LpA-I:A-II and finally cellular tracer content was determined. For both types of BHK cells, the rate of apparent lipoprotein particle uptake according to the lipid tracer ([3H]) was in substantial excess over that due to the protein tracer (125I) demonstrating selective CE uptake ([3H]-(125)I) from HDL(3), LpA-I and LpA-I:A-II. SR-BI expression increased cellular selective CE uptake from labeled HDL(3) up to 24-fold. In BHK cells without SR-BI expression, selective CE uptake was higher from LpA-I compared with LpA-I:A-II. Analogously, in BHK cells with SR-BI expression, the rate of selective CE uptake was higher from LpA-I compared with LpA-I:A-II. In summary, SR-BI significantly increases selective CE uptake from HDL(3), LpA-I and LpA-I:A-II. Concerning HDL subfractions, the rate of SR-BI-mediated selective CE uptake is greater from LpA-I compared with LpA-I:A-II and this result suggests that SR-BI preferentially facilitates the CE transfer from LpA-I lipoprotein particles to cells.

SR-BI inhibits ABCG1-stimulated net cholesterol efflux from cells to plasma HDL.

This study compares the roles of ABCG1 and scavenger receptor class B type I (SR-BI) singly or together in promoting net cellular cholesterol efflux to plasma HDL containing active LCAT. In transfected cells, SR-BI promoted free cholesterol efflux to HDL, but this was offset by an increased uptake of HDL cholesteryl ester (CE) into cells, resulting in no net efflux. Coexpression of SR-BI with ABCG1 inhibited the ABCG1-mediated net cholesterol efflux to HDL, apparently by promoting the reuptake of CE from medium. However, ABCG1-mediated cholesterol efflux was not altered in cholesterol-loaded, SR-BI-deficient (SR-BI(-/-)) macrophages. Briefly cultured macrophages collected from SR-BI(-/-) mice loaded with acetylated LDL in the peritoneal cavity did exhibit reduced efflux to HDL. However, this was attributable to reduced expression of ABCG1 and ABCA1, likely reflecting increased macrophage cholesterol efflux to apolipoprotein E-enriched HDL during loading in SR-BI(-/-) mice. In conclusion, cellular SR-BI does not promote net cholesterol efflux from cells to plasma HDL containing active LCAT as a result of the reuptake of HDL-CE into cells. Previous findings of increased atherosclerosis in mice transplanted with SR-BI(-/-) bone marrow probably cannot be explained by a defect in macrophage cholesterol efflux.

Selective uptake of HDL cholesteryl esters and cholesterol efflux from mouse peritoneal macrophages independent of SR-BI.

Scavenger receptor class B type I (SR-BI) mediates the selective uptake of HDL cholesteryl esters (CEs) and facilitates the efflux of unesterified cholesterol. SR-BI expression in macrophages presumably plays a role in atherosclerosis. The role of SR-BI for selective CE uptake and cholesterol efflux in macrophages was explored. Macrophages and HDL originated from wild-type (WT) or SR-BI knockout (KO; homozygous) mice. For uptake, macrophages were incubated in medium containing 125I-/3H-labeled HDL. For lipid removal, [3H]cholesterol efflux was analyzed using HDL as acceptor. Selective uptake of HDL CE ([3H]cholesteryl oleyl ether - 125I-tyramine cellobiose) was similar in WT and SR-BI KO macrophages. Radiolabeled SR-BI KO-HDL yielded a lower rate of selective uptake compared with WT-HDL in WT and SR-BI KO macrophages. Cholesterol efflux was similar in WT and SR-BI KO cells using HDL as acceptor. SR-BI KO-HDL more efficiently promoted cholesterol removal compared with WT-HDL from both types of macrophages. Macrophages selectively take up HDL CE independently of SR-BI. Additionally, in macrophages, there is substantial cholesterol efflux that is not mediated by SR-BI. Therefore, SR-BI-independent mechanisms mediate selective CE uptake and cholesterol removal. SR-BI KO-HDL is an inferior donor for selective CE uptake compared with WT-HDL, whereas SR-BI KO-HDL more efficiently promotes cholesterol efflux.

Angiotensin II down-regulates the SR-BI HDL receptor in proximal tubular cells.

BACKGROUND: The kidney plays an important role in the metabolism of lipoproteins, but renal cells are also a target of lipids under pathophysiological conditions contributing to organ damage and progression of disease. The majority of studies has focused on the interaction of renal cells with low-density lipoproteins. Relatively little is known of potential metabolism of high-density lipoproteins (HDL) on renal cells However, diverse pathophysiological situations, such as the nephrotic syndrome and acute renal injury, may be associated with an activated renin-angiotensin system as well as altered renal handling of HDL. Therefore, the present study sought to gain insight into the expression of the HDL receptor scavenger receptor class B type I (SR-BI) in cultured renal cells and a potential regulation by angiotensin II (ANG II). METHODS: Different renal cells lines and primary cultures (proximal tubular and mesangial cells) were screened by western blot for the expression of SR-BI. MCT cells, a mouse proximal tubular cell line, were selected for further studies. SR-BI protein and mRNA expression were determined after treatment with various doses of ANG II in the presence or absence of AT(1)- or AT(2)-receptor blocker. Uptake of HDL-associated cholesteryl ester into MCT cells was determined. Finally, rats were infused intraperitoneally with ANG II for 3-7 days, proximal tubules were isolated by differential centrifugation and SR-BI protein expression was assessed. Results. SR-BI protein was expressed in various primary cultures and permanent renal cell lines. ANG II (10(-10)-10(-6) M) treatment for 24 h induced a significant down-regulation of SR-BI protein and mRNA expression in MCT cells. This suppression was attenuated by an AT(1)-receptor antagonist whereas an AT(2)-blocker was without effect. MCT cells revealed a high selective uptake of HDL cholesteryl ester that was significantly higher than that in syngeneic mesangial cells. ANG II for 24 h significantly reduced this selective HDL cholesteryl ester uptake into MCT, but not mesangial cells. Finally, ANG II- infusion into rats for 3 and 7 days induced a significant decrease of SR-BI protein expression in isolated tubules. CONCLUSIONS: Our data show that ANG II mediates down-regulation of SR-BI expression on proximal tubular cells in vivo and in vitro. However, the effects were small and additional experiments are necessary to confirm these first observations. The attenuated SR-BI expression is functionally relevant and associated with a decrease in cholesteryl ester uptake. ANG II-mediated suppression may contribute to various pathophysiological situations, such as acute tubular injury, the nephrotic syndrome and atherosclerosis.

Hepatic lipase mediates an increase in selective uptake of HDL-associated cholesteryl esters by cells in culture independent from SR-BI.

Scavenger receptor class B type I (SR-BI) mediates the selective uptake of HDL cholesteryl esters (CEs) by the liver. Hepatic lipase (HL) promotes this lipid uptake independent from lipolysis. The role of SR-BI in this HL-mediated increase in selective CE uptake was explored. Baby hamster kidney (BHK) cells were transfected with the SR-BI cDNA yielding cells with SR-BI expression, whereas no SR-BI was detected in control cells. These cells were incubated in medium containing 125I [3H]cholesteryl oleyl ether-labeled HDL3 (d = 1.125-1.21 g/ml) and HL was absent or present. Tetrahydrolipstatin (THL) blocked lipolysis. In control BHK cells and in BHK cells with SR-BI, HDL3 selective CE uptake (3H-125I) was detectable and SR-BI promoted this uptake. In both cell types, HL mediated an increase in selective CE uptake from HDL3. Quantitatively, this HL effect was similar in control BHK cells and in BHK cells with SR-BI. These results suggest that HL promotes selective uptake independent from SR-BI. To investigate the role of cell surface proteoglycans on the HL-mediated HDL3 uptake, proteoglycan deficiency was induced by heparinase digestion. Proteoglycan deficiency decreased the HL-mediated promotion of selective CE uptake. In summary, the stimulating HL effect on HDL selective CE uptake is independent from SR-BI and lipolysis. Proteoglycans are a requisite for the HL action on selective uptake. Results suggest that (a) pathway(s) distinct from SR-BI mediate(s) selective CE uptake from HDL.