Scavenger receptor BI facilitates hepatic very low density lipoprotein production in mice

Scavenger receptor BI (SR-BI) is a selective uptake receptor for HDL cholesterol but is also involved in the catabolism of apolipoprotein (apo)B-containing lipoproteins. However, plasma levels of apoB-containing lipoproteins increase following hepatic SR-BI overexpression, suggesting that SR-BI not solely mediates their catabolism. We therefore tested the hypothesis that hepatic SR-BI impacts on VLDL production. On day 7 following adenovirus (Ad)-mediated overexpression of SR-BI, VLDL-triglyceride and VLDL-apoB production rates were significantly increased (P < 0.001), whereas VLDL production was significantly lower in SR-BI knockout mice compared with controls (P < 0.05). In mice injected with AdSR-BI, hepatic cholesterol content increased (P < 0.001), microsomal triglyceride transfer protein activity was higher (P < 0.01) and expression of sterol-regulatory element binding protein (SREBP)2 and its target genes was decreased (P < 0.01). Conversely, in SR-BI knockout mice, microsomal triglyceride transfer protein activity was lower and expression of SREBP2 target genes was increased (P < 0.01). Finally, we demonstrate in vitro in isolated primary hepatocytes as well as in vivo that cholesterol derived from HDL and taken up via SR-BI into the liver can be resecreted within VLDL. These data indicate that hepatic SR-BI expression is linked to VLDL production, and within liver, a metabolic shunt might exist that delivers HDL cholesterol, at least in part, to a pool from which cholesterol is mobilized for VLDL production. These results might have implications for HDL-based therapies against atherosclerotic cardiovascular disease, especially with SR-BI as target.     

Hepatic cholesterol and bile acid metabolism and intestinal cholesterol absorption in scavenger receptor class B type I-deficient mice

The scavenger receptor class B type I (SR-BI), which is expressed in the liver and intestine, plays a critical role in cholesterol metabolism in rodents. While hepatic SR-BI expression controls high density lipoprotein (HDL) cholesterol metabolism, intestinal SR-BI has been proposed to facilitate cholesterol absorption. To evaluate further the relevance of SR-BI in the enterohepatic circulation of cholesterol and bile salts, we studied biliary lipid secretion, hepatic sterol content and synthesis, bile acid metabolism, fecal neutral sterol excretion, and intestinal cholesterol absorption in SR-BI knockout mice. SR-BI deficiency selectively impaired biliary cholesterol secretion, without concomitant changes in either biliary bile acid or phospholipid secretion. Hepatic total and unesterified cholesterol contents were slightly increased in SR-BI-deficient mice, while sterol synthesis was not significantly changed. Bile acid pool size and composition, as well as fecal bile acid excretion, were not altered in SR-BI knockout mice. Intestinal cholesterol absorption was somewhat increased and fecal sterol excretion was slightly decreased in SR-BI knockout mice relative to controls. These findings establish the critical role of hepatic SR-BI expression in selectively controlling the utilization of HDL cholesterol for biliary secretion. In contrast, SR-BI expression is not essential for intestinal cholesterol absorption.     

Secretory phospholipase A2 increases SR-BI-mediated selective uptake from HDL but not biliary cholesterol secretion

High density lipoprotein cholesterol represents a major source of biliary cholesterol. Secretory phospholipase A2 (sPLA2) is an acute phase enzyme mediating decreased plasma HDL cholesterol levels. Clinical studies reported a link between increased sPLA2 expression and the presence of cholesterol gallstones. The aim of our study was to investigate whether the overexpression of human sPLA2 in transgenic mice affects biliary cholesterol secretion and gallstone formation. Liver weight (P < 0.01) and hepatic cholesterol content (P < 0.01) were significantly increased in sPLA2 transgenic mice compared with controls as a result of increased scavenger receptor class B type I (SR-BI)-mediated hepatic selective uptake of HDL cholesterol (P < 0.01), whereas hepatic SR-BI expression remained unchanged. However, biliary cholesterol secretion as well as fecal neutral sterol and fecal bile salt excretion remained unchanged in sPLA2 transgenic mice. Furthermore, gallstone prevalence in response to a lithogenic diet was identical in both groups. These data demonstrate that i) increased flux of cholesterol from HDL into the liver via SR-BI as a result of phospholipase modification of the HDL particle translates neither into increased biliary and fecal sterol output nor into increased gallstone formation, and ii) increased sPLA2 expression in patients with cholesterol gallstones might be a consequence rather than the underlying cause of the disease.     

Differential impact of hepatic deficiency and total body inhibition of MTP on cholesterol metabolism and RCT in mice

Because apoB-containing lipoproteins are pro-atherogenic and their secretion by liver and intestine largely depends on microsomal triglyceride transfer protein (MTP) activity, MTP inhibition strategies are actively pursued. How decreasing the secretion of apoB-containing lipoproteins affects intracellular rerouting of cholesterol is unclear. Therefore, the aim of the present study was to determine the effects of reducing either systemic or liver-specific MTP activity on cholesterol metabolism and reverse cholesterol transport (RCT) using a pharmacological MTP inhibitor or a genetic model, respectively. Plasma total cholesterol and triglyceride levels were decreased in both MTP inhibitor-treated and liver-specific MTP knockout (L-Mttp^−/−) mice (each P < 0.001). With both inhibition approaches, hepatic cholesterol as well as triglyceride content was consistently increased (each P < 0.001), while biliary cholesterol and bile acid secretion remained unchanged. A small but significant decrease in fecal bile acid excretion was observed in inhibitor-treated mice (P < 0.05), whereas fecal neutral sterol excretion was substantially increased by 75% (P < 0.001), conceivably due to decreased intestinal absorption. In contrast, in L-Mttp^−/− mice both fecal neutral sterol and bile acid excretion remained unchanged. However, while total RCT increased in inhibitor-treated mice (P < 0.01), it surprisingly decreased in L-Mttp^−/− mice (P < 0.05). These data demonstrate that: i) pharmacological MTP inhibition increases RCT, an effect that might provide additional clinical benefit of MTP inhibitors; and ii) decreasing hepatic MTP decreases RCT, pointing toward a potential contribution of hepatocyte-derived VLDLs to RCT.     

Anacetrapib reduces (V)LDL cholesterol by inhibition of CETP activity and reduction of plasma PCSK9

Recently, we showed in APOE*3-Leiden cholesteryl ester transfer protein (E3L.CETP) mice that anacetrapib attenuated atherosclerosis development by reducing (V)LDL cholesterol [(V)LDL-C] rather than by raising HDL cholesterol. Here, we investigated the mechanism by which anacetrapib reduces (V)LDL-C and whether this effect was dependent on the inhibition of CETP. E3L.CETP mice were fed a Western-type diet alone or supplemented with anacetrapib (30 mg/kg body weight per day). Microarray analyses of livers revealed downregulation of the cholesterol biosynthesis pathway (P < 0.001) and predicted downregulation of pathways controlled by sterol regulatory element-binding proteins 1 and 2 (z-scores −2.56 and −2.90, respectively; both P < 0.001). These data suggest increased supply of cholesterol to the liver. We found that hepatic proprotein convertase subtilisin/kexin type 9 (Pcsk9) expression was decreased (−28%, P < 0.01), accompanied by decreased plasma PCSK9 levels (−47%, P < 0.001) and increased hepatic LDL receptor (LDLr) content (+64%, P < 0.01). Consistent with this, anacetrapib increased the clearance and hepatic uptake (+25%, P < 0.001) of [¹⁴C]cholesteryl oleate-labeled VLDL-mimicking particles. In E3L mice that do not express CETP, anacetrapib still decreased (V)LDL-C and plasma PCSK9 levels, indicating that these effects were independent of CETP inhibition. We conclude that anacetrapib reduces (V)LDL-C by two mechanisms: 1) inhibition of CETP activity, resulting in remodeled VLDL particles that are more susceptible to hepatic uptake; and 2) a CETP-independent reduction of plasma PCSK9 levels that has the potential to increase LDLr-mediated hepatic remnant clearance.     

Postprandial lipemia enhances the capacity of large HDL2 particles to mediate free cholesterol efflux via SR-BI and ABCG1 pathways in type IIB hyperlipidemia

Lipid and cholesterol metabolism in the postprandial phase is associated with both quantitative and qualitative remodeling of HDL particle subspecies that may influence their anti-atherogenic functions in the reverse cholesterol transport pathway. We evaluated the capacity of whole plasma or isolated HDL particles to mediate cellular free cholesterol (FC) efflux, cholesteryl ester transfer protein (CETP)-mediated cholesteryl ester (CE) transfer, and selective hepatic CE uptake during the postprandial phase in subjects displaying type IIB hyperlipidemia (n = 16). Postprandial, large HDL2 displayed an enhanced capacity to mediate FC efflux via both scavenger receptor class B type I (SR-BI)-dependent (+12%; P < 0.02) and ATP binding cassette transporter G1 (ABCG1)-dependent (+31%; P < 0.008) pathways in in vitro cell systems. In addition, the capacity of whole postprandial plasma (4 h and 8 h postprandially) to mediate cellular FC efflux via the ABCA1-dependent pathway was significantly increased (+19%; P < 0.0003). Concomitantly, postprandial lipemia was associated with elevated endogenous CE transfer rates from HDL2 to apoB lipoproteins and with attenuated capacity (−17%; P < 0.02) of total HDL to deliver CE to hepatic cells. Postprandial lipemia enhanced SR-BI and ABCG1-dependent efflux to large HDL2 particles. However, postprandial lipemia is equally associated with deleterious features by enhancing formation of CE-enriched, triglyceride-rich lipoprotein particles through the action of CETP and by reducing the direct return of HDL-CE to the liver.     

ApoE promotes hepatic selective uptake but not RCT due to increased ABCA1-mediated cholesterol efflux to plasma

ApoE plays an important role in lipoprotein metabolism. This study investigated the effects of adenovirus-mediated human apoE overexpression (AdhApoE3) on sterol metabolism and in vivo reverse cholesterol transport (RCT). In wild-type mice, AdhApoE3 resulted in decreased HDL cholesterol levels and a shift toward larger HDL in plasma, whereas hepatic cholesterol content increased (P < 0.05). These effects were dependent on scavenger receptor class B type I (SR-BI) as confirmed using SR-BI-deficient mice. Kinetic studies demonstrated increased plasma HDL cholesteryl ester catabolic rates (P < 0.05) and higher hepatic selective uptake of HDL cholesteryl esters in AdhApoE3-injected wild-type mice (P < 0.01). However, biliary and fecal sterol output as well as in vivo macrophage-to-feces RCT studied with (3)H-cholesterol-loaded mouse macrophage foam cells remained unchanged upon human apoE overexpression. Similar results were obtained using hApoE3 overexpression in human CETP transgenic mice. However, blocking ABCA1-mediated cholesterol efflux from hepatocytes in AdhApoE3-injected mice using probucol increased biliary cholesterol secretion (P < 0.05), fecal neutral sterol excretion (P < 0.05), and in vivo RCT (P < 0.01), specifically within neutral sterols. These combined data demonstrate that systemic apoE overexpression increases i) SR-BI-mediated selective uptake into the liver and ii) ABCA1-mediated efflux of RCT-relevant cholesterol from hepatocytes back to the plasma compartment, thereby resulting in unchanged fecal mass sterol excretion and overall in vivo RCT.     

Susceptibility to murine cholesterol gallstone formation is not affected by partial disruption of the HDL receptor SR-BI

High density lipoprotein (HDL) promotes reverse cholesterol transport from peripheral tissues to the liver where its cholesterol is secreted preferentially into bile. The scavenger receptor class B type I (SR-BI) is believed to play a pivotal role in unloading HDL cholesterol and its ester to hepatocytes. Here, using male SR-BI "att" mice with a dysfunctional mutation in the Sr-b1 promoter, we studied whether approximately 50% of normal SR-BI expression influences gallstone susceptibility in these mice fed a lithogenic diet containing 1% cholesterol, 0.5% cholic acid and 15% butterfat. Our results showed that the disruption of SR-BI expression reduced cholesterol secretion by 37% in the chow-fed state and 10% on the lithogenic diet, and while delaying incidence slightly, did not influence cumulative susceptibility to cholesterol gallstones. The lithogenic diet induced marked increases in biliary cholesterol and phospholipid secretion rates but not of bile salts. Basal expression of hepatic SR-BI protein was dissimilar in both wild-type and SR-BI mice, and remained unaltered in response to the lithogenic diet. By two independent dual isotope methods, intestinal cholesterol absorption was unimpaired by attenuation of the SR-BI which also displays low-density expression on small intestinal enterocytes. We conclude that although HDL cholesterol is a principal source of biliary cholesterol in the basal state, uptake of cholesterol from chylomicron remnants appears to be the major contributor to biliary cholesterol hypersecretion during diet-induced cholelithogenesis in the mouse.     

SR-BI undergoes cholesterol-stimulated transcytosis to the bile canaliculus in polarized WIF-B cells

The scavenger receptor BI (SR-BI) is highly expressed in hepatocytes, where it mediates the uptake of lipoprotein cholesterol, promotes the secretion of cholesterol into bile, and protects against atherosclerosis. Despite a strong correlation between the hepatic expression of SR-BI and biliary cholesterol secretion, little is known about SR-BI trafficking in response to changes in sterol availability. Using a well characterized polarized hepatocyte cell model, WIF-B, we determine that in cholesterol-depleted cells, SR-BI is extensively located on the basolateral surface, where it can access circulating lipoproteins. However, in response to cholesterol loading, SR-BI undergoes a slow transcytosis to the apical bile canaliculus independently of lipoprotein binding and new protein synthesis. In cholesterol-replete WIF-B cells, SR-BI that resides on the canalicular membrane is dynamically associated with defined microdomains and does not rapidly recycle to and from the subapical or basolateral regions. Taken together, these data demonstrate that hepatic SR-BI transcytosis is regulated by cholesterol and suggest that SR-BI has a stationary function on the bile canaliculus.     

Type I diabetes mellitus decreases in vivo macrophage-to-feces reverse cholesterol transport despite increased biliary sterol secretion in mice

Type I diabetes mellitus (T1DM) increases atherosclerotic cardiovascular disease; however, the underlying pathophysiology is still incompletely understood. We investigated whether experimental T1DM impacts HDL-mediated reverse cholesterol transport (RCT). C57BL/6J mice with alloxan-induced T1DM had higher plasma cholesterol levels (P < 0.05), particularly within HDL, and increased hepatic cholesterol content (P < 0.001). T1DM resulted in increased bile flow (2.1-fold; P < 0.05) and biliary secretion of bile acids (BA, 10.5-fold; P < 0.001), phospholipids (4.5-fold; P < 0.001), and cholesterol (5.5-fold; P < 0.05). Hepatic cholesterol synthesis was unaltered, whereas BA synthesis was increased in T1DM (P < 0.001). Mass fecal BA output was significantly higher in T1DM mice (1.5-fold; P < 0.05), fecal neutral sterol excretion did not change due to increased intestinal cholesterol absorption (2.1-fold; P < 0.05). Overall in vivo macrophage-to-feces RCT, using [(3)H]cholesterol-loaded primary mouse macrophage foam cells, was 20% lower in T1DM (P < 0.05), mainly due to reduced tracer excretion within BA (P < 0.05). In vitro experiments revealed unchanged cholesterol efflux toward T1DM HDL, whereas scavenger receptor class BI-mediated selective uptake from T1DM HDL was lower in vitro and in vivo (HDL kinetic experiments) (P < 0.05), conceivably due to increased glycation of HDL-associated proteins (+65%, P < 0.01). In summary, despite higher mass biliary sterol secretion T1DM impairs macrophage-to-feces RCT, mainly by decreasing hepatic selective uptake, a mechanism conceivably contributing to increased cardiovascular disease in T1DM.     

The relationships of markers of cholesterol homeostasis with carotid intima-media thickness

Background

The relationship of cholesterol homeostasis and carotid intima-media thickness (cIMT) is unknown. To address this, we assessed markers of cholesterol homeostasis (serum plant sterols and cholesterol precursor concentrations as surrogate measures of cholesterol absorption and synthesis, respectively) and cIMT in a middle-aged, statin-naive population.

Methods

In this prospective study of primary prevention cIMT was measured by ultrasound in 583 hospital employees aged 25–60 years without prevalent cardiovascular disease or lipid-modifying medication. The serum concentrations of plant sterols (as markers of cholesterol absorption) were measured by gas-liquid chromatography. Lathosterol serum concentrations were quantitated to assess hepatic cholesterol synthesis.

Results

cIMT correlated positively with serum cholesterol (r = 0.22, P<0.0005) and lathosterol-to-cholesterol (r = 0.18, P<0.001). In contrast, plant sterols, as markers of cholesterol absorption, showed a weak negative correlation to cIMT measurements (r = −0.18; P<0.001 for campesterol-to-cholesterol). Stratifying subjects by serum sterol levels, we found that cIMT increased continuously over quintiles of serum cholesterol (P<0.0005) and was positively associated to serum lathosterol-to-cholesterol levels (P = 0.007), on the other hand, plant sterol levels showed a weak negative association to cIMT (P<0.001 for campesterol-to-cholesterol).

Conclusions

In this population without prevalent cardiovascular diseases or lipid-modifying medication, markers of increased endogenous cholesterol synthesis correlated positively with cIMT, while markers of cholesterol absorption showed a weakly negative correlation. These data suggest that not only total serum cholesterol levels but also differences in cholesterol homeostasis are associated with cIMT.     

The Liver-Selective Thyromimetic T-0681 Influences Reverse Cholesterol Transport and Atherosclerosis Development in Mice

Background

Liver-selective thyromimetics have been reported to efficiently reduce plasma cholesterol through the hepatic induction of both, the low-density lipoprotein receptor (LDLr) and the high-density lipoprotein (HDL) receptor; the scavenger receptor class B type I (SR-BI). Here, we investigated the effect of the thyromimetic T-0681 on reverse cholesterol transport (RCT) and atherosclerosis, and studied the underlying mechanisms using different mouse models, including mice lacking LDLr, SR-BI, and apoE, as well as CETP transgenic mice.

Methodology/Principal Findings

T-0681 treatment promoted bile acid production and biliary sterol secretion consistently in the majority of the studied mouse models, which was associated with a marked reduction of plasma cholesterol. Using an assay of macrophage RCT in mice, we found T-0681 to significantly increase fecal excretion of macrophage-derived neutral and acidic sterols. No positive effect on RCT was found in CETP transgenic mice, most likely due to the observed decrease in plasma CETP mass. Studies in SR-BI KO and LDLr KO mice suggested hepatic LDLr to be necessary for the action of T-0681 on lipid metabolism, as the compound did not have any influence on plasma cholesterol levels in mice lacking this receptor. Finally, prolonged treatment with T-0681 reduced the development of atherosclerosis by 60% in apoE KOs on Western type diet. In contrast, at an earlier time-point T-0681 slightly increased small fatty streak lesions, in part due to an impaired macrophage cholesterol efflux capacity, when compared to controls.

Conclusions/Significance

The present results show that liver-selective thyromimetics can promote RCT and that such compounds may protect from atherosclerosis partly through induction of bile acid metabolism and biliary sterol secretion. On-going clinical trials will show whether selective thyromimetics do prevent atherosclerosis also in humans.     

In vivo modulation of HDL phospholipid has opposing effects on SR-BI- and ABCA1-mediated cholesterol efflux

The effects of in vivo modulation of HDL phospholipid (PL) on scavenger receptor class BI (SR-BI)- and ATP binding cassette transporter 1 (ABCA1)-mediated efflux were examined by overexpressing either endothelial lipase (EL) or phosphatidylserine phospholipase (PS-PLA1) in human apolipoprotein A-I (apoA-I) transgenic mice. Overexpression of EL led to large reductions in the serum PL/apoA-I ratio (-60%), total cholesterol (TC; -89%), and HDL cholesterol (-91%). Relative to the serum before overexpression of EL, the efflux potential of the serum via SR-BI decreased by 90% and ABCA1-mediated efflux increased by 63%. In contrast to overexpression of EL, overexpression of PS-PLA1 led to increases in the PL/apoA-I ratio (88%), TC (78%), HDL cholesterol (57%), and HDL size. The efflux potential of the serum increased by 60% via SR-BI and decreased by 57% via ABCA1. There were significant positive correlations between SR-BI-mediated efflux and a number of serum parameters, including PL/apoA-I ratio, PL, TC, free cholesterol (FC), and HDL cholesterol. In striking contrast, the same correlations were seen with ABCA1-mediated efflux, but the relationships were inverse. In summary, in vivo modulation of HDL PL content affects ABCA1- and SR-BI-mediated efflux in a reciprocal manner. These findings indicate that the type of lipase acting on HDL in vivo will determine which FC efflux pathway the HDL serves. Additionally, the extent of lipolysis will determine the efficiency of FC removal via this pathway.     

Influence of the HDL receptor SR-BI on atherosclerosis

The scavenger receptor class B, type I (SR-BI) is an HDL receptor that mediates selective cholesterol uptake from HDL to cells. In rodents, SR-BI has a critical influence on plasma HDL-cholesterol concentration and structure, the delivery of cholesterol to steroidogenic tissues, female fertility, and biliary cholesterol concentration. SR-BI can also serve as a receptor for non-HDL lipoproteins and appears to play an important role in reverse cholesterol transport. Recent studies involving the manipulation of SR-BI expression in mice, either using adenovirus-mediated or transgenic hepatic overexpression or using homologous recombination for complete functional ablation, indicate that the expression of SR-BI protects against atherosclerosis. If SR-BI has a similar activity in humans, it may become an attractive target for therapeutic intervention.     

Liver-specific transgenic expression of cholesteryl ester hydrolase reduces atherosclerosis in Ldlr−/− mice

The liver plays a central role in the final elimination of cholesterol from the body either as bile acids or as free cholesterol (FC), and lipoprotein-derived cholesterol is the major source of total biliary cholesterol. HDL is the major lipoprotein responsible for removal and transport of cholesterol, mainly as cholesteryl esters (CEs), from the peripheral tissues to the liver. While HDL-FC is rapidly secreted into bile, the fate of HDL-CE remains unclear. We have earlier demonstrated the role of human CE hydrolase (CEH, CES1) in hepatic hydrolysis of HDL-CE and increasing bile acid synthesis, a process dependent on scavenger receptor BI expression. In the present study, we examined the hypothesis that by enhancing the elimination of HDL-CE into bile/feces, liver-specific transgenic expression of CEH will be anti-atherogenic. Increased CEH expression in the liver significantly increased the flux of HDL-CE to bile acids. In the LDLR^−/− background, this enhanced elimination of cholesterol led to attenuation of diet-induced atherosclerosis with a consistent increase in fecal sterol secretion primarily as bile acids. Taken together with the observed reduction in atherosclerosis by increasing macrophage CEH-mediated cholesterol efflux, these studies establish CEH as an important regulator in enhancing cholesterol elimination and also as an anti-atherogenic target.     

Hepatic cholesterol transport from plasma into bile: implications for gallstone disease

PURPOSE OF REVIEW: The transhepatic traffic of cholesterol from plasma lipoproteins into the bile is critical for overall cholesterol homeostasis and its alterations may lead to cholesterol gallstone formation. This review summarizes recent progress in understanding the key hepatic cholesterol metabolism-related proteins and pathways that influence biliary secretion of cholesterol. RECENT FINDINGS: In cholesterol-fed apolipoprotein E knockout mice, the availability of dietary cholesterol for biliary disposal is decreased and diet-induced gallstone formation is impaired. Scavenger receptor class B type I is relevant for cholesterol transport from plasma HDL into the bile in chow-fed mice, however its expression is not critical for biliary cholesterol secretion and gallstone formation in lithogenic diet-fed mice. Intrahepatic cholesterol transport proteins (e.g. sterol carrier protein-2, Niemann Pick type C-1 protein) also determine liver cholesterol available for biliary secretion in mice. Genetic manipulation of canalicular ATP-binding cassette transporter G5 and G8 expression in mice has established their essential role for biliary cholesterol secretion. SUMMARY: Recent studies have underscored that different proteins involved in hepatic cholesterol transport regulate the availability of cholesterol for biliary secretion. These advances may provide new avenues for prevention and treatment of various disease conditions linked to abnormal cholesterol metabolism.     

Overexpression of sterol carrier protein-2 differentially alters hepatic cholesterol accumulation in cholesterol-fed mice

Although in vitro studies suggest a role for sterol carrier protein-2 (SCP-2) in cholesterol trafficking and metabolism, the physiological significance of these observations remains unclear. This issue was addressed by examining the response of mice overexpressing physiologically relevant levels of SCP-2 to a cholesterol-rich diet. While neither SCP-2 overexpression nor cholesterol-rich diet altered food consumption, increased weight gain, hepatic lipid, and bile acid accumulation were observed in wild-type mice fed the cholesterol-rich diet. SCP-2 overexpression further exacerbated hepatic lipid accumulation in cholesterol-fed females (cholesterol/cholesteryl esters) and males (cholesterol/cholesteryl esters and triacyglycerol). Primarily in female mice, hepatic cholesterol accumulation induced by SCP-2 overexpression was associated with increased levels of LDL-receptor, HDL-receptor scavenger receptor-B1 (SR-B1) (as well as PDZK1 and/or membrane-associated protein 17 kDa), SCP-2, liver fatty acid binding protein (L-FABP), and 3α-hydroxysteroid dehydrogenase, without alteration of other proteins involved in cholesterol uptake (caveolin), esterification (ACAT2), efflux (ATP binding cassette A-1 receptor, ABCG5/8, and apolipoprotein A1), or oxidation/transport of bile salts (cholesterol 7α-hydroxylase, sterol 27α-hydroxylase, Na⁺/taurocholate cotransporter, Oatp1a1, and Oatp1a4). The effects of SCP-2 overexpression and cholesterol-rich diet was downregulation of proteins involved in cholesterol transport (L-FABP and SR-B1), cholesterol synthesis (related to sterol regulatory element binding protein 2 and HMG-CoA reductase), and bile acid oxidation/transport (via Oapt1a1, Oatp1a4, and SCP-x). Levels of serum and hepatic bile acids were decreased in cholesterol-fed SCP-2 overexpression mice, especially in females, while the total bile acid pool was minimally affected. Taken together, these findings support an important role for SCP-2 in hepatic cholesterol homeostasis.     

Hepatic overexpression of cholesteryl ester hydrolase enhances cholesterol elimination and in vivo reverse cholesterol transport

Neutral cholesteryl ester hydrolase (CEH)-mediated hydrolysis of cellular cholesteryl esters (CEs) is required not only to generate free cholesterol (FC) for efflux from macrophages but also to release FC from lipoprotein-delivered CE in the liver for bile acid synthesis or direct secretion into the bile. We hypothesized that hepatic expression of CEH would regulate the hydrolysis of lipoprotein-derived CE and enhance reverse cholesterol transport (RCT). Adenoviral-mediated CEH overexpression led to a significant increase in bile acid output. To assess the role of hepatic CEH in promoting flux of cholesterol from macrophages to feces, cholesterol-loaded and [(3)H]cholesterol-labeled J774 macrophages were injected intraperitoneally into mice and the appearance of [(3)H]cholesterol in gallbladder bile and feces over 48 h was quantified. Mice overexpressing CEH had significantly higher [(3)H]cholesterol radiolabel in bile and feces, and it was associated with bile acids. This CEH-mediated increased movement of [(3)H]cholesterol from macrophages to bile acids and feces was significantly attenuated in SR-BI(-/-) mice. These studies demonstrate that similar to macrophage CEH that rate-limits the first step, hepatic CEH regulates the last step of RCT by promoting the flux of cholesterol entering the liver via SR-BI and increasing hepatic bile acid output.