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Recently, a number of nuclear receptors have been identified as key regulators of cholesterol homeostasis. Two of these, liver X receptor alpha (LXRalpha) (NR1H3) [1] and ubiquitous receptor (UR) (NR1H2) [1], appear to be involved in cholesterol reverse transport and disposal. LXRalpha null gene mice fail to adapt metabolically to high-cholesterol diets. We have recently shown that some 6alpha-hydroxylated bile acid analogs are selective activators of LXRalpha. In this report, we show that these orally administered LXRalpha agonists have an overall hypolipidemic effect in hypercholesterolemic rats, mice and hamsters, which indicates that in these animal models, endogenous LXRalpha agonist is a limiting factor for induction of cholesterol disposal. Furthermore, in animals, these 6alpha-hydroxylated bile acid analogs exhibit a unique pharmacokinetic profile and do not increase the serum triglyceride level; therefore, they may represent a novel class of therapeutic agents for cholesterol management.

Background: Based on previous work which suggested that biliary crystals may induce inflammation in the gallbladder wall and that inflammation is an early event during the formation of pigment gallstones in the dog, studies were performed examining mucus layer thickness, myeloperoxidase activity, and interleukin-1 (IL-1) activity in the wall of mouse gallbladder during formation and growth of cholesterol gallstones., Methods and Materials: The inflammatory effects of cholesterol gallstones at 2 and 4 weeks were studied in BalB/C mice fed a crushed standard mouse chow with added cholesterol (1.0%) and cholic acid (0.5%). Results were compared to those of normal mice fed standard mouse chow. The presence or absence of crystals and stones was determined by gross and microscopic examination of bile. Myeloperoxidase and IL-1 activity in the gallbladder wall was measured using well-established bioassays. Mucus layer thickness was measured by darkfield microscopy., Results: All mice fed a lithogenic, 1.0% cholesterol/0.5% cholic acid diet developed cholesterol crystals and gallstones at 2 and 6 weeks. No control mice developed either crystals or gallstones. Myeloperoxidase and IL-1 activities, markers of an inflammatory response, increased significantly in the gallbladder of mice with crystals at 2 weeks. Myeloperoxidase activity increased two- to three-fold, and IL-1 activity sevenfold, by 6 weeks. Mucus layer thickness also progressively increased during the 6-week period., Conclusions: It is concluded that inflammation is an early event associated with the appearance of crystals and gallstones in bile.

Supersaturation of bile with cholesterol is a prerequisite of the development of gallstones. With the intention to study the integrated response of enzymes regulating hepatic cholesterol metabolism during gallstone formation we used an established model for the induction of cholesterol gallstone disease in mice. Ten mice were fed on a lithogenic diet containing 10 g cholesterol/kg and 5 g cholic acid/kg for 8 weeks and were compared with ten mice fed on a standard pellet diet. Cholesterol crystals or gallstones developed in 90% of gallbladders in treated mice. The lithogenic diet had an inhibitory effect on the rate-limiting enzyme of cholesterol biosynthesis, hepatic 3-hydroxy-3-methylglutaryl-CoA (HMG-CoA) reductase (EC 1.1.1.88) activity, 39.6 (SEM 2.8) v. 171.0 (SEM 47.3) pmol/min per mg protein. Cholesterol 7 alpha-hydroxylase (EC 1.14.13.17) activity, regulating bile acid synthesis, was decreased by 80%, and this was assumed to be due to cholic acid in the diet. The cholesterol-enriched diet also induced a tenfold increase in cholesterol esterification rate in the liver, i.e. acyl-CoA:cholesterol acyl transferase (ACAT; EC 2.3.1.26) activity. The total, as well as esterified, cholesterol contents of liver homogenates were significantly higher in cholesterol- and cholic acid-treated mice and correlated well with the ACAT activity (rs 0.72 (P < 0.005), and rs 0.68 (P < 0.01) respectively). A significantly higher ACAT activity was obtained in mice given cholesterol and cholic acid even when the enzyme was saturated with exogenous cholesterol, thus indicating an increased amount of the enzyme. The formation of gallstones is dependent on a delicate balance between lithogenic factors (increased absorption of cholesterol and reduced secretion of bile acids) and defence mechanisms (decreased synthesis and increased esterification of cholesterol). In the specific animal model studied here the two defence mechanisms cannot compensate for the increased absorption of cholesterol and the reduced synthesis of bile acids.

The effects of hyodeoxycholic (HDCA) and alpha-hyocholic acids (alpha-HCA), on cholesterol, bile acid and lipoprotein metabolism, were studied in hamsters. The animals were fed a low cholesterol control diet supplemented with 0.1% HDCA or alpha-HCA for 3 weeks. In both treated groups, the LDL-cholesterol concentration was significantly lowered and was associated with a global hypocholesterolemic effect. Moreover, hepatic cholesterol ester storage was reduced and HMGCoA reductase activity was respectively enhanced 13.5-times and 7.7-times in HDCA and alpha-HCA groups compared to controls. In contrast, cholesterol 7 alpha-hydroxylase activity and LDL-receptor activity and mass were not modified. In bile, the cholesterol saturation index was increased 5-fold (HDCA group) and 2-fold (alpha-HCA group) as a consequence of an enlarged proportion of biliary cholesterol. The two 6-hydroxylated bile acids induced an enhanced fecal excretion of neutral sterols (HDCA group: 11.6-times, alpha-HCA group: 3.2-times versus controls) which was consistent with a 59% decrease in intestinal cholesterol absorption in the HDCA group. The major effects due to bile acid treatments were a decrease in LDL-cholesterol concentration, a strong stimulation of hepatic cholesterol biosynthesis and an excessive loss of cholesterol in feces. These perturbations might be the result of the enrichment of bile with hydrophilic bile acids, leading to a limited return of endogenous cholesterol from the intestine to the liver.

The biliary output of bile acids and lipids is tightly coupled. The ability of the natural bile acid glycocholate to trigger biliary lipid secretion was compared with that of the fluorescent bile acid analogue cholyl-lysylfluorescein (cholyl-lys-F). When administered as a 5 min pulse of 2.5 mumol/min to bile acid-depleted rat livers perfused under recycling conditions, glycocholate produced well-defined peaks of phospholipid and cholesterol output, and of bile flow, which were coincident with the peak of bile acid output. Although cholyl-lys-F did trigger biliary lipid secretion, its time course of appearance was delayed and well-defined peaks of output were not observed. However, the increased biliary output of phospholipid and cholesterol was coincident with that of bile acids and, as judged by phospholipid/bile acid and cholesterol/bile acid ratios, cholyl-lys-F was as effective as glycocholate in triggering biliary lipid output. When administered to livers perfused under single pass conditions, perfusate to bile transfer of glycocholate was > 85% at infusion rates of up to 5 mumol/min whereas transfer of cholyl-lys-F showed saturation at infusion rates of > 0.2 mumol/min; the time course of biliary output of both bile acids was similar. Thus, under recycling conditions, cholyl-lys-F not taken up during first pass will be continually represented for transfer to bile, explaining why bile acid and lipid output did not occur as well-defined peaks.

The effect of dietary selenium and vitamin E on plasma total (TC) and high density lipoprotein cholesterol (HDLC) was evaluated in 54 Sprague Dawley rats fed cholesterol/cholic acid enriched diets. Diets 1, 2, and 3 had no added selenium (low Se) and 0 (low), 60 (adequate), and 600 (high) mg/kg dL alpha tocopheryl acetate added respectively. Sodium selenite at 0.2 mg/kg (adequate Se) was added to diets 4, 5, and 6 and at 4.0 mg/kg (toxic Se) to diet 7, 8, and 9 with the same pattern of vitamin E added to the diet as described above. TC and HDLC were measured using the Kodak Ectachem system. Rats in the low and adequate Se groups fed high vitamin E had lower TC values than rats fed lower vitamin E levels but differences were not significant. In the toxic Se groups, rats fed high vitamin E had significantly (p < 0.05) higher plasma TC values than did lower Vitamin E groups. Rats on the high vitamin E diets with low or adequate Se had significantly (p < 0.05) higher mean plasma HDLC values when compared to rats fed low or adequate vitamin E diets. HDLC values for animals on Se toxic diets were significantly (p < 0.05) lower in rats fed a low vitamin E diet. In rats fed Se deficient and adequate diets, a high vitamin E intake resulted in a decrease in TC and an increase in HDLC.(ABSTRACT TRUNCATED AT 250 WORDS)

The aim of this study was to assess whether diets enriched in cholesterol, sodium cholate and drugs known to modify liver cholesterol biosynthesis can modulate hepatic lipase (H-TGL) expression and activity in vivo. Female lean Zucker rats, known to be good responders to cholesterol, were fed for 7 days with a control C diet or the C diet supplemented (w/w) with either 2% cholesterol, 0.5% sodium cholate, 2% cholestyramine or simvastatin (0.1%) added to the cholestyramine diet or given by gavage (10 mg/rat) for 3 days. H-TGL activity decreased by 34% with cholesterol, and by 27% when both cholesterol and cholate were administered to the rats. Under these conditions, H-TGL mRNA decreased by 34% and 87%, respectively. The sharp decrease in H-TGL expression was associated with a strong increase in cholesteryl ester in total liver and in the liver microsome fraction. H-TGL activity decreased by 33% with cholestyramine and the mRNA level decreased by 47%. Simvastatin lowered H-TGL activity by 55% when added to the cholestyramine diet, probably because of a reduction in food intake. When administrated by gavage, simvastatin increased both the H-TGL activity (by 28%) and mRNA (by 23%). These variations may be linked to the availability of mevalonate-derived sterol and non-sterol products.

Hypercholesterolemia aggravates experimental progressive glomerular injury. Evidence suggests the infiltrating glomerular macrophage (M phi) is a potential effector mechanism for the noxious effects of hypercholesterolemia. Because transforming growth factor (TGF)-beta 1 is secreted by activated M phi s and also stimulates fibronectin production by glomerular cells, we evaluated the kinetics of gene expression for these moieties in glomeruli isolated from nephrotic rats at 3, 7, 11, and 42 days after the delivery of puromycin aminonucleoside (PA). We also assessed whether cholesterol feeding, which raises the glomerular M phi number, alters the glomerular mRNA levels for TGF-beta 1 and fibronectin. Glomerular mRNA levels for TGF-beta 1 and fibronectin in nephrotic rats exhibited a biphasic temporal pattern, decreasing significantly below control at 3 and 7 days after PA but increasing significantly at 11 and 42 days after PA. The upregulated gene expression for TGF-beta 1 and fibronectin at 11 days after PA temporally corresponded to the phase of mesangial M phi infiltration in this model. Cholesterol feeding to both normal and nephrotic rats significantly increased glomerular TGF-beta 1 and fibronectin mRNA levels at 11 days after PA. Immunohistochemical labeling for M phi s and intracellular TGF-beta 1 demonstrated both mesangial and cortical interstitial localization with the TGF-beta1-positive cells possessing M phi nuclear morphology. These findings identify a novel interaction between hypercholesterolemia, augmented glomerular M phi accumulation, and upregulated glomerular TGF-beta 1 and fibronectin gene expression. These perturbations within the acutely injured glomerulus constitute an early pathobiological determinant for the later development of mesangial matrix expansion and glomerulosclerosis.

The effect of taurine on the serum and liver cholesterol and triglyceride levels was studied in rats fed cholesterol plus cholic acid. Four groups of 4 weeks old rats were fed control diet, hypercholesterolemic diet (HCD), HCD + 1% taurine or HCD + 2% taurine for 8 weeks. Addition of taurine in HCD diet showed a significant reduction not only in serum total cholesterol and triglyceride levels but also in liver total cholesterol, lipid and triglyceride contents compared to the animals fed HCD alone. Histological examination of organs of these animals showed severe fatty vacuolation in livers and signet ring type vacuolation in kidneys of rats fed HCD. Taurine showed ameliorating effect on these abnormalities. The animals fed taurine in HCD also showed increased bile and sterol excretion in faeces compared to rats fed HCD alone. Taurine showed significant hypocholesterolemia in rats probably by enhancing the catabolism of cholesterol and reducing the absorption of dietary cholesterol.

In three healthy rhesus monkeys fed chenodeoxycholic (chenic) acid, there was no consistent increase in the total exchangeable cholesterol pool or input to the cholesterol pool. In three similar monkeys fed cholic acid, the total exchangeable pool increased in all animals and input to the cholesterol pool increased in two. Serum glutamic-pyruvic transaminase (SGPT) increased transiently in two animals in each group. Morphologic abnormalities (triaditis with atypical ductular proliferation) were noted in one animal; this animal was ingesting chenic acid but had normal liver test results at the time of biopsy. Biliary bile acids contained 8 to 14 percent lithocholic acid in the chenic acid group and 48 to 72 percent deoxycholic acid in the cholic acid group.

Unlabelled: We previously reported a hamster model for cholesterol gallstone formation and prophylaxis. The aim of this study was to validate a model for dissolution of cholesterol gallstones by testing bile acids used in patients. Sixty hamsters were allocated to six groups of ten; Group I received the standard diet (.8mg cholesterol/gram food) and Groups II-VI received the lithogenic regime (2.4 mg cholesterol/gram food and 15 micrograms ethinyl estradiol) for twelve weeks. During the next eight weeks, Group I remained on the standard diet, Group II on the lithogenic regime, while Group III switched to the standard diet. Groups IV-VI remained on the lithogenic regime, and received 20 mg/kg/d of CDC (Group IV), UDC (Group V) or cholic acid (Group VI). Cholesterol gallstones were found in 90% of hamsters on the lithogenic regime, even after return to the standard diet, in 80% of those receiving cholic acid, and in none receiving the standard diet, CDC or UDC. CDC and UDC but not cholic acid inhibited hepatic HMG-CoA reductase activity (p less than 0.01) and desaturated bile (p less than 0.01). The highest HMG-CoA reductase (p less than 0.02) occurred after return from the lithogenic regime to the standard diet., Conclusions: 1) A new model for cholesterol gallstone dissolution has been validated; 2) CDC and UDC, in contrast to cholic acid, decreased HMG-CoA reductase, desaturated bile and dissolved gallstones as in patients; and 3) Return from the lithogenic regime to the standard diet did not desaturate bile or dissolve gallstones, but did increase HMG-CoA reductase as found in gallstone patients.

The increase in serum cholesterol after feeding a diet containing 2% (w/w) of cholesterol and 0.5% of cholate for 13 days was 200 and 800% in two hypo- and two hyper-responsive inbred strains of rats, respectively. While remaining on the high-cholesterol, high-cholate diet for longer periods, the level of serum cholesterol dropped in the hyper-responsive strains, and after 8 weeks on the diet one hyper-responsive strain had similar serum cholesterol concentrations as the two hypo-responsive strains. The feeding of a semipurified diet, containing 1% (w/w) of cholesterol and 20% of fat, did not discriminate between the two hypo- and hyper-responsive strains with respect to the response of serum cholesterol. The activities in plasma of the indicators for liver function, aspartate amino transferase and alkaline phosphatase, were significantly increased in all strains after feeding the high-cholesterol, high-cholate diet. Only alkaline phosphatase was increased by the semipurified diet. Evidence is presented that in the four inbred strains of rats the differential cholesterolemic response to the high-cholesterol, high-cholate diet is not related to the baseline serum lipoprotein profile, liver cholesterol accumulation, fecal bile acid excretion, and the total activities and patterns of esterases in serum, liver and small intestine.