Contraceptive steroids alter the steady-state kinetics of bile acids

Contraceptive steroids increase the ratio of cholic acid to chenodeoxycholic acid in bile. This alteration may contribute to the development of cholesterol gallstones. The objective of this study was to measure the effects of contraceptive steroids on bile acid kinetics and to relate them to changes in cholesterol metabolism. Steady-state kinetics of bile acids were measured in 15 healthy women, on and off contraceptive steroids. Cholic acid synthesis increased 30.3% (P less than 0.025) and its pool increased by 37.4% (P less than 0.025). Chenodeoxycholic acid synthesis decreased 6.4% (P = 0.08) and its pool decreased by 11.8% (P less than 0.05) during use of contraceptive steroids. The fractional turnover rates of both primary bile acids did not change. The changes in kinetics of the primary bile acids were related to alterations in biliary lipid and cholesterol metabolism, separately reported. (J. Lipid Res. 1987. 28: 828-839). During use of contraceptive steroids, total bile acid pool and total bile acid synthesis correlated directly with cholesterol synthesis, assayed in mononuclear leukocytes (r = 0.50 and r = 0.54, respectively) but not with the plasma clearance of chylomicron remnants, measured with retinyl palmitate. The data indicate that contraceptive steroids directly alter the hepatic synthesis of bile acids and suggest that newly synthesized cholesterol may be a preferred substrate for bile acid synthesis during use of contraceptive steroids.     

Evidence for a common biliary cholesterol and VLDL cholesterol precursor pool in rat liver.

Hepatic free cholesterol levels are influenced by cholesterol synthesis and ester formation, which, in turn, might regulate cholesterol secretion into bile and plasma. We manipulated the rates of hepatic cholesterol synthesis and esterification and measured biliary and very low density lipoprotein (VLDL) cholesterol secretion, and bile acid synthesis. Mevalonate decreased HMG CoA reductase by 80%, increased acyl coenzyme A: cholesterol acyltransferase (ACAT) by 60% and increased [3H]oleate incorporation into microsomal and VLDL cholesteryl esters by 174% and 122%, respectively. Microsomal and biliary free cholesterol remained constant at the expense of increased microsomal and VLDL cholesteryl ester content. Mevalonate did not change bile acid synthesis. 25-OH cholesterol decreased HMG-CoA reductase by 39%, increased ACAT by 24%, but did not effect 7 alpha-hydroxylase. 25-OH cholesterol increased [3H]oleate in microsomal and VLDL cholesterol esters by 71% and 120%. Biliary cholesterol decreased by 40% and VLDL cholesteryl esters increased by 83%. A small and unsustained decrease in bile acid synthesis (14CO2 release) occurred after 25-OH cholesterol. After orotic acid feeding, HMG-CoA reductase increased 352%, and [3H]oleate in microsomal and VLDL cholesteryl esters decreased by 43% and 89%. Orotic acid decreased all VLDL components including free cholesterol (68%) and cholesteryl esters (55%), and increased biliary cholesterol by 160%. No change in bile acid synthesis occurred. Hepatic cholesterol synthesis and esterification appear to regulate a cholesterol pool available for both biliary and VLDL secretion. Changing cholesterol synthesis and esterification did not alter bile acid synthesis, suggesting that either this common bile/VLDL secretory pool is functionally distinct from the cholesterol pool used for bile salt synthesis, or that free cholesterol availability in this precursor pool is not a major determinant of bile acid synthesis.     

Effects of lovastatin on biliary lipid secretion and bile acid metabolism in humans

Lovastatin, an inhibitor of HMG-CoA reductase, lowers cholesterol saturation of bile. To determine the mechanism of this effect and further define the role of cholesterol synthesis in regulation of biliary lipid metabolism, we studied ten human volunteers in a control period and again after 5-6 weeks on lovastatin, 40 mg b.i.d. Mean sterol production from acetate in mononuclear leukocytes fell from 1.18 to 0.84 pmol/min per 10(6) cells on lovastatin (P less than 0.02). Concomitantly there was reduction in mean biliary secretion of cholesterol from 143 to 96 mumol/h (P less than 0.02). On lovastatin, mean pool size of bile acids by the Lindstedt method fell from 3193 to 2917 mumol (one-sided P = 0.05) and mean pool size by the one-sample method fell from 5158 to 4091 mumol (P less than 0.002). Lovastatin had no effect on mean fractional turnover rate of either cholic acid (0.77 vs. 0.74 day-1) or chenodeoxycholic acid (0.51 vs. 0.54 day-1). Mean total bile acid synthesis was lower on lovastatin (1443 vs. 1240 mumol/day), but the difference did not quite achieve statistical significance. In humans, inhibition of cholesterol synthesis by lovastatin lowers biliary cholesterol saturation by reducing cholesterol secretion into bile. Bile acid pool size, and perhaps bile acid synthesis, are also reduced by this inhibition.     

Role of cholesterol synthesis in regulation of bile acid synthesis and biliary cholesterol secretion in humans

We used lovastatin, a specific inhibitor of HMG-CoA reductase, to study the role of cholesterol synthesis in regulation of both bile acid synthesis, measured by release of 14CO2 from [26-14C]cholesterol, and biliary cholesterol secretion, measured by standard marked perfusion techniques, in humans. Six volunteers were studied in each of four periods: a) control; b) 6-10 hours after a single 40 mg oral dose of lovastatin to study acute effects; c) after 5-6 weeks of lovastatin 40 mg orally twice a day to study steady-state effects; and d) 24 h after cessation of chronic lovastatin. Mean bile acid synthesis fell to 69% of control (P less than 0.01) after single-dose lovastatin and remained at 83% of control after 5-6 weeks on lovastatin (P less than 0.05). After withdrawal of lovastatin, mean bile acid synthesis was 88% of control (NS). Mean biliary cholesterol secretion did not change after single-dose lovastatin (103% of control), but fell to 81% of control during chronic lovastatin treatment (P less than 0.05). After withdrawal of lovastatin, mean cholesterol secretion remained at 80% of control (P less than 0.05). These data suggest that in humans cholesterol synthesis is an immediate regulator of bile acid synthesis. Cholesterol synthesis also regulates biliary cholesterol secretion, but the effect is not immediate and therefore may be indirect.     

Cholecystectomy prevents expansion of the bile acid pool and inhibition of cholesterol 7alpha-hydroxylase in rabbits fed cholesterol

To study the effect of cholecystectomy on the regulation of classic and alternative bile acid syntheses, gallbladder-intact (n = 20) and cholecystectomized (n = 20) New Zealand White rabbits were fed either chow or chow with 2% cholesterol (3 g/day). After 10 days, bile fistulas were constructed in half of each rabbit group to recover and measure the bile acid pool and biliary bile acid flux. After cholesterol feeding, the bile acid pool size increased from 268 +/- 55 to 444 +/- 77 mg (P < 0.01) with a 2-fold rise in the biliary bile acid flux in intact rabbits but did not expand the bile acid pool (270 +/- 77 vs. 276 +/- 62 mg), nor did the biliary bile acid flux increase in cholecystectomized rabbits. Ileal apical sodium-dependent bile acid transporter protein increased 46% from 93 +/- 6 to 136 +/- 23 units/mg (P < 0.01) in the intact rabbits but did not change in cholecystectomized rabbits (104 +/- 14 vs. 99 +/- 19 units/mg) after cholesterol feeding. Cholesterol 7alpha-hydroxylase activity was inhibited 59% (P < 0.001) while cholesterol 27-hydroxylase activity rose 83% (P < 0.05) after cholesterol feeding in the intact rabbits but neither enzyme activity changed significantly in cholesterol-fed cholecystectomized rabbits. Fecal bile acid outputs reflecting bile acid synthesis increased significantly in the intact but not in the cholecystectomized rabbits fed cholesterol.Removal of the gallbladder prevented expansion of the bile acid pool after cholesterol feeding as seen in intact rabbits because ileal bile acid transport did not increase. As a result, cholesterol 7alpha-hydroxylase was not inhibited.     

Thyroid hormone differentially augments biliary sterol secretion in the rat. II. The chronic bile fistula model.

To further define thyroid hormone effects on bile acid synthesis and biliary lipid secretion, studies were done in chronic bile fistula rats. Euthyroid and methimazole-hypothyroid rats, with and without triiodothyronine (T3) injection, had total bile diversion for timed bile collections. With interrupted enterohepatic circulation, cholesterol absorption is negligible and bile acid secretion equals bile acid synthesis rate. Hypothyroid rats had diminished levels of bile acid synthesis and biliary secretion of cholesterol and phospholipid. Single dose T3 injection produced a 13-fold increase in bile cholesterol secretion and a 3-fold increase in phospholipid secretion, both initiated 12 h after T3. Bile acid synthesis increased by 50%, but the increase did not begin until 24 h after T3. Neither hypothyroidism nor T3 treatment abolished diurnal rhythms of bile acid synthesis and biliary lipid secretion. Inhibition of cholesterol synthesis with lovastatin resulted in a persistent 33% decrease in bile acid synthesis in euthyroid and hypothyroid rats, while bile cholesterol secretion only transiently decreased. Inhibition of cholesterol synthesis did not alter T3-induced bile cholesterol secretion, with a 10-fold increase seen. However, bile acid synthesis was not stimulated by T3 in the presence of lovastatin. We conclude that facilitated bile acid synthesis and biliary cholesterol secretion are early effects of T3 and may account for the hypocholesterolemia of T3. Cholesterol synthesis does not appear to be required for the T3-induced bile cholesterol secretion.     

Taurine increases bile acid pool size and reduces bile saturation index in the hamster

There is evidence that increased availability of taurine enhances the proportion of taurine-conjugated bile acids in bile. To explore the possibility that taurine treatment could also influence hepatic cholesterol and bile acid metabolism, we fed female hamsters for 1 week and measured both the biliary lipid content and the microsomal level of the rate-limiting enzymes of cholesterol and bile acid synthesis. In these animals the cholesterol 7 alpha-hydroxylase activity was significantly greater in respect to controls (P less than 0.05). The total HMG-CoA reductase activity, as well as that of the active form, was similarly increased. The stimulation of 7 alpha-hydroxycholesterol synthesis was associated with an expansion of the bile acid pool size in taurine-fed animals. Taurine feeding was observed to induce an increase in bile flow as well as in the rate of excretion of bile acids, whereas the secretion rate of cholesterol in bile was decreased. As a consequence, the saturation index was significantly lower in taurine-fed animals (P less than 0.05). The possible mechanisms through which taurine exhibits the modification of the enzyme activities and of the biliary lipid composition are discussed.     

Bile salts in submicellar concentrations promote bidirectional cholesterol transfer (exchange) as a function of their hydrophobicity

Cholesterol, despite its poor solubility in aqueous solutions, exchanges efficiently between membranes. Movement of cholesterol between different subcellular membranes in the hepatocyte is necessary for assembly of lipoproteins, biliary cholesterol secretion, and bile acid synthesis. Factors which initiate and facilitate transfer of cholesterol between different membranes in the hepatocyte are incompletely understood. It is known that cholesterol secretion into the bile is linked to bile salt secretion. In the present study, we investigated the effects of bile salts of different physicochemical properties at submicellar concentrations (150- 600 microM) on the transfer of [14C]cholesterol from hepatocytes, or crude hepatocellular membranes (donors), to rat high density lipoproteins (acceptor). Bile salts included taurine conjugates of ursodeoxycholic acid (TUDCA), hyodeoxycholic acid (THDCA), cholic acid (TCA), chenodeoxycholic acid (TCDCA), and deoxycholic acid (TDCA). High density lipoprotein (HDL) was separated from hepatocellular membranes and the transfer of [14C]cholesterol from the membranes to HDL was quantitatively determined. In the absence of HDL, [14C]cholesterol remained confined to the membrane fraction. Following addition of HDL, [4-14C]cholesterol in the HDL fraction increased linearly over time. Addition of hydrophilic bile salts (TUDCA and THDCA) increased transfer of [4-14C]cholesterol to HDL only minimally. By contrast, more hydrophobic bile salts stimulated transfer of labeled cholesterol to HDL, and their potency increased in order of increasing hydrophobicity (TCA less than TCDCA less than TDCA). Both for single bile salts and mixtures of bile salts at a total bile salt concentration of 0.30 mM, the rate of cholesterol transfer exhibited a strong linear correlation with a bile salt monomeric hydrophobicity index (r = 0.95; P less than 0.001).(ABSTRACT TRUNCATED AT 250 WORDS)     

Cholesterol synthesis in the perfused liver of pregnant hamsters

Pregnancy is a risk factor for the development of cholesterol gallstones. In pregnant women, biliary cholesterol saturation and secretion are increased. To investigate whether this was due to increased cholesterol synthesis, we studied hepatic cholesterol synthesis in Syrian Golden hamsters. Female controls and animals 10- to 14-days pregnant were studied. The studies were performed in the in situ perfused hamster liver. Cholesterol synthesis was determined by measuring the incorporation of 3H2O added to the perfusate into hepatic, perfusate, and bile cholesterol during a 90-min period. In both pregnant groups, bile flow decreased significantly, but biliary cholesterol concentration increased only in the 14-day pregnant group. The cholesterol synthesis rate averaged (mean +/- SD) 172 +/- 27, 127 +/- 37, and 552 +/- 79 nmol X hr-1 X g liver-1 in controls, 10-day, and 14-day pregnant animals, respectively. The 14-day pregnant animals secreted a markedly higher fraction (47.3 +/- 11.3 vs. 11.1 +/- 13.4%; P less than 0.01) of newly synthesized cholesterol into bile but not into perfusate. Chenodeoxycholate, but not cholate, synthesis rate was decreased in both pregnant groups. We conclude from our studies that hepatic cholesterol synthesis increases towards the end of pregnancy in the hamster and that more newly synthesized cholesterol is secreted into bile at that time. This could at least partially explain the increased biliary cholesterol saturation and secretion observed in women in the third trimenon, and explain pregnancy as a risk factor in the development of cholesterol gallstones.     

Disruption of the sterol carrier protein 2 gene in mice impairs biliary lipid and hepatic cholesterol metabolism.

Hepatic up-regulation of sterol carrier protein 2 (Scp2) in mice promotes hypersecretion of cholesterol into bile and gallstone formation in response to a lithogenic diet. We hypothesized that Scp2 deficiency may alter biliary lipid secretion and hepatic cholesterol metabolism. Male gallstone-susceptible C57BL/6 and C57BL/6(Scp2(-/-)) knockout mice were fed a standard chow or lithogenic diet. Hepatic biles were collected to determine biliary lipid secretion rates, bile flow, and bile salt pool size. Plasma lipoprotein distribution was investigated, and gene expression of cytosolic lipid-binding proteins, lipoprotein receptors, hepatic regulatory enzymes, and intestinal cholesterol absorption was measured. Compared with chow-fed wild-type animals, C57BL/6(Scp2(-/-)) mice had higher bile flow and lower bile salt secretion rates, decreased hepatic apolipoprotein expression, increased hepatic cholesterol synthesis, and up-regulation of liver fatty acid-binding protein. In addition, the bile salt pool size was reduced and intestinal cholesterol absorption was unaltered in C57BL/6(Scp2(-/-)) mice. When C57BL/6(Scp2(-/-)) mice were challenged with a lithogenic diet, a smaller increase of hepatic free cholesterol failed to suppress cholesterol synthesis and biliary cholesterol secretion increased to a much smaller extent than phospholipid and bile salt secretion. Scp2 deficiency did not prevent gallstone formation and may be compensated in part by hepatic up-regulation of liver fatty acid-binding protein. These results support a role of Scp2 in hepatic cholesterol metabolism, biliary lipid secretion, and intracellular cholesterol distribution.     

Regulation of bile acid synthesis. III. Correlation between biliary bile salt hydrophobicity index and the activities of enzymes regulating cholesterol and bile acid synthesis in the rat

Hepatic bile acid synthesis is thought to be under negative feedback control by bile salts in the enterohepatic circulation, acting at the level of cholesterol 7 alpha-hydroxylase (C7 alpha H), the initial and rate-limiting step in the bile acid biosynthetic pathway. Bile salts also suppress the activity of the rate-limiting enzyme for cholesterol synthesis, 3-hydroxy-3-methylglutaryl-coenzyme A reductase (HMG-CoA-R). The mechanisms of these regulatory effects are poorly understood, and one or both may be indirect. Previous data suggest that the hydrophilic-hydrophobic balance of bile salts, a major determinant of their cholesterol solubilizing properties, also determines their potency as regulators of bile acid and cholesterol synthesis. To further evaluate the relationship between the physicochemical and regulatory properties of bile acids, we altered the composition of the bile salt pool of rats by feeding one or more of seven different bile acids (1% w/w for 14 days). We then determined the mean hydrophilic-hydrophobic balance (hydrophobicity index) of the bile salts in bile, and correlated this with the specific activities of C7 alpha H and HMG-CoA-R, and of acyl-CoA:cholesterol acyltransferase (ACAT), a third hepatic microsomal enzyme which regulates cholesterol esterification. In all instances following bile acid feeding, conjugates of the fed bile acid(s) became the predominant bile salts in bile. Highly significant negative linear correlations (each P less than 0.0001) were found between the hydrophobicity indices of biliary bile salts and the activities of C7 alpha H (r = 0.79) or HMG-CoA-R (r = 0.63). By contrast, no significant correlation could be demonstrated between ACAT activity and the hydrophobicity index of biliary bile salts. The correlation between activities of HMG-CoA-R and C7 alpha H was also highly significant (r = 0.81; P less than 0.0001). No significant correlation existed between ACAT and either HMG-CoA-R or C7 alpha H. Microsomal free cholesterol was not consistently altered by bile acid feeding. Thus, the potency of circulating bile salts as suppressors of the enzymes regulating bile acid and cholesterol synthesis increases with increasing hydrophobicity. The hydrophobic-hydrophilic balance of the bile salt pool may play an important role in the regulation of cholesterol and bile acid synthesis.     

High cholesterol absorption efficiency and rapid biliary secretion of chylomicron remnant cholesterol enhance cholelithogenesis in gallstone-susceptible mice

The study of chylomicron pathway through which it exerts its metabolic effects on biliary cholesterol secretion is crucial for understanding how high dietary cholesterol influences cholelithogenesis. We explored a relationship between cholesterol absorption efficiency and gallstone prevalence in 15 strains of inbred male mice and the metabolic fate of chylomicron and chylomicron remnant cholesterol in gallstone-susceptible C57L and gallstone-resistant AKR mice. Our results show a positive and significant (P<0.0001, r=0.87) correlation between percent cholesterol absorption and gallstone prevalence rates. Compared with AKR mice, C57L mice displayed significantly greater absorption of cholesterol from the small intestine, more rapid plasma clearance of chylomicrons and chylomicron remnants, higher activities of lipoprotein lipase and hepatic lipase, greater hepatic uptake of chylomicron remnants, and faster secretion of chylomicron remnant cholesterol from plasma into bile. All of these increased susceptibility to cholesterol gallstone formation in C57L mice. We conclude that genetic variations in cholesterol absorption efficiency are associated with cholesterol gallstone formation in inbred mice and cholesterol absorbed from the intestine provides an important source for biliary hypersecretion. Differential metabolism of the chylomicron remnant cholesterol between C57L and AKR mice clearly plays a crucial role in the formation of lithogenic bile and gallstones.     

Hepatic overexpression of sterol carrier protein-2 inhibits VLDL production and reciprocally enhances biliary lipid secretion

We examined in vivo a role for sterol carrier protein-2 (SCP-2) in the regulation of lipid secretion across the hepatic sinusoidal and canalicular membranes. Recombinant adenovirus Ad.rSCP2 was used to overexpress SCP-2 in livers of mice. We determined plasma, hepatic, and biliary lipid concentrations; hepatic fatty acid (FA) and cholesterol synthesis; hepatic and biliary phosphatidylcholine (PC) molecular species; and VLDL triglyceride production. In Ad.rSCP2 mice, there was marked inhibition of hepatic fatty acids and cholesterol synthesis to <62% of control mice. Hepatic triglyceride contents were decreased, while cholesterol and phospholipids concentrations were elevated in Ad.rSCP2 mice. Hepatic VLDL triglyceride production fell in Ad.rSCP2 mice to 39% of control values. As expected, biliary cholesterol, phospholipids, bile acids outputs, and biliary PC hydrophobic index were significantly increased in Ad.rSCP2 mice. These studies indicate that SCP-2 overexpression in the liver markedly inhibits lipid synthesis as well as VLDL production, and alters hepatic lipid contents. In contrast, SCP-2 increased biliary lipid secretion and the proportion of hydrophobic PC molecular species in bile. These effects suggest a key regulatory role for SCP-2 in hepatic lipid metabolism and the existence of a reciprocal relationship between the fluxes of lipids across the sinusoidal and canalicular membranes.     

Cholesterol and bile acid dynamics: comparative aspects

While the cholesterol concentration in a given tissue is similar in the rat, pig or man, the relative importance of the processes regulating the input (absorption and synthesis) and output (faecal cholesterol and bile acid excretions) of the cholesterol system is very different from one species to another. The rat, whose cholesterolaemia does not significantly increase after cholesterol addition to the diet ("hyporesponding" animal), successfully adapts its bile acid biosynthesis to variations in cholesterol input. This process accounts for 80 to 85% of cholesterol output, faecal cholesterol excretion being a minor process. The latter results from a low liver cholesterol secretion in the bile due to the low hydrophobicity of its main bile acids. Furthermore, in this animal a high intestinal synthesis of cholesterol and apolipoproteins (particularly B48) is observed. The latter are secreted as very light lipoproteins (chylomicrons and VLDL) with a faster plasma turnover than the VLDL (apoB100, E...) secreted by the liver. The "remnants" of rat VLDL are essentially very rapidly taken up by the liver; their interplasmatic transformation pathway into IDL and LDL is not very significant (less than or equal to 10%). Man, who has a more significant hypercholesterolaemia after exogenous cholesterol ingestion ("hyperresponding" subject) seems to have a less modulable capacity for transforming cholesterol into bile acids. This process accounts for only 50% of cholesterol output, faecal cholesterol excretion being quantitatively just as significant. Cholesterol concentration and the cholesterol/bile acid ratio are much higher in human than in rat bile, the main bile acids being more hydrophobic. While both the intestine and liver contribute to cholesterogenesis, the relative importance of the latter is probably greater in man than in the rat. Moreover, a larger fraction of plasma VLDL is transformed into IDL and LDL, the latter representing the main plasma cholesterol carrier. Determining whether the differences between the biodynamics of cholesterol processes in the rat and in man can be generalised to mammals with low or high sensitivities to hypercholesterolaemia and atherosclerosis seems to be a fundamental research objective for the next few years.     

Inhibition of cholesterol absorption by SCH 58053 in the mouse is not mediated via changes in the expression of mRNA for ABCA1, ABCG5, or ABCG8 in the enterocyte

Intestinal cholesterol absorption is a major determinant of plasma low density lipoprotein-cholesterol (LDL-C) concentrations. Ezetimibe (SCH 58235) and its analogs SCH 48461 and SCH 58053 are novel potent inhibitors of cholesterol absorption whose mechanism of action is unknown. These studies investigated the effect of SCH 58053 on cholesterol metabolism in female 129/Sv mice. In mice fed a low cholesterol rodent diet containing SCH 58053, cholesterol absorption was reduced by 46% and fecal neutral sterol excretion was increased 67%, but biliary lipid composition and bile acid synthesis, pool size, and pool composition were unchanged. When the dietary cholesterol content was increased either 10- or 50-fold, those animals given SCH 58053 manifested lower hepatic and biliary cholesterol concentrations than did their untreated controls. Cholesterol feeding increased the relative mRNA level for adenosine triphosphate-binding cassette transporter A1 (ABCA1), ABC transporter G5 (ABCG5), and ABC transporter G8 (ABCG8) in the jejunum, and of ABCG5 and ABCG8 in the liver, but the magnitude of this increase was generally less if the mice were given SCH 58053. We conclude that the inhibition of cholesterol absorption effected by this new class of agents is not mediated via changes in either the size or composition of the intestinal bile acid pool, or the level of mRNA expression of proteins that facilitate cholesterol efflux from the enterocyte, but rather may involve disruption of the uptake of luminal sterol across the microvillus membrane.     

Ileal bile acid transport regulates bile acid pool, synthesis, and plasma cholesterol levels differently in cholesterol-fed rats and rabbits

We investigated the effect of ileal bile acid transport on the regulation of classic and alternative bile acid synthesis in cholesterol-fed rats and rabbits. Bile acid pool sizes, fecal bile acid outputs (synthesis rates), and the activities of cholesterol 7alpha-hydroxylase (classic bile acid synthesis) and cholesterol 27-hydroxylase (alternative bile acid synthesis) were related to ileal bile acid transporter expression (ileal apical sodium-dependent bile acid transporter, ASBT). Plasma cholesterol levels rose 2.1-times in rats (98 +/- 19 mg/dl) and 31-times (986 +/- 188 mg/dl) in rabbits. The bile acid pool size remained constant (55 +/- 17 mg vs. 61 +/- 18 mg) in rats but doubled (254 +/- 46 to 533 +/- 53 mg) in rabbits. ASBT protein expression did not change in rats but rose 31% (P < 0.05) in rabbits. Fecal bile acid outputs that reflected bile acid synthesis increased 2- and 2.4-times (P < 0.05) in cholesterol-fed rats and rabbits, respectively. Cholesterol 7alpha-hydroxylase activity rose 33% (24 +/- 2.4 vs. 18 +/- 1.6 pmol/mg/min, P < 0.01) and mRNA levels increased 50% (P < 0.01) in rats but decreased 68% and 79%, respectively, in cholesterol-fed rabbits. Cholesterol 27-hydroxylase activity remained unchanged in rats but rose 62% (P < 0.05) in rabbits. Classic bile acid synthesis (cholesterol 7alpha-hydroxylase) was inhibited in rabbits because an enlarged bile acid pool developed from enhanced ileal bile acid transport. In contrast, in rats, cholesterol 7alpha-hydroxylase was stimulated but the bile acid pool did not enlarge because ASBT did not change. Therefore, although bile acid synthesis was increased via different pathways in rats and rabbits, enhanced ileal bile acid transport was critical for enlarging the bile acid pool size that exerted feedback regulation on cholesterol 7alpha-hydroxylase in rabbits.     

Evaluation of an isotope ratio method for measuring biliary cholesterol secretion

We have evaluated an isotope ratio method for measuring biliary cholesterol secretion. Secretion was measured in eight nonhuman primates by analysis of radioactivity of feces and plasma 4 weeks after intravenous administration of a single dose of [3H]cholesterol. For the test, [14C]cholesterol was fed in known amounts daily for 10 days. The ratio of isotopes in feces (14C/3H) was equivalent to the ratio of total radioactivity that entered the intestine from diet and bile. Assuming biliary cholesterol specific activity equals plasma cholesterol specific activity, the mass of cholesterol secreted daily in the bile could be calculated. In paired experiments in four animals we were able to directly compare biliary secretion by the new method with mass measurement by intraduodenal intubation (Grundy, S. M., and A. L. Metzger. 1972. Gastroenterology. 52: 2100-1216). The two methods correlated well in these four animals (r = 0.97). We further noted that bile secretion by the new method (eight animals) and by the mass method (four animals) correlated well with body weight (r = 0.94 for weight vs secretion by the isotope ratio method; r = 0.97 for weight vs secretion by the mass ratio method). Ranges of body weight and secretion for the eight animals were 4.2-13.1 kg and 3.6-11.4 mg/hr, respectively. The slopes of the two regression lines for body weight vs. secretion measured by the two methods did not differ significantly from one another (F(1.8) = 1.42; 0.25 less than P less than 0.50).(ABSTRACT TRUNCATED AT 250 WORDS)     

Hepatic metabolism and secretion of a cholesterol-enriched lipoprotein fraction

A potentially important source of cholesterol secreted in bile is cholesterol-rich lipoproteins. However, the fate of the cholesterol carried in these lipoproteins after hepatic uptake has not been investigated. We harvested an apoE- and cholesterol-rich lipoprotein fraction (d 1.02-1.06 g/ml) from hypercholesterolemic rats and examined the acute effects of these lipoproteins on hepatic cholesterol metabolism, very low density lipoprotein (VLDL) secretion, and biliary lipid secretion. Administration of a lipoprotein bolus (20 mg of cholesterol) to rats resulted in a significant decrease in 3-hydroxy-3-methylglutaryl coenzyme A reductase activity and a significant increase in acyl-coenzyme A:cholesterol acyltransferase activity over controls at 1 hr. Hepatic cholesteryl ester content increased 400% with no change in hepatic free cholesterol content or biliary cholesterol secretion. These cholesterol-rich lipoproteins delivered in the isolated perfused liver effected a fivefold increase in hepatic VLDL secretion with no change in composition. Therefore, cholesterol-rich lipoproteins do not acutely alter biliary cholesterol secretion. Rather, the majority of the cholesterol delivered to the liver in these lipoproteins is either esterified and stored as cholesteryl ester or resecreted as free and esterified cholesterol in hepatic VLDL.     

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.     

Biliary lipid secretion in hypercholesterolemia.

A report on the effects of primary bile acid ingestion alone or in combination with plant sterols on serum cholesterol levels, biliary lipid secretion, and bile acid metabolism. Biliary bile acid and cholesterol secretion were measured in four patients with type IIa hypercholesterolemia before and after randomized treatment periods. During these periods either a bile acid mixture (cholic-chenodeoxycholic 2:1, a proportion similar to that endogenously synthesized in health), at a level of 20 mg/kg, or the same mixture plus sitosterols, 200 mg/kg, was fed. Serum cholesterol and the cholesterol saturation of fasting-state bile was also measured. Pretreatment biliary lipid secretion was within normal limits. Bile acid kinetic measurements were also recorded before treatment and showed that cholic acid synthesis was disproportionately decreased relative to that of chenodeoxycholic acid, a finding previously reported by others. Administration of the bile acid mixture increased biliary bile acid secretion in 3 of 4 patients, but did not influence biliary cholesterol secretion. The combination of sitosterol-bile acid, however, caused a relative decrease in cholesterol secretion in bile, and fasting-state bile became unsaturated in all patients. No change in fecal neutral sterol excretion occurred during the beta-sitosterol-bile acid regimen, suggesting that simultaneous bile acid feeding blocks the compensatory increase in cholesterol synthesis known to be induced by beta-sitosterol feeding in hypercholesterolemic patients. Serum cholesterol levels also fell modestly during the sitosterol-bile acid regimen, the decrease averaging 15%. We conclude that the abnormally low rate of bile acid synthesis in patients with type IIa hyperlipoproteinemia does not influence biliary lipid secretion; that increasing the input of the two primary bile acids into the enterohepatic circulation does not increase biliary cholesterol secretion or lower serum cholesterol levels in such patients; and that the usual increase in cholesterol synthesis induced by beta-sitosterol feeding does not occur if bile acids are administered simultaneously.