Contribution of newly synthesized cholesterol to rat plasma and bile determined by mass isotopomer distribution analysis: bile-salt flux promotes secretion of newly synthesized cholesterol into bile (original) (raw)
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Biochimica et Biophysica Acta (BBA) - Molecular and Cell Biology of Lipids, 2000
A new stable isotope procedure has been developed and validated in rats, applying [1-13 C]acetate infusion to quantify the production of bile salts from de novo synthesized cholesterol making use of the mass isotopomer distribution analysis (MIDA) principle. Ions (m/z) 458^461, 370^373 and 285^288 were monitored by GC/MS (EI-mode) for the methyl trimethylsilylether derivatives of cholate, chenodeoxycholate and L-muricholate, respectively. Rats with intact exteriorized enterohepatic circulation and rats with chronic bile diversion were infused with [1-13 C]acetate for up to 14 h. After 10 h of infusion the enterohepatic circulation of the intact group was interrupted to deplete the existing bile salt pool (acute bile diversion). The fractions of biliary cholesterol and individual bile salts derived from newly synthesized cholesterol were determined by MIDA at t = 14 h. In rats with acute bile diversion, these fractions were 20, 25, 27 and 23% for biliary cholesterol, cholate, chenodeoxycholate and L-muricholate, respectively. After bile diversion for 8 days to induce hepatic cholesterol and bile salt synthesis, these fractions increased significantly to 32, 47, 41 and 47%, respectively. Calculated enrichments of the acetyl-CoA precursor pools were similar for all bile salts and biliary cholesterol within the two rat groups. However, chronic enterohepatic interruption decreased the acetyl-CoA pool size almost two-fold. We conclude that MIDA is a validated new stable isotope technique for studying the synthetic pathway from acetyl-CoA to bile salts. This technique provides an important new tool for studying bile salt metabolism in humans using stable isotopes. ß
Hepatology, 2003
The e&cts of newly synthesized cholesterol availability on bile acid synthesis are largely unknown, particularly in humans. The present study was aimed to study the changes induced on bile acid synthesis by simvastatin, a competitive inhibitor of hydroxymethyl glutaryl-CoA (HMG-CoA) reductase, the rate-limiting enzyme of cholesterol synthesis, during pharmacologic interruption of the enterohepatic circulation. S i x patients with primary hypercholesterolemia were studied in basal conditions, after treatment with the bile acid binding resin cholestyramine alone (8-16 g/d for 6-8 weeks) and subsequently in combination with simvastatin (40 mg/d for 6-8 weeks). Cholesterol 7a-hydroxylation rate, a measure of total bile acid synthesis, was assayed in vim by tritium release analysis. Serum lathosterol levels were assayed by gas chromatographymass spectrometry as a measure of cholesterol synthesis. Serum total and low-density lipoprotein-cholesterol were reduced significantly after cholestyramine (by 26% and 30%, respectively) and during combined treatment (by 47% and 55%). 7a-Hydroxylation rates increased nearly 4-fold with cholestyramine alone; addition of simvastatin induced a significant decrease of hydroxylation rates (cholestyramine alone, 1,591 2 183 mg/d; plus simvastatin, 1,098 f 232 mg/dj mean 2 SEM; P < .05). Hydroxylation rates significantly correlated with serum lathosterou cholesterol ratio (T = 0.79, P < .05). In conclusion, in conditions of chronic stimulation bile acid synthesis may be affected by changes in newly synthesized cholesterol availability. The finding might relate to the degree of substrate saturation of microsomal cholesterol 7a-hydroxylase; alternatively, newly synthesized cholesterol might induce a stimulatory effkct on cholesterol 7a-hydroxylase transcription. (HEPATOLOGY 2003;38:939-946.) B ile acid production is a major mechanism whereby cholesterol is eliminated from the organism and, therefore, represents a crucial event in the maintenance of cholesterol homeostasis.1-3 Two metabolic path-Abbreviations: HMG-CoA, hydroxymethyl glutaryl-CoA; LDL, low-density lipoprotein; HDL, high-density lipoprotein.
Regulation of biliary cholesterol secretion in the rat. Role of hepatic cholesterol esterification
Journal of Clinical Investigation, 1984
Although the significance of the enterohepatic circulation of bile salts in the solubilization and biliary excretion of cholesterol is well established, little is known about the intrahepatic determinants of biliary cholesterol output. Studies were undertaken to elucidate some of these determinants in the rat. Feeding 1% diosgenin for 1 wk increased biliary cholesterol output and saturation by 400%. Bile flow, biliary bile salt, phospholipid and protein outputs remained in the normal range. When ethynyl estradiol (EE) was injected into these animals, biliary cholesterol output decreased to almost normal levels under circumstances of minor changes in the rates of biliary bile salt and phospholipid outputs. Similarly, when chylomicron cholesterol was intravenously injected into diosgenin-fed animals, biliary cholesterol output significantly decreased as a function of the dose of chylomicron cholesterol administered.
Kinetics of biliary secretion of chylomicron remnant cholesterol (esters) in the rat
European Journal of Biochemistry, 1993
Chylomicrons labelled with [3H]cholesterol/[3H]cholesterol esters in a ratio of 25.5 : 74.5, were rapidly removed from rat serum in vivo, and taken up predominantly by the parenchymal liver cells (88.2% of the hepatic uptake at 15 min after injection). Lactoferrin reduced the liver uptake of chylomicron remnants by 72%, at 20 min after injection. It appeared that the free cholesterol which is present in the chylomicrons is not readily exchanged within the used time period with other cholesterol pools in the animal. Between 10-60 min after injection of 3H-labelled chylomicrons, cholesterol esters are hydrolysed in the liver. Appearance of radioactivity in bile was rapid and at 3,24 and 72 h after injection, 13.4 %, 44.0 % and 70.0 %, respectively, of the injected dose appeared in bile, mainly as bile acids (> 90 %). Lactoferrin reduced the biliary secretion of radioactivity, especially during the first hour after injection. The total amount of radioactivity recovered was 58.0 % of the injected dose at 72 h after injection. After injection of P-migrating very low-density lipoprotein labelled with [3H]cholesteroY[3H]cholesterol esters in a ratio of 23.5 : 76.5, the maximum amount of radioactivity secreted in bile was much lower than with chylomicrons (2.6 % cf. 5.2% at 1 h after injection), although the kinetics of the initial liver association and cholesterol ester hydrolysis were even more rapid. B i l i q accumulation of radioactivity was also lower with 50.5 % of the injected dose recovered at 72 h after injection. It can be concluded from these studies that the processing of chylomicron remnant cholesterol components in the liver and the subsequent secretion in the bile mainly as bile acids is very efficient. The efficient liver uptake of chylomicron remnants by the liver remnant receptor is thereby essential to achieve this high percentage of removal, thus protecting against extrahepatic cholesterol (ester) deposition.
Journal of Clinical Investigation, 1988
The functional interrelationship between biliary cholesterol secretion, sinusoidal lipoprotein cholesterol secretion and bile salt synthesis was studied in the rat. Diosgenin, fructose, and colestipol in the diet were used to, respectively, influence biliary cholesterol output, VLDL production and bile salt synthesis. In the acute bile fistula rat, biliary cholesterol output was 700% increased by diosgenin and 50% decreased by fructose. In the rats fed both diosgenin and fructose, biliary cholesterol secretion was increased only by 200%, whereas biliary bile salts and phospholipid outputs were unchanged.
Journal of Clinical Investigation, 1978
was examined. The fractional inhibition of cholesterol synthesis found after administration of an amount of cholesterol sufficient to raise the hepatic cholesterol ester content by 1 mg/g equalled only -0.36 when bile acid synthesis was increased by biliary diversion but was -0.92 when bile acid synthesis was suppressed by bile acid feeding. It is concluded that (a) bile acids are not direct effectors of the rate of hepatic cholesterol synthesis, (b) most of the inhibitory activity seen with bile acid feeding is mediated through increased cholesterol absorption, and (c) bile acids do have an intrahepatic effect in that they regulate hepatic cholesterol synthesis indirectly by altering the flow of cellular cholesterol to bile acids. J. Clin. Invest.
Separate pools for endogenous and exogenous cholesterol in rat intestines
Atherosclerosis, 1973
Endogenous and exogenous cholesterol were pulse labelled by intraperitoneal injection of [2-sH]mevalonate and by gastric intubation of [4J%]cholesterol in rats with common bile and thoracic lymph duct fistulae. It took 3 hours for some of the exogenous cholesterol to appear in the large intestines, although a significant fraction was still present in the stomachs of the rats. The specific activity of exogenous cholesterol reached peak values and then declined at rates which were much faster than those for endogenous cholesterol in thoracic duct lymph. The free/esterified specific activity ratios for the two isotopes were different in the lymph cholesterol but they were similar for cholesterol in different segments along the length of the intestinal tract. In additional studies, cholesterol-fed rats (with cannulae in the common bile and thoracic lymph ducts) were infused intraduodenally with normal rat bile to which [4-14C]cholesterol and [2-sH]mevalonate had been added. The specific activity for both isotopes in the lymph cholesterol increased rapidly and stabilized by 16 hours at values many times greater than the specific activity of liver, bile, or plasma cholesterol. On removal of radioisotopes from the infusate there was an exponential decline for up to 30 hours in the specific activity of both isotopes. As in other experiments, the rate of decline for 14C was up to three times greater than that for sH. The specific activity ratios of free/esterified cholesterol in the lymph were also consistently different for the two isotopes. These results suggest that absorption of dietary cholesterol is considerably more efficient than the absorption of endogenous cholesterol synthesized in the intestines. These data also imply that the cholesterol derived from the two sources exists in different pools in the rat intestines.
Cholesterol Secretion Into Bile
2013
Recently we demonstrated that dietary fish oil (FO) causes changes in intrahepatic cholesterol transport and hypersecretion of cholesterol into bile in rats (J Clin. Znuest. 88: 943-951, 1991). We have now investigated in more detail the relationship between cholesterol and bile acid secretion in rats with chronic bile diversion fed purified diets supplemented (9% wt/wt) with either FO or corn oil (CO) for 2 weeks. Effects of 'This work was presented at the Annual Meeting of the American Gastroenterological Association, May 15-21 1993, Boston, MA, and has been published in abstract form (Garfromferolo~. 1993. 104: A993). The hepatobiliary pathway is the main route for sity lipoprotein; ACAT, acyl-CoA:cholesterol acyltransferase.