Biomedical subjects
Publications and source records attributed to S Hauser.
The population exposure to those living on the island of Pantelleria, Italy, was estimated by measuring the natural gamma background. Gamma spectra of natural rocks and measurements of absorbed dose in air were taken. A correlation was found between the mean gamma exposure rate and the mean values of natural radionuclide concentrations in the investigated rocks.
We measured hepatic cholesterol 7 alpha-hydroxylase activity, mass, and catalytic efficiency (activity/unit mass) in bile fistula rats infused intraduodenally with taurocholate and its 7 beta-hydroxy epimer, tauroursocholate, with or without mevalonolactone to supply newly synthesized cholesterol. Enzyme activity was measured by an isotope incorporation assay and enzyme mass by densitometric scanning of immunoblots using rabbit anti-rat liver cholesterol 7 alpha-hydroxylase antisera. Cholesterol 7 alpha-hydroxylase activity increased 6-fold, enzyme mass 34%, and catalytic efficiency 5-fold after interruption of the enterohepatic circulation for 48 h. When taurocholate was infused to the bile acid-depleted animals at a rate equivalent to the hepatic bile acid flux (27 mumol/100-g rat/h), cholesterol 7 alpha-hydroxylase activity and enzyme mass declined 60 and 61%, respectively. Tauroursocholate did not significantly decrease cholesterol 7 alpha-hydroxylase activity, mass and catalytic efficiency. The administration of mevalonolactone, which is converted to cholesterol, modestly increased cholesterol 7 alpha-hydroxylase activity and enzyme mass in the bile acid-depleted rats. However, when taurocholate was infused together with mevalonolactone, cholesterol 7 alpha-hydroxylase activity and catalytic efficiency were markedly depressed while enzyme mass did not change as compared with bile acid-depleted rats. These results show that (a) hepatic bile acid depletion increases bile acid synthesis mainly by activating cholesterol 7 alpha-hydroxylase with only a small rise in enzyme mass, (b) replacement with taurocholate for 24 h decreases both cholesterol 7 alpha-hydroxylase activity and mass proportionally, (c) when cholesterol is available (mevalonolactone supplementation), the infusion of taurocholate results in the formation of a catalytically less active cholesterol 7 alpha-hydroxylase, and (d) tauroursocholate, the 7 beta-hydroxy epimer of taurocholate, does not inhibit cholesterol 7 alpha-hydroxylase. Thus, bile acid synthesis is modulated by the catalytic efficiency and mass of cholesterol 7 alpha-hydroxylase. The enterohepatic flux of 7 alpha-hydroxylated bile acids and the formation of hepatic cholesterol apparently control cholesterol 7 alpha-hydroxylase by different mechanisms.
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Some relationships between "natural" physiological autoreactivity and antibody responses against the acetylcholine receptor (AChR) were investigated. Collections of unselected monoclonal antibodies were derived from nonmanipulated newborn mice of susceptible (BALB.B) and resistant (BALB/c) mouse strains, and screened for reactivity with heterologous AChR. Five in 200 BALB/c but none in 200 BALB.B clones were found reactive. Four of these anti-AChR antibodies were analyzed for connectivity to nine other antibodies in the same collection, and three of them found to react with a particular natural anti-idiotype. The functional in vivo significance of such natural idiotypic complementarities in the anti-AChR response could then be investigated. Treatment of BALB/c newborn animals with low (100 ng) or high (100 micrograms) doses of natural anti-idiotype significantly suppressed their anti-AChR antibody response when immunized as adults. The utilization of natural anti-idiotype reagents with broad reactivity patterns might overcome the known idiotype heterogeneity of the autoimmune antibody response to AChR and provide a strategy in prophylactic and therapeutic manipulation of disease development.
The 7 alpha-hydroxylation of two cholesterol analogues, sitosterol and cholestanol, and their effect on the 7 alpha-hydroxylation of cholesterol were measured in rat and human hepatic microsomes. In untreated rat liver microsomes, the 7 alpha-hydroxylation of cholesterol was higher than that of cholestanol (1.4-fold) and sitosterol (30-fold). After removal of endogenous sterols from the microsomes by acetone treatment, the 7 alpha-hydroxylation of cholesterol was similar to that of cholestanol and only fourfold higher than that of sitosterol. Cholestanol and sitosterol competitively inhibited cholesterol 7 alpha-hydroxylase in both rat and human liver microsomes, with cholestanol the more potent inhibitor. Patients with sitosterolemia with xanthomatosis, who have elevated microsomal cholestanol and sitosterol, showed reduced cholesterol 7 alpha-hydroxylase activity relative to the activity in control subjects (13.9 and 14.7 vs. 20.3 +/- 0.9 pmol/nmol P-450 per min, P less than 0.01). Enzyme activity in these patients was 40% higher when measured in microsomes from which competing sterols had been removed. Ileal bypass surgery in one sitosterolemic patient decreased plasma cholestanol and sitosterol concentrations and resulted in a 30% increase in hepatic microsomal cholesterol 7 alpha-hydroxylase activity. Cholesterol 7 alpha-hydroxylase appears to have a specific apolar binding site for the side chain of cholesterol and is affected by the presence of cholestanol and sitosterol in the microsomal substrate pool. Reduced bile acid synthesis in sitosterolemia with xanthomatosis may be related to the inhibition of cholesterol 7 alpha-hydroxylase activity by endogenous cholesterol analogues.
To determine cognitive disturbances in recent demyelinating disease, we studied 21 patients with definite or probable multiple sclerosis (MS) of less than two years' duration and nine patients with recently isolated optic neuritis. None had any clinical or social evidence of cognitive impairment. Mild to moderate cognitive impairment. Mild to moderate cognitive impairment was present in 18 (60%) of 30 cases, affecting visual and/or verbal efficiency. These abnormalities were statistically significant when compared with the results of a control group of 29 patients. There was no correlation with a depressive status, between the presence of cognitive impairment and either the degree of handicap or the activity of the disease. The frequency of cognitive dysfunction (60%) appears to be comparable to that reported in other series in which MS evolution is over ten years. The natural history of cognitive functions in MS has to be identified. Neuropsychologic tests could be useful in the diagnosis of monosymptomatic or paucisymptomatic forms of MS (ie, visual or medullary).
In patients with cerebrotendinous xanthomatosis (CTX), diminished cholic acid production is associated with incomplete oxidation of the cholesterol side chain and the excretion of C(25)-hydroxy bile alcohols. The aims of this investigation were 1) to provide quantitative information on the pool size and production rate of chenodeoxycholic acid by the isotope dilution technique; and 2) to investigate the possible existence of a block in chenodeoxycholic acid synthesis and explain the absence of chenodeoxycholic acid precursors in CTX. After the injection of [24-(14)C]chenodeoxycholic acid, measurements of chenodeoxycholic acid pool size and production rate in a CTX subject were, respectively, 1/20 and 1/6 as great as controls. Further, three potential precursors of chenodeoxycholic acid, namely [G-(3)H]7alpha-hydroxy-4-cholesten-3-one, [G-(3)H]5beta-cholestane-3alpha,7alpha,25-triol, and [G-(3)H]5beta-cholestane-3alpha,7alpha,26-triol, were administered to the CTX and control subjects and the specific activity curves of [G-(3)H]cholic acid and [G-(3)H]chenodeoxycholic acid were constructed and compared. In the control subjects, the two bile acids decayed exponentially, but in the CTX patient maximum specific activities were abnormally delayed, indicating the hindered transformation of precursor into bile acid. These results show that chenodeoxycholic acid synthesis is small in CTX and that the conversion of 7alpha-hydroxy-4-cholesten-3-one, 5beta-cholestane-3alpha,7alpha,25-triol, and 5beta-cholestane-3alpha,7alpha,26-triol to both chenodeoxycholic acid and cholic acid were similarly impaired.
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This paper describes a pathway of cholic acid synthesis, in man and in the rat, which involves 25-hydroxylated intermediates and is catalyzed by microsomal and soluble enzymes. The subcellular localization, stereospecificity, and other properties of the enzymes involved were studied with liver fractions of normolipidemic subjects, cerebrotendinous xanthomatosis patients, and rats. 5beta-Cholestane-3alpha,7alpha,12alpha,25-tetrol was converted to 5beta-cholestane-3alpha,7alpha,12alpha,24beta,25-pentol by the microsomal fraction in the presence of NADPH and O2. 5beta-Cholestane-3alpha,7alpha,12alpha,24alpha,25-pentol, 5beta-cholestane-3alpha,7alpha,12alpha,-23xi,25-pentol, and 5beta-cholestane-3alpha,7alpha,12alpha,25,26-pentol were also formed. In the presence of NAD, 5beta-cholestane-3alpha,7alpha,12alpha,24beta,25-pentol, but not the other 5beta-cholestanepentols formed, was converted to cholic acid by soluble enzymes in good yield. These experiments demonstrate the existence of a pathway for side-chain degradation in cholic acid synthesis which does not involve hydroxylation at C-26 or the participation of mitochondria.
The specific activity (concentration) of microsomal HMG CoA reductase of intestinal crypt cells was studied in rats fed sterols and bile acids, either singly or in combination. It was found that the basal activity of the reductase was not suppressed by the administration of relatively large amounts of bile acid (taurocholate or taurochenodeoxycholate). Bile acids reduced the specific activity of the reductase only in rats in which the activity of the enzyme had first been enhanced by biliary diversion or by sitosterol feeding. In addition, bile acid feeding abolished the diurnal elevation of reductase activity that normally occurs between midnight and 2 a.m. In no case did bile acids reduce enzyme activity below basal levels. A pronounced (60%) reduction of intestinal HMG CoA reductase activity was observed in rats fed cholesterol and bile acid in combination. This reduction in activity could not be ascribed to an increase in intestinal bile acid flux but was associated with an increase in sterol concentration within the intestinal crypt cells. These results indicate that dietary sterols and bile acids both play a role in the regulation of intestinal HMG CoA reductase.
Specific activities of the hepatic microsomal enzymes 3-hydroxy-3-methylglutaryl CoA (HMG CoA) reductase and cholesterol 7alpha-hydroxylase were studied in rats fed sterols and bile acids. The administration of bile acids (taurocholate, taurodeoxycholate, taurochenodeoxycholate) at a level of 1% of the diet for 1 wk reduced the activity of HMG CoA reductase. Taurocholate and taurodeoxycholate, but not taurochenodeoxycholate, inhibited cholesterol 7alpha-hydroxylase. Dietary sitosterol produced increases in the specific activity of HMG CoA reductase (3.6-fold) and cholesterol 7alpha-hydroxylase (1.4-fold), and biliary cholesterol concentrations in this group more than doubled. Compared with controls fed the stock diet, the simultaneous administration of sitosterol and taurochenodeoxycholate resulted in a 60% decrease of HMG CoA reductase activity and no change in cholesterol 7alpha-hydroxylase activity or biliary cholesterol concentration. Rats fed sitosterol plus taurocholate had nearly normal HMG CoA reductase activity, but cholesterol 7alpha-hydroxylase was inhibited and biliary cholesterol remained high. Bile acid secretion rates and biliary bile acid composition were similar in controls and sterol-fed animals. In all groups receiving bile acids, biliary secretion of bile acids was nearly doubled and bile acid composition was shifted in the direction of the administered bile acid. It is concluded that the composition of the bile acid pool influences the hepatic concentrations of the rate-controlling enzymes of bile acid synthesis.
Methods were developed for the determination of HMG CoA (3-hydroxy-3-methylglutaryl CoA) reductase activity in subcellular fractions of intestinal mucosa and liver of Wistar strain rats. In the liver, reductase activity was located exclusively in the microsomal fraction. In the intestinal mucosa, activity was found in both mitochondrial and microsomal fractions of crypt cells but not of villi. The microsomal HMG CoA reductases of liver and intestinal mucosa had similar kinetic characteristics and pH optima. However, the activity of the hepatic enzyme differed with age and sex of the experimental animals while that of the intestinal crypt cells did not. Cholestyramine treatment enhanced the activity of the microsomal HMG CoA reductase in both liver and intestinal mucosa. Reductase activity of the intestinal crypt cells was elevated in both jejunum and ileum. The greatest stimulation, both relatively and absolutely, was observed in the distal half of the jejunum.
The effect of phenobarbital administration on the in vitro activity of cholesterol 7alpha-hydroxylase was investigated in two strains of rats. In rats of the Wistar strain the daily injection of phenobarbital (100 mg/kg per day ip for 5 days) produced a 33% increase in hepatic microsomal protein and a sixfold stimulation of specific activity of the enzyme. In rats of the Charles River colony (Sprague-Dawley derived) identical treatment with phenobarbital resulted in a 45% increase in hepatic microsomal protein and no change in the specific activity of cholesterol 7alpha-hydroxylase. Plasma phenobarbital concentrations were three to four times greater in the Wistar rats, suggesting that these strains differ also in their capacity to metabolize phenobarbital.
HMG CoA reductase activity of rat intestinal mucosa has a diurnal rhythm which coincides with the diurnal variation of the hepatic HMG CoA reductase but has a lower amplitude. The rhythmic variation of the intestinal reductase was present in both jejunal and ileal crypt cell microsomes and was not abolished by cholestyramine administration.
The production of bile salts by rat liver is regulated by a feedback mechanism, but it is not known which enzyme controls endogenous bile acid synthesis. In order to demonstrate the biochemical site of this control mechanism, bile fistula rats were infused intravenously with (14)C-labeled bile acid precursors, and bile acid biosynthesis was inhibited as required by intraduodenal infusion of sodium taurocholate. The infusion of taurocholate (11-14 mg/100 g of rat per hr) inhibited the incorporation of acetate-1-(14)C, mevalonolactone-2-(14)C, and cholesterol-4-(14)C into bile acids by approximately 90%. In contrast, the incorporation of 7alpha-hydroxycholesterol-4-(14)C into bile acids was reduced by less than 10% during taurocholate infusion. These results indicate that the regulation of bile acid biosynthesis is exerted via cholesterol 7alpha-hydroxylase provided that hepatic cholesterol synthesis is adequate.
The hepatic biosynthesis of bile salts in the rat has been shown to be controlled homeostatically by the quantity of bile salt returning to the liver via the portal circulation. The feedback mechanism was demonstrated in two kinds of experiments. In the first, rats with bile fistulas were infused intraduodenally with sodium taurocholate 12 hr after surgery. If the rate of infusion was greater than 10 mg per 100 g rat per hr, the increase in bile acid output normally observed in bile fistula rats was prevented. In the second type of experiment, the rats were infused with taurocholate 48-72 hr after biliary diversion, when bile acid output had reached a maximal value. Provided the rate of infusion exceeded 10 mg per 100 g rat per hr, bile acid secretion returned to the low levels observed in intact rats. Previous attempts to demonstrate the feedback control have been unsuccessful because too little bile salt was infused. The taurocholate pool of the experimental animals was measured as approximately 15 mg per 100 g rat; it was calculated from this and the above results that this pool circulated 10-13 times daily.
In a study of the mechanism whereby 5alpha-bile acids are formed from cholestanol, the 7alpha-hydroxylation of cholestanol was investigated in rat liver preparations in vitro. It was found that in the presence of NADPH and oxygen, rat liver microsomes catalyzed the 7alpha-hydroxylation of cholestanol to the same extent as that of cholesterol. The rate of the hydroxylation was enhanced by prior treatment of the experimental rats with cholestyramine (a bile acid sequestrant) or by establishment of bile fistulas-i.e., by partial or complete removal of bile acids from the enterohepatic circulation. The 7-hydroxylation reaction was further stimulated by pretreatment of the animals with phenobarbital, a drug known to produce increased biosynthesis of hepatic endoplasmic membranes. The 7alpha-hydroxylase was inhibited by the reaction product, by sterols with 7-keto or 7beta-hydroxyl groups, and also by mono- and dihydroxy bile acids of the 5beta-series, although cholic acid or taurocholate produced no inhibition unless added in high concentrations. The results of these studies are in accord with the concept that the presence of a Delta(5)-double bond is not required for the enzymatic formation of the 7alpha-hydroxy derivative. The rate of this hydroxylation reaction in vitro appears to depend on the concentration of bile salts in the enterohepatic circulation of the experimental animals from whom the microsomes were obtained.
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