Fat-modified diets influence serum concentrations of cholesterol precursors and plant sterols in hypercholesterolemic subjects

Serum noncholesterol sterols, cholesterol precursors and plant sterols, are indicators of cholesterol absorption and synthesis. Serum plant sterol concentrations correlate positively with cholesterol absorption, but have also been found to correlate with dietary unsaturated to saturated fatty acid ratios. We studied the concentration of serum noncholesterol sterols during four different fat-modified diets, (1) high-fat, saturated fat-enriched (control), (2) reduced-fat, sunflower oil-enriched (SO-enriched), (3) rapeseed oil-enriched (RO-enriched), and (4) reduced-fat, saturated fat-enriched (reduced-fat), followed for 6 months in hypercholesterolemic subjects in a parallel design. The proportion of lathosterol (micrograms per 100 mg cholesterol), a precursor of cholesterol synthesis, increased significantly (P < .05) in both SO-enriched (mean +/- SD 147 +/- 57 v 167 +/- 76, 0 v 6 months) and RO-enriched (147 +/- 54 v 157 +/- 52) groups, where the reduction in low-density lipoprotein (LDL) cholesterol was also significant. The proportion of sitosterol, a plant sterol, decreased significantly in the control group (137 +/- 48 v 122 +/- 42), and the proportion of another plant sterol, campesterol, increased in the RO-enriched group (280 +/- 141 v 333 +/- 162), reflecting changes in the use of vegetable oils in these two groups rather than increased cholesterol absorption. In the whole study population, the proportion of linoleic and alpha-linolenic acid (a marker of the use of RO) in cholesterol esters (CEs) correlated (P < .001) with the proportion of sitosterol (r = .43) and campesterol (r = .36) in serum at the end of the study. In conclusion, serum cholesterol precursors were found to be useful indicators of cholesterol metabolism, but changes in serum plant sterols reflected dietary changes rather than cholesterol metabolism during long-term dietary intervention with fat-modified diets.     

Effect of pectin on serum cholesterol, fecal bile acids and biliary lipids in normolipidemic and hyperlipidemic individuals

Pectin, 40-50 g/day for two weeks administered to nine normolipidemic and hyperlipidemic patients, had no effect on serum triglycerides but did cause a significant decrease in the serum total and unesterified cholesterol of hypercholesterolemic subjects in particular. This was associated with increased excretion of fecal bile acids and total steroids and increased concentration of plasma methyl sterols. Thus, the serum cholesterol reduction by pectin appears to be caused by increased cholesterol elimination into stools as bile acids which is then balanced by enhanced cholesterol synthesis. The composition of biliary bile acids and lipids was not changed and secondary bile acids and sterols decreased inconsistently in feces. The measurement of fecal dry weight suggested that the bulk of the pectin was degraded by bacteria during passage through the intestine. Consequently fecal mass and dry weight were not consistently increased, suggesting that pectin may not be an ideal fibre for increasing fecal bulk in functional colonic disorders.     

Nonselective nerve fibre damage in peripheral nerves after experimental thermocoagulation

BACKGROUND: The insoluble material in supersaturated bile is prerequisite for the formation of gallstones. We therefore studied the biliary precipitable and soluble cholesterol and noncholesterol sterols, including the cholesterol precursor sterols (including lanosterol and lathosterols), and the plant sterols campesterol and sitosterol, and cholestanol, which usually reflect cholesterol synthesis and absorption, respectively, before and after a 6-month treatment with ursodeoxycholic acid (UCDA), 15.4 +/- 4 mg/kg/day (standard error of the mean) or simvastatin (40 mg/day) in 21 patients with cholesterol gallstones, to obtain further information about the factors contributing to the formation of gallstones. METHODS: The sediment and supernatant fractions of duodenal bile samples were separated by ultracentrifugation and analyzed with gas-liquid chromatography. RESULTS: At the base line (n = 21) 50% +/- 3% of biliary cholesterol and a variable amount of the noncholesterol sterols (from 14% of lanosterol to 62% of cholestanol) were in the sediment fraction. The pattern of the noncholesterol sterols in the sediment resembled that of gallstones described previously. At base line body mass index was positively related to the percentage of precipitable cholesterol in bile (r = 0.46, P < 0.05), and the serum sitosterol proportion negatively related to the molar percentage of biliary cholesterol and positively to that of bile acids (r = -0.46 and r = 0.50, P < 0.05 for both). UDCA decreased the precipitable percentage of cholesterol from 46% to 31% (P < 0.03) and simvastatin from 57% to 42% (P = 0.05). Both drugs also decreased the precipitable percentages of lathosterols and cholestanol while increasing that of lanosterol. In relation to cholesterol, the sediment to supernatant ratios of all methylsterols were increased, whereas those of polar lathosterols tended to decrease during UDCA treatment. CONCLUSIONS: Patients with high body mass index have more precipitable cholesterol in their bile. Although both UDCA and simvastatin decreased the precipitable cholesterol, the bile still contained one-third of its cholesterol in the sedimentable form.     

Phase II study of mitoxantrone by 14-day continuous infusion with granulocyte colony-stimulating factor (GCSF) support in patients with metastatic breast cancer and limited prior therapy

The study aim was to compare the ratio of vitamin E to serum cholesterol with the serum vitamin E level alone as a measure of vitamin E status in patients with different degrees of liver dysfunction. Assessment of serum vitamin E and total serum cholesterol was performed in 85 patients with liver cirrhosis at Child's stage A (n = 26), B (n = 26), and C (n = 33) and 50 patients with noncirrhotic liver disease. As surrogate markers of liver function, 7alpha-hydroxycholesterol and prealbumin concentrations and the plasma prothrombin time were determined. Mean serum vitamin E concentrations in Child A, B, and C patients were 27.4%, 36.9%, and 37.3% lower, respectively, than in healthy controls (P<.01). Twelve of 26 Child A, 14 of 26 Child B, and 14 of 33 Child C patients had vitamin E deficiency with respect to the absolute values, i.e., serum levels less than 13.76 micromol/L (5% percentile of healthy controls). In contrast, only two of 26 Child A, five of 26 Child B, and five of 33 Child C patients (P<.01 for Child A/B and P<.05 for Child C) were vitamin E-deficient according to the serum vitamin E to cholesterol ratio, i.e., less than 2.86 micromol/mmol. Serum vitamin E was correlated significantly with prealbumin, 7alpha-hydroxycholesterol, and the plasma prothrombin time, but the vitamin E to cholesterol ratio was not. Correcting serum vitamin E for total serum cholesterol in patients with liver cirrhosis leads to the phenomenon of reduced serum vitamin E levels inadvertently shifted toward normal values. In patients with liver cirrhosis, the absolute vitamin E concentration correlates better with the typical clinical and biochemical findings of the disease than the vitamin E to cholesterol ratio. Therefore, a considerable number of patients with advanced liver cirrhosis might actually be vitamin E-deficient.     

Absorption, metabolism, and serum concentrations of cholesterol in vegetarians: effects of cholesterol feeding

Serum concentrations and metabolism of cholesterol were studied in vegetarians basally and during a dietary cholesterol load. Cholesterol absorption efficiency was normal and synthesis was slightly enhanced, even though serum cholesterol precursors were not increased. The serum concentrations of total and low-density-lipoprotein cholesterol were decreased proportionally to the reduced intake and absolute absorption of cholesterol. Fecal plant sterols were negatively correlated with the absorption efficiency of cholesterol and positively with fecal sterols and cholesterol synthesis, suggesting interference of high plant sterol intakes with cholesterol absorption. Cholesterol saturation and bile acid composition of the bile were not changed. The increased serum plant sterol-cholesterol ratios were positively related to the intake and negatively to the biliary secretion of plant sterols. Cholesterol feeding increased absolute cholesterol absorption and serum concentrations of total and low-density-lipoprotein cholesterol, did not change absorption efficiency or synthesis of cholesterol, but increased fecal cholestanol excretion.     

Impaired absorption of cholesterol and bile acids in patients with an ileoanal anastomosis

BACKGROUND: No data exist on cholesterol absorption in patients with an ileoanal anastomosis (IAA). AIMS: To study cholesterol absorption and its effects on cholesterol and bile acid metabolism in patients with an IAA. PATIENTS AND METHODS: Cholesterol absorption, and serum, biliary, and faecal lipids were studied in 24 patients with an IAA and 20 controls. RESULTS: Fractional cholesterol absorption was significantly lower in the patients (36% versus 47% in controls). Surprisingly, the calculated intestinal influx of endogenous cholesterol was reduced so that the absolute absorption of cholesterol was decreased; elimination of cholesterol as faecal neutral steroids remained normal. Thus, the slightly increased cholesterol synthesis was mainly due to increased faecal bile acid excretion, which, in turn, was associated with reduced absorption and biliary secretion of bile acids. Serum total and low density lipoprotein (LDL) cholesterol and LDL triglycerides were lower in the patients. Molar percentage and saturation index of biliary cholesterol were slightly higher in patients with an IAA. Proportions of secondary bile acids in bile and faeces were diminished, and faecal unidentified bile acids were higher in patients. CONCLUSIONS: Cholesterol absorption is significantly impaired in patients with an IAA, and is closely related to changes in serum and biliary lipids observed in these patients.     

Effects of 25-hydroxycholesterol and 7-ketocholesterol, inhibitors of sterol synthesis, administered orally to mice

When 25-hydroxycholesterol or 7-ketocholesterol was fed to mice with the diet, growth was suppressed and mature mice lost weight. The effect of the 7-ketone upon body weight was effectively counteracted by cholesterol whereas cholestanol and beta-sitosterol were ineffective. Growth repression due to 25-hydroxycholesterol was only partially relieved by cholesterol. The effects of 25-hydroxycholesterol and 7-ketocholesterol upon body weight were related to an apparent effect upon appetite. However the sterols were not unpalatable since diets containing them were not rejected in favor of control diet. Intestinal sterol synthesis was inhibited soon after the administration of dietary 7-ketocholesterol or 25-hydroxycholesterol but inhibition decreased with prolonged feeding. When fed by gavage, the sterols suppressed intestinal sterol synthesis as soon as 2 h after administration. In contrast, cholesterol administered by gavage did not affect intestinal sterol synthesis during a 24 h test period. When fed with the diet 25-hydroxycholesterol and 7-ketocholesterol did not depress hepatic cholesterol synthesis beyond the low levels found in pair-fed controls. Inhibition of intestinal sterol synthesis was accompanied by a decrease in the concentration of cholesterol in the intestinal mucosa and, usually, by a drop in the molar ratio of cholesterol to phospholipids.     

Incorporation of 4-14C-labeled cholesterol into the cholesterol ester fractions of plasma lipoproteins in patients with cirrhosis of the liver (author's transl)

In vivo incorporation studies with 4-14C-cholesterol were performed in five patients with cirrhosis of the liver. All patients showed low concentrations of total cholesterol, esterified cholesterol, and a low activity of the lecithin: cholesterol acyltransferase activity (LCAT). The pattern of incorporation corresponded to the results known from hypercholesterolemic, normocholesterolemic and hypertriglyceridemic patients. Low cholesterol ester concentrations and low LCAT activity do not modify the pattern of incorporation into the lipoprotein ester fractions.     

Activities of recombinant human cytochrome P450c27 (CYP27) which produce intermediates of alternative bile acid biosynthetic pathways

The primary physiological significance of cytochrome P450c27 (CYP27) has been associated with its role in the degradation of the side chain of C27 steroids in the hepatic bile acid biosynthesis pathway, which begins with 7alpha-hydroxylation of cholesterol in liver. However, recognition that in humans P450c27 is a widely or ubiquitously expressed mitochondrial P450, and that there are alternative pathways of bile acid synthesis which begin with 27-hydroxylation of cholesterol catalyzed by P450c27, suggests the need to reevaluate the role of this enzyme and its catalytic properties. 27-Hydroxycholesterol was thought to be the only product formed upon reaction of P450c27 with cholesterol. However, the present study demonstrates that recombinant human P450c27 is also able to further oxidize 27-hydroxycholesterol giving first an aldehyde and then 3beta-hydroxy-5-cholestenoic acid. Kinetic data indicate that in a reconstituted system, after 27-hydroxycholesterol is formed from cholesterol, it is released from the P450 and then competes with cholesterol for reentry the enzyme active site for further oxidation. Under subsaturating substrate concentrations, the efficiencies of oxidation of 27-hydroxycholesterol and 3beta-hydroxy-5-cholestenal to the acid by human P450c27 are greater than the efficiency of hydroxylation of cholesterol to 27-hydroxycholesterol indicating that the first hydroxylation step in the overall conversion of cholesterol into 3beta-hydroxy-5-cholestenoic acid is rate-limiting. Interestingly, 3beta-hydroxy-5-cholestenoic acid was found to be further metabolized by the recombinant human P450c27, giving two monohydroxylated products with the hydroxyl group introduced at different positions on the steroid nucleus.     

Pruritus as a presenting symptom of chronic hepatitis C

Pruritus is a common symptom in cholestatic liver disease but is rare in chronic hepatitis C. Eight patients with chronic hepatitis C and severe pruritus were compared with regard to biochemical, serological, and histological features to eight disease controls with primary biliary cirrhosis and seven with cirrhosis due to hepatitis C. Among those with severe pruritus associated with chronic hepatitis C, serum aminotransferases were raised in all, alkaline phosphatase in four, and gamma-glutamyl-transpeptidase levels in all except one. Serum cholylglycine levels were elevated in seven of eight patients. Liver biopsies showed moderate to severe fibrosis in all patients and cirrhosis in five. Compared to control subjects with cirrhosis due to hepatitis C but no pruritus, ductopenia, and cholestatic changes were prominent, although less so than in controls with primary biliary cirrhosis. Chronic hepatitis C with moderate to severe fibrosis may result in low-grade cholestasis with pruritus, possibly in association with bile duct disappearance.     

Decreased feedback regulation of low density lipoprotein receptor activity by sterols in leukemic cells from patients with acute myelogenous leukemia

Leukemic cells from patients with acute myelogenous leukemia (AML) have higher low density lipoprotein (LDL) receptor activity than normal white blood and bone marrow cells. The underlying mechanism behind this is unclear. We studied the inhibitory effect of sterols on induction of LDL-receptor activity in leukemic cells from 27 patients with AML and in white blood cells from 13 healthy individuals. The high affinity degradation rate of 125I-labeled LDL was determined in mononuclear blood cells directly after isolation from blood and after incubation for 2 days in medium with 10% lipoprotein-deficient serum with or without various concentrations of 25-hydroxycholesterol + cholesterol. The median sterol concentration for 50% inhibition (IC50) of induction was more than five times higher for leukemic cells than for normal mononuclear cells. At the highest sterol concentration (0.400 microg/mL 25-hydroxycholesterol + 8 microg/mL cholesterol), the LDL-receptor activity was abolished in cells from all healthy individuals while the induction of LDL-receptor activity in cells from three AML patients was unaffected. The LDL-receptor activity of leukemic cells, directly after isolation from blood, correlated with IC50 values (r = 0.53, P = 0.007) and WBC counts (r = 0.72, P = 0.0001) but not with cellular cholesterol levels. The results demonstrate decreased feedback regulation of LDL-receptor activity by sterols in AML cells and support the conclusion that elevated LDL-receptor activity is associated with sterol resistance and cell proliferation. The findings are of potential interest for diagnosis and specific treatment of leukemia.     

Sitostanol ester margarine in dietary treatment of children with familial hypercholesterolemia

In familial hypercholesterolemia (FH) the lowering of serum cholesterol levels should be started in childhood in order to prevent coronary artery disease later in life. However, treatment of children is problematic. We studied the effects of sitostanol (3 g/day) ester dissolved in rapeseed oil margarine as a hypocholesterolemic agent in one homozygous and 14 heterozygous children with FH maintained on a low cholesterol diet for 6 weeks, using a double-blind crossover design. Absorption and synthesis of cholesterol were evaluated by measuring serum plant sterol and cholesterol precursor proportions to cholesterol by gas-liquid chromatography. The compliance was good, and the children could not distinguish by taste the two margarines without and with sitostanol ester. Sitostanol margarine significantly reduced serum total, intermediate density (IDL), and low density lipoprotein (LDL) cholesterol by 11, 26, and 15%, respectively, and increased HDL/LDL cholesterol ratio by 27%. The proportions of serum delta 8-cholestenol, lathosterol, and desmosterol were significantly increased by 36, 19, and 18%, and those of serum cholestanol, campesterol, and sitosterol were significantly decreased by 9, 42 and 29%, respectively, suggesting that cholesterol absorption was decreased and synthesis was compensatorily increased. High basal precursor sterol proportions predicted a high decrease in LDL cholesterol levels. In conclusion, partial replacement of normal dietary fat consumption by sitostanol ester margarine appears to be an effective and safe hypocholesterolemic treatment in children with FH.     

Initial clinical experience with the Ahmed Glaucoma Valve implant in pediatric patients

There is evidence that a low-density lipoprotein (LDL) subfraction profile of increased concentrations of small, dense LDL particles is less common among trained than among sedentary normocholesterolemic men, but it is still uncertain whether there is a similar association in hypercholesterolemia also. Therefore, we determined the lipid and apolipoprotein concentration and composition of six LDL subfractions (density gradient ultracentrifugation) in 20 physically fit, regularly exercising (>three times per week) hypercholesterolemic men and 20 sedentary hypercholesterolemic controls. Trained (maximal oxygen consumption [VO2max], 57.3 +/- 7.4 mL/kg/min) and sedentary (VO2max, 37.5 +/- 8.8 mL/kg/min) individuals (aged 35 +/- 11 years; body mass index [BMI], 23.9 +/- 2.7 kg/m2) were matched for LDL apolipoprotein (apo) B levels (108 +/- 23 and 112 +/- 36 mg/dL, respectively). Trained subjects had significantly lower serum triglyceride (P < .05) and very-low-density lipoprotein (VLDL) cholesterol levels (P < .05) and higher high-density lipoprotein 2 (HDL2) cholesterol levels (P < .01) than sedentary controls. LDL particle distribution showed that trained individuals had significantly less small, dense LDL (d = 1.040 to 1.063 g/mL) and more large LDL (d = 1.019 to 1.037 g/mL) subfraction particles than sedentary controls, despite equal total LDL particle number. Analysis of LDL composition showed that LDL particles of hypercholesterolemic trained men had a higher free cholesterol content than LDL of untrained hypercholesterolemic men. Small, dense LDL in hypercholesterolemic trained men were richer in phospholipids than those in sedentary controls. These data demonstrate the significant influence of aerobic fitness on lipoprotein subfraction concentration and composition, thereby emphasizing the role of exercise in the treatment and risk reduction of hypercholesterolemia.     

Effect of estrogen and progesterone on the expression of hepatic and extrahepatic sterol 27-hydroxylase in baboons (Papio sp)

Sterol 27-hydroxylase plays an important role in cholesterol metabolism in hepatic and extrahepatic tissues. To determine whether female sex steroid hormones influence its expression, we measured plasma and hepatic 27-hydroxycholesterol, hepatic mRNA levels, activity of sterol 27-hydroxylase, and adrenal mRNA levels of this enzyme in baboons (n = 6 per group) treated with placebo, estrogen, estrogen + progesterone, and progesterone. We also measured hepatic cholesterol concentration and hepatic acyl coenzyme A:cholesterol acyltransferase (ACAT) activity to determine their relationship with hepatic sterol 27-hydroxylase activity. Plasma 27-hydroxycholesterol concentration was increased by estrogen and estrogen + progesterone and was negatively correlated with plasma (P = .090) and LDL (P = .026) cholesterol concentrations. Similarly, hepatic sterol 27-hydroxylase activity was increased by estrogen and estrogen + progesterone and was negatively correlated with plasma (P = .056) and LDL (P = .052) cholesterol concentrations but was positively correlated with hepatic and plasma 27-hydroxycholesterol concentrations (P < .001). Hepatic ACAT activity was increased by progesterone (P < .004) and was positively correlated with plasma (P = .002) and LDL (P = .009) cholesterol concentrations but was negatively correlated with hepatic sterol 27-hydroxylase activity (P = .035). Hepatic and adrenal gland mRNA levels for sterol 27-hydroxylase were increased by estrogen alone or in combination with progesterone (P < .05). Hepatic sterol 27-hydroxylase activity was positively correlated with hepatic mRNA levels (P < .001), an observation suggesting that estrogen increases the activity of sterol 27-hydroxylase by increasing its synthesis. Hepatic cholesterol concentration was not influenced by the hormone treatment. These observations suggest that estrogen alone or in combination with progesterone increases the synthesis of sterol 27-hydroxylase in hepatic and extrahepatic tissues, and the increased activity of hepatic sterol 27-hydroxylase resulting from the increased synthesis is associated with a hypolipidemic effect on plasma LDL levels. Furthermore, progesterone alone increases the hepatic ACAT activity, but given in combination with estrogen progesterone does not have the same effect on hepatic ACAT activity. The effect of estrogen on hepatic ACAT activity may be mediated by sterol 27-hydroxylase and its effect on cholesterol metabolism (decreased cholesterol synthesis and increased output of cholesterol in the bile) in liver.     

Quantitative estimations of the contribution of different bile acid pathways to total bile acid synthesis in the rat

BACKGROUND & AIMS: Cholesterol degradation to bile acids occurs via "classic" or "alternative" bile acid biosynthetic pathways. The aim of this study was to assess the contributions of these two pathways to total bile acid synthesis in vivo. METHODS: Rats with biliary fistulas were infused with squalestatin for 24 and 48 hours; specific activities of cholesterol 7 alpha-hydroxylase (C7 alpha H) and sterol 27-hydroxylase (S27H) and rates of bile acid synthesis were determined. RESULTS: Continuous squalestatin infusion (15 micrograms/h) decreased C7 alpha H specific activities to 4% and 12% of paired biliary fistula controls at 24 and 48 hours, respectively (P < 0.05) without any changes in S27H specific activities (82% and 95% of controls). At 24 hours, bile acid synthesis decreased to 43% (P < 0.05) but returned to 87% at 48 hours (P = NS). Cholic acid synthesis decreased at 24 hours but returned to control levels at 48 hours. Similar changes in C7 alpha H, S27H, and bile acid synthesis were observed in primary rat hepatocytes after addition of squalestatin (1.0 mumol/L). CONCLUSIONS: In the face of persistent suppression of C7 alpha H and the classic pathway, an alternative pathway becomes a main pathway of bile acid synthesis capable of generating cholic and chenodeoxycholic acids. The observed induction of bile acid synthesis via an alternative pathway or pathways represents an important mechanism for maintenance of cholesterol homeostasis in the rat.     

Peroxide biosensors and mediated electrochemical regeneration of redox enzymes

BACKGROUND/AIMS/METHODS: Apolipoprotein E polymorphism, affecting intestinal absorption and biliary secretion of bile acids, might also contribute to the variable course and response to drug treatment of primary biliary cirrhosis. To test this possibility, we studied the apo E gene frequency, and the expression and response to drug therapy in different apo E isoforms of 88 patients with primary biliary cirrhosis, randomized to ursodeoxycholic acid, colchicine or placebo treatments for 2 years. RESULTS: The frequency of the epsilon2 allele was 2.4 times higher (p<0.01) in the patients with primary biliary cirrhosis compared with the Finnish population. At entry the patients with the epsilon4 allele were significantly younger (p<0.01) than those with other epsilon alleles, while the severity of primary biliary cirrhosis was similar in the three apolipoprotein E phenotypes. Liver enzymes, acute hepatic inflammation, serum total and low density lipoprotein cholesterol were decreased by ursodeoxycholic acid only in the patients with the epsilon4 and homozygous epsilon3 alleles, but not in those with the epsilon2 allele. Improvements of liver enzyme tests by ursodeoxycholic acid were more marked in the patients with the epsilon4 than other epsilon alleles. CONCLUSIONS: The present data show that in primary biliary cirrhosis the epsilon2 allele is overrepresented, and suggest that the expression of primary biliary cirrhosis and response of the disease to ursodeoxycholic acid treatment are closely related to the apo E polymorphism.     

Detection of retroviral antibodies in primary biliary cirrhosis and other idiopathic biliary disorders

BACKGROUND: Retroviruses have been implicated in the aetiology of various autoimmune diseases. We used immunoblots as a surrogate test to find out whether retroviruses play a part in the development of primary biliary cirrhosis. METHODS: We did western blot tests for HIV-1 and the human intracisternal A-type particle (HIAP), on serum samples from 77 patients with primary biliary cirrhosis, 126 patients with chronic liver disease, 48 patients with systemic lupus erythematosus, and 25 healthy volunteers. FINDINGS: HIV-1 p24 gag seroreactivity was found in 27 (35%) of 77 patients with primary biliary cirrhosis, 14 (29%) of 48 patients with systemic lupus erythematosus, 14 (50%) of 28 patients with chronic viral hepatitis, and nine (39%) of 23 patients with either primary sclerosing cholangitis or biliary atresia, compared with only one (4%) of 24 patients with alcohol-related liver disease or alpha1-antitrypsin-deficiency liver disease, and only one (4%) of 25 healthy volunteers (p=0.003). Western blot reactivity to more than two HIAP proteins was found in 37 (51%) of patients with primary biliary cirrhosis, in 28 (58%) of patients with systemic lupus erythematosus, in 15 (20%) of patients with chronic viral hepatitis, and in four (17%) of those with other biliary diseases. None of the 23 patients with either alcohol-related liver disease or alpha1-antitrypsin deficiency, and only one of the healthy controls showed the same reactivity to HIAP proteins (p<0.0001). Our results showed a strong association between HIAP seroreactivity and the detection of autoantibodies to double-stranded DNA. HIAP seroreactivity was also strongly associated with the detection of mitochondrial, nuclear, and extractable nuclear antigens. INTERPRETATION: The HIV-1 and HIAP antibody reactivity found in patients with primary biliary cirrhosis and other biliary disorders may be attributable either to an autoimmune response to antigenically related cellular proteins or to an immune response to uncharacterised viral proteins that share antigenic determinants with these retroviruses.     

Phytosterolaemia in a Norwegian family: diagnosis and characterization of the first Scandinavian case

Phytosterolaemia (sitosterolaemia) is a very rare inherited sterol storage disease characterized by tendon and tuberous xanthomas and by a predisposition to atherosclerosis. We here describe the first Scandinavian case. The 14-year-old female patient was found to have markedly elevated circulating levels of plant sterols (sitosterol, sitostanol, campesterol, stigmasterol), and the levels of these sterols were 20-50 times higher than in her healthy sister and heterozygous parents. In addition to the usual serum plant sterols we found a new major sterol in the patient tentatively identified as episterol or fecosterol (24-methyliden-cholest-7 (or 8)-en-3 beta-ol). A newly developed method based on the use of deuterium labelled cholesterol and plant sterols was used to measure sterol absorption in the patient and her relatives. Absorption of sitosterol averaged 20% in the patient and ranged from 4 to 8% in the relatives. Absorption of campesterol averaged 31% in the patient and ranged from 15 to 18% in her relatives. Absorption of cholesterol averaged 63% in the patient and ranged from 35 to 45% in the relatives. Cholesterol synthesis appeared to be reduced in the patient and was 46-52% of that of her relatives.     

Improved assay of hepatic microsomal cholesterol 7 alpha-hydroxylase activity by the use of hydroxypropyl-beta-cyclodextrin and an NADPH-regenerating system

Cholesterol 7 alpha-hydroxylase, the key enzyme in bile acid synthesis, has been implicated in atherosclerosis and gallstone disease. The aim of this study was to check if the use of hydroxypropyl-beta-cyclodextrin (HPBCD), a vehicle for solubilizing cholesterol, augmented the rate of 7 alpha-hydroxycholesterol formation in hamster liver microsomes compared to classical assays in which labeled cholesterol was delivered in Tween 80. We observed that [14C]cholesterol carried by HPBCD enhanced the sensitivity of the assay tenfold. However, linearity of 7 alpha-hydroxycholesterol formation with time was short because of the rapid transformation of 7 alpha-hydroxycholesterol into 7 alpha-hydroxy-cholesten-3-one and 7 alpha,12 alpha-dihydroxy-cholesten-3-one when NADPH alone was present in the incubation medium. In order to avoid the transformation of 7 alpha-hydroxycholesterol into 7 alpha-hydroxy-cholesten-3-one, which is essentially NAD(+)-dependent, but is also NADP(+)-dependent, NADPH (1 mmol/l) plus an NADPH-regenerating system must be present in the medium. In this improved assay, the optimal pH was 7.4 and the apparent Km for control and cholestyramine-fed hamsters had a similar value of 315 mumol/l; linearity in the formation of 7 alpha-hydroxycholesterol was also apparent after a relatively short time period (10 min), but with a markedly greater slope of the curve. With a short incubation time (6 min), microsomes from livers of hamsters (five and nine weeks old) that were fed with a commercial ground diet yielded rates of 7 alpha-hydroxycholesterol formation of 115 +/- 10 and 150 +/- 16 pmol/min.mg protein, respectively, whereas microsomes from hamsters fed with a lithogenic sucrose-rich diet (five weeks old) yielded rates of 7 alpha-hydroxycholesterol formation of 77 +/- 7 pmol/min.mg protein, which were significantly lower (-33%) than those of corresponding control hamsters. This improved cholesterol 7 alpha-hydroxylase assay is very sensitive, simple and rapid, and does not necessitate sophisticated equipment. It can be particularly useful for determining cholesterol 7 alpha-hydroxylase activity in liver biopsies from dyslipidemic or lithiasic patients.     

Regulation of rat hepatic 3beta-hydroxysterol delta7-reductase: substrate specificity, competitive and non-competitive inhibition, and phosphorylation/dephosphorylation

The mechanism for the catalytic reduction of the double bond at C-7, 8 in 7-dehydrocholesterol by 3beta-hydroxysterol Delta7-reductase was investigated by testing structurally related sterols as substrates and potential inhibitors. The hepatic smooth endoplasmic reticulum was identified as the site of enzyme activity. All putative substrates contained 27 carbons, but differed from 7-dehydrocholesterol by the addition of either an ethyl substituent at C-24 (7-dehydrositosterol), a double bond at C-22 with a methyl substituent at C-24 (ergosterol), epimerization of the hydroxyl from the 3beta- to 3alpha-configuration (7-dehydroepicholesterol), or a saturated double bond at C-5,6 (lathosterol). Two non-steroidal compounds that inhibit 3beta-hydroxysterol Delta7-reductase in vivo (AY 9944 and BM 15.766) were also tested. Ergosterol, 7-dehydrositosterol, and 7-dehydroepicholesterol were reduced at C-7, 8 to form brassicasterol, sitosterol, and epicholesterol, respectively, but 75% less efficiently than 7-dehydrocholesterol. Increasing concentrations of these sterols competitively inhibited 3beta-hydroxysterol Delta7-reductase activity. The double bond at C-7,8 in lathosterol was not reduced. AY 9944 and BM 15.766 inhibited 3beta-hydroxysterol Delta7-reductase activity non-competitively. 3beta-Hydroxysterol-Delta7-reductase activity declined after microsomes were exposed to alkaline phosphatase, and enzyme activity was increased by phosphorylation with Mg2+, and ATP. These results demonstrate that the reduction of the double bond at C-7,8 requires binding of the enzyme protein with the B-ring of the sterol substrate that contains a double bond at C-5,6. The reaction is hindered by substituents located on the apolar side-chain and epimerization of the hydroxyl group in ring A to a 3alpha-configuration. 3beta-Hydroxysterol Delta7-reductase exists in two forms: an active phosphorylated form and an inactive dephosphorylated form.