Cortisol metabolism after weight loss: associations with 11 ß-HSD type 1 and markers of obesity in women.

OBJECTIVE: Increased glucocorticoid metabolite excretion and enhanced expression and activity of 11ß-hydroxysteroid dehydrogenase type 1 in adipose tissue are closely correlated with obesity and its detrimental consequences. Weight loss ameliorates the latter. The aim of this study was to explore whether increased glucocorticoid exposure in obesity is improved with substantial weight loss and thus is a consequence rather than a cause of obesity. DESIGN AND PATIENTS: A prospective cohort study in 31 women. MEASUREMENTS: 11ß-HSD type 1 expression and activity, urinary glucocorticoid metabolite excretion, body composition including regional adipose tissue depots and insulin resistance by HOMA-IR before and 2 years after gastric bypass surgery. RESULTS: After weight loss, excretion of cortisol and cortisone metabolites decreased. Both cortisol and cortisone metabolite excretion correlated with central obesity, where the intraabdominal fat depot showed the strongest association. Cortisol metabolites correlated with 11ß-HSD type 1 activity in abdominal subcutaneous adipose tissue. The ratio of cortisol to cortisone metabolites [(5α-tetrahydrocortisol (5αTHF) + tetrahydrocortisol (THF) + α-cortol)/(tetrahydrocortisone (THE) + α-cortolone)] and the ratio of 5α-THF/THF both decreased after stable weight loss, reflecting a downregulation of the net activities of 11ß-HSD type 1 and 5α-reductase. CONCLUSION: Long-term weight loss in women is not only followed by reduced glucocorticoid production, but also favourably decreases the global and tissue-specific activity of the cortisol-activating enzyme 11 ß-HSD type 1, possibly contributing to the health benefits of bariatric surgery.

Picropodophyllin induces downregulation of the insulin-like growth factor 1 receptor: potential mechanistic involvement of Mdm2 and beta-arrestin1.

The insulin-like growth factor 1 receptor (IGF-1R) is crucial for growth and survival of malignant cells. Experience in targeting IGF-1R in cancer models has shown that strategies promoting downregulation of the receptor are much more efficient in inducing apoptosis than those inhibiting the IGF-1R activity. Recently, we found that the cyclolignan picropodophyllin (PPP) inhibits phosphorylation of IGF-1R and activation of downstream signaling without interfering with the highly homologous insulin receptor (IR). Furthermore, PPP treatment caused strong regression of tumor grafts and prolonged survival of animals with systemic tumor disease. Here we demonstrate that PPP also downregulates the IGF-1R, whereas the IR and several other receptors were not affected. PPP-induced IGF-1R downregulation required expression of the MDM2 E3 ligase, which recently was found to ubiquitinate and cause degradation of the IGF-1R. In addition knockdown of beta-arrestin1, the adaptor molecule known to bridges MDM2 and IGF-1R, prevented downregulation of the receptor and significantly decreased PPP-induced cell death. All together these data suggest that PPP downregulates IGF-1R by interfering with the action of beta-arrestin1/MDM2 as well as the achieved receptor downregulation contributes to the apoptotic effect of PPP.

Role of pituitary hormones on 17alpha-ethinylestradiol-induced cholestasis in rat.

Estrogens cause intrahepatic cholestasis in susceptible women during pregnancy, after administration of oral contraceptives, or during postmenopausal hormone replacement therapy. 17alpha-Ethinylestradiol (EE) is a synthetic estrogen widely used to cause experimental cholestasis in rodents with the aim of examining molecular mechanisms involved in this disease. EE actions on the liver are thought to be mediated by estrogen receptor alpha (ERalpha) and pituitary hormones. We tested this hypothesis by analyzing metabolic changes induced by EE in livers from hypophysectomized (HYPOX) and hypothyroid rats. Microarray studies revealed that the number of genes regulated by EE was increased almost 4-fold in HYPOX rat livers compared with intact males. Little overlap was apparent between the effects of EE in intact and HYPOX rats, demonstrating that pituitary hormones play a critical role in the hepatic effects of EE. Consistently, hypophysectomy protects the liver against induction by EE of serum bilirubin and alkaline phosphatase, two markers of cholestasis and hepatotoxicity and modulates the effects of EE on several genes involved in bile acid homeostasis (e.g., FXR, SHP, BSEP, and Cyp8b1). Finally, we demonstrate a novel mechanism of action of EE through binding and negative regulation of glucocorticoid receptor-mediated transcription. In summary, pituitary- and ERalpha-independent mechanisms contribute to development of EE-induced changes in liver transcriptome. Such mechanisms may be relevant when this model of EE-induced cholestasis is evaluated. The observation that the pharmacological effects of estrogen in liver differ in the absence or presence of the pituitary could be clinically relevant, because different drugs that block actions of pituitary hormones are now available.

The insulin-like growth factor-1 receptor inhibitor PPP produces only very limited resistance in tumor cells exposed to long-term selection.

The cyclolignan PPP was recently demonstrated to inhibit the activity of insulin-like growth factor-1 receptor (IGF-1R), without affecting the highly homologous insulin receptor. In addition, PPP caused complete regression of xenografts derived from various types of cancer. These data highlight the use of this compound in cancer treatment. However, a general concern with antitumor agents is development of resistance. In light of this problem, we aimed to investigate whether malignant cells may develop serious resistance to PPP. After trying to select 10 malignant cell lines, with documented IGF-1R expression and apoptotic responsiveness to PPP treatment (IC50s less than 0.1 microM), only two survived an 80-week selection but could only tolerate maximal PPP doses of 0.2 and 0.5 microM, respectively. Any further increase in the PPP dose resulted in massive cell death. These two cell lines were demonstrated not to acquire any essential alteration in responsiveness to PPP regarding IGF-1-induced IGF-1R phosphorylation. Neither did they exhibit any increase in expression of the multidrug resistance proteins MDR1 or MRP1. Consistently, they did not exhibit decreased sensitivity to conventional cytostatic drugs. Rather, the sensitivity was increased. During the first half of the selection period, both cell lines responded with a temporary and moderate increase in IGF-1R expression, which appeared to be because of an increased transcription of the IGF-1R gene. This increase in IGF-1R might be necessary to make cells competent for further selection but only up to a PPP concentration of 0.2 and 0.5 microM. In conclusion, malignant cells develop no or remarkably weak resistance to the IGF-1R inhibitor PPP.

Insulin-like growth factor-I receptor activity is essential for Kaposi's sarcoma growth and survival.

Kaposi's sarcoma (KS) is a highly vascular tumour and is the most common neoplasm associated with human immunodeficiency virus (HIV-1) infection. Growth factors, in particular vascular endothelial growth factor (VEGF), have been shown to play an important role in its development. The role of insulin-like growth factors (IGFs) in the pathophysiology of different tumours led us to evaluate the role of IGF system in KS. The IGF-I receptors (IGF-IR) were identified by immunohistochemistry in biopsies taken from patients with different AIDS/HIV-related KS stages and on KSIMM cells (an established KS-derived cell line). Insulin-like growth factor-I is a growth factor for KSIMM cells with a maximum increase of 3H-thymidine incorporation of 130 +/- 27.6% (P < 0.05) similar to that induced by VEGF and with which it is additive (281 +/- 13%) (P < 0.05). Moreover, specific blockade of the receptor (either by alpha IR3 antibody or by picropodophyllin, a recently described selective IGF-IR tyrosine phosphorylation inhibitor) induced KSIMM apoptosis, suggesting that IGF-IR agonists (IGF-I and -II) mediate antiapoptotic signals for these cells. We were able to identify an autocrine loop essential for KSIMM cell survival in which IGF-II is the IGF-IR agonist secreted by the cells. In conclusion, IGF-I pathway inhibition is a promising therapeutical approach for KS tumours.

Lipoprotein metabolism in patients with anorexia nervosa: a case-control study investigating the mechanisms leading to hypercholesterolaemia.

Hypercholesterolaemia is a common finding in patients with anorexia nervosa (AN). To investigate the type, frequency and pathophysiological mechanisms of changes in lipoprotein metabolism in AN we performed a cross-sectional study in fifty-eight female patients (mean age 24.2 years, BMI 15.3 (sd 1.5) kg/m(2)) and fifty-eight healthy age-matched controls (CO; BMI 22.2 (sd 1.7) kg/m(2)). Total cholesterol and LDL-cholesterol were higher in AN (5.5 (sd 1.3) v. 5.0 (sd 0.8) mmol/l, P=0.023; 3.6 (sd 1.1) v. 3.2 (sd 0.7) mmol/l, P=0.025 respectively). LDL particles were significantly more enriched in cholesterol and triacylglycerol in AN. In multiple regression analysis with LDL-cholesterol as the dependent and BMI, total body fat ( %), lathosterol:cholesterol ratio (endogenous cholesterol synthesis), 7alpha-hydroxy-4-cholesten-3-one (bile acid synthesis), non-esterified glycerol, free triiodothyronine and free thyroxine as independent variables, BMI was the only significant predictor in CO (R(2) 0.36, overall P=0.001). In AN the variability of LDL-cholesterol was significantly predicted by total body fat, free thyroxine, BMI, free triiodothyronine and non-esterified glycerol (R(2) 0.55, overall P<0.001). Subgroup analysis between restricting (AN-R) and binge-eating-purging patients (AN-B) indicated that in AN-R changes in lipoproteins, BMI and total body fat were more pronounced. AN-R patients had lower bile acid synthesis than AN-B (P=0.02). We conclude that elevated cholesterol concentrations in AN are generally due to an increase in LDL-cholesterol, which is mostly determined by the severe loss of body fat and the resulting changes in thyroid hormones, increased lipolysis and decreased endogenous cholesterol synthesis with resulting decrease in LDL removal. The clinical subtype of AN plays a major role in the mechanisms leading to hypercholesterolaemia.

Influence of rifampin on serum markers of cholesterol and bile acid synthesis in men.

OBJECTIVE: It has been demonstrated in preliminary studies that rifampin, a semisynthetic antibiotic and known inducer of hepatic cytochrome P450 3A4, reduces serum concentrations of total bile acids only in individuals with liver disease and elevated serum bile acid levels. METHODS: We studied the effect of rifampin on concentrations of surrogate serum markers of cholesterol and bile acid synthesis as well as of cholesterol absorption in 10 male subjects before and after administration of rifampin (600 mg/day) for 6 days. Cholesterol and its precursors were analyzed by gas-liquid chromatography (GLC), bile acid intermediates and individual bile acids by isotope-dilution methods using GLC-mass spectrometry (MS) or by high-performance liquid chromatography (HPLC). RESULTS: Treatment with rifampin resulted in a 70% increase (p = 0.008) of the serum concentration of the bile acid precursor 7alpha-hydroxy-4-cholesten-3-one, which is a marker for bile acid production. Serum total cholesterol was not altered, however, treatment with rifampin elevated the ratio of lathosterol to cholesterol, an indicator of cholesterol synthesis, by 23% (p = 0.037). Interestingly, serum concentration of total bile acids decreased slightly by 29% (p = 0.022), mainly due to a lowering of the secondary bile acid, deoxycholic acid (-60%; p = 0.005). CONCLUSION: A 6-day treatment with rifampin induces a reduction of deoxycholic serum concentrations in healthy men associated with a moderate increase of serum markers of bile acid and endogenous cholesterol synthesis.

Subjects with essential hypertension are more sensitive to the inhibition of 11 beta-HSD by liquorice.

In this intervention study, we have investigated if hypertensive patients are more sensitive to liquorice-induced inhibition of 11 beta-hydroxysteroid dehydrogenase (11 beta-HSD) type 2 than normotensive (NT) subjects and if the response depends on gender. Healthy volunteers and patients with essential hypertension (HT), consumed 100 g of liquorice daily, for 4 weeks, corresponding to a daily intake of 150 mg glycyrrhetinic acid. Office, 24-h ambulatory blood pressure (BP) and blood samples were measured before, during and after liquorice consumption. Effect on cortisol metabolism was evaluated by determining the urinary total cortisol metabolites and urinary free cortisol/free cortisone quotient (Q). The mean rise in systolic BP with office measurements after 4 weeks of liquorice consumption was 3.5 mmHg (p<0.06) in NT and 15.3 mmHg (p=0.003) in hypertensive subjects, the response being different (p=0.004). The mean rise in diastolic BP was 3.6 mmHg (p=0.01) in NT and 9.3 mmHg (p<0.001) in hypertensive subjects, the response also being different (p=0.03). Liquorice induced more pronounced clinical symptoms in women than in men (p=0.0008), although the difference in the effect on the BP was not significant. The increase in Q was prominent (p<0.0001) and correlated to the rise in BP (p=0.02). The rise in BP was not dependant on age, the change in plasma renin activity or weight. We conclude that patients with essential HT are more sensitive to the inhibition of 11 beta-HSD by liquorice than NT subjects, and that this inhibition causes more clinical symptoms in women than in men.

Cholesterol catabolism in patients with acute myelogenous leukemia and hypocholesterolemia: suppressed levels of a circulating marker for bile acid synthesis.

Hypocholesterolemia is a frequent finding in patients with acute myelogenous leukemia (AML) and in other types of malignancies. Since bile acids are major excretion products of cholesterol, the hepatic degradation of cholesterol to bile acids was investigated in AML patients by analyzing a circulating marker for bile acid synthesis. In addition, plasma levels of a marker for cholesterol synthesis were determined. The plasma levels of 7alpha-hydroxy-4-cholesten-3-one, reflecting bile acid production, were markedly lower in patients with AML than in healthy controls. The median levels were 3.3 and 18.5ng/ml (P<0.0001) in the AML patients (n=29) and the healthy subjects (n=16), respectively. The plasma levels of 7-dehydrocholesterol, reflecting hepatic cholesterol synthesis, were similar for the AML patients and the controls. The results show that the conversion of cholesterol to bile acids was suppressed in AML patients, a phenomenon that may result in a decreased intestinal absorption of cholesterol and subsequent hypocholesterolemia.

Effects of treatment with deoxycholic acid and chenodeoxycholic acid on the hepatic synthesis of cholesterol and bile acids in healthy subjects.

The degradation of cholesterol to bile acids is regulated by a negative-feedback mechanism by the bile acids, especially the hydrophobic bile acids, returning to the liver via the portal vein. Chenodeoxycholic acid (CDCA) is a potent suppressor of the cholesterol 7alpha-hydroxylase, the rate-determining enzyme in bile acid formation. CDCA may also suppress hepatic 3-hydroxy-3-methyl glutaryl coenzyme A (HMG CoA) reductase, the rate-limiting enzyme in cholesterol synthesis. Conflicting reports have appeared regarding the suppression on bile acid synthesis by the most hydrophobic bile acid of human bile, deoxycholic acid (DCA). To study the suppressive effects of CDCA and DCA on hepatic cholesterol and bile acid synthesis in humans, 10 healthy subjects were treated with CDCA or DCA for 3 weeks in a randomized cross-over study with a washout period of 4 weeks in between. Serum levels of 7alpha-hydroxy-4-cholesten-3-one, reflecting cholesterol 7alpha-hydroxylase activity, and 7-dehydrocholesterol, reflecting HMG CoA reductase activity, and bile acids were repeatedly measured during the study periods. After 3 weeks of treatment with CDCA or DCA, CDCA constituted 70% and DCA 74% of the total serum bile acids, respectively. CDCA and DCA decreased the serum levels of 7alpha-hydroxy-4-cholesten-3-one by 80% and 75%, respectively. Negative correlations between the percentages of CDCA and DCA and the serum concentration of 7alpha-hydroxy-4-cholesten-3-one were obtained. CDCA reduced the serum level of 7-dehydrocholesterol by 29%, whereas treatment with DCA tended to increase the level of 7-dehydrocholesterol. Treatment of healthy subjects with CDCA and DCA reduces bile acid synthesis. CDCA also inhibits cholesterol synthesis, whereas DCA does not.

Changing food knowledge, food choice, and dietary fiber consumption by using tailored messages.

The aim of this research was to compare the effects of tailored or general nutrition messages on individuals' food knowledge, food consumption, and fiber intake. Past research in the area of food choice has applied the Theory of Reasoned Action or the Theory of Planned Behavior to identify the determinants of choice. Neither theory, however, is structured to allow the researcher to examine a person's attitudes toward a wide range of foods and then to specify which food(s) from among the set of foods to include in a nutrition message. In contrast, the Behavioral Alternatives Model (BAM) allows the researcher to examine attitudes toward behavioral alternatives (i.e. foods) and to identify which foods are most likely to be selected from among a set of foods. We used the BAM to specify the foods to include in a tailored nutrition message. The results indicated that respondents who received a tailored message derived from the BAM had more food-related knowledge and dietary changes in the recommended direction than the respondents who received a general message.

Bile acid synthesis in cultured human hepatocytes: support for an alternative biosynthetic pathway to cholic acid.

The biosynthesis of bile acids by primary cultures of normal human hepatocytes has been investigated. A general and sensitive method for the isolation and analysis of sterols and bile acids was used, based on anion exchange chromatography and gas chromatography-mass spectrometry (GC/MS). Following incubation for 5 days, 8 oxysterols and 8 C(27)- or C(24)-bile acids were identified in media and cells. Cholic and chenodeoxycholic acids conjugated with glycine or taurine were by far the major steroids found, accounting for 70% and 24% of the total, respectively, being consistent with bile acid synthesis in human liver. Small amounts of sulfated 3beta-hydroxy-5-cholenoic acid and 3beta,7alpha-dihydroxy-5beta-cholanoic acid were also detected. Nine steroids were potential bile acid precursors (2% of total), the major precursors being 7alpha, 12alpha-dihydroxy-3-oxo-4-cholenoic acid and its 5beta-reduced form. These 2 and 5 other intermediates formed a complete metabolic sequence from cholesterol to cholic acid (CA). This starts with 7alpha-hydroxylation of cholesterol, followed by oxidation to 7alpha-hydroxy-4-cholesten-3-one and 12alpha-hydroxylation. Notably, 27-hydroxylation of the product 7alpha, 12alpha-dihydroxy-4-cholesten-3-one and further oxidation and cleavage of the side chain precede A-ring reduction. A-Ring reduction may also occur before side-chain cleavage, but after 27-hydroxylation, yielding 3alpha,7alpha, 12alpha-trihydroxy-5beta-cholestanoic acid as an intermediate. The amounts of the intermediates increased in parallel to those of CA during 4 days of incubation. Suppressing 27-hydroxylation with cyclosporin A (CsA) resulted in a 10-fold accumulation of 7alpha, 12alpha-dihydroxy-4-cholesten-3-one and a decrease of the production of CA and its acidic precursors. These results suggest that the observed intermediates reflect an alternative biosynthetic pathway to CA, which may be quantitatively significant in the cells.

Endocrine effects of the podophyllotoxine derivative drug CPH 82 (Reumacon) in patients with rheumatoid arthritis.

CPH 82 is a non-steroid antirheumatic drug containing two benzylidenated podophyllotoxin glucosides with no affinity for the glucocorticoid receptor. Treatment with CPH 82 as single drug therapy significantly decreased serum and urinary cortisol and cortisol metabolites, serum adrenal androgens and urinary androgen metabolites, plasma ACTH and serum interleukin-6 (IL-6) and tumor necrosis factor alpha (TNF-alpha), and increased serum levels of sex hormone-binding globulin (SHBG). Significant positive correlations were found between serum TNF-alpha and plasma ACTH and between serum IL-6 and TNF-alpha on the one hand and serum and urinary cortisol and cortisol metabolites on the other. The initial action of CPH 82 on adrenal steroidogenesis may be a reduction in cytokine levels due to the drugs' antiinflammatory effect. This causes decreased ACTH stimulation, resulting in a reduced adrenocortical steroid secretion. Accumulation of the drug in the adrenal cortex may also affect adrenal steroidogenesis. The elevated SHBG levels may be caused by a weak estrogenic activity of the drug.

Deoxycholic acid treatment in patients with cholesterol gallstones: failure to detect a suppression of cholesterol 7alpha-hydroxylase activity.

UNLABELLED: Hillebrant C-G, Nyberg B, Angelin B, Axelson M, Björkhem I, Rudling M, Einarsson C (Huddinge University Hospital and Karolinska Hospital, Karolinska Institute, Stockholm, Sweden). Deoxycholic acid treatment in patients with cholesterol gallstones: failure to detect a suppression of cholesterol 7alpha-hydroxylase activity. J Intern Med 1999; 246: 399-407. OBJECTIVES: Based on animal studies, hydrophobic bile acids have been postulated to be particularly strong inhibitors of bile acid synthesis. The present study was undertaken to characterize in humans the effects of one of the most hydrophobic of the common bile acids, deoxycholic acid (DCA), on the transcriptional regulation and activity of the cholesterol 7alpha-hydroxylase, on hepatic cholesterol metabolism and on biliary lipid metabolism and plasma lipids. DESIGN, SUBJECTS AND SETTINGS: Thirteen patients with cholesterol gallstone disease were treated with DCA (750 mg day-1) for 3 weeks prior to cholecystectomy. Blood samples were collected before and during treatment. At operation, a liver biopsy and gallbladder bile were obtained. Twenty-eight untreated gallstone patients undergoing cholecystectomy served as controls. The study was carried out at a university hospital. RESULTS: Deoxycholic acid comprised 72 +/- 6% (mean +/- SEM) of total biliary bile acids in DCA-treated patients (n = 8), and 21 +/- 2% in the controls (n = 16; P < 0.001). Cholesterol saturation of gallbladder bile averaged 102% in both treated (n = 7) and untreated (n = 16) patients. Cholesterol 7alpha-hydroxylase and HMG CoA reductase activities and mRNA levels were not different between DCA-treated and untreated gallstone patients. The LDL receptor mRNA levels were similar in both groups of patients. Plasma levels of total cholesterol were lowered by 10% upon DCA treatment (P < 0.05). CONCLUSIONS: Treatment with DCA did not significantly affect mRNA levels and activity of hepatic cholesterol 7alpha-hydroxylase or HMG CoA reductase in patients with cholesterol gallstones. There was no effect on the saturation of gallbladder bile, Thus, the present study could not verify that the hydrophobicity of the bile acid pool is a major factor regulating human hepatic cholesterol 7alpha-hydroxylase activity.

Effect of inhibiting HMG-CoA reductase on 7 alpha-hydroxy-4-cholesten-3-one, a marker of bile acid synthesis: contrasting findings in patients with and without prior up-regulation of the latter pathway.

BACKGROUND: Atorvastatin is a potent inhibitor of 3-hydroxy-3-methyl-glutaryl coenzyme A reductase, but its effect on bile acid synthesis is unknown. The objectives of the study were to determine the effect of atorvastatin on bile acid synthesis in patients in whom this process had not been or had been previously up-regulated by pharmacological or surgical means. MATERIALS AND METHODS: Four patients with heterozygous familial hypercholesterolaemia (FH) and partial ileal bypass (PIB) and 19 FH heterozygotes without PIB were treated with placebo, atorvastatin 10 mg and atorvastatin 40 mg daily, each regimen for 4 weeks. The non-PIB group was subsequently treated with bile acid (BA) sequestrant 8-16 g daily followed by co-administration of atorvastatin 10 mg, each for 4 weeks. Plasma 7 alpha-hydroxy-4-cholesten-3-one (7 alpha-HCO), a well-validated marker of BA synthesis was measured using high-performance liquid chromatography with UV detection. RESULTS: The plasma 7 alpha-HCO concentration was tenfold higher with placebo in the PIB than in the non-PIB group (418.5 ng mL-1 vs. 39.6 ng mL-1 p = 0.0001). Levels decreased in PIB patients treated with atorvastatin 10 mg and 40 mg daily (350.1 ng mL-1 and 174.0 ng mL-1, P = 0.007 respectively) but did not change significantly in the non-PIB group (44.7 ng mL-1 and 28.3 ng mL-1 respectively). Administration of BA sequestrant to non-PIB patients increased 7 alpha-HCO to 197.4 ng mL-1; this decreased to 106.0 ng mL-1 during co-administration of atorvastatin 10 mg daily (P = 0.0001). CONCLUSION: Atorvastatin decreases the rate of BA synthesis only if the latter is up-regulated by PIB or BA sequestrants, presumably by limiting the supply of newly synthesized free cholesterol.

Failure of cortisone acetate treatment in congenital adrenal hyperplasia because of defective 11beta-hydroxysteroid dehydrogenase reductase activity.

Congenital adrenal hyperplasia in children is often treated with cortisone acetate and fludrocortisone. It is known that certain patients with congenital adrenal hyperplasia require very high substitution doses of cortisone acetate, and a few patients do not respond to this treatment at all. A patient with 21-hydroxylase deficiency, for whom elevated pregnanetriol (P3) levels in urine were not suppressed during treatment with cortisone acetate (65 mg/m2 x day), was examined. The activation of cortisone to cortisol was assessed by measuring urinary metabolites of cortisone and cortisol. The patient's inability to respond to treatment with cortisone acetate was found to be caused by a low conversion of cortisone to cortisol, assumed to be secondary to low 11beta-hydroxysteroid dehydrogenase activity (11-oxoreductase deficiency). All exons and exon/intron junctions of the 11beta-hydroxysteroid dehydrogenase type1 gene (HSD11L) were sequenced without finding any mutations, but a genetic lesion in the promoter or other regulatory regions cannot be ruled out. The deficient 11-oxoreductase activity seems to have been congenital, in this case, but can possibly be attributable to a down-regulation of the enzyme activity. The results support the use of hydrocortisone, rather than cortisone acetate, for substitution therapy in adrenal insufficiency.

HPV16-E7 expression causes fluorodeoxyuridine-mediated radiosensitization in SW620 human colon cancer cells.

We have reported that HT29 colon cancer cells, which are radiosensitized by fluorodeoxyuridine (FdUrd), exhibit a greater increase in cyclin E-dependent kinase activity and progress further into S phase in the presence of FdUrd than do SW620 colon cancer cells, which are only minimally sensitized by this drug (Cancer Res 56: 3203, 1996). Although these findings suggested that the ability to progress into S phase in the presence of FdUrd permits cells to be radiosensitized, we wished to test this hypothesis by attempting to drive SW620 human colon cells into S phase by transducing them with the HPV16-E7 gene. Two-parameter flow cytometry showed that E7-transduced cells progressed through S phase after radiation and FdUrd treatment more rapidly than SW620 parental cells. We found that E7-transduced SW620 cells were significantly radiosensitized by FdUrd (100 nmol/L, 14 hours) with an enhancement ratio for 2 clones of 1.47 +/- 0.03 and 1.51 +/- 0.14, compared with 1.24 +/- 0.04 in SW620 parental cells. These data strongly support the hypothesis that dysregulation of S-phase progression is an important factor in FdUrd-mediated radiosensitization.

The level of 7-dehydrocholesterol in plasma reflects the activity of 3-hydroxy-3-methylglutaryl coenzyme A reductase in the human liver.

Plasma levels of the cholesterol precursor 7-dehydrocholesterol (7-DHC) were compared with activities of the rate-limiting enzyme in cholesterol biosynthesis, 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase assayed in liver biopsies from patients undergoing cholecystectomy. Some patients were treated with cholestyramine, deoxycholic acid or chenodeoxycholic acid prior to surgery in order to alter the activity of the enzyme. The median level of 7-DHC and the activity of HMG-CoA reductase in the untreated group were 55 ng/ml and 98 pmol/min/mg protein, respectively. The sterol levels and enzyme activities were increased in patients treated with cholestyramine (85 ng/ml and 439 pmol/min/mg protein) and deoxycholic acid (86 ng/ml and 173 pmol/min/mg protein) and decreased in patients treated with chenodeoxycholic acid (38 ng/ml and 51 pmol/min/mg protein). There was a strong positive correlation (rs=0.75, P<0.0005) between the plasma levels of 7-DHC and the activities of hepatic HMG-CoA reductase in these patients. This correlation was further improved when the plasma levels of 7-DHC were expressed relative to those of cholesterol (rs=0.90, P<0.0001). The results show that the level of 7-DHC in plasma reflects the activity of HMG-CoA reductase in the liver.

Effects of short-term treatment with pravastatin on the hepatic synthesis of cholesterol and bile acids in gallstone patients.

BACKGROUND: HMG-CoA reductase inhibitors are now the therapy of choice in the treatment of hypercholesterolaemia. The effects of long-term treatment with these substances on plasma lipoproteins, cholesterol metabolism and biliary secretion of lipids have been extensively studied in humans. Much less is known about the effects of short-term treatment. The aim of this study was to determine the time course of the effects of HMG-CoA reductase inhibitors on plasma lipoprotein levels as well as cholesterol and bile acid synthesis in gallstone patients. METHODS: Thirty-six patients undergoing elective cholecystectomy were included in the study. Except for the gallstone disease, these patients were otherwise healthy. Four groups of subjects were treated with the HMG-CoA reductase inhibitor pravastatin (Pravachol), 20 mg twice daily for 12, 24, 48 and 72 h preoperatively. Plasma lipoproteins and plasma levels of lathosterol and 7 alpha-hydroxy-4-cholesten-3-one were determined before initiation of pravastatin treatment and on the morning of the day of the operation, lathosterol reflecting hepatic HMG-CoA reductase activity and 7 alpha-hydroxy-4-cholesten-3-one the activity of cholesterol 7 alpha-hydroxylase, the rate-determining enzyme in bile acid synthesis. RESULTS: All treatment groups displayed a significant decrease in total cholesterol and low-density lipoprotein (LDL)-cholesterol, by about 12% and 17% respectively. Lathosterol was reduced by about 50% in all treatment groups. Of great interest was the finding that 7 alpha-hydroxy-4-cholesten-3-one was unaffected in all treatment groups. CONCLUSION: The results show that short-term pravastatin treatment in gallstone patients rapidly inhibits cholesterol synthesis and lowers plasma LDL-cholesterol levels without effects on bile acid synthesis.