Sandrine Dubrac

Insulin injections enhance cholesterol gallstone incidence by changing the biliary cholesterol saturation index and apo A-I concentration in hamsters fed a lithogenic diet

BACKGROUND/AIMS A link between insulin and cholesterol gallstone disease has often been suspected but never demonstrated. The aim was to evaluate the direct implication of insulin in the gallbladder cholesterol gallstone formation process. METHODS Hamsters fed with a soft-inducing lithogenic diet, enriched with sucrose, were injected daily, for 1 week, either with long-acting insulin or saline (controls). RESULTS Insulin injections doubled the cholesterol gallstone incidence. The cholesterol saturation index (CSI) of bile significantly increased (+19%) and biliary apolipoprotein A-I (apo A-I) decreased, both in concentration (-71%) and the proportion relative to the total biliary proteins (-25%). No modifications in the biliary bile acid composition were noticed. Hepatic HMGCoA reductase activity was higher (+341%), CYP7A1 activity was lower (-52%), whereas CYP27A1 and CYP7B1 were not affected. The hepatic low-density liprotein (LDL)-receptor and SR-BI masses did not vary. The hepatic total cholesterol content increased (+42%). Fasting plasma phospholipid and triglyceride concentrations significantly decreased (-15 and -60%, respectively), but the cholesterol concentration remained constant. CONCLUSIONS These results suggest that insulin injections enhance cholesterol gallstone incidence by increasing the CSI of bile and decreasing the concentration and proportion of a biliary anti-nucleating protein, apo A-I. Insulin modulates the major enzymes of cholesterol and bile acid metabolisms in vivo.

Assay of microsomal oxysterol 7α-hydroxylase activity in the hamster liver by a sensitive method: in vitro modulation by oxysterols

A method of assaying hepatic cytochrome P-450, oxysterol 7alpha-hydroxylase (CYP7B), was developed by combining the use of 25-[26,27-(3)H]hydroxycholesterol as a substrate and hydroxypropyl-beta-cyclodextrin as a substrate vehicle. When these assay conditions were tested, an undesirable transformation was observed of the reaction product, 7alpha,25-dihydroxycholesterol, into 3-oxo-7alpha,25-dihydroxy-4-cholesten by the activity of 3beta-hydroxy-Delta(5)-C(27) steroid oxydoreductase, a microsomal NAD(+) and NADP(+) dependent enzyme of bile acid metabolism. A great improvement was reached by using a continuous NADPH generating system which constantly re-transforms NADP(+) into NADPH, thus inhibiting this activity. This improved CYP7B assay, comparable to our previously described assay for cholesterol 7alpha-hydroxylase (CYP7A), allowed a 3-fold increase of the apparent enzyme activity. The possibility to simultaneously measure CYP7A and CYP7B activities on the same microsomal preparation was investigated. A marked decrease (-33%) in the CYP7B activity was noticed, while that of CYP7A remained unchanged. The CYP7B activity was observed to be inhibited by cholesterol (-30%) and also by the oxysterols 7alpha-hydroxycholesterol (-21%), 7beta-hydroxycholesterol (-25%) and epicoprostanol (-20%), and by cyclosporin A (-26%). It can be concluded that this sensible and easy to perform CYP7B assay allows to observe, at least in vitro, a modulation of the enzyme activity by oxysterols.

Diet-dependent effects of insulin infusion on the hepatic lipoprotein receptors and the key enzymes of bile acid synthesis in the hamster.

The effects of an induced hyperinsulinemia on both the cholesterol and bile acid metabolisms were analyzed in the hamster. The role of dietary sucrose as modulator of these effects was evaluated by feeding the animals with two semi-synthetic diets containing a low (SD, 20%) and a high (LD, 62.5%) sucrose proportion. Hamsters fed under basal nutritional conditions (chow diet, CD) were also used. LD enabled the consequences of an insulin infusion on cholesterol gallstone formation to be evaluated. Subcutaneous osmotic pumps were implanted in all the animals and delivered either 3 IU/day of insulin (insulin groups: CDI, SDI, LDI) or saline (control groups: CDC, SDC, LDC). Several parameters bound to lipid metabolism were measured. The plasma cholesterol concentration remained constant in all the insulin treated groups compared to the controls. Phospholipid and triglyceride concentrations decreased in both the plasma and liver in the CDI and SDI groups. A lower SR-BI mass (around 50%) was found in the liver of CDI and SDI hamsters with concomitant higher hydroxy-methyl-glutaryl coenzyme A reductase activity. The LDL-receptor mass and cholesterol 7alpha-hydroxylase activity in the LDI group were both decreased (-47%, -71% respectively). No variations in the cholesterol gallstone incidence were observed. In conclusion, chronic insulin infusion in growing hamsters induced similar effects on cholesterol metabolism in the CD and SD groups but different ones, between diets containing a low (SD) and a high (LD) sucrose proportion. The distribution of triglycerides and phospholipids in the plasma, liver and bile was also affected by the insulin infusion.

In vivo gallbladder bile diffusion coefficient measurement by diffusion‐weighted echo planar imaging in hamster fed normal and lithogenic diets

It is shown that in vivo measurement of bile water apparent diffusion coefficient (ADC) by diffusion‐weighted echo‐planar imaging (EPI) in hamster gallbladder is possible providing motion artifact‐free ADC values. These ADC values are used to estimate bile viscosity variation induced by normal diets, cholesterol gallstone‐inducing diets, and an antilithiasic drug, and to determine if a link exists between bile viscosity and cholesterol gallstone formation. Measurements were performed at 4.7 T with respiratory triggering in five groups of hamsters fed a commercial (RC) or a semisynthetic (SSD) diet, a SSD containing 0.2% hyodeoxycholic acid (SSD+HDC) and two lithogenic diets (LD5, LD10). ADC decreased significantly in LD10 (2.15 ± 0.07 × 10−3 mm2s−1) and SSD+HDC (2.03 ± 0.04) compared to RC (2.40 ± 0.05) but not in the most lithogenic LD5 diet (2.33 ± 0.06). No direct relationship was found between bile viscosity and gallstone incidence; however, viscosity seems to be related to lipid contents of diets. Magn Reson Med 43:854–859, 2000.