Yasushi Sunami

A Combination Therapy With Simvastatin and Ursodeoxycholic Acid Is More Effective for Cholesterol Gallstone Dissolution Than Is Ursodeoxycholic Acid Monotherapy

Inhibitors of 3-hydroxy,3-methylglutaryl coenzyme A (HMG-CoA) reductase have been reported to decrease the cholesterol saturation index (CSI) in duodenal bile in humans and to prevent formation of cholesterol gallstones in animal studies. We performed a prospective study to evaluate the role of HMG-CoA reductase inhibitors as gallstone-dissolving agents. Fifty patients with radiolucent gallstones in a gallbladder opacifying at drip infusion cholecystography were treated with either 10 mg/day simvastatin plus 600 mg/day ursodeoxycholic acid (group 1, n=26) or 600 mg/day ursodeoxycholic acid alone (group 2, n=24) for 12 months. The ratio of solitary to multiple gallstone cases was 21:29. Plasma lipid levels were assessed and ultrasonographic examination of the gallbladder was performed at baseline and at 3-month intervals during treatment. Duodenal bile sampling was performed in five patients in each group at baseline and after 12 months of treatment. Plasma cholesterol decreased significantly in group 1 but not in group 2. In solitary gallstone cases, no significant difference in dissolution rates was observed between groups 1 (3 of 9, 33%) and 2 (4 of 12, 33%). In contrast, the dissolution rate in multiple gallstone cases was significantly higher in group 1 (12 of 17, 71%) than in group 2 (3 of 12, 25%) (p < 0.01). Bile cholesterol saturation index was significantly decreased (p < 0.01) but did not significantly differ between the two groups. These results suggest that combination therapy with simvastatin and ursodeoxycholic acid is more effective for cholesterol gallstone dissolution than ursodeoxycholic acid monotherapy in patients with multiple gallstones.

Phospholipid alterations in hepatocyte membranes and transporter protein changes in cholestatic rat model.

Biliary components are transported by hepatic adenosine triphosphate-binding cassette (ABC) transporters that are located in canalicular membranes. Physiological transporter function is related to membrane fluidity, which is modulated by the phospholipid composition of the lipid bilayer. We hypothesized that cholestasis may alter transporter function by modifying phospholipid species to protect the cell from cholestatic damage. Therefore, we examined the expression of ABC transport proteins and their mRNA levels in canalicular membrane vesicles isolated from rat liver 6 hr or three days after bile duct ligation. Membrane lipid composition and membrane fluidity of both sinusoidal and canalicular membrane vesicles were also examined. By 6 hr after bile duct ligation, we found a clear increase of mdr2 and bsep mRNA. These changes were associated with an increase of mdr-Pgp and with a clear decrease of mrp2 protein, and small decrease of bsep protein. In addition, mdr1b mRNA showed a strong increase by three days after bile duct ligation. Canalicular membrane fluidity decreased in a marked time-dependent manner, whereas sinusoidal membranes showed biphasic changes: increased fluidity at 6 hr and a decrease at three days. These changes were closely related to the changes of membrane lipid constitution; the saturated/unsaturated fatty acid ratio increased for phosphatidylcholine in canalicular membrane and the reverse occurred in sinusoidal membrane, and those for sphingomyelin showed the opposite pattern. We conclude that cholestasis causes modulation of ABC transporters as well as that of the lipid constitution in lipid bilayer. These may confer cytoprotective resistance to hepatocytes against cholestatic stress.

Factors affecting gallstone recurrence after successful extracorporeal shock wave lithotripsy.

Ninety-six patients treated successively for symptomatic cholelithiasis with extracorporeal shock wave lithotripsy (ESWL) and oral bile acid therapy consisting of ursodeoxycholic acid in daily dosages of 600 mg were prospectively followed for gallstone recurrence for a median of 13 months. Ultrasonography was performed to detect stone recurrence at 3, 6, and 12 months, and then yearly after the termination of therapy. Recurrent stones were found in 17 patients (18%). The cumulative probability of gallstone recurrence was 15.8% at 12 months, 26.1% at 24 months, and 30.7% at 36 months. The probability of stone recurrence over the entire period of observation was not dependent on stone number, whereas the median interval to detection of recurrence was significantly shorter in the patients with multiple stones (2 months) than in those with solitary stones (8 months) (p < 0.05). The rate of impaired gallbladder contractility was higher in patients with recurrence (8/15, 53.3%) when compared with those with no recurrence (15/72, 20.8%) (p < 0.01). Neither age, gender, or stone characteristics predicted stone recurrence. Only one patient with a recurrence reported biliary pain. Of the 15 patients with recurrent stones who opted for further nonsurgical treatment, complete stone disappearance was achieved in 10. Impaired gallbladder function may predict gallstone recurrence after ESWL.

Role of Phospholipase A2 in Cholesterol Gallstone Formation Is Associated with Biliary Phospholipid Species Selection at the Site of Hepatic Excretion

Phospholipase A2 plays a role in cholesterol gallstone development by hydrolyzing bile phospholipids into lysolecithin and free fatty acids. Lysolecithin and polyunsaturated free fatty acids are known to stimulate the synthesis and/or secretion of gallbladder mucin via a prostanoid pathway, leading to enhancing cholesterol crystal nucleation and growth, and therefore, the action of phospholipase A2 is associated, in part, with bile phospholipid fatty acid. To clarify this hypothesis, we evaluated the effect on bile lipid metastability in vitro of replacing phospholipids with lysolecithin and various free fatty acids. Supersaturated model biles were created with an identical composition (cholesterol saturation index, 1.8; egg yolk lecithin, 34 mM; taurocholate, 120 mM; cholesterol, 25 mM) except for 5%, 10%, or 20% replacement of egg yolk lecithin with a combination of palmitoyl–lysolecithin and a free fatty acid (palmitate, stearate, oleate, linoleate, or arachidonate), followed by time-sequentially monitoring of vesicles and cholesterol crystals using spectrophotometer and video-enhanced differential contrast microscopy. Replacement with hydrophilic fatty acids (linoleate and arachidonate) reduced vesicle formation and promoted cholesterol crystallization, whereas an enhanced cholesterol-holding capacity was evident after replacement with hydrophobic fatty acids (palmitate and stearate). These results indicate that the effect of phospholipase A2 on bile lithogenecity is modulated by the fatty acid species in bile phospholipids, and therefore, that the role of phospholipase A2 in cholesterol gallstone formation is dependent, in part, on biliary phospholipid species selection at the site of hepatic excretion.

Effects of Cerivastatin Sodium, a New HMG-CoA Reductase Inhibitor, on Biliary Lipid Metabolism in Patients with Hypercholesterolemia

The use of 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitors has become common in the treatment of hypercholesterolemia. The present uncontrolled study was undertaken to determine the effect of cerivastatin sodium (BAY w 6228), a new HMG-CoA reductase inhibitor, on biliary lipid levels in patients with hypercholesterolemia. Twenty-one hypercholesterolemic patients (World Health Organization type IIa = 16 patients; type IIb = 5 patients) received placebo during a 4- to 6-week observation period, after which they received cerivastatin sodium 0.2 mg/d for 12 weeks. Fasting blood samples were drawn for the measurement of serum lipid levels early in the morning before the start of treatment and once a month for each of the 12 weeks of cerivastatin sodium treatment. Gallbladder bile samples were aspirated with a duodenal tube by cerulein stimulation to assess bile lithogenicity. Serum total cholesterol levels decreased markedly after 12 weeks. However, no significant difference was found in the molar percentage composition of biliary lipids (e.g., cholesterol, phospholipids, and total bile acids) or in individual biliary bile acids. Consequently, no significant change in bile cholesterol saturation index was found. The index values before and after 12 weeks of treatment were 0.81 +/- 0.38 and 0.80 +/- 0.47, respectively, whereas when patients were grouped by type of hypercholesterolemia, there was a tendency toward decreased lithogenicity in patients with type IIb but not type IIa hypercholesterolemia. We concluded that cerivastatin sodium was an effective cholesterol-lowering drug that did not appear to worsen biliary lipid metabolism and that may decrease lithogenicity in patients with type IIb hypercholesterolemia.

Is a role of phospholipase A(2) in cholesterol gallstone formation phospholipid species-dependent?

Phospholipase A(2) plays a role in cholesterol gallstone formation by hydrolyzing bile phospholipids into lysolecithin and free fatty acids. This study investigated its effects on cholesterol crystallization in model bile systems. Supersaturated model bile solutions with different cholesterol saturation indexes (1.2, 1.4, and 1.6) were prepared using cholesterol, taurocholate, and egg yolk phosphatidylcholine, soybean phosphatidylcholine, palmitoyl-oleoyl phosphatidylcholine, or palmitoyl-linoleoyl phosphatidylcholine. Then the effect of digestion of phosphatidylcholine by phospholipase A(2) on bile metastability was assessed by spectrophotometry and video-enhanced differential contrast microscopy. Addition of phospholipase A(2) caused the release of free fatty acids in a time-dependent manner. Cholesterol crystallization was enhanced by an increased crystal growth rate in model bile containing hydrophilic species such as soybean or palmitoyl-linoleoyl phosphatidylcholine, consisting predominantly of polyunsaturated fatty acids. Because phospholipase A(2) enhanced cholesterol crystallization in bile containing hydrophilic phosphatidylcholine species, but not hydrophobic phosphatidylcholine species, release of polyunsaturated fatty acids by hydrolysis may be responsible for such enhancement. Therefore, the role of phospholipase A(2) in cholesterol gallstone formation depends on the phospholipid species present in bile, so that phospholipid species selection during hepatic excretion is, in part, crucial to the cholesterol stone formation.

Gallbladder dysfunction enhances physical density but not biochemical metastability of biliary vesicles.

The gallbladder role in cholesterol gallstone pathogenesis occurs through modulation of bile cholesterol metastability. The present study characterized the effects of concentrating bile on cholesterol crystallization through vesicle transformation, crystal habits, and potentiation of effector substances. Supersaturated model biles with total lipid concentrations of 12, 9, 6, and 3 g/dl were prepared with identical molar ratios (taurocholate–egg yolk phosphatidylcholine–cholesterol: 71:18:11). Bile metastability was assessed spectrophotometrically, and morphology of vesicle and crystal was sequentially scanned by video-enhanced differential contrast microscopy. The effects of replacing 30% of egg yolk phosphatidylcholine with soy bean phosphatidylcholine, 30% of taurocholate with taurodeoxycholate or tauroursodeoxycholate, and addition of concanavalin A-binding glycoprotein on each model bile were examined. By lowering total lipid concentration, cholesterol crystallization was retarded with less fusion and aggregation of vesicles. The effects of substances promoting cholesterol crystallization were enhanced with lesser bile. By replacing 30% of taurocholate with tauroursodeoxycholate, cholesterol crystallization was markedly inhibited in all concentrations, forming stable liquid-crystals. Impaired water absorption by the gallbladder may stabilize vesicles and inhibit rapid cholesterol crystallization, but the potential of cholesterol crystallization effector substances must be modified to alter bile cholesterol metastability.

Impaired gallbladder mucosal function in aged gallstone patients suppresses gallstone recurrence after successful extracorporeal shockwave lithotripsy.

Background:  Absorption of water, as well as emptying of bile, are important functions of the gallbladder. We studied the changes of gallbladder function with age in gallstone patients and their influence on the outcome of extracorporeal shockwave lithotripsy (ESWL).

Dose-Dependent Conjugation of Sulfobromophthalein and Hepatic Transit Time in Bile Fistula Rats

Sulfobromophthalein (BSP) is selectively taken up by the liver and secreted into the bile as unconjugated and conjugated forms. Our previous study demonstrated that unconjugated BSP, but not conjugated BSP, caused the dissociation of biliary lipid secretion from that of bile acids, suggesting that the hepatic BSP conjugation rate partly regulated biliary lipid secretion. To evaluate the mechanisms through which biliary lipid secretion is regulated by exogenous organic anions, we intravenously administered BSP to male Sprague–Dawley rats at various doses either continuously or as a bolus. Then the relationship of the dose of BSP to its conjugation rate, hepatic transit time, and biliary lipid secretion was determined. BSP decreased biliary secretion of cholesterol and phospholipids in a dose-dependent manner without affecting bile acid secretion. In contrast, the proportion of conjugated BSP in bile was associated with the dose. Although the serum clearance of BSP after bolus infusion was constant regardless of the dose administered (50 or 200 nmol/100 g), BSP secretion was delayed with increasing doses: unconjugated BSP was secreted predominantly in the early phase (0–15 min after bolus injection), and conjugated BSP was the predominant form in the late phase (15–30 min). Pretreatment with colchicine reduced the conjugation rate and hepatic transit time of BSP, suggesting that the microtubule-dependent vesicle pathway plays a role in biliary excretion and conjugation of BSP. We conclude that biliary lipid secretion is influenced by organic anions with an affinity for bile acids such as BSP and that this effect is dependent upon the hepatic metabolic rate, i.e., conjugation rate. The hepatic transit time also plays a key role in this process by influencing metabolism.

Tauroursodeoxycholate and taurochenodeoxycholate stabilize bile lipid metastability through different mechanisms: relation to phospholipid fatty acid composition

Abstract Lecithin–cholesterol vesicles are present in bile and play a role in cholesterol metastability. Cholesterol holding capacity of vesicles is regulated by lecithin hydrophobicity. In the present study, the effect of therapeutic bile salts on biliary lipid secretion rate and phospholipid fatty acid composition was evaluated in bile duct-fistula rats to determine whether bile salts modulate biliary phospholipid species to, thereby, regulate a physico-chemical metastability of bile cholesterol. Rats were depleted with bile salt pool by overnight biliary diversion and reinfused intravenously with sodium taurocholate (TC) at a constant rate (200 nmol/min per 100 g b.w.) for 3 h, followed by infusion of either sodium taurochenodeoxycholate (TCDC) or tauroursodeoxycholate (TUDC) at a compatible rate. TCDC-infusion increased the biliary secretion rate of cholesterol and phospholipids with a decrease in molar ratio of cholesterol to phospholipid ( C / P ) when compared to values under TC-infusion, and this was associated with an increase in the molar ratio of phospholipid saturated to unsaturated fatty acids ( S / U ). In contrast, TUDC-infusion decreased such a biliary lipid secretion rate with an increase in C / P , and this was associated with a decrease in a molar ratio of arachidonate in phospholipids. These findings indicate that TCDC enhances a cholesterol packing density of biliary particulate species by decreasing C / P ratio and increasing S / U , whereas TUDC improves bile lithogenecity by decreasing biliary cholesterol and phospholipid secretion rates and reducing bile arachidonyl phospholipids.