Variants and haplotypes in Flap endonuclease 1 and risk of gallbladder cancer and gallstones: a population-based study in China.

The role of FEN1 genetic variants on gallstone and gallbladder cancer susceptibility is unknown. FEN1 SNPs were genotyped using the polymerase chain reaction-restriction fragment length polymorphism method in blood samples from 341 gallbladder cancer patients and 339 healthy controls. The distribution of FEN1-69G > A genotypes among controls (AA, 20.6%; GA, 47.2% and GG 32.2%) was significantly different from that among gallbladder cancer cases (AA, 11.1%; GA, 48.1% and GG, 40.8%), significantly increased association with gallbladder cancer was observed for subjects with both the FEN1-69G > A GA (OR = 1.73, 95% CI = 1.01-2.63) and the FEN1-69G > A GG (OR = 2.29, 95% CI = 1.31-3.9). The distribution of FEN1 -4150T genotypes among controls (TT, 21.8%;GT, 49.3% and GG 28.9%) was significantly different from that among gallbladder cancer cases (TT, 12.9%; GT, 48.4% and GG 38.7%), significantly increased association with gallbladder cancer was observed for subjects with both the FEN1-4150T GT(OR = 1.93, 95% CI = 1.04-2.91) and the FEN1-4150T GG(OR = 2.56, 95% CI = 1.37-5.39). A significant trend towards increased association with gallbladder cancer was observed with potentially higher-risk FEN1-69G > A genotypes (P < 0.001, χ2 trend test) and FEN14150G > T (P < 0.001, χ2 trend test) in gallstone presence but not in gallstone absence (P = 0.81, P = 0.89, respectively). In conclusion, this study revealed firstly that FEN1 polymorphisms and haplotypes are associated with gallbladder cancer risk.

PON3 knockout mice are susceptible to obesity, gallstone formation, and atherosclerosis.

We report the engineering and characterization of paraoxonase-3 knockout mice (Pon3KO). The mice were generally healthy but exhibited quantitative alterations in bile acid metabolism and a 37% increased body weight compared to the wild-type mice on a high fat diet. PON3 was enriched in the mitochondria-associated membrane fraction of hepatocytes. PON3 deficiency resulted in impaired mitochondrial respiration, increased mitochondrial superoxide levels, and increased hepatic expression of inflammatory genes. PON3 deficiency did not influence atherosclerosis development on an apolipoprotein E null hyperlipidemic background, but it did lead to a significant 60% increase in atherosclerotic lesion size in Pon3KO mice on the C57BL/6J background when fed a cholate-cholesterol diet. On the diet, the Pon3KO had significantly increased plasma intermediate-density lipoprotein/LDL cholesterol and bile acid levels. They also exhibited significantly elevated levels of hepatotoxicity markers in circulation, a 58% increase in gallstone weight, a 40% increase in hepatic cholesterol level, and increased mortality. Furthermore, Pon3KO mice exhibited decreased hepatic bile acid synthesis and decreased bile acid levels in the small intestine compared with wild-type mice. Our study suggests a role for PON3 in the metabolism of lipid and bile acid as well as protection against atherosclerosis, gallstone disease, and obesity.

TG-interacting factor 1 acts as a transcriptional repressor of sterol O-acyltransferase 2.

Acat2 [gene name: sterol O-acyltransferase 2 (SOAT2)] esterifies cholesterol in enterocytes and hepatocytes. This study aims to identify repressor elements in the human SOAT2 promoter and evaluate their in vivo relevance. We identified TG-interacting factor 1 (Tgif1) to function as an important repressor of SOAT2. Tgif1 could also block the induction of the SOAT2 promoter activity by hepatocyte nuclear factor 1α and 4α. Women have ∼ 30% higher hepatic TGIF1 mRNA compared with men. Depletion of Tgif1 in mice increased the hepatic Soat2 expression and resulted in higher hepatic lipid accumulation and plasma cholesterol levels. Tgif1 is a new player in human cholesterol metabolism.

Evaluation of lymphocytes CD4+ and CD8+ and expression of ZAP-70 kinase on CD3+ and CD19+ lymphocytes in obese patients undergoing laparoscopic cholecystectomy.

BACKGROUND: Obesity has become a global epidemic and a leading metabolic disease in the world. Laparoscopic surgeries may influence the function of the immunologic system. The percentages of CD4+ and CD8+ T lymphocyte cells have been described as prognostic factors for patients undergoing abdominal surgeries. This study aimed to evaluate the changes in CD4+ and CD8+ T lymphocyte cells, the ratio of CD4+ to CD8+ cells, and the ZAP-70 kinase expression on T CD3+ and B CD19+ cells in obese and normal-weight individuals undergoing laparoscopic cholecystectomy (LC). METHODS: The study group consisted of 46 asymptomatic patients with gallstones shown by ultrasound examination but without signs of any gallbladder complications. The patients underwent planned LC. Blood samples were obtained at three times, and the percentages of studied cells were measured by flow cytometry. Patients were enrolled to two groups: N group (body mass index [BMI], ≤ 25 kg/m(2)) and O group (BMI, ≥ 30 kg/m(2)). For statistical analysis, the Mann-Whitney U test and the Wilcoxon matched-pairs signed-ranks test were used. All p values lower than 0.05 were considered significant. RESULTS: The percentage of CD4+ T cells did not differ between the N and O groups before or after the surgery. Only in the N group did the percentage of CD4+ lymphocytes increase from 0 to 48 h. A higher percentage of CD8+ lymphocytes was observed in the O group postoperatively than in the N group. Differences of ZAP-70 kinase expression in the O group were observed at 24 and 48 h of the study. Decreased expression of ZAP-70 kinase was shown in the N group at both 0-24 and 24-48 h. In the O group, this tendency was noted at 24-48 h. CONCLUSIONS: Immunologic activation after LC was confirmed in both weight groups. However, higher modulation, more typical for open surgeries, was observed in the obese group.

Disruption of the murine protein kinase Cbeta gene promotes gallstone formation and alters biliary lipid and hepatic cholesterol metabolism.

The protein kinase C (PKC) family of Ca(2+) and/or lipid-activated serine-threonine protein kinases is implicated in the pathogenesis of obesity and insulin resistance. We recently reported that protein kinase Cß (PKCß), a calcium-, diacylglycerol-, and phospholipid-dependent kinase, is critical for maintaining whole body triglyceride homeostasis. We now report that PKCß deficiency has profound effects on murine hepatic cholesterol metabolism, including hypersensitivity to diet-induced gallstone formation. The incidence of gallstones increased from 9% in control mice to 95% in PKCß(-/-) mice. Gallstone formation in the mutant mice was accompanied by hyposecretion of bile acids with no alteration in fecal bile acid excretion, increased biliary cholesterol saturation and hydrophobicity indices, as well as hepatic p42/44(MAPK) activation, all of which enhance susceptibility to gallstone formation. Lithogenic diet-fed PKCß(-/-) mice also displayed decreased expression of hepatic cholesterol-7α-hydroxylase (CYP7A1) and sterol 12α-hydroxylase (CYP8b1). Finally, feeding a modified lithogenic diet supplemented with milk fat, instead of cocoa butter, both increased the severity of and shortened the interval for gallstone formation in PKCß(-/-) mice and was associated with dramatic increases in cholesterol saturation and hydrophobicity indices. Taken together, the findings reveal a hitherto unrecognized role of PKCß in fine tuning diet-induced cholesterol and bile acid homeostasis, thus identifying PKCß as a major physiological regulator of both triglyceride and cholesterol homeostasis.

Study of serum lipase, alpha-amylase and pancreatic amylose in gall-stone diseases.

Silent gall-stone causes significant morbidity and mortality and its incidence in India as well as in whole world is on the rise. It has positive correlation with development of carcinoma gall bladder. So far no predictive study has been done to show its correlation with biochemical markers. The present study has been aimed to establish whether simple enzymatic markers can predict association with cholelithiasis. Study group has been selected from the patients attending general surgery OPD of a tertiary healthcare centre with complaints of vague abdominal pain, flatulence and dyspepsia. A total of 61 cases (male = 18, female = 43) were studied and data matched with age and sex matched control. The biochemical markers studied are serum alkaline phosphatase, serum lipase, serum alpha-amylase and serum pancreatic amylase. Patients with obstructive cholelithiasis, duct stones, pancreatic insufficiency and malignancy are excluded from the study. The results were analysed by Student's t-test. Alkaline phosphatase in all the above mentioned cases was not significantly different from the control group (40 female, 21 male healthy individuals). A significant association was found out with serum alpha-amylase (p < 0.05) and a highly significant association was found out with pancreatic amylase (p < 0.001). Results of serum lipase however were inconclusive (p = 0.1). Pancreatic amylase can be estimated at a reasonable cost and costwise may prove to be a marker of gall-stone diseases which are in many cases silent preventing further complications and chances of Malignancy especially where alkaline phosphatase isinconclusive.

ACAT2 and human hepatic cholesterol metabolism: identification of important gender-related differences in normolipidemic, non-obese Chinese patients.

OBJECTIVE: ACAT2 is a major cholesterol esterification enzyme specifically expressed in hepatocytes and may control the amount of hepatic free (unesterified) cholesterol available for secretion into bile or into HDL. This study aims to further elucidate physiologic roles of ACAT2 in human hepatic cholesterol metabolism. METHODS AND RESULTS: Liver biopsies from 40 normolipidemic, non-obese gallstone patients including some gallstone-free patients (female/male, 18/22) were collected and analyzed for microsomal ACAT2 activity, protein and mRNA expression. Plasma HDL-cholesterol (HDL-C) was significantly higher in females than in males, while triglycerides were significantly lower. ACAT2 activity in females was significantly lower than observed in males, regardless of the presence of gallstone disease. Moreover, the activity of ACAT2 correlated negatively with plasma levels of HDL-C (r=-0.57, P<0.05) and with Apo AI (r=-0.49, P<0.05). CONCLUSION: This is the first description of a gender-related difference in hepatic ACAT2 activity in normolipidemic non-obese Chinese patients suggesting a possible role for ACAT2 in the regulation of cholesterol metabolism in humans. The negative correlation between ACAT2 activity and HDL-C or Apo AI may reflect this regulation. Since ACAT2 activity generally has been found to be pro-atherogenic in animal models, the observed sex-related difference may contribute to female protection from complications of coronary heart disease (CHD).

Structural examination of tryptase-, and VIP-positive mast cells in the common bile duct of patients with lithiasis.

The morphology of tryptase-, and vasoactive intestinal polypeptide (VIP)-positive mast cells was examined immunohistochemically in 38 common bile ducts collected from patients with secondary chronic cholangitis and varying degrees of inflammatory activity. Mast cells numbers in chronic exacerbated and chronic sclerotic cholangitis were significantly higher as compared with those in controls (72.4 cells/mm2 and 25.2 cells/mm2 vs. 5.9 cells/mm2; p < 0.0001, Student's t test). The increased number of tryptase-positive mast cells in chronic exacerbated cholangitis correlated with the severeness of inflammatory infiltration. In cases of chronic exacerbated cholangitis, the increased number of mast cells was detected in conjunction with active fibroplasia. In chronic sclerotic cholangitis mast cells were lower in number as compared with exacerbated cholangitis and were observed in relation with inactive fibrosis. Numerous VIP-positive mast cells were found in all patients with cholangitis. Ultrastructural immunocytochemistry showed tryptase positivity to be localized over either electron-dense or particulate granules with a mean diameter of 0.261+/-0.073 microm or 0.171+/-0.053 microm, respectively. VIP positivity was formed as a finely or coarsely granular pattern over larger electron-dense granules of 0.475+/-0.14 microm in diameter. Tryptase-positive mast cells were located mainly in and around surface and glandular epithelium. The involvement of tryptase- and VIP-positive mast cells in inflammation, fibrosis and epithelial reactions in the common bile duct is discussed.

Pigment gallstone pathogenesis: slime production by biliary bacteria is more important than beta-glucuronidase production.

Pigment stones are thought to form as a result of deconjugation of bilirubin by bacterial beta-glucuronidase, which results in precipitation of calcium bilirubinate. Calcium bilirubinate is then aggregated into stones by an anionic glycoprotein. Slime (glycocalyx), an anionic glycoprotein produced by bacteria causing foreign body infections, has been implicated in the formation of the precipitate that blocks biliary stents. We previously showed that bacteria are present within the pigment portions of gallstones and postulated a bacterial role in pigment stone formation through beta-glucuronidase or slime production. Ninety-one biliary bacterial isolates from 61 patients and 12 control stool organisms were tested for their production of beta-glucuronidase and slime. The average slime production was 42 for biliary bacteria and 2.5 for stool bacteria (P <0.001). Overall, 73% of biliary bacteria and 8% of stool bacteria produced slime (optical density >3). In contrast, only 38% of biliary bacteria produced beta-glucuronidase. Eighty-two percent of all patients, 90% of patients with common bile duct (CBD) stones, 100% of patients with primary CBD stones, and 93% of patients with biliary tubes had one or more bacterial species in their stones that produced slime. By comparison, only 47% of all patients, 60% of patients with CBD stones, 62% of patients with primary CBD stones, and 50% of patients with biliary tubes had one or more bacteria that produced beta-glucuronidase. Most biliary bacteria produced slime, and slime production correlated better than beta-glucuronidase production did with stone formation and the presence of biliary tubes or stents. Patients with primary CBD stones and biliary tubes had the highest incidence of slime production. These findings suggest that bacterial slime is important in gallstone formation and the blockage of biliary tubes.

[Bile composition and antibiotic excretion. Observations with T-drainage]. / Gallenzusammensetzung und Antibiotikaausscheidung. Beobachtungen an der T-Drainage.

Patients comparable in disease, therapy and serum bilirubin concentration were either treated with mezlocillin intravenously or not at all. The bile of each patient was collected either from a T-drainage or from a percutaneously placed drainage into the bile ducts. The concentrations of GGT and AP, which were liberated by destroyed liver cells, and of bilirubin and mezlocillin, which were secreted actively, were analysed. Those patients who had normal serum bilirubin concentrations had a significantly higher biliary bilirubin excretion than those with high serum bilirubin level. The maximum excretion was after 4 hours. While the biliary concentration of bilirubin decreased, the concentration of secreted mezlocillin increased. Due to destroyed liver cells those patients with pathologically elevated blood bilirubin levels had a 50-fold lower mezlocillin excretion than those with normal blood values.

Choledocholithiasis: a comparison between the clinical presentations of multiple and solitary stones in the common bile duct.

The clinical presentations of 20 patients with four or more choledochal stones were compared with those of 68 patients who had one to three choledochal stones, investigated during the same time period. All patients underwent endoscopic retrograde cholangiopancreatography (ERCP) and endoscopic sphincterotomy. Patients with multiple choledochal stones usually presented with insidious onset of painless jaundice, simulating malignant bile duct obstruction, in contrast to the abrupt onset of cholangitis or pain experienced by patients with one to three stones. The latter patients had an increased number of duodenal diverticula, higher bilirubins, smaller stones, and fewer positive stones as detected by ultrasound of the bile ducts. Cholesterol crystals were more numerous in duodenal aspirates of patients with multiple choledochal stones. We conclude that multiple choledochal stones have a unique, more smoldering clinical presentation, and that ERCP is the diagnostic procedure of choice. Endoscopic sphincterotomy is an efficient, simple, and safe alternative to surgery when there is no cholecystitis.

Stones in the common bile duct: experience with medical dissolution therapy.

Thirty-one patients with radiolucent common bile duct stones received medical treatment. Nineteen had Rowachol, a terpene preparation, eight (42%) achieving complete stone disappearance within 3 to 48 months. Fifteen (including 3 of the above) took Rowachol with bile acid (chenodeoxycholic in 11, ursodeoxycholic in 4) for 3 to 60 months: 11 (73%) achieved complete dissolution within 18 months. Persistent symptoms and complications settled on conservative management: 8 (25%) patients required admission (2 biliary colic, 1 obstructive jaundice, 4 cholangitis, 1 pancreatitis). One patient died of a myocardial infarction during recovery from pancreatitis; the other continued treatment, 2 achieving complete dissolution/disappearance. Oral dissolution therapy with Rowachol and bile acids should be considered when endoscopic sphincterotomy or surgery is not feasible, but careful attention to potential complications is required while stones persist.

Biliary sequelae of endoscopic sphincterotomy.

Twenty five patients were reviewed a mean of 36 months after successful endoscopic sphincterotomy for the removal of bile duct stones. All the patients had improved symptomatically but 20% had episodes of mild abdominal pain and a similar number had elevated serum gamma glutamyltranspeptidase activities (up to 3 times normal). In 12 patients (50%) biliary gas was demonstrated indicating reflux of duodenal contents. Clinical cholangitis did not occur. Aspiration liver biopsy revealed mild portal tract fibrosis and inflammation in patients with biliary reflux. Biliary reflux was significantly associated with mild upper abdominal pain (P less than 0.05). This study has shown that mild abnormalities of biliary function persist after endoscopic sphincterotomy. The long term consequence of these changes is unclear.

[Correlation between hyperamylasemia and acute pancreatitis]. / Correlazioni tra iperamilasemia e pancreatite acuta.

It is often difficult to differentiate acute pancreatitis (A.P.) from some other acute abdominal diseases, when there is an elevated serum amylase. In contrast, the renal clearance of amylase, expressed as a percentage of creatinine clearance, can separate patients with A.P. from patients with acute colecistitis, common duct stone without pancreatitis, hyperamylasemia after biliary surgery, acute peptic ulcer and acute salivary diseases.

What is involved in endoscopic sphincterotomy for gallstones?

This study reports our first year's experience of endoscopic sphincterotomy for common bile duct stones. Forty patients were considered for the procedure and it was attempted in 37. An effective endoscopic sphincterotomy was eventually achieved in 32 (86 cases per cent), 7 after a "pre-cut". Twenty-three patients passed stones spontaneously (72 per cent), 6 could not be reassessed and 3 still had stones present (9 per cent). Bed occupancy was 9.5 +/- 5.9 days. Complications occurred in 30 per cent, the most common being pancreatitis in 6 cases with 1 death. It is concluded that endoscopic sphincterotomy offers a valuable alternative to surgery in the management of common bile duct stones.

Radiomanometric guides to common bile duct exploration.

The accuracy of operative cholangiograms can be increased by radiomanometry. This study identified 17 patients among 198 studied whose choledocholiths would have likely been missed by routine cholangiography. These are almost invariably small stones the fate of which would be conjectural if left behind. In the present study, if the cholangiogram was negative and the passage pressure within normal limits, calculi were seen only once. If the cholangiogram was positive for choledocholithiasis and the passage pressure elevated, stones were almost invariably found (95 per cent). Such accuracy cannot be expected from either roentgenography alone or determination solely of pressure relationships.

Diagnostic importance of changes in circulating concentrations of immunoreactive trypsin.

A specific and sensitive radioimmunoassay (R.I.A.) has been developed which makes possible the determination of serum or plasma trypsin concentrations despite the presence of trypsin inhibitors, which have invaldiated previously available enzymatic techniques. The assay was most precise at about 300 microng trypsin standard Ag5 per litre of serum, a value comparable with the mean in 76 healthy volunteers (273 microng/1) and in 20 hospital patients with non-pancreatic disease (266 microng/1). Markedly raised concentrations (970-6500 microng/1) were found in all 14 patients with acute pancreatitis and in 8 patients with chronic renal failure (580-1360 microng/1). Abnormal concentrations were found in 11 of 16 patients (69%) with pancreatic cancer (8 high, 3 low) and in 15 of 23 patients (65%) with chronic pancreatitis (3 high, 12 low). Patients with jaundice had normal or marginally lower than normal concentrations unless pancreatic disease or common-duct gallstones were present.