Hif-1α expression targets the TMA/Fmo3/TMAO axis to participate in gallbladder cholesterol stone formation in individuals living in plateau regions.

The incidence of gallbladder cholesterol stones (GCS) increases rapidly among people living in high-altitude hypoxic environments compared to those in normoxic areas. Upregulation of hepatic hypoxia inducible factor 1α (Hif-1α) plays a key role in the formation of GCS. High plasma trimethylamine-N-oxide (TMAO) levels are positively correlated with the occurrence of GCS. We hypothesized that HIF-1α may upregulate TMAO levels by promoting the transcription of flavin-containing monooxygenase 3 (Fmo3), which eventually leads to GCS formation. Our study shows that in women, high plasma total cholesterol and apolipoprotein B were positively correlated with cholecystolithiasis and hypoxia. Hif-1α binds to the Fmo3 promoter and promotes Fmo3 expression. Hypoxia and lithogenic diet induce the expression of Hif-1α, Fmo3, TMAO and cholesterol tube transporters in the livers of mice, disturb the proportion of bile and plasma components, and induce the formation of GCS. In cell experiments, silencing Hif-1α downregulates the expression of Fmo3, TMAO and cholesterol tube transporters. In a mouse model of hypoxic cholecystolithiasis, silencing Hif-1α downregulates the expression of related genes, restores the proportion of bile and plasma lipid components, and reduces the formation of GCS. Our study shows that Hif-1α binds to the promoter region of Fmo3 and promotes Fmo3 transcription. Thus, it mediates the transcriptional activation of the TMA/Fmo3/TMAO pathway, upregulates the expression of ATP-binding cassettes (Abc) g5 and g8, and participates in the regulation of the occurrence of GCS in the plateau region.

Aquaporin-1 and 8 expression in the gallbladder mucosa might not be associated with the development of gallbladder stones in humans.

BACKGROUND: Cholecystolithiasis is a highly prevalent condition in the Western world. Gallbladder stone-related conditions represent the second most common gastrointestinal pathology. Cholesterol stones represent over 80% of gallstones. Cholesterol stones develop secondary to crystallization of bile cholesterol. Water resorption from gallbladder bile via aquaporin in the gallbladder mucosa might play a role in the development of cholesterol stones. This study investigated the expression of Aquaporin-1 (AQP1) and Aquaporin-8 (AQP8) in the human gallbladder mucosa and their possible association with the formation of gallbladder stones. METHODS: The expression of AQP1 and AQP8 in the gallbladder mucosa was examined via immunohistochemical staining. The expression of both AQP1 and AQP8 in the gallbladder mucosa of stone carriers (study group) was compared to that of nonstone carriers (control group). RESULTS: Eighty-four gallbladder specimens from 44 male (52·2%) and 40 female (47·6%) patients were analysed. The study group included 47 specimens from stone carriers, while 37 specimens from stone-free gallbladders were included in the control group. Immunostaining for both AQP1 and AQP8 was positive in 80 cases. AQP1 was expressed both over the apical and intercellular membrane, while AQP8 was expressed only over the apical membrane. A similar distribution was recorded in specimens from the cystic duct. Immunostaining with AQP1 was generally stronger in comparison with AQP8. No significant (P > 0·05) relationship was found between aquaporin expression and the presence or absence of gallbladder stones. CONCLUSION: AQP1 and AQP8 are both expressed in the gallbladder and cystic duct mucosa. However, their role in the development of gallbladder stones is still to be proven.

THE RELATIONSHIP OF THE IMMUNE AND METABOLIC DISORDERS DURING VARIOUS METHODS OF MULTICOMPONENT GENERAL ANESTHESIA IN LAPAROSCOPIC CHOLECYSTECTOMY.

BACKGROUND: In addition to operating injury in the pathogenesis of immunological and metabolic disorders after surgical interventions anesthesia plays an important role. THE AIM: to establish the relationship of the immune and metabolic disorders during various methods ofmulticomponent general anesthesia in conditions of laparoscopic cholecystectomy in patients with cholelithiasis. MATERIALS AND METHODS: Under constant observation there were 68 women admitted to the hospital for surgical treatment of cholelithiasis. Patients were divided into 3 groups depending on multicomponent general anesthesia (halothane, propofol, sevoflurane). We determined the concentration of cytokines (TNFa, IL-la, IL-i/8, IL-4, IL-iRA, IL-2, IFNy), components of the complement system (C,, C3, C4, C, and C, factor H, C,-inhibitor), the activity of neutrophilsperipheral blood, the concentration of the products ofperoxidation, catalase, superoxide dismutase in blood plasma. RESULTS: The level of immune-inflammation and metabolic disorders in patients with cholelithiasis was higher in patients operated with the use of halothane. The use of sevoflurane has had the most positive effect on the studied indices. CONCLUSION: The close correlation between the investigated immune and metabolic parameters on the background of the use of different schemes of multicomponent general anesthesia in patients with cholelithiasis have let to the conclusion that in the conditions of use of sevoflurane has the least place a "tension" immune and oxidative status.

Limits and perspective of oral therapy with statins and aspirin for the prevention of symptomatic cholesterol gallstone disease.

The prevalence of gallstones disease in Western countries is 10 - 15%. Gallstones can be one of two types - cholesterol or pigment - with cholesterol gallstones representing nearly the 80% of the total. Cholesterol and pigment gallstones have different predisposing factors: cholesterol gallstones are related to supersaturated bile in cholesterol, whereas black pigment gallstones are related to hyperbilirubinbilia factors (hemolysis, etc.); these are necessary, but not sufficient, factors to produce gallstones in vivo. Gall bladder mucosa factors (gall bladder secretion of mucin, local bile stasis and production of endogenous biliary ß-glucuronidase) may coexist with the aforementioned factors and facilitate gallstone nucleation and growth. The gold-standard treatment for symptomatic gallstones is laparoscopic cholecystectomy. Several studies have reported a significant reduction in the onset of symptomatic gallstones disease in patients undergoing chronic therapy with statins, which can reduce bile cholesterol saturation. Aspirin, which has been shown to reduce the local production of gall bladder mucins (mucosal or parietal factors of gallstone formation) in animal experimental models, does not appear to reduce the risk of symptomatic gallstones disease when tested alone. The new horizon of oral therapy for the prevention of symptomatic gallstone disease needs to evaluate the long-term effect of statins and chronic aspirin administration in patients with dyslipidemia and/or atherosclerosis.

[Clinical and laboratory evaluation of the efficiency of parenteral nutrition in destructive forms of acute calculous cholecystitis].

A clinical and laboratory studies were undertaken to evaluate the efficiency of perioperative parenteral nutrition in destructive forms of acute calculous cholecystitis. Based on the general and biochemical blood analyses and the time course of changes in humoral immunological parameters, the authors concluded that enteral tube nutrition had a positive impact on the postoperative period. A nasojejunal access reducing the risk of acute pancreatitis, which was used during tube feeding, showed benefits.

[Current concepts of lithogenetic mechanism in the gallbladder and the role of biliary sludge in this process].

A review of current data on cholelithiasis is presented. The concept of staged development of the pathological process in the biliary system is considered starting from dysfunction of gallbladder and sphincter apparatus of extrahepatic biliary ducts via chronic acalculous cholecystitis to chronic calculous cholecystitis. Other issues discussed include biliary acid metabolism; varieties, composition and formation of calculi in the gallbladder; biliary sludge and its role in cholelithogenesis. Debatable questions of sludge nature and composition as well as certain terminological problems are considered.

[Disorders of lipid metabolism in patients with cholelithiasis and gallbladder cholesterosis depending on apolipoprotein E genotype].

The work was aimed to study blood lipid spectrum in 133 patients with cholelithiasis (CL) and 159 with gallbladder cholesterosis (GC) as well as apoE genotypes (based on restriction fragment polymorphism) in 49 and 36 respectively. Lipid composition was shown to significantly differ in the two conditions. LDL cholesterol was increased in GC and TG in CL. A rise in LDL cholesterol in both groups was apparent before the age of 30 yr (34.6 +/- 8.4 and 52.6 +/- 12.9% respectively), that in TG and VLDL after 40 yr. E3/3 genotype (norm) was identified in 75.5 +/- 6.2% of the patients with CL and in 83.4 +/- 6.2% in those with GC (p < 0.05). e4 allele (mutation) equally frequently occurred in 10.2 +/- 4.3 and 8.1 +/- 4.5% of patients with CL and GC (p > 0.5), e2 allele in 14.5 +/- 5.0 and 8.1 +/- 4.5% (p < 0.05). These data suggest that patients of both groups equally frequently suffered disturbances in metabolism of saturated (e2 allele) and polyunsaturated (e4 allele) fatty acids predisposing for hypercholesterolemia and hyperlipidemia. They explain why CL is frequently associated with cholesterosis and GC with the formation of caliculi. However, the absence of significant correlation between CL, GC and alleles e2, e4 suggests participation of other factors in pathogenesis of these diseases (LP(a), LDL heterogeneity).

An immunohistochemical study of osteopontin in pigment gallstone formation.

Mucin glycoproteins from the gallbladder epithelium are thought to contribute to the matrix or nucleus of gallstones and other biomineralization systems. The involved acidic glycoproteins have been reported in bile and gallstones. In addition, osteopontin (Opn) is a noncollagenous acidic bone matrix glycoprotein that possesses calcium-binding properties. To investigate the role of Opn in pigment gallstone formation, the involvement of Opn in pigment gallstone formation was studied immunohistochemically in the gallbladder wall and in the stones. Staining for Opn was strongly positive in the epithelium of stone-laden gallbladders and in their stones. The stone-laden gallbladders were infiltrated by macrophages, which intensely stained for Opn. Sections of the pigment stones, under low magnification, showed a lamellar pattern of Opn immunolabeling and showed a reticular pattern under high magnification. Our results indicate that Opn, an acidic glycoprotein from the gallbladder epithelium, seems to be involved in lithiasis. Opn from macrophages and/or the epithelium seems to help form the matrix protein.

Polysaccharides and mucin 5AC (MUC5AC) expression in gallbladder mucosa of young patients with gallstones as evaluated by spatial visualization and quantification.

The study aimed at examination of tissue expression of polysaccharides and secretory mucin 5AC (MUC5AC) in young patients (up to 25 years of age) with a symptomatic gallstones. For comparison, patients most frequently subjected to cholecystectomy were studied, i.e. patients of approximately 50 years of age with the same diagnosis. In quantitative studies on tissue expression of both mucus components, the modern technique of spatial visualization was applied for the first time. Application of the technique permitted to demonstrate significant positive relationships between expression of glycoproteins (immunocytochemical ABC technique for detection of MUC5AC) and expression of sugar components in mucus (PAS technique) and to confirm suitability of the technique for quantitative appraisal of both histochemical and immunocytochemical reactions. An even higher expression of polysaccharides in the entire mucosa and of MUC5AC was detected in gallbladder epithelium of 50-year-old patients, as compared to young patients with symptomatic gallstones. In the young patients, expression of polysaccharides correlated with inflammatory activity (grading), width of gallbladder wall and PLT level in peripheral blood. A significantly higher expression of polysaccharides in gallbladder epithelium was demonstrated in young patients admitted in the emergency mode to the hospital. These correlations in young patients may suggest a role of both mucus components in pathogenesis of cholelithiasis in this age group. A quantitative appraisal of mucus component expression in the two parts of gallbladder mucosa (epithelium vs. entire mucosa) using spatial visualization technique permitted to more accurately compare production of glycoproteins and of polysaccharides in patients with cholelithiasis and to demonstrate additional correlations of a potential clinical significance.

High concentration of kynurenic acid in bile and pancreatic juice.

Kynurenic acid (KYNA) is an agonist of the G-protein-coupled receptor GPR35, which is predominantly expressed in gastrointestinal tissues. The aim of this study was to determine the content of KYNA in gastric juice, bile and pancreatic juice and intestinal content. KYNA was determined by means of high performance liquid chromatography. The mean concentrations of KYNA in human gastric juice is 9.91 +/- 0.71 nM in contrast to human bile (832.5 +/- 204.1 and 306.8 +/- 35.2 nM) obtained from patients with cholecystolithiasis and obstructive jaundice, respectively. In pigs, the KYNA levels in bile and pancreatic juice are 1,113.3 +/- 63.34 and 757.0 +/- 394.4 nM, respectively. The KYNA concentration increases along the digestive system, reaching 1,638 nM in the colon content. We suggest that the liver and pancreas affect the content of kynurenic acid in the lumen of the digestive tract.

MDR3 mutations associated with intrahepatic and gallbladder cholesterol cholelithiasis: an update.

BACKGROUND: The recurrent microlithiasis represents one of the most frequent clinical forms of lithiasis of the bile ducts. This affection is characterized by the presence of cholesterolic microgallstones on hepatic canaliculars, and belongs to a heterogeneous group of autosomal recessive liver disorders. Radiological diagnosis can be confirmed by analysis of MDR3 gene, coding a protein involved in physiologic translocation of phospholipids in bile. Discovery of MDR3 mutations is of particular interest, since normally associated with good effectiveness of medication by ursodesoxycholic acid. AIM: To review MDR3 mutations in humans associated with recurrent cholesterol microlithiasis and to suggest a practical approach for MDR3 gene analysis. RESULTS: 48 mutations of MDR3 gene have been reported in humans to date, from which 43 (89.5%) in the coding region, and 5 splice site mutations have been associated to cholesterol cholelithiasis. 21 (43.8%) of the 43 precited mutations are located in only 8 exons on 28, near transmembrane or nucleotide binding domains of the protein. From the 22 remaining described mutations, 9 (18.8%) are restricted to exon 14. We suggest therefore to start analysis of MDR3 gene by screening exons 6, 7, 9, 10, 12, 14, 17, 23 and 24 with an appropriate protocol in this diagnosis associated with effective treatment. In conclusion such therapeutic orientation is valuable, since recurrent cholesterolic microlithiasis occurs relatively early in life, and by the fact that recurrence of symptoms may occur despite cholecystectomy, or shock-wave therapy.

[Common occurrence of non-alcoholic fatty liver disease and cholecystolithiasis]. / A nem alkoholos zsírmáj és az epekövesség együttes elofordulásának megfigyelése.

INTRODUCTION: Non-Alcoholic Fatty Liver Disease is an acquired metabolic disease of the liver caused by accumulation of triglycerides in hepatocytes that is followed by necrobiotic inflammatory reaction, fibrosis and cirrhosis. Obesity, insulin resistance, diabetes mellitus and hyperlipidaemia are important pathogenetic factors of the process. It is known that among patients with cholecystolithiasis and diabetes mellitus in their anamnesis complications of cholecystolithiasis occur much more frequently like among patients without diabetes. AIM: The aim of the study is observation of the incidence of cholecystolithiasis and its complications in patients with Non-Alcoholic Fatty Liver Disease and comparison of cholecystolithiasis incidence between healthy population and population with Non-Alcoholic Fatty Liver Disease. METHODS: Abdominal ultrasonographical findings were analysed in patients hospitalised at our department and in outpatients, patients with severe accompanied diseases were excluded of the analysis. The analysed basic file of patients could be considered as a selected file. The independence of the two examined variables was measured by chi(2) test. RESULTS: Steatosis was described in 38% of the examined patients, cholecystolithiasis was described in 16% of patients. Cholecystolithiasis and its complications occur two times more frequently in patients with Non-Alcoholic Fatty Liver Disease (33%) like Non-Alcoholic Fatty Liver Disease in patients with cholecystolithiasis (16%). Complications of cholecystolithiasis occur more frequently among patients with Non-Alcoholic Fatty Liver Disease like in healthy individuals. The chi(2) test did not bring significant results concerning the independence of cholecystolithiasis and Non-Alcoholic Fatty Liver Disease. CONCLUSION: Pathogenetic factors of Non-Alcoholic Fatty Liver Disease participate in the pathogenesis of cholecystolithiasis. Their common pathogenetic factors bring about that the formation of cholecystolithiasis is probably faster than the progression of steatosis.

Role of caveolae in the pathogenesis of cholesterol-induced gallbladder muscle hypomotility.

Muscle cells from human gallbladders (GB) with cholesterol stones (ChS) exhibit a defective contraction, excess cholesterol (Ch) in the plasma membrane, and lower binding of CCK-1 receptors. These abnormalities improved after muscle cells were incubated with Ch-free liposomes that remove the excess Ch from the plasma membrane. The present studies were designed to investigate the role of caveolin-3 proteins (Cav-3) in the pathogenesis of these abnormalities. Muscle cells from GB with ChS exhibit higher Ch levels in the plasma membrane that were mostly localized in caveolae and associated with parallel increases in the expression of Cav-3 in the caveolae compared with that in GB with pigment stones (PS). The overall number of CCK-1 receptors in the plasma membrane was not different between muscle cells from GB with ChS and PS, but they were increased in the caveolae in muscle cells from GB with ChS. Treatment of muscle cells from GB with ChS with a Galpha(i3) protein fragment increased the total binding of CCK-1 receptors (from 8.3 to 11.2%) and muscle contraction induced by CCK-8 (from 11.2 to 17.3% shortening). However, Galpha(q/11) protein fragment had no such effect. Moreover, neither fragment had any effect on muscle cells from GB with PS. We conclude that the defective contraction of muscle cells with excessive Ch levels in the plasma membrane is due to an increased expression of Cav-3 that results in the sequestration of CCK-1 receptors in the caveolae, probably by inhibiting the functions of Galpha(i3) proteins.

Oxidised- and total non-protein bound glutathione and related thiols in gallbladder bile of patients with various gastrointestinal disorders.

BACKGROUND: Glutathione is a tripeptide composed of glutamate, cysteine and glycine, accomplishing a broad range of vital functions. Synthesis of glutathione and cysteine is performed mainly in the liver, whereas most other tissues are supplied with these thiols via sinusoidal efflux into the blood. Since canalicular efflux also occurs, thiols may be present in human bile. However, thiol composition of human gallbladder bile is largely unknown, which makes it difficult to speculate on the exact function of thiols in bile. In this study we report on the levels of non-protein bound thiols in gallbladder bile of patients with various gastrointestinal disorders. METHODS: Gallbladder bile was obtained after cholecystectomy from 30 patients who were operated for pancreatic cancer, duodenal cancer, chronic pancreatitis or cholecystolithiasis. Bile was analysed for non-protein bound total- and oxidised glutathione and related thiols, by high performance liquid chromatography. RESULTS: A more than 100-fold inter-individual variation in non-protein bound thiol levels was found in human gallbladder bile of patients with a variety of gastrointestinal disorders. Bile did contain high amounts of cysteine, whereas much lower levels of glutathione, cysteinylglycine and homocysteine were detected. Most thiols were present in their oxidised forms. CONCLUSION: Thiols are present in considerable amounts in human gallbladder bile of patients with various gastrointestinal disorders, levels of cysteine being much higher than those of glutathione and other thiols. Most thiols were in their oxidised forms, which may indicate the presence of considerable chemical- or oxidative stress in the patients studied here.

Pharmacokinetics of cefepime in bile and gall bladder tissue after prophylactic administration in patients with extrahepatic biliary diseases.

The purpose of this study was to determine the cefepime concentrations in serum, bile and gall bladder tissue after administration of a single dose in patients with extrahepatic biliary diseases for pre-operative antimicrobial prophylaxis. During a 3-year period (1999-2002), 30 patients aged above 18 years with extrahepatic biliary diseases (acute and chronic cholecystitis and symptomatic cholelithiasis) were included in the study. Cefepime concentrations were determined by the agar microbiological diffusion method. A significant correlation between serum and gall bladder tissue concentrations of cefepime with the sampling interval was observed (r2 = 0.771, P<0.0001), whereas no correlation between serum and bile fluid concentrations of the drug was noted. In patients with non-functioning gall bladder, very low tissue levels of cefepime were detected. During the time of surgery, serum and gall bladder tissue concentrations of cefepime exceeded the minimum inhibitory concentration for 90% of the organisms (MIC90) for most common pathogens. Cefepime has the required pharmacokinetic properties to be considered for pre-operative antimicrobial prophylaxis in patients undergoing biliary tract surgery.

Detection of cancer cells and gene expression of cytokines in the peritoneal cavity in patients with gastric cancer.

BACKGROUND: The gene expression of the cytokines interleukin-2 (IL-2) and IL-10 in peritoneal washings was examined in relation to the presence of cancer cells in the peritoneal cavity in patients with gastric cancer. METHODS: Total RNA was extracted from 50-ml peritoneal wash samples from 124 patients (gastric cancer, n = 110; controls, n = 14). Carcinoembrionic antigen (CEA) messenger RNA (mRNA) was used to identify the number of cancer cells in peritoneal wash samples by a real-time reverse transcription-polymerase chain reaction (RT-PCR) method, which method was also used to assay the IL-2 and IL-10 gene expression levels. RESULTS: In the 14 control samples, CEA mRNA was not detected, while CEA mRNA was detected in 2 of the 51 stage I gastric cancer patients. Thus, the specificity of this method for the detection of cancer cells in peritoneal wash samples was 97% (63/65). The CEA-based real-time RT-PCR method demonstrated greater prognostic impact than the traditional cytological method. IL-2 gene expression in peritoneal wash samples that were CEA mRNA-positive was suppressed compared with that in peritoneal wash samples that were CEA mRNA-negative, while IL-10 gene expression did not differ according to the CEA mRNA findings. CONCLUSION: The detection of small numbers of cancer cells in peritoneal wash samples from patients with advanced gastric cancer is a good marker for peritoneal metastatic recurrence. In the peritoneal cavity, cancer cells may escape from immune surveillance by controlling the expression of cytokines.

Large-scale identification of human biliary proteins from a cholesterol stone patient using a proteomic approach.

Gallbladder bile, one of the most important body fluids, is composed of water, inorganic ions, conjugated bile salts, phospholipids, cholesterol, bilirubin, mucin and proteins. The separation and identification of bile proteins remain difficult due to the complexity of this matrix. In the present study, human gallbladder bile was obtained from a cholesterol stone patient, and the proteins were isolated and purified by dialysis, precipitation and delipidation procedures. The resulting proteins were divided into several aliquots. One aliquot was subjected to two-dimensional gel electrophoresis (2DE). The protein spots were then in-gel digested and analyzed by liquid chromatography coupled with tandem mass spectrometry (LC/MS/MS). Another aliquot was directly digested and analyzed by a combination of strong cation-exchange (SCX) and reversed-phase (RP) chromatography prior to tandem mass spectrometry (2D-LC/MS/MS). Eventually, 48 and 218 unique proteins were identified from 2DE/MS and 2D-LC/MS/MS, respectively, resulting in a total of 222 unique identified proteins. Of the 218 proteins identified by 2D-LC/MS/MS, 92 were identified based on more than one unique tryptic peptide, and, of the total 222 proteins, 98 were identified based on more than one unique tryptic peptide.

Gallbladder Na+/H+ exchange activity is up-regulated prior to cholesterol crystal formation.

BACKGROUND: Gallbladder Na+ and H2O absorption are increased prior to gallstone formation and may promote cholesterol nucleation. Na+/H+ exchange (NHE) isoforms NHE2 and NHE3 are involved in gallbladder Na+ transport in prairie dogs. We examined whether increased gallbladder Na+ absorption observed during early gallstone formation is the result of NHE up-regulation. MATERIALS AND METHODS: Native gallbladder and primary cultures of gallbladder epithelial cells (GBECs) harvested from prairie dogs fed nonlithogenic (CON) or 1.2% cholesterol diet for varying lengths of time to induce cholesterol-saturated bile (PreCRYS), cholesterol crystals (CRYS), or gallstones (GS) were used. NHE activity was assessed by measuring dimethylamiloride-inhibitable 22Na+ uptake under H+ gradient in primary GBECs. HOE-694 was used to determine NHE2 and NHE3 contributions. NHE protein and mRNA expression were examined by Western and Northern blots, respectively. RESULTS: Gallbladder total NHE activity was 25.1 +/- 1.3 nmol mg protein(-1) min(-1) in the control and increased during gallstone formation peaking at the PreCRYS stage (98.4 +/- 3.9 nmol mg protein(-1) min(-1)). There was a shift in NHE activity from NHE2 to NHE3 as the animals progressed from no stones through the PreCRYS and CRYS stages to gallstones. The increase in NHE activity was partly caused by an increased Vmax without any change in K(Na)m. Both NHE2 and NHE3 protein increased moderately during the PreCRYS stage without increases in mRNA expression. CONCLUSIONS: Increased gallbladder Na+ absorption observed prior to crystal formation is in part caused by an increase NHE activity which is not fully accounted for by an increase in NHE proteins and mRNA levels but may be explained by enhanced localization in the membranes and/or altered regulation of NHE.

Gallstone disease in non-alcoholic fatty liver: prevalence and associated factors.

BACKGROUND: Insulin resistance is a risk factors for non-alcoholic fatty liver disease (NAFLD) and for gallstone disease (GD). Aims of the present study were to assess the prevalence of and factors associated with GD in unselected patients with NAFLD. METHODS: A total of 161 consecutive patients with NAFLD diagnosed through compatible ultrasonography in the absence of known etiologies of liver disease (in all patients) and/or confirmed histologically (in 61 patients), was studied. Gallstone disease was diagnosed through ultrasound scanning or on the basis of previous cholecystectomy. Anthropometric and biochemical variables and concurrent diseases were compared in 32 NAFLD-GD patients and in 129 NAFLD patients without GD (controls) according to gender. RESULTS: The overall prevalence of GD was 19.88%, higher in female patients (P < 0.05), who were older (P < 001). The overall percentage of GD increased with age (P < 0.05). The GD patients had higher uric acid (men), total cholesterol and apolipoprotein B (apo-B) serum concentrations (women; P < 0.05); women also had a higher prevalence of hypertriglyceridemia (P < 0.05). The age-corrected odds ratio of having GD by tertiles increased significantly with increasing uric acid (men) and with increasing total cholesterol, triglycerides and apo-B (women). At univariate continuous analysis GD was associated with insulin 120 min and uric acid in male patients; and with body mass index, insulin 120 min, apo-B, total cholesterol and triglycerides in female patients. On multivariate analysis it was found that among these factors only uric acid in men and apo-B in women were independently associated with GD in NAFLD. CONCLUSIONS: The prevalence of GD in NAFLD is more elevated than reported in the general population. The factors independently associated with GD in NAFLD are different from those reported in the general population and vary according to the gender.