SERPINA1 and HSD17B13 Gene Variants in Patients with Liver Fibrosis and Cirrhosis.

BACKGROUND AND AIMS: Two single nucleotide polymorphisms (SNPs) in SERPINA1 (Pi*Z rs28929474 and Pi*S rs17580) are risk factors for developing liver cirrhosis. A recent study identified a common SNP in HSD17B13 (rs72613567) that conferred protection from chronic liver disease. The aim of the present study was to test these associations in a cohort of Lithuanian patients with liver fibrosis or cirrhosis. METHODS: The study included 302 patients with cirrhosis, 127 patients with liver fibrosis (METAVIR stages I-III) and 548 controls, all from Lithuania. SNPs were genotyped by quantitative PCR, using TaqMan allelic discrimination assays. Adjusted p value of ≤ 0.016 was considered significant. RESULTS: Genotype distributions of SERPINA1 and HSD17B13 SNPs were in Hardy-Weinberg equilibrium. SERPINA1 Pi*Z was not associated with liver fibrosis or cirrhosis. HSD17B13 rs10433937 (in high linkage disequilibrium with rs72613567; r 2 =0.96) also showed no overall association with liver disease, but the GG- genotype was associated with reduced risk of liver fibrosis (aOR 0.37, p=0.03). SERPINA1 Pi*S was associated with higher risk of developing hepatic fibrosis (aOR 3.42, p=0.001) and cirrhosis (aOR 2.59, p=0.02). CONCLUSIONS: We found that SERPINA1 Pi*S variant conferred an increased risk of developing liver fibrosis, while SERPINA1 Pi*Z and HSD17B13 rs10433937 were not associated with liver fibrosis or cirrhosis of different aetiology.

TM6SF2 and MBOAT7 Gene Variants in Liver Fibrosis and Cirrhosis.

Previous large-scale genetic studies identified single nucleotide polymorphisms (SNPs) of the TM6SF2 and MBOAT7 genes as risk factors for alcoholic liver cirrhosis and non-alcoholic fatty liver disease. In this study, we tried to evaluate the association between TM6SF2 variant rs58542926 and MBOAT7 variant rs641738 and the risk of hepatic fibrosis or liver cirrhosis of different etiology. In parallel, we also aimed to evaluate whether these two SNPs modify the effects of the PNPLA3 rs738409 risk variant for the development of hepatic fibrosis and liver cirrhosis. The study was conducted at the Department of Gastroenterology, Lithuanian University of Health Sciences Hospital, and included 334 patients with liver cirrhosis, 128 patients with liver fibrosis, and 550 controls. SNPs were genotyped by quantitative PCR, using TaqMan allelic discrimination assays. Overall, TM6SF2 rs58542926 as well as MBOAT7 rs641738 were not linked to hepatic fibrosis, alcohol or hepatitis C virus induced liver cirrhosis in an Eastern European population. These genetic variations also did not mediate the effect of PNPLA3 rs738409 SNP for liver developing liver fibrosis or liver cirrhosis.

PNPLA3 and RNF7 Gene Variants are Associated with the Risk of Developing Liver Fibrosis and Cirrhosis in an Eastern European Population.

BACKGROUND AND AIMS: Genome-wide association studies have revealed an association between the risk of developing liver fibrosis or cirrhosis and the single nucleotide polymorphisms (SNPs) of the PNPLA3, RNF7, MERTK and PCSK7 genes. We aimed to validate these results in an Eastern European population. METHODS: We evaluated the associations between the PNPLA3 (rs738409), RNF7 (rs16851720), MERTK (rs4374383) and PCSK7 (rs236918) variants and liver fibrosis and cirrhosis in a series of consecutive patients recruited at the Department of Gastroenterology, Lithuanian University of Health Sciences Hospital, during the period 2012-2015. The study included 317 individuals with liver cirrhosis, 154 individuals with liver fibrosis, and 498 controls. The studied SNPs were determined using RT-PCR TaqMan assays. RESULTS: MERTK and PCSK7 SNPs were not associated with liver fibrosis or cirrhosis. The PNPLA3 SNP rs738409 was associated with a higher risk of developing liver fibrosis (aOR: 1.65, P=0.001) and cirrhosis (aOR: 1.92, P=5.57*10-7). PNPLA3 genotypes were also associated with higher risk of developing liver fibrosis and cirrhosis in dominant (aOR: 1.98, P=2.20*10-5; aOR: 1.67, P=0.008, respectively) and recessive (aOR: 3.94, P=5.16*10-5; aOR: 3.02, P=0.003, respectively) models. RNF7 rs16851720 was associated with liver cirrhosis comparing CC vs. AA + CA genotypes (aOR: 0.26, P=0.020). CONCLUSION: Our study showed that PNPLA3 rs738409 and RNF7 rs16851720 confer an increased risk of developing liver fibrosis and cirrhosis in this Eastern European population, while the MERTK and PCSK7 SNPs are not associated with these conditions.

Association of HFE gene C282Y and H63D mutations with liver cirrhosis in the Lithuanian population.

BACKGROUND AND OBJECTIVE: Liver cirrhosis is the end-stage disease of chronic liver injury. Due to differences in the natural course of chronic liver diseases, identification of genetic factors that influence individual outcomes is warranted. HFE-linked hereditary hemochromatosis (HH) predisposes disease progression to cirrhosis; however, the role of heterozygous C282Y or H63D mutations in the development of cirrhosis in the presence of other etiological factors is still debated. The aim of this study was to determine the association between heterozygous C282Y and H63D mutations and non-HH liver cirrhosis in Lithuanian population. MATERIALS AND METHODS: The patient cohort consisted of 209 individuals. Diagnosis of cirrhosis was confirmed by clinical, laboratory parameters, liver biopsy, and radiological imaging. Control samples were obtained from 1005 randomly selected unrelated healthy individuals. HFE gene mutations were determined using the PCR-RFLP method. RESULTS: The most common causes of cirrhosis were hepatitis C (33.9%), hepatitis B (13.6%), and alcohol (25.8%). C282Y allele was associated with the presence of cirrhosis (OR=2.07; P=0.005); this was also observed under recessive model for C282Y (OR=2.06, P=0.008). The prevalence of C282Y allele was higher in cirrhotic men than in controls (7.0% vs. 2.8%, P=0.002). The carriage of H63D risk allele (OR=1.54; P=0.02), heterozygous C282Y/wt and homozygous H63D/H63D genotypes were associated with liver cirrhosis in males (OR=2.48, P=0.008, and OR=4.13, P=0.005, respectively). CONCLUSIONS: Heterozygous C282Y mutation of the HFE gene was associated with liver cirrhosis in the Lithuanian population. In gender-related analysis, heterozygous C282Y and homozygous H63D mutations were linked to liver cirrhosis in men, not in women.

Liver and spleen transient elastography predicts portal hypertension in patients with chronic liver disease: a prospective cohort study.

BACKGROUND: To assess correlation between liver or spleen stiffness measurement by transient elastography (TE) and hepatic venous pressure gradient (HVPG) in patients with chronic liver disease as well find optimal and rule in/rule out cut-offs for prognosis of clinically significant (CSPH) and severe (SPH) portal hypertension. METHODS: In this prospective study patients with different chronic liver diseases were included. TE was performed at the same day prior to HVPG measurement. HVPG was measured using catheter tip occlusion technique. Based on HVPG, patients were categorized into groups of CSPH and SPH. Cut-off values were established by applying ROC curve analysis. RESULTS: The study included 107 consecutive patients referred for HVPG measurement or transjugular liver biopsy. Successful spleen TE was performed in 99 of the patients. Liver and spleen TE strongly correlated with HVPG, r = 0.75 and r = 0.62, respectively. Accuracy to detect CSPH was 88.7% for liver stiffness of 17.4 kPa and 77.7% for spleen stiffness of 47.6 kPa. Accuracy to detect SPH was 83.1% for liver stiffness of 20.6 kPa and 77.7 % for spleen stiffness of 50.7 kPa. Liver stiffness <11.4 kPa could rule out CSPH with 55.2% specificity and >21.9 kPa rule in CSPH with 74.4% sensitivity. Liver stiffness <12.1 kPa could rule out SPH with 50.0% specificity and >35 kPa rule in SPH with 58.2% sensitivity. CONCLUSIONS: Liver and spleen stiffness correlate with HVPG and could be used to predict CSPH or SPH. Spleen elastography was not superior to liver elastography in predicting portal hypertension.

Gene polymorphisms of micrornas in Helicobacter pylori-induced high risk atrophic gastritis and gastric cancer.

BACKGROUND AND AIMS: MicroRNAs (miRNAs) are known for their function as translational regulators of tumor suppressor or oncogenes. Single nucleotide polymorphisms (SNPs) in miRNAs related genes have been shown to affect the regulatory capacity of miRNAs and were linked with gastric cancer (GC) and premalignant gastric conditions. The purpose of this study was to evaluate potential associations between miRNA-related gene polymorphisms (miR-27a, miR-146a, miR-196a-2, miR-492 and miR-608) and the presence of GC or high risk atrophic gastritis (HRAG) in European population. METHODS: Gene polymorphisms were analyzed in 995 subjects (controls: n = 351; GC: n = 363; HRAG: n = 281) of European descent. MiR-27a T>C (rs895819), miR-146a G>C (rs2910164), miR-196a-2 C>T (rs11614913), miR-492 G>C (rs2289030) and miR-608 C>G (rs4919510) SNPs were genotyped by RT-PCR. RESULTS: Overall, SNPs of miRNAs were not associated with the presence of GC or HRAG. We observed a tendency for miR-196a-2 CT genotype to be associated with higher risk of GC when compared to CC genotype, however, the difference did not reach the adjusted P-value (odds ratio (OR) - 1.46, 95% confidence interval (CI) 1.03-2.07, P = 0.032). MiR-608 GG genotype was more frequent in GC when compared to controls (OR -2.34, 95% CI 1.08-5.04), but significance remained marginal (P = 0.029). A similar tendency was observed in a recessive model for miR-608, where CC + CG vs GG genotype comparison showed a tendency for increased risk of GC with OR of 2.44 (95% CI 1.14-5.22, P = 0.021). The genotypes and alleles of miR-27a, miR-146a, miR-196a-2, miR-492 and miR-608 SNPs had similar distribution between histological subtypes of GC and were not linked with the presence of diffuse or intestinal-type GC. CONCLUSIONS: Gene polymorphisms of miR-27a, miR-146a, miR-196a-2, miR-492, miR-492a and miR-608 were not associated with the presence of HRAG, GC or different histological subtypes of GC in European subjects.

Acute and 14-day hepatic venous pressure gradient response to carvedilol and nebivolol in patients with liver cirrhosis.

BACKGROUND AND OBJECTIVE: Alternative drug therapies are needed for the treatment of portal hypertension. The aim of this randomized study was to evaluate and compare the effects of carvedilol and nebivolol on the hepatic venous pressure gradient (HVPG) response in the patients with liver cirrhosis. MATERIAL AND METHODS: In total, 20 cirrhotic patients were randomized into 2 groups and treated with carvedilol (n=10) or nebivolol (n=10). HVPG was measured at baseline, 60 minutes after the administration of carvedilol (25 mg) or nebivolol (5 mg), and after 14 days of carvedilol (25 mg) or nebivolol (5 mg) administered daily. RESULTS. Carvedilol significantly reduced HVPG from 22.2 mm Hg (SD, 4.4) to 15.2 mm Hg (SD, 3.7) after 60 minutes and to 16.4 mm Hg (SD, 2.9) after 14 days (P<0.01). Nebivolol reduced HVPG from 19.7 mm Hg (SD, 2.5) to 15.7 mm Hg (SD, 2.6) and 16.7 mm Hg (SD, 3.2), respectively (P<0.02). Carvedilol effectively decreased HVPG in a greater proportion of the patients after an acute probe (88% vs. 57%) and after 14 days of the treatment (88% vs. 28%, P<0.05) in comparison with nebivolol. CONCLUSION: Carvedilol and nebivolol reduce HVPG in cirrhotic patients; however, the effect of carvedilol on the HVPG reduction might be superior to that of nebivolol, especially after 14 days of treatment.

Hepatic venous pressure gradient measurement in patients with liver cirrhosis: a correlation with disease severity and variceal bleeding.

UNLABELLED: The aim of present study was to evaluate relationships between degree of portal hypertension, severity of the disease, and bleeding status in patients with liver cirrhosis. PATIENTS AND METHODS: All study patients with liver cirrhosis underwent hepatic venous pressure gradient measurements, endoscopy, clinical and biochemical evaluation. Liver function was evaluated according to Child-Turcotte-Pugh (Child's) scoring system. Patients with decompensated cirrhosis (presence of severe ascites, acute variceal bleeding occurring within 14 days, hepatorenal syndrome, cardiopulmonary disorders, transaminase levels >10 times higher the upper normal limit), active alcohol intake, use of antiviral therapy and/or beta-blockers were excluded from the study. RESULTS: One hundred twenty-eight patients with liver cirrhosis (male/female, 67/61; mean age, 53.8+/-12.7 years) were included into the study. Etiology of cirrhosis was viral hepatitis, alcoholic liver disease, cryptogenic and miscellaneous reasons in 57, 49, 14, and 8 patients, respectively. Child's stages A, B, and C of liver cirrhosis were established in 28 (21.9%), 70 (54.9%), and 30 (23.4%) patients, respectively. The mean hepatic venous pressure gradient significantly differed among patients with different Child's classes: 13.8+/-5.3 mm Hg, 17.3+/-4.6 mm Hg, and 17.7+/-5.05 mm Hg in Child's A, B, and C classes, respectively (P=0.003). The mean hepatic venous pressure gradient in patients with grade I, II, and III varices was 14.8+/-4.5, 16.1+/-4.3, and 19.3+/-4.7 mm Hg, respectively (P=0.0001). Since nonbleeders had both small and large esophageal varices, patients with large varices were analyzed separately. The mean hepatic venous pressure gradient in patients with large (grade II and III) varices was significantly higher than that in patients with small (grade I) varices (17.8+/-4.8 mm Hg vs 14.6+/-4.8 mm Hg, P=0.007). Thirty-four (26.6%) patients had a history of previous variceal bleeding; all of them had large (20.6% - grade II, and 79.4% - grade III) varices. In patients with large varices, the mean hepatic venous pressure gradient was significantly higher in bleeders than in nonbleeders (18.7+/-4.7 mm Hg vs 15.9+/-4.7 mm Hg, P=0.006). CONCLUSIONS: Hepatic venous pressure gradient correlates with severity of liver disease, size of varices, and bleeding status. Among cirrhotics with large esophageal varices, bleeders have a significantly higher hepatic venous pressure gradient than nonbleeders. Hepatic venous pressure gradient measurement is useful in clinical practice selecting cirrhotic patients at the highest risk of variceal bleeding and guiding to specific therapy.

High frequency of the c.3207C>A (p.H1069Q) mutation in ATP7B gene of Lithuanian patients with hepatic presentation of Wilson's disease.

AIM: To investigate the prevalence of the ATP7B gene mutation in patients with hepatic presentation of Wilson's disease (WD) in Lithuania. METHODS: Eleven unrelated Lithuanian families, including 13 WD patients were tested. Clinically WD diagnosis was established in accordance to the Leipzig scoring system. Genomic DNA was extracted from whole venous blood using a salt precipitation method. Firstly, the semi-nested polymerase chain reaction (PCR) technique was used to detect the c.3207C>A (p.H1069Q) mutation. Patients not homozygous for the c.3207C>A (p.H1069Q) mutation were further analyzed. The 21 exons of the WD gene were amplified in a thermal cycler (Biometra T3 Thermocycler, Gottingen, Germany). Direct sequencing of the amplified PCR products was performed by cycle sequencing using fluorescent dye terminators in an automatic sequencer (Applied Biosystems, Darmstadt, Germany). RESULTS: Total of 13 WD patients (mean age 26.4 years; range 17-40; male/female 3/10) presented with hepatic disorders and 16 their first degree relatives (including 12 siblings) were studied. Some of WD patients, in addition to hepatic symptoms, have had extrahepatic disorders (hemolytic anemia 3; Fanconi syndrome 1; neurophsychiatric and behavioural disorder 2). Liver biopsy specimens were available in all of 13 WD patients (8 had cirrhosis; 1-chronic hepatitis; 3-acute liver failure, 1-liver steatosis). Twelve of 13 (92.3%) WD patients had the c.3207C>A (p.H1069Q) mutation, 6 of them in both chromosomes, 6 were presented as compound heterozygotes with additional c.3472-82delGGTTTAACCAT, c.3402delC, c.3121C>T (p.R1041W) or unknown mutations. For one patient with liver cirrhosis and psychiatric disorder (Leipzig score 6), no mutations were found. Out of 16 first degree WD relatives, 11 (68.7%) were heterozygous for the c.3207C>A (p.H1069Q) mutation. Two patients with fulminant WD died from acute liver failure and 11 are in full remission under penicillamine or zinc acetate treatment. Three women with WD successfully delivered healthy babies. CONCLUSION: The c.3207C>A (p.H1069Q) missense mutation is the most characteristic mutation for Lithuanian patients with WD. Even 92.3% of WD patients with hepatic presentation of the disease are homozygous or compound heterozygotes for the p.H1069Q mutation.

Prevalence of hepatitis B virus chronic carriers and risk factors for hepatitis B virus infection among Lithuanian army soldiers.

The aim of this study was to evaluate the prevalence of hepatitis B serological markers (hepatitis B virus (HBV) superficial antigen (HBsAg)) and risk factors for HBV infection among Lithuanian Army soldiers. The study was carried out in Lithuanian military subunits in 2003. Serum samples were draw from 1,830 soldiers (average age, 21.6 (0.707) years) and tested for hepatitis B infection markers (HBsAg). Questionnaires were used to obtain information about risk factors associated with HBV infection. A total of 1.97% of soldiers was seropositive for HBsAg. The prevalence rate of HBV infection was related to military subunit (p > 0.05). Most of the HBsAg-positive soldiers (53.8%) served 4 to 6 months. Among soldiers who were offered to use drugs, the prevalence of HBsAg was 4.3%; in the remaining group, the prevalence was 1.9%. No association was found between other risk factors for HBV infection and the prevalence rate of the hepatitis B marker. Study data proved the need for health promotion, prophylactic vaccination, and monitoring programs at the Lithuanian Armed Forces.

Improvement of liver histopathology in patients with hepatitis C after interferon and ribavirin combination therapy.

UNLABELLED: Aim of the study was to evaluate the effect of interferon and ribavirin combination therapy on liver histopathological outcomes. MATERIAL AND METHODS: Pre-treatment and post 24-week treatment liver biopsy specimens were available in 68 naive patients, 37 nonresponders and 18 relapsers after interferon monotherapy. For all patients paired liver biopsies (6-month interval) were assessed for necroinflammation (according to the method by K. Ishak), fibrosis (according to METAVIR score) and steatosis at the end of 24-week treatment. RESULTS: Virological end-of-treatment response was: 36.8% in naive patients, 24.3% in nonresponders and 22.2% in relapsers. Out of 38 patients, who achieved virological end-of-treatment response, sustained virological response was in 65.8%. There was obvious drop of histological activity features scores in all treated patients at the end of 24-week treatment period. According to the baseline findings, only confluent necrosis was found to be significantly lower in patients, who achieved virological end-of- treatment response (p<0.05). The fibrosis and steatosis has not been influenced at least by 24-week treatment success. But in patients, who achieved sustained virological response, fibrosis was lower at baseline and after 24-week of interferon and ribavirin therapy (p<0.001). In conclusion, assessment of liver histopathology has real value in the evaluation of therapeutic response. Grade of pre-treatment confluent necrosis could predict virological end-of-treatment response. Lower stage of fibrosis both at baseline and after 24-week treatment period seems to be a predictor of sustained virological response.

Interferon alpha-2b in combination with ribavirin for the treatment of chronic hepatitis C: assessment of virological, biochemical and histological treatment response.

UNLABELLED: Aim of the study is to establish the efficacy of treatment of chronic hepatitis C with interferon alpha-2b and ribavirin in treatment-naive patients, in non-responders to interferon monotherapy, in relapsers after interferon monotherapy, and to evaluate the possible predictors of treatment response. MATERIAL AND METHODS: One hundred thirty one patients with chronic hepatitis C were included and assigned to 24-week course of interferon alpha-2b with ribavirin. At the end of 24-week combination therapy biochemical, histological, virological and complete end-of-treatment responses were evaluated. Patients with virological end-of-treatment response were treated with interferon alpha-2b and ribavirin for further 24 weeks and followed-up for the next 6 months after stopping therapy. RESULTS: End-of-treatment response was assessed in 119 patients: 57.1% were treatment-naive, 28.6% were non-responders, and 14.3%--relapsers. Overall virological end-of-treatment response rate was 35.3%, biochemical--64.7%, histological--80.9% and complete--33.3%. Treatment-naive patients have better end-of-treatment response than non-naive patients in all dimensions, but statistical significance is reached only evaluating histological end-of-treatment response. Sustained virological response of overall 119 patients was in 23.6%. There was no statistically significant difference in sustained virological response rate among all patient groups. None of pre-treatment demographic, clinical, biochemical and morphological parameters was found as possible response predicting factors. Hepatitis C virus genotype was assessed in 40 patients. Hepatitis C virus genotype 1 was found in 34 (85%), genotype 2 in 3 (7.5%), and genotype 3 in 3 (7.5%) patients. In conclusion, in our studied population with high prevalence of hepatitis C virus genotype 1 (85%), there is low sustained virological response rate (23.6%) to interferon alpha-2b in combination with ribavirin therapy.

Results of treatment of primary liver cancer at Kaunas University of Medicine Hospital.

Though the results of treatment of primary liver cancer depend on many circumstances, the opportunity to perform a curative liver resection remains the main point in prognosis on survival. The aim of the study was to examine our first experience in the treatment of liver cancer. From 1996 to 2001 we observed 54 patients with liver cancer: 46 hepatocellular and 6 cholangiocellular carcinomas, 1 malignant carcinoid, and 1 carcinosarcoma. In presence of liver cirrhosis (21 patients, 38.8%) hepatic function was evaluated using Child Pugh classification. Lesions were multiple in 28 cases and single in 26 cases. Ten patients (18.5%) were radically resected, 12 patients (22.2%) were managed by laparotomy and biopsy, 2 by percutaneous ethanol injections, 1 by trans-ileocolic portal vein embolization + hepatic artery embolization. There were 7 deaths (28%) and 18 complications (72%) after the surgical treatment. The survival results of patients who underwent resection were better (median 240 days) compared with palliative treatment group (median 113.3 days); by Log-Rank test p=0.208. CONCLUSION. The use of liver resections in patients affected by single or monolateral liver cancer is effective and potentially radical treatment. Mortality and morbidity rate is high. Alternative therapies can be conveniently considered in case of multicentric Child B-C patients.