Low recurrence rate of hepatocellular carcinoma following ledipasvir and sofosbuvir treatment in a real-world chronic hepatitis C patients cohort.

The aims of the present study were to evaluate the efficacy and tolerability of ledipasvir/sofosbuvir (LDV/SOF) with or without ribavirin in the treatment of chronic hepatitis C (CHC) in patients with advanced liver disease and to analyse whether the use of LDV/SOF treatment is associated with a new occurrence of hepatocellular carcinoma (HCC) during and after LDV/SOF treatment. The Turkish Early Access Program provided LDV/SOF treatment to a total of 200 eligible CHC patients with advanced liver disease. The median follow-up period was 22 months. All patients were Caucasian, 84% were infected with genotype 1b, and 24% had a liver transplantation before treatment. The sustained virological response (SVR12) was 86.0% with ITT analysis. SVR12 was similar among patients with Child-Pugh classes A, B and C disease and transplant recipients. From baseline to SVR12, serum ALT level and MELD score were significantly improved (P < 0.001). LDV/SOF treatment was generally well tolerated. Only one patient developed a new diagnosed HCC. Seventeen of the 35 patients, who had a history of previous HCC, developed HCC recurrence during the LDV/SOF treatment or by a median follow-up of 6 months after treatment. HCC recurrence was less commonly observed in patients who received curative treatment for HCC compared with those patients who received noncurative treatment (P = 0.007). In conclusion, LDV/SOF with or without ribavirin is an effective and tolerable treatment in CHC patients with advanced liver disease. Eradication is associated with improvements in liver function and a reduced risk of developing a new occurrence of HCC.

Interferon alpha induced intrahepatic pSTAT1 inversely correlate with serum HCV RNA levels in chronic HCV infection.

BACKGROUND: The Jak-STAT signaling of hepatitis C virus (HCV) infected hepatocyte is critical for the antiviral action of endogenously produced interferon (IFN) as well as exogenously administered interferon alpha (IFN-α). The activation of cellular Jak-STAT signaling by IFN-α results in the phosphorylation and nuclear translocation of pSTAT1 and pSTAT2 proteins to induce antiviral gene transcription. Clinical studies show that chronic HCV patients with high viral load show poor response to interferon alpha and ribavirin combination therapy. AIM: We seek to determine whether the IFN-α induced activation of pSTAT1 and pSTAT2 in hepatocytes isolated from liver biopsy of patients chronically infected with hepatitis C virus could be related to the viral load. METHOD: Hepatocytes were isolated from liver biopsies of 18 chronic HCV patients using the collagen digestion method. Induction of pSTAT1 protein in the isolated hepatocyte was measured after IFN-α treatment. The fold change in the levels of pStat1 in the cell lysates due to IFN-treatment was measured by Western blot analysis followed by densitometry analysis. RESULTS: Results of our study indicate that IFN-α induced pSTAT1 levels vary in chronically infected hepatocytes from chronic HCV patients. Semi-quantitative analysis of the pSTAT1 bands revealed a median induction of 7.4-fold in non-infected primary hepatocytes and 2.3-fold in chronic hepatitis C patients (p < 0.001). Total STAT1 levels were not significantly different between treated and untreated primary hepatocytes. We also found a significantly inverse correlation between the intrahepatic pSTAT1 inductions with the serum HCV RNA levels. CONCLUSION: We have developed an antibody based Western blot detection method to measure intrahepatic pStat1 and pStat2 levels to assess the cellular response to exogenous IFN-alpha. Our results indicate that pStat1 activation is a good indicator to assess the level of HCV replication in chronic HCV patients.

Does Genetic Variation in PNPLA3, TM6SF2 and HSD17B13 have a Role in the Development or Prognosis of Hepatocellular Carcinoma in Turkish Patients with Hepatitis B?

BACKGROUND AND AIMS: Progression to hepatocellular carcinoma (HCC) is restricted by viral suppression in chronic hepatitis B (CHB); however, some patients still progress despite antiviral therapy. Presence of single nucleotide polymorphisms (SNPs) such as PNPLA3 rs738409 and TM6SF2 rs58542926 are associated with the development and progression of steatotic liver disease to HCC, whereas a splice variant in HSD17B13 rs72613567:TA has been shown to be protective. We investigated the role of these SNPs in the development or prognosis of HCC in pure CHB etiology, in the absence of hepatic steatosis, remains unknown. MATERIALS: We analysed PNPLA3 rs738409, TM6SF2 rs58542926, and HSD17B13 rs72613567 SNPs in a prospectively recruited cohort (n=323) consisting of healthy controls, CHB and CHB-HCC patients without hepatic steatosis. SNPs were determined by PCR analysis and associations for the alleles and genotypes were investigated using adjusted-logistic regression analyses. The overall survival (OS) data were collected from CHB-HCC patients for survival analysis. RESULTS: The genotype and allelic distribution of PNPLA3 rs738409, TM6SF2 rs58542926, and HSD17B13 rs72613567 were similar between healthy controls, CHB, and CHB-HCC groups. No genotype, allele or haplotype analysis was found to be associated with increased risk for CHB-HCC. Survival analysis revealed no genotype or allele to be associated with OS in patients with CHB-HCC. CONCLUSIONS: We could not demonstrate any association of PNPLA3 rs738409, TM6SF2 rs58542926, and HSD17B13 rs72613567 with the development or prognosis of CHB-HCC, supporting the initial hypothesis that they should be considered specific hotspots for liver diseases characterized with hepatic steatosis.

Relationship between neutrophil gelatinase-associated lipocalin (NGAL) levels and inflammatory bowel disease type and activity.

BACKGROUND AND AIM: Neutrophil gelatinase associated lipocalin (NGAL) is a recently identified molecule, which is bacteriostatic, has tissue destructive effects and is pro-inflammatory with chemoattractant molecule binding properties. Our aim was to investigate the relationship between serum NGAL levels and the type and level of disease activity of IBD. METHODS: A total of 92 patients [43 with Crohn's disease (CD) and 49 with ulcerative colitis (UC)], and 30 age- and sex-matched healthy controls (HC) were included in this study. Serum NGAL levels were measured using ELISA. RESULTS: Serum NGAL levels were elevated in the IBD group [median 171, range (57-312) ng/mL] compared to the HC group [107 (45-234) ng/mL] (p<0.0001) and were elevated in UC patients [188 (74-312) ng/mL] compared to CD patients [168 (57-279) ng/mL] (p=0.006). When NGAL levels were further analysed based on localization of the CD and UC, the levels in ulcerative pancolitis [233 (144-312) ng/mL] were significantly higher (p=0.004) than the left-sided colitis [156 (103-309) ng/mL]. Similarly, NGAL levels were significantly higher in colonic CD [207 (125-249) ng/mL] than ileal CD [114 (78-210) ng/mL], and also in ileocolonic CD [198 (57-279) ng/mL] than ileal CD (p=0.033). When CD and UC groups were further categorized as active and inactive according to clinical and endoscopic activity indices, serum NGAL concentrations did not differ between inquiescent versus active stages. When a cut-off level of 129 ng/mL was used to distinguish IBD from HC, a sensitivity of 76.1% and a specificity of 60.9% was reached. CONCLUSIONS: The serum NGAL levels in the IBD group was significantly higher than the HC group. Serum NGAL levels were higher in more extensive colonic involvement.

Evaluation of the effectiveness of drug-eluting transarterial chemoebolization in hepatocellular carcinoma.

Background and Aim: Transarterial Chemoembolization (TACE) therapy is currently considered as first option therapy in the intermediate stage HCC. The purpose of our study is to assess the efficacy and prognostic factors related to the DEB- TACE therapy. Materials and Methods: The data from 133 patients with unresecetable HCC who were treated with DEB-TACE and followed between January 2011-March 2018 were retrospectively evaluated. To assess the efficacy of therapy, control imagings were performed at 30th and 90th days after the procedure. Response rates, survival outcomes, and prognostic factors were investigated. Results: According to the Barcelona staging system, 16 patients (13%) were in the early stage, 58 patients (48%) were in the intermediate stage and 48 patients (39%) were in the advanced stage. There were complete response (CR) in 20 patients (17%), partial response (PR) in 36 patients (32%), stable disease (SD) in 24 patients (21%) and progressed disease (PD) in 35 (30%) patients. Median follow-up time was 14 months (range 1-77 months). Median PFS and OS were 4 months and 11 months, respectively. In multivariate analysis, posttreatment AFP ≥400 ng/ml was found to be an independent prognostic factor on both PFS and OS. Child-Pugh classification and tumor size >7 cm were independent prognostic factors on OS. Conclusion: DEB-TACE is effective and a tolerable treatment method for unresectable HCC patients.

Free fatty acids induce ER stress and block antiviral activity of interferon alpha against hepatitis C virus in cell culture.

BACKGROUND: Hepatic steatosis is recognized as a major risk factor for liver disease progression and impaired response to interferon based therapy in chronic hepatitis C (CHC) patients. The mechanism of response to interferon-alpha (IFN-α) therapy under the condition of hepatic steatosis is unexplored. We investigated the effect of hepatocellular steatosis on hepatitis C virus (HCV) replication and IFN-α antiviral response in a cell culture model. METHODS: Sub-genomic replicon (S3-GFP) and HCV infected Huh-7.5 cells were cultured with a mixture of saturated (palmitate) and unsaturated (oleate) long-chain free fatty acids (FFA). Intracytoplasmic fat accumulation in these cells was visualized by Nile red staining and electron microscopy then quantified by microfluorometry. The effect of FFA treatment on HCV replication and IFN-α antiviral response was measured by flow cytometric analysis, Renilla luciferase activity, and real-time RT-PCR. RESULTS: FFA treatment induced dose dependent hepatocellular steatosis and lipid droplet accumulation in the HCV replicon cells was confirmed by Nile red staining, microfluorometry, and by electron microscopy. Intracellular fat accumulation supports replication more in the persistently HCV infected culture than in the sub-genomic replicon (S3-GFP) cell line. FFA treatment also partially blocked IFN-α response and viral clearance by reducing the phosphorylation of Stat1 and Stat2 dependent IFN-ß promoter activation. We show that FFA treatment induces endoplasmic reticulum (ER) stress response and down regulates the IFNAR1 chain of the type I IFN receptor leading to defective Jak-Stat signaling and impaired antiviral response. CONCLUSION: These results suggest that intracellular fat accumulation in HCV cell culture induces ER stress, defective Jak-Stat signaling, and attenuates the antiviral response, thus providing an explanation to the clinical observation regarding how hepatocellular steatosis influences IFN-α response in CHC.

PDCD1 polymorphisms are not associated with Takayasu's arteritis in Turkey.

OBJECTIVES: Takayasu's arteritis (TA) is a chronic arterial inflammation of unknown etiology involving mainly the aorta and its major branches. Based on the associations of programmed death-1 (PD-1) protein encoding gene (PDCD1) with connective tissue diseases and vasculitides, PDCD1 polymorphisms are studied for susceptibility to TA in this study. METHODS: The study group is made up of TA patients (n=229) fulfilling the 1990 ACR classification criteria and compared to 193 healthy controls (HC). PD-1.3, PD-1.5 and PD-1.6 single nucleotide polymorphisms of PDCD1 gene are genotyped by polymerase chain reaction and restriction analysis (PCR-RFLP). RESULTS: The distribution of PD-1.5 polymorphism in TA patients and HC revealed a similar presence of TT genotype in patients and controls (13.3% vs. 11.4%). PD-1.3 and PD-1.6 were less polymorphic and did not differ between the groups. Rare AA genotype of PD-1.3 (1.4% vs. 1.0%) and AG genotype of PD-1.6 was again similarly (22.4% vs. 19.2%) present in TA and HC. CONCLUSIONS: PD-1.3, 1.5 and 1.6 polymorphisms of PDCD1 gene, which were shown to be associated with various autoimmune disorders and vasculitides, are not associated with a susceptibility to TA in Turkish population.

Proposal and Validation of a Novel Scoring System for Hepatocellular Carcinomas Beyond Curability Borders.

Optimal scoring system for clinical prognostic factors in patients with unresectable hepatocellular carcinoma (HCC) is currently uncertain. We aimed to develop and externally validate an easy to use tool, particularly for this population, and named it the "unresectable hepatocellular carcinoma prognostic index" (UHPI). We evaluated the data of patients with treatment-naive unresectable HCC who were diagnosed in the training center from 2010 to 2019 (n = 209). A simple prognostic model was developed by assigning points for each covariate in proportion to the beta coefficients in the Cox multivariable model. Predictive performance and distinction ability of the UHPI were further evaluated in an independent European validation cohort (n = 147) and compared with 11 other available models. A simple scoring system was derived, assigning 0.5/1/2 scores for six independent covariates including, the Child-Pugh score, Eastern Cooperative Oncology Group performance status, maximum tumor size, vascular invasion or extrahepatic metastasis, lymph node involvement, and alpha-fetoprotein. The UHPI score, ranging from 0 to 6, showed superior performance in prognosis prediction and outperformed 11 other staging or prognostic models, giving the highest homogeneity (c-index, 6-month and 1-year area under the receiver operator characteristic curves), lowest Akaike information criterion, and -2 log-likelihood ratio values. The UHPI score allocated well the risk of patients with unresectable HCC for mortality within the first year, using two cut-off values (low-risk, <0.5; intermediate-risk, 0.5-2; high-risk, >2). Conclusion: The UHPI score can predict prognosis better than other systems in subjects with unresectable HCC and can be used in clinical practice or trials to estimate the 6-month and 1-year survival probabilities for this group.

Mechanism of HCV's resistance to IFN-α in cell culture involves expression of functional IFN-α receptor 1.

The mechanisms underlying the Hepatitis C virus (HCV) resistance to interferon alpha (IFN-α) are not fully understood. We used IFN-α resistant HCV replicon cell lines and an infectious HCV cell culture system to elucidate the mechanisms of IFN-α resistance in cell culture. The IFN-α resistance mechanism of the replicon cells were addressed by a complementation study that utilized the full-length plasmid clones of IFN-α receptor 1 (IFNAR1), IFN-α receptor 2 (IFNAR2), Jak1, Tyk2, Stat1, Stat2 and the ISRE-luciferase reporter plasmid. We demonstrated that the expression of the full-length IFNAR1 clone alone restored the defective Jak-Stat signaling as well as Stat1, Stat2 and Stat3 phosphorylation, nuclear translocation and antiviral response against HCV in all IFN-α resistant cell lines (R-15, R-17 and R-24) used in this study. Moreover RT-PCR, Southern blotting and DNA sequence analysis revealed that the cells from both R-15 and R-24 series of IFN-α resistant cells have 58 amino acid deletions in the extracellular sub domain 1 (SD1) of IFNAR1. In addition, cells from the R-17 series have 50 amino acids deletion in the sub domain 4 (SD4) of IFNAR1 protein leading to impaired activation of Tyk2 kinase. Using an infectious HCV cell culture model we show here that viral replication in the infected Huh-7 cells is relatively resistant to exogenous IFN-α. HCV infection itself induces defective Jak-Stat signaling and impairs Stat1 and Stat2 phosphorylation by down regulation of the cell surface expression of IFNAR1 through the endoplasmic reticulum (ER) stress mechanisms. The results of this study suggest that expression of cell surface IFNAR1 is critical for the response of HCV to exogenous IFN-α.

Vitamin E has a dual effect of anti-inflammatory and antioxidant activities in acetic acid-induced ulcerative colitis in rats.

BACKGROUND: Increased free radical production, decreased antioxidant capacity and excessive inflammation are well-known features in the pathogenesis of inflammatory bowel disease. Vitamin E is a powerful antioxidant and a scavenger of hydroxyl radicals, and it has been shown to have anti-inflammatory activities in tissues. We investigated the effects of vitamin E on inflammatory activities using an acetic acid (AA)-induced ulcerative colitis model in rats. METHODS: Wistar rats were divided into 4 groups. Acetic acid was given to 2 groups of animals to induce colitis while the other 2 groups received saline intrarectally. One AA-induced colitis group and 1 control group received vitamin E (30 U/kg/d) intraperitoneally and the pair groups received saline. After 4 days, we evaluated colonic changes biochemically by measuring proinflammatory cytokine levels in tissue homogenates and by histopathologic examination. RESULTS: Acetic acid caused colonic mucosal injury, whereas vitamin E administration suppressed these changes in the AA-induced colitis group (p < 0.001). Administration of AA resulted in increased levels of tumour necrosis factor-α, interleukin-1ß, interleukin-6, myeloperoxidase and malondialdehyde, and decreased levels of glutathione and superoxide dismutase; vitamin E reversed these effects (all p < 0.001). CONCLUSION: Our study proposes that vitamin E is an effective anti-inflammatory and antioxidant and may be a promising therapeutic option for ulcerative colitis.

Programmed Cell Death 1 and Hepatocellular Carcinoma: An Epochal Story.

In recent years, immune-based therapies have emerged as novel pillars for hepatocellular carcinoma (HCC). The rationale of immune-checkpoint inhibitors (ICIs) trial in HCC originated from the fact that the tumor cells and the infiltrating stromal and immune cells promote an immunosuppressive tumor microenvironment, including the up-regulation of immune checkpoint molecules on their surface. Antibody-based blockage targeting inhibitory checkpoint molecules on cytotoxic T cells, including programmed cell death-1 (PD-1) or its counterpart on antigen-presenting cells has shown strong anti-tumor activity in a subset of HCC patients. Single nucleotide polymorphisms (SNP) of PD-1 gene may affect the PD-1 expression or function, which eventually can cause dysfunctionality of immune balance. Based on the inhibitory role of PD-1 in anti-tumor responses, it has been investigated in several studies as a candidate to test for genetic susceptibility of individuals to HCC. The present paper highlights the knowledge on cross-talks for liver immunology and HCC course, recent studies investigating the role of functional SNPs of PD-1 gene in Turkish HCC population, and the data on already investigated PD-1 inhibitor molecules in clinical trials.

Surveillance Strategies for Hepatocellular Carcinoma: Recent Advances and the Shifting Paradigm.

Currently, international liver societies recommend screening at-risk individuals for HCC (patients with cirrhosis regardless of etiology, and/or chronic hepatitis B virus, and/or advanced liver fibrosis) with biannual abdominal ultrasound (USG) with or without alpha-fetoprotein (AFP). The global acceptance of USG in surveillance relies on the absence of risks, non-invasiveness, and lower costs. However, the suboptimal performance of USG ± AFP in reaching direct and indirect goals of HCC surveillance highlights the need for alternative surveillance strategies. Several studies targeted contrast-enhanced magnetic resonance imaging techniques, but the main barriers for their entrance to surveillance programs have been concerns about cost-effectivity and long scan times. Overall, the HCC risk stratification appears at hand by several validated multiple score systems, but their optimal performance is obtained only in populations who show highly homogenous clinical, pathological, epidemiologic, etiologic, and therapeutic characteristics, and this limitation poses a major drawback to their sustainable use in clinical practice. We need globally validated and molecular integrated risk stratification tools to shape the future tailored HCC surveillance decision algorithms. A dynamic process for HCC surveillance algorithms awaits us owing to the expected further prospective studies focusing on risk-stratified screening strategy.

Hepatocellular Carcinoma in Non-cirrhotic Liver Arises with a More Advanced Tumoral Appearance: A Single-Center Cohort Study.

BACKGROUND: A small proportion of all hepatocellular carcinomas (HCCs) arise in a non-cirrhotic liver (NCL). However, our knowledge about the HCCs developing in a NCL is scarce. This study was undertaken to investigate the characteristics and survival course of this patient group. METHODS: We retrospectively analyzed the database of patients with HCC at a tertiary center during a 10-year period (2009-2019). All demographic, clinical, laboratory, and tumoral features with survival outcomes were compared between the HCC-CL and HCC-NCL groups. RESULTS: Out of 384 HCC cases, 11.2% (n = 43) had no cirrhosis. The dominant etiology in the HCC-NCL group was hepatitis B virus (n = 26, 60.5%), followed by non-alcoholic fatty liver disease (n = 10, 23.2%), and hepatitis C virus (n = 7, 16.3%). The maximum tumor diameter was approximately 2 times larger in the HCC-NCL group (HCC-NCL: 90 mm vs. HCC-CL: 46.5 mm, P < .001). The proportion of patients with vascular (HCC-NCL: 27.9% vs. HCC-CL: 8.6%, P < .001) and extrahepatic invasion (HCC-NCL: 14% vs. HCC-CL: 3%, P = .001) were prominently higher in the HCC-NCL group. Patients with HCC-NCL were less often detected in early-curable stages (BCLC 0-A) than those in the HCC-CL group (HCC-NCL: 16.3% vs. HCC-CL: 34.9%, P = .004). The overall survival was not different between the 2 groups (HCC-NCL: 19.4 ± 9.8 months vs. HCC-CL: 17.5 ± 2.3 months, P = .581). CONCLUSION: HCC in NCL is diagnosed at more advanced tumoral stages with larger tumor size and more often with vascular and extrahepatic spread. Despite the preserved liver functions, the overall survival is not prolonged in HCCs without cirrhosis, due to the late recognition.

High incidence of hepatitis B core antibody positivity in metabolic-associated fatty liver disease-related cirrhosis.

Background and Aim: The coexistence of metabolic-associated fatty liver disease (MAFLD) in the course of chronic hepatitis B virus infection increases liver-related morbidity. A positive correlation was found between positive hepatitis B core antibody (anti-HBc) and the risk of cirrhosis and hepatocellular carcinoma (HCC) in MAFLD. The relationship between anti-HBc positivity and MAFLD progression to fibrosis, cirrhosis, and liver-related outcomes was determined. Materials and Methods: This is a retrospective study including 242 patients with biopsy-proven MAFLD, 130 patients with clinically diagnosed MAFLD-related cirrhosis, and 62 patients with MAFLD-related or cryptogenic HCC. Anti-HBc antibody results were compared with clinical outcomes. Results: Anti-HBc positivity was associated with fibrosis severity (p=0.005). Anti-HBc was positive in 19 (20.2%), 33 (25.8%), 53 (35.3%), and 27 (43.5%) patients with F0-F1 fibrosis, F2-F3 fibrosis, cirrhosis (F4), and HCC, respectively. Median steatosis score was grade 3 in anti-HBc positive patients and grade 2 in negative patients (p=0.07). Anti-HBc positivity was not associated with significant fibrosis (≥F2), cirrhosis, and any liver related complications including HCC. Conclusion: Higher anti-HBc positivity was found in MAFLD patients with advanced fibrosis and cirrhosis compared to patients with early stage fibrosis. No relation was found between anti-HBc positivity and development of cirrhosis, HCC or other liver related complications.

Recommendations for Reopening Endoscopy Units in Turkey: A Set-up Plan from a Tertiary Center in Istanbul.

In the midst of Coronavirus-19 (COVID-19) pandemic, endoscopic procedures have been separated for only urgent and semi-urgent cases for the last few months to prevent transmission in endoscopy units. This approach will perhaps resolve the burden of elective procedures in the months ahead of us. As we observe a downtrend in new cases of COVID-19 in Turkey, a strategy for reopening endoscopy units is required. We are stepping into a time period where we should not only re-provide the essential services to our patients but also maintain the safety of healthcare workers and preserve the valuable personal protective equipment as well. Herein, we aim to share the available knowledge in performing endoscopy during the pandemic and the set-up plan of a tertiary center in Istanbul for reopening the endoscopy unit in the era of the COVID-19 pandemic.

Characteristics of Newly Diagnosed Hepatocellular Carcinoma Patients Across Turkey: Prospective Multicenter Observational 3K Registry Study.

AIMS: To evaluate patient profile for epidemiological and clinicopathological characteristics and potential risk/prognostic factors in newly diagnosed hepatocellular carcinoma (HCC) patients across Turkey. METHODS: A total of 547 patients (mean (SD) age 62.6 (10.3) years, 81.9% were males) were included in this registry study. Data on patient characteristics, etiologies of HCC, laboratory values, and tumor characteristics and stages were recorded at study enrollment. RESULTS: HBV infection (68.2%) was the leading etiology, followed by HCV infection (17.2%), HDV infection (5.5%), alcohol (6.4%), and NAFLD (3.5%), as the major etiologies. Considering that 51.6% of the patients had >5 cm HCC, 44% were Child-Pugh B/C and 57% were BCLC B-D, it appears that a significant group of HCC patients were diagnosed at advanced stages. Of 540 patients, 271 (50.2%) were referred or applied with the diagnosis of HCC. Patients with HCC at presentation had larger tumor size (median (min-max) 6.6 (0-30) vs. 4.8 (0-90) cm, P < .001) and more advanced BCLC stage (Stage C-D in 40.8% vs. 26.4%, respectively, P = .005), compared to patients who were diagnosed during follow-up. CONCLUSIONS: Our findings revealed that HBV infection was the leading etiology and a moderate-to-advanced disease was evident in more than half of patients at the time of diagnosis. HCC patients diagnosed at follow-up had smaller tumor size and earlier BCLC stage.

Intracellular expression of IRF9 Stat fusion protein overcomes the defective Jak-Stat signaling and inhibits HCV RNA replication.

Interferon alpha (IFN-α) binds to a cell surface receptor that activates the Jak-Stat signaling pathway. A critical component of this pathway is the translocation of interferon stimulated gene factor 3 (a complex of three proteins Stat1, Stat2 and IRF9) to the nucleus to activate antiviral genes. A stable sub-genomic replicon cell line resistant to IFN-α was developed in which the nuclear translocation of Stat1 and Stat2 proteins was prevented due to the lack of phosphorylation; whereas the nuclear translocation of IRF9 protein was not affected. In this study, we sought to overcome defective Jak-Stat signaling and to induce an antiviral state in the IFN-α resistant replicon cell line by developing a chimera IRF9 protein fused with the trans activating domain (TAD) of either a Stat1 (IRF9-S1C) or Stat2 (IRF9-S2C) protein. We show here that intracellular expression of fusion proteins using the plasmid constructs of either IRF9-S1C or IRF9-S2C, in the IFN-α resistant cells, resulted in an increase in Interferon Stimulated Response Element (ISRE) luciferase promoter activity and significantly induced HLA-1 surface expression. Moreover, we show that transient transfection of IRF9-S1C or IRF9-S2C plasmid constructs into IFN-α resistant replicon cells containing sub-genomic HCV1b and HCV2a viruses resulted in an inhibition of viral replication and viral protein expression independent of IFN-α treatment. The results of this study indicate that the recombinant fusion proteins of IRF9-S1C, IRF9-S2C alone, or in combination, have potent antiviral properties against the HCV in an IFN-α resistant cell line with a defective Jak-Stat signaling.

Intracytoplasmic stable expression of IgG1 antibody targeting NS3 helicase inhibits replication of highly efficient hepatitis C Virus 2a clone.

BACKGROUND: Hepatitis C virus (HCV) infection is a major public health problem with more than 170 million cases of chronic infections worldwide. There is no protective vaccine currently available for HCV, therefore the development of novel strategy to prevent chronic infection is important. We reported earlier that a recombinant human antibody clone blocks viral NS3 helicase activity and inhibits replication of HCV 1b virus. This study was performed further to explore the mechanism of action of this recombinant antibody and to determine whether or not this antibody inhibits replication and infectivity of a highly efficient JFH1 HCV 2a virus clone. RESULTS: The antiviral effect of intracellular expressed antibody against the HCV 2a virus strain was examined using a full-length green fluorescence protein (GFP) labeled infectious cell culture system. For this purpose, a Huh-7.5 cell line stably expressing the NS3 helicase gene specific IgG1 antibody was prepared. Replication of full-length HCV-GFP chimera RNA and negative-strand RNA was strongly inhibited in Huh-7.5 cells stably expressing NS3 antibody but not in the cells expressing an unrelated control antibody. Huh-7.5 cells stably expressing NS3 helicase antibody effectively suppressed infectious virus production after natural infection and the level of HCV in the cell free supernatant remained undetectable after first passage. In contrast, Huh-7.5 cells stably expressing an control antibody against influenza virus had no effect on virus production and high-levels of infectious HCV were detected in culture supernatants over four rounds of infectivity assay. A recombinant adenovirus based expression system was used to demonstrate that Huh-7.5 replicon cell line expressing the intracellular antibody strongly inhibited the replication of HCV-GFP RNA. CONCLUSION: Recombinant human anti-HCV NS3 antibody clone inhibits replication of HCV 2a virus and infectious virus production. Intracellular expression of this recombinant antibody offers a potential antiviral strategy to inhibit intracellular HCV replication and production.

Effects of vitamin D level on survival in patients with hepatocellular carcinoma.

Background and Aim: Hepatocellular carcinoma (HCC) is a life-threatening condition of the liver, often concurrent with vitamin D deficiency. In this study, we investigated the relationship between HCC patients' vitamin D levels and overall survival. Materials and Methods: We retrospectively enrolled patients that were being followed on their HCC diagnosis. We collected and examined data on patients' 25-OH vitamin D levels one month before diagnosis or at any point thereafter. We took levels below 10 ng/mL to indicate severe deficiency, levels between 10 ng/mL and 20 ng/mL to indicate moderate deficiency, and levels between 20 ng/mL and 30 ng/mL to indicate mild deficiency. We then analyzed the effects of vitamin D levels on patients' survival for each of these brackets. Results: We included 85 patients in our survival analyses. We found 9 ng/mL to be the significant cutoff vitamin D level for survival. Vitamin D levels were lower in cases of advanced disease. Univariate analysis showed that advanced Barcelona Clinic Liver Cancer (BCLC) grades, vitamin D levels below 9 ng/mL, and alpha-fetoprotein (AFP) levels above 400 ng/dL had a negative significant effect on survival. Multivariate analysis showed that only advanced BCLC grades and AFP levels above 400 ng/dL had a negative significant effect. Conclusion: In our study's cohort, HCC grades and AFP levels had a substantial negative impact on patients' overall survival. We found no connection, however, between vitamin D levels and overall survival.

Effect of carbon dioxide versus room air insufflation on post-colonoscopic pain: A prospective, randomized, controlled study.

BACKGROUND/AIMS: Room air (RA) and carbon dioxide (CO2) are widely used to insufflate the colon to examine the mucosa in colonoscopy. Pain, discomfort, and bloating can be seen during and after colonoscopy secondary to bowel distention. This study aimed to investigate the effect of CO2 on post-procedure pain sensation (PPPS) in comparison with RA. MATERIALS AND METHODS: Patients were randomly assigned to the RA and CO2 insufflation groups in a 1:1 ratio. The visual analog scale (VAS) was used to measure the pain before and after the colonoscopy. VAS score of 0 was accepted as the absence of pain and above 0 was accepted as the presence of pain. The primary outcome was to investigate the effect of CO2 insufflation on PPPS. Secondary outcomes were to investigate the other contributing factors affecting PPPS and the effect of CO2 on PPPS in patients with inflammatory bowel disease (IBD). RESULTS: A total of 204 patients were enrolled in the study. No significant difference in PPPS was seen between the 2 groups at any point in time after the colonoscopy. Furthermore, there was no significant difference in pain sensation between the CO2 and RA groups in patients with IBD. When we investigated the other contributing factors to pain sensation, body-mass index (BMI) was found to be significant at 30 minutes and BMI and colonoscopy time were found to be significant at 6 hours afterwards. CONCLUSION: We found no favorable effect of CO2 insufflation on PPPS in colonoscopy, including in patients with IBD.