For details about semiquantitative and real-time polymerase chain reaction (PCR) reactions, see the Supporting Materials and Methods. Cells at 70% confluence were transiently transfected with 50 nM small interfering RNA (siRNA) for 8 hours

using TransIT-siQuest following the manufacturer’s instructions Kinase Inhibitor Library (Mirus, Madison, WI). For stable transfection of short hairpin RNA (shRNA), cells at 50%-60% confluence were transfected with MATra-A reagent (IBA, Germany) according to the manufacturer’s recommendation (15 minutes on the magnet plate, 2 μg/mL of shRNA plasmid). Four different plasmids of TGFBRI shRNA were transfected separately or combined, as well as a control shRNA. Protocols used were as described.[18] For siRNA sequences and further experimental details, see the Supporting Materials and Methods. Cell motility was examined by two different GPCR Compound Library methods: (1) a wound-healing assay[16] and (2) real-time migration assay through the xCELLigence system (Roche Applied Science). For the wound-healing assay, cells were grown at basal conditions to 95% confluence and monolayers were scratched with a pipette tip (0 hours). Cell migration was recorded by phase contrast microscopy (Olympus IX-70) at 48 hours after wound scratch. For real-time monitoring of cell migration, the xCELLigence system was used; 4 × 104 cells/well were seeded onto the top

chamber of a CIM plate, which features microelectronic sensors integrated on the underside of the microporous membrane of a Boyden-like chamber. CIM plates were placed onto the Real-Time Cell Analyzer (RTCA) station (xCELLigence System, Roche, Mannheim, Germany). Cell migration was continuously monitored by measuring changes in the electrical impedance at the electrode/cell interface, as a population of cells migrated from the top to the bottom chamber. Continuous values are represented as cell index (CI), a dimensionless parameter that reflects a relative change in measured electrical impedance, and quantified as a slope (h−1) of the first 5 hours. Male mice at day 15 of age received intraperitoneal injections of DEN (5 mg/kg)

diluted in saline buffer, control animals were injected with saline buffer intraperitoneally. At 6, 9, and 12 months of age, mice were sacrificed and their livers removed. For histological studies, liver lobes were fixed in 4% paraformaldehyde check details overnight and paraffin-embedded for immunohistochemistry staining. Total RNA was isolated from frozen tissues to analyze gene expression by real-time quantitative PCR. Three to four animals/condition and two different tissue pieces/animal were processed for RNA extraction. All data represent at least three experiments and are expressed as the mean ± SEM. Differences between groups were compared using either Student t test or one-way analysis of variance (ANOVA) associated with Dunnett’s test. Statistical significance was assumed when P < 0.05.

8 In addition to the ability of HCV to trigger the TLR3 pathway,9, 10 the increased number of Th17 cells appears to be associated with the severity of liver inflammation in chronic HCV patients and treatment of infected patients with pegylated BGB324 nmr interferon (IFN)-α and ribavirin reduced the level of Th17-related cytokines.11 As one of the crucial factors for Th17 differentiation, thymic stromal lymphopoietin (TSLP), a member of the common γ-chain cytokine, is capable of activating (conditioning) DCs, thereby stimulating naïve T cells to differentiate into Th2 cells.12 In addition, DCs treated with both TSLP and poly (I:C) activate naïve T cells and

differentiate into Th2 and Th17 cells.8, 13 Thus, TSLP-activated DCs, which are known to be strong inducers of Th2 responses, can simultaneously induce Th17 cells under certain pathological conditions. In this report EPZ 6438 we demonstrate that the infection of hepatic cells in vitro by HCV triggers robust TSLP production and this HCV-induced production of TSLP is regulated in an nuclear factor kappa B (NFκB)-dependent manner. TSLP secreted by HCV-infected cells activates and conditions human

monocyte-derived DCs to enhance the production of Th17 differentiating cytokines, TGF-β, IL-6, and IL-21 by the DCs. Moreover, the addition of TSLP neutralizing antibody to the coculture of monocytes/DCs with HCV-infected hepatocytes blocks the production of these cytokines. Consistent with these data, we find that the hepatocyte-derived TSLP is readily detected in liver biopsies from chronic HCV patients. Our studies suggest a novel role for the hepatocyte-derived TSLP in the generation of CD4+ Th17 effector T cells through its ability to condition DCs to drive CD4+ Th17 differentiation. The potential implications of these findings in the development of HCV-induced chronic progressive liver disease are discussed. DC, dendritic

cell; HCV, hepatitis C virus; TSLP, thymic stromal lymphopoietin. Human see more hepatoma cell lines, Huh 7.5.1, were maintained in Dulbecco’s modified Eagle’s medium (DMEM) with 10% fetal bovine serum (FBS) and penicillin/streptomycin (100 μg/mL). THP-1 cells purchased from the American Tissue Culture Collection (ATCC) were cultured in RPMI 1640 and supplements as recommended by ATCC. Liver biopsies and peripheral blood samples from chronic HCV or control patients were obtained from Dr. Hugo Rosen (University of Colorado). Blood samples were also obtained from the Virginia Blood Services. All information of age, gender, and HCV genotype were previously described.14, 15 For infection of cells with secreted HCV, Huh 7.5.1 permissive cells were seeded at 3 × 106 cells in a T75 plate for 24 hours. Cells were infected with 4 × 104 FFU (multiplicity of infection [MOI] of 0.01) of JFH-1 producing cell supernatant and cultured for 10 days in DMEM-10% FCS media.

Results: 14/980 (1.4%) of patients with IBD at EH had at least one episode of symptomatic CDI (2005–2014). Of 14 IBD patients, 8 (57%) had ulcerative colitis, 6 (43%) had Crohn’s, mean age was 46 y, 9 (65%) were females, with mean Charlson comorbidity index score for the IBD patient cohort was 1.2. 0/14 (0 %) patients had a recent course of broad-spectrum antibiotics. 12 (86%) patients were on immunosuppressants at time

of CDI, 5/14 (35 %) had therapy for IBD escalated during admission. Matching ensured no significant differences in age, sex or Charlson comorbidity index between the IBD patients and non-IBD controls (each p > 0.30). IBD patients with CDI were significantly Roxadustat different from non-IBD controls with CDI in multiple aspects including that IBD patients were more likely to have acquired CDI as an outpatient (Fisher exact test, p = 0.003), had not received recent antibiotics (including broad spectrum antibiotics known to increase risk of CDI) (p < 0.001), been on immunosuppressant therapy (p = 0.002), presented with abdominal pain (p = 0.05) and failed initial treatment with metronidazole (p = 0.003). Also, compared with non-IBD controls, those with IBD on average had more bowel actions/ day at presentation (mean 6 vs 2, t-test, p < 0.05), required

longer duration of antibiotics before clinical improvement (mean 23 vs 10 days, selleck compound p < 0.05) and tended to have lower serum CRP but higher albumin (mean 15 vs 26 (p = 0.09) 37 vs 33 (p = 0.07) respectively. There was no significant difference in length

of inpatient stay caused by CDI between the IBD and non-IBD groups (mean 7.0 vs 5.8, p = 0.30). Conclusion: CDI was relatively uncommon in IBD with a prevalence of approximately 1%, but when it occurs it is more likely outpatient acquired, less responsive to standard antibiotic therapy and duration and associated with concurrent immunosuppression rather than the typical recent use of broad spectrum antibiotics. Consideration should be given to early or even first line use of oral vancomycin in IBD patients see more with CDI given the poor, slower response to metronidazole, but further studies are needed. DK TIAO,1 J JEGANATHAN,1 A CHEN,2 J CHANG,3 CP SELINGER,3 RWL LEONG3 1Faculty of Medicine, The University of Sydney, 2Faculty of Medicine, The University of New South Wales, 3Gastroenterology and Liver Services, Concord Hospital, Sydney, NSW, Australia Background: Inflammatory bowel diseases (IBD) often require chronic maintenance medical therapy. Non-adherence occurs in up to 40% of patients, and is associated with increased relapse rates. The Medication Belief Model of perceived medication necessity versus concerns aims to shift patients’ attitudes towards ‘acceptance’ (high necessity and low concerns).

This was suggested by the known interaction with the KRTK motif of thrombospondin, the major activator of TGF-β in vivo,40 leading us BAY 57-1293 solubility dmso to propose that the KTFR sequence could play a similar

role in ADAMTS1 (Fig. 3). The inhibition of ADAMTS1-mediated activation of TGF-β by KTFR peptides indicates that the mechanism of interaction is similar—if not identical—to that reported for thrombospondin-mediated activation.24 In this case, a “conformational” mechanism is in full agreement with the results of our experiments using proteolysis-deficient ADAMTS1 mutants and protease inhibitors, which show that TGF-β activation by ADAMTS1 is independent of the proteolytic activity of the latter. Although it might occur via similar interactions, activation of TGF-β by ADAMTS1 and thrombospondin are unlikely to overlap in vivo. Thrombospondin is expressed in freshly isolated human HSCs,22 but we show that their activation induces a decrease in thrombospondin expression Navitoclax mw counterbalanced by a dramatic increase in ADAMTS1 expression. The physiological relevance of this activation process is fully supported by our finding that depleting ADAMTS1 in HSCs strongly diminishes the release of TGF-β-dependent transcriptional activity. A conformational model of TGF-β

activation by ADAMTS1 predicts that interfering with KTFR/SLKL interactions should lead to a decrease see more in available active TGF-β. We demonstrate that such a mechanism is, indeed, at play in vivo, using a murine model of induced liver fibrosis. In full agreement with the activation pathway described above, injection of the KTFR peptide in CCl4-treated mice that develop liver fibrosis reduces the levels of biological markers associated with hepatic damage (Fig. 7). This conclusion is further borne out by the demonstration, using highly sensitive SHG analysis,41 that concomitant injection of KTFR dramatically reduces collagen deposition

associated with the early onset of fibrosis (Fig. 8). The full conservation of KTFR and LSKL motifs between humans and mice suggests that this important proof of concept can be extrapolated to humans, identifying ADAMTS1 as a new therapeutic target during chronic liver injury. Many factors have been implicated in TGF-β activation, including thrombospondin, proteases (e.g., plasmin, thrombin, and MMP), and integrins, but also heat, acid, reactive oxygen species, and mechanical force. All these components build up an environmental network, in which the role of each one is obviously part of the sum and depends on dynamic tissue changes, especially as the liver proceeds from a healthy to a fibrotic state.

According to the Los Alamos HCV database,27 this variant Alectinib nmr is uncommon in the HCV population, being present in just one of 352 genotype 1 NS5B sequences in the database. The level of antiviral activity, resistance profile, and subtype 1a/1b activity observed for filibuvir in these studies compares favorably to other NNIs currently in development. Maximum reductions in HCV RNA reported for NNIs of HCV range from 0.6-3.7 log10 IU/mL,28 and the activity observed with filibuvir is well within this range. Many NNIs demonstrate differential antiviral activity against 1a and 1b subtypes. However, filibuvir, as well as other NNIs that target the Thumb 2 site of the enzyme (e.g., VCH-795),23

seem to demonstrate equivalent antiviral activity against 1a and 1b subtypes, which may be a function of the particular binding site. Safety or tolerability concerns associated

with other NNIs under development, such as QT prolongation, gastrointestinal AEs, hepatotoxicity, and rash, were not observed in either of these filibuvir studies. In conclusion, data from the two studies presented here show that filibuvir is a potent inhibitor of HCV replication in vivo and is well tolerated in HCV genotype 1–infected patients, supporting further clinical evaluation. Filibuvir is currently being evaluated in combination with pegIFN and RBV in treatment-naive patients. The authors gratefully acknowledge all the patients who participated in the study, all the investigators, nursing staff, and research support staff involved

in the study, and the research team at Pfizer Global buy BIBW2992 Research and Development. see more The authors acknowledge Charles Craig for critical reading of the manuscript and Marilyn Lewis for help with the NS5B genotypic analysis. The authors also acknowledge the editorial assistance of Sarah Maloney, Caroline Masterman, and Susanne Gilbert of KnowledgePoint360 Group during the development of this publication, which was funded by Pfizer, Inc. “
“Pretreatment up-regulation of hepatic interferon (IFN)-stimulated genes (ISGs) has a stronger association with the treatment-resistant interleukin (IL)28B minor genotype (MI; TG/GG at rs8099917) than with the treatment-sensitive IL28B major genotype (MA; TT at rs8099917). We compared the expression of ISGs in the liver and blood of 146 patients with chronic hepatitis C who received pegylated IFN and ribavirin combination therapy. Gene expression profiles in the liver and blood of 85 patients were analyzed using an Affymetrix GeneChip (Affymetrix, Santa Clara, CA). ISG expression was correlated between the liver and blood of the MA patients, whereas no correlation was observed in the MI patients. This loss of correlation was the result of the impaired infiltration of immune cells into the liver lobules of MI patients, as demonstrated by regional gene expression analysis in liver lobules and portal areas using laser capture microdissection and immunohistochemical staining.

Methods: Patients with Crohn’s disease, with

more than five years of clinical follow-up, managed at the Royal Brisbane and Women’s hospital between 1994 and 2014 had objective clinical and laboratory data collected. An objective selleck products poor outcome was defined as the development of a fistula, a bowel stenosis or a bowel perforation. Cox regression was used to analyse the association between this outcome and serial laboratory values (CRP, platelet count, albumin level, fecal calprotectin, serum ferritin, serum haemoglobin), measured in the complication free period leading up to the development of the outcome. Recognized predictors of poor outcome were added to the model to assess independence of laboratory values. Results: 366 patients were reviewed and 311 had more INK 128 mw than five years of follow-up. 185 had a complete clinical, biochemical and genetic record, yielding 2092 years of patient follow-up. 82 outcome events were observed occurring after a median of 5.54 years, in 167 abdominal surgeries, 485 cross sectional imaging procedures and 708 colonoscopies. 4927 haemoglobin levels, 4928 platelet levels, 4242 albumin levels, 3373 CRP levels, 968 ferritin levels and 733 fecal calprotectin levels were analyzed.

A consistent haemoglobin <105 (male) or 90 (female) (hazard ratio 2.29, p < 0.001), platelet count >360 (HR 2.66, p < 0.001), albumin level <32 (HR 7.047, p < 0.001), CRP > 10 (HR 1.92, p = 0.002), ESR > 18 (HR 1.67, p = 0.02) and ferritin

<150 (HR 6.19, p = 0.013) correlated significantly with a poor outcome on univariate analysis. After multivariate analysis with inclusion of recognized predictor variables, haemoglobin level <105 (male) or 90 (female) (HR 2.16, p = 0.0016), albumin level <32 (HR 3.30, p = 0.01) and platelet count >360 (HR 1.91, p = 0.025) maintained an independent this website association with outcome. ATG16L1 AG or GG genotype (HR 2.79, p = 0.047) , continued smoking (HR 1.76, p = 0.016) and L1 or L3 Montreal location at diagnosis (HR 2.32, p = 0.015) were also independently associated with outcome in the final model. Conclusion: Longitudinally measured haemoglobin level, albumin level and platelet count correlate with subsequent development of an objective poor outcome in patients with Crohn’s disease. Serial monitoring of these values may aid in therapeutic decision making. Continued smoking, L1 or L3 Montreal location at diagnosis and ATG16L1 AG or GG genotype were also associated with poor outcome.

5. All data are given as means ± standard deviation (SD). The SAS 9.1 program (SAS Institute, Cary, NC) was used for all data processing. Demographic data and baseline characteristics were checked for normal distribution with Kolmogorov-Smirnov goodness-of-fit tests. Significant differences were evaluated in contingency tables using either Fisher’s exact tests or χ2-tests. Continuous variables were compared between groups using Student’s t tests or

Mann-Whitney rank sum tests. Two-sided P < 0.05 were considered significant, unless previous studies indicated that lower or higher values were to be expected; then, one-sided tests were performed. Pearson's correlation coefficient was used to evaluate univariate associations between the sitosterol:lathosterol and campesterol: lathosterol selleck compound ratios and different ethnic groups, age, BMI and gender. Analysis of covariance was performed to obtain estimates Alpelisib mouse of sitosterol:lathosterol and campesterol: lathosterol ratios between ethnic groups adjusted for age,

BMI and gender. The area under the receiver operating characteristic curve (AUC) was calculated for each serum surrogate marker of cholesterol synthesis and absorption as well as the specified ratios; the P-values for the AUC indicate significance in comparison to 0.5. For ABCG5/8 genotypes, Hardy-Weinberg equilibrium was checked using exact tests (http://ihg.gsf.de/cgi-bin/hw/hwa1.pl).14 Potential associations between gene variants

and cholelithiasis were tested in contingency tables (genotypes: Armitrage’s trend tests; alleles: χ2-tests). Table 1 presents the baseline demographic data of the study cohorts. In Germans and Chilean Hispanics, cases and matched controls are similar in age, gender, and BMI distribution. Compared with the Chilean Hispanic cohort, the German patients are older, leaner, and include more men. The small Amerindian Chilean cohort (Mapuche) was composed only of women who were relatively younger than the Hispanics and Germans. Of note, serum total cholesterol and low-density lipoprotein (LDL) cholesterol concentrations were learn more lower in Mapuches compared with Hispanics independently of gallstone status. Serum lipids are similar in cases and controls in the German cohort. In contrast, in the Chilean Hispanic cohort total and LDL cholesterol levels are significantly lower in cases compared with controls. In the Chilean Hispanic and Amerindian cohorts, fasting glucose and insulin levels were also determined, yielding comparable results between cases and controls. As shown in Table 1, the IR-HOMA indexes are high and in the range of insulin resistance for both cases and controls, but they do not differ between groups.

5. All data are given as means ± standard deviation (SD). The SAS 9.1 program (SAS Institute, Cary, NC) was used for all data processing. Demographic data and baseline characteristics were checked for normal distribution with Kolmogorov-Smirnov goodness-of-fit tests. Significant differences were evaluated in contingency tables using either Fisher’s exact tests or χ2-tests. Continuous variables were compared between groups using Student’s t tests or

Mann-Whitney rank sum tests. Two-sided P < 0.05 were considered significant, unless previous studies indicated that lower or higher values were to be expected; then, one-sided tests were performed. Pearson's correlation coefficient was used to evaluate univariate associations between the sitosterol:lathosterol and campesterol: lathosterol Z VAD FMK ratios and different ethnic groups, age, BMI and gender. Analysis of covariance was performed to obtain estimates selleck products of sitosterol:lathosterol and campesterol: lathosterol ratios between ethnic groups adjusted for age,

BMI and gender. The area under the receiver operating characteristic curve (AUC) was calculated for each serum surrogate marker of cholesterol synthesis and absorption as well as the specified ratios; the P-values for the AUC indicate significance in comparison to 0.5. For ABCG5/8 genotypes, Hardy-Weinberg equilibrium was checked using exact tests (http://ihg.gsf.de/cgi-bin/hw/hwa1.pl).14 Potential associations between gene variants

and cholelithiasis were tested in contingency tables (genotypes: Armitrage’s trend tests; alleles: χ2-tests). Table 1 presents the baseline demographic data of the study cohorts. In Germans and Chilean Hispanics, cases and matched controls are similar in age, gender, and BMI distribution. Compared with the Chilean Hispanic cohort, the German patients are older, leaner, and include more men. The small Amerindian Chilean cohort (Mapuche) was composed only of women who were relatively younger than the Hispanics and Germans. Of note, serum total cholesterol and low-density lipoprotein (LDL) cholesterol concentrations were selleck chemical lower in Mapuches compared with Hispanics independently of gallstone status. Serum lipids are similar in cases and controls in the German cohort. In contrast, in the Chilean Hispanic cohort total and LDL cholesterol levels are significantly lower in cases compared with controls. In the Chilean Hispanic and Amerindian cohorts, fasting glucose and insulin levels were also determined, yielding comparable results between cases and controls. As shown in Table 1, the IR-HOMA indexes are high and in the range of insulin resistance for both cases and controls, but they do not differ between groups.

Statistically significant findings were found in Caucasians but not in Asians or in Hispanics.

The pooled OR (95%CI, P-value) in Caucasians for −511 T carriers versus CC and for IL-1RN *2 carriers versus L/L were 1.33 (1.04–1.71, P = 0.023) and 1.31 (1.07–1.61, P = 0.010), respectively. When gastric carcinoma was classified into non-cardia (or distal) and cardia subtypes, statistically significant findings were found among non-cardia gastric cancer on the grounds that the pooled OR (95%CI, P-value) for IL-1B −511 T carriers versus CC and for IL-1RN *2 carriers versus L/L were 1.31 (1.04–1.64, P = 0.020) selleck chemicals llc and 1.47 (1.21–1.79, P = 0.000), respectively. When gastric carcinoma was classified into intestinal, diffuse, or mixed subtypes in terms of histopathology, statistically significant findings were found among intestinal type gastric carcinoma on the grounds that the pooled OR (95%CI, P-value) for IL-1B −511 T carriers versus CC, IL-1B −31 CC plus TT versus CT, and IL-1RN *2 carriers versus L/L were 1.55 (1.05–2.28, P = 0.026), 0.73 (0.60–0.89, P = 0.002), and 1.66 (1.23–2.25, P = 0.001), respectively. When genotyping techniques

were considered, statistically significant findings were found in PCR-RFLP for IL-1B −511 T carriers versus CC and LY294002 order in genotyping methods other than PCR-RFLP for IL-1B −31 CC plus TT versus CT on the grounds that pooled OR (95%CI, P-value) were 1.21 (1.03–1.42, P = 0.018) for the former and 0.87 (0.77–0.98, selleck screening library P = 0.023) for the latter. First, both fixed-effects models and random-effects models, if homogeneity was indicated (Q-test P-value was no less than 0.1), were employed and recorded and their results were compared simultaneously

due to the need for sensitivity analysis (Table 1). Except for the fact that the 95%CI were a little narrower using the fixed-effects models, the results of both models were similar in the case that Q-test P-value was no less than 0.1, indicating the robust stability of the outcomes theoretically in the absence of heterogeneity. I-squared statistic value suggested a weak to moderate to strong variation in all meta-analyses. Second, meta-analyses were conducted repeatedly when each particular study had been removed. The results indicated that fixed-effects estimates and/or random-effects estimates before and after the deletion of each study were similar at large, suggesting high stability of the meta-analysis results. The cumulative meta-analyses of associations were conducted for each of the polymorphic loci with overall gastric carcinoma in chronological order. The inclinations toward significant stable associations were evident with each accumulation of more data over time, although associations were initially much stronger. The 95%CI became increasingly narrower in the increasing sample size order, indicating that the precision of estimates was progressively boosted with the continual addition of even more cases.

3), 50 mM KCl, Tween-20 0.01%, 0.2 mM deoxyribonucleotides, 2-4 pmol of each

primer, 2 mM MgCl2, and 0.5 units hot-start Taq DNA polymerase (RighTaq, Euroclone, Milan, Italy). Samples containing 10 ng of genomic DNA were subjected to 40 cycles of denaturation (at 95°C for 30 seconds), annealing (at 62°C for 30 seconds), and elongation (at 72°C for 30 seconds) using a Techne TC-412 thermal cycler. In a total volume of 20 μL, 10 μL of the amplicons were digested with 1 unit of the BstU-I restriction endonuclease (New England Biolabs, Hitchin, UK) at 60°C overnight. The digest fragments were 135, 82, and 25 bp for the C allele and 160 and 82 bp for the T allele variant. The fragments were resolved by electrophoresis on a 3.5% agarose gel after staining with ethidium bromide. As mentioned above, 144 out of 211 patients (68.2%) underwent a liver biopsy before starting therapy. Selleck PD-1/PD-L1 inhibitor Grade and stage were scored according to the Ishak system.17 All patients were treated with a combination therapy of PEG-IFN plus ribavirin. One hundred fifty-three patients (72.5%) received peginterferon alfa-2b (PegIntron, Schering-Plough, New Jersey, USA) at a dosage of 1.5 μg/kg/week, and 58 patients (27.5%) received peginterferon alfa-2a (Pegasys, Roche, Basel, Switzerland) at a dosage of 180 μg per week. In patients infected with HCV genotypes 1, 4, and 5, ribavirin (either Rebetol, Schering-Plough,

or Copegus, Roche) was administered according to body weight (1,000 mg/day for patients weighing <75 kg, 1,200 mg/day for patients weighing ≥75 kg); in the case of infection by genotypes 2 and 3, a single ribavirin PLX3397 supplier dose of 800 mg/day was used. The duration of therapy was 48 weeks for genotypes 1, 4, and 5 and 24 weeks for genotypes 2 and 3. Rapid viral response (RVR) was defined as an

undetectable serum HCV RNA (<50 IU/mL) level 4 weeks after starting therapy. Complete early viral selleck kinase inhibitor response (cEVR) was defined as an undetectable serum HCV RNA level 12 weeks after starting therapy. The end of treatment viral response (EOT) was defined as an undetectable serum HCV RNA level after completing the treatment schedule. Sustained viral response (SVR) was defined as an undetectable serum HCV RNA level at 24 weeks after stopping antiviral therapy. Patients who achieved EOT but reverted to a detectable HCV RNA level after stopping therapy were considered relapsers. Dropout was defined as discontinuation of antiviral therapy due to adverse effects. The stopping rule consisted of therapy discontinuation in HCV 1-, 4- and 5-infected patients who either failed to obtain a reduction in serum HCV RNA concentration of at least 2 log compared with baseline at week 12 or had a detectable serum HCV RNA level after 24 weeks of therapy.18-20 Patients who met stopping rule criteria for therapy discontinuation were defined as nonresponders.