Omar Galdame

Gender susceptibility to chronic hepatitis C virus infection associated with interleukin 10 promoter polymorphism.

ABSTRACT Elevated levels of interleukin 10 (IL-10) were previously described for chronically hepatitis C virus (HCV)-infected patients. We determined by a sequence-specific oligonucleotide probing technique the IL-10 promoter genotypes in 286 Argentinean HCV patients grouped according to disease outcome. The GG genotype (position −1082) is known to be associated with high IL-10 production, GA is considered an intermediate producer, and AA is associated with low IL-10 production. We found an increase in frequency of the GG genotype in female patients who do not eliminate the virus (RNA+). In these patients, the GG frequency was 0.19, versus 0.10 in controls (P = 0.03). This association became more significant in those RNA+ female patients with elevated hepatic transaminases (GG frequency of 0.25; P = 0.0013). Additionally, this genotype frequency was higher in noncirrhotic female patients than in controls (GG frequency for noncirrhotic female patients was 0.31; P = 0.009). In RNA− patients, the GA frequency was elevated compared with that in controls (GA frequency of 0.76 in RNA− patients versus 0.48 in controls; P = 0.01), that in all HCV patients (GA frequency of 0.43; P = 0.001), and that in RNA+ patients (GA frequency of 0.40; P = 0.0005). We conclude that a gender effect is observed with women carrying the GG high IL-10 producer genotype. The higher levels of IL-10 present in those individuals are associated with a higher risk of an inefficient clearance of the HCV and the development of a chronic HCV infection together with a lower risk of progression to cirrhosis in female patients.

The Role of Serum Biomarkers in Predicting Fibrosis Progression in Pediatric and Adult Hepatitis C Virus Chronic Infection

Background/Aims Liver biopsy represents the gold standard for damage evaluation, but noninvasive serum markers that mirror liver fibrosis progression are actual goals both in adults and especially in children. The aim was to determine specific serum markers that correlate with liver fibrosis progression during chronic HCV infection. Methods Liver biopsies and concomitant serum samples from 22 pediatric and 22 adult HCV patients were analyzed. Histological parameters were evaluated. On serum TGF-ß1, tissue inhibitor of matrix metalloprotein inhibitor-1 (TIMP-1), hyaluronic acid (HA) and aminoterminal peptide of procollagen type III (PIIINP) were tested. Results Significant fibrosis (F≥2) and advanced fibrosis (F≥3) represented 64% and 20%, respectively in children; while 54% F≥2 and 23% F≥3 in adults. Hyaluronic acid (p = 0.011) and PIIINP (p = 0.016) were related to worse fibrosis stages only in adults, along with TIMP-1 (p = 0.039) just in children; but TGF-ß1 was associated with mild fibrosis (p = 0.022) in adults. The AUROC of TIMP-1 in children to discriminate advanced fibrosis was 0.800 (95%IC 0.598–0.932). In adults, the best AUROCs were that of HA, PIIINP and TGF-ß1 [0.929 (IC95% 0.736–0.994), 0.894 (IC95% 0.689–0.984) and 0.835 (IC95% 0.617–0.957)], respectively. In children, according to the cut off (165.7 ng/mL) value for TIMP-1, biopsies could have been avoided in 72% (18/25). Considering the cut off for HA (109.7 ng/mL), PIIINP (9.1 µg/L), and TGF-ß1 (10,848.3 pg/mL), biopsies could have been avoided in 87% (19/22) of adult patients by using HA and 73% (16/22) using PIIINP or TGF-ß1. Conclusions In adults given the diagnostic accuracy of HA, PIIINP, TGF-ß1, their combination may provide a potential useful tool to assess liver fibrosis. This first pediatric study suggests that TIMP-1 is clinically useful for predicting liver fibrosis in HCV patients.

Serum Apoptosis Markers Related to Liver Damage in Chronic Hepatitis C: sFas as a Marker of Advanced Fibrosis in Children and Adults While M30 of Severe Steatosis Only in Children

Background Liver biopsy represents the gold standard for evaluating damage and progression in patients with chronic hepatitis C (CHC); however, developing noninvasive tests that can predict liver injury represents a growing medical need. Considering that hepatocyte apoptosis plays a role in CHC pathogenesis; the aim of our study was to evaluate the presence of different apoptosis markers that correlate with liver injury in a cohort of pediatric and adult patients with CHC. Methods Liver biopsies and concomitant serum samples from 22 pediatric and 22 adult patients with CHC were analyzed. Histological parameters were evaluated. In serum samples soluble Fas (sFas), caspase activity and caspase-generated neoepitope of the CK-18 proteolytic fragment (M30) were measured. Results sFas was associated with fibrosis severity in pediatric (significant fibrosis p = 0.03, advanced fibrosis p = 0.01) and adult patients (advanced fibrosis p = 0.02). M30 levels were elevated in pediatric patients with severe steatosis (p = 0.01) while in adults no relation with any histological variable was observed. Caspase activity levels were higher in pediatric samples with significant fibrosis (p = 0.03) and they were associated with hepatitis severity (p = 0.04) in adult patients. The diagnostic accuracy evaluation demonstrated only a good performance for sFas to evaluate advanced fibrosis both in children (AUROC: 0.812) and adults (AUROC: 0.800) as well as for M30 to determine steatosis severity in children (AUROC: 0.833). Conclusions Serum sFas could be considered a possible marker of advanced fibrosis both in pediatric and adult patient with CHC as well as M30 might be a good predictor of steatosis severity in children.

Characterization of the basal core promoter and precore regions in anti-HBe-positive inactive carriers of hepatitis B virus

BACKGROUND The study of hepatitis B virus (HBV) genomic heterogeneity has become a major issue in investigations aimed at understanding the relationship between HBV mutants and the wide spectrum of clinical and pathological conditions associated with HBV infection. Although most chronically infected HBV patients are inactive carriers, several virological aspects of this state remain unclear. METHODS In order to determine the prevalence and clinical significance of mutations in the basal core promoter (BCP) and precore (pC) regions among inactive carriers, the nucleotide sequences from 41 inactive carriers were analyzed and compared with those from 29 individuals with chronic active hepatitis. RESULTS Genotypes A (24.3%), D (37.1%), F1b (12.9%), and F4 (18.6%) were the most prevalent. Mutations in the BCP/pC regions were observed in most of the inactive carriers (92.7%) and in most of the patients with chronic active hepatitis (93.1%). The prevalence of mutation 1764(A) was significantly higher in patients with chronic active hepatitis (65.5%) than in inactive carriers (36.6%) (p=0.038), whereas the prevalences of mutations at the other positions analyzed were not significantly different. Older patients (>50 years) showed BCP/pC patterns with a higher number of substitutions. Mutations were found to be biased by genotype: the 1896(A) mutation was highly prevalent in genotypes D and F4, while alternative substitutions in the pC region were more prevalent in genotypes A and F1b. CONCLUSIONS Mutations in the BCP/pC regions are the hallmark of chronic anti-HBe-positive individuals; nevertheless, the even distribution of mutations in active and inactive carriers suggests that BCP/pC mutations may occur during HBV infection not strictly related to the HBV infection activity.

HBV genotype F: natural history and treatment.

The analysis of the HBV genome revealed the existence of 10 genotypes, named A-J. Evidence of the influence of the different genotypes in the natural history and treatment response to nucleoside/nucleotide analogues or interferon-based regimens is scant. HBV genotype F is one of the most prevalent circulating genotypes in South America and the Arctic Circle. Since most of the available information on HBV is from Asia, the US and Europe, it reflects their predominant genotypes: A, B, C and D. To date, the evidence is not fully confirmed, but it appears that genotype F chronic hepatitis B is associated with a more aggressive course of liver disease, reflected by higher histological indexes, a higher risk of development of hepatocellular carcinoma and a higher rate of liver-related mortality. In terms of treatment response, the available data is, unfortunately, even more limited; however, what data is available suggests acceptable and similar response rates to pegylated interferon-α2a in genotype F compared to genotype A. Response rates to nucleoside/nucleotide analogues is not influenced by genotype. The review of this limited data sheds light on the necessity to conduct further studies in South America and the Arctic Circle in order to better understand the different aspects of HBV genotype F, especially in relation to treatment response.

Hepatitis B virus genotype distribution and genotype-specific BCP/preCore substitutions in acute and chronic infections in Argentina.

Aim In order to assess Hepatitis B Virus genotype (g) and subgenotype (sg) implications in the course of infection, 234 HBsAg positive patients in different infection stages were characterized (66 acute infections, 63 HBeAg positive chronic infections and 105 anti-HBe positive chronic infections). Results Overall, sgA2 (17.9%), gD (20.9%), sgF1b (34.2%) and sgF4 (19.7%) were the most prevalent. Subgenotype F1b was overrepresented in acute and chronic HBeAg infections (56.1%), whereas gD was the most frequent (40.0%) in anti-HBe positive chronic infections. Among chronic infections, HBeAg positivity rates were 50.0, 12.5, 62.8 and 35.3% for sgA2, gD, sgF1b and sgF4, respectively (p <0.05). A bias toward BCP/preCore mutations was observed among genotypes. In anti-HBe positive chronic infections, sgF1b was more prone to have A1762T/G1764A mutation than sgA2, sgF4 and gD (75.0, 40.0, 33.3 and 31.8%, p<0.005), whereas in the pC region, gD and sgF4 were more likely to have G1896A than sgA2 and sgF1b (81.0, 72.7, 0.0 and 31.3%, p <0.001). The unexpected low frequency of the G1896A mutation in the sgF1b (despite carrying 1858T) prompted us to perform a further analysis in order to identify genotype-specific features that could justify the pattern mutations observed. A region encompassing nucleotides 1720 to 1920 showed the higher dissimilarity between sgF1b and sgF4. Genotypes and subgenotypes carrying the 1727G, 1740C and 1773T polymorphisms were prevented to mutate position 1896. Discussion HBeAg seroconversion is a critical event in the natural history of HBV infection. Differences in the HBeAg positivity rate might be relevant since different studies have observed that delayed HBeAg seroconversion is associated with a more severe clinical course of infection, highlighting the critical role that genotypes/subgenotypes might play in the progression of HBV infection. Polymorphisms in the regions 1720 to 1920 could be involved in the molecular mechanisms underlying seroconversion of each genotype/subgenotype.

Full-Length Genome Characterization of Hepatitis B Virus Genotype H Strain Isolated from Serum Samples Collected from Two Chronically Infected Patients in Argentina

Eight genotypes (A to H) of hepatitis B virus (HBV) have been described so far ([11][1]). Each genotype seems to have a restricted geographical distribution; therefore, it constitutes an invaluable tool in tracing HBV molecular evolution and the pattern and modes in which HBV spreads ([5][2]).

Hepatitis C in Argentina: epidemiology and treatment.

Hepatitis C is the leading cause of chronic hepatitis, cirrhosis, and liver cancer in Argentina, where from 1.5% to 2.5% of adults are infected. Most of the infections were acquired 30–50 years ago. It is estimated that more than half of infected individuals are not aware of their infection. Even though the prevalence in blood donors has decreased to 0.45% at present, many high-prevalence populations still exist, where the reported prevalence ranges from 2.2% to 7.1%. Therapy is recommended for patients with fibrosis, in order to prevent disease progression, hepatic decompensation, and hepatocellular carcinoma. Great advances were achieved in the treatment of genotype 1 infection since the development and release of boceprevir and telaprevir. When either of these protease inhibitors is associated with peginterferon plus ribavirin, the sustained virological response (SVR) rate improves from 40%–50% to 67%–75%. For genotype 2 and 3 infection, treatment with peginterferon plus ribavirin is still the standard of care, with SVR rates of 70%–90%. There are significant new antivirals in development, and some of them are close to being released. These drugs will most likely be the future standard of care for all genotypes, and will be incorporated in better-tolerated and highly effective all-oral regimes. The impact that these new therapies might have in health-related economics is unpredictable, especially in developing countries. Each country must carefully evaluate the local situation in order to implement proper screening and treatment programs. Difficult-to-treat patients, such as those with decompensated cirrhosis, patients in hemodialysis, and those with other significant comorbidities, might not be able to receive these new therapeutic approaches and their management will remain challenging.

Distinctive intrahepatic characteristics of paediatric and adult pathogenesis of chronic hepatitis C infection

Mechanisms leading to liver damage in chronic hepatitis C (CHC) are being discussed, but both the immune system and the virus are involved. The aim of this study was to evaluate intrahepatic viral infection, apoptosis and portal and periportal/interface infiltrate in paediatric and adult patients to elucidate the pathogenesis of chronic hepatitis C. HCV-infected, activated caspase-3(+) and TUNEL(+) hepatocytes, as well as total, CD4(+), CD8(+), Foxp3(+) and CD20(+) lymphocytes infiltrating portal and periportal/interface tracts were evaluated in 27 paediatric and 32 adult liver samples by immunohistochemistry or immunofluorescence. The number of infected hepatocytes was higher in paediatric than in adult samples (p 0.0078). In children, they correlated with apoptotic hepatocytes (activated caspase-3(+) r = 0.74, p < 0.0001; TUNEL(+) r = 0.606, p 0.0017). Also, infected (p = 0.026) and apoptotic hepatocytes (p = 0.03) were associated with the severity of fibrosis. In adults, activated caspase-3(+) cell count was increased in severe hepatitis (p = 0.009). Total, CD4(+), CD8(+) and Foxp3(+) lymphocyte count was higher in adult samples (p < 0.05). Paediatric CD8(+) cells correlated with infected (r = 0.495, p 0.04) and TUNEL(+) hepatocytes (r = 0.474, p = 0.047), while adult ones correlated with activated caspase-3(+) hepatocytes (r = 0.387, p 0.04). In adults, CD8(+) was associated with hepatitis severity (p < 0.0001) and correlated with inflammatory activity (CD8(+) r = 0.639, p 0.0003). HCV, apoptosis and immune response proved to be involved in CHC pathogenesis of both paediatric and adult patients. However, liver injury in paediatric CHC would be largely associated with a viral cytopathic effect mediated by apoptosis, while in adults it would be mainly associated with an exacerbated immune response.

Identification of a new clade of hepatitis B virus genotype F.

Hepatitis B virus (HBV) is classified into eight main genotypes (A-H) and several subgenotypes. Here, three new genotype F complete genome sequences isolated from patients from Buenos Aires city are reported. The new sequences form a separate monophyletic group from the previously known subgenotype F4 strains. Based on results of phylogenetic, genetic distance and evolutionary analyses, the name F4b is proposed for these isolates and F4a for the formerly known as F4. The identification of new clusters allows deepening the knowledge about the diversification process and evolutionary history of HBV.