Antibody Vh Repertoire Differences between Resolving and Chronically Evolving Hepatitis C Virus Infections

Despite the production of neutralizing antibodies to hepatitis C virus (HCV), many patients fail to clear the virus and instead develop chronic infection and long-term complications. To understand how HCV infection perturbs the antibody repertoire and to identify molecular features of antibody genes associated with either viral clearance or chronic infection, we sequenced the V(D)J region of naïve and memory B cells of 6 persons who spontaneously resolved an HCV infection (SR), 9 patients with a newly diagnosed chronically evolving infection (CE), and 7 healthy donors. In both naïve and memory B cells, the frequency of use of particular antibody gene subfamilies and segments varied among the three clinical groups, especially between SR and CE. Compared to CE, SR antibody genes used fewer VH, D and JH gene segments in naïve B cells and fewer VH segments in memory B cells. SR and CE groups significantly differed in the frequency of use of 7 gene segments in naïve B cell clones and 3 gene segments in memory clones. The nucleotide mutation rates were similar among groups, but the pattern of replacement and silent mutations in memory B cell clones indicated greater antigen selection in SR than CE. Greater clonal evolution of SR than CE memory B cells was revealed by analysis of phylogenetic trees and CDR3 lengths. Pauciclonality of the peripheral memory B cell population is a distinguishing feature of persons who spontaneously resolved an HCV infection. This finding, previously considered characteristic only of patients with HCV-associated lymphoproliferative disorders, suggests that the B cell clones potentially involved in clearance of the virus may also be those susceptible to abnormal expansion

Completion of Hepatitis C Virus Replication Cycle in Heterokaryons Excludes Dominant Restrictions in Human Non-liver and Mouse Liver Cell Lines

Hepatitis C virus (HCV) is hepatotropic and only infects humans and chimpanzees. Consequently, an immunocompetent small animal model is lacking. The restricted tropism of HCV likely reflects specific host factor requirements. We investigated if dominant restriction factors expressed in non-liver or non-human cell lines inhibit HCV propagation thus rendering these cells non-permissive. To this end we explored if HCV completes its replication cycle in heterokaryons between human liver cell lines and non-permissive cell lines from human non-liver or mouse liver origin. Despite functional viral pattern recognition pathways and responsiveness to interferon, virus production was observed in all fused cells and was only ablated when cells were treated with exogenous interferon. These results exclude that constitutive or virus-induced expression of dominant restriction factors prevents propagation of HCV in these cell types, which has important implications for HCV tissue and species tropism. In turn, these data strongly advocate transgenic approaches of crucial human HCV cofactors to establish an immunocompetent small animal model

A Novel Small Molecule Inhibitor of Hepatitis C Virus Entry

Small molecule inhibitors of hepatitis C virus (HCV) are being developed to complement or replace treatments with pegylated interferons and ribavirin, which have poor response rates and significant side effects. Resistance to these inhibitors emerges rapidly in the clinic, suggesting that successful therapy will involve combination therapy with multiple inhibitors of different targets. The entry process of HCV into hepatocytes represents another series of potential targets for therapeutic intervention, involving viral structural proteins that have not been extensively explored due to experimental limitations. To discover HCV entry inhibitors, we utilized HCV pseudoparticles (HCVpp) incorporating E1-E2 envelope proteins from a genotype 1b clinical isolate. Screening of a small molecule library identified a potent HCV-specific triazine inhibitor, EI-1. A series of HCVpp with E1-E2 sequences from various HCV isolates was used to show activity against all genotype 1a and 1b HCVpp tested, with median EC50 values of 0.134 and 0.027 µM, respectively. Time-of-addition experiments demonstrated a block in HCVpp entry, downstream of initial attachment to the cell surface, and prior to or concomitant with bafilomycin inhibition of endosomal acidification. EI-1 was equally active against cell-culture adapted HCV (HCVcc), blocking both cell-free entry and cell-to-cell transmission of virus. HCVcc with high-level resistance to EI-1 was selected by sequential passage in the presence of inhibitor, and resistance was shown to be conferred by changes to residue 719 in the carboxy-terminal transmembrane anchor region of E2, implicating this envelope protein in EI-1 susceptibility. Combinations of EI-1 with interferon, or inhibitors of NS3 or NS5A, resulted in additive to synergistic activity. These results suggest that inhibitors of HCV entry could be added to replication inhibitors and interferons already in development

Musculoskeletal fitness and balance in older individuals (65–85 years) and its association with steps per day: a cross sectional study

BACKGROUND: There is limited normative, objective data combining musculoskeletal fitness (MSF), balance and physical activity (PA) among older adults. The aims were therefore to; 1) describe MSF and balance in older Norwegian adults focusing on age- and sex-related differences; 2) investigate the associations among MSF, balance and objectively-assessed PA levels. METHODS: This was part of a national multicenter study. Participants (65–85 years) were randomly selected from the national population registry. We used ActiGraph GT1M accelerometers to measure PA. Balance and MSF were assessed using: one leg standing (OLS), handgrip strength (HG), static back extension (SBE), sit and reach (SR), back scratch right, left arm over (BSR, BSL). Univariate analyses of variance were used to assess sex differences within the different MSF and balance tests and for comparisons among multiple age groups. Linear regression analysis was used to investigate how PA (expressed in 1000 steps increments) was associated with MSF and balance. RESULTS: 85 women and 76 men were included. Mean age (standard deviation (SD)) was 73.2 (5.4) years for women and 72.3 (4.8) years for men. The youngest participants (65–69 years) had significantly better mean OLS- and SBE results compared with older participants. Women (65–85 years) had significantly better mean SR, BSR, BSL and SBE results compared with men (65–85 years). Men had significantly better mean HG results compared with women. No sex differences in mean OLS results were observed. A daily increment of 1000 steps was associated with better mean test scores for OLS- and SBE tests (b = 1.88, 95 % CI: 0.85 to 2.90 (p ≤ 0.001) and b = 4.63, 95 % CI: 1.98 to 7.29 (p = 0.001), respectively). CONCLUSION: The youngest (65–69 years) had better static balance and muscular endurance in trunk extensors compared with older participants. Older women (65–85 years) had better joint flexibility than older men (65–85 years), whereas older men had better handgrip strength than older women. A higher PA level was associated with better static balance and muscular endurance in trunk extensors in older individuals. This study provides important normative data, and further investigation of trunk endurance and static balance as key foci for PA interventions in elderly is warranted

Effects of long-term oral testosterone undecanoate therapy on urinary symptoms: data from a 1-year, placebo-controlled, dose-ranging trial in aging men with symptomatic hypogonadism

Background: There has been a longstanding question as to whether testosterone therapy could precipitate or worsen urinary symptoms in aging men. We investigated the effects of 1-year oral testosterone undecanoate (TU) therapy on urinary symptoms in aging, hypogonadal men. Methods: A total of 322 men 50 years with symptomatic testosterone deficiency participated in a 1-year, randomized, multicenter, double-blind trial. Patients received placebo or oral TU 80mg/day, 160mg/day, or 240mg/day. Results and limitations: Compared with placebo, treatment with oral TU at doses of 80mg/day and 160mg/day resulted in no significant change in IPSS urinary symptoms or quality of life (QoL) scores. Treatment with oral TU 240mg/day led to a statistically significant, but clinically insignificant, improvement in IPSS total score and a significant improvement in IPSS QoL score. None of the TU doses tested had a significant effect on PSA or PV. Conclusions: Long-term oral TU therapy had no deleterious effects on IPSS total score and did not change PV and PSA in aging, hypogonadal men. Oral TU therapy at a dose of 240mg/day may even improve IPSS QoL score