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Differential modulation of innate immune responses in human primary cells by influenza A viruses carrying human or avian non-structural 1 proteins.

The influenza A virus (IAV) non-structural protein 1 (NS1) contributes to disease pathogenesis through the inhibition of host innate immune responses. Dendritic cells (DCs) release interferons (IFN), pro-inflammatory cytokines and promote the adaptive immunity upon viral infection. In order to characterize the strain-specific effects of IAV NS1 on human DC activation, we infected human DCs with a panel of recombinant viruses with the same backbone (A/Puerto Rico/08/1934) expressing different NS1 from human and avian origin. We found that these viruses induced a clearly distinct phenotype in DCs. Specifically, viruses expressing NS1 from human IAV (either H1N1 or H3N2) induced higher levels of expression of type I (IFNα and IFNβ) and type III (IFNλ1-3) than viruses expressing avian IAV NS1 proteins (H5N1, H7N9 and H7N2), but the differences observed in the expression of pro-inflammatory cytokines like TNFα or IL-6 were not significant. In addition, using Imaging Flow Cytometry, we found that human and avian NS1 segregate based on their subcellular trafficking dynamics, which might be associated with the different innate immune profile induced in DCs by viruses expressing those NS1 proteins. Innate immune responses induced by our panel of IAV recombinant viruses were also characterized in Normal Human Bronchial Epithelial cells and the results were consistent with those in DCs. Altogether, our results reveal an increased ability of NS1 from avian viruses to antagonize innate immune responses in human primary cells as compared to NS1 from human viruses which could contribute to the severe disease induced by avian IAV in humans. Influenza A viruses (IAV) cause seasonal epidemics which result in an important health and economic burden. Wild aquatic birds are the natural host of IAV. However, IAV can infect diverse hosts, including humans, domestic poultry, pigs, and others. IAV circulating in animals occasionally cross the species barrier infecting humans, which results in mild to very severe disease. In some cases, these viruses can acquire the ability to transmit among humans and initiate a pandemic. The non-structural (NS) 1 protein of IAV is an important antagonist of the innate immune response. In this study, using recombinant viruses and primary human cells, we show that NS1 proteins from human and avian hosts show intrinsic differences in the modulation of the innate immunity in human dendritic cells and epithelial cells, as well as different cellular localization dynamics in infected cells.

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[Low-levels of HBsAg quantification at 48-week in HBeAg-negative chronic hepatitis B patients are the advantageous population for HBsAg clearance].

To analyze the therapeutic effect on HBeAg-negative chronic hepatitis B patients treated with Peg-IFNα-2a combined with NAs to obtain the influencing factors for predicting HBsAg clearance. A retrospective study was conducted to investigate the effect of pegylated interferon alpha-2a combined with nucleoside analogues (lamivudine/adefovir dipivoxil) on HBeAg-negative chronic hepatitis B. The treatment course was 96 weeks. Patients were followed up 120 weeks after the treatment. HBsAg clearance at 120 weeks was taken as the objective of the study. Logistic regression and receiver operating characteristic curve analysis screened the related factors affecting HBsAg clearance. (2) test was used to compare count data. 111 patients were treated with pegylated interferon alpha-2a combined with nucleoside analogues, and 107 patients completed the scheduled course of treatment and follow-up. HBsAg clearance rate at120 week was 29.0% (31/107). The influencing factors for analysis were: (1) gender had no effect on HBsAg clearance rate; age and baseline levels of HBV DNA and alanine aminotransferase had no significant effect on HBsAg clearance; low baseline level of HBsAg (< 3.023 lgIU/ml) was beneficial to HBsAg clearance. The area under the working characteristic curve of the subjects was 0.746, the positive predictive value was 44.4%, and the negative predictive value was 86.8%. (2) HBsAg quantification or decline in 24 weeks and 48 weeks of treatment had a good predictive effect on HBsAg clearance, and the 48 weeks predicted value was higher than 24 weeks. When the HBsAg quantification was≤2.070 lgIU/ml at 48 weeks, the area under the receiver operating characteristic curve was 0.931, the positive predictive value was 52.8%, and the negative predictive value was 94.4%. When HBsAg decreased from baseline to≥0.991 lgIU/ml, the area under the receiver operating characteristic curve was 0.888, the positive predictive value was 50.8%, and the negative predictive value was 97.9%. (3) The analysis of HBsAg subgroup levels at 48 weeks suggested that the "interval analysis" can forecast HBsAg clearance more exactly than "nodal analysis" .The final HBsAg clearance rate of 100 IU/ml < HBsAg≤1 000 IU/ml, 10 IU/ml < HBsAg≤100 IU/ml and HBsAg≤10 IU/ml groups reached 6.7%, 31.8% and 67.7%, respectively. (4) The ALT abnormal group in the course of treatment obtained a higher HBsAg clearance rate (48.0%, 12/25). 96-weeks long-term treatment with pegylated interferon-alpha -alpha-2a combined with nucleoside analogues for HBeAg-negative chronic hepatitis B has a good predictive value for HBsAg clearance at baseline and during treatment. The "interval level" of HBsAg at 48-weeks is more accurate in predicting HBsAg clearance, suggesting that HBeAg-negative chronic hepatitis B patients with low HBsAg levels at 48-weeks are the advantageous populations with HBsAg clearance. These patients are worthy of prolonged treatment to pursue "clinical cure".

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Treatment of Elderly Patients with Chronic Hepatitis C: A Retrospective Cohort Study.

The prevalence of chronic hepatitis C increases in elderly patients. The aims of this study were to identify the factors associated with hepatocellular carcinoma (HCC) and end-stage liver disease development and to evaluate the efficacy and safety of pegylated interferon (PEG-IFNα) plus ribavirin (RBV) therapy in elderly patients. A retrospective cohort study included all consecutive pa-tients with hepatitis C virus (HCV) infection treated with PEG-IFNα+RBV between 2003 and 2013. Elderly patients had a higher frequency of poor prognostic factors including genotype 1 infec-tion, high fibrosis, and high fibrosis index based on four factors (FIB-4) score. The sustained virologic response (SVR) rate for genotype 1 was significantly lower (35.8% vs. 57.1%), while the frequency of PEG-IFNα (27.2% vs. 7.8%), RBV dose reduction (19.6% vs. 9.7%) and treatment discontinuation (13.0% vs. 4.1%) was significantly higher in elderly patients. However, age was not associated with SVR in multivariate analysis, and comparable SVR rates were achieved when adjusted for fibrosis score (Ishak ≤3: 66.7% vs. 69.8%). During the follow-up, HCC was diagnosed in 18 elderly patients (3 SVR+, 4 SVR- and 9 untreated patients). In conclusion, selected elderly patients can achieve comparable SVR rates as younger patients, but with a higher rate of side effects. Since complications of HCV infection occur more frequently in elderly patients, they should be given priority for antiviral therapy.

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Efficacy and safety of combined high-dose interferon and red light therapy for the treatment of human papillomavirus and associated vaginitis and cervicitis: A prospective and randomized clinical study.

We evaluated the efficacy and safety of combined high-dose interferon (IFN) and red light therapy for the treatment of subclinical and latent human papillomavirus (HPV) infections.

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Establishment of a reverse genetics system for duck Tembusu virus to study virulence and screen antiviral genes.

Recently, a newly emerged avian flavivirus, duck Tembusu virus (TMUV), was identified as the causative agent of a serious duck viral disease in Asia. Its rapid spread and expanded host range have raised substantial concerns regarding its potential threat to non-avian hosts, including humans. In this study, we report an infectious cDNA clone for a clinical strain CQW1 isolated from Southwest China, which is representative of the disease outbreak in the Chinese mainland. We generated a full-length cDNA clone pACYC FL-TMUV, which is infectious, and this cDNA clone-derived recombinant TMUV (rTMUV) showed comparative growth kinetics in both BHK21 cells and DEF cells compared with parental TMUV (pTMUV). In addition, rTMUV also showed the same high virulence in 9-day-old duck embryos as that in pTMUV, suggesting that rTMUV possessed similar properties to the natural virus both in vitro and in vivo. Based on the cDNA-clone, we first generated a reporter TMUV (TMUV-RLuc) carrying a Renilla luciferase (RLuc) gene. The luciferase kinetics of TMUV-RLuc were determined both in BHK21 and DEF cells. It seems that TMUV-RLuc grew well in vitro; however, the insertion of the RLuc gene attenuated viral replication in vitro. The higher viral titres of TMUV-RLuc were observed in BHK21 compared with that in DEF cells. The antiviral effects of exogenous-expressed duck RIG-I, MDA5, STING, MAVS, TBK1, IFNα and IFNγ were studied in vitro by using TMUV-RLuc. Our reverse genetics system will provide a multicomponent platform for the pathogenesis study of duck TMUV and the development of molecular countermeasures against duck TMUV infection.

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Serum miRNAs Predicting Sustained HBs Antigen Reduction 48 Weeks after Pegylated Interferon Therapy in HBe Antigen-Negative Patients.

The therapeutic goal for hepatitis B virus (HBV) infection is HBs antigen (HBsAg) seroclearance, which is achieved through 48-week pegylated interferon (Peg-IFN) therapy. This study aimed to identify predictive biomarkers for sustained HBsAg reduction by analyzing serum microRNAs. Twenty-two consecutive chronic HBV infection patients negative for HBe antigen (HBeAg) with HBV-DNA levels <5 log copies/mL, alanine aminotransferase (ALT) <100 U/L, and compensated liver functions, were enrolled. The patients were subcutaneously injected with Peg-IFNα-2a weekly for 48 weeks (treatment period), followed by the 48-week observation period. HBsAg 1-log drop relative to baseline levels recorded at the end of the observation period was considered effective. Sera were obtained at weeks 0 and 24 during the treatment period analyzed for microRNAs. The microRNA (miRNA) antiviral activity was evaluated in vitro using Huh7/sodium taurocholate cotransporting polypeptide (NTCP) cells. As a result, six patients achieved the HBsAg 1-log drop after the observation periods. Comparison of serum microRNA levels demonstrated that high miR-6126 levels at week 24 predicted HBsAg 1-log drop. Furthermore, miR-6126 reduced HBsAg in culture medium supernatants and intracellular HBV-DNA quantities in Huh7/NTCP cells. In conclusion, high serum miR-6126 levels during Peg-IFN therapy predicted the HBsAg 1-log drop 48 weeks after the completion of therapy. In vitro assays revealed that miR-6126 was able to suppress HBsAg production and HBV replication.

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[Changes and clinical significance of γδT cells in peripheral blood of patients with chronic hepatitis B during pegylated interferon α-2a treatment].

To observe the changes of γδT cells in the peripheral blood of patients with chronic hepatitis B (CHB) during pegylated interferon α-2a treatment, and to analyze the correlation between clinical indicators and curative effects. Peripheral blood of hepatitis B e antigen (HBeAg)-positive CHB patients were collected at different time points of Peg-IFNα-2a treatment, including 17 patients at 0 weeks, 20 patients at 12 weeks, 20 patients at 24 weeks, and 16 patients at 48 weeks. From these 11 patients, blood samples were frequently observed at 0, 12, 24, and 48 weeks of treatment. The frequencies of γδT and its subpopulation cells Vδ1T, Vδ2T, effector memory γδT (γδTem), central memory γδT (γδTcm), initial γδT (γδTnaive) and terminal differentiation effect γδT (γδTeff) cells in peripheral blood were detected by flow cytometry. Liver function, serum HBV markers and HBV DNA levels were measured simultaneously. SPSS 23.0 statistical software was used to analyze the differences in cell proportions at each treatment time point, and the correlation between cell proportions and alanine aminotransferase (ALT), HBsAg, HBeAg or HBV DNA levels. In addition, the correlation between the proportions of γδT and its subpopulation cells and the response to Peg-IFNα-2a treatment in the 11 patients with continuous follow-up were analyzed. The percentage of γδT and Vδ2T cells in peripheral blood of patients with CHB decreased gradually during the period of 0-48 weeks of Peg-IFNα-2a treatment. The percentages of γδT cells and Vδ2T cells at 48 weeks were 6.89% (5%, 8.15%), 4.61% (2.16%, 6.50%), respectively; significantly lower than the 0 week [12.5% ​​(7.73%, 19%), 6.59% (3.86%, 13.62%)], the differences were statistically significant ( < 0.05). The proportions of Vδ1T, γδTem, γδTcm, γδTnaive, or γδTeff subpopulations were not statistically different at each time points (all > 0.05). At the same time, the levels of ALT, HBsAg, HBeAg or HBV DNA were positively correlated with the ratio of γδT or Vδ2T cells ( < 0.05). Among the 11 patients with continuous followed- up, the proportion of γδTem cells in responders was significantly lower than that of non-responders at each time points, and the difference was statistically significant ( < 0.05). There was no statistically significant difference between the two groups (all > 0.05). The proportion of γδT cells in the course of CHB treatment with Peg-IFNα-2a reduces the liver inflammation by decreasing the replication of HBV virus. Chronic hepatitis B patients with a lower proportion of effector memory (γδTem) cells may be more likely to get better response with Peg-IFNα-2a.

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Defective Dendritic Cell Cytotoxic Activity of High-Grade Glioma Patients' Results from the Low Expression of Membrane TNFα and Can Be Corrected In Vitro by Treatment with Recombinant IL-2 or Exogenic Double-Stranded DNA.

Besides initiation of tumor-specific T cell immunity, dendritic cells (DCs) are endowed with tumoricidal activity. Previously, we showed that monocyte-derived DCs of high-grade glioma patients generated in the presence of interferon alpha (IFNα) (IFN-DCs) have impaired cytotoxic activity against tumor necrosis factor alpha (TNFα)-sensitive HEp-2 tumor cells. Herein, we demonstrate that decreased transmembrane TNFα (tmTNFα) expression, but not soluble TNFα (sTNFα) production by high-grade glioma patient IFN-DCs, determines the defective tumoricidal activity against TNFα-sensitive HEp-2 cells. Blocking TNFα-converting enzyme or stimulation of patient IFN-DCs with rIL-2 or dsDNA enhances tmTNFα expression on IFN-DCs and significantly increases their cytotoxicity. Decreased tmTNFα expression on patient IFN-DCs is not caused by downregulation of pNFκB. Neither rIL-2 nor dsDNA upregulates tmTNFα expression on patient IFN-DCs via an increase of pNFκB. The current study shows an important role of tmTNFα as mediator of IFN-DC tumoricidal activity and as molecular target for the restoration of defective DC killer activity in high-grade glioma patients.

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Effectiveness of Sofosbuvir, Ribavirin and PEG-IFNα-2a in the Treatment of Naïve Egyptian Patients With Chronic Hepatitis C Virus Genotype 4.

Egypt is one of the largest epidemic areas of hepatitis C virus (HCV) in the world. Its prevalent genotype is 4 with a majority of subtype 4a. In 2013, the Food and Drug Administration approved a new direct-acting antiviral drug (sofosbuvir) to treat patients with chronic HCV infection. In Egypt, the patients are already being treated with sofosbuvir in conjunction with ribavirin and pegylated interferon alfa-2a (PEG-IFNα-2a) for 12 weeks since 2015. The present study was planned to explain the efficacy of this treatment regimen against the HCV genotype 4a in Egyptian patients and its pretreatment predictive factors of virological response.

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Pretreatment microRNA levels can predict HBsAg clearance in CHB patients treated with pegylated interferon α-2a.

To investigate the predictive capability of microRNAs (miRNAs) prior treatment for HBsAg clearance in chronic hepatitis B (CHB) treated with pegylated interferon α-2a (PEG-IFNα-2a).

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