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Nucleotide analogue improves interferon responsiveness in HBV-infected human hepatocytes
Masataka Tsuge1,3, Nobuhiko Hiraga1,2, Eisuke Murakami1,2, Michio Imamura1,2, Hiromi Abe1,2, Daiki Miki1,2, Hidenori Ochi1,2, C. Nelson Hayes1,2, Kazuaki Chayama1,2;
1Department of Gastroenterology and Metabolism, Hiroshima university, Hiroshima, Japan; 2Liver Research Project Center, Hiroshima university, Hiroshima, Japan; 3Natural Science Center for Basic Research and Development, Hiroshima university, Hiroshima, Japan
Background: It has been reported that interferon treatment could reduce HBs antigen (HBsAg) production in patients with chronic hepatitis B virus (HBV) infection. However, only limited HBsAg reduction is observed in patients treated with interferon therapy. One cause of this limitation may be that interferon responsiveness in human hepatocytes is suppressed by HBV infection, and, therefore, interferon stimulated genes (ISGs) are not induced sufficiently to promote anti-viral effects. In the present study, we analyzed whether the suppression of HBV replication using nucleotide analogues (NAs) could improve interferon responsiveness in HBV infected human hepatocytes. Methods: Thirty-seven chronic hepatitis B patients were enrolled. Twenty patients underwent sequential interferon therapy, which included 6 months of conventional interferon therapy, running from one month prior to discontinuation until 5 months after discontinuation of NA therapy. The remaining 17 patients underwent interferon mono-therapy. Serum HBsAg titers were measured every year for 5 years after interferon therapy. To confirm the clinical results, we performed an in vitro study using T23 cells, which were generated from HepG2 cells stably transfected with an HBV expression plasmid. T23 cells were treated with or without the NA entecavir for 5 days. The cells were then treated with IFN for 6 hours and harvested. To evaluate the interferon responsiveness, ISG mRNA levels were quantified by real time PCR. Results: In the clinical study, more than 1 Log IU/ml reduction of HBsAg titer was achieved in 11 of 37 patients (interferon mono-therapy: 2, Sequential therapy: 9). By univariate analysis, the following factors, gender, serum HBsAg level, the existence of HBeAg, and prior NA therapy, were associated with HBsAg reduction (P=0.007, P=0.027, P=0.031, P=0.037, respectively). From the clinical results, it was predicted that interferon responsiveness might be improved by prior NA therapy. To verify these results, in vitro experiments were performed. In the absence of HBV, the ISGs MxA and OAS1 were significantly induced by interferon treatment (19.2-fold, 9.7-fold, respectively). However, in T23 cells, inductions of these ISGs was suppressed (P=0.0495, P=0.0495, respectively). After entecavir treatment, interferon responsiveness was restored and ISG induction increased (P=0.0495, P=0.0339, respectively). Conclusions: Prior NA therapy could improve interferon responsiveness in HBV infected human hepatocytes. To improve the anti-viral effects in chronic hepatitis B patients, it might be necessary to revise the way of using NAs and interferons. 作者: StephenW 时间: 2014-10-14 11:08