Tianbao Wang1, Xinwei Wang1, Ya Zhuo1, Changyun Si1, Lu Yang2, Lijun Meng2, Bin Zhu3. 1. Infectious Disease Department of the First Affiliated Hospital of Xinxiang Medical University, Weihui, 453100, China. 2. Gastroenterology Department of the First Affiliated Hospital of Xinxiang Medical University, Weihui, 453100, China. 3. Infectious Disease Department of the First Affiliated Hospital of Xinxiang Medical University, Weihui, 453100, China. Electronic address: zhubinxxmu@gmail.com.
Abstract
ETHNOPHARMACOLOGICAL RELEVANCE: Hepatitis B virus (HBV) infection frequently results in both acute and chronic hepatitis and poses serious threats to human health worldwide. Despite the availability of effective HBV vaccine and anti-HBV drugs, apparently inevitable side effects and resistance have limited its efficiency, thus prompt the search for new anti-HBV agents. The traditional Chinese medicine Radix Isatidis has been used for thousands of years, mainly for the treatment of viral and bacterial infection diseases including hepatitis. AIM OF THE STUDY: In this study, antiviral activities of a Radix Isatidis (Isatis indigotica Fortune) polysaccharide (RIP) were evaluated in vitro model using the HepG2.2.15 cell line and the underlying mechanism was elucidated with the aim of developing a novel anti-HBV therapeutic agent. MATERIALS AND METHODS: Structure features of the purified polysaccharide RIP were investigated by a combination of chemical and instrumental analysis. Drug cytotoxicity was assessed using the MTT assay. The contents of HBsAg, HBeAg, intracellular and extracellular IFN-α level were measured using respective commercially available ELISA kit. The HBV DNA expression was evaluated by real-time quantitative polymerase chain reaction (PCR) and the relevant proteins involved in TFN/JAK/STAT signaling pathways were examined by western blot assay. RESULTS: MTT assay showed that RIP had no toxicity on HepG2.2.15 cell line below the concentration 400 μg/ml at Day 3, 6 and 9. Furthermore, RIP at the concentration of 50, 100 and 200 μg/ml significantly reduced extracellular and intracellular level of HBsAg, HBeAg and HBV DNA in HepG2.2.15 cells in a time and dose-dependent manner. Moreover, RIP also enhanced the production of IFN-α in HepG2.2.15 cell via activation of JAK/STAT signal pathway and induction of antiviral proteins, as evidenced by the increased protein expression of p-STAT-1, p-STAT-2, p-JAK1, p-TYK2, OAS1, and Mx in HepG2.2.15 cells. In addition, the over expression of SOCS-1 and SOCS-3 was significantly abolished under same conditions. CONCLUSIONS: These results suggested that the HBV inhibitory effect of RIP was possibly due to the activation of IFN-α-dependent JAK/STAT signal pathway and induction of the anti-HBV protein expression.
ETHNOPHARMACOLOGICAL RELEVANCE: Hepatitis B virus (HBV) infection frequently results in both acute and chronic hepatitis and poses serious threats to human health worldwide. Despite the availability of effective HBV vaccine and anti-HBV drugs, apparently inevitable side effects and resistance have limited its efficiency, thus prompt the search for new anti-HBV agents. The traditional Chinese medicine Radix Isatidis has been used for thousands of years, mainly for the treatment of viral and bacterial infection diseases including hepatitis. AIM OF THE STUDY: In this study, antiviral activities of a Radix Isatidis (Isatis indigotica Fortune) polysaccharide (RIP) were evaluated in vitro model using the HepG2.2.15 cell line and the underlying mechanism was elucidated with the aim of developing a novel anti-HBV therapeutic agent. MATERIALS AND METHODS: Structure features of the purified polysaccharideRIP were investigated by a combination of chemical and instrumental analysis. Drug cytotoxicity was assessed using the MTT assay. The contents of HBsAg, HBeAg, intracellular and extracellular IFN-α level were measured using respective commercially available ELISA kit. The HBV DNA expression was evaluated by real-time quantitative polymerase chain reaction (PCR) and the relevant proteins involved in TFN/JAK/STAT signaling pathways were examined by western blot assay. RESULTS:MTT assay showed that RIP had no toxicity on HepG2.2.15 cell line below the concentration 400 μg/ml at Day 3, 6 and 9. Furthermore, RIP at the concentration of 50, 100 and 200 μg/ml significantly reduced extracellular and intracellular level of HBsAg, HBeAg and HBV DNA in HepG2.2.15 cells in a time and dose-dependent manner. Moreover, RIP also enhanced the production of IFN-α in HepG2.2.15 cell via activation of JAK/STAT signal pathway and induction of antiviral proteins, as evidenced by the increased protein expression of p-STAT-1, p-STAT-2, p-JAK1, p-TYK2, OAS1, and Mx in HepG2.2.15 cells. In addition, the over expression of SOCS-1 and SOCS-3 was significantly abolished under same conditions. CONCLUSIONS: These results suggested that the HBV inhibitory effect of RIP was possibly due to the activation of IFN-α-dependent JAK/STAT signal pathway and induction of the anti-HBV protein expression.
Authors: Jianru Liu; Yan Huang; Xinzhe Lou; Bei Liu; Wenyi Liu; Na An; Rong Wu; Xiangying Ouyang Journal: BMJ Open Date: 2021-11-02 Impact factor: 2.692