1.Effect Of Bicyclol On JAK/STAT Signal Pathway In Con A-intoxicated And HBV Transgenic Mice 2.Protective Effect And Related Mechanisms Of Bicyclol On Nephrotoxicity And Hepatotoxicity Induced By Cisplatin In Mice

Autoimmune,viral and drug-induced liver injuries are caused by multiple pathological factors.Although the mechanisms of above liver injuries remain unclear, oxidative stress and inflammation are considered as two important factors in the liver pathological process.Janus Kinase(JAK)/signal transducer and activator of transcription(STAT) signal pathway were believed to be involved in immune and inflammatory pathogenesis.In addition,oxidative stress can also activate mitogen-activated protein kinase(MAPKs) signal pathway in different liver injured models.Bicyclol(4,4'-dimethoxy-5,6,5',6'-Bis(dimethylene-dioxy)-2-hydroxymethyl-2 '-methoxy carbonyl biphenyl:Fig.1),a novel synthetic anti-hepatitis drug,has been widely used in patients with chronic viral hepatitis B(HBV) in China.Previous studies have shown that bicyclol can protected damaged cell membrane and nuclear deoxyribonucleic acid(DNA) induced by several chemical toxins through the antioxidation and reduction of apoptosis.The purpose of present study was to investigate the involvement of JAK/STAT signal pathway in Concanavalin A(Con A) induced liver injury/HBV infection,and the influence of bicyclol on JAK/STAT signal pathway in mice.In addition,the protective effects and related mechanism of bicyclol on nephrotoxicity and hepatotoxicity induced by cisplatin were also observed in normal and H22 tumor bearing mice.Part one:Effect of bicyclol on JAK/STAT signal pathway in Con A-intoxicated and HBV transgenic mice1.Hepatoprotective mechanism of bicyclol on liver injury induced by Con A in mice:role of TNF-α/NF-κB and IFN-γ/STAT1 signal pathwayCon A,as a T cell mitogen,is known to induce T cell-dependent liver injury in mice.Tumor necrosis factor-α(TNF-α) and Interferon gamma(IFN-γ) released from activated T cells are considered to play critical roles in liver pathological injury. Besides direct hepatotoxic,one additional pathway by IFN-γcontribution to liver damage is through the activation of JAK/STAT signal transduction.The aim of present study is to investigate the effect of bicyclol on TNF-α/nuclear factor(NF)-κB and IFN-γ/STAT1 signal pathway in Con A-intoxicated mice. Mice were pretreated with bicyclol(200mg/kg,po) for consecutive 4 days before injection with Con A(25mg/kg).The result showed that liver histopathological changes,elevated serum level of alanine aminotransferase(ALT)/aspartate aminotransferase(AST) and nitric oxide(NO) were markedly inhibited by bicyclol in Con A intoxicated mice.The increasing of inflammatory cytokines including TNF-αand IFN-γwere suppressed after bicyclol treatment.Bicyclol also inhibited the over expression of liver iNOS and activation of STAT1,while suppressors of cytokine signal(SOCS1) was further enhanced.Moreover,the expression of proapoptotic protein interferon regulation factor(IRF-1) induced by Con A was suppressed by bicyclol.The expression of liver NF-κB was elevated after Con A injection and bicyclol treatment significantly down-regulate the over expression of NF-κB. Intercellular adhesion molecule(ICAM-1) and lymphocyte function-associated antigen-1(LFA-1),two of adhesion molecule in hepatocytes and non-parenchymal cells were enhanced after Con A intoxication.Bicyclol was found to reduce the immunoreactivity of ICAM-1 and LFA-1 significantly.Microarray analysis showed that 287 genes exhibited differential expression in bicyclol group,in which 121 genes were up-regulated and 161 genes were down-regulated comparing with Con A intoxicated mice.The differential gene expression after bicyclol treatment was involved in the biotransformation,protein synthesis,degradation and circadian rhythm,proliferation and signal transduction.In conclusion,hepatoprotective action of bicyclol on Con A induced liver injury was partially due to the down regulation of imflammatory cytokines including TNF-αand IFN-γ.The regulatory effect of bicyclol on the IFN-γ/STAT1 signal pathway plays a key role in its protection against Con A induced liver injury.2.Effects of bicyclol on HBV DNA replications and IFN-γ/STAT1 signal pathway in HBV transgenic miceIt has been reported that Anti-HBV drugs inhibit HBV DNA polymerase directly, and induce IFN synthesis/activate IFN signal pathway.The latter mechanism was related to the activation and translocation of IFN-γ/STAT1.The aim of the study is to investigate the effect of bicyclol on HBV DNA replication and relevance between hepatoprotection and modulation on IFN-γ/STAT1 signal pathway in HBV DNA transgenic mice and HepG2.2.1.5 cells.HBV DNA transgenic mice received bicyclol(800 mg/kg,po,qd×90).The result showed that there was no change on the IFN-γ/STAT1 signal pathway in transgenic mice,comparing with control animals.Bicyclol can inhibit the replication of HBV DNA in both in vivo and in vitro models,however,had no effect on liver IFN-γ/STAT1 signal pathway in transgenic mice.Part two:Protective effects and related mechanisms of bicyclol on hepatotoxicity and nephrotoxicity induced by cisplatin in miceCisplatin(cis-diaminedichloroplatinum),a platinum-containing anticancer drug,is one of the most commonly used cytotoxic agents in the treatment of a variety of solid malignant tumors in clinical.In spite of its clinical usefulness,there are many occasions in which it is difficult to continue the administration of the drug due to its nephrotoxicity and hepatotoxicity.The present study is to investigate the protective effects of bicyclol against cisplatin-induced nephrotoxicity,hepatotoxicity and the possible mechanisms in mice.Bicyclol showed significant renal protection as evidenced by the decrease of elevated serum creatine(CRE) and blood urea nitrogen(BUN),and improvement of histopathological injury induced by cisplatin.The liver damage indicated by the histopathologic changes,elevated serum levels of ALT and AST were also observed after cisplatin treatment.Bicyclol not only protect the nephrotoxicity,but also significantly reduced the hepatotoxicity induced by cisplatin in certain extends.The formation of liver and kidney malondialdehyde(MDA) with a concomitant reduction of reduced glutathione(GSH) were also inhibited by bicyclol,while the decreasing of liver and kidney antioxidative enzymes including superoxide dismutase(SOD), catalase(CAT) and glutathione peroxidase(GSH-px) were all improved by bicyclol in cisplatin treated mice,respectively.Bicyclol also inhibited the increases of liver, kidney and serum NO induced by cisplatin.In addition,the over expressions of liver and kidney inducible nitric oxide synthase(iNOS) and nitrotyrosine(3-NT) were suppressed by bicyclol treatment.Cisplatin-induced phosphorylation of liver extracellular regulated protein kinases(ERK) 1/2,Ets-like protein 1(Elk-1),p38 MAPK and p53 were abrogated by pretreatment with bicyclol.The administration of bicyclol had no affect on the antitumor capacity of cisplatin in mice bearing hepatocarcinoma 22(H22).The results demonstrate that bicyclol has a remarkable protective effect against cisplatin-induced nephrotoxicity and hepatotoxicity in mice.The hepatoprotective effect of bicyclol was mainly due to the inhibition of MAPKs phosphorylation and reversion of imbalance between oxidant and antioxidant.