Epigenetic Regulation And Functional Mechanisms Of Histone Deacetylasc 1 In Gastric Cancer

Background:Gastric Cancer (GC) is one of the four most common malignant tumors that threatens people’s health seriously. There are many epigenetic changes while GC is developing. Epigenetic changes could lead to abnormity of signal pathways which influence growth, proliferation, differentiation, apoptosis of cancer cells as well as their reactions to therapy. Histone Deacetylase 1(HDAC1) plays a critical role in epigenetic regulation networks and it is up-regulated in various malignant cancers. However how HDAC1 influences gastric cancer cells’apoptosis and their reactions to chemotherapy is worth deeper investigation.Objective:First part:Researches have already shown that doxorubicin and HDAC inhibitors (HDACi) have synergistic action, however the molecular mechanism of their interaction remains unclear. The purpose of our research is to clarify how HDAC1 is epigenetically regulated by doxorubicin in gastric cancer and how HDAC1 plays its role helping doxorubicin to treat gastric cancer. Second part:Researches have already shown that HDACI is up-regulated in gastric cancer, but the molecular mechanism of how HDAC1 acts in the development progress of gastric cancer is required to be understood. The aim of this part research is to discover the molecular mechanism of how HDAC1 controls gastric cancer’s apoptosis and the epigenetic mechanism of how HDACi exert their roles in gastric cancer therapy. We decide to explain the epigenetic molecular mechanism, basically in two ways, of how HDAC1 promotes development and drug-resistance in gastric cancer:how HDAC1 influences the reaction of gastric cancer to chemotherapy and how HDAC1 influences the spontaneous apoptosis of gastric cancer cells.Methods:First part:First western blot was used to analyse HDAC1 protein expression before and after doxorubicin treatment in the gastric cancer cells. Second the possible microRNAs which may target HDAC1 after doxorubicin treatment in gastric cancer was analysed by online software and RT-qPCR, and then verified the regulation of microRNAs to HDAC1 from functional and structural aspects. Third MTS analysis was used to detect the effect of HDAC1 on sensitivity to doxorubicin. Second part:First western blot and flowcytometry analysis were used to detect the caspase 2-dependent apoptosis signal pathway which was activated by HDACi TrichostatinA (TSA) in gastric cancer. Second RT-qPCR and western blot method were used to analyse expression of CRADD after TSA treatment. Third we detected the function of CRADD in TSA-induced apoptosis in gastric cancer. Further immunohistochemistry was used to analyse CRADD expression in gastric cancer tissues. Next we verified by colony formation analysis that CRADD inhibited the growth of gastric cancer cells. Finally chromatin immunoprecipitation was used to prove that HDAC1 directly regulated CRADD transcription in gastric cancer cells.Results:First part:The research reveals the epigenetic regulation mechanism of how HDACl was regulated by doxorubicin and its influence on reactions to doxorubicin in gastric cancer. We find that miR-520h expression is up-regulated by doxorubicin and targets to HDACl 3’UTR, and then specifically down-regulates the expression of HDACl protein. This will make chromatin loosen, promote DNA-doxorubicin interaction and DNA damage induced by doxorubicin, finally will kill gastric cancer cells. Second part:The research elaborates the epigenetic regulation mechanism and function of HDAC1 in controlling cell apoptosis signal pathway. We find that HDAC1 increases transcription of CRADD, activates caspase 2 and finally promotes caspase 2-dependent cell apoptosis.Conclusion:This paper explains the epigenetic regulation and function of HDAC1 in gastric cancer from different points of view, and further provides new epigenetic theory basics of HDACi in cancer therapy.