Study On The Immunogenic Apoptosis Of CT26Colorectal Cancer Cells Induced By Clostridium Difficile Toxin A

Apoptosis was originally thoμght to be immunologically silent or tolerogenic, but recentresults suggest that tumor cells undergoing apoptosis in a stressful and/or inflammatorymicroenvironment could be highly immunogenic. Inducing immunogenic apoptosis of tumorcells may open new stratrigies for cancer disease therapy. Clostridium difficile toxin A (TcdA),one of the main pathogenic factors which were released by Clostridium difficile, can eliciteapoptotic death of wide types of cells, and process a potent proinflammatory activities.Because of its potent cytotoxic and proinflammatory activities, we investigated whether TcdAcan induce immunogenic apoptosis of tumor cells.We investigate the anti-tumor activity of TcdA using CT26colorectal cancer cell model.Firstly, we explore whether TcdA could induce apoptosis and promote the changes of heatshock proteins in respect to the expression or location in cells. Morphological changes wereobserved by microscope. Apoptosis and caspase activation were analyzed by flow cytometryor western assay. CRT exposure on cell surface and HSP70secretion were examined usingflow cytometry and western assay respectively. Secondly, we investigate whether TcdAintoxicated CT26cells can provoke the immunostimulatory effects of DCs by detecting DCphagocytosis, DC mature and T cell activation stimulated by tumor-loaded DCs in vitro. Wenext explore the induction of anti-tumor activity in Balb/C mice by TcdA treated tumor cells.Moreover, we investigate whether the anti-tumor activity is mediated by T-cell response,specific and long-lasting.As our data,(1) TcdA treated CT26cells undergoing apoptosis and caspase activity wasinvolved in this process;(2) CRT cell surface translocation and HSP70secretion of TcdAintoxicated CT26cells may indicate the immunogenic properties of apoptotic CT26cells;(3)TcdA treated CT26cells can stimulate the immunostimulatory effects of DCs;(4)immunization with TcdA-treated CT26cells can stimulate T cell response and pretend micefrom tumor challenge, and the anti-tumor immunity is specific and long lasting.Conclusion: Clostridium difficile toxin A can induce apoptotic death of CT26colorectalcancer cells and the apoptotic cells stimulate potent anti-tumor immunity.This study is the first one to confirm that TcdA can induce immunogenic apoptosis of tumor cells, indicating the important roles of TcdA in the application of tumor therapy.