Experimental Study In Vitro And Vivo On The Anti-Pancreatic Carcer Of DC Vaccine Plused With HSP70-polypeptide Complexes Purified From Mouse Pancreatic Carcinoma Cells

Objective:To establish a new method of purifying heat shock protein70-polypeptide complexes(HSP70-PCs) from pancreatic cancer tumor. Research on the use of HSP70-PCs to build dendritic cell(DC) polypeptide vaccine, to observe the effect of HSP70-PCs to the proliferation and activation of the CTL. And then proceed immunotherapy to the tumor-bearing mice,to observe the tumor size and survival of mice.Methods:HSP70-PCs from lumps of tumor-bearing mouse innoculated with pancreas cancer cells(MPC83) were purified successively by ConA-Sepharose affinity chromatography and ADP-Agarose affinity chromatography.The protein of HSP70-PCs was quantified by SDS-PAGE and Bradford assay,to calculate the rate of the proteins derived from the above method.Bone marrow-derived DC plused with HSP70-PCs were used to immunize Kunming mice.The response of cytotoxic T lymphocyte(CTL) was detected by MTT method,to analyze the effect of DC sensitized by HSP70-PCs on proliferation and activation of CTL.MPC83 tumor-bearing model of mice was established,which were treated by using dendritic cell pulsed with HSP70-PCs vaccine.To observe the inhibition of dendritic cell tumor vaccine HSP70-PCs to the growth of tumor and the effect of survival to the tumor-bearing mice.Results:A stained protein with a molecular weight of about 70KD was confirmed by SDS-PAGE to be HSP70-PCs,an average of 1g wet weight of tumor tissue obtained 79.95μg HSP70-PCs;HSP70-PCs at 10-12μg/mL could be achieved the strongest effect to stimulate the dendritic cell,DC modified with HSP70-PCs could enhance the capacity of proliferation and activation for T cell,DCs plused with HSP70-PCs could induce the specific cytotoxic activity of CTL prominently. Dendritic cell vaccine pulsed with HSP70-PCs could significantly inhibit tumor growth and prolong survival time of tumor-bearing mice.Conclusions:The high-purity,retention of tumor antigen information HSP70-PCs of mice pancreatic carcinoma cells can be separated and purified by using hypobaric affinity chromatography rapidly and conveniently.T cells stimulated by DC modified with HSP70-PCs show more specific cytotoxic activity to MPC83.DC plused with HSP70-PCs have an obvious therapeutic effect on the established pancreatic carcinoma-bearing mice.