Radiofrequency Hyperthermia Promotes The Therapeutic Effects On Pancreatic Cancer Of Mesenchymal Stem Cell-mediated Heat Shock Protein Promoter-controlled HSV-TK

Research Objectives:To investigate radiofrequency hyperthermia(RFH)promotes the therapeutic effects on pancreatic cancer of heat shock protein pomoter-controlled HSV-TK gene(PHSP-TK),and the therapeutic effects on pancreatic cancer of rat bone marrow mesenchymal stem cell-mediated(rBM-MSC)PHSP-TK.To further lay the foundation for exploring RFH combined MSC-mediated new gene therapy to treat pancreatic cancer.Research Methods:A lentiviral vector packaged with PHSP-TK gene transfected the follow cells:human pancreatic cancer cell lines(CFPAC-1 and BXPC-3),and rBM-MSC.Then we got CFPAC-1-TK,BXPC-3-TK and rBM-MSC-TK.Inverted fluorescence microscope and flow cytometry analysis were used to examinated PHSP-TK gene.The experiment to investigate RFH promotes herapeutic effects on pancreatic cancer of PHSP-TK gene were performed as following:①pancreatic cancer group(Control),②pancreatic cancer therapy of RFH group(RFH),③25%pancreatic cancer transduced PHSP-TK gene group(25%PHSP-TK),(4)25%pancreatic cancer transduced PHSP-TK combining RFH group(25%PHSP-TK +RFH),⑤50%pancreatic cancer transduced PHSP-TK gene group(50%PHSP-TK),⑥50%pancreatic cancer transduced PHSP-TK combining RFH group(50%PHSP-TK +RFH group),⑦75%pancreatic cancer transduced PHSP-TK gene group(75%PHSP-TK),⑧75%pancreatic cancer transduced PHSP-TK combining RFH group(75%PHSP-TK +RFH group).The experiment to investigate the tumoricidal effect on pancreatic cancer of rBM-MSC-mediated PHSP-TK gene were performed as following:①Pancreatic cancer group(Control),②pancreatic cancer therapy of RFH group(RFH),③pancreatic cancer cells and rBM-MSC cocultured at the ratio of 1:1 group(Pancreatic cancer +rBM-MSC),④pancreatic cancer cells and rBM-MSC-TK cocultured at the ratio of 1:1 group(pancreatic cancer +rBM-MSC-TK),⑤pancreatic cancer cells and BM-rMSC-TK cocultured at the rato of 1:1 combining RFH group(pancreatic cancer+rBM-MSC-TK+RFH).Cell proliferation and apoptosis were quantified to evaluated the tumoricidal effect of PHSP-TK gene and rBM-MSC-mediated PHSP-TK gene on pancreatic cancer.Results:The results of inverted fluorescence microscope and flow cytometry analysis indicated that the PHSP-TK gene expressed in quantity in human pancreatic cancer cell lines and rBM-MSC.And the expression ratio of PHSP-TK gene were higher after RFH than the cells without RFH(P<0.001).Pancreatic cancer cell proliferation and apoptosis results:CCK8 revealed cell survival ratio of PHSP-TK+RFH group were lower than PHSP-TK and RFH group(P<0.05).Cell apoptosis results further revealed that apoptotic index of PHSP-TK+RFH group were higher than PHSP-TK group and RFH group(P<0.05).The experiment results of rBM-MSC:②rBM-MSC which were spindle-shaped were successfully isolated,and the positive rate of surface antigen CD29,CD54,CD90 and CD45 were 96.59%、94.91%、99.43%and 3.21%respectively,And rBM-MSC had the ability of adiogenesis and osteogenic differentiation.②CCK8 revealed that cell survival ratio of pancreatic cancer + rBM-MSC-TK group was lower than pancreatic cancer + rBM-MSC group,control group(P<0.05).And cell survival ratio of pancreatic cancer + rBM-MSC-TK+RFH group was lower than pancreatic cancer + rBM-MSC-TK group,control group and RFH group(P<0.05).③Cell apoptosis results revealed apoptotic index of pancreatic cancer + rBM-MSC-TK+ RFH group was higher than the pancreatic cancer+ rBM-MSC-TK group(P<0.05).Conclusion:Our study indicated that PHSP-TK gene could transfect pancreatic cancer cells and rBM-MSC,and RFH promoted tumoricidal effect of the PHSP-TK on pancreatic cancer.To further lay the foundation for exploring RFH combined MSC targeted new gene therapy to treat pancreatic cancer.