Effects Of Wt-p53,B7-1 And GM-CSF Genes Transfection On Proliferation, Apoptosis And Immunogenicity Of SH-SY5Y Cell Lines

Background and ObjectNeuroblastoma(NB)is a kind of malignant tumor which usually happened in children.The effect of conventional therapy for NB is unsatisfied.The occurrence and development of tumor involve many kinds of gene abnormality.Multi-gene therapy is more reasonable than single gene therapy.Two of the main cause of tumorigenesis are the mutation of anti-oncogene and immune escape.NB is a poorly immunogenic tumor and the function of the wild-type p53(wt-p53)gene in SH-SY5Y cells is inactivated.Hence,in the present study the effect of cotransduction of wt-p53,B7-1 and GM-CSF genes mediated by recombinant adenovirus on cell growth, apoptosis and immunogenicity of NB cell line SH-SY5Y was studied.MethodThe NB cell line SH-SY5Y were transfected with recombinant adenovirus mediating green fluorescent protein(Ad-GFP)separately at 50,100,200 multiplicity of infection(MOI).The suitable MOI was determined after the transfer efficiency was tested by flow cytometry(FCM).We transfected SH-SY5Y cells separately with recombinant adenovirus,recombinant adenovirus mediating human wild type p53 (wt-p53)gene and BB-102.We separately named them with SH/Ad,SH/p53 and SH/BB-102.48 hours after gene transfection into SH-SY5Y cells,we investigated the protein expression of p53,B7-1 and GM-CSF by Western blot,FCM and enzyme linked immunosorbent assay(ELASA)Cell number was counted and growth curves were drawed.Both cell cycle and apoptosis were tested by FCM.Lymphocyte proliferation and cytokine secretory was evaluated by mixed lymphocyte culturing.Data were collected and SPSS 12.0 was used in statistical analysis.ResultResults indicated that the transfer efficiency of the recombinant adenovirus to SH-SY5Y was high,and 100MOI was the suitable MOI.After transfection with BB-102,p53,B7-land GM -CSF genes were expressed.Growth of the cells were inhibited,apoptosis and cell cycle arrest was induced in the SH/Ad and SH/BB-102 groups.Proliferation of lymphocytes was stimulated and cytoldne increased vigorously in the SH/BB-102 group.Conclusionwt-p53,B7-1 and GM-CSF genes can be effectively expressed by SH-SY5Y cells transfected with recombinant adenovirus.Growth of the cells were inhibited,apoptosis and cell cycle arrest was induced,and immunogenicity was greatly enhanced.Our results indicate that clinical application of the in vitro BB-102-modifide tumor cell vaccine might be feasible and effective in clinical treatment of NB.