The Effects Of Arg1 On Proliferation, Apoptosis And The Mapk-sigal Pathway Of 293t Cell

Objective To study the anti-arsenic effects of ARG1 gene by obseving the effects of ARG1 gene on the proliferation, apoptosis and MAPK-sigal path way of the 293T cells.Methods (1) The plasmid was indentified with PCR,the incision enzyme of EcoRland BamHl, and sequenced; (2) The plasmids of pcDNA3.1-IE and pcDNA-IE-ARG1 was transfected in 293T cells by the Lipofectamine2000, then the transfectio of ARG1 gene was observed by the Immunofluorescence and Real Time PCR;(3) The effcts of ARG1 on proliferation of cells were observed by MTT assay at the 0,1,2,4,8, 16,32μmol/L NaAsO2 incubated with arsenic for 48 h respectively,; (4) The 0,1,4,8μmol/L arsenic were chosen to incubated with the cells for 48 h respectively, and then we detected the apoptotosis with Annexin V FITC/PI by the Flow Cytometry, and the expression of p-JNK and p-ERK by Western Blot respectively.Results (1)The length and sequence of ARG1 gene was correct; (2) The transfected efficiency of the ARG1 gene was about 75-80% and the expression of ARG1 gene, expressed green fluorescence protein, of 293T cels was on plasmalemma,but was not in the pcDNA3.1-IEI; (3) The expression of ARG1 in experimental group was about 10 times than that in the control group (p<0.05); (4) The inhibitory rate of proliferation of the experimental group was apparently lower than the control group among 1-8μmol/L arsenic (p<0.05),but when the concentrations of arsenic was high than 8μmol/L, the inhibitory rate of proliferation of cells were not observed to be different among the tree goups (p>0.05); (5) The apoptotic rate of experimental group cells was observed apparently lower than that of the control group among 0-8μmol/L NaAsO2 (p<0.05); (6) Along with the NaAsO2 concentrations increasing, the expression of p-JNK of 239T cells increased before and after transfection, but it was lower than that of the control group cells (p<0.05). The expression of p-ERK of the experimental group cells was increasing among 0-4μmol/L, but it was lower at 8μmol/L and gradually degraded along with the NaAsO2 concentrations increasing.Conclusion The arsenic-resesistance of ARG1 gene acted in≤8μmol/L arsenic and the MAPK-signal pathway.maybe involved in the arsenic-resesistance of ARg1 ARG1 gene.