The Study On FOXO3a In The Proliferation And Apoptosis Of Gastrointestinal Stromal Tumor

Objective: Gastrointestinal stromal tumor(GIST)is the most common mesenchymal tumor in the digestive tract.Forkhead box O3a(FOXO3a)plays an important role in the process of cell proliferation,apoptosis,stress,differentiation and so on.This research investigated the expression of FOXO3 a in the GIST tissues and studied the effect of FOXO3 a on the cell proliferation and apoptosis in GIST-T1 cells in vitro in order to provide new clues for the treatment of GIST.Methods: The expression of FOXO3 a protein in 32 cases of paraffin-embedded GIST and its normal tissues was detected by immunohistochemistry,and the correlation with GIST was discussed.The inclusion criteria were the first GIST surgical treatment in our hospital in 2010 and the targeted therapy,radiotherapy and chemotherapy were not performed before.2.The FOXO3 a plasmid with FOXO3 a overexpression was contructed by overlap extension PCR and enzyme-digested and enzyme-linked.3.The constructed plasmid was transfected into GIST-T1 cells with the electroporation method.The cells were observed with fluorescence microscopy after 24-48 h and Western blot was used to evaluate the transfection effect.4.The effect of FOXO3 a on the proliferation of GIST-T1 cells was observed by CCK-8 method.5.Cell cycle and late cell apoptosis were detected by flow cytometry(PI).6.The effect of FOXO3 a on apoptosis and cycle-related proteins was observed by Western blot.Results:1.The expression of FOXO3 a in GIST tissues was lower than that in normal ones(P <0.05).2.The overexpression of FOXO3 a plasmids was successfully constructed and verified by double digestion and sequencing.3.With GIST-T1 cell number 2×106,plasmid concentration 1μg /μl at least,20μg at a voltage of 570 V for30ms in a volume of 120μl or the cell number 1.5×105,plasmid concentration 1μg /μl at least,3μg at a voltage of 360 V for 30 ms in a volume of 20μl,the expression level of FOXO3 a in the overexpression group was higher than that of the control groups,and the transfection efficiency is over 70%.4.The proliferation ability of the overexpression one was decreased(P <0.05).5.The GIST-T1 cells with FOXO3 a overexpression showed more late apoptosis than the control group(P <0.05)and the other cell cycle stages were not different(P > 0.05).6.The expressions of the Bim,p27kip1 and cleaved PARP were higher and the expression of the cyclin D1 was lower in the overexpression group.Conclusion: 1.The expression level of FOXO3 a in GIST is lower.2.FOXO3 a can inhibit the proliferation of GIST-T1 cells,which also promotes their apoptosis.3.The FOXO3 a may be used as a tumor suppressor in GIST.