Effects of Rac1 on the biological behaviors of RAW264.7 cells

PENG Jingjing; QI Chen; ZHANG Danyi; SU Zhaoliang; WANG Shengjun; XU Huaxi

doi:10.13431/j.cnki.immunol.j.20170099

Abstract：This study was performed to analyze the expression level of Rac1 in RAW264.7 cells,and discuss the effect of Rac1 vector and specific siRNA on the biological activity of RAW264.7 cells.The reverse transcription method was used to clone Rac1 cDNA and construct Rac1 expression vector,and the protein expression level of Rac1 was detected by Western blotting.Reverse transcription-quantitative polymerase chain reaction (RT-qPCR) was used to determinate Rac1 mRNA,while flow cytometry analysis was used to detect the changes of CD80,CD86 and MHC Ⅱ expression,and cell migration ability was observed by transwell method.In addition,ELISA was used to detect the levels of cytokines in supernatants.Data showed that Rac1 vector was obtained,the expression of Rac1 mRNA/protein were significantly up-regulated in Rac 1-transfected RAW264.7 cells,and the ability of migration was enhanced.But there were no significant changes in the expression of CD80,CD86 and MHC Ⅱ in Rac1-transfected RAW264.7 cells,however,these three kinds of membrane molecules could be up-regulated by Rac 1 inhibitors or specific siRNA.Moreover,the IL-1 [β level in supernatants of Rac 1-transfected RAW264.7 cells was lower than that of other groups,but there were no difference in the levels of IL-6,IL-33 or TNF-α.Taken together,the positive regulation of Rac1 may contribute to the migration of RAW264.7 cells and inhibit the expression of IL-1β,but exert no significant effect on antigen presentation.