MEF2D Promotes The Proliferation,migration And Infiltration Of Hepatoma Cells PLC/PRF5 And SMMC7721 By Negatively Regulating MIG6 Expression

Objective The study showed that myocyte enhancer factor 2D(MEF2D)could promote proliferation,migration and infiltration of hepatoma cells PLC/PRF5 and SMMC7721 by negatively regulating the expression of mitogen-induced gene 6(MIG6).Methods The mRNA levels of MEF2D and MIG6 were detected by real time fluorescence quantitative PCR in hcclm3,SMMC7721,PLC/PRF5,Hep G2,7703,Huh7,Bel-7402 cell lines and we drew a linear correlation diagram with“MEF2D mRNA expression”on the horizontal axis and“MIG6 mRNA expression”on the vertical one in these hepatoma cell lines.The expression of MEF2D and MIG6 were down-regulated by small interfering RNA(si RNA),then the proliferation,migration and infiltration were observed by 7Sea-Cell Counting Kit(7Sea-CCK),wound healing assay and Transwell in PLC/PRF5 cell lines.The expression of MEF2D and MIG6 were detected by western blot after MEF2D down-regulated by si RNA in PLC/PRF5 cell lines or up-regulated by plasmid in SMMC7721 cell lines.The experimental were divided into seven groups:negative group(NC),si MIG6 group,si MEF2D group,si2M(Both MEF2D and MIG6 were down-regulated)group,blank group,empty vector plasmid group and MEF2D group.Results We found that the level of MEF2D mRNA was negatively correlated with the expression of MIG6 mRNA in hepatoma cell lines(r =-0.78).Next,the results analyzed by 7Sea-CCK showed that the proliferation ability was significantly enhanced at72 h after si MIG6 treatment compared with NC group in PLC/PRF5 cell lines(OD=3.73±0.18,t=3.84,P=0.02),while there was no change at 24 h and 48 h.However,the proliferation ability was significantly suppressed at 72 h under MEF2D knockdown condition in PLC/PRF5 cell lines compared with NC group(OD=1.79±0.22,t=3.95,P=0.02),but the proliferation ability was significantly increased at 72 h under MEF2D and MIG6 both knockdown condition compared si MEF2D group(OD=2.99±0.03,t=4.83,P<0.01).Furthermore,wound healing assay revealed that the cell mobility in si MIG6 group significantly was increased [cell mobility=(51±4)%,t=3.22,P<0.05] at 48 hcompared with negative control and the cell mobility in si MEF2D group was significantly reduced [cell mobility=(19±3),t=7.70,P<0.01] in PLC/PRF5 cell lines,but the cell mobility in si2 M group was significantly increased at 48 h compared with si MEF2D group[cell mobility=(46±3)%,t=6.99,P<0.01].Next,transwell revealed that the ability of infiltration in si MIG6 group signification was increased(t=4.02,P<0.05)compared with negative control and the ability of infiltration in si MIG6 group signification was reduced(t=3.57,P<0.05)in PLC/PRC5 cell lines,but the ability of infiltration in si2 M group was significantly increased compared with si MEF2D group(t=5.25,P<0.01).The result of western blot showed that,compared with NC group,the expression of MIG6 was significantly increased under MEF2D si RNA treatment in PLC/PRF5 cell lines(t=7.86,P<0.001),while its level was significantly decreased by the elevated level of MEF2D in SMMC7721 cell lines(t=4.61,P=0.004).Conclusion The expression of MEF2D was negatively correlated with the level of MIG6 PLC/PRF5 and SMMC7721 cell lines.Thus,MEF2D promotes the proliferation,migration and infiltration of PLC/PRF5 and SMMC7721 cell lines possibly through negatively regulating MIG6 expression.