Impact of CD74 Gene Transfection on Characteristics of Human Umbilical Vein Endothelial Cell ECV304

Objective To explore the transfecting conditions of the cluster of differentiation(CD74) gene in human umbilical vein endothelial cells and impact of CD74 on cell characteristics and functions,and lay the basis for biotherapy based on CD74 expression modulation. Methods PCMV-CD74 enhanced green fluorescent protein(EGFP) plasmid was transformed into E.coli,extracted,enzyme-cut and sequenced.The EGFP fluorescent plasmid was transfected to ECV304 and the transfection efficiency was detected.The expressions of CD74 and HLA-DR before and after transfecting ECV304 were detected by immunohistochemical and Western blot.The expression of CD74 mRNA in transfected ECV304,and the expressions of function-associated genes including human leukocyte antigens A(HLA-A),human leukocyte antigens DR(HLA-DR) and interferon-γ receptor(IFNGR) before and after transfecting ECV304 was detected by real-time reverse transcriptase polymerase chain reaction(RT-PCR).Data was analysed by real-time RT-PCR software. Results The △Ct of HLA-A was 9.4881 and 12.1097 before and after CD74 transfection respectively,and 2-△△Ct was 6.1543,which indicated significant difference in the expression of HLA-A before and after CD74 transfection.The △Ct before and after CD74 transfection,and 2-△△Ct of IFNGR was 4.9828,5.5228 and 1.4539,and of HLA-DR was 14.1098,14.7924 and 1.3036,which implied no significant difference before and after transfection. Conclusion Human umbilical vein endothelial cell ECV304 can express HLA-A,HLA-DR,IFNGR and CD74 mRNA.PCMV-CD74 transfection can increase the expressions of CD74 gene,membrane molecules and HLA-A gene,but can't increase the expressions of HLA-DR and IFNGR.