人β防御素-2稳定表达细胞株的建立及其表达产物对耐药大肠埃希菌抗菌活性研究

Antimicrobial activities against E. coli by the stable cell line transfected with recombinant hBD-2 gene eukaryotic expression plasmid

目的建立稳定表达人β防御素2(hBD2)的COS-7细胞株,并检测其表达产物对耐药大肠埃希菌的抗菌活性。方法将重组hBD2真核表达载体pCMV-hBD2通过脂质体转染COS-7细胞,筛选稳定表达hBD2单克隆细胞株;经RT-PCR和Western blot检测稳定转染后目的基因在转录水平和蛋白水平的表达;用微量稀释法检测稳定转染后细胞培养上清液对耐药大肠埃希菌的抗菌作用。结果筛选出稳定表达hBD2的COS-7细胞株,检测到目的基因在转录水平和蛋白水平均有表达,且其表达产物使耐药大肠埃希菌存活率降至15%。结论建立了稳定表达hBD2的COS-7细胞株,其表达产物对耐药大肠埃希菌具有一定的抗菌活性。

In recent years, antibiotic resistance is one of the world＇s most pressing public health problems.This study was designed to detect the of the human beta defensin 2（hBD2）. Firstly, plasmid vector pCMV-hBD2containing recombinant hBD2 gene was constructed and transfected to COS-7 cells, and the cell lines stably expressing hBD2 clone was selected. Secondly, RT-PCR and Western blot were applied to identify mRNA and protein levels of hBD2. Furthermore, the supernatant solution of the stable cells was mixed with E. coli solution to detected antimicrobial activities. Data showed that hBD2 was expressed successfully. And hBD2 expression product could decrease the survival rate of E. coli to 15%. In conclusion, this study construct stable cell lines of expressing hBD2, and the hBD2 expression products demonstrate the antimicrobial activity against drug-resistant E. coli.