Effects Of Genetic Engineering Peptides On MRNA Expression Of Cytokines And Toll-like Receptors In Mammary Epithelial Cells Invaded By Escherichia Coli

This experiment studied on cell culture of Holstein cows'mammary epithelial cells.The bovine mammary epithelial cells were invaded by Escherichia Coli to establelish the model of clinical medicine in vitro. Genetic engineering peptides﹙PH-SA﹚was used to cure the mammary epithelial cells.The treatment effects of drug were determined,which base on the change of Toll-like receptors and inflammation factors in the experiment.At the same time, provide theory basis for using the new antibiotics treat and prevent mastitis.The experiments have cultured the third generation mammary epithelial cells. The mammary epithelial cells were invaded by 107CFU/mL Escherichia Coli﹙O111:K58﹚, after 2 hour, PH-SA was used to treat the mammary epithelial cells. The mammary epithelial cells were collected at 6h, 12h and 24h. The mRNA expression of TLR-4, TLR-2, IL-lβand IL-8 were determined by Real-time PCR method.PH-SA was a fusion protein by linking the C-terminal of colicin Ia, with the the pheromone AgrD1 of Staphylocoecos aureus, binded to the bacterial cytomembrane in target cell, and can kill bacteria through forming ion channel. The results of RT-PCR showed that IL-lβmRNA expression level increased significantly﹙P<0.05﹚at 6 h, and the gene IL-8 mRNA express significantly at 24 h﹙P<0.05﹚, but the TLR-2 mRNA expression hasn't change in the experiment﹙P>0.05﹚.However, the TLR-4 mRNA expression level increased at 6 and 12 h﹙P<0.05﹚. With the extension of sterilization time, find that the most of E.coli were dead. PH-SA can depress Toll-like receptors and decrease the mRNA expression levels of inflammatory factor. In contrast, the gentamicin was failed to decrease the mRNA expression levels of IL-8 and TLR-4. In conclusion, the cure effect of PH-SA was better than gentamicin.