The Effect of Streptococcus uberis Infection on Amino Acid Metabolism in Mammary Epithelial Cells
Metabolism is the cornerstone of all physiological activities,and amino acid metabo-lism,as one of the three major substance metabolisms,is involved in a variety of physiological activities of the host organism.It is closely related to the function of host immune cells and has an irreplaceable role in the development of diseases.To investigate the effect of Streptococcus uberis(S.uberis)infection on amino acid metabolism in mammary epithelial cells,S.uberis and mammary epithelial cell line(EpH4-Ev)were used for quantitative analysis of amino acids of cells by targeted amino acid metabolomics techniques in this study.The key enzymes expression of key amino acid(arginine)metabolic pathway was also validated by RT-qPCR and Western blot.The results showed that the levels of glycine,threonine,ornithine,asparagine and gluta-mine were significantly up-regulated(P＜0.05),and the levels of glutamate,proline,arginine,serine,histidine,valine,isoleucine,alanine,tyrosine,tryptophan,lysine and aspartate were significantly down-regulated(P＜0.05)in S.uberis-infected cells,with arginine content was most significantly changed(P＜0.000 1,VIP＞1.2).Two glycogenic amino acids,asparagine and glutamine,were significantly increased(P＜0.05),two ketogenic amino acids,lysine and leucine,were significantly decreased(P＜0.05).Further exploration of the arginine catabolic pathway revealed that S.uberis infection of mammary epithelial cells significantly upregulated the mRNA and protein expression levels of NOS2,a key enzyme of arginine metabolism(P＜0.05),and significantly downregulated the expression level of ARG1(P＜0.05).Compared with the control group,the expression of pro-inflammatory factors IL-1β and TNF-α in the cells chal-lenged with S.uberis were significantly up-regulated(P＜0.05).In summary,S.uberis infec-tion caused disruption of amino acid metabolism in mammary epithelial cells,accelerated intracel-lular arginine depletion,and initiated inflammatory responses.The results suggest that arginine has an important role in host resistance to S.uberis infection,and can be used as a breakthrough for early diagnosis and future control of mastitis in dairy cows.
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