Effect of Plasma-derived Exosomes on LPS-induced MAC-T Cell Inflammation in Dairy Cows with Mastitis
[Objective]Exosomes are key factors in cellular communication and establishment of tissue microenvironment,and they have a regulatory role in inflammatory processes.This study was aimed to investigate the role of plasma-derived exosomes from dairy cows in the regulation of lipopolysaccharide(LPS)-induced inflammation in mammary epithelial cells(MAC-T)of dairy cows.[Method]In this study,venous blood was collected from healthy and mastitis-affected dairy cows,and plasma was obtained after centrifugation,and plasma-derived exosomes were isolated by ultracentrifugation.The expression of pro-inflammatory factors after incubation of plasma-derived exosomes from dairy cows was detected by Real-time quantitative PCR.Western blotting was used to verify the expression of NF-κB pathway proteins TLR4,MyD88 and TNF-α after incubation of plasma-derived exosomes from dairy cows.The effects of plasma-derived exosomes from dairy cows on the proliferation as well as apoptosis of MAC-T cells were verified by CCK8 and Hoechst staining.[Result]Real-time quantitative PCR results showed that plasma-derived exosomes from dairy cows with mastitis significantly down-regulated the expression of pro-inflammatory factors IL-6,IL-23,IL-1α,NFRK,MyD88,TNF-α and IL-1β genes mRNA compared with LPS group(P＜0.05).Plasma-derived exosomes from healthy cows significantly inhibited mRNA expression of the IL-1β gene(P＜0.05).Western blotting results showed that the plasma-derived exosomes from dairy cows with mastitis significantly decreased the expression of the NF-κB pathway proteins TLR4,MyD88 and TNF-α(P＜0.05).NF-κB pathway protein expression in plasma-derived exosomes from healthy cows was not significantly different from that of the LPS group(P＞0.05).CCK8 and Hoechst staining results showed that plasma-derived exosomes from dairy cows with mastitis could promote cell proliferation and alleviate apoptosis compared with LPS group.Plasma-derived exosomes from healthy cows restored some cell viability.[Conclusion]The results indicated that plasma-derived exosomes from dairy cows with mastitis could alleviate LPS-induced inflammation of MAC-T cells through NF-κB pathway.This study would provide a theoretical basis for the anti-inflammatory role played by plasma-derived exosomes in dairy cow mastitis,and at the same time laid the foundation for the clinical application of plasma-derived exosomes.
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