Effect of Staphylococcus aureus and Staphylococcus epidermidis on the Secretion of Inflammatory Factors by Neutrophils in Dairy Cows
[Objective]The purpose of this experiment was to investigate the effects of Staphylococcus aureus and Staphylococcus epidermidis on the activity of neutrophils(PMNs)and the secretion of inflammatory factors in peripheral blood of dairy cows.[Method]The morphology and cell viability of PMNs in peripheral blood of dairy cows cultured in vitro were identified by Giemsa and Trypan blue staining,and the optimal time of action of PMNs was determined.The peripheral blood PMNs infection model induced by Staphylococcus aureus and Staphylococcus epidermidis was established to determine the killing ability of peripheral blood PMNs to the two bacteria.PMNs activity was determined by CCK-8 method,and the levels of inflammatory factors tumor necrosis factor-α(TNF-α),interleukin-1β(IL-1β),IL-8 and IL-10 were detected by ELISA.[Result]The results showed that the purity of PMNs from peripheral blood of dairy cows isolated and cultured in vitro reached 99％,and the survival rate decreased gradually with the extension of culture time,and the survival rate reached more than 85％within 24 h.The logarithmic growth periods of Staphylococcus aureus and Staphylococcus epidermidis were 4-7 h and 4-8 h,respectively.When MOI was 10,the survival rate of Staphylococcus aureus at 2,4 and 6 h was extremely significantly higher than that of Staphylococcus epidermidis(P＜0.01).Compared with control group,the contents of TNF-α and IL-8 in PMNs infected by Staphylococcus aureus and Staphylococcus epidermidis were extremely significantly or significantly increased at different time points(P＜0.01 or P＜0.05).The content of IL-10 in cells was extremely significantly or significantly increased at 3,6,12 and 24 h after infection(P＜0.01 or P＜0.05).The content of IL-1β in cells was extremely significantly or significantly increased at 6,9,12 and 24 h after infection(P＜0.01 or P＜0.05).Compared with Staphylococcus aureus group,the content of TNF-α in Staphylococcus epidermidis group at 3 h of infection was extremely significantly decreased(P＜0.01),and the contents of IL-10 and IL-8 were extremely significantly or significantly increased(P＜0.01 or P＜0.05).After 6 h of infection,the contents of TNF-α and IL-1β in Staphylococcus epidermidis group were extremely significantly decreased(P＜0.01).The contents of TNF-α and IL-8 in cells were extremely significantly or significantly increased at 9 h after infection(P＜0.01 or P＜0.05).[Conclusion]Both Staphylococcus aureus and Staphylococcus epidermidis could evade phagocytic and killing effects of PMNs in peripheral blood of dairy cows,and the cell viability was significantly reduced after infection.Staphylococcus aureus and Staphylococcus epidermidis could mediate the innate immune response by regulating the secretion of inflammatory factors in PMNs of dairy cows.This result laid a foundation for screening candidate factors related to Staphylococcus aureus mastitis resistance by targeting PMNs in dairy cows.
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