| Heat stress have a negative impact on production performance, reproductive performance and immune performance of dairy cattle, and explore the ways to decrease or relieve heat stress was always one of the major topics of dairy cattle breeding. To research the effect of exogenous metallothionein (MT) on apoptosis rate and cell cycle of lymphocytes in peripheral blood of dairy cattle under heat stress,20lactating Chinese Holstein cows were selected and randomly divided into group A, group B, group C and group D, Zn-MT in the levels of0,4.0,8.0and12.0mg were supplemented by intravenous injection respectively. Blood samples were collected at1st (before injection),21st,36th,51st and61st day, and the apoptosis rate, cell cycle and subsets distribution as well as the levels of cell cycle and apoptosis related genes expression and related protein phosphorylation of lymphocytes in peripheral blood of dairy cattle were determined. The results showed that:1) Effect of MT on cell cycle, apoptosis rate and subsets distribution of lymphocytes in peripheral blood of dairy cattle under heat stressThe apoptosis rate of lymphocytes in group B and C were lower than those in group A by26.63%(P>0.05) and24.84%(P>0.05), respectively. The number of lymphocytes at G0/G1phage in B and C groups were decreased and the number of lymphocytes at S and G2/M phages were tend to increase, but significant differences were not be observed(P>0.05). The number of CD3+in three trial groups were greater than those in group A by7.02%(P>0.05),5.45%(P>0.05) and3.85%(P>0.05), while the number of CD4+in experimental groups were higher than those in control group by31.04%(P<0.05),35.68%(P<0.05) and39.34%(P<0.05), respectively. The number of CD8+and the levels of CD4+/CD8+in trial groups were increased observably, but significant differences(P<0.05) was observed in the levels of CD4+/CD8+between group A and C only. It suggested that exogenous Zn-MT can decrease apoptosis rate, and regulate cell cycle and subsets distribution in dairy cattle in a dose-dependent manner.2) Effect of MT on the expression levels of cell cycle related and apoptosis related genes of lymphocytes in peripheral blood of dairy cattle under heat stressThe expression levels of cytc, Fas, Apaf-1and p53genes of lymphocytes in peripheral blood were decreased after injection of MT, and the expression level of cytc in group B was lower than control group significantly(P<0.05), the expression level of Fas in group D and the expression levels of Apaf-1and p53in group B, C and D were decreased highly significantIy(P<0.01) compared to group A. The expression levels of Cyclin A, Cyclin D and Cyclin E in group B and C were increased or the same as control group, the expression level of Cyclin A in group B and C was great than group A highly significantly(P<0.01), and the trend of decrease was observed in group D. The expression levels of CDK2, CDK4and CDK6in experimental groups were higher than control group significantly(P<0.05) or highly significantly(P<0.01), except CDK6in group C. It suggested that exogenous Zn-MT can decrease the expression level of apoptotic genes and increased the expression level of positive regulator in cell cycle effectively in a dose-dependent manner.3) Effect of MT on the expression levels of p53and p105protein, and phosphoryl-ation of p53, p65and p105proteinThe expression levels of p53and p105protein, and phosphorylation of p53, p65and p105protein were decreased after injection of MT. The expression level and phosphorylation of p53protein in group B and C were lower than control group by35.00%(P<0.01),32.00%(P<0.01) and18.00%(P<0.01),12.00%(P<0.01) respectively. The expression levels and phosphorylation of p105protein in group B, C and D were lower than group A by25.00%(P<0.01),20.00%(P<0.01),10.00%(P<0.05) and10.00%(P<0.01),8.00%(P<0.05),4.00%(P>0.05)respectively. The phosphorylation of p65protein in experimental groups were lower than control group by16.00%(P<0.01),11.00%(P<0.01) and5.00%(P>0.05) respectively. It suggested that exogenous Zn-MT can decrease the apoptosis rate of lymphocytes in peripheral blood by inhibiting the expression levels and phosphorylation of pro-apoptotic proteins, and dose-dependent manner was exerted.In conclusion, exogenous MT can decrease the apoptosis rate of lymphocytes by inhibiting the expression levels of pro-apoptotic genes and proteins as well as the phosphorylation of pro-apoptotic proteins. Exogenous MT also can promote the regular cell cycle by increaseing the expression levels of positive regulator, while improve the subsets distribution of lymphocytes, and finally enhance the immunity of dairy cattle under heat stress. The dosage in4.0and8.0mg were batter than12.0mg in this study. |
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