钙依赖性蛋白激酶4在柔嫩艾美耳球虫感染鸡盲肠组织中水平变化及其对细胞凋亡的影响OA

This study aimed to investigate the expression profile of Eimeria tenella calcium-dependent protein kinase 4(CDPK4)across its developmental cycle in chickens,as well as its regulatory role in apoptosis in chicken fibroblast DF-1 cells.Recombinant CDPK4 protein was prokaryotically expressed and purified using an Escherichia coli system,and murine polyclonal antibodies against CDPK4 were prepared.Immuno-fluorescence staining was employed to examine the temporal expression of CDPK4 in cecal tissues from chickens infected with E.tenella at different time points.Meanwhile,the recombinant plasmid pEGFP-N1-CDPK4 was constructed and transfected into DF-1 cells.The regulatory effects of CDPK4 on the expression of key apoptotic mediators and Caspase-3 enzymatic activity in DF-1 cells were evaluated using RT-qPCR,We stern blot,and colorimetric Caspase-3 activity assays.The results showed that CDPK4 expression in cecal tissues of E.tenella-infected chickens increased gradually with parasite develop-ment and proliferation.In DF-1 cells expressing CDPK4,the mRNA and protein expression levels of the anti-apoptotic factor Bcl-2 were upregulated significantly(P＜0.01),whereas the mRNA and protein expression levels of the pro-apoptotic factor Bax were downregulated significantly(P＜0.01),leading to an elevated Bcl-2/Bax ratio.Caspase-3 enzymatic activity was reduced significantly(P＜0.05),indicating that CDPK4 suppresses apoptosis in DF-1 cells.