Peresistent Chlamydophila Psittaci Infection Of HeLa Cell Inhibits Apoptosis Via Bcl-2 Family Proteins And The ERK Survival Pathway

Background and Objective: Apoptosis plays a regulatory pivotal machining of pathogen invade and pathopoiesia in human organisms.Our groups research indicates Chlamydophila psittaci(Cps) infected HeLa cell via inhibition of host cell apoptosis. But its mechanism is still unknown. This study was to evaluate the mechanism of apoptosis after deal with recombinant human IFN-γ(rh IFN-γ). Observe the influence of HeLa cell related proteins. And sequentially, preliminarily clarify the regulatory mechanism by Cps persistent infection.Methods:HeLa cell culture in vitro, and infected Cps(MOI 2). 35ng/m L rh IFN-γ adds with Cps infected host cells,and introduce Cps persistent infection. Cps persistent infected with various time points: 0, 4, 8, 12, 24 h,and collected with experimental samples in various time points. After Hochest33258 and TUNEL staining, we detect apoptotic phenomenon of host cell. Nucleus morphology, mitochondrial membrane potential(MMP)and cytochrome c(Cyt.C) were detected by immunofluorescence assay(IFA). And contemporary, expression levels of Fas、Fas-L、Caspase 8、Caspase 3、Cyt.C、t Bid、Bad、Bim、Mcl-1 and ERK phosphorylation were detected by Western Blotting.Results:(1)Cps 6BC persistent infected HeLa cells at 0, 4, 8, 12, 24 h,we observed Hochest33258 and TUNEL staining, we can showed the inhibition of host cells by apoptotic rate. Cps 6BC infection after 24 h,apoptotic phenomenon was recover but still lower than controls.( 2)Caspase 3 expression levels was up-regulate Cps persistent infection after4 h. But its activated state(Cleaved Caspase 3) was down-regulate in a time-dependent manner.(3)Caspase 3 expression level was up-regulate Cps persistent infection after 4h. But its activated state(Cleaved Caspase3) was down-regulate in a time-dependent manner.(4)Mcl-1 and Bad expression level are lower in uninfected cell. Cps persistent infection after 4h, Mcl-1 growing gradually and continuing to 24 h. And contemporary, Bad significantly increase and expression constant in4~24h.( 5) Inhibits the CCCP introduce the release of mitochondrial Cyt.C by Cps persistent infection, and continuing to 24 h. Furthermore,This factor also reduce the effects of MMP.(6)Along with the infection time, Bim EL expression decreased and Bims expression increased in HeLa cell. And contemporary, Bim, t Bid, Cyt.C were decreased in HeLa cell(. 7)Along with the infection time, the apoptotic receptor Fas, Fas-L and Caspase 8 expression levels were decreased in a time-dependent manner.(8) Cps 6BC infection after 4~24h, The phosphorylation of survival pathway ERK1/2 was up-regulted, and up-related the ERK phosphorylation level of STS introdced cell.Conclusion: Cps 6BC persistent infection induce up-regulation of Bcl-2family protein Mcl-1, t Bid, Bim and ERK phosphorylation level, and inhibition of Cyt.C release to inhibition of host cells apoptosis.