Establishment Of Cell Model And Relative Genes Expression Of Chlamydophila Psittaci Persistent Infection Induced By Interferon-γ

Objective: To establish the cell culture model of Chlamydophila psittaci6BCpersistent infection induced by gamma interferon （IFN-γ） in vitro, and detect theexpressin of relative genes at mRNA and protein levels.Methods:HeLa cells were infected with C. psittaci6BC and recombinant human IFN-γ（rhIFN-γ） was added to the medium at5to55ng/mL, then the cell cultures were har-vested30h later, or C. psittaci6BC was treated with35ng/mL rhIFN-γ and harve-sted at12,24,36,48or60h later. The infectivity of C. psittaci6BC treated by rhIFN-γ were detected by indirect immunefluorescence assay （IFA）. And the chlamydiainclusion body’s morphology was detected with IFA and transmission electron micros-cope （TEM） in rhIFN-γ induced persistent infection. After rhIFN-γ treated24h, theinducer was removing, and enough L-tryptopha was added, then the infectivity andthe inclusion body’s morphology of the C. psittaci6BC were detected by IFA or TEM.The mRNA and protein expression levels of relative C. psittaci6BC gene underpersistent infection state were assessed by real-time PCR. The expression of CPSIT₀844and CPSIT₀594during the transition of C. psittaci to the persistent wereanalysed by western blot.Results:After treated with different concentrations of rhIFN-γ, the infectivity of C.psittaci6BC was reduced and presented a dose-dependent relationship. And theinfectivity was also declined to different levels after treated with35ng/mL IFN-γ fordifferent time, especially declined dramatically at36,48and60h later. IFA and TEMshowed that the inclusion bodies appeared to be smaller, RBs were larger and the numbers of the infectious EB were less than those seen with acute, untreated C.psittaci infection. When IFN-γ was removed and L-tryptophan was added, theinfectivity and inclusion body’s morphologyof C. psittaci6BC could returned to thestate of acute infection.Realtime-PCR showed that the relative mRNA expression levels of CPSIT₀844,CPSIT₀846, CPSIT₀959, CPSIT₀594, CPSIT₀310, CPSIT₀870, CPSIT₀057and CPSIT₀208were changed during IFN-γ-induced C. psittaci persistent infection.Among which CPSIT₀870encoding product for50S ribosomal protein was signifi-cantly upregulated at2～36h p.i, the CPSIT₀959encoding cystein desulfurase wassignificantly downregulated at36and48h p.i. The type III secretion system encodinggenes CPSIT₀594and CPSIT₀844were significantly upregulated at36h and48hp.i, while the CPSIT₀846was significantly downregulated at36h p.i. The membraneproteins encoding genes CPSIT₀057and CPSIT₀310were upregulated at2～48hp.i, and CPSIT₀208was downregulated at48h p.i. The changes of protein expressionfor CPSIT₀844and CPSIT-0594was consistent with their mRNA transcriptionlevel.Conclusion:1. The C. psittaci persistent infection cell model was successfully established bylow concentration of rhIFN-γ.2. The expression levels of CPSIT₀844, CPSIT₀846, CPSIT₀959, CPSIT₀594,CPSIT₀310, CPSIT₀870, CPSIT₀057and CPSIT₀208were changed duringrhIFN-γ-induced C. psittaci6BC persistent infection, which may be concernedwith the transition of C. psittaci to persistent infection state.