Host-cell death pathways in L929 cells induced by Chlamydia muridarum infection

Abstract

Objective To identify the host-cell death pathways (apoptosis, autophagy or necrosis) in L929 cells at the time point of 48 hours post infection (h.p.i.) with Chlamydia muridarum. Methods L929 cells were infected with Chlamydia muridarum at a multiplicity of infection (MOI) of 0.85 for 48 hours. Nuclear fragmentation was observed under fluorescence microscopy following staining L929 cells with DAPI (4′, 6-diamidino-2-phenylindole). L929 cells were stained with propidium iodide (PI) plus Annexin Ⅴ and then analyzed by fluorescence-activated cell sorting (FACS) to clarify whether apoptosis or necrosis occurred after Chlamydia muridarum infection. L929 cells were transiently transfected with GFP-LC3 and observed under fluorescent microscopy to analyze cell autophagy. Western blot assay was performed to detect LC3 protein for further analysis of autophagy. Results Apoptosis was not induced in L929 cells by Chlamydia muridarum infection at 48 h. p.i. as no significant nuclear fragmentation was observed. Results of FACS showed that most cells died due to necrosis. Moreover, fluorescent dots of GFP-LC3 formed after infecting transfected L929 cells with Chlamydia muridarum. An increased ratio of LC3Ⅰ to LC3Ⅱ in the L929 cells infected with Chlamydia muridarum was detected by Western blot assay, indicating that autophagy occurred during Chlamydia muridarum infection. Conclusion Necrosis and autophagy rather than apoptosis are induced in L929 cells 48 hours after infection with Chlamydia muridarum. Key words: Chlamydia; L929 cell; Apoptosis; Necrosis; Autophagy