Mycoplasma Genitalium Lipid-assoxiated Membrane Proteins Induce Human Monocytic Cells Express Proinflammatory Cytokines And Apoptosis By Activating Nuclear Factor κB

Objectives: This study was designed to investigated the potential pathogenicity of Mycoplasma genitalium (M. genitalium) and its molecular mechanisms responsible for the induction of proinflammatory cytokines and cell apoptosis in human monocytic cells (THP-1) stimulated by lipid-associated membrane proteins (LAMPs) prepared from M. genitalium. LAMPs were utilized to study the production of tumor necrosis factorα(TNF-α) , interleukin-1β(IL-1β) and interleukin-6 (IL-6) and the expression of their mRNA. The activation of nuclear factor kappa B (NF-κB) and cell apoptosis in THP-1 cells stimulated by LAMPs were also detected.Methods: THP-1 cells were stimulated with M.genitalium LAMPs to analyze the production of TNF-α,IL-1βand IL-6 by ELISA and the expression of their mRNA was detected by RT-PCR. The activity of transcriptional factors, NF-κB, was examined by Electrophoretic Mobility Shift Assay (EMSA). The effects of pyrrolidine dithiocarbamate (PDTC), an inhibitor of NF-κB, on the production of TNF-α,IL-1βand IL-6, the expression of mRNA and apoptosis were also examined in THP-1 cells treated with M.genitalium LAMPs. Cell apoptosis was detected in THP-1 cells by Annexin- V-FITC-propidium iodide (PI) staining and acriding orange (AO)-ethidium bromide (EB) staining.Result: M.genitalium LAMPs stimulated THP-1 cells to produce TNF-α,IL-1βand IL-6 in dose- and time-dependent manner. There was an optimal concentration of LAMPs at 3μg/mL in the induction of TNF-α(1885.29±58.62)pg/mL, IL-1β(256.20±2.03)pg/mL, IL-6(35.29±0.49)pg/mL;when the concentration of LAMPs added from 3μg/mL to 5μg/mL, whereas the level of proinflammatory cytokines decreased.The TNF-α,IL-1βand IL-6 could be detected in the conditioned media after 6h by stimulation with LAMPs and reached peak levels at 12h. The mRNA levels and production of TNF-α,IL-1βand IL-6 were inhibited by LAMPs combination with PDTC, an inhibitor of NF-κB. The activation of NF-κB was triggered in THP-1 cells after 2h treated with LAMPs. Cell apoptosis were detected in THP-1 cells treated with LAMPs and apoptosis could be partly inhibited by PDTC。Conclusion:1. M.genitalium LAMPs could stimulate THP-1 cells to produce TNF-α,IL-1βand IL-6 and apoptosis;2. M.genitalium LAMPs could activate NF-κB;3. The expression of proinflammatory cytokines and the apoptosis induced by M.genitalium LAMPs in THP-1 cells were associated with the activation of NF-κB.