Mycobacterium Tuberculosis LpqT Protein Can Enhance The Survival Of M.smegmatis In Vivo By Inhibiting Inflammatory Cytokine Expression And Apoptosis Of Macrophage

Tuberculosis(TB)is achronic infectious disease caused by Mycobacterium tuberculosis(MTB).Macrophages are the major host cells of MTB,as well as the first line of defense against the bacteria.MTB cellular components are their main pathogenic substances,through which they inter fere with the host immune system to evade clearance.A variety of MTB cell wall lipoproteins can bind to macrophage membrane receptors and regulate their signaling pathways to facilitate the survival of the bacteria.LpqT is a conserved lipid protein of about 25 kDa whose function is still unknown.A high through-put screening of H37 Rv transposon mutation library showed that LpqTmutant had a much lower survival rate than that of normal control,suggesting LpqT might be able to support the survival of MTB in macrophages.To investigate the functions of LpqT on Mtb survival,we design and carry out the project,and achieved the following results:(1)We successfully constructed the LpqT knockout M.smegmatisbacterial strain as well as one that overexpressed LpqT.(2)In order to investigate the effect of LpqT on the survival of Mycobacterium smegmatis in vivo,we infect the macrophage cell line and BalB/c mice with LpqT knockout or overexpression M.smegmatis strains,and bacterial loads in macrophage or animal tissues were determined.The results showed that LpqT can promote the survival of M.smegmatis in both macrophages cell line and animal tissues,indicating that LpqT can promote the survival of M.smegmatis invivo.(4)To investigate the effect of LpqT on the immunological functions of macrophage,we infected RAW264.7 cells with the LpqT knockout or overexpression bacteria strains,and detected the expression of various cytokines by Real time PCR and ELISA.It was found that LpqT inhibited the expression of IL-1 and Il-6 in macrophages.We also examined the apoptosis related proteins including cleaved Caspase3 finding that LpqT could inhibit the apoptosis ofmacrophages(4)To investigate the mechanism by which LpqT inhibits cytokine expression in macrophage,we stimulate the RAW264.7 cells wtih the LpqT knockout or overexpression bacteria strains,then detect the levels of phosphorylated JNK,Erk1/2,and protein p38.The results show that LpqT can inhibit JNK,P38 and Erk1/2phosphorylation,both of which belongs to MAPK signal pathways.According to ourresults,we get the conclusion that LpqT can promote the survival of M.smegmatis in macrophage by inhibiting inflammatory cytokine expression and apoptosisin macrophages,and this is at least partially due to its inhibition of Mycobacterium activated MAPK signal pathways.