Infection Of Mice And Macrophages With Mycobacterium Tuberculosis H37Ra And BCG

Objectives:1. To investigate the colonization of H37Ra or BCG in mice, and special immune response after the immunization of mice with H37Ra or BCG . 2. To explore the product of nitric oxide(NO), viability of bacteria, and the survival of macrophages after the infection of macrophages with H37Ra or BCG. Methods:1. BALB/c mice were subcutaneously immunized with H37Ra and BCG respectively. At various intervals(15 days, 30 days, 60 days) after immunization, the spleens and lungs of mice were isolated, dissociated and suitable dilutions of the homogenates were plated on Lowenstein medium. The numbers of colony forming unit(CFU) were counted 18 days after incubation. 30 days and 60 days after immunization, the spleen lymphocytes of mice were cultured with PPD in vitro. Lymphocyte transformation test was detected by MTT assay. The product of IFN-γ in the supernatants of culture was determined by ELISA. 2. Infection of BALB/c mice peritoneum-derived macrophages, RAW264.7 cells and THP-1 cells with H37Ra or BCG, the product of NO in the supernatants of culture was measured by using the Griess reagent after 24 hours. At 0 day, 4 days and 7 days after infection, macrophages were lysed with 1% Triton X-100, and bacterial numbers were determined by plating on Lowenstein medium, and at each time point the survival of macrophages was measured by MTT assay. Results:1. H37Ra and BCG can exist in the spleens and lungs of BALB/c mice for 60 days at least. The stimulation index(SI) of spleen lymphocytes and the IFN-γ product in group immunized with H37Ra or BCG were statistically larger than that unimmunized 30 days or 60 days after immunization(P<0.05); The product of IFN-γ in group immunized with H37Ra was statistically larger than that immunized with BCG (P<0.05). 2. The NO product of BALB/c mice peritoneum-derived macrophages, RAW264.7 or THP-1 cells induced by H37Ra or BCG were significant larger than the control(P<0.05); there was no significant difference in NO product between group induced by H37Ra and that induced by BCG(P>0.05). H37Ra and BCG can multiply in the macrophages. The survival of macrophages decreased after infection of H37Ra or BCG. Bacteria destroyed macrophage monolayers, and the effects of damage were more and more significant when the infection delayed. Conclusions:1. H37Ra and BCG can colonize in BALB/c mice and induce special anti-tuberculosis cell immune response in mice; the immunizing capacity of H37Ra was slightly better than BCG. 2. H37Ra and BCG can activate macrophages, and they were some level of virulence to macrophages; H37Ra was less virulent than BCG.