Study On The Immune Efficacy With Mycobacterium Tuberculosis H37Ra And Organic Selenium

Objectives:1. To investigate the colonization of bacteria in mice, and the special cellular immune efficacy after immunization with Mycobacterium tuberculosis(MTB) H37Ra and organic selenium. 2. To construct target cells expressing Ag85B protein of MTB, and to investigate the specific cytotoxic activity of spleen lymphocyte, and the immune mechanism after mice immunized with MTB H37Ra and organic selenium.Methods: 1.C57BL/6 female mice were divided into six Group:⑴H37Ra+Se group,⑵BCG+Se group,⑶H37Ra group,⑷BCG group,⑸Se group and⑹PBS control group.Mice were immunized intracutaneously with H37Ra or BCG after fed with organic selenium 1 week, then continued to fed 4 weeks. At various intervals(4 weeks, 8 weeks)after immunization, to detect the index of spleen and thymus and colonization of bacteria in mice. 2. the spleen lymphocytes of mice were collected and cultured with PPD in vitro. Lymphocyte transformation test was detected by MTT assay .The production of IFN-γand IL-2 in the supernatants of mice was tested by ELISA. 3. To construct target cells expressing Ag85B protein of MTB, and detected the cytotoxic activity of spleen lymphocyte in immunized mice by the method of MTT assay. 4.At 8 weeks after immunization, the expression of spleen lymphocytes perfrion,granzymeB mRNA were measured by RT-PCR.Results: 1. At various intervals(4 weeks, 8 weeks)after immunization, the the spleen index,the thyme index and colonization of bacteria in mice in H37Ra+Se group slightly higher than BCG+Se group,H37Ra group, but no statistically difference(P>0.05). 2.SI and the production of IFN-γ,IL-2 in H37Ra+Se group statistically higher than H37Ra group(P<0.05), but compared with BCG+Se group, there were no statistically difference(P>0.05) except the production of IFN-γ(P<0.05). The results of 8 weeks were lower than that of 4 weeks. 3. Ag85B protein were expressed in the positive cells. The cytotoxic activity of spleen lymphocyte in the group immunized with MTB H37Ra and organic selenium was significantly higher than that of H37Ra group (P<0.05), and slightly higher than BCG+Se group (P>0.05). 4.The expression of perfrion,granzymeB mRNA in H37Ra+Se group was statistically higher than those in PBS control group(P<0.05), and slightly higher than BCG+Se group(P>0.05); the expression of perfrion mRNA in H37Ra+Se group was significantly higher than H37Ra group(P<0.05), but the expression of granzymeB mRNA was no difference between the two groups(P>0.05).Conclusions: 1.Colonization of H37Ra in the spleens and lungs could last 60 days after mice were immunized with MTB H37Ra and organic selenium. 2. The immune efficacy of immunization with MTB H37Ra and organic selenium was better than vaccinated only with H37Ra or BCG, and slightly better than immunization with BCG and organic selenium. 3. The target cells expressing MTB Ag85B proteinwas constructed successfully and to detect the cytotoxic activity of spleen lymphocyte in immunized mice by the method of MTT assay, would provide a better method for study the cytotoxic activity of MTB vaccine. 4. The cytotoxic activity of spleen lymphocyte were enhanced after mice immunized with MTB H37Ra and organic selenium, which may be relative to the expression of perfrion and granzymeB.