Preliminary Study On The Apoptosis Mechanism Of Macrophages Induced By Brucella Virulent Strain And Vaccine Strain

Object:Brucellosis (Brucellosis) by a zoonotic infectious disease caused by members of the Brucella genus (Brucella), usually capable of long-term sustainability of the infection to humans and animals, cause genital injury, once the infection is difficult to cure. Cases of brucellosis in the worldwide outbreak, caused a huge threat to human health and safety and agricultural economy and loss, also can be as a terrorist warfare agents.Brucella are intracellular parasites, the Gram-negative bacteria, primarily to nourish in embryonic layer nucleus macrophage cells as a host cell. The brucella not produce exotoxin, its virulence is mainly manifested in the invasion force, as well as in host survival and ability to reproduce. Brucella inhibit macrophage apoptosis is capable of growth and reproduction in the host body and not by the body’s immune system to identify clear the main reason. Therefore, through to the brucella induced macrophage apoptosis mechanism research is the basis for the study of brucellosis of pathogenic mechanism.Methods:(1) Establishment of Brucella strains induced macrophage apoptosis mechanism to provide the best multiplicity of infection Brucella virulent strain16M, the vaccine strain M5-90infected macrophage RAW264.7model.(2) Brucella virulent strain16M vaccine strain M5-90best multiplicity of infection infected macrophages RAW264.7, compare the strength of strain induced macrophage apoptosis differences.(3) Based on the strength of strain-induced macrophage apoptosis differences, selected strains promote apoptosis, inquiring of caspase3,8,9induced apoptosis.(4) Detection of apoptosis-related factors AIF, Bcl-2, Bax, Apaf-1, Bcl-xl in the strength of strain-induced macrophage apoptosis expression difference in the amount.Results:(1) Successful establishment of Brucella induced macrophage apoptosis multiplicity of infection of50:1.(2) Early stage of the infection, Brucella vaccine strain M5-90can promote macrophage apoptosis.(3) Caspase3,9Brucella vaccine strain M5-90-induced macrophage apoptosis plays a role in promoting.(4) ELISA results show that the expression of macrophage apoptosis induced by the vaccine strain M5-90, TNF-a and Cytc amount increased, and higher than the virulent strain16M induced. Show the AIF expression of an upward trend in real-time quantitative PCR results and the vaccine strain M5-90-induced expression expression levels higher than16M; ratio of Bax/Bcl-2ratio induced M5-90ratio is greater than16M; Apaf-1expression level there is no significant change in trend, but the M5-90-induced expression always higher than16M; Bcl-xl expression levels increased with the time of infection, increasing expression levels higher than M5-90-induced16M. Confocal laser display Cytc positioning display Cytc belong pack slurry expression in macrophages within and M5-90induced expression was significantly higher than16M, and with increasing time after infection growth.Conclusion:(1) The multiplicity of infection50:1infected macrophages, the early stages of infection, Brucella vaccine strains M5-90can promote apoptosis.(2) Caspase3,9M5-90-induced macrophage apoptosis play an important pro-apoptotic role.(3) Vaccine strain M5-90infected macrophages induced TNF-a Cytc, AIF, induced the upregulation of Bax/Bcl-2Apaf-1expression levels higher than16M, contrary expression of Bcl-xl.