布鲁氏菌效应分子在持续性感染中的作用机制研究

Brucellosis as a chronic infectious disease caused by Brucella. Brucella cause persistent infection by inducing autophagy, apoptosis in host in vivo, which is an important cause of brucellosis of hard to kill. Previous research found 2 Brucella secreted effector molecules VceA and VceC, and confirmed that plays an important role in Brucella in host cells growth and pathogenicity, but its host cell autophagy / apoptosis whether the role is not clear. The study of the construction of Brucella 2308 standard strains of VceA and VceC gene deletion mutant; respectively by deletion strains of Brucella infection in trophoblast cells and SPF mice, using confocal microscopy, flow cytometry, QRT-PCR, ELISA, CFU and other detection methods in different stages, infecting the cell supernatant of infected mice. infected cells and cytokines, apoptosis, autophagy and apoptosis rate was detected Brucella effector molecules VceA and VceC in the regulation of autophagy and apoptosis in persistent infection, which will reveal and provide scientific basis in the Brucella infection mechanism and the development of new drugs.

布鲁氏菌病是由布鲁氏菌引起的一种慢性感染性疾病，利用细胞自噬、抑制细胞凋亡在宿主体内造成持续性感染，是布鲁氏菌病难以消灭的重要原因。研究发现了布鲁氏菌外分泌的效应分子VceA和VceC对布鲁氏菌在宿主细胞内的生长繁殖和致病具有重要作用，但其对宿主细胞的自噬/凋亡是否有调控作用尚不清楚。本研究拟通过构建布鲁氏菌2308标准株VceA和VceC基因缺失株；布鲁氏菌标准株和缺失株分别侵染胚胎滋养层细胞和感染SPF小鼠，采用共聚焦显微镜、流式细胞仪、QRT-PCR、ELISA、CFU等检测手段，在不同感染阶段对细胞液上清、感染细胞及感染鼠的细胞因子、自噬/凋亡分子、细胞凋亡率等进行检测，研究布鲁氏菌效应分子VceA和VceC在细胞自噬/凋亡中的调控作用，为揭示布鲁氏菌的持续性感染机制和开发新型治疗药物提供科学依据。

获得布鲁氏菌国际标准强毒株2308株VceA和VceC基因缺失突变株；证实在侵染不同的时间段 VceA 蛋白抑制细胞自噬；VceC 缺失株侵染细胞后表现为相反趋势，则 VceC 蛋白是促进细胞自噬。VceA 和 VceC 缺失株侵染后 Caspase-3 的高表达， Bax/Bcl-2 比值的增大，细胞凋亡率显著升高，细胞中 Cyt-C 的释放量增多，VceA 和 VceC 两蛋白均在不同程度上促进细胞凋亡。培养博士研究生2名，培养硕士研究生3名。发表SCI收录论文4篇，国内核心期刊论文1篇。超额完成了既定的研究目标。