鲍曼不动杆菌荚膜厚度对鼠巨噬细胞炎症复合体活化的影响

Activation of inflammasome in infection by Acinetobacter baumannii with different capsule thickness

目的:探讨荚膜厚度不同的鲍曼不动杆菌激活鼠巨噬细胞Raw 264.7炎症复合体能力的差异。方法:采用刚果红染色法观察鲍曼不动杆菌荚膜,筛选出荚膜厚度差异显著的两株菌株。将两株菌分别感染Raw 264.7,以实时定量PCR法分析感染6 h的细胞NOD样受体家族3(NOD-like receptors,NLRP3)、Caspase-1和IL-1β基因表达量;流式细胞术分析感染1、3和6 h的细胞活性氧表达量。结果:刚果红染色可以清晰地观察到鲍曼不动杆菌周围一圈不着色的荚膜,选择出荚膜厚度差异显著的两株鲍曼不动杆菌用于感染细胞。不同荚膜厚度的鲍曼不动杆菌感染均可刺激Raw 264.7细胞NLRP3、Caspase-1和IL-1βmRNA表达上调,其中厚荚膜菌株诱导细胞NLRP3和IL-1βmRNA表达能力显著高于薄荚膜菌株(P <0.05);不同荚膜厚度的鲍曼不动杆菌感染3 h和6 h时,细胞活性氧表达量均显著高于对照组(P <0.05),且厚荚膜菌株诱导活性氧表达能力显著高于薄荚膜菌株(P <0.05)。结论:不同荚膜厚度的鲍曼不动杆菌活化鼠巨噬细胞炎症复合体能力存在差异,厚荚膜菌株诱导炎症复合体活化的能力更强。

Objective: To investigate the activation of Raw 264. 7 cell inflammasome by Acinetobacter baumannii ( A. baumannii) with different capsule thickness. Methods: A. baumannii capsule was stained by Congo red,and selected two strains with significant differences in capsule thickness. The Raw 264. 7 cells were infected with two strains of bacteria,and the cells culture was collected in 6 h. The gene expression of NOD-like receptors ( NLRP3),caspase-1 and IL-1β were determined by real-time PCR. The level of reactive oxygen species( ROS) in Raw 264. 7 cells was determined by flow cytometry in 1 h,3 h and 6 h. Results: The capsule around A. baumannii could not be stained by Congo red,and it was easy to be observed. The expression of NLRP3,caspase-1 and IL-1β mRNA in Raw 264. 7 cells infected by two strains of A. baumannii were increased significantly compared with normal cell control.The expression of NLRP3 and IL-1β in thick capsular strain was significantly higher than that of the thin capsular strain( P < 0. 05),and there was no significant difference in caspase-1 expression between the two strains. Both the two strains could up-regulate the ROS production of Raw 264. 7 cells. The ROS production in two strains infection group was significantly higher than that of normal cells control in 3 h and 6 h( P < 0. 05),and the ROS production in the thick capsular strain group was significantly higher than that of the thin capsular strain group ( P < 0. 05). Conclusion: A. baumannii with different capsule thickness showed different activation abilities of inflammasome in Raw 264. 7 cell,and the activation ability of inflammasome induced by the thick capsule strain was stronger.