双歧杆菌脂磷壁酸激活巨噬细胞活性机制的研究

Study on the activating mechanisms of macrophages by LTA of Bifidobacteria

目的 从信号分子PKC和细胞内游离Ca2 + ([Ca2 + ]i)这一途径探索青春型双歧杆菌的脂磷壁酸 (lipoteichoicacid ,LTA)激活小鼠腹腔巨噬细胞的机制 ,同时观察巨噬细胞之间缝隙连接通讯(GJIC)的变化。方法 γ 3 2 PATP磷酸转移法测定巨噬细胞总PKC活性 ;激光共聚焦显微镜检测 [Ca2 + ]i浓度的变化 ;激光漂白后荧光恢复技术 (FRAP)观察GJIC的状态。结果 (1)LTA能明显提高巨噬细胞总PKC活性 ,呈剂量依赖性 ;(2 )LTA可显著升高巨噬细胞 [Ca2 + ]i的水平 ,并且EDTA和维拉帕米处理组、肝素和普鲁卡因处理组以及EDTA、维拉帕米、肝素和普鲁卡因处理组巨噬细胞内 [Ca2 + ]i亦升高 ,但明显低于对照组 (P <0 .0 1)。 (3)巨噬细胞被LTA刺激后 ,其平均荧光恢复率明显高于对照组(P <0 .0 1)。结论 LTA可通过提高PKC活性 ,升高 [Ca2 +

Objective To explore the mechanisms of mouse peritoneal macrophages activated by lipoteichoic acid(LTA) of Bifidobacteria adolescence the pathway of protein kinase C(PKC) and intercellular free calcium ion ([Ca 2+ ]i), and to identify the changes of gap junctional intercellular communication (GJIC) among macrophages. Methods Total PKC activity of macrophages was detected by γ 32 P ATP phosphate transferring method; The changes of [Ca 2+ ]i was examined by laser scanning confocal microscopy; The state of GJIC was observed by Fluorescene recovery after photobleaching(FRAP). Results (1) LTA obviously elevated total PKC activity of macrophages in a dose dependant manner. (2) LTA significantly enhanced the level of [Ca 2+ ]i of macrophages. Furthermore, the [Ca 2+ ]i concentration of macrophages increased in EDTA and verapamil preparation group, heparin and procaine treated group, EDTA and verapamil heparin and procaine treated group, but was significantly lower when compared with control group ( P <0.01). (3) After macrophages being stimulated by LTA, its average fluorescent recovery ratio was significantly higher than that in control group ( P <0.01). Conclusion LTA could activate macrophages by elevating PKC activity, increasing [Ca 2+ ]i level as well as promoting the function of GJIC.