中国病理生理杂志
中國病理生理雜誌
중국병리생리잡지
Chinese Journal of Pathophysiology
2015年
11期
2059-2064
,共6页
张宇晴%王新敏%王婵%王飞雨%王小芳%赵瑾%吴芳%吴江东%季榕%张万江%章乐
張宇晴%王新敏%王嬋%王飛雨%王小芳%趙瑾%吳芳%吳江東%季榕%張萬江%章樂
장우청%왕신민%왕선%왕비우%왕소방%조근%오방%오강동%계용%장만강%장악
Mcl-1%细胞凋亡%结核分枝杆菌%H37Rv%巨噬细胞
Mcl-1%細胞凋亡%結覈分枝桿菌%H37Rv%巨噬細胞
Mcl-1%세포조망%결핵분지간균%H37Rv%거서세포
Mcl-1%Apoptosis%Mycobacterium tuberculosis%H37Rv%Macrophages
目的：探讨Mcl－1信号通路阻断剂在结核分枝杆菌H37Rv感染小鼠模型中对Mcl－1表达、巨噬细胞凋亡情况及结核分枝杆菌的影响。方法：小鼠腹腔注射H37 Rv菌悬液，建立感染小鼠模型，针对Mcl－1的信号通路选用JAK／STAT信号通路阻断剂AG490、MAPK信号通路阻断剂PD98059和PI3K信号通路阻断剂LY294002用腹腔注射方式作用于各组感染小鼠模型，分为H37Rv感染组、AG490处理组、PD98059处理组、LY294002处理组和对照组。通过细胞抗酸染色观察结核分枝杆菌H37Rv感染小鼠腹腔巨噬细胞的动物模型是否建立成功；通过免疫细胞化学检测结核分枝杆菌H37Rv感染巨噬细胞的Mcl－1表达情况，使用流式细胞技术检测各组巨噬细胞的凋亡率，采用结核分枝杆菌菌落计数来判断巨噬细胞凋亡对结核分枝杆菌的清除效果。结果：细胞抗酸染色结果可见感染的巨噬细胞内散在排列的红色短小抗酸结核分枝杆菌。免疫细胞化学结果显示H37Rv感染组、AG490处理组和LY294002处理组中的Mcl－1蛋白为强阳性表达，PD98059处理组中Mcl－1蛋白为弱阳性表达，对照组Mcl－1蛋白为阴性表达。流式细胞术检测发现H37Rv感染组巨噬细胞凋亡率较对照组高，PD98059处理组的凋亡率显著高于各组，差异显著（P＜0．05）。结核分枝杆菌菌落计数结果显示PD98059处理组对H37Rv菌株抑菌作用最明显。结论：Mcl－1信号通路阻断剂通过抑制JAK／STAT、MAPK和PI3K信号通路增加结核分枝杆菌H37Rv感染巨噬细胞的凋亡率，抑制结核分枝杆菌生长；其中，MAPK信号通路干扰Mcl－1的作用最明显，感染的巨噬细胞凋亡率最高，抑菌作用最强。
目的：探討Mcl－1信號通路阻斷劑在結覈分枝桿菌H37Rv感染小鼠模型中對Mcl－1錶達、巨噬細胞凋亡情況及結覈分枝桿菌的影響。方法：小鼠腹腔註射H37 Rv菌懸液，建立感染小鼠模型，針對Mcl－1的信號通路選用JAK／STAT信號通路阻斷劑AG490、MAPK信號通路阻斷劑PD98059和PI3K信號通路阻斷劑LY294002用腹腔註射方式作用于各組感染小鼠模型，分為H37Rv感染組、AG490處理組、PD98059處理組、LY294002處理組和對照組。通過細胞抗痠染色觀察結覈分枝桿菌H37Rv感染小鼠腹腔巨噬細胞的動物模型是否建立成功；通過免疫細胞化學檢測結覈分枝桿菌H37Rv感染巨噬細胞的Mcl－1錶達情況，使用流式細胞技術檢測各組巨噬細胞的凋亡率，採用結覈分枝桿菌菌落計數來判斷巨噬細胞凋亡對結覈分枝桿菌的清除效果。結果：細胞抗痠染色結果可見感染的巨噬細胞內散在排列的紅色短小抗痠結覈分枝桿菌。免疫細胞化學結果顯示H37Rv感染組、AG490處理組和LY294002處理組中的Mcl－1蛋白為彊暘性錶達，PD98059處理組中Mcl－1蛋白為弱暘性錶達，對照組Mcl－1蛋白為陰性錶達。流式細胞術檢測髮現H37Rv感染組巨噬細胞凋亡率較對照組高，PD98059處理組的凋亡率顯著高于各組，差異顯著（P＜0．05）。結覈分枝桿菌菌落計數結果顯示PD98059處理組對H37Rv菌株抑菌作用最明顯。結論：Mcl－1信號通路阻斷劑通過抑製JAK／STAT、MAPK和PI3K信號通路增加結覈分枝桿菌H37Rv感染巨噬細胞的凋亡率，抑製結覈分枝桿菌生長；其中，MAPK信號通路榦擾Mcl－1的作用最明顯，感染的巨噬細胞凋亡率最高，抑菌作用最彊。
목적：탐토Mcl－1신호통로조단제재결핵분지간균H37Rv감염소서모형중대Mcl－1표체、거서세포조망정황급결핵분지간균적영향。방법：소서복강주사H37 Rv균현액，건립감염소서모형，침대Mcl－1적신호통로선용JAK／STAT신호통로조단제AG490、MAPK신호통로조단제PD98059화PI3K신호통로조단제LY294002용복강주사방식작용우각조감염소서모형，분위H37Rv감염조、AG490처리조、PD98059처리조、LY294002처리조화대조조。통과세포항산염색관찰결핵분지간균H37Rv감염소서복강거서세포적동물모형시부건립성공；통과면역세포화학검측결핵분지간균H37Rv감염거서세포적Mcl－1표체정황，사용류식세포기술검측각조거서세포적조망솔，채용결핵분지간균균락계수래판단거서세포조망대결핵분지간균적청제효과。결과：세포항산염색결과가견감염적거서세포내산재배렬적홍색단소항산결핵분지간균。면역세포화학결과현시H37Rv감염조、AG490처리조화LY294002처리조중적Mcl－1단백위강양성표체，PD98059처리조중Mcl－1단백위약양성표체，대조조Mcl－1단백위음성표체。류식세포술검측발현H37Rv감염조거서세포조망솔교대조조고，PD98059처리조적조망솔현저고우각조，차이현저（P＜0．05）。결핵분지간균균락계수결과현시PD98059처리조대H37Rv균주억균작용최명현。결론：Mcl－1신호통로조단제통과억제JAK／STAT、MAPK화PI3K신호통로증가결핵분지간균H37Rv감염거서세포적조망솔，억제결핵분지간균생장；기중，MAPK신호통로간우Mcl－1적작용최명현，감염적거서세포조망솔최고，억균작용최강。
[ ABSTRACT] AIM: To explore the effects of Mcl-1 signal pathway blockers on Mcl-1 expression, macrophage apoptosis and Mycobacterium tuberculosis in the model of mice infected with Mycobacterium tuberculosis H37Rv.METH-ODS:A mouse infection model was established by intraperitoneal injection of H37Rv suspension.The signaling pathway blockers AG490, PD98059 and LY294002 for JAK/STAT, MAPK and PI3K, respectively, were intraperitoneally injected into the mice infected with H37Rv.Cell acid-fast staining was used to observe whether the mouse peritoneal macrophages infected with H37Rv were successfully established.Immunocytochemical method was employed to detect Mcl-1 expression in the mouse peritoneal macrophages infected with H37Rv.The apoptotic rate in each group was measured by flow cytomer-ty.The scavenging capacity of apoptotic macrophages against H37Rv was determined by Mycobacterium tuberculosis colony counting.RESULTS:The result of cell acid-fast staining revealed the existence of dispersive arrangement of red short anti-acid Mycobacterium tuberculosis within infected macrophages.The result of cell immunocytochemistry showed strongly posi-tive expression of Mcl-1 protein in H37Rv infection group, AG490 treatment group and LY294002 treatment group, weakly positive expression of Mcl-1 protein in PD98059 treatment group, and negative expression of Mcl-1 protein in control group. The result of flow cytometry found that the macrophage apoptotic rate in H37Rv infection group was higher than that in con-trol group, while that in PD98059 treatment group was high than that in other groups with statistically significant differences (P<0.05).The result of Mycobacterium tuberculosis colony counting showed that PD98059 treatment had the most signifi-cant inhibitory effect on H37Rv strain.CONCLUSION: Mcl-1 signaling pathway blockers increase the apoptotic rate of macrophages infected with Mycobacterium tuberculosis H37Rv and inhibit the growth of Mycobacterium tuberculosis by inhibi-ting the signaling pathways of JAK/STAT, MAPK and PI3K, among which the MAPK has the most obvious interfering effect on Mcl-1, and leads to the highest apoptotic rate of infected macrophages and the strongest bacteriostasis.
