中华危重病急救医学
中華危重病急救醫學
중화위중병급구의학
Chinese Critical Care Medicine
2015年
6期
504-508
,共5页
多器官功能障碍综合征%深肌层Cajal间质细胞%细胞凋亡%Bax%Bcl-2
多器官功能障礙綜閤徵%深肌層Cajal間質細胞%細胞凋亡%Bax%Bcl-2
다기관공능장애종합정%심기층Cajal간질세포%세포조망%Bax%Bcl-2
Multiple organ dysfunction syndrome%Interstitial cells of Cajal in deep muscular layer%Apoptosis%Bax%Bcl-2
目的：观察多器官功能障碍综合征（MODS）大鼠小肠深肌层Cajal间质细胞（ICC-DMP）的凋亡现象，以及表型标志酪氨酸激酶受体（c-kit）、线粒体凋亡信号通路Bcl-2和Bax表达的变化，揭示MODS状态下胃肠运动功能障碍的机制。方法健康成年Wistar大鼠40只，按随机数字表法分为对照组（20只）和MODS组（20只）。经腹腔注射8×108 cfu/mL大肠杆菌混悬液1 mL制备细菌性腹膜炎致MODS大鼠模型；对照组给予等量生理盐水。制模24 h后，取大鼠上段小肠肌层制作全厚标本，分别应用透射电镜扫描、免疫荧光双标染色、激光扫描共聚焦显微镜，观察两组大鼠小肠ICC-DMP的超微结构、网络结构及c-kit和Bax/Bcl-2的表达。结果大体标本观察显示：对照组大鼠胃肠蠕动正常；MODS组大鼠麻痹性胃肠梗阻症状明显。透射电镜下观察显示：ICC-DMP中间丝结构清晰，无肿胀线粒体，染色质分布均匀，核周聚集少量异染色质；MODS组ICC-DMP中间丝模糊，线粒体肿胀明显，细胞核内有大量染色体聚集。共聚焦显微镜下观察显示：对照组ICC-DMP网络结构清晰，c-kit和Bcl-2表达强烈且区域重合，而Bax表达较弱并呈散点样分布；MODS组ICC-DMP数量较对照组显著减少（个/HP：15.80±2.30比25.70±3.97，t＝6.819，P＝0.000），网络完整性损伤明显；c-kit和Bcl-2的表达较对照组明显降低〔c-kit（荧光强度）：129.56±36.90比307.23±40.07，t＝10.314， P＝0.000；Bcl-2（荧光强度）：103.23±25.19比378.92±43.79，t＝17.259，P＝0.000〕，而Bax表达较对照组明显增加（荧光强度：270.94±36.98比92.57±20.92，t＝-13.277，P＝0.000）。结论 MODS致胃肠动力障碍的机制可能与ICC-DMP超微结构损伤、c-kit表型改变和线粒体凋亡途径的激活密切相关。
目的：觀察多器官功能障礙綜閤徵（MODS）大鼠小腸深肌層Cajal間質細胞（ICC-DMP）的凋亡現象，以及錶型標誌酪氨痠激酶受體（c-kit）、線粒體凋亡信號通路Bcl-2和Bax錶達的變化，揭示MODS狀態下胃腸運動功能障礙的機製。方法健康成年Wistar大鼠40隻，按隨機數字錶法分為對照組（20隻）和MODS組（20隻）。經腹腔註射8×108 cfu/mL大腸桿菌混懸液1 mL製備細菌性腹膜炎緻MODS大鼠模型；對照組給予等量生理鹽水。製模24 h後，取大鼠上段小腸肌層製作全厚標本，分彆應用透射電鏡掃描、免疫熒光雙標染色、激光掃描共聚焦顯微鏡，觀察兩組大鼠小腸ICC-DMP的超微結構、網絡結構及c-kit和Bax/Bcl-2的錶達。結果大體標本觀察顯示：對照組大鼠胃腸蠕動正常；MODS組大鼠痳痺性胃腸梗阻癥狀明顯。透射電鏡下觀察顯示：ICC-DMP中間絲結構清晰，無腫脹線粒體，染色質分佈均勻，覈週聚集少量異染色質；MODS組ICC-DMP中間絲模糊，線粒體腫脹明顯，細胞覈內有大量染色體聚集。共聚焦顯微鏡下觀察顯示：對照組ICC-DMP網絡結構清晰，c-kit和Bcl-2錶達彊烈且區域重閤，而Bax錶達較弱併呈散點樣分佈；MODS組ICC-DMP數量較對照組顯著減少（箇/HP：15.80±2.30比25.70±3.97，t＝6.819，P＝0.000），網絡完整性損傷明顯；c-kit和Bcl-2的錶達較對照組明顯降低〔c-kit（熒光彊度）：129.56±36.90比307.23±40.07，t＝10.314， P＝0.000；Bcl-2（熒光彊度）：103.23±25.19比378.92±43.79，t＝17.259，P＝0.000〕，而Bax錶達較對照組明顯增加（熒光彊度：270.94±36.98比92.57±20.92，t＝-13.277，P＝0.000）。結論 MODS緻胃腸動力障礙的機製可能與ICC-DMP超微結構損傷、c-kit錶型改變和線粒體凋亡途徑的激活密切相關。
목적：관찰다기관공능장애종합정（MODS）대서소장심기층Cajal간질세포（ICC-DMP）적조망현상，이급표형표지락안산격매수체（c-kit）、선립체조망신호통로Bcl-2화Bax표체적변화，게시MODS상태하위장운동공능장애적궤제。방법건강성년Wistar대서40지，안수궤수자표법분위대조조（20지）화MODS조（20지）。경복강주사8×108 cfu/mL대장간균혼현액1 mL제비세균성복막염치MODS대서모형；대조조급여등량생리염수。제모24 h후，취대서상단소장기층제작전후표본，분별응용투사전경소묘、면역형광쌍표염색、격광소묘공취초현미경，관찰량조대서소장ICC-DMP적초미결구、망락결구급c-kit화Bax/Bcl-2적표체。결과대체표본관찰현시：대조조대서위장연동정상；MODS조대서마비성위장경조증상명현。투사전경하관찰현시：ICC-DMP중간사결구청석，무종창선립체，염색질분포균균，핵주취집소량이염색질；MODS조ICC-DMP중간사모호，선립체종창명현，세포핵내유대량염색체취집。공취초현미경하관찰현시：대조조ICC-DMP망락결구청석，c-kit화Bcl-2표체강렬차구역중합，이Bax표체교약병정산점양분포；MODS조ICC-DMP수량교대조조현저감소（개/HP：15.80±2.30비25.70±3.97，t＝6.819，P＝0.000），망락완정성손상명현；c-kit화Bcl-2적표체교대조조명현강저〔c-kit（형광강도）：129.56±36.90비307.23±40.07，t＝10.314， P＝0.000；Bcl-2（형광강도）：103.23±25.19비378.92±43.79，t＝17.259，P＝0.000〕，이Bax표체교대조조명현증가（형광강도：270.94±36.98비92.57±20.92，t＝-13.277，P＝0.000）。결론 MODS치위장동력장애적궤제가능여ICC-DMP초미결구손상、c-kit표형개변화선립체조망도경적격활밀절상관。
Objective To observe the apoptosis of interstitial cells of Cajal in deep muscular layer ( ICC-DMP ) of small intestine in rats with multiple organ dysfunction syndrome ( MODS ) as a result of bacterial peritonitis, and the expression of c-kit ( an ICC phenotype marker ) and Bax/Bcl-2, in order to investigate the mechanism of gastrointestinal motility dysfunction in MODS. Methods According to the random number table, 40 Wistar rats were randomly divided into two groups:control group ( n=20 ) and MODS group ( n=20 ). The MODS model in rats was reproduced by intraperitoneal injection of 8×108 cfu/mL Escherichia coli suspension 1 mL, and the control group was given the same amount of normal saline. After 24 hours, the upper small intestine was harvested for examination. Ultrastructure of ICC-DMP was observed using electron microscope. The network structure of ICC-DMP and the expression of c-kit and Bax/Bcl-2 were observed and determined with immunofluorescence and laser scanning confocal microscope. Results Macroscopic observation revealed that the gastrointestinal motility of rats was normal in the control group. Compared with the control group, gastro intestine was significantly expanded with parulytic ileus in MODS group. It was shown by transmission electron microscopy that intermediate filament structure of ICC-DMP was clear without swelling of mitochondria; chromatin distributed uniformly with small amounts of heterochromatin aggregated in perinuclear. Compared with the control group, intermediate filament structure of ICC-DMP was fuzzy, and mitochondria were swollen obviously in MODS group;chromatin was assembled in nucleus centre. It was shown by laser scanning confocal microscope that the network structure of ICC-DMP was clear, the expression of c-kit and Bcl-2 was strongly and overlapping;the expression of Bax was weak and scatter distributed. Compared with control group, ICC-DMP quantity in MODS group was significantly reduced ( cells/HP: 15.80±2.30 vs. 25.70±3.97, t = 6.819, P = 0.000 ), and ICC network was incomplete. The expression of c-kit and Bcl-2 was significantly decreased as compared with control group [ c-kit ( fluorescence intensity ):129.56±36.90 vs. 307.23±40.07, t=10.314, P=0.000;Bcl-2 ( fluorescence intensity ):103.23±25.19 vs. 378.92±43.79, t=17.259, P=0.000 ], whereas, the expression of Bax was significantly increased ( fluorescence intensity:270.94±36.98 vs. 92.57±20.92, t=-13.277, P=0.000 ). Conclusion The mechanism of gastrointestinal motility dysfunction in MODS maybe closely related to ultrastructural damage of ICC-DMP, changes of c-kit phenotypic and activation of mitochondrial apoptosis pathway.
