CAJ | 학술논문

目的探讨大鼠肠缺血再灌注时肠组织微小RNA(miRNA)表达的变化.方法成年雄性SD大鼠12只,体重230～250 g,采用随机数字表法分为2组(n=6):假手术组(S组)和缺血再灌注组(Ⅰ/R组).采用阻断肠系膜上动脉60 mim再灌注120 min的方法制备肠缺血再灌注损伤模型.再灌注结束后观察小肠组织病理学结果,行Chiu评分,测定小肠含水率和肠黏膜组织乳酸含量.对肠黏膜组织进行miRNA表达谱芯片分析,采用qRT-PCR法验证差异表达的miRNA.另取成年雄性SD大鼠24只,体重230～ 250 g,采用随机数字表法分为4组(n=6):miR-378抑制剂组(An组)、miR-378激动剂组(Ag组)、抑制剂对照组(AnC组)和激动剂对照组(AgC组)分别经尾静脉注射miR-378抑制剂antagomir、激动剂agomir、antagomir阴性对照、agomir阴性对照溶液100μl(10nmol/ml),24 h后制备肠缺血再灌注损伤模型.再灌注结束后观察小肠组织病理学结果,并行Chiu评分,测定肠含水率和小肠黏膜乳酸含量.结果与S组比较,其余5组Chiu评分、肠含水率和肠黏膜乳酸含量升高(P＜0.01).与Ⅰ/R组比较,An组Chiu评分、肠含水率和肠黏膜乳酸含量升高,Ag组上述指标降低(P＜0.05),AnC组及AgC组上述指标差异无统计学意义(P＞0.05).An组肠组织病理学损伤较Ⅰ/R组加重,Ag组肠组织病理学损伤较Ⅰ/R组减轻.miRNA芯片分析共筛选出19个差异表达的miRNA,其中miR-292-3p表达上调,包括miR-378在内的18个miRNA表达下调(P＜0.05).qRT-PCR验证9个miRNA表达下调(P＜0.05或0.01),与芯片分析结果一致.结论大鼠肠缺血再灌注时肠组织有19个miRNA表达发生变化,且明确miR-378表达下凋与大鼠肠缺血再灌注损伤有关.
목적 탐토대서장결혈재관주시장조직미소RNA(miRNA)표체적변화.방법 성년웅성SD대서12지,체중230～250 g,채용수궤수자표법분위2조(n=6):가수술조(S조)화결혈재관주조(Ⅰ/R조).채용조단장계막상동맥60 mim재관주120 min적방법제비장결혈재관주손상모형.재관주결속후관찰소장조직병이학결과,행Chiu평분,측정소장함수솔화장점막조직유산함량.대장점막조직진행miRNA표체보심편분석,채용qRT-PCR법험증차이표체적miRNA.령취성년웅성SD대서24지,체중230～ 250 g,채용수궤수자표법분위4조(n=6):miR-378억제제조(An조)、miR-378격동제조(Ag조)、억제제대조조(AnC조)화격동제대조조(AgC조)분별경미정맥주사miR-378억제제antagomir、격동제agomir、antagomir음성대조、agomir음성대조용액100μl(10nmol/ml),24 h후제비장결혈재관주손상모형.재관주결속후관찰소장조직병이학결과,병행Chiu평분,측정장함수솔화소장점막유산함량.결과 여S조비교,기여5조Chiu평분、장함수솔화장점막유산함량승고(P＜0.01).여Ⅰ/R조비교,An조Chiu평분、장함수솔화장점막유산함량승고,Ag조상술지표강저(P＜0.05),AnC조급AgC조상술지표차이무통계학의의(P＞0.05).An조장조직병이학손상교Ⅰ/R조가중,Ag조장조직병이학손상교Ⅰ/R조감경.miRNA심편분석공사선출19개차이표체적miRNA,기중miR-292-3p표체상조,포괄miR-378재내적18개miRNA표체하조(P＜0.05).qRT-PCR험증9개miRNA표체하조(P＜0.05혹0.01),여심편분석결과일치.결론 대서장결혈재관주시장조직유19개miRNA표체발생변화,차명학miR-378표체하조여대서장결혈재관주손상유관.
Objective To investigate the changes in the expression of intestinal microRNAs (miRNAs) during intestinal ischemia-reperfusion (Ⅰ/R) in rats.Methods Twelve adult male SpragueDawley rats,weighing 230-250 g,were randomly divided into 2 groups (n =6 each) using a random number table:sham operation group (group S) and Ⅰ/R group.Intestinal Ⅰ/R was induced by 60 min occlusion of superior mesenteric artery followed by 120 min reperfusion in group Ⅰ/R.At the end of reperfusion,the intestinal specimens were obtained for microscopic examination of pathological changes which were graded using Chiu's scoring system and for determination of small intestinal water content and intestinal mucosal lactic acid content,miRNA microarray chip analysis was performed in the intestinal mucosal tissues,and quantitative real-time PCR was used to verify the differentially expressed miRNAs.Another 24 adult male Sprague-Dawley rats,weighing 230-250 g,were randomly divided into 4 groups (n =6 each) using a random number table:miR-378 inhibitor antagomir group (group An),miR-378 agonist agomir group (group Ag),inhibitor control group (group AnC) and agonist control group (group AgC).Antagomir,agomir,antagomir negative control and agomir negative control solutions l00 μl (10 nmol/ml) were injected via the tail vein in An,Ag,AnC and AgC groups,respectively.Intestinal Ⅰ/R was then produced 24 h later.The intestinal specimens were obtained for microscopic examination of pathological changes which were graded using Chiu's scoring system and for determination of small intestinal water content and intestinal mucosal lactic acid content.Results Compared with group S,Chiu's score,intestinal water content and intestinal mucosal lactic acid content were significantly increased in the other 5 groups.Compared with group Ⅰ/R,Chiu's score,intestinal water content and intestinal mucosal lactic acid content were were significantly increased in group An,the parameters mentioned above were decreased in group Ag,and no significant change was found in the parameters mentioned above in AnC and AgC groups.The pathological damage was aggravated in group An and was attenuated in group Ag as compared with group Ⅰ/R.miRNA microarray chip analysis revealed that a total of 19 miRNAs were found to be differentially expressed.Among the 19 differentially expressed miRNAs,the expression of miR-292-3p was significantly up-regulated,and the expression of 18 miRNAs including miR-378 was down-regulated.The expression of 9 miRNAs verified by quantitative real-time PCR was down-regulated,which was consistent with the results of miRNA microarray chip analysis.Conclusion Nineteen miRNA expression changes after Ⅰ/R,and it is confirmed that down-regulation of miR-378 expression is related to the intestinal Ⅰ/R injury in rats.