CAJ | 학술논문

背景:缺血预处理可诱发机体内源性保护机制,可全面有效地防治器官移植缺血再灌注损伤.在胰腺移植过程中冷、热缺血均可导致移植胰腺缺血再灌注损伤,线粒体结构及功能与胰腺病变密切相关,近些年研究发现,线粒体DNA存在修复体系,其与线粒体DNA损伤之间的平衡决定了疾病的发生和转归.目的:观察缺血预处理对大鼠胰腺移植缺血再灌注损伤时的细胞凋亡的影响,分析线粒体DNA修复酶8-氧鸟嘌呤DNA糖基化酶和氧化应激在其中的变化规律及可能途径.方法:纳入健康雄性SD大鼠50只,其中20只为供体,10只为假手术组,另20只糖尿病造模后分为缺血再灌注组和缺血预处理组,每组10只.假手术组只行开、关腹手术,缺血再灌注组和缺血预处理组行异位全胰十二指肠移植.缺血再灌注组对应供体大鼠于获取供胰前以4℃ UW液灌洗20 min:缺血预处理组对应供体大鼠于获取供胰前阻断腹上动脉5 min,再灌注5 min,共2次.供胰均控制热缺血时间为15 min,冷缺血时间为180 min.再灌注后12 h检测血浆淀粉酶活性、血糖浓度及Caspase-3,9活化水平,流式细胞法检测腺泡细胞凋亡率,罗丹明123法检测线粒体膜电位,二氯荧光素法检测线粒体过氧化氢产生速率,高效液相色谱法检测线粒体DNA中8-氧鸟嘌呤质量浓度,荧光定量聚合酶链反应法检测8-氧鸟嘌呤DNA糖基化酶mRNA的表达,Westenn-blotting法检测细胞色素C释放、磷酸化Akt及线粒体8-氧鸟嘌呤DNA糖基化酶蛋白表达水平.结果与结论:缺血预处理可降低线粒体氧化应激,提高Akt磷酸化水平,从而上调8-氧鸟嘌呤DNA糖基化酶表达,减少线粒体DNA氧化损伤,抑制腺泡细胞凋亡,减轻移植胰缺血再灌注损伤.
배경:결혈예처리가유발궤체내원성보호궤제,가전면유효지방치기관이식결혈재관주손상.재이선이식과정중랭、열결혈균가도치이식이선결혈재관주손상,선립체결구급공능여이선병변밀절상관,근사년연구발현,선립체DNA존재수복체계,기여선립체DNA손상지간적평형결정료질병적발생화전귀.목적:관찰결혈예처리대대서이선이식결혈재관주손상시적세포조망적영향,분석선립체DNA수복매8-양조표령DNA당기화매화양화응격재기중적변화규률급가능도경.방법:납입건강웅성SD대서50지,기중20지위공체,10지위가수술조,령20지당뇨병조모후분위결혈재관주조화결혈예처리조,매조10지.가수술조지행개、관복수술,결혈재관주조화결혈예처리조행이위전이십이지장이식.결혈재관주조대응공체대서우획취공이전이4℃ UW액관세20 min:결혈예처리조대응공체대서우획취공이전조단복상동맥5 min,재관주5 min,공2차.공이균공제열결혈시간위15 min,랭결혈시간위180 min.재관주후12 h검측혈장정분매활성、혈당농도급Caspase-3,9활화수평,류식세포법검측선포세포조망솔,라단명123법검측선립체막전위,이록형광소법검측선립체과양화경산생속솔,고효액상색보법검측선립체DNA중8-양조표령질량농도,형광정량취합매련반응법검측8-양조표령DNA당기화매mRNA적표체,Westenn-blotting법검측세포색소C석방、린산화Akt급선립체8-양조표령DNA당기화매단백표체수평.결과여결론:결혈예처리가강저선립체양화응격,제고Akt린산화수평,종이상조8-양조표령DNA당기화매표체,감소선립체DNA양화손상,억제선포세포조망,감경이식이결혈재관주손상.
BACKGROUND: Ischemic preconditioning (IPC) can induce endogenous protection mechanism, which effectively prevent ischemia/reperfusion injury following organ transplantation. Cold and warm ischemia may induce ischemia/reperfusion injury of pancreas transplantation, and apoptosis of pancreatic acinar cells is one of the important reasons of pancreas graft functional defect after transplantation. Mitochondrial DNA has repair system, and its balance with mitochondrial DNA injury influences disease occurrence and outcome.OBJECTIVE: To observe the effect of IPC on apoptosis of transplanted pancreatic acinar cells, and the possible role of reactive oxygen (ROS) and mitochondrial DNA repair enzyme.METHODS: A total of 50 health, male, Sprague-Dawley rats were randomly divided into three groups: sham operated (n = 10), donors (n = 20) and recipients (n = 20). The recipients were randomly divided into ischemia/reperfusion group (IR, n = 10) and IPC group (n = 10). The sham operated group was subjected to abdominal open and close operation. IR group and IPC group received establishment of diabetic model by streptozotocin injection. IR rats received whole pancreatic-duodenal transplantation alone. IPC rats received whole pancreatic-duodenal transplantation exposed ischemic preconditioning with 5 minutes ischemia and 5 minutes reperfusion twice. All grafts were keep with warm ischemia time 15 minutes and cold ischemia (in 4 ℃ UW preservation solution) time 180 minutes. Twelve hours after reperfusion, serum amylase, blood glucose, Caspase-3, -9 activity were detected. Pancreatic acinar cell apoptosis was measured by flow cytometry. Mitochondrial cross-membrane potential (Δψ) was measured by monitoring the fluorescence spectrum of rhodamine 123. Mitochondrial H2O2 generation was determined using dichlorofluorescein as a probe. 8-oxodG in mitochondrial DNA (mtDNA) was measured with HPLC system. Release of cytochrome C, phosphorylation of Akt and mitochondrial OGG1 protein expression were determined by Western-blotting. RESULTS AND CONCLUSION: The ischemia preconditioning can relieve the pancreatic acinar cell apoptosis in pancreas graft and relieve IR injury by decreasing mitochondrial oxidative stress, mtDNA injury, and increasing phosphorylation of Akt and mitochondrial OGG1 expression.