大鼠肺移植供肺缺血再灌注的自噬水平变化

发布时间：2017-12-26 19:40

  本文关键词：大鼠肺移植供肺缺血再灌注的自噬水平变化　出处：《南昌大学》2015年硕士论文　论文类型：学位论文

  更多相关文章： 自噬 肺移植 缺血再灌注损伤 大鼠

【摘要】：目的:实验通过建立大鼠原位肺移植模型,利用实验动物学及分子生物学的技术,阐明大鼠肺移植供肺灌注、冷缺血、再灌注后自噬水平变化,为调控自噬水平,减轻肺移植缺血再灌注损伤提供理论依据。方法:1、分组并建立模型:SPF级雄性SD大鼠,共60只,质量300~350g。随机挑选12只作单纯灌注冷缺血处理,其中6只作为单纯灌注冷缺血0h对照,另外6只作单纯灌注冷缺血后保存6h处理;其余48只分为供、受体,受体略大于供体,供、受体间质量差不超过20g,进行左肺移植手术。本实验共分五组:(1)CON组即灌注后,冷缺血保存0h组;(2)ISC6组即灌注后,冷缺血保存6h组;(3)REP2组即灌注后,先冷缺血保存6h,移植后再灌注2h组;(4)REP6组即灌注后,先冷缺血保存6h,移植后再灌注6h组;(5)REP12组即灌注后,先冷缺血保存6h,移植后再灌注12h组。其中CON,ISC6组各6例,REP2,REP6,REP12组各移植6对大鼠。2、分子生物学检测:免疫荧光双染LC3,LAMP-2,检测自噬流,Western blot检测LC3-II/LC3-I。结果:1、成功建立了一种经过改良了肺移植技术的大鼠原位左肺移植模型;2、通过免疫荧光双染LC3,LAMP-2,发现:CON组自噬流最低,ISC6组自噬流稍升高,REP2,REP6组自噬流显著升高,其中REP6组自噬流最高,REP12组自噬流降低。3、通过Western blot检测LC3蛋白表达发现:CON组LC3蛋白表达最低,ISC6组LC3蛋白表达稍升高,REP2,REP6组LC3蛋白表达显著升高,其中REP6组LC3蛋白表达最高,REP12组LC3蛋白表达降低。结论:大鼠肺移植供肺灌注、冷缺血6h后自噬水平开始增高,再灌注2h的自噬水平明显增高,再灌注6h达到高峰,再灌注12h后自噬减轻。
[Abstract]:Objective: through the establishment of experimental model of orthotopic lung transplantation in rats, the use of laboratory animal science and molecular biology technology, clarify the rat lung transplantation for pulmonary perfusion and cold ischemia after reperfusion, the level of autophagy changes, for the regulation of autophagy, alleviate the ischemia reperfusion injury in lung transplantation and provide a theoretical basis for the. Methods: 1, group and model: grade SPF male SD rats, with a total of 60 rats, with a mass of 300~350g. 12 randomly selected only for simple perfusion cold ischemia treatment, including 6 as simple perfusion of cold ischemia for 0h control, the other 6 only for simple perfusion cold ischemia preserved after 6h treatment; the remaining 48 were divided into donor and receptor, receptor is slightly larger than the donor between donor and recipient of poor quality does not exceed 20g, left lung transplantation surgery. The experiments were divided into five groups: (1) CON group after reperfusion, cold ischemia 0h group; (2) ISC6 group after reperfusion, cold ischemia 6h group; (3) REP2 group after reperfusion, first cold ischemia 6h, reperfusion after transplantation in 2H group; REP6 group (4) the first cold ischemia after reperfusion, 6h after transplantation, reperfusion 6h group; (5) REP12 group after reperfusion, first cold ischemia 6h, reperfusion after transplantation in 12h group. 6 cases in each group of CON, group ISC6, REP2, REP6, and REP12 were transplanted in 6 pairs of rats. 2, molecular biological detection: immunofluorescence double staining LC3, LAMP-2, detection of autophagic flow, Western blot detection of LC3-II/LC3-I. Results: 1, successfully established a modified rat model of orthotopic left lung transplantation and lung transplantation technology; 2, by double immunofluorescence staining LC3, LAMP-2, CON group found that autophagy flow is the lowest, ISC6 group REP2, autophagy flux was slightly higher in REP6 group significantly increased the flow of autophagy, autophagy group REP6 flow the highest REP12 group decreased autophagy flow. 3, the expression of LC3 protein was detected by Western blot. It was found that the expression of LC3 protein was the lowest in group CON, the expression of LC3 protein increased slightly in ISC6 group, and the expression of LC3 protein in REP2 and REP6 group increased significantly, among which the expression of LC3 protein in the REP6 group was the highest and the expression of group C was decreased. Conclusion: autophagy level is increased after lung transplantation and cold ischemia 6h in rat lung transplantation. The autophagy level of 2h after reperfusion is significantly higher than that of reperfusion, and the peak of 6h is reached after reperfusion, and autophagy is alleviated after 12h reperfusion.
【学位授予单位】：南昌大学
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：R655.3

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