异丙酚对肝冷缺血再灌注所致大鼠肾损伤的影响

发布时间：2018-04-08 21:36

本文选题：异丙酚　切入点：缺血再灌注损伤　出处：《天津医科大学》2017年硕士论文

【摘要】：目的:临床上肝移植或其他复杂肝脏外科手术中必然会涉及肝缺血再灌注这一病理过程,肝缺血再灌注不仅能够损害肝脏本身,还可能诱发远隔脏器损伤,严重影响患者生存预后。如何保护远隔脏器,并且减少再灌注后损伤的发生,已成为近年来研究的热点。本研究主要研究大鼠肝冷缺血再灌注过程中JAK/STAT信号通路相关蛋白的表达情况,并探讨异丙酚能否通过调节JAK/STAT信号通路对肝冷缺血再灌注后肾损伤产生影响,以揭示肝冷缺血再灌注诱发肾损伤过程中的可能机制,为肝缺血再灌注后远隔脏器损伤的防治提供新思路。方法:SD成年雄性健康大鼠,周龄8~10周,体重220~250 g,随机分为4组(n=8):假手术组(Sham组);肝冷缺血再灌注改良模型组(I/R组);异丙酚组(Pro组);JAK2激酶抑制剂AG490组(AG490组)。Pro组大鼠行右侧股静脉置管连接异丙酚微量输液泵后,于再灌注前5 min泵注异丙酚20 mg·kg-1·h-1持续30 min。AG490组于建立模型前30 min腹腔注射AG490 10 mg/kg。再灌注6 h后,经下腔静脉采集血样后处死大鼠,使用全自动生化分析仪检测大鼠血清BUN、Cr含量变化,ELISA法测定TNF-α和IL-6的浓度。取肾组织标本,测定肾组织MDA含量及SOD水平。HE染色与TUNEL法观察肾组织病理学变化与细胞凋亡情况,并对肾小管损伤程度进行评分、计算凋亡指数。免疫组织化学法检测肾组织细胞内Cleaved Caspase-3的表达情况;Western blot法检测p-JAK2、p-STAT1和p-STAT3的蛋白表达水平。应用SPSS21.0统计软件分析实验结果,P0.05具有统计学差异。结果:与Sham组比较,I/R组大鼠肾功能衰退明显,血清BUN和Cr浓度明显升高,炎症反应及氧化应激增强,血清TNF-α、IL-6浓度和肾组织MDA含量明显升高,SOD活性降低(P0.05)。光镜下可见I/R组大鼠多数肾小管上皮细胞水肿、空泡变性,肾小管扩张,管腔狭窄,炎症细胞浸润,间质扩张淤血,肾小管损伤评分较Sham组显著升高(P0.05)。Sham组肾组织细胞TUNEL标记的凋亡细胞较少,而I/R组肾组织凋亡细胞显著增多,凋亡指数明显升高,且肾组织细胞中Cleaved Caspase-3的表达水平升高(P0.05)。I/R组大鼠肾组织p-JAK2、p-STAT1和p-STAT3的蛋白水平显著上调(P0.05)。与I/R组比较,Pro组及AG490组大鼠肾功能明显改善,血清BUN、Cr浓度明显降低,炎症反应减轻,血清TNF-α、IL-6的浓度显著降低,氧化应激缓解,肾组织MDA含量降低且SOD活性升高(P0.05)。Pro组与AG490组大鼠肾脏病理损伤较I/R组明显减轻,肾小管损伤评分降低,TUNEL标记的肾组织凋亡细胞数量减少,凋亡指数显著降低,且肾组织中Cleaved Caspase-3的表达明显减少(P0.05)。与此同时,Pro组及AG490组p-JAK2、p-STAT1和p-STAT3蛋白水平与I/R组比较显著下调(P0.05)。Pro组与AG490组各项指标差异无统计学意义。结论:(1)肝冷缺血再灌注可导致大鼠肾组织损伤,其机制与炎症反应、氧化应激和细胞凋亡等多种因素有关;(2)肝冷缺血再灌注能够激活JAK/STAT信号通路,抑制此通路可显著减轻肝冷缺血再灌注导致的肾损伤;(3)异丙酚能够减轻大鼠肝冷缺血再灌注导致的肾损伤,机制可能与抑制JAK/STAT信号通路从而减轻氧化应激、缓解炎症反应,抑制肾组织细胞凋亡有关。
[Abstract]:Objective: the clinical liver transplantation or other complex liver surgery must be involved in hepatic ischemia reperfusion of the pathological process, hepatic ischemia reperfusion can damage the liver itself, may also induce distant organ damage, seriously affect the prognosis. How to protect the remote organs, and reduce the occurrence of injury after reperfusion has become the research hotspot in recent years. The expression of JAK/STAT signaling pathway related protein in the research of cold ischemia reperfusion in rat liver, and to explore the effect of propofol can produce kidney injury by regulating JAK/STAT signal pathway on liver ischemia reperfusion injury, in order to reveal the possible mechanism of liver cold ischemia and reperfusion induced renal injury in the process and provide new ideas for the prevention of remote organ injury after liver ischemia reperfusion in rats. Methods: SD rats, 8~10 weeks, weighing 220~250 g, were randomly divided into The 4 group (n=8): sham operation group (group Sham); liver cold ischemia reperfusion model group (group I/R); Propofol group (Pro group); JAK2 kinase inhibitor AG490 group (group AG490).Pro group rats underwent right femoral vein catheter connected with propofol infusion pump, before reperfusion 5 min pump propofol 20 mg - kg-1 - H-1 for 30 min.AG490 group to model 30 min before intraperitoneal injection of AG490 10 mg/kg. 6 h after reperfusion, blood samples were collected from the inferior vena cava after rats were sacrificed, using automatic biochemical analyzer to detect the serum BUN, Cr content, determination of TNF- alpha and IL-6 ELISA method. Renal tissue samples, determination of renal tissue MDA content and SOD level of.HE and TUNEL staining method was used to observe the pathological changes and cell apoptosis, and the score of the degree of renal tubular injury, apoptosis index is calculated. The expression of immunohistochemical detection of kidney cells Cleaved Caspase-3 P-JAK2 Western blot; detection method, the expression level of p-STAT1 and p-STAT3 protein. The results were analyzed by SPSS21.0 statistical software, P0.05 has a statistically significant difference. Results: compared with Sham group, the renal function of I/R rats decline significantly, serum BUN and Cr were significantly increased, inflammatory reaction and oxidative stress increased, serum alpha TNF- the content of MDA, IL-6 concentration and renal tissue were significantly increased, SOD activity decreased (P0.05). Under visible edema, rats in the I/R group the majority of renal tubular epithelial cells light vacuolar degeneration, tubular dilatation, stenosis, inflammatory cell infiltration, interstitial congestion expansion, renal tubular injury score were significantly higher than that of Sham group (P0.05) a few apoptotic cells in renal tissues in.Sham group of TUNEL labeled cells, and apoptosis in the I/R group of renal tissue increased significantly, apoptosis index was significantly increased, and the expression of Cleaved in renal tissue cells in water Ping Shenggao Caspase-3 (P0.05) in kidney of rats in.I/R group P-JAK2, p-STAT1 protein level and p-STAT3 significantly increased (P0.05). Compared with I/R group, AG490 group and renal function of rats in group Pro were significantly improved, serum BUN, Cr concentrations were significantly decreased, inflammatory reaction, serum TNF-, IL-6 concentrations were significantly decreased, alleviate oxidative stress, MDA content of renal tissue decreased the increased activity of SOD (P0.05).Pro group and AG490 group rat kidney pathological damage significantly reduced compared with group I/R, renal tubular injury score decreased, TUNEL marked the number of apoptotic cells in renal tissue decreased, the apoptosis index decreased significantly, and the expression of Cleaved in renal tissue Caspase-3 decreased significantly (P0.05). At the same time, Pro group and AG490 group. P-JAK2, p-STAT1 and p-STAT3 protein levels compared with I/R group significantly decreased (P0.05) there was no significant difference between.Pro group and AG490 group index. Conclusion: (1) liver cold ischemia reperfusion can lead to kidney injury in rats, and the mechanism of anti inflammation Be related to various factors of oxidative stress and apoptosis; (2) liver cold ischemia reperfusion can activate JAK/STAT signaling pathway, inhibition of renal injury in this pathway can significantly reduce the liver cold ischemia reperfusion induced renal injury; (3) propofol can relieve the liver cold ischemia reperfusion in rats caused by the mechanism might be related to the inhibition the JAK/STAT signaling pathway to reduce oxidative stress, relieve inflammation, inhibit the apoptosis of renal tissue.

【学位授予单位】：天津医科大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R614

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