远程缺血预处理诱导脑保护的作用研究来自Notch、S-100B、NSE观察的证据

发布时间：2018-05-03 07:52

本文选题：远程缺血预处理 + 脑保护　；参考：《第四军医大学》2012年硕士论文

【摘要】：脑卒中(stroke)是一种突然起病的脑血液循环障碍性疾病。随着外科技术的推广应用，重要脏器的手术越来越多，围手术期脑卒中的发生率也随之增加，脑的缺血再灌注损伤是其最主要原因，虽然临床上目前使用很多措施但效果不理想。尤其是心脏手术后脑损伤发生率较高，表现为一系列的脑功能紊乱，如中风、脑病及认知功能下降等。其中中风的发生率为1%～3%，而认知功能障碍的发生率在术后1月可高达30%～65%，在术后5月仍有20%～40%。体外循环状态下脑的栓塞和血流灌注不足所致的脑缺血／再灌注损伤被认为是术后脑损伤的主要原因。近年研究发现肢体远程缺血预处理对脑和脊髓具有保护作用，可以明显减轻缺血再灌注导致的神经组织损伤。由于远程缺血预处理是一种简单、安全的预处理方式，与经典缺血预处理相比，更能方便安全地应用于临床患者。研究发现，生物体内普遍存在高度保守的Notch信号通路，Notch信号在神经系统中与神经元分化、成熟、再生、功能形成、以及神经系统疾病存在着密切的关系。那么，Notch信号通路是否参与了远程缺血预处理诱导神经保护过程？神经组织细胞在缺血远程缺血预处理过程中具有何种反应？这些都是有待阐明的相关机制问题。本研究对于以上问题进行了实验研究，提供了远程缺血预处理诱导脑保护机制的Notch、S-100B、NSE初步证据。实验一远程缺血预处理诱导脑保护与Notch分子表达变化的实验研究目的探讨Notch信号在远程缺血预处理诱导脑保护效应的可能参与作用。方法采用大脑中动脉阻闭（MCAO）法制备大鼠局灶性脑缺血模型，将36只雄性SD大鼠随机分为假手术组(Sham)、MCAO组和肢体远程缺血预处理（RIPC）组+MCAO组（n=10），分别观察各组大鼠脑梗死灶的大小和神经功能学评分，并用免疫组织化学染色以及Western blot、Real-Time PCR检测再灌注2h、24h、72h大鼠脑组织纹状体区Notch信号通路中NICD（Notch intracellular domain）及靶基因HES-1表达的变化。结果1.RIPC组脑梗死容积小于MCAO组，具有统计学差异（P＜0.05）。2.其神经功能学评分也明显优于MCAO组（P＜0.05）。3.免疫荧光：RIPC组2h、24hNICD表达明显少于MCAO组。4.Western blot：RIPC组NICD表达量在再灌注2h、24h降低，MCAO组NICD表达量在再灌注2h、24h强于同时间点RIPC组（P＜0.05），但两组均高于假手术组（P＜0.05）。5.Real-Time PCR：受体Notch-1mRNA表达在MCAO组再灌注2h、24h增高，RIPC组Notch-1表达量在再灌注2h、24h低于同时间点MCAO组（P＜0.05），，靶基因HES-1mRNA表达MCAO组在再灌注2h、24h逐步增高，RIPC组表达量在再灌注2h、24h低于同时间点MCAO组（P＜0.05）。但再灌注72h两组表达量无统计学差异。结论肢体远程缺血预处理对脑的缺血再灌注损伤具有保护作用，肢体远程预处理可以使Notch信号通路相关分子Notch-1、NICD、HES-1表达发生变化，肢体远程预处理诱导脑缺血耐受的机制可能与Notch信号表达下降有关。实验二远程缺血预处理对心脏围手术期脑缺血再灌注损伤保护作用研究目的观察远程缺血预处理对心脏手术体外循环（CPB）所致脑缺血再灌注损伤的保护作用及其检测评价。方法80例择期行心脏瓣膜置换术患者在麻醉诱导前随机分成两组：肢体远程缺血预处理组和对照组，每组各40例。肢体远程缺血预处理使用充气式止血带对右上肢实施3次5分钟缺血、5分钟再灌注，充气压力为200mmHg。检测麻醉诱导前、CPB前、术毕及开放后6、24、48、72h血清S-100B和神经元特异性烯醇化酶（NSE）浓度。术前及术后1W、3M、6M进行认知功能评估。结果远程缺血预处理组开放后6、24、48、72h血清S-100B和开放后24、48、72h的NSE释放明显减少（P＜0.05）。尽管两组患者认知功能评估虽不统计学差异，但远程预处理组评分优于对照组。结论肢体远程缺血预处理减轻心脏围手术期患者神经组织细胞损伤引起的血清S-100B和NSE释放，提示其对缺血再灌注患者的脑组织可能具有一定的保护作用。
[Abstract]:Cerebral apoplexy (stroke) is a sudden onset of cerebral blood circulation disorder. With the popularization and application of surgical techniques, the operation of important organs is more and more, the incidence of cerebral apoplexy in the perioperative period is increasing, and cerebral ischemia reperfusion injury is the most important reason. Although many measures are used at present, the effect is not satisfactory. The incidence of brain injury after cardiac surgery is high, showing a series of cerebral dysfunction, such as stroke, encephalopathy and cognitive decline, among which the incidence of stroke is 1% to 3%, and the incidence of cognitive dysfunction can reach 30% ~ 65% in January after operation, and there is still 20% ~ 40%. of cerebral embolism and blood flow irrigation in May after operation. Cerebral ischemia / reperfusion injury caused by insufficient injection is considered to be the main cause of postoperative brain injury.
In recent years, remote ischemic preconditioning has been found to have protective effects on the brain and spinal cord, which can obviously reduce the nerve tissue damage caused by ischemia-reperfusion. Because remote ischemic preconditioning is a simple and safe preconditioning method, it is more convenient and safe to apply to clinical patients compared with classical ischemic preconditioning. There are highly conserved Notch signaling pathways in the body. Notch signals are differentiated from neurons in the nervous system, maturation, regeneration, function formation, and nervous system diseases. Is the Notch signaling pathway involved in the long distance ischemic preconditioning induced neuroprotection process? Neural tissue cells are short of ischemia. What is the reaction in the process of blood preconditioning? These are relevant mechanisms to be elucidated. This study has conducted experimental studies on the above problems and provided preliminary evidence of Notch, S-100B, and NSE for the mechanism of brain protection induced by remote ischemic preconditioning.
Experimental study on the changes of Notch expression induced by remote ischemic preconditioning
Objective to investigate the possible role of Notch signaling in remote ischemic preconditioning induced brain protection.
Methods the rat model of focal cerebral ischemia was prepared by middle cerebral artery occlusion (MCAO) method. 36 male SD rats were randomly divided into sham operation group (Sham), group MCAO and +MCAO group (n=10) of limb remote ischemic preconditioning (RIPC) group. The size of cerebral infarction and the score of neurologic function were observed in each group, and immunohistochemical staining was used. Western blot and Real-Time PCR were used to detect the changes of NICD (Notch intracellular domain) and the expression of target genes in the Notch signaling pathway in the striatum of the brain tissue of the rats with 2H, 24h and 72h. Results the volume of cerebral infarction was less than that of the group. Immunofluorescence: the expression of 2h, 24hNICD in group RIPC was significantly less than that in group.4.Western blot:RIPC of group MCAO, NICD expression was reduced to 2h, 24h decreased, NICD expression of MCAO group was in reperfusion 2h, but the two groups were higher than those in the sham group (0.05). 24h increased, the expression of Notch-1 in group RIPC was 2h, 24h was lower than that in group MCAO (P < 0.05) at the same time point (P < 0.05). The expression of target gene HES-1mRNA in MCAO group was gradually increased in 2H and 24h, and the expression of RIPC group was lower than that in the same time. There was no statistical difference between the two groups. It has protective effect on cerebral ischemia reperfusion injury. Remote preconditioning can change the expression of Notch-1, NICD, and HES-1 in Notch signaling pathway. The mechanism of remote preconditioning induced cerebral ischemia tolerance may be related to the decrease of Notch signal expression.
Experiment two the protective effect of remote ischemic preconditioning on cerebral ischemia-reperfusion injury during perioperative period
Objective To observe the protective effect of remote ischemic preconditioning on cerebral ischemia reperfusion injury induced by cardiac surgery extracorporeal circulation (CPB). Methods 80 patients undergoing elective cardiac valve replacement were randomly divided into two groups before induction of anesthesia: limb remote ischemic preconditioning group and treatment group, 40 cases in each group. The inflatable tourniquet was used for 3 times 5 minutes of ischemia, 5 minutes of reperfusion, and inflatable pressure of 200mmHg. before anesthesia induction, before CPB, 6,24,48,72h serum S-100B and neuron specific enolase (NSE) concentration. Preoperative and postoperative 1W, 3M, 6M were evaluated for cognitive function. Results remote ischemic preconditioning group was open. The release of 6,24,48,72h serum S-100B and the release of NSE after open 24,48,72h decreased significantly (P < 0.05). Although the cognitive function assessment of the two groups was not statistically different, the distance preconditioning group was superior to the control group. Conclusion limb remote ischemic preconditioning alleviated the serum S-100B and NS caused by the injury of nerve tissue cells in the perioperative patients. The release of E suggests that it may have some protective effects on cerebral tissue in patients with ischemia-reperfusion.

【学位授予单位】：第四军医大学
【学位级别】：硕士
【学位授予年份】：2012
【分类号】：R363

【参考文献】