p-Akt和caspase-3参与远程创伤预处理脑保护作用
发布时间：2019-02-21 08:56
【摘要】：目的：采用大鼠局灶性脑缺血再灌注模型探讨远程创伤预处理(remote preconditioning of trauma, RPCT)对脑缺血再灌注损伤的影响,研究p-Akt和caspase-3在远程创伤预处理脑保护机制中的作用。 方法：(一)70只健康雄性SD大鼠,体重250-300g,随机分成七组(n=10)：假手术组(Sham组)、缺血再灌注组(I/R组)、RPCT0.5h组、RPCT1h组、RPCT2h组、RPCT4h组和RPCT24h组。用以测定脑梗死容积及神经功能缺损评分。(二)40只健康雄性SD大鼠随机分成四组(n=10):Sham、I/R组、RPCT1h组和RPCT24h组。用以组织形态学、免疫组化及Western Blot法检测。①Sham组：仅分离大鼠颈部血管,不插入拴线阻塞大脑中动脉。②I/R组：采用Longa线栓法制作MCAO模型方法建立局灶性脑缺血再灌注模型。③RPCT组：在MCAO开始前分别0.5h、1h、2h、4h和24h做2cm腹部正中切口并切开腹壁全层,随即缝合伤口。各组大鼠均于缺血再灌注后观察24h,在24h对大鼠行神经功能缺损评分；行TTC染色法测定脑梗死容积；HE染色法观察脑组织形态；Nissl染色法观察神经细胞存活情况；免疫组化及Western Blot法观察并测定缺血半影区Akt、p-Akt及caspase-3蛋白的表达。 结果：①TTC染色：Sham组未见脑梗死；与I/R组比较,RPCT1h组、RPCT2h组、RPCT4h组及RPCT24h组脑梗死容积显著减小(P0.05),RPCT0.5h组与I/R组脑梗死容积无显著差异(P0.05)。②神经功能缺损评分：RPCT1h组、RPCT2h组、RPCT4h组和RPCT24h组Garcia评分较I/R组显著提高(P0.05),RPCT0.5h组与I/R组Garcia评分无显著差异；各组Longa五分制评分无显著差异(P0.05)。③HE染色：Sham组大鼠大脑皮质神经元形态正常；与I/R组相比,RPCT1h组及RPCT24h组缺血半影区神经元数目较多,层次较清楚,细胞形态较完整,细胞间质水肿减轻；I/R组、RPCT1h组及RPCT24h组缺血中心区均出现细胞坏死,神经元数目减少,核固缩深染,组织疏松。④Nissl染色：Sham组大脑皮质神经元形态正常；与I/R组相比,RPCT1h组及RPCT24h组缺血半影区存活神经元数量显著增加(P0.05)。⑤免疫组化及Western Blot结果：Akt在缺血半影区表达稳定,各组间无统计学差异(P0.05)；I/R组、RPCT1h组及RPCT24h组p-Akt蛋白和caspase-3蛋白在缺血半影区的表达均显著高于Sham组(P0.05)；与I/R组相比,RPCT1h组及RPCT24h组缺血半影区的p-Akt表达显著增加(P0.05),而caspase-3表达显著减少(P0.05)。 结论：远程创伤预处理能够减轻大鼠局灶性脑缺血再灌注损伤,其机制可能与激活Akt蛋白、抑制caspase-3表达有关。
[Abstract]:Aim: to investigate the effects of remote preconditioning (remote preconditioning of trauma, RPCT) on cerebral ischemia-reperfusion injury in rats with focal cerebral ischemia-reperfusion, and to investigate the role of p-Akt and caspase-3 in the protective mechanism of remote traumatic preconditioning (RIP). Methods: (1) 70 healthy male SD rats, weighing 250-300 g, were randomly divided into seven groups: sham operation group (Sham group), ischemia reperfusion group (I / R group), RPCT0.5h group, RPCT1h group, RPCT2h group, RPCT4h group and RPCT24h group. To determine the volume of cerebral infarction and neurological impairment score. (2) 40 healthy male SD rats were randomly divided into four groups: Sham,I/R group, RPCT1h group and RPCT24h group. For histomorphology, immunohistochemistry and Western Blot detection. 1Sham group: isolated only the cervical vessels of rats, Middle cerebral artery occlusion was not inserted into the middle cerebral artery. 2I/R group: the model of focal cerebral ischemia-reperfusion was established by using Longa thread occlusion method. The 3RPCT group: 0.5 h before the start of MCAO, 1 h and 2 h, respectively, in the 3RPCT group, the model of focal cerebral ischemia and reperfusion was established by using the method of Longa thread embolization. 2cm median abdominal incision was performed at 4 h and 24 h, and the whole abdominal wall was cut. Then the wound was sutured. All the rats in each group were observed for 24 hours after ischemia and reperfusion. The neurological deficit score was evaluated at 24 hours; the volume of cerebral infarction was measured by TTC staining; the morphology of brain tissue was observed by HE staining; the survival of nerve cells was observed by Nissl staining. The expression of Akt,p-Akt and caspase-3 in ischemic penumbra was observed by immunohistochemistry and Western Blot method. Results: 1TTC staining: no cerebral infarction was found in Sham group. Compared with I / R group, cerebral infarction volume in RPCT1h group, RPCT2h group, RPCT4h group and RPCT24h group was significantly decreased (P0.05), but there was no significant difference in cerebral infarction volume between RPCT0.5h group and I / R group (P0.05). The Garcia score of RPCT4h and RPCT24h group was significantly higher than that of I / R group (P0.05). There was no significant difference in Garcia score between RPCT0.5h group and I / R group. There was no significant difference in Longa score in each group (P0.05). 3HE staining: the cerebral cortex neurons in Sham group were normal. Compared with I / R group, RPCT1h group and RPCT24h group had more neurons in ischemic penumbra, more clear layers, more complete cell morphology and less interstitial edema. In I / R group, RPCT1h group and RPCT24h group, there were cell necrosis, decreased number of neurons, deep staining of nucleus pyknosis and loose tissue. 4Nissl staining: the morphology of cerebral cortex neurons in Sham group was normal. Compared with I / R group, the number of surviving neurons in ischemic penumbra of RPCT1h group and RPCT24h group was significantly increased (P0.05). 5 the results of immunohistochemistry and Western Blot showed that the expression of Akt was stable in ischemic penumbra area, and there was no statistical difference among the groups (P0.05). The expression of p-Akt protein and caspase-3 protein in I / R group, RPCT1h group and RPCT24h group were significantly higher than those in Sham group (P0.05). Compared with I / R group, the expression of p-Akt in ischemic penumbra of RPCT1h group and RPCT24h group was significantly increased (P0.05), while the expression of caspase-3 was significantly decreased (P0.05). Conclusion: remote trauma preconditioning can attenuate focal cerebral ischemia-reperfusion injury in rats, and its mechanism may be related to activation of Akt protein and inhibition of caspase-3 expression.
【学位授予单位】：中南大学
【学位级别】：硕士
【学位授予年份】：2014
【分类号】：R743.3
本文编号：2427405
[Abstract]:Aim: to investigate the effects of remote preconditioning (remote preconditioning of trauma, RPCT) on cerebral ischemia-reperfusion injury in rats with focal cerebral ischemia-reperfusion, and to investigate the role of p-Akt and caspase-3 in the protective mechanism of remote traumatic preconditioning (RIP). Methods: (1) 70 healthy male SD rats, weighing 250-300 g, were randomly divided into seven groups: sham operation group (Sham group), ischemia reperfusion group (I / R group), RPCT0.5h group, RPCT1h group, RPCT2h group, RPCT4h group and RPCT24h group. To determine the volume of cerebral infarction and neurological impairment score. (2) 40 healthy male SD rats were randomly divided into four groups: Sham,I/R group, RPCT1h group and RPCT24h group. For histomorphology, immunohistochemistry and Western Blot detection. 1Sham group: isolated only the cervical vessels of rats, Middle cerebral artery occlusion was not inserted into the middle cerebral artery. 2I/R group: the model of focal cerebral ischemia-reperfusion was established by using Longa thread occlusion method. The 3RPCT group: 0.5 h before the start of MCAO, 1 h and 2 h, respectively, in the 3RPCT group, the model of focal cerebral ischemia and reperfusion was established by using the method of Longa thread embolization. 2cm median abdominal incision was performed at 4 h and 24 h, and the whole abdominal wall was cut. Then the wound was sutured. All the rats in each group were observed for 24 hours after ischemia and reperfusion. The neurological deficit score was evaluated at 24 hours; the volume of cerebral infarction was measured by TTC staining; the morphology of brain tissue was observed by HE staining; the survival of nerve cells was observed by Nissl staining. The expression of Akt,p-Akt and caspase-3 in ischemic penumbra was observed by immunohistochemistry and Western Blot method. Results: 1TTC staining: no cerebral infarction was found in Sham group. Compared with I / R group, cerebral infarction volume in RPCT1h group, RPCT2h group, RPCT4h group and RPCT24h group was significantly decreased (P0.05), but there was no significant difference in cerebral infarction volume between RPCT0.5h group and I / R group (P0.05). The Garcia score of RPCT4h and RPCT24h group was significantly higher than that of I / R group (P0.05). There was no significant difference in Garcia score between RPCT0.5h group and I / R group. There was no significant difference in Longa score in each group (P0.05). 3HE staining: the cerebral cortex neurons in Sham group were normal. Compared with I / R group, RPCT1h group and RPCT24h group had more neurons in ischemic penumbra, more clear layers, more complete cell morphology and less interstitial edema. In I / R group, RPCT1h group and RPCT24h group, there were cell necrosis, decreased number of neurons, deep staining of nucleus pyknosis and loose tissue. 4Nissl staining: the morphology of cerebral cortex neurons in Sham group was normal. Compared with I / R group, the number of surviving neurons in ischemic penumbra of RPCT1h group and RPCT24h group was significantly increased (P0.05). 5 the results of immunohistochemistry and Western Blot showed that the expression of Akt was stable in ischemic penumbra area, and there was no statistical difference among the groups (P0.05). The expression of p-Akt protein and caspase-3 protein in I / R group, RPCT1h group and RPCT24h group were significantly higher than those in Sham group (P0.05). Compared with I / R group, the expression of p-Akt in ischemic penumbra of RPCT1h group and RPCT24h group was significantly increased (P0.05), while the expression of caspase-3 was significantly decreased (P0.05). Conclusion: remote trauma preconditioning can attenuate focal cerebral ischemia-reperfusion injury in rats, and its mechanism may be related to activation of Akt protein and inhibition of caspase-3 expression.
【学位授予单位】：中南大学
【学位级别】：硕士
【学位授予年份】：2014
【分类号】：R743.3
【参考文献】
相关期刊论文 前2条
1 ;Greater stress protein expression enhanced by combined prostaglandin A_1 and lithium in a rat model of focal ischemia[J];Acta Pharmacologica Sinica;2007年08期
2 Uwe Grutzner;Melanie Keller;Michael Bach;Alexandra K Kiemer;Herbert Meissner;Manfred Bilzer;Stefan Zahler;Alexander L Gerbes;Angelika M Vollmar;;PI 3-kinase pathway is responsible for antiapoptotic effects of atrial natriuretic peptide in rat liver transplantation[J];World Journal of Gastroenterology;2006年07期
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