【作者】 申发平；

【导师】 孙保亮；

【作者基本信息】 泰山医学院 ， 老年医学， 2009， 硕士

【摘要】 研究目的:1、探讨CGRP经鼻给药后对SAH后CVS的缓解作用。2、探讨经鼻应用CGRP对SAH后血管新生的影响和机制。研究方法:1、选用健康成年雄性Wister大鼠,采用枕大池二次注血法建立SAH模型,将动物随机分为正常对照组、SAH组、经鼻NS SAH组、经鼻CGRP SAH组。用体式显微镜及活体循环检测系统在第二次SAH 72h观察基底动脉(BA)管径;用激光多普勒血流计在第一次SAH制作前,第二次SAH鼻腔给药后1/2h,1h,6h,12h各测量一次顶叶皮层局部脑血流量(rCBF),对大鼠局部脑血流量进行动态观察。2、建立大鼠SAH模型,将动物随机分为正常对照组、SAH组、经鼻NS SAH组、经鼻CGRP SAH组。于第二次SAH后72h,制备新鲜脑组织冠状冰冻切片,用荧光免疫组化法检测皮层和海马CD31的表达,用图像分析仪计数新生血管密度;用逆转录RT-PCR技术测定大脑皮层、海马等部位血管内皮生长因子(vascular endothelial growth factor,VEGF)mRNA表达;用West blot、免疫荧光法检测皮层和海马VEGF的表达。研究结果:1、各组动物第二次SAH后72h,SAH组和NS SAH组大鼠BA明显痉挛,管径缩窄严重,与正常对照组有显著性差异(P<0.01);CGRP SAH组大鼠BA轻微痉挛,管径挛缩不明显,与SAH组和NS SAH组有显著性差异(P<0.01)。各组大鼠BA管径分别为正常对照组的48.18%、47.69%、93.13%。第二次SAH后SAH组和NS SAH组,rCBF显著降低,与正常对照组有显著性差异(P<0.01);CGRP SAH组rCBF降低的程度小,与SAH组和NS SAH组有显著性差异(P<0.01)。各组在第二次SAH后rCBF降低,1 h下降至最低值,在12 h内无明显恢复趋势。2、第二次SAH后72h,SAH组、经鼻NS SAH组与正常对照组比较皮层和海马CD31、VEGF表达增强,有较多阳性细胞(P<0.01);经鼻CGRP SAH组表达最强,有大量的免疫阳性细胞分部,与SAH组和NS SAH组有显著性差异(P<0.01)。SAH组和NS SAH组与正常对照组比较皮层和海马VEGF mRNA、VEGF蛋白表达增加(P<0.01);经鼻CGRP SAH组表达最强,与SAH组和NS SAH组有显著性差异(P<0.01)。研究结论:1、经鼻给予CGRP能缓解SAH后BA血管痉挛,增加顶叶皮层局部脑血流量(rCBF),改善SAH后脑循环,增加脑部血液供应,对SAH后CVS有缓解作用。2、经鼻给予CGRP能促进大脑皮层和海马CD31表达,增加大脑皮层和海马VEGF mRNA及VEGF蛋白表达,增强VEGF分泌和释放,通过VEGF、CD31血管新生作用,促进SAH后血管生成,增加脑部血流量,改善脑循环,防治SAH后CVS。更多还原

【Abstract】 OBJECTIVES:1.To investigate the effect of CGRP given intranasally on relief CVS following SAH.2.To investigate the effect and mechanism of CGRP given intranasally on new cerebral vessel generation after SAH.METHOD:1. In this study, adult male Wister rats were used as experimental animals. Cisterna injection twice of freshly autologous arterial blood was used to induce SAH in rats. The rats were divided into : normal control group, SAH group, per nasal NS SAH group, per nasal CGRP SAH group. Somatotype microscope and living body circulation measure system were used to observe the caliber of BA in 72h after second SAH ; To measure parietal cortex rCBF in before first SAH, in 1/2h, 1h, 6h, 12h after second SAH given intranasally by laser-Doppler flow meter, developmently observe regional cerebral blood floe of rats.2. The SAH model in rats was established , the rats were divided into : normal control group, SAH group, per nasal NS SAH group, per nasal CGRP SAH group. Rats were sacrificed at 72h after second SAH, the brain were taken out and made into the coronal frozen section. Immunohistochemical staining was used to detect the expression of CD31 and VEGF in cortex and hippocamp respectively; the expression of VEGF mRNA and the protein expression of VEGF in cortex and hippocamp of brain were assessed by technic of TR-PCR and West blot respectively.RESULT: 1.In 72h after second SAH, the BA in SAH group and NS SAH group obviously cramped, the caliber of BA was significantly coarctat compared with that in normal group (P<0.01); The BA in CGRP SAH group slighely cramped, the caliber of BA was lightly coarctat compared with that in SAH group and NS SAH group(P<0.01); the BA caliber of every group is 48.18% 47.69% 93.13% of normal group respectively. The rCBF in SAH group and NS SAH group was evidently degraged compared with normal group(P<0.01); the rCBF in CGRP SAH group was smally step down compared with SAH group and NS SAH group (P<0.01); The rCBF cut down after second SAH every group, nadir in 1h, did not obviously return within 12h.2. In 72h after second SAH, the expression of CD31, VEGF enhance respectively in cortex and hippocamp in SAH group and NS SAH group that have more macs cell compared with normal group(P<0.01); the expression of CD31, VEGF in cortex and hippocamp in CGRP SAH group that abound with immue masculin cell is the most obvious contrasted to SAH group and NS SAH group (P<0.01); the expression of VEGF mRNA and the protein expression of VEGF in cortex and hippocamp in SAH group and NS SAH group increased respectively, cnotrast with that in normal group (P<0.01); the expression of VEGF mRNA and the protein expression of VEGF in cortex and hippocamp in CGRP SAH group was the most maximun respectively, cnotrast with that in SAH group and NS SAH group (P<0.01).CONCLUSION:1.The CGRP via intranasal pathway can relieve BA vasospasm after SAH, increase rCBF, improve cerebral circulation, raise brain blood provision, play an important role in releasing CVS following SAH.2. The CGRP given intranaseny can stimulate CD31 expression in cortex and hippocamp, increase the density of new blood vessels, enhance the expression of VEGF cell VEGF mRNA VEGF protein, VEGF that can promote new blood generation is bloodvessel regulatory factor. Thus, the CGRP via intranasal pathway can promote new cerebral vessels generation, increase cerebral blood flow , improve cerebral circulation.更多还原