【作者】 郭予大；

【导师】 游潮；

【作者基本信息】 四川大学 ， 外科学， 2005， 博士

【摘要】 血管壁细胞外基质的大量破坏是颅内动脉瘤形成的重要因素,而细胞外基质的破坏与基质金属蛋白酶(matrix metalloproteinase,MMPs)尤其MMP-9过度表达密切相关,同时伴有MMP的抑制剂(tissue inhibitor of metalloproteinase,TIMP)表达升高,但二者的平衡失调致使MMP-9的超量表达参与了颅内动脉瘤的发生及其发展。 巨噬细胞作为重要的炎症分子通过诱导MMP-9的表达参与了腹主动脉瘤的形成,CD68作为巨噬细胞的分子标志常用于判断组织内炎症反应的程度。此外研究表明MMP-9的过度表达与转录因子c-Jun有关,本研究拟探讨诱导颅内动脉瘤血管壁内MMP-9的过度表达相关调控机制,从颅内动脉瘤MMP-9诱导产生的早期阶段阻断其后继病理反应,为临床开发新药或检测特异性病理分子标记物提示颅内动脉瘤形成早期提供有用的实验依据。 自Hashimoto首次运用结扎大鼠一侧颈总动脉+切除一侧肾脏+术后(高盐饮食+β-丙晴氨)喂养2月等综合措施成功建立颅内动脉瘤动物模型后,该模型几经改进和完善,发现肾性高血压和血流动力学异常是制作颅内动脉瘤发病机制模型的两大基本条件,制作的动物模型与人类更多还原

【Abstract】 The initiation and development of the intracranial aneurysm are involved in the serious destruction of the extracellular matrix, demolished extracellular structural proteins like elastin and collagen are strongly associated with overexpression of matrix metalloproteinase(MMPs) especially of MMP-9, meanwhile with elevated expression of tissue inhibitor of metalloproteinase(TIMPs) in cerebral aneurysm, however, imbalance between MMPs and TIMPs due to MMP-9 overexpression contribute to the formation and development of the intracranial aneurysm.Macrophage usually acted as an important inflammatory molecule implicated in the progress of abdominal aortic aneurysm through induction of MMP-9, CD68 is regarded as macrophage identification which used to judge the extent of inflammatory reaction, in addtion, research manifest that overexpression of MMP-9 is related with transcription factor c-Jun. Our study focuses on exploring the regulation mechanism of MMP-9 overexpression of vascular tissue in intracranial aneurysm, strategies are adopted to preventing early pathological cascade reaction in MMP-9production in the cerebral aneurysm, and providing the useful experimental data for developing new drugs or detecting specific molecular marker suggested early intracranial aneurysmHashimoto firstly reported that cerebral aneurysms were induced after two months in rats treated with unilateral ligation of the common carotid artery and hypertension produced by renal resection as well as high saline and beta-aminopropionitrile feeding. With the development and improvement of this model, recently literature reported that intracranial aneurysm model were created by ligations of the left common carotid arteried and posterior branches of bilateral renal arteried with high salt diet, the model have two great advantages including high induction achievement and pathological features extremely similar to those in ruptured human cerebral aneurysm, however, there are few animal model in domestic regarding to the pathogenesis of intracranial aneurysm. Therefore, we firstly induce rat cerebral aneurysm with stablilization and resemble human pathology and wonder to know the expression of MMP-9 in the animal specimens, traditional approach of separating posterior branches of both renal artery via abdominal incision in rat is improved to investigate the pathogenesis of the cerebral aneurysm.Objective: To investigate the expression of CD68 -. c-Jun and MMP-9mRNA as well as related clinical significance in intracranial aneurysm. Two different operatival approaches were applied to establishment of animal model, operation results were compared in the two approaches, meanwhile, the expression of MMP-9 and related significance in rat specimen were analyzed.Methods: 1: the morphological alterations of extracellular matrix in the hunan intracranial aneurysm were observed by conventional light microscope and transmission electron microscope. 2: the expression of CD68 and immunoreactivity of c-Jun wre measured by immunohistochemical SP method. 3: the gene expression levels of MMP-9mRNA were detected by real time TaqRT-PCR quantify method. 4: Animal model of cerebral aneurysm were induced by ligations of the left common carotid artery and posterior branches both renal arteried as well as 2% saline drinking through abdominal and dorsal incisions , the rates of kidney resection > infection, mortality and total failure were compared in two groups of abdominal and dorsal incisions. 5: the expression of MMP-9 in rat specimens were measured by using immunohistochemical SP method.Results: 1: the numbers of elastic fiber in vascular wall obvious diminished, the elastic fiber become shorter, arrangement in disorder, some districts are substituted by numerous without cell or hyalinization of fabric component, there are abundant amorphous floccus existed in the fibrocytes. 2: Macrophage inflammatory infiltration with different degree appeared in every case of intracranial aneurysm(CD68 positive), and there was a parallel tendency between the degree of injury of extracellular matrix with various macrophage infiltration. 3: Immunoreactivity scores of phosphorylated c-Jun were increased in the cerebral aneurysm and abdominal aortic aneurysm compared with normal vessel(1.92±0.51 versus 0.17±0.41 and 1.67±0.52 versus 0.17+0.41 respectively, p<0.0\). 4: the A Ct of the gene expression of MMP-9 in the cerebral aneurysm and abdominal aortic aneurysm were (3.67+1.11) and (3.17 + 0.75) respectively, theACt of control group is 7.00± 1.26, their expression were significantly upregulated in aneurysms compared with that of the normal artery( 10.06-fold and 14.22-fold respectively). 5: Various aneurysmal changes were detected in 13 rats, including 4 preaneurysm and 9 early aneurysms, pathological alterations from induced aneurysmal sample show that internal elastic lamina disrupt or become thinning and even disappear, arrangement in disorder of the elastic fiber raritas, vacuolus degeneration of endothelial cell or shedding, the number of vascular smooth muscle cell decreased with the characteristics of apoptosis. 6: The positive MMP-9 expression is observed obviously in rat cerebral aneurysm compared without occurance in control group. 7: the rates of kidney resection and infection of control group were higher in abdominal group than those in dorsal group (0.025however, the relationship between CD68 and c-Jun should be further elucidated. 4: Increased BP combined with the hyperperfusion of one-lateraled willis loop are two key requirements for the establishment of intracranial aneurysm, the expression of MMP-9 is probably associated with the destruction of extracellular matrix. 5: It is an high safety and convenient method that the intracranial aneurysm were induced by ligation of posterior branches of both renal artery via bilateral dorsal approach.更多还原