2-甲氧雌二醇对硬脑膜动静脉瘘形成中血管新生的抑制作用

发布时间：2018-07-13 15:10

【摘要】：目的：硬脑膜动静脉瘘(DAVF)是脑血管病中的一大顽症,其病理基础是硬脑膜血管新生致动静脉短路。我们前期研究提示“静脉窦高压—脑组织低灌注—硬脑膜血管新生—DAVF形成”这一发病过程,干预血管新生有望成为防治DAVF的新途径。2-甲氧雌二醇(2-ME)具有强效抗血管生成作用,临床研究显示2-ME具有显著的血管新生抑制作用,进而抑制肿瘤的生长,因而提示它可能对DAVF血管新生过程同样有干预作用。本研究旨在通过大鼠静脉窦高压模型模拟人DAVF的病理生理过程,并进一步验证2-ME对大鼠静脉窦高压模型所致的硬膜血管新生的干预作用。方法：该研究首先检测患者DAVF组织中血管新生因子(ID-1、HIF-1α、VEGF、 MMPs等)的表达,并与大鼠颅内静脉高压模型中硬膜的组织病理改变相比较。之后我们进一步通过Elisa,免疫组化,免疫印迹,RT-PCR等方法研究2-ME对该模型中硬脑膜血管新生因子转录和表达的影响,以及对血管生成活跃程度的影响。结果：我们发现人DAVF病灶的组织切片与大鼠颅内静脉高压模型的硬膜组织切片中血管新生因子的表达变化具有一致性,发现2-ME能够一定程度抑制颅内静脉高压所致的VEGF、MMP-2/9、HIF-1α及ID-1基因和蛋白表达,并抑制硬脑膜血管新生活跃程度。这一效应在撤药后可维持2周。结论：2-ME对于大鼠静脉窦高压引发的硬膜血管新生具有抑制作用,提示其对于人DAVF有潜在的治疗价值。第一部分大鼠颅内静脉窦高压模型的建立及组织病理学验证目的：硬脑膜动静脉瘘(DAVF)是后天性疾病,许多患者发病前存在各种原因导致的静脉窦高压。通过文献复习,我们认为许多血管新生因子可能参与DAVF的形成,而这些血管新生因子的表达变化是否也在出现颅内静脉高压大鼠模型中,目前仍不清楚。本试验通过比较人DAVF病灶的组织切片与大鼠颅内静脉高压模型的硬膜组织切片中VEGF、MMP-2/9、HIF-1α和ID-1的表达,验证大鼠颅内静脉高压模型的硬膜组织病理学变化。方法：实验选取18例人DAVF标本进行免疫组化染色,并对6只大鼠行颅内静脉高压造模,12周后对硬膜进行切片染色,比较血管新生因子VEGF、MMP-2/9、HIF-1α和ID-1的表达程度。结果：人DAVF病灶的组织切片与大鼠颅内静脉高压模型的硬膜组织切片中血管新生因子均为高表达,具有一致性。结论：大鼠颅内静脉高压模型能够代表人DAVF的组织的部分病理特征。第二部分2-甲氧雌二醇对颅内静脉高压大鼠硬脑膜血管新生的抑制作用目的：硬脑膜动静脉瘘是一种复杂的脑血管疾病,可致颅内出血。近来我们发现慢性颅内低灌注能够引发硬膜血管新生,或进一步诱发硬脑膜动静脉瘘。甲氧雌二醇作为一种雌激素的代谢产物,有很强的血管新生抑制作用,已被用于抗肿瘤治疗中。因此本试验目的在于验证2-甲氧雌二醇对于颅内静脉高压大鼠硬膜血管新生的抑制作用。方法：试验中采用78只雄性SD大鼠,动物模型采用颅内静脉高压模型,造模后分为2-ME早晚期干预,并于不同时间节点取材,通过免疫组化、Elisa,蛋白印迹及RT-PCR方法分析2-ME血管新生抑制效果。结果：通过组织学分析、血清学分析、蛋白印迹及RT-PCR可以证实,2-ME能够一定程度抑制颅内静脉高压所致的VEGF、MMP-2/9、HIF-1α及ID-1的表达,进而抑制硬膜血管新生。这一效应在撤药后可维持2周。结论：2-ME对于大鼠静脉窦高压引发的硬膜血管新生具有抑制作用,提示其对人DAVF具有潜在的治疗价值。
[Abstract]:Objective: dural arteriovenous fistula (DAVF) is a major disease of cerebrovascular disease. Its pathological basis is the short circuiting of the vena cava in the dura mater vessels. Our early study suggests the process of "venous sinus hypertension - cerebral hypoperfusion - dural angiogenesis - DAVF formation", and the intervention of angiogenesis is expected to be a new way to prevent and cure DAVF .2- methoxy estradiol (2-ME) has a strong anti angiogenesis effect. Clinical studies have shown that 2-ME has significant angiogenesis inhibition and thus inhibits tumor growth, thus suggesting that it may also interfere with the angiogenesis process of DAVF. This study aims to simulate the pathophysiology of human DAVF through the rat model of venous sinus hypertension. To further verify the intervention effect of 2-ME on the neonatal vascular neovascularization induced by the rat venous sinus pressure model. Methods: the study first detected the expression of angiogenic factors (ID-1, HIF-1 a, VEGF, MMPs, etc.) in the DAVF tissues of the patients, and compared with the histopathological changes in the dura of the intracranial venous hypertension model in rats. Step through Elisa, immunohistochemistry, Western blot, RT-PCR and other methods to study the effect of 2-ME on the transcription and expression of the neovascularization factor in the dura mater, and the effect on the activity of angiogenesis. Results: we found the tissue section of the human DAVF focus and the neovascularization in the dural tissue section of the rat model of intracranial venous hypertension. The expression of 2-ME can inhibit the expression of VEGF, MMP-2/9, HIF-1 alpha and ID-1 gene and protein to a certain extent, and inhibit the new level of the dural blood vessel. This effect can be maintained for 2 weeks after withdrawal. Conclusion: 2-ME is a new type of dural angiogenesis induced by venous sinus hypertension in rats. The inhibitory effect suggests that it has a potential therapeutic value for human DAVF. The first part of the rat model of intracranial venous sinus hypertension and histopathological examination: DAVF is a acquired disease, and many patients have various causes of venous sinus hypertension before the onset of the disease. We think a lot of blood through literature review. Guan Xinsheng factor may be involved in the formation of DAVF, and whether the expression of these angiogenic factors is also in the rat model of intracranial venous hypertension is still unclear. This trial was conducted by comparing the expression of VEGF, MMP-2/9, HIF-1, and ID-1 in the tissue sections of human DAVF lesions and the intracranial venous hypertension model of rats. To verify the pathological changes in the dura tissue of the rat model of intracranial venous hypertension. Methods: 18 specimens of DAVF were selected for immunohistochemical staining and 6 rats were treated with intracranial venous hypertension. After 12 weeks, the epidural sections were stained to compare the expression of angiogenic factor VEGF, MMP-2/9, HIF-1 A and ID-1. Results: human DAVF disease. Conclusion: the intracranial venous pressure model in rats can represent some pathological features of the tissue of human DAVF. The second part of 2- methoxy estradiol can inhibit the neovascularization of the dura mater in the intracranial venous hypertension rats. Objective: the dural arteriovenous fistula (dural arteriovenous fistula) is a complex cerebrovascular disease which can cause intracranial hemorrhage. Recently, we found that chronic intracranial hypoperfusion can cause epidural angiogenesis, or further induce dural arteriovenous fistula. Methoxy estradiol is a metabolic product of estrogen, and has a strong inhibitory effect on angiogenesis. The purpose of this experiment is to verify the inhibitory effect of 2- methoxy estradiol on the epidural angiogenesis in intracranial venous hypertension rats. Methods: 78 male SD rats were used in the experiment. The model of intracranial vein high pressure was used in the animal model, and the model was divided into 2-ME early and late stage. Immunohistochemical, Elisa, Western blot and RT-PCR methods were used to analyze the effect of 2-ME angiogenesis. Results: through histological analysis, serological analysis, Western blot and RT-PCR, 2-ME could inhibit the expression of VEGF, MMP-2/9, HIF-1 A and ID-1 caused by intracranial venous hypertension to a certain extent, and then the effect of the inhibition of epidural angiogenesis. It can be maintained for 2 weeks after the withdrawal. Conclusion: 2-ME has a inhibitory effect on the neovascularization of the dural vessels induced by the high pressure of the venous sinus in rats, suggesting that it has potential therapeutic value for human DAVF.
【学位授予单位】：复旦大学
【学位级别】：博士
【学位授予年份】：2014
【分类号】：R743

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