蛋白酶体在腹主动脉瘤发病中的作用及分子病理机制研究
发布时间：2018-10-09 13:03
【摘要】：蛋白酶体在腹主动脉瘤发病中的作用及分子病理机制研究研究背景及目的：腹主动脉瘤(AAA)是老龄化社会的常见病,并且随着人群平均年龄日趋增加,AAA的发病率渐呈增高趋势。AAA危害巨大,一旦破裂致大出血,则死亡率达60～80%,造成生命及财产的巨大损失。炎症(淋巴细胞及巨噬细胞的浸润活化)、胶原及弹力纤维的破坏、平滑肌细胞(SMC)数量质量、和动脉壁新生血管的形成等均是AAA发生的关键环节。但是调节动脉壁炎症及SMC功能的内在分子机制仍不完全清楚。近年来,许多研究提示,泛素蛋白酶体系统,特别是蛋白酶体的活性变化在细胞炎症因子的释放及细胞凋亡等现象中起着关键作用。蛋白酶体是负责降解细胞内靶蛋白的主要“构件”,一种能够水解多聚泛素化蛋白的功能复合体。蛋白酶体活性变化多能够参与到调控炎症反应、影响SMC的凋亡等机制中。蛋白酶体活性的专门抑制剂硼替佐米(Bortezomib, BTZ)已经发现可通过调控细胞内蛋白酶体活性影响炎症细胞及SMC的功能变化,但是蛋白酶体在调节AAA形成相关的病理机制的作用,尚未见报道。因此,本研究拟在人及动物模型的主动脉组织中检测动脉壁内蛋白酶体活性变化,通过BTZ抑制主动脉内蛋白酶体活性,探讨蛋白酶体在AAA形成过程中参与炎症及SMC表型变化的相关作用及分子机制。实验方法：(1)收集人AAA病变及对照动脉组织；(2)通过对雄性缺陷小鼠(ApoE-/-)皮下埋植血管紧张素Ⅱ (AngⅡ)缓释泵(1,000 ng/min/kg)的方法,复制动物AAA模型；药物处理则进行腹腔注射BTZ (50μg/kg,每周2次),因此小鼠分成四组：Control, BTZ, AngⅡ, AngⅡ+BTZ。采用免疫染色、PCR分析的方法分析动脉组织内蛋白酶体各亚基表达水平(包括β1、β2、β5、βli、β2i、β5i),通过试剂盒测定蛋白酶体各蛋白酶活性(包括半胱天冬氨酸酶、胰蛋白酶、糜蛋白酶3种酶活性)；通过测量血管超声、鼠尾监测血压及大体解剖观察各组小鼠体内动脉大体形态及血压变化；通过HE染色、VVG染色和免疫组织化学染色法观察各组小鼠组织结构的变化和炎症细胞浸润(包括巨噬细胞和CD4+T细胞)；通过PCR分析、免疫组织染色分析各组小鼠动脉壁内炎症因子表达水平(包括TNF-α, IFN-γ,IL-4,IL-6和MCP-1)；通过免疫荧光染色aSMA与TUNEL或pCNA共染色确定主动脉壁内SMC的凋亡或增值状态,通过PCR分析、免疫组织染色和Westemblot方法分析各组小鼠动脉壁内SMC表型标识物表达水平及心肌素(myocardin)明确调控SMC表型的分子机制。结果：1.人AAA标本中蛋白酶体亚基表达水平及活性均增加；2.Ang II刺激诱导ApoE-/-小鼠建立成功诱导建立AAA动物模型,检测动脉内蛋白酶体表达及活性情况,随着疾病进展,蛋白酶体亚基表达水平及活性均增；3.BTZ抑制蛋白酶体活性后显著抑制AngⅡ诱导的小鼠AAA形成(发生率及直径)；4.蛋白酶体抑制后抑制AngⅡ诱导的①动脉壁内基质金属蛋白酶的表达及活性；②血管重塑及弹力纤维降解；③血管壁内炎症；5.蛋白酶体抑制后抑制AngⅡ诱导的血管壁内SMC增殖但并不影响其凋亡；6.蛋白酶体抑制后逆转AngⅡ诱导动脉壁内SMC表型转化。结论：我们证明了蛋白酶体的活化在促进体内AAA形成了关键的作用。其发挥作用与血管紧张素Ⅱ诱导的炎性细胞浸润,MMP活性增加,及SMC表型转化相关。低剂量的蛋白酶体抑制剂BTZ有效抑制这些改变。这些发现,明确了蛋白酶体参与AAA发生的病理生理学机制,也使得BTZ是可用于治疗AAA形成一个新的治疗靶点。
[Abstract]:Background and purpose of the research and research on the role and molecular pathological mechanism in the pathogenesis of abdominal aortic aneurysm (AAA): abdominal aortic aneurysm (AAA) is a common disease in aging society, and the incidence of AAA is increasing as the average age of the population increases. The AAA hazard is huge, and the death rate can reach 60-80%, resulting in great loss of life and property, once the major bleeding is broken. The destruction of inflammation (infiltration and activation of lymphocytes and macrophages), the destruction of collagen and elastic fibers, the mass of smooth muscle cells (SMC), and the formation of neovascularization in arterial wall are the key links of AAA. However, the intrinsic molecular mechanism of arterial wall inflammation and SMC function is still unclear. In recent years, many studies have suggested that the activity of ubiquitin-proteolysis system, especially the proteasomes, plays a key role in the release of cell inflammatory factors and apoptosis. Proteasomes are the main 鈥淐omponent鈥，
本文编号：2259428
[Abstract]:Background and purpose of the research and research on the role and molecular pathological mechanism in the pathogenesis of abdominal aortic aneurysm (AAA): abdominal aortic aneurysm (AAA) is a common disease in aging society, and the incidence of AAA is increasing as the average age of the population increases. The AAA hazard is huge, and the death rate can reach 60-80%, resulting in great loss of life and property, once the major bleeding is broken. The destruction of inflammation (infiltration and activation of lymphocytes and macrophages), the destruction of collagen and elastic fibers, the mass of smooth muscle cells (SMC), and the formation of neovascularization in arterial wall are the key links of AAA. However, the intrinsic molecular mechanism of arterial wall inflammation and SMC function is still unclear. In recent years, many studies have suggested that the activity of ubiquitin-proteolysis system, especially the proteasomes, plays a key role in the release of cell inflammatory factors and apoptosis. Proteasomes are the main 鈥淐omponent鈥，
本文编号：2259428
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