低切应力诱导血管细胞活化激酶C受体1的表达及其意义
发布时间：2018-07-31 18:36
【摘要】：血管重建（remodeling）是心血管疾病共有的发病基础和基本的病理过程。血流切应力变化是影响血管重建的重要因素，与动脉粥样硬化的发生发展密切相关。血管内皮细胞(endothelial cells, ECs)直接承载血流切应力刺激，并通过细胞间相互作用影响血管平滑肌细胞（vascular smooth muscle cells, VSMCs）。研究ECs与VSMCs间的相互交流，以及切应力下的细胞功能变化，有助于阐明血管重建的力学生物学机制。 我们通过前期蛋白质组学的工作，得到了正常切应力（normalshear stress, NSS,15dyn/cm2）和低切应力（low shear stress，LowSS，5dyn/cm2）条件下体外培养24h的血管组织差异表达的蛋白质谱。对比发现，活化激酶C受体1（receptor for activated C kinase1，RACK1）在低切应力作用下的血管组织中呈高表达，提示RACK1可能参与了LowSS诱导的血管重建。 本文应用ECs/VSMCs联合培养流动腔系统，获得静态（static）、NSS和LowSS条件下联合培养大鼠ECs和VSMCs表达RACK1及Akt磷酸化水平。同时，为探讨应力调控的上述变化是通过ECs与VSMCs的直接接触或是通过旁分泌作用，本文设计了ECs//VSMCs隔开培养的体系，同样检测静态、NSS和LowSS条件下ECs和VSMCs的RACK1表达状况及Akt磷酸化水平。此外，本文还在静态条件下，应用RNA干扰（RNA interference, RNAi）技术特异性抑制VSMCs内RACK1表达，探讨RACK1对细胞凋亡和Akt磷酸化水平的作用。 结果显示：①切应力与VSMCs对ECs的RACK1表达均无显著影响；②LowSS与细胞直接接触共同诱导VSMCs表达RACK1，NSS、LowSS与旁分泌共同阻遏VSMCs表达RACK1；③NSS和LowSS都可能抑制ECs的Akt磷酸化；④旁分泌参与NSS对VSMCs的Akt磷酸化的促进作用；⑤抑制VSMCs的RACK1表达下调细胞凋亡和caspase-3表达，上调Akt磷酸化。 上述结果表明，LowSS条件下，ECs通过直接接触作用诱导VSMCs的RACK1表达；切应力对ECs与VSMCs的Akt磷酸化水平的诱导作用不同，NSS和LowSS均抑制ECs的Akt磷酸化，，但NSS的抑制作用更强，而NSS则促进VSMCs的Akt磷酸化；RACK1可能通过PI3K/Akt信号通路参与切应力对细胞凋亡的调控。结果提示，ECs与VSMCs之间的相互接触和联系，对维持血管的正常结构与功能的稳定有重要作用。
[Abstract]:Revascularization of (remodeling) is the common pathogenesis and basic pathological process of cardiovascular disease. The change of blood flow shear stress is an important factor affecting vascular remodeling and is closely related to the occurrence and development of atherosclerosis. Vascular endothelial cells (endothelial cells, ECs) directly carry shear stress stimulation and influence vascular smooth muscle cells (VSMC) (vascular smooth muscle cells, VSMCs). Through intercellular interaction. The study of the interaction between ECs and VSMCs, as well as the changes of cell function under shear stress, is helpful to elucidate the mechanics and biological mechanism of vascular remodeling. We obtained the differentially expressed protein profiles of vascular tissues cultured in vitro for 24 hours under normal shear stress (normalshear stress,) 15 dyn / cm 2) and low shear stress (5 dyn / cm 2). It was found that activated kinase C receptor 1 (receptor for activated C kinase1 rack1) was highly expressed in vascular tissues under low shear stress, suggesting that RACK1 might be involved in the vascular remodeling induced by LowSS. In this paper, the expression of RACK1 and Akt phosphorylation of ECs and VSMCs in rats co-cultured with ECs/VSMCs and LowSS were obtained. At the same time, in order to investigate whether the above changes in stress regulation are through the direct contact between ECs and VSMCs or through paracrine action, the system of isolated culture of ECs//VSMCs was designed in this paper. The expression of RACK1 and the level of Akt phosphorylation in ECs and VSMCs were also detected under the static condition of NSS and LowSS. In addition, under static conditions, RNA interference (RNA interference, RNAi) technique was used to specifically inhibit the expression of RACK1 in VSMCs, and to explore the effect of RACK1 on apoptosis and Akt phosphorylation. The results showed that the RACK1 expression of ECs was not significantly affected by the shear stress of 1: 1 and VSMCs. The direct contact between VSMCs and cells induced VSMCs expression of rack1, NSS, low SS and paracrine to inhibit VSMCs expression of rack1n3NSS and LowSS, both of which might inhibit Akt phosphorylation of ECs. 4 paracrine plays an important role in the promotion of Akt phosphorylation of VSMCs by NSS. 5 inhibits the expression of RACK1 in VSMCs and down-regulates the expression of caspase-3 and apoptosis, and up-regulates the phosphorylation of Akt. The results indicated that the RACK1 expression of VSMCs was induced by direct contact between ECs and VSMCs under LowSS condition, and that shear stress induced Akt phosphorylation level of ECs and VSMCs was different. Both LowSS and ECs inhibited Akt phosphorylation of ECs, but NSS inhibited ECs phosphorylation more strongly. NSS promotes the Akt phosphorylation of VSMCs, RACK1, which may be involved in the regulation of apoptosis by shear stress through PI3K/Akt signaling pathway. The results suggest that the contact and connection between ECs and VSMCs play an important role in maintaining the normal structure and function of blood vessels.
【学位授予单位】：上海交通大学
【学位级别】：硕士
【学位授予年份】：2012
【分类号】：R363
本文编号：2156474
[Abstract]:Revascularization of (remodeling) is the common pathogenesis and basic pathological process of cardiovascular disease. The change of blood flow shear stress is an important factor affecting vascular remodeling and is closely related to the occurrence and development of atherosclerosis. Vascular endothelial cells (endothelial cells, ECs) directly carry shear stress stimulation and influence vascular smooth muscle cells (VSMC) (vascular smooth muscle cells, VSMCs). Through intercellular interaction. The study of the interaction between ECs and VSMCs, as well as the changes of cell function under shear stress, is helpful to elucidate the mechanics and biological mechanism of vascular remodeling. We obtained the differentially expressed protein profiles of vascular tissues cultured in vitro for 24 hours under normal shear stress (normalshear stress,) 15 dyn / cm 2) and low shear stress (5 dyn / cm 2). It was found that activated kinase C receptor 1 (receptor for activated C kinase1 rack1) was highly expressed in vascular tissues under low shear stress, suggesting that RACK1 might be involved in the vascular remodeling induced by LowSS. In this paper, the expression of RACK1 and Akt phosphorylation of ECs and VSMCs in rats co-cultured with ECs/VSMCs and LowSS were obtained. At the same time, in order to investigate whether the above changes in stress regulation are through the direct contact between ECs and VSMCs or through paracrine action, the system of isolated culture of ECs//VSMCs was designed in this paper. The expression of RACK1 and the level of Akt phosphorylation in ECs and VSMCs were also detected under the static condition of NSS and LowSS. In addition, under static conditions, RNA interference (RNA interference, RNAi) technique was used to specifically inhibit the expression of RACK1 in VSMCs, and to explore the effect of RACK1 on apoptosis and Akt phosphorylation. The results showed that the RACK1 expression of ECs was not significantly affected by the shear stress of 1: 1 and VSMCs. The direct contact between VSMCs and cells induced VSMCs expression of rack1, NSS, low SS and paracrine to inhibit VSMCs expression of rack1n3NSS and LowSS, both of which might inhibit Akt phosphorylation of ECs. 4 paracrine plays an important role in the promotion of Akt phosphorylation of VSMCs by NSS. 5 inhibits the expression of RACK1 in VSMCs and down-regulates the expression of caspase-3 and apoptosis, and up-regulates the phosphorylation of Akt. The results indicated that the RACK1 expression of VSMCs was induced by direct contact between ECs and VSMCs under LowSS condition, and that shear stress induced Akt phosphorylation level of ECs and VSMCs was different. Both LowSS and ECs inhibited Akt phosphorylation of ECs, but NSS inhibited ECs phosphorylation more strongly. NSS promotes the Akt phosphorylation of VSMCs, RACK1, which may be involved in the regulation of apoptosis by shear stress through PI3K/Akt signaling pathway. The results suggest that the contact and connection between ECs and VSMCs play an important role in maintaining the normal structure and function of blood vessels.
【学位授予单位】：上海交通大学
【学位级别】：硕士
【学位授予年份】：2012
【分类号】：R363
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