CELSR1促进血管内皮细胞迁移与血管形成作用的研究
发布时间：2018-09-06 14:43
【摘要】：【背景和目的】 全基因组关联研究发现CELSR1基因是日本人群缺血性卒中的易感基因，我们课题组前期实验也发现CELSR1基因rs6007897和rs4044210与缺血性卒中具有显著相关性，与大动脉粥样硬化具有相关性；CELSR1是Wnt/PCP通路的核心成员，Wnt/PCP通路在动脉粥样硬化过程中起作用。本研究观察过表达和沉默CELSR1基因的人主动脉内皮细胞（human aortic endothelial cells，HAECs）血管内皮细胞的影响，来阐述CELSR基因在动脉粥样硬化过程中的作用。 【方法】 首先利用构建干扰CELSR1表达的质粒，通过慢病毒载体感染HAEC细胞，利用puromycin筛选出表达有CELSR1shRNA的HAEC的细胞株；利用TALEA技术将携带过表达CELSR1质粒转染HAEC，利用puromycin筛选出过表达CELSR1的HAEC的细胞株；采用Westernblot法、实时荧光定量PCR法以及MTT法和划痕实验、Transwell实验、成环实验检测过表达和沉默细胞株的CELSR1的表达及HAECs增殖和迁移、成环能力。 【结果】 CELSR1shRNA序列可有效抑制HACE细胞CELSR1的表达导入CELSR1序列可明显提高CELSR1在HAEC细胞中的表达，差异有统计学意义(P0.05)。MTT实验结果示，干扰CELSR1表达的HAEC细胞生长明显受到抑制,差异有统计学意义(P0.05)；过表达CELSR1的HAEC细胞与对照组相比，生长速度加快，差异有统计学意义(P0.05)。划痕实验及Transwel实验结果示，干扰CELSR1表达的HAEC细胞迁移速度较对照组减慢，差异有统计学意义(P0.05)，过表达CELSR1的HAEC细胞迁移速度与对照组相比，迁移速度增快，差异有统计学意义(P0.05)。成环实验示，干扰CELSR1表达的HAEC细胞较对照组的成环数低于对照组，差异有统计学意义(P0.05)。过表达CELSR1的HAEC细胞的成环数高于对照组，差异有统计学意义(P0.05)。 【结论】 CELSR1shRNA能够下调HAEC的CELSR1表达，有效地抑制HAEC的增殖和迁移、成环；转染CELSR1能上调HAEC内源性CELSR1的表达，有效地促进HAEC的增殖和迁移、成环，提示CELSR1可能在血管生成过程中发挥重要作用。
[Abstract]:[background and objective] Genome-wide association studies have found that CELSR1 gene is a susceptible gene for ischemic stroke in Japanese population. Our previous study also found that CELSR1 gene rs6007897 and rs4044210 have a significant correlation with ischemic stroke. CELSR1 is a core member of the Wnt/PCP pathway, which plays an important role in atherosclerosis. The purpose of this study was to investigate the effects of human aortic endothelial cells (human aortic endothelial cells,HAECs) expressing and silencing CELSR1 gene on vascular endothelial cells. To explain the role of CELSR gene in atherosclerosis. [methods] first, the plasmid interfering with CELSR1 expression was constructed, and HAEC cells were infected by lentivirus vector. Puromycin was used to screen the cell lines expressing HAEC with CELSR1shRNA; HAEC, was transfected with overexpressed CELSR1 plasmid by TALEA technique; HAEC cell lines expressing CELSR1 were screened by puromycin; Westernblot method, real-time fluorescence quantitative PCR method, MTT method and scratch test were used to screen HAEC cell lines. The expression of CELSR1 and the proliferation and migration of HAECs in overexpression and silencing cell lines were detected by loop forming assay. [results] CELSR1shRNA sequence could effectively inhibit the expression of CELSR1 in HACE cells and induce CELSR1 sequence to increase the expression of CELSR1 in HAEC cells. The difference was statistically significant (P0.05). The growth of HAEC cells which interfered with the expression of CELSR1 was significantly inhibited (P0.05); the growth rate of HAEC cells overexpression of CELSR1 was faster than that of the control group (P0.05). The results of scratch test and Transwel test showed that the migration speed of HAEC cells interfering with CELSR1 expression was slower than that of control group (P0.05). The migration speed of HAEC cells with overexpression of CELSR1 was faster than that of control group (P0.05). The ring-forming experiment showed that the number of HAEC cells interfering with CELSR1 expression was lower than that of the control group (P0.05). The number of HAEC cells with overexpression of CELSR1 was significantly higher than that of control group (P0.05). [conclusion] CELSR1shRNA can down-regulate the expression of HAEC CELSR1 and effectively inhibit the proliferation and migration of HAEC. Transfection of CELSR1 can up-regulate the expression of endogenous CELSR1 in HAEC and effectively promote the proliferation and migration of HAEC, suggesting that CELSR1 may play an important role in angiogenesis.
【学位授予单位】：福建医科大学
【学位级别】：硕士
【学位授予年份】：2014
【分类号】：R743.3
本文编号：2226673
[Abstract]:[background and objective] Genome-wide association studies have found that CELSR1 gene is a susceptible gene for ischemic stroke in Japanese population. Our previous study also found that CELSR1 gene rs6007897 and rs4044210 have a significant correlation with ischemic stroke. CELSR1 is a core member of the Wnt/PCP pathway, which plays an important role in atherosclerosis. The purpose of this study was to investigate the effects of human aortic endothelial cells (human aortic endothelial cells,HAECs) expressing and silencing CELSR1 gene on vascular endothelial cells. To explain the role of CELSR gene in atherosclerosis. [methods] first, the plasmid interfering with CELSR1 expression was constructed, and HAEC cells were infected by lentivirus vector. Puromycin was used to screen the cell lines expressing HAEC with CELSR1shRNA; HAEC, was transfected with overexpressed CELSR1 plasmid by TALEA technique; HAEC cell lines expressing CELSR1 were screened by puromycin; Westernblot method, real-time fluorescence quantitative PCR method, MTT method and scratch test were used to screen HAEC cell lines. The expression of CELSR1 and the proliferation and migration of HAECs in overexpression and silencing cell lines were detected by loop forming assay. [results] CELSR1shRNA sequence could effectively inhibit the expression of CELSR1 in HACE cells and induce CELSR1 sequence to increase the expression of CELSR1 in HAEC cells. The difference was statistically significant (P0.05). The growth of HAEC cells which interfered with the expression of CELSR1 was significantly inhibited (P0.05); the growth rate of HAEC cells overexpression of CELSR1 was faster than that of the control group (P0.05). The results of scratch test and Transwel test showed that the migration speed of HAEC cells interfering with CELSR1 expression was slower than that of control group (P0.05). The migration speed of HAEC cells with overexpression of CELSR1 was faster than that of control group (P0.05). The ring-forming experiment showed that the number of HAEC cells interfering with CELSR1 expression was lower than that of the control group (P0.05). The number of HAEC cells with overexpression of CELSR1 was significantly higher than that of control group (P0.05). [conclusion] CELSR1shRNA can down-regulate the expression of HAEC CELSR1 and effectively inhibit the proliferation and migration of HAEC. Transfection of CELSR1 can up-regulate the expression of endogenous CELSR1 in HAEC and effectively promote the proliferation and migration of HAEC, suggesting that CELSR1 may play an important role in angiogenesis.
【学位授予单位】：福建医科大学
【学位级别】：硕士
【学位授予年份】：2014
【分类号】：R743.3
【参考文献】
中国期刊全文数据库 前3条
1 曲宜波;周立兵;;非典型钙粘蛋白Celsr1-3在神经系统发育和PCP中的功能[J];暨南大学学报(自然科学与医学版);2012年04期
2 郭超伟;宋凯;李萍;;Notch和Wnt信号通路在血管形成中的协同调控作用[J];中国生物化学与分子生物学报;2010年06期
3 柳华;季一飞;熊利;张翼;;缺血性脑卒中的遗传研究进展[J];西部医学;2013年06期
，本文编号：2226673
本文链接：https://www.wllwen.com/yixuelunwen/shenjingyixue/2226673.html
最近更新
教材专著
