Slit2-N对VEGF165诱导的HUVEC及HMVEC细胞增殖的抑制作用及其机制

发布时间：2018-04-06 21:31

  本文选题：SLIT2-N蛋白　切入点：VEGF165　出处：《重庆医科大学》2017年硕士论文

【摘要】：目的:探索Slit2-N蛋白对VEGF165诱导的HUVEC及HMVEC细胞增殖的影响及其机制,为寻找治疗CNV新的靶点提供思路和线索。方法:1.体外培养HUVEC及HMVEC细胞株。2.CCK-8筛选Slit2-N蛋白及VEGF165蛋白作用于HUVEC及HMVEV细胞的有效工作浓度。3.CCK-8检测Slit2-N蛋白对VEGF165诱导的HUVEC及HMVEC细胞增殖的影响。4.Annexin-V FITC/PI双染流式法检测HUVEC及HMVEC细胞空白对照组、Slit2-N组、VEGF165组和Slit2-N and VEGF165组细胞的凋亡情况。5.CFSE流式法检测HUVEC及HMVEC细胞空白对照组、Slit2-N组、VEGF165组和Slit2-N and VEGF165组各组细胞增殖情况。6.Western Blot法检测HUVEC及HMVEC细胞空白对照组、Slit2-N组、VEGF165组和Slit2-N and VEGF-165组各组细胞中p-AKT,AKT,p-ERK1/2,ERK1/2表达情况。7.多组间单因素比较采用单因素方差分析。结果:1.CCK-8结果显示,在48小时VEGF165能引起HUVEC和HMVEC细胞明显增殖的最小浓度分别为25ng/ml和500p M,分别筛选为工作浓度;48小时Slit2-N不引起HUVEC和HMVEC活力下降的最大浓度均为16n M,筛选为工作浓度。2.细胞接受处理后48小时CCK-8结果提示,Slit2-N组与Control组之间无显著差异(HUVEC和HMVEC均p0.05),VEGF165组较Control组明显上升(HUVEC:P0.01,HMVEC:P0.001),Slit2-N and VEGF165组与VEGF165组之间差异显著(HUVEC:p0.05,HMVEC:p0.001)。3.Annexin-V FITC/PI双染流式结果显示细胞早期凋亡率在HUVEC和HMVEC中,24小时,48小时,72小时VEGF165组与Slit2-N and VEGF165组之间均无明显差异(P0.05)。4.在细胞接受处理后48小时CFSE染色流式结果显示HUVEC细胞Control组,Slit2-N组,VEGF165组,Slit2-N and VEGF165组中增殖指数分别为(31.63±0.28),(31.88±0.92),(42.12±1.68),(32.71±0.32);HMVEC各组增殖指数分别为(12.54±1.08),(13.39±0.15),(15.74±0.51),(12.89±0.90),Slit2-N组与Control组与之间无明显差异(HUVEC和HMVEC中均p0.05),VEGF165组与Control组之间有显著差异(HUVEC:P0.001,HMVEC:P0.01),Slit2-N andVEGF165组与VEGF165组之间有显著差异(HUVEC:p0.001,HMVEC:p0.01);5.Western Blot结果显示,p-AKT的相对表达量在Slit2-N组与Control组之间无显著差异(HUVEC和HMVEC均p0.05),VEGF165组较Control组明显上升(HUVEC:p0.001,HMVEC:p0.01),Slit2-N and VEGF165组较VEGF165组明显下调(HUVEC:p0.001,HMVEC:p0.01);p-ERK 1/2的相对表达量在Slit2-N与Control组之间无明显差异(HUVEC和HMVEC均p0.05),VEGF165组较Control组明显上升(HUVEC:p0.001,HMVEC:p0.001),Slit2-N and VEGF165组与VEGF165组之间无明显差异(HUVEC和HMVEC均p0.05)。结论:Slit2-N可能通过下调p-AKT的相对表达量,而非p-ERK1/2,抑制VEGF165诱导的HUVEC及HMVEC的细胞增殖,为寻找治疗CNV新的靶点提供了思路和线索。
[Abstract]:Aim: to explore the effect of Slit2-N protein on the proliferation of HUVEC and HMVEC cells induced by VEGF165 and its mechanism.Method 1: 1.Screening the effective working concentration of Slit2-N protein and VEGF165 protein on HUVEC and HMVEV cells in Vitro cultured HUVEC and HMVEC Cell Line .3.The effect of Slit2-N protein on VEGF165 induced proliferation of HUVEC and HMVEC cells. 4. Annexin-V FITC/PI double staining flow cytometry to detect HUVEC and HMVEC cellsThe expression of p-AKTT tachycardia and p-ERK1 / 2 ERK1 / 2 in VEGF165 group and Slit2-N and VEGF-165 group was 7. 7.Single factor analysis of variance (ANOVA) was used to compare single factors among groups.Results 1. The results of CCK-8 showed that the minimum concentration of VEGF165 induced significant proliferation of HUVEC and HMVEC cells at 48 hours was 25ng/ml and 500p / M, respectively, and the maximum concentration of Slit2-N that could not induce the decrease of HUVEC and HMVEC activity was 16nM.In HUVEC and HMVEC, there was no significant difference between VEGF165 group and Slit2-N and VEGF165 group.鍦ㄧ粏鑳炴帴鍙楀�勭悊鍚�48灏忔椂CFSE鏌撹壊娴佸紡缁撴灉鏄剧ずHUVEC缁嗚優Control缁，

本文编号：1719012