康柏西普联合PDGF受体抑制剂对缺氧条件下视网膜色素上皮细胞增殖、迁移及VEGF表达的影响

发布时间：2018-01-26 01:54

本文关键词： 康柏西普血小板衍生生长因子受体抑制剂缺氧人视网膜色素上皮细胞血管内皮生长因子　出处：《眼科新进展》2017年12期 　论文类型：期刊论文

【摘要】：目的探讨康柏西普联合血小板衍生生长因子(platelet derived growth factor,PDGF)受体抑制剂对CoCl_2诱导的人视网膜色素上皮细胞ARPE-19细胞增殖、迁移以及血管内皮生长因子(vascular endothelial growth factor,VEGF)m RNA及蛋白表达的影响。方法体外培养ARPE-19细胞,取对数生长良好的细胞,用不同浓度CoCl_2诱导培养,细胞增殖与CCK-8法检测ARPE-19细胞增殖活性,筛选最佳CoC l2浓度用于构建缺氧模型;将培养的ARPE-19细胞分为正常组、缺氧组、不同浓度康柏西普组,CCK-8法检测不同浓度康柏西普对缺氧损伤后ARPE-19细胞增殖的影响,筛选最佳康柏西普浓度;将培养的ARPE-19细胞分为正常组、缺氧组、不同浓度PDGF受体抑制剂组,CCK-8法检测不同浓度PDGF受体抑制剂对缺氧损伤后ARPE-19细胞增殖的影响,筛选最佳PDGF受体抑制剂浓度;将培养的ARPE-19细胞分为正常组、缺氧组、20 mg·L~(-1)康柏西普组、20μmol·L~(-1)PDGF受体抑制剂组、康柏西普联合PDGF受体抑制剂组(最佳药物浓度组合);CCK-8法检测缺氧损伤后各组ARPE-19细胞增殖活性,Transwell小室检测细胞的迁移情况,ELISA法检测细胞上清液中VEGF蛋白浓度,应用实时荧光定量PCR法检测细胞VEGF mR NA的表达水平。结果 CCK-8筛选结果显示,100μmol·L~(-1)CoC l2处理组ARPE-19细胞的吸光度值最大(1.063±0.031),设立为缺氧细胞模型浓度。CCK-8筛选结果显示,20 mg·L~(-1)康柏西普组细胞增殖率为(94.58±3.80)%、20μmol·L~(-1)PDGF受体抑制剂组细胞增殖率为(96.72±5.44)%,效果均较其他相应组好,选取该浓度进行后续实验。析因分析发现缺氧损伤后各药物处理组(康柏西普、PDGF受体抑制剂、康柏西普联合PDGF受体抑制剂)与缺氧组比较,细胞增殖活性下降,细胞迁移数降低,VEGF蛋白浓度亦下降,其中康柏西普联合PDGF受体抑制剂组下降最明显,康柏西普组及两药联合处理组ARPE-19细胞VEGF mR NA表达较缺氧组降低。结论缺氧损伤后可诱导ARPE-19细胞增殖、迁移,VEGF蛋白相对表达量增加,VEGF mR NA表达上调;康柏西普联合PDGF受体抑制剂对缺氧损伤后ARPE-19细胞的增殖、迁移及VEGF蛋白、VEGF mR NA的表达有抑制作用,作用效果优于单独应用康柏西普。
[Abstract]:Objective to investigate the combination of Compactsep and platelet derived growth factor. PDGF receptor inhibitor could induce the proliferation of human retinal pigment epithelium (ARPE-19) cells induced by CoCl_2. Migration and vascular endothelial growth factor. Methods ARPE-19 cells were cultured in vitro. The cells with good logarithmic growth were cultured with different concentrations of CoCl_2. Cell proliferation and CCK-8 assay were used to detect the proliferative activity of ARPE-19 cells, and the optimal concentration of CoC l2 was selected to construct anoxic model. The cultured ARPE-19 cells were divided into three groups: normal group, hypoxia group, and different concentrations of Compactopril (CCK-8). The effects of different concentrations of Compactopril on the proliferation of ARPE-19 cells after hypoxia injury were detected by CCK-8 method. Screening of the best concentration of Compactopril; The cultured ARPE-19 cells were divided into normal group hypoxia group and different concentration of PDGF receptor inhibitor group. The effects of different concentrations of PDGF receptor inhibitors on the proliferation of ARPE-19 cells after hypoxia injury were detected by CCK-8 assay, and the best concentration of PDGF receptor inhibitors was selected. The cultured ARPE-19 cells were divided into two groups: normal group, hypoxia group (20 mg 路L ~ (-1)) Compactopril group (n = 20 渭 mol 路L ~ (-1)). Compactopril combined with PDGF receptor inhibitor group (best drug concentration combination); CCK-8 assay was used to detect the proliferation activity of ARPE-19 cells in each group after hypoxia injury. ELISA assay was used to detect the concentration of VEGF protein in supernatant and real-time fluorescence quantitative PCR method was used to detect the expression level of VEGF MRNA. The maximum absorbance of ARPE-19 cells was 1.063 卤0.031 in 100 渭 mol 路L ~ (-1) ARPE-19 cells treated with CoCl _ (2). The results of CCK-8 screening showed that the cell proliferation rate was 94.58 卤3.80% in the Compactopril group (20 mg 路L ~ (-1) 路L ~ (-1)). The cell proliferation rate of 20 渭 mol 路L ~ (-1) PDGF receptor inhibitor group was 96.72 卤5.44, and the effect was better than that of other corresponding groups. This concentration was selected for follow-up experiments. The results of factorial analysis showed that each drug treatment group (Compactopril PDGF-receptor inhibitor, Compactopril combined with PDGF receptor inhibitor) was compared with the hypoxia group after hypoxia injury. The cell proliferation activity decreased, the cell migration decreased and the concentration of PDGF protein decreased, especially in the Compactopril combined with PDGF receptor inhibitor group. Compared with hypoxia group, the expression of VEGF mRNA in ARPE-19 cells in Compactopril group and combined treatment group was lower than that in hypoxia group. Conclusion the proliferation and migration of ARPE-19 cells can be induced by hypoxia injury. The relative expression of VEGF protein was increased and the expression of VEGF mRNA was up-regulated. Compactopril combined with PDGF receptor inhibitor can inhibit the proliferation, migration and the expression of VEGF mRNA in ARPE-19 cells after hypoxia injury. The effect was better than that of Compathep alone.
【作者单位】：徐州医科大学研究生学院;徐州医科大学附属医院眼科;
【分类号】：R774
【正文快照】： 眼部新生血管性疾病的共同病理改变是病理性1.2方法新生血管形成。研究表明,细胞内稳定的氧含量是1.2.1 ARPE-19细胞的培养采用含体积分数细胞生存的关键,在缺血、缺氧、炎症等因素刺激下,10%胎牛血清的DMEM/F12培养基在37℃、体积分脉络膜毛细血管血液灌注不良,视网膜色素上

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