载脂蛋白A-I模拟肽D-4F拮抗血管平滑肌细胞增殖及迁移的机制研究

发布时间：2018-05-11 12:15

  本文选题：载脂蛋白A-I模拟肽D-4F + 氧化型低密度脂蛋白(ox-LDL)　； 参考：《福建医科大学》2015年硕士论文

【摘要】：目的:探讨载脂蛋白A-I模拟肽D-4F拮抗血管平滑肌细胞(vascular smooth muscle cells,VSMCs)增殖及迁移的机制。方法:大鼠胸主动脉VSMCs分离与培养。分别给予D-4F及氧化型低密度脂蛋白(oxidized low-density lipoprotein,ox-LDL)处理VSMCs,CCK-8(Cell counting kit-8)试剂盒来测定细胞增殖,transwell小室和划痕实验来评估细胞迁移,而活性氧探针,即双氢乙酰乙酸一二氯荧光黄(dihydrochloride acetyl acid dichloride fluorescent yellow,DCFH-DA)测定VSMCs中活性氧簇(reactive oxygen species,ROS)。利用western blotting检测D-4F对VSMCs中P13K/Akt信号通路磷酸化激活和血红素加氧酶-1(heme oxygenase-1,HO-1)蛋白表达。利用P13K/Akt抑制剂LY294002预孵育VSMCs,western blotting检测HO-1蛋白表达。利用HO-1抑制剂Znpp拮抗VSMCs中HO-1的活性,再次检测D-4F对ox-LDL诱导的ROS释放以及VSMCs增殖和迁移的抑制作用。结果:D-4F在体外能够抑制ox-LDL诱导的血管平滑肌细胞(VSMCs)的增殖和迁移。D-4F能够诱导VSMCs中P13K/Akt磷酸化激活,D-4F上调VSMCs中HO-1蛋白的表达,PI3K/Akt抑制剂LY294002能够拮抗D-4F诱导的VSMCs中HO-1蛋白表达。此外D-4F能够有效的抑制ox-LDL诱导的VSMCs中ROS的释放,进而抑制ox-LDL诱导的VSMCs的增殖和迁移。HO-1抑制剂Znpp能够拮抗D-4F的抗氧化与抗增殖和抗迁移活性。结论:载脂蛋白A-I模拟肽D-4F通过激活PI3K/Akt通路上调VSMCs中HO-1蛋白的表达,以此来抑制ox-LDL诱导的ROS释放,拮抗VSMCs增殖和迁移。
[Abstract]:Aim: to investigate the mechanism of antagonistic effect of apolipoprotein A-I mimic peptide D-4F on proliferation and migration of vascular smooth muscle cells in vascular smooth muscle cells (VSMCs). Methods: VSMCs was isolated and cultured from rat thoracic aorta. D-4F and oxidized low density lipoprotein (LDL) -treated VSMCsCCK-8 cell counting kit-8 were used to measure cell proliferation, transwell chamber and scratch test to evaluate cell migration, and reactive oxygen probe (Ros). That is, dihydroacetoacetic acid, dichlorofluorescein dihydrochloride acetyl acid dichloride fluorescent yellow.DCFH-DAA was used to determine reactive oxygen species, oxygen speciesrossuch as Ros in VSMCs. Western blotting was used to detect the activation of phosphorylation of P13K/Akt signaling pathway and the expression of hemhemoxygenase-1 (HO-1) protein in VSMCs by D-4F. P13K/Akt inhibitor LY294002 preincubated VSMCswestern blotting to detect the expression of HO-1 protein. HO-1 inhibitor Znpp was used to antagonize the activity of HO-1 in VSMCs. The inhibitory effects of D-4F on ROS release induced by ox-LDL and VSMCs proliferation and migration were detected again. Results D-4F could inhibit the proliferation and migration of vascular smooth muscle cells induced by ox-LDL. D-4F could induce P13K/Akt phosphorylation in VSMCs to activate the expression of HO-1 protein in VSMCs. PI3K / Akt inhibitor LY294002 could antagonize the expression of HO-1 protein in VSMCs induced by D-4F. In addition, D-4F could effectively inhibit the release of ROS in VSMCs induced by ox-LDL, and then inhibit the proliferation and migration of VSMCs induced by ox-LDL. Znpp, an inhibitor of HO-1, could antagonize the antioxidant, anti-proliferation and anti-migration activities of D-4F. Conclusion: apolipoprotein A-I mimic peptide D-4F up-regulates the expression of HO-1 protein in VSMCs by activating the PI3K/Akt pathway, thereby inhibiting the ROS release induced by ox-LDL and antagonizing the proliferation and migration of VSMCs.
【学位授予单位】：福建医科大学
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：R541.4

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