线粒体凋亡通路在棕榈酸诱导的血管内皮细胞凋亡中的作用
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摘要
目的探讨线粒体凋亡通路在棕榈酸诱导的人脐静脉血管内皮细胞(human umbilical vein endothelial cells,HUVECs)凋亡中的作用。方法不同浓度棕榈酸(0.1、0.2、0.4、0.8 mmol·L~(-1))作用HUVECs(0、12、24、48 h),MTT法检测HUVECs增殖能力;免疫荧光法检测HUVECs细胞内活性氧(reactive oxygen species,ROS)的表达水平;免疫印迹法检测HUVECs中AIF、Cyt-C、cleaved caspase-3、Bcl-2、Bax等线粒体凋亡通路蛋白的表达水平;TUNEL法检测细胞内凋亡水平。结果 0.4 mmol·L~(-1)棕榈酸作用HUVECs 24 h时,细胞增殖率明显降低;AIF、Cyt-C、cleaved caspase-3、Bax/Bcl-2的水平明显升高(P<0.05),细胞凋亡水平明显增高(P<0.05);线粒体通透性抑制剂环孢素A (ciclosporine A, CsA)预处理组HUVECs凋亡水平明显下调(P<0.05)。结论棕榈酸所致血管内皮细胞损伤的发病机制可能与线粒体凋亡相关通路密切关联,此研究对脂毒性心肌损伤的防治有重要意义。
Aim To explore the role of mitochondrial apoptosis pathway in palmitic acid(PA)-induced cell apoptosis in human umbilical vein endothelial cells(HUVECs). Methods HUVECs were exposed to different concentrations of PA(0.1, 0.2, 0.4, 0.8 mmol·L~(-1)) for 24 h and different time points of 0.4 mmol·L~(-1) PA(0, 12, 24, 48 h). Cell viability was measured by MTT. The expression of reactive oxygen species(ROS) in HUVECs was detected by immunofluorescence. The level of intracellular apoptosis was detected by TUNEL assay. The protein expressions of mitochondrial apoptosis pathway proteins such as AIF, Cyt-C, cleaved caspase-3, Bcl-2, Bax were determined by Western blot. Results HUVECs exposed to PA at 0.4 mmol·L~(-1) for 24 h showed a decrease in their viability and an increase in the level of AIF, Cyt-C, cleaved caspase-3, Bax/Bcl-2(P<0.05). Cell apoptosis level was significantly up-regulated(P<0.05). The intracellular apoptosis levels in the vascular endothelial cells treatment with mitochondrial apoptosis pathway inhibitor ciclosporine A(CsA) were significantly lower than those of PA group(P<0.05). Conclusion Activated mitochondrial apoptosis pathway might play an important role in PA-induced cell apoptosis in vascular endothelial cells.
Aim To explore the role of mitochondrial apoptosis pathway in palmitic acid(PA)-induced cell apoptosis in human umbilical vein endothelial cells(HUVECs). Methods HUVECs were exposed to different concentrations of PA(0.1, 0.2, 0.4, 0.8 mmol·L~(-1)) for 24 h and different time points of 0.4 mmol·L~(-1) PA(0, 12, 24, 48 h). Cell viability was measured by MTT. The expression of reactive oxygen species(ROS) in HUVECs was detected by immunofluorescence. The level of intracellular apoptosis was detected by TUNEL assay. The protein expressions of mitochondrial apoptosis pathway proteins such as AIF, Cyt-C, cleaved caspase-3, Bcl-2, Bax were determined by Western blot. Results HUVECs exposed to PA at 0.4 mmol·L~(-1) for 24 h showed a decrease in their viability and an increase in the level of AIF, Cyt-C, cleaved caspase-3, Bax/Bcl-2(P<0.05). Cell apoptosis level was significantly up-regulated(P<0.05). The intracellular apoptosis levels in the vascular endothelial cells treatment with mitochondrial apoptosis pathway inhibitor ciclosporine A(CsA) were significantly lower than those of PA group(P<0.05). Conclusion Activated mitochondrial apoptosis pathway might play an important role in PA-induced cell apoptosis in vascular endothelial cells.
引文
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[13] Meng X, Gao X, Zhang Z,et al. Protective effect and mechanism of rat recombinant S100 calcium-binding protein A4 on oxidative stress injury of rat vascular endothelial cells [J]. Oncol Lett, 2018, 16(3): 3614-22.
[14] Su G, Sun G, Liu H,et al. Downregulation of miR-34a promotes endothelial cell growth and suppresses apoptosis in atherosclerosis by regulating Bcl-2 [J]. Heart Vessels, 2018, 33(10): 1185-94.
[15] Yu X, Yu R Q, Gui D,et al. Hexavalent chromium induces oxidative stress and mitochondria-mediated apoptosis in isolated skin fibroblasts of Indo-Pacific humpback dolphin [J]. Aquat Toxicol, 2018, 203:179-86.
[2] 祝晓庆, 杨洁, 邵娟,等. 复方总黄酮对ApoE基因敲除小鼠动脉粥样硬化的影响[J]. 中国药理学通报, 2017, 33(2): 180-4. Zhu X Q, Yang J, Shao J,et al. Effect of compound flavonoids on atherosclerosis in ApoE-/- mice [J]. Chin Pharmacol Bull, 2017, 33(2): 180-4.
[3] Morton J, Bao S, Vanags L Z,et al. Strikingly different atheroprotective effects of apolipoprotein A-I in early-versus late-stage atherosclerosis [J]. JACC Basic Transl Sci, 2018, 3(2): 187-99.
[4] Wang H M, Gao J H, Lu J L. Pravastatin improves atherosclerosis in mice with hyperlipidemia by inhibiting TREM-1/DAP12 [J]. Eur Rev Med Pharmacol Sci, 2018, 22(15): 4995-5003.
[5] 杜瑞雪,叶平,蔡剑鸣,等. 长期调脂治疗对颈动脉粥样硬化斑块成分的影响[J]. 中华老年心脑血管病杂志, 2018, 20(7): 696-9. Du R X, Ye P, Cai J M,et al. Effect of long-term lipid lowering therapy on components of carotid atherosclerotic plaques [J]. Chin J Geriatr Heart Brain Ves Dis, 2018, 20(7): 696-9.
[6] 王艳春,王雪梅,秦和伟,刘俊保. 基于血脂代谢紊乱探讨益气调脂八味汤对CD40-CD40L系统的调控作用[J]. 中华中医药学刊, 2017, 35(6): 1509-12. Wang Y C, Wang X M, Qin H W,Liu J B. Investigation on Yiqi Tiaozhi Bawei decoction on CD40-CD40L system based on dyslipidemia [J]. Chin Arch Tradit Chin Med, 2017, 35(6): 1509-12.
[7] 侯卓奇, 李刚, 张至. 高密度脂蛋白胆固醇的前世今生和未来[J]. 中华高血压杂志, 2017, 25(12): 1185-9. Hou Z Q, Li G, Zhang Z. The past and present of high-density lipoprotein cholesterol [J]. Chin J Hypertens, 2017, 25(12): 1185-9.
[8] 黄馥菡, 施向东, 范德墉,等. 血浆活化蛋白C与高血压肾病患者动脉粥样硬化的相关性[J]. 中华高血压杂志, 2016, 24(5): 470-2. Huang F H, Shi X D, Fan D Y,et al. Correlation between plasma activated protein C and atherosclerosis in patients with hypertensive nephropathy [J]. Chin J Hypertens, 2016, 24(5): 470-2.
[9] Yan X, Gou Z, Li Y,et al. Fibroblast growth factor 21 inhibits atherosclerosis in apoE-/- mice by ameliorating Fas-mediated apoptosis[J]. Lipids Health Dis, 2018, 17(1): 203.
[10] Zhang W, Feng J, Cheng B,et al. Oleanolic acid protects against oxidative stress-induced human umbilical vein endothelial cell injury by activating AKT/eNOS signaling [J]. Mol Med Rep, 2018, 18(4): 3641-8.
[11] Wang X, Mao R, Chen W. FSD-C10 shows therapeutic effects in suppressing oxidized low-density lipoprotein (ox-LDL)-induced human brain microvascular endothelial cells apoptosis via Rho-associated coiled-coil kinase (ROCK)/mitogen- activated protein kinase (MAPK) signaling [J]. Med Sci Monit, 2018, 24: 5509-16.
[12] Sui X, Yu S, Dou L, et al. miR-291b-3p mediated ROS-induced endothelial cell dysfunction by targeting HUR [J]. Int J Mol Med, 2018, 42(5): 2383-92.
[13] Meng X, Gao X, Zhang Z,et al. Protective effect and mechanism of rat recombinant S100 calcium-binding protein A4 on oxidative stress injury of rat vascular endothelial cells [J]. Oncol Lett, 2018, 16(3): 3614-22.
[14] Su G, Sun G, Liu H,et al. Downregulation of miR-34a promotes endothelial cell growth and suppresses apoptosis in atherosclerosis by regulating Bcl-2 [J]. Heart Vessels, 2018, 33(10): 1185-94.
[15] Yu X, Yu R Q, Gui D,et al. Hexavalent chromium induces oxidative stress and mitochondria-mediated apoptosis in isolated skin fibroblasts of Indo-Pacific humpback dolphin [J]. Aquat Toxicol, 2018, 203:179-86.
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