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目的:观察三氧化二砷(As2O3)对血管内皮细胞增殖、凋亡及VCAM-1/ICAM-1表达的影响,探讨As2O3对血管内皮细胞增殖生长以及炎症反应的影响。方法:人脐静脉内皮细胞(HUVEC)体外培养,以不同As2O3浓度及时间对其进行干预。采用CCK-8测定细胞增殖活性,流式细胞仪AnnexinⅤ/PI双染法检测细胞的凋亡率,实时荧光定量PCR检测VCAM-1mRNA表达,酶联免疫吸附试验(ELISA)检测细胞间黏附分子(VCAM-1)及血管细胞黏附分子(ICAM-1)的表达情况。结果:当As2O3浓度在3μmol.L-1时HUVEC培养24 h的的凋亡率为(0.134±0.03)%,48 h为(3.305±0.53)%,72 h为(3.748±0.84)%(P<0.05),凋亡率均在一较低水平。当As2O3浓度>3μmol.L-1时HUVEC凋亡率明显增加(P<0.01)。不同浓度As2O3作用HUVEC48 h后检测上清液中ICAM-1与VCAM-1浓度时发现1μmol.L-1时VCAM-1表达即开始增加(123.32±3.78 mmol.L-1,P<0.01),而HUVEC表达ICAM-1含量与对照组相比差异并不明显(38.94±2.59 mmol.L-1,P>0.05),随着As2O3浓度的增加,HUVEC表达ICAM-1/VCAM-1的量均增加但敏感性不同。对照组及(1.0、2.0、3.0、4.0、5.0)μmol.L-1As2O3作用于HUVEC 48 h实时荧光定量PCR法检测VCAM-1mRNA表达量明显增加,与对照组相比实验组的表达量分别为(1.657±0.287,1.858±0.241,2.321±0.280,3.012±0.235,3.508±0.342)(P<0.01)。结论:As2O3可直接降低细胞活性,诱导细胞凋亡,并且呈一定的时间-浓度依赖性。在较低浓度时VCAM-1/ICAM-1的表达在一个相对较低的水平,随着As2O3浓度的逐渐升高,内皮细胞凋亡率增高,VCAM-1/ICAM-1表达增加,并且VCAM-1/ICAM-1对As2O3的敏感性呈现一定的差异性。
Objective: To observe the effects of arsenic trioxide (As2O3) on the proliferation and apoptosis of vascular endothelial cells and the expression of VCAM-1 / ICAM-1, and to explore the effect of As2O3 on the proliferation and the proliferation of vascular endothelial cells. Methods: Human umbilical vein endothelial cells (HUVECs) were cultured in vitro with different As2O3 concentration and time intervention. The cell proliferation activity was measured by CCK-8, the apoptosis rate was detected by flow cytometry Annexin Ⅴ / PI double staining, the expression of VCAM-1 mRNA was detected by real-time fluorescence quantitative PCR, and the expression of intercellular adhesion molecule VCAM-1) and vascular cell adhesion molecule (ICAM-1) expression. Results: The apoptotic rates of HUVEC cultured for 24 h were (0.134 ± 0.03)% and (3.305 ± 0.53)% for 48 h and (3.748 ± 0.84)% for 72 h, respectively, when the concentration of As2O3 was 3 μmol·L-1 <0.05), the apoptosis rate was at a lower level. When As2O3 concentration> 3μmol.L-1, the apoptosis rate of HUVEC increased significantly (P <0.01). When the concentrations of ICAM-1 and VCAM-1 in supernatant were detected by As2O3 after 48 h of HUVEC, the expression of VCAM-1 began to increase (123.32 ± 3.78 mmol.L-1, P <0.01) when 1 μmol·L-1 was detected. The level of ICAM-1 in HUVEC was not significantly different from that in control group (38.94 ± 2.59 mmol.L-1, P> 0.05). The amount of ICAM-1 / VCAM-1 expressed in HUVEC increased with the increase of As2O3 concentration Increased but different sensitivities. Compared with the control group, the expression of VCAM-1 mRNA in the control group and (1.0,2.0,3.0,4.0,5.0) μmol.L-1As2O3 treated with HUVEC for 48 h by real-time fluorescence quantitative PCR increased significantly compared with the control group (1.657 ± 0.287,1.858 ± 0.241,2.321 ± 0.280,3.012 ± 0.235,3.508 ± 0.342) (P <0.01). Conclusion: As2O3 can directly decrease cell viability and induce apoptosis, and in a time-concentration-dependent manner. At a lower concentration, VCAM-1 / ICAM-1 expression was at a relatively low level. With the gradual increase of As2O3 concentration, the apoptosis rate of endothelial cells increased and the expression of VCAM-1 / ICAM-1 increased. -1 / ICAM-1 As2O3 sensitivity showed some differences.