普萘洛尔对体外培养血管瘤内皮细胞血管内皮生长因子和基质金属蛋白酶-2表达的影响

The influence of propranolol on expression of vascular endothelial growth factor and matrix metalloproteinases-2 from in vitro hemangioma endothelial cells

目的观察普萘洛尔对体外培养血管瘤内皮细胞血管内皮生长因子、基质金属蛋白酶一2的影响，探讨普萘洛尔治疗血管瘤的机制。方法用组织块结合酶消化法原代培养血管瘤内皮细胞，并用第Ⅷ凝血因子相关抗原进行鉴定。待细胞处于对数生长期，换无血清培养基孵育48h使细胞同步化，然后分别加入0．5μmol／L和1μmol／L的普萘洛尔作为干预组，而对照组不加任何干预因素，继续孵育24h，用Real—timePCR法检测血管内皮生长因子、基质金属蛋白酶-2的表达水平，同期检测血管瘤血管内皮细胞凋亡率，并分析血管内皮生长因子mRNA、基质金属蛋白酶-2mRNA的表达水平与血管瘤血管内皮细胞凋亡的相关性。结果0．5μmol／L普萘洛尔组和1．0μmol／L普萘洛尔组血管瘤内皮细胞凋亡率分别为（22．42±0．83）％，（24．36±1．42）％，比对照组（15．72±0．59）％明显升高（P〈0．01）；血管内皮生长因子mRNA相对表达量分别为1．02±0．42、0．93±0．22，比对照组1．61±0．52降低（P〈0．05），基质金属蛋白酶-2mRNA相对表达量分别为0．77±0．18、0．67±0．27，比对照组1．47±0．57降低（P〈0．05），但0．5μmol／L普萘洛尔组血管内皮生长因子mRNA、基质金属蛋白酶-2mRNA与1．0μmol／L普萘洛尔组比较，差异均无统计学意义（P〉0．05）。结论普萘洛尔可以促进体外培养的血管瘤内皮细胞凋亡，其机制可能是通过下调血管瘤内皮细胞血管内皮生长因子、基质金属蛋白酶-2的表达，从而促进血管瘤内皮细胞凋亡。

Objective To investigate the influence of propranolol on expression of vascular en- dothelial growth factor （VEGF） and matrix metalloproteinases-2 （MMP-2） from in vitro hemangioma endothelial cells, aiming to explore the mechanism of propranolol in treatment of hemangioma. Methods The hemangioma vascular endothelial cells were harvested from the specimens arising from infant with hemangioma, and cultured via tissue explants combined with trypsin digestion approach. The ex- pression of the factor W related antigen in endothelial cells was identified via immunohistochemistry assay. When cells were in the logarithmic phase, the serum-free medium was added and incubated for 48 hours for synchronization. 0. 5 μmol/L and 1.0 μmol/L concentration of propranolol was added respectively and 0μmol/L group was set for control. The expression level of VEGF and MMP-2 were detected using RT-PCR after 24 hours incubation, meanwhile the cell apoptosis in each group was de- tected using flow cytometry, then further analysis the correlation between the apoptosis and expression of them. Results The apoptosis rate in 0. 5μmol/L and 1.0μmol/L group was respectively 22. 42 ± 0. 83 and 24. 36 ± l. 42 percent, compared with control （15.72± 0. 59 percent）, they were significantly increased （P〈0. 01 ）. While in term of VEGF mRNA expression level, they were respectively 1.02 ±11. 42 and 0. 93 ± 0. 22 in （1. 5 μmol/L and 1.0μmol/L group. Compared with the control （1.61 ± 0.52）. they decreased remarkably. Regarding to the MMP-2 mRNA expression level, they were respectivc, ly 0. 77 ± 0. 18 and 0. 67 ± 0. 27 in 0. 5 μmol/L and 1.0 μmol/L group. In contrast to control （1.47 ± 0. 57）, they also decreased significantly. However, compared with the 0. 5μmol/L and 1.0 μmol/L groups, statistical difference in both VEGF and MMP-2 was not seen （P）0. 05）. Conclusions Propranolol can promote apoptosis of in vitro hemangioma vascular endothelial cells. The potential mechanism is that they can down-regulate the expression of VEGF and MMP-2, through which the cell apoptosis occurs.