Shandong Provincial Key Laboratory of Animal Cells and Developmental Biology, School of Life Science, Shandong University, Ji-nan, China.
Acta Pharmacol Sin. 2011 Feb;32(2):209-16. doi: 10.1038/aps.2010.201.
To investigate the effect of N-benzyl-5-phenyl-1H-pyrazole-3-carboxamide (BPC) on angiogenesis in human umbilical vein endothelial cells (HUVECs).
Capillary-like tube formation on matrigel and cell migration analyses were performed in the absence of serum and fibroblast growth factor (FGF-2). Reactive oxygen species (ROS) were measured using a fluorescent probe, 2', 7'- dichlorodihydrofluorescein (DCHF). The nitric oxide (NO) production of HUVECs was examined using a NO detection kit. Morphological observation under a phase contrast microscope, a viability assay using 3-[4, 5-dimethylthiazol-2-yl]-2, 5-diphenyl-tetrazolium (MTT) and a lactate dehydrogenase (LDH) activity analysis by a detection kit were performed to evaluate the toxicity of BPC on HUVECs in the presence of serum and FGF-2. The level of hypoxia-inducible factor 1α (HIF-1α) and the release of vascular endothelial growth factor (VEGF) were measured by Western blot and ELISA, respectively.
In the absence of serum and FGF-2, cells treated with BPC (5-20 μmol/L) rapidly aligned with one another and formed tube-like structures within 12 h. In the presence of serum and FGF-2, cells treated with BPC for 24, 48 and 72 h had no changes in morphology, viability or LDH release compared with the control group. Cell migration in the BPC-treated group was significantly increased compared with the control group. During this process, NO production and ROS level were elevated dramatically, and the levels of HIF-1α and VEGF were increased dependent on the generation of ROS.
BPC most effectively promoted angiogenesis and migration in HUVECs in the absence of FGF-2 and serum.
研究 N-苄基-5-苯基-1H-吡唑-3-甲酰胺（BPC）对人脐静脉内皮细胞（HUVEC）血管生成的影响。
在无血清和成纤维细胞生长因子（FGF-2）的情况下，通过在基质胶上进行毛细血管样管形成和细胞迁移分析来研究。使用荧光探针 2'，7'-二氯二氢荧光素（DCHF）测量活性氧（ROS）。使用一氧化氮（NO）检测试剂盒检查 HUVEC 的 NO 产生。通过相差显微镜进行形态观察，使用 3-[4,5-二甲基噻唑-2-基]-2,5-二苯基四唑（MTT）进行细胞活力测定以及使用检测试剂盒进行乳酸脱氢酶（LDH）活性分析，评估在血清和 FGF-2 存在下 BPC 对 HUVEC 的毒性。通过 Western blot 和 ELISA 分别测量缺氧诱导因子 1α（HIF-1α）的水平和血管内皮生长因子（VEGF）的释放。
在无血清和 FGF-2 的情况下，用 BPC（5-20 μmol/L）处理的细胞在 12 小时内迅速彼此对齐并形成管状结构。在有血清和 FGF-2 的情况下，与对照组相比，用 BPC 处理 24、48 和 72 小时的细胞形态、活力或 LDH 释放均无变化。BPC 处理组的细胞迁移显著增加。在此过程中，NO 产生和 ROS 水平显著升高，HIF-1α 和 VEGF 的水平随 ROS 的产生而增加。
在无 FGF-2 和血清的情况下，BPC 最有效地促进了 HUVEC 的血管生成和迁移。
