Department of Cardiology and Angiology, University Hospital Giessen and Marburg, D‑35392 Giessen, Germany.
Department of Radiotherapy, University Hospital Essen, D‑45147 Essen, Germany.
Int J Mol Med. 2018 Nov;42(5):2811-2818. doi: 10.3892/ijmm.2018.3828. Epub 2018 Aug 16.
In addition to being an important component of the gap junction, connexin 43 (Cx43) has been shown to regulate other cellular functions, including cell proliferation. This regulatory role of Cx43 may be important in therapeutic situations, including wound healing or ischemic injuries. Caveolin‑1 (Cav‑1) has been shown to regulate angiogenesis. The aim of the present study was to analyze whether Cx43 counter‑regulates Cav‑1 in controlling the proliferation and migration of endothelial cells. The inhibition of Cx43 with niflumic acid, flufenamic acid and 18‑α‑glycyrrhetinic acid in cultured human umbilical vein endothelial cells resulted in decreased phosphorylation of extracellular signal‑regulated kinase (ERK)1/2 and increased expression of Cav‑1, as shown by western blot analysis. Furthermore, the inhibition of Cx43 resulted in a 50±7% decrease in cell proliferation, determined using a crystal violet assay, a 48±5% decrease in migration, determined using a migration assay, and a 49±6% decrease in endothelial tube formation, determined using a Matrigel assay, compared with the control. Similar results were obtained following specific inhibition of Cx43 by mimetic peptides (Gap26 and Gap27). Inhibition of the mitogen‑activated protein kinase kinase/ERK pathway with PD‑98059 resulted in an increased expression of Cav‑1 and a reduction in the expression of Cx43. Furthermore, cell proliferation, migration and tube formation in endothelial cells were impaired. By contrast, downregulation of the protein expression of Cav‑1 by small interference RNA resulted in increased expression of Cx43 and phosphorylation of ERK1/2. Accordingly, the number of cells in the Cav‑1 treated‑group increased by 35±5% compared with the controls. The data of the present study showed that Cav‑1 suppressed cell proliferation by inhibiting the activity of Cx43, which is upstream of ERK1/2. The downregulation of Cav‑1 protein resulted in loss of the inhibitory activity of Cav‑1 on cell proliferation and led to increased cell proliferation. This counter‑regulatory effect of Cx43 may be of importance in therapeutic angiogenesis.
除了作为间隙连接的重要组成部分外，连接蛋白 43（Cx43）已被证明可调节其他细胞功能，包括细胞增殖。Cx43 的这种调节作用在治疗情况下可能很重要，包括伤口愈合或缺血性损伤。窖蛋白 1（Cav-1）已被证明可调节血管生成。本研究旨在分析 Cx43 是否通过调节 Cav-1 来控制内皮细胞的增殖和迁移。用尼氟酸、氟芬那酸和 18-α-甘草次酸抑制培养的人脐静脉内皮细胞中的 Cx43，通过 Western blot 分析显示，细胞外信号调节激酶（ERK）1/2 的磷酸化减少，Cav-1 的表达增加。此外，用结晶紫测定法测定，细胞增殖减少 50±7%；用迁移测定法测定，迁移减少 48±5%；用 Matrigel 测定法测定，内皮管形成减少 49±6%，与对照组相比。用模拟肽（Gap26 和 Gap27）特异性抑制 Cx43 也得到了类似的结果。用 PD-98059 抑制丝裂原激活的蛋白激酶激酶/ERK 通路导致 Cav-1 表达增加和 Cx43 表达减少。此外，内皮细胞的增殖、迁移和管形成受损。相比之下，通过小干扰 RNA 下调 Cav-1 的蛋白表达导致 Cx43 的表达增加和 ERK1/2 的磷酸化。因此，与对照组相比，Cav-1 处理组的细胞数量增加了 35±5%。本研究的数据表明，Cav-1 通过抑制 ERK1/2 上游的 Cx43 活性来抑制细胞增殖。Cav-1 蛋白的下调导致 Cav-1 对细胞增殖的抑制活性丧失，导致细胞增殖增加。Cx43 的这种反向调节作用在治疗性血管生成中可能很重要。
