Shono T, Ono M, Izumi H, Jimi S I, Matsushima K, Okamoto T, Kohno K, Kuwano M
Department of Biochemistry, Kyushu Unviersity School of Medicine, Fukuoka, Japan.
Mol Cell Biol. 1996 Aug;16(8):4231-9. doi: 10.1128/MCB.16.8.4231.
Oxygen radicals are induced under various pathologic conditions associated with neovascularization. Oxygen radicals modulate angiogenesis in cultured human microvascular endothelial cells by an unknown mechanism. Treatment of human microvascular endothelial cells for 15 min with 0.1 to 0.5 mM hydrogen peroxide (H2O2) or 100 U of tumor necrosis factor alpha per ml induced tubular morphogenesis in type I collagen gels. Gel shift assays with nuclear extracts demonstrated that H2O2 increases the binding activities of two transcription factors, NF-kappaB and AP-1, but not of Spl. Tumor necrosis factor alpha increased the binding activities of all three factors. A supershift assay with specific antibodies against JunB, JunD, and c-Jun (Jun family) showed that the antibody against c-Jun supershifted the AP-1 complex after H2O2 treatment. Coadministration of the antisense sequence of NF-kappaB inhibited H2O2-dependent tubular morphogenesis, and the antisense c-Jun oligonucleotide caused partial inhibition. The angiogenic factor responsible for H2O2-induced tubular morphogenesis was examined. Cellular mRNA levels of vascular endothelial growth factor and interleukin-8 (IL-8), but not those of transforming growth factor alpha, were increased after treatment with 0.5 mM H2O2. Coadministration of anti-IL-8 antibody inhibited tubular morphogenesis enhanced by H2O2, and IL-8 itself also enhanced the formation of tube-like structures. Treatment with antisense NF-kappaB oligonucleotide completely blocked H2O2-dependent IL-8 production by endothelial cells. The tubular morphogenesis of vascular endothelial cells after treatment with oxidative stimuli and its possible association with NF-kappaB and IL-8, is examined.
在与新血管形成相关的各种病理条件下会诱导产生氧自由基。氧自由基通过未知机制调节培养的人微血管内皮细胞中的血管生成。用0.1至0.5 mM过氧化氢（H2O2）或每毫升100 U肿瘤坏死因子α处理人微血管内皮细胞15分钟，可诱导I型胶原凝胶中的管状形态发生。用核提取物进行的凝胶迁移试验表明，H2O2增加了两种转录因子NF-κB和AP-1 的结合活性，但不增加Spl的结合活性。肿瘤坏死因子α增加了所有三种因子的结合活性。用针对JunB、JunD和c-Jun（Jun家族）的特异性抗体进行的超迁移试验表明，抗c-Jun抗体在H2O2处理后使AP-1复合物发生超迁移。共给予NF-κB的反义序列可抑制H2O2依赖性管状形态发生，而反义c-Jun寡核苷酸则引起部分抑制。研究了负责H2O2诱导的管状形态发生的血管生成因子。用0.5 mM H2O2处理后，血管内皮生长因子和白细胞介素-8（IL-8）的细胞mRNA水平升高，但转化生长因子α的mRNA水平未升高。共给予抗IL-8抗体可抑制H2O2增强的管状形态发生，IL-8本身也增强了管状结构的形成。用反义NF-κB寡核苷酸处理可完全阻断内皮细胞H2O2依赖性IL-8的产生。研究了氧化刺激处理后血管内皮细胞的管状形态发生及其与NF-κB和IL-8的可能关联。
