Pathology and Laboratory Medicine Services, (S-113), Department of Veterans Affairs Puget Sound Health Care System, University of Washington, 1660 S Columbian Way, Seattle, WA 98108, USA.
Arterioscler Thromb Vasc Biol. 2011 May;31(5):1151-9. doi: 10.1161/ATVBAHA.111.223917. Epub 2011 Mar 3.
The goal of this study was to define the role of tumor necrosis factor-α (TNFα) in the cascade of gene activation that regulates aortic angiogenesis in response to injury.
Angiogenesis was studied by culturing rat or mouse aortic rings in collagen gels. Gene expression was evaluated by quantitative reverse transcription-polymerase chain reaction, microarray analysis, immunocytochemistry, and ELISA. TNFα gene disruption and recombinant TNFα or blocking antibodies against vascular endothelial growth factor (VEGF) or TNF receptors were used to investigate TNFα-mediated angiogenic mechanisms. Resident aortic macrophages were depleted with liposomal clodronate. Angiogenesis was preceded by overexpression of TNFα and TNFα-inducible genes. Studies with isolated cells showed that macrophages were the main source of TNFα. Angiogenesis, VEGF production, and macrophage outgrowth were impaired by TNFα gene disruption and promoted by exogenous TNFα. Antibody-mediated inhibition of TNF receptor 1 significantly inhibited angiogenesis. The proangiogenic effect of TNFα was suppressed by blocking VEGF or by ablating aortic macrophages. Exogenous TNFα, however, maintained a limited proangiogenic capacity in the absence of macrophages and macrophage-mediated VEGF production.
Overexpression of TNFα is required for optimal VEGF production and angiogenesis in response to injury. This TNFα/VEGF-mediated angiogenic pathway requires macrophages. The residual capacity of TNFα to stimulate angiogenesis in macrophage-depleted aortic cultures implies the existence of a VEGF-independent alternate pathway of TNFα-induced angiogenesis.
本研究旨在确定肿瘤坏死因子-α（TNFα）在调节损伤后主动脉血管生成的基因激活级联反应中的作用。
通过在胶原凝胶中培养大鼠或小鼠主动脉环来研究血管生成。通过定量逆转录聚合酶链反应、微阵列分析、免疫细胞化学和 ELISA 评估基因表达。使用 TNFα 基因敲除和重组 TNFα或针对血管内皮生长因子（VEGF）或 TNF 受体的阻断抗体来研究 TNFα 介导的血管生成机制。用脂质体氯膦酸盐耗尽固有主动脉巨噬细胞。血管生成之前是 TNFα 和 TNFα 诱导基因的过度表达。对分离细胞的研究表明，巨噬细胞是 TNFα 的主要来源。TNFα 基因敲除和外源性 TNFα促进血管生成、VEGF 产生和巨噬细胞增生，而 TNFα 基因敲除则抑制血管生成。TNF 受体 1 的抗体介导抑制显著抑制血管生成。TNFα 的促血管生成作用被阻断 VEGF 或消除主动脉巨噬细胞所抑制。然而，在外源 TNFα 存在的情况下，在没有巨噬细胞和巨噬细胞介导的 VEGF 产生的情况下，仍保持有限的促血管生成能力。
TNFα 的过表达是损伤后最佳 VEGF 产生和血管生成所必需的。这种 TNFα/VEGF 介导的血管生成途径需要巨噬细胞。在耗尽巨噬细胞的主动脉培养物中 TNFα 刺激血管生成的剩余能力暗示了 TNFα 诱导的血管生成存在一种 VEGF 非依赖的替代途径。
