Combination of antiangiogenesis and immunotherapy may be an effective strategy for treatment of solid tumors. Our previous work reported that activation of CD137 signaling promotes intraplaque angiogenesis. A number of studies have demonstrated that vascular endothelial growth factor receptor 2 (VEGFR2) is a key target for angiogenesis. However, it is unknown whether CD137-mediated angiogenesis is related to VEGFR2. In this study, we investigated the effect of CD137 on the VEGFR2 expression and explored the underlying mechanisms of CD137-mediated angiogenesis. Knock-out of CD137 in ApoE^-/- mice significantly decreased neovessel density in atherosclerotic plaques. CD137 silencing or inhibition attenuated endothelial cell (ECs) proliferation, migration, and tube formation. We found activation of CD137 signaling for increased VEGFR2 transcription and translation steadily. Moreover, CD137 signaling activated phosphorylated VEGFR2 (Tyr1175) and the downstream Akt/eNOS pathway, whereas neutralizing CD137 signaling weakened the activation of VEGFR2 and the downstream Akt/eNOS pathway. The aortic ring assay further demonstrated that CD137 signaling promoted ECc sprouting. Inhibition of VEGFR2 by siRNA or XL184 (cabozantinib) and inhibition of downstream signaling by LY294002 (inhibits AKT activation) and L-NAME (eNOS inhibitor) remarkably abolished proangiogenic effects of CD137 signaling both in vitro and ex vivo. In addition, the condition medium from CD137-activated ECs and vascular endothelial growth factor A (VEGFA) had similar effects on ECs that expressed high VEGFR2. Additionally, activating CD137 signaling promoted endothelial secretion of VEGFA, while blocking CD137 signaling attenuated VEGFA secretion. In conclusion, activation of CD137 signaling promoted sprouting angiogenesis by increased VEGFA secretion and the VEGFR2/Akt/eNOS pathway. These findings provide a basis for stabilizing intraplaque angiogenesis through VEGFR2 intervatioin, as well as cancer treatment via combination of CD137 agonists and specific VEGFR2 inhibitors.

中文翻译：

---

激活 CD137 信号传导通过增加 VEGFA 分泌和 VEGFR2/Akt/eNOS 途径促进发芽血管生成

联合抗血管生成和免疫治疗可能是治疗实体瘤的有效策略。我们之前的工作报告了 CD137 信号的激活促进斑块内血管生成。许多研究表明，血管内皮生长因子受体 2 (VEGFR2) 是血管生成的关键目标。然而，尚不清楚 CD137 介导的血管生成是否与 VEGFR2 相关。在这项研究中，我们研究了 CD137 对 VEGFR2 表达的影响，并探讨了 CD137 介导的血管生成的潜在机制。敲除 ApoE 中的 CD137 ^-/-小鼠显着降低动脉粥样硬化斑块中的新血管密度。CD137 沉默或抑制减弱了内皮细胞 (EC) 的增殖、迁移和管形成。我们发现 CD137 信号的激活稳定增加了 VEGFR2 的转录和翻译。此外，CD137 信号激活磷酸化 VEGFR2 (Tyr1175) 和下游 Akt/eNOS 通路，而中和 CD137 信号减弱了 VEGFR2 和下游 Akt/eNOS 通路的激活。主动脉环试验进一步证明 CD137 信号促进了 ECc 的萌发。siRNA 或 XL184（卡博替尼）对 VEGFR2 的抑制以及 LY294002（抑制 AKT 激活）和 L-NAME（eNOS 抑制剂）对下游信号的抑制显着消除了体外和离体 CD137 信号传导的促血管生成作用。此外，来自 CD137 激活的 ECs 和血管内皮生长因子 A (VEGFA) 的条件培养基对表达高 VEGFR2 的 ECs 具有相似的影响。此外，激活 CD137 信号促进了 VEGFA 的内皮分泌，而阻断 CD137 信号减弱了 VEGFA 的分泌。总之，CD137 信号传导的激活通过增加 VEGFA 分泌和 VEGFR2/Akt/eNOS 途径促进发芽血管生成。这些发现为通过 VEGFR2 干预稳定斑块内血管生成以及通过 CD137 激动剂和特定 VEGFR2 抑制剂的组合治疗癌症提供了基础。同时阻断 CD137 信号减弱了 VEGFA 的分泌。总之，CD137 信号传导的激活通过增加 VEGFA 分泌和 VEGFR2/Akt/eNOS 途径促进发芽血管生成。这些发现为通过 VEGFR2 干预稳定斑块内血管生成以及通过 CD137 激动剂和特定 VEGFR2 抑制剂的组合治疗癌症提供了基础。同时阻断 CD137 信号减弱了 VEGFA 的分泌。总之，CD137 信号传导的激活通过增加 VEGFA 分泌和 VEGFR2/Akt/eNOS 途径促进发芽血管生成。这些发现为通过 VEGFR2 干预稳定斑块内血管生成以及通过 CD137 激动剂和特定 VEGFR2 抑制剂的组合治疗癌症提供了基础。