肿瘤坏死因子-α (TNFα) 是一种主要细胞因子，可诱导内皮细胞中趋化因子和粘附分子的表达，例如细胞间粘附分子 1 (ICAM-1) 和血管细胞粘附分子 1 (VCAM-1)，从而启动血管炎症反应。在这项研究中，我们确定了神经毡蛋白-1 (NRP1)（几种结构不同的配体的共同受体）作为 TNFα 诱导的内皮细胞炎症反应的调节剂。 NRP1 shRNA 表达抑制 TNFα 刺激的白细胞粘附以及人脐静脉内皮细胞 (HUVEC) 中 ICAM-1 和 VCAM-1 的表达。同样，它减少了 TNFα 诱导的 MAPK p38 磷酸化，但没有显着影响其他 TNF 诱导的信号通路，例如经典的 NFκB 和 AKT 通路。免疫荧光染色证明 NRP1 与 TNF 的两种受体 TNFR1 和 TNFR2 共定位。免疫共沉淀进一步证实NRP1分别与TNFR1和TNFR2位于相同的蛋白复合物或膜区室中。然而，NRP1 表达的调节既不影响细胞膜中的 TNFR 水平，也不影响 TNFα 的受体结合亲和力。尽管从蛋白质对接模型来看，NRP1 和 TNFα/TNFR1 之间的直接界面似乎是可能的，但无细胞微孔板和培养细胞中的结合测定并不支持直接相互作用。此外，在 HUVEC 中，TNFα 通过 TNFR1-NFκB 通路以时间依赖性方式下调 NRP1。总而言之，我们的研究揭示了血管内皮细胞中 NRP1 和 TNFα 之间的新型相互串扰。版权所有 © 2024 Wang, Wang, Anany, Füllsack, Huo, Dutta, Ji, Hoeppner, Kilari, Misra, Caulfield, Vander Kooi, Wajant 和 Mukhopadhyay 。

Tumor necrosis factor-α (TNFα) is a master cytokine which induces expression of chemokines and adhesion molecules, such as intercellular adhesion molecule 1 (ICAM-1) and vascular cell adhesion molecule 1 (VCAM-1), in endothelial cells to initiate the vascular inflammatory response. In this study, we identified neuropilin-1 (NRP1), a co-receptor of several structurally diverse ligands, as a modulator of TNFα-induced inflammatory response of endothelial cells. NRP1 shRNA expression suppressed TNFα-stimulated leukocyte adhesion and expression of ICAM-1 and VCAM-1 in human umbilical vein endothelial cells (HUVECs). Likewise, it reduced TNFα-induced phosphorylation of MAPK p38 but did not significantly affect other TNF-induced signaling pathways, such as the classical NFκB and the AKT pathway. Immunofluorescent staining demonstrated co-localization of NRP1 with the two receptors of TNF, TNFR1 and TNFR2. Co-immunoprecipitation further confirmed that NRP1 was in the same protein complex or membrane compartment as TNFR1 and TNFR2, respectively. Modulation of NRP1 expression, however, neither affected TNFR levels in the cell membrane nor the receptor binding affinities of TNFα. Although a direct interface between NRP1 and TNFα/TNFR1 appeared possible from a protein docking model, a direct interaction was not supported by binding assays in cell-free microplates and cultured cells. Furthermore, TNFα was shown to downregulate NRP1 in a time-dependent manner through TNFR1-NFκB pathway in HUVECs. Taken together, our study reveals a novel reciprocal crosstalk between NRP1 and TNFα in vascular endothelial cells.Copyright © 2024 Wang, Wang, Anany, Füllsack, Huo, Dutta, Ji, Hoeppner, Kilari, Misra, Caulfield, Vander Kooi, Wajant and Mukhopadhyay.