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Thioredoxin-interacting protein mediates sustained VEGFR2 signaling in endothelial cells required for angiogenesis.

ABSTRACT: Thioredoxin-interacting protein (TXNIP) is an ?-arrestin protein whose function is important for the regulation of vascular endothelial growth factor receptor 2 (VEGFR2) signaling and endothelial cell survival. Because VEGFR2 is critical for angiogenesis, we explored the role of TXNIP in VEGF-induced angiogenesis.TXNIP knockdown inhibited VEGF-induced endothelial cell tube formation and proliferation in cultured human umbilical vein endothelial cell. To elucidate the mechanism by which TXNIP altered VEGFR2 signaling in human umbilical vein endothelial cell, we studied phosphorylation of VEGFR2, phospholipase C gamma-1 (PLC?1), endothelial NO synthase, and Akt (known as protein kinase B). TXNIP knockdown significantly decreased phosphorylation of VEGFR2 and PLC?1 at times >5 minutes, but phosphorylation was unchanged at 2 minutes, as was Akt and endothelial NO synthase phosphorylation. Cell-surface biotinylation assay showed that TXNIP knockdown significantly attenuated VEGFR2 internalization. These results suggested that TXNIP was required for sustained VEGFR2 signaling, which is mediated largely by internalized VEGFR2. Rab5 knockdown to inhibit the trafficking and fusion of early endosomes significantly blocked VEGF-induced VEGFR2 internalization and phosphorylation of VEGFR2 and PLC?1. Immunofluorescence and coimmunoprecipitation showed that TXNIP was part of a complex that included Rab5 and VEGFR2. Finally, TXNIP knockdown prevented the association of VEGFR2 and Rab5.Our results show that TXNIP is essential for VEGFR2 internalization in Rab5 positive endosomes, which is required for endothelial cell growth and angiogenesis.

SUBMITTER: Park SY

PROVIDER: S-EPMC3629821 | biostudies-literature | 2013 Apr

REPOSITORIES: biostudies-literature

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Thioredoxin-interacting protein mediates sustained VEGFR2 signaling in endothelial cells required for angiogenesis.

Park Shin-Young SY Shi Xi X Pang Jinjiang J Yan Chen C Berk Bradford C BC

Arteriosclerosis, thrombosis, and vascular biology 20130207 4

<h4>Objective</h4>Thioredoxin-interacting protein (TXNIP) is an α-arrestin protein whose function is important for the regulation of vascular endothelial growth factor receptor 2 (VEGFR2) signaling and endothelial cell survival. Because VEGFR2 is critical for angiogenesis, we explored the role of TXNIP in VEGF-induced angiogenesis.<h4>Approach and results</h4>TXNIP knockdown inhibited VEGF-induced endothelial cell tube formation and proliferation in cultured human umbilical vein endothelial cell ...[more]

PMID: 23393387

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Project description:AimsThioredoxin-interacting protein (TXNIP) contributes to cellular redox-state homeostasis via binding and inhibiting thioredoxin (TRX). Increasing evidence suggests that cellular redox homeostasis regulates vascular endothelial growth factor (VEGF)-mediated signaling. This study aims to examine the redox-dependant role of TXNIP in regulating VEGF-mediated S-glutathionylation and angiogenic signaling. TXNIP-knockout mice (TKO) or wild-type (WT) treated with the reduced glutathione (GSH)-precursor, N-acetyl cysteine (WT-NAC, 500?mg/kg) were compared to WT using hypoxia-induced neovascularization model.ResultsIn response to hypoxia, retinas from TKO and WT-NAC mice showed significant decreases in reparative revascularization and pathological neovascularization with similar VEGF expression compared with WT. VEGF failed to stimulate vascular sprouting from aortic rings of TKO compared to WT mice. TKO mice or WT+NAC experienced reductive stress as indicated by twofold increase in TRX reductase activity and fourfold increase in reduced-GSH levels compared with WT. In human microvascular endothelial (HME) cells, VEGF stimulated co-precipitation between vascular endothelial growth factor receptor 2 (VEGFR2) with low molecular weight protein tyrosine phosphatase (LMW-PTP). Silencing TXNIP expression blunted VEGF-induced oxidation of GSH and S-glutathionylation of the LMW-PTP in HME cells. These effects were associated with impaired VEGFR2 phosphorylation that culminated in inhibiting cell migration and tube formation. Overexpression of TXNIP restored VEGFR2 phosphorylation and cell migration in TKO-endothelial cells.InnovationTXNIP expression is required for VEGF-mediated VEGFR2 activation and angiogenic response in vivo and in vitro. TXNIP expression regulates VEGFR-2 phosphorylation via S-glutathionylation of LMW-PTP in endothelial cells.ConclusionOur results provide novel mechanistic insight into modulating TXNIP expression as a potential therapeutic target in diseases characterized by aberrant angiogenesis.

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Thioredoxin-interacting protein mediates sustained VEGFR2 signaling in endothelial cells required for angiogenesis.

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Thioredoxin-interacting protein mediates sustained VEGFR2 signaling in endothelial cells required for angiogenesis.

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