Endothelial FoxO1 is an intrinsic regulator of thrombospondin 1 expression that restrains angiogenesis in ischemic muscle
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Peripheral artery disease (PAD) is characterized by chronic muscle ischemia. Compensatory angiogenesis is minimal within ischemic muscle despite an increase in angiogenic factors. This may occur due to the prevalence of angiostatic factors. Regulatory mechanisms that could evoke an angiostatic environment during ischemia are largely unknown. Forkhead box O (FoxO) transcription factors, known to repress endothelial cell proliferation in vitro, are potential candidates. Our goal was to determine whether FoxO proteins promote an angiostatic phenotype within ischemic muscle. FoxO1 and the angiostatic matrix protein thrombospondin 1 (THBS1) were elevated in ischemic muscle from PAD patients, or from mice post-femoral artery ligation. Mice with conditional endothelial cell-directed deletion of FoxO proteins (Mx1Cre +, FoxO1,3,4 L/L , referred to as FoxOΔ) were used to assess the role of endothelial FoxO proteins within ischemic tissue. FoxO deletion abrogated the elevation of FoxO1 and THBS1 proteins, enhanced hindlimb blood flow recovery and improved neovascularization in murine ischemic muscle. Endothelial cell outgrowth from 3D explant cultures was more robust in muscles derived from FoxOΔ mice. FoxO1 overexpression induced THBS1 production, and a direct interaction of endogenous FoxO1 with the THBS1 promoter was detectable in primary endothelial cells. We provide evidence that FoxO1 directly regulates THBS1 within ischemic muscle. Altogether, these findings bring novel insight into the regulatory mechanisms underlying the repression of angiogenesis within peripheral ischemic tissues.
KeywordsEndothelium Ischemia Angiogenesis Peripheral artery disease Skeletal muscle Capillary
Peripheral artery disease
Forkhead box O
Capillary to muscle fiber ratio
We appreciate the technical assistance of Ms. Justyna Kopycinska and Mr. Emmanuel Nwadozi with animal surgeries and imaging, and Mr. Sammy Liu with endothelial cell isolations. This research was supported by the Canadian Institutes of Health Research (IMH-107537 to T.L.H.), the Heart and Stroke Foundation of Canada (NA7059 to T.L.H.) and the National Institutes of Health Grant (R01 AG034995 to I.P.).
Conflict of interest
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