Objective- Angiogenesis, entire step from endothelial cells (ECs) sprouts to vascular maturation, is a critical response to ischemia. To form functional mature vessels, interactions between ECs and pericytes are essential. Ninj1 (ninjurin1) is an adhesion molecule that contributes to the pathogenesis of neuroinflammation. We recently demonstrated that Ninj1 is expressed in pericytes during angiogenesis. However, the role of Ninj1 in angiogenesis under pathophysiological ischemic conditions has not yet been elucidated. Approach and Results- Ninj1 was detected in microvessels, and its expression was enhanced in ischemic tissues after mouse hindlimb ischemia. Knockdown of Ninj1 was performed by injection of biodegradable microspheres releasing Ninj1-small interfering RNA into muscle tissues. Alternatively, pericyte-specific Ninj1 knockout was induced by tamoxifen treatment of NG2-CreERT/Ninj1-flox mice. Ninj1 knockdown/knockout reduced the formation of blood-circulating functional vessels among total CD31+ microvessels within ischemic tissues and subsequently attenuated color Doppler-assessed blood flow recovery. Ninj1 overexpression enhanced expression of Anpt (angiopoietin) 1, whereas Ninj1 knockdown enhanced the endogenous Anpt1 antagonist, Anpt2 expression in pericytes and inhibited the association of pericytes with ECs and subsequent formation of capillary-like structure, that is, EC tube surrounded with pericytes in 3-dimensional gel culture. Conclusions- Our data demonstrate that Ninj1 is involved in the formation of functional matured vessels through the association between pericytes and ECs, resulting in blood flow recovery from ischemia. These findings further the current our understanding of vascular maturation and may support the development of therapeutics for ischemic diseases.
Keywords: angiogenesis; endothelial cells; ischemia; pericytes; peripheral vascular diseases.