Thrombospondin-1 Activation of Signal-Regulatory Protein-α Stimulates Reactive Oxygen Species Production and Promotes Renal Ischemia Reperfusion Injury

Ischemia reperfusion injury (IRI) causes tissue and organ injury, in part, through alterations in tissue blood flow and the production of reactive oxygen species. The cell surface receptor signal-regulatory protein-alpha (SIRP-alpha) is expressed on inflammatory cells and suppresses phagocytosis, but the function of SIRP-alpha in IRI has not been determined. We reported previously that the matricellular protein thrombospondin-1 is upregulated in IRI. Here, we report a novel interaction between thrombospondin-1 and SIRP-alpha on nonphagocytic cells. In cell-free experiments, thrombospondin-1 bound SIRP-alpha. In vascular smooth muscle cells and renal tubular epithelial cells, treatment with thrombospondin-1 led to phosphorylation of SIRP-alpha and downstream activation of Src homology domain 2 containing phosphatase-1. Thrombospondin-1 also stimulated phosphorylation of p47(Phox) (an organizer subunit for nicotinamide adenine dinucleotide phosphate (NADPH) oxidase 1/2) and increased production of superoxide, both of which were abrogated by knockdown or antibody blockade of SIRP-alpha. In rodent aortic rings, treatment with thrombospondin-1 increased the production of superoxide and inhibited nitric oxide mediated vasodilation in a SIRP-alpha dependent manner. Renal IRI upregulated the thrombospondin-1 SIRP-alpha signaling axis and was associated with increased superoxide production and cell death. A SIRP-alpha antibody that blocks thrombospondin-1 activation of SIRP-alpha mitigated the effects of renal IRI, increasing blood flow, suppressing production of reactive oxygen species, and preserving cellular architecture. A role for CD47 in SIRP-alpha activation in these pathways is also described. Overall, these results suggest that thrombospondin-1 binding to SIRP-alpha on nonphagocytic cells activates NADPH oxidase, limits vasodilation, and promotes renal IRI.