Shear stress activation of nitric oxide synthase and increased NO levels in human red blood cells

Since the discovery of nitric oxide (NO) as a vasoactive molecule, red blood cells (RBC) have been considered to participate in NO-mediated control of the circulation. The classical role attributed to RBC was scavenging of NO, thereby impacting the local bioavailability of this important regulator of vascular tone^1^. RBC have been shown to be a source of NO, primarily via its transport bound to haemoglobin^2, 3^. Under specific conditions, haemoglobin plays an active role in converting NO derivatives (e.g., nitrite) to NO^4, 5^, with this NO originating from RBC being an effective modulator of vascular smooth muscle tone^6^. Interestingly, RBC contain a NO synthase (NOS) protein^7^, can actively synthesize NO using L-arginine as a substrate^8^, and can export NO under appropriate conditions^8, 9^. It has been previously hypothesized that RBC NOS may be activated by shear forces acting on the cell^10^, and we have recently shown that RBC NOS phosphorylation can be enhanced by subjecting RBC in suspension to shearing forces^9^: NO concentration in the suspending medium was increased subsequent to flow of RBC suspensions through five [mu]m pores^11^. We have now directly demonstrated increased RBC NOS activity and intracellular NO levels in immobilized RBC exposed to well-defined fluid shear stress. Immunostaining for serine 1177 phosphorylation and the NO-sensitive fluorescent probe diaminofluorescein were employed. Our results suggest that RBC deformation in constricted vessels may increase NO levels and favor vasodilation, thereby providing an important role for RBC in regulating the circulation