Circulating microparticles carry a functional endothelial nitric oxide synthase that is decreased in patients with endothelial dysfunction.

Abstract

Microparticles (MPs) are circulating membrane particles of less than a micrometer in diameter shed from endothelial and blood cells. Recent literature suggests that MPs are not just functionally inert cell debris but may possess biological functions and mediate the communication between vascular cells. As a significant proportion of MPs originate from platelets and endothelial cells, we hypothesized that MPs may harbor functional enzymes including an endothelial NO synthase (eNOS). Using immunoprecipitation and Western blot analysis, we found that human circulating MPs carry an eNOS. Ca(2+) and l-arginine-dependent NOS activity of crude enzyme extract from MPs was determined by measuring the conversion of [(3)H]-L-arginine to [(3)H]-citrulline and NOS-dependent nitrite production. NOS-dependent NO production in intact MPs was assessed by the NO-specific fluorescent probe MNIP-Cu. In patients with cardiovascular disease, endothelial dysfunction was associated with an increase in the total number of circulating MPs as well as a significant decrease in the expression and activity of eNOS in MPs. No difference in reactive oxygen species was noted in MPs isolated from either group. Our data further support the concept that circulating MPs may not only retain phenotypic markers but also preserve the functionality of enzymes of the cells they originate from, including eNOS.