LC-MS/MS Confirms That COX-1 Drives Vascular Prostacyclin Whilst Gene Expression Pattern Reveals Non-Vascular Sites of COX-2 Expression

There are two schools of thought regarding the cyclooxygenase (COX) isoformactive in the vasculature. Using urinary prostacyclin markers some groups haveproposed that vascular COX-2 drives prostacyclin release. In contrast, we andothers have found that COX-1, not COX-2, is responsible for vascularprostacyclin production. Our experiments have relied on immunoassays to detectthe prostacyclin breakdown product, 6-keto-PGF_1α and antibodies todetect COX-2 protein. Whilst these are standard approaches, used by manylaboratories, antibody-based techniques are inherently indirect and have beencriticized as limiting the conclusions that can be drawn. To address thisquestion, we measured production of prostanoids, including6-keto-PGF_1α, by isolated vessels and in the circulationin vivo using liquid chromatography tandem massspectrometry and found values essentially identical to those obtained byimmunoassay. In addition, we determined expression from theCox2 gene using a knockin reporter mouse in whichluciferase activity reflects Cox2 gene expression. Using thiswe confirm the aorta to be essentially devoid of Cox2 drivenexpression. In contrast, thymus, renal medulla, and regions of the brain and gutexpressed substantial levels of luciferase activity, which correlated well withCOX-2-dependent prostanoid production. These data are consistent with theconclusion that COX-1 drives vascular prostacyclin release and puts the sparseexpression of Cox2 in the vasculature in the context of therest of the body. In doing so, we have identified the thymus, gut, brain andother tissues as target organs for consideration in developing a newunderstanding of how COX-2 protects the cardiovascular system.