Prostaglandin production by COX-1 and -2-deficient mouse hearts

Nicholas S. Kirkby1,2, Philip D. M. Leadbeater1, Timothy D. Warner2, Jane A. Mitchell1. 1National Heart & Lung Institute, Imperial College, London SW3 6LY, United Kingdom, 2William Harvey Research Institute, Barts& the London School of Medicine, London EC1M 6BQ, United Kingdom.

Introduction It is widely held that COX2 is the primary source of the protective prostaglandin, PGI2, in the cardiovascular system because COX2 but not COX1 inhibition lowers urinary levels of its metabolites. Despite this, in healthy endothelial cells, and many other tissues, COX2 protein is rare in comparison to COX1. We sought to clarify the role of COX1 and COX2 in prostaglandin release in endocardium and whole mouse heart by measuring COX activity in mice deficient in each isoform.

Methods Prostaglandin production was measured in intact pieces of free left ventricular wall (∼10mg), homogenates of whole hearts, or by the interior of ex vivo cannulated left ventricles. All were freshly isolated from male and female wild type, COX1-/- and COX2-/- mice (n = 4-6 per group). Ventricle wall pieces and cannulated hearts were allowed to equilibrate for 60mins in DMEM before replacing the media and allowing PGI2 to accumulate for further 30mins. PGI2 was then measured as its stable metabolite, 6-keto-PGF1a. Separate hearts were homogenised and incubated for 20mins before determination of PGE2 content of the supernatent. Data were analysed by 1-way ANOVA with Dunnett’s post-hoc test.

Results All preparations of hearts from WT mice yielded detectable levels of the prostaglandins of interest. In incubations of intact pieces of left ventricular wall (n = 6) 6-keto-PGF1a accumulation was not altered by COX2-deficiency (WT: 3.56±0.80ng/ml ; COX2: 4.84±0.71ng/ml; p>0.05) but was almost abolished by COX1-deficiency (0.19±0.02ng/ml; p<0.01). In perfused left ventricle preparations (n = 6) 6-keto-PGF1a accumulation was also similar between WT (1.71±0.40ng/ml) and COX2-/- mice hearts (1.30±0.31ng/ml; p>0.05), but significantly reduced in those from COX1-/- mice (0.03±0.00ng/ml; p<0.01) mice. Similarly, PGE2 release by whole heart homogenates (n = 4) was reduced in COX1-/- (WT: 0.69±0.18ng/ml; COX1: 0.14±0.07ng/ml; p<0.05) but not COX2-/- mice (1.13±0.15ng/ml; p>0.05).

Conclusion The primary COX isoform responsible for PGI2 and PGE2 production in whole mouse heart and within the left ventricle is COX1 with COX2 providing little contribution in these healthy tissues.