[op.3b.02] ageing and lack of estrogens activates cyclooxygenases pathway increasing superoxide anion production in response to thromboxane a2

Objective: The role of ageing in the development of vascular disease is currently associated with a rise in oxidative stress. Thromboxane A2 (TXA2) is an inflammatory and oxidant endogenous vasoconstrictor which action increases with ageing and lack of estrogens, both conditions found in menopause. Our aim was to determine superoxide anion (O2-) levels in response to TXA2 in aortic vascular tissue from senescent and ovariectomized mice and the role of cyclooxygenase as source of O2-. Design and method: Senescence accelerated mice (SAM) were used in this study. 5 month-old SAM prone (SAMP8, n = 7) and SAM resistant (SAMR1, n = 7) were separated in three groups: Sham-operated, ovariectomized (OVX) and ovariectomized treated with estradiol (OVE). Estrogen replacement was given by subcutaneous implant of an Alzet osmotic pump (10 μg/Kg/day of 17ß-estradiol). 28 days after surgery, mice were sacrificed, aorta was isolated and included in OCT. Aortic sections (10 μm-thickness) were incubated with U46619 (10–8 M), a stable analogue of TXA2, in the absence (control) and in the presence of 4-hidroxy-TEMPO (tempol, 10–3 M), a reactive oxygen species scavenger; GR-3219 (10–8 M), a TXA2 receptor (TP) antagonist; and indomethacin (10–5 M), a cyclooxygenase inhibitor. Vascular O2- production was detected by fluorescent microscopy after dihydroethidium staining in aortic frozen sections. Mean of fluorescence intensity was quantified with ImageJ software. Results: TXA2 increased vascular O2- production in aortic sections of SAMR1. This increment was higher in SAMP8. In aorta from OVX groups, TXA2 produced a further increased in O2-. In all the groups, the increase of O2- levels in response to U46619 was inhibited by tempol, GR-3219, and indomethacin, suggesting a specific role of cyclooxygenases after TP receptor activation. The effects were estrogen-dependent as O2- levels were reversed in OVE mice. Conclusions: TXA2, through TP receptor, induces an increment in vascular O2- production mediated by cyclooxygenase activation. The O2- production induced by TXA2 is enhanced by senescence and by the lack of estrogens. Estrogen administration reduces partially the O2- production in OVX mice.