Effect of 17beta-oestradiol on cytokine-induced nitric oxide production in rat isolated aorta.

1. Studies were performed on isolated aortic rings without endothelium to investigate the effect of 17beta-oestradiol on cytokine-induced nitric oxide production by the inducible nitric oxide synthase (iNOS). 2. Treatment of the isolated aortic rings with interleukin-1beta (IL-1beta, 20 micro ml(-1)) led to the expression of iNOS mRNA and protein, as well as significant nitrite accumulation in the incubation media and suppression of phenylephrine (1 nM-10 microM)-evoked contraction. 3. Cycloheximide (1 microM), a protein synthesis inhibitor, prevented iNOS protein expression, nitrite accumulation and the suppression of contractility by IL-1beta on the isolated aortic rings. 17Beta-oestradiol (1 nM-10 microM) and the partial oestrogen receptor agonist 4-OH-tamoxifen (1 nM-10 microM) produced concentration-dependent inhibition of IL-1beta-induced nitrite accumulation and restored vasoconstrictor responsiveness to phenylephrine, similar to the iNOS inhibitor aminoguanidine (100 microM). 4. Semiquantitative PCR demonstrated decreased iNOS mRNA in the IL-1beta-induced and 17beta-oestradiol-treated rings. Western blot analysis of rat aorta homogenates revealed that 17beta-oestradiol treatment resulted in a reduction in IL-1beta-induced iNOS protein level. 5. Incubation with tumour necrosis factor alpha (TNF alpha, 1 ng ml(-1)) resulted in significant nitrite accumulation in the incubation media and suppression of the smooth muscle contractile response to phenylephrine, similar to IL-1beta. The effects of TNF alpha were also inhibited by co-incubation of the rings with 17beta-oestradiol and 4-OH-tamoxifen (1 microM). 6. The anti-transforming growth factor-beta1 (TGF-beta1) antibody, which inhibited TGF-beta1-induced suppression of nitrite production from IL-1beta-treated vascular rings, did not affect the inhibitory action of 17beta-oestradiol, suggesting that the effect of oestrogen on iNOS inhibition was not mediated by TGF-beta1. 7. These results show that the ovarian sex steroid, 17beta-oestradiol is a modulator of cytokine-induced iNOS activity in rat vascular smooth muscle and its mechanism of action involves decrease of iNOS mRNA and protein.