Altered arachidonic acid metabolism via COX-1 and COX-2 contributes to the endothelial dysfunction of penile arteries from obese Zucker rats

Abstract

Background and purpose: The aim of the current study was to investigate the role of arachidonic acid (AA) metabolism via cyclooxygenase (COX) in the endothelial dysfunction of penile arteries from pre-diabetic, obese Zucker rats (OZR).

Experimental approach: Penile arteries from OZR and from lean Zucker rats (LZR) were mounted in microvascular myographs to assess vascular function and COX expression was determined by immunohistochemistry.

Key results: Acetylcholine (ACh) and AA elicited relaxations that were impaired in arteries from OZR. Inhibition of both COX-1 and COX-2 reduced the relaxant effects of ACh and AA in LZR but not in OZR. Inhibitors of COX-1 and of the TXA(2)/PGH(2) (TP) receptor enhanced the relaxations induced by AA in both LZR and OZR, whereas COX-2 inhibition enhanced these responses only in OZR. TP receptor blockade did not restore ACh relaxant responses in arteries from OZR. Inhibition of COX-1 increased basal tension in OZR and this contraction was blunted by TP receptor blockade. The vasoconstrictor responses to noradrenaline were augmented by indomethacin and by COX-2 inhibition in LZR but not in OZR. Immunohistochemical staining showed that both COX-1 and COX-2 are expressed in the endothelium of penile arteries from both LZR and OZR.

Conclusions and implications: Vasoactive prostanoids were formed via constitutively active COX-1 and COX-2 pathways in normal rat penile arteries. Under conditions of insulin resistance, the release and/or effects of vasodilator prostanoids were impaired, contributing to the blunted endothelium-dependent vasodilatation and to the enhanced vasoconstriction.

Figure 2

Effects of the selective COX-1 inhibitor SC-560 (1 µM) (A, B) and the selective COX-2 inhibitor NS-398 (1 µM) (C, D) on the average relaxant responses to acetylcholine (ACh) in penile arteries from LZR (A, C) and OZR (B, D). Results are expressed as percentage of the pre-contraction induced by phenylephrine (Phe). Data are shown as the means ± SEM of 8–10 arteries. *P < 0.05, **P < 0.01 versus control before treatment. COX, cyclooxygenase; LZR, lean Zucker rats; OZR, obese Zucker rats.

Figure 1

Effect of the inhibitor of COX, indomethacin (indo) (1 µM) and the inhibitor of the NO synthesis, L-NNA (100 µM) on the relaxant responses to acetylcholine (ACh) in penile arteries from LZR (A) and OZR (B). Results are expressed as percentage of the pre-contraction induced by phenylephrine (Phe). Data are shown as the means ± SEM of 6–13 arteries. *P < 0.05, **P < 0.01, ***P < 0.001 versus control before treatment. †P < 0.01; ††P < 0.001 versus indomethacin-treated. COX, cyclooxygenase; L-NNA, N^ω-nitro-L-arginine; LZR, lean Zucker rats; NO, nitric oxide; OZR, obese Zucker rats.

Figure 3

Relaxant responses to AA in penile arteries from LZR and OZR. (A, B) Representative traces showing the AA-induced relaxations in penile arteries from (A) LZR and (B) OZR. (C) Average concentration–response curves for the relaxation to AA. Data are shown as the means ± SEM of 37 and 39 arteries (one to two per animal). **P < 0.01, ***P < 0.001 versus LZR. AA, arachidonic acid; LZR, lean Zucker rats; OZR, obese Zucker rats.

Figure 4

Effect of the COX-1 inhibitor SC-560 (1 µM) (A, B), the COX-2 inhibitor NS-398 (1 µM) (C, D) and the TXA₂/PGH₂ receptor antagonist ICI-192 (1 µM) (E, F) on the average concentration–response curves for the relaxation to AA. Results are expressed as percentage of the precontraction induced by phenylephrine (Phe). Data are shown as means ± SEM of 10 arteries (A, B) and 11 arteries (C, D) and 8 arteries (E, F). *P < 0.05, **P < 0.01, ***P < 0.001 versus controls in the absence of treatment. COX, cyclooxygenase.

Figure 5

Effect of the COX-1 inhibitor SC-560 (1 µM) (A) and of the COX-2 inhibitor NS-398 (1 µM) (B) alone or in the presence of the TXA₂/PGH₂ receptor antagonist ICI-192 (1 µM) on the basal tone of penile arteries from LZR and OZR. Results are shown as the increase in baseline tension (ΔBL), expressed as % of the contraction to 120 mM K⁺ (% of KPSS), after addition of the antagonists. Data are shown as the means ± SEM of 10 arteries (A, B). ††P < 0.01 versus OZR. ***P < 0.001 versus controls in the absence of treatment. COX, cyclooxygenase; LZR, lean Zucker rats; OZR, obese Zucker rats.

Figure 6

Effect of the TXA₂/PGH₂ receptor with ICI-192 (1 µM) on the contractile effects of the of TXA₂ analogue U-46619 [A, B; expressed as % of the contraction to 120 mM K⁺ (% of KPSS)] and on the relaxant responses to acetylcholine (ACh) (C, D) in penile arteries from LZR (A, C) and OZR (B, D). Data are shown as the means ± SEM of 10 arteries (A, B), 8 arteries (C) and 6 arteries (D). *P < 0.05, ***P < 0.001 versus controls in the absence of treatment. LZR, lean Zucker rats; OZR, obese Zucker rats.

Figure 7

Effects of the selective COX-1 inhibitor SC-560 (1 µM) (A, B) and of the selective COX-2 inhibitor NS-398 (1 µM) (C, D) on the average contractile responses to noradrenaline (NA) in penile arteries from LZR (A, C) and OZR (B, D). Results are expressed as percentage of the maximum contraction induced by NA in each artery. Data are shown as the means ± SEM of seven to eight arteries (A, B) and 9–12 arteries (C, D). *P < 0.05, **P < 0.01 versus control before treatment. COX, cyclooxygenase; LZR, lean Zucker rats; OZR, obese Zucker rats.

Figure 8

Immunohistochemical staining of COX-1 and COX-2 in the endothelium of penile arteries from LZR and OZR. (A) Haematoxylin and eosin-stained cross sections showing the vascular structure of penile arteries from LZR and OZR. Inmunohistochemical demonstration of COX-1 (B) and COX-2 (C) in LZR and OZR. (B) COX-1 isoform was widely distributed throughout the endothelial lining of penile arteries and absent in the smooth muscle layer. Additional COX-1 immunostaining was also found in the trabecular tissue, probably associated with macrophages and mast cell-like cells (asterisk). (C) COX-2 immunolabeling was sparse and restricted to small foci of endothelial cells marked with arrows. A diffuse COX-2 immunostaining was also observed in the smooth muscle layer. COX, cyclooxygenase; LZR, lean Zucker rats; OZR, obese Zucker rats.