We investigated whether responsiveness to dinucleotide uridine adenosine tetraphosphate (Up4A) was altered in aortas from type 2 diabetic Otsuka Long-Evans Tokushima Fatty (OLETF) rats compared with those from age-matched control Long-Evans Tokushima Otsuka (LETO) rats at the chronic stage of disease. In OLETF aortas, we observed the following: (1) Up4A-induced contractions were lower than those in the LETO aortas under basal conditions, (2) slight relaxation occurred due to Up4A, but this was not observed in phenylephrine-precontracted LETO aortas, (3) acetylcholine-induced relaxation was reduced (vs. LETO), and (4) prostanoid release (prostaglandin (PG)F2α, thromboxane (Tx)A2 metabolite, and PGE2) due to Up4A was decreased (vs. LETO). Endothelial denudation suppressed Up4A-induced contractions in the LETO group, but increased the contractions in the OLETF group. Under nitric oxide synthase (NOS) inhibition, Up4A induced contractions in phenylephrine-precontracted aortas; this effect was greater in the LETO group (vs. the OLETF group). The relaxation response induced by Up4A was unmasked by cyclooxygenase inhibitors, especially in the LETO group, but this effect was abolished by NOS inhibition. These results suggest that the relaxant component of the Up4A-mediated response was masked by prostanoids in the LETO aortas and that the LETO and OLETF rats presented different contributions of the endothelium to the response.

Although substantial evidence suggests that an increase in the level of trimethylamine-N-oxide (TMAO) is associated with the risk of cardiovascular diseases, including atherosclerosis, chronic kidney diseases, and hypertension, the direct effect of TMAO on vascular endothelial function remains unclear. Therefore, we investigated the acute effects of TMAO on endothelium-dependent relaxation induced by acetylcholine (ACh) in the superior mesenteric arteries and femoral arteries of rat. In endothelium-intact preparations, it was observed that TMAO (300 µmol/L for 60 min) did not affect ACh-induced relaxation in either of the two arteries. In endothelium-derived hyperpolarizing factor (EDHF)-mediated relaxation under nitric oxide synthase (NOS) and cyclooxygenase (COX) inhibitions by N ω-nitro-L-arginine (L-NNA) and indomethacin, respectively, TMAO specifically impairs the relaxation in femoral arteries but not in the superior mesenteric arteries. Under the inhibitory actions of NOS and as well as blockade of intermediate-conductance calciumactivated potassium channel (IK Ca) (by TRAM-34) and small-conductance calcium-activated potassium channel (SK Ca) (by apamin), which are putative sources of EDHF, ACh-induced relaxation was low, and there were no differences between the control and TMAO-treated groups with respect to both arteries. In femoral arteries, TMAO slightly reduces ACh-induced relaxation in the presence of indomethacin (preserved NO and EDHF signals) but does not affect ACh-induced NO-mediated relaxation under the combined presence of indomethacin, TRAM-34, and apamin. These results suggest that acute treatment with TMAO specifically impairs EDHF-mediated relaxation in the femoral arteries but not in the superior mesenteric arteries. These novel observations show that TMAO is a causative factor in the development of peripheral arterial disease.

Gut microbiota have been emerging as important contributors to the regulation of host homeostasis. Accordingly, several substances converted by gut microbiota can have beneficial or adverse effects on human health. Among them, S-equol, which is produced from the isoflavone daidzein in the human and animal gut by certain microbiota, exerts estrogenic and antioxidant activities. Indoxyl sulfate, which is metabolized in the liver from indole converted from dietary tryptophan by bacterial tryptophanases in the colon, is known as a protein-bound uremic toxin. Trimethylamine N-oxide, which is generated via the oxidization of gut microbiota-derived trimethylamine by hepatic flavin monooxygenases, is known as an accelerator of atherosclerosis. The aforementioned gut-derived substances could be potential regulators of systematic tissue/organ function, including the vascular system. Macro- and microvascular complications of cardiovascular and metabolic diseases, including atherosclerosis, hypertension, and diabetes, occur systemically and represent the principal cause of morbidity and mortality. Vascular endothelial and smooth muscle dysfunction play pivotal roles in the development and progression of vasculopathies. We herein review the link between the aforementioned gut-derived substances and endothelial and vascular smooth muscle cell function. This information will provide a conceptual framework that would allow the development of novel preventive and/or therapeutic approaches against vasculopathies.

Upon stimulation, endothelial cells release various factors to regulate the vascular tone. In particular, vasorelaxing factors, called endothelium-derived relaxing factors (EDRFs), are altered in the production and/or release, as well as their signaling every vessel and under pathophysiological states, including cardiovascular, kidney, and metabolic diseases. Although indoxyl sulfate is known as a protein-bound uremic toxin and circulating levels are elevated in the impaired kidney functions, direct impact on the vascular function, especially EDRF's signaling, remains unclear. In this study, we hypothesize that acute exposure to indoxyl sulfate could alter vascular relaxation in the rat superior mesenteric artery. Accordingly, we measured acetylcholine (ACh)-induced endothelium-dependent relaxation in the absence and presence of several inhibitors to divide into each EDRF, including nitric oxide (NO), vasodilator prostaglandins (PGs), and endotheliumderived hyperpolarizing factor (EDHF). Indoxyl sulfate reduced the sensitivity to ACh but not sodium nitroprusside. Under cyclooxygenase (COX) inhibition or inhibitions of COX plus source of EDHF, such as small (SK Ca)-and intermediate (IK Ca)-conductance calcium-activated K channels, the decreased sensitivity to ACh in indoxyl sulfate exposed vessel was still preserved. However, under inhibition of NO synthase (NOS) or inhibitions of NOS and COX, the difference of sensitivity to ACh between vehicle and indoxyl sulfate was eliminated. These findings indicated that acute exposure of indoxyl sulfate in the rat superior mesenteric artery specifically explicitly impaired NO signaling but not EDHF or vasodilator PGs.