Polyphenols prevent ageing-related impairment in skeletal muscle mitochondrial function through decreased reactive oxygen species production.

Ageing is associated with skeletal muscle impairment. Changes in mitochondrial homeostasis are thought to play a key role in this process. This study examined whether chronic intake of polyphenols (PPs), which are known to be modulators of oxidative stress, might prevent the age-related decline of mitochondrial functions in skeletal muscle. Three groups of 10 Wistar rats were investigated. Rats aged 16 weeks were compared with rats aged 40 weeks that were given 75 mg kg(-1) day(-1) PPs or solvent in the drinking water starting at week 16. Mitochondrial respiratory chain complex activities were measured in saponin-skinned fibres of soleus muscles using glutamate-malate (V(max)), succinate (V(succ)) and N,N,N',N'-tetramethyl-p-phenylenediamine dihydrochloride-ascorbate (V(TMPD)). Production of reactive oxygen species was assessed using dihydroethidium staining. Transcript levels of genes involved in antioxidant defence were determined using RT-PCR. Ageing reduced muscle V(max) (from 8.8 ± 0.45 to 6.17 ± 0.51 µmol O(2) min(-1) g(-1), -30.5%, P < 0.01), V(TMPD) (from 20.67 ± 1.24 to 16.55 ± 1.16 µmol O(2) min(-1) g(-1), -19.9%, P < 0.05), increased production of reactive oxygen species (from 100 ± 9.9 to 351.1 ± 31.7%) and decreased transcripts of mitochondrial superoxide dismutase 2 (-59.3%, P < 0.01), peroxisome proliferator-activated receptor γ coactivator-1ß (PGC-1ß; -61.5%, P < 0.05) and sirtuin 1 (-54.2%, P < 0.05). Chronic PP intake normalized V(max) (8.63 ± 0.63 µmol O(2) min(-1) g(-1)), decreased production of reactive oxygen species (141.7 ± 16.7%, P < 0.001) and enhanced antioxidant defence (superoxide dismutase 2 expression, +151.3%, P < 0.05) and PGC-1ß expression (+185.7%, P < 0.05) in comparison to age-matched untreated rats. The present data indicate that regular intake of PPs starting at a young age prevents age-related mitochondrial respiratory impairment in skeletal muscle, probably through decreased oxidative stress and enhancement of PGC-1ß expression.

Natural product extract of Dicksonia sellowiana induces endothelium-dependent relaxations by a redox-sensitive Src- and Akt-dependent activation of eNOS in porcine coronary arteries.

BACKGROUND/AIMS: The consumption of polyphenol-rich food is associated with a decreased mortality from coronary diseases. This study examined whether a standardized hydroalcoholic extract of Dicksonia sellowiana (HEDS) triggered endothelium-dependent relaxations in porcine coronary artery rings and characterized the underlying mechanism. METHODS: The phosphorylation level of Src, Akt and eNOS was assessed by Western blot analysis, the formation of reactive oxygen species by dihydroethidine staining and the level of eNOS Ser1177 phosphorylation by immunohistochemical staining in sections of coronary arteries. RESULTS: HEDS-induced endothelium-dependent relaxations were strongly reduced by Nω-nitro-L-arginine, an eNOS inhibitor, and by its combination with charybdotoxin plus apamin, inhibitors of endothelium-derived hyperpolarizing factor-mediated responses. These relaxations were markedly reduced by MnTMPyP (a membrane-permeant mimetic of superoxide dismutase), polyethylene glycol catalase (PEG-catalase; a membrane-permeant analog of catalase), and by wortmannin (an inhibitor of PI3-kinase). HEDS-induced sustained phosphorylation of Akt and eNOS in endothelial cells was abolished by MnTMPyP, PEG-catalase and wortmannin. Oral administration of HEDS induced a significant decrease of mean arterial pressure in spontaneously hypertensive rats. CONCLUSION: These findings indicate that HEDS caused endothelium-dependent relaxations of coronary artery rings through the redox-sensitive activation of the endothelial PI3-kinase/Akt pathway leading to the subsequent activation of eNOS by phosphorylation. HEDS also has antihypertensive properties.

Red wine polyphenols improve an established aging-related endothelial dysfunction in the mesenteric artery of middle-aged rats: role of oxidative stress.

Aging is associated with blunted endothelium-dependent relaxations and vascular oxidative stress. Our previous study has indicated that daily intake of red wine polyphenols (RWPs) by young rats retards aging-related endothelial dysfunction in middle-aged rats. The aim of the present study is to determine whether intake of RWPs also improves an established endothelial dysfunction in middle-aged rats and, if so, to determine the underlying mechanism. Middle-aged rats (51 weeks) received either solvent (3% ethanol), RWPs extract (100mg/kg/day) or the antioxidant and NADPH oxidase inhibitor apocynin (100mg/kg/day) in the drinking water for 4 weeks. Vascular reactivity of mesenteric artery rings from control young (12 weeks) and middle-aged rats was assessed in organ chambers. The expression level of endothelial NO synthase (eNOS), arginase I, angiotensin II receptors (AT1R and AT2R), NADPH oxidase subunits and nitrotyrosines was assessed by immunohistochemistry, and the vascular formation of reactive oxygen species (ROS) by dihydroethidine. Aging is associated with blunted endothelium-dependent relaxations, an excessive vascular formation of ROS and peroxynitrites, and an up-regulation of eNOS, arginase I, NADPH oxidase subunits (nox-1, p22phox), and AT1R and AT2R expression. RWPs and apocynin treatments improved endothelial dysfunction, normalized oxidative stress and the expression of the different proteins in the mesenteric artery of middle-aged rats. The present findings indicate that aging is associated with blunted endothelium-dependent relaxations involving an increased oxidative stress, and that these responses are improved by the intake of RWPs or apocynin for 4weeks most likely by normalizing the expression of eNOS, arginase I, NADPH oxidase and angiotensin receptors.

Chronic intake of red wine polyphenols by young rats prevents aging-induced endothelial dysfunction and decline in physical performance: role of NADPH oxidase.

Aging is associated with oxidative stress-mediated endothelial dysfunction and decline in physical performance, which promote cardiovascular diseases. This study examined whether chronic intake of red wine polyphenols (RWPs), a rich source of natural antioxidants, prevents aging-related impairment of vascular function and physical exercise capacity. Vascular reactivity from 12, 20 and 40 week-old rats was assessed in organ chambers. Rats received from week 16 to 40 either solvent, RWPs or the antioxidant and NADPH oxidase inhibitor, apocynin. Aging was associated with blunted endothelium-dependent relaxations, oxidative stress (dihydroethidine staining), and an upregulation of eNOS, arginase I, NADPH oxidase p22phox and nox1 subunits, and AT1 and AT2 receptors (assessed by immunohistochemistry) in the mesenteric artery. RWPs and apocynin improved the endothelial dysfunction, normalized oxidative stress and the expression of the different proteins. RWPs also improved aging-related decline in physical exercise. Thus, intake of RWPs protects against aging-induced endothelial dysfunction and decline in physical performance. These effects likely involve the ability of RWPs to normalize oxidative stress and the expression of proteins involved in the formation of NO and the angiotensin II pathway.

Endothelium-derived contracting factors mediate the Ang II-induced endothelial dysfunction in the rat aorta: preventive effect of red wine polyphenols.

Angiotensin II (Ang II)-induced hypertension is associated with vascular oxidative stress and an endothelial dysfunction. This study examined the role of reactive oxygen species (ROS) and endothelium-derived contracting factors in Ang II-induced endothelial dysfunction and whether these effects are prevented by red wine polyphenols (RWPs), a rich source of natural antioxidants. Rats were infused with Ang II for 14 days. RWPs were administered in the drinking water 1 week before and during the Ang II infusion. Arterial pressure was measured in conscious rats. Vascular reactivity was assessed in organ chambers and cyclooxygenase-1 (COX-1) and COX-2 expression by Western blot and immunofluorescence analyses. Ang II-induced hypertension was associated with blunted endothelium-dependent relaxations and induction of endothelium-dependent contractions in the presence of nitro-L-arginine in response to acetylcholine (Ach). These effects were not affected by the combination of membrane permeant analogs of superoxide dismutase and catalase but were abolished by the thromboxane A(2) (TP) receptor antagonist GR32191B and the COX-2 inhibitor NS-398. The COX-1 inhibitor SC-560 also prevented contractile responses to Ach. Ang II increased the expression of COX-1 and COX-2 in the aortic wall. RWPs prevented Ang II-induced hypertension, endothelial dysfunction, and upregulation of COX-1 and COX-2. Thus, Ang II-induced endothelial dysfunction cannot be explained by an acute formation of ROS reducing the bioavailability of nitric oxide but rather by COX-dependent formation of contracting factors acting on TP receptors. RWPs are able to prevent the Ang II-induced endothelial dysfunction mostly due to their antioxidant properties.

Losartan prevents portal hypertension-induced, redox-mediated endothelial dysfunction in the mesenteric artery in rats.

BACKGROUND & AIMS: Advanced stages of portal hypertension are characterized by generalized vasodilatation and a hyperdynamic syndrome that leads to complications such as hepatopulmonary syndrome. We assessed the endothelial function--particularly the formation of nitric oxide (NO) and endothelium-derived hyperpolarizing factor (EDHF)--in rats following common bile duct ligation (CBDL) to determine the underlying mechanisms of these processes. METHODS: Reactivity of mesenteric artery rings from male Wistar rats was determined in organ chambers. The expression levels of connexins (Cx) (Cx37, Cx40, Cx43), intermediate and small conductance Ca(2+)-activated K(+) channels (IK(Ca), SK(Ca)), endothelial NO synthase (eNOS), NADPH oxidase subunits, and nitrotyrosines were assessed by immunohistochemistry in mesenteric and pulmonary arteries. The vascular formation of reactive oxygen species (ROS) was evaluated using dihydroethidine. Control rats or those that had undergone CBDL were given either the NADPH oxidase inhibitor apocynin or the angiotensin II receptor type 1 antagonist losartan. RESULTS: Decreased EDHF-mediated relaxations to acetylcholine and red wine polyphenols were observed in CBDL rats, compared with controls, whereas the level of NO-mediated relaxation was similar. Impaired EDHF-mediated relaxations were associated with reduced vascular expression of Cx37, Cx40, Cx43, IK(Ca) and SK(Ca); increased expression of eNOS and NADPH oxidase subunits; and increased vascular formation of ROS and peroxynitrites. These effects were prevented by exposure to apocynin or losartan. CONCLUSIONS: CBDL is associated with reduced EDHF-mediated relaxations in the mesenteric artery, whereas NO-mediated relaxations persisted. These findings indicate that impaired EDHF-mediated relaxation involves an excessive vascular oxidative stress, most likely following activation of angiotensin II type 1 receptors.

Crataegus special extract WS 1442 causes endothelium-dependent relaxation via a redox-sensitive Src- and Akt-dependent activation of endothelial NO synthase but not via activation of estrogen receptors.

PURPOSE: This study determined whether the Crataegus (Hawthorn species) special extract WS 1442 stimulates the endothelial formation of nitric oxide (NO), a vasoprotective factor, and characterized the underlying mechanism. METHODS AND RESULTS: Vascular reactivity was assessed in porcine coronary artery rings, reactive oxygen species (ROS) formation in artery sections by microscopy, and phosphorylation of Akt and endothelial NO synthase (eNOS) in endothelial cells by Western blot analysis. WS 1442 caused endothelium-dependent relaxations in coronary artery rings, which were reduced by N-nitro-L-arginine (a competitive inhibitor of NO synthase) and by charybdotoxin plus apamin (two inhibitors of endothelium-derived hyperpolarizing factor-mediated responses). Relaxations to WS 1442 were inhibited by intracellular ROS scavengers and inhibitors of Src and PI3-kinase, but not by an estrogen receptor antagonist. WS 1442 stimulated the endothelial formation of ROS in artery sections, and a redox-sensitive phosphorylation of Akt and eNOS in endothelial cells. CONCLUSIONS: WS 1442 induced endothelium-dependent NO-mediated relaxations of coronary artery rings through the redox-sensitive Src/PI3-kinase/Akt-dependent phosphorylation of eNOS.

Angiotensin II-induced hypertension is associated with a selective inhibition of endothelium-derived hyperpolarizing factor-mediated responses in the rat mesenteric artery.

Hypertension has been shown to be associated with impaired endothelium-derived hyperpolarizing factor (EDHF)-mediated arterial relaxation and hyperpolarization. Treatments of hypertensive rats with inhibitors of the renin-angiotensin system have been shown to restore both EDHF-mediated responses and the expression of connexins involved in the intercellular transfer of the hyperpolarization in mesenteric arteries. The present study was designed to determine whether chronic treatment of rats with angiotensin II impairs EDHF-mediated responses and the expression of connexins in the mesenteric arterial wall. Male Wistar rats were treated with angiotensin II (0.4 mg/kg/day) for 21 days using osmotic minipumps. Arterial pressure was measured by tail-cuff plethysmography. Contractile responses and membrane potential were measured in isolated mesenteric arteries. The expression of the three connexins (Cxs), Cx37, Cx40, and Cx43, was quantified in segments of mesenteric arteries by immunohistochemistry and quantitative real-time reverse transcriptase-polymerase chain reaction. Angiotensin II administration increased the mean systolic blood pressure. EDHF-mediated relaxation and hyperpolarization to acetylcholine and red wine polyphenols were significantly impaired in mesenteric arteries from angiotensin II-treated rats in comparison with control animals, whereas nitric oxide-mediated relaxation was unaltered. The expression of connexins Cx37, Cx40, and Cx43 was significantly decreased in the mesenteric artery from angiotensin II-treated rats. These findings indicate that angiotensin II-induced hypertension is associated with a selective impairment of EDHF-mediated relaxation and hyperpolarization in the rat mesenteric artery. The inhibition of EDHF-mediated responses is due, at least in part, to a decreased expression of connexins Cx37, Cx40, and Cx43 in the arterial wall.