Modulation of mitochondrial capacity and angiogenesis by red wine polyphenols via estrogen receptor, NADPH oxidase and nitric oxide synthase pathways.

Red wine polyphenolic compounds (RWPC) are reported to exert vasculoprotective properties on endothelial cells, involving nitric oxide (NO) release via a redox-sensitive pathway. This NO release involves the activation of the estrogen receptor-alpha (ERα). Paradoxical effects of a RWPC treatment occur in a rat model of post-ischemic neovascularization, where a low-dose is pro-angiogenic while a higher dose is anti-angiogenic. NO and ERα are key regulators of mitochondrial capacity, and angiogenesis is a highly energetic process associated with mitochondrial biogenesis. However, whether RWPC induces changes in mitochondrial capacity has never been addressed. We investigated the effects of RWPC at low (10(-4)g/l, LCP) and high concentration (10(-2)g/l, HCP) in human endothelial cells. Mitochondrial respiration, expression of mitochondrial biogenesis factors and mitochondrial DNA content were assessed using oxygraphy and quantitative PCR respectively. In vitro capillary formation using ECM gel(®) was also performed. Treatment with LCP increased mitochondrial respiration, with a maximal effect achieved at 48h. LCP also increased expression of several mitochondrial biogenesis factors and mitochondrial DNA content. In contrast, HCP did not affect these parameters. Furthermore, LCP modulated both mitochondrial capacity and angiogenesis through mechanisms sensitive to ER, NADPH oxidase and NO-synthase inhibitors. Finally, the inhibition of mitochondrial protein synthesis abolished the pro-angiogenic capacity of LCP. These results suggest a possible association between the modulation of mitochondrial capacity by LCP and its pro-angiogenic activity. These data provide evidence for a role of mitochondria in the regulation of angiogenesis by RWPC

Role of the mitochondria on the paradoxical effect of red wine polyphenols on angiogenesis

Red wine polyphenol (RWPC) extracts has been reported to possess vasoprotective properties that involve nitric oxide (NO) release from endothelial cells via a redox- sensitive pathway. Besides, the molecular target of RWPC to release NO has been recently revealed and it involves the activation of the estrogen receptor alpha (ERα). Paradoxical effects of RWPC have been shown with regard to angiogenesis. Indeed in a rat model of postischemic neovascularization, low- dose is pro-  whereas high dose is anti- angiogenic. NO and ERα  are key regulators of mitochondrial function. Furthermore, angiogenesis is a highly energetic process associated with mitochondrial biogenesis. However, whether RWPC induces changes in mitochondrial function has never been addressed and it is the aim of this study. The effects of RWPC at low concentration (10- 4 g/l, LCP) and high concentration (10- 2 g/l, HCP) after 48 hours time exposure were investigated on human endothelial cells. Mitochondrial respiration, expression of biogenesis factors and DNA content was assessed using oxygraphy and qRT- PCR, respectively. In vitro capillary formation using Matrigel® was performed. The mechanism involved with respect to ER using the ER- antagonist fulvestrant was studied. The involvement of both NADPH oxidase and NO synthase was addressed using apocynin and L- NA respectively. LCP, but not HCP, increased mitochondrial respiration. The effect of LCP was associated with an increase of both expression of several mitochondrial biogenesis factors (NRF- 1, NRF- 2, ERRα, Tfam, PolG) and mitochondrial DNA content whereas HCP had no effect on these parameters. All the effects of LCP on mitochondrial respiration are prevented by fulvestrant, apocynin and L- NA. LCP also promoted in vitro capillary elongation that was prevented by fulvestrant, apocynin and L- NA. Finally, the inhibition of mitochondrial protein synthesis using chloramphenicol suppressed the pro- angiogenic property of LCP. The present study highlights the implication of the axis ER, NADPH oxidase and NOS pathways on both increase mitochondrial function and capillary elongation in response to RWPC at low concentration. They explain the paradoxical effect of RWPC depending on the concentration with respect to angiogenesis, mitochondria being key targets for its pro- angiogenic properties

Estrogen Receptor α Participates to the Beneficial Effect of Red Wine Polyphenols in a Mouse Model of Obesity-Related Disorders

Red wine polyphenol extracts (polyphenols) ameliorate cardiovascular and metabolic disorders associated with obesity. Previously, we demonstrated that the alpha isoform of estrogen receptor (ERα) triggers the vascular protection of polyphenols. Here, we investigated the contribution of ERα on the effects of polyphenols on cardiovascular and metabolic alterations associated with obesity. We used ovariectomized wild type or ERα-deficient mice receiving standard (SD) or western (WD) diets, or SD and WD containing polyphenols (SD+polyphenols and WD+polyphenols, respectively) over a 12-week period. Body weight was measured during treatment. Echocardiography examination was performed before sacrifice. Blood and tissues were sampled for biochemical and functional analysis with respect to nitric oxide (NO(•)) and oxidative stress. Vascular reactivity and liver mitochondrial complexes were analyzed. In WD-fed mice, polyphenols reduced adiposity, plasma triglycerides and oxidative stress in aorta, heart, adipose and liver tissues and enhanced NO(•) production in aorta and liver. ERα deletion prevented or reduced the beneficial effects of polyphenols, especially visceral adiposity, aortic and liver oxidative stresses and NO(•) bioavailability. ERα deletion, however, had no effect on polyphenol\u27s ability to decrease the fat accumulation and oxidative stress of subcutaneous adipose tissue. Also, ERα deletion did not modify the decrease of ROS levels induced by polyphenols treatment in the visceral adipose tissue and heart from WD-fed mice. Dietary supplementation of polyphenols remarkably attenuates features of metabolic syndrome; these effects are partially mediated by ERα-dependent mechanisms. This study demonstrates the therapeutic potential of this extract in metabolic and cardiovascular alterations linked to excessive energy intake

Mitochondria As Potential Targets of Flavonoids: Focus on Adipocytes and Endothelial Cells

Obesity is a major public health problem, resulting from an excess of energy storage and/or a default of energy expenditure leading to the increased occurrence of cardiovascular risk factors that favour the development of vascular complications. As a consequence, many studies are interested to find novel therapeutic chemical including flavonoids that appear to be promising natural compounds to treat obesity and its complications. Several in vitro studies addressed the mechanisms involved that might explain their beneficial effects, on adipocytes and endothelial cells, two cell types that play major role in obesity and its vascular complications. Besides the well-described antioxidant properties of flavonoids, at least a part of their beneficial effects on these cell types might be explained by their action on the regulation of mitochondrial function. In this review, we will therefore focus on the pathophysiological role of mitochondria in regulating endothelial and adipocyte functions. In addition, we will present some of the more promising flavonoids, important in human diet, including flavanols, flavonols, isoflavones, anthocyanins, flavanones and flavones; and their potential effects to improve endothelial or adipocyte functions via the mitochondria

Effects of delphinidin on mitochondrial function in endothelial cells

Delphinidine (delph), an anthocyanin with the same pharmacological pro¿ le than the total extract of red wine polyphenol, induces vasodilatation and possess anti-apoptotic property in endothelial cells (ECs) by mechanism involving nitric oxide (NO). The later can regulate mitochondrial (mt) function. However, the link between NO, mitochondria and the upstream target of delph including the alpha isoform of the estrogen receptor (ERa) has never been assessed and is the aim of the present study. For this purpose, the effects of delph (3.10-5 M) and the ERa agonist propylpyrazole triol (PPT, 10-5 M) were conducted at two time points, 10min and 48h, on mt respiration (R) by oxygraphy and on respi- ratory chain complexes activities by spectrophotometry in Eahy926 ECs. NO production was assessed by electron paramagnetic resonance at the early time. These experiments were performed with or without the ER antagonist, fulves- trant (fulv, 3.10-5 M), or the NO synthase inhibitor, L-nitro-arginine (L-NA, 10-4 M). As expected, delph and PPT induced an early increase of NO that was prevented by fulv. Delph time-dependently increased basal R and maximal R capacity but not the non-phosphorylative R by a mechanism insensitive to fulv but sensitive to L-NA. PPT did not affect basal and non-phosphorylative R but increased the maximal R capacity by a mechanism sensitive to fulv and L-NA. These effects were associated with increased cytochrome c oxidase acti- vity. These data highlight the implication of both NO and cytochrome c oxidase activity on the modulation of mt R in response to delph and PPT in ECs. The lack of effect of ER blockade on the increase of mt R by delph supports the involvement of ER-independent mechanism although this receptor is implicated at the early increase of NO production. Thus, this study suggests a probable role of mitochondria in the effect of polyphenol in the regulation of endothelial func- tion including vasodilatation and endothelial integrity

Delphinidin inhibits VEGF induced-mitochondrial biogenesis and Akt activation in endothelial cells.

Delphinidin, an anthocyanin present in red wine, has been reported to exert vasculoprotective properties on endothelial cells, including vasorelaxing and anti-apoptotic effects. Moreover, delphinidin treatment in a rat model of post-ischemic neovascularization has been described to exert anti-angiogenic property. Angiogenesis is an energetic process and VEGF-induced angiogenesis is associated with mitochondrial biogenesis. However, whether delphinidin induces changes in mitochondrial biogenesis has never been addressed. Effects of delphinidin were investigated in human endothelial cells at a concentration described to be anti-angiogenic in vitro (10−2 g/l). mRNA expression of mitochondrial biogenesis factors, mitochondrial respiration, DNA content and enzyme activities were assessed after 48 h of stimulation. Delphinidin increased mRNA expression of several mitochondrial biogenesis factors, including NRF1, ERRα, Tfam, Tfb2m and PolG but did not affect neither mitochondrial respiration, DNA content nor enzyme activities. In presence of delphinidin, VEGF failed to increase mitochondrial respiration, DNA content, complex IV activity and Akt activation in endothelial cells. These results suggest a possible association between inhibition of VEGF-induced mitochondrial biogenesis through Akt pathway by delphinidin and its anti-angiogenic effect, providing a novel mechanism sustaining the beneficial effect of delphinidin against pathologies associated with excessive angiogenesis such as cancers

Propionyl-L-carnitine corrects metabolic and cardiovascular alterations in diet-induced obese mice and improves liver respiratory chain activity

AIMS: Obesity is a primary contributor to acquired insulin resistance leading to the development of type 2 diabetes and cardiovascular alterations. The carnitine derivate, propionyl-L-carnitine (PLC), plays a key role in energy control. Our aim was to evaluate metabolic and cardiovascular effects of PLC in diet-induced obese mice. METHODS: C57BL/6 mice were fed a high-fat diet for 9 weeks and then divided into two groups, receiving either free- (vehicle-HF) or PLC-supplemented water (200 mg/kg/day) during 4 additional weeks. Standard diet-fed animals were used as lean controls (vehicle-ST). Body weight and food intake were monitored. Glucose and insulin tolerance tests were assessed, as well as the HOMA(IR), the serum lipid profile, the hepatic and muscular mitochondrial activity and the tissue nitric oxide (NO) liberation. Systolic blood pressure, cardiac and endothelial functions were also evaluated. RESULTS: Vehicle-HF displayed a greater increase of body weight compared to vehicle-ST that was completely reversed by PLC treatment without affecting food intake. PLC improved the insulin-resistant state and reversed the increased total cholesterol but not the increase in free fatty acid, triglyceride and HDL/LDL ratio induced by high-fat diet. Vehicle-HF exhibited a reduced cardiac output/body weight ratio, endothelial dysfunction and tissue decrease of NO production, all of them being improved by PLC treatment. Finally, the decrease of hepatic mitochondrial activity by high-fat diet was reversed by PLC. CONCLUSIONS: Oral administration of PLC improves the insulin-resistant state developed by obese animals and decreases the cardiovascular risk associated to this metabolic alteration probably via correction of mitochondrial function

Pharmacological targeting of the protein synthesis mTOR/4E-BP1 pathway in cancer-associated fibroblasts abrogates pancreatic tumourchemoresistance

International audiencePancreatic ductal adenocarcinoma (PDAC) is extremely stroma-rich. Cancer-associated fibroblasts (CAFs) secrete proteins that activate survival and promote chemoresistance of cancer cells. Our results demonstrate that CAF secretome-triggered chemoresistance is abolished upon inhibition of the protein synthesis mTOR/4E-BP1 regulatory pathway which we found highly activated in primary cultures of -SMA-positive CAFs, isolated from human PDAC resections. CAFs selectively express the sst1 somatostatin receptor. The SOM230 analogue (Pasireotide) activates the sst1 receptor and inhibits the mTOR/4E-BP1 pathway and the resultant synthesis of secreted proteins including IL-6. Consequently, tumour growth and chemoresistance in nude mice xenografted with pancreatic cancer cells and CAFs, or with pieces of resected human PDACs, are reduced when chemotherapy (gemcitabine) is combined with SOM230 treatment. While gemcitabine alone has marginal effects, SOM230 is permissive to gemcitabine-induced cancer cell apoptosis and acts as an antifibrotic agent. We propose that selective inhibition of CAF protein synthesis with sst1-directed pharmacological compounds represents an anti-stromal-targeted therapy with promising chemosensitization potential

Systems biology of antioxidants

Understanding the role of oxidative injury will allow for therapy with agents that scavenge ROS (reactive oxygen species) and antioxidants in the management of several diseases related to free radical damage. The majority of free radicals are generated by mitochondria as a consequence of the mitochondrial cycle, whereas free radical accumulation is limited by the action of a variety of antioxidant processes that reside in every cell. In the present review, we provide an overview of the mitochondrial generation of ROS and discuss the role of ROS in the regulation of endothelial and adipocyte function. Moreover, we also discuss recent findings on the role of ROS in sepsis, cerebral ataxia and stroke. These results provide avenues for the therapeutic potential of antioxidants in a variety of diseases