γ-Oryzanol ameliorates fine dust-induced premature endothelial senescence and dysfunction via attenuating oxidative stress.

Various cardiovascular diseases are associated with endothelial senescence, and a recent study showed that fine dust (FD)-induced premature endothelial senescence and dysfunction is associated with increased oxidative stress. The aim of the present study was to investigate protective effect of rice bran extract (RBE) and its major component of γ-Oryzanol (γ-Ory) against FD-induced premature endothelial senescence. Porcine coronary artery endothelial cells (PCAECs) were treated with FD alone or with RBE or γ-Ory. Senescence-associated ß-galactosidase (SA-ß-gal) activity, expression of cell cycle regulatory proteins, and oxidative stress levels were evaluated. The results indicated that SA-ß-gal activity in the FD-treated PCAECs was attenuated by RBE and γ-Ory. Additionally, γ-Ory inhibited FD-induced cell cycle arrest, restored cell proliferation, and reduced the expression of cell cycle regulatory proteins. γ-Ory also inhibited oxidative stress and prevented senescence-associated NADPH oxidase and LAS activity in FD-exposed ECs suggesting that γ-Ory could protect against FD-induced ECs senescence and dysfunction.

SGLT2 inhibition ameliorates nano plastics-induced premature endothelial senescence and dysfunction.

Nano plastics (NPs) have been a significant threat to human health and are known to cause premature endothelial senescence. Endothelial senescence is considered one of the primary risk factors contributing to numerous cardiovascular disorders. Recent studies have suggested that inhibition of sodium glucose co-transporter (SGLT2) ameliorates endothelial senescence and dysfunction. Therefore, our study intends to explore the role of SGLT2 in NPs-induced endothelial senescence and dysfunction. Porcine coronary artery and its endothelial cells were treated with NPs in the presence or absence of Enavogliflozin (ENA), a SGLT2 inhibitor and then SGLTs expression, senescence markers and vascular function were evaluated. NPs significantly up-regulated SGLT2 and ENA significantly decreased NPs-induced senescence-associated-ß-gal activity, cell-cycle arrest, and senescence markers p53 and p21 suggesting that inhibition of SGLT2 prevents NPs-induced endothelial senescence. In addition, ENA decreased the formation of reactive oxygen species with the downregulation of Nox2, and p22phox. Furthermore, SGLT2 inhibition also up regulated the endothelial nitric oxide synthase expression along with improving vascular function. In conclusion, premature endothelial senescence by NPs is, at least in part, associated with SGLT2 and it could be a potential therapeutic target for preventing and/or treating environmental pollutants-induced cardiovascular disorders mediated by endothelial senescence and dysfunction.

Empagliflozin prevents angiotensin II-induced hypertension related micro and macrovascular endothelial cell activation and diastolic dysfunction in rats despite persistent hypertension: Role of endothelial SGLT1 and 2.

SGLT2 inhibitors (SGLT2i) showed pronounced beneficial effects in patients with heart failure but the underlying mechanisms remain unclear. We evaluated the effect of empagliflozin, selective SGLT2i, on hypertension-induced cardiac and vascular dysfunction. Male Wistar rats received diet with or without empagliflozin (30 mg/kg/day). After 1 week, a hypertensive dose of Ang II (0.4 mg/kg/day) was administered using osmotic mini-pumps for 4 weeks. Systolic blood pressure was determined by sphygmomanometry, the cardiac function by echocardiography and ex vivo (coronary microvascular endothelial cell activation, LV remodeling and fibrosis responses), and the systemic micro and macrovascular endothelial cell activation ex vivo. Empagliflozin treatment did not affect the Ang II-induced hypertensive response. Ang II treatment increased LV mass and induced LV diastolic dysfunction, fibrosis, collagen I and ANP expression, and infiltration of macrophages. In the vasculature, it caused eNOS upregulation in the aorta and down-regulation in mesenteric microvessels associated with increased oxidative stress, ACE, AT1R, VCAM-1, MCP-1, MMP-2, and MMP-9 and collagen I expression, increased endothelial SGLT1 staining in the aorta, mesenteric and coronary microvessels, increased SGLT1 and 2 protein levels in the aorta. All Ang II-induced cardiac and vascular responses were reduced by the empagliflozin treatment. Thus, the SGLT2i effectively attenuated the deleterious impact of Ang II-induced hypertension on target organs including cardiac diastolic dysfunction and remodeling, and endothelial cell activation and pro-atherosclerotic, pro-fibrotic and pro-remodeling responses in macro and microvessels despite persistent hypertension.

Carissa edulis Vahl (Apocynaceae) extract, a medicinal plant of Benin pharmacopoeia, induces potent endothelium-dependent relaxation of coronary artery rings involving nitric oxide.

BACKGROUND: Hypertension is a major cardiovascular risk factor that affects most countries including those of Africa. Although Carissa edulis Vahl, Diodia scandens Sw. and Cleome gynandra L. are traditionally used in Benin as antihypertensive treatments with some efficacy mentioned by the local population, their biological activity on the cardiovascular system remains poorly studied. AIM: The study investigated the vasoreactivity of the plants and assessed the underlying mechanisms using isolated arteries. STUDY DESIGN: Aqueous-ethanolic extracts of aerial parts of C. edulis, D. scandens and C. gynandra were prepared by maceration before being subjected to multi-step liquid-liquid fractionation with solvents of increasing polarity. The vasoreactivity of the extracts and fractions were assessed on isolated porcine coronary artery and rat aorta using organ chambers, the role of nitric oxide (NO) using NG-nitro-L-arginine (NO synthase inhibitor), prostanoids using indomethacin (cyclooxygenases inhibitor) and endothelium-dependent hyperpolarization using TRAM-34 plus UCL 1684 (inhibitors of calcium-dependent K+ channels), and the vascular uptake of polyphenols using Neu reagent. RESULTS: The aqueous-ethanolic crude extract of C. edulis (CECE) induced potent relaxations that were exclusively endothelium-dependent and more pronounced than those to D. scandens and C. gynandra. The n-butanolic fraction of C. edulis (CEBF) was more active than the cyclohexane, dichloromethane, and ethyl acetate fractions. The relaxation induced by CECE and CEBF were inhibited by NG-nitro-L-arginine and affected neither by TRAM-34 plus UCL 1684 nor by indomethacin. CEBF induced sustained endothelium-dependent relaxations for at least 60 min, and inhibited, in a concentration-dependent manner, contractions to KCl, CaCl2, U46619 and serotonin in rings with endothelium. Analysis of CEBF by LCHRMS indicated the presence of polyphenols, terpenes, and alkaloids. Exposure of coronary artery and aorta rings to CEBF caused the accumulation of polyphenols predominantly in the endothelium. CONCLUSION: C. edulis leaf extract induced pronounced endothelium-dependent relaxations and inhibited contractile responses by stimulating the endothelial formation of NO. LCHRMS analysis of the most active fraction, the butanolic fraction, revealed the presence of numerous compounds including polyphenols, terpenes, and alkaloids. The polyphenols of CEBF accumulated preferentially in the endothelium of the arterial wall. Thus, these observations support the folkloric use of C. edulis in hypertension.

Effects of polystyrene nanoplastics on endothelium senescence and its underlying mechanism.

Global plastic use has increased rapidly, and environmental pollution associated with nanoplastics (NPs) has been a growing concern recently. However, the impact and biological mechanism of NPs on the cardiovascular system are not well characterized. This study aimed to assess the possibility that NPs exposure promotes premature endothelial cell (EC) senescence in porcine coronary artery ECs and, if so, to elucidate the underlying mechanism. Treatment of ECs with NPs promoted the acquisition of senescence markers, senescence-associated ß-galactosidase activity, and p53, p21, and p16 protein expression, resulting in the inhibition of proliferation. In addition, NPs impaired endothelium-dependent vasorelaxation associated with decreased endothelial nitric oxide synthase (eNOS) expression. NPs enhanced reactive oxygen species formation in ECs, and increased oxidative stress levels were associated with the induction of NADPH oxidases expression, followed by the subsequent downregulation of Sirt1 expression. The characteristics of EC senescence and dysfunction caused by NPs are prevented by an antioxidant (N-acetylcysteine), an NADPH oxidase inhibitor (apocynin), and a Sirt1 activator (resveratrol). These findings indicate that NPs induced premature EC senescence, at least in part, through the redox-sensitive eNOS/Sirt1 signaling pathway. This study suggested the effects and underlying mechanism of NPs on the cardiovascular system, which may provide pharmacological targets to prevent NPs-associated cardiovascular diseases.

Molecular Mechanisms Responsible for In Vitro Cytotoxic Attributes of Conyza bonariensis Extract against Lymphoblastic Leukaemia Jurkat Cells.

BACKGROUND: Conyza bonariensis is known to have anti-cancer properties. OBJECTIVE: The current study investigated the in vitro pro-apoptotic properties of Conyza bonariensis (C. bonariensis) towards human lymphoblastic leukemia Jurkat cells. METHODS: Ariel parts of C. bonariensis were macerated in a non-polar (n-Hexane) solvent. MTS cell viability assay was employed to determine the cytotoxic activity of the extract towards human leukemia Jurket cells and normal Peripheral Blood Mononuclear Cells (PBMCs). The phytochemical composition of the extract was screened using HPLC method. Flow cytometric studies (FACS) were conducted to explore the pro-apoptotic potential of the extract. Western blot studies were employed to identify the molecular targets involved in the induction of apoptosis. RESULTS: The n-hexane extract showed selective cytotoxic activity towards Jurkat cells. FACS analysis indicated that the extract induced early and late apoptosis in Jurkat cells. Western blot studies revealed that the extract downregulated the expression of DNMT1, SIRT1, and UHRF1 with a simultaneous up-regulation of p73 and caspases-3 proteins expression. HPLC characterization of the extract revealed the presence of phenolic compounds. CONCLUSION: Overall, these findings demonstrate that the anti-cancer effects of a Conyza bonariensis extract towards human lymphoblastic leukemia Jurkat cells are due to the modulation of the activity of multiple oncogenic and tumor suppressor proteins. Phenolic contents of the extract are proposed to be responsible for these activities.

In Vitro Impact of Pro-Senescent Endothelial Microvesicles on Isolated Pancreatic Rat Islets Function.

BACKGROUND: Ischemia-driven islet isolation procedure is one of the limiting causes of pancreatic islet transplantation. Ischemia-reperfusion process is associated with endothelium dysfunction and the release of pro-senescent microvesicles. We investigated whether pro-senescent endothelial microvesicles prompt islet senescence and dysfunction in vitro. MATERIAL AND METHODS: Pancreatic islets were isolated from male young rats. Replicative endothelial senescence was induced by serial passaging of primary porcine coronary artery endothelial cells, and microvesicles were isolated either from young passage 1 (P1) or senescent passage 3 (P3) endothelial cells. Islet viability was assessed by fluorescence microscopy, apoptosis by flow cytometry, and Western blot. Function was assessed by insulin secretion and islet senescence markers p53, p21, and p16 by Western blot. Microvesicles were stained by the PKH26 lipid fluorescent probe and their islet integration assessed by microscopy and flow cytometry. RESULTS: Regardless of the passage, half microvesicles were integrated in target islets after 24 hours incubation. Insulin secretion significantly decreased after treatment by senescent microvesicles (P3: 1.7 ± 0.2 vs untreated islet: 2.7 ± 0.2, P < .05) without altering the islet viability (89.47% ± 1.69 vs 93.15% ± 0.97) and with no significant apoptosis. Senescent microvesicles significantly doubled the expression of p53, p21, and p16 (P < .05), whereas young microvesicles had no significant effect. CONCLUSION: Pro-senescent endothelial microvesicles specifically accelerate the senescence of islets and alter their function. These data suggest that islet isolation contributes to endothelial driven islet senescence.

Angiotensin II-induced upregulation of SGLT1 and 2 contributes to human microparticle-stimulated endothelial senescence and dysfunction: protective effect of gliflozins.

BACKGROUND: Sodium-glucose cotransporter 2 (SGLT2) inhibitors reduced cardiovascular risk in type 2 diabetes patients independently of glycemic control. Although angiotensin II (Ang II) and blood-derived microparticles are major mediators of cardiovascular disease, their impact on SGLT1 and 2 expression and function in endothelial cells (ECs) and isolated arteries remains unclear. METHODS: ECs were isolated from porcine coronary arteries, and arterial segments from rats. The protein expression level was assessed by Western blot analysis and immunofluorescence staining, mRNA levels by RT-PCR, oxidative stress using dihydroethidium, nitric oxide using DAF-FM diacetate, senescence by senescence-associated beta-galactosidase activity, and platelet aggregation by aggregometer. Microparticles were collected from blood of patients with coronary artery disease (CAD-MPs). RESULTS: Ang II up-regulated SGLT1 and 2 protein levels in ECs, and caused a sustained extracellular glucose- and Na+-dependent pro-oxidant response that was inhibited by the NADPH oxidase inhibitor VAS-2780, the AT1R antagonist losartan, sotagliflozin (Sota, SGLT1 and SGLT2 inhibitor), and empagliflozin (Empa, SGLT2 inhibitor). Ang II increased senescence-associated beta-galactosidase activity and markers, VCAM-1, MCP-1, tissue factor, ACE, and AT1R, and down-regulated eNOS and NO formation, which were inhibited by Sota and Empa. Increased SGLT1 and SGLT2 protein levels were observed in the rat aortic arch, and Ang II- and eNOS inhibitor-treated thoracic aorta segments, and were associated with enhanced levels of oxidative stress and prevented by VAS-2780, losartan, Sota and Empa. CAD-MPs promoted increased levels of SGLT1, SGLT2 and VCAM-1, and decreased eNOS and NO formation in ECs, which were inhibited by VAS-2780, losartan, Sota and Empa. CONCLUSIONS: Ang II up-regulates SGLT1 and 2 protein expression in ECs and arterial segments to promote sustained oxidative stress, senescence and dysfunction. Such a sequence contributes to CAD-MPs-induced endothelial dysfunction. Since AT1R/NADPH oxidase/SGLT1 and 2 pathways promote endothelial dysfunction, inhibition of SGLT1 and/or 2 appears as an attractive strategy to enhance the protective endothelial function.

Fluorescent nanocarriers targeting VCAM-1 for early detection of senescent endothelial cells.

Endothelial senescence has been identified as an early event in the development of endothelial dysfunction, a hallmark of cardiovascular disease. This study developed theranostic nanocarriers (NC) decorated with VCAM-1 antibodies (NC-VCAM-1) in order to target cell surface VCAM-1, which is overexpressed in senescent endothelial cells (ECs) for diagnostic and therapeutic purposes. Incubation of Ang II-induced premature senescent ECs or replicative senescent ECs with NC-VCAM-1 loaded with lipophilic fluorescent dyes showed higher fluorescence signals than healthy EC, which was dependent on the NC size and VCAM-1 antibodies concentration, and not observed following masking of VCAM-1. NC loaded with omega 3 polyunsaturated fatty acid (NC-EPA:DHA6:1) were more effective than native EPA:DHA 6:1 to prevent Ang II-induced VCAM-1 and p53 upregulation, and SA-ß-galactosidase activity in coronary artery segments. These theranostic NC might be of interest to evaluate the extent and localization of endothelial senescence and to prevent pro-senescent endothelial responses.

Significance of neutrophil microparticles in ischaemia-reperfusion: Pro-inflammatory effectors of endothelial senescence and vascular dysfunction.

Endothelial senescence is an emerging cause of vascular dysfunction. Because microparticles are effectors of endothelial inflammation and vascular injury after ischaemia-reperfusion, we examined leucocyte-derived microparticles of spleen origin as possible contributors. Microparticles were generated from primary rat splenocytes by either lipopolysaccharide or phorbol-myristate-acetate/calcium ionophore, under conditions mimicking innate and adaptive immune responses. Incubation of primary porcine coronary endothelial cells with either type of microparticles, but not with those from unstimulated splenocytes, leads to a similar threefold raise in senescence-associated ß-galactosidase activity within 48 hours, indicating accelerated senescence, to endothelial oxidative stress, and a fivefold and threefold increase in p21 and p16 senescence markers after 24 hours. After 12-hour incubation, the endothelial-dependent relaxation of coronary artery rings was reduced by 50%, at distinct optimal microparticle concentration. In vitro, microparticles were pro-thrombotic by up-regulating the local angiotensin system, by prompting tissue factor activity and a secondary generation of pro-coagulant endothelial microparticles. They initiated an early pro-inflammatory response by inducing phosphorylation of NF-κB, MAP kinases and Akt after 1 hour, and up-regulated VCAM-1 and ICAM-1 at 24 hours. Accordingly, VCAM-1 and COX-2 were also up-regulated in the coronary artery endothelium and eNOS down-regulated. Lipopolysaccharide specifically favoured the shedding of neutrophil- and monocyte-derived microparticles. A 80% immuno-depletion of neutrophil microparticles reduced endothelial senescence by 55%, indicating a key role. Altogether, data suggest that microparticles from activated splenocytes prompt early pro-inflammatory, pro-coagulant and pro-senescent responses in endothelial cells through redox-sensitive pathways. The control of neutrophil shedding could preserve the endothelium at site of ischaemia-reperfusion-driven inflammation and delay its dysfunction.

Unveiling the Role of Inflammation and Oxidative Stress on Age-Related Cardiovascular Diseases.

The global population above 60 years has been growing exponentially in the last decades, which is accompanied by an increase in the prevalence of age-related chronic diseases, highlighting cardiovascular diseases (CVDs), such as hypertension, atherosclerosis, and heart failure. Aging is the main risk factor for these diseases. Such susceptibility to disease is explained, at least in part, by the increase of oxidative stress, in which it damages cellular components such as proteins, DNA, and lipids. In addition, the chronic inflammatory process in aging "inflammaging" also contributes to cell damage, creating a stressful environment which drives to the development of CVDs. Taken together, it is possible to identify the molecular connection between oxidative stress and the inflammatory process, especially by the crosstalk between the transcription factors Nrf-2 and NF-κB which are mediated by redox signalling and are involved in aging. Therapies that control this process are key targets in the prevention/combat of age-related CVDs. In this review, we show the basics of inflammation and oxidative stress, including the crosstalk between them, and the implications on age-related CVDs.

Empagliflozin improved systolic blood pressure, endothelial dysfunction and heart remodeling in the metabolic syndrome ZSF1 rat.

BACKGROUND: Empagliflozin (empa), a selective sodium-glucose cotransporter (SGLT)2 inhibitor, reduced cardiovascular mortality and hospitalization for heart failure in patients with type 2 diabetes at high cardiovascular risk independent of glycemic control. The cardiovascular protective effect of empa was evaluated in an experimental model of metabolic syndrome, the obese ZSF1 rat, and its' lean control. METHODS: Lean and obese ZSF1 rats were either non-treated or treated with empa (30 mg/kg/day) for 6 weeks. Vascular reactivity was assessed using mesenteric artery rings, systolic blood pressure by tail-cuff sphygmomanometry, heart function and structural changes by echocardiography, and protein expression levels by Western blot analysis. RESULTS: Empa treatment reduced blood glucose levels from 275 to 196 mg/dl in obese ZSF1 rats whereas normoglycemia (134 mg/dl) was present in control lean ZSF1 rats and was unaffected by empa. Obese ZSF1 rats showed increased systolic blood pressure, and blunted endothelium-dependent relaxations associated with the appearance of endothelium-dependent contractile responses (EDCFs) compared to control lean rats. These effects were prevented by the empa treatment. Obese ZSF1 rats showed increased weight of the heart and of the left ventricle volume without the presence of diastolic or systolic dysfunction, which were improved by the empa treatment. An increased expression level of senescence markers (p53, p21, p16), tissue factor, VCAM-1, SGLT1 and SGLT2 and a down-regulation of eNOS were observed in the aortic inner curvature compared to the outer one in the control lean rats, which were prevented by the empa treatment. In the obese ZSF1 rats, no such effects were observed. The empa treatment reduced the increased body weight and weight of lungs, spleen, liver and perirenal fat, hyperglycemia and the increased levels of total cholesterol and triglycerides in obese ZSF1 rats, and increased blood ketone levels and urinary glucose excretion in control lean and obese ZSF1 rats. CONCLUSION: Empa reduced glucose levels by 28% and improved both endothelial function and cardiac remodeling in the obese ZSF1 rat. Empa also reduced the increased expression level of senescence, and atherothrombotic markers at arterial sites at risk in the control lean, but not obese, ZSF1 rat.

Oral Intake of EPA:DHA 6:1 by Middle-Aged Rats for One Week Improves Age-Related Endothelial Dysfunction in Both the Femoral Artery and Vein: Role of Cyclooxygenases.

In humans, aging is associated with endothelial dysfunction and an increased risk of venous thromboembolism. Although intake of eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) at a ratio of 6:1 by old rats improved the endothelial dysfunction in arteries, the impact on veins remains unclear. Eight-month-old male Wistar rats were either untreated or orally administered corn oil, EPA:DHA 1:1, or EPA:DHA 6:1 (500 mg/kg/d) for seven days. Vascular reactivity was studied by myography. In middle-aged femoral artery rings, acetylcholine caused a partial relaxation at low concentrations and a contractile response at high concentrations, whereas in the old femoral vein only a partial relaxation was observed. The EPA:DHA 6:1 treatment blunted the contractile response to acetylcholine in the middle-aged femoral artery and both EPA:DHA 6:1 and 1:1 increased the relaxation to acetylcholine in the old femoral vein. No such effects were observed with corn oil. Both the non-selective cyclooxygenase inhibitor indomethacin and the COX-1 inhibitor SC-560 increased the relaxation to acetylcholine in the middle-aged femoral artery whereas the COX-2 inhibitor NS-398 increased that in the middle-aged femoral vein. In conclusion, our results indicate that aging is associated with an endothelial dysfunction in the femoral artery and vein, which can be improved by EPA:DHA 6:1 treatment-most likely via a cyclooxygenase-dependent mechanism.

Angiotensin II-induced redox-sensitive SGLT1 and 2 expression promotes high glucose-induced endothelial cell senescence.

High glucose (HG)-induced endothelial senescence and dysfunction contribute to the increased cardiovascular risk in diabetes. Empagliflozin, a selective sodium glucose co-transporter2 (SGLT2) inhibitor, reduced the risk of cardiovascular mortality in type 2 diabetic patients but the protective mechanism remains unclear. This study examines the role of SGLT2 in HG-induced endothelial senescence and dysfunction. Porcine coronary artery cultured endothelial cells (ECs) or segments were exposed to HG (25 mmol/L) before determination of senescence-associated beta-galactosidase activity, protein level by Western blot and immunofluorescence staining, mRNA by RT-PCR, nitric oxide (NO) by electron paramagnetic resonance, oxidative stress using dihydroethidium and glucose uptake using 2-NBD-glucose. HG increased ECs senescence markers and oxidative stress, down-regulated eNOS expression and NO formation, and induced the expression of VCAM-1, tissue factor, and the local angiotensin system, all these effects were prevented by empagliflozin. Empagliflozin and LX-4211 (dual SGLT1/2 inhibitor) reduced glucose uptake stimulated by HG and H2 O2 in ECs. HG increased SGLT1 and 2 protein levels in cultured ECs and native endothelium. Inhibition of the angiotensin system prevented HG-induced ECs senescence and SGLT1 and 2 expression. Thus, HG-induced ECs ageing is driven by the local angiotensin system via the redox-sensitive up-regulation of SGLT1 and 2, and, in turn, enhanced glucotoxicity.

Intake of omega-3 formulation EPA:DHA 6:1 by old rats for 2 weeks improved endothelium-dependent relaxations and normalized the expression level of ACE/AT1R/NADPH oxidase and the formation of ROS in the mesenteric artery.

Omega-3 polyunsaturated fatty acids (PUFAs) including eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) have been shown to protect the cardiovascular system, in part, by stimulating the endothelial formation of nitric oxide (NO). EPA:DHA 6:1 has been identified as a potent omega 3 PUFA formulation to induce endothelium-dependent vasorelaxation and activation of endothelial NO synthase (eNOS). This study examined whether intake of EPA:DHA 6:1 (500 mg/kg/day) for 2 weeks improves an established endothelial dysfunction in old rats (20 months old), and, if so, the underlying mechanism was subsequently determined. In the main mesenteric artery rings, an endothelial dysfunction characterized by a blunted NO component, an abolished endothelium-dependent hyperpolarization component, and increased endothelium-dependent contractile responses (EDCFs) are observed in old rats compared to young rats. Age-related endothelial dysfunction was associated with increased vascular formation of reactive oxygen species (ROS) and expression of eNOS, components of the local angiotensin system, senescence markers, and cyclooxygenase-2 (COX-2), and the downregulation of COX-1. The EPA:DHA 6:1 treatment improved the NO-mediated relaxation, reduced the EDCF-dependent contractile response and the vascular formation of ROS, and normalized the expression level of all target proteins in the old arterial wall. Thus, the present findings indicate that a 2-week intake of EPA:DHA 6:1 by old rats restored endothelium-dependent NO-mediated relaxations, most likely, by preventing the upregulation of the local angiotensin system and the subsequent formation of ROS.

Thrombin Induces Angiotensin II-Mediated Senescence in Atrial Endothelial Cells: Impact on Pro-Remodeling Patterns.

BACKGROUND: Besides its well-known functions in hemostasis, thrombin plays a role in various non-hemostatic biological and pathophysiologic processes. We examined the potential of thrombin to promote premature atrial endothelial cells (ECs) senescence. METHODS AND RESULTS: Primary ECs were isolated from porcine atrial tissue. Endothelial senescence was assessed by measuring beta-galactosidase (SA-ß-gal) activity using flow cytometry, oxidative stress using the redox-sensitive probe dihydroethidium, protein level by Western blot, and matrix metalloproteinases (MMPs) activity using zymography. Atrial endothelial senescence was induced by thrombin at clinically relevant concentrations. Thrombin induced the up-regulation of p53, a key regulator in cellular senescence and of p21 and p16, two cyclin-dependent kinase inhibitors. Nicotinamide adenine dinucleotide phosphate NADPH oxidase, cyclooxygenases and the mitochondrial respiration complex contributed to oxidative stress and senescence. Enhanced expression levels of vascular cell adhesion molecule (VCAM)-1, tissue factor, transforming growth factor (TGF)-ß and MMP-2 and 9 characterized the senescence-associated secretory phenotype of atrial ECs. In addition, the pro-senescence endothelial response to thrombin was associated with an overexpression of both angiotensin converting enzyme and AT1 receptors and was inhibited by perindoprilat and losartan. CONCLUSIONS: Thrombin promotes premature ageing and senescence of atrial ECs and may pave the way to deleterious remodeling of atrial tissue by a local up-regulation of the angiotensin system and by promoting pro-inflammatory, pro-thrombotic, pro-fibrotic and pro-remodeling responses. Hence, targeting thrombin and/or angiotensin systems may efficiently prevent atrial endothelial senescence.

EPA:DHA 6:1 is a superior omega-3 PUFAs formulation attenuating platelets-induced contractile responses in porcine coronary and human internal mammary artery by targeting the serotonin pathway via an increased endothelial formation of nitric oxide.

At arterial sites of endothelial denudation and dysfunction, activated platelets contribute to vascular injury through the release of potent contracting factors such as serotonin (5-HT). This study evaluated whether omega-3 polyunsaturated fatty acids (PUFAs), known to protect the vascular system, are able to prevent platelets-induced contractile responses in isolated arteries and, if so, to investigate the underlying mechanism and the importance of the omega-3 PUFAs formulation. Porcine coronary arteries (PCA), human internal mammary arteries (IMA) and washed human platelets were prepared and vascular reactivity was studied in organ chambers. In PCA rings, aggregating platelets caused concentration-dependent contractions that were significantly inhibited by the 5-HT2A receptor antagonist ketanserin, and by EPA:DHA 6:1 but not EPA:DHA 1:1 at 0.4% v/v. EPA:DHA 6:1 also prevented the 5-HT-induced contractions but affected only slightly those to the thromboxane A2 analogue U46619. The inhibitory effect of EPA:DHA 6:1 on platelets-induced contractions was not observed in rings without endothelium, and prevented by an eNOS inhibitor but not by inhibitors of endothelium-dependent hyperpolarization. In IMA rings, EPA:DHA 6:1 but not EPA:DHA 1:1 at 0.4% v/v significantly prevented the 5-HT-induced contraction, and induced greater endothelium-dependent relaxations than bradykinin and acetylcholine sensitive to an eNOS inhibitor. EPA:DHA 6:1 strongly inhibits platelets- and 5-HT-induced contractions in PCA rings and those to 5-HT in IMA rings most likely through an increased endothelial formation of NO. These findings suggest that the omega-3 PUFAs EPA:DHA 6:1 formulation may be of interest to prevent platelets-induced vascular injury at arterial sites of endothelial dysfunction.

Urapidil, but not dihydropyridine calcium channel inhibitors, preserves the hypoxic pulmonary vasoconstriction: an experimental study in pig arteries.

Hypoxic pulmonary vasoconstriction (HPV) is a protective mechanism maintaining blood oxygenation by redirecting blood flow from poorly ventilated to well-ventilated areas in the lung. Such a beneficial effect is blunted by antihypertensive treatment with dihydropyridine calcium channel inhibitors. The aim of the present study was to evaluate the effect of urapidil, an antihypertensive agent acting as an α1 adrenergic antagonist and a partial 5-HT1A agonist, on HPV in porcine proximal and distal pulmonary artery rings, and to characterize underlying mechanisms. Rings from proximal and distal porcine pulmonary artery were suspended in organ chambers and aerated with a 95% O2 + 5% CO2 gas mixture. HPV was induced by changing the gas to a 95% N2 + 5% CO2 mixture following a low level of pre-contraction with U46619. Hypoxia induced a contractile response in both proximal and distal pulmonary artery rings. This effect is observed in the presence of a functional endothelium and is inhibited by a soluble guanylyl cyclase inhibitor (ODQ), a NO scavenger (carboxy-PTIO), and by catalase in proximal pulmonary artery rings. The endothelium-dependent HPV is prevented by nicardipine and clevidipine but remained unaffected by urapidil in both proximal and distal pulmonary artery rings. These findings indicate that urapidil, in contrast to nicardipine and clevidipine, preserves the hypoxia-triggered vasoconstriction in isolated pulmonary arteries. They further indicate the involvement of the NO-guanylyl cyclase pathway and H2 O2 in HPV. Further research is warranted to determine the potential clinical relevance of the preserved hypoxia-induced pulmonary vasoconstriction by urapidil.

Tambulin is a major active compound of a methanolic extract of fruits of Zanthoxylum armatum DC causing endothelium-independent relaxations in porcine coronary artery rings via the cyclic AMP and cyclic GMP relaxing pathways.

BACKGROUND: Zanthoxylum armatum DC (Z. armatum), belonging to Rutaceae family, has been traditionally used for the treatment of various diseases such as hypertension, abdominal pain, headache, fever, high altitude sickness, diarrhea, dysentery, and as a tonic, condiment, and an anthelmintic treatment. HYPOTHESIS: The present study aims to evaluate the vasorelaxant effect of a methanolic extract of the fruits of Z. armatum, isolate the active components and characterize the underlying mechanism. STUDY DESIGN: A methanolic extract of fruits of Z. armatum was prepared and its vasorelaxant effect was studied using porcine coronary artery rings. Thereafter, the methanolic extract was analyzed, and a major compound was isolated and its structure elucidated (tambulin). Different pharmacological tools were used to characterize the vasorelaxant effect of tambulin. RESULTS: The methanolic extract and the isolated tambulin caused similar endothelium-independent relaxations of porcine coronary artery rings with and without endothelium indicating a direct relaxing effect at the vascular smooth muscle. Tambulin did not affect the relaxation curves to the endothelium-dependent vasodilators, bradykinin and the calcium ionophore A23187 in rings with endothelium. Tambulin (1 µM) slightly but significantly shifted leftwards the concentration-relaxation curve to the endothelium-independent vasodilators, sodium nitroprusside (SNP), forskolin (FC) and isoproterenol but not those to soluble guanylyl cyclase activators (YC-1 and BAY 41-2272) and K+ channel openers (levcromakalim and 1-EBIO). Pretreatment with tambulin inhibited, in a concentration-dependent manner, contractions to KCl, serotonin (5-HT), CaCl2 and U46619 in coronary artery rings without endothelium. Both the protein kinase A (H-89, 10 µM) and the protein kinase G (Rp-8-br-cyclic GMPS, 30 µM) inhibitors significantly reduced relaxations to tambulin in coronary artery rings without endothelium. CONCLUSION: The present findings indicate that tambulin isolated from Z. armatum (fruits) is a major active principle inducing vasorelaxation through a direct effect at the vascular smooth muscle and involving both the cyclic AMP and/or cyclic GMP relaxing pathways.

[Significance of hawthorn extract in general practice - a current positioning]. / Stellenwert von Weißdornextrakt in der hausärztlichen Praxis ­ eine aktuelle Standortbestimmung.

BACKGROUND: Traditionally, hawthorn extract has been used for preventive and curative support in mild forms of age-related cardiovascular problems. There are now solid data demonstrating pre-clinical effects and mechanisms of action on a molecular-biological and cellular level that appear to be of particular interest in influencing vascular ageing and in arterial vascular disorders. METHOD: The review presents the results of a meeting of experts that took place to work out a current assessment of the therapeutic suitability of hawthorn extract in the treatment of cardiovascular disease. RESULTS AND CONCLUSIONS: Although currently no general recommendation can be given on the use of hawthorn extract in cardiac insufficiency, its use is indicated for typical challenges arising in general practice, where particularly patients with functional cardiorespiratory complaints present, possibly those with cardiac insufficiency with preserved heart function for whom there has thus far been no effective therapy apart from exercise. This recommendation is supported by the findings of studies on the safety and very good tolerability of hawthorn extract, particularly for therapy adjuvant to standard practice.