Fenugreek extract improves diabetes-induced endothelial dysfunction via the arginase 1 pathway.

Endothelial dysfunction (ED) is an initiating trigger and key factor in vascular complications, leading to disability and mortality in individuals with diabetes. The research concerning therapeutic interventions for ED has gained considerable interest. Fenugreek, a commonly used edible plant in dietary consumption, has attracted significant attention due to its management of diabetes and its associated complications. The research presented in this study examines the potential therapeutic benefits of fenugreek in treating ED and investigates the underlying mechanism associated with its effects. The analysis on fenugreek was performed using 70% ethanol extract, and its chemical composition was analyzed using ultrahigh-performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF/MS). In total, we identified 49 compounds present in the fenugreek extract. These compounds encompass flavonoids, saponins, and phospholipids. Then, the models of ED in streptozotocin-induced diabetic mice and high glucose-induced isolated rat aortas were established for research. Through vascular function testing, it was observed that fenugreek extract effectively improved ED induced by diabetes or high glucose. By analyzing the protein expression of arginase 1 (Arg1), Arg activity, Arg1 immunohistochemistry, nitric oxide (NO) level, and the protein expression of endothelial nitric oxide synthase (eNOS), p38 mitogen-activated protein kinase (p38 MAPK), and p-p38 MAPK in aortas, this study revealed that the potential mechanism of fenugreek extract in anti-ED involves the downregulation of Arg1, leading to enhanced NO production. Furthermore, analysis of serum exosomes carrying Arg activity indicates that fenugreek may decrease the activity of Arg transported by serum exosomes, potentially preventing the increase in Arg levels triggered by the uptake of serum exosomes by vascular endothelial cells. In general, this investigation offers valuable observations regarding the curative impact of fenugreek extract on anti-ED in diabetes, revealing the involvement of the Arg1 pathway in its mechanism.

Protective effect of alizarin on vascular endothelial dysfunction via inhibiting the type 2 diabetes-induced synthesis of THBS1 and activating the AMPK signaling pathway.

BACKGROUND: In this study, we investigated the protective effects of alizarin (AZ) on endothelial dysfunction (ED). AZ has inhibition of the type 2 diabetes mellitus (T2DM)-induced synthesis of thrombospondin 1 (THBS1). Adenosine 5'-monophosphate- activated protein kinase (AMPK), particularly AMPKα2 isoform, plays a critical role in maintaining cardiac homeostasis. PURPOSE: The aim of this study was to investigate the ameliorative effect of AZ on vascular injury caused by T2DM and to reveal the potential mechanism of AZ in high glucose (HG)-stimulated human umbilical vein endothelial cells (HUVECs) and diabetic model rats. STUDY DESIGN: HUVECs, rats and AMPK-/- transgenic mice were used to investigate the mitigating effects of AZ on vascular endothelial dysfunction caused by T2DM and its in vitro and in vivo molecular mechanisms. METHODS: In type 2 diabetes mellitus rats and HUVECs, the inhibitory effect of alizarin on THBS1 synthesis was verified by immunohistochemistry (IHC), immunofluorescence (IF) and Western blot (WB) so that increase endothelial nitric oxide synthase (eNOS) content in vitro and in vivo. In addition, we verified protein interactions with immunoprecipitation (IP). To probe the mechanism, we also performed AMPKα2 transfection. AMPK's pivotal role in AZ-mediated prevention against T2DM-induced vascular endothelial dysfunction was tested using AMPKα2-/- mice. RESULTS: We first demonstrated that THBS1 and AMPK are targets of AZ. In T2DM, THBS1 was robustly induced by high glucose and inhibited by AZ. Furthermore, AZ activates the AMPK signaling pathway, and recoupled eNOS in stressed endothelial cells which plays a protective role in vascular endothelial dysfunction. CONCLUSIONS: The main finding of this study is that AZ can play a role in different pathways of vascular injury due to T2DM. Mechanistically, alizarin inhibits the increase in THBS1 protein synthesis after high glucose induction and activates AMPKα2, which increases NO release from eNOS, which is essential in the prevention of vascular endothelial dysfunction caused by T2DM.

Flavonoid Compounds in Hippophae rhamnoides L. Protect Endothelial Cells from Oxidative Damage Through the PI3K/AKT-eNOS Pathway.

Sea buckthorn, a traditional medicinal plant, has been used for several years in China for the prevention and treatment of various diseases, a practice closely associated with its significant antioxidant activity. The aim of this study was to investigate the protective effects of sea buckthorn flavonoids on vascular endothelial cells in an oxidative stress environment. We isolated and extracted active compounds from sea buckthorn and investigated their impact on endothelial nitric oxide synthase (eNOS) activity through the PI3K/AKT-eNOS signaling pathway through a combination of network pharmacology and cellular experiments, elucidating the regulatory effects of these compounds on endothelial cell functions. Three flavonoids, named Fr.4-2-1, Fr.4-2-2 and Fr.4-2-3, were obtained from sea buckthorn. The results of network pharmacology indicated that they might exert their effects by regulating the PI3K-AKT signaling pathway. In vitro results showed that all three flavonoids were effective in alleviating the degree of oxidative stress in cells, among which Fr.4-2-1 exerted its antioxidant effects by modulating the PI3K/AKT-eNOS pathway. Flavonoids in sea buckthorn can effectively inhibit oxidative stress-induced cellular damage, preserving the integrity and functionality of endothelial cells, which is crucial for maintaining vascular health and function.

Absorbed Bioactive Compounds Replicate Guanxin II-Induced Endothelium-Associated in/ex vivo Vasodilation.

OBJECTIVE: To develop an interference-free and rapid method to elucidate Guanxin II (GX II)'s representative vasodilator absorbed bioactive compounds (ABCs) among enormous phytochemicals. METHODS: The contents of ferulic acid, tanshinol, and hydroxysafflor yellow A (FTA) in GX II/rat serum after the oral administration of GX II (30 g/kg) were detected using ultra-performance liquid chromatography-mass spectrometry. Totally 18 rats were randomly assigned to the control group (0.9% normal saline), GX II (30 g/kg) and FTA (5, 28 and 77 mg/kg) by random number table method. Diastolic coronary flow velocity-time integral (VTI), i.e., coronary flow or coronary flow-mediated dilation (CFMD), and endothelium-intact vascular tension of isolated aortic rings were measured. After 12 h of exposure to blank medium or 0.5 mmol/L H2O2, endothelial cells (ECs) were treated with post-dose GX II of supernatant from deproteinized serum (PGSDS, 300 µL PGSDS per 1 mL of culture medium) or FTA (237, 1539, and 1510 mg/mL) for 10 min as control, H2O2, PGSDS and FTA groups. Nitric oxide (NO), vascular endothelial growth factor (VEGF), endothelin-1 (ET-1), superoxide dismutase (SOD), malondialdehyde (MDA) and phosphorylated phosphoinositide 3 kinase (p-PI3K), phosphorylated protein kinase B (p-AKT), phosphorylated endothelial nitric oxide synthase (p-eNOS) were analyzed. PGSDS was developed as a GX II proxy of ex vivo herbal crude extracts. RESULTS: PGSDS effectively eliminates false responses caused by crude GX II preparations. When doses equaled the contents in GX II/its post-dose serum, FTA accounted for 98.17% of GX II -added CFMD and 92.99% of PGSDS-reduced vascular tension. In ECs, FTA/PGSDS was found to have significant antioxidant (lower MDA and higher SOD, P<0.01) and endothelial function-protective (lower VEGF, ET-1, P<0.01) effects. The increases in aortic relaxation, endothelial NO levels and phosphorylated PI3K/Akt/eNOS protein induced by FTA/PGSDS were markedly abolished by NG-nitro-L-arginine methyl ester (L-NA, eNOS inhibitor) and wortmannin (PI3K/AKT inhibitor), respectively, indicating an endothelium-dependent vasodilation via the PI3K/AKT-eNOS pathway (P<0.01). CONCLUSION: This study provides a strategy for rapidly and precisely elucidating GX II's representative in/ex vivo cardioprotective absorbed bioactive compounds (ABCs)-FTA, suggesting its potential in advancing precision ethnomedicine.

Black Soybean Seed Coat Extract Improves Endothelial Function and Upregulates Oxidative Stress Marker Expression in Healthy Volunteers by Stimulating Nitric Oxide Production in Endothelial Cells.

Black soybean seed coat extract (BE) contains multiple bioactive polyphenols, including flavan-3-ols and anthocyanins. BE improves endothelial function; however, it is unclear whether BE protects endothelial cells from senescence. In this study, we examined the effects of BE on endothelial cell senescence and vascular function in healthy individuals. High concentrations of glucose were used to induce senescence in bovine aortic endothelial cells incubated with BE. Senescence, vascular function, and oxidative stress markers were measured. Incubation with BE remarkably inhibited senescence-associated ß-galactosidase and lactate dehydrogenase activities and dose dependently reduced intracellular reactive oxygen species levels in bovine aortic endothelial cells. BE treatment increased the levels of endothelial nitric oxide synthase (eNOS) mRNA and endothelial nitric oxide (NO) metabolites and increased the mRNA expression of klotho, a gene associated with an antiaging phenotype. To examine the effects of BE in humans, we conducted a clinical study using the second derivative of the fingertip photoplethysmogram to investigate vascular function and aging in 24 healthy volunteers. The participants consumed BE supplements (100 mg/day) or a placebo for 2 weeks. When compared with the placebo group, the BE group showed considerably improved vascular function, NO metabolite levels, and oxidative stress. These results suggest that BE supplementation improves endothelial function, possibly through antioxidant activity and NO production, and may consequently reduce the cardiovascular risk associated with aging. BE supplementation may be an effective and safe approach to reduce the risk of atherosclerosis and cardiovascular disease; however, additional studies investigating chronic vascular inflammation are needed.

Danhong formula alleviates endothelial dysfunction and reduces blood pressure in hypertension by regulating MicroRNA 24 - Phosphatidylinositol 3-Kinase-Serine/Threonine Kinase- Endothelial Nitric Oxide Synthase axis.

ETHNOPHARMACOLOGICAL RELEVANCE: Essential hypertension (EH) is one of the important risk factors of cardio-cerebrovascular diseases, and it can significantly increase the incidence and mortality of acute myocardial infarction, cerebral infarction and hemorrhage. Danhong Formula (DHF) was consisting of Radix et Rhizoma Salviae Miltiorrhizae (Salvia miltiorrhiza Bge., Labiatae, Danshen in Chinese) and Flos Carthami (Carthamus tinctorius L., Compositae, Honghua in Chinese) (Plant names have been checked with http://www.the plant list.org on June 28th, 2023) was approved by State Food and Drug Administration of China, that has been used for thousands of years in the treatment of cardiovascular diseases in China with proven safety and efficacy. Though our previous studies have found that DHF improved endothelial dysfunction (ED) and decreased high blood pressure (BP), the underlying mechanisms of its antihypertensive effect still remain unclear. AIM OF THE STUDY: This study investigated whether DHF regulated MicroRNA 24- Phosphatidylinositol 3-Kinase-Serine/Threonine Kinase- Endothelial Nitric Oxide Synthase (miR-24 - PI3K/AKT/eNOS) axis to produce antihypertensive effect and improve endothelial dysfunction. MATERIALS AND METHODS: Firstly, the chemical components of DHF were analyzed by UHPLC-MS. After that, BP was continuously monitored within the 1st, 3rd, and 4th week in SHR to evaluate the antihypertensive effect of DHF intraperitoneal injection. In addition, not only the contents of serum nitric oxide (NO), prostacyclin (PGI2), and angiotensin II (Ang II) were detected, but also the isolated aorta ring experiment was conducted to evaluate the vasomotoricity to evaluate of DHF on improving endothelial dysfunction. Key proteins or mRNA expression associated with miR-24 - PI3K/AKT/eNOS axis in aorta were detected by capillary Western blot, immunohistochemistry or RT-PCR to explore the underlying mechanisms. Index of NO, Ang II PGI2 and key proteins or mRNA expression were also conducted in miR-24-3p over-expression HUVECs model. RESULTS: Compared with SHR control group, DHF (4 mL/kg/day, 2 mL/kg/day, 1 mL/kg/day) treatment significantly reduced high BP in SHR and selectively increased acetylcholine (Ach) induced vasodilation, but not sodium nitroprusside (SNP) in a manner of concentration dependency in isolated aorta ring. DHF (4 mL/kg/day, 1 mL/kg/day) treatment was accompanying an increment of NO and PGI2, and lowering AngII in SHR. Moreover, DHF treatment significantly up-regulated expression of p-PI3K, p-AKT, mTOR, eNOS and p-eNOS, but down-regulated miR-24-3p expression in aorta. Compared with miR-24-3p over-expression HUVECs model group, DHF treatment inhibited miR- 24-3p expression and up-regulated p-PI3K, p-AKT, mTOR and eNOS mRNA expression. Similarly, DHF treatment increased PI3K, AKT, mTOR and eNOS protein expression in HUVECs by Western blot. CONCLUSIONS: These findings suggest that DHF alleviates endothelial dysfunction and reduces high BP in SHR mediated by down-regulating miR-24 via ultimately facilitating up-regulation of PI3K/AKT/eNOS axis. This current study firstly demonstrates a potential direction for antihypertensive mechanism of DHF from microRNA aspect and will promote its clinical applications.

Angelica gigas extract inhibits acetylation of eNOS via IRE1α sulfonation/RIDD-SIRT1-mediated posttranslational modification in vascular dysfunction.

Angelica gigas NAKAI (AG) is a popular traditional medicinal herb widely used to treat dyslipidemia owing to its antioxidant activity. Vascular disease is intimately linked to obesity-induced metabolic syndrome, and AG extract (AGE) shows beneficial effects on obesity-associated vascular dysfunction. However, the effectiveness of AGE against obesity and its underlying mechanisms have not yet been extensively investigated. In this study, 40 high fat diet (HFD) rats were supplemented with 100-300 mg/kg/day of AGE to determine its efficacy in regulating vascular dysfunction. The vascular relaxation responses to acetylcholine were impaired in HFD rats, while the administration of AGE restored the diminished relaxation pattern. Endothelial dysfunction, including increased plaque area, accumulated reactive oxygen species, and decreased nitric oxide (NO) and endothelial nitric oxide synthase (eNOS) Ser1177 phosphorylation, were observed in HFD rats, whereas AGE reversed endothelial dysfunction and its associated biochemical signaling. Furthermore, AGE regulated endoplasmic reticulum (ER) stress and IRE1α sulfonation and its subsequent sirt1 RNA decay through controlling regulated IRE1α-dependent decay (RIDD) signaling, ultimately promoting NO bioavailability via the SIRT1-eNOS axis in aorta and endothelial cells. Independently, AGE enhanced AMPK phosphorylation, additionally stimulating SIRT1 and eNOS deacetylation and its associated NO bioavailability. Decursin, a prominent constituent of AGE, exhibited a similar effect in alleviating endothelial dysfunctions. These data suggest that AGE regulates dyslipidemia-associated vascular dysfunction by controlling ROS-associated ER stress responses, especially IRE1α-RIDD/sirt1 decay and the AMPK-SIRT1 axis.

Progranulin Maintains Blood Pressure and Vascular Tone Dependent on EphrinA2 and Sortilin1 Receptors and Endothelial Nitric Oxide Synthase Activation.

Background The mechanisms determining vascular tone are still not completely understood, even though it is a significant factor in blood pressure management. Many circulating proteins have a significant impact on controlling vascular tone. Progranulin displays anti-inflammatory effects and has been extensively studied in neurodegenerative illnesses. We investigated whether progranulin sustains the vascular tone that helps regulate blood pressure. Methods and Results We used male and female C57BL6/J wild type (progranulin+/+) and B6(Cg)-Grntm1.1Aidi/J (progranulin-/-) to understand the impact of progranulin on vascular contractility and blood pressure. We found that progranulin-/- mice display elevated blood pressure followed by hypercontractility to noradrenaline in mesenteric arteries, which is restored by supplementing the mice with recombinant progranulin. In ex vivo experiments, recombinant progranulin attenuated the vascular contractility to noradrenaline in male and female progranulin+/+ arteries, which was blunted by blocking EphrinA2 or Sortilin1. To understand the mechanisms whereby progranulin evokes anticontractile effects, we inhibited endothelial factors. N(gamma)-nitro-L-arginine methyl ester (nitric oxide synthase inhibitor) prevented the progranulin effects, whereas indomethacin (cyclooxygenase inhibitor) affected only the contractility in arteries incubated with vehicle, indicating that progranulin increases nitric oxide and decreases contractile prostanoids. Finally, recombinant progranulin induced endothelial nitric oxide synthase phosphorylation and nitric oxide production in isolated mesenteric endothelial cells. Conclusions Circulating progranulin regulates vascular tone and blood pressure via EphrinA2 and Sortilin1 receptors and endothelial nitric oxide synthase activation. Collectively, our data suggest that deficiency in progranulin is a cardiovascular risk factor and that progranulin might be a new therapeutic avenue to treat high blood pressure.

Chronic SIRT1 supplementation in diabetic mice improves endothelial function by suppressing oxidative stress.

AIMS: Enhancing SIRT1 activity exerts beneficial cardiovascular effects. In diabetes, plasma SIRT1 levels are reduced. We aimed to investigate the therapeutic potential of chronic recombinant murine SIRT1 (rmSIRT1) supplementation to alleviate endothelial and vascular dysfunction in diabetic mice (db/db). METHODS AND RESULTS: Left internal mammary arteries obtained from patients undergoing coronary artery bypass grafting with or without a diagnosis of diabetes were assayed for SIRT1 protein levels. Twelve-week-old male db/db mice and db/+ controls were treated with vehicle or rmSIRT1 intraperitoneally for 4 weeks, after which carotid artery pulse wave velocity (PWV) and energy expenditure/activity were assessed by ultrasound and metabolic cages, respectively. Aorta, carotid, and mesenteric arteries were isolated to determine endothelial and vascular function using the myograph system.Arteries obtained from diabetic patients had significantly lower levels of SIRT1 relative to non-diabetics. In line, aortic SIRT1 levels were reduced in db/db mice compared to db/+ mice, while rmSIRT1 supplementation restored SIRT1 levels. Mice receiving rmSIRT1 supplementation displayed increased physical activity and improved vascular compliance as reflected by reduced PWV and attenuated collagen deposition. Aorta of rmSIRT1-treated mice exhibited increased endothelial nitric oxide (eNOS) activity, while endothelium-dependent contractions of their carotid arteries were significantly decreased, with mesenteric resistance arteries showing preserved hyperpolarization. Ex vivo incubation with reactive oxygen species (ROS) scavenger Tiron and NADPH oxidase inhibitor apocynin revealed that rmSIRT1 leads to preserved vascular function by suppressing NADPH oxidase (NOX)-related ROS synthesis. Chronic rmSIRT1 treatment resulted in reduced expression of both NOX1 and NOX4, in line with a reduction in aortic protein carbonylation and plasma nitrotyrosine levels. CONCLUSIONS: In diabetic conditions, arterial SIRT1 levels are significantly reduced. Chronic rmSIRT1 supplementation improves endothelial function and vascular compliance by enhancing eNOS activity and suppressing NOX-related oxidative stress. Thus, SIRT1 supplementation may represent novel therapeutic strategy to prevent diabetic vascular disease.

Natural Bioactive Compounds Targeting NADPH Oxidase Pathway in Cardiovascular Diseases.

Cardiovascular disease (CVD) is the leading cause of death worldwide, in both developed and developing countries. According to the WHO report, the morbidity and mortality caused by CVD will continue to rise with the estimation of death going up to 22.2 million in 2030. NADPH oxidase (NOX)-derived reactive oxygen species (ROS) production induces endothelial nitric oxide synthase (eNOS) uncoupling and mitochondrial dysfunction, resulting in sustained oxidative stress and the development of cardiovascular diseases. Seven distinct members of the family have been identified of which four (namely, NOX1, 2, 4 and 5) may have cardiovascular functions. Currently, the treatment and management plan for patients with CVDs mainly depends on the drugs. However, prolonged use of prescribed drugs may cause adverse drug reactions. Therefore, it is crucial to find alternative treatment options with lesser adverse effects. Natural products have been gaining interest as complementary therapy for CVDs over the past decade due to their wide range of medicinal properties, including antioxidants. These might be due to their potent active ingredients, such as flavonoid and phenolic compounds. Numerous natural compounds have been demonstrated to have advantageous effects on cardiovascular disease via NADPH cascade. This review highlights the potential of natural products targeting NOX-derived ROS generation in treating CVDs. Emphasis is put on the activation of the oxidases, including upstream or downstream signalling events.

Myocardial angiogenesis induced by concurrent vitamin D supplementation and aerobic-resistance training is mediated by inhibiting miRNA-15a, and miRNA-146a and upregulating VEGF/PI3K/eNOS signaling pathway.

This study aimed to investigate the effects of co-treatment of aerobic-resistance training (ART), vitamin D3 (VD3) on cardiovascular function considering the involvement of microRNA-15a and microRNA-146a, vascular endothelial growth factor (VEGF), phosphatidylinositol-3 kinase (PI3K), and endothelial nitric oxide synthase (eNOS) after myocardial infarction (MI) in rats. To induce MI, male Wistar rats subcutaneously received isoproterenol for 2 days, then MI was confirmed by echocardiography. MI rats were divided into six groups (n = 8/group). MI + VD3, MI + sesame oil (Veh), MI + ART, MI + VD3 + ART, and MI + Veh + ART, and received the related treatments for 8 weeks. Exercise tests, echocardiography, real-time quantitative polymerase chain reaction (qRT-PCR), western blotting, and histological staining were performed after the end of treatments. The highest ejection fraction (EF%), fractional shortening (FS%), exercise capacity (EC), and maximal load test (MLT) amounts were observed in the groups treated with VD3, ART, and VD3 + ART (P < 0.05). These were accompanied by a significantly increased angiogenesis post-MI. Furthermore, the levels of circulating microRNA-15a and microRNA-146a were significantly decreased in these groups compared to MI rats that were together with a significant upregulation of cardiac VEGF, PI3K, and eNOS expression. Overall, the best results were observed in the group treated with VD3 + ART. Concurrent VD3 supplementation and ART attenuated microRNA-15a and microRNA-146a and induced angiogenesis via VEGF/PI3K/eNOS axis. This data demonstrate that concurrent VD3 supplementation and ART is a more efficient strategy than monotherapy to improve cardiac function post-MI.

Berberine promotes the viability of random skin flaps via the PI3K/Akt/eNOS signaling pathway.

Random skin flaps are often used in reconstruction operations. However, flap necrosis is still a common postoperative complication. Here, we investigated whether berberine (C20 H19 NO5 , BBR), a drug with antioxidant activity, improves the survival rate of random flaps. Fifty-four rats were divided into three groups: control, BBR and BBR + L -NAME groups (L -NAME, L -NG -Nitro-arginine methyl ester). The survival condition and the percentage of survival area of the flaps were evaluated on the seventh day after surgery. After animals were sacrificed, angiogenesis, apoptosis, oxidative stress and inflammation levels were assessed by histological and protein analyses. Our findings suggest that berberine promotes flap survival. The level of angiogenesis increased; the levels of oxidative stress, inflammation and apoptosis decreased; the levels of phosphoinositide 3-kinase (PI3K), phospho-Akt (p-Akt) and phospho-endothelial nitric oxide synthase (p-eNOS) increased in the flap tissue; and L -NAME reversed the effects of berberine on random skin flaps. Statistical analysis showed that the BBR group results differed significantly from those of the control and the BBR + L -NAME groups (p < .05). Our results confirm that berberine is an effective drug for significantly improving the survival rate of random skin flaps by promoting angiogenesis, inhibiting inflammation, attenuating oxidative stress, and reducing apoptosis through the PI3K/Akt/eNOS signaling pathway.

Effects of resveratrol supplementation on nitric oxide-mediated vascular outcomes in hypertension: A systematic review.

Hypertension is associated with endothelial dysfunction and decreased nitric oxide (NO). It has been proposed that decreasing oxidative stress may help regulate blood pressure by increasing NO concentration. Therefore, the purpose of this systematic review was to determine whether the antioxidant resveratrol effects NO-mediated vascular outcomes in hypertension. A comprehensive literature search of PubMed and EBSCOhost databases was conducted using the terms: "resveratrol" and "nitric oxide or NO" and "hypertension or high blood pressure." Searches were not restricted for year of publication or study design but limited to full-text studies from scholarly, peer-reviewed journals. Ten animal studies published between 2005 and 2017 were identified. Human studies did not meet criteria and were not included. Articles were critically assessed using the Academy of Nutrition and Dietetics' Evidence Analysis Library Quality Criteria Worksheet. All studies evaluated resveratrol supplementation and at least one NO outcome measure including: circulating NO metabolites, endothelial nitric oxide synthase (eNOS) expression, eNOS phosphorylation, and eNOS uncoupling. All but one study assessed blood pressure. Nine of ten studies reported positive significant results of resveratrol supplementation on NO outcomes, and in all but one study, this was seen concomitantly with decreases in blood pressure. Resveratrol supplementation shows promise for improving NO-mediated vascular outcomes and improving blood pressure. Translation to human studies is warranted, with dose of resveratrol considered, as the human equivalency doses are not consistent amongst animal studies. Additionally, a standard battery of tests examining NO-mediated vascular outcomes is needed to ensure generalizability among studies to determine dose-duration effects.

The Beneficial Impact of Zinc Supplementation on the Vascular Tissue of the Abdominal Aorta under Repeated Intoxication with Cadmium: A Study in an In Vivo Experimental Model.

In an in vivo rat model of human exposure to cadmium (Cd; 5 and 50 mg/L, 6 months), whether the supplementation with zinc (Zn; 30 and 60 mg/L, increasing its daily intake by 79% and 151%, respectively) protects against the unfavourable impact of this xenobiotic on the vascular tissue of the abdominal aorta was investigated. The treatment with Cd led to oxidative stress and increased the concentrations of pro-inflammatory interleukin 1ß (IL-1ß), total cholesterol (TC), triglycerides (TG), and endothelial nitric oxide synthase (eNOS) and decreased the concentration of anti-inflammatory interleukin 10 (IL-10) in the vascular tissue. Cd decreased the expression of intercellular adhesion molecule-1 (ICAM-1), platelet endothelial cell adhesion molecule-1 (PECAM-1), and L-selectin on the endothelial cells. The administration of Zn prevented most of the Cd-induced alterations or at least weakened them (except for the expression of adhesive molecules). In conclusion, Zn supplementation may protect from the toxic impact of Cd on the blood vessels and thus exert a beneficial influence on the cardiovascular system. The increase in the intake of Zn by 79% may be sufficient to provide this protection and the effect is related to the antioxidative, anti-inflammatory, and antiatherogenic properties of this essential element.

Grape Pomace Polyphenols as a Source of Compounds for Management of Oxidative Stress and Inflammation-A Possible Alternative for Non-Steroidal Anti-Inflammatory Drugs?

Flavonoids, stilbenes, lignans, and phenolic acids, classes of polyphenols found in grape pomace (GP), were investigated as an important alternative source for active substances that could be used in the management of oxidative stress and inflammation. The benefic antioxidant and anti-inflammatory actions of GP are presented in the literature, but they are derived from a large variety of experimental in vitro and in vivo settings. In these in vitro works, the decrease in reactive oxygen species, malondialdehyde, and thiobarbituric acid reactive substances levels and the increase in glutathione levels show the antioxidant effects. The inhibition of nuclear factor kappa B and prostaglandin E2 inflammatory pathways and the decrease of some inflammatory markers such as interleukin-8 (IL-8) demonstrate the anti-inflammatory actions of GP polyphenols. The in vivo studies further confirmed the antioxidant (increase in catalase, superoxide dismutase and glutathione peroxidase levels and a stimulation of endothelial nitric oxide synthase -eNOS gene expression) and anti-inflammatory (inhibition of IL-1𝛼, IL-1ß, IL-6, interferon-𝛾, TNF-α and C-reactive protein release) activities. Grape pomace as a whole extract, but also different individual polyphenols that are contained in GP can modulate the endogenous pathway responsible in reducing oxidative stress and chronic inflammation. The present review analyzed the effects of GP in oxidative stress and inflammation, suggesting that it could become a valuable therapeutic candidate capable to reduce the aforementioned pathological processes. Grape pomace extract could become an adjuvant treatment in the attempt to reduce the side effects of the classical anti-inflammatory medication like non-steroidal anti-inflammatory drugs (NSAIDs).

Role of BH4 deficiency as a mediator of oxidative stress-related endothelial dysfunction in menopausal women.

Endothelial function (brachial artery flow-mediated dilation [FMD]) is reduced in estrogen-deficient postmenopausal women, mediated, in part, by reduced nitric oxide (NO) bioavailability, secondary to tetrahydrobiopterin (BH4) deficiency and oxidative stress. FMD is increased, but not fully restored, in postmenopausal women after acute intravenous vitamin C (VITC; superoxide scavenger) or oral BH4 supplementation. In vitro studies demonstrate that coadministration of VITC with BH4 prevents endothelial nitric oxide synthase (eNOS) uncoupling and reductions in NO by peroxynitrite. To investigate mechanisms of endothelial dysfunction in women, we assessed the separate and combined effects of VITC and BH4 to determine whether coadministration of VITC + BH4 improves FMD in healthy postmenopausal women (n = 19, 58 ± 5 yr) to premenopausal (n = 14, 36 ± 9 yr) levels, with exploratory testing in perimenopausal women (n = 8, 51 ± 3 yr). FMD was measured during acute intravenous infusions of saline (control) and VITC (∼2-3 g) ∼3 h after a single dose of oral BH4 (KUVAN, 10 mg/kg body wt) or placebo (randomized crossover, separated by ∼1 mo). Under the placebo condition, FMD was reduced in postmenopausal compared with premenopausal women during the saline infusion (5.6 ± 0.7 vs. 11.6 ± 0.9%, P < 0.001) and increased in postmenopausal women during VITC (+3.5 [1.4, 5.6]%, P = 0.001) and acute BH4 (+1.8 [0.37, 3.2]%, P = 0.01) alone. Coadministration of VITC + BH4 increased FMD in postmenopausal women (+3.0 [1.7, 4.3]%, P < 0.001), but FMD remained reduced compared with premenopausal women (P = 0.02). Exploratory analyses revealed that VITC + BH4 did not restore FMD in perimenopausal women to premenopausal levels (P = 0.045). Coadministration of VITC + BH4 does not restore FMD in menopausal women, suggesting that additional mechanisms may be involved.NEW & NOTEWORTHY Endothelial function is reduced across the menopausal stages related to increased oxidative stress associated with estrogen deficiency. In vitro studies demonstrate that coadministration of VITC with BH4 prevents endothelial nitric oxide synthase (eNOS) uncoupling and reductions in NO by peroxynitrite; however, this remains untested in humans. We demonstrate that the coadministration of BH4 + VITC does not restore endothelial function in perimenopausal and postmenopausal women to the level of premenopausal women, suggesting that other mechanisms contribute.

The mechanism of Yinchenhao decoction in treating obstructive-jaundice-induced liver injury based on Nrf2 signaling pathway.

BACKGROUND: Obstructive jaundice (OJ) is caused by bile excretion disorder after partial or complete bile duct obstruction. It may cause liver injury through various mechanisms. Traditional Chinese medicine (TCM) has a lot of advantages in treating OJ. The recovery of liver function can be accelerated by combining Chinese medicine treatment with existing clinical practice. Yinchenhao decoction (YCHD), a TCM formula, has been used to treat jaundice. Although much progress has been made in recent years in understanding the mechanism of YCHD in treating OJ-induced liver injury, it is still not clear. AIM: To investigate chemical components of YCHD that are effective in the treatment of OJ and predict the mechanism of YCHD. METHODS: The active components and putative targets of YCHD were predicted using a network pharmacology approach. Gene Ontology biological process and Kyoto Encyclopedia of Genes and Genomes path enrichment analysis were carried out by cluster profile. We predicted the biological processes, possible targets, and associated signaling pathways that YCHD may involve in the treatment of OJ. Thirty male Sprague-Dawley rats were randomly divided into three groups, each consisting of 10 rats: the sham group (Group S), the OJ model group (Group M), and the YCHD-treated group (Group Y). The sham group only received laparotomy. The OJ model was established by ligating the common bile duct twice in Groups M and Y. For 1 wk, rats in Group Y were given a gavage of YCHD (3.6 mL/kg) twice daily, whereas rats in Groups S and M were given the same amount of physiological saline after intragastric administration daily. After 7 d, all rats were killed, and the liver and blood samples were collected for histopathological and biochemical examinations. Total bilirubin (TBIL), direct bilirubin (DBIL), alanine aminotransferase (ALT), and aspartate transaminase (AST) levels in the blood samples were detected. The gene expression levels of inducible nitric oxide synthase (iNOS) and endothelial nitric oxide synthase (eNOS), and the nucleus positive rate of NF-E2 related factor 2 (Nrf2) protein were measured. Western blot analyses were used to detect the protein and gene expression levels of Nrf2, Kelch-like ECH-associated protein 1, NAD(P)H quinone dehydrogenase 1 (NQO1), and glutathione-S-transferase (GST) in the liver tissues. One-way analysis of variance was used to evaluate the statistical differences using the statistical package for the social sciences 23.0 software. Intergroup comparisons were followed by the least significant difference test and Dunnett's test. RESULTS: The effects of YCHD on OJ involve biological processes such as DNA transcription factor binding, RNA polymerase II specific regulation, DNA binding transcriptional activator activity, and nuclear receptor activity. The protective effects of YCHD against OJ were closely related to 20 pathways, including the hepatitis-B, the mitogen-activated protein kinase, the phosphatidylinositol 3-kinase/protein kinase B, and tumor necrosis factor signaling pathways. YCHD alleviated the swelling and necrosis of hepatocytes. Following YCHD treatment, the serum levels of TBIL (176.39 ± 17.03 µmol/L vs 132.23 ± 13.88 µmol/L, P < 0.01), DBIL (141.41 ± 14.66 µmol/L vs 106.43 ± 10.88 µmol/L, P < 0.01), ALT (332.07 ± 34.34 U/L vs 269.97 ± 24.78 U/L, P < 0.05), and AST (411.44 ± 47.64 U/L vs 305.47 ± 29.36 U/L, P < 0.01) decreased. YCHD promoted the translocation of Nrf2 into the nucleus (12.78 ± 0.99 % vs 60.77 ± 1.90 %, P < 0.001). After YCHD treatment, we found a decrease in iNOS (0.30 ± 0.02 vs 0.20 ± 0.02, P < 0.001) and an increase in eNOS (0.18 ± 0.02 vs 0.32 ± 0.02, P < 0.001). Meanwhile, in OJ rats, YCHD increased the expressions of Nrf2 (0.57 ± 0.03 vs 1.18 ± 0.10, P < 0.001), NQO1 (0.13 ± 0.09 vs 1.19 ± 0.07, P < 0.001), and GST (0.12 ± 0.02 vs 0.50 ± 0.05, P < 0.001), implying that the potential mechanism of YCHD against OJ-induced liver injury was the upregulation of the Nrf2 signaling pathway. CONCLUSION: OJ-induced liver injury is associated with the Nrf2 signaling pathway. YCHD can reduce liver injury and oxidative damage by upregulating the Nrf2 pathway.

Bergapten mediated inflammatory and apoptosis through AMPK/eNOS/AKT signaling pathway of isoproterenol-induced myocardial infarction in Wistar rats.

Bergapten (BeG) is explored for its anti-inflammatory and antioxidant properties. Myocardial infarction (MI) is reported to be one of the leading cardiovascular diseases characterized by mitochondrial dysfunction and apoptosis. The main purpose of this study is to assess the cardiopreventive effects of BeG (50 mg/kg) in isoproterenol (ISO)-induced MI in Wistar rats. The increased infarct size after ISO induction was reduced simultaneously on treatment with BeG. Similarly, augmented levels of cardiac biomarkers, namely cardiac troponin T, creatine kinase (CK), cardiac troponin I, and CK-MB were also suppressed by BeG. The increased rate of lipid hydroperoxides and thiobarbituric acid reactive substances owing to the oxidative stress caused by free radical generation in ISO-induced rats were also inhibited by BeG. Antioxidants reduce oxidative stress by scavenging free radicals. ISO induction reduces these antioxidant enzymes glutathione peroxidase, catalase, superoxide dismutase, and glutathione, and levels causing oxidative cardiac damage to the heart tissue. BeG supplementation improved these enzymes synthesis preventing potential damage to the myocardium. Inflammation caused by ISO pretreatment increased the secretion of proinflammatory cytokines in ISO-induced rats. Pretreatment with BeG suppressed these inflammatory cytokines to a normal level in ISO + BeG-treated rats. The histopathological examination of the morphological characteristics showed that the intensity of cardiac damage caused by ISO induction was less in BeG pretreated rats with less inflammatory cells and no necrosis. BeG also showed promising results in the molecular alteration of AMP-activated protein kinase/endothelial nitric oxide synthase/protein kinase B signaling molecules. These observations emphasize the cardioprotective effects of BeG and its potential use as a drug in the near future.

Influence of electroacupuncture on ghrelin and the phosphoinositide 3-kinase/protein kinase B/endothelial nitric oxide synthase signaling pathway in spontaneously hypertensive rats.

OBJECTIVE: To investigate the influence of electroacupuncture (EA) on ghrelin and the phosphoinositide 3-kinase/protein kinase B/endothelial nitric oxide synthase (PI3K/Akt/eNOS) signaling pathway in spontaneously hypertensive rats (SHRs). METHODS: Eight Wistar-Kyoto rats were used as the healthy blood pressure (BP) control (normal group), and 32 SHRs were randomized into model group, EA group, EA plus ghrelin group (EA + G group), and EA plus PF04628935 group (a potent ghrelin receptor blocker; EA + P group) using a random number table. Rats in the normal group and model group did not receive treatment, but were immobilized for 20 min per day, 5 times a week, for 4 continuous weeks. SHRs in the EA group, EA + G group and EA + P group were immobilized and given EA treatment in 20 min sessions, 5 times per week, for 4 weeks. Additionally, 1 h before EA, SHRs in the EA + G group and EA + P group were intraperitoneally injected with ghrelin or PF04628935, respectively, for 4 weeks. The tail-cuff method was used to measure BP. After the 4-week intervention, the rats were sacrificed by cervical dislocation, and pathological morphology of the abdominal aorta was observed using hematoxylin-eosin (HE) staining. Enzyme-linked immunosorbent assay (ELISA) was used to detect the levels of ghrelin, nitric oxide (NO), endothelin-1 (ET-1) and thromboxane A2 (TXA2) in the serum. Isolated thoracic aortic ring experiment was performed to evaluate vasorelaxation. Western blot was used to measure the expression of PI3K, Akt, phosphorylated Akt (p-Akt) and eNOS proteins in the abdominal aorta. Further, quantitative real-time polymerase chain reaction (qRT-PCR) was conducted to measure the relative levels of mRNA expression for PI3K, Akt and eNOS in the abdominal aorta. RESULTS: EA significantly reduced the systolic BP (SBP) and diastolic BP (DBP) (P < 0.05). HE staining showed that EA improved the morphology of the vascular endothelium to some extent. Results of ELISA indicated that higher concentrations of ghrelin and NO, and lower concentrations of ET-1 and TXA2 were presented in the EA group (P < 0.05). The isolated thoracic aortic ring experiment demonstrated that the vasodilation capacity of the thoracic aorta increased in the EA group. Results of Western blot and qRT-PCR showed that EA increased the abundance of PI3K, p-Akt/Akt and eNOS proteins, as well as expression levels of PI3K, Akt and eNOS mRNAs (P < 0.05). In the EA + G group, SBP and DBP decreased (P < 0.05), ghrelin concentrations increased (P < 0.05), and the concentrations of ET-1 and TXA2 decreased (P < 0.05), relative to the EA group. In addition, the levels of PI3K and eNOS proteins, the p-Akt/Akt ratio, and the expression of PI3K, Akt and eNOS mRNAs increased significantly in the EA + G group (P < 0.05), while PF04628935 reversed these effects. CONCLUSION: EA effectively reduced BP and protected the vascular endothelium, and these effects may be linked to promoting the release of ghrelin and activation of the PI3K/Akt/eNOS signaling pathway.

Ginkgo biloba L. extract prevents steroid-induced necrosis of the femoral head by rescuing apoptosis and dysfunction in vascular endothelial cells via the PI3K/AKT/eNOS pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Ginkgo biloba L. extract (EGb) is one of the world's most extensively used herbal medicines. Due to the diverse pharmacological properties of EGb, it has been used in the treatment of neurological illnesses, as well as cardiovascular and cerebrovascular ailments. However, the effect and pharmacological mechanism of EGb on steroid-induced necrosis of the femoral head (SINFH) are still unclear. AIM OF THE STUDY: SINFH remains a challenging problem in orthopedics. Previous investigations have shown that EGb has the potential to reduce the occurrence of SINFH. The goal was to determine the effect and mechanism of EGb in preventing SINFH by inhibiting apoptosis and improving vascular endothelial cells (VECs) functions. MATERIALS AND METHODS: CCK-8, nitric oxide (NO) production and flow cytometry were used to determine the cell apoptosis and function. The scratch and angiogenesis tests assessed migration and tube formation. Western blot analysis detected the expressions of apoptosis-related proteins and PI3K/AKT/eNOS pathway-related proteins. Apoptosis and angiogenesis were also detected treated with the inhibitors. A mouse model of SINFH was established. Paraffin section was used to determine the necrotic pathology and apoptosis. Vessels in the femoral heads were assessed by immunofluorescence staining. RESULTS: When stimulated by methylprednisolone (MPS), cell viability, NO generation and tube formation were decreased, the apoptotic rate increased. Simultaneously, MPS decreased the expression levels of p-PI3K, p-AKT, and p-eNOS. EGb increased the expression levels of these proteins, restrained apoptosis, and restored cell functions. The addition of the inhibitors decreased anti-apoptotic effect and angiogenesis. In addition, when compared to the model mice, there were fewer empty lacunae and normal trabecular arrangement after taking different doses of EGb. The protective effect was also confirmed by the vascular quantitative analysis in vivo. CONCLUSION: This study established that EGb increased endothelial cell activity and inhibited apoptosis and function loss induced by MPS, elucidating the effect and molecular mechanism of EGb on early SINFH.