Near-Infrared II Photobiomodulation Preconditioning Ameliorates Stroke Injury via Phosphorylation of eNOS.

BACKGROUND: The current management of patients with stroke with intravenous thrombolysis and endovascular thrombectomy is effective only when it is timely performed on an appropriately selected but minor fraction of patients. The development of novel adjunctive therapy is highly desired to reduce morbidity and mortality with stroke. Since endothelial dysfunction is implicated in the pathogenesis of stroke and is featured with suppressed endothelial nitric oxide synthase (eNOS) with concomitant nitric oxide deficiency, restoring endothelial nitric oxide represents a promising approach to treating stroke injury. METHODS: This is a preclinical proof-of-concept study to determine the therapeutic effect of transcranial treatment with a low-power near-infrared laser in a mouse model of ischemic stroke. The laser treatment was performed before the middle cerebral artery occlusion with a filament. To determine the involvement of eNOS phosphorylation, unphosphorylatable eNOS S1176A knock-in mice were used. Each measurement was analyzed by a 2-way ANOVA to assess the effect of the treatment on cerebral blood flow with laser Doppler flowmetry, eNOS phosphorylation by immunoblot analysis, and stroke outcomes by infarct volumes and neurological deficits. RESULTS: Pretreatment with a 1064-nm laser at an irradiance of 50 mW/cm2 improved cerebral blood flow, eNOS phosphorylation, and stroke outcomes. CONCLUSIONS: Near-infrared II photobiomodulation could offer a noninvasive and low-risk adjunctive therapy for stroke injury. This new modality using a physical parameter merits further consideration to develop innovative therapies to prevent and treat a wide array of cardiovascular diseases.

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.

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.

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.

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.

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.

Hypericum perforatum L. protects against renal function decline in ovariectomy rat model by regulating expressions of NOS3 and AKT1 in AGE-RAGE pathway.

BACKGROUND: Hypericum perforatum L. (HPL) is a potential traditional Chinese medicine. It could promotes menopausal 'kidney-yin deficiency syndrome' that characterized by renal function decline. However, its potential pharmacological effect and mechanism remains unknown. OBJECTIVE: The aim of this study was to investigate whether HPL can improve menopausal renal function decline and to explore its mechanism of action. METHODS: The mainly ingredients of HPL were identified using UPLC-Q-TOF-MS/MS approach, and the potential therapeutic targets of HPL for renal function decline were chose via network pharmacology technique. The key therapeutic metabolites were selected through non-targeted metabolomic and chemometric methods. Then, the network were constructed and the key targets and metabolites were screened. At last, the validation experiments and mechanism exploring were adopted by using Immunofluorescence, enzyme-linked immunosorbent assay (ELISA), real-time PCR (RT-PCR), and western blotting assays. RESULTS: mainly ingredients of HPL were identified and determined 17 compounds and 29 targets were chose as mainly active compounds and potential therapeutic targets. Based on OVX induced renal decline rat model, after chemometric analysis, 59 endo-metabolites were selected as key therapeutic metabolites, and AGE-RAGE signal pathway in diabetes complications was enriched as the key pathway. By constructing a "disease-component-target" network, Hyperoside, Quercetrin, and quinic were selected as the key therapeutic compounds, and the AKT1 and NOS3 were selected as the key therapeutic targets. The results of ELISA, RT-PCR and western blot experiments indicated that HPL could rescue the abnormal expressions both of AKT1 and NOS3, as well as their related metabolites distortion. CONCLUSION: Our findings indicated that HPL regulated expression of AKT1 and NOS3 through modulating AGE-RAGE signaling pathway in OVX stimulated rats` renal dysfunction, implicating the potential values of HPL in menopause syndromes therapy.

Pentaclethra macrophylla leaves reduce blood glucose level, improve sexual behavior and modulate critical enzymes associated with diabetes-erectile dysfunction morbidity in rats.

OBJECTIVES: This research work studied the phenolic composition of Pentaclethra macrophylla (PM), the inclusion of dietary supplementation of PM leaves on sexual functions and its connection to inhibit enzymes (arginase and phosphodiesterase-5) and nitric oxide level, linked to type 2 diabetes-induced erectile dysfunction in rats. METHODS: Gallic acid, chlorogenic and ellagic acids, Kaempferol, and epicatechin etc. was spotted with High performance liquid chromatography-diode array detector from PM extract. Twenty-five (25) rats were used for the study. Five rats were placed with basal diet; diets not supplemented with PM leaves (normal rat group) while twenty rats were made diabetic by feeding them with high fat diet for two weeks, prior to single injection with 35 mg/kg of streptozotocin (STZ). After checking with glucometer, experimental animals with blood glucose level >250 mg/dL were accepted as diabetic. The diabetic rats were subsequently divided into four groups of five rats each (n=5). The diabetic rats were placed on basal diet, or diets supplemented with PM leaves (10 % or 5 % inclusion) or sildenafil citrate (SC). RESULTS: The result revealed that PM supplemented diets caused significant (p<0.05) reduction in blood glucose level, and augmented erectile function by inhibiting arginase and PDE5 activities as well as enhancing nitric oxide level. CONCLUSIONS: In conclusion, dietary inclusion of PM leaves could serve as a potent nutraceutical source in hyperglycemia induced erectile dysfunction management.

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.

Exploring the mechanism of the antithrombotic effects of Pueraria lobata and Pueraria lobata var. thomsonii based on network pharmacology.

ETHNOPHARMACOLOGICAL RELEVANCE: Pueraria lobata (Willd.) Ohwi and Pueraria lobata var. thomsonii (Benth.) Maesen are nutritious medicine food homology plants that are widely used in the food and health products industry and are excellent natural materials for the development of new health foods, with great potential for domestic and foreign markets. Clinically, P. lobata and P. thomsonii are used to treat coronary heart disease, atherosclerosis, cerebral infarction and other cardiovascular diseases, and antithrombotic actions may be their core effect in the treatment of thrombotic diseases. However, the underlying mechanisms of the antithrombotic properties of P. lobata and P. thomsonii have not been clarified. METHODS: First, P. lobata and P. thomsonii were identified by high-performance liquid chromatography (HPLC). An arteriovenous bypass thrombosis rat model was established. Thrombus dry‒wet weight, platelet accumulation rate and the four coagulation indices, including activated partial thromboplastin time (APTT), prothrombin time (PT), thrombin time (TT) and fibrinogen (FIB), were detected in plasma to manifest the P. lobata and P. thomsonii antithrombotic function. Network pharmacology and molecular docking methods were used to obtain key targets and verify reliability. David 6.8 was used for GO and KEGG analyses to explore pathways and potential targets for P. lobata and P. thomsonii antithrombotic functions. Prostaglandin I2 (PGI2), thromboxane A2 (TXA2), cyclooxygenase 2 (COX-2), myeloperoxidase (MPO) and endothelial nitric oxide synthase (eNOS) were tested by enzyme-linked immunosorbent assay (ELISA). RESULTS: The results indicated that P. lobata and P. thomsonii can reduce thrombus dry‒wet weight and platelet accumulation in rats and inhibit TT, APTT, FIB, and PT. A comprehensive network pharmacology approach successfully identified 9 active ingredients in P. lobata and P. thomsonii. The main active ingredients include polyphenols, amino acids and flavonoids. A total of 15 antithrombotic function targets were obtained, including 3 key targets (PTGS2, NOS3, MPO). Pathway analysis showed 10 significant related pathways and 29 biological processes. P. lobata and P. thomsonii inhibited platelet aggregation by upregulating PGI2 and downregulating TXA2, inhibited PTGS2 to reduce inflammation, and increased the level of eNOS to promote vasodilation. In addition, P. lobata and P. thomsonii alleviated oxidative stress by increasing SOD levels and significantly decreasing MDA contents. CONCLUSION: The results of the study further clarify the antithrombotic mechanism of action of P. lobata and P. thomsonii, which provides a scientific basis for the development of new drugs for thrombogenic diseases and lays the foundation for the development of P. lobata and P. thomsonii herbal resources and P. lobata and P. thomsonii health products.

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.