Vascular Relaxation Research Articles

The impact of thyroid hormone dysfunction on ischemic heart disease.

Thyroid hormones have a central role in cardiovascular homeostasis. In myocardium, these hormones stimulate both diastolic myocardial relaxation and systolic myocardial contraction, have a pro-angiogenic effect and an important role in extracellular matrix maintenance. Thyroid hormones modulate cardiac mitochondrial function. Dysfunction of thyroid axis impairs myocardial bioenergetic status. Both overt and subclinical hypothyroidism are associated with a higher incidence of coronary events and an increased risk of heart failure progression. Endothelial function is also impaired in hypothyroid state, with decreased nitric oxide-mediated vascular relaxation. In heart disease, particularly in ischemic heart disease, abnormalities in thyroid hormone levels are common and are an important factor to be considered. In fact, low thyroid hormone levels should be interpreted as a cardiovascular risk factor. Regarding ischemic heart disease, during the late post-myocardial infarction period, thyroid hormones modulate left ventricular structure, function and geometry. Dysfunction of thyroid axis might even be more prevalent in the referred condition since there is an upregulation of type 3 deiodinase in myocardium, producing a state of local cardiac hypothyroidism. In this focused review, we summarize the central pathophysiological and clinical links between altered thyroid function and ischemic heart disease. Finally, we highlight the potential benefits of thyroid hormone supplementation as a therapeutic target in ischemic heart disease.

Resistance Arteries from Low‐Capacity Running Rats Exhibit Diminished Acetylcholine‐Induced Relaxation in Comparison to High‐Capacity Running Rats: Effects of Native and Allografted Perivascular Adipose Tissue on Vascular Function

Previous research has demonstrated that perivascular adipose tissue (PVAT) plays a significant metabolic role in vascular function by secreting a variety of bioactive substances in both healthy and diseased states. PVAT has been largely recognized to cause an anti‐contractile response; this anti‐contractile action is lost in cardiovascular diseases, including hypertension. Acknowledging the importance of PVAT in vascular health, we hypothesized that the impact of PVAT on vascular function would be different in healthy and diseased states. Specifically, we investigated the potential therapeutic and harmful effects of native PVAT and allografted PVAT on vascular function from an animal model of metabolic syndrome compared to an animal model for longer lifespan. Our animal model (male, 20–24‐weeks old) study used low‐capacity running rats (LCR) as a model for metabolic syndrome, high‐capacity running rats (HCR) as a model for an “ultra‐healthy” state and high response trainer rats (HRT) served as a control. The intrinsic and trained aerobic capacity of HRT rats is higher than LCR rats but lower than HCR rats. Mesenteric resistance arteries (MRA) and MRA PVAT from HRT rats, LCR rats, and HCR rats were extracted. The MRA were isolated and mounted in a wire myograph. Concentration‐response curves to acetylcholine (1nM – 30μM) were performed in the absence and presence of an animal's native mesenteric PVAT or LCR/HCR grafted mesenteric PVAT. T‐test *p < 0.05; n=3–4. We observed a dramatic change in PVAT structure between control, LCR, and HCR animals (Figure 1). The efficacy of acetylcholine, an endothelium‐dependent vasodilator of MRA, was diminished in LCR rats compared to HRT rats and HCR rats (Emax: HRT: 97.88 ± 0.8314 vs. LCR: 77.43 ± 7.038* vs. HCR: 97.70 ± 1.942, p < 0.05). As has been previously shown, the presence of native PVAT increased the HRT MRA sensitivity to acetylcholine‐induced relaxation. However, the presence of native PVAT diminished the LCR MRA sensitivity to acetylcholine‐induced relaxation. This sensitivity was restored by allografting HCR PVAT onto the LCR MRA. Although no changes were observed in HCR MRA relaxation in the presence of its native PVAT, allografting LCR PVAT onto the HCR MRA decreased the HCR MRA relaxation. This data suggests that LCR rats present with an impaired PVAT that contributes to diminished vascular relaxation. On the other hand, HCR rats present with an enhanced PVAT that could potentially improve vascular relaxation. This study demonstrates the potential therapeutic effect of PVAT from HCR rats.Support or Funding InformationThis work was supported by National Institutes of Health (NIH) (NIGMS: K99GM11888).This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Testosterone Replacement Enhances Internal Pudendal Artery Relaxation to Reverse Erectile Dysfunction in a Rat Model of Androgen Deprivation Therapy

Introduction/ObjectivesAndrogen deprivation therapy (ADT) is frequently used to manage prostate cancer; however, erectile dysfunction (ED) and cardiovascular disease are common side effects. After cessation of ADT, testosterone (T) levels may take years to recover leaving men prone to cardiovascular disease and long‐term ED. The internal pudendal arteries (IPA) supply blood to the penis and vascular injury to these vessels can lead to ED. This study will determine if castration impairs vascular function in systemic (aorta and mesenteric arteries) and penile vasculature (IPA), and if T can restore erectile and vascular physiology. We hypothesize the IPA will develop impaired relaxation prior to systemic arteries following castration, and T therapy will recover both vascular and erectile function.MethodsMale Sprague Dawley rats (12 wks) were divided into 3 groups (n=8/grp): control (CON), CAST (6 wks castration), and CAST+T (6 wks castration+T (3mg/kg) in last 2 wks). Erections were assessed via cavernous nerve stimulation and measurement of intracavernosal to mean arterial pressures (ICP/MAP). Aortas, mesenteric arteries, and IPA were excised, cut (2 mm) and mounted into tissue baths. Contractility to high concentration potassium solution (KCl), electrical field stimulation (EFS), and phenylephrine (PE) were measured. Endothelial dependent acetylcholine (ACh) relaxation and endothelial independent relaxation to DEA NONOate were assessed. Non‐adrenergic non‐cholinergic (NANC) relaxation was evaluated by EFS in the presence of guanethidine and atropine. Serum testosterone levels were measured via ELISA. Androgen receptor protein expression was measured in IPA.ResultsCAST markedly impaired erectile function (ICP/MAP; CON: 0.83±0.03, CAST: 0.31±0.02; p<0.05) and decreased IPA ACh relaxation (CON: 50.4±3.5%, CAST: 31.2±2.3%; p<0.05). DEA NONOate relaxation was unchanged with CAST. CAST diminished IPA NANC relaxation (CON: 40.8±5.1%, CAST: 22.1±3.9%; p<0.05). However, CAST did not impact IPA contraction. Interestingly, CAST did not change vascular contraction or relaxation in aortas or mesenteric arteries. IPA androgen receptor expression decreased after CAST (CON: 1±0.17, CAST: 0.39±0.03; p<0.05). T supplementation restored serum T (T (ng/ml); CON: 4.2±0.8, CAST: 0.1±0.02, CAST+T: 32.9±3.2: p<0.05), erectile function (ICP/MAP; CAST+T 0.74±0.01 p<0.05) and improved ACh relaxation in IPA to values greater than CON (CAST+T: 63.9±2.9%; p<0.05). Similarly, CAST+T enhanced IPA NANC relaxation to levels greater than CON (CAST+T: 69.2±1.4%; p<0.05). T supplementation did not impact IPA contractions and had no effect on aortic or mesenteric vasoreactivity.ConclusionsCastration induces severe IPA vascular damage, yet systemic vasculature is unaffected. T therapy dramatically enhances IPA endothelial dependent and NANC‐mediated relaxation. Though T therapy safety in cancer survivors remains controversial, a growing body of literature is finding no correlation between T therapy and cancer progression. In certain populations of prostate cancer survivors, T replacement is warranted.Support or Funding InformationBSOM start‐up grant, SMSNA, AUAThis abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

ATP‐sensitive K+ Channel Inhibition via Glibenclamide Impairs Maximal Aerobic Capacity and Critical Speed of Healthy Rats without Compromising Cardiac Function

Glibenclamide (GLI) enhances insulin release for Type II diabetes patients by inhibiting pancreatic ATP‐sensitive K+ (KATP) channels. However, systemic KATP channel inhibition decreases maximal aerobic capacity (V̇O2max) during treadmill running in rats; where V̇O2max is determined by cardiac output (Q̇) and O2 utilization within contracting skeletal muscle (arterial‐venous O2 difference). In vitro studies demonstrate that KATP channels enhance isolated cardiomyocyte and vascular smooth muscle relaxation. In vivo studies show that GLI induces vasoconstriction, decreases blood flow, and impairs O2 delivery‐utilization matching in contracting skeletal muscle. Whether high‐intensity exercise tolerance (i.e. the speed‐duration relationship with critical speed, CS, and distance covered above CS, D′) is dependent upon cardiac and/or peripheral (i.e., vascular) KATP channel function in vivo is unknown.PURPOSETo determine the role of KATP channels in establishing cardiac function (see below), maximal aerobic capacity (V̇O2max) and sustaining submaximal exercise tolerance (CS). We hypothesized that acute KATP channel inhibition via GLI would decrease V̇O2max and CS, but not decrease cardiac function.METHODSTen adult female Sprague‐Dawley rats were assessed during the proestrus cycle in randomized order with and without GLI. GLI (10 mg kg−1 in DMSO i.p.) was administered 30–60 min prior to resting Doppler echocardiography and exercise tests. Doppler echocardiography measured Q̇, heart rate (HR), stroke volume (SV), ejection fraction (EF), fractional shortening (FS) and rates of left ventricular contraction and relaxation. V̇O2max and CS tests utilized a motorized treadmill at a 5% incline. V̇O2max was measured via plexiglass metabolic chamber connected to an O2 analyzer with increasing treadmill speed (5–10 m min−1) every minute until V̇O2 plateaued with increases in treadmill speed. Multiple (i.e., ≥5) constant speed runs to exhaustion were used to resolve CS and D′.RESULTSAt rest GLI did not alter SV, EF, FS, nor rates of left ventricular contraction and relaxation compared to control (p>0.05 for all). However, GLI did reduce HR (321 ± 8 vs 300 ± 7 b min−1) and V̇ (217 ± 23 vs 192 ± 13 mL min−1; p<0.05 for both). Despite HR and Q̇ reductions at rest, neither were significantly correlated with decreases in V̇O2max or CS (p>0.05 for both). GLI significantly reduced V̇O2max (69 ± 2 vs 72 ± 1 mL O2 kg−1 min−1), CS (32 ± 1 vs 36 ± 1 m min−1; n=8), and D′ (100 ± 6 vs 86 ± 8 m; n=7) compared to control (all p<0.05).CONCLUSIONSConsistent with our hypothesis, KATP channels are vital in establishing maximal aerobic capacity (GLI decreased V̇O2max ~4%) and exercise tolerance (GLI decreased CS ~12%). Therefore, we speculate that HR regulation via KATP channel activity is overridden during exercise. These data support that vascular KATP channel function is important for matching O2 delivery to O2 requirements within skeletal muscles and this plays a pivotal role in determining exercise tolerance. Thus GLI treatment may potentiate the exercise intolerance of diabetic and other cardiovascular disease patients.Support or Funding InformationHL108328This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Sodium nitroprusside encapsulated in Silica nanoparticles as a strategy for safer nitric oxide delivery

Sodium nitroprusside (SNP) has been used as a nitric oxide (NO) donor in medicine for several decades, even before any knowledge of the biological role of NO. This medicine is used in cases of emergency of hypertension due to its fast release of NO causing fast drop in blood pressure. Unfortunately, this compound also releases cyanide (CN-) as a side product. Nanoencapsulated nitroprusside was synthesized by using silica nanoparticles functionalized with primary amine (MPSi-NH2) (the final compound was named APTES). The compound was bioassayed using vascular tissues from Wistar rats, albine guinea-pigs and New Zealand rabbits. All the protocols were approved by the Ethic committee of the Ceará State University under the protocol # 2897836/15. The thoracic aorta, pulmonary artery or corpora cavernosa were carefully removed and mounted in a 5 mL organ bath containing Krebs–Henseleit solution at 37°C, gassed with 95% O2: 5% CO2, attached to isometric transducers coupled to a powerlab data acquisition system. After equilibration, the tissues were precontracted with phenylephrine (PE, 10−6–10−5 M) and after a plateau was achieved increasing concentrations of SNP or APTES (10−10 to 10−5 M) were cumulatively added in intervals of 5 minutes. The relaxant effect was measured as percentage of the maximal contraction induced by phenylephrine. In addition, we also measured the level of phosphorylation of Ser239 of VASP, a useful marker for monitoring cGMP-dependent protein kinase activation and signaling. SNP APTEs relaxed rat aortic rings with similar potency than non-modified SNP with a concentration necessary to induce 50% vasodilation (CE50) of 26 nM [95% confidence interval=20–33 nM] compared to 14 nM [95% confidence interval=7,9–21 nM] for non-modified SNP. However, the maximal vasodilation induced by SNP APTEs (177.4±7.1%) was higher than the maximal relaxation induced by nonmodified SNP (143.8±9.6%). When comparing different vascular beds, i.e., guinea pig thoracic aorta vs. pulmonary arteries vs. rabbit corpora cavernosa, SNP APTEs was shown to have similar potency and efficacy (119.6±5.7% and 130nM vs. 108±6.3% and 275nM vs. 116±4.5% and 98 nM, respectively). Interestingly, SNP increase VASP phosphorylation by 3.8-fold and APTES increased by 4.13-fold (P<0.05 vs. control). Our synthetic strategy to make functionalized silica nanoparticles showed quite efficient to incorporate sodium nitroprusside, where we succeeded to have up to 320 umol g−1. Remarkably, we noticed a minor release of cyanide or even NP, despite full effect of NO was observed during a series of biological assays. Indeed, EC50 and maximal response for relaxation in aorta, pulmonary arteries and Rabbit corpora cavernosa were quite similar. Support or Funding Information CAPES, CNPq and FUNCAP Vasorelaxant Effect of Sodium nitroprusside encapsulated in Silica nanoparticles Original tracings depicting the strong and potent vascular relaxant effect of SNP APTES (panel A). Effect of SNP APTES on the phenylephrine precontracted aortic rings compared with non-modified sodium nitroprusside (Panel B). Panel C depicts the Effect of 1 μM SNP or SNP APTES compared with the addition of vehicle only in the level of phosphorylation of the residues Serine 239 of the protein Vasodilator-stimulated phosphoprotein (VASP) a substrate of activated cGMP-activated protein kinase (PKG) *p<0.05 vs SNP at the same concentration This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

AdipoRon, adiponectin receptor agonist, improves vascular function in the mesenteric arteries of type 2 diabetic mice

BackgroundAdiponectin is one of the most abundant adipokines secreted from adipose tissue. An orally active synthetic adiponectin receptor agonist, adipoRon has been suggested to ameliorate insulin resistance, myocardial apoptosis, and pancreatic tumor. It has been reported that adiponectin directly induces vascular relaxation however, the chronic effect of adipoRon in the vascular dysfunction in type 2 diabetes has not been studied yet. Thus, in this study, we examined whether adipoRon improves vascular function in type 2 diabetes and what mechanism is involved.MethodsTen to 12‐week old male type 2 diabetic (db−/db−) mice were treated with adiponectin receptor agonist (adipoRon, 10 mg/kg/everyday, by oral gavage) for 2 weeks. Isolated mesenteric arteries were mounted in the arteriography and arterial diameter was measured. And western blot analysis was assessed.ResultsPressure‐induced myogenic response was significantly increased, whereas endothelium‐dependent relaxation was significantly reduced in the mesenteric arteries from type 2 diabetic mice. Interestingly, treatment of adipoRon normalized potentiated myogenic response. However, endothelium‐dependent relaxation was not affected by treatment of adipoRon. The expression levels of adiponectin receptor 1, 2 and adaptor protein phosphotyrosine interacting with PH domain and leucine zipper 1, 2 (APPL 1, 2) were increased in the mesenteric arteries from Type 2 diabetic mice and treatment of adipoRon did not affect them. Interestingly, adipoRon treatment increased the phosphorylation level of AMP‐activated protein kinase (AMPK) and decreased phosphorylation of myosin phosphatase target subunit 1 (MYPT1) in the type 2 diabetic mice while there was no change in the level of eNOS phosphorylation.ConclusionThe treatment of adipoRon improves vascular function in the mesenteric arteries from type 2 diabetic mice through endothelium‐independent mechanism. We suggest that myosin light chain phosphatase (MLCP) activation through reduced phosphorylation of MYPT1 might be the dominant mechanism in the adipoRon‐induced vascular effect.Support or Funding InformationThis work was supported by the Basic Science Research Program of the National Research Foundation (NRF) of Korea funded by the Ministry of Education, Science and Technology (NRF‐2018R1D1A1B07041820).This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Mas1 Receptor Knockout Rats as a Novel Model to Study Vascular Function in Salt‐Sensitive Hypertension

Hypertension is a critical risk factor for coronary and peripheral arterial disease, stroke and heart failure. It has been shown that activation of the Mas1 receptor (Mas1R) for angiotensin (1–7) [Ang (1–7)] has potent beneficial effects to offset the impaired vascular relaxation caused by increased oxidative stress, reduced nitric oxide (NO) availability, and down regulation of antioxidant defense mechanisms that occur during salt‐induced angiotensin II (ANG II) suppression. The recent development of a Mas 1R‐knockout rat (Mas 1R KO) in the Dahl salt sensitive (SS) genetic background is a crucial development, because this genetic model will allow for more detailed studies of Mas 1R function at the whole animal, molecular, and cellular level in salt‐sensitive rats. The aim of this study was to provide initial phenotyping of vascular responses in middle cerebral arteries (MCA) of wild type (WT) and Mas1R KO rats. Mas 1R KO− and WT rats were maintained on a low salt (LS; 0.4 % NaCl) diet to evaluate endothelial function without elevated blood pressure. MCA were isolated to assess vascular function ex vivo using the endothelium‐dependent vasodilator acetylcholine (ACh), exogenously added Ang (1–7), the non‐peptide Ang‐(1–7) mimic AVE 0991, and the NO donor sodium nitroprusside (SNP). ACh induced a concentration‐dependent dilation in isolated arteries from Mas 1R WT rats but not in Mas 1R KO arteries (p&lt;0.05). Arteries stimulated with Ang (1–7) and AVE 0991 exhibited a concentration‐dependent vasodilation in Mas 1R WT but not in Mas 1R KO rats. However, endothelium‐independent vessel relaxation by SNP was unaltered in KO vessels compared to WT controls. The vasodilator effect of Ang (1–7) and AVE 0991 were eliminated by the Ang (1–7) receptor antagonist A779 and by inhibition of NO synthase with L‐NAME. Dilation of MCA in response to the AT2 receptor agonist cGP42112A was unaltered in Mas 1R KO rats. Our results indicate that Mas 1R activation has an important role in maintaining endothelial cell function in WT rats. Use of the novel Mas 1R KO genetic rat model provides a powerful tool to investigate Mas 1R functions at the molecular and cellular level. Further study of the mechanisms by which Mas1 receptors induce protective effects in the endothelium will be important since Mas 1R receptor analogs such as AVE 0991 and Ang (1–7) may offer a potentially novel therapeutic strategy for the management of hypertension.Support or Funding InformationThis work was supported in part by grants: R21‐OD024781‐01, R01‐HL128242 and T32‐GM089586 from the National Institutes of Health, Bethesda, MDThis abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Sirtuin 1 (SIRT1) Content and Angiotensin II Receptor Type I (AT1) Expression in Human Umbilical Vein Endothelial Cells (HUVECs) in Response to Resveratrol: Pregnancy Induced Hypertension (PIH) vs Normotensive pregnancy (NTP)

Oxidative stress and endothelial dysfunction within uteroplacental unit are inherent components initiating PIH. The normal decrease of vascular angiotensin II sensitivity is lost in PIH. The polyphenol stilbene compound resveratrol (3,5,4′‐trihydroxy‐trans‐stilbene) boosts endothelium‐dependent vasorelaxation enhancing endothelial nitrix oxide synthase (eNOS) production. In fact, resveratrol acts as a secondary regulator influencing more complex biological response that includes the sirtuin system. SIRT1 is highly expressed in endothelial cells and activated/modulated by resveratrol has been shown to target eNOS for deacetylation resulting in enhanced NO‐dependent vascular relaxation. Moreover, anti‐hypertensive effects of resveratrol may be a consequence of the angiotensin II receptor type I (AT1) modulation by SIRT1. The aim of the study was to examine comparatively the effect of resveratrol on SIRT level in HUVECs isolated after PIH vs. NTP. In addition, AT1 expressions were compared. HUVECs were isolated immediately after cesarean section delivery from PIH (N = 24) and NTP (N = 24) patients (group I and group II, respectively). The umbilical cord segments was subjected to collagenase digestion and the HUVECs were cultured in vitro under normoxia. After achieving the confluence, HUVECs were exposed to resveratrol (50, 100 and 250 μM) administered into the culture media in the respective subgroups A, B and C during the 7 day period. SIRT1 levels were determined in HUVECs culture lysates obtained in the days 1,3 and 7, using ELISA. After termination of the cultures AT1 were immunostained in formalin fixed and paraffin embedded sections. To estimate the mean expression of AT1, quantitative immunohostochemistry was applied. The initial AT1 expression was estimated in the respective controls. After 3 and 7 days of the culture, in both groups the mean SIRT1 levels were significantly (p &lt; 0.05) higher in the subgroups B and C, compared to the initial levels at day 1. However, the mean SIRT1 levels in group I were 4.6‐fold and 2.4‐fold lower (the subgroups B and C, respectively) than observed in normotensive group II. The initial expression of AT1 was significantly higher in group I and amounted to 170% of the average value in the normotensive group II. The mean expression of AT1 was significantly (p &lt; 0.05) downregulated in group I (the subgroups B and C), while in group II was nearly unaffected. These results may suggest potential beneficial effect of SIRT1 activation by resveratrol with regard to antihypertensive therapy in PIH.Support or Funding InformationWUM grant: 2M2‐W1‐17This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Redox Pioneer: Professor Hideo Kimura.

Dr. Hideo Kimura is recognized as a redox pioneer because he has published an article in the field of antioxidant and redox biology that has been cited >1000 times, and 29 articles that have been cited >100 times. Since the first description of hydrogen sulfide (H2S) as a toxic gas 300 years ago, most studies have been devoted to its toxicity. In 1996, Dr. Kimura demonstrated a physiological role of H2S as a mediator of cognitive function and cystathionine β-synthase as an H2S-producing enzyme. In the following year, he showed H2S as a vascular smooth muscle relaxant in synergy with nitric oxide and its production by cystathionine γ-lyase in vasculature. Subsequently he reported the cytoprotective effect of H2S on neurons against oxidative stress. Since then, studies on H2S have unveiled numerous physiological roles such as the regulation of inflammation, cell growth, oxygen sensing, and senescence. He also discovered polysulfides (H2Sn), which have a higher number of sulfur atoms than H2S and are one of the active forms of H2S, as potent signaling molecules produced by 3-mercaptopyruvate sulfurtransferase. H2Sn regulate ion channels and transcription factors to upregulate antioxidant genes, tumor suppressors, and protein kinases to, in turn, regulate blood pressure. These findings led to the re-evaluation of other persulfurated molecules such as cysteine persulfide and glutathione persulfide. Dr. Kimura is a pioneer of studies on H2S and H2Sn as signaling molecules.It is fortunate to come across a secret of nature and pick it up. —Prof. Hideo Kimura

The Role of Epoxyeicosatrienoic Acids in Diabetes Mellitus-Induced Impaired Vascular Relaxation of Aortic Rings in Ovariectomized Sprague-Dawley Rats.

Aim The present study was aimed at determining if type 1 diabetes mellitus (DM) affects vascular function and at elucidating the mechanisms mediating vasorelaxation in both nonovariectomized and ovariectomized Sprague-Dawley (SD) rats. Materials and Methods Eighty female SD rats were divided into four groups: nonovariectomized healthy (non-OVX-CTR) and diabetic (non-OVX-DM) rats and ovariectomized healthy (OVX-CTR) and diabetic (OVX-DM) rats. Bilateral ovariectomy was performed at the age of 5 weeks, and type 1 DM was induced by streptozotocin at the age of 6 weeks. At the age of 12 weeks, acetylcholine-induced relaxation (AChIR) was assessed in aortic rings in the absence/presence of L-NAME, Indomethacin, and MS-PPOH. Aortic tissue mRNA expression of eNOS, iNOS, COX-1, COX-2, thromboxane synthase 1 (TBXAS1), CYP4A1, CYP4A3, and CYP2J3, as well as plasma oxidative stress, was measured. Results AChIR did not differ in non-OVX-DM rats compared to non-OVX-CTR ones. AChIR was significantly reduced in the OVX-DM group compared to the OVX-CTR group. MS-PPOH did not reduce AChIR in OVX-DM rats as it did in OVX-CTR ones. CYP4a3 mRNA expression in OVX-DM rats was significantly lower compared to that in the OVX-CTR group. Conclusions Female sex hormones may protect vasorelaxation in type 1 diabetic rats. Type 1 diabetes impairs vasorelaxation in response to ACh in ovariectomized rats (but not in nonovariectomized rats) by affecting vasorelaxation pathways mediated by EETs.

Whey hydrolysate-based ingredient with dual functionality: From production to consumer's evaluation.

The aim of the present study concerns the development, characterization and sensory evaluation of a dual-functional whey hydrolysate. Four concentrations of commercial pepsin (0.48%, 0.95%, 1.43%, 1.91% w/w) were evaluated. The hydrolyses curves and the Reversed-Phase High Performance Liquid Chromatography analyses showed a direct relationship between enzyme concentration and degree of hydrolysis. Through mass spectrometry 21 peptides were identified and 5 of them have never been described in the literature before. The hydrolysate produced (PC3) induced a vascular relaxation of 65.02% in phenylephrine-contracted rat aortic rings. PC3 powder presented a homogeneous aspect with a mean particle size of 86.39 μm, high water solubility (>92%) in a wide pH range (1-12) and an increase of 33% in oil absorption capacity, when compared to the unhydrolyzed product. Sensory analysis showed a high acceptance (7.6 in a 9-point hedonic scale) of the hydrolysate among 100 consumers. The results brought the possibility of developing a whey hydrolysate with high vasorelaxant activity, great technological properties and sensory appeal, as an interesting dual-functional ingredient to be incorporated into food products.

Antihypertensive effects of the Vitis vinifera grape skin (ACH09) extract consumption elicited by functional improvement of P1 (A1) and P2 (P2X1) purinergic receptors in diabetic and hypertensive rats

Cross-talk between hypertension and diabetes associated with red wine consumption triggering cardioprotective and antihypertensive effects have been well established; nevertheless, the role of isolated total polyphenols from extracts of Vitis vinifera grape skin (ACH09) and the possible correlation with cardiovascular functionality mediated by purinergic receptors remain poorly understood. We evaluate how ACH09 extract causes an antihypertensive effect by modulation of purinergic receptors. Male Wistar Kyoto (WKY), streptozotocin-induced diabetic WKY and SHR (spontaneously hypertensive) rats were subjected to intake of ACH09 extract 200 mg/kg/v.o. for 21 days. Systolic blood pressure (SBP), blood glucose levels were measured; cardiovascular functionality of sympathetic neurotransmission, represented by purinergic (P1-A1/P2-P2X1) and adrenergic receptors (α1/β1), was evaluated by isometric contraction. Compared to their respective controls, we found that ACH09 extract exposure decreased SBP in both groups, primarily by functional improvement of the following receptors: (1) P1 purinergic (A1), augmenting vascular endothelium-dependent relaxation; (2) P2 purinergic (P2X1), decreasing vasoconstriction; and (3) P1 purinergic (A1), increasing atrial negative inotropism. In conclusion, these findings suggest that dietary supplementation with polyphenolic-rich foods (lentil, citrus fruit and/or red wine) may inspire new pharmacologic and dietetic therapeutic approaches for treatment of hypertension and diabetes.

Acute changes in morphology and renal vascular relaxation function after renal denervation using temperature-controlled radiofrequency catheter

BackgroundResistant hypertension and renal sympathetic hyperactivity are closely linked, and catheter-based renal denervation (RDN) is regarded as a new treatment strategy. However, the acute changes in vascular morphology and relaxation function have yet to be evaluated, and these may be important for the efficacy and safety of the procedure. In this study, we explored these questions by conventional temperature-controlled cardiac radiofrequency catheter-based RDN in a pig model.MethodsSix mini-pigs were randomly divided into the renal denervation (RDN) group (n = 3) and the Sham-RDN group (n = 3). Animals in the RDN group underwent unilateral radiofrequency ablation, and those in the Sham-RDN group underwent the same procedure except for the ablation. The pigs were examined by angiography pre- and post-RDN and were euthanized immediately thereafter. Renal arteries were processed for histological and molecular biology analyses as well as for in vitro vascular tension testing.ResultsCompared with the Sham-RDN group, the RDN caused vascular intima and media injury, renal nerve vacuolization, mild collagen fiber hyperplasia and elastic fiber cleavage (all p < 0.05). The RDN group also significantly exhibited nitric oxide synthase pathway inhibition and decreased endothelium-independent vascular relaxation function Compared to the Sham-RDN group (all p < 0.05).ConclusionsIn this porcine model, renal artery denervation led to vascular wall injury and endothelial dysfunction in the acute phase, which negatively affected vascular relaxation function. Thus, this process may be detrimental to the prognosis and progress of hypertension patients.

High-Fat Diet-Induced Obesity Model Does Not Promote Endothelial Dysfunction via Increasing Leptin/Akt/eNOS Signaling.

Experimental studies show that the unsaturated high-fat diet-induced obesity promotes vascular alterations characterized by improving the endothelial L-arginine/Nitric Oxide (NO) pathway. Leptin seems to be involved in this process, promoting vasodilation via increasing NO bioavailability. The aim of this study was to test the hypothesis that unsaturated high-fat diet-induced obesity does not generate endothelial dysfunction via increasing the vascular leptin/Akt/eNOS signaling. Thirty-day-old male Wistar rats were randomized into two groups: control (C) and obese (Ob). Group C was fed a standard diet, while group Ob was fed an unsaturated high-fat diet for 27 weeks. Adiposity, hormonal and biochemical parameters, and systolic blood pressure were observed. Concentration response curves were performed for leptin or acetylcholine in the presence or absence of Akt and NOS inhibitor. Our results showed that an unsaturated high-fat diet promoted a greater feed efficiency (FE), elevation of body weight and body fat (BF), and an adiposity index, characterizing a model of obesity. However, comorbidities frequently associated with experimental obesity were not visualized, such as glucose intolerance, dyslipidemia and hypertension. The evaluation of the endothelium-dependent relaxation with acetylcholine showed no differences between the C and Ob rats. After NOS inhibition, the response was completely abolished in the Ob group, but not in the C group. Furthermore, Akt inhibition completely blunted vascular relaxation in the C group, but not in the Ob group, which was more sensitive to leptin-induced vascular relaxation. L-NAME incubation abolished the relaxation in both groups at the same level. Although Akt inhibitor pre-incubation reduced the leptin response, group C was more sensitive to its effect. In conclusion, the high-unsaturated fat diet-induced obesity improved the vascular reactivity to leptin and does not generate endothelial dysfunction, possibly by the increase in the vascular sensitivity to leptin and increasing NO bioavailability. Moreover, our results suggest that the increase in NO production occurs through the increase in NOS activation by leptin and is partially mediated by the Akt pathway.

Angiotensin-(1-7) induced vascular relaxation in spontaneously hypertensive rats

Enhanced vasoconstriction and decreased vasodilatation due to endothelial dysfunction contribute to the progression of hypertension. Angiotensin (Ang)-(1-7) plays important roles in regulating the cardiovascular activity. The current study aimed to investigate the roles of Ang-(1-7) in modulating blood pressure, vascular tension and its signal pathway in spontaneously hypertensive rats (SHR). The effects of intravenous injection of drugs were determined in rats with anesthesia in vivo. Mesenteric artery (MA), coronary artery (CA) and pulmonary artery (PA) were isolated from rats and isometric tension measurements in arteries were performed. Compared with Wistar-Kyoto rats (WKY), the high K+ induced vasoconstriction was enhanced and acetylcholine-induced vasodilatation were attenuated in the MA, CA and PA in SHR. Intravenous injection of Ang-(1-7) decreased, while A-779 increased mean arterial pressure and abolished the hypotensive effect of Ang-(1-7) in SHR. Ang-(1-7) caused dose-dependent relaxation in MA, CA and PA in SHR, which was inhibited by pretreatment with Mas receptor antagonist A-779, nitric oxide (NO) synthase inhibitor l-NAME, guanylate cyclase inhibitor ODQ and protein kinase G (PKG) inhibitor DT-2. The Mas receptor expression, NO, cGMP and PKG levels of the three above arteries of SHR were lower than that of WKY. Ang-(1-7) increased the NO, cGMP and PKG levels in arteries from SHR, which was blocked by A-779. Activation of the Mas receptor by Ang-(1-7) relaxes the MA, CA, and PA through the NO-cGMP-PKG pathway, which contributes to the decrease of arterial pressure in SHR.

Signaling by hydrogen sulfide (H2S) and polysulfides (H2Sn) in the central nervous system

Hydrogen sulfide (H2S) is a signaling molecule used to modify neuronal transmission, regulate vascular tone, protect tissues from oxidative stress, sense oxygen, and generate ATP. Hydrogen polysulfides (H2Sn) have recently been identified as signaling molecules that mediate the activation of ion channels, regulation of tumor growth, and the transcriptional regulation of oxidative stress; some of which were previously ascribed to H2S. Cystathionine β-synthetase (CBS), cystathionine γ-lyase (CSE), and 3-mercaptopyruvate sulfurtransferase (3MST) are known as H2S-producing enzymes. 3MST also produces H2Sn and other persulfurated molecules such as cysteine persulfide, glutathione persulfide, and persulfurated proteins. The chemical interaction of H2S and nitric oxide (NO) also produces H2Sn, which may be the mechanism underlying the synergistic effect of H2S and NO that was initially reported on vascular relaxation. H2Sn and other persulfurated molecules elicit their effect via S-sulfuration (S-sulfhydration) of specific cysteine residues of the target proteins. This review article focuses on the production and roles of H2Sn as well as H2S in the central nervous system.

Class I histone deacetylase inhibitor MS-275 attenuates vasoconstriction and inflammation in angiotensin II-induced hypertension

ObjectiveNon-selective histone deacetylase (HDAC) inhibitors are known to improve hypertension. Here, we investigated the therapeutic effect and regulatory mechanism of the class I HDAC selective inhibitors, MS-275 and RGFP966, in angiotensin (Ang) II-induced hypertensive mice.Methods and resultsMS-275 inhibited the activity of HDAC1, HDAC2, and HDAC3, while RGFP966 weakly inhibited that of HDAC3 in a cell-free system. MS-275 and RGFP966 treatment reduced systolic blood pressure and thickness of the aorta wall in Ang II-induced hypertensive mice. MS-275 treatment reduced aorta collagen deposition, as determined by Masson’s trichrome staining. MS-275 decreased the components of the renin angiotensin system and increased vascular relaxation of rat aortic rings via the nitric oxide (NO) pathway. NO levels reduced by Ang II were restored by MS-275 treatment in vascular smooth muscle cells (VSMCs). However, MS-275 dose (3 mg·kg-1·day-1) was not enough to induce NO production in vivo. In addition, MS-275 did not prevent endothelial nitric oxide synthase (eNOS) uncoupling in the aorta of Ang II-induced mice. Treatment with MS-275 failed to inhibit Ang II-induced expression of NADPH oxidase (Nox)1, Nox2, and p47phox. MS-275 treatment reduced proinflammatory cytokines such as tumor necrosis factor (TNF)-α, interleukin (IL)-1β, and monocyte chemoattractant protein (MCP)-1, as well as adhesion molecules. Histological analysis showed that Ang II-induced macrophage infiltration was reduced by MS-275 and RGFP966 administration.ConclusionsOur results indicate that class I HDAC selective inhibitors may be good therapeutic agents for the treatment of hypertension through the regulation of vascular remodeling and vasoconstriction, as well as inflammation.

The role of NO-cGMP pathway inhibition in vascular endothelial-dependent smooth muscle relaxation disorder of AT1-AA positive rats: protective effects of adiponectin

Angiotensin II type 1 receptor autoantibodies (AT1-AA) cause endothelial-dependent smooth muscle relaxation disorder. It is well understood that impairment of the NO–cGMP signaling pathway is one of the mechanisms of endothelial-dependent smooth muscle relaxation disorder. However, it is still unclear whether AT1-AA induces endothelial-dependent smooth muscle relaxation disorder via the impairment of the NO–cGMP signaling pathway. In addition, adiponectin exerts vascular endothelial protection through the NO–cGMP signaling pathway. Therefore, the purpose of this investigation was to assess the mechanism of vascular endothelial-dependent smooth muscle relaxation disorder induced by AT1-AA and the role of adiponectin in attenuating this dysregulation.Serum endothelin-1 (ET-1), adiponectin and AT1-AA were detected by enzyme-linked immunosorbent assay. In preeclamptic patients, there was an increased level of AT1-AA, which was positively correlated with ET-1 and negatively correlated with adiponectin, as elevated levels of ET-1 suggested endothelial injury. AT1-AA-positive animal models were actively immunized with the second extracellular loop of the angiotensin II type 1 receptor (AT1R-ECII) for eight weeks. In thoracic aortas of AT1-AA positive rats, ET-1 was elevated, endothelium-dependent vasodilation was decreased. Paradoxically, as the upstream element of the NO-cGMP signaling pathway, NO production was not decreased but increased, and the ratio of p-VASP/VASP, an established biochemical endpoint of NO-cGMP signaling pathway, was reduced. Moreover, the levels of nitrotyrosine and gp91phox which indicate the presence of peroxynitrite (ONOO-) and superoxide anion (O2·−) were increased. Pretreatment with the ONOO- scavenger FeTMPyP or O2·−scavenger Tempol normalized vasorelaxation. Key enzymes responsible for NO synthesis were also assessed. iNOS protein expression was increased, but p-eNOS(Ser1177)/eNOS was reduced. Preincubation with the iNOS inhibitor 1400 W or eNOS agonist nebivolol restored vasorelaxation. Further experiments showed levels of p-AMPKα (Thr172)/AMPKα, which controls iNOS expression and eNOS activity, to have been reduced. Furthermore, levels of the upstream component of AMPK, adiponectin, was reduced in sera of AT1-AA positive rats and supplementation of adiponectin significantly decreased ET-1 contents, improved endothelial-dependent vasodilation, reduced NO production, elevated p-VASP/VASP, inhibited protein expression of nitrotyrosine and gp91phox, reduced iNOS overexpression, and increased eNOS phosphorylation at Ser1177 in the thoracic aorta of AT1-AA positive rats.These results established that impairment NO–cGMP pathway may aggravate the endothelial-dependent smooth muscle relaxation disorder in AT1-AA positive rats and adiponectin improved endothelial-dependent smooth muscle relaxation disorder by enhancing NO–cGMP pathway. This discovery may shed a novel light on clinical treatment of vascular diseases associated with AT1-AA.

Pharmacological Characterization of a Novel 5-Hydroxybenzothiazolone-Derived β 2-Adrenoceptor Agonist with Functional Selectivity for Anabolic Effects on Skeletal Muscle Resulting in a Wider Cardiovascular Safety Window in Preclinical Studies.

The anabolic effects of β 2-adrenoceptor (β 2-AR) agonists on skeletal muscle have been demonstrated in various species. However, the clinical use of β 2-AR agonists for skeletal muscle wasting conditions has been limited by their undesired cardiovascular effects. Here, we describe the preclinical pharmacological profile of a novel 5-hydroxybenzothiazolone (5-HOB) derived β 2-AR agonist in comparison with formoterol as a representative β 2-AR agonist that have been well characterized. In vitro, 5-HOB has nanomolar affinity for the human β 2-AR and selectivity over the β 1-AR and β 3-AR. 5-HOB also shows potent agonistic activity at the β 2-AR in primary skeletal muscle myotubes and induces hypertrophy of skeletal muscle myotubes. Compared with formoterol, 5-HOB demonstrates comparable full-agonist activity on cAMP production in skeletal muscle cells and skeletal muscle tissue-derived membranes. In contrast, a greatly reduced intrinsic activity was determined in cardiomyocytes and cell membranes prepared from the rat heart. In addition, 5-HOB shows weak effects on chronotropy, inotropy, and vascular relaxation compared with formoterol. In vivo, 5-HOB significantly increases hind limb muscle weight in rats with attenuated effects on heart weight and ejection fraction, unlike formoterol. Furthermore, changes in cardiovascular parameters after bolus subcutaneous treatment in rats and rhesus monkeys are significantly lower with 5-HOB compared with formoterol. In conclusion, the pharmacological profile of 5-HOB indicates superior tissue selectivity compared with the conventional β 2-AR agonist formoterol in preclinical studies and supports the notion that such tissue-selective agonists should be investigated for the safe treatment of muscle-wasting conditions without cardiovascular limiting effects.

Randomized study of the effects of cocoa-rich chocolate on the ventricle–arterial coupling and vascular function of young, healthy adults

The aim of this study was to evaluate and explore the benefits of long-term dark chocolate intake in young, healthy adults by measuring cardiovascular function. A randomized study was conducted with 30 healthy participants ages 18 to 27 y. Half of the participants ingested a 20-g dose of lower cocoa chocolate (LCC; ∼55%; 12.6± 1.4 mg equivalent of epicatechin/g) and the others ingested a daily dose of 20 g of higher cocoa chocolate (HCC; ∼90%; 18.2± 2.6 mg equivalent of epicatechin/g). A baseline evaluation was performed before the participants started ingesting the assigned chocolate for a 30-d period, after which a final evaluation was performed. Each evaluation included heart ultrasonography, carotid-femoral pulse wave velocity (PWV) and carotid pulse wave analysis, flow-mediated slowing (FMS), and an analysis of the ventricular-arterial coupling (VAC), which reflects the matching between the aorta and the left ventricle (ratio of arterial elastance to left ventricle elastance). The baseline evaluation presented similar values within normal range in both groups. The positive vascular effects were overall more distinct in the group eating the HCC. No structural modifications on the heart were found after the intervention, notwithstanding cardiac function was improved on certain functional parameters in the HCC group only. A statistically significant improvement was depicted over the brachial and central systolic and pulse pressures in the HCC group, and a trend for improvement in the reflected waves component (Aix) and the FMS was also observed in the HCC, but not in the LCC group. VAC parameters were similar at baseline between groups, but showed a significant improvement in the HCC group after intervention, increasing from 0.674 to 0.719 (P = 0.004), so that the post-intervention VAC was significantly higher in the HCC group than in the LCC group (P < 0.05). In addition, significant variation was observed in both groups regarding arterial and left ventricle elastances, stroke work, and potential energy, with greater mean differences identified in the HCC group. This study demonstrated that regular consumption of HCC has beneficial effects on the cardiovascular system in young, healthy adults, improving vascular function by reducing central brachial artery pressures and promoting vascular relaxation, and thus enhancing the matching of the arterial system with the left ventricle.