Endothelium-Independent Relaxation of Vascular Smooth Muscle Induced by Persimmon-Derived Polyphenol Phytocomplex in Rats

The vasorelaxant effect of polyphenols is well known, and the mortality rate due to coronary artery disease is low in people who consume polyphenol-containing foods. We aimed to elucidate the mechanism by which polyphenols derived from persimmon juice (PJ) and persimmon leaves (PLs) induce vasorelaxation and suppress vasocontraction in the superior mesenteric arteries isolated from male Sprague Dawley rats. Vasocontraction was induced with 1 µM phenylephrine, and polyphenol-induced vasorelaxation was expressed as a percentage of the previous tone induced by phenylephrine. PJ powder (100 mg/L) induced higher levels of vasorelaxation (mean ± standard error of the mean, 88.6% ± 4.4%) than PLs powder (1 g/L; 72.0% ± 10.8%). Nitric oxide pathway inhibitors (NG-nitro-L-arginine methyl ester + carboxy-PTIO) did not affect persimmon-derived polyphenol-induced vasorelaxation, whereas potassium chloride, tetraethylammonium, and potassium-channel inhibitors did. Vasorelaxation was endothelium independent with both extracts. Phenylephrine-induced vasocontraction was suppressed by pretreatment with PJ and PLs powder, even when inositol triphosphate-mediated Ca2+ release and extracellular Ca2+ influx were inhibited. These results suggest that persimmon-derived polyphenol phytocomplex cause vasorelaxation and inhibit vasocontraction through hyperpolarization of smooth muscle cells. Persimmon-derived polyphenols may be able to prevent cardiovascular diseases caused by abnormal contraction of blood vessels.

COX-2/sEH Dual Inhibitor PTUPB Alleviates CCl4-Induced Liver Fibrosis and Portal Hypertension

Background: 4-(5-phenyl-3-{3-[3-(4-trifluoromethylphenyl)-ureido]-propyl}-pyrazol-1-yl) -benzenesulfonamide (PTUPB), a dual cyclooxygenase-2 (COX-2)/soluble epoxide hydrolase (sEH) inhibitor, was found to alleviate renal, pulmonary fibrosis and liver injury. However, few is known about the effect of PTUPB on liver cirrhosis. In this study, we aimed to explore the role of PTUPB in liver cirrhosis and portal hypertension (PHT).Method: Rat liver cirrhosis model was established via subcutaneous injection of carbon tetrachloride (CCl4) for 16 weeks. The experimental group received oral administration of PTUPB (10 mg/kg) for 4 weeks. We subsequently analyzed portal pressure (PP), liver fibrosis, inflammation, angiogenesis, and intra- or extrahepatic vascular remodeling. Additionally, network pharmacology was used to investigate the possible mechanisms of PTUPB in live fibrosis.Results: CCl4 exposure induced liver fibrosis, inflammation, angiogenesis, vascular remodeling and PHT, and PTUPB alleviated these changes. PTUPB decreased PP from 17.50 ± 4.65 to 6.37 ± 1.40 mmHg, reduced collagen deposition and profibrotic factor. PTUPB alleviated the inflammation and bile duct proliferation, as indicated by decrease in serum interleukin-6 (IL-6), liver cytokeratin 19 (CK-19), transaminase, and macrophage infiltration. PTUPB also restored vessel wall thickness of superior mesenteric arteries (SMA) and inhibited intra- or extrahepatic angiogenesis and vascular remodeling via vascular endothelial growth factor (VEGF), von Willebrand factor (vWF), etc. Moreover, PTUPB induced sinusoidal vasodilation by upregulating endothelial nitric oxide synthase (eNOS) and GTP-cyclohydrolase 1 (GCH1). In enrichment analysis, PTUPB engaged in multiple biological functions related to cirrhosis, including blood pressure, tissue remodeling, immunological inflammation, macrophage activation, and fibroblast proliferation. Additionally, PTUPB suppressed hepatic expression of sEH, COX-2, and transforming growth factor-β (TGF-β).Conclusion: 4-(5-phenyl-3-{3-[3-(4-trifluoromethylphenyl)-ureido]-propyl}-pyrazol-1-yl)- benzenesulfonamide ameliorated liver fibrosis and PHT by inhibiting fibrotic deposition, inflammation, angiogenesis, sinusoidal, and SMA remodeling. The molecular mechanism may be mediated via the downregulation of the sEH/COX-2/TGF-β.

Therapeutic Potential of Sunitinib in Ameliorating Endothelial Dysfunction in Type 2 Diabetic Rats

<b><i>Introduction:</i></b> Sunitinib, a multi-targeted tyrosine kinase receptor inhibitor used to treat renal-cell carcinoma and gastrointestinal stromal tumor, was recently shown to have a beneficial effect on metabolism in type 2 diabetes (T2D). Endothelial dysfunction is a key factor behind macro- and microvascular complications in T2D. The effect of sunitinib on endothelial function in T2D remains, however, unclear. We therefore tested the hypothesis that sunitinib ameliorates endothelial dysfunction in T2D. <b><i>Methods:</i></b> Sunitinib (2 mg/kg/day, by gavage) was administered to T2D Goto-Kakizaki (GK) rats for 6 weeks, while water was given to GK and Wistar rats as controls. Hemodynamic, inflammatory, and metabolic parameters as well as endothelial function were measured. <b><i>Results:</i></b> Systolic, mean arterial blood pressures, plasma tumor necrosis factor α levels, kidney weight to body weight (BW) ratio, and glucose levels were higher, while BW was lower in GK rats than in Wistar rats. Six-week treatment with sunitinib in GK rats did not affect these parameters but suppressed the increase in glucose levels. Endothelium-dependent relaxations were reduced in both aortas and mesenteric arteries isolated from GK as compared to Wistar rats, which was markedly reversed in both types of arteries from GK rats treated with sunitinib. <b><i>Conclusions:</i></b> This study demonstrates that sunitinib has a glucose-lowering effect and ameliorates endothelial dysfunction in both conduit and resistance arteries of GK rats.

Food Restriction Augmented Alpha1 Adrenergic Mediated Contraction in Mesenteric Arteries

Food restriction (FR) enhances the sensitivity to cardiopulmonary reflexes and alpha1 adrenoreceptors in the female, despite hypotension. The effect of male FR on cardiopulmonary and systemic vascular function is not well understood. This study examines the effects of FR on cardiopulmonary, isolated mesenteric arterial function and potential underlying mechanisms. We hypothesized that FR decreased eNOS activity in mesenteric arteries. Male Sprague Dawley (SD) rats were randomly divided into three groups: (1) control (n=30), (2) 20 percent of food reduction (FR20, n=30), and (3) 40 percent of food reduction (FR40, n=30) for five weeks. Non-invasive blood pressure was measured twice a week. Pulmonary arterial pressure (PAP) was measured using isolated/perfused lungs in rats. The isolated vascular reactivity was assessed in double-wire myograph. After five weeks, food restricted rats exhibited a lower mean arterial pressure and heart rate, however, only FR40 groups exhibited statistically significant differences. The basal tone of PAP and various vasoconstrictors did not show significant differences in pulmonary circulation between each group. We observed that food restriction were enhanced the sensitivity (EC50) in response to α1-adrenoreceptors (phenylephrine, PhE)-induced vasoconstriction, but not to serotonin, U46619, and high K+ in the mesenteric arteries. FR reduced endothelium-dependent relaxation via decreased function of endothelial nitric oxide synthase (eNOS)-nitric oxide (NO) pathway in the mesenteric arteries. PhE-mediated vasoconstriction in mesenteric arteries was eliminated in the presence of eNOS inhibitor (L-NAME). In addition, incubation with NOX2/4 inhibitors (apocynin, GKT137831, VAS2870) and reactive oxygen species (ROS) scavenger inhibitor (Tiron) were eliminated the differences of PhE-mediated vasoconstriction but not to cyclooxygenase inhibitor (indomethacin) in the mesenteric artery. Augmentation of alpha1 adrenergic mediated contraction via inhibition of eNOS-NO pathway by increased activation of ROS through NOX2/4 in response to FR. Reduced eNOS-NO signaling might be a pathophysiological counterbalance to prevent hypovolemic shock in response to FR.

Cystamine reduces vascular stiffness in Western diet-fed female mice

Consumption of diets high in fat, sugar and salt (Western diet, WD) is associated with accelerated arterial stiffening, a major independent risk factor for cardiovascular disease (CVD). Obese women are more prone to develop arterial stiffening leading to more frequent and severe CVD compared to men. As tissue transglutaminase (TG2) has been implicated in vascular stiffening, our goal herein was to determine the efficacy of cystamine, a non-specific TG2 inhibitor, at reducing vascular stiffness in female mice chronically fed a WD. Three experimental groups of female mice were created. One was fed regular chow diet (CD) for 43 weeks starting at four weeks of age. The second was fed a WD for the same 43 weeks, whereas a third cohort was fed WD, but also received cystamine (216 mg/kg/d) in the drinking water during the last eight weeks on the diet (WD+C). All vascular stiffness parameters assessed, including aortic pulse wave velocity and the incremental modulus of elasticity of isolated femoral and mesenteric arteries, were significantly increased in WD- vs. CD-fed mice, and reduced in WD+C vs. WD-fed mice. These changes coincided with respectively augmented and diminished vascular wall collagen and F-actin content, with no associated effect in blood pressure. In cultured human vascular smooth muscle cells, cystamine reduced TG2 activity, F-actin/G-actin ratio, collagen compaction capacity and cellular stiffness. We conclude that cystamine treatment represents an effective approach to reduce vascular stiffness in female mice in the setting of WD consumption, likely due to its TG2 inhibitory capacity.

Neflamapimod Induces Vasodilation in Resistance Mesenteric Arteries by Inhibiting p38 Mitogen-Activated Protein Kinase Alpha (MAPKα) and Heat-Shock Protein 27 (Hsp27) Phosphorylation

Neflamapimod, a selective inhibitor of p38 MAPKα, is under clinical investigation for its efficacy in Alzheimer’s disease (AD) and dementia with Lewy Bodies (DLB). Here, we investigated if neflamapimod-mediated acute inhibition of p38 MAPKα could induce vasodilation in resistance-size rat mesenteric arteries. Our pressure myography data demonstrated that neflamapimod produced a dose-dependent vasodilation in mesenteric arteries. Our Western blotting data revealed that acute neflamapimod treatment significantly reduced the phosphorylation of p38 MAPKα and its downstream target heat-shock protein 27 (Hsp27) that is involved in cytoskeletal reorganization and smooth muscle contraction. Likewise, non-selective inhibition of p38 MAPK by SB203580 attenuated p38 MAPKα and Hsp27 phosphorylation, and induced vasodilation. Endothelium denudation or pharmacological inhibition of endothelium-derived vasodilators such as nitric oxide (NO) and prostacyclin (PGI2) had no effect on such vasodilation. Neflamapimod-evoked vasorelaxation remained unaltered by the inhibition of smooth muscle cell K+ channels. Altogether, our data for the first time demonstrates that in resistance mesenteric arteries, neflamapimod inhibits p38 MAPKα and phosphorylation of its downstream actin-associated protein Hsp27, leading to vasodilation. This novel finding may be clinically significant and is likely to improve systemic blood pressure and cognitive deficits in AD and DLB patients for which neflamapimod is being investigated.

Aging Affects KV7 Channels and Perivascular Adipose Tissue-Mediated Vascular Tone

Aging is an independent risk factor for hypertension, cardiovascular morbidity, and mortality. However, detailed mechanisms linking aging to cardiovascular disease are unclear. We studied the aging effects on the role of perivascular adipose tissue and downstream vasoconstriction targets, voltage-dependent KV7 channels, and their pharmacological modulators (flupirtine, retigabine, QO58, and QO58-lysine) in a murine model. We assessed vascular function of young and old mesenteric arteries in vitro using wire myography and membrane potential measurements with sharp electrodes. We also performed bulk RNA sequencing and quantitative reverse transcription-polymerase chain reaction tests in mesenteric arteries and perivascular adipose tissue to elucidate molecular underpinnings of age-related phenotypes. Results revealed impaired perivascular adipose tissue-mediated control of vascular tone particularly via KV7.3–5 channels with increased age through metabolic and inflammatory processes and release of perivascular adipose tissue-derived relaxation factors. Moreover, QO58 was identified as novel pharmacological vasodilator to activate XE991-sensitive KCNQ channels in old mesenteric arteries. Our data suggest that targeting inflammation and metabolism in perivascular adipose tissue could represent novel approaches to restore vascular function during aging. Furthermore, KV7.3–5 channels represent a promising target in cardiovascular aging.

Cholesterol-Induced Suppression of Endothelial Kir Channels Is a Driver of Impairment of Arteriolar Flow-Induced Vasodilation in Humans

Dyslipidemia-induced endothelial dysfunction is an important factor in the progression of cardiovascular disease; however, the underlying mechanisms are unclear. Our recent studies demonstrated that flow-induced vasodilation (FIV) is regulated by inwardly rectifying K + channels (Kir2.1) in resistance arteries. Furthermore, we showed that hypercholesterolemia inhibits Kir2.1-dependent vasodilation. In this study, we introduced 2 new mouse models: (1) endothelial-specific deletion of Kir2.1 to demonstrate the role of endothelial Kir2.1 in FIV and (2) cholesterol-insensitive Kir2.1 mutant to determine the Kir2.1 regulation in FIV under hypercholesterolemia. FIV was significantly reduced in endothelial-specific Kir2.1 knock-out mouse mesenteric arteries compared with control groups. In cholesterol-insensitive Kir2.1 mutant mice, Kir2.1 currents were not affected by cyclodextrin and FIV was restored in cells and arteries, respectively, with a hypercholesterolemic background. To extend our observations to humans, 16 healthy subjects were recruited with LDL (low-density lipoprotein)-cholesterol ranging from 51 to 153 mg/dL and FIV was assessed in resistance arteries isolated from gluteal adipose. Resistance arteries from participants with >100 mg/dL LDL (high-LDL) exhibited reduced FIV as compared with those participants with <100 mg/dL LDL (low-LDL). A significant negative correlation was observed between LDL cholesterol and FIV in high-LDL. Expressing dominant-negative Kir2.1 in endothelium blunted FIV in arteries from low-LDL but had no further effect on FIV in arteries from high-LDL. The Kir2.1-dependent vasodilation more negatively correlated to LDL cholesterol in high-LDL. Overexpressing wild-type Kir2.1 in endothelium fully recovered FIV in arteries from participants with high-LDL. Our data suggest that cholesterol-induced suppression of Kir2.1 is a major mechanism underlying endothelial dysfunction in hypercholesterolemia.

Dual Role of Thrombospondin-1 in Flow-Induced Remodeling

(1) Background: Chronic increases in blood flow, as in cardiovascular diseases, induce outward arterial remodeling. Thrombospondin-1 (TSP-1) is known to interact with matrix proteins and immune cell-surface receptors, but its contribution to flow-mediated remodeling in the microcirculation remains unknown. (2) Methods: Mesenteric arteries were ligated in vivo to generate high- (HF) and normal-flow (NF) arteries in wild-type (WT) and TSP-1-deleted mice (TSP-1−/−). After 7 days, arteries were isolated and studied ex vivo. (3) Results: Chronic increases in blood flow induced outward remodeling in WT mice (increasing diameter from 221 ± 10 to 280 ± 10 µm with 75 mmHg intraluminal pressure) without significant effect in TSP-1−/− (296 ± 18 to 303 ± 14 µm), neutropenic or adoptive bone marrow transfer mice. Four days after ligature, pro inflammatory gene expression levels (CD68, Cox2, Gp91phox, p47phox and p22phox) increased in WT HF arteries but not in TSP-1−/− mice. Perivascular neutrophil accumulation at day 4 was significantly lower in TSP-1−/− than in WT mice. (4) Conclusions: TSP-1 origin is important; indeed, circulating TSP-1 participates in vasodilation, whereas both circulating and tissue TSP-1 are involved in arterial wall thickness and diameter expansion.