Alteration of Piezo1 signaling in type 2 diabetic mice: focus on endothelium and BKCa channel.

Piezo1 mechanosensitive ion channel plays a important role in vascular physiology and disease. This study aimed to elucidate the altered signaling elicited by Piezo1 activation in the arteries of type 2 diabetes. Ten- to 12-week-old male C57BL/6 (control) and type 2 diabetic mice (db-/db-) were used. The second-order mesenteric arteries (~ 150 µm) were used for isometric tension experiments. Western blot analysis and immunofluorescence staining were performed to observe protein expression. Piezo1 was significantly decreased in mesenteric arteries of type 2 diabetic mice compared to control mice, as analyzed by western blot and immunofluorescence staining. Piezo1 agonist, Yoda1, concentration-dependently induced relaxation of mesenteric arteries in both groups. Interestingly, the relaxation response was significantly greater in control mice than in db-/db- mice. The removal of endothelium reduced relaxation responses induced by Yoda1, which was greater in control mice than db-/db- mice. Furthermore, the relaxation response was reduced by pre-treatment with various types of K+ channel blockers in endothelium-intact arteries in control mice. In endothelium-denuded arteries, pre-incubation with charybdotoxin, an Ca2+-activated K+ channel (BKCa channel) blocker, significantly attenuated Yoda1-induced relaxation in db-/db- mice, while there was no effect in control mice. Co-immunofluorescence staining showed co-localization of Piezo1 and BKCa channel was more pronounced in db-/db- mice than in control mice. These results indicate that the vascular responses induced by Piezo1 activation are different in the mesenteric resistance arteries in type 2 diabetic mice.

Relaxant Effects of Piperine and Piperlongumine from Piper longum Fruits on Porcine Coronary Artery.

Since ancient times, Piper longum Linn. fruits have been recognized for exhibiting various effects, including the diaphoretic effects linked to enhanced blood flow. Piperine and piperlongumine coexist in Piper longum Linn. fruits, although the cardiovascular effects of both compounds remain elusive. We investigated their action of piperine and piperlongumine in porcine coronary arteries, comparing them to the Ca2+ channel antagonist diltiazem. Piperlongumine, unlike piperine or diltiazem, concentration-dependently inhibited basal contractile tone in endothelium-denuded coronary arteries. All three compounds inhibit tonic contractions induced by high potassium chloride (KCl) concentrations. The order of relaxation potency indexed by the half-maximal effective concentration (EC50) were as follows: diltiazem > piperlongumine > piperine. These effects were not different between endothelium-intact and -denuded preparations. In endothelial-denuded preparations, pretreatment with these compounds not only inhibited KCl-induced tonic contractions attenuated calcium chloride (CaCl2)-induced ones in a Ca2+-free medium. Histamine-induced phasic contractions in a Ca2+-free medium containing intracellular Ca2+ chelator was completely suppressed by selective inositol trisphosphate receptor antagonist and piperlongumine, whereas piperine or diltiazem do not have the same effect. These findings suggest that piperine and piperlongumine similar to diltiazem cause vasorelaxation by inhibiting both KCl- and CaCl2-induced contractions in coronary arteries, possibly through the inhibition of voltage-dependent Ca2+ channels. Piperlongumine inhibits histamine-induced contractions in a Ca2+-free medium, which is associated with the intracellular Ca2+ signaling pathway, suggesting that the relaxant effect of piperlongumine differs from that of piperine.

Effect of Torilis japonica Fruit Extract for Endothelium-Independent Vasorelaxation and Blood Pressure Lowering in Rats.

Torilis japonica (TJ) fruit, is a herb that is traditionally used for erectile dysfunction (ED). Given the shared mechanisms of ED and hypertension through vascular smooth muscle, we hypothesized that TJ would be effective in vasodilation and blood pressure reduction. This study confirmed the authenticity of TJ samples via DNA barcoding and quantified the main active compound, torilin, using HPLC. TJ was extracted with distilled water (TJW) and 50% ethanol (TJE), yielding torilin contents of 0.35 ± 0.01% and 2.84 ± 0.02%, respectively. Ex vivo tests on thoracic aortic rings from Sprague-Dawley rats showed that TJE (3-300 µg/mL) induced endothelium-independent, concentration-dependent vasodilation, unlike TJW. Torilin caused concentration-dependent relaxation with an EC50 of 210 ± 1.07 µM. TJE's effects were blocked by a voltage-dependent K+ channel blocker and alleviated contractions induced by CaCl2 and angiotensin II. TJE inhibited vascular contraction induced by phenylephrine or KCl via extracellular CaCl2 and enhanced inhibition with nifedipine, indicating involvement of voltage-dependent and receptor-operated Ca2+ channels. Oral administration of TJE (1000 mg/kg) significantly reduced blood pressure in spontaneously hypertensive rats. These findings suggest TJ extract's potential for hypertension treatment through vasorelaxant mechanisms, though further research is needed to confirm its efficacy and safety.

Vasorelaxant and Blood Pressure-Lowering Effects of Cnidium monnieri Fruit Ethanol Extract in Sprague Dawley and Spontaneously Hypertensive Rats.

Cnidium monnieri (L.) Cusson, a member of the Apiaceae family, is rich in coumarins, such as imperatorin and osthole. Cnidium monnieri fruit (CM) has a broad range of therapeutic potential that can be used in anti-bacterial, anti-cancer, and sexual dysfunction treatments. However, its efficacy in lowering blood pressure through vasodilation remains unknown. This study aimed to assess the potential therapeutic effect of CM 50% ethanol extract (CME) on hypertension and the mechanism of its vasorelaxant effect. CME (1-30 µg/mL) showed a concentration-dependent vasorelaxation on constricted aortic rings in Sprague Dawley rats induced by phenylephrine via an endothelium-independent mechanism. The vasorelaxant effect of CME was inhibited by blockers of voltage-dependent and Ca2+-activated K+ channels. Additionally, CME inhibited the vascular contraction induced by angiotensin II and CaCl2. The main active compounds of CM, i.e., imperatorin (3-300 µM) and osthole (1-100 µM), showed a concentration-dependent vasorelaxation effect, with half-maximal effective concentration values of 9.14 ± 0.06 and 5.98 ± 0.06 µM, respectively. Orally administered CME significantly reduced the blood pressure of spontaneously hypertensive rats. Our research shows that CME is a promising treatment option for hypertension. However, further studies are required to fully elucidate its therapeutic potential.

Ketone body 3-hydroxybutyrate elevates cardiac output through peripheral vasorelaxation and enhanced cardiac contractility.

The ketone body 3-hydroxybutyrate (3-OHB) increases cardiac output and myocardial perfusion without affecting blood pressure in humans, but the cardiovascular sites of action remain obscure. Here, we test the hypothesis in rats that 3-OHB acts directly on the heart to increase cardiac contractility and directly on blood vessels to lower systemic vascular resistance. We investigate effects of 3-OHB on (a) in vivo hemodynamics using echocardiography and invasive blood pressure measurements, (b) isolated perfused hearts in Langendorff systems, and (c) isolated arteries and veins in isometric myographs. We compare Na-3-OHB to equimolar NaCl added to physiological buffers or injection solutions. At plasma concentrations of 2-4 mM in vivo, 3-OHB increases cardiac output (by 28.3±7.8%), stroke volume (by 22.4±6.0%), left ventricular ejection fraction (by 13.3±4.6%), and arterial dP/dtmax (by 31.9±11.2%) and lowers systemic vascular resistance (by 30.6±11.2%) without substantially affecting heart rate or blood pressure. Applied to isolated perfused hearts at 3-10 mM, 3-OHB increases left ventricular developed pressure by up to 26.3±7.4 mmHg and coronary perfusion by up to 20.2±9.5%. Beginning at 1-3 mM, 3-OHB relaxes isolated coronary (EC50=12.4 mM), cerebral, femoral, mesenteric, and renal arteries as well as brachial, femoral, and mesenteric veins by up to 60% of pre-contraction within the pathophysiological concentration range. Of the two enantiomers that constitute racemic 3-OHB, D-3-OHB dominates endogenously; but tested separately, the enantiomers induce similar vasorelaxation. We conclude that increased cardiac contractility and generalized systemic vasorelaxation can explain the elevated cardiac output during 3-OHB administration. These actions strengthen the therapeutic rationale for 3-OHB in heart failure management.

Synthesis of 6-alkoxy and 6-hydroxy-alkyl amine derivatives of braylin as vasorelaxing agents.

Braylin (10b) is a 8,8-dimethyl chromenocoumarin present in the plants of the family Rutaceae and Meliaceae and possesses vasorelaxing and anti-inflammatory activities. In this study, six 6-alkoxy (10b, 15-19), and twelve 6-hydroxy-alkyl amine (20a-20l) derivatives of braylin (11 and 12) were synthesized to delineate its structural requirement for vasorelaxing activity. The synthesized compounds were evaluated for vasorelaxation response in preconstricted intact rat Main Mesenteric Artery (MMA). The compounds showed l-type VDCC channel blockade depended and endothelium-independent vasorelaxation within the range of Emax < 50.00-96.70 % at 30 µM. Amongst all, 6-alkoxy derivatives were more active than 6-hydroxy-alkyl amine derivatives. The structural refinements about braylin showed that deletion of its methoxy group or homologation beyond ethoxy group presented deleterious effect on vasorelaxation response of braylin. Interestingly, substituting the ethoxy group in 10b presented the best activity and selectivity towards l-type VDCC channel blockade, a specific target cardiovascular function.

Endothelium-Independent Vasorelaxant Effects of Sudachitin and Demethoxysudachitin, Polymethoxyflavone from the Peel of Citrus sudachi on Isolated Rat Aorta.

Although polymethoxyflavones have been reported to exhibit various pharmacological actions, the effects of polymethoxyflavones sudachitin and demethoxysudachitin from the peel of Citrus sudachi on the cardiovascular system have not been clarified. This study investigated the mechanisms of vasorelaxation induced by sudachitin and demethoxysudachitin in rat aorta. Both compounds inhibited phenylephrine-induced contractions in a concentration-dependent manner. This was also observed in the case of potassium chloride (KCl)-induced contractions although the inhibitory effect was weak. In both contraction types, no differences were found in the inhibitory effects of sudachitin and demethoxysudachitin between endothelium-intact and -denuded aorta. The relaxant effects of sudachitin in endothelium-intact aortas were not affected by the nitric oxide synthase inhibitor N-nitro-L-arginine methyl ester hydrochloride (L-NAME) or the cyclooxygenase inhibitor indomethacin. In endothelium-denuded aorta, propranolol did not affect the relaxant effect of sudachitin. Both the adenylate cyclase activator forskolin- and soluble guanylate cyclase activator sodium nitroprusside-induced relaxant effects were potentiated by preincubation of sudachitin. Furthermore, the relaxant effect of sudachitin was not affected by the adenylate and guanylate cyclase inhibitors SQ22536 and or 1H-[1,2,4]Oxadiazolo[4,3-a]quinoxaline-1-one (ODQ), respectively. Finally, we examined the effect of phosphodiesterase inhibition. Phosphodiesterase inhibitors (3-isobutyl-1-methylxanthine, cilostamide or sildenafil) alone, sudachitin alone, and a combination of phosphodiesterase inhibitors with sudachitin exhibited relaxant effects, while the lack of any interaction between each phosphodiesterase inhibitor and sudachitin indicated an additive effect between the two substance categories. These results suggest that sudachitin and demethoxysudachitin cause endothelial-independent relaxation, and that the mechanism of vasorelaxation by sudachitin is associated with the enhancement of cAMP- and guanosine 3',5'-cyclic monophosphate (cGMP)-dependent pathways.

Enhancement of Sphingomyelinase-Induced Endothelial Nitric Oxide Synthase-Mediated Vasorelaxation in a Murine Model of Type 2 Diabetes.

Sphingolipids are important biological mediators both in health and disease. We investigated the vascular effects of enhanced sphingomyelinase (SMase) activity in a mouse model of type 2 diabetes mellitus (T2DM) to gain an understanding of the signaling pathways involved. Myography was used to measure changes in the tone of the thoracic aorta after administration of 0.2 U/mL neutral SMase in the presence or absence of the thromboxane prostanoid (TP) receptor antagonist SQ 29,548 and the nitric oxide synthase (NOS) inhibitor L-NAME. In precontracted aortic segments of non-diabetic mice, SMase induced transient contraction and subsequent weak relaxation, whereas vessels of diabetic (Leprdb/Leprdb, referred to as db/db) mice showed marked relaxation. In the presence of the TP receptor antagonist, SMase induced enhanced relaxation in both groups, which was 3-fold stronger in the vessels of db/db mice as compared to controls and could not be abolished by ceramidase or sphingosine-kinase inhibitors. Co-administration of the NOS inhibitor L-NAME abolished vasorelaxation in both groups. Our results indicate dual vasoactive effects of SMase: TP-mediated vasoconstriction and NO-mediated vasorelaxation. Surprisingly, in spite of the general endothelial dysfunction in T2DM, the endothelial NOS-mediated vasorelaxant effect of SMase was markedly enhanced.

Cracking the Endothelial Calcium (Ca2+) Code: A Matter of Timing and Spacing.

A monolayer of endothelial cells lines the innermost surface of all blood vessels, thereby coming into close contact with every region of the body and perceiving signals deriving from both the bloodstream and parenchymal tissues. An increase in intracellular Ca2+ concentration ([Ca2+]i) is the main mechanism whereby vascular endothelial cells integrate the information conveyed by local and circulating cues. Herein, we describe the dynamics and spatial distribution of endothelial Ca2+ signals to understand how an array of spatially restricted (at both the subcellular and cellular levels) Ca2+ signals is exploited by the vascular intima to fulfill this complex task. We then illustrate how local endothelial Ca2+ signals affect the most appropriate vascular function and are integrated to transmit this information to more distant sites to maintain cardiovascular homeostasis. Vasorelaxation and sprouting angiogenesis were selected as an example of functions that are finely tuned by the variable spatio-temporal profile endothelial Ca2+ signals. We further highlighted how distinct Ca2+ signatures regulate the different phases of vasculogenesis, i.e., proliferation and migration, in circulating endothelial precursors.

The Phthalic Selenoanhydride Decreases Rat Blood Pressure and Tension of Isolated Mesenteric, Femoral and Renal Arteries.

Phthalic selenoanhydride (R-Se) solved in physiological buffer releases various reactive selenium species including H2Se. It is a potential compound for Se supplementation which exerts several biological effects, but its effect on the cardiovascular system is still unknown. Therefore, herein we aimed to study how R-Se affects rat hemodynamic parameters and vasoactive properties in isolated arteries. The right jugular vein of anesthetized Wistar male rats was cannulated for IV administration of R-Se. The arterial pulse waveform (APW) was detected by cannulation of the left carotid artery, enabling the evaluation of 35 parameters. R-Se (1-2 µmol kg-1), but not phthalic anhydride or phthalic thioanhydride, transiently modulated most of the APW parameters including a decrease in systolic and diastolic blood pressure, heart rate, dP/dtmax relative level, or anacrotic/dicrotic notches, whereas systolic area, dP/dtmin delay, dP/dtd delay, anacrotic notch relative level or its delay increased. R-Se (~10-100 µmol L-1) significantly decreased the tension of precontracted mesenteric, femoral, and renal arteries, whereas it showed a moderate vasorelaxation effect on thoracic aorta isolated from normotensive Wistar rats. The results imply that R-Se acts on vascular smooth muscle cells, which might underlie the effects of R-Se on the rat hemodynamic parameters.

Perivascular Adipose Tissue Compensation for Endothelial Dysfunction in the Superior Mesenteric Artery of Female SHRSP.Z-Leprfa/IzmDmcr Rats.

Regulation of arterial tone by perivascular adipose tissue (PVAT) differs between sexes. In male SHRSP.Z-Leprfa/IzmDmcr rats (SHRSP.ZF), PVAT exerts a compensatory relaxation effect for the loss of endothelium-mediated vasorelaxation, which occurs during the early stages of metabolic syndrome. However, this effect deteriorates by 23 weeks of age. Here, therefore, we compared the effects of PVAT in female and male SHRSP.ZF. Acetylcholine-induced relaxation in superior mesenteric artery without PVAT did not differ between 23-week-old females and males. However, the presence of PVAT enhanced relaxation in 23-week-old females, but not in males. The mRNA levels of angiotensin II type 1 receptor (AT1R) in PVAT did not differ between sexes, but AT1R-associated protein (ATRAP) and apelin levels were higher in females than in males. We observed a positive relationship between differences in artery relaxation with and without PVAT and ATRAP or apelin mRNA levels. In 30-week-old females, PVAT-enhanced relaxation disappeared, and mRNA levels of AT1R increased, while apelin levels decreased compared to 23-week-old females. These results demonstrated that in SHRSP.ZF, PVAT compensation for endothelium dysfunction extended to older ages in females than in males. Apelin and AT1R/ATRAP expression in PVAT may be predictors of favorable effects.

Endothelial thioredoxin interacting protein (TXNIP) modulates endothelium-dependent vasorelaxation in hyperglycemia.

Endothelial dysfunction, hallmarked by an imbalance between vasoconstriction and vasorelaxation, is associated with diabetes. Thioredoxin Interacting protein (TXNIP), controlled by an exquisitely glucose sensitive gene, is increasingly recognized for its role in diabetes. However, the role of TXNIP in modulating diabetes-related endothelial dysfunction remains unclear. To elucidate the role of TXNIP, we generated two novel mouse strains; endothelial-specific TXNIP knockout (EKO) and a Tet-O inducible, endothelial-specific TXNIP overexpression (EKI). Hyperglycemia was induced by streptozotocin (STZ) treatment in floxed control (fl/fl) and EKO mice. Doxycycline (DOX) was given to EKI mice to induce endothelial TXNIP overexpression. The ablation of endothelial TXNIP improved glucose tolerance in EKO mice. Acetylcholine-induced, endothelium-dependent vasorelaxation was impaired in STZ-treated fl/fl mice while this STZ impaired vasorelaxation was attenuated in EKO mice. Hyperglycemia induction of NLRP3 and reductions in Akt and eNOS phosphorylation were also mitigated in EKO mice. Overexpression of endothelial TXNIP did not impair glucose tolerance in DOX-treated EKI mice, however induction of endothelial TXNIP led to impaired vasorelaxation in EKI mice. This was associated with increased NLRP3 and reduced Akt and eNOS activation. In conclusion, deletion of endothelial TXNIP is protective against and overexpression of endothelial TXNIP induces endothelial dysfunction; thus, endothelial TXNIP plays a critical role in modulating endothelial dysfunction.

In Vitro Model for Ischemic Stroke: Functional Analysis of Vascular Smooth Muscle Cells.

The Neurovascular Unit (NVU) is formed by vascular and neural cells controlling the cerebral hyperaemia. All the components are anatomically and functionally linked to each other, resulting in a highly efficient regulation of the cerebral blood flow, which, when interrupted, can lead to stroke. An ischemic stroke (IS) is the most common type of stroke with high rates of morbidity, mortality and disability. Therefore, it is of extreme importance to protect the functional and structural integrity of the NVU in patients with IS, understanding the mechanisms involved and how it affects each component of the NVU. Thus, the aim of this work is to analyse how the vascular smooth muscle cells from the rat middle cerebral artery function/react after an ischemic event. To mimic this event, primary cortical cultures were challenged to oxygen and glucose deprivation (OGD) for 4 h and 6 h, and the smooth muscle cells (SMCs) contractility was analysed after exposure to different media previously conditioned by the cortical cultures upon reperfusion. The results show a dual effect on the SMCs response to the vasorelaxant agent, only for cells exposed to the reperfusion media conditioned by neuron-glia cultures challenged by OGD, leading to increased relaxation of the SMCs for OGD 4 h, whereas for OGD 6 h the effect is reversed leading to contraction of the SMCs. These differences demonstrate that the astrocytes mediate the vasoactive response of vascular smooth muscle by releasing factors into the reperfusion medium, and the hypoxia time is fundamental for a beneficial/harmful response by the vascular smooth muscle.

Cardiovascular characterization of the novel organic mononitrate NDIBP in rats.

Organic nitrates are widely used to restore endogenous nitric oxide (NO) levels reduced by endothelial nitric oxide synthase dysfunction. However, these drugs are associated with undesirable side effects, including tolerance. This study aims to investigate the cardiovascular effects of the new organic nitrate 1,3-diisobutoxypropan-2-yl nitrate (NDIBP). Specifically, we assessed its effects on blood pressure, vascular reactivity, acute toxicity, and the ability to induce tolerance. In vitro and ex vivo techniques showed that NDIBP released NO both in a cell-free system and in isolated mesenteric arteries preparations through a process catalyzed by xanthine oxidoreductase. NDIBP also evoked endothelium-independent vasorelaxation, which was significantly attenuated by 2-phenyl-4,4,5,5,-tetramethylimidazoline-1-oxyl 3-oxide (PTIO, 300 µM), a nitric oxide scavenger; 1-H-[1,2,4] oxadiazolo-[4,3-a]quinoxalin-1-one (ODQ, 10 µM), a soluble guanylyl cyclase inhibitor; tetraethylammonium (TEA, 3 mM), a potassium channel blocker; febuxostat (500 nM), a xanthine oxidase inhibitor; and proadifen (10 µM), an inhibitor of cytochrome P450 enzyme. Furthermore, this organic nitrate did not induce tolerance in isolated vessels and presented low toxicity following acute oral administration. In vivo changes on cardiovascular parameters were assessed using normotensive and renovascular hypertensive rats. NDIBP evoked a reduction of blood pressure that was significantly higher in hypertensive animals. Our results suggest that NDIBP acts as a NO donor, inducing blood pressure reduction without having the undesirable effects of tolerance. Those effects seem to be mediated by activation of NO-sGC-cGMP pathway and positive modulation of K+ channels in vascular smooth muscle.

Rosa damascena Miller essential oil relaxes rat thoracic aorta through the NO-cGMP-dependent pathway.

AIM: This study aimed to investigate the effects of Rosa damascena Mill. essential oil on the vascular activity of rat thoracic aorta and its underlying mechanisms. METHODS: Experiments were performed using the isolated tissue bath model and Wistar rats. 0.1, 1, 10, and 100 µg/mL concentrations of rose oil were administered in all groups. To determine the vasoactive effects of rose oil, submaximal contractions were conducted by applying 10-5 M PE and 45 mM KCl separately in both endothelium-intact and -denuded segments. Time-matched distilled water groups were formed for control. To evaluate the role of endothelium-derived vasodilative factors, endothelium-intact segments were incubated with nitric oxide synthase inhibitor L-NAME, soluble guanylate cyclase inhibitor ODQ, and a non-selective cyclooxygenase inhibitor INDO. The statistical significance level was considered as p < 0.05. RESULTS: 1, 10, and 100 µg/mL rose oil doses led to vasorelaxation in thoracic aortas precontracted with 10-5 M PE (p: 0.029, p: 0.000, p: 0.000, respectively). In precontracted thoracic aortas with 45 mM KCl, the significant effect of rose oil persisted, albeit slightly diminished. When the endothelium was removed, the relaxant effect of rose oil was partially reduced, but still significant (p: 0.035, p: 0.028, p: 0.000, respectively). Preincubations with L-NAME and ODQ significantly attenuated rose oil-induced relaxation of endothelium-intact aortas precontracted with 10-5 M PE. In contrast, preincubation INDO did not modulate rose oil-induced relaxation. CONCLUSION: In conclusion, it was shown for the first time that rose oil can significantly mediate vasorelaxation in both PE and KCl precontracted rat thoracic aortas. Rose oil induced vasodilation with or without endothelium in a concentration-dependent manner. It was also shown that rose oil-induced vasorelaxant effects were reduced by L-NAME or ODQ pretreatment, but not modulated by INDO. These results demonstrated that rose oil-induced endothelium-dependent vasodilation is mediated by the NO-cGMP-dependent pathway.

Dystrophin deficiency impairs vascular structure and function in the canine model of Duchenne muscular dystrophy.

Duchenne muscular dystrophy (DMD) is a muscle-wasting disease caused by dystrophin deficiency. Vascular dysfunction has been suggested as an underlying pathogenic mechanism in DMD. However, this has not been thoroughly studied in a large animal model. Here we investigated structural and functional changes in the vascular smooth muscle and endothelium of the canine DMD model. The expression of dystrophin and endothelial nitric oxide synthase (eNOS), neuronal NOS (nNOS), and the structure and function of the femoral artery from 15 normal and 16 affected adult dogs were evaluated. Full-length dystrophin was detected in the endothelium and smooth muscle in normal but not affected dog arteries. Normal arteries lacked nNOS but expressed eNOS in the endothelium. NOS activity and eNOS expression were reduced in the endothelium of dystrophic dogs. Dystrophin deficiency resulted in structural remodeling of the artery. In affected dogs, the maximum tension induced by vasoconstrictor phenylephrine and endothelin-1 was significantly reduced. In addition, acetylcholine-mediated vasorelaxation was significantly impaired, whereas exogenous nitric oxide-induced vasorelaxation was significantly enhanced. Our results suggest that dystrophin plays a crucial role in maintaining the structure and function of vascular endothelium and smooth muscle in large mammals. Vascular defects may contribute to DMD pathogenesis. © 2021 The Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Mechanism of canagliflozin-induced vasodilation in resistance mesenteric arteries and the regulation of systemic blood pressure.

Canagliflozin, a sodium glucose co-transporter 2 (SGLT2) inhibitor, is reported to produce beneficial cardiovascular effects including a reduction in arterial contractility, and blood pressure. However, whether canagliflozin could directly relax resistance mesenteric arteries, underlying molecular mechanism and its role in regulating systemic blood pressure remain unclear. Here, we investigated the mechanism of regulation of small mesenteric artery contractility and its relevance for blood pressure regulation. Our pressure myography data showed that canagliflozin application rapidly produces a concentration-dependent vasodilation in mesenteric arteries. Such vasodilation was inhibited by concurrent inhibition of smooth muscle cell voltage-gated K+ channels KV1.5 (by 1 µM DPO-1), KV2.1 (by 100 nM guangxitoxin), and KV7 (by 10 µM linopirdine) but not by the inhibition of small-, intermediate-, and large-conductance Ca2+-activated K+ channels (SKCa by 1 µM apamin, IKCa 10 µM TRAM-34, and BKCa by 10 µM paxilline, respectively), ATP-sensitive (KATP) channels (by 10 µM glibenclamide), or SERCA pump (by 0.1 µM thapsigargin). Inhibition of SGLTs (by 1 µM phlorizin or the inhibition of endothelial signaling did not alter canagliflozin-evoked vasodilation. Consistently, acute canagliflozin treatment (4 mg/kg body weight) lowered systemic blood pressure in vivo. Overall, our data suggests that canagliflozin stimulates KV1.5, KV2.1, and KV7 channels, leading to vasodilation and a reduction of systemic blood pressure.

Soluble Guanylate Cyclase-Mediated Relaxation in Aortas from Rats with Renovascular Hypertension.

Soluble guanylate cyclase (sGC) plays an important role in nitric oxide (NO)-mediated regulation of vascular tone; however, NO bioavailability is often reduced in diseased blood vessels. Accumulating evidence suggests that a shift of sGC from the NO-sensitive form to the NO-insensitive form could be an underlying cause contributing to this reduction. Herein, we investigated the impact of renovascular hypertension on NO-sensitive and NO-insensitive sGC-mediated relaxation in rat aortas. Renovascular hypertension was induced by partially clipping the left renal artery (2-kidneys, 1-clip; 2K1C) for 10 weeks. Systolic, diastolic, and mean arterial pressures were significantly increased in the 2K1C group when compared with the sham group. In addition, plasma thiobarbituric acid reactive substances and aortic superoxide generation were significantly enhanced in the 2K1C group when compared with those in the sham group. The vasorelaxant response of isolated aortas to the sGC stimulator BAY 41-2272 (NO-sensitive sGC agonist) was comparable between the sham and 2K1C groups. Likewise, the sGC activator BAY 60-2770 (NO-insensitive sGC agonist)-induced relaxation did not differ between the sham and 2K1C groups. In addition, the cGMP mimetic 8-Br-cGMP (protein kinase G agonist) induced similar relaxation in both groups. Furthermore, there were no differences in BAY 41-2272-stimulated and BAY 60-2770-stimulated cGMP generation between the groups. These findings suggest that the balance between NO-sensitive and NO-insensitive forms of sGC is maintained during renovascular hypertension. Therefore, sGC might not be responsible for the reduced NO bioavailability observed during renovascular hypertension.

Schwarzinicine A inhibits transient receptor potential canonical channels and exhibits overt vasorelaxation effects.

This study investigated the vasorelaxant effects of schwarzinicine A, an alkaloid recently reported from Ficus schwarzii Koord. Regulation of calcium homeostasis in vascular smooth muscle cells (VSMC) is viewed as one of the main mechanisms for controlling blood pressure. L-type voltage-gated calcium channel (VGCC) blockers are commonly used for controlling hypertension. Recently, the transient receptor potential canonical (TRPC) channels were found in blood vessels of different animal species with evidence of their roles in the regulation of vascular contractility. In this study, we studied the mechanism of actions of schwarzinicine A focusing on its regulation of L-type VGCC and TRPC channels. Schwarzinicine A exhibited the highest vasorelaxant effect (123.1%) compared to other calcium channel blockers. It also overtly attenuated calcium-induced contractions of the rat isolated aortae in a calcium-free environment showing its mechanism to inhibit calcium influx. Fluorometric intracellular calcium recordings confirmed its inhibition of hTRPC3-, hTRPC4-, hTRPC5- and hTRPC6-mediated calcium influx into HEK cells with IC50 values of 3, 17, 19 and 7 µM, respectively. The evidence gathered in this study suggests that schwarzinicine A blocks multiple TRPC channels and L-type VGCC to exert a significant vascular relaxation response.

GPR18-Mediated Relaxation of Human Isolated Pulmonary Arteries.

GPR18 receptor protein was detected in the heart and vasculature and appears to play a functional role in the cardiovascular system. We investigated the effects of the new GPR18 agonists PSB-MZ-1415 and PSB-MZ-1440 and the new GPR18 antagonist PSB-CB-27 on isolated human pulmonary arteries (hPAs) and compared their effects with the previously proposed, but unconfirmed, GPR18 ligands NAGly, Abn-CBD (agonists) and O-1918 (antagonist). GPR18 expression in hPAs was shown at the mRNA level. PSB-MZ-1415, PSB-MZ-1440, NAGly and Abn-CBD fully relaxed endothelium-intact hPAs precontracted with the thromboxane A2 analog U46619. PSB-CB-27 shifted the concentration-response curves (CRCs) of PSB-MZ-1415, PSB-MZ-1440, NAGly and Abn-CBD to the right; O-1918 caused rightward shifts of the CRCs of PSB-MZ-1415 and NAGly. Endothelium removal diminished the potency and the maximum effect of PSB-MZ-1415. The potency of PSB-MZ-1415 or NAGly was reduced in male patients, smokers and patients with hypercholesterolemia. In conclusion, the novel GPR18 agonists, PSB-MZ-1415 and PSB-MZ-1440, relax hPAs and the effect is inhibited by the new GPR18 antagonist PSB-CB-27. GPR18, which appears to exhibit lower activity in hPAs from male, smoking or hypercholesterolemic patients, may become a new target for the treatment of pulmonary arterial hypertension.