Vasomotion in human arteries and their regulations based on ion channel regulations: 10 years study.

Vasomotion is the oscillation of vascular tone which gives rise to flow motion of blood into an organ. As is well known, spontaneous contractile organs such as heart, GI, and genitourinary tract produce rhythmic contraction. It imposes or removes pressure on their vessels alternatively for exchange of many substances. It was first described over 150 years ago, however the physiological mechanism and pathophysiological implications are not well understood. This study aimed to elucidate underlying mechanisms and physiological function of vasomotion in human arteries. Conventional contractile force measurement, immunohistochemistry, and Western blot analysis were employed to study human left gastric artery (HLGA) and uterine arteries (HUA). RESULTS: Circular muscle of HLGA and/or HUA produced sustained tonic contraction by high K+ (50 mM) which was blocked by 2 µM nifedipine. Stepwise stretch and high K+ produced nerve-independent spontaneous contraction (vasomotion) (around 45% of tested tissues). Vasomotion was also produced by application of BayK 8644, 5-HT, prostagrandins, oxytocin. It was blocked by nifedipine (2 µM) and blockers of intracellular Ca2+ stores. Inhibitors of Ca2+ -activated Cl- channels (DIDS and/or niflumic acid) and ATP-sensitive K+ (KATP ) channels inhibited vasomotion reversibly. Metabolic inhibition by sodium cyanide (NaCN) and several neuropeptides also regulated vasomotion in KATP channel-sensitive and -insensitive manner. Finally, we identified TMEM16A Ca2+ -activated Cl- channels and subunits of KATP channels (Kir 6.1/6.2 and sulfonylurea receptor 2B [SUR2B]), and c-Kit positivity by Western blot analysis. We conclude that vasomotion is sensitive to TMEM16A Ca2+ -activated Cl- channels and metabolic changes in human gastric and uterine arteries. Vasomotion might play an important role in the regulation of microcirculation dynamics even in pacemaker-related autonomic contractile organs in humans.

Palmitate decreases migration and proliferation and increases oxidative stress and inflammation in smooth muscle cells: role of the Nrf2 signaling pathway.

Fatty acids are essential to cell functionality and may exert diverging vascular effects including migration, proliferation, oxidative stress, and inflammation. This study examined the effect of palmitate on human coronary artery smooth muscle cell (HCASMC) function. An in vitro wound-healing assay indicated that palmitate decreased HCASMC migration in dose- and time-dependent manners. Furthermore, bromodeoxyuridine incorporation assays indicated that palmitate decreased HCASMC proliferation in a dose-response manner. Palmitate also increased reactive oxygen species formation, malondialdehyde content, and intracellular lipid droplets accompanied with increased fatty acid binding protein 4 expression. Moreover, palmitate induced gene expression (monocyte chemoattractant protein 1, matrix metalloproteinase-2, IL-1ß, IL-6, IL-8, and TNF-α) and intracellular protein content (plasminogen activator inhibitor-1 and urokinase plasminogen activator) of inflammatory mediators. Finally, we showed that palmitate activates the transcription factor Nrf2 and the upstream kinases ERK1/2 and Akt in HCASMCs. The inhibitor of Nrf2, trigonelline, significantly attenuated palmitate-induced HCASMC expression of the Nrf2 target gene NQO1. These findings indicate that palmitate might be critically related to HCASMC function by slowing cell migration and proliferation and inducing lipid-laden cells, oxidative stress, and inflammation in part by activation of the Nrf2 transcription factor. Palmitate's activation of proinflammatory Nrf2 signaling may represent a novel mechanism mediating the proatherogenic actions of saturated fatty acids.

Melanocortin-1 Receptor Positively Regulates Human Artery Endothelial Cell Migration.

BACKGROUND/AIMS: Melanocortin receptors (MCRs) belong to a hormonal signalling pathway with multiple homeostatic and protective actions. Microvascular and umbilical vein endothelial cells (ECs) express components of the melanocortin system, including the type 1 receptor (MC1R), playing a role in modulating inflammation and vascular tone. Since ECs exhibit a remarkable heterogeneity, we investigated whether human artery ECs express any functional MCR and whether its activation affects cell migration. METHODS: We used reverse transcription real-time PCR to examine the expression of melanocortin system components in primary human artery ECs. We assessed MC1R protein expression and activity by western blot, immunohistochemistry, cAMP production, and intracellular Ca²âº mobilization assays. We performed gap closure and scratch tests to examine cell migration after stimulation with alpha-melanocyte-stimulating hormone (α-MSH), the receptor highest-affinity natural ligand. We assessed differential time-dependent transcriptional changes in migrating cells by microarray analysis. RESULTS: We showed that human aortic ECs (HAoECs) express a functionally active MC1R. Unlike microvascular ECs, arterial cells did not express the α-MSH precursor proopiomelanocortin, nor produced the hormone. MC1R engagement with a single pulse of α-MSH accelerated HAoEC migration both in the directional migration assay and in the scratch wound healing test. This was associated with an enhancement in Ca²âº signalling and inhibition of cAMP elevation. Time-course genome-wide expression analysis in HAoECs undergoing directional migration allowed identifying dynamic co-regulation of genes involved in extracellular matrix-receptor interaction, vesicle-mediated trafficking, and metal sensing - which have all well-established influences on EC motility -, without affecting the balance between pro- and anticoagulant genes. CONCLUSION: Our work broadens the knowledge on peripherally expressed MC1R. These results indicate that the receptor is constitutively expressed by arterial ECs and provide evidence of a novel homeostatic function for MC1R, whose activation may participate in preventing/healing endothelial dysfunction or denudation in macrovascular arteries.

[Effects of Aging on Endothelium-dependent Vasodilation of Human Artery].

OBJECTIVE: To determine the effects of aging on endothelium-dependent vasodilation of human artery. METHODS: Vessel tension changes induced by acetylcholine (ACh) and endothelial-derived hyperpolarizing factor (EDHF) on gastroepiploic artery rings were recorded in 15 patients with stomach cancer aged between 51 and 83 years. The mRNA expressions of endothelial nitric oxide synthase (eNOS), cyclooxygenase (COX), cystathionineγ lyase (CSE) and the C-type natriuretic peptide (CNP) in the artery vessels were detected by qRT-PCR. RESULTS: Both endothelium-dependent and EDHF induced vasodilation decreased with age (P<0.05). The mRNA expressions of eNOS, CSE and CNP also decreased with age (P<0.05), except COX. CONCLUSION: Aging could impair the function of endothelium-dependent vasodilation and EDHF-induced vasodialtion of human artery, possibly due to decreased NO, H2S and CNP in artery.

The atherogenic actions of LPC on vascular smooth muscle cells and its LPA receptor mediated mechanism.

Lysophosphatidylcholine (LPC) is a bioactive lipid constituent of oxidized low density lipoprotein (ox-LDL). It regulates various cellular functions, including migration of circulating monocytes, expression of endothelial adhesion molecules, proliferation and migration of vascular smooth muscle cells (VSMCs). LPC can also be hydrolyzed into lysophosphatidic acid (LPA) by autotaxin (ATX) which possesses lysophospholipase D (lyso-PLD) activity. The aim of this study was to explore the effects of LPC on proliferation and migration of human artery smooth muscle cells (HASMCs) and the involvement of LPC-ATX-LPA pathway in these processes. In vitro, we found that LPC and LPA stimulated HASMCs proliferation and migration. Knockdown of LPA1 by siRNA and inhibit Gi protein with pertussis toxin (PTX) showed the contrary results. Silencing of LPC receptor genes did not significantly affect the LPC induced proliferation and migration. We detected the higher expressed mRNA and protein of ATX in HASMCs, and measured lyso-PLD activity. In atherosclerotic rabbit model, we observed high LPC level and high lyso-D activity in blood, and high expression of LPA1 in aorta walls. We also found that neointima appeared to be thickened and mRNA expressions of LPA1 appeared to be increased. These results revealed that LPC was converted into LPA by ATX to induce the proliferation and migration in HASMCs through LPA1/Gi/o/MAP Kinase signaling pathway. Our research suggested that LPC-ATX-LPA system contributed to the atherogenic action induced by ox-LDL. LPA1 antagonist may be considered as a potential therapeutic and preventative drug for cardiovascular disease.

Vasodilator activity of hydrogen sulfide (H2S) in human mesenteric arteries.

The role of endogenous H2S has been highlighted as a gaseous transmitter. The vascular smooth muscle inhibitory effects of H2S have been characterized in isolated aorta and mesenteric arteries in rats and mice. Our study was aimed at investigating the vascular effects of H2S on human isolated mesenteric arteries and examining the underlying mechanisms involved. All experiments were performed on rings (4-8mm long) of human mesenteric arteries obtained from patients undergoing abdominal surgery. Ethical approval was obtained from the Ethics Committee of the University Hospital of the University of Florence (app. N. 2015/0024947). The effect of NaHS, an H2S donor, was determined using noradrenaline pre-contracted human isolated mesenteric rings. NaHS evoked a concentration-dependent relaxation (EC50 57µM). In contrast, homocysteine, an endogenous precursor of H2S, failed to affect human isolated mesenteric rings. Vasorelaxant response to NaHS was reduced by endothelium removal, application of the nitric oxide synthase inhibitor L-NAME and ODQ inhibitor of cyclic GMP. SQ 22536, an adenylate-cyclase inhibitor, failed to block NaHS-induced vasorelaxation. Inhibition of endogenous prostanoid production by indomethacin significantly reduced NaHS induced vasorelaxation. The role of potassium channels was also examined: blockers of the Ca2+-dependent potassium channel, charybdotoxin and apamin, failed to have any influence on the relaxant response to NaHS on this vascular tissue. In summary, H2S induced relaxation of isolated rings of human mesenteric arteries. Endothelium-dependent related mechanisms with the stimulation of ATP-sensitive potassium channels represents important cellular mechanisms for H2S effect on human mesenteric arteries.

Neuropeptide Y: Direct vasoconstrictor and facilitatory effects on P2X1 receptor-dependent vasoconstriction in human small abdominal arteries.

Neuropeptide Y (NPY) is co-released with norepinephrine and ATP by sympathetic nerves innervating arteries. Circulating NPY is elevated during exercise and cardiovascular disease, though information regarding the vasomotor function of NPY in human blood vessels is limited. Wire myography revealed NPY directly stimulated vasoconstriction (EC50 10.3 ± 0.4 nM; N = 5) in human small abdominal arteries. Maximum vasoconstriction was antagonised by both BIBO03304 (60.7 ± 6%; N = 6) and BIIE0246 (54.6 ± 5%; N = 6), suggesting contributions of both Y1 and Y2 receptor activation, respectively. Y1 and Y2 receptor expression in arterial smooth muscle cells was confirmed by immunocytochemistry, and western blotting of artery lysates. α,ß-meATP evoked vasoconstrictions (EC50 282 ± 32 nM; N = 6) were abolished by suramin (IC50 825 ± 45 nM; N = 5) and NF449 (IC50 24 ± 5 nM; N = 5), suggesting P2X1 mediates vasoconstriction in these arteries. P2X1, P2X4 and P2X7 were detectable by RT-PCR. Significant facilitation (1.6-fold) of α,ß-meATP-evoked vasoconstrictions was observed when submaximal NPY (10 nM) was applied between α,ß-meATP applications. Facilitation was antagonised by either BIBO03304 or BIIE0246. These data reveal NPY causes direct vasoconstriction in human arteries which is dependent upon both Y1 and Y2 receptor activation. NPY also acts as a modulator, facilitating P2X1-dependent vasoconstriction. Though in contrast to the direct vasoconstrictor effects of NPY, there is redundancy between Y1 and Y2 receptor activation to achieve the facilitatory effect.

Stretch and stress distributions in the human artery based on two-layer model considering residual stresses.

The objective is to know the stress distributions in the arterial walls under residual stresses based on two-layer model. Human common carotid arteries were analysed to show stress distributions at physiological and supraphysiological intraluminal pressures. The analyses for the loaded states were performed with stretch ratios with reference to a Riemannian stress-free configuration which is a 3D non-Euclidean manifold due to the nonzero Riemann curvature tensor. The experimental data obtained by other literature were used for the common carotid arteries to analyse the stretch and stress distributions in the arterial wall although kinematics is different from the literature. The stretches and stresses were calculated for the unloaded state, i.e. the residual stretches and stresses. And those at the axial stretch ratio 1.1 with reference to the unloaded state were calculated at the intraluminal pressures 16, 50, and 100 kPa. The stresses increased from the inner surface to the outer surface at all pressures analysed. These results suggest that in the human arteries the mechanical loads are mainly supported with the adventitia even though the media and intima play an important role to control of physiological functions.

TIM­3 inhibits PDGF­BB­induced atherogenic responses in human artery vascular smooth muscle cells.

Increasing evidence suggests that T­cell immunoglobulin and mucin domain 3 (TIM­3) displays anti­atherosclerotic effects, but its role in vascular smooth muscle cells (VSMCs) has not been reported. The present study aimed to investigate the function of TIM­3 and its roles in human artery VSMCs (HASMCs). A protein array was used to investigate the TIM­3 protein expression profile, which indicated that TIM­3 expression was increased in the serum of patients with lower extremity arteriosclerosis obliterans disease (LEAOD) compared with healthy individuals. Immunohistochemistry and western blotting of arterial tissue further revealed that TIM­3 expression was increased in LEAOD artery tissue compared with normal artery tissue. Additionally, platelet­derived growth factor­BB (PDGF­BB) displayed a positive correlation with TIM­3 expression in HASMCs. TIM­3 decreased the migration and proliferation of PDGF­BB­induced HASMCs, and anti­TIM­3 blocked the effects of TIM­3. The effect of TIM­3 on the proliferation and migration of HASMCs was further investigated using LV­TIM­3­transduced cells. The results revealed that TIM­3 also inhibited PDGF­BB­induced expression of the inflammatory factors interleukin­6 and tumor necrosis factor­α by suppressing NF­κB activation. In summary, the present study revealed that TIM­3 displayed a regulatory role during the PDGF­BB­induced inflammatory reaction in HASMCs, which indicated that TIM­3 may display anti­atherosclerotic effects.

Vascular and molecular pharmacology of the metabolically stable CGRP analogue, SAX.

The main purpose of this study was to compare in vitro pharmacological properties of human αCGRP (CGRP) and a recently discovered metabolically stable CGRP analogue, SAX, in isolated rat and human artery segments. In rat, CGRP and SAX induced similar vasodilatory responses in isolated mesenteric artery with the potency of SAX being lower than that of CGRP (vasodilatory pEC50 8.2 ±â€¯0.12 and 9.0 ±â€¯0.11, respectively). A corresponding difference in receptor binding affinity of SAX and CGRP was determined in rat cerebral membranes (pKi 8.3 ±â€¯0.19 and 9.3 ±â€¯0.14, respectively). CGRP and SAX-induced vasodilation was antagonised with similar potencies by the CGRP receptor antagonist BIBN4096BS supporting a uniform receptor population for the agonists. In human tissue, SAX and CGRP induced similar pharmacological responses with different potencies in subcutaneous artery (vasodilatory pEC50 8.8 ±â€¯0.18 and 9.5 ±â€¯0.13, respectively) and human recombinant receptors (cAMP signalling pEC50 9.1 ±â€¯0.16 and 10.2 ±â€¯0.19). Like in the rat mesenteric artery, both SAX and CGRP-responses were inhibited by the CGRP receptor antagonist BIBN4096BS with similar antagonistic potencies. In conclusion, all pharmacological characteristics of SAX and CGRP in human and rat sources points towards action via a uniform BIBN4096BS sensitive receptor population with the potency of SAX being 5-10 fold lower than that of CGRP.

The Effect of Strain Hardening on the Dynamic Response of Human Artery Segments.

BACKGROUND: When subjected to time-dependent blood pressure, human arteries undergo large deformations, exhibiting mainly nonlinear hyperelastic type of response. The mechanical response of arteries depends on the health of tissues that comprise the artery walls. Typically, healthy arteries exhibit convex strain hardening under tensile loads, atherosclerotic parts exhibit stiffer response, and aneurysmatic parts exhibit softening response. In reality, arterial dynamics is the dynamics of a propagating pulse, originating in heart ventricle, propagating along aorta, bifurcating, etc. Artery as a whole cannot be simulated as a lump ring, however its cross section can be simulated as a vibrating ring having a phase lag with respect to the other sections, creating a running pressure wave. A full mathematical model would require fluid-solid interaction modeling continuity of blood flow in a compliant vessel and a momentum equation. On the other hand, laboratory testing often uses small-length arteries, the response of which is covered by the present work. In this way, material properties that change along the artery length can be investigated. OBJECTIVE: The effect of strain hardening on the local dynamic response of human arteries (excluding the full fluid-structure interaction) is examined through appropriate hyperelastic models related to the health condition of the blood vessel. Furthermore, this work aims at constituting a basis for further investigation of the dynamic response of arteries accounting for viscosity. METHOD: The governing equation of motion is formulated for three different hyperelastic material behaviors, based on the constitutive law proposed by Skalak et al., Hariton, and Mooney-Rivlin, associated with the hardening behavior of healthy, atherosclerotic, and aneurysmatic arteries, respectively. The differences between these modelling implementations are caused by physiology, since aneurysmatic arteries are softer and often sclerotic arteries are stiffer than healthy arteries. The response is investigated by proper normalization of the involved material parameters of the arterial walls, geometry of the arteries, load histories, time effects, and pre-stressing. The effect of each problem parameter on the arterial response has been studied. The peak response of the artery segment is calculated in terms of radial displacements, principal elongations, principal stresses, and strain-energy density. The validity of the proposed analytical models is demonstrated through comparison with previous studies that investigate the dynamic response of arterial models. RESULTS: Important metrics that can be useful to vascular surgery are the radial deformation and the maximum strain-energy density along with the radial resonance frequencies. These metrics are found to be influenced heavily by the nonlinear strain-hardening characteristics of the model and the longitudinal pre-stressing. CONCLUSION: The proposed formulation permits a systematic and generalizable investigation, which, together with the low computational cost of analysis, makes it a valuable tool for calculating the response of healthy, atherosclerotic, and aneurysmatic arteries. The radial resonance frequencies can explain certain murmures developed in stenotic arteries.

A personalized-model-based central aortic pressure estimation method.

Central Aortic Pressure (CAP) can be used to predict cardiovascular structural damage and cardiovascular events, and the development of simple, well-validated and non-invasive methods for CAP waveforms estimation is critical to facilitate the routine clinical applications of CAP. Existing widely applied methods, such as generalized transfer function (GTF-CAP) method and N-Point Moving Average (NPMA-CAP) method, are based on clinical practices, and lack a mathematical foundation. Those methods also have inherent drawback that there is no personalisation, and missing individual aortic characteristics. To overcome this pitfall, we present a personalized-model-based central aortic pressure estimation method (PM-CAP)in this paper. This PM-CAP has a mathematical foundation: a human aortic network model is proposed which is developed based on viscous fluid mechanics theory and could be personalized conveniently. Via measuring the pulse wave at the proximal and distal ends of the radial artery, the least square method is then proposed to estimate patient-specific circuit parameters. Thus the central aortic pulse wave can be obtained via calculating the transfer function between the radial artery and central aorta. An invasive validation study with 18 subjects comparing PM-CAP with direct aortic root pressure measurements during percutaneous transluminal coronary intervention was carried out at the Beijing Hospital. The experimental results show better performance of the PM-CAP method compared to the GTF-CAP method and NPMA-CAP method, which illustrates the feasibility and effectiveness of the proposed method.