Upregulation of NOR-1 in calcified human vascular tissues: impact on osteogenic differentiation and calcification.

Cardiovascular calcification is a significant public health issue whose pathophysiology is not fully understood. NOR-1 regulates critical processes in cardiovascular remodeling, but its contribution to ectopic calcification is unknown. NOR-1 was overexpressed in human calcific aortic valves and calcified atherosclerotic lesions colocalizing with RUNX2, a factor essential for osteochondrogenic differentiation and calcification. NOR-1 and osteogenic markers were upregulated in calcifying human valvular interstitial cells (VICs) and human vascular smooth muscle cells (VSMCs). Gain- and loss-of-function approaches demonstrated that NOR-1 negatively modulates the expression of osteogenic genes relevant for the osteogenic transdifferentiation (RUNX2, IL-6, BMP2, and ALPL) and calcification of VICs. VSMCs from transgenic mice overexpressing NOR-1 in these cells (TgNOR-1VSMC) expressed lower basal levels of osteogenic genes (IL-6, BMP2, ALPL, OPN) than cells from WT littermates, and their upregulation by a high-phosphate osteogenic medium (OM) was completely prevented by NOR-1 transgenesis. Consistently, this was associated with a dramatic reduction in the calcification of both transgenic VSMCs and aortic rings from TgNOR-1VSMC mice exposed to OM. Atherosclerosis and calcification were induce in mice by the administration of AAV-PCSK9D374Y and a high-fat/high-cholesterol diet. Challenged-TgNOR-1VSMC mice exhibited decreased vascular expression of osteogenic markers, and both less atherosclerotic burden (assessed in whole aorta and lesion size in aortic arch and brachiocephalic artery) and less vascular calcification (assessed either by near-infrared fluorescence imaging or histological analysis) than WT mice. Our data indicate that NOR-1 negatively modulates the expression of genes critically involved in the osteogenic differentiation of VICs and VSMCs, thereby restraining ectopic cardiovascular calcification.

LRRC8A anion channels modulate vascular reactivity via association with myosin phosphatase rho interacting protein.

Leucine-rich repeat containing 8A (LRRC8A) volume regulated anion channels (VRACs) are activated by inflammatory and pro-contractile stimuli including tumor necrosis factor alpha (TNFα), angiotensin II and stretch. LRRC8A associates with NADPH oxidase 1 (Nox1) and supports extracellular superoxide production. We tested the hypothesis that VRACs modulate TNFα signaling and vasomotor function in mice lacking LRRC8A exclusively in vascular smooth muscle cells (VSMCs, Sm22α-Cre, Knockout). Knockout (KO) mesenteric vessels contracted normally but relaxation to acetylcholine (ACh) and sodium nitroprusside (SNP) was enhanced compared to wild type (WT). Forty-eight hours of ex vivo exposure to TNFα (10 ng/mL) enhanced contraction to norepinephrine (NE) and markedly impaired dilation to ACh and SNP in WT but not KO vessels. VRAC blockade (carbenoxolone, CBX, 100 µM, 20 min) enhanced dilation of control rings and restored impaired dilation following TNFα exposure. Myogenic tone was absent in KO rings. LRRC8A immunoprecipitation followed by mass spectroscopy identified 33 proteins that interacted with LRRC8A. Among them, the myosin phosphatase rho-interacting protein (MPRIP) links RhoA, MYPT1 and actin. LRRC8A-MPRIP co-localization was confirmed by confocal imaging of tagged proteins, Proximity Ligation Assays, and IP/western blots. siLRRC8A or CBX treatment decreased RhoA activity in VSMCs, and MYPT1 phosphorylation was reduced in KO mesenteries suggesting that reduced ROCK activity contributes to enhanced relaxation. MPRIP was a target of redox modification, becoming oxidized (sulfenylated) after TNFα exposure. Interaction of LRRC8A with MPRIP may allow redox regulation of the cytoskeleton by linking Nox1 activation to impaired vasodilation. This identifies VRACs as potential targets for treatment or prevention of vascular disease.

Heightened apoptotic priming of vascular cells across tissues and life span predisposes them to cancer therapy-induced toxicities.

Although major organ toxicities frequently arise in patients treated with cytotoxic or targeted cancer therapies, the mechanisms that drive them are poorly understood. Here, we report that vascular endothelial cells (ECs) are more highly primed for apoptosis than parenchymal cells across many adult tissues. Consequently, ECs readily undergo apoptosis in response to many commonly used anticancer agents including cytotoxic and targeted drugs and are more sensitive to ionizing radiation and BH3 mimetics than parenchymal cells in vivo. Further, using differentiated isogenic human induced pluripotent stem cell models of ECs and vascular smooth muscle cells (VSMCs), we find that these vascular cells exhibit distinct drug toxicity patterns, which are linked to divergent therapy-induced vascular toxicities in patients. Collectively, our results demonstrate that vascular cells are highly sensitive to apoptosis-inducing stress across life span and may represent a "weakest link" vulnerability in multiple tissues for development of toxicities.

Differential Effects of Platelet Factor 4 (CXCL4) and Its Non-Allelic Variant (CXCL4L1) on Cultured Human Vascular Smooth Muscle Cells.

Platelet factor 4 (CXCL4) is a chemokine abundantly stored in platelets. Upon injury and during atherosclerosis, CXCL4 is transported through the vessel wall where it modulates the function of vascular smooth muscle cells (VSMCs) by affecting proliferation, migration, gene expression and cytokine release. Variant CXCL4L1 is distinct from CXCL4 in function and expression pattern, despite a minor three-amino acid difference. Here, the effects of CXCL4 and CXCL4L1 on the phenotype and function of human VSMCs were compared in vitro. VSMCs were found to constitutively express CXCL4L1 and only exogenously added CXCL4 was internalized by VSMCs. Pre-treatment with heparin completely blocked CXCL4 uptake. A role of the putative CXCL4 receptors CXCR3 and DARC in endocytosis was excluded, but LDL receptor family members appeared to be involved in the uptake of CXCL4. Incubation of VSMCs with both CXCL4 and CXCL4L1 resulted in decreased expression of contractile marker genes and increased mRNA levels of KLF4 and NLRP3 transcription factors, yet only CXCL4 stimulated proliferation and calcification of VSMCs. In conclusion, CXCL4 and CXCL4L1 both modulate gene expression, yet only CXCL4 increases the division rate and formation of calcium-phosphate crystals in VSMCs. CXCL4 and CXCL4L1 may play distinct roles during vascular remodeling in which CXCL4 induces proliferation and calcification while endogenously expressed CXCL4L1 governs cellular homeostasis. The latter notion remains a subject for future investigation.

Role of smooth muscle activation in the static and dynamic mechanical characterization of human aortas.

Experimental data and a suitable material model for human aortas with smooth muscle activation are not available in the literature despite the need for developing advanced grafts; the present study closes this gap. Mechanical characterization of human descending thoracic aortas was performed with and without vascular smooth muscle (VSM) activation. Specimens were taken from 13 heart-beating donors. The aortic segments were cooled in Belzer UW solution during transport and tested within a few hours after explantation. VSM activation was achieved through the use of potassium depolarization and noradrenaline as vasoactive agents. In addition to isometric activation experiments, the quasistatic passive and active stress-strain curves were obtained for circumferential and longitudinal strips of the aortic material. This characterization made it possible to create an original mechanical model of the active aortic material that accurately fits the experimental data. The dynamic mechanical characterization was executed using cyclic strain at different frequencies of physiological interest. An initial prestretch, which corresponded to the physiological conditions, was applied before cyclic loading. Dynamic tests made it possible to identify the differences in the viscoelastic behavior of the passive and active tissue. This work illustrates the importance of VSM activation for the static and dynamic mechanical response of human aortas. Most importantly, this study provides material data and a material model for the development of a future generation of active aortic grafts that mimic natural behavior and help regulate blood pressure.

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). Women with obesity are more prone to develop arterial stiffening leading to more frequent and severe CVD compared with men. As tissue transglutaminase (TG2) has been implicated in vascular stiffening, our goal herein was to determine the efficacy of cystamine, a nonspecific 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 wk starting at 4 wk of age. The second was fed a WD for the same 43 wk, whereas a third cohort was fed WD, but also received cystamine (216 mg/kg/day) in the drinking water during the last 8 wk 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- versus CD-fed mice, and reduced in WD + C versus 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 because of its TG2 inhibitory capacity.NEW & NOTEWORTHY This study evaluates the novel role of transglutaminase 2 (TG2) inhibition to directly treat vascular stiffness. Our data demonstrate that cystamine, a nonspecific TG2 inhibitor, improves vascular stiffness induced by a diet rich in fat, fructose, and salt. This research suggests that TG2 inhibition might bear therapeutic potential to reduce the disproportionate burden of cardiovascular disease in females in conditions of chronic overnutrition.

Mecanosensores en el cambio de fenotipo de la musculatura lisa vascular / Mechanosensors and phenotype changes in vascular smooth muscle

Resumen: Introducción: Las células de la musculatura lisa vascular (CMLV) se caracterizan por mantener cierto grado de desdiferenciación, variando su fenotipo entre el contráctil y el secretor, de acuerdo con las necesidades del tejido, y el contráctil predominante en condiciones fisiológicas. Cualquier alteración del estímulo mecánico, ya sea en el flujo sanguíneo o la tensión mecánica ejercida sobre las CMLV, conducen a cambios de su fenotipo y remodelamiento de la vasculatura, lo que puede constituir el punto de inflexión de varias patologías relevantes en la salud pública como, por ejemplo, la hipertensión arterial. Objetivo: Realizar una revisión sobre los mecanosensores y las vías transduccionales conocidas e implicadas en el cambio de fenotipo de las CMLV. Metodología: Se realizó una búsqueda sistemática en las bases de datos PubMed, Scopus, Google Académico y Scielo sobre la mantención y cambio de fenotipo de las células de la musculatura lisa vascular asociado principalmente a el estrés mecánico, la participación de los mecanosensores más relevantes y las vías de señalización involucrados en este proceso. Conclusión: Los mecanosensores implicados en el cambio de fenotipo de las CMLV contemplan principalmente receptores acoplados a proteína G, moléculas de adhesión y canales iónicos activados por estiramiento. Los estudios se han concentrado en la activación o inhibición de vías como las proteínas quinasas activadas por mitógenos (MAPK), la vía AKT, mTOR y factores transcripcionales que regulan la expresión de genes de diferenciación y/o desdiferenciación, como las miocardinas. Existen además otros receptores involucrados en la respuesta al estrés mecánico, como los receptores tirosina quinasas. A pesar de la importancia que reviste el conocimiento de los mecanosensores y las vías implicadas en el cambio de fenotipo de las CMLV, así como el papel que cumplen en el establecimiento de patologías vasculares, es aún escaso el conocimiento que se tiene sobre los mismos.

Abstract: Introduction: Vascular smooth muscle cells (VS- MCs) are characterized by maintaining a certain de- gree of dedifferentiation. VSMCs may vary their phenotype between contractile and secretory according to tissue needs. Under physiological conditions, the predominant phenotype is contractile. Any alteration of the mechanical stimulus, either in the blood flow or the mechanical stress exerted on the VSMCs, leads to changes in their phenotype and remodeling of the vasculature. These changes can constitute the turning point in several hypertension and other diseases relevant in public health. Objective: To review the main mechanosensor and transduction pathways involved changes in VSMCs phenotype. Methods: A systematic search of PubMed, Scopus, Google Scholar and Scielo databases was carried out to ascertain the state of the art regarding the maintenance and change of VSMCs phenotype mainly associated with mechanical stress. Additionally, the participation of the most relevant mechanosensors and the signaling pathways involved in this process are discussed. Conclusion: The mechanosensors involved in the change in VSMCs phenotype mainly contempla- te G-protein-coupled receptors, adhesion molecules, and stretch-activated ion channels. Studies have been focused on the activation or inhibition of MAPK, AKT, mTOR, pathways and transcriptional factors that regulate the expression of differentiation and/or des differentiation genes such as Myocardins. There are also other receptors involved in the response to mechanical stress such as the tyrosine kinases receptor. Although the importance of understanding mechanosensors, the signaling pathways involved in VSMC phenotype switching and their role in the establishment of vascular pathologies, knowledge about them is limited.

MicroRNA-137 Inhibited Hypoxia-Induced Proliferation of Pulmonary Artery Smooth Muscle Cells by Targeting Calpain-2.

The proliferation of pulmonary artery smooth muscle cells (PASMCs) is an important cause of pulmonary vascular remodeling in pulmonary hypertension (PH). It has been reported that miR-137 inhibits the proliferation of tumor cells. However, whether miR-137 is involved in PH remains unclear. In this study, male Sprague-Dawley rats were subjected to 10% O2 for 3 weeks to establish PH, and rat primary PASMCs were treated with hypoxia (3% O2) for 48 h to induce cell proliferation. The effect of miR-137 on PASMC proliferation and calpain-2 expression was assessed by transfecting miR-137 mimic and inhibitor. The effect of calpain-2 on PASMC proliferation was assessed by transfecting calpain-2 siRNA. The present study found for the first time that miR-137 was downregulated in pulmonary arteries of hypoxic PH rats and in hypoxia-treated PASMCs. miR-137 mimic inhibited hypoxia-induced PASMC proliferation and upregulation of calpain-2 expression in PASMCs. Furthermore, miR-137 inhibitor induced the proliferation of PASMCs under normoxia, and knockdown of calpain-2 mRNA by siRNA significantly inhibited hypoxia-induced proliferation of PASMCs. Our study demonstrated that hypoxia-induced downregulation of miR-137 expression promoted the proliferation of PASMCs by targeting calpain-2, thereby potentially resulting in pulmonary vascular remodeling in hypoxic PH.

Role of H2S in Regulation of Vascular Tone in Metabolic Disorders.

We studied the effect of the H2S donor (NaHS, 1-500 µM) on the contractile responses of isolated aortic smooth muscle segments from rats with metabolic syndrome induced by high-fat, high-carbohydrate diet. It was found that the vasorelaxing effect of NaHS (5-100 µM) decreased in under conditions of MS. The endothelial NO synthase inhibitor L-NAME (100 µM) suppressed the effect of NaHS, while cystathionine-gamma-lyase inhibitor PAG (100 µM) decreased the vasodilating effects of acetylcholine (0.1-100 µM). Application of endogenous NO precursor L-arginine (1 mM) potentiated in the effects of H2S donor NaHS. Thus, the contractile activity of vascular smooth muscles in metabolic syndrome is determined by not only the effect of H2S, but also the influence of NO.

Loss of GATA4 C-Terminus by p.S335X Mutation Modulates Coronary Artery Vascular Smooth Muscle Cell Phenotype.

Coronary artery disease (CAD) has been the leading cause of morbidity and mortality worldwide, and its pathogenesis is closely related with the proliferation and migration of vascular smooth muscle cell (VSMC). We previously reported a truncated GATA4 protein lacking C-terminus induced by p.S335X mutation in cardiomyocyte from ventricular septal defect (VSD) patients. However, it is still unclear whether GATA4 p.S335X mutation could influence the development of CAD. GATA4 wild-type (WT) and p.S335X mutant (MU) overexpression plasmids were constructed and transfected transiently into rat coronary artery smooth muscle cell (RCSMC) to observe the proliferative and migratory abilities by MTS and wound healing assay, respectively. PCR array was used to preliminarily detect the expression of phenotypic modulation-related genes, and QRT-PCR was then carried out to verify the screened differentially expressed genes (DEGs). The results showed that, when stimulated by fetal bovine serum (10%) for 24 h or tumor necrosis factor-α (10 or 30 ng/ml) for 10 or 24 h, deletion of GATA4 C-terminus by p.S335X mutation in GATA4 enhanced the proliferation of RCSMC, without alteration of the migration capability. Twelve DEGs, including Fas, Hbegf, Itga5, Aimp1, Cxcl1, Il15, Il2rg, Il7, Tnfsf10, Il1r1, Irak1, and Tlr3, were screened and identified as phenotypic modulation-related genes. Our data might be beneficial for further exploration regarding the mechanisms of GATA4 p.S335X mutation on the phenotypic modulation of coronary VSMC.

Myogenic autoregulation in bone marrow arterioles and in vivo intramedullary pressure in femora of conscious, female Long Evans rats.

OBJECTIVES: The ability to regulate skeletal blood flow is critical for the maintenance of bone. The myogenic response is essential for regulating tissue blood flow. Myogenic responsiveness in bone marrow arterioles has not yet been determined. Furthermore, the literature is disparate regarding intramedullary pressures (IMP) within bone. The purposes of this study were to (1) determine whether bone marrow arterioles have myogenic activity and (2) assess if the autoregulatory zone corresponds with IMP. Also, this study provides detailed methodology on dissecting and isolating bone marrow arterioles for functional assessment. METHODS: Experiment 1: Femoral shafts of female Long Evans rats were catheterized to assess in vivo IMP. Experiment 2: Bone marrow arterioles from female Long Evans rats were cannulated. Active and passive myogenic responses were determined. RESULTS: In vivo intramedullary pressure averaged 32 ± 3 mmHg, intramedullary pulse pressure averaged 5.28 ± 0.03 mmHg, and the mean maximal diameter and wall thickness of the bone marrow arterioles were 96 ± 7 µm and 18 ± 2 µm, respectively. An active myogenic response was observed and differed (p < .001) from the passive curve. CONCLUSION: Bone marrow arterioles have myogenic responsiveness and the autoregulatory zone corresponded with the range of IMP (15-51 mmHg) within the femoral diaphysis of conscious animals.

MOVAS Cells: A Versatile Cell Line for Studying Vascular Smooth Muscle Cell Cholesterol Metabolism.

Cholesterol metabolism is paramount to cells. Aberrations to cholesterol metabolism affects cholesterol homeostasis, which may impact the risk of several diseases. Recent evidence has suggested that vascular smooth muscle cell (VSMC) cholesterol metabolism may play a role in atherosclerosis. However, there is scant in vitro mechanistic data involving primary VSMC that directly tests how VSMC cholesterol metabolism may impact atherosclerosis. One reason for this lack of data is due to the impracticality of gene manipulation studies in primary VSMC, as cultured primary VSMC become senescent and lose their morphology rapidly. However, there are no immortalized VSMC lines known to be suitable for studying VSMC cholesterol metabolism. The purpose of this study was to determine whether MOVAS cells, a commercially available VSMC line, are suitable to use for studying VSMC cholesterol metabolism. Using immunoblotting and immunofluorescence, we showed that MOVAS cells express ABCA1, ABCG1, and SREBP-2. We also determined that MOVAS cells efflux cholesterol to apoAI and HDL, which indicates functionality of ABCA1/ABCG1. In serum-starved MOVAS cells, SREBP-2 target gene expression was increased, confirming SREBP-2 functionality. We detected miR-33a expression in MOVAS cells and determined this microRNA can silence ABCA1 and ABCG1 via identifying conserved miR-33a binding sites within ABCA1/ABCG1 3'UTR in MOVAS cells. We showed that cholesterol-loading MOVAS cells results in this cell line to transdifferentiate into a macrophage-like cell, which also occurs when VSMC accumulate cholesterol. Our characterization of MOVAS cells sufficiently demonstrates that they are suitable to use for studying VSMC cholesterol metabolism in the context of atherosclerosis.

Glabridin Relaxes Vascular Smooth Muscles by Activating BKCa Channels and Inhibiting Phosphodiesterase in Human Saphenous Vein.

The aim of the current study was to investigate the pharmacological activity of glabridin on the isolated human saphenous vein (SV) and explore the underlying mechanisms. Samples of patients' SVs were removed during bypass surgery, and 4-mm lengths of the vessels were placed in Krebs solution at +4°C and hung in an isolated organ bath to assess their contraction/relaxation responses. The contraction/relaxation responses were recorded to observe if the cyclic guanosine monophosphate (cGMP)/protein kinase G (PKG) pathway mediates the relaxant effect of glabridin after treatment with blockers like ODQ (a guanylate cyclase inhibitor), KT5823 (a PKG inhibitor), isobutylmethylxanthine [IBMX, a phosphodiesterase (PDE) inhibitor], and cantharidin [Cant, a myosin light-chain phosphatase (MLCP) inhibitor]. Moreover, nitric oxide (NO), cGMP, and PKG levels in SV tissues were determined by ELISA after incubation with glabridin, N(ω)-nitro-L-arginine methyl ester (L-Name, a NO synthetase inhibitor), phenylephrine (PE), ODQ, IBMX, and KT5823. The results showed that glabridin relaxed the vascular smooth muscle of human SV pretreated with PE in a dose-dependent manner, which was independent of the endothelium. The vasorelaxant effect of glabridin was only inhibited by iberiotoxin (IbTX), Cant, and KT5823. Glabridin increased cGMP and PKG levels in SV homogenates, whereas it did not alter the NO level. The enhancing effects of cGMP and PKG levels by glabridin were abolished by ODQ and KT5823. In conclusion, glabridin has a vasorelaxant effect, which is associated with the activation of BKCa channels and inhibition of PDE.

Vascular smooth muscle-derived Trpv1+ progenitors are a source of cold-induced thermogenic adipocytes.

Brown adipose tissue (BAT) and beige fat function in energy expenditure in part due to their role in thermoregulation, making these tissues attractive targets for treating obesity and metabolic disorders. While prolonged cold exposure promotes de novo recruitment of brown adipocytes, the exact sources of cold-induced thermogenic adipocytes are not completely understood. Here, we identify transient receptor potential cation channel subfamily V member 1 (Trpv1)+ vascular smooth muscle (VSM) cells as previously unidentified thermogenic adipocyte progenitors. Single-cell RNA sequencing analysis of interscapular brown adipose depots reveals, in addition to the previously known platelet-derived growth factor receptor (Pdgfr)α-expressing mesenchymal progenitors, a population of VSM-derived adipocyte progenitor cells (VSM-APC) expressing the temperature-sensitive cation channel Trpv1. We demonstrate that cold exposure induces the proliferation of Trpv1+ VSM-APCs and enahnces their differentiation to highly thermogenic adipocytes. Together, these findings illustrate the landscape of the thermogenic adipose niche at single-cell resolution and identify a new cellular origin for the development of brown and beige adipocytes.

BCL11B Regulates Arterial Stiffness and Related Target Organ Damage.

[Figure: see text].

Involvement of sarco/endoplasmic reticulum Ca2+-ATPase (SERCA) in mPRα (PAQR7)-mediated progesterone induction of vascular smooth muscle relaxation.

Progesterone acts directly on vascular smooth muscle cells (VSMCs) through activation of membrane progesterone receptor α (mPRα)-dependent signaling to rapidly decrease cytosolic Ca2+ concentrations and induce muscle relaxation. However, it is not known whether this progesterone action involves uptake of Ca2+ by the sarco/endoplasmic reticulum (SR) and increased sarco/endoplasmic reticulum Ca2+-ATPase (SERCA) activity. The present results show that treatment of cultured human VSMCs with progesterone and the selective mPR agonist Org OD-02-0 (OD 02-0) but not with the nuclear PR agonist R5020 increased SERCA protein expression, which was blocked by knockdown of mPRα with siRNA. Moreover, treatments with progesterone and OD 02-0, but not with R5020, increased phospholamban (PLB) phosphorylation, which would result in disinhibition of SERCA function. Progesterone and OD 02-0 significantly increased Ca2+ levels in the SR and caused VSMC relaxation. These effects were blocked by pretreatment with cyclopiazonic acid (CPA), a SERCA inhibitor, and by knockdown of SERCA2 with siRNA, suggesting that SERCA2 plays a critical role in progesterone induction of VSMC relaxation. Treatment with inhibitors of inhibitory G proteins (Gi, NF023), MAP kinase (AZD 6244), Akt/Pi3k (wortmannin), and a Rho activator (calpeptin) blocked the progesterone- and OD 02-0-induced increase in Ca2+ levels in the SR and SERCA expressions. These results suggest that the rapid effects of progesterone on cytosolic Ca2+ levels and relaxation of VSMCs through mPRα involve regulation of the functions of SERCA2 and PLB through Gi, MAP kinase, and Akt signaling pathways and downregulation of RhoA activity.NEW & NOTEWORTHY The rapid effects of progesterone on cytosolic Ca2+ levels and relaxation of VSMCs through mPRα involve regulation of the functions of SERCA2 and PLB through Gi, MAP kinase, and Akt signaling pathways and downregulation of RhoA activity.

Galectin-3 Modulates Macrophage Activation and Contributes Smooth Muscle Cells Apoptosis in Abdominal Aortic Aneurysm Pathogenesis.

Galectin-3 (Gal-3) is a 26-kDa lectin that regulates many aspects of inflammatory cell behavior. We assessed the hypothesis that increased levels of Gal-3 contribute to abdominal aortic aneurysm (AAA) progression by enhancing monocyte chemoattraction through macrophage activation. We analyzed the plasma levels of Gal-3 in 76 patients with AAA (AAA group) and 97 controls (CTL group) as well as in angiotensin II (Ang-II)-infused ApoE knockout mice. Additionally, conditioned media (CM) were used to polarize THP-1 monocyte to M1 macrophages with or without Gal-3 inhibition through small interfering RNA targeted deletion to investigate whether Gal-3 inhibition could attenuate macrophage-induced inflammation and smooth muscle cell (SMC) apoptosis. Our results showed a markedly increased expression of Gal-3 in the plasma and aorta in the AAA patients and experimental mice compared with the CTL group. An in vitro study demonstrated that the M1 cells exhibited increased Gal-3 expression. Gal-3 inhibition markedly decreased the quantity of macrophage-induced inflammatory regulators, including IL-8, TNF-α, and IL-1ß, as well as messenger RNA expression and MMP-9 activity. Moreover, Gal-3-deficient CM weakened SMC apoptosis through Fas activation. These findings prove that Gal-3 may contribute to AAA progression by the activation of inflammatory macrophages, thereby promoting SMC apoptosis.

Impact of the menstrual cycle on peripheral vascular function in premenopausal women: systematic review and meta-analysis.

Fluctuations in endogenous hormones estrogen and progesterone during the menstrual cycle may offer vasoprotection for endothelial and smooth muscle (VSM) function. While numerous studies have been published, the results are conflicting, leaving our understanding of the impact of the menstrual cycle on vascular function unclear. The purpose of this systematic review and meta-analysis was to consolidate available research exploring the role of the menstrual cycle on peripheral vascular function. A systematic search of MEDLINE, Web of Science, and EMBASE was performed for articles evaluating peripheral endothelial and VSM function across the natural menstrual cycle: early follicular (EF) phase versus late follicular (LF), early luteal, mid luteal, or late luteal. A meta-analysis examined the effect of the menstrual cycle on the standardized mean difference (SMD) of the outcome measures. Analysis from 30 studies (n = 1,363 women) observed a "very low" certainty of evidence that endothelial function increased in the LF phase (SMD: 0.45, P = 0.0001), with differences observed in the macrovasculature but not in the microvasculature (SMD: 0.57, P = 0.0003, I2 = 84%; SMD: 0.21, P = 0.17, I2 = 34%, respectively). However, these results are partially explained by differences in flow-mediated dilation [e.g., discrete (SMD: 0.86, P = 0.001) vs. continuous peak diameter assessment (SMD: 0.25, P = 0.30)] and/or menstrual cycle phase methodologies. There was a "very low" certainty that endothelial function was largely unchanged in the luteal phases, and VSM was unchanged across the cycle. The menstrual cycle appears to have a small effect on macrovascular endothelial function but not on microvascular or VSM function; however, these results can be partially attributed to methodological differences.

TRPP2 and STIM1 form a microdomain to regulate store-operated Ca2+ entry and blood vessel tone.

BACKGROUND: Polycystin-2 (TRPP2) is a Ca2+ permeable nonselective cationic channel essential for maintaining physiological function in live cells. Stromal interaction molecule 1 (STIM1) is an important Ca2+ sensor in store-operated Ca2+ entry (SOCE). Both TRPP2 and STIM1 are expressed in endoplasmic reticular membrane and participate in Ca2+ signaling, suggesting a physical interaction and functional synergism. METHODS: We performed co-localization, co-immunoprecipitation, and fluorescence resonance energy transfer assay to identify the interactions of TRPP2 and STIM1 in transfected HEK293 cells and native vascular smooth muscle cells (VSMCs). The function of the TRPP2-STIM1 complex in thapsigargin (TG) or adenosine triphosphate (ATP)-induced SOCE was explored using specific small interfering RNA (siRNA). Further, we created TRPP2 conditional knockout (CKO) mouse to investigate the functional role of TRPP2 in agonist-induced vessel contraction. RESULTS: TRPP2 and STIM1 form a complex in transfected HEK293 cells and native VSMCs. Genetic manipulations with TRPP2 siRNA, dominant negative TRPP2 or STIM1 siRNA significantly suppressed ATP and TG-induced intracellular Ca2+ release and SOCE in HEK293 cells. Inositol triphosphate receptor inhibitor 2-aminoethyl diphenylborinate (2APB) abolished ATP-induced Ca2+ release and SOCE in HEK293 cells. In addition, TRPP2 and STIM1 knockdown significantly inhibited ATP- and TG-induced STIM1 puncta formation and SOCE in VSMCs. Importantly, knockdown of TRPP2 and STIM1 or conditional knockout TRPP2 markedly suppressed agonist-induced mouse aorta contraction. CONCLUSIONS: Our data indicate that TRPP2 and STIM1 are physically associated and form a functional complex to regulate agonist-induced intracellular Ca2+ mobilization, SOCE and blood vessel tone. Video abstract.

Hsa_circ_0029589 knockdown inhibits the proliferation, migration and invasion of vascular smooth muscle cells via regulating miR-214-3p and STIM1.

AIMS: Circular RNAs (circRNAs) are relevant to atherosclerosis progression. However, the role and mechanism of circRNA hsa_circ_0029589 (circ_0029589) in atherosclerosis are not fully understood. This research aims to explore the function and mechanism of circ_0029589 in oxidized low-density lipoprotein (ox-LDL)-caused vascular smooth muscle cells (VSMCs) injury in vitro. MAIN METHODS: VSMCs were challenged via ox-LDL to mimic atherosclerosis-like injury in vitro. Circ_0029589, microRNA-214-3p (miR-214-3p) and stromal interaction molecule 1 (STIM1) abundances were detected via quantitative reverse transcription polymerase chain reaction or western blot. Cell proliferation was investigated via cell viability, cycle, apoptosis and proliferation-associated protein levels. Cell migration and invasion were assessed via transwell analysis. The relationship between miR-214-3p and circ_0029589 or STIM1 was tested via dual-luciferase reporter analysis and RNA immunoprecipitation. KEY FINDINGS: Circ_0029589 level was enhanced in ox-LDL-challenged VSMCs. Circ_0029589 interference constrained cell proliferation, migration and invasion in ox-LDL-challenged VSMCs. miR-214-3p was targeted by circ_0029589 and miR-214-3p knockdown weakened the suppressive function of circ_0029589 silence on VSMCs proliferation, migration and invasion. STIM1 was targeted via miR-214-3p and miR-214-3p could suppress VSMCs proliferation, migration and invasion via decreasing STIM1. Moreover, circ_0029589 modulated STIM1 level by miR-214-3p. SIGNIFICANCE: Circ_0029589 knockdown repressed proliferation, migration and invasion of VSMCs challenged via ox-LDL by regulating miR-214-3p and STIM1, indicating that circ_0029589 might play important role in atherosclerosis.