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BACKGROUND: The N6-methyladenosine (m6A) modification of RNA and its regulators have important roles in the pathogenesis of pulmonary hypertension (PH). Ythdf2 (YTH N6-methyladenosine RNA binding protein 2) is best known for its role in degrading m6A-modified mRNAs such as Hmox1 mRNA, which leads to alternative activation of macrophages in PH. Recent studies have also linked Ythdf2 to the proliferation of pulmonary artery smooth muscle cells (PASMCs). However, its specific roles in PASMCs and downstream targets during the development of PH remain unclear. METHODS: The expression and biological function of Ythdf2 in PASMCs were investigated in human and experimental models of PH. Smooth muscle cell-specific Ythdf2-deficient mice were used to assess the roles of Ythdf2 in PASMCs in vivo. Proteomic analysis, m6A sequencing, and RNA immunoprecipitation analysis were used to screen for potential downstream targets. RESULTS: Ythdf2 was significantly upregulated in human and rodent PH-PASMCs, and smooth muscle cell-specific Ythdf2 deficiency ameliorated PASMC proliferation, right ventricular hypertrophy, pulmonary vascular remodeling, and PH development. Higher expression of Ythdf2 promoted PASMC proliferation and PH by paradoxically stabilizing Myadm mRNA in an m6A-dependent manner. Loss of Ythdf2 decreased the expression of Myadm in PASMCs and pulmonary arteries, both in vitro and in vivo. Additionally, silencing Myadm inhibited the Ythdf2-dependent hyperproliferation of PASMCs by upregulating the cell cycle kinase inhibitor p21. CONCLUSIONS: We have identified a novel mechanism where the increased expression of Ythdf2 stimulates PH-PASMC proliferation through an m6A/Myadm/p21 pathway. Strategies targeting Ythdf2 in PASMCs might be useful additions to the therapeutic approach to PH. [ABSTRACT FROM AUTHOR]

Objective To explore the pathological features of lung tissue in mild chronic obstructive pulmonary disease (COPD) and their association with inflammatory factors. Methods A total of 70 patients who underwent surgery for small lung nodule were prospectively included, and were divided into the normal group (n=10), the mild COPD group (n=50) and the moderate and severe COPD group (n=10). The pathological changes of lung tissue were evaluated after HE, Masson and EVG staining. The expression levels of SMA, Actin and CD31 proteins were detected by immunohistochemistry staining. Tumor necrosis factor α (TNF-α), interleukin-10 (IL-10) protein and mRNA levels were detected by immunohistochemistry and qPCR. Results Pulmonary tissue in mild COPD showed widening of alveolar septum, dilation of small airways, mild thickening of blood vessel wall and inflammatory reaction dominated by lymphocyte infiltration. Immunohistochemistry staining showed that contents of SMA and Actin proteins in mild COPD lung tissue were higher than those in the normal group (P＜0.05). In addition, the TNF- α mRNA and the positive rate of TNF- α in lung tissue of mild COPD were significantly higher than those in the normal group, while the IL-10 mRNA was significantly lower than that of the normal group (all P＜0.05). SMA and Actin were positively correlated with the positive expression of inflammatory cytokine TNF-α, but negatively correlated with the positive expression of IL-10 (all P＜0.05). Conclusion The main pathological changes of lung tissue in mild COPD include small lung blood vessel remodeling ocharacterized by thickening of small blood vessel smooth muscle layer and lymphocyte-dominated inflammatory response, while the increase of pro-inflammatory factor TNF-α and decrease of anti-inflammatory factor IL-10 are associated with pathological changes of COPD. [ABSTRACT FROM AUTHOR]

BACKGROUND: Abdominal aortic aneurysm (AAA) is a potentially life-threatening vascular condition, but approved medical therapies to prevent AAA progression and rupture are currently lacking. Sphingolipid metabolism disorders are associated with the occurrence and development of AAA. It has been discovered that ganglioside GM3, a sialic acid–containing type of glycosphingolipid, plays a protective role in atherosclerosis, which is an important risk factor for AAA; however, the potential contribution of GM3 to AAA development has not been investigated. METHODS: We performed a metabolomics study to evaluated GM3 level in plasma of human patients with AAA. We profiled GM3 synthase (ST3GAL5) expression in the mouse model of aneurysm and human AAA tissues through Western blotting and immunofluorescence staining. RNA sequencing, affinity purification and mass spectrometry, proteomic analysis, surface plasmon resonance analysis, and functional studies were used to dissect the molecular mechanism of GM3-regulating ferroptosis. We conditionally deleted and overexpressed St3gal5 in smooth muscle cells (SMCs) in vivo to investigate its role in AAA. RESULTS: We found significantly reduced plasma levels of GM3 in human patients with AAA. GM3 content and ST3GAL5 expression were decreased in abdominal aortic vascular SMCs in patients with AAA and an AAA mouse model. RNA sequencing analysis showed that ST3GAL5 silencing in human aortic SMCs induced ferroptosis. We showed that GM3 interacted directly with the extracellular domain of TFR1 (transferrin receptor 1), a cell membrane protein critical for cellular iron uptake, and disrupted its interaction with holo-transferrin. SMC-specific St3gal5 knockout exacerbated iron accumulation at lesion sites and significantly promoted AAA development in mice, whereas GM3 supplementation suppressed lipid peroxidation, reduced iron deposition in aortic vascular SMCs, and markedly decreased AAA incidence. CONCLUSIONS: Together, these results suggest that GM3 dysregulation promotes ferroptosis of vascular SMCs in AAA. Furthermore, GM3 may constitute a new therapeutic target for AAA. [ABSTRACT FROM AUTHOR]

Potassium ion (K+) channel is a transmembrane protein located on cell membrane. The K+ channels of vascular smooth muscle cells play an important role in regulating vascular tension, cell excitability and proliferation through membrane potential. The dysfunction of K+ channels in pulmonary artery smooth muscle cells (PASMCs) is closely related to the pathological process of hypoxic pulmonary hypertension (HPH), and K+ channels are expected to become the therapeutic target of HPH. In this artical, types of K+ channels in PASMCs, the research progress of K+ channels in HPH and drugs that interfere with HPH were reviewed, in order to provide new ideas for the pathogenesis research and drug development of HPH. [ABSTRACT FROM AUTHOR]

Background: Activation of vascular smooth muscle cell (VSMC) inflammation is vital to initiate vascular disease. The role of human-specific long noncoding RNAs in VSMC inflammation is poorly understood. Methods: Bulk RNA sequencing in differentiated human VSMCs revealed a novel human-specific long noncoding RNA called inflammatory MKL1 (megakaryoblastic leukemia 1) interacting long noncoding RNA (INKILN). INKILN expression was assessed in multiple in vitro and ex vivo models of VSMC phenotypic modulation as well as human atherosclerosis and abdominal aortic aneurysm. The transcriptional regulation of INKILN was verified through luciferase reporter and chromatin immunoprecipitation assays. Loss-of-function and gain-of-function studies and multiple RNA–protein and protein–protein interaction assays were used to uncover a mechanistic role of INKILN in the VSMC proinflammatory gene program. Bacterial artificial chromosome transgenic mice were used to study INKILN expression and function in ligation injury–induced neointimal formation. Results: INKILN expression is downregulated in contractile VSMCs and induced in human atherosclerosis and abdominal aortic aneurysm. INKILN is transcriptionally activated by the p65 pathway, partially through a predicted NF-κB (nuclear factor kappa B) site within its proximal promoter. INKILN activates proinflammatory gene expression in cultured human VSMCs and ex vivo cultured vessels. INKILN physically interacts with and stabilizes MKL1, a key activator of VSMC inflammation through the p65/NF-κB pathway. INKILN depletion blocks interleukin-1β–induced nuclear localization of both p65 and MKL1. Knockdown of INKILN abolishes the physical interaction between p65 and MKL1 and the luciferase activity of an NF-κB reporter. Furthermore, INKILN knockdown enhances MKL1 ubiquitination through reduced physical interaction with the deubiquitinating enzyme USP10 (ubiquitin-specific peptidase 10). INKILN is induced in injured carotid arteries and exacerbates ligation injury–induced neointimal formation in bacterial artificial chromosome transgenic mice. Conclusions: These findings elucidate an important pathway of VSMC inflammation involving an INKILN /MKL1/USP10 regulatory axis. Human bacterial artificial chromosome transgenic mice offer a novel and physiologically relevant approach for investigating human-specific long noncoding RNAs under vascular disease conditions. [ABSTRACT FROM AUTHOR]

Background: Vascular MR (mineralocorticoid receptor) expression increases with age driving aging-associated vascular stiffness and hypertension. MR has two isoforms (1α and 1β) with distinct 5'-untranslated and promoter sequences (P1 and P2), but the gene regulatory mechanisms remain unknown. We investigated mechanisms driving MR gene transcriptional regulation in aging human smooth muscle cells (SMC).Methods: MR was quantified in aortic tissue and primary human aortic SMC (HASMC) comparing adult and aged donors and adult HASMC treated with H2O2, to induce aging. Predicted transcription factor (TF) binding sites in the MR gene were validated using chromatin immunoprecipitations and reporter assays. The impact of TF inhibitors on MR isoforms and fibrosis gene target was examined.Results: Expression of both MR mRNA isoforms increased with donor age or H2O2 treatment in HASMCs. HIF1α (hypoxia-inducible factor) and the inflammatory TF NFκB (nuclear factor kappa B) both increased with age in HASMCs and are predicted to bind MR promoters. H2O2 induced HIF1α and NFκB expression and DNA binding of HIF1α to the MR P1 promoter and of NFκB to both MR promoters in HASMCs. HIF1α inhibition decreased MR-1α isoform expression while NFκB inhibition decreased both MR isoforms. HIF1α, NFκB, and MR inhibition decreased the expression of a SMC-MR target gene implicated in vascular fibrosis. In human aortic tissues, expression of HIF1α and NFκB each positively correlated with donor age and MR expression (P<0.0001).Conclusions: These data implicate the inflammatory TF, NFκB, and oxidative stress-induced TF, HIF1α, in regulating SMC MR transcription in aging HASMCs, which drives aging-related vascular stiffness and cardiovascular disease. [ABSTRACT FROM AUTHOR]

Background: The long isoform of the Wnk1 (with-no-lysine [K] kinase 1) is a ubiquitous serine/threonine kinase, but its role in vascular smooth muscle cells (VSMCs) pathophysiology remains unknown., Methods: AngII (angiotensin II) was infused in Apoe -/- to induce experimental aortic aneurysm. Mice carrying an Sm22-Cre allele were cross-bred with mice carrying a floxed Wnk1 allele to specifically investigate the functional role of Wnk1 in VSMCs., Results: Single-cell RNA-sequencing of the aneurysmal abdominal aorta from AngII-infused Apoe -/- mice revealed that VSMCs that did not express Wnk1 showed lower expression of contractile phenotype markers and increased inflammatory activity. Interestingly, WNK1 gene expression in VSMCs was decreased in human abdominal aortic aneurysm. Wnk1 -deficient VSMCs lost their contractile function and exhibited a proinflammatory phenotype, characterized by the production of matrix metalloproteases, as well as cytokines and chemokines, which contributed to local accumulation of inflammatory macrophages, Ly6C hi monocytes, and γδ T cells. Sm22Cre+Wnk1 lox/lox mice spontaneously developed aortitis in the infrarenal abdominal aorta, which extended to the thoracic area over time without any negative effect on long-term survival. AngII infusion in Sm22Cre+Wnk1 lox/lox mice aggravated the aortic disease, with the formation of lethal abdominal aortic aneurysms. Pharmacological blockade of γδ T-cell recruitment using neutralizing anti-CXCL9 (anti-CXC motif chemokine ligand 9) antibody treatment, or of monocyte/macrophage using Ki20227, a selective inhibitor of CSF1 receptor, attenuated aortitis. Wnk1 deletion in VSMCs led to aortic wall remodeling with destruction of elastin layers, increased collagen content, and enhanced local TGF-β (transforming growth factor-beta) 1 expression. Finally, in vivo TGF-β blockade using neutralizing anti-TGF-β antibody promoted saccular aneurysm formation and aorta rupture in Sm22 Cre+ Wnk1 lox/lox mice but not in control animals., Conclusion: Wnk1 is a key regulator of VSMC function. Wnk1 deletion promotes VSMC phenotype switch toward a pathogenic proinflammatory phenotype, orchestrating deleterious vascular remodeling and spontaneous severe aortitis in mice., Competing Interests: None.

Background: The N6-methyladenosine (m 6 A) modification of RNA and its regulators have important roles in the pathogenesis of pulmonary hypertension (PH). Ythdf2 (YTH N6-methyladenosine RNA binding protein 2) is best known for its role in degrading m 6 A-modified mRNAs such as Hmox1 mRNA, which leads to alternative activation of macrophages in PH. Recent studies have also linked Ythdf2 to the proliferation of pulmonary artery smooth muscle cells (PASMCs). However, its specific roles in PASMCs and downstream targets during the development of PH remain unclear., Methods: The expression and biological function of Ythdf2 in PASMCs were investigated in human and experimental models of PH. Smooth muscle cell-specific Ythdf2 -deficient mice were used to assess the roles of Ythdf2 in PASMCs in vivo. Proteomic analysis, m 6 A sequencing, and RNA immunoprecipitation analysis were used to screen for potential downstream targets., Results: Ythdf2 was significantly upregulated in human and rodent PH-PASMCs, and smooth muscle cell-specific Ythdf2 deficiency ameliorated PASMC proliferation, right ventricular hypertrophy, pulmonary vascular remodeling, and PH development. Higher expression of Ythdf2 promoted PASMC proliferation and PH by paradoxically stabilizing Myadm mRNA in an m 6 A-dependent manner. Loss of Ythdf2 decreased the expression of Myadm in PASMCs and pulmonary arteries, both in vitro and in vivo. Additionally, silencing Myadm inhibited the Ythdf2 -dependent hyperproliferation of PASMCs by upregulating the cell cycle kinase inhibitor p21., Conclusions: We have identified a novel mechanism where the increased expression of Ythdf2 stimulates PH-PASMC proliferation through an m 6 A/Myadm/p21 pathway. Strategies targeting Ythdf2 in PASMCs might be useful additions to the therapeutic approach to PH., Competing Interests: None.

Background: Atherosclerosis is driven by the infiltration of the arterial intima by diverse immune cells and smooth muscle cells (SMCs). CD8 + T cells promote lesion growth during atherosclerotic lesion development, but their role in advanced atherosclerosis is less clear. Here, we studied the role of CD8 + T cells and their effects on SMCs in established atherosclerosis., Methods: CD8 + T cells were depleted in (SMC reporter) low-density lipoprotein receptor-deficient ( Ldlr -/- ) mice with established atherosclerotic lesions. Atherosclerotic lesion formation was examined, and single-cell RNA sequencing of aortic SMCs and their progeny was performed. Additionally, coculture experiments with primary aortic SMCs and CD8 + T cells were conducted., Results: Although we could not detect differences in atherosclerotic lesion size, an increased plaque SMC content was noted in mice after CD8 + T-cell depletion. Single-cell RNA sequencing of aortic lineage-traced SMCs revealed contractile SMCs and a modulated SMC cluster, expressing macrophage- and osteoblast-related genes. CD8 + T-cell depletion was associated with an increased contractile but decreased macrophage and osteoblast-like gene signature in this modulated aortic SMC cluster. Conversely, exposure of isolated aortic SMCs to activated CD8 + T cells decreased the expression of genes indicative of a contractile SMC phenotype and induced a macrophage and osteoblast-like cell state. Notably, CD8 + T cells triggered calcium deposits in SMCs under osteogenic conditions. Mechanistically, we identified transcription factors highly expressed in modulated SMCs, including Runx1 , to be induced by CD8 + T cells in cultured SMCs in an IFNγ (interferon-γ)-dependent manner., Conclusions: We here uncovered CD8 + T cells to control the SMC phenotype in atherosclerosis. CD8 + T cells promote SMC dedifferentiation and drive SMCs to adopt features of macrophage-like and osteoblast-like, procalcifying cell phenotypes. Given the critical role of SMCs in atherosclerotic plaque stability, CD8 + T cells could thus be explored as therapeutic target cells during lesion progression., Competing Interests: None.

Background: Abdominal aortic aneurysm (AAA) is a severe aortic disease without effective pharmacological approaches. The nuclear hormone receptor LXRα (liver X receptor α), encoded by the NR1H3 gene, serves as a critical transcriptional mediator linked to several vascular pathologies, but its role in AAA remains elusive., Methods: Through integrated analyses of human and murine AAA gene expression microarray data sets, we identified NR1H3 as a candidate gene regulating AAA formation. To investigate the role of LXRα in AAA formation, we used global Nr1h3 -knockout and vascular smooth muscle cell-specific Nr1h3 -knockout mice in 2 AAA mouse models induced with angiotensin II (1000 ng·kg·min; 28 days) or calcium chloride (CaCl 2 ; 0.5 mol/L; 42 days)., Results: Upregulated LXRα was observed in the aortas of patients with AAA and in angiotensin II- or CaCl 2 -treated mice. Global or vascular smooth muscle cell-specific Nr1h3 knockout inhibited AAA formation in 2 mouse models. Loss of LXRα function prevented extracellular matrix degeneration, inflammation, and vascular smooth muscle cell phenotypic switching. Uhrf1 , an epigenetic master regulator, was identified as a direct target gene of LXRα by integrated analysis of transcriptome sequencing and chromatin immunoprecipitation sequencing. Susceptibility to AAA development was consistently enhanced by UHRF1 (ubiquitin-like containing PHD and RING finger domains 1) in both angiotensin II- and CaCl 2 -induced mouse models. We then determined the CpG methylation status and promoter accessibility of UHRF1-mediated genes using CUT&Tag (cleavage under targets and tagmentation), RRBS (reduced representation bisulfite sequencing), and ATAC-seq (assay for transposase-accessible chromatin with sequencing) in vascular smooth muscle cells, which revealed that the recruitment of UHRF1 to the promoter of miR-26b led to DNA hypermethylation accompanied by relatively closed chromatin states, and caused downregulation of miR-26b expression in AAA. Regarding clinical significance, we found that underexpression of miR-26b-3p correlated with high risk in patients with AAA. Maintaining miR-26b-3p expression prevented AAA progression and alleviated the overall pathological process., Conclusions: Our study reveals a pivotal role of the LXRα/UHRF1/miR-26b-3p axis in AAA and provides potential biomarkers and therapeutic targets for AAA., Competing Interests: Disclosures None.

Competing Interests: Disclosures P. Maffia reports consulting fees for Orion Biotechnology.

The implementation of human induced pluripotent stem cell (hiPSC) models has introduced an additional tool for identifying molecular mechanisms of disease that complement animal models. Patient-derived or CRISPR/Cas9-edited induced pluripotent stem cells differentiated into smooth muscle cells (SMCs) have been leveraged to discover novel mechanisms, screen potential therapeutic strategies, and model in vivo development. The field has evolved over almost 15 years of research using hiPSC-SMCs and has made significant strides toward overcoming initial challenges such as the lineage specificity of SMC phenotypes. However, challenges both specific (eg, the lack of specific markers to thoroughly validate hiPSC-SMCs) and general (eg, a lack of transparency and consensus around methodology in the field) remain. In this review, we highlight the recent successes and remaining challenges of the hiPSC-SMC model., Competing Interests: Disclosures None.

Objective: Rare disease Background: Uterine lipoleiomyoma is a rare benign tumor composed of varying proportions of smooth muscle fibers and mature adipocytes, without identification of hemorrhage, necrosis, or cyto-architectural atypia. It is part of the leiomyomas category, with an incidence of 0.03-0.2%. The pathogenesis of this lesional category is still unclear, but there are several theories that could explain the occurrence. Magnetic resonance imaging is the most useful diagnostic imaging method. There are a number of pathologies whose exclusion is necessary, with the differential diagnosis being made mainly based on microscopic examination and completed with immunohisto-chemical tests. Their treatment, when necessary, is surgical, with an excellent post-therapeutic evolution and prognosis. Case Report: We present the case of an elderly, postmenopausal patient who presented with abdominal pain, abnormal vaginal bleeding, and pollakiuria. The associated pathologies of the patient correspond to those mentioned in the literature, the particularities of the case being given by the large size of the tumor and the association with 2 other typical leiomyomas. Immunohistochemical markers used to exclude other diagnoses (desmin, h-caldesmon, S100, calretinin, MDM2, CD34) confirmed the diagnosis of uterine lipoleiomyoma. Because the patient was symptomatic and a large nodular mass was identified by ultrasound, surgical treatment was performed. Conclusions: Although it is a benign lesion with an excellent prognosis, the pathogenetic mechanisms are not fully known. Theories of pathogenesis range from misplacing embryonic adipocytes to connective tissue fatty degeneration, and further studies are needed to establish the origin of this lesion. [ABSTRACT FROM AUTHOR]

Ethnopharmacological Relevance: Tea (Camellia sinensis) is a favorite drink worldwide. Tea extracts and green tea main component (-)-epigallocatechin gallate (EGCG) are recommended for various vascular diseases. Anji white tea is a very popular green tea. Its vascular effect profile, the mechanisms, and the contribution of EGCG to its integrated effect need elucidation., Aim: To characterize the vasomotion effects of Anji white tea and EGCG, and to explore possible involvement of voltage-gated Ca 2+ channels (VGCCs) and voltage-gated K + (Kv) channels in their vasomotion effects., Materials and Methods: Anji white tea water soaking solution (AJWT) was prepared as daily tea-making process and concentrated to a concentration amounting to 200 mg/ml of dry tea leaves. The tension of rat arteries including aorta, coronary artery (RCA), cerebral basilar artery (CBA), intrarenal artery (IRA), intrapulmonary artery (IPA) and mesenteric artery (MA) was recorded with myographs. In arterial smooth muscle cells (ASMCs) freshly isolated from RCA, the levels of intracellular Ca 2+ were measured with Ca 2+ -sensitive fluorescent probe fluo 4-AM, and Kv currents were recorded with patch clamp. The expressions of VGCCs and Kv channels were assayed with RT-qPCR and immunofluorescence staining., Results: At 0.4-12.8 mg/ml of dry tea leaves, AJWT profoundly relaxed all tested arteries precontracted with various vasoconstrictors about half with a small transient potentiation on the precontractions before the relaxation. KCl-induced precontraction was less sensitive than precontractions induced by phenylephrine (PE), U46619 and serotonin (5-HT). IPA was less sensitive to the relaxation compared with other arteries. AJWT pretreatment for 1 h, 24 h and 72 h time-dependently inhibited the contractile responses of RCAs. In sharp contrast, at equivalent concentrations according to its content in AJWT, EGCG intensified the precontractions in most small arteries, except that it induced relaxation in PE-precontracted aorta and MA, U46619-precontracted aorta and CBA. EGCG pretreatment for 1 h and 24 h did not significantly affect RCA contractile responses. In RCA ASMCs, AJWT reduced, while EGCG enhanced, intracellular Ca 2+ elevation induced by depolarization which activates VGCCs. Patch clamp study showed that both AJWT and EGCG reduced Kv currents. RT-qPCR and immunofluorescence staining demonstrated that both AJWT and EGCG reduced the expressions of VGCCs and Kv channels., Conclusion: AJWT, but not EGCG, consistently induces vasorelaxation. The vasomotion effects of either AJWT or EGCG vary with arterial beds and vasoconstrictors. Modulation of VGCCs, but not Kv channels, contributes to AJWT-induced vasorelaxation. It is suggested that Anji white tea water extract instead of EGCG may be a promising food supplement for vasospastic diseases., Competing Interests: Declaration of competing interest We certify that there is no conflict of interest regarding this manuscript., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Competing Interests: Disclosures None.

Background: Pulmonary hypertension (PH) is a progressive and life-threatening disease characterized by pulmonary vascular remodeling, which involves aberrant proliferation and apoptosis resistance of the pulmonary arterial smooth muscle cells (PASMCs), resembling the hallmark characteristics of cancer. In cancer, the HMGB2 (high-mobility group box 2) protein promotes the pro-proliferative/antiapoptotic phenotype. However, the function of HMGB2 in PH remains uninvestigated., Methods: Smooth muscle cell (SMC)-specific HMGB2 knockout or HMGB2-OE (HMGB2 overexpression) mice and HMGB2 silenced rats were used to establish hypoxia+Su5416 (HySu)-induced PH mouse and monocrotaline-induced PH rat models, respectively. The effects of HMGB2 and its underlying mechanisms were subsequently elucidated using RNA-sequencing and cellular and molecular biology analyses. Serum HMGB2 levels were measured in the controls and patients with pulmonary arterial (PA) hypertension., Results: HMGB2 expression was markedly increased in the PAs of patients with PA hypertension and PH rodent models and was predominantly localized in PASMCs. SMC-specific HMGB2 deficiency or silencing attenuated PH development and pulmonary vascular remodeling in hypoxia+Su5416-induced mice and monocrotaline-treated rats. SMC-specific HMGB2 overexpression aggravated hypoxia+Su5416-induced PH. HMGB2 knockdown inhibited PASMC proliferation in vitro in response to PDGF-BB (platelet-derived growth factor-BB). In contrast, HMGB2 protein stimulation caused the hyperproliferation of PASMCs. In addition, HMGB2 promoted PASMC proliferation and the development of PH by RAGE (receptor for advanced glycation end products)/FAK (focal adhesion kinase)-mediated Hippo/YAP (yes-associated protein) signaling suppression. Serum HMGB2 levels were significantly increased in patients with PA hypertension, and they correlated with disease severity, predicting worse survival., Conclusions: Our findings indicate that targeting HMGB2 might be a novel therapeutic strategy for treating PH. Serum HMGB2 levels could serve as a novel biomarker for diagnosing PA hypertension and determining its prognosis., Competing Interests: Disclosures None.

Background: Thromboembolic events secondary to rupture or erosion of advanced atherosclerotic lesions is the global leading cause of death. The most common and effective means to reduce these major adverse cardiovascular events, including myocardial infarction and stroke, is aggressive lipid lowering via a combination of drugs and dietary modifications. However, we know little regarding the effects of reducing dietary lipids on the composition and stability of advanced atherosclerotic lesions, the mechanisms that regulate these processes, and what therapeutic approaches might augment the benefits of lipid lowering., Methods: Smooth muscle cell lineage-tracing Apoe -/- mice were fed a high-cholesterol Western diet for 18 weeks and then a zero-cholesterol standard laboratory diet for 12 weeks before treating them with an IL (interleukin)-1β or control antibody for 8 weeks. We assessed lesion size and remodeling indices, as well as the cellular composition of aortic and brachiocephalic artery lesions, indices of plaque stability, overall plaque burden, and phenotypic transitions of smooth muscle cell and other lesion cells by smooth muscle cell lineage tracing combined with single-cell RNA sequencing, cytometry by time-of-flight, and immunostaining plus high-resolution confocal microscopic z-stack analysis., Results: Lipid lowering by switching Apoe -/- mice from a Western diet to a standard laboratory diet reduced LDL cholesterol levels by 70% and resulted in multiple beneficial effects including reduced overall aortic plaque burden, as well as reduced intraplaque hemorrhage and necrotic core area. However, contrary to expectations, IL-1β antibody treatment after diet-induced reductions in lipids resulted in multiple detrimental changes including increased plaque burden and brachiocephalic artery lesion size, as well as increasedintraplaque hemorrhage, necrotic core area, and senescence as compared with IgG control antibody-treated mice. Furthermore, IL-1β antibody treatment upregulated neutrophil degranulation pathways but downregulated smooth muscle cell extracellular matrix pathways likely important for the protective fibrous cap., Conclusions: Taken together, IL-1β appears to be required for the maintenance of standard laboratory diet-induced reductions in plaque burden and increases in multiple indices of plaque stability., Competing Interests: Disclosures None.

Vascular calcification is an important factor in the high morbidity and mortality of Cardiovascular and cerebrovascular diseases. Vascular damage caused by calcification of the intima or media impairs the physiological function of the vascular wall. Inflammation is a central factor in the development of vascular calcification. Macrophages are the main inflammatory cells. Dynamic changes of macrophages with different phenotypes play an important role in the occurrence, progression and stability of calcification. This review focuses on macrophage polarization and the relationship between macrophages of different phenotypes and calcification environment, as well as the mechanism of interaction, it is considered that macrophages can promote vascular calcification by releasing inflammatory mediators and promoting the osteogenic transdifferentiation of smooth muscle cells and so on. In addition, several therapeutic strategies aimed at macrophage polarization for vascular calcification are described, which are of great significance for targeted treatment of vascular calcification., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Stromal derived factor 1 (SDF1) has been shown to be involved in the pathogenesis of pulmonary artery hypertension (PAH). However, the detailed molecular mechanisms remain unclear. To address this, we utilized primary cultured rat pulmonary artery smooth muscle cells (PASMCs) and monocrotaline (MCT)-induced PAH rat models to investigate the mechanisms of SDF1 driving PASMCs proliferation and pulmonary arterial remodeling. SDF1 increased runt-related transcription factor 2 (Runx2) acetylation by Calmodulin (CaM)-dependent protein kinase II (CaMKII)-dependent HDAC4 cytoplasmic translocation, elevation of Runx2 acetylation conferred its resistance to proteasome-mediated degradation. The accumulation of Runx2 further upregulated osteopontin (OPN) expression, finally leading to PASMCs proliferation. Blocking SDF1, suppression of CaMKII, inhibition the nuclear export of HDAC4 or silencing Runx2 attenuated pulmonary arterial remodeling and prevented PAH development in MCT-induced PAH rat models. Our study provides novel sights for SDF1 induction of PASMCs proliferation and suggests that targeting SDF1/CaMKII/HDAC4/Runx2 axis has potential value in the management of PAH., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Ex vivo tissue culture of the human corpus cavernosum (CC) can be used to explore the tissue structural changes and complex signaling networks. At present, artificial CC-like tissues based on acellular or three-dimensional (3D)-printed scaffolds are used to solve the scarcity of primary penis tissue samples. However, inconvenience and high costs limit the wide application of such methods. Here, we describe a simple, fast, and economical method of constructing artificial CC-like tissue. Human CC fibroblasts (FBs), endothelial cells (ECs), and smooth muscle cells (SMCs) were expanded in vitro and mixed with Matrigel in specific proportions. A large number of bubbles were formed in the mixture by vortexing combined with pipette blowing, creating a porous, spongy, and spatial structure. The CC FBs produced a variety of signaling factors, showed multidirectional differentiation potential, and grew in a 3D grid in Matrigel, which is necessary for CC-like tissue to maintain a porous structure as a cell scaffold. Within the CC-like tissue, ECs covered the surface of the lumen, and SMCs were located inside the trabeculae, similar to the structure of the primary CC. Various cell components remained stable for 3 days in vitro , but the EC content decreased on the 7 th day. Wingless/integrated (WNT) signaling activation led to lumen atrophy and increased tissue fibrosis in CC-like tissue, inducing the same changes in characteristics as in the primary CC. This study describes a preparation method for human artificial CC-like tissue that may provide an improved experimental platform for exploring the function and structure of the CC and conducting drug screening for erectile dysfunction therapy., (Copyright © 2024 Copyright: © The Author(s)(2024).)

Fine particulate matter (PM2.5) has been linked to increased severity and incidence of airway diseases, especially chronic obstructive pulmonary disease (COPD) and asthma. Airway remodeling is an important event in both COPD and asthma, and airway smooth muscle cells (ASMCs) are key cells which directly involved in airway remodeling. However, it was unclear how PM2.5 affected ASMCs. This study investigates the effects of PM2.5 on airway smooth muscle and its mechanism. We first showed that inhaled particulate matter was distributed in the airway smooth muscle bundle, combined with increased airway smooth muscle bundle and collagen deposition in vivo. Then, we demonstrated that PM2.5 induced up-regulation of collagen-I and alpha-smooth muscle actin (α-SMA) expression in rat and human ASMCs in vitro. Next, we found PM2.5 led to rat and human ASMCs senescence and exhibited senescence-associated secretory phenotype (SASP) by autophagy-induced GATA4/TRAF6/NF-κB signaling, which contributed to collagen-I and α-SMA synthesis as well as airway smooth muscle remodeling. Together, our results provided evidence that SASP induced by PM2.5 in airway smooth muscle cells prompted airway remodeling., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Diabetes mellitus (DM) is a significant risk factor for peripheral arterial disease (PAD), and PAD is an independent predictor of cardiovascular disorders (CVDs). Growing evidence suggests that long non-coding RNAs (lncRNAs) significantly contribute to disease development and underlying complications, particularly affecting smooth muscle cells (SMCs). So far, no study has focused on transcriptome analysis of lncRNAs in PAD patients with and without DM. Tissue samples were obtained from our Vascular Biobank. Due to the sample's heterogeneity, expression analysis of lncRNAs in whole tissue detected only ACTA2-AS1 with a 4.9-fold increase in PAD patients with DM. In contrast, transcriptomics of SMCs revealed 28 lncRNAs significantly differentially expressed between PAD with and without DM (FDR < 0.1). Sixteen lncRNAs were of unknown function, six were described in cancer, one connected with macrophages polarisation, and four were associated with CVDs, mainly with SMC function and phenotypic switch (NEAT1, MIR100HG, HIF1A-AS3, and MRI29B2CHG). The enrichment analysis detected additional lncRNAs H19, CARMN, FTX, and MEG3 linked with DM. Our study revealed several lncRNAs in diabetic PAD patients associated with the physiological function of SMCs. These lncRNAs might serve as potential therapeutic targets to improve the function of SMCs within the diseased tissue and, thus, the clinical outcome., (© 2024. The Author(s).)

Vascular smooth muscle cells (VSMCs) contribute to neointimal hyperplasia (NIH) after vascular injury, a common feature of vascular remodelling disorders. Suramin is known to exert antitumour effects by inhibiting the proliferation of various tumour cells; however, its effects and mechanism on VSMCs remain unclear. This study investigated the effects of suramin on human aortic smooth muscle cells (HASMCs), rat aortic smooth muscle cells (RASMCs) and NIH to examine its suitability for the prevention of vascular remodelling disorders. In vitro, suramin administration reduced platelet-derived growth factor type BB (PDGF-BB)-stimulated proliferation, migration, and dedifferentiation of VSMCs through a transforming growth factor beta receptor 1 (TGFBR1)/Smad2/3-dependent pathway. Suramin dramatically inhibited NIH ligation in the left common carotid artery (LCCA) vivo. Therefore, our results indicate that suramin protects against the development of pathological vascular remodelling by attenuating VSMCs proliferation, migration, and phenotypic transformation and may be used as a potential medicine for the treatment of NIH., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Abdominal aortic aneurysm (AAA), a vascular degenerative disease, is a potentially life-threatening condition characterised by the loss of vascular smooth muscle cells (VSMCs), degradation of extracellular matrix (ECM), inflammation, and oxidative stress. Despite the severity of AAA, effective drugs for treatment are scarce. At low doses, terazosin (TZ) exerts antiapoptotic and anti-inflammatory effects in several diseases, but its potential to protect against AAA remains unexplored. Herein, we investigated the effects of TZ in two AAA animal models: Angiotensin II (Ang II) infusion in Apoe -/- mice and calcium chloride application in C57BL/6J mice. Mice were orally administered with TZ (100 or 1000 μg/kg/day). The in vivo results indicated that low-dose TZ alleviated AAA formation in both models. Low-dose TZ significantly reduced aortic pulse wave velocity without exerting an apparent antihypertensive effect in the Ang II-induced AAA model. Paternally expressed gene 3 (Peg3) was identified via RNA sequencing as a novel TZ target. PEG3 expression was significantly elevated in both mouse and human AAA tissues. TZ suppressed PEG3 expression and reduced the abundance of matrix metalloproteinases (MMP2/MMP9) in the tunica media. Functional experiments and molecular analyses revealed that TZ (10 nM) treatment and Peg3 knockdown effectively prevented Ang II-induced VSMC senescence and apoptosis in vitro. Thus, Peg3, a novel target of TZ, mediates inflammation-induced VSMC apoptosis and senescence. Low-dose TZ downregulates Peg3 expression to attenuate AAA formation and ECM degradation, suggesting a promising therapeutic strategy for AAA., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Pulmonary arterial hypertension (PAH) causes pulmonary vascular remodeling, increasing pulmonary vascular resistance (PVR) and leading to right heart failure and death. Matrix stiffening early in the disease promotes remodeling in pulmonary artery smooth muscle cells (PASMCs), contributing to PAH pathogenesis. Our research identified YAP and TAZ as key drivers of the mechanobiological feedback loop in PASMCs, suggesting targeting them could mitigate remodeling. However, YAP/TAZ are ubiquitously expressed and carry out diverse functions, necessitating a cell-specific approach. Our previous work demonstrated that targeting non-canonical IKB kinase TBK1 reduced YAP/TAZ activation in human lung fibroblasts. Here, we investigate non-canonical IKB kinases TBK1 and IKKε in pulmonary hypertension (PH) and their potential to modulate PASMC pathogenic remodeling by regulating YAP/TAZ. We show that TBK1 and IKKε are activated in PASMCs in a rat PH model. Inflammatory cytokines, elevated in PAH, activate these kinases in human PASMCs. Inhibiting TBK1/IKKε expression/activity significantly reduces PAH-associated PASMC remodeling, with longer-lasting effects on YAP/TAZ than treprostinil, an approved PAH therapy. These results show that non-canonical IKB kinases regulate YAP/TAZ in PASMCs and may offer a novel approach for reducing vascular remodeling in PAH., (© 2024 The Authors. Physiological Reports published by Wiley Periodicals LLC on behalf of The Physiological Society and the American Physiological Society.)

Alginate oligosaccharide (AOS) may delay aging by decreasing oxidative stress, but the effects on vascular aging remain unclear. Here, we evaluate the effect of AOS on vascular aging and investigate the underlying mechanisms. Twenty-month-old rats acted as the natural aging model in vivo. Senescence of human aortic vascular smooth muscle cells (HA-VSMCs) was induced in vitro using angiotensin II (AngII). The aging rats and senescent cells were treated with AOS, followed by assessment of aging makers, oxidative stress, and aging-induced vascular remodeling. AOS treatment alleviated vascular aging and HA-VSMC senescence and decreased the levels of oxidative stress and vascular remodeling-associated indicators. AOS upregulated the expression of glutathione peroxidase 7 (GPX7) in aging rats and GPX7 depletion disrupted the geroprotective effect of AOS. AOS increased the nuclear translocation of nuclear factor erythroid-2-related factor (Nrf2) protein, which interacts with GPX7 protein to induce its expression. In conclusion, AOS alleviates vascular aging and HA-VSMC senescence and reduces aging-related vascular remodeling via the GPX7 antioxidant pathway, which may provide new avenues for treating aging-associated diseases., Competing Interests: Declarations of interest The authors declare that there are no conflicts of interest., (Copyright © 2024. Published by Elsevier Inc.)

Accumulating evidence shows that the abnormal proliferation and migration of vascular smooth muscle cells (VSMCs) can significantly affect the long-term prognosis of coronary artery bypass grafting. This study aimed to explore the factors affecting the proliferation, migration, and phenotypic transformation of VSMCs. First, we stimulated VSMCs with different platelet-derived growth factor-BB (PDGF-BB) concentrations, analyzed the expression of phenotype-associated proteins by Western blotting, and examined cell proliferation by scratch wound healing and the 5-ethynyl-2-deoxyuridine (EdU) assay. VSMC proliferation was induced most by PDGF-BB treatment at 20 ng/mL. miR-200a-3p decreased significantly in A7r5 cells stimulated with PDGF-BB. The overexpression of miR-200a-3p reversed the downregulation of α-SMA (p < 0.001) and the upregulation of vimentin (p < 0.001) caused by PDGF-BB. CCK8 and EdU analyses showed that miR-200a-3p overexpression could inhibit PDGF-BB-induced cell proliferation (p < 0.001). However, flow cytometric analysis showed that it did not significantly increase cell apoptosis. Collectively, the overexpression of miR-200a-3p inhibited the proliferation and migration of VSMCs induced by PDGF-BB, partly by affecting phenotypic transformation-related proteins, providing a new strategy for relieving the restenosis of vein grafts., (© 2024 Wiley Periodicals LLC.)

Objective: Abnormal proliferation and migration of biomechanical force-induced venous smooth muscle cells (VSMCs) is a major cause to limit the efficacy of coronary artery bypass grafting (CABG) for coronary heart disease (CHD). Scutellarin is the main active ingredient of Erigeron Breviscapus, and has broad-spectrum pharmacological effects. Therefore, the present study was proposed to investigate the effect of Scutellarin on VSMCs under tensile stress., Methods: After interfering with VSMCs at different tensile stresses, the optimal tensile stress was screened. In a tensile stress environment, 100 μM Scutellarin and Hesperetin (p38 MAPK pathway activator) was used to treatment with VSMCs. CCK-8, EDU, Wound healing, flow cytometry and western blotting assays were used to detect cell proliferation, migration, apoptosis, and the expression of apoptosis-related proteins (Caspase3, Bcl2 and Bax)., Results: Tensile stress with 10% significantly enhanced the activity, wound-healing ratio, and EDU+ cells of VSMCs, and decreased their apoptosis ratio. Moreover, it upregulated Bcl2 expression, and downregulated cleaved-Caspase3 and Bax expression of VSMCs. Hence, 10% tensile stress was selected to creates a tensile stress environment for VSMCs. Interestingly, 100 μM Scutellarin alleviated the effect of 10% tensile stress on the phenotype of VSMCs. Notably, 10% tensile stress increased the phosphorylation level of p38 MAPK (Thr180 +Tyr182) in VSMCs, which was restricted by Scutellarin. Further, Hesperetin restored the effect of Scutellarin on the phenotype of VSMCs., Conclusion: Scutellarin alleviates tension stress-induced proliferation and migration of VSMCs via suppressing p38 MAPK pathway. Scutellarin may be used as an adjunctive strategy for future GABG treatment in CHD patients., (Copyright © 2024. Published by Elsevier Ltd.)

Objective: To explore the role of interleukin (IL)-17 in connective tissue disease-associated pulmonary arterial hypertension (CTD-PAH) and to investigate its possible mechanism on pulmonary artery smooth muscle cells (PASMCs)., Methods: Enzyme-linked immunosorbent assay (ELISA) were used to compare levels of serum IL-17 in patients with CTD-PAH and healthy controls (HCs). After treatment for 3 months, the serum IL-17 levels were tested in CTD-PAH. ELISA and immunohistochemistry were used to compare levels of serum IL-17 and numbers of pulmonary artery IL-17 + cells, respectively, in a rat model of monocrotaline-induced PAH and untreated rats. Proliferation, migration, and inflammatory factors expression of PASMCs were assessed after stimulation with different concentrations of IL-17 for various time periods. Proteins in the mitogen-activated protein kinase (MAPK) pathway were examined by western blot., Results: Levels of IL-17 were upregulated in patients with CTD-PAH compared to HCs. After 3 months of treatment, serum IL-17 levels were downregulated with pulmonary artery pressure amelioration. Moreover, serum IL-17 levels and numbers of IL-17 + cells infiltrating lung arterioles were increased in PAH model rats. IL-17 could dose- and time-dependently promote proliferation and migration of PASMCs as well as time-dependently induce IL-6 and intercellular cell adhesion molecule-1 (ICAM-1) expression. The levels of MKK6 increased after IL-17 treatment. Inhibition of MAPK decreased proliferation of PASMCs., Conclusion: Levels of IL-17 may increase in CTD-PAH, and IL-17 promotes proliferation, migration, and secretion of IL-6 and ICAM in PASMCs, respectively, which likely involves the p-38 MAPK pathway., (© 2024 The Authors. Immunity, Inflammation and Disease published by John Wiley & Sons Ltd.)

Background: Hyperproliferation, inflammation, and mitochondrial abnormalities in pulmonary artery smooth muscle cells (PASMCs) underlie the pathological mechanisms of vascular remodeling in pulmonary arterial hypertension (PAH). Cytoplasmic mtDNA activates the cGAS-STING-NFκB pathway and secretes pro-inflammatory cytokines that may be involved in the pathogenesis of PAH. Calcitonin gene-related peptide (CGRP) acts as a vasodilator to regulate patterns of cellular energy metabolism and has vasodilatory and anti-inflammatory effects., Methods: The role of the cGAS-STING-NFκB signaling pathway in PAH vascular remodeling and the regulation of CGRP in the cGAS-STING-NFκB signaling pathway were investigated by echocardiography, morphology, histology, enzyme immunoassay, and fluorometry., Results: Monocrotaline (MCT) could promote right heart hypertrophy, pulmonary artery intima thickening, and inflammatory cell infiltration in rats. Cinnamaldehyde (CA)-induced CGRP release alleviates MCT-induced vascular remodeling in PAH. CGRP reduces PDGF-BB-induced proliferation, and migration, and downregulates smooth muscle cell phenotypic proteins. In vivo and in vitro experiments confirm that the mitochondria of PASMCs were damaged during PAH, and the superoxide and mtDNA produced by injured mitochondria activate the cGAS-STING-NFκB pathway to promote PAH process, while CGRP could play an anti-PAH role by protecting the mitochondria and inhibiting the cGAS-STING-NFκB pathway through PKA., Conclusion: This study identifies that CGRP attenuates cGAS-STING-NFκB axis-mediated vascular remodeling in PAH through PKA., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Deoxynivalenol (DON) is one of the frequent Fusarium mycotoxins and poses a serious threat to public health worldwide. DON-induced weight loss is tightly connected with its ability to decrease feed intake by influencing gastrointestinal tract (GIT) motility. Our previous reports indicated that DON interfered with intestinal motility by injuring the contractility of enteric smooth muscle cells (SMC). Here, we further explored the potential mechanisms by employing a complementary method of transcriptomics and proteomics using the porcine enteric smooth muscle cell line (PISMC) as an experimental model. The transcriptomic and proteomic data uncover that the expression of numerous extracellular matrix (ECM) proteins and multiple integrin subunits were downregulated in PISMC under DON exposure, suppressing the ECM-integrin receptor interaction and its mediated signaling. Furthermore, DON treatment could depress actin polymerization, as reflected by the upregulated expression of Rho GTPase-activating proteins and cofilin in PISMC. Meanwhile, the expression levels of downstream contractile apparatus genes were significantly inhibited after challenge with DON. Taken together, the current results suggest that DON inhibits enteric SMC contractility by regulating the ECM-integrin-actin polymerization signaling pathway. Our findings provide novel insights into the potential mechanisms behind the DON toxicological effects in the GIT of humans and animals., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Background: Metabolic alterations provide substrates that influence chromatin structure to regulate gene expression that determines cell function in health and disease. Heightened proliferation of smooth muscle cells (SMC) leading to the formation of a neointima is a feature of pulmonary arterial hypertension (PAH) and systemic vascular disease. Increased glycolysis is linked to the proliferative phenotype of these SMC.Methods: RNA sequencing was applied to pulmonary arterial SMC (PASMC) from PAH patients with and without a BMPR2 (bone morphogenetic receptor 2) mutation versus control PASMC to uncover genes required for their heightened proliferation and glycolytic metabolism. Assessment of differentially expressed genes established metabolism as a major pathway, and the most highly upregulated metabolic gene in PAH PASMC was aldehyde dehydrogenase family 1 member 3 (ALDH1A3), an enzyme previously linked to glycolysis and proliferation in cancer cells and systemic vascular SMC. We determined if these functions are ALDH1A3-dependent in PAH PASMC, and if ALDH1A3 is required for the development of pulmonary hypertension in a transgenic mouse. Nuclear localization of ALDH1A3 in PAH PASMC led us to determine whether and how this enzyme coordinately regulates gene expression and metabolism in PAH PASMC.Results: ALDH1A3 mRNA and protein were increased in PAH versus control PASMC, and ALDH1A3 was required for their highly proliferative and glycolytic properties. Mice with Aldh1a3 deleted in SMC did not develop hypoxia-induced pulmonary arterial muscularization or pulmonary hypertension. Nuclear ALDH1A3 converted acetaldehyde to acetate to produce acetyl coenzyme A to acetylate H3K27, marking active enhancers. This allowed for chromatin modification at NFYA (nuclear transcription factor Y subunit α) binding sites via the acetyltransferase KAT2B (lysine acetyltransferase 2B) and permitted NFY-mediated transcription of cell cycle and metabolic genes that is required for ALDH1A3-dependent proliferation and glycolysis. Loss of BMPR2 in PAH SMC with or without a mutation upregulated ALDH1A3, and transcription of NFYA and ALDH1A3 in PAH PASMC was β-catenin dependent.Conclusions: Our studies have uncovered a metabolic-transcriptional axis explaining how dividing cells use ALDH1A3 to coordinate their energy needs with the epigenetic and transcriptional regulation of genes required for SMC proliferation. They suggest that selectively disrupting the pivotal role of ALDH1A3 in PAH SMC, but not endothelial cells, is an important therapeutic consideration. [ABSTRACT FROM AUTHOR]

Vascular smooth muscle cells (VSMCs) have long been associated with phenotypic modulation/plasticity or dedifferentiation. Innovative technologies in cell lineage tracing, single-cell RNA sequencing, and human genomics have been integrated to gain unprecedented insights into the molecular reprogramming of VSMCs to other cell phenotypes in experimental and clinical atherosclerosis. The current thinking is that an apparently small subset of contractile VSMCs undergoes a fate switch to transitional, multipotential cells that can adopt plaque-destabilizing (inflammation, ossification) or plaque-stabilizing (collagen matrix deposition) cell states. Several candidate mediators of such VSMC fate and state changes are coming to light with intriguing implications for understanding coronary artery disease risk and the development of new treatment modalities. Here, we briefly summarize some technical and conceptual advancements derived from 2 publications in Circulation and another in Nature Medicine that, collectively, illuminate new research directions to further explore the role of VSMCs in atherosclerotic disease. [ABSTRACT FROM AUTHOR]

How are p300 and CBP regulated in SMCs; in particular, what makes p300 decrease with injury and CBP increase with injury or aging? Keywords: Editorials; chromatin assembly and disassembly; muscle, smooth, vascular; histone acetyltransferases; myocytes, smooth muscle; phenotype EN Editorials chromatin assembly and disassembly muscle, smooth, vascular histone acetyltransferases myocytes, smooth muscle phenotype 1738 1740 3 06/03/22 20220607 NES 220607 B Article, see p 1720 b Despite an identical genome in every cell, differential gene expression determines call fate: cardiomyocyte versus leukocyte and vascular smooth muscle versus endothelial cell. This is confirmed in vivo using novel mouse models with CBP or p300 specifically deleted from SMCs: Loss of p300 promotes neointima formation, and loss of CBP attenuates the SMC response to injury. [Extracted from the article]

Background: Although sex differences in coronary artery disease are widely accepted with women developing more stable atherosclerosis than men, the underlying pathobiology of such differences remains largely unknown. In coronary artery disease, recent integrative systems biological studies have inferred gene regulatory networks (GRNs). Within these GRNs, key driver genes have shown great promise but have thus far been unidentified in women.Methods: We generated sex-specific GRNs of the atherosclerotic arterial wall in 160 women and age-matched men in the STARNET study (Stockholm-Tartu Atherosclerosis Reverse Network Engineering Task). We integrated the female GRNs with single-cell RNA-sequencing data of the human atherosclerotic plaque and single-cell RNA sequencing of advanced atherosclerotic lesions from wild type and Klf4 knockout atherosclerotic smooth muscle cell (SMC) lineage-tracing mice.Results: By comparing sex-specific GRNs, we observed clear sex differences in network activity within the atherosclerotic tissues. Genes more active in women were associated with mesenchymal cells and endothelial cells, whereas genes more active in men were associated with the immune system. We determined that key drivers of GRNs active in female coronary artery disease were predominantly found in (SMCs by single-cell sequencing of the human atherosclerotic plaques, and higher expressed in female plaque SMCs, as well. To study the functions of these female SMC key drivers in atherosclerosis, we examined single-cell RNA sequencing of advanced atherosclerotic lesions from wild type and Klf4 knockout atherosclerotic SMC lineage-tracing mice. The female key drivers were found to be expressed by phenotypically modulated SMCs and affected by Klf4, suggesting that sex differences in atherosclerosis involve phenotypic switching of plaque SMCs.Conclusions: Our systems approach provides novel insights into molecular mechanisms that underlie sex differences in atherosclerosis. To discover sex-specific therapeutic targets for atherosclerosis, an increased emphasis on sex-stratified approaches in the analysis of multi-omics data sets is warranted. [ABSTRACT FROM AUTHOR]

Objective: Atherosclerosis (AS) is a chronic inflammatory disease of the arterial wall, in which Human Vascular Smooth Muscle Cells (HVSMCs) are involved. Nevertheless, the functions and mechanisms of circRNAs in oxidized Low-Density Lipoprotein (ox-LDL)-induced vascular smooth muscle cells remain unclear., Methods: Circ-ABCA1 expression was measured in the models of AS. Then, in the vitro model, oligonucleotide transfection was performed, followed by an analysis of VSMC proliferation, migration, inflammation, and phenotypic switch. Also, in the in vivo model, mice were injected with shRNA lentivirus, followed by histological examination of aortic tissues. Finally, the interaction of circ-ABCA1, miR-885-5p, and ROCK2 was identified., Results: Circ-ABCA1, was confirmed to be overexpressed in ox-LDL-induced VSMCs and mouse models of AS. Functionally, silencing circ-ABCA1 via oligonucleotide transfection suppressed VSMC proliferation, migration, inflammation, and phenotypic switch in vitro and prevented AS development in mice in vivo. Mechanistically, circ-ABCA1 absorbed miR-885-5p, which targeted ROCK2., Conclusion: Taken together, the data from this study suggest that circ-ABCA1 mediates cellular inflammation and phenotype switching through the miR-885-5p/ROCK2 axis in ox-LDL-induced VSMCs, and the circ-ABCA1/miR-885-5p/ROCK2 axis is a new potential biomarker for the treatment of AS., Competing Interests: Conflicts of Interest The authors declare no conflicts of interest., (Copyright © 2024 HCFMUSP. Published by Elsevier España, S.L.U. All rights reserved.)

Background: Injury to contractile organs such as the heart, vasculature, urinary bladder and gut can stimulate a pathological response that results in loss of normal contractility. PDGF and TGFβ are among the most well studied initiators of the injury response and have been shown to induce aberrant contraction in mechanically active cells of hollow organs including smooth muscle cells (SMC) and fibroblasts. However, the mechanisms driving contractile alterations downstream of PDGF and TGFβ in SMC and fibroblasts are incompletely understood, limiting therapeutic interventions., Methods: To identify potential molecular targets, we have leveraged the analysis of publicly available data, comparing transcriptomic changes in mechanically active cells stimulated with PDGF and TGFβ. Additional Analysis of publicly available data sets were performed on SMC and fibroblasts treated in the presence or absence of the MYC inhibitor JQ1. Validation of in silico findings were performed with qPCR, immunoblots, and collagen gel contraction assays measure the effect of JQ1 on cytoskeleton associated genes, proteins and contractility in mechanically active cells. Likelihood ratio test and FDR adjusted p-values were used to determine significant differentially expressed genes. Student ttest were used to calculate statistical significance of qPCR and contractility analyses., Results: Comparing PDGF and TGFβ stimulated SMC and fibroblasts identified a shared molecular profile regulated by MYC and members of the AP-1 transcription factor complex. Additional in silico analysis revealed a unique set of cytoskeleton-associated genes that were sensitive to MYC inhibition with JQ1. In vitro validation demonstrated JQ1 was also able to attenuate TGFβ and PDGF induced changes to the cytoskeleton and contraction of smooth muscle cells and fibroblasts in vitro., Conclusions: These findings identify MYC as a key driver of aberrant cytoskeletal and contractile changes in fibroblasts and SMC, and suggest that JQ1 could be used to restore normal contractile function in hollow organs., (© 2024. The Author(s).)

Non-immune cells, like innate immune cells, can develop a memory-like phenotype in response to priming with microbial compounds or certain metabolites, which enables an enhanced response to a secondary unspecific stimulus. This paper describes a step-by-step protocol for the induction and analysis of trained immunity in human endothelial and smooth muscle cells. We then describe steps for cell culture with cryopreserved vascular cells, subcultivation, and induction of trained immunity. We then provide detailed procedures for downstream analysis using ELISA and qPCR. For complete details on the use and execution of this protocol, please refer to Sohrabi et al. (2020) 1 and Shcnack et al. 2 ., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2023 The Author(s). Published by Elsevier Inc. All rights reserved.)

In the search for novel, bi-functional compounds acting as Ca V 1.2 channel blockers and K + channel stimulators, which represent an effective therapy for hypertension, 3,3'-O-dimethylquercetin was isolated for the first time from Brazilian Caatinga green propolis. Its effects were investigated through electrophysiological, functional, and computational approaches. In rat tail artery myocytes, 3,3'-O-dimethylquercetin blocked Ba 2+ currents through Ca V 1.2 channels (I Ba1.2 ) in a concentration-dependent manner, with the inhibition being reversed upon washout. The compound also shifted the voltage dependence of the steady-state inactivation curve to more negative potentials without affecting the slope of the inactivation and activation curves. Furthermore, the flavonoid stimulated K Ca 1.1 channel currents (I KCa1.1 ). In silico simulations provided additional evidence for the binding of 3,3'-O-dimethylquercetin to K Ca 1.1 and Ca V 1.2 channels and elucidated its mechanism of action. In depolarized rat tail artery rings, the flavonoid induced a concentration-dependent relaxation. Moreover, in rat aorta rings its antispasmodic effect was inversely related to the transmembrane K + gradient. In conclusion, 3,3'-O-dimethylquercetin demonstrates effective in vitro vasodilatory properties, encouraging the exploration of its scaffold to develop novel derivatives for potential use in the treatment of hypertension., Competing Interests: Declaration of competing interest The authors declare no competing financial interest., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Ischemic stroke produces the highest adult disability. Despite successful recanalization, no-reflow, or the futile restoration of the cerebral perfusion after ischemia, is a major cause of brain lesion expansion. However, the vascular mechanism underlying this hypoperfusion is largely unknown, and no approach is available to actively promote optimal reperfusion to treat no-reflow. Here, by combining two-photon laser scanning microscopy (2PLSM) and a mouse middle cerebral arteriolar occlusion (MCAO) model, we find myogenic vasomotion deficits correlated with post-ischemic cerebral circulation interruptions and no-reflow. Transient occlusion-induced transient loss of mitochondrial membrane potential (ΔΨm) permanently impairs mitochondria-endoplasmic reticulum (ER) contacts and abolish Ca 2+ oscillation in smooth muscle cells (SMCs), the driving force of myogenic spontaneous vasomotion. Furthermore, tethering mitochondria and ER by specific overexpression of ME-Linker in SMCs restores cytosolic Ca 2+ homeostasis, remotivates myogenic spontaneous vasomotion, achieves optimal reperfusion, and ameliorates neurological injury. Collectively, the maintaining of arteriolar myogenic vasomotion and mitochondria-ER contacts in SMCs, are of critical importance in preventing post-ischemic no-reflow., (© 2024. The Author(s).)

Competing Interests: Conflict of Interest: No conflict of interest was declared by the authors.

Many vascular disorders arise as a result of dysfunctional smooth muscle cells. Tissue engineering strategies have evolved as key approaches to generate functional vascular smooth muscle cells for use in cell-based precision and personalized regenerative medicine approaches. This article highlights some of the challenges that exist in the field and presents some of the prospects for translating research advancements into therapeutic modalities. The article emphasizes the need for better developing synergetic intracellular and extracellular cues in the processes to generate functional vascular smooth muscle cells from different stem cell sources for use in tissue engineering strategies.

Primary cell cultures are essential tools for elucidating the physiopathological mechanisms of the cardiovascular system. Therefore, a primary culture growth protocol of cardiovascular smooth muscle cells (VSMCs) obtained from human abdominal aortas was standardized. Ten abdominal aorta samples were obtained from patients diagnosed with brain death who were organ and tissue donors with family consent. After surgical ablation to capture the aorta, the aortic tissue was removed, immersed in a Custodiol® solution, and kept between 2 and 8 °C. In the laboratory, in a sterile environment, the tissue was fragmented and incubated in culture plates containing an enriched culture medium (DMEM/G/10% fetal bovine serum, L-glutamine, antibiotics and antifungals) and kept in an oven at 37 °C and 5% CO 2 . The aorta was removed after 24 h of incubation, and the culture medium was changed every six days for twenty days. Cell growth was confirmed through morphological analysis using an inverted optical microscope (Nikon®) and immunofluorescence for smooth muscle alpha-actin and nuclei. The development of the VSMCs was observed, and from the twelfth day, differentiation, long cytoplasmic projections, and adjacent cell connections occurred. On the twentieth day, the morphology of the VSMCs was confirmed by actin fiber immunofluorescence, which is a typical characteristic of VSMCs. The standardization allowed VSMC growth and the replicability of the in vitro test, providing a protocol that mimics natural physiological environments for a better understanding of the cardiovascular system. Its use is intended for investigation, tissue bioengineering, and pharmacological treatments., (© 2023. The Author(s), under exclusive licence to Springer Nature B.V.)