Smooth Muscle Cell Migration Research Articles

Abstract P1075: Elucidate The Mechanism Driving The Imbalance Between BMPs And Activin/Gdfs Signaling In Pulmonary Arterial Hypertension

Transforming Growth Factor-beta (TGF-β) family signaling is involved in Pulmonary Arterial Hypertension (PAH) in humans. PAH is a rare, highly morbid disorder characterized by dysregulated pulmonary vascular remodeling, elevated pulmonary vascular resistance, and the progressive obliteration of the pulmonary arteriolar circulation. The progression of PAH is largely influenced by the proliferation and migration of the pulmonary smooth muscle cells (PASMCs) and pulmonary endothelial cells (PAECs). While the exact mechanism is not understood, there is evidence that germline mutations in the BMPRII gene, one of the TGF-β family type II receptors, is present in PAH patients. Due to the functional similarity of BMPRII and type II Activin receptors ACVR2A and ACVR2B, they interact with overlapping TGF-β family ligands to activate either the SMAD 1/5/8 or SMAD 2/3 signaling. In the pulmonary vasculature, the BMP-9 ligand regulates cell proliferation by binding to type I receptors, ALK-1 and BMPRII, to activate SMAD 1/5/8 signaling. In PAH, there is a decrease in SMAD 1/5/8 signaling and an increase in SMAD 2/3 signaling. However, what causes the signaling switch is still unknown. In our lab, we found out that BMPRII and ACVR2A bind TGF-β ligands with nearly identical geometry based on our crystal structures. Both of the receptors contain a conserved hydrophobic hot spot within the ligand binding pocket. We found that the BMPRII ligand binding domain may exhibit greater flexibility than ACVR2A. We also investigated eight soluble BMPRII extracellular domain mutants and expressed them as BMPRII-Fc fusion proteins (BMPRII-Fc variants). Using surface plasmon resonance, we observed four of the BMPRII-Fc variants bind to high affinity ligands, such as BMP9, BMP10, Activin B and Activin A. In reporter gene assays, we found these BMPRII-Fc variants act as a ligand trap and attenuate BMP9 and BMP10-mediated SMAD1/5/8 signaling in HEK293 cells. Interestingly, they did not inhibit Activin B-mediated SMAD2/3 signaling. In conclusion, our findings show gain of function of specific BMPRII mutations and suggest that imbalances between SMAD1/5/8 and SMAD2/3 signaling may lead to PAH progression.

Local intraluminal delivery of a smooth muscle-targeted RNA ligand inhibits neointima growth in a porcine model of peripheral vascular disease

Anti-proliferative agents have been the primary therapeutic drug of choice to inhibit restenosis after endovascular treatment. However, recent safety and efficacy concerns for patients who underwent peripheral artery disease revascularization have demonstrated the need for alternative therapeutics. The aim of this investigation was to investigate the efficacy of a cell-specific RNA aptamer inhibiting vascular smooth muscle cell proliferation and migration. First, the impact of the RNA aptamer (Apt 14) on the wound healing of primary cultured porcine vascular smooth muscle cells (VSMCs) was examined in response to a scratch wound injury. We then evaluated the effect of local luminal delivery of Apt 14 on neointimal formation in a clinically relevant swine iliofemoral injury model. In contrast with a non-selected control aptamer (NSC) that had no impact on VSMC migration, Apt 14 attenuated the wound healing of primary cultured porcine VSMCs to platelet-derived growth factor-BB. Histological analysis of the Apt 14-treated arteries demonstrated a significant reduction in neointimal area percent diameter stenosis compared with arteries treated with saline and NSC controls. The findings of this study suggest that aptamers can function as selective inhibitors and thus provide more fine-tuning to inhibit selective pathways responsible for neointimal hyperplasia.

Platelet TGF-β1 inhibits the migration and proliferation of smooth muscle cells in aneurysms

BackgroundThe study explored the role of platelet TGF-β1 from the perspective of inhibiting the excessive proliferation, migration and invasion of murine aortic vascular smooth muscle cells (MASMCs). MethodThe platelets were first extracted from C57BL/6 mice, and the TGF-β1 protein was obtained after the purification of protein. In vitro, the concentrations of angiotensin Ⅱ (Ang Ⅱ) and TGF-β1 for intervention were screened by testing the viability of MASMCs, followed by the analysis concerning the effects of platelets, Ang Ⅱ and TGF-β1 on the proliferation, migration, invasion, and the expressions of pathway-related proteins in MASMCs. In vivo, an Ang Ⅱ-induced mouse model was established. TGF-β1 was injected into the tail of mice as a therapeutic agent, and its action mechanism was further verified by the treatment of inhibitor SB505124. The results of the cell experiment were validated by evaluating the maximum diameter of abdominal aorta, the proportion of total weight, the changes of both pathology and the expressions of pathway-related proteins in the mice. Result0.5 ng/mL Ang Ⅱ and 15 ng/mL TGF-β1 were chosen for treatment. The following results of cell functional experiments and Western blot assay demonstrated that Ang Ⅱ promoted the proliferation, migration and invasion of MASMCs via regulating related pathways, the effects of which were evidently reversed by TGF-β1 and platelets. Consistent results were also observed in the animal experiments, where TGF-β1 effectively alleviated Ang Ⅱ-induced abdominal aortic injury in mice. ConclusionTGF-β1 in platelets inhibits Ang Ⅱ-induced proliferation, migration and invasion of MASMCs.

PPARγ activation inhibits PDGF-induced pulmonary artery smooth muscle cell proliferation and migration by modulating TERT.

Vascular remodeling is a significant feature of pulmonary artery hypertension (PAH), and is characterized by abnormal proliferation and migration of pulmonary arterial smooth muscle cells (PASMCs). Telomerase reverse transcriptase (TERT), as a determining factor for controlling telomerase activity, has been proven to be associated with cell proliferation. This study aims to explore whether TERT mediates the proliferation and migration of PASMCs and the underlying molecular mechanism. Primary PASMCs from Sprague-Dawley (SD) rats were used in this experiment. Cell proliferation and migration were evaluated by Cell Counting Kit-8, EdU incorporation assay and transwell assay, respectively. Telomerase activity was assessed with a rat TE ELISA kit. Small interfering RNA (siRNA) transfection was conducted to silence c-MYC expression. The protein levels of p-Akt, c-MYC, PPARγ and TERT were determined through western blotting. Our work demonstrates that PDGF upregulated TERT expression and telomerase activation by activating Akt and upregulating of c-MYC in PASMCs. Inhibition of Akt with LY294002, knockdown of c-MYC by siRNA or suppression of telomerase activity with BIBR1532 repressed PDGF-induced PASMC proliferation and migration. Furthermore, activation of peroxisome proliferator-activated receptor γ (PPARγ) with pioglitazone suppressed PDGF-induced TERT expression and telomerase activation, leading to inhibition of PASMC proliferation and migration.

Drug-Eluting Stents.

Endovascular revascularization strategies have advanced tremendously over the years and are now often considered first line for treatment of peripheral arterial disease. Drug-eluting stents (DESs) have been developed as one of the tools to overcome the limitations of elastic recoil and neointimal hyperplasia observed with balloon angioplasty and bare metal stents. While these stents have been extremely successful in coronary revascularization, they have not translated as effectively to the peripheral arteries which differ in their unique mechanical environments and differences in vessel and lesion composition. DESs, through their embedded pharmaceutical agent, seek to inhibit vascular smooth muscle cell (VSMC) proliferation and migration. Paclitaxel, sirolimus, and its derivatives (-limus family) achieve VSMC inhibition through unique mechanisms. Several clinical trials have been performed to evaluate the use of DES in the femoropopliteal and infrapopliteal territory and have demonstrated overall decrease in revascularization rates and improved clinical outcomes.

Effect of talaporfin sodium- mediate red laser- assisted photodynamic therapy on arterial biomechanical parameters in the rabbit carotid artery model of neointimal hyperplasia

Background and Aims : Smooth muscle cell proliferation and migration from the medial layer into the intima, following an intravascular damage like angioplasty, causes neointimal hyperplasia. There exists a range of treatment protocols for neointimal which can be divided into two main groups: invasive and non-invasive. In this study, we aimed to introduce a novel non-invasive treatment (Photodynamic therapy) for neointimal hyperplasia reduction, for which the evaluation procedure (a new computerized ultrasound image processing) is be based on a non-invasive method.

Extracellular vesicles enriched in PCSK9 are indicative of pro-atherogenic phenotype - in vitro and in vivo evidence

Background and Aims : Extracellular vesicles (EVs) are secreted into the extracellular space by several cell types. EVs carry proteins, lipids, and nucleic acids. EVs play a significant role in the process of atherosclerotic cardiovascular diseases (ASCVD). In the pathophysiology of ASCVD, proprotein convertase subtilisin/kexin type 9 (PCSK9) appears to play a crucial role, e.g., PCSK9 influences vascular smooth muscle cells (SMC) differentiation, migration and proliferation. Aim: To unveil the impact of EVs derived from human SMC overexpressing PCSK9 on the inflammatory milieu in in vitro (THP-1 and derived-macrophages) and in vivo (zebrafish) models.

Endothelial Foxp1 Regulates Neointimal Hyperplasia Via Matrix Metalloproteinase-9/Cyclin Dependent Kinase Inhibitor 1B Signal Pathway.

BackgroundThe endothelium is essential for maintaining vascular physiological homeostasis and the endothelial injury leads to the neointimal hyperplasia because of the excessive proliferation of vascular smooth muscle cells. Endothelial Foxp1 (forkhead box P1) has been shown to control endothelial cell (EC) proliferation and migration in vitro. However, whether EC‐Foxp1 participates in neointimal formation in vivo is not clear. Our study aimed to investigate the roles and mechanisms of EC‐Foxp1 in neointimal hyperplasia.Methods and ResultsThe wire injury femoral artery neointimal hyperplasia model was performed in Foxp1 EC‐specific loss‐of‐function and gain‐of‐function mice. EC‐Foxp1 deletion mice displayed the increased neointimal formation through elevation of vascular smooth muscle cell proliferation and migration, and the reduction of EC proliferation hence reendothelialization after injury. In contrast, EC‐Foxp1 overexpression inhibited the neointimal formation. EC‐Foxp1 paracrine regulated vascular smooth muscle cell proliferation and migration via targeting matrix metalloproteinase‐9. Also, EC‐Foxp1 deletion impaired EC repair through reduction of EC proliferation via increasing cyclin dependent kinase inhibitor 1B expression. Delivery of cyclin dependent kinase inhibitor 1B‐siRNA to ECs using RGD (Arg‐Gly‐Asp)‐peptide magnetic nanoparticle normalized the EC‐Foxp1 deletion‐mediated impaired EC repair and attenuated the neointimal formation. EC‐Foxp1 regulates matrix metalloproteinase‐9/cyclin dependent kinase inhibitor 1B signaling pathway to control injury induced neointimal formation.ConclusionsOur study reveals that targeting EC‐Foxp1‐matrix metalloproteinase‐9/cyclin dependent kinase inhibitor 1B pathway might provide future novel therapeutic interventions for restenosis.

Smooth Muscle Myosin Localizes at the Leading Edge and Regulates the Redistribution of Actin-regulatory Proteins during Migration.

Airway smooth muscle cell migration plays an essential role in airway development, repair, and remodeling. Smooth muscle myosin II has been traditionally thought to localize in the cytoplasm solely and regulates cell migration by affecting stress fiber formation and focal adhesion assembly. In this study, we unexpectedly found that 20-kDa myosin light chain (MLC20) and myosin-11 (MYH11), important components of smooth muscle myosin, were present at the edge of lamellipodia. The knockdown of MLC20 or MYH11 attenuated the recruitment of c-Abl, cortactinProfilin-1 (Pfn-1), and Abi1 to the cell edge. Moreover, myosin light chain kinase (MLCK) colocalized with integrin β1 at the tip of protrusion. The inhibition of MLCK attenuated the recruitment of c-Abl, cortactin, Pfn-1, and Abi1 to the cell edge. Furthermore, MLCK localization at the leading edge was reduced by integrin β1 knockdown. Taken together, our results demonstrate that smooth muscle myosin localizes at the leading edge and orchestrates the recruitment of actin-regulatory proteins to the tip of lamellipodia. Mechanistically, integrin β1 recruits MLCK to the leading edge, which catalyzes MLC20 phosphorylation. Activated myosin regulates the recruitment of actin-regulatory proteins to the leading edge, and promotes lamellipodial formation and migration.

BRD7 restrains TNF-α-induced proliferation and migration of airway smooth muscle cells by inhibiting notch signaling

Objective: Bromodomain-containing protein 7 (BRD7) is a key component of the switch/sucrose non-fermentable complex that participates in chromatin remodeling and transcriptional regulation. Although the emerging role of BRD7 in the pathophysiology of various diseases has been observed, its role in asthma remains unknown. Here, we assessed the function of BRD7 as a mediator of airway remodeling in asthma using an in vitro model. Methods: Airway smooth muscle cells (ASMCs) were challenged with tumor necrosis factor-α (TNF-α) to establish an in vitro airway remodeling model. Protein levels were examined using western blotting. Cell proliferation was measured using the cell counting kit-8 and 5-ethynyl-2′-deoxyuridine assays. Cell migration was assessed using a transwell migration assay. Results: Exposure to TNF-α dramatically decreased BRD7 levels in ASMCs. BRD7 remarkably decreased TNF-α-induced proliferation and migration of ASMCs. In contrast, ASMCs with BRD7 deficiency were more sensitive to TNF-α-induced pro-proliferative and pro-migratory effects. Mechanistically, BRD7 could repress the expression of Notch1 and block the Notch pathway in TNF-α-challenged cells. Notably, reactivation of Notch signaling substantially reversed the BRD7 overexpression-mediated effects, whereas restraining Notch signaling abolished BRD7-depletion-mediated effects on TNF-α-challenged cells. Conclusions: BRD7 inhibits the proliferation and migration of ASMCs elicited by TNF-α by downregulating the Notch pathway. This study indicates that BRD7 may exert a suppressive effect on airway remodeling during asthma.

Exosomes Derived from AT2R-Overexpressing BMSC Prevent Restenosis After Carotid Artery Injury by Attenuating the Injury-Induced Neointimal Hyperplasia

Restenosis is a severe complication after percutaneous transluminal coronary angioplasty which limits the long-term efficacy of the intervention. In this study, we investigated the efficiency of exosomes derived from AT2R-overexpressing bone mesenchymal stem cells on the prevention of restenosis after carotid artery injury. Our data showed that AT2R-EXO promoted the proliferation and migration of vascular endothelial cells and maintained the ratio of eNOS/iNOS. On the contrary, AT2R-EXO inhibited the proliferation and migration of vascular smooth muscle cells. In vivo study proved that AT2R-Exo were more effectively accumulated in the injured carotid artery than EXO and Vehicle-EXO controls. AT2R-EXO treatment could improve blood flow of the injured carotid artery site more effectively. Further analysis revealed that AT2REXO prevents restenosis after carotid artery injury by attenuating the injury-induced neointimal hyperplasia. Our study provides a novel and more efficient exosome for the treatment of restenosis diseases after intervention.

Methylation-mediated silencing of PTPRD induces pulmonary hypertension by promoting pulmonary arterial smooth muscle cell migration via the PDGFRB/PLCγ1 axis.

Objective:Pulmonary hypertension is a lethal disease characterized by pulmonary vascular remodeling and is mediated by abnormal proliferation and migration of pulmonary arterial smooth muscle cells (PASMCs). Platelet-derived growth factor BB (PDGF-BB) is the most potent mitogen for PASMCs and is involved in vascular remodeling in pulmonary hypertension development. Therefore, the objective of our study is to identify novel mechanisms underlying vascular remodeling in pulmonary hypertension.Methods:We explored the effects and mechanisms of PTPRD downregulation in PASMCs and PTPRD knockdown rats in pulmonary hypertension induced by hypoxia.Results:We demonstrated that PTPRD is dramatically downregulated in PDGF-BB-treated PASMCs, pulmonary arteries from pulmonary hypertension rats, and blood and pulmonary arteries from lung specimens of patients with hypoxic pulmonary arterial hypertension (HPAH) and idiopathic PAH (iPAH). Subsequently, we found that PTPRD was downregulated by promoter methylation via DNMT1. Moreover, we found that PTPRD knockdown altered cell morphology and migration in PASMCs via modulating focal adhesion and cell cytoskeleton. We have demonstrated that the increase in cell migration is mediated by the PDGFRB/PLCγ1 pathway. Furthermore, under hypoxic condition, we observed significant pulmonary arterial remodeling and exacerbation of pulmonary hypertension in heterozygous PTPRD knock-out rats compared with the wild-type group. We also demonstrated that HET group treated with chronic hypoxia have higher expression and activity of PLCγ1 in the pulmonary arteries compared with wild-type group.Conclusion:We propose that PTPRD likely plays an important role in the process of pulmonary vascular remodeling and development of pulmonary hypertension in vivo.

WNT1-inducible signalling pathway protein 1 stabilizes atherosclerotic plaques in apolipoprotein-E-deficient mice via the focal adhesion kinase/mitogen-activated extracellular signal-regulated kinase/extracellular signal-regulated kinase pathway.

The migration, proliferation and apoptosis of vascular smooth muscle cells (VSMCs) are critical for plaque stability. WNT-inducible signalling pathway protein-1 (WISP1), a member of the CCN family of extracellular matrix proteins, can expedite the migration and proliferation of VSMCs. However, its underlying mechanism and relationship with atherosclerosis remain elusive. The relationship between WISP1 and apoptosis of VSMCs has not been determined previously. In the study, we aimed to investigate the relationship between WISP1 and plaque stability and its related mechanism.ApoE-/- mice were divided following groups: the null lentivirus (NC), lentivirus WISP1 (IvWISP1) and WISP1-shRNA (shWISP1) groups. Immunofluorescence, Oil Red O and Masson's staining of the carotid arteries were performed. Transwell wound healing assay, CCK8 assay, and TdT-mediated dUTP nick-end labeling (TUNEL) staining were performed using VSMCs. The levels of WISP1, P38, C-Jun N-terminal kinase, extracellular signal-regulated kinase (ERK), mitogen-activated extracellular signal-regulated kinase (MEK), focal adhesion kinase (FAK), phosphatidylinositol 3-kinase (PI3K), Akt (also known as PKB, protein kinase B), mammalian target of rapamycin (mTOR), cleaved caspase3, Bcl2 and Bax were detected by western blotting. The relative area of lipids and monocytes/macrophages in the shWISP1 group increased compared with that of the NC group. However, the relative area of smooth muscle cell and collagen in the IvWISP1 group increased compared with that in the NC group. Therefore, WISP1 could stabilize atherosclerotic plaques. Besides, WISP1 accelerate the migration and proliferation of VSMCs via integrin α5β1 and FAK/MEK/ERK signalling pathways. In addition, WISP1 can inhibit the apoptosis of VSMCs via the PI3K/Akt/mTOR pathway. WISP1 not only inhibits the apoptosis of VSMCs via the PI3K/Akt/mTOR pathway but also enhances the migration and proliferation of VSMCs via the integrin α5β1 and FAK/MEK/ERK pathways. Therefore, WISP1 could enhance the stability of atherosclerotic plaques.

The Lnc-RNA APPAT Suppresses Human Aortic Smooth Muscle Cell Proliferation and Migration by Interacting With MiR-647 and FGF5 in Atherosclerosis

Purpose: LncRNA-Atherosclerotic plaque pathogenesis-associated transcript (APPAT) could be detected in circulating blood and has been demonstrated to correlate with the development of atherosclerosis in our previous work. It could be a potential noninvasive biomarker for earlier diagnoses of clinical cardiovascular disease. Moreover, the expression of miR-647 increased in ox-LDL-treated vascular smooth muscle cells and peripheral blood of patients with coronary heart disease. A negative correlation between APPAT and miR-647 was confirmed, and FGF5 was screened as molecular target of miR-647. However, it is largely unclear how APPAT, miR-647, and FGF5 interact and function in disease development. Here, we aim to explore the underlying molecular mechanism in this progression. Materials and Methods: APPAT, miR-647, and FGF5 expression levels were detected by quantitative reverse transcription polymerase chain reaction; cell proliferation was detected by EdU incorporation assay; cell migration was detected by wound-healing assay; the molecular interaction of APPAT/FGF5 with miR-647 was verified by dual-luciferase reporter assay; the western blot was performed to determine the gene expression at protein levels; subcellular localizations of APPAT and miR-647 were observed by fluorescence in situ hybridization; cytosolic and nucleus fractionation assay was performed to further detect the distribution of miR-647. Results: APPAT and miR-647 have inverse effects on human aortic smooth muscle cells’ (HASMCs) proliferation and migration. APPAT negatively regulated the cell activity, whereas miR-647 did it in a positive way (p<0.05). Three pairs of molecular interplay were found: mutual negative regulation between APPAT and miR-647, APPAT downregulated FGF5, miR-647 regulation on FGF5 (p<0.05). Subcellular location assay confirmed the molecular interaction of APPAT and miR-647. Conclusions: APPAT could suppress the migration and proliferation of ox-LDL-treated HASMCs via interacting with miR-647 and FGF5. We revealed a nontypical competing endogenous RNA mechanism of long noncoding RNA in the progression of atherosclerosis.

The role of extravillous trophoblasts and uterine NK cells in vascular remodeling during pregnancy.

Successful embryo implantation requires both a receptive endometrium and competent blastocysts. After implantation, the maternal decidua undergoes a series of changes, including uterine spiral artery (SA) remodeling to accommodate the fetus and provide nutrients and oxygen for the fetus to survive. Uterine spiral arteries transform from small-diameter, high-resistance arteries to large-diameter and low-resistance arteries during pregnancy. This transformation includes many changes, such as increased permeability and dilation of vessels, phenotypic switching and migration of vascular smooth muscle cells (VSMCs), transient loss of endothelial cells (ECs), endovascular invasion of extravillous trophoblasts (EVTs), and presence of intramural EVT, which are regulated by uterine NK (uNK) cells and EVTs. In this review, we mainly focus on the separate and combined roles of uNK cells and EVTs in uterine SA remodeling in establishing and maintaining pregnancy. New insight into related mechanisms will help us better understand the pathogenesis of pregnancy complications such as recurrent pregnancy loss (RPL) and preeclampsia (PE).

Mulberry Leaf and Neochlorogenic Acid Alleviates Glucolipotoxicity-Induced Oxidative Stress and Inhibits Proliferation/Migration via Downregulating Ras and FAK Signaling Pathway in Vascular Smooth Muscle Cell.

Mulberry leaf (Morus alba L.) has been used as a health food and in traditional medicine to treat several metabolic diseases, including diabetes, hypertension, and hyperlipidemia. However, the mechanism by which mulberry leaf and its functional components mediate atherosclerosis remains unclear. This study aimed to determine the effect of mulberry leaf extract (MLE) and its major component, neochlorogenic acid (nCGA), on the proliferation and migration of rat aortic vascular smooth muscle cells (VSMCs, A7r5 cell line) under diabetic cultured conditions (oleic acid and high glucose, OH). Our findings showed that MLE and nCGA significantly inhibited cell proliferation and migration in A7r5 cells as determined by a scratch wound assay and a Transwell assay. Furthermore, we observed MLE and nCGA inhibited cell proliferation and migration, such as reducing the phosphoinositide 3-kinases (PI3K)/protein kinase B (Akt), focal adhesion kinase (FAK), and small GTPase proteins using Western blot analysis. In conclusion, we confirmed the anti-atherosclerotic effects of MLE and nCGA in reducing vascular smooth muscle cell (VSMC) migration and proliferation under diabetic cultured conditions via inhibition of FAK/small GTPase proteins, PI3K/Akt, and Ras-related signaling.

Smooth muscle cell specific NEMO deficiency inhibits atherosclerosis in ApoE−/− mice

The development of atherosclerotic plaques is the result of a chronic inflammatory response coordinated by stromal and immune cellular components of the vascular wall. While endothelial cells and leukocytes are well-recognised mediators of inflammation in atherosclerosis, the role of smooth muscle cells (SMCs) remains incompletely understood. Here we aimed to address the role of canonical NF-κB signalling in SMCs in the development of atherosclerosis. We investigated the role of NF-κB signalling in SMCs in atherosclerosis by employing SMC-specific ablation of NEMO, an IKK complex subunit that is essential for canonical NF-κB activation, in ApoE−/− mice. We show that SMC-specific ablation of NEMO (NEMOSMCiKO) inhibited high fat diet induced atherosclerosis in ApoE−/− mice. NEMOSMCiKO/ApoE−/− mice developed less and smaller atherosclerotic plaques, which contained fewer macrophages, decreased numbers of apoptotic cells and smaller necrotic areas and showed reduced inflammation compared to the plaques of ApoE−/− mice. In addition, the plaques of NEMOSMCiKO/ApoE−/− mice showed higher expression of α-SMA and lower expression of the transcriptional factor KLF4 compared to those of ApoE−/− mice. Consistently, in vitro, NEMO-deficient SMCs exhibited reduced proliferation and migration, as well as decreased KLF4 expression and lower production of IL-6 and MCP-1 upon inflammatory stimulus (TNF or LPS) compared to NEMO-expressing SMCs. In conclusion, NEMO-dependent activation of NF-κB signalling in SMCs critically contributes to the pathogenesis of atherosclerosis by regulating SMC proliferation, migration and phenotype switching in response to inflammatory stimuli.

Inhibition of KIR2.1 decreases pulmonary artery smooth muscle cell proliferation and migration

The investigation of effective therapeutic drugs for pulmonary hypertension (PH) is critical. KIR2.1 plays crucial roles in regulating cell proliferation and migration, and vascular remodeling. However, researchers have not yet clearly determined whether KIR2.1 participates in the proliferation and migration of pulmonary artery smooth muscle cells (PASMCs) and its role in pulmonary vascular remodeling (PVR) also remains elusive. The present study aimed to examine whether KIR2.1 alters PASMC proliferation and migration, and participates in PVR, as well as to explore its mechanisms of action. For the in vivo experiment, a PH model was established by intraperitoneally injecting Sprague-Dawley rats monocrotaline (MCT). Hematoxylin and eosin staining revealed evidence of PVR in the rats with PH. Immunofluorescence staining and western blot analysis revealed increased levels of the KIR2.1, osteopontin (OPN) and proliferating cell nuclear antigen (PCNA) proteins in pulmonary blood vessels and lung tissues following exposure to MCT, and the TGF-β1/SMAD2/3 signaling pathway was activated. For the in vitro experiments, the KIR2.1 inhibitor, ML133, or the TGF-β1/SMAD2/3 signaling pathway blocker, SB431542, were used to pre-treat human PASMCs (HPASMCs) for 24 h, and the cells were then treated with platelet-derived growth factor (PDGF)-BB for 24 h. Scratch and Transwell assays revealed that PDGF-BB promoted cell proliferation and migration. Immunofluorescence staining and western blot analysis demonstrated that PDGF-BB upregulated OPN and PCNA expression, and activated the TGF-β1/SMAD2/3 signaling pathway. ML133 reversed the proliferation and migration induced by PDGF-BB, inhibited the expression of OPN and PCNA, inhibited the TGF-β1/SMAD2/3 signaling pathway, and reduced the proliferation and migration of HPASMCs. SB431542 pre-treatment also reduced cell proliferation and migration; however, it did not affect KIR2.1 expression. On the whole, the results of the present study demonstrate that KIR2.1 regulates the TGF-β1/SMAD2/3 signaling pathway and the expression of OPN and PCNA proteins, thereby regulating the proliferation and migration of PASMCs and participating in PVR.

ATF4 promotes brain vascular smooth muscle cells proliferation, invasion and migration by targeting miR-552-SKI axis

BackgroundStudies have indicated vascular smooth muscle cells (VSMCs) played a crucial role in atherosclerosis and microRNAs (miRNAs) played key roles in biological functions of VSMCs. Whereas, the potential function and mechanism of miR-552 in VSMCs remains unclear. Our aim was to explore the role of miR-552 on VSMCs and underlying mechanism.Material/MethodsMTT assay and transwell assay were used to measure the proliferation, invasion, and migration of human brain VSMCs (HBVSMCs) and mice VSMCs (mVSMCs), respectively. Bioinformatics tools and luciferase assay were adopted to verify the association between miR-552 and SKI. Rescue experiments were employed to assess the interaction of miR-552 and SKI in modulating biological functions in HBVSMCs and mVSMCs. The expression level of transcription factors (TFs)was measured via qRT-PCR assay. The effect of ATF4 on miR-552 and SKI expression was tested by qRT-PCR or western blot assay. Finally, chromatin immunoprecipitation (ChIP) assay and JASPAR databases were used to analyze the regulatory linkage between ATF4 and miR-552.ResultsWe found that miR-552 was upregulated in HBVSMCs treated with PDGF-bb and miR-552 overexpression could promote proliferation, invasion, and migration of HBVSMCs and mVSMCs, whereas, miR-552 knockdown had the opposite impact. In addition, we also found that SKI was a direct target of miR-552, which reversed miR-552-mediated proliferation, invasion, and migration in HBVSMCs and mVSMCs. Furthermore, we also discovered that miR-552 overexpression promoted the effects of ATF4 elevation on proliferation, migration and invasion of HBVSMCs and mVSMCs, but, miR-552 decline had the opposite impact.ConclusionsATF4-miR-552-SKI axis played critical roles in the proliferation and migration of HBVSMCs and mVSMCs, which were closely involved in atherosclerosis (AS). Therefore, our findings might offer a novel therapeutic target for AS.

Delivery of miR-654-5p via SonoVue Microbubble Ultrasound Inhibits Proliferation, Migration, and Invasion of Vascular Smooth Muscle Cells and Arterial Thrombosis and Stenosis through Targeting TCF21.

Background Abnormal proliferation of vascular smooth muscle cells (VSMCs) is an important cause of vascular stenosis. The study explored the mechanism of inhibition of vascular stenosis through the molecular mechanism of smooth muscle cell phenotype transformation. Methods Coronary heart disease-related genes were screened by bioinformatics, and the target genes of miR-654-5p were predicted by dual-luciferase method and immunofluorescence method. miR-654-5p mimic stimulation and transfection of TCF21 and MTAP into cells. SonoVue microbubble sonication was used to deliver miR-654-5p into cells. Cell proliferation, migration, and invasion were detected by CCK-8, wound scratch, and Transwell. HE and IHC staining were performed to study the effect of miR-654-5p delivery via SonoVue microbubble ultrasound on vessel stenosis in a model of arterial injury. Gene expression was determined by qRT-PCR and WB. Results TCF21 and MTAP were predicted as the target genes of miR-654-5p. Cytokines induced smooth muscle cell proliferation, migration, and invasion and promoted miR-654-5p downregulation; noticeably, downregulated miR-654-5p was positively associated with the cell proliferation and migration. Overexpression of TCF21 promoted proliferation, invasion, and migration, and mimic reversed such effects. miR-654-5p overexpression delivered by SonoVue microbubble ultrasound inhibited proliferation, migration, and invasion of cells. Moreover, in arterial injury model, we found that SonoVue microbubble ultrasound transmitted miR-654-5p into the arterial wall to inhibit arterial thrombosis and stenosis, while TCF21 was inhibited. Conclusion Ultrasound delivery of miR-654-5p via SonoVue microbubbles was able to inhibit arterial thrombosis and stenosis by targeting TCF21.