Smooth Muscle Cell Phenotypic Switching Research Articles

Effect of monocytes on NLRP3 inflammasome activation in vascular smooth muscle cells phenotypic switch and foam cells formation in atherosclerosis

Effect of monocytes on NLRP3 inflammasome activation in vascular smooth muscle cells phenotypic switch and foam cells formation in atherosclerosis

Krüppel-like factor 4-mediated smooth muscle cell phenotype switching to a galectin-3 positive subclass is a detrimental event in the pathogenesis of atherosclerotic cardiovascular disease

Krüppel-like factor 4-mediated smooth muscle cell phenotype switching to a galectin-3 positive subclass is a detrimental event in the pathogenesis of atherosclerotic cardiovascular disease

MiR-126-5p promotes contractile switching of aortic smooth muscle cells by targeting VEPH1 and alleviates Ang II-induced abdominal aortic aneurysm in mice

AbstractAbdominal aortic aneurysm (AAA) is a potential lethal disease that is defined by an irreversible dilatation (>50%) of the aorta. During AAA expansion, the aortic wall is often remodeled, which is featured by extracellular matrix (ECM) degeneration, medial and adventitial inflammation, depletion and phenotypic switching of vascular smooth muscle cells (SMCs). Recent studies have suggested microRNAs as vital regulators for vascular SMC function. Our earlier work demonstrated an anti-AAA role of miR-126-5p in ApoE−/− mice infused with angiotensin (Ang) II. The present study aimed to further elucidate its role in AAA pathogenesis with a focus on aortic SMC phenotypic switching. Ventricular zone expressed PH domain containing 1 (VEPH1) was identified as a novel negative regulator for vascular SMC differentiation by our group, and its expression was negatively correlated to miR-126-5p in mouse abdominal aortas based on the present microarray data. In vivo, in addition attenuating Ang II infusion-induced aortic dilation and elastin degradation, miR-126-5p agomirs also significantly reduced the expression of VEPH1. In vitro, to induce synthetic transition of human aortic smooth muscle cells (hAoSMCs), cells were stimulated with 1 μM Ang II for 24 h. Ectopic overexpression of miR-126-5p restored the differentiation of hAoSMCs—the expression of contractile/differentiated SMC markers, MYH11, and α-SMA, increased, whilst that of synthetic/dedifferentiated SMC markers, PCNA and Vimentin, decreased. Both mus and homo VEPH1 genes were validated as direct targets for miR-126-5p. VEPH1 re-expression impaired miR-126-5p-induced differentiation of hAoSMCs. In addition, Ang II-induced upregulation in matrix metalloproteinase (MMP)-9 and MMP2, two key proteases responsible for ECM degradation, in mouse aortas and hAoSMCs was reduced by miR-126-5p overexpression as well. Collectively, these results reveal an important, but previously unexplored, role of miR-126-5p in inhibiting AAA development-associated aortic SMC dedifferentiation.MiR-126-5p expression decreases abdominal aorta dilation in mice with Ang II-induced abdominal aortic aneurysm (AAA), and its agomirs limit experimental AAA formation. MiR-126-5p inhibits Ang II- and PDGF-BB-induced dedifferentiation of aortic smooth muscle cells (AoSMCs) in vitro. MiR-126-5p promotes contractile switching of AoSMCs exposed to Ang II by targeting VEPH1.

MicroRNA-92a -mediated endothelial to mesenchymal transition controls vein graft neointimal lesion formation

PurposeLong-term failure of vein grafts due to neointimal hyperplasia remains an important problem in coronary artery bypass graft surgery. Endothelial to mesenchymal transition (EndMT) contributes to vein graft vascular remodeling. However, there is little study on microRNA-mediated EndMT contributions to neointimal formation in vein graft. We hypothesized that microRNA-92a (miR-92a) might play an important role in determining EndMT contributions to neointimal formation. MethodsmiR-92a and EndMT-related proteins detected by qRT-PCR and Western blot in vitro and in vivo. Adeno-associated virus 6 (AAV6) delivery gene therapy was used to inhibit neointimal formation in vivo. The intimal hyperplasia of vein grafts was measured by HE staining, the expression of EndMT-related protein in vein grafts was measured by immunofluorescence. Immunohistochemistry and luciferase assay were used to detect potential targets of miR-92a. ResultsThe expression of miR-92a was found to be upregulated in neointimal hyperplasic lesions after vein grafting. Using cultured human umbilical vein endothelial cells (HUVECs), we show that TGF-β1 treatment of HUVECs significantly increased miR-92a expression and induced EndMT, characterized by suppression of endothelial-specific markers (CD31 and VE-cadherin) and an increase in mesenchymal-specific markers (a-SMA and vimentin), while inhibition of miR-92a expression blunted EndMT in cultured HUVECs. Furthermore, AAV6 mediated miR-92a suppression gene therapy effectively resulted in decreased EndMT and less neointimal formation in vein grafts in vivo. We further identified that integrin alpha 5 (ITGA5) is a potential target gene involved in the development of neointima formation in these vein grafts. ConclusionThis data suggests that neointimal formation does not solely rely on vascular smooth muscle cell phenotypic switching but is also related to EndMT, and miR-92a-mediated EndMT is an important mechanism underlying neointimal formation in vein grafts.

Suppression of miR-4463 promotes phenotypic switching in VSMCs treated with Ox-LDL

Vascular smooth muscle cell (VSMC) phenotypic switching is a hallmark of vascular remodeling that contributes to atherosclerotic diseases. MicroRNA 4463 (miR-4463) has been implicated in the development of arteriosclerosis obliterans, whereas the underlying mechanisms in VSMCs have not been fully addressed. In this study, we assessed whether miR-4463 is involved in the phenotypic switching process in VSMCs. Oxidized low-density lipoprotein (Ox-LDL, 50mg/L) was used to simulate the oxidative stress condition, and miR-4463 expression in VSMCs was detected by a quantitative polymerase chain reaction. To determine the effect of Ox-LDL-mediated regulation of miR-4463 on the phenotypic switching of VSMCs, cell counting kit-8, cell migration assays, and cytoskeleton test were performed. After using specific antagonists of c-Jun N-terminal kinase (JNK) and extracellular signal-regulated kinase (ERK), the relationship between miR-4463 and its downstream signaling proteins was explored. Ox-LDL induced oxidative stress to promote VSMC transformation from contraction to secretion, which clearly decreased the level of miR-4463. Then, downregulated miR-4463 enhanced the migration and phenotypic transformation of VSMCs and activated the phosphorylation of JNK and ERK; these effects were increased after Ox-LDL induction. As expected, inhibiting the two signaling proteins blocked the effect of the miR-4463 inhibitor combined with Ox-LDL. In addition, inhibition of miR-4463 led to the upregulation of basic fibroblast growth factor (bFGF) expression. The results of this study demonstrate that miR-4463 is a novel regulator of VSMC function in hypoxic conditions and modulates VSMC phenotypic switching via the JNK and ERK signaling pathways; bFGF may be the target gene of miR-4463.

LncRNA-modulated autophagy in plaque cells: a new paradigm of gene regulation in atherosclerosis?

The development of atherosclerosis is accompanied by the functional deterioration of plaque cells, which leads to the escalation of endothelial inflammation, abnormal vascular smooth muscle cell phenotype switching and the accumulation of lipid-laden macrophages within vascular walls. Autophagy, a highly conserved homeostatic mechanism, is critical for the delivery of cytoplasmic substrates to lysosomes for degradation. Moderate levels of autophagy prevent atherosclerosis by safeguarding plaque cells against apoptosis, preventing inflammation, and limiting the lipid burden, whereas excessive autophagy exacerbates cell damage and inflammation and thereby accelerates the formation of atherosclerotic plaques. Increasing lines of evidence suggest that long noncoding RNAs can be either beneficial or detrimental to atherosclerosis development by regulating the autophagy level. This review summarizes the research progress related to 1) the significant role of autophagy in atherosclerosis and 2) the effects of the lncRNA-mediated modulation of autophagy on the plaque cell fate, inflammation levels, proliferative capacity, and cholesterol metabolism and subsequently on atherogenesis.

Transcriptional Programming in Arteriosclerotic Disease: A Multifaceted Function of the Runx2 (Runt-Related Transcription Factor 2).

Despite successful therapeutic strategies in the prevention and treatment of arteriosclerosis, the cardiovascular complications remain a major clinical and societal issue worldwide. Increased vascular calcification promotes arterial stiffness and accelerates cardiovascular morbidity and mortality. Upregulation of the Runx2 (Runt-related transcription factor 2), an essential osteogenic transcription factor for bone formation, in the cardiovascular system has emerged as an important regulator for adverse cellular events that drive cardiovascular pathology. This review discusses the regulatory mechanisms that are critical for Runx2 expression and function and highlights the dynamic and complex cross talks of a wide variety of posttranslational modifications, including phosphorylation, acetylation, ubiquitination, and O-linked β-N-acetylglucosamine modification, in regulating Runx2 stability, cellular localization, and osteogenic transcriptional activity. How the activation of an array of signaling cascades by circulating and local microenvironmental factors upregulates Runx2 in vascular cells and promotes Runx2-mediated osteogenic transdifferentiation of vascular smooth muscle cells and expression of inflammatory cytokines that accelerate macrophage infiltration and vascular osteoclast formation is summarized. Furthermore, the increasing appreciation of a new role of Runx2 upregulation in promoting vascular smooth muscle cell phenotypic switch, and Runx2 modulated by O-linked β-N-acetylglucosamine modification and Runx2-dependent repression of smooth muscle cell-specific gene expression are discussed. Further exploring the regulation of this key osteogenic transcription factor and its new perspectives in the vasculature will provide novel insights into the transcriptional regulation of vascular smooth muscle cell phenotype switch, reprograming, and vascular inflammation that promote the pathogenesis of arteriosclerosis.

Interleukin-11 is important for vascular smooth muscle phenotypic switching and aortic inflammation, fibrosis and remodeling in mouse models

Transforming growth factor beta-1 (TGFβ1) is a major driver of vascular smooth muscle cell (VSMC) phenotypic switching, an important pathobiology in arterial disease.We performed RNA-sequencing of TGFβ1-stimulated human aortic or arterial VSMCs which revealed large and consistent upregulation of Interleukin 11 (IL11). IL11 has an unknown function in VSMCs, which highly express the IL11 receptor alpha, suggestive of an autocrine loop. In vitro, IL11 activated ERK signaling, but inhibited STAT3 activity, and caused VSMC phenotypic switching to a similar extent as TGFβ1 or angiotensin II (ANGII) stimulation. Genetic or therapeutic inhibition of IL11 signaling reduced TGFβ1- or ANGII-induced VSMC phenotypic switching, placing IL11 activity downstream of these factors. Aortas of mice with Myh11-driven IL11 expression were remodeled and had reduced contractile but increased matrix and inflammatory genes expression. In two models of arterial pressure loading, IL11 was upregulated in the aorta and neutralizing IL11 antibodies reduced remodeling along with matrix and pro-inflammatory gene expression. These data show that IL11 plays an important role in VSMC phenotype switching, vascular inflammation and aortic pathobiology.

The Role of Inflammation and Myeloperoxidase-Related Oxidative Stress in the Pathogenesis of Genetically Triggered Thoracic Aortic Aneurysms.

Genetically triggered thoracic aortic aneurysms (TAAs) are usually considered to exhibit minimal levels of inflammation. However, emerging data demonstrate that specific features of an inflammatory response can be observed in TAA, and that the extent of the inflammatory response can be correlated with the severity, in both mouse models and in human studies. Myeloperoxidase (MPO) is a key mediator of the inflammatory response, via production of specific oxidative species, e.g., the hypohalous acids. Specific tissue modifications, mediated by hypohalous acids, have been documented in multiple cardiovascular pathologies, including atherosclerosis associated with coronary artery disease, abdominal aortic, and cerebral aneurysms. Similarly, data are now emerging that show the capacity of MPO-derived oxidative species to regulate mechanisms important in TAA pathogenesis, including alterations in extracellular matrix homeostasis, activation of matrix metalloproteinases, induction of endothelial dysfunction and vascular smooth muscle cell phenotypic switching, and activation of ERK1/2 signaling. The weight of evidence supports a role for inflammation in exacerbating the severity of TAA progression, expanding our understanding of the pathogenesis of TAA, identifying potential biomarkers for early detection of TAA, monitoring severity and progression, and for defining potential novel therapeutic targets.

Insulin-like growth factor-binding protein 3 inhibits angiotensin II-induced aortic smooth muscle cell phenotypic switch and matrix metalloproteinase expression.

What is the central question of this study? Insulin-like growth factor 1 and its major binding protein insulin-like growth factor binding protein 3 (IGFBP3) are involved in collagen deregulation in several cardiovascular diseases: what is the role of IGFBP3 in thoracic aortic dissection and does it regulate aortic smooth muscle cells' phenotypic switch? What is the main finding and its importance? IGFBP3 inhibits aortic smooth muscle cells' phenotypic switch from a contractile to a synthetic phenotype, decreases matrix metalloproteinase 9 activation and suppresses elastin degradation. These findings provide a better understanding of the pathogenesis of thoracic aortic dissection. Thoracic aortic dissection (TAD) is characterized by aortic media degeneration and is a highly lethal disease. An aortic smooth muscle cell (AoSMC) phenotypic switch is considered a key pathophysiological change in TAD. Insulin-like growth factor binding protein 3 (IGFBP3) was found to be downregulated in aortic tissues of TAD patients. The present work aimed to study the function of IGFBP3 in AoSMCs' phenotypic switch and matrix metalloproteinase (MMP) expression. We established a mouse model of TAD by angiotensin (Ang) II infusion to β-aminopropionitrile-administrated mice, and found decreased IGFBP3 expression accompanied by aortic dilatation and elastin degradation in vivo. Further, mouse (m)AoSMCs were isolated from mouse thoracic aorta and treated with Ang II. Ang II induced downregulation of IGFBP3 in vitro. To further study the function of IGFBP3, primary mAoSMCs were infected with adenovirus expressing IGFBP3 followed by Ang II induction. Enforced upregulation of IGFBP3 decreased MMP9 expression and activation as well as increasing tissue inhibitor of metalloproteinase (TIMP) 1 expression in Ang II-induced mAoSMCs. No difference was observed in MMP2 and TIMP3 expression. IGFBP3 suppressed subsequent Ang II-induced elastin degradation in vitro. IGFBP3 inhibited Ang II-induced mAoSMCs' phenotypic switch as evidenced by increased smooth muscle actin α-2 (ACTA2) and myosin heavy chain 11 (MYH11) expression and decreased secreted phosphoprotein 1 (SPP1) and vimentin expression. Taken together, the present study demonstrates the role of IGFBP3 in preserving AoSMCs' contractile state and reducing MMP9 activation and thus promoting elastic fibre synthesis, which provides a better understanding of the pathogenesis of TAD.

Phenotypic Switching of Atherosclerotic Smooth Muscle Cells is Regulated by Activated PARP1-Dependent TET1 Expression.

Aim: During the development of atherosclerosis, the vascular smooth muscle cells (SMCs) undergo phenotypic switching from contractile phenotype to synthetic phenotype. This study aimed at examining the role of DNA modification mediated by the oxidative stress dependent ten eleven translocation enzymes (TETs) expression at early stage of phenotypic switching. Methods: Based on the in vitro SMCs calcification model, DNA damage, phenotypic switching and 5-hydroxymethylcytosine (5hmC) were examined by comet assay, alkaline DNA unwinding assay, immunofluorescence staining, Dot blotting and Western blotting. Then Western blotting and qRT-PCR were performed to analyze the TETs expression and the relationship between the activity of poly(ADP-ribose) polymerase 1 (PARP1) and TETs expression. We further alter 5hmC modification by inhibition of TET1 or PARP1 to rescue the phenotypic switching of SMCs using immunofluorescence staining, Dot blotting and qRT-PCR. We performed immunochemistry staining to examine the activated PARP1-TET1 pathway in vivo . Results: The phenotypic switching was observed in the SMCs cultured with calcification medium as the expression of the cell markers of contractile SMCs decreased and cell proliferation increased. In contrast, PAR and 5hmC were markedly increased in SMCs with calcification due to DNA damage. Our study further demonstrated that oxidative stress-activated PARP1, promotes TET1 expression and 5hmC increase during the phenotypic switching. Inhibition of TET1 or PARP1 can rescue the phenotypic switching of SMCs with calcification. Conclusion: Our study demonstrated the important role of PARylation dependent 5hmC, in SMCs phenotypic switching. It raises the possibility to target TET1 and PARP1 for atherosclerosis treatment.

Single-Cell Genomics Reveals a Novel Cell State During Smooth Muscle Cell Phenotypic Switching and Potential Therapeutic Targets for Atherosclerosis in Mouse and Human.

Smooth muscle cells (SMCs) play significant roles in atherosclerosis via phenotypic switching, a pathological process in which SMC dedifferentiation, migration, and transdifferentiation into other cell types. Yet how SMCs contribute to the pathophysiology of atherosclerosis remains elusive. To reveal the trajectories of SMC transdifferentiation during atherosclerosis and to identify molecular targets for disease therapy, we combined SMC fate mapping and single-cell RNA sequencing of both mouse and human atherosclerotic plaques. We also performed cell biology experiments on isolated SMC-derived cells, conducted integrative human genomics, and used pharmacological studies targeting SMC-derived cells both in vivo and in vitro. We found that SMCs transitioned to an intermediate cell state during atherosclerosis, which was also found in human atherosclerotic plaques of carotid and coronary arteries. SMC-derived intermediate cells, termed "SEM" cells (stem cell, endothelial cell, monocyte), were multipotent and could differentiate into macrophage-like and fibrochondrocyte-like cells, as well as return toward the SMC phenotype. Retinoic acid (RA) signaling was identified as a regulator of SMC to SEM cell transition, and RA signaling was dysregulated in symptomatic human atherosclerosis. Human genomics revealed enrichment of genome-wide association study signals for coronary artery disease in RA signaling target gene loci and correlation between coronary artery disease risk alleles and repressed expression of these genes. Activation of RA signaling by all-trans RA, an anticancer drug for acute promyelocytic leukemia, blocked SMC transition to SEM cells, reduced atherosclerotic burden, and promoted fibrous cap stability. Integration of cell-specific fate mapping, single-cell genomics, and human genetics adds novel insights into the complexity of SMC biology and reveals regulatory pathways for therapeutic targeting of SMC transitions in atherosclerotic cardiovascular disease.

Activation of M3AChR (Type 3 Muscarinic Acetylcholine Receptor) and Nrf2 (Nuclear Factor Erythroid 2-Related Factor 2) Signaling by Choline Alleviates Vascular Smooth Muscle Cell Phenotypic Switching and Vascular Remodeling.

Phenotypic switching of vascular smooth muscle cells (VSMCs) plays a critical role in atherosclerosis, vascular restenosis, and hypertension. Choline exerts cardioprotective effects; however, little is known about its effects on VSMC phenotypic switching and vascular remodeling. Here, we investigated whether choline modulates VSMC phenotypic changes and explored the underlying mechanisms. Approach and Results: In cultured VSMCs, choline promoted Nrf2 (nuclear factor erythroid 2-related factor 2) nuclear translocation, inducing the expression of HO-1 (heme oxygenase-1) and NQO-1 (NAD[P]H quinone oxidoreductase-1). Consequently, choline ameliorated Ang II (angiotensin II)-induced increases in NOX (NAD[P]H oxidase) expression and the mitochondrial reactive oxygen species level, thereby attenuating Ang II-induced VSMC phenotypic switching, proliferation, and migration, presumably via M3AChRs (type 3 muscarinic acetylcholine receptors). Downregulation of M3AChR or Nrf2 diminished choline-mediated upregulation of Nrf2, HO-1, and NQO-1 expression, as well as inhibition of VSMC phenotypic transformation, suggesting that M3AChR and Nrf2 activation are responsible for the protective effects of choline. Moreover, activation of the Nrf2 pathway by sulforaphane suppressed Ang II-induced VSMC phenotypic switching and proliferation, indicating that Nrf2 is a key regulator of VSMC phenotypic switching and vascular homeostasis. In a rat model of abdominal aortic constriction in vivo, choline attenuated VSMC phenotypic transformation and vascular remodeling in a manner related to activation of the Nrf2 pathway. These results reveal that choline impedes VSMC phenotypic switching, proliferation, migration, and vascular remodeling by activating M3AChR and Nrf2-antioxidant signaling and suggest a novel role for Nrf2 in VSMC phenotypic modulation.

Targeting Heme Oxygenase-1 in the Arterial Response to Injury and Disease.

Heme oxygenase-1 (HO-1) catalyzes the degradation of heme into carbon monoxide (CO), iron, and biliverdin, which is rapidly metabolized to bilirubin. The activation of vascular smooth muscle cells (SMCs) plays a critical role in mediating the aberrant arterial response to injury and a number of vascular diseases. Pharmacological induction or gene transfer of HO-1 improves arterial remodeling in animal models of post-angioplasty restenosis, vascular access failure, atherosclerosis, transplant arteriosclerosis, vein grafting, and pulmonary arterial hypertension, whereas genetic loss of HO-1 exacerbates the remodeling response. The vasoprotection evoked by HO-1 is largely ascribed to the generation of CO and/or the bile pigments, biliverdin and bilirubin, which exert potent antioxidant and anti-inflammatory effects. In addition, these molecules inhibit vascular SMC proliferation, migration, apoptosis, and phenotypic switching. Several therapeutic strategies are currently being pursued that may allow for the targeting of HO-1 in arterial remodeling in various pathologies, including the use of gene delivery approaches, the development of novel inducers of the enzyme, and the administration of unique formulations of CO and bilirubin.

Mutual regulation between β-TRCP mediated REST protein degradation and Kv1.3 expression controls vascular smooth muscle cell phenotype switch

Background and aimsPhenotypic switch of vascular smooth muscle cells (VSMC) plays a key role in the pathogenesis of atherosclerosis and restenosis after artery intervention. Transcription repressor element 1-silencing transcription factor (REST) has been identified as key regulator of VSMC proliferation. In the present study, we sought to investigate the potential association of E3-ubiquitin ligase β-TRCP mediated REST protein degradation with Kv1.3 expression during VSMC phenotypic switch. MethodsProtein and mRNA expression was measured in ex vivo and in vitro models. Protein interaction and ubiquitination were analyzed by immunoprecipitation assays. ChIP assays were performed to assess the relationship between REST and targeted DNA binding site. ResultsWe found that the expression level of E3-ubiquitin ligase β-TRCP is significantly increased during VSMC phenotypic switch. REST protein ubiquitination mediated by β-TRCP is critical for VSMC proliferation and migration. We also found that the gene KCNA3 encoding potassium channel protein Kv1.3 contains a functional REST binding site and is repressed by REST. Downregulation of REST by β-TRCP and consequently upregulation of Kv1.3 are important events during VSMC phenotypic switch. Furthermore, upregulated Kv1.3 accelerates β-TRCP modulated REST degradation through Erk1/2 signaling. ConclusionsOur results reveal a fundamental role for regulatory interactions between β-TRCP modulated REST degradation and Kv1.3 in the control of the multilayered regulatory programs required for VSMC phenotype switch.

Correction to: miR-128-3p Is a Novel Regulator of Vascular Smooth Muscle Cell Phenotypic Switch and Vascular Diseases

Correction to: miR-128-3p Is a Novel Regulator of Vascular Smooth Muscle Cell Phenotypic Switch and Vascular Diseases

Tanshinone II A attenuates vascular remodeling through klf4 mediated smooth muscle cell phenotypic switching

The aim of this study is to investigate the therapeutic role of Tanshinone II A, a key integrant from salvia miltiorrhiza, against pathological vascular remodeling. Completed ligation of mouse left common carotid arteries animal model and rat smooth muscle cells used to investigate the role of Tanshinone II A in regulating pathological vascular remodeling through hematoxylin and eosin staining, immunohistochemistry staining, immunofluorescence staining, adenovirus infection, real time PCR and western blotting. Our data demonstrated that Tanshinone II A treatment suppresses vascular injury-induced neointima formation. In vitro studies on rat smooth muscle cell indicated that Tanshinone II A treatment attenuates PDGF-BB induced cell growth, and promotes smooth muscle cell differentiated marker genes expression that induced by rapamycin treatment. Tanshinone II A treatment significant inhibits rat smooth muscle cell proliferation and migration. Tanshinone II A promotes KLF4 expression during smooth muscle phenotypic switching. Overexpression of KLF4 exacerbates Tanshinone II A mediated smooth muscle cell growth inhibition. Tanshinone II A plays a pivotal role in regulating pathological vascular remodeling through KLF4 mediated smooth muscle cell phenotypic switching. This study demonstrated that Tanshinone II A is a potential therapeutic agent for vascular diseases.

S-(–)-equol alleviates stenosis of the injured carotid artery in Sprague Dawley rats by preventing the vascular smooth muscle cell phenotypic switch via inhibition of the MAPKp38-NFκBp65 signaling

Phenotypic switching of the vascular smooth muscle cells (VSMCs) is closely related to an in-stent restenosis (ISR). This study aimed to investigate whether S-(–)-equol prevented the phenotypic switch of the VSMCs as a potential treatment of an ISR. The carotid arteries of female Sprague Dawley (SD) rats with or without a carotid injury and ovariectomy were harvested after 4 weeks of treatment with S-(–)-equol. Stenosis of the carotid artery and two phenotype-related proteins-α smooth muscle actin (αSMA) and osteopontin (OPN)-were determined. The proliferation and migration capacities of VSMCs were determined by the CCK-8 and transwell assays, respectively. The expressions of αSMA, OPN, MAPKp38, p-MAPKp38, NF-κBp65, and p-NF-κBp65 were detected by a western blot. S-(–)-equol alleviated carotid stenosis and prevented the VSMC phenotypic switch in female SD rats with a carotid artery injury and a bilateral ovariectomy. S-(–)-equol inhibited the phenotypic switch of VSMCs, which was induced by PDGF-BB, and enhanced the proliferation and migration of VSMCs. The effects of S-(–)-equol on VSMCs were confirmed to be related to the inactivation of the GPER-MAPKp38-NF-κBp65 signaling in vitro and in vivo. Our results indicate that S-(–)-equol affects carotid stenosis by preventing the phenotypic switch of VSMCs via the GPER-MAPKp38-NF-κBp65 signaling pathway.

Pyrogallol-Phloroglucinol-6,6-Bieckolon Attenuates Vascular Smooth Muscle Cell Proliferation and Phenotype Switching in Hyperlipidemia through Modulation of Chemokine Receptor 5.

Hyperlipidemia induces vascular smooth muscle cell (VSMC) proliferation and phenotype switching from contractile to synthetic. This process is involved in arterial remodeling via the chemokine ligand 5 (CCL5)/chemokine receptor 5 (CCR5) pathway. Arterial remodeling is related to atherosclerosis or intimal hyperplasia. The purpose of this study was to evaluate whether pyrogallol-phloroglucinol-6,6-bieckol (PPB) from E. cava reduces VSMC proliferation and phenotype switching via the CCL5/CCR5 pathway. The CCL5/CCR5 expression, VSMC proliferation and phenotypic alterations were evaluated using a cell model of VSMC exposed in hyperlipidemia, and an animal model of mice fed a high-fat-diet (HFD). The expression of CCL5/CCR5 increased in both the cell and animal models of hyperlipidemia. Treatment with PPB decreased CCL5/CCR5 expression in both models. The expression of contractile markers of VSMCs, including alpha-smooth muscle actin (α-SMA), smooth muscle myosin heavy chain (SM-MHC), and smooth muscle protein 22 alpha (SM22α), were decreased by hyperlipidemia and restored after treatment with PPB. The silencing of CCR5 attenuated the effects of PPB treatment. VSMC proliferation and the intima-media thickness of the aortas, increased with HFD and decreased after treatment with PPB. The VSMC proliferation ratio and messenger ribonucleic acid (mRNA) expression of cell cycle regulatory factors increased in the in vitro model and were restored after treatment with PPB. PPB treatment reduced VSMC proliferation and phenotype switching induced by hyperlipidemia through inhibition of the CCL5/CCR5 pathway.

Exosomal LINC01005 derived from oxidized low-density lipoprotein-treated endothelial cells regulates vascular smooth muscle cell phenotypic switch.

Phenotype switch of vascular smooth muscle cells (VSMCs) plays an important role in the development of atherosclerosis (AS). Endothelial cells can regulate VSMC phenotypic switch by secreting exosomes, crucial mediators of intracellular communication. This study aimed to determine whether exosomal LINC01005 from oxidized low-density lipoprotein (ox-LDL)-treated human umbilical vein endothelial cells (HUVECs) plays a role in regulating VSMC phenotypic switch and to validate the underlying molecular mechanism. Exosomes were extracted from ox-LDL-treated HUVECs (ox-LDL-Exo) and then administered into VSMCs. VSMC phenotypic switch was assessed by determining VSMC phenotypic markers using western blot. VSMC cell proliferation and migration were evaluated by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and wound healing assay, respectively. The interaction between miR-128-3p and LINC01005 or Krüppel-like factor 4 (KLF4) was analyzed by luciferase reporter assay. ox-LDL-Exo contained high expression of LINC01005. Inhibition of LINC01005 expression in ox-LDL-Exo abrogated the ox-LDL-Exo-induced VSMC phenotypic switch, proliferation, and migration. Furthermore, LINC01005 acted as a sponge of miR-128-3p to upregulate KLF4 expression. Moreover, miR-128-3p overexpression and KLF4 silencing in VSMCs attenuated the ox-LDL-Exo-induced VSMC phenotypic switch, proliferation, and migration. Collectively, exosomal LINC01005 from ox-LDL-treated HUVECs promotes VSMC phenotype switch, proliferation, and migration by regulating the miR-128-3p/KLF4 axis.