Smooth Muscle Cell Plasticity Research Articles

Pulse Pressure, Arterial Stiffness, and End-Organ Damage

Whereas larger arteries participate in the dampening of blood pressure (BP) oscillations resulting from intermittent ventricular ejection, smaller arteries steadily deliver an adequate supply of blood from the heart to the peripheral organs. Numerous active mechanisms are involved in this process. Cyclic stress acts differently from steady stress, inducing stronger and stiffer material of the vessel wall than under static conditions. Cyclic strain participates in the phenotypic plasticity of smooth muscle cells, initiates transduction mechanisms and induces the transcriptional profile of mechanically induced genes. Finally, the autoregulatory mechanism protecting the brain, heart and kidney from cardiovascular (CV) damage differ markedly according to their localization. Whereas the heart is dependent on pulsatile forces, owing to the diastolic perfusion of coronary arteries, the brain and the kidney are rather influenced by steady mechanical forces. For the kidney, the transmission of pulsatile pressure may greatly contribute to glomerular sclerosis in the elderly.

Physiological and pathological role of the ubiquitin-proteasome system in the vascular smooth muscle cell

Vascular smooth muscle cell (VSMC) plasticity implies a capacity for rapid change and adaptability through processes requiring protein turnover. The ubiquitin-proteasome system (UPS) is at the core of protein turnover as the main pathway for the degradation of proteins related to cell-cycle regulation, signalling, apoptosis, and differentiation. This review briefly addresses some structural UPS aspects under the perspective of VSMC (patho)biology. The UPS loss-of-function promotes direct cell effects and many indirect effects related to the adaptation to apoptosis/survival signalling, oxidative stress, and endoplasmic reticulum stress. The UPS regulates redox homeostasis and is redox-regulated. Also, the UPS closely interacts with endoplasmic reticulum (ER) homeostasis as the effector of un/misfolded protein degradation, and ER stress is strongly involved in atherosclerosis. Inhibition of cell cycle-controlling ubiquitin ligases or the proteasome reduces VSMC proliferation and prevents modulation of their synthetic phenotype. Proteasome inhibition also strongly promotes VSMC apoptosis and reduces neointima. In atherosclerosis models, proteasome inhibitors display vasculoprotective effects and reduce inflammation. However, worsening of atherosclerosis or vascular dysfunction has also been reported. Proteasome inhibitors sensitize VSMC to increased ER stress-mediated cell death and suppress unfolded protein response signalling. Taken together, these observations show that the UPS has powerful effects in the control of VSMC phenotype and survival signalling. However, more profound knowledge of mechanisms is needed in order to render the UPS an operational therapeutic target.

Genes that Confer the Identity of the Renin Cell

Renin-expressing cells modulate BP, fluid-electrolyte homeostasis, and kidney development, but remarkably little is known regarding the genetic regulatory network that governs the identity of these cells. Here we compared the gene expression profiles of renin cells with most cells in the kidney at various stages of development as well as after a physiologic challenge known to induce the transformation of arteriolar smooth muscle cells into renin-expressing cells. At all stages, renin cells expressed a distinct set of genes characteristic of the renin phenotype, which was vastly different from other cell types in the kidney. For example, cells programmed to exhibit the renin phenotype expressed Akr1b7, and maturing cells expressed angiogenic factors necessary for the development of the kidney vasculature and RGS (regulator of G-protein signaling) genes, suggesting a potential relationship between renin cells and pericytes. Contrary to the plasticity of arteriolar smooth muscle cells upstream from the glomerulus, which can transiently acquire the embryonic phenotype in the adult under physiologic stress, the adult juxtaglomerular cell always possessed characteristics of both smooth muscle and renin cells. Taken together, these results identify the gene expression profile of renin-expressing cells at various stages of maturity, and suggest that juxtaglomerular cells maintain properties of both smooth muscle and renin-expressing cells, likely to allow the rapid control of body fluids and BP through both contractile and endocrine functions.

Smooth Muscle Biomechanics and Plasticity: Relevance for Vascular Calibre and Remodelling

Blood vessel structure and calibre are not static. Rather, vessels remodel continuously in response to their biomechanical environment. Vascular calibre is dictated by the amount, composition and organization of the elastic extracellular matrix. In addition, the amount and organization of contractile smooth muscle cell (SMC) also need to be regulated. The SMCs are organized such that maximum contractile force generally occurs at diameters slightly below the diameter at full dilation and physiological pressure. Thus, in a remodelling vessel, not only the matrix but also the SMCs need to undergo structural adaptation. Surprisingly little is known on the adaptation of SMC contractile properties in the vasculature. The purpose of this review is to explore this SMC plasticity in the context of vascular remodelling. While not much work on this has been carried out on blood vessels, SMC plasticity is more extensively studied on other hollow structures such as airway and bladder. We therefore include studies on bladder and airway SMCs because of their possible relevance for vascular SMC behaviour. Here, plasticity is thought to form an adaptation allowing maintained function despite large volume changes. In blood vessels, the general match of active and passive diameter-tension relations suggests that SMC plasticity is part of normal vascular physiological adaptation. Vascular SMCs display similar processes and forms of adaptation as seen in nonvascular SMCs. This may become particularly relevant under strong vasoconstriction, when inward cytoskeletal adaptation possibly prevents immediate full dilation. This may contribute to structural inward remodelling as seen in hypertension and flow reduction.

MicroRNA-26a is a novel regulator of vascular smooth muscle cell function.

Aberrant smooth muscle cell (SMC) plasticity has been implicated in a variety of vascular disorders including atherosclerosis, restenosis, and abdominal aortic aneurysm (AAA) formation. While the pathways governing this process remain unclear, epigenetic regulation by specific microRNAs (miRNAs) has been demonstrated in SMCs. We hypothesized that additional miRNAs might play an important role in determining vascular SMC phenotype. Microarray analysis of miRNAs was performed on human aortic SMCs undergoing phenotypic switching in response to serum withdrawal, and identified 31 significantly regulated entities. We chose the highly conserved candidate miRNA-26a for additional studies. Inhibition of miRNA-26a accelerated SMC differentiation, and also promoted apoptosis, while inhibiting proliferation and migration. Overexpression of miRNA-26a blunted differentiation. As a potential mechanism, we investigated whether miRNA-26a influences TGF-β-pathway signaling. Dual-luciferase reporter assays demonstrated enhanced SMAD signaling with miRNA-26a inhibition, and the opposite effect with miRNA-26a overexpression in transfected human cells. Furthermore, inhibition of miRNA-26a increased gene expression of SMAD-1 and SMAD-4, while overexpression inhibited SMAD-1. MicroRNA-26a was also found to be downregulated in two mouse models of AAA formation (2.5- to 3.8-fold decrease, P < 0.02) in which enhanced switching from contractile to synthetic phenotype occurs. In summary, miRNA-26a promotes vascular SMC proliferation while inhibiting cellular differentiation and apoptosis, and alters TGF-β pathway signaling. MicroRNA-26a represents an important new regulator of SMC biology and a potential therapeutic target in AAA disease.

Endothelin‐1 induces smooth muscle cell plasticity in small mesenteric arteries

AimSmall arteries show eutrophic inward remodeling in response to many physiological and pathological stimuli. We hypothesized that early events include repositioning and adapted architecture of smooth muscle cells, reflected by shifts in active length‐tension relations.MethodsRat mesenteric arteries were incubated overnight in a wire myograph at low (0.4 D100) or high (1.1 D100) distension with or without 10 nM endothelin‐1 (ET‐1). D100 is the passive diameter at an equivalent pressure of 100 mmHg. Before and after incubation, active (125 mM K+) and passive force‐length relationships were determined by stepwise distension of the vessel.ResultsSegments (n=10) incubated with ET‐1 at 0.4 D100 showed a marked leftward shift of the force‐length curve. Optimal force developed at 0.90±0.02 D100 (day 0) versus 0.84±0.02 D100 (day 1, p=0.03). This shift did not occur in the absence of ET‐1. At 1.1 D100 incubation, slight rightward shifts occurred.Conclusionmaintained activation at low diameter shifts tension‐generating properties, possibly forming an early event in inward remodeling. This study was performed within the framework of the Dutch Top Institute Pharma project T2‐108.

MicroRNA control of podosome formation in vascular smooth muscle cells in vivo and in vitro

Smooth muscle cell (SMC) plasticity plays an important role during development and in vascular pathologies such as atherosclerosis and restenosis. It was recently shown that down-regulation of microRNA (miR)-143 and -145, which are coexpressed from a single promoter, regulates the switch from contractile to synthetic phenotype, allowing SMCs to migrate and proliferate. We show in this study that loss of miR-143/145 in vitro and in vivo results in the formation of podosomes, which are actin-rich membrane protrusions involved in the migration of several cell types, including SMCs. We further show that platelet-derived growth factor (PDGF) mediates podosome formation in SMCs through the regulation of miR-143/145 expression via a pathway involving Src and p53. Moreover, we identify key podosome regulators as targets of miR-143 (PDGF receptor alpha and protein kinase C epsilon) and miR-145 (fascin). Thus, dysregulation of the miR-143 and -145 genes is causally involved in the aberrant SMC plasticity encountered during vascular disease, in part through the up-regulation of an autoregulatory loop that promotes podosome formation.

MicroRNAs in Angiogenesis and Vascular Smooth Muscle Cell Function

The Human Genome Project was barely completed in 2003 before a new quest was launched: this time, in a search for short stretches of RNA called microRNA. The belief that these miniature molecules, which had only been discovered a decade earlier, were major regulators of genetic function gained momentum in the intervening years and sparked a worldwide interest in the physiologic and pathologic roles of microRNAs in the human body. These single-stranded, noncoding molecules of RNA, spanning to lengths of only 25 nucleotides, appear to modulate a range of cellular events by modifying the primary function of messenger RNA (mRNA): protein synthesis. The first regulatory RNA, lin-4, was discovered by Victor Ambros in 1993. Later, Andy Fire, Craig Mello, and David Baulcombe discovered small interfering RNA (siRNAs) and established the role of small noncoding mRNA in regulating cell function under a variety of conditions. The field has exploded in directions that no one could have predicted. We now know that small RNAs are expressed in most eukaryotic cells and regulate a remarkable number of cellular functions.1 The exact mechanism of action of microRNAs is unclear; however, it is believed that these molecules bind to select, noncoding regions of mRNA through traditional Watson and Crick base pairing, repressing or increasing expression of the genetic transcript and its corresponding protein. It is theorized that each molecule of microRNA regulates more than 100 distinct molecules of mRNA, with unknown but potentially far-reaching effects in the human body.2 Already multiple reports implicate an array of distinct microRNAs in cardiac and skeletal muscle myogenesis, apoptosis, regeneration, hypertrophy, fibrosis, and cardiac function. Investigators use microRNA expression profiling to identify specific microRNAs in a host of human and animal cells. Quantitative mass spectrometry measures protein levels in cells following the introduction of a specific microRNA …

The knockout of miR-143 and -145 alters smooth muscle cell maintenance and vascular homeostasis in mice: correlates with human disease

Mechanisms controlling vascular smooth muscle cell (VSMC) plasticity and renewal still remain to be completely elucidated. A class of small RNAs called microRNAs (miRs) regulate gene expression at the post-transcriptional level. Here we demonstrate a critical role of the miR-143/145 cluster in SMC differentiation and vascular pathogenesis, also through the generation of a mouse model of miR-143 and -145 knockout. We determined that the expression of miR-143 and -145 is decreased in acute and chronic vascular stress (transverse aortic constriction and in aortas of the ApoE knockout mouse). In human aortic aneurysms, the expression of miR-143 and -145 was significantly decreased compared to control aortas. In addition, overexpression of miR-143 and -145 decreased neointimal formation in a rat model of acute vascular injury. An in-depth analysis of the miR-143/145 knockout mouse model demonstrated that this miR cluster is expressed mostly in the SMC compartment, both during development and post-natally, in vessels and SMC-containing organs. Loss of miR-143 and miR-145 expression induces structural modifications of the aorta, due to an incomplete differentiation of VSMCs. In conclusion, our results demonstrate that the miR-143/145 gene cluster plays a critical role during SMC differentiation and strongly suggest its involvement in the reversion of the VSMC differentiation phenotype that occurs during vascular disease.

Smooth(ing) Muscle Differentiation by MicroRNAs

In a recent report in Nature, Cordes et al. (2009) demonstrate that miR-143 and miR-145 modulate smooth muscle cell (SMC) plasticity in part by regulating key transcription factors involved in SMC fate determination.

MiR-145 and miR-143 regulate smooth muscle cell fate and plasticity

SUMMARYmicroRNAs are regulators of myriad cellular events, but evidence for a single microRNA that can efficiently differentiate multipotent cells into a specific lineage or regulate direct reprogramming of cells into an alternate cell fate has been elusive. Here, we show that miR-145 and miR-143 are co-transcribed in multipotent cardiac progenitors before becoming localized to smooth muscle cells, including neural crest stem cell–derived vascular smooth muscle cells. miR-145 and miR-143 were direct transcriptional targets of serum response factor, myocardin and Nkx2.5, and were downregulated in injured or atherosclerotic vessels containing proliferating, less differentiated smooth muscle cells. miR-145 was necessary for myocardin-induced reprogramming of adult fibroblasts into smooth muscle cells and sufficient to induce differentiation of multipotent neural crest stem cells into vascular smooth muscle. Furthermore, miR-145 and miR-143 cooperatively targeted a network of transcription factors, including Klf4, myocardin, and Elk-1 to promote differentiation and repress proliferation of smooth muscle cells. These findings demonstrate that miR-145 can direct the smooth muscle fate and that miR-145 and miR-143 function to regulate the quiescent versus proliferative phenotype of smooth muscle cells.

Real‐time imaging of Hsp27‐cfp redistribution in response to oxidative stress

The small heat shock protein, hsp27, has been suggested to regulate actin cytoskeleton structure and modulate the interaction of actin and myosin, given its biochemical properties and high expression levels in smooth muscle cells (SMCs) under stress. This may contribute to the mechanical plasticity of SMCs. It is proposed that hsp27's putative interaction with the actin cytoskeleton and its subcellular localization is regulated by its phosphorylation via the p38 MAP kinase pathway (MAPK). This pathway is activated by mechanical and oxidative stress, like hydrogen peroxide, and can be specifically inhibited with SB203580 (SB). We have established a real‐time oxidative stress smooth muscle cell imaging protocol utilizing time lapse microscopy to examine cellular redistribution of hsp27‐cfp in response to 200μM hydrogen peroxide +/‐ SB. Immunoanalysis of endogenous and ectopically expressed hsp27 demonstrates its phosphorylation and p38 activation. Interestingly, Hsp27‐cfp translocated to the perinuclear region with oxidative stress only with SB treatment. These results illustrate the dynamic and complex nature of hsp27 in the oxidative stress response of vascular smooth muscle cells.

Directing phenotype of vascular smooth muscle cells using electrically stimulated conducting polymer

Vascular smooth muscle cells (VSMCs) isolated from rabbit aorta and immortalised A7r5 cells were cultured on conducting polypyrrole (PPy) substrates and were subjected to a 50 μA sinusoidal electrical stimulation at 0.05, 5 and 500 Hz. These substrates were doped with hyaluronic acid and coated with collagen IV followed by Matrigel ® in order to mimic the basement membrane and encourage cell attachment. Increased proliferation and expression of smooth muscle phenotype markers (smooth muscle α-actin and smooth muscle myosin heavy chain) were observed in cultures stimulated at 5 and 500 Hz. This increased proliferation and expression of contractile proteins were found to be significantly decreased when L-type voltage-gated calcium channels (VGCC) were blocked with the drug nifedipine. To the best of our knowledge, this is the first work that demonstrates that VSMCs cultured on a conducting polymer substrate and subject to electrical stimulation not only exhibit enhanced proliferation but can be simultaneously encouraged to increase contractile protein expression. This behaviour is somewhat contrary to the classical definition of smooth muscle contractile and synthetic phenotypes that, in general, requires a modulation in phenotype as a prerequisite for smooth muscle proliferation. This interesting result highlights both the inherent plasticity of vascular smooth muscle cells and the potential of electrical stimulation via conducting polymer substrates to manipulate their behaviour.

Regulation of smooth muscle excitation and contraction

Smooth muscle cells (SMC) make up the muscular portion of the gastrointestinal (GI) tract from the distal oesophagus to the internal anal sphincter. Coordinated contractions of these cells produce the motor patterns of GI motility. Considerable progress was made during the last 20 years to understand the basic mechanisms controlling excitation-contraction (E-C) coupling. The smooth muscle motor is now understood in great molecular detail, and much has been learned about the mechanisms that deliver and recover Ca2+ during contractions. The majority of Ca2+ that initiates contractions comes from the external solution and is supplied by voltage-dependent Ca2+ channels (VDCC). VDCC are regulated largely by the effects of K+ and non-selective cation conductances (NSCC) on cell membrane potential and excitability. Ca2+ entry is supplemented by release of Ca2+ from IP(3) receptor-operated stores and by mechanisms that alter the sensitivity of the contractile apparatus to changes in cytoplasmic Ca2+. Molecular studies of the regulation of smooth muscle have been complicated by the plasticity of SMC and difficulties in culturing these cells without dramatic phenotypic changes. Major questions remain to be resolved regarding the details of E-C coupling in human GI smooth muscles. New discoveries regarding molecular expression that give GI smooth muscle their unique properties, the phenotypic changes that occur in SMC in GI motor disorders, tissue engineering approaches to repair or replace defective muscular regions, and molecular manipulations of GI smooth muscles in animals models and in cell culture will be topics for exciting investigations in the future.

The effects of matrix stiffness and RhoA on the phenotypic plasticity of smooth muscle cells in a 3-D biosynthetic hydrogel system

Studies using 2-D cultures have shown that the mechanical properties of the extracellular matrix (ECM) influence cell migration, spreading, proliferation, and differentiation; however, cellular mechanosensing in 3-D remains under-explored. To investigate this topic, a unique biomaterial system based on poly(ethylene glycol)-conjugated fibrinogen was adapted to study phenotypic plasticity in smooth muscle cells (SMCs) as a function of ECM mechanics in 3-D. Tuning the compressive modulus between 448 and 5804 Pa modestly regulated SMC cytoskeletal assembly in 3-D, with spread cells in stiff matrices having a slightly higher degree of F-actin bundling after prolonged culture. However, vinculin expression in all 3-D conditions was qualitatively low and was not assembled into the classic focal adhesions typically seen in 2-D cultures. Given the evidence that RhoA-mediated cytoskeletal contractility represents a critical node in mechanosensing, we molecularly upregulated contractility by inducing SMCs to express constitutively active RhoA. In these cells, F-actin bundling and total vinculin expression increased, and focal adhesion-like structures began to emerge, consistent with RhoA's mechanism of action in cells cultured on 2-D substrates. Furthermore, SMC proliferation in 3-D did not depend significantly on matrix stiffness, and was reduced by constitutive activation of RhoA irrespective of ECM mechanical properties. Conversely, the expression of contractile markers globally increased with constitutive RhoA activation and depended on 3-D matrix stiffness only in cells with heightened RhoA activity. Combined, these data suggest that the synergistic effects of ECM mechanics and RhoA activity on SMC phenotype in 3-D are distinct from those in 2-D, and highlight the importance of studying the mechanical role of cell–matrix interactions in tunable 3-D environments.

Control of Phenotypic Plasticity of Smooth Muscle Cells by Bone Morphogenetic Protein Signaling through the Myocardin-related Transcription Factors

Vascular smooth muscle cells (VSMCs), unlike other muscle cells, do not terminally differentiate. In response to injury, VSMCs change phenotype, proliferate, and migrate as part of the repair process. Dysregulation of this plasticity program contributes to the pathogenesis of several vascular disorders, such as atherosclerosis, restenosis, and hypertension. The discovery of mutations in the gene encoding BMPRII, the type II subunit of the receptor for bone morphogenetic proteins (BMPs), in patients with pulmonary arterial hypertension (PAH) provided an indication that BMP signaling may affect the homeostasis of VSMCs and their phenotype modulation. Here we report that BMP signaling potently induces SMC-specific genes in pluripotent cells and prevents dedifferentiation of arterial SMCs. The BMP-induced phenotype switch requires intact RhoA/ROCK signaling but is not blocked by inhibitors of the TGFbeta and PI3K/Akt pathways. Furthermore, nuclear localization and recruitment of the myocardin-related transcription factors (MRTF-A and MRTF-B) to a smooth muscle alpha-actin promoter is observed in response to BMP treatment. Thus, BMP signaling modulates VSMC phenotype via cross-talk with the RhoA/MRTFs pathway, and may contribute to the development of the pathological characteristics observed in patients with PAH and other obliterative vascular diseases.

Rapamycin Promotes Vascular Smooth Muscle Cell Differentiation through Insulin Receptor Substrate-1/Phosphatidylinositol 3-Kinase/Akt2 Feedback Signaling

The phenotypic plasticity of mature vascular smooth muscle cells (VSMCs) facilitates angiogenesis and wound healing, but VSCM dedifferentiation also contributes to vascular pathologies such as intimal hyperplasia. Insulin/insulin-like growth factor I (IGF-I) is unique among growth factors in promoting VSMC differentiation via preferential activation of phosphatidylinositol 3-kinase (PI3K) and Akt. We have previously reported that rapamycin promotes VSMC differentiation by inhibiting the mammalian target of rapamycin (mTOR) target S6K1. Here, we show that rapamycin activates Akt and induces contractile protein expression in human VSMC in an insulin-like growth factor I-dependent manner, by relieving S6K1-dependent negative regulation of insulin receptor substrate-1 (IRS-1). In skeletal muscle and adipocytes, rapamycin relieves mTOR/S6K1-dependent inhibitory phosphorylation of IRS-1, thus preventing IRS-1 degradation and enhancing PI3K activation. We report that this mechanism is functional in VSMCs and crucial for rapamycin-induced differentiation. Rapamycin inhibits S6K1-dependent IRS-1 serine phosphorylation, increases IRS-1 protein levels, and promotes association of tyrosine-phosphorylated IRS-1 with PI3K. A rapamycin-resistant S6K1 mutant prevents rapamycin-induced Akt activation and VSMC differentiation. Notably, we find that rapamycin selectively activates only the Akt2 isoform and that Akt2, but not Akt1, is sufficient to induce contractile protein expression. Akt2 is required for rapamycin-induced VSMC differentiation, whereas Akt1 appears to oppose contractile protein expression. The anti-restenotic effect of rapamycin in patients may be attributable to this unique pattern of PI3K effector regulation wherein anti-differentiation signals from S6K1 are inhibited, but pro-differentiation Akt2 activity is promoted through an IRS-1 feedback signaling mechanism.

CGMP-Dependent Protein Kinase I and Smooth Muscle Relaxation

See related article, pages 1096–1103 The maintenance of vascular tone is central to the regulation of blood pressure and tissue perfusion and plays a role in the pathogenesis of hypertension and atherosclerosis. Vascular tone is determined by the balance of vasodilator and vasoconstrictor stimuli. After several decades of research, the NO/cGMP/cGMP-dependent protein kinase (cGK) pathway is now recognized as an important mediator of vasodilation. However, the mechanisms by which cGK causes smooth muscle relaxation continue to be an important question. Smooth muscle contraction and relaxation are tightly coupled to the phosphorylation and dephosphorylation, respectively, of the regulatory myosin light chain.1 Myosin light chain phosphorylation state is determined by the relative activities of myosin light chain kinase (MLCK) and myosin light chain phosphatase (MLCP). MLCK phosphorylates MLC leading to contraction,2 and MLCP dephosphorylates MLC, leading to relaxation3 (Figure). Both MLCK and MLCP activities are highly regulated. MLCK activity is activated by the binding of calcium/calmodulin and thus is the primary mechanism linking intracellular calcium concentration to smooth muscle contractility.4 MLCP activity is regulated by both vasodilator and vasoconstrictor stimuli, and is therefore responsible for much of the calcium-independent regulation of contractility (reviewed in5). Figure. MLC phosphorylation determines smooth muscle contractility. Contractile agonists lead to inositol 1,4,5 triphosphate (IP3) production or activation of RhoA (RhoA-GTP). IP3 binding to its receptor in the sarcoplasmic reticulum leads to release of Ca2+. Ca2+/calmodulin binds to and activates MLCK, which in turn phosphorylates MLC (calcium-dependent contraction). Activated RhoA binds to and activates ROCK, leading to phosphorylation and inhibition of MLCP, inhibiting MLC dephosphorylation (calcium-independent contraction). cGKI mediates relaxation by inhibiting both calcium-dependent and -independent contraction. cGKIα activates MLCP by a direct interaction and by inhibition of RhoA, and activates RGS2 to inhibit Gαq …

MCAT Got YouR TEF?

See related article, pages 883–892 Adult smooth muscle cells are highly plastic and can exhibit a range of phenotypes in response to different environmental and developmental cues. Their phenotypes can range from quiescent highly contractile cells with high levels of characteristic contractile protein isoforms, to highly proliferative cells that secrete large amounts of extracellular matrix and express only low levels of smooth muscle-specific isoforms of contractile proteins. These two states are often referred to as differentiated and dedifferentiated or phenotypically modulated states.1 In reality the situation is more complex, with smooth muscle cells likely existing in a continuum of phenotypes between these two extremes. This smooth muscle cell plasticity often makes the unequivocal identification of smooth muscle cells challenging, particularly for the more dedifferentiated smooth muscle cells that are very similar to fibroblasts. The situation is further complicated by the existence of cell types with phenotypes part way between a fibroblast and a fully differentiated adult smooth muscle cell, namely myoepithelial and myofibroblast cells, and pericytes.2–4 A major challenge to developmental biologists is determining how these cells relate to each other, determining whether they are derived from common or distinct precursors, and whether myofibroblasts or pericytes can become smooth muscle cells. One approach to begin to answer these questions is to determine the molecular mechanisms that control the phenotype of each of these cell types. A new study by Gan and colleagues,5 described in this issue of Circulation Research , provides definitive molecular evidence that distinguishes myofibroblasts from adult smooth muscle cells by the distinct transcriptional mechanisms that they use to direct expression of a shared molecular marker, smooth muscle α-actin. In this elegant study, the authors demonstrate that the binding of RTEF-1 to MCAT elements within the smooth muscle α-actin promoter is required for transcription …

Phenotypical Plasticity of Vascular Smooth Muscle Cells—Effect ofIn VitroandIn VivoShear Stress for Tissue Engineering of Blood Vessels

Vascular smooth muscle cells (vSMCs) can switch between a contractile (differentiated) and a synthetic (dedifferentiated) phenotype. Synthetic, proliferative vSMCs are observed during embryogenesis, wound repair, and tissue engineering. The potential of isolated vSMCs to reverse this phenotypic modulation depends strictly on culture conditions. Previous studies have demonstrated that applied shear stress is an important signal for vSMC phenotype. The objective of this study was to determine whether applied shear stress is capable of triggering re-differentiation of vSMCs in tissue-engineered aortas. vSMCs were isolated from ovine arteries. Cells were cultured statically or exposed to two- (2D) and three-dimensional (3D) shear stress after seeding on a tubular matrix. For 3D in vivo testing, grafts were seeded additionally with endothelial cells and implanted in the descending aorta. Particular attention was paid to the expression pattern of vSMC markers, cell ultra-structure, matrix remodeling activity, and proliferative activity. Cultured vSMCs de-differentiated during static in vitro culture, but 2D and 3D in vitro shear stress promoted re-expression of vSMC markers. During in vivo culture, vSMCs progressed toward a fully differentiated phenotype. Cells were expressing markers of differentiated vSMCs and resembled a morphologically contractile vSMC phenotype. Matrix remodeling activity and proliferative activity decreased. This study demonstrates the phenotypic plasticity of vSMCs and their ability to return to a differentiated phenotype under shear stress conditions. These results are crucial for tissue engineering of blood vessels, because they indicate for the first time the in vitro potential to regain physiological functionality of isolated vSMCs.