Contractile Process Research Articles

Crystal structures of cables formed by the acetylated and unacetylated forms of the Schizosaccharomyces pombe tropomyosin orthologue TpmCdc8

Cables formed by head-to-tail polymerization of tropomyosin, localized along the length of sarcomeric and cytoskeletal actin filaments, play a key role in regulating a wide range of motile and contractile processes. The stability of tropomyosin cables, their interaction with actin filaments and the functional properties of the resulting co-filaments are thought to be affected by N-terminal acetylation of tropomyosin. Here, we present high–resolution structures of cables formed by acetylated and unacetylated Schizosaccharomyces pombe tropomyosin orthologue TpmCdc8. The crystal structures represent different types of cables, each consisting of TpmCdc8 homodimers in a different conformation. The structures show how the interactions of the residues in the overlap junction contribute to cable formation and how local structural perturbations affect the conformational dynamics of the protein and its ability to transmit allosteric signals. In particular, N-terminal acetylation increases the helicity of the adjacent region, which leads to a local reduction in conformational dynamics and consequently to less fraying of the N-terminal region. This creates a more consistent complementary surface facilitating the formation of specific interactions across the overlap junction.

Therapeutic Potential of 1-(2-Chlorophenyl)-6,7-dimethoxy-3-methyl-3,4-dihydroisoquinoline.

The synthesized compound 1-(2-chlorophenyl) 6-7-dimethoxy-3-methyl-3,4-dihydroisoquinoline (DIQ) was investigated as a biological agent. Its potential to affect muscle contractility was predicted through in silico PASS analysis. Based on the in silico analysis, its capabilities were experimentally investigated. The study aimed to investigate the effects of DIQ on the ex vivo spontaneous contractile activity (CA) of smooth muscle (SM) tissue. DIQ was observed to reduce the strength of Ca2+-dependent contractions in SM preparations (SMP), possibly by increasing cytosolic Ca2+ levels through the activation of a voltage-gated L-type Ca2+ channel. DIQ potently affected calcium currents by modulating the function of muscarinic acetylcholine receptors (mAChRs) and 5-hydroxytryptamine (5-HT) receptors at a concentration of 50 μM. Immunohistochemical tests showed a 47% reduction in 5-HT2A and 5-HT2B receptor activity in SM cells and neurons in the myenteric plexus (MP), further confirming the effects of DIQ. Furthermore, a significant inhibition of neuronal activity was observed when the compound was co-administered with 5-HT to SM tissues. The conducted experiments confirm the ability of the isoquinoline analog to act as a physiologically active molecule to control muscle contractility and related physiological processes.

Post-translational modifications of vertebrate striated muscle myosin heavy chains.

Post-translational modifications (PTMs) play a crucial role in regulating the function of many sarcomeric proteins, including myosin. Myosins comprise a family of motor proteins that play fundamental roles in cell motility in general and muscle contraction in particular. A myosin molecule consists of two myosin heavy chains (MyHCs) and two pairs of myosin light chains (MLCs); two MLCs are associated with the neck region of each MyHC's N-terminal head domain, while the two MyHC C-terminal tails form a coiled-coil that polymerizes with other MyHCs to form the thick filament backbone. Myosin undergoes extensive PTMs, and dysregulation of these PTMs may lead to abnormal muscle function and contribute to the development of myopathies and cardiovascular disorders. Recent studies have uncovered the significance of PTMs in regulating MyHC function and showed how these PTMs may provide additional modulation of contractile processes. Here, we discuss MyHC PTMs that have been biochemically and/or functionally studied in mammals' and rodents' striated muscle. We have identified hotspots or specific regions in three isoforms of myosin (MYH2, MYH6, and MYH7) where the prevalence of PTMs is more frequent and could potentially play a significant role in fine-tuning the activity of these proteins.

Recent Advances in Maturation of Induced pluripotent Stem Cells-derived Cardiomyocytes

Induced pluripotent stem cells -derived cardiomyocytes (iPSC-CMs) increasingly become an important tool for studying genetic heart diseases, drug screening and heart cell therapy. However, iPSC-CMs differentiated by the traditional ways are immature and the challenge of producing fully matured iPSC-CMs that functionally resemble adult cardiomyocytes persists. Immature iPSC-CMs exhibit characteristics typical of fetal-stage cells, including less developed contractility, electrophysiological properties, and metabolic processes. These limitations restrict their clinical applications, especially in regenerative medicine. Over the past decade, numerous strategies have been developed to enhance the maturation of iPSC-CMs. These include prolonged culture periods, biochemical induction using specific hormones and energy substrates, and the application of biophysical stimuli such as electrical and mechanical stimulation. In addition, advances in biomaterials, such as extracellular matrices and hydrogels, have been crucial in replicating the physiological environment needed to support iPSC-CM development. Together, these methods aim to produce cardiomyocytes that more closely mimic adult heart cells in structure and function. This review explores recent progress in optimizing the maturation of iPSC-CMs and highlights their potential impact on future applications in cell therapy and heart disease modeling.

Load sharing between synergistic muscles characterized by a ligand-binding approach and elastography

The skeletal muscle contraction is determined by cross-bridge formation between the myosin heads and the actin active sites. When the muscle contracts, it shortens, increasing its longitudinal shear elastic modulus ({mu }_{L}). Structurally, skeletal muscle can be considered analogous to the molecular receptors that form receptor–ligand complexes and exhibit specific ligand-binding dynamics. In this context, this work aims to apply elastography and the ligand-binding framework to approach the possible intrinsic mechanisms behind muscle synergism. Based on the short-range stiffness principle and the acoustic–elasticity theory, we define the coefficient C, which is directly related to the fraction saturation of molecular receptors and links the relative longitudinal deformation of the muscle to its {mu }_{L}. We show that such a coefficient can be obtained directly from {mu }_{L} estimates, thus calculating it for the biceps brachii, brachioradialis, and brachialis muscles during isometric elbow flexion torque (τ) ramps. The resulting Cleft(tau right) curves were analyzed by conventional characterization methods of receptor–ligand systems to study the dynamical behavior of each muscle. The results showed that, depending on muscle, Cleft(tau right) exhibits typical ligand-binding dynamics during joint torque production. Therefore, the above indicates that these different behaviors describe the longitudinal shortening pattern of each muscle during load sharing. As a plausible interpretation, we suggested that this could be related to the binding kinetics of the cross-bridges during their synergistic action as torque increases. Likewise, it shows that elastography could be useful to assess contractile processes at different scales related to the change in the mechanical properties of skeletal muscle.

Thapsigargin-resistant thiacalix[4]arene C-1087-sensitive component of the contractile activity in rat myometrium reflects the functioning of plasma membrane calcium pump

Background. According to existing knowledge, thiacalix[4]arene С-1087 is highly capable of inhibiting Са2+-pump (Са2+, Mg2+-АТРase) of the plasma membrane; at the same time it inhibits the functioning of Са2+-pump of the sarcoplasmic reticulum of uterine smooth muscles to some degree. The aim of this research was to study the effects of С-1087 on the concentration of Ca2+ ions and contractile activity of the rat myometrium cells using an inhibitor of Са2+-pump of the sarcoplasmic reticulum – thapsigargin. Materials and Methods. The experiments were conducted using outbred white non-pregnant rats. The contractile activity in the preparations of longitudinal SM of uterine horns with preserved endothelium was registered in the isometric mode. To determine the changes in [Ca2+]i level, myocytes were treated with probes Hoechst 33342 (to test the nucleus of the cell) and fluo-4 AM (to test the change in Са2+-concentration in the cell). Results. The tenzometric studies with the subsequent mechanokinetic analysis demonstrated that under the action of thapsigargin (0.5 µM), thiacalix[4]arene C-1087 (10 µM) caused considerable changes in the kinetics of the spontaneous contractile activity processes in the myometrium of rats, including the decrease in the maximal contraction velocity and the increase in the maximal relaxation velocity. By means of confocal microscopy with Ca2+-sensitive fluorescent probe fluo-4, it was demonstrated that the application of thiacalix[4]arene С-1087 to immobilized myocytes of the uterus against the background of thapsigargin caused a transient spike of Са2+-signal with the subsequent turn of the intracellular concentration of Ca ions to the stable increased level. The effects of С-1087 under the action of thapsigargin regarding the relaxation phase in the spontaneous myometrium contractions were removed after the preliminary blocking of nitric oxide synthases L-NAME (100 µM). Under the action of L-NAME, thiacalix[4]arene С-1087 (10 µM) caused complete inhibition of the relaxation process in the contraction of myometrium preparations, induced by high-potassium solution (80 mM). Conclusions. The primary reason for changes in the contractile activity and Ca2+-signal in uterine myocytes under the effect of thiacalix[4]arene С-1087 is its ability to inhibit Са2+-pump of the plasma membrane; further С-1087-induced changes in the smooth muscle tissues may be caused by the increased level of Са2+ concentration in myocytes. The obtained results demonstrate thiacalix[4]arene С-1087 is a promising compound for the elaboration of pharmacological preparations for modulating the contractile activity in smooth muscles, including myometrium.

Automated Dual-Mode Cell Monitoring To Simultaneously Explore Calcium Dynamics and Contraction-Relaxation Kinetics within Drug-Treated Stem Cell-Derived Cardiomyocytes.

This manuscript proposes a new dual-mode cell imaging system for studying the relationships between calcium dynamics and the contractility process of cardiomyocytes derived from human-induced pluripotent stem cells. Practically, this dual-mode cell imaging system provides simultaneously both live cell calcium imaging and quantitative phase imaging based on digital holographic microscopy. Specifically, thanks to the development of a robust automated image analysis, simultaneous measurements of both intracellular calcium, a key player of excitation-contraction coupling, and the quantitative phase image-derived dry mass redistribution, reflecting the effective contractility, namely, the contraction and relaxation processes, were achieved. Practically, the relationships between calcium dynamics and the contraction-relaxation kinetics were investigated in particular through the application of two drugs─namely, isoprenaline and E-4031─known to act precisely on calcium dynamics. Specifically, this new dual-mode cell imaging system enabled us to establish that calcium regulation can be divided into two phases, an early phase influencing the occurrence of the relaxation process followed by a late phase, which although not having a significant influence on the relaxation process affects significantly the beat frequency. In combination with cutting-edge technologies allowing the generation of human stem cell-derived cardiomyocytes, this dual-mode cell monitoring approach therefore represents a very promising technique, particularly in the fields of drug discovery and personalized medicine, to identify compounds likely to act more selectively on specific steps that compose the cardiomyocyte contractility.

NR4A1 modulates intestinal smooth muscle cell phenotype and dampens inflammation-associated intestinal remodeling.

Stricture formation is a common complication of Crohn's disease (CD), driven by enhanced deposition of extracellular matrix (ECM) and expansion of the intestinal smooth muscle layers. Nuclear receptor subfamily 4 group A member 1 (NR4A1) is an orphan nuclear receptor that exhibits anti-proliferative effects in smooth muscle cells (SMCs). We hypothesized that NR4A1 regulates intestinal SMC proliferation and muscle thickening in the context of inflammation. Intestinal SMCs isolated from Nr4a1+/+ and Nr4a1-/- littermates were subjected to shotgun proteomic analysis, proliferation, and bioenergetic assays. Proliferation was assessed in the presence and absence of NR4A1 agonists, cytosporone-B (Csn-B) and 6-mercaptopurine (6-MP). In vivo, we compared colonic smooth muscle thickening in Nr4a1+/+ and Nr4a1-/- mice using the chronic dextran sulfate sodium (DSS) model of colitis. Second, SAMP1/YitFc mice (a model of spontaneous ileitis) were treated with Csn-B and small intestinal smooth muscle thickening was assessed. SMCs isolated from Nr4a1-/- mice exhibited increased abundance of proteins related to cell proliferation, metabolism, and ECM production, whereas Nr4a1+/+ SMCs highly expressed proteins related to the regulation of the actin cytoskeleton and contractile processes. SMCs isolated from Nr4a1-/- mice exhibited increased proliferation and alterations in cellular metabolism, whereas activation of NR4A1 attenuated proliferation. In vivo, Nr4a1-/- mice exhibited increased colonic smooth muscle thickness following repeated cycles of DSS. Activating NR4A1 with Csn-B, in the context of established inflammation, reduced ileal smooth muscle thickening in SAMP1/YitFc mice. Targeting NR4A1 may provide a novel approach to regulate intestinal SMC phenotype, limiting excessive proliferation that contributes to stricture development in CD.

Interface extension is a continuum property suggesting a linkage between AP contractile and DV lengthening processes.

In the early Drosophila embryo, the elongation of the anterior-posterior (AP) body axis is driven by cell intercalation in the germband epithelium. Neighboring cells intercalate through the contraction of AP interfaces (between AP neighbors) into higher-order vertices, which then resolve through the extension of new dorsal-ventral (DV) interfaces (between DV neighbors). Although interface contraction has been extensively studied, less is known about how new interfaces are established. Here we show that DV interface elongation behaviors initiate at the same time as AP contractions, and that DV interfaces which are newly created from resolution of higher-order vertices do not appear to possess a unique 'identity;' instead, all horizontal interfaces undergo lengthening, elongating through ratchetlike sliding behaviors analogous to those found in AP interfaces. Cortical F-actin networks are essential for high area oscillation amplitudes required for effective ratcheting. Our results suggest that, contrary to canonical models, the elongation of new DV interfaces is not produced by a mechanistically separate process. Instead, medial myosin populations drive oscillating radial forces in the cells to generate transient force asymmetries at all tricellular vertices, which-combined with planar polarized stabilization-produce directional ratcheted sliding to generate both AP interface contraction and DV interface elongation.

High-intensity interval training remodels the proteome and acetylome of human skeletal muscle.

Exercise is an effective strategy in the prevention and treatment of metabolic diseases. Alterations in the skeletal muscle proteome, including post-translational modifications, regulate its metabolic adaptations to exercise. Here, we examined the effect of high-intensity interval training (HIIT) on the proteome and acetylome of human skeletal muscle, revealing the response of 3168 proteins and 1263 lysine acetyl-sites on 464 acetylated proteins. We identified global protein adaptations to exercise training involved in metabolism, excitation-contraction coupling, and myofibrillar calcium sensitivity. Furthermore, HIIT increased the acetylation of mitochondrial proteins, particularly those of complex V. We also highlight the regulation of exercise-responsive histone acetyl-sites. These data demonstrate the plasticity of the skeletal muscle proteome and acetylome, providing insight into the regulation of contractile, metabolic and transcriptional processes within skeletal muscle. Herein, we provide a substantial hypothesis-generating resource to stimulate further mechanistic research investigating how exercise improves metabolic health.

High Fat High Sucrose Diet Modifies Uterine Contractility and Cervical Resistance in Pregnant Rats: The Roles of Sex Hormones, Adipokines and Cytokines.

Background: In obesity, the adipose tissue becomes a very significant endocrine organ producing different factors called adipokines, such as leptin, adiponectin and kisspeptin; however, no data are available about their actions on uterine contraction in obese pregnant rats. Our aim was to study the impact of obesity on pregnant uterine contraction in a rat model. Methods: Obesity was induced by the consumption of a high fat high sucrose diet (HFHSD) for 9 weeks, including pregnancy. Glucose tolerance, sex hormone, cytokine and adipokine levels were measured. Uterine contractions and cervical resistance, as well as their responses to adipokines, were tested along with the expressions of their uterine receptors. Results: HFHSD increased body weight, and altered glucose tolerance and fat composition. The uterine leptin and kisspeptin pathway affect increased. The levels of proinflammatory cytokines were reduced, while the plasma level of progesterone was increased, resulting in weaker uterine contractions, and improving the uterine relaxing effects of adipokines. HFHSD reduced cervical resistance, but the core effect of adipokines is difficult to determine. Conclusions: Obesity in pregnant rats reduces uterine contractility and cytokine-induced inflammatory processes, and therefore obese pregnant rat methods are partially applicable for modelling human processes.

UV-B Filter Octylmethoxycinnamate Is a Modulator of the Serotonin and Histamine Receptors in Human Umbilical Arteries.

Every day, people use personal care products containing UV filters. Although their use initially showed a protective role, toxicity is a concern for human health as several UV filters are endocrine-disrupting chemicals (EDCs). Exposure to EDCs may induce cardiovascular diseases and can affect the health of sensitive people, such as pregnant women. Currently, the world’s most widely used UV-B filter is octylmethoxycinnamate (OMC), an EDC. However, the disruptive effects on pregnant women are little known. The present work proposed to understand how long-term exposure to OMC affects vascular homeostasis. Endothelium-denuded human umbilical artery (HUA) rings were incubated in an organ bath system. Long-term effects of exposure to OMC (0.001–50 μmol/L) were evaluated on the contractile responses of HUA to the application of the contractile agents, serotonin (5-HT) and histamine (Hist). To investigate in more detail the vascular mode of action of OMC, through which it impairs the vascular homeostasis of HUA, the activity and expression of different 5-HT and Hist-receptors involved in contractility processes were studied. Our findings pointed out an increase in the reactivity of HUA to 5-HT and Hist due to OMC exposure. These alterations in reactivity may be precursors of preeclampsia development and/or gestational hypertension.

FXR1 Decreases Blood Pressure by Regulating Vascular Contractility

IntroductionHypertension is a major risk factor for cardiovascular disease and stroke. Optimally treated hypertensive patients still have a 50% greater cardiovascular risk than untreated normotensive subjects. Despite current medication, half a million deaths in the United States include hypertension as a primary contributing cause in 2018 presenting a need for additional targets. Vascular smooth muscle cells (VSMCs) are the fulcrum of vascular disease, particularly hypertension. VSMCs play a critical role in vascular contractility and the regulation of blood pressure. Fragile X‐related protein (FXR1) is a muscle‐enhanced RNA binding protein and we previously found siRNA knock down of FXR1 increases inflammatory mRNA stability. Overexpression of FXR1 decreases inflammatory mRNA stability in VSMC. Little is known concerning FXR1 protein binding partners and its role in vascular disease. The specific aim of this study is to test the hypothesis that FXR1 regulates vascular contractility by RNA stability and protein interactions.ResultsWe generated a novel, VSMC‐specific FXR1 conditional knock out mouse (FXR1VSMC/VSMC) in order to establish an in vivo role of FXR1 in vascular disease. Preliminary data indicates that these mice show decreased diastolic (P < 0.05) blood pressure at baseline compared to controls. Gene ontology of RNA immuno‐precipitation sequencing analysis in human VSMCs identified that FXR1 binds to mRNA that participate in VSMC contractility and regulation of blood pressure‐ related pathways. Although considered an RNA stability protein, mass‐spectrometry identified that FXR1 interacts with proteins related to contractile processes such as cell migration, adhesion and stress fiber formation. FXR1 depletion in human VSMCs decreases mRNA abundance of contractile machinery. Future studies will determine the effect of FXR1 depletion on mRNA stability of these transcripts. Additionally, siRNA knock down of FXR1 decreased VSMC adhesion, migration and collagen gel contraction corroborating in vivo observations.ConclusionThese data are the first to suggest FXR1 regulates blood pressure and vascular contractility potentially by two mechanisms: mRNA stability and functional activity by protein‐protein interactions. The findings support FXR1 activity may represent a target for therapeutic invention to regulate blood pressure.

Fat mass and obesity-associated gene (FTO) hypermethylation induced by decabromodiphenyl ethane causing cardiac dysfunction via glucolipid metabolism disorder

Decabromodiphenyl ethane (DBDPE) is a major alternative to BDE-209 owing to its lower toxicity. However, the mass production and increased consumption of DBDPE in recent years have raised concerns related to its adverse health effects. However, the effect and mechanism of DBDPE on cardiotoxicity have rarely been studied. In the present study, we investigated the impacts of DBDPE on the cardiovascular system in male SD rats and then explored the underlying mechanisms to explain the cardiotoxicity of DBDPE using AC16 cells. Under in vivo conditions, male rats were administered with an oral dosage of DBDPE at 0, 5, 50, and 500 mg/kg/day for 28 days, respectively. Histopathological analysis demonstrated that DBDPE induced cardiomyocyte injury and fibrosis, and ultrastructural observation revealed that DBDPE could induce mitochondria damage and dissolution. DBDPE could thus decrease the level of MYH6 and increase the level of SERCA2, which are the two key proteins involved in the maintenance of homeostasis during myocardial contractile and diastolic processes. Furthermore, DBDPE could increase the serum levels of glucose and low-density lipoprotein but decrease the content of high-density lipoprotein. In addition, DBDPE could activate the PI3K/AKT/GLUT2 and PPARγ/RXRα signaling pathways in AC16 cells. In addition, DBDPE decreased the UCP2 level and ATP synthesis in mitochondria both under in vitro and in vivo conditions, consequently leading to apoptosis via the Cytochrome C/Caspase-9/Caspase-3 pathway. Bisulfite sequencing PCR (BSP) identified the hypermethylation status of fat mass and obesity-associated gene (FTO). 5-aza exerted the opposite effects on the PI3K/AKT/GLUT2, PPARγ/RXRα, and Cytochrome C/Caspase-9/Caspase-3 signaling pathways induced by DBDPE in AC16 cells. In addition, the DBDPE-treated altered levels of UCP2, ATP, and apoptosis were also found to be significantly reversed by 5-aza in AC16 cells. These results suggested that FTO hypermethylation played a regulative role in the pathological process of DBDPE-induced glycolipid metabolism disorder, thereby contributing to the dysfunction of myocardial contraction and relaxation through cardiomyocytes fibrosis and apoptosis via the mitochondrial-mediated apoptotic pathway resulting from mitochondrial dysfunction.

The nuclear receptor ERR cooperates with the cardiogenic factor GATA4 to orchestrate cardiomyocyte maturation

Estrogen-related receptors (ERR) α and γ were shown recently to serve as regulators of cardiac maturation, yet the underlying mechanisms have not been delineated. Herein, we find that ERR signaling is necessary for induction of genes involved in mitochondrial and cardiac-specific contractile processes during human induced pluripotent stem cell-derived cardiomyocyte (hiPSC-CM) differentiation. Genomic interrogation studies demonstrate that ERRγ occupies many cardiomyocyte enhancers/super-enhancers, often co-localizing with the cardiogenic factor GATA4. ERRγ interacts with GATA4 to cooperatively activate transcription of targets involved in cardiomyocyte-specific processes such as contractile function, whereas ERRγ-mediated control of metabolic genes occurs independent of GATA4. Both mechanisms require the transcriptional coregulator PGC-1α. A disease-causing GATA4 mutation is shown to diminish PGC-1α/ERR/GATA4 cooperativity and expression of ERR target genes are downregulated in human heart failure samples suggesting that dysregulation of this circuitry may contribute to congenital and acquired forms of heart failure.

Synaptopodin stress fiber and contractomere at the epithelial junction.

The apical junction of epithelial cells can generate force to control cell geometry and perform contractile processes while maintaining barrier function and adhesion. Yet, the structural basis for force generation at the apical junction is not fully understood. Here, we describe two synaptopodin-dependent actomyosin structures that are spatially, temporally, and structurally distinct. The first structure is formed by the retrograde flow of synaptopodin initiated at the apical junction, creating a sarcomeric stress fiber that lies parallel to the apical junction. Contraction of the apical stress fiber is associated with either clustering of membrane components or shortening of junctional length. Upon junction maturation, apical stress fibers are disassembled. In mature epithelial monolayer, a motorized "contractomere" capable of "walking the junction" is formed at the junctional vertex. Actomyosin activities at the contractomere produce a compressive force evident by actin filament buckling and measurement with a new α-actinin-4 force sensor. The motility of contractomeres can adjust junctional length and change cell packing geometry during cell extrusion and intercellular movement. We propose a model of epithelial homeostasis that utilizes contractomere motility to support junction rearrangement while preserving the permeability barrier.

A single-cell atlas of the myometrium in human parturition.

Parturition is a well-orchestrated process characterized by increased uterine contractility, cervical ripening, and activation of the chorioamniotic membranes; yet, the transition from a quiescent to a contractile myometrium heralds the onset of labor. However, the cellular underpinnings of human parturition in the uterine tissues are still poorly understood. Herein, we performed a comprehensive study of the human myometrium during spontaneous term labor using single-cell RNA sequencing (scRNA-Seq). First, we established a single-cell atlas of the human myometrium and unraveled the cell type–specific transcriptomic activity modulated during labor. Major cell types included distinct subsets of smooth muscle cells, monocytes/macrophages, stromal cells, and endothelial cells, all of which communicated and participated in immune (e.g., inflammation) and nonimmune (e.g., contraction) processes associated with labor. Furthermore, integrating scRNA-Seq and microarray data with deconvolution of bulk gene expression highlighted the contribution of smooth muscle cells to labor-associated contractility and inflammatory processes. Last, myometrium-derived single-cell signatures can be quantified in the maternal whole-blood transcriptome throughout pregnancy and are enriched in women in labor, providing a potential means of noninvasively monitoring pregnancy and its complications. Together, our findings provide insights into the contributions of specific myometrial cell types to the biological processes that take place during term parturition.

Abstract 13129: FXR1 Regulates Blood Pressure by Altering Vascular Contractility

Introduction: Hypertension is a major risk factor for heart disease and stroke. Despite current medication, half a million deaths in the United States include hypertension as a primary contributing cause, presenting a need for additional targets. Vascular smooth muscle cells (VSMCs) play a central role in vascular contractility and the regulation of blood pressure (BP). Fragile X-related protein (FXR1) is a muscle-enhanced RNA binding protein. Knock down of FXR1 increases inflammatory mRNA stability, and overexpression of FXR1 decreases inflammatory mRNA stability in VSMC. FXR1 knock out mice have a muscle-specific phenotype. While little is known concerning FXR1 protein binding partners and its role in vascular disease, its muscle-enhanced expression suggests a specialized role in VSMC. The specific aim of this study is to test the hypothesis that FXR1 regulates vascular contractility by regulation of RNA stability and protein-protein interactions. Results: To characterize an in vivo role of FXR1 in vascular disease, we generated a novel, VSMC-specific FXR1 conditional knock out mouse (FXR1 VSMC/VSMC ). Preliminary data indicates that these mice are hypotensive as they show decreased systolic (P < 0.001) and diastolic (P < 0.01) BP, and increased resting heart rate (P < 0.05) at baseline compared to controls. To identify mRNA binding targets, gene ontology of RNA immuno-precipitation sequencing analysis (RIPseq) in human VSMCs identified that FXR1 preferentially binds to mRNA that participate in VSMC contractility and regulation of BP. Although considered an RNA stability protein, mass-spectrometry identified that FXR1 interacts with proteins related to VSMC contractile processes such as cell migration, adhesion and stress fiber formation. At a functional level, siRNA knock down of FXR1 decreased VSMC migration and collagen gel contraction corroborating in vivo observations. Conclusion: These data show that deletion of FXR1 decrease BP and VSMC contractile processes. They are the first to suggest FXR1 regulates BP and vascular contractility potentially by two mechanisms: mRNA stability and functional activity by protein-protein interactions. The findings support FXR1 activity may represent a target for therapeutic invention to regulate BP.

Intercalate or invaginate: PI(3,4,5)P3 governs a membrane constriction switch in cell shaping.

Although contractile processes, from tissue invagination to cell intercalation, utilize diverse ratcheting mechanisms, little is known about how ratcheting becomes engaged at specific cell surfaces. In this issue of Developmental Cell, Maio etal. demonstrate that PI(3,4,5)P3 is a paramount regulator of the Sbf/RabGEF-Rab35 ratchet mechanism.

Excessive adventitial stress drives inflammation-mediated fibrosis in hypertensive aortic remodelling in mice.

Hypertension induces significant aortic remodelling, often adaptive but sometimes not. To identify immuno-mechanical mechanisms responsible for differential remodelling, we studied thoracic aortas from 129S6/SvEvTac and C57BL/6 J mice before and after continuous 14-day angiotensin II infusion, which elevated blood pressure similarly in both strains. Histological and biomechanical assessments of excised vessels were similar at baseline, suggesting a common homeostatic set-point for mean wall stress. Histology further revealed near mechano-adaptive remodelling of the hypertensive 129S6/SvEvTac aortas, but a grossly maladaptive remodelling of C57BL/6 J aortas. Bulk RNA sequencing suggested that increased smooth muscle contractile processes promoted mechano-adaptation of 129S6/SvEvTac aortas while immune processes prevented adaptation of C57BL/6 J aortas. Functional studies confirmed an increased vasoconstrictive capacity of the former while immunohistochemistry demonstrated marked increases in inflammatory cells in the latter. We then used multiple computational biomechanical models to test the hypothesis that excessive adventitial wall stress correlates with inflammatory cell infiltration. These models consistently predicted that increased vasoconstriction against an increased pressure coupled with modest deposition of new matrix thickens the wall appropriately, restoring wall stress towards homeostatic consistent with adaptive remodelling. By contrast, insufficient vasoconstriction permits high wall stresses and exuberant inflammation-driven matrix deposition, especially in the adventitia, reflecting compromised homeostasis and gross maladaptation.