Project description:The Notch signaling pathway is a highly conserved pathway involved in cell fate determination in embryonic development and also functions in the regulation of physiological processes in several systems. It plays an especially important role in vascular development and physiology by influencing angiogenesis, vessel patterning, arterial/venous specification, and vascular smooth muscle biology. Aberrant or dysregulated Notch signaling is the cause of or a contributing factor to many vascular disorders, including inherited vascular diseases, such as cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy, associated with degeneration of the smooth muscle layer in cerebral arteries. Like most signaling pathways, the Notch signaling axis is influenced by complex interactions with mediators of other signaling pathways. This complexity is also compounded by different members of the Notch family having both overlapping and unique functions. Thus, it is vital to fully understand the roles and interactions of each Notch family member in order to effectively and specifically target their exact contributions to vascular disease. In this chapter, we will review the Notch signaling pathway in vascular smooth muscle cells as it relates to vascular development and human disease.
Project description:The role of the endothelium is not just limited to acting as an inert barrier for facilitating blood transport. Endothelial cells (ECs), through expression of a repertoire of angiocrine molecules, regulate metabolic demands in an organ-specific manner. Insulin flux across the endothelium to muscle cells is a rate-limiting process influencing insulin-mediated lowering of blood glucose. Here, we demonstrate that Notch signaling in ECs regulates insulin transport to muscle. Notch signaling activity was higher in ECs isolated from obese mice compared to non-obese. Sustained Notch signaling in ECs lowered insulin sensitivity and increased blood glucose levels. On the contrary, EC-specific inhibition of Notch signaling increased insulin sensitivity and improved glucose tolerance and glucose uptake in muscle in a high-fat diet-induced insulin resistance model. This was associated with increased transcription of Cav1, Cav2, and Cavin1, higher number of caveolae in ECs, and insulin uptake rates, as well as increased microvessel density. These data imply that Notch signaling in the endothelium actively controls insulin sensitivity and glucose homeostasis and may therefore represent a therapeutic target for diabetes.
Project description:The first and second branchiomeric (branchial arch) muscles are craniofacial muscles that derive from branchial arch mesoderm. In mammals, this set of muscles is indispensable for jaw movement and facial expression. Defects during embryonic development that result in congenital partial absence of these muscles can have significant impact on patients' quality of life. However, the detailed molecular and cellular mechanisms that regulate branchiomeric muscle development remains poorly understood. Herein we investigated the role of retinoic acid (RA) signaling in developing branchiomeric muscles using mice as a model. We administered all-trans RA (25 mg/kg body weight) to Institute of Cancer Research (ICR) pregnant mice by gastric intubation from E8.5 to E10.5. In their embryos at E13.5, we found that muscles derived from the first branchial arch (temporalis, masseter) and second branchial arch (frontalis, orbicularis oculi) were severely affected or undetectable, while other craniofacial muscles were hypoplastic. We detected elevated cell death in the branchial arch mesoderm cells in RA-treated embryos, suggesting that excessive RA signaling reduces the survival of precursor cells of branchiomeric muscles, resulting in the development of hypoplastic craniofacial muscles. In order to uncover the signaling pathway(s) underlying this etiology, we focused on Pitx2, Tbx1, and MyoD1, which are critical for cranial muscle development. Noticeably reduced expression of all these genes was detected in the first and second branchial arch of RA-treated embryos. Moreover, elevated RA signaling resulted in a reduction in Dlx5 and Dlx6 expression in cranial neural crest cells (CNCCs), which disturbed their interactions with branchiomeric mesoderm cells. Altogether, we discovered that embryonic craniofacial muscle defects caused by excessive RA signaling were associated with the downregulation of Pitx2, Tbx1, MyoD1, and Dlx5/6, and reduced survival of cranial myogenic precursor cells.
Project description:In skeletal muscle, the major isoform of β-adrenergic receptor (β-AR) is β2-AR and the minor isoform is β1-AR, which is opposite to the situation in cardiac muscle. Despite extensive studies in cardiac muscle, the physiological roles of the β-AR subtypes in skeletal muscle are not fully understood. Therefore, in this work, we compared the effects of chronic β1- or β2-AR activation with a specific β1-AR agonist, dobutamine (DOB), or a specific β2-AR agonist, clenbuterol (CB), on masseter and cardiac muscles in mice. In cardiac muscle, chronic β1-AR stimulation induced cardiac hypertrophy, fibrosis and myocyte apoptosis, whereas chronic β2-AR stimulation induced cardiac hypertrophy without histological abnormalities. In masseter muscle, however, chronic β1-AR stimulation did not induce muscle hypertrophy, but did induce fibrosis and apoptosis concomitantly with increased levels of p44/42 MAPK (ERK1/2) (Thr-202/Tyr-204), calmodulin kinase II (Thr-286) and mammalian target of rapamycin (mTOR) (Ser-2481) phosphorylation. On the other hand, chronic β2-AR stimulation in masseter muscle induced muscle hypertrophy without histological abnormalities, as in the case of cardiac muscle, concomitantly with phosphorylation of Akt (Ser-473) and mTOR (Ser-2448) and increased expression of microtubule-associated protein light chain 3-II, an autophagosome marker. These results suggest that the β1-AR pathway is deleterious and the β2-AR is protective in masseter muscle. These data should be helpful in developing pharmacological approaches for the treatment of skeletal muscle wasting and weakness.
Project description:Muscle cell fusion is a multistep process involving cell migration, adhesion, membrane remodeling and actin-nucleation pathways to generate multinucleated myotubes. However, molecular brakes restraining cell-cell fusion events have remained elusive. Here we show that transforming growth factor beta (TGFβ) pathway is active in adult muscle cells throughout fusion. We find TGFβ signaling reduces cell fusion, regardless of the cells' ability to move and establish cell-cell contacts. In contrast, inhibition of TGFβ signaling enhances cell fusion and promotes branching between myotubes in mouse and human. Exogenous addition of TGFβ protein in vivo during muscle regeneration results in a loss of muscle function while inhibition of TGFβR2 induces the formation of giant myofibers. Transcriptome analyses and functional assays reveal that TGFβ controls the expression of actin-related genes to reduce cell spreading. TGFβ signaling is therefore requisite to limit mammalian myoblast fusion, determining myonuclei numbers and myofiber size.
Project description:Mesodermal tissue with heart forming potential (cardiogenic mesoderm) is induced during gastrulation. This cardiogenic mesoderm later differentiates into heart muscle tissue (myocardium) and non-muscular heart tissue. Inhibition of Wnt/beta-catenin signaling is known to be required early for induction of cardiogenic mesoderm; however, the identity of the inhibiting Wnt signal itself is still elusive. We have identified Wnt6 in Xenopus as an endogenous Wnt signal, which is expressed in tissues close to and later inside the developing heart. Our loss-of-function experiments show that Wnt6 function is required in the embryo to prevent development of an abnormally large heart muscle. We find, however, that Wnt6 is not required as expected during gastrulation stages, but later during organogenesis stages just before cells of the cardiogenic mesoderm begin to differentiate into heart muscle (myocardium). Our gain-of-function experiments show that Wnt6 and also activated canonical Wnt/beta-catenin signaling are capable of restricting heart muscle development at these relatively late stages of development. This repressive role of Wnt signaling is mediated initially via repression of cardiogenic transcription factors, since reinstatement of GATA function can rescue expression of other cardiogenic transcription factors and downstream cardiomyogenic differentiation genes.
Project description:Cells need to sense their environment to ensure accurate targeting to specific destinations. This occurs in developing muscles, which need to attach to tendon cells before muscle contractions can begin. Elongating myotube tips form filopodia, which are presumed to have sensory roles, and are later suppressed upon building the attachment site. Here, we use live imaging and quantitative image analysis of lateral transverse (LT) myotubes in Drosophila to show that filopodia suppression occurs as a result of integrin signaling. Loss of the integrin subunits αPS2 and βPS (also known as If and Mys, respectively, in flies) increased filopodia number and length at stages when they are normally suppressed. Conversely, inducing integrin signaling, achieved by the expression of constitutively dimerised βPS cytoplasmic domain (diβ), prematurely suppressed filopodia. We discovered that the integrin signal is transmitted through the protein G protein-coupled receptor kinase interacting ArfGAP (Git) and its downstream kinase p21-activated kinase (Pak). Absence of these proteins causes profuse filopodia and prevents the filopodial inhibition mediated by diβ. Thus, integrin signaling terminates the exploratory behavior of myotubes seeking tendons, enabling the actin machinery to focus on forming a strong attachment and assembling the contractile apparatus.
Project description:Muscle remodeling is an important physiological process that promotes adaptive changes in cytoarchitecture and protein composition after exercise, aging, or disease conditions. Numerous transcription factors have been reported to regulate skeletal muscle homeostasis. NF-kappaB is a major pleiotropic transcription factor modulating immune, inflammatory, cell survival, and proliferating responses; however, its role in muscle development, physiology, and disease has just started to be elucidated. The current review article aims to summarize the literature on the role of NF-kappaB signaling in skeletal muscle pathophysiology, investigated over the last years using in vitro and more recently in vivo systems. Understanding the exact role of NF-kappaB in muscle cells will allow better therapeutic manipulations in the setting of human muscle diseases.
Project description:BACKGROUND:Skeletal muscle growth and regeneration depend on the activation of satellite cells, which leads to myocyte proliferation, differentiation and fusion with existing muscle fibers. Skeletal muscle cell proliferation and differentiation are tightly coordinated by a continuum of molecular signaling pathways. The striated muscle activator of Rho signaling (STARS) is an actin binding protein that regulates the transcription of genes involved in muscle cell growth, structure and function via the stimulation of actin polymerization and activation of serum-response factor (SRF) signaling. STARS mediates cell proliferation in smooth and cardiac muscle models; however, whether STARS overexpression enhances cell proliferation and differentiation has not been investigated in skeletal muscle cells. RESULTS:We demonstrate for the first time that STARS overexpression enhances differentiation but not proliferation in C2C12 mouse skeletal muscle cells. Increased differentiation was associated with an increase in the gene levels of the myogenic differentiation markers Ckm, Ckmt2 and Myh4, the differentiation factor Igf2 and the myogenic regulatory factors (MRFs) Myf5 and Myf6. Exposing C2C12 cells to CCG-1423, a pharmacological inhibitor of SRF preventing the nuclear translocation of its co-factor MRTF-A, had no effect on myotube differentiation rate, suggesting that STARS regulates differentiation via a MRTF-A independent mechanism. CONCLUSION:These findings position STARS as an important regulator of skeletal muscle growth and regeneration.
Project description:The platelet-derived growth factor receptors alpha and beta (PDGFRα and PDGFRβ) mark fibroadipogenic progenitor cells/fibroblasts and pericytes in skeletal muscle, respectively. While the role that these cells play in muscle growth and development has been evaluated, it was not known whether the PDGF receptors activate signaling pathways that control transcriptional and functional changes during skeletal muscle hypertrophy. To evaluate this, we inhibited PDGFR signaling in mice subjected to a synergist ablation muscle growth procedure, and performed analyses 3 and 10 days after induction of hypertrophy. The results from this study indicate that PDGF signaling is required for fiber hypertrophy, extracellular matrix production, and angiogenesis that occur during muscle growth.