miR-486 improves fibrotic activity in myocardial infarction by targeting SRSF3/p21-Mediated cardiac myofibroblast senescence.

The regulation of fibrotic activities is key to improving pathological remodelling post-myocardial infarction (MI). Currently, in the clinic, safe and curative therapies for cardiac fibrosis and improvement of the pathological fibrotic environment, scar formation and pathological remodelling post-MI are lacking. Previous studies have shown that miR-486 is involved in the regulation of fibrosis. However, it is still unclear how miR-486 functions in post-MI regeneration. Here, we first demonstrated that miR-486 targeting SRSF3/p21 mediates the senescence of cardiac myofibroblasts to improve their fibrotic activity, which benefits the regeneration of MI by limiting scar size and post-MI remodelling. miR-486-targeted silencing has high potential as a novel target to improve fibrotic activity, cardiac fibrosis and pathological remodelling.

Cardiac telocytes exist in the adult Xenopus tropicalis heart.

Recent research has revealed that cardiac telocytes (CTs) play an important role in cardiac physiopathology and the regeneration of injured myocardium. Recently, we reported that the adult Xenopus tropicalis heart can regenerate perfectly in a nearly scar-free manner after injury via apical resection. However, whether telocytes exist in the X tropicalis heart and are affected in the regeneration of injured X tropicalis myocardium is still unknown. The present ultrastructural and immunofluorescent double staining results clearly showed that CTs exist in the X tropicalis myocardium. CTs in the X tropicalis myocardium were mainly twined around the surface of cardiomyocyte trabeculae and linked via nanocontacts between the ends of the telopodes, forming a three-dimensional network. CTs might play a role in the regeneration of injured myocardium.

BDNF and NGF signals originating from sensory ganglia promote cranial motor axon growth.

After exiting the hindbrain, branchial motor axons reach their targets in association with sensory ganglia. The trigeminal ganglion has been shown to promote motor axon growth from rhombomeres 2/3 and 4/5, but it is unknown whether this effect is ganglion specific and through which signals it is mediated. Here, we addressed these questions by co-cultures of ventral rhombomere 8 explants with cranial and spinal sensory ganglia in a collagen gel matrix. Our results show that all cranial sensory ganglia and even a trunk dorsal root ganglion can promote motor axon growth and that ganglia isolated from older embryos had a stronger effect on the axonal growth than younger ones. We found that brain-derived neurotrophic factor (BDNF) and nerve growth factor (NGF) are necessary and sufficient for this effect. Altogether, our results demonstrate that the promoting effect of sensory ganglia on cranial motor axon growth is stage dependent, but not ganglion specific and is mediated by BDNF and NGF signals.

Early moderate exercise benefits myocardial infarction healing via improvement of inflammation and ventricular remodelling in rats.

Thus far, the cellular and molecular mechanisms related to early (especially within 24 hours after acute myocardial infarct (MI)) exercise-mediated beneficial effects on MI have not yet been thoroughly established. In the present study, we demonstrated that acute MI rats that underwent early moderate exercise training beginning one day after MI showed no increase in mortality and displayed significant improvements in MI healing and ventricular remodelling, including an improvement in cardiac function, a decrease in infarct size, cardiomyocyte apoptosis, cardiac fibrosis and cardiomyocyte hypertrophy, and an increase in myocardial angiogenesis, left ventricular wall thickness and the number of cardiac telocytes in the border zone. Integrated miRNA-mRNA profiling analysis performed by the ingenuity pathway analysis system revealed that the inhibition of the TGFB1 regulatory network, activation of leucocytes and migration of leucocytes into the infarct zone comprise the molecular mechanism underlying early moderate exercise-mediated improvements in cardiac fibrosis and the pathological inflammatory response. The findings of the present study demonstrate that early moderate exercise training beginning one day after MI is safe and leads to significantly enhanced MI healing and ventricular remodelling. Understanding the mechanism behind the positive effects of this early training protocol will help us to further tailor suitable cardiac rehabilitation programmes for humans.

Development of the shoulder girdle musculature.

The muscles of the shoulder region are important for movements of the upper limbs and for stabilizing the girdle elements by connecting them to the trunk. They have a triple embryonic origin. First, the branchiomeric shoulder girdle muscles (sternocleidomastoideus and trapezius muscles) develop from the occipital lateral plate mesoderm using Tbx1 over the course of this development. The second population of cells constitutes the superficial shoulder girdle muscles (pectoral and latissimus dorsi muscles), which are derived from the wing premuscle mass. This muscle group undergoes a two-step development, referred to as the "in-out" mechanism. Myogenic precursor cells first migrate anterogradely into the wing bud. Subsequently, they migrate in a retrograde manner from the wing premuscle mass to the trunk. SDF-1/CXCR4 signaling is involved in this outward migration. A third group of shoulder muscles are the rhomboidei and serratus anterior muscles, which are referred to as deep shoulder girdle muscles; they are thought to be derived from the myotomes. It is, however, not clear how myotome cells make contact to the scapula to form these two muscles. In this review, we discuss the development of the shoulder girdle muscle in relation to the different muscle groups.

A novel interaction between ATOH8 and PPP3CB.

ATOH8 is a bHLH transcription factor playing roles in a variety of developmental processes such as neurogenesis, differentiation of pancreatic precursor cells, development of kidney and muscle, and differentiation of endothelial cells. PPP3CB belongs to the catalytic subunit of the serine/threonine phosphatase, calcineurin, which can dephosphorylate its substrate proteins to regulate their physiological activities. In our study, we demonstrated that ATOH8 interacts with PPP3CB in vitro with different approaches. We show that the conserved catalytic domain of PPP3CB interacts with both the N-terminus and the bHLH domain of ATOH8. Although the interaction domain of PPP3CB is conserved among all isoforms of calcineurin A, ATOH8 selectively interacts with PPP3CB instead of PPP3CA, probably due to the unique proline-rich region present in the N-terminus of PPP3CB, which controls the specificity of its interaction partners. Furthermore, we show that inhibition of the interaction with calcineurin inhibitor, cyclosporin A (CsA), leads to the retention of ATOH8 to the cytoplasm, suggesting that the interaction renders nuclear localization of ATOH8 which may be critical to control its activity as transcription factor.

CCR9+CD4+ T cells are associated with disease activity in patients with rheumatoid arthritis.

An increase in CD4+ T cells in the synovium is closely linked to the pathogenesis of rheumatoid arthritis (RA). We aimed to identify the possible causes of the elevated CD4+ T cell levels and to explore the factors influencing disease activity in RA. Fifty-five RA patients, including 28 with active RA (ARA), 27 with inactive RA, and 22 healthy controls, were recruited for this study. The proportion of CCR9+CD4+ T cells and the expression of chemokine receptor 9 (CCR9) on CD4+ T cells were analyzed by flow cytometry. Enzyme-linked immunosorbent assay and chemiluminescent immunoassay were used to evaluate interleukin (IL)-17A and IL-6 levels, respectively. The proportion of CCR9+CD4+ T cells and the expression of CCR9 on CD4+ T cells increased significantly in peripheral blood (PB) and synovial fluid (SF) in ARA compared to those in inactive RA. Furthermore, SF contained more CCR9+CD4+ T cells, IL-6, and IL-17A than PB in RA patients. Moreover, CD4+ T cells in the PB of patients with RA, especially ARA, expressed more CCR9 and secreted more IL-6 and IL-17A after activation. Here, we also demonstrated that both the percentage of CCR9+ cells in CD4+ T cells and the expression of CCR9 on circulating CD4+ T cells were positively correlated with erythrocyte sedimentation rate, hypersensitive C-reactive protein, rheumatoid factor, and anti-cyclic citrullinated peptide antibody. CCR9+CD4+ T cells are elevated in PB and SF, and are associated with disease activity in patients with RA.

The dermomyotome ventrolateral lip is essential for the hypaxial myotome formation.

BACKGROUND: The myotome is the primitive skeletal muscle that forms within the embryonic metameric body wall. It can be subdivided into an epaxial and hypaxial domain. It has been shown that the formation of the epaxial myotome requires the dorsomedial lip of the dermomyotome (DML). Although the ventrolateral lip (VLL) of the dermomyotome is believed to be required for the formation of the hypaxial myotome, experimentally evidence for this statement still needs to be provided. Provision of such data would enable the resolution of a debate regarding the formation of the hypaxial dermomyotome. Two mechanisms have been proposed for this tissue. The first proposes that the intermediate dermomyotome undergoes cellular expansion thereby pushing the ventral lateral lip in a lateral direction (translocation). In contrast, the alternative view holds that the ventral lateral lip grows laterally. RESULTS: Using time lapse confocal microscopy, we observed that the GFP-labelled ventrolateral lip (VLL) of the dermomyotome grows rather than translocates in a lateral direction. The necessity of the VLL for lateral extension of the myotome was addressed by ablation studies. We found that the hypaxial myotome did not form after VLL ablation. In contrast, the removal of an intermediate portion of the dermomyotome had very little effect of the hypaxial myotome. These results demonstrate that the VLL is required for the formation of the hypaxial myotome. CONCLUSION: Our study demonstrates that the dermomyotome ventrolateral lip is essential for the hypaxial myotome formation and supports the lip extension model. Therefore, despite being under independent signalling controls, both the dorsomedial and ventrolateral lip fulfil the same function, i.e. they extend into adjacent regions permitting the growth of the myotome.

The occipital lateral plate mesoderm is a novel source for vertebrate neck musculature.

In vertebrates, body musculature originates from somites, whereas head muscles originate from the cranial mesoderm. Neck muscles are located in the transition between these regions. We show that the chick occipital lateral plate mesoderm has myogenic capacity and gives rise to large muscles located in the neck and thorax. We present molecular and genetic evidence to show that these muscles not only have a unique origin, but additionally display a distinct temporal development, forming later than any other muscle group described to date. We further report that these muscles, found in the body of the animal, develop like head musculature rather than deploying the programme used by the trunk muscles. Using mouse genetics we reveal that these muscles are formed in trunk muscle mutants but are absent in head muscle mutants. In concordance with this conclusion, their connective tissue is neural crest in origin. Finally, we provide evidence that the mechanism by which these neck muscles develop is conserved in vertebrates.

Echocardiographic assessment of Xenopus tropicalis heart regeneration.

BACKGROUND: Recently, it was reported that the adult X. tropicalis heart can regenerate in a nearly scar-free manner after injury via apical resection. Thus, a cardiac regeneration model in adult X. tropicalis provides a powerful tool for recapitulating a perfect regeneration phenomenon, elucidating the underlying molecular mechanisms of cardiac regeneration in an adult heart, and developing an interventional strategy for the improvement in the regeneration of an adult heart, which may be more applicable in mammals than in species with a lower degree of evolution. However, a noninvasive and rapid real-time method that can observe and measure the long-term dynamic change in the regenerated heart in living organisms to monitor and assess the regeneration and repair status in this model has not yet been established. RESULTS: In the present study, the methodology of echocardiographic assessment to characterize the morphology, anatomic structure and cardiac function of injured X. tropicalis hearts established by apex resection was established. The findings of this study demonstrated for the first time that small animal echocardiographic analysis can be used to assess the regeneration of X. tropicalis damaged heart in a scar-free perfect regeneration or nonperfect regeneration with adhesion manner via recovery of morphology and cardiac function. CONCLUSIONS: Small animal echocardiography is a reliable, noninvasive and rapid real-time method for observing and assessing the long-term dynamic changes in the regeneration of injured X. tropicalis hearts.

Temporal sequence in the formation of midline dermis and dorsal vertebral elements in avian embryos.

Somites compartmentalize into a dorsal epithelial dermomyotome and a ventral mesenchymal sclerotome. While sclerotomes give rise to vertebrae and intervertebral discs, dermomyotomes contribute to skeletal muscle and epaxial dermis. Bone morphogenetic protein (BMP)-signals from the lateral mesoderm induce the lateral portion of the dermomyotome to form chondrogenic precursor cells, forming the cartilage of the scapula blade. The fact that BMPs are expressed in the roof plate of the neural tube where they induce cartilage formation led to the question why cells migrating from the medial part of the dermomyotome do not undergo chondrogenic differentiation and do not contribute to the dorsal part of the vertebrae. In the present study, we traced dermomyotomal derivatives by using the quail-chick marker technique. Our study reveals a temporal sequence in the formation of the vertebral cartilage and the midline dermis. The dorsal mesenchyme overlying the roof plate of the neural tube is formed prior to the de-epithelialization of the dermomyotome. Dermomyotomal cells start to migrate medially into the sub-ectodermal space to form the midline dermis after chondrogenesis of the dorsal mesenchyme has occurred. This time delay between chondrogenesis of the dorsal vertebra and dermal formation allows an undisturbed development of these two tissue components within a narrow region of the embryo.

Ablation of cardiomyocyte-derived BDNF during development causes myocardial degeneration and heart failure in the adult mouse heart.

Objective: Brain-derived neurotrophic factor (BDNF) and its receptor TrkB-T1 were recently found to be expressed in cardiomyocytes. However, the functional role of cardiomyocyte-derived BDNF in heart pathophysiology is not yet fully known. Recent studies revealed that BDNF-TrkB pathway plays a critical role to maintain integrity of cardiac structure and function, cardiac pathology and regeneration of myocardial infarction (MI). Therefore, the BDNF-TrkB pathway may be a novel target for myocardial pathophysiology in the adult heart. Approach and results: In the present study, we established a cardiomyocyte-derived BDNF conditional knockout mouse in which BDNF expression in developing cardiomyocytes is ablated under the control of the Myosin heavy chain 6 (MYH6) promoter. The results of the present study show that ablation of cardiomyocyte-derived BDNF during development does not impair survival, growth or reproduction; however, in the young adult heart, it causes cardiomyocyte death, degeneration of the myocardium, cardiomyocyte hypertrophy, left atrial appendage thrombosis, decreased cardiac function, increased cardiac inflammation and ROS activity, and metabolic disorders, leading to heart failure (HF) in the adult heart and eventually resulting in a decrease in the one-year survival rate. In addition, ablation of cardiomyocyte-derived BDNF during the developmental stage leads to exacerbation of cardiac dysfunction and poor regeneration after MI in adult hearts. Conclusion: Cardiomyocyte-derived BDNF is irreplaceable for maintaining the integrity of cardiac structure and function in the adult heart and regeneration after MI. Therefore, the BDNF-TrkB pathway will be a novel target for myocardial pathophysiology in the adult heart.

Identification of cGAS as an innate immune sensor of extracellular bacterium Pseudomonas aeruginosa.

Cyclic GMP-AMP synthase (cGAS) is reported essential for detecting intracellular bacteria. However, it remains to be determined whether and how cGAS is involved in extracellular bacterial infection. Here, we report that cGAS is essential for mediating type I interferon (IFN) production in infection by multiple extracellular pathogens, including Pseudomonas aeruginosa, Klebsiella pneumoniae, and Staphylococcus aureus. In addition, the canonical cGAS-stimulator of interferon gene (STING)-IFN axis is required for protecting mice from P. aeruginosa-induced mouse acute pulmonary infection, confirmed in cGAS pathway-specific gene deficiency mouse models. cGAS -/- and STING -/- mice exhibited reduced type I IFNs production, excessive inflammatory response accompanied with decreased resistance to P. aeruginosa challenge. Unfolded protein response was also modulated by cGAS through IRF3 and type I IFNs under P. aeruginosa infection. Collectively, these findings uncover the importance of cGAS in initiating immune responses against extracellular bacterial infection.

Cardiac telocytes inhibit cardiac microvascular endothelial cell apoptosis through exosomal miRNA-21-5p-targeted cdip1 silencing to improve angiogenesis following myocardial infarction.

Promotion of cardiac angiogenesis in ischemic myocardium is a critical strategy for repairing and regenerating the myocardium after myocardial infarction (MI). Currently, effective methods to aid in the survival of endothelial cells, to avoid apoptosis in ischemic myocardium and to achieve long-term cardiac angiogenesis are still being pursued. Here, we investigated whether cardiac telocyte (CT)-endothelial cell communication suppresses apoptosis and promotes the survival of endothelial cells to facilitate cardiac angiogenesis during MI. Methods: CT exosomes were isolated from CT conditioned medium, and their miRNA profile was characterized by small RNA sequencing. A rat model of left anterior descending coronary artery ligation (LAD)-mediated MI was assessed with histology for infarct size and fibrosis, immunostaining for angiogenesis and cell apoptosis and echocardiography to evaluate the therapeutic effects. Cardiac microvascular endothelial cells (CMECs) and the LAD-MI model treated with CT exosomes or CT exosomal miRNA-21-5p in vitro and in vivo were assessed with cellular and molecular techniques to demonstrate the underlying mechanism. Results: CTs exert therapeutic effects on MI via the potent paracrine effects of CT exosomes to facilitate the inhibition of apoptosis and survival of CMECs and promote cardiac angiogenesis. A novel mechanism of CTs is revealed, in which CT-endothelial cell communication suppresses apoptosis and promotes the survival of endothelial cells in the pathophysiological myocardium. CT exosomal miRNA-21-5p targeted and silenced the cell death inducing p53 target 1 (Cdip1) gene and thus down-regulated the activated caspase-3, which then inhibited the apoptosis of recipient endothelial cells under ischemic and hypoxic conditions, facilitating angiogenesis and regeneration following MI. Conclusions: The present study is the first to show that CTs inhibit cardiac microvascular endothelial cell apoptosis through exosomal miRNA-21-5p-targeted Cdip1 silencing to improve angiogenesis in myocardial infarction. It is believed that these novel findings and the discovery of cellular and molecular mechanisms will provide new opportunities to tailor novel cardiac cell therapies and cell-free therapies for the functional and structural regeneration of the injured myocardium.

Commitment of chondrogenic precursors of the avian scapula takes place after epithelial-mesenchymal transition of the dermomyotome.

BACKGROUND: Cells of the epithelially organised dermomyotome are traditionally believed to give rise to skeletal muscle and dermis. We have previously shown that the dermomyotome can undergo epithelial-mesenchymal transition (EMT) and give rise to chondrogenic cells, which go on to form the scapula blade in birds. At present we have little understanding regarding the issue of when the chondrogenic fate of dermomyotomal cells is determined. Using quail-chick grafting experiments, we investigated whether scapula precursor cells are committed to a chondrogenic fate while in an epithelial state or whether commitment is established after EMT. RESULTS: We show that the hypaxial dermomyotome, which normally forms the scapula, does not generate cartilaginous tissue after it is grafted to the epaxial domain. In contrast engraftment of the epaxial dermomyotome to the hypaxial domain gives rise to scapula-like cartilage. However, the hypaxial sub-ectodermal mesenchyme (SEM), which originates from the hypaxial dermomyotome after EMT, generates cartilaginous elements in the epaxial domain, whereas in reciprocal grafting experiments, the epaxial SEM cannot form cartilage in the hypaxial domain. CONCLUSIONS: We suggest that the epithelial cells of the dermomyotome are not committed to the chondrogenic lineage. Commitment to this lineage occurs after it has undergone EMT to form the sub-ectodermal mesenchyme.

Cxcr4 and Sdf-1 are critically involved in the formation of facial and non-somitic neck muscles.

The present study shows that the CXCR4/SDF-1 axis regulates the migration of second branchial arch-derived muscles as well as non-somitic neck muscles. Cxcr4 is expressed by skeletal muscle progenitor cells in the second branchial arch (BA2). Muscles derived from the second branchial arch, but not from the first, fail to form in Cxcr4 mutants at embryonic days E13.5 and E14.5. Cxcr4 is also required for the development of non-somitic neck muscles. In Cxcr4 mutants, non-somitic neck muscle development is severely perturbed. In vivo experiments in chicken by means of loss-of-function approach based on the application of beads loaded with the CXCR4 inhibitor AMD3100 into the cranial paraxial mesoderm resulted in decreased expression of Tbx1 in the BA2. Furthermore, disrupting this chemokine signal at a later stage by implanting these beads into the BA2 caused a reduction in MyoR, Myf5 and MyoD expression. In contrast, gain-of-function experiments based on the implantation of SDF-1 beads into BA2 resulted in an attraction of myogenic progenitor cells, which was reflected in an expansion of the expression domain of these myogenic markers towards the SDF-1 source. Thus, Cxcr4 is required for the formation of the BA2 derived muscles and non-somitic neck muscles.

The hypaxial origin of the epaxially located rhomboid muscles.

In vertebrates, skeletal muscles of the body are made up of epaxial and hypaxial muscles based on their innervation and relative position to the vertebral column. The epaxial muscles are innervated by the dorsal branches of the spinal nerves and comprise the intrinsic (deep) back muscles, while the hypaxial muscles are innervated by the ventral branches of the spinal nerves including the plexus and consist of a heterogeneous group of intercostal, abdominal, and limb as well as girdle muscles. The canonical view holds that the epaxial muscles are derived from the medial halves of the somites, whereas the hypaxial muscles are all derived from the lateral somitic halves. The rhomboid muscles are situated dorsal to the vertebral column and therefore in the domain typically occupied by epaxial muscles. However, they are innervated by a ventral branch of the brachial plexus called the N. dorsalis scapulae. Due to the apparent inappropriate position of the muscle in relation to its innervation we investigated its origin to help clarify this issue. To study the embryonic origin of the rhomboid muscles, we followed derivatives of the medial and lateral somite halves using quail-chick chimeras. Our results showed that the rhomboid muscles are made up of cells derived mainly from the lateral portion of the somite. Therefore the rhomboid muscles which lie within the epaxial domain of the body, originate from the hypaxial domain of the somites. However their connective tissue is derived from both medial and lateral somites.

Effect of an R69C mutation in the myelin protein zero gene on myelination and ion channel subtypes.

BACKGROUND: Most mutations in the myelin protein zero gene (MPZ) typically cause a severe demyelinating/dysmyelinating neuropathy that begins in infancy or an adult-onset axonal neuropathy. Axonal degeneration in the late-onset H10P mutation may be caused by the disruption of axoglial interaction. OBJECTIVE: To evaluate sural nerve biopsy samples from a patient with early-onset Charcot-Marie-Tooth disease type 1B caused by an arg69-to-cys (R69C) mutation. DESIGN AND PARTICIPANTS: Biopsies of sural nerves were performed 20 years apart in a patient with an R69C mutation (early onset). In addition, peripheral nerves were obtained from autopsy material from a patient with a T95M mutation (late onset). These nerves were analyzed using light microscopy of semithin sections, teased nerve fiber immunohistochemical analysis, electron microscopy, and immunologic electron microscopy. MAIN OUTCOME MEASURES: Pathological changes in sural nerve. RESULTS: Both R69C biopsy samples showed prominent demyelination and onion bulb formation, unlike the late-onset T95M mutation, which showed primarily axonal degeneration with no onion bulbs. The sural biopsy sample obtained 20 years earlier from the R69C patient showed minimal difference from the present sample, consistent with the lack of clinical progression during the 2 decades. Teased fiber immunohistochemical analysis of R69C revealed voltage-gated sodium channel subtype 1.8 expressions at the nodes of Ranvier around the areas of segmental demyelination. Internodal length in all R69C nerve fibers was invariably short (>94% of all internodes are <150 mum). CONCLUSIONS: Morphologic abnormalities in this early-onset R69C neuropathy were severe in childhood but progressed very slowly after adolescence. The switch to voltage-gated sodium channel subtype 1.8 expression at the nodes may provide clues into the pathogenesis of this case of early-onset neuropathy, and the short internodes may contribute to the extremely slowed conduction velocities in this case (<10 m/s).

The unique axon trajectory of the accessory nerve is determined by intrinsic properties of the neural tube in the avian embryo.

The accessory nerve is a cranial nerve, composed of only motor axons, which control neck muscles. Its axons ascend many segments along the lateral surface of the cervical spinal cord and hindbrain. At the level of the first somite, they pass ventrally through the somitic mesoderm into the periphery. The factors governing the unique root trajectory are unknown. Ablation experiments at the accessory nerve outlet points have shown that somites do not regulate the trajectory of the accessory nerve fibres. Factors from the neural tube that may control the longitudinal pathfinding of the accessory nerve fibres were tested by heterotopic transplantations of an occipital neural tube to the cervical and thoracic level. These transplantations resulted in a typical accessory nerve trajectory in the cervical and thoracic spinal cord. In contrast, cervical neural tube grafts were unable to give rise to the typical accessory nerve root pattern when transplanted to occipital level. Our results show that the formation of the unique axon root pattern of the accessory nerve is an intrinsic property of the neural tube.