Transgenic expression of the HERV-W envelope protein leads to polarized glial cell populations and a neurodegenerative environment.

The human endogenous retrovirus type W (HERV-W) has been identified and repeatedly confirmed as human-specific pathogenic entity affecting many cell types in multiple sclerosis (MS). Our recent contributions revealed the encoded envelope (ENV) protein to disturb myelin repair by interfering with oligodendroglial precursor differentiation and by polarizing microglial cells toward an axon-damage phenotype. Indirect proof of ENV's antiregenerative and degenerative activities has been gathered recently in clinical trials using a neutralizing anti-ENV therapeutic antibody. Yet direct proof of its mode of action can only be presented here based on transgenic ENV expression in mice. Upon demyelination, we observed myelin repair deficits, neurotoxic microglia and astroglia, and increased axon degeneration. Experimental autoimmune encephalomyelitis activity progressed faster in mutant mice equally accompanied by activated glial cells. This study therefore provides direct evidence on HERV-W ENV's contribution to the overall negative impact of this activated viral entity in MS.

Human Endogenous Retrovirus K Envelope in Spinal Fluid of Amyotrophic Lateral Sclerosis Is Toxic.

OBJECTIVE: Human endogenous retroviruses have been implicated in neurodegenerative diseases including amyotrophic lateral sclerosis (ALS). Expression of human endogenous retrovirus K (HERV-K) subtype HML-2 envelope (Env) in human neuronal cultures and in transgenic mice results in neurotoxicity and neurodegeneration, and mice expressing HML-2 Env display behavioral and neuromuscular characteristics resembling ALS. This study aims to characterize the neurotoxic properties of HML-2 Env. METHODS: Env neurotoxicity was detected in ALS cerebrospinal fluid and confirmed using recombinant Env protein in a cell-based assay and a mouse model. The mechanism of neurotoxicity was assessed with immunoprecipitation followed by mass spectrometry and Western blot, and by screening a panel of inhibitors. RESULTS: We observed that recombinant HML-2 Env protein caused neurotoxicity resulting in neuronal cell death, retraction of neurites, and decreased neuronal electrical activity. Injection of the Env protein into the brains of mice also resulted in neuronal cell death. HML-2 Env protein was also found in the cerebrospinal fluid of patients with sporadic ALS. The neurotoxic properties of the Env and the cerebrospinal fluid could be rescued with the anti-Env antibody. The Env was found to bind to CD98HC complexed to ß1 integrin on the neuronal cell surface. Using a panel of compounds to screen for their ability to block Env-induced neurotoxicity, we found that several compounds were protective and are linked to the ß1 integrin pathway. INTERPRETATION: HERV-K Env is released extracellularly in ALS and causes neurotoxicity via a novel mechanism. Present results pave the way for new treatment strategies in sporadic ALS. ANN NEUROL 2022;92:545-561.

Type I Interferon Receptor Signaling Drives Selective Permissiveness of Astrocytes and Microglia to Measles Virus during Brain Infection.

Fatal neurological syndromes can occur after measles virus (MeV) infection of the brain. The mechanisms controlling MeV spread within the central nervous system (CNS) remain poorly understood. We analyzed the role of type I interferon (IFN-I) receptor (IFNAR) signaling in the control of MeV infection in a murine model of brain infection. Using organotypic brain cultures (OBC) from wild-type and IFNAR-knockout (IFNARKO) transgenic mice ubiquitously expressing the human SLAM (CD150) receptor, the heterogeneity of the permissiveness of different CNS cell types to MeV infection was characterized. In the absence of IFNAR signaling, MeV propagated significantly better in explant slices. In OBC from IFNAR-competent mice, while astrocytes and microglia were infected on the day of explant preparation, they became refractory to infection with time, in contrast to neurons and oligodendrocytes, which remained permissive to infection. This selective loss of permissiveness to MeV infection was not observed in IFNARKO mouse OBC. Accordingly, the development of astrogliosis related to the OBC procedure was exacerbated in the presence of IFNAR signaling. In the hippocampus, this astrogliosis was characterized by a change in the astrocyte phenotype and by an increase of IFN-I transcripts. A proteome analysis showed the upregulation of 84 out of 111 secreted proteins. In the absence of IFNAR, only 27 secreted proteins were upregulated, and none of these were associated with antiviral activities. Our results highlight the essential role of the IFN-I response in astrogliosis and in the permissiveness of astrocytes and microglia that could control MeV propagation throughout the CNS.IMPORTANCE Measles virus (MeV) can infect the central nervous system (CNS), with dramatic consequences. The mechanisms controlling MeV invasion of the CNS remain ill-defined since most previous data were obtained from postmortem analysis. Here, we highlight for the first time the crucial role of the type I interferon (IFN-I) response not only in the control of CNS invasion but also in the early permissiveness of glial cells to measles virus infection.

TGFß and FGF promote tendon progenitor fate and act downstream of muscle contraction to regulate tendon differentiation during chick limb development.

The molecular programme underlying tendon development has not been fully identified. Interactions with components of the musculoskeletal system are important for limb tendon formation. Limb tendons initiate their development independently of muscles; however, muscles are required for further tendon differentiation. We show that both FGF/ERK MAPK and TGFß/SMAD2/3 signalling pathways are required and sufficient for SCX expression in chick undifferentiated limb cells, whereas the FGF/ERK MAPK pathway inhibits Scx expression in mouse undifferentiated limb mesodermal cells. During differentiation, muscle contraction is required to maintain SCX, TNMD and THBS2 expression in chick limbs. The activities of FGF/ERK MAPK and TGFß/SMAD2/3 signalling pathways are decreased in tendons under immobilisation conditions. Application of FGF4 or TGFß2 ligands prevents SCX downregulation in immobilised limbs. TGFß2 but not FGF4 prevent TNMD and THBS2 downregulation under immobilisation conditions. We did not identify any intracellular crosstalk between both signalling pathways in their positive effect on SCX expression. Independently of each other, both FGF and TGFß promote tendon commitment of limb mesodermal cells and act downstream of mechanical forces to regulate tendon differentiation during chick limb development.

Knockdown of col22a1 gene in zebrafish induces a muscular dystrophy by disruption of the myotendinous junction.

The myotendinous junction (MTJ) is the major site of force transfer in skeletal muscle, and defects in its structure correlate with a subset of muscular dystrophies. Col22a1 encodes the MTJ component collagen XXII, the function of which remains unknown. Here, we have cloned and characterized the zebrafish col22a1 gene and conducted morpholino-based loss-of-function studies in developing embryos. We showed that col22a1 transcripts localize at muscle ends when the MTJ forms and that COLXXII protein integrates the junctional extracellular matrix. Knockdown of COLXXII expression resulted in muscular dystrophy-like phenotype, including swimming impairment, curvature of embryo trunk/tail, strong reduction of twitch-contraction amplitude and contraction-induced muscle fiber detachment, and provoked significant activation of the survival factor Akt. Electron microscopy and immunofluorescence studies revealed that absence of COLXXII caused a strong reduction of MTJ folds and defects in myoseptal structure. These defects resulted in reduced contractile force and susceptibility of junctional extracellular matrix to rupture when subjected to repeated mechanical stress. Co-injection of sub-phenotypic doses of morpholinos against col22a1 and genes of the major muscle linkage systems showed a synergistic gene interaction between col22a1 and itga7 (α7ß1 integrin) that was not observed with dag1 (dystroglycan). Finally, pertinent to a conserved role in humans, the dystrophic phenotype was rescued by microinjection of recombinant human COLXXII. Our findings indicate that COLXXII contributes to the stabilization of myotendinous junctions and strengthens skeletal muscle attachments during contractile activity.

Stable and bicistronic expression of two genes in somite- and lateral plate-derived tissues to study chick limb development.

BACKGROUND: Components of the limb musculoskeletal system have distinct mesoderm origins. Limb skeletal muscles originate from somites, while the skeleton and attachments (tendons and connective tissues) derive from limb lateral plate. Despite distinct mesoderm origins, the development of muscle, skeleton and attachments is highly coordinated both spatially and temporally to ensure complete function of the musculoskeletal system. A system to study molecular interactions between somitic-derived tissues (muscles) and lateral-plate-derived tissues (skeletal components and attachments) during limb development is missing. RESULTS: We designed a gene delivery system in chick embryos with the ultimate aim to study the interactions between the components of the musculoskeletal system during limb development. We combined the Tol2 genomic integration system with the viral T2A system and developed new vectors that lead to stable and bicistronic expression of two proteins at comparable levels in chick cells. Combined with limb somite and lateral plate electroporation techniques, two fluorescent reporter proteins were co-expressed in stoichiometric proportion in the muscle lineage (somitic-derived) or in skeleton and their attachments (lateral-plate-derived). In addition, we designed three vectors with different promoters to target muscle cells at different steps of the differentiation process. CONCLUSION: Limb somite electroporation technique using vectors containing these different promoters allowed us to target all myogenic cells, myoblasts or differentiated muscle cells. These stable and promoter-specific vectors lead to bicistronic expression either in somitic-derived myogenic cells or lateral plate-derived cells, depending on the electroporation sites and open new avenues to study the interactions between myogenic cells and tendon or connective tissue cells during limb development.

HERV-W ENV transcription in B cells predicting symptomatic COVID-19 and risk for long COVID can express a full-length protein despite stop codon in mRNA from chromosome X via a ribosome readthrough.

The human genome comprises 8% of endogenous retroviruses (HERVs). Though HERVS contribute to physiological functions, copies retained pathogenic potential. The HERV-W ENV protein was shown expressed in patients with worse COVID-19 symptoms and post-COVID syndrome. A significant detection of the mRNA encoding HERV-W ENV from patients with COVID-19 in B cells from RNAseq reads obtained from peripheral blond mononuclear cells. This data stratified with increased COVID-19 symptoms or with post-acute sequelae of COVID-19 (long COVID) after 3 months. The HERV-W ENV-U3R RNA was confirmed to display the best alignment with chromosome X ERVWE2 locus. However, a stop codon precluding its translation was re-addressed after recent understandings of ribosome readthrough mechanisms. Experimental results evidenced that this HERV gene can effectively express a full-length protein in the presence of molecules allowing translation via a readthrough mechanism at the ribosome level. Results not only confirm HERV-W ENV RNA origin in these patients but show for the first time how a defective HERV copy can be translated into a complete protein when specific factors make it possible at the ribosome level. The present proof of concept now requires further studies to identify the factors involved in this newly understood mechanism, following SARS-CoV-2 exposure.

Patients with psychosis spectrum disorders hospitalized during the COVID-19 pandemic unravel overlooked SARS-CoV-2 past infection clustering with HERV-W ENV expression and chronic inflammation.

Epidemiology has repeatedly associated certain infections with a risk of further developing psychiatric diseases. Such infections can activate retro-transposable genetic elements (HERV) known to trigger immune receptors and impair synaptic plasticity of neuroreceptors. Since the HERV-W ENV protein was recently shown to co-cluster with pro-inflammatory cytokines in a subgroup of patients with schizophrenia or bipolar disorder, we questioned the influence of the COVID-19 pandemic on patients with psychosis spectrum disorders (PSD). Present results revealed that (i) SARS-CoV-2 serology shows high prevalence and titers of antibodies in PSD, (ii) HERV-W ENV is detected in seropositive individuals only and (iii) SARS-CoV-2 and HERV-W ENV positivity co-clustered with high serum levels of pro-inflammatory cytokines in psychotic patients. These results thus suggest that SARS-CoV-2 infection in many patients with psychotic disorders now admitted in the psychiatry department did not cause severe COVID-19. They also confirm the previously reported association of elevated serum pro-inflammatory cytokines and HERV-W ENV in a subgroup of psychotic patients. In the context of the COVID-19 pandemic, this cluster is only found in SARS-CoV-2 seropositive PSD cases, suggesting a dominant influence of this virus on HERV-W ENV and cytokine expression, and/or patients' greater susceptibility to SARS-CoV-2 infection. Further investigation on an interplay between this viral infection and the clinical evolution of such PSD patients is needed. However, this repeatedly defined subgroup of psychotic patients with a pro-inflammatory phenotype and HERV expression calls for a differential therapeutic approach in psychoses, therefore for further precision medicine development.

SARS-CoV-2 awakens ancient retroviral genes and the expression of proinflammatory HERV-W envelope protein in COVID-19 patients.

Patients with COVID-19 may develop abnormal inflammatory response, followed in some cases by severe disease and long-lasting syndromes. We show here that in vitro exposure to SARS-CoV-2 activates the expression of the human endogenous retrovirus (HERV) HERV-W proinflammatory envelope protein (ENV) in peripheral blood mononuclear cells from a subset of healthy donors, in ACE2 receptor and infection-independent manner. Plasma and/or sera of 221 COVID-19 patients from different cohorts, infected with successive SARS-CoV-2 variants including the Omicron, had detectable HERV-W ENV, which correlated with ENV expression in T lymphocytes and peaked with the disease severity. HERV-W ENV was also found in postmortem tissues of lungs, heart, gastrointestinal tract, brain olfactory bulb, and nasal mucosa from COVID-19 patients. Altogether, these results demonstrate that SARS-CoV-2 could induce HERV-W envelope protein expression and suggest its involvement in the immunopathogenesis of certain COVID-19-associated syndromes and thereby its relevance in the development of personalized treatment of patients.

HERV-W ENV antigenemia and correlation of increased anti-SARS-CoV-2 immunoglobulin levels with post-COVID-19 symptoms.

Due to the wide scope and persistence of COVID-19´s pandemic, post-COVID-19 condition represents a post-viral syndrome of unprecedented dimensions. SARS-CoV-2, in line with other infectious agents, has the capacity to activate dormant human endogenous retroviral sequences ancestrally integrated in human genomes (HERVs). This activation was shown to relate to aggravated COVID-19 patient´s symptom severity. Despite our limited understanding of how HERVs are turned off upon infection clearance, or how HERVs mediate long-term effects when their transcription remains aberrantly on, the participation of these elements in neurologic disease, such as multiple sclerosis, is already settling the basis for effective therapeutic solutions. These observations support an urgent need to identify the mechanisms that lead to HERV expression with SARS-CoV-2 infection, on the one hand, and to answer whether persistent HERV expression exists in post-COVID-19 condition, on the other. The present study shows, for the first time, that the HERV-W ENV protein can still be actively expressed long after SARS-CoV-2 infection is resolved in post-COVID-19 condition patients. Moreover, increased anti-SARS-CoV-2 immunoglobulins in post-COVID-19 condition, particularly high anti-SARS-CoV-2 immunoglobulin levels of the E isotype (IgE), seem to strongly correlate with deteriorated patient physical function (r=-0.8057, p<0.01). These results indicate that HERV-W ENV antigenemia and anti-SARS-CoV-2 IgE serology should be further studied to better characterize post-COVID-19 condition pathogenic drivers potentially differing in subsets of patients with various symptoms. They also point out that such biomarkers may serve to design therapeutic options for precision medicine in post-COVID-19 condition.

Development of the zebrafish myoseptum with emphasis on the myotendinous junction.

Zebrafish myosepta connect two adjacent muscle cells and transmit muscular forces to axial structures during swimming via the myotendinous junction (MTJ). The MTJ establishes transmembrane linkages system consisting of extracellular matrix molecules (ECM) surrounding the basement membrane, cytoskeletal elements anchored to sarcolema, and all intermediate proteins that link ECM to actin filaments. Using a series of zebrafish specimens aged between 24 h post-fertilization and 2 years old, the present paper describes at the transmission electron microscope level the development of extracellular and intracellular elements of the MTJ. The transverse myoseptum development starts during the segmentation period by deposition of sparse and loosely organized collagen fibrils. During the hatching period, a link between actin filaments and sarcolemma is established. The basal lamina underlining sarcolemma is well differentiated. Later, collagen fibrils display an orthogonal orientation and fibroblast-like cells invade the myoseptal stroma. A dense network of collagen fibrils is progressively formed that both anchor myoseptal fibroblasts and sarcolemmal basement membrane. The differentiation of a functional MTJ is achieved when sarcolemma interacts with both cytoskeletal filaments and extracellular components. This solid structural link between contractile apparatus and ECM leads to sarcolemma deformations resulting in the formation of regular invaginations, and allows force transmission during muscle contraction. This paper presents the first ultrastructural atlas of the zebrafish MTJ development, which represents an useful tool to analyse the mechanisms of the myotendinous system formation and their disruption in muscle disorders.

Human Endogenous Retrovirus Type W Envelope from Multiple Sclerosis Demyelinating Lesions Shows Unique Solubility and Antigenic Characteristics.

In multiple sclerosis (MS), human endogenous retrovirus W family (HERV-W) envelope protein, pHERV-W ENV, limits remyelination and induces microglia-mediated neurodegeneration. To better understand its role, we examined the soluble pHERV-W antigen from MS brain lesions detected by specific antibodies. Physico-chemical and antigenic characteristics confirmed differences between pHERV-W ENV and syncytin-1. pHERV-W ENV monomers and trimers remained associated with membranes, while hexamers self-assembled from monomers into a soluble macrostructure involving sulfatides in MS brain. Extracellular hexamers are stabilized by internal hydrophobic bonds and external hydrophilic moieties. HERV-W studies in MS also suggest that this diffusible antigen may correspond to a previously described high-molecular-weight neurotoxic factor secreted by MS B-cells and thus represents a major agonist in MS pathogenesis. Adapted methods are now needed to identify encoding HERV provirus(es) in affected cells DNA. The properties and origin of MS brain pHERV-W ENV soluble antigen will allow a better understanding of the role of HERVs in MS pathogenesis. The present results anyhow pave the way to an accurate detection of the different forms of pHERV-W ENV antigen with appropriate conditions that remained unseen until now.

Evidence of the pathogenic HERV-W envelope expression in T lymphocytes in association with the respiratory outcome of COVID-19 patients.

BACKGROUND: Despite an impressive effort in clinical research, no standard therapeutic approach for coronavirus disease 2019 (COVID-19) patients has been established, highlighting the need to identify early biomarkers for predicting disease progression and new therapeutic interventions for patient management. The present study aimed to evaluate the involvement of the human endogenous retrovirus -W envelope (HERV-W ENV) in severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection considering recent findings that HERVs are activated in response to infectious agents and lead to various immunopathological effects. We analysed HERV-W ENV expression in blood cells of COVID-19 patients in correlation with clinical characteristics and have discussed its potential role in the outcome of the disease. METHODS: We analysed HERV-W ENV expression in blood samples of COVID-19 patients and healthy donors by flow cytometry and quantitative reverse transcriptase PCR analysis, and evaluated its correlation with clinical signs, inflammatory markers, cytokine expression, and disease progression. FINDINGS: HERV-W ENV was highly expressed in the leukocytes of COVID-19 patients but not in those of healthy donors. Its expression correlated with the markers of T-cell differentiation and exhaustion and blood cytokine levels. The percentage of HERV-W ENV-positive lymphocytes correlated with inflammatory markers and pneumonia severity in COVID-19 patients. Notably, HERV-W ENV expression reflects the respiratory outcome of patients during hospitalization. INTERPRETATION: Given the known immuno- and neuro-pathogenicity of HERV-W ENV protein, it could promote certain pathogenic features of COVID-19 and therefore serve as a biomarker to predict clinical progression of disease and open to further studies for therapeutic intervention. FUNDING: Information available at the end of the manuscript.

Coxsackievirus-B4 Infection Can Induce the Expression of Human Endogenous Retrovirus W in Primary Cells.

Human Endogenous Retrovirus W Envelope (HERV-W ENV) mRNA or protein can be found in peripheral blood mononuclear cells (PBMCs) and exocrine pancreas of patients with type 1 diabetes (T1D). Further, previous observations have shown an association between enteroviral infection and development of T1D; specifically, coxsackievirus-B (CV-B) has been detected in the blood and pancreas of patients with T1D. Notably, viruses can activate HERV-W expression. Hence, we evaluated the effect of CV-B4 infection on HERV-W ENV mRNA expression. Primary human pancreatic ductal cells were obtained from five brain-dead donors. In the pancreatic cells of three donors, the HERV-W ENV mRNA level measured using RT-qPCR was upregulated upon CV-B4 infection. The HERV-W ENV protein was detected in the infected cells using the immunoblot assay. In human PBMCs inoculated with CV-B4 or when CV-B4 was incubated with an enhancing serum, the HERV-W ENV mRNA level was higher than the background RNA level. In monocyte-derived macrophages obtained from 5 of 13 donors, the HERV-W ENV mRNA level was higher in cultures inoculated with CV-B4 than in the control. Therefore, CV-B4 can upregulate or induce the transcription of a certain HERV-W ENV copy (or copies) in primary cell cultures, such as monocytes, macrophages, and pancreatic cells.

Zebrafish collagen XII is present in embryonic connective tissue sheaths (fascia) and basement membranes.

Connective tissues ensure the cohesion of the tissues of the body, but also form specialized structures such as tendon and bone. Collagen XII may enhance the stability of connective tissues by bridging collagen fibrils, but its function is still unclear. Here, we used the zebrafish model to visualize its expression pattern in the whole organism. The zebrafish col12a1 gene is homologous to the small isoform of the tetrapod col12a1 gene. In agreement with the biochemical data reported for the small isoform, the zebrafish collagen XII alpha1 chain was characterized as a collagenase sensitive band migrating at approximately 200 kDa. Using newly generated polyclonal antibodies and anti-sense probes, we performed a comprehensive analysis of its expression in developing zebrafish. Collagen XII exhibited a much broader expression pattern than previously thought: it was ubiquitously expressed in the connective tissue sheaths (fascia) that encase the tissues and organs of the body. For example, it was found in sclera, meninges, epimysia and horizontal and vertical myosepta. Collagen XII was also detected in head mesenchyme, pharyngeal arches and within the spinal cord, where it was first expressed within and then at the lateral borders of the floor plate and at the dorsal midline. Furthermore, double immunofluorescence staining with laminin and immunogold electron microscopy revealed that collagen XII is associated with basement membranes. These data suggest that collagen XII is implicated in tissue cohesion by stabilizing fascia and by linking fascia to basement membranes.

Induction of Proinflammatory Multiple Sclerosis-Associated Retrovirus Envelope Protein by Human Herpesvirus-6A and CD46 Receptor Engagement.

The aberrant expression of human endogenous retrovirus (HERV) elements of the HERV-W family has been associated with different diseases, including multiple sclerosis (MS). In particular, the expression of the envelope protein (Env) from the multiple sclerosis-associated retrovirus (MSRV), a member of HERV-W family and known for its potent proinflammatory activity, is repeatedly detected in the brain lesions and blood of MS patients. Furthermore, human herpesvirus 6 (HHV-6) infection has long been suspected to play a role in the pathogenesis of MS and neuroinflammation. We show here that both HHV-6A and stimulation of its receptor, transmembrane glycoprotein CD46, induce the expression of MSRV-Env. The engagement of extracellular domains SCR3 and SCR4 of CD46-Cyt1 isoform was required for MSRV-env transactivation, limiting thus the MSRV-Env induction to the CD46 ligands binding these domains, including C3b component of complement, specific monoclonal antibodies, and both infectious and UV-inactivated HHV-6A, but neither HHV-6B nor measles virus vaccine strain. Induction of MSRV-Env required CD46 Cyt-1 singling and was abolished by the inhibitors of protein kinase C. Finally, both membrane-expressed and secreted MSRV-Env trigger TLR4 signaling, displaying thus a proinflammatory potential, characteristic for this viral protein. These data expand the specter of HHV-6A effects in the modulation of the immune response and support the hypothesis that cross-talks between exogenous and endogenous viruses may contribute to inflammatory diseases and participate in neuroinflammation. Furthermore, they reveal a new function of CD46, known as an inhibitor of complement activation and receptor for several pathogens, in transactivation of HERV env genes, which may play an important role in the pathogenesis of inflammatory diseases.

[Endogenous retroviral sequences in the human genome can play a physiological or pathological role]. / Des séquences rétrovirales endogènes dans le génome humain peuvent jouer un rôle physiologique ou pathologique.

Human endogenous retroviruses (HERV) represent a large part of our genome and the few elements that have retained a potential of expression still remain "dormant" in physiological conditions. In some instances, they can be awakened by environmental factors activating their expression. The best studied conditions of HERV activation are infections caused by microorganisms such as viruses of the Herpesvirus family. This activation can thus lead to the expression of pathogenic proteins such as envelope proteins belonging to the HERV-W and HERV-K families, respectively involved in Multiple Sclerosis (MS) and amyotrophic lateral sclerosis (ALS). Endogenous retroviral proteins can also acquire a physiological function beneficial for humans. This is the case of Syncytin-1 from the HERV-W family, that is involved in placenta formation.

Transcription factor EGR1 directs tendon differentiation and promotes tendon repair.

Tendon formation and repair rely on specific combinations of transcription factors, growth factors, and mechanical parameters that regulate the production and spatial organization of type I collagen. Here, we investigated the function of the zinc finger transcription factor EGR1 in tendon formation, healing, and repair using rodent animal models and mesenchymal stem cells (MSCs). Adult tendons of Egr1-/- mice displayed a deficiency in the expression of tendon genes, including Scx, Col1a1, and Col1a2, and were mechanically weaker compared with their WT littermates. EGR1 was recruited to the Col1a1 and Col2a1 promoters in postnatal mouse tendons in vivo. Egr1 was required for the normal gene response following tendon injury in a mouse model of Achilles tendon healing. Forced Egr1 expression programmed MSCs toward the tendon lineage and promoted the formation of in vitro-engineered tendons from MSCs. The application of EGR1-producing MSCs increased the formation of tendon-like tissues in a rat model of Achilles tendon injury. We provide evidence that the ability of EGR1 to promote tendon differentiation is partially mediated by TGF-ß2. This study demonstrates EGR1 involvement in adult tendon formation, healing, and repair and identifies Egr1 as a putative target in tendon repair strategies.

The development of the myotendinous junction. A review.

The myotendinous junction (MTJ) is a complex specialized region located at the muscle-tendon interface that represents the primary site of force transmission. Despite their different embryologic origins, muscle and tendon morphogenesis occurs in close spatial and temporal association. After muscle attachment, muscle and tendon constitute a dynamic and functional integrated unit that transduces muscle contraction force to the skeletal system. We review here the current understanding of MTJ formation describing changes during morphogenesis and focusing on the crosstalk between muscle and tendon cells that leads to the development of a functional MTJ. Molecules involved in the formation of the linkage, both at the tendon side and at the muscle side of the junction are described. Much of this knowledge comes from studies using different animal models such as mice, zebrafish and Drosophila where powerful methods for in vivo imaging and genetic manipulations can be used to enlighten this developmental process.