Biology subjects

Mendias, C. L.

Publications and source records attributed to Mendias, C. L..

11 recordsLinked to original sources

bioRxiv2023

Increased Ca2+ signaling through CaV1.2 induces tendon hypertrophy with increased collagen fibrillogenesis and biomechanical properties

Tendons are tension-bearing tissues transmitting force from muscle to bone for body movement. This mechanical loading is essential for tendon development, homeostasis, and healing after injury. While Ca2+ signaling has been studied extensively for its roles in mechanotransduction, regulating muscle, bone and cartilage development and homeostasis, knowledge about Ca2+ signaling and the source of Ca2+ signals in tendon fibroblast biology are largely unknown. Here, we investigated the function of Ca2+ signaling through CaV1.2 voltage-gated Ca2+ channel in tendon formation. Using a reporter mouse, we found that CaV1.2 is highly expressed in tendon during development and downregulated in adult homeostasis. To assess its function, we generated ScxCre;CaV1.2TS mice that express a gain-of-function mutant CaV1.2 channel (CaV1.2TS) in tendon. We found that tendons in the mutant mice were approximately 2/3 larger and had more tendon fibroblasts, but the cell density of the mutant mice decreased by around 22%. TEM analyses demonstrated increased collagen fibrillogenesis in the hypertrophic tendon. Biomechanical testing revealed that the hypertrophic Achilles tendons display higher peak load and stiffness, with no changes in peak stress and elastic modulus. Proteomics analysis reveals no significant difference in the abundance of major extracellular matrix (ECM) type I and III collagens, but mutant mice had about 2-fold increase in other ECM proteins such as tenascin C, tenomodulin, periostin, type XIV and type VIII collagens, around 11-fold increase in the growth factor of TGF-{beta} family myostatin, and significant elevation of matrix remodeling proteins including Mmp14, Mmp2 and cathepsin K. Taken together, these data highlight roles for increased Ca2+ signaling through CaV1.2 on regulating expression of myostatin growth factor and ECM proteins for tendon collagen fibrillogenesis during tendon formation.

molecular biology↗

bioRxiv2020

Scleraxis is required for the growth of adult tendons in response to mechanical loading

Scleraxis is a basic helix-loop-helix transcription factor that plays a central role in promoting tenocyte proliferation and matrix synthesis during embryonic tendon development. However, the role of scleraxis in the growth and adaptation of adult tendons is not known. We hypothesized that scleraxis is required for tendon growth in response to mechanical loading, and that scleraxis promotes the specification of progenitor cells into tenocytes. We conditionally deleted scleraxis in adult mice using a tamoxifen-inducible Cre-recombinase expressed from the Rosa26 locus (Scx{Delta}), and then induced tendon growth in Scx+ and Scx{Delta} adult mice via plantaris tendon mechanical overload. Compared to the wild type Scx+ group, Scx{Delta} mice demonstrated blunted tendon growth. Transcriptional and proteomic analyses revealed significant reductions in cell proliferation, protein synthesis, and extracellular matrix genes and proteins. Our results indicate that scleraxis is required for mechanically-stimulated adult tendon growth by causing the commitment of CD146+ pericytes into the tenogenic lineage, and by promoting the initial expansion of newly committed tenocytes and the production of extracellular matrix proteins.

cell biology↗

bioRxiv2019

Widespread diversity in the transcriptomes of functionally divergent limb tendons

Tendon is a functionally important connective tissue that transmits force between skeletal muscle and bone. Previous studies have evaluated the architectural designs and mechanical properties of different tendons throughout the body. However, less is known about the underlying transcriptional differences between tendons which may dictate their designs and properties. Therefore, our objective was to develop a comprehensive atlas of the transcriptome of limb tendons in adult mice and rats using systems biology techniques. We selected the Achilles, forepaw digit flexor, patellar, and supraspinatus tendons due to their divergent functions and high rates of injury and tendinopathies in patients. Using RNA sequencing data, we generated the Comparative Tendon Transcriptional Database (CTTDb) that identified substantial diversity in the transcriptomes of tendons both within and across species. Approximately 30% of transcripts were differentially regulated between tendons of a given species, and nearly 60% of the transcripts present in anatomically similar tendons were different between species. Many of the genes that differed between tendons and across species are important in tissue specification and limb morphogenesis, tendon cell biology and tenogenesis, growth factor signaling, and production and maintenance of the extracellular matrix. This study indicates that tendon is a surprisingly heterogenous tissue with substantial genetic variation based on anatomical location and species. Key PointsO_LITendon is a hypocellular, matrix-rich tissue that has been excluded from comparative transcriptional atlases. These atlases have provided important knowledge about biological heterogeneity between tissues, and our manuscript addresses this important gap. C_LIO_LIWe performed measures on four of the most studied tendons, the Achilles, forepaw flexor, patellar, and supraspinatus tendons of both mice and rats. These tendons are functionally distinct and are also among the most commonly injured, and therefore of important translational interest. C_LIO_LIApproximately one-third of the transcriptome was differentially regulated between Achilles, forepaw flexor, patellar, and supraspinatus tendons within either mice or rats. Nearly two thirds of the transcripts that are expressed in anatomically similar tendons were different between mice and rats. C_LIO_LIThe overall findings from this study identified that although tendons across the body share a common anatomical definition based on their physical location between skeletal muscle and bone, tendon is a surprisingly genetically heterogeneous tissue. C_LI

systems biology↗

bioRxiv2019

A single-cell transcriptional atlas identifies extensive heterogeneity in the cellular composition of tendons

Tendon is a connective tissue that transmits forces between muscles and bones. Cellular heterogeneity is increasingly recognized as an important factor in the biological basis of tissue homeostasis and disease, but little is known about the diversity of cells that populate tendon. Our objective was to explore the heterogeneity of cells in mouse Achilles tendons using single-cell RNA sequencing. We assembled a transcriptomic atlas and identified 11 distinct cell types in tendons, including 3 previously undescribed populations of fibroblasts. Using trajectory inference analysis, we provide additional support for the notion that pericytes are progenitor cells for the fibroblasts that compose adult tendons. We also modeled cell-interactions and identified ligand-receptor pairs involved in tendon homeostasis. Our findings highlight notable heterogeneity between and within tendon cell populations, which may contribute to our understanding of tendon extracellular matrix assembly and maintenance, and inform the design of therapies to treat tendinopathies.

cell biology↗

bioRxiv2019

Prostaglandin D2 Signaling is Not Involved in Rat Hindlimb Tendon Healing

Injured tendons heal through the formation of a fibrovascular scar that has inferior mechanical properties compared to native tendon tissue. Reducing inflammation that occurs as a result of the injury could limit scar formation and improve functional recovery of tendons. Prostaglandin D2 (PGD2) plays an important role in promoting inflammation in some injury responses and chronic disease processes, and the inhibition of PGD2 has improved healing and reduced disease burden in animal models and early clinical trials. Based on these findings, we sought to determine the role of PGD2 signaling in the healing of injured tendon tissue. We tested the hypothesis that a potent and specific inhibitor of hematopoietic PGD synthase (HPGDS), GSK2894631A, would improve the recovery of tendons of adult male rats following an acute tenotomy and repair. To test this hypothesis, we performed a full-thickness plantaris tendon tenotomy followed by immediate repair and treated rats twice daily with either 0mg/kg, 2mg/kg, or 6mg/kg of GSK2894631A. Tendons were collected either 7 or 21 days after surgical repair, and mechanical properties of tendons were assessed along with RNA sequencing and histology. While there were some differences in gene expression across groups, the targeted inhibition of HPGDS did not impact the functional repair of tendons after injury as HPGDS expression was surprisingly low in injured tendons. These results indicate that PGD2 signaling does not appear to be important in modulating the repair of injured tendon tissue.

physiology↗

bioRxiv2019

The MRL/MpJ Mouse Strain Is Not Protected From Muscle Atrophy And Weakness After Rotator Cuff Tear

Chronic rotator cuff tears are a common source of shoulder pain and disability. Patients with rotator cuff tears often have substantial weakness, fibrosis, and fat accumulation which limit successful surgical repair and postoperative rehabilitation. The Murphy Roths Large (MRL) strain of mice have demonstrated superior healing and protection against pathological changes in several disease and injury conditions. We tested the hypothesis that, compared to the commonly used C57Bl/6 (B6) strain, MRL mice would have less muscle fiber atrophy and fat accumulation, and be protected against the loss in force production that occurs after cuff tear. Adult male mice B6 and MRL mice were subjected to a rotator cuff tear, and changes in muscle fiber contractility and histology were measured. RNA sequencing, and shotgun metabolomics and lipidomics were also performed. Muscles were harvested one month after tear. B6 and MRL mice had a 40% reduction in relative muscle force production after rotator cuff tear. RNA sequencing identified an increase in fibrosis-associated genes and a reduction in mitochondrial metabolism genes. Markers of glycolytic metabolism increased in B6 mice, while MRL mice appeared to increase amino acid metabolism after tear. There was an accumulation of lipid after injury, although there was a divergent response between B6 and MRL mice in the types of lipid species that accrued. There were strain-specific differences between the transcriptome, metabolome, and lipidome of B6 and MRL mice, but these differences did not protect MRL mice from weakness and pathological changes after rotator cuff tear.

physiology↗

bioRxiv2019

Insulin-like growth factor 1 receptor signaling in tenocytes is required for adult tendon growth

Tenocytes serve to synthesize and maintain collagen fibrils and other matrix proteins in tendon. The underlying biological mechanisms of postnatal tendon growth and repair are not well understood. Insulin-like growth factor 1 (IGF1) plays an important role in the growth and remodeling of numerous tissues, but less is known about IGF1 in tendon. We hypothesized that IGF1 signaling is required for proper tendon growth in response to mechanical loading through regulation of collagen synthesis and cell proliferation. We conditionally deleted the IGF1 receptor (IGF1R) in scleraxis (Scx) expressing tenocytes, and compared to control Scx:IGF1R+ mice, Scx:IGF1R{Delta} mice demonstrated reduced tenocyte proliferation and smaller tendons in response to mechanical loading. Additionally, we identified that both the PI3K/Akt and ERK pathways are activated downstream of IGF1 and interact in a coordinated manner to regulate cell proliferation and protein synthesis. These studies indicate that IGF1 signaling is required for proper postnatal tendon growth.

physiology↗

bioRxiv2019

Adaptive and Innate Immune Cell Responses in Tendons and Lymph Nodes After Tendon Injury and Repair

Tendon injuries are a common clinical condition with limited treatment options. The cellular components of the innate system, such as neutrophils and macrophages, have been well studied in tendon injuries. However the adaptive immune system, comprised of specialized lymphocytes, plays an important role in orchestrating the healing of numerous tissues but less is known about these cells in tendon healing. To gain a greater understanding of the biological processes that regulate tendon healing, we sought to determine how the cellular components of the adaptive and innate immune system respond to a tendon injury using two-month old male mice. We determined that the lymphatic vasculature is present in the epitenon and superficial regions of Achilles tendons. We then created an acute Achilles tenotomy followed by repair, and collected tendons and draining lymph nodes one, two, and four weeks after injury. Using flow cytometry and histology, after tendon injury we observed a robust adaptive immune cell response that followed an initial innate immune cell response. There was an accumulation of monocytes, neutrophils, and macrophages one week after injury that declined thereafter. Dendritic cells and CD4+ T cells peaked two weeks after injury, while B cells and CD8+ T cells progressively increased over time. In parallel, immune cells of the draining popliteal lymph node demonstrated a similarly coordinated response to the injury. These results suggest that there is an adaptive immune response to tendon injury and adaptive immune cells may play a role in regulating tendon healing.

physiology↗

bioRxiv2019

Evaluating Muscle Fiber Contractility and the Transcriptome, Proteome, Metabolome, and Lipidome of the mdx/mTR Mouse Model of Duchenne Muscular Dystrophy

Duchenne muscular dystrophy (DMD) is a progressive neuromuscular disease characterized by extensive muscle weakness. Patients with DMD lack a functional dystrophin protein, which transmits force and organizes the cytoskeleton of skeletal muscle. Multiomic studies evaluate combined changes in the transcriptome, proteome, and metabolome, and have been proposed as a way to obtain novel insight about disease processes from preclinical models. We therefore sought to use this approach to study pathological changes in dystrophic muscles. We evaluated hindlimb muscles of male mdx/mTR mice, which lack a functional dystrophin protein and have deficits in satellite cell abundance and proliferative capacity. Wild type (WT) C57BL/6J mice served as controls. Muscle fiber contractility was measured, along with changes in the transcriptome using RNA sequencing, and in the proteome, metabolome, and lipidome using mass spectroscopy. While mdx/mTR mice displayed gross pathological changes and continued cycles of degeneration and regeneration, we found no differences in fiber contractility between strains. However, there were numerous changes in the transcriptome and proteome related to protein balance, contractile elements, extracellular matrix, and metabolism. There was only a 53% agreement in fold change data between the proteome and transcriptome, highlighting the need to study protein abundance along with gene expression measures. Numerous changes in markers of skeletal muscle metabolism were observed, with dystrophic muscles exhibiting elevated glycolytic metabolites. These findings highlight the utility of multiomics in studying muscle disease, and provide additional insight into the pathological changes in dystrophic muscles that might help to guide evidence-based exercise prescription in DMD patients.

physiology↗

bioRxiv2018

Reduced mitochondrial lipid oxidation leads to fat accumulation in myosteatosis

Myosteatosis is the pathological accumulation of lipid that can occur in conjunction with atrophy and fibrosis following skeletal muscle injury. Little is known about the mechanisms by which lipid accumulates in myosteatosis, but many clinical studies have demonstrated the degree of lipid infiltration negatively correlates with muscle function and regeneration. Our objective was to determine the pathological changes that result in lipid accumulation in injured muscle fibers. We used a rat model of rotator cuff injury in this study, as the rotator cuff muscle group is particularly prone to the development of myosteatosis after injury. Muscles were collected from uninjured controls, or 10, 30, or 60 days after injury, and analyzed using a combination of muscle fiber contractility assessments, RNA sequencing, and undirected metabolomics, lipidomics and proteomics, along with bioinformatics techniques, to identify potential pathways and cellular processes that are dysregulated after rotator cuff tear. Bioinformatics analyses indicated that mitochondrial function was likely disrupted after injury. Based on these findings, and given the role that mitochondria play in lipid metabolism, we then performed targeted biochemical and imaging studies and determined that mitochondrial dysfunction and reduced fatty acid oxidation likely leads to the accumulation of lipid in myosteatosis.

physiology↗

bioRxiv2017

Postnatal Tendon Growth and Remodeling Requires Platelet-Derived Growth Factor Receptor Signaling

Platelet-derived growth factor receptor (PDGFR) signaling plays an important role in the fundamental biological activities of many cells that compose musculoskeletal tissues. However, little is known about the role of PDGFR signaling during tendon growth and remodeling in adult animals. Using the hindlimb synergist ablation model of tendon growth, our objectives were to determine the role of PDGFR signaling in the adaptation of tendons subjected to a mechanical growth stimulus, as well as to investigate the biological mechanisms behind this response. We demonstrate that both PDGFRs, PDGFR and PDGFR{beta}, are expressed in tendon fibroblasts, and that the inhibition of PDGFR signaling suppresses the normal growth of tendon tissue in response to mechanical growth cues due to defects in fibroblast proliferation and migration. We also identify that membrane type-1 matrix metalloproteinase (MT1-MMP) as an essential proteinase for the migration of tendon fibroblasts through their extracellular matrix. Furthermore, we report that MT1-MMP translation is regulated by PI3K/Akt signaling, while ERK1/2 controls post-translational trafficking of MT1-MMP to the plasma membrane of tendon fibroblasts. Taken together, these findings demonstrate that PDGFR signaling is necessary for postnatal tendon growth and remodeling, and that MT1-MMP is a critical mediator of tendon fibroblast migration and a potential target for the treatment of tendon injuries and diseases.

physiology↗