Pharmacological Antagonism of Ccr2+ Cell Recruitment to Facilitate Regenerative Tendon Healing.

Successful tendon healing requires sufficient deposition and remodeling of new extracellular matrix at the site of injury, with this process mediating in part through fibroblast activation via communication with macrophages. Moreover, resolution of healing requires clearance or reversion of activated cells, with chronic interactions with persistent macrophages impairing resolution and facilitating the conversion the conversion to fibrotic healing. As such, modulation of the macrophage environment represents an important translational target to improve the tendon healing process. Circulating monocytes are recruited to sites of tissue injury, including the tendon, via upregulation of cytokines including Ccl2, which facilitates recruitment of Ccr2+ macrophages to the healing tendon. Our prior work has demonstrated that Ccr2-/- can modulate fibroblast activation and myofibroblast differentiation. However, this approach lacked temporal control and resulted in healing impairments. Thus, in the current study we have leveraged a Ccr2 antagonist to blunt macrophage recruitment to the healing tendon in a time-dependent manner. We first tested the effects of Ccr2 antagonism during the acute inflammatory phase and found that this had no effect on the healing process. In contrast, Ccr2 antagonism during the late inflammatory/ early proliferative period resulted in significant improvements in mechanical properties of the healing tendon. Collectively, these data demonstrate the temporally distinct impacts of modulating Ccr2+ cell recruitment and Ccr2 antagonism during tendon healing and highlight the translational potential of transient Ccr2 antagonism to improve the tendon healing process.

Development of a nanoparticle-based tendon-targeting drug delivery system to pharmacologically modulate tendon healing.

Satisfactory healing following acute tendon injury is marred by fibrosis. Despite the high frequency of tendon injuries and poor outcomes, there are no pharmacological therapies in use to enhance the healing process. Moreover, systemic treatments demonstrate poor tendon homing, limiting the beneficial effects of potential tendon therapeutics. To address this unmet need, we leveraged our existing tendon healing spatial transcriptomics dataset and identified an area enriched for expression of Acp5 (TRAP) and subsequently demonstrated robust TRAP activity in the healing tendon. This unexpected finding allowed us to refine and apply our existing TRAP binding peptide (TBP) functionalized nanoparticle (NP) drug delivery system (DDS) to facilitate improved delivery of systemic treatments to the healing tendon. To demonstrate the translational potential of this DDS, we delivered niclosamide (NEN), an S100a4 inhibitor. While systemic delivery of free NEN did not alter healing, TBP-NPNEN enhanced both functional and mechanical recovery, demonstrating the translational potential of this approach to enhance the tendon healing process.

Identification of Periostin as a critical niche for myofibroblast dynamics and fibrosis during tendon healing.

Tendon injuries are a major clinical problem, with poor patient outcomes caused by abundant scar tissue deposition during healing. Myofibroblasts play a critical role in the initial restoration of structural integrity after injury. However, persistent myofibroblast activity drives the transition to fibrotic scar tissue formation. As such, disrupting myofibroblast persistence is a key therapeutic target. While myofibroblasts are typically defined by the presence of αSMA+ stress fibers, αSMA is expressed in other cell types including the vasculature. As such, modulation of myofibroblast dynamics via disruption of αSMA expression is not a translationally tenable approach. Recent work has demonstrated that Periostin-lineage (PostnLin) cells are a precursor for cardiac fibrosis-associated myofibroblasts. In contrast to this, here we show that PostnLin cells contribute to a transient αSMA+ myofibroblast population that is required for functional tendon healing, and that Periostin forms a supportive matrix niche that facilitates myofibroblast differentiation and persistence. Collectively, these data identify the Periostin matrix niche as a critical regulator of myofibroblast fate and persistence that could be targeted for therapeutic manipulation to facilitate regenerative tendon healing.

Development of a Nanoparticle-Based Tendon-Targeting Drug Delivery System to Pharmacologically Modulate Tendon Healing.

Tendon regeneration following acute injury is marred by a fibrotic healing response that prevents complete functional recovery. Despite the high frequency of tendon injuries and the poor outcomes, including functional deficits and elevated risk of re-injury, there are currently no pharmacological therapies in clinical use to enhance the healing process. Several promising pharmacotherapies have been identified; however, systemic treatments lack tendon specificity, resulting in poor tendon biodistribution and perhaps explaining the largely limited beneficial effects of these treatments on the tendon healing process. To address this major unmet need, we leveraged our existing spatial transcriptomics dataset of the tendon healing process to identify an area of the healing tendon that is enriched for expression of Acp5. Acp5 encodes tartrate-resistant acid phosphatase (TRAP), and we demonstrate robust TRAP activity in the healing tendon. This unexpected finding allowed us to refine and apply our existing TRAP binding peptide (TBP) functionalized nanoparticle (NP) drug delivery system (DDS) to facilitate improved delivery of systemic treatments to the healing tendon. To demonstrate the translational potential of this drug delivery system, we delivered the S100a4 inhibitor, Niclosamide to the healing tendon. We have previously shown that genetic knockdown of S100a4 enhances tendon healing. While systemic delivery of Niclosamide did not affect the healing process, relative to controls, TBP-NP delivery of Niclosamide enhanced both functional and mechanical outcome measures. Collectively, these data identify a novel tendon-targeting drug delivery system and demonstrate the translational potential of this approach to enhance the tendon healing process.

Identification of Periostin as a critical niche for myofibroblast dynamics and fibrosis during tendon healing.

Tendon injuries are a major clinical problem, with poor patient outcomes caused by abundant scar tissue deposition during healing. Myofibroblasts play a critical role in the initial restoration of structural integrity after injury. However, persistent myofibroblast activity drives the transition to fibrotic scar tissue formation. As such, disrupting myofibroblast persistence is a key therapeutic target. While myofibroblasts are typically defined by the presence of αSMA+ stress fibers, αSMA is expressed in other cell types including the vasculature. As such, modulation of myofibroblast dynamics via disruption of αSMA expression is not a translationally tenable approach. Recent work has demonstrated that Periostin-lineage (PostnLin) cells are a precursor for cardiac fibrosis-associated myofibroblasts. In contrast to this, here we show that PostnLin cells contribute to a transient αSMA+ myofibroblast population that is required for functional tendon healing, and that Periostin forms a supportive matrix niche that facilitates myofibroblast differentiation and persistence. Collectively, these data identify the Periostin matrix niche as a critical regulator of myofibroblast fate and persistence that could be targeted for therapeutic manipulation to facilitate regenerative tendon healing.

Epitenon-derived cells comprise a distinct progenitor population that contributes to both tendon fibrosis and regeneration following acute injury.

Flexor tendon injuries are common and heal poorly owing to both the deposition of function- limiting peritendinous scar tissue and insufficient healing of the tendon itself. Therapeutic options are limited due to a lack of understanding of the cell populations that contribute to these processes. Here, we identified a bi-fated progenitor cell population that originates from the epitenon and goes on to contribute to both peritendinous fibrosis and regenerative tendon healing following acute tendon injury. Using a combination of genetic lineage tracing and single cell RNA-sequencing (scRNA-seq), we profiled the behavior and contributions of each cell fate to the healing process in a spatio-temporal manner. Branched pseudotime trajectory analysis identified distinct transcription factors responsible for regulation of each fate. Finally, integrated scRNA-seq analysis of mouse healing with human peritendinous scar tissue revealed remarkable transcriptional similarity between mouse epitenon- derived cells and fibroblasts present in human peritendinous scar tissue, which was further validated by immunofluorescent staining for conserved markers. Combined, these results clearly identify the epitenon as the cellular origin of an important progenitor cell population that could be leveraged to improve tendon healing.

Scleraxis-lineage cells are required for tendon homeostasis and their depletion induces an accelerated extracellular matrix aging phenotype.

Aged tendons have disrupted homeostasis, increased injury risk, and impaired healing capacity. Understanding mechanisms of homeostatic disruption is crucial for developing therapeutics to retain tendon health through the lifespan. Here, we developed a novel model of accelerated tendon extracellular matrix (ECM) aging via depletion of Scleraxis-lineage cells in young mice (Scx-DTR). Scx-DTR recapitulates many aspects of tendon aging including comparable declines in cellularity, alterations in ECM structure, organization, and composition. Single-cell RNA sequencing demonstrated a conserved decline in tenocytes associated with ECM biosynthesis in aged and Scx-DTR tendons, identifying the requirement for Scleraxis-lineage cells during homeostasis. However, the remaining cells in aged and Scx-DTR tendons demonstrate functional divergence. Aged tenocytes become pro-inflammatory and lose proteostasis. In contrast, tenocytes from Scx-DTR tendons demonstrate enhanced remodeling capacity. Collectively, this study defines Scx-DTR as a novel model of accelerated tendon ECM aging and identifies novel biological intervention points to maintain tendon function through the lifespan.

Defining the spatial-molecular map of fibrotic tendon healing and the drivers of Scleraxis-lineage cell fate and function.

Tendon injuries heal via a scar-mediated response, and there are no biological approaches to promote more regenerative healing. Mouse flexor tendons heal through the formation of spatially distinct tissue areas: a highly aligned tissue bridge between the native tendon stubs that is enriched for adult Scleraxis-lineage cells and a disorganized outer shell associated with peri-tendinous scar formation. However, the specific molecular programs that underpin these spatially distinct tissue profiles are poorly defined. In the present study, we combine lineage tracing of adult Scleraxis-lineage cells with spatial transcriptomic profiling to define the overarching molecular programs that govern tendon healing and cell-fate decisions. Pseudotime analysis identified three fibroblast trajectories (synthetic, fibrotic, and reactive) and key transcription factors regulating these fate-switching decisions, including the progression of adult Scleraxis-lineage cells through the reactive trajectory. Collectively, this resource defines the molecular mechanisms that coordinate the temporo-spatial healing phenotype, which can be leveraged to inform therapeutic candidate selection.

CCR2 is expressed by tendon resident macrophage and T cells, while CCR2 deficiency impairs tendon healing via blunted involvement of tendon-resident and circulating monocytes/macrophages.

During tendon healing, macrophages are thought to be a key mediator of scar tissue formation, which prevents successful functional restoration of the tendon. However, macrophages are critical for successful tendon healing as they aid in wound debridement, extracellular matrix deposition, and promote fibroblast proliferation. Recent work has sought to better define the multi-faceted functions of macrophages using depletion studies, while other studies have identified a tendon resident macrophage population. To begin to delineate the functions of tendon-resident versus circulation-derived macrophages, we examined the tendon healing phenotype in Chemokine Receptor 2 (CCR2) reporter (CCR2GFP/+ ), and knockout mice. CCR2 is a chemokine receptor primarily found on the surface of circulating bone marrow-derived monocytes, with CCR2 being an important mediator of macrophage recruitment to wound environments. Surprisingly, CCR2GFP/+ cells were present in the tendon during adult homeostasis, and single-cell RNA sequencing identified these cells as tendon-resident macrophages and T cells. During both homeostasis and healing, CCR2 knockout resulted in a substantial decrease in CCR2GFP+ cells and pan-macrophages. Additionally, loss of CCR2 resulted in reduced numbers of myofibroblasts and impeded functional recovery during late healing. This study highlights the heterogeneity of tendon-resident and recruited immune cells and their contributions following injury, and establishes an important role for CCR2 in modulating both the adult tendon cell environment and tendon healing process.

The Adrenal Cortisol Response to Increasing Ambient Temperature in Polar Bears (Ursus maritimus).

Our objective was to identify the upper ambient temperature threshold that triggers an increase in cortisol in response to increased thermoregulatory demands in polar bears. The results reported here include endocrine data collected over two years from 25 polar bears housed in 11 accredited zoological institutions across North America. The effects of ambient temperature, sex, age group (juvenile, adult, elderly), breeding season and humidity on fecal cortisol metabolite (FCM) concentrations (N = 8439 samples) were evaluated using linear mixed models. Ambient temperatures were placed into five different categories: <5 °C, 6−10 °C, 11−15 °C, 16−20 °C, and >20 °C. Ambient temperature and humidity had a significant (p < 0.05) effect on FCM concentrations with significant (p < 0.05) interactions of sex, age and breeding season. Once biotic factors were accounted for, there was a significant (p < 0.05) increase in FCM concentrations associated with ambient temperatures above 20 °C in adult polar bears. The implications of these findings for the management of both zoo and wild polar bears are discussed.

Impact of isolation method on cellular activation and presence of specific tendon cell subpopulations during in vitro culture.

Tendon injuries are common and heal poorly, due in part to a lack of understanding of fundamental tendon cell biology. A major impediment to the study of tendon cells is the absence of robust, well-characterized in vitro models. Unlike other tissue systems, current tendon cell models do not account for how differences in isolation methodology may affect the activation state of tendon cells or the presence of various tendon cell subpopulations. The objective of this study was to characterize how common isolation methods affect the behavior, fate, and lineage composition of tendon cell cultures. Tendon cells isolated by explant exhibited reduced proliferative capacity, decreased expression of tendon marker genes, and increased expression of genes associated with fibroblast activation compared to digested cells. Consistently, explanted cells also displayed an increased propensity to differentiate to myofibroblasts compared to digested cells. Explanted cultures from multiple different tendons were substantially enriched for the presence of scleraxis-lineage (Scx-lin+) cells compared to digested cultures, while the overall percentage of S100a4-lineage (S100a4-lin+) cells was dependent on both isolation method and tendon of origin. Neither isolation methods preserved the ratios of Scx-lin+ or S100a4-lin+ to non-lineage cells seen in tendons in vivo. Combined, these data indicate that further refinement of in vitro cultures models is required in order to more accurately understand the effects of various stimuli on tendon cell behavior. Statement of clinical significance: The development of informed in vitro tendon cell models will facilitate enhanced screening of potential therapeutic candidates to improve tendon healing.

A novel murine muscle loading model to investigate Achilles musculotendinous adaptation.

Achilles tendinopathy is a debilitating condition affecting the entire spectrum of society and a condition that increases the risk of tendon rupture. Effective therapies remain elusive, as anti-inflammatory drugs and surgical interventions show poor long-term outcomes. Eccentric loading of the Achilles muscle-tendon unit is an effective physical therapy for treatment of symptomatic human tendinopathy. Here, we introduce a novel mouse model of hindlimb muscle loading designed to achieve a tissue-targeted therapeutic exercise. This model includes the application of tissue (muscle and tendon)-loading "doses," coupled with ankle dorsiflexion and plantarflexion, inspired by human clinical protocols. Under computer control, the foot was rotated through the entire ankle joint range of motion while the plantar flexors simultaneously contracted to simulate body mass loading, consistent with human therapeutic exercises. This approach achieved two key components of the heel drop and raise movement: ankle range of motion coupled with body mass loading. Model development entailed the tuning of parameters such as footplate speed, number of repetitions, number of sets of repetitions, treatment frequency, treatment duration, and treatment timing. Initial model development was carried out on uninjured mice to define a protocol that was well tolerated and nondeleterious to tendon biomechanical function. When applied to a murine Achilles tendinopathy model, muscle loading led to a significant improvement in biomechanical outcome measures, with a decrease in cross-sectional area and an increase in material properties, compared with untreated animals. Our model facilitates the future investigation of mechanisms whereby rehabilitative muscle loading promotes healing of Achilles tendon injuries.NEW & NOTEWORTHY We introduce a novel mouse model of hindlimb muscle loading designed to achieve a tissue-targeted therapeutic exercise. This innovative model allows for application of muscle loading "doses," coupled with ankle dorsiflexion and plantarflexion, inspired by human loading clinical treatment. Our model facilitates future investigation of mechanisms whereby rehabilitative muscle loading promotes healing of Achilles tendon injuries.

Scleraxis-lineage cell depletion improves tendon healing and disrupts adult tendon homeostasis.

Despite the requirement for Scleraxis-lineage (ScxLin) cells during tendon development, the function of ScxLin cells during adult tendon repair, post-natal growth, and adult homeostasis have not been defined. Therefore, we inducibly depleted ScxLin cells (ScxLinDTR) prior to tendon injury and repair surgery and hypothesized that ScxLinDTR mice would exhibit functionally deficient healing compared to wild-type littermates. Surprisingly, depletion of ScxLin cells resulted in increased biomechanical properties without impairments in gliding function at 28 days post-repair, indicative of regeneration. RNA sequencing of day 28 post-repair tendons highlighted differences in matrix-related genes, cell motility, cytoskeletal organization, and metabolism. We also utilized ScxLinDTR mice to define the effects on post-natal tendon growth and adult tendon homeostasis and discovered that adult ScxLin cell depletion resulted in altered tendon collagen fibril diameter, density, and dispersion. Collectively, these findings enhance our fundamental understanding of tendon cell localization, function, and fate during healing, growth, and homeostasis.

Effects of tamoxifen on tendon homeostasis and healing: Considerations for the use of tamoxifen-inducible mouse models.

The use of tamoxifen-inducible models of Cre recombinase in the tendon field is rapidly expanding, resulting in an enhanced understanding of tendon homeostasis and healing. However, the effects of tamoxifen on the tendon are not well-defined, which is particularly problematic given that tamoxifen can have both profibrotic and antifibrotic effects in a tissue-specific manner. Therefore, in the present study, we examined the effects of tamoxifen on tendon homeostasis and healing in male and female C57Bl/6J mice. Tamoxifen-treated mice were compared to corn oil (vehicle)-treated mice. In the "washout" treatment regimen, mice were treated with tamoxifen or corn oil for 3 days beginning 1 week prior to undergoing complete transection and surgical repair of the flexor digitorum longus tendon. In the second regimen, mice were treated with tamoxifen or corn oil beginning on the day of surgery, daily through day 2 postsurgery, and every 48 hours thereafter (D0-2q48) until harvest. All repaired tendons and uninjured contralateral control tendons were harvested at day 14 postsurgery. Tamoxifen treatment had no effect on tendon healing in male mice, regardless of the treatment regimen, while Max load was significantly decreased in female repairs in the Tamoxifen washout group, relative to corn oil. In contrast, D0-2q48 corn oil treatment in female mice led to substantial disruptions in tendon homeostasis, relative to washout corn oil treatment. Collectively, these data clearly define the functional effects of tamoxifen and corn oil treatment in the tendon and inform future use of tamoxifen-inducible genetic models.

NF-κB activation persists into the remodeling phase of tendon healing and promotes myofibroblast survival.

Although inflammation is necessary during the early phases of tissue repair, persistent inflammation contributes to fibrosis. Acute tendon injuries often heal through a fibrotic mechanism, which impedes regeneration and functional recovery. Because inflammation mediated by nuclear factor κB (NF-κB) signaling is implicated in this process, we examined the spatial, temporal, and cell type-specific activation profile of canonical NF-κB signaling during tendon healing. NF-κB signaling was maintained through all phases of tendon healing in mice, including the remodeling phase, and tenocytes and myofibroblasts from the Scleraxis (Scx) lineage were the predominant populations that retained NF-κB activation into the late stages of repair. We confirmed persistent NF-κB activation in myofibroblasts in human tendon scar tissue. Deleting the canonical NF-κB kinase, IKKß, in Scx-lineage cells in mice increased apoptosis and the deposition of the matrix protein periostin during the late stages of tendon repair, suggesting that persistent NF-κB signaling may facilitate myofibroblast survival and fibrotic progression. Consistent with this, myofibroblasts in human tendon scar samples displayed enhanced prosurvival signaling compared to control tissue. Together, these data suggest that NF-κB may contribute to fibrotic tendon healing through both inflammation-dependent and inflammation-independent functions, such as NF-κB-mediated cell survival.

Effects of Type II Diabetes Mellitus on Tendon Homeostasis and Healing.

Over 300,000 tendon repairs are performed annually in the United States to repair damage to tendons as a result of either acute trauma or chronic tendinopathy. Individuals with type II diabetes mellitus (T2DM) are four times more likely to experience tendinopathy, and up to five times more likely to experience a tendon tear or rupture than non-diabetics. As nearly 10% of the US population is diabetic, with an additional 33% pre-diabetic, this is a particularly problematic health care challenge. Tendon healing in general is challenging and often unsatisfactory due to the formation of mechanically inferior scar-tissue rather than regeneration of native tendon structure. In T2DM tendons, there is evidence of an amplified scar tissue response, which may be associated with the increased the risk of rupture or impaired restoration of range of motion. Despite the dramatic effect of T2DM on tendon function and outcomes following injury, there are few therapies available to promote improved healing in these patients. Several recent studies have enhanced our understanding of the pro-inflammatory environment of T2DM healing and have assessed potential treatment approaches to mitigate pathological progression in pre-clinical models of diabetic tendinopathy. This review discusses the current state of knowledge of diabetic tendon healing from molecular to mechanical disruptions and identifies promising approaches and critical knowledge gaps as the field moves toward identification of novel therapeutic strategies to maintain or restore tendon function in diabetic patients. © 2019 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 38:13-22, 2020.

Urinary bladder matrix does not improve tenogenesis in an in vitro equine model.

Extracellular matrix (ECM) is responsible for tendon strength and elasticity. Healed tendon ECM lacks structural integrity, leading to reinjury. Porcine urinary bladder matrix (UBM) provides a scaffold and source of bioactive proteins to improve tissue healing, but has received limited attention for treating tendon injuries. The objective of this study was to evaluate the ability of UBM to induce matrix organization and tenogenesis using a novel in vitro model. We hypothesized that addition of UBM to tendon ECM hydrogels would improve matrix organization and cell differentiation. Hydrogels seeded with bone marrow cells (n = 6 adult horses) were cast using rat tail tendon ECM ± UBM, fixed under static tension and harvested at 7 and 21 days for construct contraction, cell viability, histology, biochemistry, and gene expression. By day 7, UBM constructs contracted significantly from baseline, whereas control constructs did not. Both control and UBM constructs contracted significantly by day 21. In both groups, cells remained viable over time and changed from round and randomly oriented to elongated along lines of tension with visible compaction of the ECM. There were no differences over time or between treatments for nuclear aspect ratio, DNA, or glycosaminoglycan content. Decorin, MMP-13, and Scleraxis expression increased significantly over time, but not in response to UBM treatment. Mohawk expression was constant over time. COMP expression decreased over time in both groups. Using a novel ECM hydrogel model, substantial matrix organization and cell differentiation occurred; however, addition of UBM failed to induce greater matrix organization than tendon ECM alone. This article is protected by copyright. All rights reserved.

Urinary Bladder Matrix Does Not Improve Tenogenesis in an In Vitro Equine Model.

Extracellular matrix (ECM) is responsible for tendon strength and elasticity. Healed tendon ECM lacks structural integrity, leading to reinjury. Porcine urinary bladder matrix (UBM) provides a scaffold and source of bioactive proteins to improve tissue healing, but has received limited attention for treating tendon injuries. The objective of this study was to evaluate the ability of UBM to induce matrix organization and tenogenesis using a novel in vitro model. We hypothesized that addition of UBM to tendon ECM hydrogels would improve matrix organization and cell differentiation. Hydrogels seeded with bone marrow cells (n = 6 adult horses) were cast using rat tail tendon ECM ± UBM, fixed under static tension and harvested at 7 and 21 days for construct contraction, cell viability, histology, biochemistry, and gene expression. By day 7, UBM constructs contracted significantly from baseline, whereas control constructs did not. Both control and UBM constructs contracted significantly by day 21. In both groups, cells remained viable over time and changed from round and randomly oriented to elongated along lines of tension with visible compaction of the ECM. There were no differences over time or between treatments for nuclear aspect ratio, DNA, or glycosaminoglycan content. Decorin, matrix metalloproteinase 13, and scleraxis expression increased significantly over time, but not in response to UBM treatment. Mohawk expression was constant over time. Cartilage oligomeric matrix protein expression decreased over time in both groups. Using a novel ECM hydrogel model, substantial matrix organization and cell differentiation occurred; however, the addition of UBM failed to induce greater matrix organization than tendon ECM alone. © 2019 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 37:1848-1859, 2019.

Cell non-autonomous functions of S100a4 drive fibrotic tendon healing.

Identification of pro-regenerative approaches to improve tendon healing is critically important as the fibrotic healing response impairs physical function. In the present study we tested the hypothesis that S100a4 haploinsufficiency or inhibition of S100a4 signaling improves tendon function following acute injury and surgical repair in a murine model. We demonstrate that S100a4 drives fibrotic tendon healing primarily through a cell non-autonomous process, with S100a4 haploinsufficiency promoting regenerative tendon healing. Moreover, inhibition of S100a4 signaling via antagonism of its putative receptor, RAGE, also decreases scar formation. Mechanistically, S100a4 haploinsufficiency decreases myofibroblast and macrophage content at the site of injury, with both cell populations being key drivers of fibrotic progression. Moreover, S100a4-lineage cells become α-SMA+ myofibroblasts, via loss of S100a4 expression. Using a combination of genetic mouse models, small molecule inhibitors and in vitro studies we have defined S100a4 as a novel, promising therapeutic candidate to improve tendon function after acute injury.