Biology and physiology of tendon healing.

Tendon disorders affect people of all ages, from elite and recreational athletes and workers to elderly patients. After an acute injury, 3 successive phases are described to achieve healing: an inflammatory phase followed by a proliferative phase, and finally by a remodeling phase. Despite this process, healed tendon fails to recover its original mechanical properties. In this review, we proposed to describe the key factors involved in the process such as cells, transcription factors, extracellular matrix components, cytokines and growth factors and vascularization among others. A better understanding of this healing process could help provide new therapeutic approaches to improve patients' recovery while tendon disorders management remains a medical challenge.

Neonatal Achilles Tendon Microstructure is Negatively Impacted by Decorin and Biglycan Knockdown After Injury and During Development.

Interest in studying neonatal development and the improved healing response observed in neonates is increasing, with the goal of using this work to create better therapeutics for tendon injury. Decorin and biglycan are two small leucine-rich proteoglycans that play important roles in collagen fibrillogenesis to develop, maintain, and repair tendon structure. However, little is known about the roles of decorin and biglycan in early neonatal development and healing. The goal of this study was to determine the effects of decorin and biglycan knockdown on Achilles tendon structure and mechanics during neonatal development and recovery of these properties after injury of the neonatal tendon. We hypothesized that knockdown of decorin and biglycan would disrupt the neonatal tendon developmental process and produce tendons with impaired mechanical and structural properties. We found that knockdown of decorin and biglycan in an inducible, compound decorin/biglycan knockdown model, both during development and after injury, in neonatal mice produced tendons with reduced mechanical properties. Additionally, the collagen fibril microstructure resembled an immature tendon with a large population of small diameter fibrils and an absence of larger diameter fibrils. Overall, this study demonstrates the importance of decorin and biglycan in facilitating tendon growth and maturation during neonatal development.

Collagen V haploinsufficiency in female murine patellar tendons results in altered matrix engagement and cellular density, demonstrating decreased healing.

Collagen V (Col5) is a quantitatively minor component of collagen fibrils comprising tendon, however, plays a crucial role in regulation of development and dynamic healing processes. Clinically, patients with COL5a1 haploinsufficiency, known as classic Ehlers-Danlos Syndrome (cEDS), present with hyperextensible skin, joint instability and laxity, with females more likely to be affected. Previous studies in Col5-deficient mice indicated that reduced Col5a1 expression leads to a reduction in stiffness, fibril deposition, and altered fibril structure. Additionally, Col5-deficient male tendons demonstrated altered healing compared to wild-type tendons, however female mice have not yet been studied utilizing this model. Along with clinical differences between sexes in cEDS patient populations, differences in hormone physiology may be a factor influencing tendon health. Therefore, the objective of this study was to utilize a Col5a1+/ - female mouse model, to determine the effect of Col5 on tendon cell morphology, cell density, tissue composition, and mechanical properties throughout healing. We hypothesized that reduction in Col5 expression would result in an abnormal wound matrix post-injury, resulting in reduced mechanical properties compared to normal tendons. Following patellar tendon surgery, mice were euthanized at 1, 3, and 6-week post-injury. Col5-deficient tendons demonstrated altered and decreased healing compared to WT tendons. The lack of resolution in cellularity by 6-week post-injury in Col5-deficient tendons influenced the decreased mechanical properties. Stiffness did not increase post-injury in Col5-deficient mice, and collagen fiber realignment was delayed during mechanical loading. Therefore, increased Col5a1 expression post-injury is necessary to re-establish matrix engagement and cellularity throughout tendon healing.

Long-Term Effects of Reproduction and Lactation on the Rat Supraspinatus Tendon and Proximal Humerus.

During pregnancy and breastfeeding, women undergo hormonal fluctuations required for fetal development, parturition, and infant growth. These changes have secondary consequences on the maternal musculoskeletal system, increasing the risk for joint pain and osteoporosis. Though hormone levels return to prepregnancy levels postpartum, women may experience lasting musculoskeletal pain. Sex disparities exist in the prevalence of musculoskeletal disorders, but it remains unclear how reproductive history may impact sex differences. Specifically, the effects of both reproductive history and sex on the rotator cuff have not been studied. Pregnancy and lactation affect bone microstructure, suggesting possible impairments at the enthesis of rotator cuff tendons, where tears commonly occur. Therefore, our objective was to evaluate how reproductive history affects sex differences of the supraspinatus tendon and proximal humerus using male, virgin female, and female rats with a history of reproduction (referred to as reproductive females). We hypothesized tendon mechanical properties and humeral bone microstructure would be inferior in reproductive females compared to virgin females. Results showed sex differences independent of reproductive history, including greater tendon midsubstance modulus but lower subchondral bone mineral density (BMD) in females. When considering reproductive history, reproductive rats exhibited reduced tendon insertion site modulus and trabecular bone micro-architecture compared to virgin females with no differences from males. Overall, our study identified long-term changes in supraspinatus tendon mechanical and humeral trabecular bone properties that result following pregnancy and lactation, highlighting the importance of considering reproductive history in investigations of sex differences in the physiology and pathology of rotator cuff injuries.

Novel application of in vivo microdialysis in a rat Achilles tendon acute injury model.

Tendon injury and healing involve intricate changes to tissue metabolism, biology, and inflammation. Current techniques often require animal euthanasia or tissue destruction, limiting assessment of dynamic changes in tendon, including treatment response, disease development, rupture risk, and healing progression. Microdialysis, a minimally invasive technique, offers potential for longitudinal assessment, yet it has not been applied to rat tendon models. Therefore, the objective of this study is to adapt a novel application of an in vivo assay, microdialysis, using acute injury as a model for extreme disruption of the tendon homeostasis. We hypothesize that microdialysis will be able to detect measurable differences in the healing responses of acute injury with high specificity and sensitivity. Overall results suggest that microdialysis is a promising in vivo technique for longitudinal assessment for this system with strong correlations between extracellular fluid (ECF) and dialysate concentrations and reasonable recovery rates considering the limitations of this model. Strong positive correlations were found between dialysate and extracellular fluid (ECF) concentration for each target molecule of interest including metabolites, inflammatory mediators, and collagen synthesis and degradation byproducts. These results suggest that microdialysis is capable of detecting changes in tendon healing following acute tendon injury with high specificity and sensitivity. In summary, this is the first study to apply microdialysis to a rat tendon model and assess its efficacy as a direct measurement of tendon metabolism, biology, and inflammation.NEW & NOTEWORTHY This study adapts a novel application of microdialysis to rat tendon models, offering a minimally invasive avenue for longitudinal tendon assessment. Successfully detecting changes in tendon healing after acute injury, it showcases strong correlations between extracellular fluid and dialysate concentrations. The results highlight the potential of microdialysis as a direct measure of tendon metabolism, biology, and inflammation, bypassing the need for animal euthanasia and tissue destruction.

Preclinical tendon and ligament models: Beyond the 3Rs (replacement, reduction, and refinement) to 5W1H (why, who, what, where, when, how).

Several tendon and ligament animal models were presented at the 2022 Orthopaedic Research Society Tendon Section Conference held at the University of Pennsylvania, May 5 to 7, 2022. A key objective of the breakout sessions at this meeting was to develop guidelines for the field, including for preclinical tendon and ligament animal models. This review summarizes the perspectives of experts for eight surgical small and large animal models of rotator cuff tear, flexor tendon transection, anterior cruciate ligament tear, and Achilles tendon injury using the framework: "Why, Who, What, Where, When, and How" (5W1H). A notable conclusion is that the perfect tendon model does not exist; there is no single gold standard animal model that represents the totality of tendon and ligament disease. Each model has advantages and disadvantages and should be carefully considered in light of the specific research question. There are also circumstances when an animal model is not the best approach. The wide variety of tendon and ligament pathologies necessitates choices between small and large animal models, different anatomic sites, and a range of factors associated with each model during the planning phase. Attendees agreed on some guiding principles including: providing clear justification for the model selected, providing animal model details at publication, encouraging sharing of protocols and expertise, improving training of research personnel, and considering greater collaboration with veterinarians. A clear path for translating from animal models to clinical practice was also considered as a critical next step for accelerating progress in the tendon and ligament field.

Investigating the temporal roles of decorin and biglycan in tendon healing.

The small leucine-rich proteoglycans, decorin and biglycan, are minor components of the tendon extracellular matrix that regulate fibrillogenesis and matrix assembly. Our study objective was to define the temporal roles of decorin and biglycan during tendon healing using inducible knockout mice to include genetic knockdown at specific phases of healing: time of injury, the proliferative phase, and the remodeling phase. We hypothesized that knockdown of decorin or biglycan would adversely affect tendon healing, and that by prescribing the timing of knockdown, we could elucidate the temporal roles of these proteins during healing. Contrary to our hypothesis, decorin knockdown did not affect tendon healing. However, when biglycan was knocked down, either alone or coupled with decorin, tendon modulus was increased relative to wild-type mice, and this finding was consistent among all induction timepoints. At 6 weeks postinjury, we observed increased expression of genes associated with the extracellular matrix and growth factor signaling in the biglycan knockdown and compound decorin-biglycan knockdown tendons. Interestingly, these groups demonstrated opposing trends in gene expression as a function of knockdown-induction timepoint, highlighting distinct temporal roles for decorin and biglycan. In summary, this study finds that biglycan plays multiple functions throughout tendon healing, with the most impactful, detrimental role likely occurring during late-stage healing. Statement of clinical importance: This study helps to define the molecular factors that regulate tendon healing, which may aid in the development of new clinical therapies.

Knockdown of biglycan reveals an important role in maintenance of structural and mechanical properties during tendon aging.

Biglycan, a small leucine-rich proteoglycan (SLRP), is involved in collagen fibrillogenesis and also acts as a signaling molecule. Although decorin has been considered as the primary SLRP in developing and maintaining tendon structure and mechanics, more recent work using inducible knockdown models suggests that biglycan is involved in tendon homeostasis. The purpose of the study was to determine the role of biglycan in tendon homeostasis to maintain mechanical and structural integrity in aged mice. Aged (485 days old) female Bgn+/+ control (wild type [WT], n = 16) and 16 bitransgenic conditional Bgnflox/flox mice (I-Bgn-/- , n = 16) with a tamoxifen-inducible Cre (driven by ROSA) were utilized. After biglycan knockdown, the transgenic model demonstrated effective knockdown of the target gene without any compensation from other SLRPs or type I collagen. Patellar tendon cellularity was not modified after biglycan knockdown. However, biglycan knockdown had an impact on collagen fibrillogenesis with a higher percentage of small diameter fibrils (25-45 nm) and a lower percentage of medium size fibrils (150-165 nm) in I-Bgn-/- tendons. Biglycan knockdown also induced a reduction in the midsubstance modulus and maximum stress compared to WT. Stress relaxation was reduced at 4% strain in I-Bgn-/- tendons but no changes were observed in dynamic modulus and tan delta. As in mature tendons (120 days old), this study showed significant effects of biglycan knockdown on mechanical and structural properties of aged tendons only 30 days after knockdown. These data suggest that biglycan has a major role in maintaining homeostasis in aged tendon.

Post-injury tendon mechanics are not affected by tamoxifen treatment.

PURPOSE: A growing interest in the mechanisms that govern tendon healing has resulted in the develop-ment of tools, such as the tamoxifen-inducible mouse knockdown model, to address these questions. However, tamoxifen is a selective estrogen receptor modulator and may interfere with the tendon healing process. The objective of this study was to evaluate the effects of tamoxifen on post-injury tendon mechanics in wild-type mice. METHODS: The mice underwent treatment at the time of injury using an established mouse injury model and the injured tendons were evaluated 3 weeks post-injury. The treatment contained tamoxifen suspended in corn oil and was compared to a treatment with only corn oil, as well as mice with no treatment. Tendons were evaluated by measuring the quasi-static and viscoelastic mechanics, collagen fiber realignment, cellularity, and nuclear morphology. RESULTS: Mechanical testing of the tendons post-injury revealed no changes to viscoelastic mechanics, quasi-static mechanics, or collagen realignment during loading after tamoxifen treatment with the dosage regimen utilized (three daily injections of 4.5 mg/40 g body weight). Additionally, histological analysis revealed no changes to cellularity or cell nuclear shape. CONCLUSION: Overall, this study revealed that tamoxifen treatment at the time of tendon injury did not result in changes to tendon mechanics or the histological parameters at 3 weeks post-injury.

Targeted conditional collagen XII deletion alters tendon function.

Collagen XII is a fibril-associated collagen with interrupted triple helices (FACIT). This non-fibrillar collagen is a homotrimer composed of three α1(XII) chains assembled into a collagenous molecule with a C terminal collagenous domain and a large N terminal non-collagenous domain. During tendon development and growth, collagen XII is broadly expressed throughout the extracellular matrix and enriched pericellularly around tenocytes. Tendons in a global Col12a1 -/- knockout model demonstrated disrupted fibril and fiber structure and disordered tenocyte organization, highlighting the critical regulatory roles of collagen XII in determining tendon structure and function. However, muscle and bone also are affected in the collagen XII knockout model. Therefore, secondary effects on tendon due to involvement of bone and muscle may occur in the global knockout. The global knockout does not allow the definition of intrinsic mechanisms involving collagen XII in tendon versus extrinsic roles involving muscle and bone. To address this limitation, we created and characterized a conditional Col12a1-null mouse model to permit the spatial and temporal manipulation of Col12a1 expression. Collagen XII knockout was targeted to tendons by breeding conditional Col12a1 flox/flox mice with Scleraxis-Cre (Scx-Cre) mice to yield a tendon-specific Col12a1-null mouse line, Col12a1 Δten/Δten . Both mRNA and protein expression in Col12a1 Δten/Δten mice decreased to near baseline levels in flexor digitorum longus tendons (FDL). Collagen XII immuno-localization revealed an absence of reactivity in the tendon proper, but there was reactivity in the cells of the surrounding peritenon. This supports a targeted knockout in tenocytes while peritenon cells from a non-tendon lineage were not targeted and retained collagen XII expression. The tendon-targeted, Col12a1 Δten/Δten  mice had significantly reduced forelimb grip strength, altered gait and a significant decrease in biomechanical properties. While the observed decrease in tendon modulus suggests that differences in tendon material properties in the absence of Col12a1 expression underlie the functional deficiencies. Together, these findings suggest an intrinsic role for collagen XII critical for development of a functional tendon.

Decorin knockdown is beneficial for aged tendons in the presence of biglycan expression.

Decorin and biglycan are two major small leucine-rich proteoglycans (SLRPs) present in the tendon extracellular matrix that facilitate collagen fibrillogenesis, tissue turnover, and cell signal transduction. Previously, we demonstrated that knockout of decorin prevented the decline of tendon mechanical properties that are associated with aging. The objective of this study was to determine the effects of decorin and biglycan knockdown on tendon structure and mechanics in aged tendons using tamoxifen-inducible knockdown models. We hypothesized that the knockdown of decorin and compound knockdown of decorin and biglycan would prevent age-related declines in tendon mechanics and structure compared to biglycan knockdown and wild-type controls, and that these changes would be exacerbated as the tendons progress towards geriatric ages. To achieve this objective, we created tamoxifen-inducible mouse knockdown models to target decorin and biglycan gene inactivation without the abnormal tendon development associated with traditional knockout models. Knockdown of decorin led to increased midsubstance modulus and decreased stress relaxation in aged tendons. However, these changes were not sustained in the geriatric tendons. Knockdown in biglycan led to no changes in mechanics in the aged or geriatric tendons. Contrary to our hypothesis, the compound decorin/biglycan knockdown tendons did not resemble the decorin knockdown tendons, but resulted in increased viscoelastic properties in the aged and geriatric tendons. Structurally, knockdown of SLRPs, except for the 570d I-Dcn -/- /Bgn -/- group, resulted in alterations to the collagen fibril diameter relative to wild-type controls. Overall, this study identified the differential roles of decorin and biglycan throughout tendon aging in the maintenance of tendon structural and mechanical properties and revealed that the compound decorin and biglycan knockdown phenotype did not resemble the single gene decorin or biglycan models and was detrimental to tendon properties throughout aging.

The Non-pregnant and Pregnant Human Cervix: a Systematic Proteomic Analysis.

Appropriate timing of cervical remodeling (CR) is key to normal term parturition. To date, mechanisms behind normal and abnormal (premature or delayed) CR remain unclear. Recent studies show regional differences exist in human cervical tissue structure. While the entire cervix contains extracellular matrix (ECM), the internal os is highly cellular containing 50-60% cervical smooth muscle (CSM). The external os contains 10-20% CSM. Previously, we reported ECM rigidity and different ECM proteins influence CSM cell function, highlighting the importance of understanding not only how cervical cells orchestrate cervical ECM remodeling in pregnancy, but also how changes in specific ECM proteins can influence resident cellular function. To understand this dynamic process, we utilized a systematic proteomic approach to understand which soluble ECM and cellular proteins exist in the different regions of the human cervix and how the proteomic profiles change from the non-pregnant (NP) to the pregnant (PG) state. We found the human cervix proteome contains at least 4548 proteins and establish the types and relative abundance of cellular and soluble matrisome proteins found in the NP and PG human cervix. Further, we report the relative abundance of proteins involved with elastic fiber formation and ECM organization/degradation were significantly increased while proteins involved in RNA polymerase I/promoter opening, DNA methylation, senescence, immune system, and compliment activation were decreased in the PG compared to NP cervix. These findings establish an initial platform from which we can further comprehend how changes in the human cervix proteome results in normal and abnormal CR.

Increasing Vascular Response to Injury Improves Tendon Early Healing Outcome in Aged Rats.

Tendon injuries positively correlate with patient age, as aging has significant effects on tendon homeostatic maintenance and healing potential after injury. Vascularity is also influenced by age, with both clinical and animal studies demonstrating reduced blood flow in aged tissues. However, it is unknown how aging effects vascularity following tendon injury, and if this vascular response can be modulated through the delivery of angiogenic factors. Therefore, the objective of this study is to evaluate the vascular response following Achilles tendon injury in adult and aged rats, and to define the alterations to tendon healing in an aged model following injection of angiogenic factors. It was determined that aged rat Achilles tendons have a reduced angiogenesis following injury. Further, the delivery of vascular endothelial growth factor, VEGF, caused an increase in vascular response to tendon injury and improved mechanical outcome in this aged population. This work suggests that reduced angiogenic potential with aging may be contributing to impaired tendon healing response and that the delivery of angiogenic factors can rescue this impaired response. This study was also the first to relate changes in vascular response in an aged model using in vivo measures of blood perfusion to alterations in healing properties.

Biglycan has a major role in maintenance of mature tendon mechanics.

Decorin and biglycan are two small leucine-rich proteoglycans (SLRPs) that regulate collagen fibrillogenesis and extracellular matrix assembly in tendon. The objective of this study was to determine the individual roles of these molecules in maintaining the structural and mechanical properties of tendon during homeostasis in mature mice. We hypothesized that knockdown of decorin in mature tendons would result in detrimental changes to tendon structure and mechanics while knockdown of biglycan would have a minor effect on these parameters. To achieve this objective, we created tamoxifen-inducible mouse knockdown models targeting decorin or biglycan inactivation. This enables the evaluation of the roles of these SLRPs in mature tendon without the abnormal tendon development caused by conventional knockout models. Contrary to our hypothesis, knockdown of decorin resulted in minor alterations to tendon structure and no changes to mechanics while knockdown of biglycan resulted in broad changes to tendon structure and mechanics. Specifically, knockdown of biglycan resulted in reduced insertion modulus, maximum stress, dynamic modulus, stress relaxation, and increased collagen fiber realignment during loading. Knockdown of decorin and biglycan produced similar changes to tendon microstructure by increasing the collagen fibril diameter relative to wild-type controls. Biglycan knockdown also decreased the cell nuclear aspect ratio, indicating a more spindle-like nuclear shape. Overall, the extensive changes to tendon structure and mechanics after knockdown of biglycan, but not decorin, provides evidence that biglycan plays a major role in the maintenance of tendon structure and mechanics in mature mice during homeostasis.

Coordinate roles for collagen VI and biglycan in regulating tendon collagen fibril structure and function.

Tendon is a vital musculoskeletal tissue that is prone to degeneration. Proper tendon maintenance requires complex interactions between extracellular matrix components that remain poorly understood. Collagen VI and biglycan are two matrix molecules that localize pericellularly within tendon and are critical regulators of tissue properties. While evidence suggests that collagen VI and biglycan interact within the tendon matrix, the relationship between the two molecules and its impact on tendon function remains unknown. We sought to elucidate potential coordinate roles of collagen VI and biglycan within tendon by defining tendon properties in knockout models of collagen VI, biglycan, or both molecules. We first demonstrated co-expression and co-localization of collagen VI and biglycan within the healing tendon, providing further evidence of cooperation between the two molecules during nascent tendon matrix formation. Deficiency in collagen VI and/or biglycan led to significant reductions in collagen fibril size and tendon mechanical properties. However, collagen VI-null tendons displayed larger reductions in fibril size and mechanics than seen in biglycan-null tendons. Interestingly, knockout of both molecules resulted in similar properties to collagen VI knockout alone. These results indicate distinct and non-additive roles for collagen VI and biglycan within tendon. This work provides better understanding of regulatory interactions between two critical tendon matrix molecules.

Pulsed electromagnetic field therapy alters early healing in a rat model of rotator cuff injury and repair: Potential mechanisms.

Rotator cuff repair failure remains common due to poor tendon healing, particularly at the enthesis. We previously showed that pulsed electromagnetic field (PEMF) therapy improved the mechanical properties of the rat supraspinatus tendon postoperatively. However, little is known about the mechanisms behind PEMF-dependent contributions to improved healing in this injury model. The objective of this study was to determine the influence of PEMF treatment on tendon gene expression and cell composition, as well as bone microarchitecture and dynamic bone metabolism during early stages of healing. We hypothesized that PEMF treatment would amplify tendon-healing related signaling pathways while mitigating inflammation and improve bone metabolism at the repair site. Rats underwent rotator cuff injury and repair followed by assignment to either control (non-PEMF) or PEMF treatment groups. Gene and protein expression as well as tendon and bone histological assessments were performed 3, 7, 14, 21, and 28 days after injury. Gene expression data demonstrated an upregulation in the bone morphogenetic protein 2 signaling pathway and increases in pro-osteogenic genes at the insertion, supporting important processes to re-establish the tendon-bone interface. PEMF also downregulated genes related to a fibrotic healing response. Anti-inflammatory effects were demonstrated by both gene expression and macrophage phenotype. PEMF significantly increased the rate of kinetic bone formation directly adjacent to the tendon enthesis as well as the number of cuboidal surface osteoblasts (active osteoblasts) in the humeral head. This study has provided insight into how PEMF affects cellular and molecular processes in the supraspinatus tendon and adjacent bone after injury and repair.

Inhibition of glucose use improves structural recovery of injured Achilles tendon in mice.

Injured tendons do not regain their native structure except at fetal or very young ages. Healing tendons often show mucoid degeneration involving accumulation of sulfated glycosaminoglycans (GAGs), but its etiology and molecular base have not been studied substantially. We hypothesized that quality and quantity of gene expression involving the synthesis of proteoglycans having sulfated GAGs are altered in injured tendons and that a reduction in synthesis of sulfated GAGs improves structural and functional recovery of injured tendons. C57BL6/j mice were subjected to Achilles tendon tenotomy surgery. The injured tendons accumulated sulfate proteoglycans as early as 1-week postsurgery and continued so by 4-week postsurgery. Transcriptome analysis revealed upregulation of a wide range of proteoglycan genes that have sulfated GAGs in the injured tendons 1 and 3 weeks postsurgery. Genes critical for enzymatic reaction of initiation and elongation of chondroitin sulfate GAG chains were also upregulated. After the surgery, mice were treated with the 2-deoxy-d-glucose (2DG) that inhibits conversion of glucose to glucose-6-phosphate, an initial step of glucose metabolism as an energy source and precursors of monosaccharides of GAGs. The 2DG treatment reduced accumulation of sulfated proteoglycans, improved collagen fiber alignment, and reduced the cross-sectional area of the injured tendons. The modulus of the 2DG-treated groups was higher than that in the vehicle group, but not of statistical significance. Our findings suggest that mucoid degeneration in injured tendons may result from the upregulated expression of genes involved the synthesis of sulfate proteoglycans and can be inhibited by reduction of glucose utilization.

Nonsurgical treatment reduces tendon inflammation and elevates tendon markers in early healing.

Operative treatment is assumed to provide superior outcomes to nonoperative (conservative) treatment following Achilles tendon rupture, however, this remains controversial. This study explores the effect of surgical repair on Achilles tendon healing. Rat Achilles tendons (n = 101) were bluntly transected and were randomized into groups receiving repair or non-repair treatments. By 1 week after injury, repaired tendons had inferior mechanical properties, which continued to 3- and 6-week post-injury, evidenced by decreased dynamic modulus and failure stress. Transcriptomics analysis revealed >7000 differentially expressed genes between repaired and non-repaired tendons after 1-week post-injury. While repaired tendons showed enriched inflammatory gene signatures, non-repaired tendons showed increased tenogenic, myogenic, and mechanosensitive gene signatures, with >200-fold enrichment in Tnmd expression. Analysis of gastrocnemius muscle revealed elevated MMP activity in tendons receiving repair treatment, despite no differences in muscle fiber morphology. Transcriptional regulation analysis highlighted that the highest expressed transcription factors in repaired tendons were associated with inflammation (Nfκb, SpI1, RelA, and Stat1), whereas non-repaired tendons expressed markers associated with tissue development and mechano-activation (Smarca1, Bnc2, Znf521, Fbn1, and Gli3). Taken together, these data highlight distinct differences in healing mechanism occurring immediately following injury and provide insights for new therapies to further augment tendons receiving repaired and non-repaired treatments.