Tendon injuries are common and can dramatically impair patient mobility and productivity, resulting in a significant socioeconomic burden and reduced quality of life. Because the tendon healing process results in the formation of a fibrotic scar, injured tendons never regain the mechanical strength of the uninjured tendon, leading to frequent re-injury. Many tendons are also prone to the development of peritendinous adhesions and excess scar formation, which further reduce tendon function and lead to chronic complications. Despite this, there are currently no treatments that adequately improve the tendon healing process due in part to a lack of information regarding the contributions of various cell types to tendon healing and how their activity may be modulated for therapeutic value. In this review, we summarize recent efforts to identify and characterize the distinct cell populations involved at each stage of tendon healing. In addition, we examine the mechanisms through which different cell populations contribute to the fibrotic response to tendon injury, and how these responses can be affected by systemic factors and comorbidities. We then discuss gaps in our current understanding of tendon fibrosis and highlight how new technologies and research areas are shedding light on this clinically important and intractable challenge. A better understanding of the complex cellular environment during tendon healing is crucial to the development of new therapies to prevent fibrosis and promote tissue regeneration.

During tendon healing, it is postulated that tendon cells drive tissue regeneration, whereas extrinsic cells drive pathologic scar formation. Tendon cells are frequently described as a homogenous, fibroblast population that is positive for the marker Scleraxis (Scx). It is controversial whether tendon cells localize within the forming scar tissue during adult tendon healing. We have previously demonstrated that S100 calcium‐binding protein A4 (S100a4) is a driver of tendon scar formation and marks a subset of tendon cells. The relationship between Scx and S100a4 has not been explored. In this study, we assessed the localization of Scx lineage cells (ScxLin) following adult murine flexor tendon repair and established the relationship between Scx and S100a4 throughout both homeostasis and healing. We showed that adult ScxLin localize within the scar tissue and organize into a cellular bridge during tendon healing. Additionally, we demonstrate that markers Scx and S100a4 label distinct populations in tendon during homeostasis and healing, with Scx found in the organized bridging tissue and S100a4 localized throughout the entire scar region. These studies define a heterogeneous tendon cell environment and demonstrate discrete contributions of subpopulations during healing. These data enhance our understanding and ability to target the cellular environment of the tendon.—Best, K. T., Loiselle, A. E. Scleraxis lineage cells contribute to organized bridging tissue during tendon healing and identify a subpopulation of resident tendon cells. FASEB J. 33, 8578–8587 (2019). http://www.fasebj.org

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.

Despite the requirement for Scleraxis-lineage (Scx Lin ) cells during tendon development, the function of Scx Lin cells during adult tendon repair, post-natal growth, and adult homeostasis have not been defined. Therefore, we crossed Scx-Cre and Diphtheria Toxin Receptor (DTR) mouse models to inducibly deplete Scx Lin cells prior to acute flexor tendon injury and repair (ScxLin DTR ) and hypothesized that ScxLin DTR mice would exhibit worse healing than wildtype littermates. Surprisingly, depletion of Scx Lin cells resulted in increased biomechanical properties without detriments to tendon gliding function at 28 days post-repair, indicative of regeneration. These improvements in biomechanical properties correspond to increased αSMA+ myofibroblasts in ScxLin DTR repairs, indicating that increased myofibroblast-mediated matrix remodeling may be responsible for accelerated recovery of tensile strength. Furthermore, we have defined the relative timing of Scx Lin cell localization relative to collagen deposition in bridging tissue of the repair and demonstrated a predominance of non-Scx Lin cells in the early bridging tissue, consistent with lack of bridging tissue disruption in ScxLin DTR repairs. Lastly, we utilized ScxLin DTR mice to assess effects of tendon cell depletion on post-natal tendon growth and adult tendon homeostasis. Collectively, these findings enhance our understanding of tendon cell localization, function, and fate during healing, growth, and homeostasis.3 Introduction:Despite the significant efforts toward improving tendon healing and regeneration, the specific cellular contributions during tendon healing have not been extensively characterized(1). While many studies have examined the potential of using various stem cell populations to promote healing, originating from both tendon intrinsic(2) and extrinsic sources(3), little focus has been directed toward defining the functions and therapeutic potential of tendon cells during tendon healing following an acute injury. Tendon cells are increasingly being recognized as a heterogenous population of cells where many, but not all, express the gene Scleraxis (Scx)(4-6).Understanding the localization and function of tendon cell subpopulations during healing could be instrumental in discovering why tendon heals using scar-mediated processes, resulting in poor patient outcomes, and could be used to develop pro-regenerative approaches to improve healing.Scx, a basic helix-loop-helix transcription factor, is currently the most well-characterized marker that the field possesses to study tendon (7). Scx has been utilized to examine tendon biology and development(8-10), healing and regeneration(4, 11-13), differentiation(14-17), and mechano-transduction (18,19). Functionally, Scx expression has previously been shown to drive matrix production and remodeling (11,20,21), epithelial-tomesenchymal transition(22), development of force-transmitting tendons(8), and effect focal adhesion morphology (19). Despite the effort to understand the functions of Scx as a transcr...

Flexor tendon injuries heal with excessive scar tissue that limits range of motion and increases incidence of re-rupture. The molecular mechanisms that govern tendon healing are not well defined. Both the canonical nuclear factor kappa B (NF-κB) and mitogen activated protein kinase (MAPK) pathways have been implicated in tendon healing. The gene NFKB1 (proteins p105/p50) is involved in both NF-κB and MAPK signaling cascades. In the present study, we tested the hypothesis that global NFKB1 deletion would increase activation of both NF-κB and MAPK through loss of signaling repressors, resulting in increased matrix deposition and altered biomechanical properties. As hypothesized, NFKB1 deletion increased activation of both NF-κB and MAPK signaling. While gliding function was not affected, NFKB1 deletion resulted in tendons that were significantly stiffer and trending towards increased strength by four weeks post-repair. NFKB1 deletion resulted in increased collagen deposition, increase macrophage recruitment, and increased presence of myofibroblasts. Furthermore, NFKB1 deletion increased expression of matrix-related genes ( Col1a1 , Col3a1 ), macrophage-associated genes ( Adgre1 , Ccl2 ), myofibroblast markers ( Acta2 ), and general inflammation ( Tnf ). Taken together, these data suggest that increased activation of NF-κB and MAPK via NFKB1 deletion enhance macrophage and myofibroblast content at the repair, driving increased collagen deposition and biomechanical properties.