Tendon injury occurs at high frequency and is difficult to repair. Identification of human stem cells being able to regenerate tendon will greatly facilitate the development of regenerative medicine for tendon injury. Genetic and functional analyses identify human CD29+/CD56+ myogenic progenitors with tenogenic differentiation potential in vitro and in vivo. Transplantation of human CD29+/CD56+ myogenic progenitors contributes to injured tendon repair and thus improves locomotor function. Interestingly, the tendon differentiation potential in mouse muscle stem cells is minimal and the higher TGFβ signaling level may be the key for the distinct feature of human CD29+/CD56+ myogenic progenitors. The discovery of bi-potential CD29+/CD56+ myogenic progenitors highlights their potential as a novel adult stem cell source for tendon regeneration.

Tendons are force-transmitting structures which facilitate musculoskeletal functioning. Characterizing variations between different anatomical tendons, regions within tendons, as well as between the sexes and with age can improve understanding of tendon physiology and pathology. A systematic search of the literature was conducted to identify and summarize microscopic structural (histological) variations in normal/healthy tendons in relation to these variables (Tendon, Region, Age, Sex, and Other). Regional differences within individual tendons have been investigated in numerous studies, however investigations comparing histological variations between a range of different tendons are sparse, with most focusing on a few select tendons. When injured, ageing tendons typically have a greater degree of pathological changes than younger tendons, but few studies have documented variations in tendon histology throughout typical (uninjured) ageing or across large age spans. Similarly, sex-related observations of tendon structure are underreported. This narrative review summarizes studies on these topics and explores interactions between these variables, as well as the implications of these in the context of selecting control samples for studies of tendon pathology. Future studies should endeavour to improve knowledge of tendon structural variations-specifically focusing on normal tendons-to facilitate understanding of tendon structure-function relationships, physiological mechanisms involved in tendon damage/healing, and to aid clinical research and practice.

Age-related diseases are often linked to chronic inflammation. Senescent cells secrete inflammatory cytokines, chemokines and matrix metalloproteinases, collectively referred to as the senescence-associated secretory phenotype (SASP). The current study discovered that aging leads to the accumulation of senescent tendon stem/progenitor cells (TSPCs) in tendon tissue, resulting in the development of a SASP. Conditioned medium from aged TSPCs induced cellular inflammation in young TSPCs. In addition, we found that Canopy homolog 2 (CNPY2) expression is reduced during tendon aging. CNPY2 deficiency causes TSPCs senescence and SASP. Our findings showed that the NF-κB signaling pathway is activated in CNPY2 knockdown TSPCs, pharmacological inhibition of NF-κB signaling pathway with BMS-345541 attenuated SASP of senescent TSPCs, which indicated that CNPY2 regulates TSPCs SASP might through NF-κB signaling pathway. Our findings suggested that CNPY2 plays an important role in TSPCs senescence and SASP, CNPY2 could be a promising target for age-related tendon disorders.

Previously, chitosan reduces the senescence-related phenotypes in human foreskin fibroblasts through the transforming growth factor beta (TGF-β) pathway, and enhances the proliferation and migration capabilities of these cells are demonstrated. In this study, we examined whether the senescence-delaying effect of chitosan could be applied to primary knee-related fibroblasts, such as human synovial membrane derived cells (SCs) and anterior cruciate ligament fibroblasts (ACLs). These two types of cells were obtained from donors who needed ACL reconstruction or knee replacement. We found that chitosan treatment effectively reduced aging-associated β-galactosidase (SA-β-gal)-positive cells, downregulated the expression of senescence-related proteins pRB and p53, and enhanced the 5-bromo-2'-deoxyuridine (BrdU) incorporation ability of SCs and ACLs. Moreover, chitosan could make SCs secret more glycosaminoglycans (GAGs) and produce type I collagen. The ability of ACLs to close the wound was also enhanced, and the TGF-β and alpha smooth muscle actin (αSMA) protein expression decreased after chitosan treatment. In summary, chitosan not only delayed the senescence but also enhanced the functions of SCs and ACLs, which is beneficial to the application of chitosan in cell expansion in vitro and cell therapy.

Our aim was to investigate occurrence of senescent cells directly in tendon tissue biopsies from patients with chronic shoulder tendinopathies, and to correlate senescence with Enhancer of zeste 2 (EZH2) expression, the functional subunit of the epigenetic master regulator polycomb repressive complex.

Human proximal long head of biceps tendons from patients with different chronic shoulder pathologies (n = 22), and controls from patients with humerus fracture (n = 6) and pathology (n = 4), were histologically scored for degeneration and analyzed for gene and protein expression of tendon specific factors, senescence markers, and EZH2. Tissues were further exposed to senotherapeutic compounds and the USA Food and Drugs Administration-approved selective EZH2 inhibitor EPZ-6438 and their senescence-associated secretory phenotype (SASP) assessed.

Expression of senescence markers (CDKN2A/p16, CDKN2D/p19) and EZH2 was significantly higher in tendinopathies compared to fracture or healthy tissue controls and positively correlated with the degree of tissue degeneration. Immunofluorescent stainings demonstrated colocalization of p16 and p19 with EZH2 in tenocytes. Treatment of tendon biopsies with EPZ-6438 reduced secretion of a panel of SASP factors, including interleukin-6 (IL6), IL8, matrix metalloproteinase-3 (MMP3) or GRO1, similarly to the senotherapeutic compound AG490.

We demonstrate that senescence traits accumulate in pathological tendon tissues and positively correlate with tissue degeneration. Increased expression of CDKN2A/p16 and CDKN2D/p19 coincides with EZH2 expression, while its inhibition decreased the secretion of SASP factors, indicating a possible regulatory role of EZH2 in tenocyte senescence in tendinopathies. Reduction of cellular senescence, e.g. with EPZ-6438, opens ways to new potential therapeutic approaches for enhancing regeneration in chronic tendinopathies.

Tendinopathy is a condition characterized by persistent tendon pain, structural damage, and compromised functionality. Presently, the treatment for tendinopathy remains a formidable challenge, partly because of its unclear pathogenesis. Tendon stem/progenitor cells (TSPCs) are essential for tendon homeostasis, regeneration, remodeling, and repair. An innovative theory has been previously proposed, with insufficient evidence, that the erroneous differentiation of TSPCs may constitute one of the fundamental mechanisms underpinning tendinopathy. Over the past few years, there has been accumulating evidence for plausibility of this theory. In this review, we delve into alterations in the differentiation potential of TSPCs and the underlying mechanisms in the context of injury-induced tendinopathy, diabetic tendinopathy, and age-related tendinopathy to provide updated evidence on the erroneous differentiation theory. Despite certain limitations inherent in the existing body of evidence, the erroneous differentiation theory emerges as a promising and highly pertinent avenue for understanding tendinopathy. In the future, advanced methodologies will be harnessed to further deepen comprehension of this theory, paving the way for prospective developments in clinical therapies targeting TSPCs for the management of tendinopathy.

The fluoroquinolone (FQ) class of antibiotics includes the world's most prescribed antibiotics such as ciprofloxacin, levofloxacin, and ofloxacin that are known for their low bacterial resistance. This is despite their potential to trigger severe side effects, such as myopathy, hearing loss, tendinopathy, and tendon rupture. Thus, healthcare organizations around the world have recommended limiting the prescription of FQs. Tendinopathy is a common name for maladies that cause pain and degeneration in the tendon tissue, which can result in tendon rupture. Whilst there are several identified effects of FQ on tendons, the exact molecular mechanisms behind FQ-mediated tendon rupture are unclear. Previous research studies indicated that FQ-mediated tendinopathy and tendon rupture can be induced by changes in gene expression, metabolism, and function of tendon resident cells, thus leading to alterations in the extracellular matrix. Hence, this review begins with an update on FQs, their mode of action, and their known side effects, as well as summary information on tendon tissue structure and cellular content. Next, how FQs affect the tendon tissue and trigger tendinopathy and tendon rupture is explored in detail. Lastly, possible preventative measures and promising areas for future research are also discussed. Specifically, follow-up studies should focus on understanding the FQ-mediated tendon changes in a more complex manner and integrating in vitro with in vivo models. With respect to in vitro systems, the field should move towards three-dimensional models that reflect the cellular diversity found in the tissue.

Although older adult patients have a higher retear rate after rotator cuff (RC) repair, the influence of aging on the healing process remains unclear. During mouse enthesis development, a multipotent progenitor coexpressing scleraxis (Scx) and SRY-box 9 (Sox9) contributes to enthesis formation. Scx+/Sox9+ cells may act as enthesis progenitors even during postnatal healing, and their number decreases with maturation. However, the pathophysiology of decreased RC healing ability due to aging remains unclear. We aimed to evaluate the effects of aging on tendon-to-bone healing after surgical RC repair in rats through biomechanical and histological analyses of Scx+ and Sox9+ progenitors in a ScxGFP transgenic rat model.

This was a controlled laboratory study. Adult Sprague-Dawley rats (n = 111) underwent unilateral surgery for supraspinatus tendon repair immediately after transection. The rats were divided into aged (95 week old) and control (12 week old) groups. The effects of aging were assessed using biomechanical tests at 6 weeks postoperatively and histologically at 2 and 6 weeks postoperatively. ScxGFP transgenic rats were used for the immunohistochemical assessment of Scx- and Sox9-expressing progenitors during the repair process. Sox9, Scx, and tenomodulin expression was assessed using real-time reverse transcription polymerase chain reaction.

In the biomechanical test at 6 weeks postoperatively, the aged group had lower ultimate load to failure (control: 20.4 ± 6.1 N, aged: 14.9 ± 6.6 N, P = .007), stiffness (control: 16.1 ± 6.2 N/mm, aged: 12.6 ± 5.5 N/mm, P = .023), and ultimate stress to failure (control: 6.0 ± 3.4 N/mm2, aged: 3.4 ± 2.5 N/mm2, P < .001) than the control group. The total histological score of the aged group was lower than that of the control group at 6 weeks postoperatively (control: 8.8 ± 0.8, aged: 5.8 ± 0.4, P = .029). Immunohistochemistry tests showed that the percentages of Sox9+ (control: 6.6 ± 1.1, aged: 2.0 ± 1.0, P = .029) and Scx+/Sox9+ (control: 3.6 ± 0.8, aged: 1.1 ± 0.6, P = .029) cells at the reparative site were lower in the aged group than in the control group at 2 weeks postoperatively.

In a rat RC tendon-to-bone healing model, the decreased recruitment of Scx+/Sox9+ enthesis-related progenitor cells in the early phase may be associated with delayed reparative tissue remodeling in aging animals. These findings encourage the development of therapeutic strategies that biologically promote healing and reduce postoperative retears in older adult patients.

Musculoskeletal injuries, including tendinopathies, present a significant clinical burden for aging populations. While the biological drivers of age-related declines in tendon function are poorly understood, it is well accepted that dysregulation of extracellular matrix (ECM) remodeling plays a role in chronic tendon degeneration. Senescent cells, which have been associated with multiple degenerative pathologies in musculoskeletal tissues, secrete a highly pro-inflammatory senescence-associated secretory phenotype (SASP) that has potential to promote ECM breakdown. However, the role of senescent cells in the dysregulation of tendon ECM homeostasis is largely unknown. To assess this directly, we developed an in vitro model of induced cellular senescence in murine tendon explants. This novel technique enables us to study the isolated interactions of senescent cells and their native ECM without interference from age-related systemic changes. We document multiple biomarkers of cellular senescence in induced tendon explants including cell cycle arrest, apoptosis resistance, and sustained inflammatory responses. We then utilize this in vitro senescence model to compare the ECM remodeling response of young, naturally aged, and induced-senescent tendons to an altered mechanical stimulus. We found that both senescence and aging independently led to alterations in ECM-related gene expression, reductions in protein synthesis, and tissue compositional changes. Furthermore, MMP activity was sustained, thus shifting the remodeling balance of aged and induced-senescent tissues towards degradation over production. Together, this demonstrates that cellular senescence plays a role in the altered mechano-response of aged tendons and likely contributes to poor clinical outcomes in aging populations.

Rotator cuff tears (RCT) are the most common cause of shoulder pain among adults. "Rotator cuff" refers to the four muscles that cover the shoulder joint: supraspinatus, infraspinatus, subscapularis, and teres minor. These muscles help maintain the rotational movement and stability of the shoulder joint. RCT is a condition in which one or more of these four muscles become ruptured or damaged, causing pain in the arms and shoulders. RCT results from degenerative changes caused by chronic inflammation of the tendons and consequent tendon tissue defects. This phenomenon occurs because of the exhaustion of endogenous tendon stem cells. Tendon regeneration requires rejuvenation of these endogenous tendon stem/progenitor cells (TSPCs) prior to their growth phase. TSPCs exhibit clonogenicity, multipotency, and self-renewal properties; they express classical stem cell markers and genes associated with the tendon lineage. However, specific markers for TSPC are yet to be identified. In this review, we introduce novel TSPC markers and discuss various strategies for TSPC reprogramming. With further research, TSPC reprogramming technology could be adapted to treat age-related degenerative diseases, providing a new strategy for regenerative medicine.

Dysregulated adipokine production is an influencing factor for the homeostatic imbalance of tendons. High levels of serum leptin may be a potential link between increasing adiposity and tendinopathy, while the detailed mechanistic explanation was not well-defined. In this study, we investigated the regulatory role of leptin in the tendon stem/progenitor cells (TSPCs) and the molecular mechanism within, and determined the effect of high levels of leptin on tendon recovery. We demonstrated that leptin reduced the viability of isolated rat TSPCs in a dose-dependent way, accompanied with increased transdifferentiation and altered gene expression of a series of extracellular matrix (ECM) enzymatic modulators. Also, we found that leptin could dose-dependently promote TSPCs senescence, while exhibiting limited effect in apoptotic or autophagic induction. Mechanistic study evidenced that leptin treatment increased the AKT/mTOR signaling activity and elevated the expression of leptin receptor (LEPR) in TSPCs, without marked change in MAPK or STAT5 activation. Further, we confirmed that rapamycin treatment, but not AKT inhibition, effectively reduced the leptin-promoted TSPCs senescence. In a rat model with Achilles wounding, exposure to leptin profoundly delayed tendon healing, which was effectively rescued with rapamycin treatment. Our results suggested that leptin could cause intrinsic cellular deficits in TSPCs and impede tendon repair through the AKT/mTOR signaling pathway. These findings evidenced for an important role of elevated leptin levels in the care of tendinopathy and tendon tears.

The rate of retear after surgical repair remains high. Mesenchymal stem cells (MSCs) have been extensively employed in regenerative medicine for several decades. However, safety and ethical concerns constrain their clinical application. Tendon Stem/Progenitor Cells (TSPCs)-derived exosomes have emerged as promising cell-free therapeutic agents. Therefore, urgent studies are needed to investigate whether TSPC-Exos could enhance tendon-bone healing and elucidate the underlying mechanisms. In this study, TSPC-Exos were found to promote the proliferation, migration, and expression of fibrogenesis markers in BMSCs. Furthermore, TSPC-Exos demonstrated an ability to suppress the polarization of M1 macrophages while promoting M2 macrophage polarization. In a rat model of rotator cuff repair, TSPC-Exos modulated inflammation and improved the histological structure of the tendon-bone interface, the biomechanical properties of the repaired tendon, and the function of the joint. Mechanistically, TSPC-Exos exhibited high expression of miR-21a-5p, which regulated the expression of PDCD4. The PDCD4/AKT/mTOR axis was implicated in the therapeutic effects of TSPC-Exos on proliferation, migration, and fibrogenesis in BMSCs. This study introduces a novel approach utilizing TSPC-Exos therapy as a promising strategy for cell-free therapies, potentially benefiting patients with rotator cuff tear in the future.

Rotator cuff tear (RCT) is the leading cause of shoulder pain, primarily associated with age-related tendon degeneration. This study aimed to elucidate the potential differential gene expressions in tendons across different age groups, and to investigate their roles in tendon degeneration.

Linear regression and differential expression (DE) analyses were performed on two transcriptome profiling datasets of torn supraspinatus tendons to identify age-related genes. Subsequent functional analyses were conducted on these candidate genes to explore their potential roles in tendon ageing. Additionally, a secondary DE analysis was performed on candidate genes by comparing their expressions between lesioned and normal tendons to explore their correlations with RCTs.

We identified 49 genes in torn supraspinatus tendons associated with advancing age. Among them, five age-related genes showed DE in lesioned tendons compared to normal tendons. Functional analyses and previous studies have highlighted their specific enrichments in biological functions, such as muscle development (e.g. myosin heavy chain 3 (MYH3)), transcription regulation (e.g. CCAAT enhancer binding brotein delta (CEBPD)), and metal ion homeostasis (e.g. metallothionein 1X (MT1X)).

This study uncovered molecular aspects of tendon ageing and their potential links to RCT development, offering insights for targeted interventions. These findings enhance our understanding of the mechanisms of tendon degeneration, allowing potential strategies to be made for reducing the incidence of RCT.

Tendon defect repair remains a tough clinical procedure that hinders functional motion in patients. Electrohydrodynamic (EHD) three-dimensional (3D) printing, as a novel strategy, can controllably fabricate biomimetic micro/nanoscale architecture, but the hydrophobic and bioinert nature of polymers might be adverse to cell-material interplay. In this work, 3D EHD printed polycaprolactone (PCL) was immobilized on basic fibroblast growth factor (bFGF) using polydopamine (PDA), and the proliferation and tenogenic differentiation of tendon stem/progenitor cells (TSPCs) in vitro was researched. A subcutaneous model was established to evaluate the effects of tenogenesis and immunomodulation. We then investigated the in situ implantation and immunomodulation effects in an Achilles tendon defect model. After immobilization of bFGF, the scaffolds profoundly facilitated proliferation and tenogenic differentiation; however, PDA had only a proliferative effect. Intriguingly, the bFGF immobilized on EHD printed PCL indicated a synergistic effect on the highest expression of tenogenic gene and protein markers at 14 days, and the tenogenesis may be induced by activating the transforming growth factor-β (TGF-β) signal pathway in vitro. The subcutaneous engraftment study confirmed a tendon-like structure, similar to that of the native tendon, as well as an M2 macrophage polarization effect. Additionally, the bioactive scaffold exhibited superior efficacy in new collagen formation and repair of Achilles tendon defects. Our study revealed that the topographic cues alone were insufficient to trigger tenogenic differentiation, requiring appropriate chemical signals, and that appropriate immunomodulation was conducive to tenogenesis. The tenogenesis of TSPCs on the bioactive scaffold may be correlated with the TGF-β signal pathway and M2 macrophage polarization.

Energy storing tendons such as the human Achilles and equine superficial digital flexor tendon (SDFT) are prone to injury, with incidence increasing with aging, peaking in the 5th decade of life in the human Achilles tendon. The interfascicular matrix (IFM), which binds tendon fascicles, plays a key role in energy storing tendon mechanics, and aging alterations to the IFM negatively impact tendon function. While the mechanical role of the IFM in tendon function is well-established, the biological role of IFM-resident cell populations remains to be elucidated. Therefore, the aim of this study was to identify IFM-resident cell populations and establish how these populations are affected by aging. Cells from young and old SDFTs were subjected to single cell RNA-sequencing, and immunolabelling for markers of each resulting population used to localise cell clusters. Eleven cell clusters were identified, including tenocytes, endothelial cells, mural cells, and immune cells. One tenocyte cluster localised to the fascicular matrix, whereas nine clusters localised to the IFM. Interfascicular tenocytes and mural cells were preferentially affected by aging, with differential expression of genes related to senescence, dysregulated proteostasis and inflammation. This is the first study to establish heterogeneity in IFM cell populations, and to identify age-related alterations specific to IFM-localised cells.

Adult tendon stem/progenitor cells (TSPCs) are essential for tendon maintenance, regeneration, and repair, yet they become susceptible to senescence with age, impairing the self-healing capacity of tendons. In this study, we employ a recently developed deep-learning-based efficacy prediction system to screen potential stemness-promoting and senescence-inhibiting drugs from natural products using the transcriptional signatures of stemness. The top-ranked candidate, prim-O-glucosylcimifugin (POG), a saposhnikovia root extract, could ameliorate TPSC senescent phenotypes caused by long-term passage and natural aging in rats and humans, as well as restore the self-renewal and proliferative capacities and tenogenic potential of aged TSPCs. In vivo, the systematic administration of POG or the local delivery of POG nanoparticles functionally rescued endogenous tendon regeneration and repair in aged rats to levels similar to those of normal animals. Mechanistically, POG protects TSPCs against functional impairment during both passage-induced and natural aging by simultaneously suppressing nuclear factor-κB and decreasing mTOR signaling with the induction of autophagy. Thus, the strategy of pharmacological intervention with the deep learning-predicted compound POG could rejuvenate aged TSPCs and improve the regenerative capacity of aged tendons.

Tendon aging is associated with an increasing prevalence of tendon injuries and/or chronic tendon diseases, such as tendinopathy, which affects approximately 25% of the adult population. Aged tendons are often characterized by a reduction in the number and functionality of tendon stem/progenitor cells (TSPCs), fragmented or disorganized collagen bundles, and an increased deposition of glycosaminoglycans (GAGs), leading to pain, inflammation, and impaired mobility. Although the exact pathology is unknown, overuse and microtrauma from aging are thought to be major causative factors. Due to the hypovascular and hypocellular nature of the tendon microenvironment, healing of aged tendons and related injuries is difficult using current pain/inflammation and surgical management techniques. Therefore, there is a need for novel therapies, specifically cellular therapy such as cell rejuvenation, due to the decreased regenerative capacity during aging. To augment the therapeutic strategies for treating tendon-aging-associated diseases and injuries, a comprehensive understanding of tendon aging pathology is needed. This review summarizes age-related tendon changes, including cell behaviors, extracellular matrix (ECM) composition, biomechanical properties and healing capacity. Additionally, the impact of conventional treatments (diet, exercise, and surgery) is discussed, and recent advanced strategies (cell rejuvenation) are highlighted to address aged tendon healing. This review underscores the molecular and cellular linkages between aged tendon biomechanical properties and the healing response, and provides an overview of current and novel strategies for treating aged tendons. Understanding the underlying rationale for future basic and translational studies of tendon aging is crucial to the development of advanced therapeutics for tendon regeneration.

Tendon injury is one of the prevalent disorders of the musculoskeletal system in orthopedics and is characterized by pain and limitation of joint function. Due to the difficulty of spontaneous tendon healing, and the scar tissue and low mechanical properties that usually develops after healing. Therefore, the healing of tendon injury remains a clinical challenge. Although there are a multitude of approaches to treating tendon injury, the therapeutic effects have not been satisfactory to date. Recent studies have shown that stem cell therapy has a facilitative effect on tendon healing. In particular, tendon stem/progenitor cells (TSPCs), a type of stem cell from tendon tissue, play an important role not only in tendon development and tendon homeostasis, but also in tendon healing. Compared to other stem cells, TSPCs have the potential to spontaneously differentiate into tenocytes and express higher levels of tendon-related genes. TSPCs promote tendon healing by three mechanisms: modulating the inflammatory response, promoting tenocyte proliferation, and accelerating collagen production and balancing extracellular matrix remodeling. However, current investigations have shown that TSPCs also have a negative effect on tendon healing. For example, misdifferentiation of TSPCs leads to a "failed healing response," which in turn leads to the development of chronic tendon injury (tendinopathy). The focus of this paper is to describe the characteristics of TSPCs and tenocytes, to demonstrate the roles of TSPCs in tendon healing, while discussing the approaches used to culture and differentiate TSPCs. In addition, the limitations of TSPCs in clinical application and their potential therapeutic strategies are elucidated.

Tendinopathy is a medical condition that includes a spectrum of inflammatory and degenerative tendon changes caused by traumatic or overuse injuries. The pathological mechanism of tendinopathy has not been well defined, and no ideal treatment is currently available. Platelet-rich plasma (PRP) is an autologous whole blood derivative containing a variety of cytokines and other protein components. Various basic studies have found that PRP has the therapeutic potential to promote cell proliferation and differentiation, regulate angiogenesis, increase extracellular matrix synthesis, and modulate inflammation in degenerative tendons. Therefore, PRP has been widely used as a promising therapeutic agent for tendinopathy. However, controversies exist over the optimal treatment regimen and efficacy of PRP for tendinopathy. This review focuses on the specific molecular and cellular mechanisms by which PRP manipulates tendon healing to better understand how PRP affects tendinopathy and explore the reason for the differences in clinical trial outcomes. This article has also pointed out the future direction of basic research and clinical application of PRP in the treatment of tendinopathy, which will play a guiding role in the design of PRP treatment protocols for tendinopathy.

Age-related tendon disorders are closely linked with tendon stem/progenitor cell (TSPC) senescence. However, the underlying mechanisms of TSPC senescence and promising therapeutic strategies for rejuvenation of TSPC senescence remain unclear. In this study, the senescent state of TSPCs increased with age. It was also verified that the AMPK inhibition/mTOR activation is correlated with the senescent state of TSPCs. Furthermore, a low dose of metformin mitigated TSPC senescence and restored senescence-related functions, including proliferation, colony-forming ability, migration ability and tenogenic differentiation ability at the early stage of aging. The protective effects of metformin on TSPCs were regulated through the AMPK/mTOR axis. An in vivo study showed that metformin treatment postpones tendon aging and enhances AMPK phosphorylation but reduces mTOR phosphorylation in a natural aging rat model. Our study revealed new insight and mechanistic exploration of TSPC senescence and proposed a novel therapeutic treatment for age-related tendon disorders by targeting the AMPK/mTOR axis at the early stage of aging.

Tendon wounds are a worldwide health issue affecting millions of people annually. Due to the characteristics of tendons, their natural restoration is a complicated and lengthy process. With the advancement of bioengineering, biomaterials, and cell biology, a new science, tissue engineering, has developed. In this field, numerous ways have been offered. As increasingly intricate and natural structures resembling tendons are produced, the results are encouraging. This study highlights the nature of the tendon and the standard cures that have thus far been utilized. Then, a comparison is made between the many tendon tissue engineering methodologies proposed to date, concentrating on the ingredients required to gain the structures that enable appropriate tendon renewal: cells, growth factors, scaffolds, and scaffold formation methods. The analysis of all these factors enables a global understanding of the impact of each component employed in tendon restoration, thereby shedding light on potential future approaches involving the creation of novel combinations of materials, cells, designs, and bioactive molecules for the restoration of a functional tendon.

This study aimed to characterize aging-induced tendinopathy in mouse Achilles tendon and also to assess the treatment effects of metformin (Met) on aging tendon. We showed that compared to young tendon, aging tendon was in an inflammatory and senescent state as shown by increased expression of inflammatory disulfide HMGB1 (dsHMGB1), inflammatory macrophage marker CD68, and senescent cell markers SA-β-gal, p53, and p16. Moreover, aging tendon was degenerated marked by accumulation of proteoglycans and lipids in its interior. However, treatment of aging tendon by intraperitoneal (IP) injection of Met, a specific inhibitor of HMGB1, reduced dsHMGB1 levels, decreased the expression of CD68, SA-β-gal, CCN1, and p16 in vitro and in vivo. Furthermore, Met treatment also increased the number of NS, SSEA-1, and CD73 positive stem cells in culture and improved the tendon structure in aging mouse. These findings of this study indicate that Met exerts anti-inflammatory and anti-senescent effects on aging tendon.

Degenerative tendinopathy, a condition causing movement restriction due to high pain, highly impacts productivity and quality of life. The healing process is a complex phenomenon and involves a series of intra-cellular and inter-cellular processes. Proliferation and differentiation of the tenocyte is a major and essential process to heal degenerative tendinopathy. The recent development in microRNA (miRNA)-mediated reprogramming of the cellular function through specific pathways opened door for the development of new regenerative therapeutics. Based on information about gene expression and regulation of tendon injury and healing, we attempted to evaluate the combinatorial effect of selected miRNAs for better healing of degenerative tendinopathy.

The present study was designed to evaluate the combinatorial effect of two miRNAs (has-miR-140 and has-miR-135) in the healing process of the tendon. Publicly available information/data were retrieved from appropriate platforms such as PubMed. Only molecular data, directly associated with tendinopathies, including genes/proteins and miRNAs, were used in this study. The miRNAs involved in tendinopathy were analyzed by a Bioinformatics tools (e.g., TargetScan, miRDB, and the RNA22v2). Interactive involvement of the miRNAs with key proteins involved in tendinopathy was predicted by the Insilco approach.

Based on information available in the public domain, tendon healing-associated miRNAs were predicted to explore their therapeutic potentials. Based on computation analysis, focusing on the potential regulatory effect on tendon healing, the miR-135 and miR-140 were selected for this study. These miRNAs were found as key players in tendon healing through Rho-associated coiled-coil containing protein kinase 1 (ROCK1), IGF-1/PI3K/Akt, PIN, and Wnt signaling pathways. It was also predicted that these miRNAs may reprogram the cells to induce proliferation and differentiation activity. Many miRNAs are likely to regulate genes important for the tendinopathy healing process, and the result of this study allows an approach for miRNA-mediated regeneration of the tenocyte for tendon healing. Based on computational analysis, the role of these miRNAs in different pathways was established, and the results provided insights into the combinatorial approach of miRNA-mediated cell reprogramming.

In this study, the association between miRNAs and the disease was evaluated to correlate the tendinopathy genes and the relevant role of different miRNAs in their regulation. Through this study, it was established that the synergistic effect of more than one miRNA on directed reprogramming of the cell could be helpful in the regeneration of damaged tissue. It is anticipated that this study will be helpful for the design of miRNA cocktails for the orchestration of cellular reprogramming events.

Tendinopathy, the most common form of chronic tendon disorder, leads to persistent tendon pain and loss of function. Profiling the heterogeneous cellular composition in the tendon microenvironment helps to elucidate rational molecular mechanisms of tendinopathy.

In this study, through a multi-modal analysis, a single-cell RNA- and ATAC-seq integrated tendinopathy landscape was generated for the first time. We found that a specific cell subpopulation with low PRDX2 expression exhibited a higher level of inflammation, lower proliferation and migration ability, which not only promoted tendon injury but also led to microenvironment deterioration. Mechanistically, a motif enrichment analysis of chromatin accessibility showed that FOXO1 was an upstream regulator of PRDX2 transcription, and we confirmed that functional blockade of FOXO1 activity induced PRDX2 silencing. The TNF signaling pathway was significantly activated in the PRDX2-low group, and TNF inhibition effectively restored diseased cell degradation.

We revealed an essential role of diseased cells in tendinopathy and proposed the FOXO1-PRDX2-TNF axis is a potential regulatory mechanism for the treatment of tendinopathy.

Age-related tendon disorder, a primary motor system disease, is characterized by biological changes in the tendon tissue due to senescence and seriously affects the quality of life of the elderly. The pathogenesis of this disease is not well-understood. Tendon stem/progenitor cells (TSPCs) exhibit multi-differentiation capacity. These cells are important cellular components of the tendon because of their roles in tendon tissue homeostasis, remodeling, and repair. Previous studies revealed alterations in the biological characteristics and tenogenic differentiation potential of TSPCs in senescent tendon tissue, in turn contributing to insufficient differentiation of TSPCs into tenocytes. Poor tendon repair can result in age-related tendinopathies. Therefore, targeting of senescent TSPCs may restore the tenogenic differentiation potential of these cells and achieve homeostasis of the tendon tissue to prevent or treat age-related tendinopathy. In this review, we summarize the biological characteristics of TSPCs and histopathological changes in age-related tendinopathy, as well as the potential mechanisms through which TSPCs contribute to senescence. This information may promote further exploration of innovative treatment strategies to rescue TSPCs from senescence.

Trigger thumb is a pathologic condition of the digital pulleys and flexor tendons. To find a cutoff value of the cross-sectional area ratio of specific parts of the flexor pollicis longus tendon to diagnosis trigger thumb in the high-frequency ultrasound examination.

We evaluated 271 healthy volunteers and 57 patients with clinical diagnosis of trigger thumb. The cross-sectional area of the metacarpophalangeal joint of flexor pollicis longus tendon (C1) and the cross-sectional area of the midpoint of the first metacarpal of flexor pollicis longus tendon (C2) were analyzed.

There is no difference between gender, age and left and right hands in the ratio of C1 to C2 (C1/ C2). The mean of C1/ C2 in the healthy thumb was 0.983 ± 0.103, which was significantly smaller in comparison to the diseased thumb (P < 0.05). Based on the receiver operating characteristic curve, we chose the diagnostic cut-off value for the C1/ C2 to be 1.362 and 1.153 in order to differ a trigger thumb from children and adults.

The C1/ C2 of the healthy thumb was relatively stable, with a mean value of 0.983 ± 0.103. The cutoff value of C1/C2 to distinguish healthy thumb from diseased thumb in children and adults were 1.362 and 1.153, respectively.

Functional repair of tendons remains a challenge to be overcome for both clinicians and scientists. We have previously reported a three-dimensional RADA peptide hydrogel that provides a suitable microenvironment for human tendon stem/progenitor cells (TSPCs) survival and tenogenesis. In this study, we explore the potential of in vivo patellar tendon repair by human TSPC-laden RADA hydrogel in rats, which were sacrificed at 4 and 8 weeks after operation. Hind limb function test, macroscopical and histological examination, tendon cell amount and alignment analysis, and radiographic assessments were performed at several time points. Our results demonstrated that human TSPC-laden RADA hydrogel (RADA+TSPC group) boosted in vivo patellar tendon repair with better ambulatory function recovery compared with the control groups, in which tendon defects were untreated (Defect group) or treated with RADA hydrogel alone (RADA group). In addition, better macroscopic appearance and improved matrix organization in the repaired tendon with less cell amount and reduced adipocyte accumulation and blood vessel formation were observed in the RADA+TSPC group. Moreover, tendon defect treated with TSPC-laden RADA hydrogel resulted in diminished heterotopic ossification (HO) at 8 weeks postoperation, which was indicated by both X-ray examination and micro-computed tomography scan. Taken together, the combination of TSPC and nanofiber hydrogel provide an optimistic alternative method to accelerate functional tendon repair with reduced HO. Impact statement Our study clearly demonstrates the combination of tendon stem/progenitor cell and nanofiber hydrogel provide a new and optimistic tissue engineering strategy to treat tendon injury by accelerating functional tendon repair with reduced heterotopic ossification. The clinical translation is also very promising, which can provide a minimally invasive, nonsurgical, or complementary treatment methods to treat human tendon injury.

The regenerative capabilities including self-renewal, migration and differentiation potentials shift from the embryonic phase to the mature period of endogenous tendon stem/progenitor cells (TSPCs) characterize restricted functions and disabilities following tendon injuries. Recent studies have shown that tendon regeneration and repair rely on multiple specific transcription factors to maintain TSPCs characteristics and functions. Here, we demonstrate Yap, a Hippo pathway downstream effector, is associated with TSPCs phenotype and regenerative potentials through gene expression analysis of tendon development and repair process. Exosomes have been proven an efficient transport platform for drug delivery. In this study, purified exosomes derived from donor platelets are loaded with recombinant Yap1 protein (PLT-Exo-Yap1) via electroporation to promote the stemness and differentiation potentials of TSPCs in vitro. Programmed TSPCs with Yap1 import maintain stemness and functions after long-term passage in vitro. The increased oxidative stress levels of TSPCs are related to the phenotype changes in duplicative senescent processes. The results show that treatment with PLT-Exo-Yap1 significantly protects TSPCs against oxidative stressor-induced stemness loss and senescence-associated secretory phenotype (SASP) through the NF-κB signaling pathway. In addition, we fabricate an Exos-Yap1-functioned GelMA hydrogel with a parallel-aligned substrate structure to enhance TSPCs adhesion, promote cell stemness and force regenerative cells toward the tendon lineage for in vitro and in vivo tendon regeneration. The application of Exos-Yap1 functioned implant assists new tendon-like tissue formation with good mechanical properties and locomotor functions in a full-cut Achilles tendon defect model. Thus, PLT-Exo-Yap1-functionalized GelMA promotes the rejuvenation of TSPCs to facilitate functional tendon regeneration. STATEMENT OF SIGNIFICANCE: This is the first study to explore that the hippo pathway downstream effector Yap is involved in tendon aging and repair processes, and is associated with the regenerative capabilities of TSPCs. In this syudy, Platelet-derived exosomes (PLT-Exos) act as an appropriate carrier platform for the delivery of recombinant Yap1 into TSPCs to regulate Yap activity. Effective Yap1 delivery inhibit oxidative stress-induced senescence associated phenotype of TSPCs by blocking ROS-mediated NF-κb signaling pathway activation. This study emphasizes that combined application of biomimetic scaffolds and Yap1 loaded PLT-Exos can provide structural support and promote rejuvenation of resident cells to assist functional regeneration for Achilles tendon defect, and has the prospect of clinical setting.

Primary tenocytes rapidly undergo senescence and a phenotypic drift upon in vitro monolayer culture, which limits tendon research. The Ink4a/Arf locus encodes the proteins p16^Ink4a/Arf and p14^ARF (p19^ARF in mice) that regulate cell cycle progression and senescence. We here established an immortalized cell line using tenocytes isolated from Ink4a/Arf deficient mice (Ink4a/Arf^-/-). These cells were investigated at three distinct time points, at low (2-5), intermediate (14-17) and high (35-44) passages. Wild-type cells at low passage (2-5) served as controls. Ink4a/Arf^-/- tenocytes at all stages were comparable to wild-type cells regarding morphology, expression of tenogeneic genes (collagen type 1, 3 and 5, Scleraxis, Tenomodulin and Tenascin-C), and surface markers (CD29, CD44 and CD105) and form 3D tendon-like structures. Importantly, Ink4a/Arf^-/- tenocytes maintained their phenotypic features and proliferation potential in culture for more than 40 passages and also following freeze-thaw cycles. In contrast, wild-type tenocytes underwent senescence starting in passage 6. These data define Ink4a/Arf^-/- tenocytes as novel tool for in vitro tendon research and as valuable in vitro alternative to animal experiments.

Tendon injuries are some of the most commonly diagnosed musculoskeletal diseases. Tendon regeneration is sensitive to the topology of the substitute as it affects the cellular microenvironment and homeostasis. To bionic in vivo three-dimensional (3D) aligned microenvironment, an ordered 3D sandwich model was used to investigate the cell response in the tendon. First, high-resolution 3D printing provided parallel-grooved topographical cues on the hydrogel surface. Then the cells were seeded on its surface to acquire a 2D model. Afterward, an additional hydrogel coating layer was applied to the cells to create the 3D model. The interaction between cells and order structures in three-dimensions is yet to be explored. The study found that the tendon stem/progenitor cells (TSPCs) still maintain their ordering growth in the 3D model as in the 2D model. The study also found that the 3D-aligned TSPCs exhibited enhanced tenogenic differentiation through the PI3K-AKT signaling pathway and presented a less inflammatory phenotype than those in the 2D model. The in vivo implantation of such a 3D-aligned TSPC composite promoted tendon regeneration and mitigated heterotopic ossification in an Achilles defect model. These findings demonstrated that 3D-aligned TSPCs within a biomimetic topology environment are promising for functional tendon regeneration.

A crucial component of the musculoskeletal system, the tendon is one of the most commonly injured tissues in the body. In severe cases, the ruptured tendon leads to permanent dysfunction. Although many efforts have been devoted to seeking a safe and efficient treatment for enhancing tendon healing, currently existing treatments have not yet achieved a major clinical improvement. Here, an injectable granular hyaluronic acid (gHA)-hydrogel is engineered to deliver fibromodulin (FMOD)-a bioactive extracellular matrix (ECM) that enhances tenocyte mobility and optimizes the surrounding ECM assembly for tendon healing. The FMOD-releasing granular HA (FMOD/gHA)-hydrogel exhibits unique characteristics that are desired for both patients and health providers, such as permitting a microinvasive application and displaying a burst-to-sustained two-phase release of FMOD, which leads to a prompt FMOD delivery followed by a constant dose-maintaining period. Importantly, the generated FMOD-releasing granular HA hydrogel significantly augmented tendon-healing in a fully-ruptured rat's Achilles tendon model histologically, mechanically, and functionally. Particularly, the breaking strength of the wounded tendon and the gait performance of treated rats returns to the same normal level as the healthy controls. In summary, a novel effective FMOD/gHA-hydrogel is developed in response to the urgent demand for promoting tendon healing.

Aging is a complex and progressive process where the tissues of the body demonstrate a decreased ability to maintain homeostasis. During aging, there are substantial cellular and molecular changes, with a subsequent increase in susceptibility to pathological degeneration of normal tissue function. In tendon, aging results in well characterized alterations in extracellular matrix (ECM) structure and composition. In addition, the cellular environment of aged tendons is altered, including a marked decrease in cell density and metabolic activity, as well as an increase in cellular senescence. Collectively, these degenerative changes make aging a key risk factor for the development of tendinopathies and can increase the frequency of tendon injuries. However, inconsistencies in the extent of age-related degenerative impairments in tendons have been reported, likely due to differences in how "old" and "young" age-groups have been defined, differences between anatomically distinct tendons, and differences between animal models that have been utilized to study the impact of aging on tendon homeostasis. In this review, we address these issues by summarizing data by well-defined age categories (young adults, middle-aged, and aged) and from anatomically distinct tendon types. We then summarize in detail how aging affects tendon mechanics, structure, composition, and the cellular environment based on current data and underscore what is currently not known. Finally, we discuss gaps in the current understanding of tendon aging and propose key avenues for future research that can shed light on the specific mechanisms of tendon pathogenesis due to aging.

Tendons are mechanosensitive connective tissues responsible for the connection between muscles and bones by transmitting forces that allow the movement of the body, yet, with advancing age, tendons become more prone to degeneration followed by injuries. Tendon diseases are one of the main causes of incapacity worldwide, leading to changes in tendon composition, structure, and biomechanical properties, as well as a decline in regenerative potential. There is still a great lack of knowledge regarding tendon cellular and molecular biology, interplay between biochemistry and biomechanics, and the complex pathomechanisms involved in tendon diseases. Consequently, this reflects a huge need for basic and clinical research to better elucidate the nature of healthy tendon tissue and also tendon aging process and associated diseases. This chapter concisely describes the effects that the aging process has on tendons at the tissue, cellular, and molecular levels and briefly reviews potential biological predictors of tendon aging. Recent research findings that are herein reviewed and discussed might contribute to the development of precision tendon therapies targeting the elderly population.

Cell-laden hydrogels used in tissue engineering generally lack sufficient 3D topographical guidance for cells to mature into aligned tissues. A new strategy called filamented light (FLight) biofabrication rapidly creates hydrogels composed of unidirectional microfilament networks, with diameters on the length scale of single cells. Due to optical modulation instability, a light beam is divided optically into FLight beams. Local polymerization of a photoactive resin is triggered, leading to local increase in refractive index, which itself creates self-focusing waveguides and further polymerization of photoresin into long hydrogel microfilaments. Diameter and spacing of the microfilaments can be tuned from 2 to 30 µm by changing the coherence length of the light beam. Microfilaments show outstanding cell instructive properties with fibroblasts, tenocytes, endothelial cells, and myoblasts, influencing cell alignment, nuclear deformation, and extracellular matrix deposition. FLight is compatible with multiple types of photoresins and allows for biofabrication of centimeter-scale hydrogel constructs with excellent cell viability within seconds (<10 s per construct). Multidirectional microfilaments are achievable within a single hydrogel construct by changing the direction of FLight projection, and complex multimaterial/multicellular tissue-engineered constructs are possible by sequentially exchanging the cell-laden photoresin. FLight offers a transformational approach to developing anisotropic tissues using photo-crosslinkable biomaterials.

Tendon is a bradytrophic and hypovascular tissue, hence, healing remains a major challenge. The molecular key events involved in successful repair have to be unravelled to develop novel strategies that reduce the risk of unfavourable outcomes such as non-healing, adhesion formation, and scarring. This review will consider the diverse pathophysiological features of tendon-derived cells that lead to failed healing, including misrouted differentiation (e.g. de- or transdifferentiation) and premature cell senescence, as well as the loss of functional progenitors. Many of these features can be attributed to disturbed cell-extracellular matrix (ECM) or unbalanced soluble mediators involving not only resident tendon cells, but also the cross-talk with immigrating immune cell populations. Unrestrained post-traumatic inflammation could hinder successful healing. Pro-angiogenic mediators trigger hypervascularization and lead to persistence of an immature repair tissue, which does not provide sufficient mechano-competence. Tendon repair tissue needs to achieve an ECM composition, structure, strength, and stiffness that resembles the undamaged highly hierarchically ordered tendon ECM. Adequate mechano-sensation and -transduction by tendon cells orchestrate ECM synthesis, stabilization by cross-linking, and remodelling as a prerequisite for the adaptation to the increased mechanical challenges during healing. Lastly, this review will discuss, from the cell biological point of view, possible optimization strategies for augmenting Achilles tendon (AT) healing outcomes, including adapted mechanostimulation and novel approaches by restraining neoangiogenesis, modifying stem cell niche parameters, tissue engineering, the modulation of the inflammatory cells, and the application of stimulatory factors.Cite this article: Bone Joint Res 2022;11(8):561-574.

Embryonic tendon cells have been studied in vitro to better understand mechanisms of tendon development. Outcomes of in vitro cell culture studies are easily affected by phenotype instability of embryonic tendon cells during expansion in vitro to achieve desired cell numbers, yet this has not been characterized. In the present study, we characterized phenotype stability, expansion potential, and onset of senescence in chick embryo tendon cells from low to high cell doublings. We focused on cells of Hamburger-Hamilton stages (HH) 40 and HH42, where HH40 is the earliest stage associated with substantial increases in extracellular matrix and mechanical properties during embryonic tendon development. HH40 and HH42 cells both downregulated expression levels of tendon phenotype markers, scleraxis and tenomodulin, and exhibited onset of senescence, based on p16 and p21 expression levels, cell surface area, and percentage of β-galactosidase positive cells, before significant decreases in proliferation rates were detected. These findings showed that embryonic tendon cells destabilize phenotype and become senescent earlier than they begin to decline in proliferation rates in vitro. Additionally, embryonic stage of isolation appears to have no effect on proliferation rates, whereas later stage HH42 cells downregulate phenotype and become susceptible to senescence sooner than earlier stage HH40 cells. Based on our data, we recommend chick embryo tendon cells be used before a maximum cumulative doubling level of 12 (passage 4 in this study) to avoid phenotype destabilization and onset of senescence.