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To examine the differential mechanobiological responses of specific resident tendon cells, we developed an in vivo model of whole-body irradiation followed by injection of either tendon stem/progenitor cells (TSCs) expressing green fluorescent protein (GFP-TSCs) or mature tenocytes expressing GFP (GFP-TNCs) into the patellar tendons of wild type C57 mice. Injected mice were subjected to short term (3 weeks) treadmill running, specifically moderate treadmill running (MTR) and intensive treadmill running (ITR). In MTR mice, both GFP-TSC and GFP-TNC injected tendons maintained normal cell morphology with elevated expression of tendon related markers collagen I and tenomodulin. In ITR mice injected with GFP-TNCs, cells also maintained an elongated shape similar to the shape found in normal/untreated control mice, as well as elevated expression of tendon related markers. However, ITR mice injected with GFP-TSCs showed abnormal changes, such as cell morphology transitioning to a round shape, elevated chondrogenic differentiation, and increased gene expression of non-tenocyte related genes LPL, Runx-2, and SOX-9. Increased gene expression data was supported by immunostaining showing elevated expression of SOX-9, Runx-2, and PPARγ. This study provides evidence that while MTR maintains tendon homeostasis by promoting the differentiation of TSCs into TNCs, ITR causes the onset of tendinopathy development by inducing non-tenocyte differentiation of TSCs, which may eventually lead to the formation of non-tendinous tissues in tendon tissue after long term mechanical overloading conditions on the tendon., Competing Interests: The authors have declared that no competing interests exist.

Tendon stem/progenitor cells (TSCs) are tendon-specific adult stem cells, which play crucial roles in tendon homeostasis, repair or regeneration once tendons are injured. Additionally, their pathological role in the development of tendinopathy in response to excessive mechanical loading placed on the tendon is also implicated. Similar to other adult stem cells, TSCs also exhibit universal characteristics of stem cells including colony formation in culture, self-renewal and multidifferentiation potential. Nevertheless, once TSCs are isolated from tendinous tissues and cultured in vitro, they may quickly lose stemness by undergoing differentiation. To maintain and prolong the stemness of TSCs in culture, we have developed two effective methods, namely culturing TSCs in hypoxic condition, or in growth media supplemented with low levels of PGE 2 . Here we present these methods in detail, along with the detailed description of the procedures to isolate TSCs from tendon samples and to culture them in vitro.

Treatment of tendon-bone junction injuries is a challenge because tendon-bone interface often heals poorly and the fibrocartilage zone, which reduces stress concentration, at the interface is not formed. In this study, we used a compound called kartogenin (KGN) with platelet-rich plasma (PRP) to induce the formation of fibrocartilage zone in a rat tendon graft-bone tunnel model. The experimental rats received KGN-PRP or PRP injections in the tendon graft-bone tunnel interface. The control group received saline. After 4, 8 and 12 weeks, Safranin O staining of the tendon graft-bone tunnels revealed abundant proteoglycans in the KGN-PRP group indicating the formation of cartilage-like transition zone. Immunohistochemical and immuno-fluorescence staining revealed collagen types I (Col-I) and II (Col-II) in the newly formed fibrocartilage zone. Both fibrocartilage zone formation and maturation were healing time dependent. In contrast, the PRP and saline control groups had no cartilage-like tissues and minimal Col-I and Col-II staining. Some gaps were also present in the saline control group. Finally, pull-out strength in the KGN-PRP-treated group at 8 weeks was 1.4-fold higher than the PRP-treated group and 1.6-fold higher than the saline control group. These findings indicate that KGN, with PRP as a carrier, promotes the formation of fibrocartilage zone between the tendon graft and bone interface. Thus, KGN-PRP may be used as a convenient cell-free therapy in clinics to promote fibrocartilage zone formation in rotator calf repair and anterior cruciate ligament reconstruction, thereby enhancing the mechanical strength of the tendon-bone interface and hence the clinical outcome of these procedures. Copyright © 2017 John Wiley & Sons, Ltd., (Copyright © 2017 John Wiley & Sons, Ltd.)

Introduction: Platelet-rich plasma (PRP) is widely used to treat tendon injuries in clinics. These PRP preparations often contain white blood cells or leukocytes, and the precise cellular effects of leukocyte-rich PRP (L-PRP) on tendons are not well defined. Therefore, in this study, we determined the effects of L-PRP on tendon stem/progenitor cells (TSCs), which play a key role in tendon homeostasis and repair., Methods: TSCs isolated from the patellar tendons of rabbits were treated with L-PRP or P-PRP (pure PRP without leukocytes) in vitro, followed by measuring cell proliferation, stem cell marker expression, inflammatory gene expression, and anabolic and catabolic protein expression by using immunostaining, quantitative real-time polymerase chain reaction, Western blot, and enzyme-linked immunosorbent assay, respectively., Results: Cell proliferation was induced by both L-PRP and P-PRP in a dose-dependent manner with maximum proliferation at a 10 % PRP dose. Both PRP treatments also induced differentiation of TSCs into active tenocytes. Nevertheless, the two types of PRP largely differed in several effects exerted on TSCs. L-PRP induced predominantly catabolic and inflammatory changes in differentiated tenocytes; its treatment increased the expression of catabolic marker genes, matrix metalloproteinase-1 (MMP-1), MMP-13, interleukin-1beta (IL-1β), IL-6 and tumor necrosis factor-alpha (TNF-α), and their respective protein expression and prostaglandin E2 (PGE 2) production. In contrast, P-PRP mainly induced anabolic changes; that is, P-PRP increased the gene expression of anabolic genes, alpha-smooth muscle actin (α-SMA), collagen types I and III., Conclusions: These findings indicate that, while both L-PRP and P-PRP appear to be "safe" in inducing TSC differentiation into active tenocytes, L-PRP may be detrimental to the healing of injured tendons because it induces catabolic and inflammatory effects on tendon cells and may prolong the effects in healing tendons. On the other hand, when P-PRP is used to treat acutely injured tendons, it may result in the formation of excessive scar tissue due to the strong potential of P-PRP to induce inordinate cellular anabolic effects.

Prostaglandin E2 (PGE2) has been reported to exert different effects on tissues at low and high levels. In the present study, cell culture experiments were performed to determine the potential biphasic effects of PGE2 on human tendon stem/progenitor cells (hTSCs). After treatment with PGE2, hTSC proliferation, stemness, and differentiation were analyzed. We found that high concentrations of PGE2 (>1 ng/ml) decreased cell proliferation and induced non-tenocyte differentiation. However, at lower concentrations (<1 ng/ml), PGE2 markedly enhanced hTSC proliferation. The expression levels of stem cell marker genes, specifically SSEA-4 and Stro-1, were more extensive in hTSCs treated with low concentrations of PGE2 than in cells treated with high levels of PGE2. Moreover, high levels of PGE2 induced hTSCs to differentiate aberrantly into non-tenocytes, which was evident by the high levels of PPARγ, collagen type II, and osteocalcin expression in hTSCs treated with PGE2 at concentrations >1 ng/ml. The findings of this study reveal that PGE2 can exhibit biphasic effects on hTSCs, indicating that while high PGE2 concentrations may be detrimental to tendons, low levels of PGE2 may play a vital role in the maintenance of tendon homeostasis in vivo.

Tendon stem cells (TSCs) may be used to effectively repair or regenerate injured tendons. However, the fates of TSCs once implanted in vivo remain unclear. This study was aimed to determine the feasibility of labeling TSCs with super-paramagnetic iron oxide (SPIO) nano-particles to track TSCs in vivo using MRI. Rabbit TSCs were labeled by incubation with 50 μg/mL SPIO. Labeling efficiency, cell viability, and proliferation were then measured, and the stemness of TSCs was tested by quantitative real time RT-PCR (qRT-PCR) and immunocytochemistry. We found that the labeling efficiency of TSCs reached as high as 98%, and that labeling at 50 μg/mL SPIO concentrations did not alter cell viability and cell proliferation compared to non-labeled control cells. Moreover, the expression levels of stem cell markers (Nucleostemin, Nanog, and Oct-4) did not change in SPIO-labeled TSCs compared to non-labeled cells. Both labeled and non-labeled cells also exhibited similar differentiation potential. Finally, labeled TSCs could be detected by MRI both in vitro and in vivo. Taken together, the findings of this study show that labeling TSCs with SPIO particles is a feasible approach to track TSCs in vivo by MRI, which offers a non-invasive method to monitor repair of injured tendons.

Mechanical loading constantly acts on tendons, and a better understanding of its effects on the tendons is essential to gain more insights into tendon patho-physiology. This study aims to investigate tendon mechanobiological responses through the use of mouse treadmill running as an in vivo model and mechanical stretching of tendon cells as an in vitro model. In the in vivo study, mice underwent moderate treadmill running (MTR) and intensive treadmill running (ITR) regimens. Treadmill running elevated the expression of mechanical growth factors (MGF) and enhanced the proliferative potential of tendon stem cells (TSCs) in both patellar and Achilles tendons. In both tendons, MTR upregulated tenocyte-related genes: collagen type I (Coll. I ∼10 fold) and tenomodulin (∼3-4 fold), but did not affect non-tenocyte-related genes: LPL (adipocyte), Sox9 (chondrocyte), Runx2 and Osterix (both osteocyte). However, ITR upregulated both tenocyte (Coll. I ∼7-11 fold; tenomodulin ∼4-5 fold) and non-tenocyte-related genes (∼3-8 fold). In the in vitro study, TSCs and tenocytes were stretched to 4% and 8% using a custom made mechanical loading system. Low mechanical stretching (4%) of TSCs from both patellar and Achilles tendons increased the expression of only the tenocyte-related genes (Coll. I ∼5-6 fold; tenomodulin ∼6-13 fold), but high mechanical stretching (8%) increased the expression of both tenocyte (Coll. I ∼28-50 fold; tenomodulin ∼14-48 fold) and non-tenocyte-related genes (2-5-fold). However, in tenocytes, non-tenocyte related gene expression was not altered by the application of either low or high mechanical stretching. These findings indicate that appropriate mechanical loading could be beneficial to tendons because of their potential to induce anabolic changes in tendon cells. However, while excessive mechanical loading caused anabolic changes in tendons, it also induced differentiation of TSCs into non-tenocytes, which may lead to the development of degenerative tendinopathy frequently seen in clinical settings.

Tissues and organs in vivo are under a hypoxic condition; that is, the oxygen tension is typically much lower than in ambient air. However, the effects of such a hypoxic condition on tendon stem cells, a recently identified tendon cell, remain incompletely defined. In cell culture experiments, we subjected human tendon stem cells (hTSCs) to a hypoxic condition with 5% O2, while subjecting control cells to a normaxic condition with 20% O2. We found that hTSCs at 5% O2 had significantly greater cell proliferation than those at 20% O2. Moreover, the expression of two stem cell marker genes, Nanog and Oct-4, was upregulated in the cells cultured in 5% O2. Finally, in cultures under 5% O2, more hTSCs expressed the stem cell markers nucleostemin, Oct-4, Nanog and SSEA-4. In an in vivo experiment, we found that when both cell groups were implanted with tendon-derived matrix, more tendon-like structures formed in the 5% O2 treated hTSCs than in 20% O2 treated hTSCs. Additionally, when both cell groups were implanted with Matrigel, the 5% O2 treated hTSCs showed more extensive formation of fatty, cartilage-like and bone-like tissues than the 20% O2 treated cells. Together, the findings of this study show that oxygen tension is a niche factor that regulates the stemness of hTSCs, and that less oxygen is better for maintaining hTSCs in culture and expanding them for cell therapy of tendon injuries.

When injured, tendons tend to heal but with poor structure and compromised function. Tissue engineering is a promising approach to enhancing the quality of healing tendons. Our group and others have identified tendon stem cells (TSCs), a type of tendon-specific stem cells which may be optimal for cellular interventions seeking to restore normal structure and function to injured tendons. However, in vitro expanding of TSCs on regular plastic cell culture dishes only yields a limited number of TSCs before they lose the stemness, i.e., the self-renewal capability and multipotency. In this study, we developed a substrate material for TSCs, engineered tendon matrix (ETM) from decellularized tendon tissues. We showed that ETM in vitro was able to stimulate TSC proliferation and better preserve the stemness of TSCs than plastic culture surfaces. In vivo, implantation of ETM-TSC composite promoted tendon-like tissue formation whereas implantation of TSCs alone led to little such tissue formation. Together, the findings of this study indicate that ETM may be used to effectively expand TSCs in vitro and with TSCs, to enhance repair of injured tendons in vivo., (Copyright © 2011 Elsevier Ltd. All rights reserved.)

Background: Platelet-rich plasma (PRP) has been used to enhance tendon healing in clinical settings. However, the cellular mechanisms underlying PRP treatment of injured tendons remain unclear. The aim of this study was to determine the effects of PRP, in the form of PRP-clot releasate (PRCR), on tendon stem cells (TSCs), a newly discovered cell population in tendons., Hypothesis: The PRCR treatment promotes differentiation of TSCs into tenocytes that are activated to proliferate quickly and increase collagen production., Study Design: Controlled laboratory study., Methods: After PRCR treatment, cell morphology, expression of stem/progenitor cell marker nucleostemin, and population doubling time were examined. In addition, gene and protein analyses were performed using reverse transcription-polymerase chain reaction, immunocytochemistry, and Western blot to characterize the type of cells that had differentiated after PRCR treatment., Results: The TSCs without PRCR treatment were small and exhibited an irregular shape, whereas with increasing PRCR dosage, TSCs became large, well spread, and highly elongated with downregulation of nucleostemin expression. The PRCR treatment also markedly enhanced TSC proliferation, tenocyte-related gene and protein expression, and total collagen production, all of which indicated that PRCR treatment induced differentiation of TSCs into activated tenocytes., Conclusion: The PRCR treatment promotes differentiation of TSCs into active tenocytes exhibiting high proliferation rates and collagen production capability., Clinical Relevance: The findings of this study suggest that PRP treatment of injured tendons is "safe" as it promotes TSC differentiation into tenocytes rather than nontenocytes, which would compromise the structure and function of healing tendons by formation of nontendinous tissues. Moreover, they suggest that PRP treatment can enhance tendon healing because tenocytes induced to differentiate by PRP are activated to proliferate quickly and produce abundant collagen to repair injured tendons that have lost cells and matrix.

Background: Tendons are traditionally thought to consist of tenocytes only, the resident cells of tendons; however, a recent study has demonstrated that human and mouse tendons also contain stem cells, referred to as tendon stem/progenitor cells (TSCs). However, the differential properties of TSCs and tenocytes remain largely undefined. This study aims to characterize the properties of these tendon cells derived from rabbits., Methods: TSCs and tenocytes were isolated from patellar and Achilles tendons of rabbits. The differentiation potential and cell marker expression of the two types of cells were examined using histochemical, immunohistochemical, and qRT-PCR analysis as well as in vivo implantation. In addition, morphology, colony formation, and proliferation of TSCs and tenocytes were also compared., Results: It was found that TSCs were able to differentiate into adipocytes, chondrocytes, and osteocytes in vitro, and form tendon-like, cartilage-like, and bone-like tissues in vivo. In contrast, tenocytes had little such differentiation potential. Moreover, TSCs expressed the stem cell markers Oct-4, SSEA-4, and nucleostemin, whereas tenocytes expressed none of these markers. Morphologically, TSCs possessed smaller cell bodies and larger nuclei than ordinary tenocytes and had cobblestone-like morphology in confluent culture whereas tenocytes were highly elongated. TSCs also proliferated more quickly than tenocytes in culture. Additionally, TSCs from patellar tendons formed more numerous and larger colonies and proliferated more rapidly than TSCs from Achilles tendons., Conclusions: TSCs exhibit distinct properties compared to tenocytes, including differences in cell marker expression, proliferative and differentiation potential, and cell morphology in culture. Future research should investigate the mechanobiology of TSCs and explore the possibility of using TSCs to more effectively repair or regenerate injured tendons.

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. Clinical significance: Metformin may be used as a therapeutic to aging‐induced tendinopathy in clinics. [ABSTRACT FROM AUTHOR]

The elderly population is prone to tendinopathy due to aging-related tendon changes such as cellular senescence and a decreased ability to modulate inflammation. Aging can render tendon stem/progenitor cells (TSCs) into premature senescence. We investigated the effects of rapamycin, a specific mTOR inhibitor, on the senescence of TSCs. We first showed that after treatment with bleomycin in vitro, rat patellar TSCs (PTSCs) underwent senescence, characterized by morphological alterations, induction of senescence-associated β-galactosidase (SA-β-gal) activity, and an increase in p53, p21, and p62 protein expression. Senescence of PTSCs was also characterized by the elevated expression of MMP-13 and TNF-α genes, both of which are molecular hallmarks of chronic tendinopathy. We then showed that rapamycin treatment was able to reverse the above senescent phenotypes and increase autophagy in the senescent PTSCs. The activation of autophagy and senescence rescue was, at least partly, due to the translocation of HMGB1 from the nucleus to the cytosol that functions as an autophagy promoter. By reducing TSC senescence, rapamycin may be used as a therapeutic to inhibit tendinopathy development in the aging population by promoting autophagy. [ABSTRACT FROM AUTHOR]

Mechanical overloading is a major cause of tendinopathy, but the underlying pathogenesis of tendinopathy is unclear. Here we report that high mobility group box1 (HMGB1) is released to the tendon extracellular matrix and initiates an inflammatory cascade in response to mechanical overloading in a mouse model. Moreover, administration of glycyrrhizin (GL), a naturally occurring triterpene and a specific inhibitor of HMGB1, inhibits the tendon’s inflammatory reactions. Also, while prolonged mechanical overloading in the form of long-term intensive treadmill running induces Achilles tendinopathy in mice, administration of GL completely blocks the tendinopathy development. Additionally, mechanical overloading of tendon cells in vitro induces HMGB1 release to the extracellular milieu, thereby eliciting inflammatory and catabolic responses as marked by increased production of prostaglandin E2 (PGE2) and matrix metalloproteinase-3 (MMP-3) in tendon cells. Application of GL abolishes the cellular inflammatory/catabolic responses. Collectively, these findings point to HMGB1 as a key molecule that is responsible for the induction of tendinopathy due to mechanical overloading placed on the tendon. [ABSTRACT FROM AUTHOR]

Previous animal studies have shown that long term rat treadmill running induces over-use tendinopathy, which manifests as proteoglycan accumulation and chondrocytes-like cells within the affected tendons. Creating this animal model of tendinopathy by long term treadmill running is however time-consuming, costly and may vary among animals. In this study, we used a new approach to develop an animal model of tendinopathy using kartogenin (KGN), a bio-compound that can stimulate endogenous stem/progenitor cells to differentiate into chondrocytes. KGN-beads were fabricated and implanted into rat Achilles tendons. Five weeks after implantation, chondrocytes and proteoglycan accumulation were found at the KGN implanted site. Vascularity as well as disorganization in collagen fibers were also present in the same site along with increased expression of the chondrocyte specific marker, collagen type II (Col. II). In vitro studies confirmed that KGN was released continuously from KGN-alginate in vivo beads and induced chondrogenic differentiation of tendon stem/progenitor cells (TSCs) suggesting that chondrogenesis after KGN-bead implantation into the rat tendons is likely due to the aberrant differentiation of TSCs into chondrocytes. Taken together, our results showed that KGN-alginate beads can be used to create a rat model of tendinopathy, which, at least in part, reproduces the features of over-use tendinopathy model created by long term treadmill running. This model is mechanistic (stem cell differentiation), highly reproducible and precise in creating localized tendinopathic lesions. It is expected that this model will be useful to evaluate the effects of various topical treatments such as NSAIDs and platelet-rich plasma (PRP) for the treatment of tendinopathy. [ABSTRACT FROM AUTHOR]

Aging is known to cause tendon degeneration whereas moderate exercise imparts beneficial effects on tendons. Since stem cells play a vital role in maintaining tissue integrity, in this study we aimed to define the effects of aging and moderate exercise on tendon stem/progenitor cells (TSCs) using in vitro and in vivo models. TSCs derived from aging mice (9 and 24 months) proliferated significantly slower than TSCs obtained from young mice (2.5 and 5 months). In addition, expression of the stem cell markers Oct-4, nucleostemin (NS), Sca-1 and SSEA-1 in TSCs decreased in an age-dependent manner. Interestingly, moderate mechanical stretching (4%) of aging TSCs in vitro significantly increased the expression of the stem cell marker, NS, but 8% stretching decreased NS expression. Similarly, 4% mechanical stretching increased the expression of Nanog, another stem cell marker, and the tenocyte-related genes, collagen I and tenomodulin. However, 8% stretching increased expression of the non-tenocyte-related genes, LPL, Sox-9 and Runx-2, while 4% stretching had minimal effects on the expression of these genes. In the in vivo study, moderate treadmill running (MTR) of aging mice (9 months) resulted in the increased proliferation rate of aging TSCs in culture, decreased lipid deposition, proteoglycan accumulation and calcification, and increased the expression of NS in the patellar tendons. These findings indicate that while aging impairs the proliferative ability of TSCs and reduces their stemness, moderate exercise can mitigate the deleterious effects of aging on TSCs and therefore may be responsible for decreased aging-induced tendon degeneration. [ABSTRACT FROM AUTHOR]

Mechanical loading constantly acts on tendons, and a better understanding of its effects on the tendons is essential to gain more insights into tendon patho-physiology. This study aims to investigate tendon mechanobiological responses through the use of mouse treadmill running as an in vivo model and mechanical stretching of tendon cells as an in vitro model. In the in vivo study, mice underwent moderate treadmill running (MTR) and intensive treadmill running (ITR) regimens. Treadmill running elevated the expression of mechanical growth factors (MGF) and enhanced the proliferative potential of tendon stem cells (TSCs) in both patellar and Achilles tendons. In both tendons, MTR upregulated tenocyte-related genes: collagen type I (Coll. I ∼10 fold) and tenomodulin (∼3–4 fold), but did not affect non-tenocyte-related genes: LPL (adipocyte), Sox9 (chondrocyte), Runx2 and Osterix (both osteocyte). However, ITR upregulated both tenocyte (Coll. I ∼7–11 fold; tenomodulin ∼4–5 fold) and non-tenocyte-related genes (∼3–8 fold). In the in vitro study, TSCs and tenocytes were stretched to 4% and 8% using a custom made mechanical loading system. Low mechanical stretching (4%) of TSCs from both patellar and Achilles tendons increased the expression of only the tenocyte-related genes (Coll. I ∼5–6 fold; tenomodulin ∼6–13 fold), but high mechanical stretching (8%) increased the expression of both tenocyte (Coll. I ∼28–50 fold; tenomodulin ∼14–48 fold) and non-tenocyte-related genes (2–5-fold). However, in tenocytes, non-tenocyte related gene expression was not altered by the application of either low or high mechanical stretching. These findings indicate that appropriate mechanical loading could be beneficial to tendons because of their potential to induce anabolic changes in tendon cells. However, while excessive mechanical loading caused anabolic changes in tendons, it also induced differentiation of TSCs into non-tenocytes, which may lead to the development of degenerative tendinopathy frequently seen in clinical settings. [ABSTRACT FROM AUTHOR]

Tendon stem cells (TSCs) have been proposed to play a major role in the development of tendinopathy, which refers to pathological changes, such as calcification, in affected tendons. Using a human TSC (hTSC) culture model, this study investigated the effects of PGE2, an inflammatory mediator present in injured tendons, on hTSC proliferation and differentiation as well as the molecular mediator for such PGE2-induced effects. We found that PGE2 treatment of hTSCs decreased cell proliferation and caused osteogenic differentiation of hTSCs in a dose-dependent manner. Also, PGE2 treatment of hTSCs induced dose-dependent BMP-2 production in culture, and moreover, addition of BMP-2 to hTSC culture decreased cell proliferation and induced hTSC differentiation into osteoblasts. Finally, addition of BMP-2 antibodies to hTSC culture treated with PGE2 nearly abolished PGE2 effects on both cell proliferation and osteogenic differentiation. Taken together, the findings of this study showed that BMP-2 mediates PGE2-induced reduction of proliferation and osteogenic differentiation of hTSCs. We suggest that such a mechanism may be partially responsible for the formation of calcified tissues in tendinopathic tendons seen in clinical settings. © 2011 Orthopaedic Research Society Published by Wiley Periodicals, Inc. J Orthop Res 30:47-52, 2012 [ABSTRACT FROM AUTHOR]

Exercise is known to enhance tendon size and strength, but the stem cell-based mechanisms for such exercise-induced effects are largely unknown. This study aims to explore these mechanisms by using a mouse treadmill running model to examine the effects of exercise on newly discovered tendon stem cells (TSCs). After treadmill running, patellar TSCs (PTSCs) and Achilles TSCs (ATSCs) were isolated from the mice, and their proliferation was measured in vitro. We found that treadmill running nearly doubled proliferation rates of both PTSCs and ATSCs compared to cage control mice. Moreover, using a mixed tendon cell culture consisting of TSCs and tenocytes, cellular production of collagen was found to increase by 70% and 200% in PTSCs and ATSCs, respectively, from the treadmill running group over cells from the cage control group. These findings suggest that exercise exerts its anabolic effects on tendons at least in part by increasing proliferation to expand the pool of TSCs and also by increasing TSC-related cellular production of collagen, the predominant component of tendons. © 2010 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 28:1178-1183, 2010 [ABSTRACT FROM AUTHOR]

Tendons are constantly subjected to mechanical loading in vivo. Recently, stem cells were identified in human, mouse, and rabbit tendons, but the mechanobiological responses of tendon stem cells (TSCs) are still undefined. Using an in vitro system capable of mimicking in vivo loading conditions, it was determined that mechanical stretching increased TSC proliferation in a stretching magnitude-dependent manner. Moreover, low mechanical stretching at 4% ('clamp-to-clamp' engineering strain) promoted differentiation of TSCs into tenocytes, whereas large stretching at 8% induced differentiation of some TSCs into adipogenic, chondrogenic, and osteogenic lineages, as indicated by upregulated expression of marker genes for adipocytes, chondrocytes, and osteocytes. Thus, low mechanical stretching may be beneficial to tendons by enabling differentiation of TSCs into tenocytes to maintain tendon homeostasis. However, large mechanical loading may be detrimental, as it directs differentiation of TSCs into non-tenocytes in tendons, thus resulting in lipid accumulation, mucoid formation, and tissue calcification, which are typical features of tendinopathy at later stages. © 2009 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 28:639-643, 2010 [ABSTRACT FROM AUTHOR]

Mechanical loading is known to alter tendon structure, but its cellular mechanisms are unclear. This study aimed to determine the effect of mechanical loading on tendon cells in vivo. C57BL/6J female mice were used in a treadmill running study. The treadmill running protocol consisted of treadmill training for 1 week, followed by sustained moderate running at 13 m/min for 50 min/day, 5 days/week, for 3 weeks. Immunohistochemical staining of tendon sections of mice after treadmill running revealed that numerous cells in the tendon section expressed α-SMA, whereas in the tendon sections of control mice, only a few cells exhibited weak α-SMA signals. Furthermore, mouse patellar tendon cells (MPTCs) derived from treadmill running mice were generally larger in culture, proliferated faster, expressed a higher level of α-SMA, and formed more abundant stress fibers compared to MPTCs from control mice. In addition, MPTCs from treadmill running mice generated larger traction forces (169 ± 66.1 Pa) than those from control mice (102 ± 34.2 Pa). Finally, cells from treadmill running mice produced higher levels of total collagen (516.4 ± 92.7 µg/10,000 cells) than their counterparts (303.9 ± 34.8 µg/10,000 cells). Thus, mechanical loading via treadmill running increased the presence of myofibroblasts in mouse patellar tendons. As myofibroblasts are activated fibroblasts, their presence in the tendon following treadmill running indicates that they actively repair and remodel tendon tissue under strenuous mechanical loading, leading to known changes in tendon structure. © 2009 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 27:1373-1378, 2009 [ABSTRACT FROM AUTHOR]

To understand the variable efficacy with platelet rich plasma (PRP) treatments for tendon injury, we determined the differential effects of proteinase-activated receptor (PAR)1- or PAR4-activated PRP (PAR1-PRP, PAR4-PRP) from humans on human patellar tendon stem/progenitor cells (TSCs) and tendon healing. We show that PAR1-PRP released VEGF, whereas PAR4-PRP released endostatin. Treatment of TSCs with PAR1-PRP increased collagen I expression and matrix metalloproteinase-1 (MMP-1), but cells treated with PAR4-PRP increased less collagen I and higher MMP-2 expression. The wound area treated with PAR4-PRP formed tendon-like tissues with well-organized collagen fibers and fewer blood vessels, while PAR1-PRP treatment resulted in the formation of blood vessels and unhealed tissues. These findings indicate that differential activation of PRP leads to different effects on TSCs and tendon healing. We suggest that based on acute or chronic type of tendon injury, selective activation of PRP should be applied in clinics in order to treat injured tendons successfully. [ABSTRACT FROM AUTHOR]

Objectives: The purpose of this study is to investigate the effect of Agkistroden blomhoffi (mamushi) aqueous extract on human patellar tendon cells in vitro, to pharmacologically explain the natural medicine''s healing effect on tendon, bone and muscle injuries.Design and methods: Human patellar tendon fibroblasts (HPTF) were incubated in media containing different concentrations of mamushi aqueous extract. Cell proliferation was studied by microscopic observations and total protein, actin, collagen I, and cyclooxygenase-2 (Cox 2) expressions.Results: Mamushi aqueous extract enhanced HPTF proliferation when its concentration was lower than 333 μg/ml. Cells cultured in manushi-containing medium showed developed intercellular structure and increased protein production. However, mamushi extract higher than 500 μg/ml oppressed cell growth. At 667 μg/ml, mamushi induced Cox 2 production, a sign of cytotoxicity.Conclusion: A. blomhoffi aqueous extract was found to directly stimulate the proliferation and protein production, particularly collagen I synthesis, of HPTF in a dose-dependent manner. [Copyright &y& Elsevier]