软骨寡聚基质蛋白对BMP 2诱导骨髓间质干细胞分化的影响

 目的 探讨过表达软骨寡聚基质蛋白（cartilage oligomeric matrix protein,COMP）对BMP-2诱导骨髓间质干细胞成骨及成软骨分化的影响。方法 用BMP-2诱导骨髓间质干细胞分化,通过脂质体转染含人COMP基因的质粒使骨髓间质干细胞过表达COMP,空载质粒作为对照。以RT-PCR检测成骨相关基因Ⅰ型胶原、RUNX2、骨桥蛋白、骨钙蛋白以及成软骨相关基因Ⅱ型胶原、SOX9、蛋白聚糖的表达变化;通过碱性磷酸酶染色观察成骨过程中的碱性磷酸酶活性,茜素红染色观察成骨终末阶段矿化结节的生成情况,阿利新蓝染色观察细胞基质蛋白多糖的合成情况。结果 COMP组目的基因COMP mRNA的表达显著升高;骨桥蛋白mRNA表达水平较对照组低,呈现出一致的下调趋势（P<0.05）;Ⅰ型胶原、RUNX2、骨钙蛋白mRNA表达水平在诱导早期均高于对照组（P<0.05）,但在诱导晚期均明显低于对照组（P<0.05）;成软骨指标（Ⅱ型胶原、SOX9、蛋白聚糖）的基因表达水平强于对照组,呈一致的上调趋势（P<0.05）;SOX9 mRNA表达水平高于对照组,仅在第7天时差异具有统计学意义（P<0.05）;细胞成骨染色（碱性磷酸酶染色、茜素红染色）均弱于对照组,而阿利新蓝染色强于对照组。结论 COMP能抑制BMP-2诱导骨髓间质干细胞成骨分化,促进BMP-2诱导骨髓间质干细胞成软骨分化。     

Adipose tissue is an ideal stem cell source for tissue reconstruction of bone，cartilage，and soft tissue defects

目的 验证人脂肪基质细胞是否具有向成骨细胞、软骨细胞、脂肪细胞分化的能力,从而为骨、软骨、软组织再建寻找一种理想的干细胞来源.方法 分别用成骨向分化培养基（DMEM＋10?S＋地塞米松＋维生素C＋β-甘油磷酸）、软骨向分化培养基（DMEM＋1?S＋胰岛素＋维生素C＋转化生长因子β1）及脂肪向分化培养基（DMEM＋10?S＋地塞米松＋胰岛素＋吲哚美辛＋异丁基甲基黄嘌呤）诱导人脂肪基质细胞向成骨细胞、软骨细胞及脂肪细胞分化.用von Kossa和碱性磷酸酶染色鉴定成骨细胞分化,而软骨细胞分化和脂肪细胞分化分别用Alcian blue染色和油红O染色显示.成骨细胞、软骨细胞以及脂肪细胞特异相关或标志基因的表达用RT-PCR检测.结果 体外实验表明,人脂肪基质细胞在定向分化诱导剂的作用下可分别向成骨细胞、软骨细胞及脂肪细胞分化.结论 人脂肪基质细胞中包含有多向分化能力的干细胞,可用于今后骨、软骨、软组织的组织工程再建.     

Insulin‐like growth factor 2 (IGF‐2) potentiates BMP‐9‐induced osteogenic differentiation and bone formation

Efficient osteogenic differentiation and bone formation from mesenchymal stem cells (MSCs) should have clinical applications in treating nonunion fracture healing. MSCs are adherent bone marrow stromal cells that can self‐renew and differentiate into osteogenic, chondrogenic, adipogenic, and myogenic lineages. We have identified bone morphogenetic protein 9 (BMP‐9) as one of the most osteogenic BMPs. Here we investigate the effect of insulin‐like growth factor 2 (IGF‐2) on BMP‐9‐induced bone formation. We have found that endogenous IGF‐2 expression is low in MSCs. Expression of IGF‐2 can potentiate BMP‐9‐induced early osteogenic marker alkaline phosphatase (ALP) activity and the expression of later markers. IGF‐2 has been shown to augment BMP‐9‐induced ectopic bone formation in the stem cell implantation assay. In perinatal limb explant culture assay, IGF‐2 enhances BMP‐9‐induced endochondral ossification, whereas IGF‐2 itself can promote the expansion of the hypertropic chondrocyte zone of the cultured limb explants. Expression of the IGF antagonists IGFBP3 and IGFBP4 leads to inhibition of the IGF‐2 effect on BMP‐9‐induced ALP activity and matrix mineralization. Mechanistically, IGF‐2 is further shown to enhance the BMP‐9‐induced BMPR‐Smad reporter activity and Smad1/5/8 nuclear translocation. PI3‐kinase (PI3K) inhibitor LY294002 abolishes the IGF‐2 potentiation effect on BMP‐9‐mediated osteogenic signaling and can directly inhibit BMP‐9 activity. These results demonstrate that BMP‐9 crosstalks with IGF‐2 through PI3K/AKT signaling pathway during osteogenic differentiation of MSCs. Taken together, our findings suggest that a combination of BMP‐9 and IGF‐2 may be explored as an effective bone‐regeneration agent to treat large segmental bony defects, nonunion fracture, and/or osteoporotic fracture. © 2010 American Society for Bone and Mineral Research.     

Donor Heart Treatment With COMP‐Ang1 Limits Ischemia‐Reperfusion Injury and Rejection of Cardiac Allografts

The major cause of death during the first year after heart transplantation is primary graft dysfunction due to preservation and ischemia‐reperfusion injury (IRI). Angiopoietin‐1 is a Tie2 receptor‐binding paracrine growth factor with anti‐inflammatory properties and indispensable roles in vascular development and stability. We used a stable variant of angiopoietin‐1 (COMP‐Ang1) to test whether ex vivo intracoronary treatment with a single dose of COMP‐Ang1 in donor Dark Agouti rat heart subjected to 4‐h cold ischemia would prevent microvascular dysfunction and inflammatory responses in the fully allogeneic recipient Wistar Furth rat. COMP‐Ang1 reduced endothelial cell–cell junction disruption of the donor heart in transmission electron microscopy during 4‐h cold ischemia, improved myocardial reflow, and reduced microvascular leakage and cardiomyocyte injury of transplanted allografts during IRI. Concurrently, the treatment reduced expression of danger signals, dendritic cell maturation markers, endothelial cell adhesion molecule VCAM‐1 and RhoA/Rho‐associated protein kinase activation and the influx of macrophages and neutrophils. Furthermore, COMP‐Ang1 treatment provided sustained anti‐inflammatory effects during acute rejection and prevented the development of cardiac fibrosis and allograft vasculopathy. These results suggest donor heart treatment with COMP‐Ang1 having important clinical implications in the prevention of primary and subsequent long‐term injury and dysfunction in cardiac allografts.     

Noggin resistance contributes to the potent osteogenic capability of BMP9 in mesenchymal stem cells

Mesenchymal stem cells (MSCs) are multipotent progenitors and can differentiate into osteogenic, chondrogenic, and adipogenic lineages. Bone morphogenetic proteins (BMPs) play important roles in stem cell proliferation and differentiation. We recently demonstrated that BMP9 is a potent but less understood osteogenic factor. We previously found that BMP9‐induced ectopic bone formation is not inhibited by BMP3. Here, we investigate the effect of BMP antagonist noggin on BMP9‐induced osteogenic differentiation. BMP antagonists noggin, chording, gremlin, follistatin, and BMP3 are highly expressed in MSCs, while noggin and follistatin are lowly expressed in more differentiated pre‐osteoblast C2C12 cells. BMP9‐induced osteogenic markers and matrix mineralization are not inhibited by noggin, while noggin blunts BMP2, BMP4, BMP6, and BMP7‐induced osteogenic markers and mineralization. Likewise, ectopic bone formation by MSCs transduced with BMP9, but not the other four BMPs, is resistant to noggin inhibition. BMP9‐induced nuclear translocation of Smad1/5/8 is not affected by noggin, while noggin blocks BMP2‐induced activation of Smad1/5/8 in MSCs. Noggin fails to inhibit BMP9‐induced expression of downstream targets in MSCs. Thus, our results strongly suggest that BMP9 may effectively overcome noggin inhibition, which should at least in part contribute to BMP9's potent osteogenic capability in MSCs. © 2013 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 31:1796–1803, 2013     

Glycogen synthase kinase‐3α/β inhibition promotes in vivo amplification of endogenous mesenchymal progenitors with osteogenic and adipogenic potential and their differentiation to the osteogenic lineage

Small molecules are attractive therapeutics to amplify and direct differentiation of stem cells. They also can be used to understand the regulation of their fate by interfering with specific signaling pathways. Mesenchymal stem cells (MSCs) have the potential to proliferate and differentiate into several cell types, including osteoblasts. Activation of canonical Wnt signaling by inhibition of glycogen synthase kinase 3 (GSK‐3) has been shown to enhance bone mass, possibly by involving a number of mechanisms ranging from amplification of the mesenchymal stem cell pool to the commitment and differentiation of osteoblasts. Here we have used a highly specific novel inhibitor of GSK‐3, AR28, capable of inducing β‐catenin nuclear translocation and enhanced bone mass after 14 days of treatment in BALB/c mice. We have shown a temporally regulated increase in the number of colony‐forming units–osteoblast (CFU‐O) and –adipocyte (CFU‐A) but not colony‐forming units–fibroblast (CFU‐F) in mice treated for 3 days. However, the number of CFU‐O and CFU‐A returned to normal levels after 14 days of treatment, and the number of CFU‐F was decreased significantly. In contrast, the number of osteoblasts increased significantly only after 14 days of treatment, and this was seen together with a significant decrease in bone marrow adiposity. These data suggest that the increased bone mass is the result of an early temporal wave of amplification of a subpopulation of MSCs with both osteogenic and adipogenic potential, which is driven to osteoblast differentiation at the expense of adipogenesis. © 2011 American Society for Bone and Mineral Research.     

Comparison of Multipotent Differentiation Potentials of Murine Primary Bone Marrow Stromal Cells and Mesenchymal Stem Cell Line C3H10T1/2

Murine C3H10T1/2 cells have many features of mesenchymal stem cells (MSCs). Whether or not the multipotent differentiation capability of C3H10T1/2 cells is comparable to that of primary bone marrow–derived MSCs (BM-MSCs) was investigated in this study. For in vitro osteogenic differentiation, both BM-MSCs and C3H10T1/2 cells differentiated to osteoblastic cell lineage and showed positive staining for alkaline phosphatase (ALP) and increased mRNA expression of Runx2, Col1αI, and osteocalcin. C3H10T1/2 cells and BM-MSCs induced similar amounts of bone formation in the biomaterials. Under chondrogenic induction in the presence of TGF-β1, cell pellets of both BM-MSCs and C3H10T1/2 cells formed cartilage-like tissues with cartilage matrix components including proteoglycan, type II collagen, and aggrecan. However, C3H10T1/2 cells presented lower adipogenic differentiation potential, with only about 10% C3H10T1/2 cells (but about 70% of BM-MSCs) being committed to adipogenesis. In this study we confirmed that C3H10T1/2 cells coimplanted with osteoconductive scaffolds can form bone spontaneously in vivo and that C3H10T1/2 cells have a basal level of osteocalcin expression, suggesting that they may be a good alternative source of primary BM-MSCs for investigating osteogenic and chondrogenic differentiation in bone or cartilage tissue engineering studies. Caution is needed when using C3H10T1/2 cells for adipogenic studies as they appear to have lower adipogenic potential than BM-MSCs.     

The effect of two- and three-dimensional cell culture on the chondrogenic potential of human adipose-derived mesenchymal stem cells after subcutaneous transplantation with an injectable hydrogel

Articular cartilage is an avascular tissue composed of chondrocytes, a unique cell type responsible for abundant matrix synthesis and maintenance. When damaged, it never heals spontaneously under physiological circumstances. Therefore, the delivery of mesenchymal stem cells using hydrogel has been considered for cartilage repair. This study aims at investigating the influence of in vitro chondrogenic differentiation of human adipose tissue-derived stem cells (hATSCs) on in vivo cartilage formation when associated with a cellulose-based self-setting hydrogel (Si-HPMC). hATSCs were characterized for their proliferation, surface marker expression, and multipotency. The in vitro chondrogenic potential of hATSCs cultured within Si-HPMC in control or chondrogenic medium was evaluated by measuring COL2A1, ACAN, SOX9, and COMP expression by real-time PCR. Alcian blue and type II collagen staining were also performed. To determine whether in vitro chondrogenically differentiated hATSCs may give rise to cartilage in vivo, cells differentiated as a monolayer or in pellets were finally associated with Si-HPMC and implanted subcutaneously into nude mice. Cartilage formation was assessed histologically by alcian blue and type II collagen staining. Our data demonstrate that hATSCs exhibited proliferation and self-renewal. hATSCs also expressed typical stem cell surface markers and were able to differentiate towards the adipogenic, osteogenic, and chondrogenic lineages. Real-time PCR and histological analysis indicated that Si-HPMC enabled chondrogenic differentiation of hATSCs in inductive medium, as demonstrated by increased expression of chondrogenic markers. In addition, histological analysis of implants showed that chondrogenically differentiated hATSCs (monolayers or pellets) have the ability to form cartilaginous tissue, as indicated by the presence of sulphated glycosaminoglycans and type II collagen. This study therefore suggests that an in vitro induction of hATSCs in 2D was sufficient to obtain cartilaginous tissue formation in vivo. Si-HPMC associated with autologous hATSCs could thus be a significant tool for regenerative medicine in the context of cartilage damage.     

ERK途径在转化生长因子-β3诱导骨髓基质干细胞向软骨分化过程中的作用

目的 观察丝裂原活化蛋白激酶(MAPK)信号途径中细胞外信号调节的激酶ERK途径在转化生长因子(TGF)-β3诱导的骨髓基质干细胞(MSCs)向软骨分化的过程中的作用.方法 在体外培养大鼠的MSCs,在含有TGF-β3的诱导培养基中向软骨方向诱导分化,在诱导分化的不同时间点,分别用Western blot测定ERK1/2的表达和磷酸化.确定ERK1/2在分化过程中的变化,同时测定在分化的过程中MSCs与软骨分化相关基因的表达,之后用ERK1/2的抑制剂U0126,观察ERK1/2信号传导通路阻断后对软骨分化的影响.结果 在TGF-β3促使MSCs向软骨方向分化的过程中,ERK1/2参与了细胞的分化和相关软骨基质的合成,ERK1/2抑制剂的使用削弱和减缓了上述过程的发生.结论 ERK1/2途径在软骨分化过程中起重要作用.     

Dynamic expression of DKK1 protein in the process whereby Epimedium‐derived flavonoids up‐regulate osteogenic and down‐regulate adipogenic differentiation of bone marrow stromal cells in ovariectomized rats

Objective: To observe the dynamic expression of DKK1 protein in the process whereby Epimedium‐derived flavonoids (EFs) regulate the balance between osteogenic and adipogenic differentiation of bone marrow stromal cells in ovariectomized rats, and to provide experimental evidence for the mechanism of EFs in the treatment of postmenopausal osteoporosis. Methods: Bone marrow stromal cells from ovariectomized rats were separated and cultivated in osteoinductive or liquid medium for 15 days in vitro. EFs (10 µg/mL) were applied to both cultures. Alkaline phosphatase (ALP) staining, ALP activity determination, Oil Red O staining and fluorescence quantitative polymerase chain reaction were used to determine the influence of EFs on osteogenic and adipogenic differentiation of bone marrow stromal cells in ovariectomized rats. Moreover, in order to explore the exact mechanism of EFs on osteogenic and adipogenic differentiation of bone marrow stromal cells in ovariectomized rats, enzyme linked immunosorbent assay was used to determine the dynamic expression of DKK1 protein in this process. Results: EFs increased activity of ALP and mRNA expression of Runx2 (early osteoblast differentiation factor) and decreased mRNA expression of PPARγ‐2 (key factor of fat generation). Importantly, EFs down‐regulated expression of DKK1 protein in an osteogenic induction medium and inhibited up‐regulation of DKK1 protein in an adipogenic induction medium. Conclusion: EFs regulate the balance between osteogenic and adipogenic differentiation of bone marrow stromal cells in ovariectomized rats by down‐regulating expression of DKK1 protein. This may be an important molecular mechanism of EFs in the context of treatment of postmenopausal osteoporosis.     

骨折后外周血间充质干细胞浓度变化的实验研究

[目的]观察骨折后不同时间点外周血间充质干细胞(MSCs)浓度变化,并比较其与骨髓间充质干细胞生物学特性的异同.[方法]根据不同处理条件将SD大鼠分为:对照组和骨折组(骨折后1、3、7d共3组),每组20只.分别于骨折后1、3、7d抽取外周血,密度梯度离心法分离培养外周血MSCs,计数成纤维细胞集落形成单位(CFU-Fs)数.流式细胞仪检测细胞表面标记(CD44、CD90、CD34、CD45).成骨、成脂诱导,碱性磷酸酶、茜素红和油红染色检测其分化特性.[结果]原代外周血MSCs呈集落生长,骨折组集落数明显多于对照组,其中以骨折后3d组形成的集落数最多,具有显著差异(27.25±11.52 CFU-Fs/cuhure vs 2.80±3.96 CFU-Fs/culture,P＜0.01).外周血MSCs高表达CD44、CD90,低表达CD34、CD45,不同的是CD34小部分呈阳性(＜20％).与对照组骨髓MSCs的诱导结果相同,外周血MSCs成骨诱导后28 d出现钙结节,茜素红染色阳性;成脂诱导后21 d有大量脂滴出现,油红染色阳性.[结论]外周血体外密度梯度离心法分离培养的细胞具有多潜能分化的MSCs表面标记且可以向成骨、成脂分化.骨折后外周循环中MSCs数量明显增多,呈一定的时序性变化,可能参与骨折修复.     

High levels of β‐catenin signaling reduce osteogenic differentiation of stem cells in inflammatory microenvironments through inhibition of the noncanonical Wnt pathway

Periodontal ligament stem cells (PDLSCs), a new population of mesenchymal stem cells (MSCs), have been isolated from the periodontal ligament (PDL). The capacity of multipotency and self‐renewal makes them an excellent cell source for bone regeneration and repair. However, their bone‐regeneration ability could be awakened in inflammatory microenvironments, which may be the result of changes in their differentiation potential. Recently, genetic evidences has shown that the Wnt pathway plays an important role in bone homeostasis. In this study we have determined the specific role of β‐catenin in osteogenic differentiation of PDLSCs obtained from inflammatory microenvironments (P‐PDLSCs). The inflammatory microenvironment, while inhibiting osteogenic differentiation potential, promotes proliferation of MSCs. A higher the level of β‐catenin in P‐PDLSCs than in H‐PDLSCs (PDLSCs obtained from a healthy microenvironment) resulted in the same disparity in canonical Wnt signaling pathway activation between each cell type. Here we show that activation of β‐catenin suppresses the noncanonical Wnt/Ca²⁺ pathway, leading to increased proliferation but reduced osteogenic differentiation of P‐PDLSCs. Downregulation of the levels of β‐catenin by treatment with dickkopf‐1 (DKK‐1) leads to activation of the noncanonical Wnt/Ca²⁺ pathway, which, in turn, results in the promotion of osteogenic differentiation in P‐PDLSCs. Interestingly, β‐catenin can affect both the canonical Wnt/β‐catenin pathway and the noncanonical Wnt/Ca²⁺ pathway. Our data indicate that β‐catenin plays a central role in regulating osteogenic differentiation of MSCs in inflammatory microenvironments. Given the important role of Wnt signaling in osteogenic differentiation, it is possible that agents that can modify this pathway may be of value in bone regeneration by MSCs in chronic inflammatory microenvironments. © 2011 American Society for Bone and Mineral Research     

Multipotent mesenchymal stem cells with immunosuppressive activity can be easily isolated from dental pulp

BACKGROUND: Bone marrow mesenchymal stem cells (MSCs) are currently being investigated in preclinical and clinical settings because of their multipotent differentiative capacity or, alternatively, their immunosuppressive function. The aim of this study was to evaluate dental pulp (DP) as a potential source of MSCs instead of bone marrow (BM). METHODS: Flow cytometric analysis showed that DP-MSCs and BM-MSCs were equally SH2, SH3, SH4, CD29 and CD 166 positive. The in vitro proliferative kinetics of MSCs were measured by 3H-thymidine incorporation uptake. The immunosuppressive function of MSCs was then tested by coculturing PHA-stimulated allogeneic T cells with or without MSCs for 3 days. RESULTS: BM-MSCs could be differentiated in vitro into osteogenic, chondrogenic and adipogenic lineages. DP-MSCs showed osteogenic and adipocytic differentiation, but did not differentiate into chondrocytes. Although DP-MSCs grow rapidly in vitro between day 3 and day 8 of culture and then decrease their proliferation by day 15, BM-MSCs have a stable and continuous proliferation over the same period of time. The addition of DP-MSCs or BM-MSCs resulted in 91 +/- 4% and 75 +/- 3% inhibition of T cell response, respectively, assessed by a 3H-thymidine assay. CONCLUSIONS: Dental pulp is an easily accessible and efficient source of MSCs, with different kinetics and differentiation potentialities from MSCs as isolated from the bone marrow. The rapid proliferative capacity together with the immunoregulatory characteristics of DP-MSCs may prompt future studies aimed at using these cells in the treatment or prevention of T-cell alloreactivity in hematopoietic or solid organ allogeneic transplantation.     

Effects of continuous dexamethasone treatment on differentiation capabilities of bone marrow-derived mesenchymal cells

Human bone marrow-derived mesenchymal cells (hBMMCs) originate from cell populations in the bone marrow and are capable of differentiating along multiple mesenchymal lineages. To differentiate hBMMCs into osteoblasts, adipocytes and chondrocytes, dexamethasone has been used as a differentiation reagent. We hypothesized that dexamethasone would augment the responsiveness of BMMCs to other differentiation reagents and not define the lineage. This study investigated the effect of continuous treatment with 100 nM dexamethasone on the differentiation of BMMCs into three different lineages. hBMMCs cultured with continuous dexamethasone treatment (100 nM) exhibited higher mRNA expression levels of osteogenic markers and higher positive rates of colony forming unit assays for osteogenesis compared to hBMMCs treated with dexamethasone only during the differentiation culture. Furthermore, continuous dexamethasone treatment augmented bone formation capability of monkey-derived BMMCs in a bone induction experimental model at an extra skeletal site. In addition, continuously dexamethasone-treated hBMMCs formed larger chondrogenic pellets and expressed SOX9 at higher level than the control BMMCs. Likewise, continuous dexamethasone treatment facilitated adipogenic differentiation based on mRNA level and colony forming unit analysis. To investigate the mechanism of the augmentation of differentiation, further studies on apoptosis were conducted. The studies indicated that dexamethasone selectively induced apoptosis of some populations of hBMMCs which were thought to have poor differentiation capability.     

Comparative analysis with collagen type II distinguishes cartilage oligomeric matrix protein as a primary TGFβ-responsive gene

Objective

This study aims to investigate the regulation of expression of Cartilage oligomeric matrix protein (COMP), which is predominately expressed by chondrocytes and functions to organize the extracellular matrix. Mutations in COMP cause two skeletal dysplasias: pseudoachondroplasia and multiple epiphyseal dysplasia. The mechanism controlling COMP expression during chondrocyte differentiation is still poorly understood.

Design

Primary human bone marrow-derived stem cells were induced to differentiate into chondrocyte by pellet cultures. We then compared the temporal expression of COMP with the well-characterized cartilage-specific Type II collagen (Col2a1), and their response to transforming growth factor (TGF)β and Sox trio (Sox5, 6, and 9) stimulation.

Results

COMP and Col2a1 expression are differentially regulated by three distinct mechanisms. First, upregulation of COMP mRNA precedes Col2a1 by several days during chondrogenesis. Second, COMP expression is independent of high cell density but requires TGF-β1. Induction of COMP mRNA by TGF-β1 is detected within 2 h in the absence of protein synthesis and is blocked by specific inhibitors of the TGFβ signaling pathway; and therefore, COMP is a primary TFGβ-response gene. Lastly, while Col2a1 expression is intimately controlled by the Sox trio, overexpression of Sox trio fails to activate the COMP promoter.

Conclusion

COMP and Col2a1 expression are regulated differently during chondrogenesis. COMP is a primary response gene of TGFβ and its fast induction during chondrogenesis suggests that COMP is suitable for rapidly accessing the chondrogenic potential of stem cells.     

Editorial Commentary: Tendon-Derived Stem Cells Are in the Rotator Cuff Remnant and Decline With Age and Tear Chronicity—But the Clinical Relevance Is Not Known

The presence of tendon-derived stem cells (TDSCs) and progenitor cells in tendon tissue has been established previously. These cells are part of the mesenchymal adult stem cell line, are multipotent, and can differentiate into several mesenchymal cell lines: osteogenic, chondrogenic, adipogenic, and tenogenic. Mechanical loading may play an important role in the differentiation process and regulates cell differentiation via several signaling pathways. TDSCs can therefore differentiate into several tissues, and the potential for chondrogenic and osteogenic differentiation may be beneficial in tendon-bone regeneration. TDSCs are present in the tendon stumps, and numerous tests have shown that these cells are alive and have the potential to differentiate. Age is a predictor of TDSC activity, and in patients older than 60 years, cell viability and the potential to differentiate are reduced. Despite the theoretical potential that TDSCs may have for tendon healing and improved function, the potential for clinical applications is unclear.