In vitro and in vivo Characterization of Tendon Stem Cells and Role of Stem Cells in Tendon Healing

Tendon repair remains a great challenge due to current therapies cannot restore normal tendon function. Tendon-derived stem cells (TDSCs) have been isolated from tendon tissues and characterized in vitro in recent studies and provide new strategies for tendon repair. But what should we know about tendon stem cells before we use them to repair injured tendon?;Although stem cells that originate from different tissues share some common stem cell characteristics, they might also exhibit some tissue unique properties and hence functional differences. Therefore, we hypothesized that TDSCs have unique stemness properties compared with bone marrow-derived stem cells (BMSCs). There has been no study to compare the stemness properties of TDSCs and BMSCs. Clinical applications often require the in vitro expansion of stem cells. in vitro microenvironment also affects the stemness properties and therapeutic potential of stem cells. It is not clear if the stemness properties of TDSCs can be maintained and how long that they can be preserved during in vitro expansion. Moreover, successful stem cell-based repair therapies will require an understanding of tissue specific stem cells in vivo and their roles in the tissue repair. Tendon stem cells have not been described in details in vivo and it is unknown whether these endogenous stem cells participate in the tendon healing.;Therefore, in order to better make use of TDSCs for tendon repair, the objective of this study is to characterize the stemness properties of TDSCs compared with BMSCs and also to investigate the stemness limitation of TDSCs during culture in vitro for clinical use purpose. Furthermore, this study aims to identify the putative tendon stem cells in vivo and their role in tendon healing. This study would tell how much we should know about tendon stem cells in vitro and in vivo.;In the first part of the study, TDSCs and BMSCs were isolated from the same GFP Sprague-Dawley rat. TDSCs showed higher mensenchymal and pluripotent stem cell makers; clonogenicity; proliferative capacity; and tenogenic, osteogenic, chondrogenic, and adipogenic differentiation markers and multi-lineage differentiation potential than BMSCs. Compared with BMSCs, TDSCs shows great stemness properties and might be an alternative cell source for tendon regeneration.;In the second part of this study, the senescence-associated beta-galactosidase activity of TDSCs increased while their stem cell-related marker expression and the multi-lineage differentiation potential decreased during in vitro passaging. It suggests that researchers and clinicians need to consider the changes of stemness properties of TDSCs when multiplying them in vitro for tissue engineering.;In the third part of the study, IdU label-retaining method was used for the labeling of stem cells in vivo. We have identified quiescent stem cells as IdU label retaining cells (LRCs) at the peritenon, tendon mid-substance and tendon-bone junction. More LRCs were found at the peri-tenon and tendon-bone junction compared to the mid-substance. Some LRCs could be identified in the peri-vascular niche in the peri-tenon, suggesting that peri-vascular niche is one source of tendon stem cells. After injury, The LRC number and the expression of proliferative, tendon-related, pluripotency and pericyte-related markers in LRCs in the window wound increased, indicating that LRCs might be involved in tendon repair via cell migration, proliferation and differentiation.;In conclusion, our results have provided new findings about the understanding of tendon-derived stem cells including their stemness properties and their changes during the in vitro culture, as well as in vivo identity of tendon stem cells, which might facilitate the application of TDSCs in tissue engineering for tendon repair in the future.