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MiR-378a suppresses tenogenic differentiation and tendon repair by targeting at TGF-?2.

ABSTRACT: BACKGROUND:Tendons are a crucial component of the musculoskeletal system and responsible for transmission forces derived from muscle to bone. Patients with tendon injuries are often observed with decreased collagen production and matrix degeneration, and healing of tendon injuries remains a challenge as a result of limited understanding of tendon biology. Recent studies highlight the contribution of miR-378a on the regulation gene expression during tendon differentiation. METHODS:We examined the tendon microstructure and tendon repair with using miR-378a knock-in transgenic mice, and the tendon-derived stem cells were also isolated from transgenic mice to study their tenogenic differentiation ability. Meanwhile, the expression levels of tenogenic markers were also examined in mouse tendon-derived stem cells transfected with miR-378a mimics during tenogenic differentiation. With using online prediction software and luciferase reporter assay, the binding target of miR-378a was also studied. RESULTS:Our results indicated miR-378a impairs tenogenic differentiation and tendon repair by inhibition collagen and extracellular matrix production both in vitro and in vivo. We also demonstrated that miR-378a exert its inhibitory role during tenogenic differentiation through binding at TGF?2 by luciferase reporter assay and western blot. CONCLUSIONS:Our investigation suggests that miR-378a could be considered as a new potential biomarker for tendon injury diagnosis or drug target for a possible therapeutic approach in future clinical practice.

SUBMITTER: Liu Y

PROVIDER: S-EPMC6440014 | biostudies-literature | 2019 Mar

REPOSITORIES: biostudies-literature

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MiR-378a suppresses tenogenic differentiation and tendon repair by targeting at TGF-β2.

Liu Yang Y Feng Lu L Xu Jia J Yang Zhengmeng Z Wu Tianyi T Zhang Jiajun J Shi Liu L Zhu Dahai D Zhang Jinfang J Li Gang G

Stem cell research & therapy 20190329 1

<h4>Background</h4>Tendons are a crucial component of the musculoskeletal system and responsible for transmission forces derived from muscle to bone. Patients with tendon injuries are often observed with decreased collagen production and matrix degeneration, and healing of tendon injuries remains a challenge as a result of limited understanding of tendon biology. Recent studies highlight the contribution of miR-378a on the regulation gene expression during tendon differentiation.<h4>Methods</h4> ...[more]

PMID: 30922407

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Project description:Tendon injury is a common but tough medical problem. Unsatisfactory clinical results have been reported in tendon repair using mesenchymal stem cell (MSC) therapy, creating a need for a better strategy to induce MSCs to tenogenic differentiation. This study was designed to examine the effect of hypoxia on the tenogenic differentiation of different MSCs and their tenogenic differentiation capacities under hypoxia condition in vitro and to investigate the in vivo inductility of hypoxia in tenogenesis. Adipose tissue-derived MSCs (AMSCs) and bone marrow-derived MSCs (BMSCs) were isolated and characterized. The expression of hypoxia-induced factor-1 alpha (Hif-1?) was examined to confirm the establishment of hypoxia condition. qRT-PCR, western blot, and immunofluorescence staining were used to evaluate the expression of tendon-associated marker Col-1a1, Col-3a1, Dcn, and Tnmd in AMSCs and BMSCs under hypoxia condition, compared with Tgf-?1 induction. In vivo, a patellar tendon injury model was established. Normoxic and hypoxic BMSCs were cultured and implanted. Histological, biomechanical, and transmission electron microscopy analyses were performed to assess the improved healing effect of hypoxic BMSCs on tendon injury. Our in vitro results showed that hypoxia remarkably increased the expression of Hif-1? and that hypoxia not only promoted a significant increase in tenogenic markers in both AMSCs and BMSCs compared with the normoxia group but also showed higher inductility compared with Tgf-?1. In addition, hypoxic BMSCs exhibited higher potential of tenogenic differentiation than hypoxic AMSCs. Our in vivo results demonstrated that hypoxic BMSCs possessed better histological and biomechanical properties than normoxic BMSCs, as evidenced by histological scores, patellar tendon biomechanical parameters, and the range and average of collagen fibril diameters. These findings suggested that hypoxia may be a practical and reliable strategy to induce tenogenic differentiation of BMSCs for tendon repair and could enhance the effectiveness of MSCs therapy in treating tendon injury.

Project description:Adipose-derived stem cells (ASCs) are multipotent and immune-privileged mesenchymal cells, making them ideal candidates for therapeutic purposes to manage tendon disorders. Providing safe and regulated cell therapy products to patients requires adherence to good manufacturing practices. To this aim we investigated the in vitro tenogenic differentiation potential of ASCs using a chemically defined serum-free medium (SF) or a xenogenic-free human pooled platelet lysate medium (hPL) suitable for cell therapy and both supplemented with CTGF, TGFβ-3, BMP-12 and ascorbic acid (AA) soluble factors. Human ASCs were isolated from 4 healthy donors and they were inducted to differentiate until 14 days in both hPL and SF tenogenic media (hPL-TENO and SF-TENO). Cell viability and immunophenotype profile were analysed to evaluate mesenchymal stem cell (MSC) characteristics in both xenogenic-free media. Moreover, the expression of stemness and tendon-related markers upon cell differentiation by RT-PCR, protein staining and cytofluorimetric analysis were also performed. Our results showed the two xenogenic-free media well support cell viability of ASCs and maintain their MSC nature as demonstrated by their typical immunophenototype profile and by the expression of NANOG, OCT4 and Ki67 genes. Moreover, both hPL-TENO and SF-TENO expressed significant high levels of the tendon-related genes SCX, COL1A1, COL3A1, COMP, MMP3 and MMP13 already at early time points in comparison to the respective controls. Significant up-regulations in scleraxis, collagen and tenomodulin proteins were also demonstrated at in both differentiated SF and hPL ASCs. In conclusion, we demonstrated firstly the feasibility of both serum and xenogenic-free media tested to culture ASCs moving forward the GMP-compliant approaches for clinical scale expansion of human MSCs needed for therapeutical application of stem cells. Moreover, a combination of CTGF, BMP-12, TGFβ3 and AA factors strongly and rapidly induce human ASCs to differentiate into tenocyte-like cells.

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MiR-378a suppresses tenogenic differentiation and tendon repair by targeting at TGF-?2.

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MiR-378a suppresses tenogenic differentiation and tendon repair by targeting at TGF-β2.

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