ADSCs Promote Tenocyte Proliferation by Reducing the Methylation Level of lncRNA Morf4l1 in Tendon Injury

Abstract

Objective: Tendons are the special connective tissue that connects bones to muscles and governs joint movement in response to loads passed by muscles. The healing of tendon injuries is still a challenge. In recent years, adipose-derived mesenchymal stem cells (ADSCs) have been increasingly used for tissue regeneration, but the underlying mechanism of tendon injury still remains unclear.Methods: High-throughput sequencing was used to identify a novel lncRNA, whose expression was significantly decreased in injured tendon compared with normal tendon. Furthermore, pyrosequencing, nuclear-cytoplasmic separation, FISH assay and qRT-PCR analysis were used to verify the level of lncRNA methylation in the injured tenocytes. lncRNA was confirmed to promote the proliferation of tenocytes by flow cytometry, wound healing assay, qRT-PCR, and western blot, and the target gene of lncRNA was predicted and verified. To confirm that ADSCs could repair injured tendons, ADSCs and injured tenocytes were co-cultured in vitro, and ADSCs were injected into injured tendons in vitro, respectively.Results: The lncRNA Morf4l1 promoter methylation in injured tendons led to down-regulation of its expression and inhibition of tenocyte proliferation. LncRNA Morf4l1 promoted the expression of TGF-β2 by targeting 3′U of miR-145-5p. After co-cultured ADSCs and injured tenocytes, the expression of lncRNA Morf4l1 was up-regulated, and the proliferation of injured tenocytes in vitro was promoted. The ADSCs were injected into the injured tendon to repair the injured tendon in vivo.Conclusion: This study confirmed that ADSCs promoted tendon wound healing by reducing the methylation level of lncRNA Morf4l1.

Recent Advances in the Use of Stem Cells in Tissue Engineering and Adjunct Therapies for Tendon Reconstruction and Future Perspectives

“…In contrast to this study, the use of miR-378a knock-in transgenic mice showed that the absence of this molecule inhibited collagen and ECM production, tendon differentiation and tendon repair in vitro and in vivo [143]. A study showed that methylation of the promoter of the long non-coding RNA (lncRNA) Morf4l1 in damaged tendons led to a decrease in its expression and inhibition of tenocyte proliferation [144]. This molecule stimulated TGF-β2 via miR-145-5p.…”

Advances in non-coding RNA in tendon injuries

“…It was reported that tendon cell oxidative stress levels were reduced and cell proliferation was enhanced after lncRNA demethylation, suggesting that lncRNA demethylation rescued tendon injury. In addition, they found that adipose-derived MSCs promoted healing after tendon injury by reducing the promoter methylation level of lncRNA Morf4l1 ( Zhao et al, 2022 ). Targeting the lncRNA dnm3os repressed a range of genes associated with fibrogenesis ( Zheng et al, 2018 ).…”

Advanced strategies for constructing interfacial tissues of bone and tendon/ligament

“…251 In a related study, Zhao et al co-cultured ADSCs with injured tenocytes for 48 h, ADSCs clearly promoted proliferation rates and largely reduced levels of oxidative stress in the injured tenocytes via inhibition of methylation LncRNA Morf4l1. 252 Therefore, it is hypothesized that ADSCs mainly target the regeneration of Zone I in the injured enthesis tissue, and exert their positive effect through LncRNA Morf4l1 demethylation.…”