Effects and mechanism of mesenchymal stem cell osteogenesis-derived exosomes deliver miRNA-222-5p on tendon cell injury repair / 中华骨科杂志

ABSTRACT

Objective:To investigate the repair effects and mechanism of mesenchymal stem cell osteogenesis-derived exosomes deliver miRNA-222-5p on tendon cell injury.Methods:Mesenchymal stem cell exosomes were collected, isolated and characterized. The mice achilles tendon was collected after cutting one week later. The tendon cells were isolated and cultured to obtain a tendon cell injury model (injury group). The subjects were divided into three groups. During the process of osteogenic differentiation, mesenchymal stem cells were co-cultured with tendon cells (co-culture group). Further, miRNA-222-5p mimics were transfected into mesenchymal stem cells, the co-cultured mesenchymal stem cells and tendon cells during the process of osteogenic differentiation (miRNA-222-5p group). Clone formation and Transwell were used to detect the ability of mesenchymal stem cells to secrete exosomes and exosomes transported miRNA-222-5p to repair damaged tendon cells. qPCR, Western-blot and fluorescence staining were used to detect the expression levels of cartilage-related proteins and bone-related proteins on mRNA and protein. Western-blot was used to detect the expression of signal pathway factors.Results:The shape of the exosomes was a typical cup - like structure. The exosome proteins marker was all expressed positively ( P<0.05). After counting the colony formation and Transwell test, the number of cell communities and cell invasion were significantly increased in the co-culture group compared with those in the injury group. The number of those in miRNA-222-5p group were significantly increased compared with those in co-culture group ( P<0.05). The qPCR, Western-blot and fluorescent staining test results showed that the mRNA and protein expression levels of SOX-9, COL2A1, ACAN, BMP2, Runx2, and OPN in the co-culture group were significantly increased compared with the injury group. The mRNA and protein expression levels of the above proteins in the miRNA-222-5p group were significantly increased compared with those in co-culture group ( P<0.05). We also detected the protein expression levels of signal pathway related factors. Western-blot results showed that the expression levels of NF-κB, Erk2, STAT3, STAT5 and Smad2 in the co-culture group were significantly increased compared with those in the injury group. The expression levels of the above factors in the miRNA-222-5p group were significant increased compared with the co-culture group ( P<0.05). Conclusion:The co-culture of mesenchymal stem cells and achilles tendon cells could promote the osteogenic differentiation of achilles tendon cells. Exosomes derived from mesenchymal stem cells could further promote the osteogenic differentiation of achilles tendon cells by transporting miRNA-222-5p. The mechanism may be related to the up-regulation of signal pathway factors in injured tendons, including NF-κB, Erk2, STAT3, STAT5 and Smad2.