| Background: Tendon injury is the injury of the tendon caused by excessive activity of muscle or tendon tissue.The scar left after treatment causes the structure of tendon tissue to become fragile and easy to rupture again.The current treatment can not completely repair the injured tendon.Tendon stem cells(TSCs)which exist in tendon tissue have the advantages of low immunogenicity,rapid proliferation,strong differentiation ability of tendon lineage,and strong application potential in the field of tendon tissue regeneration.As an important part of the extracellular microenvironment,exosomes participate in various physiological and pathological processes in vivo,and its molecular mechanism is unclear.Therefore,further exploration of the effects and mechanisms of exosomes on TSCs has become an important scientific issue in the current tendon injury research.Objective: To explore the influence of TSCs-derived exosomes on the biological functions of TSCs,clarify its molecular mechanism,and provide a new theoretical basis for the treatment of tendon injuries in the future.Methods:(1)Isolation and identification of TSCs: TSCs were isolated from rat patellar tendon;the stem cell surface molecules were detected by flow cytometry.(2)Isolation and identification of exosomes: Exosomes were obtained from the culture supernatant of TSCs by ultracentrifugation;the morphological characteristics of TSCs-derived exosomes were detected by transmission electron microscopy;the particle size of TSCs-derived exosomes was detected by Malvern particle size analyzer;the expression of the specific proteins related to exosomes was detected by western blotting;the uptake of TSCs in exosomes was detected by laser confocal microscope.(3)The analysis of the contents of exosomes derived from TSCs: The protein of TSCs-derived exosome was detected by Western blot.(4)The effect of TSCs-derived exosomes on the cell proliferation and migration of TSCs: the effects of exosomes from TSCs on the proliferation and migration of TSCs were detected by cell proliferation assay and wound healing assay.(5)The molecular mechanism of the effect of TSCs-derived exosomes on the biological functions of TSCs: the expression of the proteins related to cell proliferation and migration was detected by Western blotting.Results:(1)TSCs express CD44 and CD90,but not CD11 b and CD106.(2)TSCs-derived exosomes were hemispherical with a concave surface and the particle size of44-190 nm,containing exosomal marker proteins Alix and CD63.(3)TSCs-derived exosomes contained TGF β.(4)TSCs-derived exosomes promoted the cell proliferation and migration of TSCs.(5)TSCs-derived exosomes increased the expression levels of P-ERK1/2,P-Smad2/3and MMP2 proteins of TSCs,while TGF β receptors Inhibitor(ITD 1)inhibited cell function changes and related protein expression induced by the exosomes derived from TSCs.Conclusion: TGF β of TSCs-derived exosomes promotes cell proliferation and migration of TSCs through ERK1/2 and Smad2/3 signaling pathways.The results of this study are expected to provide a new theoretical basis for tendon repair. |
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