Combination Of Antigen-Free Bovine Cancellous Bone With Exosomes Derived From Chm-1 Overexpressing Bone Marrow Mesenchymal Stem Cells For Osteochondral Defect Regeneration

ObjectiveTo investigate the effect of combination of antigen-free bovine cancellous bone(ABCB)with exosomes(Exos)derived from Chm-1 overexpressing bone marrow mesenchymal stem cells(BMSCs)for osteochondral defect regeneration.MethodsIn vitro,BMSCs and ACs from SD rats were extracted and cultured;Lentiviral vector for negative control or its Chm-1 overexpression was transfected into BMSCs;Exos from conditioned medium source of NC-BMSCs(NC-Exos)and from conditioned medium source of Chm-1-BMSCs(Chm-1-Exos)were extracted by overspeed gradient centrifugation;The expression of Chm-1 were evaluated by Western blot assay in NC-Exos and Chm-1-Exos,respectively;IL-1β induced ACs to construct osteoarthritis model(OA-ACs);Dil labeling was used to evaluate the uptake of NC-Exos and Chm-1-Exos by OA-AC;Antigenized bovine cancellous bone(ABCB)were combined with NC-Exos and Chm-1-Exos,respectively;The adhesion of OA-ACs were observed by transmission electron microscop(TEM)in NC-Exos-ABCB and Chm-1-Exos-ABCB,respectively;Cell proliferation and cell activity were evaluated by CCK-8 and Live/Dead assay;Western blot and RT-q PCR analysis were performed to assess the effect of Chm-1-Exos-ABCB on proliferation of ACs in vitro;BAY 11-7082 as the inhibitor was applied to explore the NF-κB signaling pathway involved in the Chm-1-Exos to maintain morphology of ACs;In vivo,MRI was used to evaluate the effect of Chm-1-Exos-ABCB on the repair of osteochondral defects in 6 weeks and 12 weeks after surgery.ResultsNC-Exos and Chm-1-Exos had typical characteristics of Exos.Chm-1 could achieve high expression in Chm-1-Exos.OA-ACs showed better cell activity and showed significant proliferation effect in Chm-1-Exos-ABCB.In addition,Chm-1-Exos maintained phenotypic stability of ACs in the inflammatory environment by inhibiting the NF-κB signaling pathway.In vivo experiments showed that Chm-1-Exos-ABCB had great bio-safety and could significantly promote the repair of osteochondral defects.ConclusionChm-1-Exos-ABCB can significantly promote the proliferation of OA-ACs and maintain the phenotypic stability of ACs in inflammatory environment by inhibiting NF-κB signaling pathway.Chm-1-Exos-ABCB has great bio-safety and can significantly promote the repair of osteochondral defects in OA animal models.Therefore,Chm-1-Exos-ABCB can be used as a new therapeutic strategy for the clinical treatment of osteochondral defects in osteoarthritis.