Introduction:Cartilage defect(CD)is a common complication in osteoarthritis(OA).Impairment of chondrogenesis and cellular senescence are considered as hallmarks of OA development and caused failure of cartilage repair...Introduction:Cartilage defect(CD)is a common complication in osteoarthritis(OA).Impairment of chondrogenesis and cellular senescence are considered as hallmarks of OA development and caused failure of cartilage repair in most clinical CD cases.Exploring markers for cellular senescence in CD patients might provide new perspectives for osteoarthritic CD patients.In the present study,we aim to explore senescent markers in CD patients with OA to fabricate a senescence-targeted SMSC organoid hydrogel for cartilage repair.Methods:Clinical cartilage samples from cartilage defect patients were collected.Immunofluorescence staining of senescent markers and SA-β-Gal staining were used to detect the senescence state of SMSCs and chondrocytes in cartilage defect and OA patients.MicroRNA expression profiles of SMSC organoids and H2O2-treated SMSC organoids were analyzed and compared with high-throughput microRNA sequencing.Fluorescent in situ hybridization of miRNA were used to determine the expression level of miR-24 in SMSC organoids and cartilage samples.Interaction between miR-24 and its downstream target was analyzed via qRT-PCR,immunofluorescence and luciferase assay.Senescence-targeted miR-24μS/SMSC organoid hydrogel(MSOH)was constructed for cartilage repair.Anti-senescence properties and chondrogenesis were determined in vitro for MSOH.Rats were used to evaluate the cartilage repair capacity of the MSOH hydrogel in vivo.Results:In this study,we found Osteoarthritic cartilage defect patients demonstrated upregulated cellular senescence in joint cartilage.MicroRNA sequencing demonstrated senescence marker miR-24 was negatively associated with cartilage impairment and cellular senescence in osteoarthritic CD patients.Moreover,miR-24 mimics alleviates cellular senescence to promote chondrogenesis by targeting downstream TAOK1.Also,miR-24 downregulated TAOK1 expression and promoted chondrogenesis in SMSC organoids.Senescence-targeted miR-24μS/SMSC organoid hydrogel(MSOH)was constructed and demonstrated superior chondrogenesis in vitro.Animal experiments demonstrated that MSOH hydrogel showed better cartilage repairing effects and better maintained joint function at 24 weeks with low intra-articular inflammatory response after transplantation in rat joint.Single-cell RNA-seq of generated cartilage indicated that implanted MSOH could affect chondrocyte homeostatic state and alter the chondrocyte cluster frequency by regulating cellular glycolysis and OXPHOS,impacting cell cycle and ferroptosis to alleviate cellular senescence and prevent joint degeneration.Conclusion:Osteoarthritic cartilage defect patients demonstrated upregulated cellular senescence in joint cartilage.Senescence marker miR-24 was negatively associated with cartilage impairment in osteoarthritic CD patients.miR-24 attenuates chondrocytes senescence and promotes chondrogenesis in SMSC organoids through targeting TAOK1.Senescence-targeted miR-24 microsphere/SMSC organoid composite hydrogel could successfully repair cartilage defect in osteoarthritic microenvironment via enhanced miR-24/TAOK1 signaling pathway,suggesting MSOH might be a novel therapy for cartilage repair in osteoarthritic CD patients.展开更多
Dear Editor,Human cell-based and personalized in vitro cartilage models are urgently needed for osteoarthritis treatment in pre-clinical regenerative medicine development.Cellular self-assemblies and condensations of ...Dear Editor,Human cell-based and personalized in vitro cartilage models are urgently needed for osteoarthritis treatment in pre-clinical regenerative medicine development.Cellular self-assemblies and condensations of the appropriate stem cells could initiate the formation of transient tissue structures programmed for specific organogenesis processes.1 This recapitulation of developmental events has previously been demonstrated for the formation of cardiac,epithelial and liver organoids.However,there has been very limited progress in the development of human cartilage organoids for osteoarthritis(OA).2 Here,we describe the fabrication of functional bioengineered cartilage organoid suitable for OA treatment.Briefly,agarose microwell inserts for formation of a high number of synovial mesenchymal stromal cell(SMSC)organoids with homogeneous size distribution were created as previously described by Leijten et al.33D-cultured SMSC organoids were generated and phenotypically analyzed for potential applications in OA modeling and treatment(Fig.1a).展开更多
基金funded by the China National Natural Science Funds(No.82172446).We are also grateful for the help and advice from Professor Kerong Dai from Shanghai Ninth People’s Hospital,Shanghai Jiaotong University。
文摘Introduction:Cartilage defect(CD)is a common complication in osteoarthritis(OA).Impairment of chondrogenesis and cellular senescence are considered as hallmarks of OA development and caused failure of cartilage repair in most clinical CD cases.Exploring markers for cellular senescence in CD patients might provide new perspectives for osteoarthritic CD patients.In the present study,we aim to explore senescent markers in CD patients with OA to fabricate a senescence-targeted SMSC organoid hydrogel for cartilage repair.Methods:Clinical cartilage samples from cartilage defect patients were collected.Immunofluorescence staining of senescent markers and SA-β-Gal staining were used to detect the senescence state of SMSCs and chondrocytes in cartilage defect and OA patients.MicroRNA expression profiles of SMSC organoids and H2O2-treated SMSC organoids were analyzed and compared with high-throughput microRNA sequencing.Fluorescent in situ hybridization of miRNA were used to determine the expression level of miR-24 in SMSC organoids and cartilage samples.Interaction between miR-24 and its downstream target was analyzed via qRT-PCR,immunofluorescence and luciferase assay.Senescence-targeted miR-24μS/SMSC organoid hydrogel(MSOH)was constructed for cartilage repair.Anti-senescence properties and chondrogenesis were determined in vitro for MSOH.Rats were used to evaluate the cartilage repair capacity of the MSOH hydrogel in vivo.Results:In this study,we found Osteoarthritic cartilage defect patients demonstrated upregulated cellular senescence in joint cartilage.MicroRNA sequencing demonstrated senescence marker miR-24 was negatively associated with cartilage impairment and cellular senescence in osteoarthritic CD patients.Moreover,miR-24 mimics alleviates cellular senescence to promote chondrogenesis by targeting downstream TAOK1.Also,miR-24 downregulated TAOK1 expression and promoted chondrogenesis in SMSC organoids.Senescence-targeted miR-24μS/SMSC organoid hydrogel(MSOH)was constructed and demonstrated superior chondrogenesis in vitro.Animal experiments demonstrated that MSOH hydrogel showed better cartilage repairing effects and better maintained joint function at 24 weeks with low intra-articular inflammatory response after transplantation in rat joint.Single-cell RNA-seq of generated cartilage indicated that implanted MSOH could affect chondrocyte homeostatic state and alter the chondrocyte cluster frequency by regulating cellular glycolysis and OXPHOS,impacting cell cycle and ferroptosis to alleviate cellular senescence and prevent joint degeneration.Conclusion:Osteoarthritic cartilage defect patients demonstrated upregulated cellular senescence in joint cartilage.Senescence marker miR-24 was negatively associated with cartilage impairment in osteoarthritic CD patients.miR-24 attenuates chondrocytes senescence and promotes chondrogenesis in SMSC organoids through targeting TAOK1.Senescence-targeted miR-24 microsphere/SMSC organoid composite hydrogel could successfully repair cartilage defect in osteoarthritic microenvironment via enhanced miR-24/TAOK1 signaling pathway,suggesting MSOH might be a novel therapy for cartilage repair in osteoarthritic CD patients.
基金This work was funded by the National Key R&D Program of China(No.2018YFB1105600,No.2018YFA0703000)China National Natural Science Funds(No.51631009,No.81802122)+1 种基金Chinese post-doctoral funding(No.2019M661559)the Funds from Shanghai jiao tong university for the Clinical and Translational Research Center for 3D Printing Technology.
文摘Dear Editor,Human cell-based and personalized in vitro cartilage models are urgently needed for osteoarthritis treatment in pre-clinical regenerative medicine development.Cellular self-assemblies and condensations of the appropriate stem cells could initiate the formation of transient tissue structures programmed for specific organogenesis processes.1 This recapitulation of developmental events has previously been demonstrated for the formation of cardiac,epithelial and liver organoids.However,there has been very limited progress in the development of human cartilage organoids for osteoarthritis(OA).2 Here,we describe the fabrication of functional bioengineered cartilage organoid suitable for OA treatment.Briefly,agarose microwell inserts for formation of a high number of synovial mesenchymal stromal cell(SMSC)organoids with homogeneous size distribution were created as previously described by Leijten et al.33D-cultured SMSC organoids were generated and phenotypically analyzed for potential applications in OA modeling and treatment(Fig.1a).