The purpose of this study was to investigate the repair of the osteoarthritis（OA）-induced car- tilage injury by transfecting the new TGF-β3 fusion protein （LAP-MMP-mTGF-β3） with targeted ther- apy function into ...The purpose of this study was to investigate the repair of the osteoarthritis（OA）-induced car- tilage injury by transfecting the new TGF-β3 fusion protein （LAP-MMP-mTGF-β3） with targeted ther- apy function into the bone marrow-derived mesenchymal stem cells （MSCs） in rats. The recombinant of plRES-EGFP-MMP was constructed by combination of DNA encoding MMP enzyme cutting site and eukaryotic expression vector plRES-EGFP. LAP and mTGF-β3 fragments were obtained from rat em- bryos by RT-PCR and inserted into the upstream and downstream of MMP from plRES-EGFP-MMP respectively, so as to construct the recombinant plasmid ofplRES-EGFP-LAP-MMP-mTGF-β3, plRES- EGFP-LAP-MMP-mTGF-β3 was transfected into rat MSCs. The genetically modified MSCs were cul- tured in medium with MMP-1 or not. The transfected MSCs were transplanted in the rat OA models. The OA animal models were surgically induced by anterior cruciate ligament transaction （ACLT）. The pathological changes were observed under a microscope by HE staining, Alcian blue, Safranin-fast Gre- en and graded by Mankin＇s scale, plRES-EGFP-LAP-MMP-mTGF-β3 was successfully constructed by means of enzyme cutting and sequencing, and the mTGF-β3 fusion protein （39 kD） was certified by Western blotting. Those genetically modified MSCs could differentiate into chondrocytes induced by MMP and secrete the relevant-matrix. The transfected MSCs could promote chondrogenesis and matrix production in rat OA models in vivo. It was concluded that a new fusion protein LAP-MMP-mTGF-β3 was constructed successfully by gene engineering, and could be used to repair the OA-induced cartilage injury.展开更多
基金supported by the National Natural Science Foundation of China(No.81101376)
文摘The purpose of this study was to investigate the repair of the osteoarthritis（OA）-induced car- tilage injury by transfecting the new TGF-β3 fusion protein （LAP-MMP-mTGF-β3） with targeted ther- apy function into the bone marrow-derived mesenchymal stem cells （MSCs） in rats. The recombinant of plRES-EGFP-MMP was constructed by combination of DNA encoding MMP enzyme cutting site and eukaryotic expression vector plRES-EGFP. LAP and mTGF-β3 fragments were obtained from rat em- bryos by RT-PCR and inserted into the upstream and downstream of MMP from plRES-EGFP-MMP respectively, so as to construct the recombinant plasmid ofplRES-EGFP-LAP-MMP-mTGF-β3, plRES- EGFP-LAP-MMP-mTGF-β3 was transfected into rat MSCs. The genetically modified MSCs were cul- tured in medium with MMP-1 or not. The transfected MSCs were transplanted in the rat OA models. The OA animal models were surgically induced by anterior cruciate ligament transaction （ACLT）. The pathological changes were observed under a microscope by HE staining, Alcian blue, Safranin-fast Gre- en and graded by Mankin＇s scale, plRES-EGFP-LAP-MMP-mTGF-β3 was successfully constructed by means of enzyme cutting and sequencing, and the mTGF-β3 fusion protein （39 kD） was certified by Western blotting. Those genetically modified MSCs could differentiate into chondrocytes induced by MMP and secrete the relevant-matrix. The transfected MSCs could promote chondrogenesis and matrix production in rat OA models in vivo. It was concluded that a new fusion protein LAP-MMP-mTGF-β3 was constructed successfully by gene engineering, and could be used to repair the OA-induced cartilage injury.
基金Supported by A grant from Ministero Salute-Ricerca Oncologica-RECAM-2006-353005PRIN 2007-prot.2007EN8F7T-004+1 种基金Conven-zione CEINGE-Regione Campania.POR Campania FSE 2007-2013,Project CREMEPRIN 2010-2011-prot.2010K34C45_006
文摘AIM: To investigated the molecular cause of very early-onset ulcerative colitis (UC) in an 18-mo-old affected child.
