Chuandong Wang1, Fei Xiao1, Yaokai Gan2, Weien Yuan3, Zhanjing Zhai2, Tuo Jin3, Xiaodong Chen1, Xiaoling Zhang1. 1. Department of Orthopedic Surgery, Xin Hua Hospital Affilliated to Shanghai Jiao Tong University School of Medicine (SJTUSM), Shanghai, China. 2. Shanghai Key Laboratory of Orthopaedic Implant, Department of Orthopaedic Surgery, Shanghai Ninth People's Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, China. 3. School of Pharmacy, Shanghai Jiao tong University, 800 Dongchuan Road, Shanghai, China.
Abstract
BACKGROUND/AIMS: Bone nonunion remains a challenge for orthopaedists. The technological advancements that have been made in precisely silencing target genes have provided promising methods to address this challenge. METHODS: We detected the expression levels of the bone morphogenetic protein (BMP) inhibitors Chordin, Gremlin and Noggin using realtime PCR in bone mesenchymal stem cells (BMSCs) isolated from patients with normal fracture healing and those with bone nonunion. Moreover, we detected the expression of Chordin, Gremlin and Noggin during the osteogenic differentiation of human BMSCs (hBMSCs) using real-time PCR and Western blot. We delivered Chordin siRNA to hBMSCs using a previously reported cationic polymer, polyspermine imidazole-4,5-imine (PSI), as a pH-responsive and non-cytotoxic transfection agent. The apoptosis and cellular uptake efficiency were analysed by flow cytometry. RESULTS: We identified Chordin as the most appropriate potential therapeutic target gene for enhancing the osteogenic differentiation of hBMSCs. Chordin knockdown rescued the osteogenic capacity of hBMSCs isolated from patients with bone nonunion. Highly efficient knockdown of Chordin was achieved in hBMSCs using PSI. Chordin knockdown promoted hBMSC osteogenesis and bone regeneration in vitro and in vivo. CONCLUSIONS: Our results suggest that Chordin is a potential target for improving osteogenesis and bone nonunion therapy and that responsive and non-toxic cationic polyimines such as PSI are therapeutically feasible carriers for the packaging and delivery of Chordin siRNA to hBMSCs.
BACKGROUND/AIMS: Bone nonunion remains a challenge for orthopaedists. The technological advancements that have been made in precisely silencing target genes have provided promising methods to address this challenge. METHODS: We detected the expression levels of the bone morphogenetic protein (BMP) inhibitors Chordin, Gremlin and Noggin using realtime PCR in bone mesenchymal stem cells (BMSCs) isolated from patients with normal fracture healing and those with bone nonunion. Moreover, we detected the expression of Chordin, Gremlin and Noggin during the osteogenic differentiation of human BMSCs (hBMSCs) using real-time PCR and Western blot. We delivered Chordin siRNA to hBMSCs using a previously reported cationic polymer, polyspermine imidazole-4,5-imine (PSI), as a pH-responsive and non-cytotoxic transfection agent. The apoptosis and cellular uptake efficiency were analysed by flow cytometry. RESULTS: We identified Chordin as the most appropriate potential therapeutic target gene for enhancing the osteogenic differentiation of hBMSCs. Chordin knockdown rescued the osteogenic capacity of hBMSCs isolated from patients with bone nonunion. Highly efficient knockdown of Chordin was achieved in hBMSCs using PSI. Chordin knockdown promoted hBMSC osteogenesis and bone regeneration in vitro and in vivo. CONCLUSIONS: Our results suggest that Chordin is a potential target for improving osteogenesis and bone nonunion therapy and that responsive and non-toxic cationic polyimines such as PSI are therapeutically feasible carriers for the packaging and delivery of Chordin siRNA to hBMSCs.
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