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Shanhong Fang,1,* Tianmin He,2,* Jiarun Jiang,3 Yongfeng Li,1 Peng Chen1 1Department of Orthopedics, The First Affiliated Hospital of Fujian Medical University, Fuzhou 350005, People’s Republic of China; 2Department of Vascular Surgery, The First Affiliated Hospital of Fujian Medical University, Fuzhou 350005, People’s Republic of China; 3Department of Orthopedics, The Hospital of Changle District, Fuzhou 350005, People’s Republic of China*These authors contributed equally to this workCorrespondence: Peng ChenDepartment of Orthopedics, The First Affiliated Hospital of Fujian Medical University, Taijiang District, Fuzhou 350005, People’s Republic of ChinaEmail 290633950@qq.comPurpose: Steroids are known to inhibit osteogenic differentiation and subsequent bone formation in bone mesenchymal stem cells (BMSCs). However, little is known about the role of BMSC exosomes (Exos) and tRNA-derived small RNAs (tsRNAs) in steroid-induced osteonecrosis of the femoral head (SONFH). The objective of this study was to characterize the tsRNA expression profiles of plasma Exos collected from SONFH patients and healthy individuals using small RNA sequencing and further explore the effect of BMSC Exos carrying specific tsRNAs on osteogenic differentiation.Materials and Methods: Based on insights from small RNA sequencing, five differentially expressed (DE) tsRNAs were selected for quantitative real-time polymerase chain reaction (qRT-PCR). The regulatory networks associated with interactions of the tsRNAs-mRNA-pathways were reconstructed. The osteogenesis and adipogenesis in BMSCs were detected via ALP and oil red O staining methods, respectively.Results: A total of 345 DE small RNAs were screened, including 223 DE tsRNAs. The DE tsRNAs were enriched in Wnt signaling pathway and osteogenic differentiation. We identified five DE tsRNAs, among which tsRNA-10277 was significantly downregulated in plasma Exos of SONFH patients compared to that in hea

Shanhong Fang,1,* Yongfeng Li,2,* Peng Chen1 1Department of Orthopedic Surgery, The First Affiliated Hospital of Fujian Medical University, Fuzhou 350005, China; 2Department of Bone Surgery, Fujian Medical University, Fuzhou 350005, China *These authors contributed equally to this work Background: Animal studies have demonstrated the therapeutic effect of mesenchymal stem cells (MSCs) on osteogenesis, but little is known about the functions of exosomes (Exos) released by bone MSCs (BMSCs). Here, we investigated the effect of BMSC Exos on steroid-induced femoral head necrosis (SFHN) and explored the vital genes involved in this process. Materials and methods: BMSCs were isolated from healthy and SFHN rats. BMSC Exos were isolated using the Exosome Precipitation Kit and characterized by transmission electron microscopy and Western blotting. SFHN BMSCs were incubated with Exos from healthy BMSCs. Osteogenic ability was assessed by oil red O staining and alizarine red staining. Differentially expressed genes (DEGs) induced by Exos were screened using the Osteogenesis RT2 Profiler PCR Array. The effect of upregulated Sox9 was examined using lentivirus-mediated siRNA. Results: The results revealed that BMSC Exos were 100–150 nm in size and expressed CD63. Moreover, BMSC Exo-treated SFHN cells exhibited suppressed adipogenesis compared to model cells. PCR array showed that eleven and nine genes were upregulated and downregulated, respectively, in the BMSC Exo-treated SFHN cells compared to the model group. Among the DEGs, osteogenesis-related genes, including Bmp2, Bmp6, Bmpr1b, Mmp9, and Sox9, may play important roles in SFHN. Furthermore, the DEGs were mainly involved in immune response, osteoblast differentiation, and in the transforming growth factor-β/bone morphogenetic protein signaling pathway. The level of the SOX9 protein was upregulated by Exos, and Sox9 silencing significantly decreased the osteogenic effect of BMSC Exos. Conclusion: Our data