1. Orthopedic Department, The Fifth Affiliated Hospital of Sun Yat-sen University, Zhuhai 519000, China;
2. Guangdong Provincial Key Laboratory of Orthopedics and Traumatology/Orthopedic Research Institute, The First Affiliated Hospital of Sun Yat-sen University, Guangzhou 510075, China;
3. Department of Spine Surgery, The First Affiliated Hospital of Sun Yat-sen University, Guangzhou 510080, China;
4. School of Materials Science and Engineering, South China University of Technology, Guangzhou 510640, China;
5. National Engineering Research Center for Tissue Restoration and Reconstruction, Guangzhou 510006, China;
6. Department of Burn Surgery and Gastroenterology, The First Affiliated Hospital of Nanchang University, Nanchang 330000, China
Although cartilage tissue engineering has been developed for decades, it is still unclear whether angiogenesis was the accompaniment of chondrogenesis in cartilage regeneration. This study aimed to explore the process of anti-angiogenesis during cartilage regenerative progress in cartilage repair extracellular matrix (ECM) materials under Hypoxia. C3H10T1/2 cell line, seeded as pellet or in ECM materials, was added with chondrogenic medium or DMEM medium for 21 days under hypoxia or normoxia environment. Genes and miRNAs related with chondrogenesis and angiogenesis were detected by RT-qPCR technique on Days 7, 14, and 21. Dual-luciferase report system was used to explore the regulating roles of miRNAs on angiogenesis. Results showed that the chondrogenic medium promotes chondrogenesis both in pellet and ECM materials culture. HIF1α was up-regulated under hypoxia compared with normoxia (P?<?0.05). Meanwhile, hypoxia enhanced chondrogenesis. miR-140-5p exhibited higher expression while miR-146b exhibited lower expression. The chondrogenic phenotype was more stabilized in the ECM materials in chondrogenic medium than DMEM medium, with lower VEGFα expression even under hypoxia. Dual-luciferase report assays demonstrated that miR-140-5p directly targets VEGFα by binding its 3′-UTR. Taken together, chondrogenic cytokines, ECM materials and hypoxia synergistically promoted chondrogenesis and inhibited angiogenesis. miR-140-5p played an important role in this process.
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