A novel electrospun-aligned nanoyarn/three-dimensional porous nanofibrous hybrid scaffold for annulus fibrosus tissue engineering

Abstract

Jun Ma,1,* Yunfei He,1,2,* Xilin Liu,1,* Weiming Chen,3 An Wang,1,4 Chia-Ying Lin,5,6 Xiumei Mo,3 Xiaojian Ye1 1Department of Orthopaedics, Changzheng Hospital, Second Military Medical University, Shanghai, 2Department of Spinal Surgery, Lanzhou General Hospital of Lanzhou Military Command Region, Lanzhou, 3College of Chemistry, Chemical Engineering and Biotechnology, Donghua University, 4Department of Orthopaedics, Shanghai Armed Police Force Hospital, Shanghai, China; 5Structural Tissue Evaluation and Engineering Laboratories, Department of Biomedical Engineering, 6Department of Orthopaedic Surgery, University of Cincinnati, Cincinnati, OH, USA *These authors contributed equally to this work Introduction: Herniation of the nucleus pulposus (NP) because of defects in the annulus fibrosus (AF) is a well-known cause of low back pain. Defects in the AF thus remain a surgical challenge, and efforts have been made to develop new techniques for closure and repair. In this study, we developed an electrospun aligned nanoyarn scaffold (AYS) and nanoyarn/three-dimensional porous nanofibrous hybrid scaffold (HS) for AF tissue engineering. Methods: The AYS was fabricated via conjugated electrospinning, while the aligned nanofibrous scaffold (AFS) was prepared by traditional electrospinning as a baseline scaffold. The HS was constructed by freeze-drying and cross-linking methods. Scanning electron microscopy and mechanical measurement were used to characterize the properties of these scaffolds. Bone marrow derived mesenchymal stem cells (BMSCs) were seeded on scaffolds, and cell proliferation was determined by CCK-8 assay, while cell infiltration and differentiation were assessed by histological measurement and quantitative real-time polymerase chain reaction, respectively.Results: Morphological measurements showed that AYS presented a relatively better three-dimensional structure with larger pore sizes, higher porosity, and better fibers’ alignment compared to AFS. Mechanical testing demonstrated that the tensile property of AFS and AYS was qualitatively similar to the native AF tissue, albeit to a lesser extent. When BMSCs were seeded and cultured on these scaffolds, the number of cells cultured on HS and AYS was found to be significantly higher than that on AFS and culture plate after 7 days of culture (P<0.05). In addition, cell infiltration was significantly higher in HS when compared with AFS and AYS (P<0.05). A part of BMSCs ingressed into the inner part of AYS upon long-term in vitro culture. No significant difference was observed between AFS and AYS in terms of the median infiltration depth (P>0.05). BMSCs seeded on AYS demonstrated an increased expression of COL1A1, while the expression levels of SOX-9, COL2A1, and Aggrecan were higher in HS compared to other scaffolds (P<0.05). Conclusion: These findings indicate that HS makes a proper scaffold for the AF tissue engineering as it replicates the axial compression and tensile property of AF, thereby providing a better platform for cell infiltration and cell–scaffold interaction. Keywords: electrospinning, nanoyarn, three-dimensional scaffold, cell infiltration, annulus fibrosus, tissue engineering

Keywords

• Electrospinning
• Nanoyarn
• Three-dimensional scaffold
• Cell infiltration
• Annulus fibrosus
• Tissue engineering.