INTRODUCTION

Osteochondral (OC) tissue engineeringhas been proposingbilayeredscaffolds consisting of a cartilaginous and an underlying osseous layers. These scaffolds hold unique composition, structural strength and specific biological properties according to the target tissues.¹Among natural biopolymers, silk fibroin (SF) exhibits high chemical versatility, biocompatibility and tunable mechanical properties.²On the other side, bioresorbable inorganic materials, such as β-tricalcium phosphate(β-TCP)have outstanding osteoconductivity.²The incorporation of ionic dopants into β-TCPenhance osteogenesis and neovascularization of scaffolds.³In this study, we aim to produce novel monolithic bilayered scaffolds composed by a combined enzymatically cross-linked SF hydrogels (HRP-SF) and β-TCP powdersincorporating Sr and Zn, with enhanced viscoelastic properties for OC tissue repair/regeneration.

METHODS

Hierarchical bilayered scaffolds were prepared with HRP-SF for the cartilage layer, and 80/20 (w/w) HRP-SF/undoped and ZnSr-doped β-TCP for the bone layer, using salt-leaching and b freeze-drying techniques (Fig.1a). Physicochemical characterization was assessed through FTIR, XRD, SEM, and micro-CT. Structural integrity was assessed by degradation profile studies and themechanical properties were determined after immersion in PBS. Scaffolds bioactivity was assessed by immersion in SBF up to 30 days. The in vitrocell adhesion and proliferation were evaluated by co-culturing human chondrocytes and human osteoblasts in the scaffolds up to 14 days. Biochemical characterization of ALP activity and GAGs production were also performed. 

RESULTS AND DISCUSSION

The results showed porosity index of 50-60% and highly interconnected poresof 130-140 mm. A homogeneous distribution of the β-TCP into the HRP-SF on the osseouslayer was also observed.The mechanical properties of ZnSr-doped bilayered scaffolds were superior than the undoped scaffolds. Co-cultured cells adhered and proliferated on the bilayered scaffolds (Fig.1b) and higher ALP activity was detected on the monocultured HRP-SF/undoped and ZnSr-doped β-TCP constructs. A positive effect was induced by the co-culture system for GAGs production and deposition (Fig.1c).

CONCLUSION

The structural adaptability and suitable mechanical properties of the hierarchical tissue engineered OC scaffolds, combined with the biological performance achieved using co-culturing systems, make these constructs a viable strategy for OC defects regeneration.

REFERENCES

[1] L.-P.Yan,et al.,Acta biomaterialia,12:227-241,2015.

[2] C.Vepari,D.L.Kaplan,Progress in polymer science,32:991-1007,2007.

[3] S.Pina,et al.,Cells Tissues Organs,204:150,2017.

ACKNOWLEDGMENTS

Portuguese Foundation for Science and Technology for M-ERA-NET/0001/2014 project.

Investigador FCT program (IF/00423/2012, IF/01214/2014 and IF/01285/2015).