Speaker
Description
Introduction
Cartilage tissue regeneration has been significantly advanced through the development of artificial scaffolds, including three-dimensional (3D) electrospun structures. A key challenge in designing in vitro osteochondral models is creating scaffolds with a functional barrier that mimics the native tidemark—separating cartilage and bone—while still permitting cellular communication across tissues. In this study, we aimed to develop 3D electrospun polycaprolactone (PCL) scaffolds with fiber architectures that support cell attachment, proliferation, and tissue development, while incorporating a barrier layer to limit cell migration and intermixing between cartilage and bone cells.
Methods
3D fibrous scaffolds featuring interconnected porous networks with gradient designs were fabricated using a 3D fiber printer (3Df-01C, Bious Labs, Lithuania), which integrates melt electrospinning and fused deposition modeling techniques. Surface hydrophilicity was enhanced via non-thermal plasma (NTP) treatment using a dielectric barrier discharge device (DBD-01-V, Bious Labs).
Chondrocyte cell line C28-I2 was cultured on the PCL scaffolds using CellCrown-24NX culture inserts at varying seeding densities over extended time periods. Cell proliferation and viability were assessed using the CCK-8 assay. Scaffold cross-sections were fixed and analyzed microscopically following NucBlue nuclear staining to evaluate cell distribution and scaffold colonization.
Results
The scaffolds exhibited gradient architectures with fiber diameters ranging from 12 µm to 35 µm and pore sizes from 10 µm to 50 µm. A water contact angle of approximately 50° was achieved with an NTP energy dose of 0.39 J/cm².
C28-I2 chondrocytes were successfully cultured on three different scaffold types over an 8-day period. Cells remained viable and demonstrated progressive proliferation, with the highest proliferation observed in scaffolds featuring a gradual porosity and fiber density gradient. These scaffolds supported a more uniform vertical distribution of cells. In contrast, scaffolds with a more porous upper layer exhibited cell aggregation primarily in the denser lower region. Microscopic analysis of cross-sections confirmed these findings. The tightly woven lower layer of the scaffold functioned effectively as a barrier, preventing cell migration, and mimicking the native tidemark. This barrier design shows promise for supporting spatial separation of chondrocytes and osteocytes in osteochondral constructs.
Conclusions
Electrospun PCL fibrous scaffolds provide a suitable environment for chondrocyte adhesion and proliferation. The incorporation of a dense, tightly woven barrier within a gradient porous structure presents a promising approach for engineering in vitro osteochondral models. These scaffolds could facilitate the development of complex tissue constructs for regenerative medicine applications and preclinical testing.
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