Small spheroids for head and neck cartilage tissue engineering.

The demand for cartilage reconstruction in the head and neck region arises frequently due to trauma, malignancies, and hereditary diseases. Traditional tissue engineering produces cartilage from a small biopsy by combining biomaterials and expanded cells. However, this top-down approach is associated with several limitations, including the non-uniform distribution of cells, lack of physiological cell-cell and cell-matrix interactions, and compromised mechanical properties and tissue architecture. The capacity of cells to aggregate into microtissues enables an alternative bottom-up approach to producing cartilage with or without further scaffolding support. Here we explored the optimal conditions for obtaining small spheroids from head and neck cartilage tissues. We used chondrocytes (CCs) and chondroprogenitors (CPCs) isolated from auricular and nasoseptal cartilage to prepare spheroids using ultra-low attachment (ULA) plates or micromass cultures. Different cell densities were tested to estimate the minimal cell number required for optimal spheroid formation. Furthermore, we evaluated the influence of key chondrogenic cytokines, such as transforming growth factor (TGF)-β, connective tissue growth factor (CTGF), and insulin-like growth factor (IGF)-1, on spheroid morphology and the production of cartilage extracellular matrix (ECM) components. Spheroids expressing cartilage markers were formed with 2.5 × 10