Directing the chondrogenic differentiation of human Wharton's jelly mesenchymal stem cells using spider silk-based micropattern

Abstract

Mesenchymal stem cells (MSCs)-based therapy shows promise to treat cartilage degeneration, which lacks the regenerative ability in osteoarthritic patients. Differentiation of MSCs into chondrocytes mainly uses a specific differentiation medium. However, biomechanical induction can direct the chondrogenesis of MSCs without the chondrogenic differentiation medium, particularly with micropattern. Micropattern provides physical and mechanical induction to the cell hence facilitates signal transduction that leads to cell differentiation. Human Wharton's jelly MSCs (HWJ-MSCs) were used as the material due to their low risk of allogenic transplant rejection. In this study, biomechanical induction was done using striped micropatterns of 500 μm and 1000 μm width with spidroin concentrations of 50 μg/ml and 10 mg/ml to determine the effect of cell density on HWJ-MSCs chondrogenesis. This study aimed to develop a non-toxic spider silk protein (spidroin) bioink from Argiope appensa as a biomaterial, produce micropattern using microcontact printing, and determine micropattern width and bioink concentration effect on chondrogenic differentiation markers, specifically glycosaminoglycan (GAG) and collagen type II. GAG content was measured through colorimetric analysis with Alcian Blue, whereas collagen type II expression was visualized using immunocytochemistry. MTT assay showed that the spidroin bioink was non-toxic and improved the viability of HWJ-MSCs. The combination of micropattern 1000 μm and spidroin 50 μg/ml produced the highest GAG content. The expression of collagen type II was detected in all treatment groups. Micropattern 1000 μm was able to induce chondrogenesis by increasing cell density, which played an essential role in the condensation step of chondrogenic differentiation. Spidroin 50 μg/ml was hypothesized to induce chondrogenesis via integrin signaling, influenced by cell spreading and contractility. This study concludes that based on GAG content, the optimal condition to induce chondrogenesis was using a micropattern of 1000 μm width and spidroin concentration of 50 μg/ml.