Dialysis bioreactor enables high-density, serum-free expansion of dental pulp-derived mesenchymal stromal cells with enhanced secretome.

Abstract

Dental pulp-derived mesenchymal stromal cells (DPSCs) offer a potential source for both autologous cell-based and cell-free therapies aimed at treating a wide range of diseases. However, current culture practices predominantly rely on serum-based media, which introduce variability and contain potential xenogeneic components, limiting their applicability for therapeutic use. While serum-free media containing growth factors offer an alternative solution to mitigate this issue, they are often inefficient and costly due to the removal of remaining essential growth factors during media replacement. In this study, we developed a scalable dialysis bioreactor that recycles and accumulates endogenous and exogenous growth factors, allowing for the efficient high-density propagation of DPSCs. This bioreactor achieved a cell density of approximately 4.1 × 10⁷ cells/mL DPSCs per batch in a serum-free environment, while maintaining multipotency and differentiation potential after long-term culture. DPSCs grown with this system demonstrated improved proliferation, preserved stemness and enhanced differentiation capacity into odontogenic, osteogenic, chondrogenic, adipogenic and neurogenic lineages. Additionally, the profile of secreted therapeutic proteins related to immunomodulation and tissue regeneration significantly improved compared to manual medium replacement method. By making optimal use of similar amounts of growth factors, the production efficiency significantly increased relative to traditional serum-free culture system. In summary, the dialysis-based bioreactor offers a proof of concept for a scalable platform to improve DPSCs production and optimize their secreted products for therapeutic use. This system overcomes key challenges in serum-free DPSC culture by providing a scalable, efficient and clinically relevant biomanufacturing process.