Bioreactor-Mediated Gelatin Methacryloyl Microcarrier Expansion and Human Platelet Lysate Supplementation: A Scalable Mesenchymal Stem Cell Expansion Platform

Abstract

Bioreactor-mediated expansion of human mesenchymal stem cells (hMSCs) is a promising technique for the generation of hMSCs and their secreted products for immunoregulation therapies at scales required for use in the clinic. However, there are gaps in knowledge regarding how hMSCs behave on different microcarrier surfaces during bioreactor culture, and how the type of media supplementation alters their phenotype after expansion. The goals of the experiments presented in this dissertation were two-fold: 1) to characterize the behavior of a renewable source of induced pluripotent stem cell-derived mesenchymal stem cells (ihMSCs) in a scalable bioreactor platform using gelatin-based microcarriers generated with high-throughput microfluidics; and 2) to evaluate ihMSC phenotype in monolayer culture using xenogen-free media containing human platelet lysates (hPL).

Gelatin methacryloyl (GelMA) is a photocrosslinkable derivative of gelatin that has not previously been used for microcarrier-mediated bioreactor expansion of hMSCs for immunomodulatory applications. The immunomodulatory potency of the ihMSCs and their extracellular vesicles (EVs) were evaluated in vitro. The results demonstrate the ability to rapidly produce GelMA microcarriers using a step emulsification microfluidic device and their utility for the expansion of ihMSCs in a 100 mL vertical wheel bioreactor without compromising cell identity, while also enhancing the immunomodulatory potential of cells and harvested EVs. Additionally, the culture of these ihMSCs with hPL-supplemented media improved adipogenic differentiation potential and EV production capabilities compared to ihMSCs culture in media supplemented with FBS while retaining their immunomodulatory potential.