Developing a novel approach to differentiate human induced pluripotent stem cells into mature astrocytes

In-vitro cellular models of experimental setups have become indispensable for studying different aspects of neurodegenerative diseases, from their pathogenesis to their progression and manifestation through different clinical symptoms. Specifically, human-induced pluripotent stem cells have brought groundbreaking advances in personalized medicine, regenerative medicine, disease modeling, and drug discovery. Hence, hiPSC-derived neurons and glial cells allow researchers to study various neurodegenerative diseases, while brain organoids are ideal for investigating complex neurodevelopmental disorders. The critical role of astrocytes in neurodegenerative diseases has emphasized the need for reliable hiPSC-derived astrocyte models. To date, hiPSC-based modeling of neurodegenerative diseases has primarily concentrated on neurons, as protocols for generating mature astrocytes from hiPSCs were not fully optimized. Thus, the requirement of obtaining an efficient and standardized protocol satisfying certain research expectations has become evident. Furthermore, the use of xenotropic agents, such as FBS, in the differentiation of hiPSCs to obtain mature astrocytes showed adverse effects. In particular, it induced alterations of astrocyte morphology and molecular features, thereby compromising the standards of the studies, even though it was helpful to promote the proliferation and growth of the astrocytes. In this study, starting from a protocol by Perriot et al. (2021), we optimized our method for astrocyte differentiation from hiPSC to maximize the yield of mature astrocytes. For both qualitative and quantitative analysis, we conducted an immunocytochemical method to verify the expression of specific astrocytic markers to support our findings and to demonstrate how different time points in the astrocyte maturation phase influence the number of mature astrocytes, as well as the astrocytes transitioning to full maturity. This study aims to establish a reliable approach for differentiating mature astrocytes from hiPSC, which can be further utilized to investigate astrocyte-related neurodegenerative disorders in vitro, providing expanded opportunities to explore the roles of astrocytes in neurodevelopmental diseases and to test astrocyte-targeted therapeutics.