Protocol for Tri-culture of hiPSC-Derived Neurons, Astrocytes, and Microglia

• Published in: STAR protocols Vol. 1; no. 3; p. 100190
• Main Authors: Ryan, Sean K., Jordan-Sciutto, Kelly L., Anderson, Stewart A.
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 18.12.2020; Elsevier
• Subjects: Action Potentials; Astrocytes - metabolism; Cell Communication; Cell Culture; Cell Culture Techniques - methods; Cell Differentiation; Cells, Cultured; Coculture Techniques - methods; Humans; Induced Pluripotent Stem Cells - metabolism; Microglia - metabolism; Neurons - metabolism; Neuroscience; Protocol; Stem Cells; Stem Cells; Cell Culture; Neuroscience; Cell Differentiation
• ISSN: 2666-1667; 2666-1667
• DOI: 10.1016/j.xpro.2020.100190

This protocol establishes a tri-culture of hiPSC-derived neurons, astrocytes, and microglia for the study of cellular interactions during homeostasis, injury, and disease. This system allows for mechanistic studies that can identify the roles of individual cell types in disease and injury response in a physiologically relevant, all-human system. This protocol utilizes and modifies prior differentiations. Limitations include the prolonged maturation of human astrocytes and neurons and scalability. For complete details on the use and execution of this protocol, please refer to Ryan et al. (2020). [Display omitted] •30-day tri-culture assembly once iAstrocytes and CMPs are produced•Can integrate disease iPSC lines for one to all three cell types•Reduced and tractable system allowing for mechanistic studies•Allows for better control of cell-type ratios than organoids This protocol establishes a tri-culture of hiPSC-derived neurons, astrocytes, and microglia for the study of cellular interactions during homeostasis, injury, and disease. This system allows for mechanistic studies that can identify the roles of individual cell types in disease and injury response in a physiologically relevant, all-human system. This protocol utilizes and modifies prior differentiations. Limitations include the prolonged maturation of human astrocytes and neurons and scalability.