iAstrocytes model cytokine influences on complement expression and neuronal network synchronization

Area of Science:

• Neuroscience
• Stem Cell Biology
• Genetics

Background:

• Astrocytes are crucial for neuronal development, function, and disease.
• Current methods for deriving human astrocytes from pluripotent stem cells (hPSCs) are time-consuming and complex.
• A need exists for rapid and efficient generation of functional human astrocytes for research.

Purpose of the Study:

• To develop a rapid and robust method for generating functional induced astrocytes (iAstrocytes) from hPSCs.
• To establish a genetically tractable platform for studying astrocyte-neuron interactions.
• To investigate the role of astrocytes in modulating neuronal network activity.

Main Methods:

• Developed a genomic safe-harbor knock-in system for inducible expression of astrogenic transcription factors (NFIA, NFIB, SOX9).
• Generated iAstrocytes using NFIB-SOX9 and NFIA-NFIB-SOX9 combinations across multiple hPSC lines.
• Utilized CRISPR interference (CRISPRi) for gene expression knockdown in iAstrocytes.
• Co-cultured iAstrocytes with human induced neurons (iNeurons) and performed pharmacological and genetic perturbations.

Main Results:

• Efficiently generated highly pure iAstrocyte populations expressing astrocyte-specific and synaptogenic genes.
• Demonstrated cytokine-induced expression of complement factors C3 and C4 in iAstrocytes.
• Successfully co-cultured iAstrocytes with iNeurons, promoting synchronized iNeuron network calcium activity.
• Identified specific gap junction proteins mediating astrocyte-induced neuronal network synchronization.

Conclusions:

• The developed iAstrocyte platform offers a rapid and genetically tractable model for astrocyte research.
• This system enables the dissection of human genetic and environmental influences on astrocyte-neuron interactions.
• iAstrocytes provide a valuable tool for studying neurological diseases and developing therapeutic strategies.