Distinct and Dynamic Transcriptome Adaptations of iPSC-Generated Astrocytes after Cytokine Stimulation

Astrocytes (ACs) do not only play a role in normal neurogenesis and brain homeostasis, but also in inflammatory and neurodevelopmental disorders. We studied here the different patterns of inflammatory activation triggered by cytokines in human induced pluripotent stem cell (iPSC)-derived ACs. An optimized differentiation protocol provided non-inflamed ACs. These cells reacted to TNF alpha with a rapid translocation of NF kappa B, while AC precursors showed little response. Transcriptome changes were quantified at seven time points (2-72 h) after stimulation with TNF alpha, IFN gamma or TNF alpha plus IFN gamma. TNF alpha triggered a strong response within 2 h. It peaked from 12-24 h and reverted towards the ground state after 72 h. Activation by IFN gamma was also rapid, but the response pattern differed from that of TNF alpha. For instance, several chemokines up-regulated by TNF alpha were not affected by IFN gamma. Instead, MHC-II-related antigen presentation was drastically enhanced. The combination of the two cytokines led to a stronger and more persistent response. For instance, TRIB3 up-regulation by the combination of TNF alpha plus IFN gamma may have slowed NF kappa B inactivation. Additionally, highly synergistic regulation was observed for inflammation modifiers, such as CASP4, and for STAT1-controlled genes. The combination of the cytokines also increased oxidative stress markers (e.g., CHAC1), led to phenotypic changes in ACs and triggered markers related to cell death. In summary, these data demonstrate that there is a large bandwidth of pro-inflammatory AC states, and that single markers are not suitable to describe AC activation or their modulation in disease, development and therapy.