In response to central nervous system injury or disease, astrocytes become reactive, adopting context-dependent states with altered functions. Certain inflammatory insults induce reactive astrocyte states that lose homeostatic functions and gain harmful outputs, and likely contribute to neuroinflammatory and neurodegenerative diseases. However, the cellular pathways controlling these states are not fully understood. Here, we combined single-cell transcriptomics with CRISPRi screening in human iPSC-derived astrocytes to systematically interrogate inflammatory reactivity. We found that autocrine-paracrine IL-6 and interferon signaling downstream of canonical NF-kappaB activation drove two distinct inflammatory reactive states promoted by and inhibited by STAT3, respectively. Furthermore, these states corresponded with those observed in other experimental contexts, including in vivo, and their markers were upregulated in the human brain in Alzheimer's disease and hypoxic ischemic encephalopathy. These results and the platform we established have the potential to guide the development of therapeutics to selectively modulate different aspects of inflammatory astrocyte reactivity. Competing Interest Statement M. Kampmann has filed a patent application related to CRISPRi and CRISPRa screening (PCT/US15/40449) and serves on the Scientific Advisory Board of Engine Biosciences, Casma Therapeutics, and Cajal Neuroscience, and is an advisor to Modulo Bio and Recursion Therapeutics. None of the other authors declare competing interests. Footnotes * https://www.crisprbrain.org/ * https://kampmannlab.ucsf.edu/inflammatory-reactive-astrocyte-analysis...

CRISPRi screens in human astrocytes elucidate regulators of distinct inflammatory reactive states

10.1101/2021.08.23.457400