Multi‐modal sequencing analysis of astrocytes in human and mouse reveals strategically positioned novel reactive sub‐states

Abstract

AbstractBackgroundAstrocytes can have helpful or harmful effects on neuron health and brain function in disease. While they normally provide trophic support to neurons during development and normal functioning, in response to many stimuli their heterogeneous ‘reactive’ responses can alter these functions drastically. Changes in astrocyte function depends on their ‘reactive’ sub‐state. Understanding when and where sub‐states of reactive astrocytes occur, and how these altered functions contribute to disease will pave the way for novel strategies to protect neurons.MethodWe performed combined 10x genomics single cell and spatial transcriptomics in wildtype and Alzheimer’s disease (AD) model mice, combined with single nuclei RNA sequencing of human postmortem non‐symptomatic and AD patient brains.ResultsWith improved capture rates and subsequent powering of astrocyte sequencing we highlight lowly abundant, biologically important, reactive astrocyte sub‐states that are positioned in strategic locations throughout the brain – namely at sites of entry for peripheral immune cells (e.g. adjacent to penetrating vessels in layer I of the cortex, and around the ventricles). Further, we integrate our datasets with previously published scRNAseq and snRNAseq datasets to confirm these small populations exist in other patient populations. Most surprising was that interferon‐responsive reactive astrocytes were present early progression of pathology in the 5xFAD mouse AD model, but not at later time points – suggesting important early (possibly protective) roles for astrocytes early in AD. Additionally, when comparing mouse and human datasets we find most disease pathology‐associated reactive astrocytes are located around strategic points of entry to the brain, and express many inflammation‐responsive transcripts. Probing for ‘modules’ of genes associated with inflammation‐response and reactive sub‐states of microglia and other immune cells highlights putative interactions likely integral for feedback between these two cell types.ConclusionOptimization of astrocyte capture for single cell/nuclei sequencing combined with integration of previously published datasets increased the size of datasets for analysis and power of our analysis. Our data highlight several novel reactive astrocyte sub‐states that warrant additional functional characterization and further investigation.