Spatiotemporal dynamics of mammalian brain development and neuroinflammation by multimodal tri-omics mapping

Introduction: : Understanding the spatial and temporal dynamics underlying mammalian brain development and neuroinflammation is crucial for deciphering the mechanisms that govern brain function and repair. However, integrated insights into chromatin accessibility, gene expression, and protein distribution within the same tissue context have been lacking. To address this gap, we developed spatial tri-omic technologies that simultaneously profile the epigenome, transcriptome, and proteome at near-cellular resolution. Applying these platforms to mouse and human brain tissues, we mapped the molecular landscapes of cortical development, corpus callosum myelination, and neuroinflammatory responses following demyelination. Our work reveals new principles of spatiotemporal gene regulation, myelination dynamics, and immune cell activation in the central nervous system.

Materials and

Methods: : We developed two spatial tri-omic sequencing approaches: DBiT ARP-seq, which co-profiles chromatin accessibility (ATAC-seq), RNA, and proteins, and DBiT CTRP-seq, which co-profiles histone modification (H3K27me3 CUT&Tag), RNA, and proteins from the same tissue section. Spatial barcoding was achieved using orthogonal microfluidic channels, followed by cDNA and genomic DNA library preparation for next-generation sequencing. Multiplexed immunofluorescence imaging (CODEX) was performed using a 23-plex antibody panel for mouse and a 19-plex panel for human tissues to benchmark cellular architectures. TRIC-DISCO was applied for cleared tissue 3D imaging of microglial markers in the neuroinflammation model. Data integration included generalized additive models (GAMs) for spatial-temporal modeling and label transfer with single-cell references for cell type annotation.

Results, Conclusions, and Discussions:: Spatial tri-omic mapping across mouse postnatal cortex (P0-P21) revealed persistent chromatin accessibility of layer-defining transcription factors (e.g., CUX2, TBR1) despite a marked reduction in RNA and protein expression, suggesting the presence of epigenetic memory. In the corpus callosum, we observed a dynamic, spatially patterned myelination process, with lateral to medial progression coupled to projection neuron maturation. Application to a lysolecithin-induced demyelination model demonstrated a coordinated response involving OPC proliferation, transient activation of CD11c+ microglia, and successful remyelination by 21 days post-lesion. Notably, we discovered activation of microglia not only at the lesion core but also distally along white matter tracts, indicating a previously underappreciated spatial propagation of neuroinflammatory signals. Tri-omic measurements revealed spatially resolved gene regulatory programs for repair, distinct chromatin accessibility, and repressive histone mark dynamics during inflammation and remyelination phases.
Our spatial multi-omics atlas uncovers novel principles in brain development and injury response. Chromatin accessibility serves as a stable developmental memory even as transcription declines during postnatal maturation. Oligodendrocyte lineage progression and myelination are tightly coupled with the maturation state of projection neurons. In neuroinflammation, microglial activation extends far beyond the primary lesion site, suggesting that axonal signaling may coordinate distal immune responses. These findings not only advance our understanding of brain development and repair but also offer critical insights into mechanisms underlying neuroinflammatory diseases such as multiple sclerosis.
This study introduces spatial tri-omics as a transformative framework to dissect complex biological processes across development and disease at unprecedented molecular depth and spatial resolution. By simultaneously capturing chromatin state, transcriptional output, and proteomic context, we reveal the layered regulatory mechanisms that orchestrate brain architecture and function. The identification of epigenetic trailing in cortical neurons and the lateral-to-medial remyelination patterns in the corpus callosum highlights the interplay between neuronal maturation and glial support. Furthermore, the detection of distal microglial activation emphasizes the need to consider long-range inflammatory signaling in CNS pathology. Our platforms open new avenues for exploring spatial epigenomic regulation across diverse biological systems and disease models.

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