Distinct early and late programs of fibroblast - immune crosstalk govern functional responses to brain injury

PROJECT SUMMARY/ABSTRACT Fibroblasts are stromal cells that are critical to both wound repair and the support and regulation of tissue- resident immune cells; when dysregulated, they drive pathologic scarring and maladaptive immunity. While the Central Nervous System (CNS) is an immunoprivileged site, brain and spinal cord inflammation, injury, and pathologic aging all disrupt this delicate balance, leading to increased immune cell activation and infiltration, and contributing to a range of neuropsychiatric diseases. Although fibroblasts are essentially absent from CNS parenchyma, they are abundant at CNS borders, and perturbations drive the expansion and infiltration of fibroblasts, potentially regulating both acute brain injury and the long-term generation of de novo immune niches, which themselves can alter CNS function. However, the regulatory mechanisms and impacts of CNS fibroblast- immune crosstalk are poorly understood. Our preliminary data show that lesional CNS fibroblasts expand in mouse models of brain injury, transiently adopting a myofibroblast-like state that requires fibroblast-intrinsic TGF? signaling and is spatially correlated with macrophages. Mice with inducible myofibroblast loss have enlarged CNS lesions and sub-acute CNS functional deficits. The myofibroblast program subsides, yet fibroblasts persist for months, intimately associate with late lesional CNS lymphocytes, and are necessary and sufficient to support lymphocytes. Spatial transcriptomics indicate a lesional transition from an early myofibroblast to a late immunoregulatory state. We hypothesize that CNS damage promotes early myofibroblast expansion, requiring macrophages and TGF? activation, to drive wound contraction and limit CNS functional loss, followed by a transition to an immunoregulatory fibroblast state that persists and creates de novo CNS lymphocyte niches. The specific goals of this proposal are to understand how CNS fibroblast-macrophage interactions govern responses to brain damage and establishment of lymphoid niches. Here we will focus on three related questions: (1) How do early injury-associated myofibroblasts regulate CNS damage and functional impairment? (2) What are the cells and signals that regulate CNS fibroblast state transition? (3) How do late CNS immunofibroblasts regulate T-cell accumulation and function? Successful completion of the proposed work will illuminate how brain fibroblasts regulate early CNS damage responses and late immune cell accumulation, laying the foundation for future work in humans and driving therapeutic approaches that target stromal cells to modulate CNS immunity and function.