Ependymal cell inflammatory activation in response to intracerebral hemorrhage

Neuroinflammation is a central pathological process in secondary brain injury following intracerebral hemorrhage (ICH). While inflammatory responses in perihematomal brain tissue have been extensively investigated, the contribution of ependymal cells to post-ICH neuroinflammatory responses and ventricular pathology remains poorly defined. An autologous blood-induced ICH mouse model was used in combination with single-cell RNA sequencing and spatial transcriptomic analyses to characterize transcriptional reprogramming of ependymal cells. EGFP transgenic mice were used to trace the infiltration of peripheral immune cells. Clinical data was used to assess the association between C3 and hematoma volume. Complement pathway activation was evaluated through integrated single-cell transcriptomic analysis, spatial transcriptomic analysis, immunofluorescence staining, and western blot. Bst2-deficient mice were employed to investigate the mechanisms how complement pathway is regulated in ependymal cells. Single-cell and spatial transcriptomic analysis showed impaired ciliary function and reduced capacity for homeostatic maintenance in ependymal cells. Ventricular asymmetry showed positive association with cognitive impairment at 14-days post-ICH. In parallel, ependymal cells underwent transcriptional reprogramming toward immune and inflammatory phenotypes, accompanied by ipsilateral immune cell infiltration. Cell-cell communication analysis further indicated extensive bidirectional signaling between ependymal cells and multiple immune cell populations, particularly through the complement pathway. Consistent with these findings, elevated C3 expression was detected in ipsilateral ependymal cells. Clinically, circulating C3 levels were elevated in patients with basal ganglia hemorrhage and positively correlated with hematoma volume. Knock out of Bst2 downregulated C3 expression in ependymal cells. Following ICH, ependymal cells undergo a functional transition characterized by loss of cilia-dependent functions and gain of immune and inflammatory properties, including activation of the complement pathway, particularly C3, regulated by BST2.