Immunometabolic and Functional Profiling of Monocytes in Neuroinflammation

Abstract

Monocyte-derived cells (MCs) play crucial roles in central nervous system (CNS) diseases, acting to both enhance and dampen immune-mediated pathology. The relationship between their functional and metabolic profiles in CNS pathologies is not well understood. Traditional paradigms, such as M1/M2 macrophage polarization, fail to capture in vivo heterogeneity. In this thesis, we identified significant transcriptional, metabolic, and functional diversity within CNS MCs through transcriptomic integration and meta-analysis of murine neuropathology models. Disease-specific myeloid responses were reflected in core subpopulations that expanded in different contexts and were driven by specific metabolic processes, highlighting a complex function-associated metabolic model rather than the M1/M2 paradigm.

The M1/M2 paradigm does not effectively connect phenotype and function. Spectral and conventional flow cytometry enable simultaneous investigation of metabolic and functional phenotypes at the single-cell level. We developed and validated a spectral flow cytometry-based metabolic panel that investigates eight metabolic pathways simultaneously. This tool is essential for understanding the metabolic underpinnings of myeloid responses in disease.

In West Nile virus encephalitis, a model of severe neuroinflammation, single-cell RNA sequencing and metabolic flow analysis revealed that brain MCs follow a trajectory from bone marrow through distinct metabolic profiles before differentiating into pro-inflammatory HIF1-α+ and iNOS+ M1-like populations, as well as glycolytically more quiescent antigen-presenting cells. Inhibiting glycolysis with 2-deoxy-D-glucose reduced neuroinflammation and disease signs without increasing viral load, primarily by reducing iNOS+ MC differentiation and cellular migration.

Together, this thesis highlights the complexity of myeloid cell functions and metabolism in CNS diseases, providing new therapeutic targets for severe inflammation.