EV-D68 and immune modulation in spinal cord organoids

About This Grant

Enterovirus D68 (EV-D68) was described in 1962 as a respiratory illness but has recently had increased circulation and disease severity. EV-D68 is associated with acute flaccid myelitis (AFM), paralysis in children characterized by spinal cord lesions. There are no specific treatments for AFM and most children have long-lasting neurological deficits. Neuropathogenesis of EV-D68 is likely a combination of virus- and immune- mediated cytotoxicity, though relevant models to investigate this are limited. We recently showed that neonatal mice paralyzed by EV-D68 have abundant T cell recruitment to the spinal cord. When T cells were depleted, mice were protected from paralysis, suggesting a role for T cells in AFM. The role of T cells in human AFM remains unknown. The absence of human models to study T cell interactions in the central nervous system (CNS) hinders progress in identifying viral targets, mechanisms of neural injury, and immune contribution to pathogenesis. Major histocompatibility complexes (MHC) present antigens to T cells to induce the cytotoxic response and effector and memory T cells. Cell surface MHC Class I (MHC-I) is upregulated on neurons and glial cells after CNS injury. However, MHC-I modulation during viral infection of the CNS has not been studied in a multicellular human model. While viral infections are canonically expected to increase surface MHC-I, many viruses downregulate MHC-I as an immune evasion strategy. Our data suggests that EV-D68 infection downregulates, but does not eliminate, MHC- I in infected hSCO. We developed a human spinal cord organoid (hSCO) model for EV-D68 infection from induced pluripotent stem cells (iPSC). hSCO differentiate into multiple cell types of the spinal cord, including neurons and glial cells, and can be infected by EV-D68. Importantly, hSCO are in suspension without Matrigel, a substance that has hindered incorporation of T cells into CNS organoids due to effects on T cell activation and migration. We will utilize our human organoids to understand MHC-I modulation during EV-D68 infection and to investigate interactions between T cells and EV-D68 infected cells of the human CNS. Our overarching hypothesis is that EV-D68 neuropathogenesis is mediated by effects of CD8+ T cells, which we will test by 1) defining the mechanism of EV-D68 downregulation of MHC-I and 2) examining interactions between T cells and EV-D68 infected hSCO. Results will define mechanisms of EV- D68 neurovirulence in a novel human model by elucidating interactions between infected hSCO and the adaptive immune system. Findings will increase understanding of EV-D68 pathogenesis and potentially identify new virus-specific or immune-specific therapeutic targets for AFM.