Presentations

Abstract

Introduction: High risk populations, such as children with mitochondrial diseases (MtD), are prone to develop severe morbidity and mortality from viral infections. Children with Leigh syndrome (LS), a severe MtD phenotype, represent a particularly medically vulnerable group. Respiratory infections like influenza A virus (IAV) can trigger life-threatening neurodegenerative events in LS, necessitating investigation into host-viral pathogen interactions in this disorder. At the cellular level, cells affected by LS adopt Warburg metabolism, which is marked by increased glycolysis, pentose phosphate pathway flux, and fatty acid synthesis. These metabolic characteristics are similar to metabolic reprogramming induced during IAV infection. We hypothesized that respiratory epithelial cells affected by LS would facilitate viral pathogenesis due to their baseline metabolism mirroring the alterations induced in IAV-infected cells.

Methods: To elucidate the mechanisms of augmented IAV pathogenesis in LS, we began by examining CRISPR-edited Ndufs4 knock out (KO) murine lung epithelial type I (LET1) cells, a model of LS due to complex I deficiency. Wild type (WT) and Ndufs4 KO cells were infected with a mouse adapted H3N2 subtype of human IAV, which causes mild illness in mice. Host glycosylation patterns are known to be a crucial determinant factor for viral attachment; therefore, we examined surface glycans in Ndufs4 KO cells using fluorescently tagged lectins. To better understand this enhanced viral pathogenesis in vivo, we infected Ndufs4 KO mice with nebulized IAV to induce viral pneumonia.

Results: At 24 hours post-infection, Ndufs4 KO cells displayed markedly increased viral load by RT-PCR for several IAV gene segments, including NS1, NP, PB2, M1, M2, and PA. In addition, at 10 and 20 minutes post-infection, viral attachment was significantly increased in Ndufs4 KO cells relative to WT. Ndufs4 KO cells also exhibited elevated glycoprotein/glycolipid sialylation, the primary receptors for IAV. Following infection, Ndufs4 KO mice demonstrated elevated sialic acid in the lungs by fluorescence microscopy for lectin binding. Lung viral load was also increased, and the mice displayed worse clinical severity scores and amplified weight loss relative to WT. Moreover, we observed systemic cytokine storm in IAV-infected Ndufs4 KO mice, with elevated levels of inflammatory cytokines, including IFN-γ, IP-10, IL-6, and TNF-α.

Conclusion: Increased sialylation may be attributed to metabolic reprogramming in LS cells marked by increased utilization of the hexosamine and sialic acid biosynthetic pathways. Altogether, our results suggest that aberrant glycosylation leads to increased IAV attachment, producing higher viral loads, which provokes host cytokine storm and heightened illness in LS.