Exploring The Potential Cell Type Specific Interactions of Transactive Response DNA-binding Protein-43 and SARS-CoV-2-N-Protein in COVID-19 Human Brain

The role of neuroinvasion in the development of cognitive and neuropsychiatric disturbances following acute SARS-CoV-2 (Neuro-PASC) infection remains debated. The RNA/DNA-binding protein TDP-43 (transactive response DNA-binding protein of 43 kDa) has been implicated in the pathogenesis of several viral infections and is cleaved by the main SARS-CoV-2 protease Nsp5, leading to neurotoxic TDP-43 fragments in mammalian cells. In our previous study on COVID-19 autopsy brains, we reported evidence of perivascular and vascular phosphoTDP-43 in astrocytes and endothelial cells, destabilization of the blood-brain barrier (BBB), and activation of transcellular and paracellular routes. Furthermore, comparison of Alzheimer’s disease +/- Limbic-predominant age-related TDP-43 encephalopathy (LATE) and SARS-CoV-2 post-mortem human brain showed shared features of TDP-43 microvasculopathy and blood-brain barrier disruption, suggesting a potential link between COVID and AD risk. In the present study we explored whether vascular and perivascular phosphoTDP-43 pathology in patients who died of COVID-19 was associated with presence of SARS-CoV-2- N gene and protein viral particles. In this way, detection of any cell type-specific interactions between SARS-CoV-2-N gene/protein and phosphoTDP-43 could further validate our previous studies. To test this, hippocampus and dorsolateral prefrontal cortex (dlPFC) from cognitively normal age- and sex-matched controls, with and without COVID-19, were selected from autopsy cases conducted at Yale between 2014-2018 (non-COVID), and COVID-positive cases conducted between April 2020-2022; (n=10-12 per group). We report the presence of N gene RNA copies by qRT-PCR, where RNA was extracted from formalin fixed paraffin embedded tissues (FFPE). Immunohistochemical and high-resolution confocal microscopy detected the presence of SARS-CoV-2 Virus N protein in the hippocampus and cerebral cortex of COVID-19 brains. Our analysis revealed that pTDP-43 and N protein exhibited either colocalization or distinct expression within the microvessels of brains affected by COVID-19. A direct interaction of N protein with endothelial cells labelled with CD31, correlated with a disorganized or complete loss of VE-CAD expression in microvessels. Also, strong expression of both N protein and phosphoTDP-43 was detected in astrocytes of COVID-19 brains. Within neurons, marked by MAP2, the N protein was found either in the cytoplasm or in nuclear speckles. The observation that the SARS-CoV-2 N protein undergoes nuclear import, and trafficking suggests that it may play a role in modulating host RNA metabolism pathways or potentially use or even usurp the host’s nuclear transport machinery. Therefore, here we report a novel finding on the identification of SARS-CoV-2-N-protein associated phosphoTDP-43 microvasculopathy in COVID-19 human brain. Moreover, neurovascular unit (NVU) cell type specific associations of N protein and phosphoTDP-43 were identified. The identification of N gene RNA copies and N protein confirms neuroinvasion of SARS-CoV-2 virus. This abstract was presented at the American Physiology Summit 2026 and is only available in HTML format. There is no downloadable file or PDF version. The Physiology editorial board was not involved in the peer review process.