Abstract Body: Introduction
SARS-CoV-2 causes various neurological sequelae in COVID-19 survivors including fatigue and cognitive dysfunction. Endothelial dysfunction, a key mechanism in COVID-19 illness, is also a major risk factor for vascular dementia (VaD). Clinical evidence suggests that reduced nitric oxide (NO) bioavailability is a likely pathogenic factor of endothelial dysfunction in COVID-19 patients, and eNOS levels decline with advancing age, a risk factor for both COVID-19 morbidity and VaD . We hypothesize that endothelial nitric oxide synthase (eNOS) deficiency contributes to brain endothelial dysfunction in SARS-CoV-2 infection and that SARS-CoV-2 infection accelerates the onset of VaD in eNOS-deficient mice.

Methods
6-month-old eNOS+/- (pre-cognitively impaired experimental VaD) and WT male mice were infected with 1X10⁴-pfu mouse-adapted (MA10) SARS-CoV-2 intranasally, and animals were evaluated acutely out to 3-days-postinfection (3dpi) for changes in body weight and clinical signs of illness. Viral copy numbers and nuclear capsid were also quantified in lung and brain. Quantitative PCR and immunofluorescence were used to analyze markers of brain inflammation and senescence.

Results
eNOS+/- infected mice exhibited more disease-associated weight loss (~15%) than WT-infected mice (~5 %). While infected WT and eNOS+/- had comparable pulmonary viral load, neither had detectable virus in the brain. Quantitative PCR analysis of whole brain-isolated mRNA showed increases in multiple proinflammatory mediators such as CCL2 and IL-6 and senescence markers such as p53 and p21 (eNOS^+/- >> WT). Similarly, immunofluorescent analysis showed increased Iba1 (microglia marker) fluorescent intensity in the cortex of infected mice (eNOS^+/- >> WT).

Conclusions
eNOS+/- deficiency, a clinically relevant model of VaD, worsens acute SARS-CoV-2-associated morbidity, neuroinflammation and markers of brain senescence despite comparable pulmonary viral load (to WT-infected animals) and absence of virus in the brain. While the potential effects of SARS-COV-2 on cognitive decline in this model will be assessed in future studies, this is the first experimental evidence demonstrating a link between eNOS and neuropathology associated with COVID-19.