Abstract
In addition to respiratory complications produced by SARS‐CoV‐2, accumulating evidence suggests that some neurological symptoms are associated with the disease caused by this coronavirus. In this study, we investigated the effects of the SARS‐CoV‐2 spike protein S1 stimulation on neuroinflammation in BV-2 microglia. Analyses of culture supernatants revealed an increase in the production of TNF-α, IL-6, IL-1β and iNOS/NO. S1 also increased protein levels of phospho-p65 and phospho-IκBα, as well as enhanced DNA binding and transcriptional activity of NF-κB. These effects of the protein were blocked in the presence of BAY11-7082 (1 µM). Exposure of S1 to BV-2 microglia also increased the protein levels of NLRP3 inflammasome and enhanced caspase-1 activity. Increased protein levels of p38 MAPK was observed in BV-2 microglia stimulated with the spike protein S1 (100 ng/ml), an action that was reduced in the presence of SKF 86,002 (1 µM). Results of immunofluorescence microscopy showed an increase in TLR4 protein expression in S1-stimulated BV-2 microglia. Furthermore, pharmacological inhibition with TAK 242 (1 µM) and transfection with TLR4 small interfering RNA resulted in significant reduction in TNF-α and IL-6 production in S1-stimulated BV-2 microglia. These results have provided the first evidence demonstrating S1-induced neuroinflammation in BV-2 microglia. We propose that induction of neuroinflammation by this protein in the microglia is mediated through activation of NF-κB and p38 MAPK, possibly as a result of TLR4 activation. These results contribute to our understanding of some of the mechanisms involved in CNS pathologies of SARS-CoV-2.
Highlights
The emergence of the severe acute respiratory syndrome coronavirus‐2 (SARS-CoV‐2) in 2019 has resulted in a global pandemic affecting most countries and territories
Luciferase reporter gene assays to evaluate the effects of S1 on transcriptional activity showed that S1 stimulation of BV-2 microglia that were transfected with NF-κB plasmid vector resulted in a significant (p < 0.01) increase in luciferase activity, in comparison with untreated transfected cells
In order to further confirm the roles of NF-κB in S1-induced neuroinflammation, BV-2 microglia were pre-treated with BAY11-7082 prior to stimulation with the protein for 24 h, followed by measurements of the release of pro-inflammatory cytokines (TNF-α, IL-6 and IL-1β) in culture supernatants
Summary
The emergence of the severe acute respiratory syndrome coronavirus‐2 (SARS-CoV‐2) in 2019 has resulted in a global pandemic affecting most countries and territories. The most common complications of illness due to SARS‐CoV‐2 infection are mainly related to respiratory symptoms [2,3,4]. In addition to respiratory complications of SARS-CoV-2 illness, accumulating pieces of evidence suggest that neurological symptoms are associated with COVID-19, the disease caused by the novel coronavirus [6]. These symptoms include headaches, loss of smell, confusion and strokes [7]. The SARS-CoV-2 sub-unit S1 has been shown in a study to promote loss of barrier integrity and trigger a pro-inflammatory response in a 3D model of the human blood–brain barrier [9], suggesting a possible mechanism for the neurological symptoms induced by the coronavirus. Further understanding of the cellular mechanisms involved in neurological consequences of SARS-CoV-2 infection is critical for identifying pharmacological strategies for prevention and treatment
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