Coronavirus disease 2019 (COVID-19) is a clinical syndrome caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). It is a systemic multiple-organ disease characterized by a broad spectrum of clinical manifestations, including neurological. After recovering from COVID-19, a large proportion of patients suffer from long-lasting post-COVID symptoms, called long-COVID-19 or post-acute COVID-19 syndrome (PACS). Post-COVID symptoms often include neuropsychiatric complications that last for months after the initial infection, strongly suggesting a persisting neurobiological dysfunction. In this in vitro study, researchers from the United States investigated whether the full-length recombinant SARS-CoV-2 spike (S) protein and receptor-binding domain (RBD) stimulate human microglia to release proinflammatory mediators.
Microglia are the resident mononuclear phagocytes of the central nervous system (CNS), which act as the first line of defense in the brain. These cells are highly heterogeneous in the healthy CNS and account for between 5 and 10 percent of total brain cells. Activated microglia have important functions in the CNS, especially in neuroinflammation and neurodegenerative diseases. The postmortem examination of brains from deceased individuals infected with SARS-CoV-2 revealed extensive microglial activation and neuroinflammation associated with brain pathology.
A recent brain magnetic resonance spectroscopy study that investigated the neurometabolites in patients with post-COVID persistent neuropsychiatric symptoms has shown that changes in glutamate/glutamine balance, total N-acetyl-compound levels, and myo-inositol concentration provided evidence of neuronal injury, persistent neuroinflammation and glial dysfunction in brains of individuals diagnosed with post-COVID syndrome. https://discovermednews.com/brain-proton-mr-spectroscopy-changes-in-neurometabolites-in-post-covid-patients/
SARS-CoV-2 is an enveloped, positive-sense, single-stranded RNA virus. Its genome encodes four structural proteins, namely the spike (S), envelope (E), nucleocapsid (N), and membrane (M) protein. The S protein, which appears to be a major pathogenic factor, is a glycosylated homotrimer with each monomer composed of subunits S1 and S2, separated by host cell proteases. The S1 domain comprises the N-terminal domain (NTD), the RBD with a receptor binding motif (RBM), and two C-terminal domains. The RBD is a short immunogenic fragment that facilitates the binding of the S protein to the host cell angiotensin-converting enzyme 2 (ACE2) receptor.
About the study
This in vitro analysis evaluated the influence of the full-length recombinant SARS-CoV-2 S protein and its RBD on the secretion of proinflammatory mediators such as interleukin (IL)-1β, IL-6, IL-18, the chemokine CXCL8, tumor necrosis factor-alpha (TNF-α), matrix metallopeptidase 9 (MMP-9), and S100B protein in the immortalized human microglia-SV40 cell line.
Results
The SARS-CoV-2 S protein stimulated the release of proinflammatory mediators from human microglia in a dose-dependent manner. IL-1β, TNF-α, S100B, CXCL8, IL-6, and MMP9 levels significantly increased after 24 hours of stimulation with various concentrations (1, 5, or 10 ng/ml) of the full-length recombinant S protein, compared to controls. The RBD alone significantly increased TNF-α, IL-18, and S100B levels. These findings were confirmed with the S protein and RBD from two different sources.
The authors then investigated whether the proinflammatory responses triggered by the full-length recombinant S protein or the RBD in human microglia were mediated by Toll-like receptor (TLR)2 or TLR4 signaling or via ACE2. They preincubated human microglia with anti-TLR2, anti-TLR4, and anti-ACE2 antibodies.
Only pretreatment with anti-TLR4 antibody completely suppressed the increase in IL-1β, CXCL8, IL-6, S100B, and MMP9 in human microglia stimulated for 24 hours with the full-length S protein.
Only pretreatment with anti-ACE2 antibody completely suppressed the increase in TNF-α, IL-18, and S100B in human microglia induced with RBD alone.
Conclusion
This study showed that recombinant full-length S protein and RBD alone stimulated human microglia via different receptors to secrete various proinflammatory mediators. According to the authors, these findings confirmed that the SARS-CoV-2 S protein contributes to neuroinflammation via several mechanisms involved in CNS pathologies.
This study has been published on a preprint server and is currently being peer-reviewed.
Journal Reference
Tsilioni I, Theoharides TC. Recombinant SARS-CoV-2 Spike Protein and its Receptor Binding Domain stimulate release of different pro-inflammatory mediators via activation of distinct receptors on human microglia cells. Research Square preprint. (Open Access). https://doi.org/10.21203/rs.3.rs-2394904/v1
