Garcinia kola and garcinoic acid suppress SARS-CoV-2 spike glycoprotein S1-induced hyper-inflammation in human PBMCs through inhibition of NF-κB activation (preprint)

Symptoms and complications associated with severe SARS-CoV-2 infection such as acute respiratory distress syndrome (ARDS) and organ damage have been linked to SARS-CoV-2 spike glycoprotein S1-induced increased production of pro inflammatory cytokines by immune cells. In this study, the effects of an extract of Garcinia kola seeds and garcinoic acid were investigated in SARS-CoV-2 spike glycoprotein S1-stimulated human PBMCs. Results of ELISA experiments revealed that Garcinia kola extract (6.25, 12.5 and 25 g/mL) and garcinoic acid (1.25, 2.5 and 5 M) significantly reduced SARS-CoV-2 spike glycoprotein S1-induced increased secretion of TNF, IL-6, IL-1{beta} and IL-8 in PBMCs. In-cell western assays showed that pre-treatment with Garcinia kola extract and garcinoic acid reduced elevated expressions of both phospho-p65 and phospho-I{kappa}B proteins, as well as NF-{kappa}B DNA binding capacity and NF-{kappa}B driven luciferase expression following stimulation of PBMCs with spike glycoprotein S1. Furthermore, pre-treatment of PBMCs with Garcinia kola extract prior to stimulation with SARS-CoV-2 spike glycoprotein S1 resulted in reduced damage to adjacent A549 lung epithelial cells. Gas Chromatography-Mass Spectrometry (GCMS) and HPLC-PDA confirmed the presence of garcinoic acid in the Garcinia kola extract used in this study. These results suggest that the seed of Garcinia kola and garcinoic acid are natural products which may possess pharmacological/therapeutic benefits in reducing cytokine storm during the late stage of severe SARS-CoV-2 and other coronavirus infections.

Exaggerated cytokine production in human peripheral blood mononuclear cells by recombinant SARS-CoV-2 spike glycoprotein S1 and its inhibition by dexamethasone (preprint)

An understanding of the pathological inflammatory mechanisms involved in SARS-CoV-2 virus infection is necessary in order to discover new molecular pharmacological targets for SARS-CoV-2 cytokine storm. In this study, the effects of a recombinant SARS-CoV-2 spike glycoprotein S1 was investigated in human peripheral blood mononuclear cells (PBMCs). Stimulation of PBMCs with spike glycoprotein S1 (100 ng/mL) resulted in significant elevation in the production of TNF, IL-6, IL-1{beta} and IL-8. However, pre-treatment with dexamethasone (100 nM) caused significant reduction in the release of these cytokines. Further experiments revealed that S1 stimulation of PBMCs increased phosphorylation of NF-{kappa}B p65 and I{kappa}B, and I{kappa}B degradation. DNA binding of NF-{kappa}B p65 was also significantly increased following stimulation with spike glycoprotein S1. Treatment of PBMCs with dexamethasone (100 nM) or BAY11-7082 (1 {micro}M) resulted in inhibition of spike glycoprotein S1-induced NF-{kappa}B activation. Activation of p38 MAPK by S1 was blocked in the presence of dexamethasone and SKF 86002. CRID3, but not dexamethasone pre-treatment produced significant inhibition of S1-induced activation of NLRP3/caspase-1. Further experiments revealed that S1-induced increase in the production of TNF, IL-6, IL-1{beta} and IL-8 was reduced in the presence of BAY11-7082 and SKF 86002, while CRID3 pre-treatment resulted in the reduction of IL-1{beta} production. These results suggest that SARS-CoV-2 spike glycoprotein S1 stimulated PBMCs to release pro-inflammatory cytokines through mechanisms involving activation of NF-{kappa}B, p38 MAPK and NLRP3 inflammasome. It is proposed that the clinical benefits of dexamethasone in COVID-19 is possibly due to its anti-inflammatory activity in reducing SARS-CoV-2 cytokine storm.

SARS-CoV-2 spike glycoprotein S1 induces neuroinflammation in BV-2 microglia (preprint)

The emergence of SARS-CoV-2 has resulted in a global pandemic. In addition to respiratory complications as a result of SARS-CoV-2 illness, accumulating evidence suggests that neurological and neuropsychiatric symptoms are associated with the disease caused by the virus. In this study, we investigated the effects of the SARS-CoV-2 spike glycoprotein S1 stimulation on neuroinflammation in BV-2 microglia. Analyses of culture supernatants revealed an increase in the production of TNF, IL-6, IL-1{beta} and iNOS/NO. SARS-CoV-2 spike glycoprotein S1 increased protein expressions of phospho-p65 and phospho-I{kappa}B, as well as enhancing DNA binding and transcriptional activity of NF-{kappa}B. Pro-inflammatory effects of the glycoprotein effects were reduced in the presence of BAY11-7082 (1 M). The presence of SARS-CoV-2 spike glycoprotein S1 in BV-2 microglia increased the protein expression of NLRP3, as well as caspase-1 activity. However, pre-treatment with CRID3 (1 M) or BAY11-7082 (1 M) resulted in the inhibition of NLRP3 inflammasome/caspase-1. It was also observed that CRID3 attenuated SARS-CoV-2 spike glycoprotein S1-induced increase in IL-1{beta} production. Increased protein expression of p38 MAPK was observed in BV-2 microglia stimulated with the spike glycoprotein S1, and was reduced in the presence of SKF 86002. These results have provided the first evidence demonstrating SARS-CoV-2 spike S1 glycoprotein-induced neuroinflammation in BV-2 microglia. We propose that promotion of neuroinflammation by this glycoprotein is mediated through activation of NF-{kappa}B, NLRP3 inflammasome and p38 MAPK. These results are significant because of their relevance to our understanding of neurological and neuropsychiatric symptoms observed in patients infected with SARS-CoV-2.