SARS coronavirus spike protein-induced innate immune response occurs via activation of the NF-κB pathway in human monocyte macrophages in vitro

Abstract

A purified recombinant spike (S) protein was studied for its effect on stimulating human peripheral blood monocyte macrophages (PBMC). We examined inflammatory gene mRNA abundances found in S protein-treated PBMC using gene arrays. We identified differential mRNA abundances of genes with functional properties associated with antiviral (CXCL10) and inflammatory (IL-6, IL-8) responses. We confirmed cytokine mRNA increases by real time quantitative(q) RT-PCR or ELISA. We further analyzed the sensitivity and specificity of the prominent IL-8 response. By real time qRT-PCR, S protein was shown to stimulate IL-8 mRNA accumulation in a dose dependent manner while treatment with E protein did not. Also, titration of S protein-specific production and secretion of IL-8 by ELISA showed that the dose of 5.6nM of S produced a significant increase in IL-8 (p=0.003) compared to mock-treated controls. The increase in IL-8 stimulated by a concentration of 5.6 nM of S was comparable to concentrations seen for S protein binding to ACE2 or to neutralizing monoclonal antibody suggesting a physiological relevance. An NF-κB inhibitor, TPCK (N-Tosyl-L-Phenylalanine Chloromethyl Ketone) could suppress IL-8 production and secretion in response to S protein in PBMC and THP-1 cells and in HCoV-229E virus-infected PBMC. Activation and translocation of NF-κB was shown to occur rapidly following exposure of PBMC or THP-1 cells to S protein using a highly sensitive assay for active nuclear NF-κB p65 transcription factor. The results further suggested that released or secreted S protein could activate blood monocytes thru recognition by toll-like receptor (TLR)2 ligand.

Porcine epidemic diarrhea virus infection induces NF-κB activation through the TLR2, TLR3 and TLR9 pathways in porcine intestinal epithelial cells

“…Examples of such viral proteins include the PRRSV structural protein N and the non-structural protein 2 (NSP2) (Fang et al, 2012;Fu et al, 2012;Luo et al, 2008), herpes simplex virus glycoproteins gH/gL and gB (Leoni et al, 2012), Epstein-Barr virus latent membrane protein 1 (Smirnova et al, 2011), HIV envelope glycoprotein gp120 (Bren et al, 2009), SARS-CoV proteins S and N (Dosch et al, 2009;Liao et al, 2005), and hepatitis C virus protein NSP2 (Oem et al, 2008). Virion envelope proteins could initiate NF-kB activation by binding to cellular surface receptors.…”

Pulmonary macrophages: key players in the innate defence of the airways

“…Spike (S) protein, a major structural component of SARS, induces the conversion of B cells into cells that resemble macrophages, when displayed on recombinant baculovirus 33. Dosch et al 34 showed that secreted S protein could activate blood monocytes via TLR2. Furthermore, anti-S antibodies increased infection rates of SARS coronavirus in human monocyte-derived macrophages,35 demonstrating the potential of macrophage-focused therapy during SARS infection…”

SARS coronavirus pathogenesis: host innate immune responses and viral antagonism of interferon

“…Initial models of SARS-CoV innate immune pathogenesis were viral infection of cell lines including Vero E6, Caco-2, and Huh-7 cells, as well as PBMCs; however, these systems may not yield relevant biological information consistent with SARS-CoV infection of pneumocytes, because they are not derived from lung tissues [19][20][21]. Human Airway Epithelial Cultures (HAEs) are primary cell lines of pseudostratified mucocilliary epithelium that replicate the morphological and physiological characteristics of human airways.…”