title: SARS coronavirus spike protein-induced innate immune response occurs via activation of the NF-kappaB 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 and 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 5.6nM of S was comparable to concentrations seen for S protein binding to ACE2 or to neutralizing monoclonal antibody suggesting a physiological relevance. An NF-kappaB 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-kappaB 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-kappaB p65 transcription factor. The results further suggested that released or secreted S protein could activate blood monocytes through recognition by toll-like receptor (TLR)2 ligand.
authors.affiliation: Department of Microbiology and Immunology, Division of Allergy, Immunology and Rheumatology, State University of New York at Buffalo, Buffalo, NY, United States.
authors.forename: Susan F
authors.initials: SF
authors.lastname: Dosch
authors.forename: Supriya D
authors.initials: SD
authors.lastname: Mahajan
authors.forename: Arlene R
authors.initials: AR
authors.lastname: Collins
mesh_terms: D002460:Cell Line; D002478:Cells, Cultured; D015870:Gene Expression; D006801:Humans; D015850:Interleukin-6 / Q000235:genetics / Q000276:immunology; D016209:Interleukin-8 / Q000235:genetics / Q000276:immunology; D008264:Macrophages / Q000276:immunology*; D008562:Membrane Glycoproteins / Q000276:immunology*; D045473:Severe acute respiratory syndrome-related coronavirus / Q000276:immunology*; D045169:Severe Acute Respiratory Syndrome / Q000235:genetics / Q000276:immunology* / Q000821:virology; D064370:Spike Glycoprotein, Coronavirus; D051996:Transcription Factor RelA / Q000235:genetics / Q000276:immunology*; D014759:Viral Envelope Proteins / Q000276:immunology*
chemical_list: D015850:Interleukin-6; D016209:Interleukin-8; D008562:Membrane Glycoproteins; D064370:Spike Glycoprotein, Coronavirus; D051996:Transcription Factor RelA; D014759:Viral Envelope Proteins; C578557:spike glycoprotein, SARS-CoV; C578553:spike protein, mouse hepatitis virus