Effects of toll-like receptor agonists and SARS-CoV-2 antigens on interferon (IFN) expression by peripheral blood CD3+ T cells from COVID-19 patients

Abstract

Background: Signaling by toll-like receptors (TLRs) initiates important immune responses against viral infection. The role of TLRs in severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection is not well elucidated. Thus, we investigated the interaction of TLRs agonists and SARS-COV-2 antigens with immune cells in vitro.

Material & methods: 30 coronavirus disease 2019 (COVID-19) patients (15 severe and 15 moderate) and 10 age and sex-matched healthy control (HC) were enrolled. Peripheral blood mononuclear cells (PBMCs) were isolated and activated with TLR3, 7, 8, and 9 agonists, the spike protein (SP) of SARS-CoV-2, and the receptor binding domain (RBD) of SP. Frequencies of CD3⁺IFN-β⁺ T cells, and CD3⁺IFN-γ⁺ T cells were evaluated by flow cytometry. Interferon (IFN)-β gene expression was assessed by qRT-PCR.

Results: The frequency of CD3⁺IFN-β⁺ T cells was higher in PBMCs from moderate (p < 0.0001) and severe (p = 0.009) patients at baseline in comparison with HCs. The highest increase in the frequency of CD3⁺IFN-β⁺ T cells in cell from moderate patients was induced by TLR8 agonist and SP (p < 0.0001 for both) when compared to HC, while, the highest increase of the frequency of CD3⁺IFN-β⁺ T cells in sample of severe patients was seen with TLR8 and TLR7 agonists (both p = 0.002). The frequency of CD3⁺IFN-γ⁺ T cells was significantly increased upon stimulation with TLR agonists in cell from patients with moderate and severe COVID-19, compared with HC (all p < 0.01), except with TLR7 and TLR8 agonists. The TLR8 agonist did not significantly increase the frequency of CD3⁺IFN-γ⁺ T cells in PBMCs of severe patients, but did so in cells from patients with moderate disease (p = 0.01). Moreover, IFN-β gene expression was significantly upregulated in CD3⁺T cells from moderate (p < 0.0001) and severe (p = 0.002) COVID-19 patients, compared to HC after stimulation with the TLR8 agonist, while, stimulation of T cells with SP, significantly up-regulated IFN-β mRNA expression in cells from patients with moderate (p = 0.0003), but not severe disease.

Conclusion: Stimulation of PBMCs from COVID-19 patients, especially patients with moderate disease, with TLR8 agonist and SP increased the frequency of IFN-β-producing T cells and IFN-β gene expression.

Keywords: COVID-19; IFN; SARS-CoV-2; TLR.