COVID-19 患者外周血 CD3 T 细胞中 Toll 样受体激动剂和 SARS-CoV-2 抗原对干扰素（IFN）表达的影响。

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.

机构信息

Department of Immunology, School of Medicine, Shahid Beheshti University of Medical Sciences, Tehran, Iran.

Immunology Research Center, Tabriz University of Medical Sciences, Tabriz, Iran.

出版信息

Exp Mol Pathol. 2024 Jun;137:104897. doi: 10.1016/j.yexmp.2024.104897. Epub 2024 Apr 30.

DOI:10.1016/j.yexmp.2024.104897

PMID:38691979

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 CD3IFN-β T cells, and CD3IFN-γ T cells were evaluated by flow cytometry. Interferon (IFN)-β gene expression was assessed by qRT-PCR.

RESULTS

The frequency of CD3IFN-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 CD3IFN-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 CD3IFN-β T cells in sample of severe patients was seen with TLR8 and TLR7 agonists (both p = 0.002). The frequency of CD3IFN-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 CD3IFN-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 CD3T 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.

摘要

背景

Toll 样受体（TLRs）的信号转导启动了针对病毒感染的重要免疫反应。TLRs 在严重急性呼吸综合征冠状病毒 2（SARS-CoV-2）感染中的作用尚未得到充分阐明。因此，我们研究了 TLR 激动剂和 SARS-CoV-2 抗原与体外免疫细胞的相互作用。

材料与方法

共纳入 30 名 2019 年冠状病毒病（COVID-19）患者（15 名重症和 15 名中度）和 10 名年龄和性别匹配的健康对照（HC）。分离外周血单核细胞（PBMC），并用 TLR3、7、8 和 9 激动剂、SARS-CoV-2 的刺突蛋白（SP）和 SP 的受体结合域（RBD）进行激活。通过流式细胞术评估 CD3IFN-β T 细胞和 CD3IFN-γ T 细胞的频率。通过 qRT-PCR 评估干扰素（IFN）-β 基因表达。

结果

与 HC 相比，中度（p<0.0001）和重度（p=0.009）COVID-19 患者的 PBMC 中 CD3IFN-β T 细胞的频率在基线时更高。与 HC 相比，中度患者的 CD3IFN-β T 细胞频率增加最显著的是 TLR8 激动剂和 SP（均 p<0.0001），而重度患者的 CD3IFN-β T 细胞频率增加最显著的是 TLR8 和 TLR7 激动剂（均 p=0.002）。与 HC 相比，中度和重度 COVID-19 患者经 TLR 激动剂刺激后，CD3IFN-γ T 细胞的频率均显著增加（均 p<0.01），但 TLR7 和 TLR8 激动剂除外。TLR8 激动剂不能显著增加重症患者 PBMC 中 CD3IFN-γ T 细胞的频率，但可增加中度疾病患者的细胞频率（p=0.01）。此外，与 HC 相比，TLR8 激动剂刺激后，中度（p<0.0001）和重度（p=0.002）COVID-19 患者的 CD3T 细胞中 IFN-β 基因表达显著上调，而 SP 刺激 T 细胞可显著上调中度疾病患者（p=0.0003）的 IFN-β mRNA 表达，但对重度疾病患者无影响。