Biosciences, College of Health, Medicine and Life Sciences, Brunel University London, Uxbridge UB8 3PH, UK.
School of Biosciences and Technology, Vellore Institute of Technology, Vellore 632014, India.
Viruses. 2023 May 29;15(6):1269. doi: 10.3390/v15061269.
The complement system is a key component of the innate immune response to viruses and proinflammatory events. Exaggerated complement activation has been attributed to the induction of a cytokine storm in severe SARS-CoV-2 infection. However, there is also an argument for the protective role of complement proteins, given their local synthesis or activation at the site of viral infection. This study investigated the complement activation-independent role of C1q and C4b-binding protein (C4BP) against SARS-CoV-2 infection. The interactions of C1q, its recombinant globular heads, and C4BP with the SARS-CoV-2 spike and receptor binding domain (RBD) were examined using direct ELISA. In addition, RT-qPCR was used to evaluate the modulatory effect of these complement proteins on the SARS-CoV-2-mediated immune response. Cell binding and luciferase-based viral entry assays were utilised to assess the effects of C1q, its recombinant globular heads, and C4BP on SARS-CoV-2 cell entry. C1q and C4BP bound directly to SARS-CoV-2 pseudotype particles via the RBD domain of the spike protein. C1q via its globular heads and C4BP were found to reduce binding as well as viral transduction of SARS-CoV-2 spike protein expressing lentiviral pseudotypes into transfected A549 cells expressing human ACE2 and TMPRSS2. Furthermore, the treatment of the SARS-CoV-2 spike, envelope, nucleoprotein, and membrane protein expressing alphaviral pseudotypes with C1q, its recombinant globular heads, or C4BP triggered a reduction in mRNA levels of proinflammatory cytokines and chemokines such as IL-1β, IL-8, IL-6, TNF-α, IFN-α, and RANTES (as well as NF-κB) in A549 cells expressing human ACE2 and TMPRSS2. In addition, C1q and C4BP treatment also reduced SARS-CoV-2 pseudotype infection-mediated NF-κB activation in A549 cells expressing human ACE2 and TMPRSS2. C1q and C4BP are synthesised primarily by hepatocytes; however, they are also produced by macrophages, and alveolar type II cells, respectively, locally at the pulmonary site. These findings support the notion that the locally produced C1q and C4BP can be protective against SARS-CoV-2 infection in a complement activation-independent manner, offering immune resistance by inhibiting virus binding to target host cells and attenuating the infection-associated inflammatory response.
补体系统是先天免疫反应中针对病毒和促炎事件的关键组成部分。在严重的 SARS-CoV-2 感染中，补体过度激活被归因于细胞因子风暴的诱导。然而，鉴于补体蛋白在病毒感染部位的局部合成或激活，也存在其具有保护作用的观点。本研究旨在调查 C1q 和 C4b 结合蛋白（C4BP）在 SARS-CoV-2 感染中的补体激活非依赖性作用。使用直接 ELISA 检测 C1q、其重组球形头部和 C4BP 与 SARS-CoV-2 刺突和受体结合域（RBD）的相互作用。此外，还使用 RT-qPCR 评估了这些补体蛋白对 SARS-CoV-2 介导的免疫反应的调节作用。细胞结合和基于荧光素酶的病毒进入测定用于评估 C1q、其重组球形头部和 C4BP 对 SARS-CoV-2 细胞进入的影响。C1q 和 C4BP 通过刺突蛋白的 RBD 结构域直接结合 SARS-CoV-2 假型颗粒。发现 C1q 通过其球形头部和 C4BP 降低了 SARS-CoV-2 刺突蛋白表达慢病毒假型的结合以及转导到表达人 ACE2 和 TMPRSS2 的转染 A549 细胞中的能力。此外，用 C1q、其重组球形头部或 C4BP 处理表达 SARS-CoV-2 刺突、包膜、核蛋白和膜蛋白的甲病毒假型，可降低表达人 ACE2 和 TMPRSS2 的 A549 细胞中促炎细胞因子和趋化因子（如 IL-1β、IL-8、IL-6、TNF-α、IFN-α 和 RANTES（以及 NF-κB）的 mRNA 水平。此外，C1q 和 C4BP 处理还降低了表达人 ACE2 和 TMPRSS2 的 A549 细胞中 SARS-CoV-2 假型感染介导的 NF-κB 激活。C1q 和 C4BP 主要由肝细胞合成；然而，它们也分别由巨噬细胞和肺泡 II 型细胞在肺部局部产生。这些发现支持这样一种观点，即局部产生的 C1q 和 C4BP 可以通过补体激活非依赖性方式提供对 SARS-CoV-2 感染的免疫抵抗，通过抑制病毒与靶宿主细胞的结合和减弱感染相关的炎症反应来发挥抗病毒作用。
