Publication date: Jun 23, 2025
Recent findings indicate that hyperinflammatory responses to SARS-CoV-2 are major contributors to the severity and fatality of COVID-19. Pattern recognition receptors, particularly Toll-like receptors 2 (TLR2) and 4 (TLR4), have been implicated in detecting SARS-CoV-2 proteins, especially the spike protein. However, the role of viral structural components in triggering innate immune responses remains poorly understood. HEK293 reporter cells, engineered to stably express TLR2 or TLR4, and THP-1 differentiated macrophages were exposed to various spike protein components and SARS-CoV-2 variants. Protein levels of cytokines were detected by ELISA and mRNA expression was evaluated by quantitative real-time RT-PCR. We demonstrate that the S1 subunit and full-length spike protein elicit TLR4-dependent pro-inflammatory responses. TLR4 activation was triggered by the monomeric, but not the homotrimeric, form of the SARS-CoV-2 spike protein. These findings suggest that distinct elements of the SARS-CoV-2 spike protein differentially activate TLR4 signaling pathways, driving innate immune and inflammatory responses.