Dear Editor, Accumulating clinical data suggest the main causes of death by COVID-19 include respiratory failure and the onset of sepsis. 1 Importantly, sepsis has been observed in nearly all deceased patients. 2-5 It remains elusive how SARS-CoV-2 infection results in viral sepsis in humans. Toll-like receptor 4 (TLR4) mediates antigram-negative bacterial immune responses by recognizing lipopolysaccharide (LPS) from bacteria. 6 We recently found that SARS-CoV-2 infection provoked an anti-bacterial like response at the very early stage of infection via TLR4. However, the identity of the original trigger initiating these abnormal immune responses during SARS-CoV-2 infection is unknown. Previous in silico studies predicted cell surface TLRs, especially TLR4, are most likely to be involved in recognizing molecular patterns, probably spike protein, from SARS-CoV-2 to induce inflammatory responses. 7,8 Consistently, we found that the induction of IL1B by SARS-CoV-2 was completely blocked by TLR4-specific inhibitor Resatorvid (Fig. 1a). Combined with our recent data that TLR4 signaling was activated by SARS-CoV-2, we hypothesized that spike protein could activate TLR4 pathway. A recent study has reported that trimeric SARS-CoV-2 spike proteins are high quality antigens. 9 To this end, we purified the trimeric spike protein (1-1208 aa) (Fig. 1b; Supplementary information, Fig. S1a), as this form of spike protein presents on the surface of viral particle, which most likely interacts with the proteins on the cell surface. Results of the surface plasmon resonance (SPR) assay showed that SARS-CoV-2 spike trimer directly bound to TLR4 with an affinity of~300 nM (Fig. 1b), comparable to many virus-receptor interactions. We then treated THP-1 cells, a cell line of human monocytes, with purified spike protein. IL1B was robustly induced by spike protein in a dose-dependent manner (Fig. 1c), which was comparable to LPS (Supplementary information, Fig. S1b). IL6 was also induced by spike protein (Supplementary information, Fig. S1c). As IL1B induction was much more robust than that of IL6, we chose IL1B production as a marker of immune activation. Moreover, the pseudovirus expressing spike protein can also induce IL1B production (Fig. 1d). Neutrophils also express TLR4 on their cell surface and play an important role in the development of sepsis. We utilized all-trans retinoic acid (ATRA) to treat HL-60 cell (a promyelocytic leukemia cell line), which directed those cells to differentiate into neutrophils. Spike proteins significantly induced IL1B production in HL-60 cells after ATRA treatment (Fig. 1e; Supplementary information, Fig. S1d). We treated THP-1 cells with the N-terminal domain (NTD) or the receptor-binding domain (RBD) of spike protein. Only the trimeric protein could induce IL1B and IL6 (Fig. 1f; Supplementary information, Fig. S1e). To examine if this activation was mediated by TLR4, we treated cells with Resatorvid. Resatorvid greatly blocked induction of IL1B by spike protein and LPS (Fig. 1g). Moreover, spike pro...