High mobility group box 1 knockdown inhibits EV71 replication and attenuates cell pyroptosis through TLR4/NF-B/NLRP3 axis

Enterovirus 71 (EV71) is a major causative agent of hand, foot, and mouth disease (HFMD) in children. Nowadays, there are still no effective antiviral drugs for EV71 infection. High mobility group box 1 (HMGB1) is reported to be highly expressed in HFMD patients. However, the role and underlying mechanism of HMGB1 in EV71-associated HFMD are still unclear. HMGB1 expression was detected using RT-qPCR and western blot assays. Loss- and gain-function experiments were performed to evaluate the effects of HMGB1 on EV71-infected cells. The virus titer was examined by TCID50. CCK-8 and flow cytometry assays were applied to detect the cell viability and cell cycle. Oxidative stress was determined by relative commercial kits. HMGB1 level was elevated in the serum of EV71-infected patients with HFMD and EV71-induced RD cells. EV71 infection induced the transfer of HMGB1 from the nucleus into the cytoplasm. HMGB1 knockdown inhibited virus replication, viral protein (VP1) expression and promoted antiviral factor expression. In addition, the inhibition of HMGB1 improved cell viability, protected against S phase arrest, and inhibited EV71-induced cell injury and oxidative stress, whereas HMGB1 overexpression showed the opposite effects. In terms of mechanism, HMGB1 overexpression activated the TLR4/NF-kappa B/NLRP3 signaling pathway and promoted cell pyroptosis. The inhibition of TLR4 and NF-kappa B reversed the effects of HMGB1 overexpression on virus replication, oxidative stress, and pyroptosis. In conclusion, HMGB1 knockdown inhibits EV71 replication and attenuates pyroptosis through TLR4/NF-kappa B/NLRP3 axis. EV71-infected cells induced the transfer of HMGB1 from the nucleus to the cytoplasm, activating the TLR4/NF-kappa B/NLRP3 signaling pathway, thereby promoting cell pyroptosis and oxidative stress, further enhancing viral replication.image