This study investigates the signal transduction pathway mediated by extracellular HSP70, focusing on the roles of Toll-like receptors (TLRs) TLR2 and TLR4. The authors demonstrate that HSP70-induced proinflammatory cytokine production is mediated through the MyD88/IRAK/NF-κB pathway and that both TLR2 and TLR4 are involved in this process, acting in a CD14-dependent manner. The study also shows that TLR2 and TLR4 synergize to enhance HSP70-induced cytokine production. These findings suggest that HSP70 can act as a potent adjuvant for immune responses and a powerful inducer of anti-tumor immunity, providing potential targets for developing pharmacological or molecular tools to modulate HSP70's effects in various conditions.This study investigates the signal transduction pathway mediated by extracellular HSP70, focusing on the roles of Toll-like receptors (TLRs) TLR2 and TLR4. The authors demonstrate that HSP70-induced proinflammatory cytokine production is mediated through the MyD88/IRAK/NF-κB pathway and that both TLR2 and TLR4 are involved in this process, acting in a CD14-dependent manner. The study also shows that TLR2 and TLR4 synergize to enhance HSP70-induced cytokine production. These findings suggest that HSP70 can act as a potent adjuvant for immune responses and a powerful inducer of anti-tumor immunity, providing potential targets for developing pharmacological or molecular tools to modulate HSP70's effects in various conditions.