Previous reports have indicated that <italic>Helicobacter pylori</italic> heat-shock protein 60 (<italic>H. pylori</italic>-HSP60), as an immunodominant antigen, induces interleukin (IL)-8 production in human monocytes. The exact mechanism by which <italic>H. pylori</italic>-HSP60 induces IL-8 production in monocytes has not been fully elucidated. In the present study, the downstream pathway by which <italic>H. pylori</italic>-HSP60 induces IL-8 secretion in human monocytic cell lines was investigated. Intact <italic>H. pylori</italic>, heat-killed <italic>H. pylori</italic> and <italic>H. pylori</italic> recombinant HSP60 (rHpHSP60) all induced the secretion of IL-8 and the activation of mitogen-activated protein kinase (MAPK), extracellular signal-regulated kinase (ERK) and p38, but not c-Jun N-terminal kinase (JNK), up to 24 h in NOMO1 cells. The specific inhibitors PD98059 and U0126 (for ERK1/2 signalling) and SB203580 (for p38 MAPK signalling) down-regulated IL-8 secretion from rHpHSP60-treated NOMO1 cells. An anti-Toll-like receptor (TLR)2 antibody or TLR2 small interfering RNA (siRNA) partially inhibited the secretion of IL-8, and anti-TLR2 antibody also suppressed activation of ERK and p38 MAPK in rHpHSP60-treated NOMO1 cells. These reactions were associated with nuclear factor-<italic>κ</italic>B (NF-<italic>κ</italic>B)-mediated transcriptional activation, since U0126, SB203580 and the anti-TLR2 antibody decreased NF-<italic>κ</italic>B activation. Taken together, the results suggest that ERK and p38 MAPK signalling linked to the TLR2 recognition receptor in human monocytes may be an important pathway in <italic>H. pylori</italic>-HSP60-induced IL-8 secretion.