Objective: Recently we have reported that cleaved high molecular weight kininogen (HKa) accelerates the onset of endothelial progenitor cells (EPCs) senescence by induction of reactive oxygen species (ROS). However, the mechanisms by which HKa induces production of ROS remain unknown. In this study, we have shown that HKa induces EPC senescence via stimulation of c-Jun N-terminal kinases (JNK)-related pathway. Methods and results: Treatment of human EPCs with HKa for 72 h stimulated JNK phosphorylation at Thr183/Tyr185, and FOXO4 phosphorylation at Thr451, Concomitantly, upregulated the expression of MnSOD at protein and mRNA levels in a concentration-dependent manner. HKa increased intracellular level of H2O2, without affecting the expression of catalase. To narrow down the functional domain of HKa, recombinant proteins of human HK heavy chain (HC, 19-380aa) and light chain (LC, 390-644aa) were generated. HC, but not LC, increased senescence of EPCs and intracellular ROS levels, to a similar extent with HKa. Moreover, HC at 50 nM increased FOXO4 phosphorylation at Thr451 and the protein level of MnSOD in EPCs. Conclusion: These results demonstrate that HKa accelerates the onset of EPC senescence by stimulating JNK/FOXO4/MnSOD pathway, its effect is mediated by the HC. (C) 2014 Elsevier Inc. All rights reserved.