Id1 enhances human ovarian cancer endothelial progenitor cell angiogenesis via PI3K/Akt and NF-κB/MMP-2 signaling pathways

Endothelial progenitor cells (EPCs) contribute to tumor angiogenesis and growth. We previously reported that over-expression of an inhibitor of DNA binding/differentiation 1 (Id1) in EPCs can enhance EPC proliferation, migration, and adhesion. In this study, we investigated the role of Id1 in EPC angiogenesis in patients with ovarian cancer and the underlying signaling pathway. Circulating EPCs from 22 patients with ovarian cancer and 15 healthy control subjects were cultured. Id1 and matrix metalloproteinase-2 (MMP-2) expression were analyzed by real-time reverse transcription-polymerase chain reaction (RT-PCR) and western blot. EPC angiogenesis was detected by tube formation assays. Double-stranded DNA containing the interference sequences was synthesized according to the structure of a pGCSIL-GFP viral vector and then inserted into a linearized vector. Positive clones were identified as lentiviral vectors that expressed human Id1 short hairpin RNA (shRNA). Id1 and MMP-2 expression were increased in EPCs freshly isolated from ovarian cancer patients compared to those obtained from healthy subjects. shRNA-mediated Id1 down-regulation substantially reduced EPC angiogenesis and MMP-2 expression. Importantly, transfection of EPCs with Id1 in vitro induced phosphorylation of Akt (p-Akt) via phosphoinositide 3-kinase and increased the expression of MMP-2 via NF-κB. Blockage of both pathways by specific inhibitors (LY294002 and PDTC, respectively) abrogated Id1-enhanced EPC angiogenesis. Id1 can enhance EPC angiogenesis in ovarian cancer, which is mainly mediated by the PI3K/Akt and NF-κB/MMP-2 signaling pathways. Id1 and its downstream effectors are potential targets for treatment of ovarian cancer because of their contribution to angiogenesis.