Novel Function of Cancer Stem Cell Marker ALDH1A3 in Glioblastoma: Pro-Angiogenesis through Paracrine PAI-1 and IL-8

SIMPLE SUMMARY: Hyper-angiogenesis is a characteristic of glioblastoma (GBM), and anti-angio-genesis is a crucial strategy to interfere with tumor progression in GBM therapy. We previously found a high expression of ALDH1A3 in proliferating vasculature in GBM patients, which was associated with poor prognosis. The present study generated two ALDH1A3-overexpressing GBM cells (oxGBMs) and demonstrated a potent pro-angiogenesis function of ALDH1A3 under the different conditions of co-culturing oxGBMs with endothelial cells in vitro and in an angiogenesis model in vivo. Moreover, we identified a mechanism underlying the oxALDH1A3-mediated pro-angiogenic effect involving the paracrine PAI-1 and IL-8 derived from oxGBMs. Blockage of PAI-1 or IL-8 hindered the hyper-angiogenesis phenotype resulting from oxALDH1A3. These findings defined a novel function of ALDH1A3 as an angiogenesis promoter in GBM, beyond its role as a well-known cancer stem cell marker, and highlighted ALDH1A3-PAI-1/IL-8 as a novel targeting signaling for future anti-angiogenesis therapy in GBM. ABSTRACT: Hyper-angiogenesis is a typical feature of glioblastoma (GBM), the most aggressive brain tumor. We have reported the expression of aldehyde dehydrogenase 1A3 (ALDH1A3) in proliferating vasculature in GBM patients. We hypothesized that ALDH1A3 may act as an angiogenesis promoter in GBM. Two GBM cell lines were lentivirally transduced with either ALDH1A3 (ox) or an empty vector (ev). The angiogenesis phenotype was studied in indirect and direct co-culture of endothelial cells (ECs) with oxGBM cells (oxGBMs) and in an angiogenesis model in vivo. Angiogenesis array was performed in oxGBMs. RT(2)-PCR, Western blot, and double-immunofluorescence staining were performed to confirm the expression of targets identified from the array. A significantly activated angiogenesis phenotype was observed in ECs indirectly and directly co-cultured with oxGBMs and in vivo. Overexpression of ALDH1A3 (oxALDH1A3) led to a marked upregulation of PAI-1 and IL-8 mRNA and protein and a consequential increased release of both proteins. Moreover, oxALDH1A3-induced angiogenesis was abolished by the treatment of the specific inhibitors, respectively, of PAI-1 and IL-8 receptors, CXCR1/2. This study defined ALDH1A3 as a novel angiogenesis promoter. oxALDH1A3 in GBM cells stimulated EC angiogenesis via paracrine upregulation of PAI-1 and IL-8, suggesting ALDH1A3-PAI-1/IL-8 as a novel signaling for future anti-angiogenesis therapy in GBM.