AMPK/p38 MAPK signaling selectively enhances HIF-induced VEGF-A165 expression under hypoxic and low-glucose conditions in HepG2 cells to promote endothelial cell proliferation and migration.

BACKGROUND: Angiogenesis is essential for tumor development and growth. Vascular endothelial growth factor-A (VEGF-A), a key regulator of angiogenesis, has several isoforms; however, their expression patterns and mechanisms of action are not fully understood. In this study, we aimed to investigate the expression patterns of VEGF-A isoforms and their effects on endothelial cell responses related to angiogenesis under hypoxic and low-glucose conditions, which are major features of the tumor microenvironment, using a hepatocellular carcinoma cell line (HepG2).

METHODS: We cultured human liver cancer-derived cell lines under hypoxic and low-glucose conditions and analyzed the expression patterns of VEGF-A isoforms using an original enzyme-linked immunosorbent assay system that could separately detect human VEGF-A121 and VEGF-A165, which we had previously developed.

RESULTS: The addition of low-glucose conditions to a hypoxic environment increased VEGF-A mRNA and protein expression in HepG2 cells, particularly that of VEGF-A165. Since it is known that AMP-activated protein kinase (AMPK)/p38 mitogen-activated protein kinase (p38 MAPK) signaling increases the stability of VEGF-A mRNA, we investigated the involvement of this signal and found that activation of AMPK increased VEGF-A165 protein expression, while inhibition of AMPK and p38 MAPK reduced VEGF-A165 protein expression. Furthermore, the effects of VEGF-A isoforms on human umbilical vein endothelial cells (HUVECs) were examined. VEGF-A165 activated phosphorylation signals in HUVECs and upregulated their proliferation and migration abilities compared to VEGF-A121.

CONCLUSIONS: These findings suggest that AMPK/p38 MAPK signaling selectively enhances hypoxia-inducible factor-induced VEGF-A165 expression in tumors and promotes endothelial cell proliferation and migration, which may contribute to angiogenesis in vivo.