Dissecting the signaling pathways regulating the vascular endothelial growth factor (VEGF) in brain tumor cells. (c2017)

Abstract

Angiogenesis, the fabrication of neo-blood vessels (capillaries), is one of the major hallmarks of cancer and is one of the underlying mechanisms for the tumor’s aggressiveness and metastasis. When faced with hypoxic conditions, cancer cells release various growth and angiogenic factors that lead to the activation of endothelial cells (ECs). The activated ECs then evolve into endothelial tubes that mature into capillaries which supply the tumor cells with the necessary nutrients and oxygen for their survival, proliferation and metastasis. Vascular endothelial growth factor (VEGF) is one of the most important angiogenic modulators that has been intensely studied for its potential in treating various angiogenic pathologies. However, the pathway(s) involved in VEGF’s production and secretion have not been fully deciphered yet, at least not for astrocytoma. In this study, we aim to determine the different pathways and signaling proteins involved in VEGF production in astrocytoma. The astrocytomas were subjected to either hypoxic conditions or to stimulation by the epithelial growth factor (EGF). First, we showed, by conducting experiments at different time intervals, that both hypoxia and EGF stimulation lead to an increase in the expression of HIF-1α and VEGF. Then, the time point with the highest VEGF expression was used to carry out the remaining experiments in order to identify the players behind VEGF production in these cells. The knockdown of the Rho GTPases, RhoA and RhoC separately had no effect on VEGF expression neither after hypoxia nor after EGF stimulation. However, when both RhoA and RhoC were knocked down, VEGF expression decreased after EGF stimulation. Moreover, when the astrocytoma cells were subjected to hypoxia at different times, we detected a significant increase in pERK. So, in order to confirm that the MAPK pathway is involved in VEGF expression, we added the MAPK inhibitor U0126 to these cells. U0126 resulted in a decrease in VEGF expression in the EGF stimulated cells as well as viii in the hypoxic cells. Therefore, our results show that RhoA and RhoC, as well as the MAPK pathway are involved in VEGF production downstream from the EGF receptor whereas only the MAPK pathway is involved in VEGF production under hypoxic conditions.