Ammonia mediates mitochondrial uncoupling and promotes glycolysis via HIF-1 activation in human breast cancer MDA-MB-231 cells.

It has been reported that ammonia produced by glutaminolysis activates the HIF-1 pathway in several types of cancer cells, but the underlying mechanisms remain unclear. In this study, the effects of ammonia on the activation of HIF-1 pathway and glycolysis in MDA-MB-231 breast cancer cells were investigated and the underlying mechanisms involved were elucidated. The results showed that NHCl concentration-dependently increased the protein level of HIF-1α and enhanced the transactivation activity of HIF-1 in MDA-MB-231 cells. In addition, NHCl increased the expression of GluT1 and LDHA and promoted aerobic glycolysis by activating the HIF-1 pathway. Further study revealed that NHCl increased the mitochondrial ROS level and decreased the cellular Fe level in MDA-MB-231 cells. Activation of the HIF-1 pathway induced by NHCl was inhibited by addition of the antioxidant NAC or the NADPH oxidase (NOX) inhibitor apocynin, indicating the involvement of the NOX-induced ROS generation. When MDA-MB-231 cells were treated with NHCl, the oxygen consumption of cells increased, followed by the decreased mitochondrial membrane potential and cellular ATP level, indicating the uncoupling of mitochondria. In conclusion, NHCl activated the HIF-1 signaling pathway and promoted aerobic glycolysis in MDA-MB-231 cells, likely through the promotion of mitochondrial ROS release and mitochondrial uncoupling.