Inhibition of pyruvate kinase M2 by reactive oxygen species contributes to the development of pulmonary arterial hypertension

Aims: Pulmonary arterial hypertension [1] is a proliferative disorder associated with enhanced proliferation and suppressed apoptosis of pulmonary artery smooth muscle cells (PASMCs). Reactive oxygen species (ROS) is implicated in the development of PAH and regulates the vascular tone and functions. However, which cellular signaling mechanisms are triggered by ROS in PAH is still unknown. Hence, here we wished to characterize the signaling mechanisms triggered by ROS. Methods and Results: By Western blots, we showed that increased intracellular ROS caused inhibition of the glycolytic pyruvate kinase M2 (PKM2) activity through promoting the phosphorylation of PKM2. Monocrotaline (MCT)-induced rats developed severe PAH and right ventricular hypertrophy, with a significant increase in the P-PKM2 and decrease in pyruvate kinase activity which could be attenuated with the treatments of PKM2 activators, FBP and l-serine. The antioxidant NAC, apocynin and MnTBAP had the similar protective effects in the development of PAH. In vitro assays confirmed that inhibition of PKM2 activity could modulate the flux of glycolytic intermediates in support of cell proliferation through the increased pentose phosphate pathway (PPP). Increased ROS and decreased PKM2 activity also promoted the Cav1.2 expression and intracellular calcium. Conclusion: Our data provide new evidence that PKM2 makes a critical regulatory contribution to the PAHs for the first time. Decreased pyruvate kinase M2 activity confers additional advantages to rat PASMCs by allowing them to sustain anti-oxidant responses and thereby support cell survival in PAH. It may become a novel treatment strategy in PAH by using of PKM2 activators.

A comparative study of the effects of hypoxia upon isolated arterial muscle from the rat

Contractile response of rat aorta, mesenteric artery and femoral artery to noradrenaline and potassium chloride were studied under standard and hypoxic conditions and the effect of hypoxia was dependent upon both the vessel and the stimulant. Hypoxia had less effect upon contractions to potassium chloride than those to noradrenaline. The effects of hypoxia on potassium chloride induced responses in different vessels were relatively similar although responses to noradrenaline were vessel dependent. Noradrenaline induced contractions of the femoral artery were most affected by hypoxia whilst those of the mesenteric artery were least affected. Hypoxia changed the well maintained response of the femoral artery to noradrenaline to a transient form; this effect of hypoxia was not evident in the aorta or the mesenteric artery. The aorta and mesenteric artery contracted in calcium free EGTA PSS suggesting that these vessels displayed a release component. Hypoxia reduced the magnitude of this component. The effects of verapamil on noradrenaline and potassium chloride induced responses were investigated and were found to be different to those of hypoxia. Verapamil exerted a greater effect on contractions to potassium chloride than on those to noradrenaline. The effects of hypoxia on 45calcium flux were also vessel dependent. In the mesenteric and femoral arteries hypoxia increased basal 45calcium accumulation. However, the magnitude of noradrenaline stimulated 45calcium accumulation was reduced in the femoral artery and aorta but was unchanged in the mesenteric artery. The effects of hypoxia on 45calcium accumulation were similar to verapamil only in the aorta. The results provide evidence that the effects of hypoxia may arise from alterations in calcium mobilisation processes and that differences between vessels in these processes accounts for the heterogeneity between vessels in their response to hypoxia.