Abstract 631: Complementary Roles for 3-mercaptopyruvate Sulfurtransferase and Cystathionine γ-lyase in Angiogenesis

Abstract

Introduction: Hydrogen sulfide (H 2 S) is a signaling molecule with important actions in the cardiovascular system. The major endogenous sources for H 2 S in the endothelium are cystathionine γ-lyase (CSE) and 3-mercaptopyruvate sulfotransferase (3MST). We have previously shown the CSE-derived H 2 S enhances angiogenesis, however, the role of 3MST in new blood vessel formation remains largely unexplored. Aim: To evaluate the impact of 3MST on angiogenesis and to study the potential mechanisms involved. Methods & Results: 3MST deficient EC exhibited attenuated cell growth, reduced mobility in the scratch wound assay and less sprouting in the fibrin gel bead assay. All of the above-mentioned deficits were less prominent than in cells lacking CSE. In an ex vivo sprouting assay, fewer microvessel-like structures were generated by aortic rings treated with a siRNA directed against 3MST compared to a control oligonucleotide. Similarly, sprouting from 3MST KO aortic rings was less than from wild-type mice. In line with the in vitro responses, angiogenesis in vivo in the Matrigel plug assay and in the retina was reduced in 3MST KO mice, but the decrease was of smaller magnitude compared to that seen with CSE KO mice. The angiogenic switch of EC relies mainly on glycolysis, therefore glycolytic intermediates and TCA metabolites were assessed. Glucose-6P, pyruvate, lactate, α-ketoglutarate and citrate were reduced in CSE KO cells, but not in 3MST KO cells, while fumarate and malate were reduced in 3MST KO, but not CSE KO cells. Moreover, extracellular acidification was reduced in EC lacking CSE, but was only minimally affected in 3MST KO EC. In contrast, oxygen consumption rate was more affected in 3MST KO compared to CSE KO cells. In vivo, 3MST expression was mainly observed in the stalk cells, while CSE was highly expressed in the tip cells. Interestingly, only 3MST ablation resulted in reduced pericyte coverage of retinal vessels. Conclusion: We conclude that 3MST and CSE are both required for angiogenesis. 3MST preserves the quiescence of stalk cells by targeting oxidative phosphorylation, while CSE supports tip cell behavior by enhancing their glycolytic capacity.