The new role of Tryptophan 2,3- dioxygenase in modulating human endothelial cell and endothelial precursor functions

Tryptophan-2,3-dioxygenase (TDO) and Indoleamine-2,3-dioxygenase (IDO1) are the main enzymes involved in tryptophan (Trp) catabolism to kynurenine (Kyn) via the kynurenine pathway (KP), which is known to have a role in the immune regulation. An increased plasma Kyn/Trp ratio has been found in numerous diseases, including cardiovascular diseases. Most Trp is catabolized by IDO1 and its activity has been associated with worse cardiovascular outcomes in patients with coronary artery disease. TDO is mainly expressed in the liver in physiological conditions and recently, TDO-expressing cells were identified as pericytes in numerous types of tumors and in inflammatory pulmonary lesions, suggesting its pro-angiogenic role. Since TDO and IDO1 involvement in angiogenesis is still unknown, we aimed to characterize their role in human umbilical venular endothelial cells (HUVECs) and in human endothelial colony-forming cell function (ECFCs). TDO and IDO1 expression was evaluated by real-time PCR and immunofluorescence. ELISA assay was performed to assess KP activity. Cell proliferation was evaluated by MTT test and in vitro angiogenesis was studied by Geltrex 3D capillary morphogenesis. TDO and IDO1 were expressed by both HUVECs and ECFCs. To evaluate IDO1 and TDO function, cells were treated with epacadostat and 680C91, selective inhibitors for IDO1 and TDO, respectively. TDO inhibition significantly impaired HUVEC proliferation and 3D-tube formation in response to VEGF. VEGF stimulated Akt phosphorylation which was prevented by 680C91. Conversely, ERK1/2 phosphorylation was unaffected by TDO inhibition. Kyn production was increased in response to VEGF and was prevented by the TDO inhibitor. ECFC capillary-like structures were also prevented by 680C91, while IDO1 inhibition was devoid of any effect. Our data demonstrated that HUVECs and ECFCs express the two enzymes of the KP. TDO is involved in in vitro proliferation and capillary morphogenesis, suggesting a putative pathophysiological role in angiogenesis in addition to its well-known immune modulatory effect.