Tetrahydrobiopterin Prevents Coronary Artery Malformations Induced by Pregestational Diabetes
Authors: Engineer Anish , Lim Yong Jin , Lu Xiangru , Feng Qingping 
Published: June 2021

The heart is the first functional organ to form during embryogenesis. Coronary artery development in the heart begins at embryonic day (E)9.5 and involves coordinated epithelial to mesenchymal transition (EMT) and cell differentiation, which form a complete coronary artery network by E15 in the mouse. Alterations in angiogenic gene expression, cell migration or proliferation can have implications on vasculature and heart function, as seen during diabetic embryopathy. Hyperglycemia‐induced oxidative stress can lead to the inactivation of endothelial nitric oxide synthase (eNOS), vital for heart development. Tetrahydrobiopterin (BH4), an antioxidant, is the co‐factor for eNOS, and when oxidized leads to eNOS deactivation, further increasing the oxidative environment. Treatment with BH4 has been shown to improve vascular endothelial function in diabetes. This study aims to investigate the effect of BH4 on embryonic coronary artery development during pregestational maternal diabetes.Diabetes was induced by streptozotocin (STZ, 50 mg/kg, IPx5) to adult female C57BL/6 mice. BH4 (10 mg/kg/day) was orally administered to pregnant mice. Fetal hearts were collected at E18.5 for coronary artery analysis and 3D reconstruction. Markers of EMT and oxidative stress were examined at E12.5 via immunostaining and qPCR. Immunoblotting was used to determine activity of Akt and eNOS.Marked decreases in coronary artery diameter of both the left and right main branches for the coronary tree were found in hearts from diabetic mothers, which were prevented with BH4 administration. Coronary artery abundance and volume were also decreased in diabetic hearts, and recovered to normal levels with BH4 treatment. A normal branching pattern of the vessels was abolished in hearts from diabetic mothers, and returned to normal with BH4. Notably, BH4 treatment to diabetic dams significantly decreased the incidence of CAMs in their offspring from 47.4 to 23.3%. The expression of molecular regulators of angiogenesis such as Snail1, β‐Catenin, bFGF, Aldh1a2 were decreased with maternal diabetes and reestablished with BH4 supplementation. Additionally, more cell to cell connections, measured through E‐cadherin expression, were found in diabetic hearts and returned to control levels with BH4. The epicardium of hearts from diabetic dams exhibited decreased proliferation and expression of WT1, which were both normalized with BH4 treatment. Furthermore, Akt and eNOS activity, assessed through phosphorylation of both enzymes, was reduced in embryonic hearts from diabetic dams, and was returned to basal levels with BH4 treatment. Finally, oxidative stress, in the form of lipid peroxidation was significantly increased in diabetic hearts and reduced with BH4.BH4 treatment prevents CAMs induced by pregestational diabetes. The importance of BH4‐mediated increase in Akt/eNOS activity and reduction in oxidative stress during cardiac angiogenesis provides insight into factors contributing to CAMs. The therapeutic potential of BH4 during gestation can be extrapolated through these studies.Support or Funding InformationThis study was funding in parts by grants from the Canadian Institutes of Health Research (CIHR) to Q.F.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.