Proceedings of The Physiological Society

Physiology 2016 (Dublin, Ireland) (2016) Proc Physiol Soc 37, PCA085

Poster Communications

Tetrahydrobiopterin prevents congenital heart defects during pregestational diabetes in mice

X. Lu1, A. Engineer1, T. Saiyin1, Q. Feng1

1. Physiology and Pharmacology, University of Western Ontario, London, Ontario, Canada.

Congenital heart defects (CHDs) are the leading cause of death in the first year of life. Pregestational diabetes increases the risk of CHDs over five fold. Exposure of the embryo to a hyperglycemic environment leads to oxidative stress. Tetrahydrobiopterin (BH4) is a principal co-factor required for endothelial nitric oxide synthase (eNOS) dimerization. In states of oxidative stress, BH4 is oxidized leading to eNOS uncoupling. Our recent studies show that deficiency in eNOS results in CHDs (1-3). The aim of this study was to investigate the effects of BH4 on fetal heart development in mice with pregestational diabetes. Pregestational diabetes was induced by streptozotocin (STZ, 75 mg/kg, i.p. for 3 days) to adult female C57BL/6 mice and treated with BH4 (Kuvanâ 10 mg/kg/day, p.o.). Diabetic pregnant mice without BH4 treatment served as controls. At E18.5, mice were anesthetized with a ketamine/xylazine cocktail and embryos were collected for histological analysis of heart morphology (4, 5). Our data show that pregestational diabetes resulted in a spectrum of CHDs including atrial septal defect (ASD), ventricular septal defects (VSD), atrioventricular septal defect (AVSD), and double outlet right ventricle (DORV). Notably, BH4 treatment significantly decreased the incidence of CHDs from 59.4 to 26.7% and abrogated major CHDs such as VSD, AVSD and DORV. Lineage tracing was performed with a global double fluorescent Mef2C-Cre;mT/mG mouse, where anterior second heart field (SHF) progenitors are labeled with GFP. Fate mapping revealed significantly reduced numbers of GFP+ SHF progenitors contributing to the outflow tract cushions at E9.5, endocardial cushions and ventricular walls at E12.5, indicating defects in proliferation, migration and myocardialization induced by maternal diabetes. Finally, western blot analysis revealed lower eNOS dimer to monomer ratio in the STZ-induced diabetic E12.5 ventricles, which was reversed by BH4 treatment. In conclusion, BH4 treatment recouples eNOS and prevents CHDs. Our study suggests BH4 may have therapeutic potential in the prevention of CHDs in patients with pregestational diabetes.

Where applicable, experiments conform with Society ethical requirements