Abstract 4358975: Maternal Diabetes Tips the Scale: Unmasking Hidden Notch Mutations in Outflow Tract Development

Background: Congenital heart disease (CHD) is the most common birth defect and pregestational maternal diabetes is a modifiable risk factor associated with CHD. It’s estimated that 60 million women of childbearing age currently live with diabetes worldwide. The impact of uncontrolled hyperglycemia in the mother on fetal cardiac development is largely unknown. Significance: Understanding the impact of uncontrolled maternal hyperglycemia during outflow tract development will alleviate the global burden of CHD, particularly in regions with limited access to primary and prenatal health care. Hypothesis: We hypothesize that pregestational maternal diabetes acts synergistically with otherwise non-deleterious Notch pathway mutations in second heart field (SHF) to increase the incidence of outflow tract (OFT) defects, while having no effect on arch defects. Experimental Design: Female C57BL/6 and CD1 mice were injected intraperitoneally with streptozotocin (75 mg/kg) for three consecutive days, and mating occurred when fasting blood glucose levels reached ≥300 mg/dL. To study the effects of genetic mutations, we utilized Mef2c enhancer-driven Cre/loxP system to conditionally knock out the Notch ligand Delta-like ligand 4 ( Dll4) in SHF cells. Statistical differences were determined using one-way analysis of variance (ANOVA), followed by Tukey's post-hoc test. Statistical significance was defined as p< 0.05. Results: Compared to control mice, maternal diabetes group had significantly higher HbA1c levels (6.0 vs. 3.1%) and free cholesterol levels (42.8 vs. 18.7 μg/mL), though no significant differences were observed in HDL, LDL and triacylglyceride levels. In offsprings born to non-diabetic mothers, those with SHF mutation had a slight increase in incidence of isolated VSD (12 vs. 0%) compared to wildtype. In setting of maternal diabetes, mutants exhibited 3.5-fold increase in OFT defects compared to wildtype (45 vs. 13%, p=0.03), while none of the offsprings (with or without mutation) born to non-diabetic mothers had OFT defects. In contrast, arch defects were seen in 50% of mutants born to non-diabetic mothers, and this incidence was not increased in mutant offsprings born to diabetic mothers. Conclusions: Maternal hyperglycemia synergizes with Notch haploinsufficiency to induce OFT defects, while having no effect on arch development. This suggests distinct molecular mechanisms underlying arterial endothelial cell versus cardiac cell specification in second heart field.