Abstract 11646: Impact of Fetal Hypoxia on Myocardial Molecular Profile in the Lamb Extra-Uterine Environment for Neonatal Development

Abstract

Background: Using the EXTrauterine Environment for Neonatal Development (EXTEND) system to chronically reduce myocardial oxygen delivery (DO 2 ) to levels mimicking human congenital heart disease caused increased myocardial capillarity and cardiomyocyte hypotrophy. However, molecular mechanisms for this have not been evaluated. Methods: Preterm fetal lambs were isolated by cesarean section and supported by EXTEND for 7 days in normoxia (n=5) or hypoxia (n=4) and compared to in utero gestational age matched controls (n=3). Right and left ventricle (RV, LV) molecular profiles were assessed by RNAseq, qPCR, and western blot. Differentially expressed genes (DEGs) were identified with limma/edgeR. BioPlanet enrichment analysis was performed with Enrichr. Results: DO 2 was lower in hypoxic lambs (15.5 vs 23.8 ml/kg/min, p<0.0001). RNAseq revealed no significant DEGs between normoxia and control, while hypoxia significantly altered myocardial gene expression. The directionality was similar for LV and RV (r=0.82, p<0.0001), but more statistically significant in LV (97 vs 7 DEGs, adj p<0.05). Enrichment analysis revealed Hif1-target upregulation. qPCR confirmed upregulation of Hif1 targets and apoptosis activators Bnip3 (RV log 2 fold change (FC) 0.8) and Bnip3l (LV log 2 FC 1.6) and downregulation of the antiapoptotic Angpt1 (RV log 2 FC -1.5; all p<0.05). However, by protein, Bnip3 and Bnip3l were reduced (RV FC 0.6, p<0.0001) and (LV FC 0.8, p=0.0001), Bax was equally reduced (RV and LV FC 0.65, p<0.05), and the proangiogenic/antiapoptotic hormone Angptl4 was increased (FC 1.9 RV and 2.3 LV, p<0.05). In contrast, the proangiogenic/antiapoptotic cadherin receptor Cdh13 was reduced (RV and LV FC 0.5, p<0.05). Finally, Fblim1 , a driver of cardiomyocyte hypertrophy, was decreased in LV and RV by RNAseq (logFC -0.75 and -0.63, adj p<0.05) but increased by protein (LV FC 1.9, p=0.006). Conclusions: The normoxic EXTEND system faithfully recapitulates the myocardial molecular profile of normal lamb gestation. The molecular impact of fetal myocardial hypoxia is heterogeneous and suggests negative feedback loops with alterations in angiogenesis and cell survival/size. Future studies will dissect if these changes are sufficient to alter myocardial structure.