M. Shah1, M. Phillips1, R. Frye1, S. McLean1 1University Of North Carolina – Chapel Hill,Chapel Hill, NC, USA
Introduction: The Slit3 knockout (KO) mouse reliably produces a congenital diaphragmatic hernia (CDH) phenotype, and is associated with right ventricular (RV) hypertension and pulmonary arterial hypertension (PAH). PAH occurs due to abnormal pulmonary vascular development and pulmonary vascular remodeling due to unknown mechanisms. We hypothesize that long-term exposure to PAH will induce evidence of altered right ventricular (RV) physiology.
Methods: Slit3 KO mice were bred at our institution. At 3 months of age, wild type (WT) and KO mice were harvested. RV pressures were measured by cardiac puncture. The hearts were then harvested for RNA isolation and weighed. They were placed in RNA-later solution at 4 degrees C for 24 hours and then frozen at -80 degrees C. Hearts were then thawed and RNA was harvested using Trizol reagent. Real-time quantitative PCR was performed using TaqMan reagents with probes for brain natriuretic peptide (BNP).
Results: Slit3 KO mice with CDH had elevated RV systolic (18.8 vs 34.6) and mean (10.2 vs 16.4) pressures at 3 months of age. RT-PCR of Slit3 KO and WT mouse showed significantly increased expression of BNP in RNA harvest from murine RVs, with mean relative quantification (RQ) normalized to b-actin of 0.114 vs 0.493, p<0.005. This represents a 4.35 fold increase in RV-BNP expression in Slit3 KO mice with CDH. RV weight in KO mice was also significantly increased when compared as a percentage of total body weight (TBW), (0.53% vs 0.72%), p<0.02.
Conclusion: Slit3 KO mice develop PAH, right heart strain, and RV hypertrophy when compared to WT mice. This further validates the importance of the Slit3 KO mouse model of PAH in CDH. The use of a validated mouse model of CDH would provide researchers with a novel tool for the study of PAH in CDH. The use of this model to study the mechanisms of PAH development in CDH, may provide temporal and mechanistic targets for therapeutic interventions.
