Gestational intermittent hyperoxia rescues murine genetic congenital heart disease in part.

Cardiac development is a dynamic process, temporally and spatially. When disturbed, it leads to congenital cardiac anomalies that affect approximately 1% of live births. Genetic variants in several loci lead to anomalies, with the transcription factor NKX2-5 being one of the largest. However, there are also non-genetic factors that influence cardiac malformations. We examined the hypothesis that hyperoxia may be beneficial and can rescue genetic cardiac anomalies induced by an Nkx2-5 mutation. Intermittent mild hyperoxia (40% PO2) was applied for 10 h per day to normal wild-type female mice mated with heterozygous Nkx2-5 mutant males from gestational day 8.5 to birth. Hyperoxia therapy reduced excessive trabeculation in Nkx2-5 mutant mice compared to normoxic conditions (ratio of trabecular layer relative to compact layer area, normoxia 1.84 ± 0.07 vs. hyperoxia 1.51 ± 0.04) and frequency of muscular ventricular septal defects per heart (1.53 ± 0.32 vs. 0.68 ± 0.15); however, the incidence of membranous ventricular septal defects in Nkx2-5 mutant hearts was not changed. Nkx2-5 mutant embryonic hearts showed defective coronary vessel organization, which was improved by intermittent mild hyperoxia. The results of our study showed that mild gestational hyperoxia therapy rescued genetic cardiac malformation induced by Nkx2-5 mutation in part.

Defective coronary vessel organization and reduction of VEGF-A in mouse embryonic hearts with gestational mild hypoxia.

BACKGROUND: Vasculature is formed by responding to homeostatic tissue demands including in developing hearts. Hypoxia generally stimulates vascular formation in which vascular endothelial growth factor A (VEGF-A) plays a critical role. Gestational hypoxia increases the risk of low intrauterine growth and low birth weight, both of which are known to increase the risk of the fetus developing cardiovascular defects. In fact, continuous gestational mild hypoxia (14% O2 ) from the mid-embryonic stage causes cardiac anomalies accompanied by a thinning compact layer in mice in vivo. Because coronary vasculature formation is necessary for compact layers to thicken, we hypothesized that defective coronary vessel organization is related to the thinning compact layer under gestational hypoxia conditions. RESULTS: Continuous gestational mild hypoxia (14% O2 ) applied from embryonic day 10.5 (E10.5) reduced the expression of VEGF-A mRNA and proteins by over 60% in E12.5 hearts relative to control normoxic hearts. Formation of CD31-positive vascular plexus, blood islands, and microvessels in embryonic ventricles were stunted by gestational hypoxia compared to control E12.5 hearts. CONCLUSIONS: Our results suggest that mild hypoxia (14% O2 ) does not induce coronary vessel organization or VEGF-A expression in developing mouse hearts, opposing the general effects of hypoxia-triggering vascular organization and VEGF-A expression.

Heterozygous Mylk3 Knockout Mice Partially Recapitulate Human DCM With Heterozygous MYLK3 Mutations.

Backgrounds: Recent studies identified heterozygous variants in MYLK3 gene that encodes cardiac myosin light chain kinase (cMLCK) are related to familial dilated cardiomyopathy (DCM) for the first time. Autosomal dominant traits suggest that pathogenesis of DCM could be related to heterozygous MYLK3 loss-of-function variants (haploinsufficiency). We previously generated and examined homozygous Mylk3 knockout mice that lead to heart failure. It had yet to be examined whether heterozygous Mylk3 knockout mice represent a DCM-like phenotype. Methods and Results: Heterozygous knockout (Mylk3 wild/-) mice were examined regarding cardiac function, heart histology and expression of cMLCK protein and mRNA relative to age-matched wild-type controls (Mylk3 wild/wild). At 4 months of age, cardiac contractility in heterozygous knockout mice was reduced with percent fractional shortening of 23.3 ± 1.2% compared to 30.1 ± 1.8% in control (Mylk3 wild/- vs. Mylk3 wild/wild, n = 9 each). In 4-month-old heterozygous knockout hearts, expression of cMLCK mRNA was expectedly reduced by almost half, however, protein expression was reduced by approximately 75% relative to the control wild-type (Mylk3 wild/- vs. Mylk3 wild/wild, n = 9 each). Isolated ventricular cardiomyocytes from heterozygous knockout mice were larger with increase of short-axis length relative to the cardiomyocytes from control mice. However, increase of heart failure markers as well as interstitial fibrosis were not evident in heterozygous knockout mice compared to controls. Conclusion: Heterozygous Mylk3 knockout mice show mild reduction of cardiac contractility by 4 months of age, and proteins reduced by approximately 75% relative to the control wild-type mice. These mice partly resemble human with the heterozygous MYLK3 mutation, but the reduction in cardiac contractility was milder.