Mitigation of Fetal Irradiation Injury from Mid-Gestation Total Body Radiation with Mitochondrial-Targeted GS-Nitroxide JP4-039.

Victims of a radiation terrorist event will include pregnant women and unborn fetuses. Mitochondrial dysfunction and oxidative stress are key pathogenic factors of fetal irradiation injury. The goal of this preclinical study is to investigate the efficacy of mitigating fetal irradiation injury by maternal administration of the mitochondrial-targeted gramicidin S (GS)- nitroxide radiation mitigator, JP4-039. Pregnant female C57BL/6NTac mice received 3 Gy total body ionizing irradiation (TBI) at mid-gestation embryonic day 13.5 (E13.5). Using novel time- and-motion-resolved 4D in utero magnetic resonance imaging (4D-uMRI), we found TBI caused extensive injury to the fetal brain that included cerebral hemorrhage, loss of cerebral tissue, and hydrocephalus with excessive accumulation of cerebrospinal fluid (CSF). Histopathology of the fetal mouse brain showed broken cerebral vessels and elevated apoptosis. Further use of novel 4D Oxy-wavelet MRI capable of probing in vivo mitochondrial function in intact brain revealed significant reduction of mitochondrial function in the fetal brain after 3Gy TBI. This was validated by ex vivo Oroboros mitochondrial respirometry. Maternal administration JP4-039 one day after TBI (E14.5), which can pass through the placental barrier, significantly reduced fetal brain radiation injury and improved fetal brain mitochondrial respiration. This also preserved cerebral brain tissue integrity and reduced cerebral hemorrhage and cell death. As JP4-039 administration did not change litter sizes or fetus viability, together these findings indicate JP4-039 can be deployed as a safe and effective mitigator of fetal radiation injury from mid-gestational in utero ionizing radiation exposure. One Sentence Summary: Mitochondrial-targeted gramicidin S (GS)-nitroxide JP4-039 is safe and effective radiation mitigator for mid-gestational fetal irradiation injury.

Common deletion variants causing protocadherin-α deficiency contribute to the complex genetics of BAV and left-sided congenital heart disease.

Bicuspid aortic valve (BAV) with ~1%-2% prevalence is the most common congenital heart defect (CHD). It frequently results in valve disease and aorta dilation and is a major cause of adult cardiac surgery. BAV is genetically linked to rare left-heart obstructions (left ventricular outflow tract obstructions [LVOTOs]), including hypoplastic left heart syndrome (HLHS) and coarctation of the aorta (CoA). Mouse and human studies indicate LVOTO is genetically heterogeneous with a complex genetic etiology. Homozygous mutation in the Pcdha protocadherin gene cluster in mice can cause BAV, and also HLHS and other LVOTO phenotypes when accompanied by a second mutation. Here we show two common deletion copy number variants (delCNVs) within the PCDHA gene cluster are associated with LVOTO. Analysis of 1,218 white individuals with LVOTO versus 463 disease-free local control individuals yielded odds ratios (ORs) at 1.47 (95% confidence interval [CI], 1.13-1.92; p = 4.2 × 10-3) for LVOTO, 1.47 (95% CI, 1.10-1.97; p = 0.01) for BAV, 6.13 (95% CI, 2.75-13.7; p = 9.7 × 10-6) for CoA, and 1.49 (95% CI, 1.07-2.08; p = 0.019) for HLHS. Increased OR was observed for all LVOTO phenotypes in homozygous or compound heterozygous PCDHA delCNV genotype comparison versus wild type. Analysis of an independent white cohort (381 affected individuals, 1,352 control individuals) replicated the PCDHA delCNV association with LVOTO. Generalizability of these findings is suggested by similar observations in Black and Chinese individuals with LVOTO. Analysis of Pcdha mutant mice showed reduced PCDHA expression at regions of cell-cell contact in aortic smooth muscle and cushion mesenchyme, suggesting potential mechanisms for BAV pathogenesis and aortopathy. Together, these findings indicate common variants causing PCDHA deficiency play a significant role in the genetic etiology of common and rare LVOTO-CHD.

Lamin B2 Levels Regulate Polyploidization of Cardiomyocyte Nuclei and Myocardial Regeneration.

Heart regeneration requires cardiomyocyte proliferation. It is thought that formation of polyploid nuclei establishes a barrier for cardiomyocyte proliferation, but the mechanisms are largely unknown. Here, we show that the nuclear lamina filament Lamin B2 (Lmnb2), whose expression decreases in mice after birth, is essential for nuclear envelope breakdown prior to progression to metaphase and subsequent division. Inactivating Lmnb2 decreased metaphase progression, which led to formation of polyploid cardiomyocyte nuclei in neonatal mice, which, in turn, decreased myocardial regeneration. Increasing Lmnb2 expression promoted cardiomyocyte M-phase progression and cytokinesis and improved indicators of myocardial regeneration in neonatal mice. Inactivating LMNB2 in human iPS cell-derived cardiomyocytes reduced karyokinesis and increased formation of polyploid nuclei. In primary cardiomyocytes from human infants with heart disease, modifying LMNB2 expression correspondingly altered metaphase progression and ploidy of daughter nuclei. In conclusion, Lmnb2 expression is essential for karyokinesis in mammalian cardiomyocytes and heart regeneration.