Defective parasympathetic innervation is associated with airway branching abnormalities in experimental CDH

Abstract

Developmental mechanisms leading to lung hypoplasia in congenital diaphragmatic hernia (CDH) remain poorly defined. Pulmonary innervation is defective in the human disease and in the rodent models of CDH. We hypothesize that defective parasympathetic innervation may contribute to airway branching abnormalities and, therefore, lung hypoplasia, during lung development in CDH. The murine nitrofen model of CDH was utilized to study the effect of the cholinergic agonist carbachol on embryonic day 11.5 (E11.5) lung explant cultures. Airway branching and contractions were quantified. In a subset of experiments, verapamil was added to inhibit airway contractions. Sox9 immunostaining and 5-bromo-2-deoxyuridine incorporation were used to identify and quantify the number and proliferation of distal airway epithelial progenitor cells. Intra-amniotic injections were used to determine the in vivo effect of carbachol. Airway branching and airway contractions were significantly decreased in nitrofen-treated lungs compared with controls. Carbachol resulted in increased airway contractions and branching in nitrofen-treated lungs. Nitrofen-treated lungs exhibited an increased number of proliferating Sox9-positive distal epithelial progenitor cells, which were decreased and normalized by treatment with carbachol. Verapamil inhibited the carbachol-induced airway contractions in nitrofen-treated lungs but had no effect on the carbachol-induced increase in airway branching, suggesting a direct carbachol effect independent of airway contractions. In vivo treatment of nitrofen-treated embryos via amniotic injection of carbachol at E10.5 resulted in modest increases in lung size and branching at E17.5. These results suggest that defective parasympathetic innervation may contribute to airway branching abnormalities in CDH.

Impaired Fetal Lung Development can be Rescued by Administration of Extracellular Vesicles Derived from Amniotic Fluid Stem Cells

“…1E to L, fig. S3A and B) (32,37,38). AFSC-EV administration rescued cell proliferation back to control levels and improved epithelial cell differentiation, as evidenced by a reduction of SOX9+ progenitor cell density (Fig 1E to F) and by an increased expression of SPC ( Fig.…”

MiR-449a Affects Epithelial Proliferation during the Pseudoglandular and Canalicular Phases of Avian and Mammal Lung Development

“…Transgenic pups with conditional overexpression of the miRNA 17~92 cluster in the fetal lung similarly die soon after birth or have severe respiratory distress, and manifest increased epithelial cell proliferation and increased expression of Sox9 and N-myc in the lungs [ 12 ]. Further, overproliferation of Sox9-positive cells was also described in nitrofen-treated lung explants, a validated model of CDH-associated lung hypoplasia [ 40 ].…”

Fgf10/Fgfr2b Signaling Orchestrates the Symphony of Molecular, Cellular, and Physical Processes Required for Harmonious Airway Branching Morphogenesis

“…For example, Bower et al ( 2014 ), employing a unique laser technique to specifically ablate pulmonary nerves, demonstrated that nerve tissue ablation led to reduced branching, not only in vertebrates (mouse), but in invertebrates as well ( Drosophila ), and that this effect was independent of neurotransmission. Furthermore, in mouse models of congenital diaphragmatic hernia, which is a lethal birth defect characterized by lung hypoplasia and pulmonary hypertension, embryonic lungs showed a decrease in parasympathetic innervation and ASM peristalsis, as well as impaired branching (Pederiva et al, 2012 ; Rhodes et al, 2015 ).…”