Experimental congenital diaphragmatic hernia features an alteration of DNA sensing targets cGAS and STING.

BACKGROUND: The pathogenesis of congenital diaphragmatic hernia (CDH) depends on multiple factors. Activation of the DNA-sensing cyclic-GMP-AMP-synthase (cGAS) and Stimulator-of-Interferon-Genes (STING) pathway by double-stranded DNA (dsDNA) links environmental stimuli and inflammation. We hypothesized that nitrofen exposure alters cGAS and STING in human bronchial epithelial cells and fetal rat lungs. METHODS: We used the Quant-IT™-PicoGreen™ assay to assess dsDNA concentration in BEAS-2B cells after 24 h of nitrofen-exposure and performed immunofluorescence of cGAS/STING. We used nitrofen to induce CDH and harvested control and CDH lungs at embryonic day E15, E18 and E21 for cGAS/STING immunofluorescence, RT-qPCR and RNA-Scope™ in-situ-hybridization (E18, E21). RESULTS: We found a higher concentration of dsDNA following nitrofen treatment. Nitrofen-exposure to BEAS-2B cells increased cGAS and STING protein abundance. cGAS abundance was higher in nitrofen lungs at E15, E18 and E21. RNA-Scope in-situ-hybridization showed higher cGAS and STING expression in E18 and E21 lungs. RT-qPCR revealed higher mRNA expression levels of STING in E21 nitrofen-induced lungs. CONCLUSION: Our data suggest that nitrofen-exposure increases dsDNA content which leads to stimulation of the cGAS/STING pathway in human BEAS-2B cells and the nitrofen rat model of CDH. Consequently, DNA sensing and the cGAS-STING-pathway potentially contribute to abnormal lung development in CDH. IMPACT STATEMENT: We found an alteration of DNA sensing targets cGAS and STING in human BEAS-2B cells and experimental congenital diaphragmatic hernia with higher protein abundance and mRNA expression in cells and lung sections of nitrofen-treated rat pups. This is the first study to investigate DNA sensing, a potential link between environmental stimuli and inflammation, in experimental CDH. Our study extends the knowledge on the pathogenesis of experimental CDH.

Embryology and anatomy of congenital diaphragmatic hernia.

Prenatal and postnatal treatment modalities for congenital diaphragmatic hernia (CDH) continue to improve, however patients still face high rates of morbidity and mortality caused by severe underlying persistent pulmonary hypertension and pulmonary hypoplasia. Though the majority of CDH cases are idiopathic, it is believed that CDH is a polygenic developmental defect caused by interactions between candidate genes, as well as environmental and epigenetic factors. However, the origin and pathogenesis of these developmental insults are poorly understood. Further, connections between disrupted lung development and the failure of diaphragmatic closure during embryogenesis have not been fully elucidated. Though several animal models have been useful in identifying candidate genes and disrupted signalling pathways, more studies are required to understand the pathogenesis and to develop effective preventative care. In this article, we summarize the most recent litterature on disrupted embryological lung and diaphragmatic development associated with CDH.