Abstract
Preterm infants frequently suffer from pulmonary complications due to a physiological and structural lung immaturity resulting in significant morbidity and mortality. Novel in vitro and in vivo models are required to study the underlying mechanisms of late lung maturation and to facilitate the development of new therapeutic strategies. Organoids recapitulate essential aspects of structural organization and possibly organ function, and can be used to model developmental and disease processes. We aimed at generating fetal lung organoids (LOs) and to functionally characterize this in vitro model in comparison to primary lung epithelial cells and lung explants ex vivo. LOs were generated with alveolar and endothelial cells from fetal rat lung tissue, using a Matrigel-gradient and air-liquid-interface culture conditions. Immunocytochemical analysis showed that the LOs consisted of polarized epithelial cell adhesion molecule (EpCAM)-positive cells with the apical membrane compartment facing the organoid lumen. Expression of the alveolar type 2 cell marker, RT2-70, and the Club cell marker, CC-10, were observed. Na+ transporter and surfactant protein mRNA expression were detected in the LOs. First time patch clamp analyses demonstrated the presence of several ion channels with specific electrophysiological properties, comparable to vital lung slices. Furthermore, the responsiveness of LOs to glucocorticoids was demonstrated. Finally, maturation of LOs induced by mesenchymal stem cells confirmed the convenience of the model to test and establish novel therapeutic strategies. The results showed that fetal LOs replicate key biological lung functions essential for lung maturation and therefore constitute a suitable in vitro model system to study lung development and related diseases.
Highlights
The study of fetal lung development is a challenging task
Our study focused on the generation of a biological relevant model system of fetal lung development that can be reproduced by other scientists
lung organoids (LOs) formation was compared between Matrigel-coated permeable inserts covered with cells in Matrigel exposed to air (ALI culture) and Matrigel-containing cell suspension plated at the bottom of a well and overlaid with LO medium (LO-Med) (Figure 1B)
Summary
The study of fetal lung development is a challenging task. Any model system needs to reflect the biological properties such as morphology and function, while being reproducible and, if possible, broadly accessible. ENaC expression is reduced in the preterm lungs [1], compromising AFC These pathognomonic features must be reflected in model systems of fetal lung development. Respiratory distress in preterm infants is multifactorial, including congenital infection, growth restriction and placental dysfunction, and most preterm infants were exposed to clinical interventions like antenatal corticosteroids This is not reflected in most animal models possibly leading to different responses to pathogenic challenges as well as therapeutic strategies. Primary fetal distal lung epithelial (FDLE) cells represent a widely studied in vitro model due to their ability to differentiate into polarized and functional epithelia [3]. Studies showed that fetal rat pups born 24 h prior to term birth experience respiratory distress due to structural and functional lung immaturity [4]. FDLE cells are limited in their lifetime and offer only a short time window for experiments of 2–3 days
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