Epithelial-mesenchymal co-culture model for studying alveolar morphogenesis

Abstract

Division of large, immature alveolar structures into smaller, more numerous alveoli increases the surface area available for gas exchange. Alveolar division requires precise epithelial-mesenchymal interactions. However, few experimental models exist for studying how these cell-cell interactions produce changes in 3-dimensional structure. Here we report an epithelial-mesenchymal cell co-culture model where 3-dimensional peaks form with similar cellular orientation as alveolar structures in vivo. Co-culturing fetal mouse lung mesenchyme with A549 epithelial cells produced tall peaks of cells covered by epithelia with cores of mesenchymal cells. These structures did not form when using adult lung fibroblasts. Peak formation did not require localized areas of cell proliferation or apoptosis. Mesenchymal cells co-cultured with epithelia adopted an elongated cell morphology closely resembling myofibroblasts within alveolar septa in vivo. Because inflammation inhibits alveolar formation, we tested the effects of E. coli lipopolysaccharide on 3-dimensional peak formation. Confocal and time-lapse imaging demonstrated that lipopolysaccharide reduced mesenchymal cell migration, resulting in fewer, shorter peaks with mesenchymal cells present predominantly at the base. This epithelial-mesenchymal co-culture model may therefore prove useful in future studies of mechanisms regulating alveolar morphogenesis.

Keywords:

• alveolarization
• bronchopulmonary dysplasia
• myofibroblast
• lung development

Disclosure of Potential Conflicts of Interest

No potential conflicts of interest were disclosed.

Acknowledgments

The authors would like to thank Albert Tousson (UAB), Sam Wells (Vanderbilt University), and Mark Brown (Andor) for their advice and assistance with imaging and image analysis. We thank our colleagues for their helpful comments regarding the experimental data and in writing and editing the manuscript. This work was supported by the NHLBI (HL086324 and HL097195, LSP), a March of Dimes Basil O’Connor Award (LSP), the American Lung Association (LSP), the Child Health Research Center at UAB, an Advancing Newborn Medicine Fellowship Grant from Ikaria (JDM), and a Neonatal Fellowship Grant from Discovery Laboratories (JDM).