Introduction: 

Birt-Hogg-Dubé syndrome (BHD) is a rare disease caused by mutations of the FLCN gene which encodes for folliculin. A robust high expression of FLCN is observed in alveolar type II epithelial cells (AEC2), but it is unknown how this mutation affects alveolar epithelial cell biology.

Aims and objectives:

By introducing a mutation in FLCN in human induced pluripotent stem cells (hiPSCs), differentiating them into AEC2 organoids and exposing them to different triggers, we aim to understand the mechanisms governing alveolar epithelial cell dysfunction and the BHD phenotype in the lungs.

Methods:

A CRISPR/Cas9 knock out (KO) of the FLCN gene was generated in hiPSCs and differentiated into iAEC2 organoids. Primary BHD-AEC2 were cultured to allow validation of our hiPSC model. Formation of alveolar cysts was modelled by adding forskolin. Furthermore, primary BHD-AEC2 were seeded into a Lung-on-Chip system to mimic breathing-related stresses by stretch application.

Results:

Genome editing of the hiPSC line generated a FLCN KO line. In parallel, BHD-AEC2 organoids with a FLCN KO variant showed an increase in organoid lumen size upon forskolin addition compared to control organoids (1.97 times larger versus 1.1, respectively). BHD-AEC2 were seeded in the microfluidic system and stretched at 8% for three days.

Conclusions:

We have successfully generated a FLCN KO hiPSC line and derived it to iAEC2s. Parallelly, primary BHD-AEC2 displayed significant alveolar enlargement upon forskolin addition, when compared to alveolar organoids from a control. Further characterization of its phenotype is currently ongoing, by assessment of tight junctions, forskolin-induced swelling, and responses to stretch.