Cell Culture Differentiation and Proliferation Conditions Influence the Regeneration of the Human Airway Epithelium.

The human airway mucociliary epithelium can be recapitulated using primary cells cultured in an air-liquid interface (ALI), a reliable surrogate to perform pathophysiological studies. As tremendous variations exist among media used for ALI-cultured human airway epithelial cells, the aim of our study was to evaluate the impact of several media (BEGM, PneumaCult, Half & Half, and Clancy) on cell type distribution using single-cell RNA sequencing and imaging. Our work revealed the impact of these media on cell composition, gene expression profile, cell signaling, and epithelial morphology.

Combination of CRISPR-Cas9-RNP and Single-Cell RNAseq to Identify Cell State-Specific FOXJ1 Functions in the Human Airway Epithelium.

The study of the airway epithelium in vitro is routinely performed using air-liquid culture (ALI) models from nasal or bronchial basal cells. These 3D experimental models allow to follow the regeneration steps of fully differentiated mucociliary epithelium and to study gene function by performing gene invalidation. Recent progress made with CRISPR-based techniques has overcome the experimental difficulty of this approach, by a direct transfection of ribonucleoprotein complexes combining a mix of synthetic small guide RNAs (sgRNAs) and recombinant Cas9.

The MIR34B/C genomic region contains multiple potential regulators of multiciliogenesis.

The MIR449 genomic locus encompasses several regulators of multiciliated cell (MCC) formation (multiciliogenesis). The miR-449 homologs miR-34b/c represent additional regulators of multiciliogenesis that are transcribed from another locus. Here, we characterized the expression of BTG4, LAYN, and HOATZ, located in the MIR34B/C locus using single-cell RNA-seq and super-resolution microscopy from human, mouse, or pig multiciliogenesis models.