Platform effects on regeneration by pulmonary basal cells as evaluated by single-cell RNA sequencing [array]

Cell-based therapies have shown promise for treating myriad chronic pulmonary diseases through direct application of epithelial progenitors or by way of engineered tissue grafts or whole organs. To elucidate environmental effects on epithelial regenerative outcomes in vitro, we isolated and cultured a population of pharmacologically expanded basal cells (peBC) from rat tracheas. At peak basal marker expression we simultaneously split peBC into four in vitro platforms – organoid, air-liquid interface (ALI), engineered trachea, and engineered lung. Following differentiation, these samples were evaluated using single-cell RNA sequencing (scRNAseq) and computational pipelines were developed to compare samples both globally and at the population level. A sample of native rat tracheal epithelium was also evaluated by scRNAseq as a control for engineered epithelium. Overall this work identifies platform-specific effects that support the use of engineered models to achieve the most physiologic differential outcomes in pulmonary epithelial regenerative applications. Comparison of three primary rat tracheal epithelial scrapings plated into BEGM or EpiX media and grown for 8 days