Identifying a Lung Stem Cell Subpopulation by Combining Single-Cell Morphometrics, Organoid Culture, and...

Area of Science:

• Stem cell biology
• Regenerative medicine
• Single-cell genomics

Background:

• Dissecting cellular heterogeneity in complex systems using single-cell RNA sequencing is crucial.
• Estimating proliferation and differentiation potentials of dormant tissue stem cell subpopulations remains challenging.

Purpose of the Study:

• To establish a novel single-cell analysis method, scMORN, for profiling organoid-forming capacity and differentiation potential of tissue stem cells.
• To identify and characterize stem cell subpopulations within lung epithelial stem cells.

Main Methods:

• Integration of single-cell morphometrics, organoid-forming assays, and RNA sequencing (scMORN).
• Development of a feeder-free culture system for expanding lung stem cells (basal, club, AT2).
• Single-cell transcriptome profiling and immunohistochemical analysis.

Main Results:

• Discovery of Muc5b+ and large club (ML-club) cells, a subpopulation of club cells with superior organoid formation and differentiation capacity.
• ML-club cells are localized in the intrapulmonary proximal airway and distinct from known club cell subpopulations.
• Identification of CD14 as a cell surface antigen for ML-club cells, showing enhanced engraftment and expansion in injured mouse lungs.

Conclusions:

• The scMORN method effectively identifies and characterizes stem cell subpopulations with specific potentials.
• ML-club cells represent a promising target for lung regeneration and repair.
• The scMORN approach is adaptable for discovering novel stem cell subpopulations in various tissues.