Commentary: Sca-1 and Cells of the Lung: A matter of Different Sorts

Abstract

In two separate articles published in this issue, Teisanu et al. and McQualter et al. report the use of flow cytometry and cell sorting to identify putative bronchiolar stem cells that are low in expression for the cell surface marker Sca-1 yet negative for CD34, and a mesenchymal, fibroblastic progenitor cell population from the lung that is positive for Sca-1, respectively. At first glance, these studies may seem to suggest that Sca-1 and CD34 are not markers of an epithelial stem cell population in the lung, as we previously determined in studies that identified bronchioalveolar stem cells (BASCs), and may also appear to contradict each other. However, here we point to evidence that the findings of these three studies are not mutually exclusive, and rather, that the different cell isolation and culturing protocols used in these studies have allowed for the identification of unique pulmonary cell populations. Rather than discounting previous work on BASCs, these studies reveal the existence of new methods and new cell types that will be interesting to use in future functional tests for their importance in lung biology and lung disease.

FIGURE 1. Colony morphology of sorted Sca-1^pos and Sca-1^neg cell populations

Cells were isolated from β-actin-GFP mouse lungs as described , and the viable CD45^negCD31^neg population was divided into Sca-1^pos (left panel) and Sca-1^neg (right panel) fractions and plated on irradiated MEF feeders. Colonies of epithelial (upper right) and mesenchymal (lower left) morphologies were observed in the Sca-1^pos fraction, whereas the Sca-1^neg fraction yielded mostly epithelial colonies.

FIGURE 2. Choice of antibody conjugates affects apparent size of Sca-1^pos and CD34^pos populations

(A) Lungs from C57Bl/6 mice were processed as described . Aliquots from each of four individual mice were stained with antibodies against CD45, CD31, and a combination of either Sca-1-FITC/CD34-PE or Sca-1-PE/CD34-FITC; dead cells were excluded with 7AAD. (Sca-1 antibodies were purchased from BD Pharmingen, and CD34 antibodies were purchased from eBioscience.) Representative plots of the viable CD45^negCD31^neg populations are shown. Note the high autofluorescence in the FITC channel, a characteristic trait of lung cells and notably AT2 cells as in our previous studies. (B) Total Sca-1^pos (left), total CD34^pos (middle), and Sca-1^pos CD34^pos double positive populations as percentages of the parent CD45^negCD31^neg population (black bars) or all viable cells (gray bars). (C) Percent of Sca-1^pos cells also positive for CD34; right two bar values were calculated or taken directly from data presented in the publications listed.

FIGURE 3. Proposed cell lineage relationships of lung stem/progenitor cells to differentiated progeny

(Left) Bronchioalveolar stem cells are Sca-1^pos CD34^pos cells that can self-renew on feeder cells and can differentiate into three distinct epithelial cell types on Matrigel. (Center) McQualter et al. demonstrate predominant mesenchymal potential and rare epithelial potential of Sca-1^pos cells under various in vitro culture conditions. (Right) Teisanu et al. use FACS data to infer a stem (AF^low) to progenitor (AF^hi) lineage from Sca-1^low lung cells. All cells depicted here are also CD45^neg and CD31^neg. Abbreviations: SPC, pro-surfactant protein C; CCSP, Clara cell secretory protein; AT1, alveolar type I cell; AT2, alveolar type II cell; AQ5, aquaporin 5; ALI, air-liquid interface culture; AF, autofluorescent in the AlexaFluor 647 channel; TA cell, transit-amplifying cell. Hatched lines indicate rare events and/or limited potential.