(A–E) NSG mice were treated with IT bleomycin. Two weeks later, mice received either CD44^hi or CD44^lo IPF MPCs (IPF424); 10 mice/group. Flow cytometry showing isolation of SSEA4^hi/CD44^hi and SSEA4^hi/CD44^lo IPF MPCs (A). CD44 levels were quantified in CD44^hi and CD44^lo IPF MPCs by qPCR (B; left panel) and Western blot analysis (B; middle panel) (n = 4 cell lines tested IPF259, IPF309, IPF424, and IPF327); densitometry values summarize Western blot data (right panel). Lungs were harvested at week 6. (C) Collagen content was quantified in left lungs by Sircol assay (left panel). Ten mice from the above animal experiment (5 mice each from CD44^hi and CD44^lo groups; IPF 424) were used in this analysis. Semiquantitative analysis of collagen deposition was performed by Trichrome staining (right panel). Three sections from each animal were screened. Three Trichrome stained images at low power (5×; scale bar: 500 µm) randomly selected from each section were used for quantification. Blue regions (fibrotic stain) were defined and quantified using ImageJ (right panel). (D–M) Serial 4 µm sections of right lung tissue from mice receiving CD44^hi IPF MPCs (D–G; scale bar: 200 µm) (H–M; scale bar: 50 µm). H&E and Trichrome stains assessing fibrosis and collagen deposition, respectively (D–G). IHC using an antibody recognizing human procollagen to identify human cells and assess collagen synthesis (H–J) and a CD44 antibody to determine the distribution of CD44 expressing cells (K–M). (H, I, K, and L) IHC for procollagen (H and I) and CD44 (K and L) to assess the distribution of human cells expressing collagen and CD44-expressing cells from fibrotic and nonfibrotic lung regions from mice receiving CD44^hi IPF MPCs. Data expressed as mean ± SEM. P values were determined by 2-tailed Student’s t test.