Figure 6. Retention of Injected MSCs in Mice With or Without LPS-Induced ALI.

MSCs were labeled with the cell tracing dye CFDA SE (green) prior to injection. Nuclei were stained with TO-PRO-3 (blue). Scale bars in photomicrographs = 20 μm. White arrows indicate labeled MSCs.

(A) Labeled MSCs were observed in 5 μm, PFA-fixed lung sections from LPS-injured mice sacrificed at 15 min (initial retention). Image obtained with a Leica laser scanning confocal microscope with a 20× objective.

(B) A three-dimensional lung section from an animal that received labeled MSCs. Lung was inflated, stored in OCT, and cut into 50 μm thick sections. Z-series images (30 sections, total thickness of the tissue scanned = 17.71 μm) were collected with a 63× oil objective and projected in different axes, as shown. Red autofluorescence shows outline of the alveolus.

(C) pANGPT1-transfected MSCs labeled with the green fluorescent cell tracker CFDA SE were observed in lung section from LPS-injured mice killed at 3 d. Photomicrographs were taken in z-axis with a 20× objective, then images were stacked using Leica confocal software.

(D) Lung lobes (left upper and all right lobes) from each animal were enzyme-digested into single cells before MSCs were counted by flow cytometry. n = 5 per group.