Fig. 2

BM-MSCs incubated with SMG-ECM formed cell aggregates during culture for 7–14 days. A BM-MSCs were either untreated or treated with SMG-ECM for 1 h at 37 °C before transfer to a standard tissue culture dish and culture for up to 7 days. Over time, the untreated cells formed a monolayer across the surface of the dish, while the treated cells formed cell aggregates as well as a monolayer. Bar: 100 µm (2 left panels), 200 µm (far right panels). B After culture for 7 days, as described in A, cell aggregates were hand-picked under a dissecting microscope, transferred to a fresh tissue culture dish, and the cultures continued for an additional 7 days in epithelial cell differentiation media (i.e., 14 days total). During this second 7-day culture, cell aggregate organization was maintained. Bar: 100 µm. C BrdU assay was used to measure the proliferation of BM-MSCs that were untreated (Untreated) or treated with SMG-ECM and formed aggregates (aggregates) or a monolayer (monolayer) of cells during culture for 4 or 7 days. Negative control cultures either did not receive BrdU (No BrdU) or were treated with mitomycin C (MMC) to block proliferation. Data are the mean ± SD from a representative experiment. *p < 0.05 versus no BrdU and MMC groups; each experimental group contained an n = 4–6 and the experiment was repeated 3 times with MSCs from different donors. D Comparison of calcium mobilization in untreated and SMG-ECM-treated cell aggregates (50–100 cells) in the presence of thapsigargin (Tg in calcium-free media) followed by addition of external calcium (1 mM). The results were obtained using a minimum of 50 cells and are representative of those obtained in 3 individual experiments using cells from different donors