Figure 3. Effects of BM-MSC-conditioned medium on cell migration and proliferation.

(A) migration of CD14⁺ monocytes. CD14⁺ monocytes were isolated as described in “Methods” and equal numbers of the cells were loaded to the top chambers. Control (CTL) vehicle medium, fibroblast (FB-M)- or BM-MSC (MSC-M)-conditioned medium at different concentrations were added to the bottom chambers. Cells migrated into the bottom chambers were counted. Triple wells were used. Data shown represent mean±SD of 3 independent experiments (P<0.01). (B) Keratinocyte migration. Equal numbers of murine dermal keratinocytes were added to the top chambers. Media in the bottom chambers were as indicated. Cells migrated to the down-side of the filter were stained, photographed (6 fields per well) and counted. Triple wells were used for each treatment and data shown represent mean±SD of 3 independent experiments (P<0.001). (C) Keratinocyte proliferation. 0.5×10⁵ murine dermal keratinocytes per well were incubated with vehicle-M, FB-M, MSC-M or keratinocyte SFM supplemented with EGF (5 ng/ml, EGF) for different times and cell numbers were counted. Triple wells were used for each treatment. Values shown represent mean±SD of 4 independent experiments (* P<0.01). (D) HUVEC migration. The bottom chambers contained vehicle-M, FB-M or MSC-M at various dilutions. Cells migrated to the down-side of the filter were stained, photographed (6 fields per well) and counted. Triple wells were used for each treatment and data shown represent mean±SD of three independent experiments (*P<0.01 vs vehicle-M; #P<0.01 vs FB-M) (E) HUVEC proliferation. Equal numbers of HUVECs were grown in vehicle-, FB- or MSC-conditioned basal endothelial growth medium (EGM-2) supplemented with 2% FBS or complete EGM-2 and incubated for 3 days. Cell numbers were counted. Experiments were performed in triplicate wells (n = 3, * P<0.001).