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. Author manuscript; available in PMC: 2015 May 1.
Published in final edited form as: Clin Cancer Res. 2014 May 1;20(9):2375–2387. doi: 10.1158/1078-0432.CCR-13-1415

Figure 4. hAMSCs remain multipotent and retain proliferation capacity when exposed to BTIC-CM or transduced with BMP4 in vitro.

Figure 4

(A) MTS assay of hAMSCs cultured in BTICCM or control media for 2 weeks to measure proliferating hAMSCs every 3 days. (B) EdU assay of td-tomato-hAMSCs co-cultured with BTICs for 5 days to determine proliferating hAMSCs. (C) Real-time RT-PCR was performed. Markers for adipocyte, osteocyte and chondrocyte lineages were tested. GAPDH served as a control. Other groups were normalized and compared to the undifferentiated hAMSCs group. (D) hAMSCs were cultured in control media (undifferentiated hAMSCs), differentiation media (differentiated hAMSCs), or BTIC-CM for 3 weeks. Various stains were performed to assess differentiation capabilities (scale bar, 100 μm) and (E) EdU assay of hAMSCs treated with BMP4 (50 ng/ml) for different time periods (24 hours and 48 hours) to measure hAMSC proliferation. Results were normalized and compared to the 0 hour condition. (F) EdU assay comparing proliferation of hAMSCs-Vector and hAMSCs-BMP4 cells after 5 days of culturing. Results were normalized and compared to hAMSCs-Vector. (G) hAMSCs-Vector, hAMSCs-Vector treated with BMP4 (100 ng/ml), or hAMSCs-BMP4 cultured in control media or differentiation media for 3 weeks. Various lineage stains were performed as described previously. Scale bar, 100 μm. (H-I) Transwell assays: hAMSCs-Vector (H) or hAMSCs-BMP4 (I) were seeded on top chamber. BTIC-CM or control media+2% FBS was at the bottom of the chamber. Results were normalized and compared to control.*p<0.05, **p<0.01, ***p<0.001; N.S., not significant.