Research Paper Volume 11, Issue 24 pp 12476—12496

Figure 3. Knockdown of Rmst diminishes BMP9-induced osteogenic and adipogenic differentiation of MSCs. (A) AdR-simRmst was shown to infect the iMADs with high efficiency alone or co-infect with Ad-BMP9. Images were recorded at 48h post infection. Representative images are shown. (B and C) Downregulation of Rmst reduces BMP9-induced ALP activity in iMADs. Subconfluent iMADs were infected with Ad-BMP9, Ad-GFP, and/or AdR-simRmst. ALP activity was quantitatively determined at 3, 5 and 7 days after infection (B) or stained histochemically (C). Assays were done in triplicate. “*” p<0.05 and “**” p<0.001 when compared with the Ad-BMP9 alone group. Representative images are shown. (D) Silencing Rmst leads to reduced matrix mineralization induced by BMP9 in iMADs. Subconfluent iMADs were infected with Ad-BMP9, Ad-GFP, and/or AdR-simRmst, and cultured in mineralization medium. At day 7 and day 14, the infected cells were fixed and subjected to Alizarin Red S staining. Each assay condition was done in triplicate. Representative microscope images are shown. (E) Downregulation of Rmst reduces BMP9-induced adipogenesis in iMADs. Subconfluent iMADs were infected with Ad-BMP9, Ad-GFP, and/or Ad-simRmst. At 10 days post infection, the cells were fixed and subjected to Oil Red O staining. Each assay condition was done in triplicate. Representative microscopic images are shown.