FIG. 1.

Overview of experimental approach. (A) MSCs were cultured in monolayer. The use of SMAD3 shRNA and RUNX2 overexpression was hypothesized to induce MSCs toward osteogenesis, despite the presence of TGFβ3. (B) MSCs were pretransduced with SMAD3 shRNA or SMAD3 shRNA with RUNX2 before being seeded on the woven scaffolds. Scrambled shRNA and scrambled shRNA with RUNX2 were used as control groups. Square box indicates two scaffolds cultured in the same well. Yellow and blue bars represent warp and weft fibers. (C) Schema of viral backbones used. A detailed description of the backbone components is provided in Supplementary Table S2. We expect transduced cells to emit a strong signal under the red fluorescent protein (RFP) channel (due to dsRedExpress2 in our transfer vector construct). (D) Distinct differences between NT and virally transduced scaffolds cultured in the same well. Representative microscopic images of scaffolds in culture (week 5). Only the scaffold with virally transduced cells (scrambled control in this image) emitted RFP signal, while the NT scaffold did not. Upper panel: stitched image from multiple views to capture the entire scaffold pair. Lower panel: single images of scaffold pairs. Scale bar = 500 μm. TGFβ3, transforming growth factor β; MSCs, mesenchymal stem cells; SMAD3, Mothers against DPP homolog 3; shRNA, short hairpin RNA; NT, nontransduced. Color images available online at www.liebertpub.com/tea