Adsorption of FN and VN by electrospun scaffolds.

<p>Scaffolds were coated with fetal bovine serum (A), or implanted into rat tibial osteotomies for 30 min (B). Scaffolds were then washed to remove loosely bound proteins, and proteins were subsequently desorbed by incubation in boiling SDS-containing solution. The amounts of FN and VN were evaluated by Western blot.</p

Immunostaining for phosphorylated Focal Adhesion Kinase.

<p>MSCs were seeded onto glass coverslips coated with electrospun nanofibers, or with FBS as a control. After 5 hours, cells were fixed and stained for phosphorylated Focal Adhesion Kinase (red). Cells were counterstained with DAPI to show cell nuclei (blue). Cells seeded onto PCL/col/HA scaffolds were better spread, and exhibited greater amounts of punctuate pFAK staining (site pY397) as compared with cells on PCL or PCL/HA. Cells seeded onto FBS-coated glass coverslips displayed pFAK staining in focal adhesion-type structures (white arrows), as expected for cells grown on 2D surfaces.</p

Live cell imaging of GFP-expressing MSCs seeded onto electrospun scaffolds.

<p>A) Cells were seeded onto scaffolds and imaged over varying time points. Panels a–c: PCL scaffolds; panels d–f: PCL/HA scaffolds; panels g–i: PCL/col/HA scaffolds and panels j–l: col scaffolds. Scale bar = 100 µm. B) Higher magnification images of GFP-expressing MSCs at seven hours on electrospun scaffolds (panels m–p).</p

Tensile Properties of dry and hydrated scaffolds.

<p>Values represent the average ± standard deviation calculated in the linear portion at 10% strain. The hydrated collagen scaffolds have very low mechanical properties and could not be measured by this technique.</p

SEM images of MSCs cultured on nanofibrous scaffolds for 24 hours.

<p>A) Cell spreading was observed on PCL, PCL/HA, and PCL/col/HA scaffolds, but not on 100% collagen I (col). B) Col scaffolds (without cells) were incubated in culture media for 24 hrs to allow the potential release of soluble factors, and then the solution was collected. MSCs were suspended into this conditioned media, seeded onto PCL scaffolds, and allowed adhere in the media for 24 h. Under these conditions cell spreading was extensive, suggesting that lack of cell spreading on col substrates was not due to any soluble factors released from these scaffolds.</p

MTS assay quantifying cell proliferation on electrospun scaffolds of PCL, PCL/HA or PCL/col/HA.

<p>At day one, cell number was significantly higher on PCL/HA and PCL/col/HA scaffolds in comparison to PCL. By day four, PCL/HA was still significantly higher than PCL, and PCL/col/HA was significantly higher than PCL/HA and PCL. In addition, cell number on PCL/col/HA was significantly higher on day four than day one. An * denotes p<0.05</p