Forward and Reverse primers of transcription factors and Genes used for qRT-PCR.

<p>Forward and Reverse primers of transcription factors and Genes used for qRT-PCR.</p

SEM of nanosurfaces coated with BSA, Collagen and Fibronectin.

<p>Close interactions of proteins with nanosurface of 50nm dot diameter, 20nm inter-dot spacing and 40nm nanodot height, were seen. Proteins did not appear to interact with base of the nanosurface with 100nm dot diameter due to a greater nanodot height of 100nm and an inter-dot spacing of 70nm. Scale bar = 500nm.</p

Scanning electron microscopy, Immunofluorescence staining and Statistical analysis of cell area and cell viability of MG63 cells on 50 and 100nm dot diameters.

<p>(a): SEM of cells displaying a well extended morphology on the nanodots with 50nm dot diameter in contrast to an elongated morphology on the 100nm dot diameter. Scale bar for Flat = 200, 50μm; for 50nm = 100μm, 10μm; for 100nm = 100μm, 10μm. (b): Immunofluorescence staining of cell cytoskeleton and focal adhesions. Cell morphology was analyzed by staining the cytoskeleton with Phalloidin (Red) and FITC conjugated goat anti rabbit anti-vinculin antibodies (Green). Cells displayed a well extended morphology with a plethora of focal adhesions on nanodots with 50nm dot diameter in contrast to an elongated/constricted morphology with few focal adhesions on the nanodots with 100nm dot diameter. Scale bar = 50μm. (c): Statistical analysis of cell area and cell viability of cells seeded on nanodots with 50 and 100nm dot diameter. Cells displayed a smaller cell area on nanosurfaces with 100nm dot diameter as compared to the nanosurfaces with 50nm dot diameter. Statistical analysis complimented the SEM and Immunofluorescence staining results.</p

Fold change in expression of transcription factors and genes coding for bone proteins.

<p>Fold change in expression of Alkaline phosphatase and Osterix in cells on 50nm dot diameter was twice as compared to fold change in cells on 100nm dot diameter. Additionally, Osteocalcin, RUNX and Bone Sialoprotein were also higher expressed in cells on 50nm nanodots.</p

Scanning electron microscopy, Immunofluorescence staining and statistical analysis of cells after Cytochalasin-D treatment on nanodots with different dot diameter.

<p>(a): Scanning electron microscopy of cells with/without Cytochalasin-D treatment. Cells displayed a round morphology due to actin de-polymerization by Cytochalasin-D. Very few microfilaments were seen on normal and Cytochalasin-D treated cells showing that even after the morphology transition, cells could not establish adhesions with the nanodots due to a greater nanodot height and inter-dot spacing. Normal represents untreated cells on nanosurfaces. Scale bar = 100μm. (b): Immunofluorescence staining of MG63 cells. From left to right: (i) Vinculin was stained with FITC conjugated goat anti rabbit antibodies (Green). (ii) Cytoskeleton was stained with Phalloidin (Red), and far right displays (iii) Merged images. Scale bars: 20X = 75μm, 40X = 50μm, 63X = 25μm. (c): Statistical analysis of cell area and cell viability of cells seeded on nanodots with 50 and 100nm dot diameter after Cytochalasin-D treatment. Cells displayed a smaller cell area on nanosurfaces with 100nm dot diameter as compared to the nanosurfaces with 50nm dot diameter. Cells also displayed a lower viability on the nanodots with 100nm dot diameter. Cells failed to form focal adhesions with the 100nm nanodots due to a greater height and suffered cell death through Anoikis.</p

Schematic representation of fabrication of Tantalum oxide nanodot arrays.

<p>Schematic representation of fabrication of Tantalum oxide nanodot arrays.</p

List of fabricated nanotopographies with corresponding features.

<p>List of fabricated nanotopographies with corresponding features.</p

Scanning electron microscopy of Tantalum oxide nanodots.

<p>(a), (c): Top and cross-sectional view of nanodots with 50nm diameter, 20nm inter-dot spacing and 40nm nanodot height. (b), (d): Top and cross-sectional view of nanodots with 100nm diameter, 70nm inter-dot spacing and 100nm nanodot height. Scale bar = 100nm.</p