Genetically Engineered Plant Viral Nanoparticles Direct Neural Cells

Aug 6, 2015 - An important aim of tissue engineering is to design biomimetic materials with specific cell binding motifs and precisely controlled stru...
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Figure 1. Biocompatibility of TMV materials. Cell viability and proliferation with TMV exposure. (a) N2a cells exhibited similar cell numbers regardless of TMV addition to the media at each of the tested time points. Cells exhibited significant proliferation (* P < 0.001, two-way ANOVA, n = 6) in the presence of each type of TMV mutants during the tested time, same as the positive control (culture media without TMV). (b) N2a cells cultured on various substrates coated by various TMV mutants exhibited similar proliferation rate, same as the control cultured on tissue culture plastics (TCP) and PDL coating substrates proliferation (P < 0.001, two-way ANOVA , n = 6). 189x209mm (300 x 300 DPI)

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Figure 2. Growth Patterns of differentiated N2a cells on TMV substrates. SEM images of differentiated N2a cells (24 hours in differentiated media) on TCP (a), wild-type TMV (b), RGD1 (c), RGD7 (d), P15 (e), DGEA (f), and PHSRN3 (g) substrates. (a-g) share the scale bar in (g), indicating 100 µm. (h) Plot of cell spatial distribution on TMV substrates. (i) Shchematic diagrams of the nearest neighbor analysis. In the analysis the distribution of cells can range from independent (represented by a theoretical Poisson’s distribution), to clustered, or regular. 184x199mm (300 x 300 DPI)

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Figure 3. Neurite outgrowth of N2a cells on TMV substrates. Images are typical fields of differentiated N2a cells (48 hours in differentiation media) on PDL (a) and wildtype- (b), RGD1- (c), RGD7- (d), P15- (e), DGEA- (f), PHSRN3-TMV (g) substrates. (a-g) share the scale bar in (g), indicating 100 µm. (h) Average neurite length of cells in each group (at least 200 cells were measured). Each data point is the mean ± SE of three independent experiments. ** indicates P < 0.001. 128x95mm (300 x 300 DPI)

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Figure 4. Flow assembly of TMV in capillaries. (a) SEM image of control (i. e. 0.05 mg mL-1 TMV solution was slowly added in a capillary without applying a fluid flow). SEM images of flow assembly of different concentrations of TMV on chitosan modified capillary tubes at a fixed flow rate of 200 cm s-1: (b) 0.005 mg mL-1, (c) 0.01 mg mL-1, (d) 0.03 mg mL-1, (e) 0.05 mg mL-1, (f) 0.1 mg mL-1, and (g) 0.2 mg mL-1. All SEM images share the scale bar in (g), indicating 500 nm. (h) The dependence of TMV density on assembled solution concentration. Orientation order parameter for all conditions remain above 0.85. 130x100mm (300 x 300 DPI)

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Figure 5. Orientation of neurite outgrowth on aligned TMV substrates. N2a cells generated neurites in (a) control (no virus), and substrates with aligned (b) wild-type TMV, (c) RGD1, (d) RGD7, (e) P15, (f) DGEA, and (g) PHSRN3. (a-g) share the scale bar in (g), indicating 100 µm. (h) Average neurite length of cells in each group (at least 100 neurites were measured). Each data point is the mean ± SEM. * P