# Engineering Biomimetic Gelatin Based Nanostructures as Synthetic Substrates for Cell Culture
## 4 Discussion  
Functionalization is a strategy which is used in the field of scaffold technology to increase the biocompatibility of a scaffold [44]. In this experiment, P188 was successfully co-electrospun with PCL/gelatin to produce three different functionalized scaffolds (PCL/gelatin/P188 with concentration ratio of 85:10:5, 80:10:10, and 70:10:20 %w/v) with improved cell adhesion and proliferation property. This scaffold has brought together the inherent nanofiber forming ability of PCL, the integrin mediated cell adhesion property of gelatin, and the ability of P188 to revive damaged cells [32; 38]. The presence of all the components in the novel nanofibrous scaffold was confirmed by ATR-FTIR analysis. Many studies in the past have shown the ability of high voltages to stretch the solution and decrease the diameter of the fibers [45; 46; 47]. However, no such relationship of a decrease in diameter with an increase in voltage was observed in this study (Table 2). However, the average diameter of more than 50% of the fibrous scaffolds determined using statistical analysis was estimated to be within 200 to 500 nm range (irrespective of their polymer composition), with the highest reproducibility observed for scaffolds electrospun at 22 kV. Another observation with respect to the fiber diameter  
Figure 8: The mean fluorescence indicating the Caco-2 cell viability when cultured on three different PCL/gelatin/P188 (concentration ratios of 85:10:5, 80:10:10, and 70:10:20 %w/v) nanofibrous scaffolds. \({}^{*}\)\(p<0.05\); \({}^{***}\)\(p<0.001\). is the broader diameter distribution of all the scaffolds except PCL. This is attributed to the fact that the high conductive polymer solutions are unstable in the presence of high voltages, resulting in fibers of varying diameter [45; 48]. The conductivity of PCL/gelatin and PCL/gelatin/P188 solutions are significantly higher (around 12 \(\upmu\)S) in comparison with the PCL solution (1.9 \(\pm\) 0.95 \(\upmu\)S). Among the composite fibrous scaffolds, there was a decrease in homogeneity with the decrease in concentration of PCL in the fibers. The PCL/gelatin fibers appeared more homogeneous with fewer polymeric solids when compared to that of PCL/gelatin/P188 composites. Even among PCL/gelatin/P188 composites, the one with the least concentration of PCL (PCL/gelatin/P188, 70:10:20) produced many beads when compared to other two P188 based scaffolds (PCL/gelatin/P188, 85:10:5 and 80:10:10). This could be because of the lowering of viscosity with a decrease in PCL concentration [49]. Similarly, PCL (15%) sample showed a large concentration of beads indicating an instability during electrospinning. A similar observation was made by [PERSON] et al., who determined that 15% PCL solution produced using 90/10 %v/v acetic acid/formic acid solvent system has low viscosity and low conductivity resulting in electrospinning instabilities [46]. This roughness of the scaffold is known to play a very important role in cell adhesion as a rougher surface provides a larger surface area for cell interaction compared to the smooth surface [50]. AFM analysis of PCL/gelatin/P188 composite with 20% P188 showed the highest mean roughness of 1039 \(\pm\) 265 nm. Whereas the roughness value obtained for the other two PCL/gelatin/P188 composites with 5% and 10% P188 were 679 \(\pm\) 33 nm and 683 \(\pm\) 95 nm, respectively. Although the increase in fiber diameter is known to increase the surface roughness [51; 52; 53], the high roughness of the scaffold with 20% P188 was probably due to increasing in P188 concentration. As 20% P188 based composite scaffold electrospun at 22 kv had the lowest average diameter of 306 \(\pm\) 163 nm when compared to scaffolds with 10% and 5% P188, which had an average diameter of 327 \(\pm\) 196 nm and 390 \(\pm\) 158 nm, respectively (Table 2). The mean roughness Ra of PCL/gelatin composite scaffold was 855 \(\pm\) 11 nm and that of PCL scaffold was 696 \(\pm\) 134 nm.