Polyurethane/chitosan/hyaluronic acid scaffolds: providing an optimum environment for fibroblast growth
Authors: Hashemi Seyedeh-Sara , Rajabi Seyedeh-Somayeh , Mahmoudi Reza , Ghanbari Amir , Zibara Kazem , Barmak Mehrzad Jafari 
Published: October 2020

Objective: Biodegradable polymers can replace damaged tissue components using tissue engineering techniques. The objective of this study is to determine an optimum environment for polymer scaffolds to improve the proliferation of fibroblast cells capable of wound repair. Method: In this study, the addition of polysaccharides, such as chitosan (CH) or hyaluronic acid (HA), to a polyurethane (PU) polymer was evaluated using different methods to determine if they affect scaffold morphology and cell activity of fibroblasts prepared from human foreskin tissues. Mechanical properties, such as tensile strength, contact angle and swelling test, were used to check the physical and mechanical properties of the scaffold. Fibroblast growth was also measured at 24, 48 and 72 hours. Results: Scanning electron microscopy (SEM) determined that a 3:1 ratio of PU/CH scaffold, developed by electrospinning, allowed the formation of a uniform structure in scaffold fibres. Physical mechanical tests showed that PU electrospun scaffolds were not modified by the addition of CH. The mean stretch and mean water absorption increased significantly using the PU/CH scaffold, compared with the PU scaffold. However, the mean tensile strength and the mean contact angle, used to study space and porosity, did not differ between scaffolds. Fourier transform infrared spectroscopy confirmed the functional groups (–OH, –NH and –C=O) in the PU/CH scaffold, compared with PU or CH chemical structures alone. HA was then added to CH and PU/CH scaffolds to evaluate the growth of fibroblast cells. Results showed that cell viability and the number of cells, using MTT and trypan blue exclusion assay, respectively, increased significantly at 24, 48 and 72 hours of culture in PU/CH/HA scaffold compared to HA, CH/HA, and PU/HA. Moreover, PU/HA at 48 and 72 hours also increased cell viability and cell numbers compared to HA and CH/HA scaffolds. However, scaffolds at 72 hours had limited space for cell growth. Moreover, SEM data demonstrated that fibroblasts were able to proliferate, penetrate, migrate and survive on PU/HA and PU/CH/HA three-dimensional scaffolds, especially during the first 48 hours. Furthermore, 4′,6-diamidino-2-phenylindole (DAPI) staining confirmed that fibroblasts could penetrate PU scaffolds and showed higher cell viability and lower cellular damage in PU/CH/HA, compared to CH/HA and PU/HA scaffolds. Finally, flow cytometry using CD90 and CD105 surface markers revealed that >90% of cells isolated from the human dermis were fibroblasts. Conclusion: In summary, PU/HA and PU/CH/HA scaffolds were found to be biocompatible and provided a suitable environment for the growth and proliferation of fibroblasts, which filled and covered all pores between the fibres. The new scaffold used in this study, made of synthetic and natural polymers, is a good candidate for applications in tissue engineering. It is therefore recommended to use PU in grafts or in wound dressing.