Preparation of HMWCNT/PLLA nanocomposite scaffolds for application in nerve tissue engineering and evaluation of their physical, mechanical and cellular activity properties

Abstract

This study was aimed to prepare biodegradable and porous nanocomposite scaffolds with microtubular orientation structure as a model for nerve tissue engineering by thermally induced phase separation (TIPS) method using dioxane as the solvent, crystalline poly (L-lactic acid) (PLLA) and multi-walled carbon nanotubes (MWCNTs). In order to overcome dispersion of MWCNTs in the PLLA matrix, heparinization of MWCNTs was performed. Solvent crystallization, oriented structure, the mean pore diameter and porosity percentage of the scaffolds were controlled by fundamental system parameters including temperature-gradient of the system, polymer solution concentration and carbon nanotube content. Scanning Electron Microscopy (SEM), ImageJ, software and dynamic mechanical thermal analysis (DMTA) were used to investigate the structural and mechanical properties. TEM observation was carried out for characterization of nanotube dispersion in PLLA. It was found that the scaffolds containing heparinized multi-walled carbon nanotubes (HMWCNTs) exhibited higher storage modulus, better carbon nanotube (CNT) dispersion and tubular orientation structure than those with non heparinized MWCNTs.In-vitro studies were also conducted by using murine P 19 cell line as a suitable model system to analyze neuronal differentiation over a 2-week period. Immunofluorescence and DAPI staining were used to confirm the cells' attachment and differentiation on the PLLA/HMWCNT nanocomposite scaffolds. Based on the results, we can conclude that the PLLA/HMWCNT scaffolds enhanced the nerve cell differentiation and proliferation, and therefore, acted as a positive cue to support neurite outgrowth.The PLLA scaffolds with different polymer concentration (1, 2.5 and 5 wt/v%) were prepared by solid-liquid phase separation method. The same samples containing 1, 2.5 and 5 wt% MWCNT were prepared as well. We should note that as the polymer concentration increases, the viscosity of the solution increases, and this provides better stability for nanotubes. Therefore, the minimum amount of polymer (1 wt/v%) was not used for the composite scaffolds.S. MOUNESI RAD ET AL.wileyonlinelibrary.com/journal/pat