Conductive nerve conduit with piezoelectric properties for enhanced PC12 differentiation

Abstract

Restoration of nerve tissue remains highly challenging, mainly due to the limited regeneration capacity of the nervous system and the development of fibrosis. This limitation necessitates designing new nerve guidance channel to promote nerve repairing. In this study, we developed a novel core/shell conduit to induce PC12 differentiation. Co-electrospinning method was utilized to produce a fibrous shell containing polycaprolactone/polyvinylidene fluoride PCL/PVDF, gelatin and polyaniline/graphene (PAG) nanocomposite. The core section of the conduit was filled with chitosan-gelatin hydrogel containing PAG and ZnO nanoparticles. Such conduit shows antibacterial activity, electrical conductivity and piezoelectric property. The effect of such engineered conduit on PC12 differentiation was investigated by analyzing differentiation markers Nestin and microtubule-associated protein 2 (MAP2) through immunocytochemistry and PCR-RT techniques. The result revealed that such conduit could significantly induce Nestin and MAP2 gene expression in the PC12 cells and, thus, it is a viable option for effective cell differentiation and nerve regeneration.

Figure 1

The photometric image of the conduit (a), the core–shell structure with its size (b).

Figure 2

FESEM micrographs and the mean fibers diameter of PAG0 (a), PAG2 (b) nanofibrous shell. FTIR spectra of PAG0 and PAG2 fibrous mats (shell of conduits) (c); the average contact angle of PAG2 (shell of conduit) (d); FESEM micrographs of the CS–GEL/ZnO (1%)/PAG (2%) hydrogel (e).

Figure 3

Degradation of the conduit without and with 2 wt.% PAG nanoparticles (PCN0, PCN2) after 3, 7 and 21 days. The obtained results are presented as the mean ± SD of at least three replicates. *p < 0.05, ***p < 0.001.

Figure 4

The effect of PAG nanocomposite on the output voltage of the conduit (PCN0 vs. PCN2).

Figure 5

FESEM micrographs of PC12 cell adhesion and proliferation on the PCN0 (a) and the PCN2 (b) conduits.

Figure 6

Anti-bacterial test of nanofibrous scaffold PAG0 and PAG2 without and with 2% PAG nanocomposit in shell structure.

Figure 7

Nestin gene expression in the differentiated PC12 cells on PCN0 and PCN2 neural conduction channel (a); MAP2 gene expression in the differentiated PC12 cells on PCN0 and PCN2 neural conduction channel (b). The obtained results are presented as the mean ± SD of at least three replicates. *p < 0.05, **p < 0.01, ***p < 0.001.

Figure 8

Immunostaining micrographs of PC12 cells cultured on the PCN0 conduit (neural guidance channel without PAG nanocomposite) and PCN2 conduit (neural guidance channel containing 2 wt.% PAG nanocomposite in core and shell structure).