(25c) Reinforcement of Poly(caprolactone)/Gelatin Nerve Tissue Engineering Scaffolds By Graphene Oxide Nanosheets

Damage to the nervous system is devastating to patients and although methods such as nerve autografts and allografts have been developed, they have limitations such as shortage of donor nerves or rejection by the body’s immune system¹. In recent years, engineered materials for the regeneration of the nerve tissue have been proposed. These engineered materials must possess balanced characteristics in terms of biocompatibility, biodegradability, mechanical properties, surface properties, and permeability². To this end, composites of natural and synthetic polymers are designed. It is also desirable to mimic the structure of the extracellular matrix (ECM) which has a fibrous structure. One common method for the synthesis of nanometer fibers is electrispinning³. Ghasemi et al.⁴electrospan poly(e- caprolactone) or PCL and gelatin and showed that although the 50:50 ratio of PCL/gelatin has good wettability, it lacks mechanical integrity compared to the PCL/gelatin 70:30 nanofibers.

Recently carbon nanomaterials such as graphene have gained much attention due to their unique properties. Graphene is a 2-D sheet of sp² bonded carbons with high Young’s modulus and good electrical conductivity⁵. Graphene oxide (GO) is the oxidized form of graphene which has carboxyl and carbonyl groups on the edges, and epoxide and hydroxyl functional groups on its basal plane. Due to these functional groups, GO is hydrophilic. In this work we utilize GO as nanofiller for 70:30 PCL/gelatin electrospun scaffolds and report the morphology and biocompatibility of these structures towards neural cells.

EXPERIMENTAL METHODS

Graphene oxide was prepared through a modified Hummer’s method⁶. In a typical procedure, graphite powder (0.5 g) and NaNO₃ (0.5 g) were added to H₂SO4 (23 mL). Mixture was stirred at 0°C and KMnO₄ (3 g) was gradually added. The temperature was raised to 35°C and stirred for 24 hours. Afterwards DI water (140 mL) and H₂O₂(3 mL) were added until gas evolution ceased. The mixture was allowed to rest for 1 day. The supernatant was removed and the remaining particles were washed and centrifuged with HCl (1M) and water to remove remaining salts and acids. The aqueous solution of graphite oxide was ultrasonicated for 1 hour to achieve GO dispersion. This solution was dried at 40°C to yield the solid form of graphene oxide. Two dispersions of 0.5 wt% and 1 wt% of GO are prepared in via ultrasonication. PCL and gelatin are added to these solutions with a weight ratio of 70:30, and a total concentration of 5 wt%. A control solution containing only PCL and gelatin will also be prepared. These solutions will be electrospun at optimum conditions to prepare nanofibrous scaffolds.

The chemical properties of the synthesized GO nanosheets were characterized by FT-IR. XRD was used to show interlayer spacing of the sheets. AFM and TEM analyses were performed to determine sheet thickness and morphology respectively. Morphology, fiber diameter, and porosity of the scaffolds will be determined by SEM images. Mechanical properties of the scaffolds will also be determined. In order to observe neural biocompatibility of the samples, PC 12 cells will be proliferated for 2 days and Cell viability will be determined by MTT assay.

RESULTS AND DISCUSSION

FTIR and XRD analyses confirmed successful formation of GO nanosheets. Also, TEM images (in the attached file) showed the sheet like structure of these nanoparticles.

SEM images will be obtained to confirm desirable morphology for nerve regeneration. Moreover, mechanical properties and contact angle measurements will be obtained. Biocompatibility of these samples will be checked by the viability of neural cells.

CONCLUSION

We hypothesize that the addition of GO as a nanofiller will not only enhance the mechanical properties of the scaffolds, but will also make the scaffolds less hydrophobic due to its hydrophilic nature; hence making the scaffold suitable for nerve regeneration applications.

REFERENCES

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5. Zhu Y. et al., J. Adv. Mat. 22: 3906-3924, 2010

6. Hummers W. et al., J. Amer. Chem. Soc. 80: 1339-1339, 1958