The Effect of Electrical Stimulation on Growth and Proliferation of Neural Cells Using Conductive Nanofibrous Scaffolds

Document Type : Research Paper

Authors

1 Hazrate Masoumeh University, P.O. Box: 3736175514, Qom, Iran

2 Department of Textile Engineering, Amirkabir University of Technology, P.O. Box: 15875-4413, Tehran, Iran

Abstract

Hypothesis: Nowadays, the use of scaffolds in tissue engineering to repair and regenerate human lesions, including nervous injuries has been widely considered. Also, nanofibrous scaffolds due to their structural similarity with the extracellular matrix (ECM) in the body are found to be suitable substrates for cell growth. Therefore, the main focus of the present work is on the production of conductive nanofibrous scaffolds for neural cell culture and their electrical stimulation performance.
Methods: Two biocompatible polymers including polycaprolactone (PCL) and poly(lactic-co-glycolicacid) (PLGA) were used as main materials, and polyaniline (PANI) was applied as a conductive polymer to create conductivity in the substrates. After determination and optimization of the electrospinning process factors, 4 types of nanofibrous scaffolds with 4 levels of conductive polymer (0%, 1%, 10% and 18%) were prepared. To investigate the effect of scaffolds' conductivity and electrical stimulation on the nerve cells behavior, a plate with steel electrodes was designed to apply electrical field to the scaffolds during cell culture experiments.
Findings: SEM, Dino-Lite digital microscopy, Potentiostat-Galvanostat and 3-(4,5-dimethylthiazed-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay were used to study the properties of scaffolds including hydrophilicity, conductivity, fiber diameter and the results of cell culture. By investigation of the physical properties of the scaffolds it was shown that increasing the amount of PANI in scaffolds causes significant drop in the fiber diameter and hydrophilicity. In cell culture experiment, shape and proliferation of nerve cells were studied. MTT assay and SEM images showed that electrical stimulation, proportional to the amount of polyaniline, enhanced neurite outgrowth compared to the scaffolds that were not subjected to electrical stimulation. Furthermore, proliferation of cells on conductive scaffolds (by 10% v/w of PANI) increased and subsequently the cell proliferation decreased with increasing conductive polymer content due to its toxicity.

Keywords


  1. Ahn H.S., Hwang J.Y., Kim M.S., Lee J.Y., Kim J.W., and Kim H.S., Carbon-nanotube-Interfaced Glass Fiber Scaffold for Regeneration of Transected Sciatic Nerve, Acta Biomaterialia, 13, 324-334, 2015.
  2. Chan K.M., Gordon T., Zochodne D.W., and Power H.A., Improving Peripheral Nerve Regeneration: from Molecular Mechanisms to Potential Therapeutic Targets, Experiment. Neurology, 261, 826-835, 2014.
  3. Zamani F., Amani-Tehran M., Latifi M., and Shokrgozar M.A., The Influence of Surface Nanoroughness of Electrospun PLGA Nanofibrous Scaffold on Nerve Cell Adhesion and Proliferation, J. Mater. Sci. Mater. Med.,24, 1551-1560, 2013.
  4. Rahimi-Tanha N. and Nouri M., Core/Shell Nanofibers of Silk Fibroin/Polyvinyl Alcohol: Structure and Controlled Release Behavior, Iran. J. Polym. Sci. Technol. (Persian)30, 473-488, 2018.
  5. Baghersad S.,  Mansurnezhad R., Ghasemi- Mobarakeh L., Molahosseini H., and Morshed M., Coating of Silk Fabrics by PVA/Ciprofloxacin HCl Nanofibers for Biomedical Applications, Iran. J. Polym. Sci. Technol. (Persian)29, 171-184, 2016.
  6. Norouzi M., Ghasemi-Mobarakeh L., and Morshed M., Fabrication of Antibacterial Poly(vinyl alcohol) Microfibers Mat for Wound Dressing Application, Iran. J. Polym. Sci. Technol. (Persian)29, 15-25, 2016.
  7. Nouri M., Mokhtari J., Salmani L., and Sadeghieh K., Electrospinning of Silk Fibroin/β-Cyclodextrin Nanofibers for Controlled Drug Release, Iran. J. Polym. Sci. Technol. (Persian)29, 89-100, 2016.
  8. Zamani F., Amani-Tehran M., Latifi M., Shokrgozar M., and Zaminy A., Promotion of Spinal Cord Axon Regeneration by 3D Nanofibrous Core–Sheath Scaffolds, J. Biomed. Mater. Res., 102A, 506-513, 2014.
  9. Abadi F.J.H., Amani-Tehran M., Zamani F., Nematollahi M., Ghasemi-Mobarakeh L., and Nasr-Esfahani M.H., Effect of Nanoporous Fibers on Growth and Proliferation of Cells on Electrospun Poly(ϵ-caprolactone) Scaffolds, Int. J. Polym. Mat. Polym. Biomat.,63, 57-64, 2014.
  10. Mokhtari F., Salehi M., Zamani F., Hajiani F., Zeighami F., and Latifi M., Advances in Electrospinning: The Production and Application of Nanofibres and Nanofibrous Structures, Text. Prog., 48, 119-219, 2016.
  11. Ghasemi-Mobarakeh L., Prabhakaran M.P., Morshed M., Nasr-Esfahani M.H., and Ramakrishna S., Electrical Stimulation of Nerve Cells Using Conductive Nanofibrous Scaffolds for Nerve Tissue Engineering, Tissue Eng., 15A, 3605-3619, 2009.
  12. Pedrotty D., Koh J., Davis B., Taylor D.A., Wolf P., and Niklason L.E, Engineering Skeletal Myoblasts: Roles of 3-D Culture and Electrical Stimulation, Am. J. Physiol. Heart Circ. Physiol., 288, 1620-1626, 2005.
  13. Dodel M., Nejad N.H., Bahrami S.H., Soleimani M., Amirabad L.M., and Hanaee-Ahvaz H., Electrical Stimulation of Somatic Human Stem Cells Mediated by Composite Containing Conductive Nanofibers for Ligament Regeneration, Biologicals, 46, 99-107, 2017.
  14. Shi G., Zhang Z., and Rouabhia M., The Regulation of Cell Functions Electrically Using Biodegradable Polypyrrole–Polylactide Conductors, Biomaterials, 29, 3792-3798, 2008.
  15. Ghasemi-Mobarakeh L., Prabhakaran M., and Morshed M., Application of Conductive Polymers, Scaffolds and Electrical Stimulation for Nerve Tissue Engineering, J. Tissue Eng. Regen Med., 5, 17-35, 2011.
  16. Shi G., Rouabhia M., Meng S., and Zhang Z., Electrical Stimulation Enhances Viability of Human Cutaneous Fibroblasts on Conductive Biodegradable Substrates, J. Biomed. Mater. Res., 84A, 1026-1037, 2007.
  17. Hopkins A.R., Rasmussen P.G., Characterization of Solution and Solid State Properties of Undoped and Doped Polyanilines Processed from Hexafluoro-2-propanol, Macromolecules, 29, 7838-7846, 1996.
  18. Bidez P.R., Li S., MacDiarmid A.G., Venancio E.C., Wei Y., and Lelkes P.I., Polyaniline, an Electroactive Polymer, Supports Adhesion and Proliferation of Cardiac Myoblasts, J. Biomater. Sci., Polym Edition, 17, 199-212, 2006.
  19. Zamani, F., Amani-Tehran M., Zaminy A., and Shokrgozar M., Conductive 3D Structure Nanofibrous Scaffolds for Spinal Cord Regeneration, Fiber. Polym., 18, 1874-1881, 2017.
  20. Li M., Guo Y., Wei Y., MacDiarmid A.G., and Lelkes P.I., Electrospinning Polyaniline-Contained Gelatin Nanofibers for Tissue Engineering Applications, Biomaterials, 27, 2705-2715, 2006.
  21. Armentano I., Dottori M., Fortunati E., Mattioli S., and Kenny J., Biodegradable Polymer Matrix Nanocomposites for Tissue Engineering: A Review, Polym. Degrad. Stabil., 95, 2126-2146, 2010.
  22. Zamani F., Latifi M., Amani-Tehran M., and Shokrgozar M.A., Effects of PLGA Nanofibrous Scaffolds Structure on Nerve Cell Directional Proliferation and Morphology, Fiber. Polym., 14, 568-702, 2013.
  1. Santos N.M., Cicuéndez M., Holz T.V., Silva V.S., Fernandes A.J.S., and Vila M., Diamond-Graphite Nanoplatelet Surfaces as Conductive Substrates for the Electrical Stimulation of Cell Functions, ACS Appl. Mater. Interfac, 9, 1331-1342, 2017.