Original Research Report
Polymeric ionically conductive composite matrices and electrical stimulation strategies for nerve regeneration: In vitro characterization
Ohan S. Manoukian,
Scott Stratton,
Michael R. Arul,
Joshua Moskow,
Naseem Sardashti,
Xiaojun Yu,
Swetha Rudraiah,
Sangamesh G. Kumbar,
Ohan S. Manoukian
Department of Biomedical Engineering, University of Connecticut, Storrs, Connecticut
Department of Orthopedic Surgery, University of Connecticut Health, Farmington, Connecticut
These authors contributed equally to this work.Search for more papers by this authorScott Stratton
Department of Biomedical Engineering, University of Connecticut, Storrs, Connecticut
Department of Orthopedic Surgery, University of Connecticut Health, Farmington, Connecticut
These authors contributed equally to this work.Search for more papers by this authorMichael R. Arul
Department of Orthopedic Surgery, University of Connecticut Health, Farmington, Connecticut
Search for more papers by this authorJoshua Moskow
Department of Biomedical Engineering, University of Connecticut, Storrs, Connecticut
Department of Orthopedic Surgery, University of Connecticut Health, Farmington, Connecticut
Search for more papers by this authorNaseem Sardashti
Department of Biomedical Engineering, University of Connecticut, Storrs, Connecticut
Search for more papers by this authorXiaojun Yu
Department of Chemistry, Chemical Biology and Biomedical Engineering, Stevens Institute of Technology, Hoboken, New Jersey
Search for more papers by this authorCorresponding Author
Swetha Rudraiah
Department of Orthopedic Surgery, University of Connecticut Health, Farmington, Connecticut
Department of Pharmaceutical Sciences, University of Saint Joseph, Hartford, Connecticut
Correspondence to: S. Rudraiah; e-mail: [email protected] or S. G. Kumbar; e-mail: [email protected]Search for more papers by this authorCorresponding Author
Sangamesh G. Kumbar
Department of Biomedical Engineering, University of Connecticut, Storrs, Connecticut
Department of Orthopedic Surgery, University of Connecticut Health, Farmington, Connecticut
Correspondence to: S. Rudraiah; e-mail: [email protected] or S. G. Kumbar; e-mail: [email protected]Search for more papers by this authorOhan S. Manoukian,
Scott Stratton,
Michael R. Arul,
Joshua Moskow,
Naseem Sardashti,
Xiaojun Yu,
Swetha Rudraiah,
Sangamesh G. Kumbar,
Ohan S. Manoukian
Department of Biomedical Engineering, University of Connecticut, Storrs, Connecticut
Department of Orthopedic Surgery, University of Connecticut Health, Farmington, Connecticut
These authors contributed equally to this work.Search for more papers by this authorScott Stratton
Department of Biomedical Engineering, University of Connecticut, Storrs, Connecticut
Department of Orthopedic Surgery, University of Connecticut Health, Farmington, Connecticut
These authors contributed equally to this work.Search for more papers by this authorMichael R. Arul
Department of Orthopedic Surgery, University of Connecticut Health, Farmington, Connecticut
Search for more papers by this authorJoshua Moskow
Department of Biomedical Engineering, University of Connecticut, Storrs, Connecticut
Department of Orthopedic Surgery, University of Connecticut Health, Farmington, Connecticut
Search for more papers by this authorNaseem Sardashti
Department of Biomedical Engineering, University of Connecticut, Storrs, Connecticut
Search for more papers by this authorXiaojun Yu
Department of Chemistry, Chemical Biology and Biomedical Engineering, Stevens Institute of Technology, Hoboken, New Jersey
Search for more papers by this authorCorresponding Author
Swetha Rudraiah
Department of Orthopedic Surgery, University of Connecticut Health, Farmington, Connecticut
Department of Pharmaceutical Sciences, University of Saint Joseph, Hartford, Connecticut
Correspondence to: S. Rudraiah; e-mail: [email protected] or S. G. Kumbar; e-mail: [email protected]Search for more papers by this authorCorresponding Author
Sangamesh G. Kumbar
Department of Biomedical Engineering, University of Connecticut, Storrs, Connecticut
Department of Orthopedic Surgery, University of Connecticut Health, Farmington, Connecticut
Correspondence to: S. Rudraiah; e-mail: [email protected] or S. G. Kumbar; e-mail: [email protected]Search for more papers by this authorAbstract
Stem cell strategies and the use of electrical stimulation (ES) represent promising new frontiers for peripheral nerve regeneration. Composite matrices were fabricated by coating electrospun polycaprolactone/cellulose acetate micro–nanofibers with chitosan and ionically conductive (IC) polymers including, sulfonated polyaniline, and lignin sulfonate. These composite matrices were characterized for surface morphology, coating uniformity, ionic conductivity, and mechanical strength to explore as scaffold materials for nerve regeneration in conjunction with ES. Composite matrices measured conductivity in the range of 0.0049–0.0068 mS/m due to the uniform coating of sulfonated polymers on the micro–nanofibers. Thin films (2D) and composite fiber matrices (3D) of IC polymers seeded with human mesenchymal stem cells (hMSCs) were electrically stimulated at 0.5 V, 20 Hz for 1 h daily for 14 days to study the changes in cell viability, morphology, and expression of the neuronal-like phenotype. In vitro ES lead to changes in hMSCs' fibroblast morphology into elongated neurite-like structures with cell bodies for ES-treated and positive control growth factor-treated groups. Immunofluorescent staining revealed the presence of neuronal markers including β3-tubulin, microtubule-associated protein 2, and nestin in response to ES. © 2018 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater 107B: 1792–1805, 2019.
REFERENCES
- 1Taylor CA, Braza D, Rice JB, Dillingham T. The incidence of peripheral nerve injury in extremity trauma. Am J Phys Med Rehabil 2008; 87: 381–385.
- 2Grinsell D, Keating C. Peripheral nerve reconstruction after injury: A review of clinical and experimental therapies. Biomed Res Int 2014; 2014: 1–13.
- 3Anderson M, Shelke NB, Manoukian OS, Yu X, McCullough LD, Kumbar SG. Peripheral nerve regeneration strategies: Electrically stimulating polymer based nerve growth conduits. Crit Rev Biomed Eng 2015; 43:131–159.
- 4Al-Majed AA, Neumann CM, Brushart TM, Gordon T. Brief electrical stimulation promotes the speed and accuracy of motor axonal regeneration. J Neurosci 2000; 20: 2602–2608.
- 5Kline DG, Hudson AR. Nerve Injuries: Operative Results for Major Nerve Injuries, Entrapments, and Tumors. Philadelphia, PA, USA: Saunders; 1995.
- 6Manoukian O, Ahmad A, Marin C, James R, Mazzocca A, Kumbar S. Bioactive nanofiber dressings for wound healing. In: Ågren MS, editor. Wound Healing Biomaterials-Volume 2: Functional Biomaterials. Cambridge, UK: Elsevier; 2016. p. 451.
10.1016/B978-1-78242-456-7.00022-2 Google Scholar
- 7Lee P, Manoukian OS, Zhou G, Wang Y, Chang W, Yu X, Kumbar SG. Osteochondral scaffold combined with aligned nanofibrous scaffolds for cartilage regeneration. RSC Adv 2016; 6: 72246–72255.
- 8Stratton S, Manoukian OS, Patel R, Wentworth A, Rudraiah S, Kumbar SG. Polymeric 3D printed structures for soft-tissue engineering. J Appl Polym Sci 2017;135:45569.
- 9Manoukian OS, Matta R, Letendre J, Collins P, Mazzocca AD, Kumbar SG. Electrospun nanofiber scaffolds and their hydrogel composites for the engineering and regeneration of soft tissues. Methods Mol Biol 2017;1570: 261–278.
- 10Keilhoff G, Goihl A, Langnäse K, Fansa H, Wolf G. Transdifferentiation of mesenchymal stem cells into Schwann cell-like myelinating cells. Eur J Cell Biol 2006; 85: 11–24.
- 11Kingham PJ, Kalbermatten DF, Mahay D, Armstrong SJ, Wiberg M, Terenghi G. Adipose-derived stem cells differentiate into a Schwann cell phenotype and promote neurite outgrowth in vitro. Exp Neurol 2007; 207: 267–274.
- 12Caddick J, Kingham PJ, Gardiner NJ, Wiberg M, Terenghi G. Phenotypic and functional characteristics of mesenchymal stem cells differentiated along a Schwann cell lineage. Glia 2006; 54: 840–849.
- 13Brohlin M, Mahay D, Novikov LN, Terenghi G, Wiberg M, Shawcross SG, Novikova LN. Characterisation of human mesenchymal stem cells following differentiation into Schwann cell-like cells. Neurosci Res 2009; 64: 41–49.
- 14Mahay D, Terenghi G, Shawcross SG. Schwann cell mediated trophic effects by differentiated mesenchymal stem cells. Exp Cell Res 2008; 314: 2692–2701.
- 15Tohill M, Mantovani C, Wiberg M, Terenghi G. Rat bone marrow mesenchymal stem cells express glial markers and stimulate nerve regeneration. Neurosci Lett 2004; 362: 200–203.
- 16Janssen J. Advantages and disadvantages of GH/IGF-I combination treatment. Rev Endocr Metabol Disord 2009; 10: 157–162.
- 17Lee K, Silva EA, Mooney DJ. Growth factor delivery-based tissue engineering: General approaches and a review of recent developments. J Roy Soc Interface 2011; 8: 153–170.
- 18Brushart TM, Hoffman PN, Royall RM, Murinson BB, Witzel C, Gordon T. Electrical stimulation promotes motoneuron regeneration without increasing its speed or conditioning the neuron. J Neurosci 2002; 22: 6631–6638.
- 19Asensio-Pinilla E, Udina E, Jaramillo J, Navarro X. Electrical stimulation combined with exercise increase axonal regeneration after peripheral nerve injury. Exp Neurol 2009; 219: 258–265.
- 20Al-Majed AA, Brushart TM, Gordon T. Electrical stimulation accelerates and increases expression of BDNF and trkB mRNA in regenerating rat femoral motoneurons. Eur J Neurosci 2000; 12: 4381–4390.
- 21Kotwal A, Schmidt CE. Electrical stimulation alters protein adsorption and nerve cell interactions with electrically conducting biomaterials. Biomaterials 2001; 22: 1055–1064.
- 22Lee J-W, Serna F, Schmidt CE. Carboxy-endcapped conductive polypyrrole: Biomimetic conducting polymer for cell scaffolds and electrodes. Langmuir 2006; 22: 9816–9819.
- 23Haan N, Song B. Therapeutic application of electric fields in the injured nervous system. Adv Wound Care (New Rochelle) 2014; 3: 156–165.
- 24Salmons S, Ashley Z, Sutherland H, Russold MF, Li F, Jarvis JC. Functional electrical stimulation of denervated muscles: Basic issues. Artif Organs 2005; 29: 199–202.
- 25Guimard NK, Gomez N, Schmidt CE. Conducting polymers in biomedical engineering. Prog Polym Sci 2007; 32: 876–921.
- 26Guo B, Glavas L, Albertsson A-C. Biodegradable and electrically conducting polymers for biomedical applications. Prog Polym Sci 2013; 38(9): 1263–1286.
- 27Huang J, Zhang Y, Lu L, Hu X, Luo Z. Electrical stimulation accelerates nerve regeneration and functional recovery in delayed peripheral nerve injury in rats. Eur J Neurosci 2013; 38: 3691–3701.
- 28Huang L, Zhuang X, Hu J, Lang L, Zhang P, Wang Y, Chen X, Wei Y, Jing X. Synthesis of biodegradable and electroactive multiblock polylactide and aniline pentamer copolymer for tissue engineering applications. Biomacromolecules 2008; 9: 850–858.
- 29Rivers TJ, Hudson TW, Schmidt CE. Synthesis of a novel, biodegradable electrically conducting polymer for biomedical applications. Adv Funct Mater 2002; 12: 33–37.
- 30Balint R, Cassidy NJ, Cartmell SH. Conductive polymers: Towards a smart biomaterial for tissue engineering. Acta Biomater 2014; 10: 2341–2353.
- 31Langer R, Vacanti JP. Tissue engineering. Science 1993; 260: 920–926.
- 32Croisier F, Jérôme C. Chitosan-based biomaterials for tissue engineering. Eur Polym J 2013; 49: 780–792.
- 33Sakai S, Hashimoto I, Kawakami K. Synthesis of an agarose-gelatin conjugate for use as a tissue engineering scaffold. J Biosci Bioeng 2007; 103: 22–26.
- 34Jacobsen PAL, Rafaelsen J, Nielsen JL, Juhl MV, Theilgaard N, Larsen KL. Distribution of grafted β-cyclodextrin in porous particles for bone tissue engineering. Micropor Mesopor Mater 2013; 168: 132–141.
- 35Müller FA, Müller L, Hofmann I, Greil P, Wenzel MM, Staudenmaier R. Cellulose-based scaffold materials for cartilage tissue engineering. Biomaterials 2006; 27: 3955–3963.
- 36Duarte ARC, Mano JF, Reis RL. Preparation of starch-based scaffolds for tissue engineering by supercritical immersion precipitation. J Supercrit Fluids 2009; 49: 279–285.
- 37Wang M-D, Zhai P, Schreyer DJ, Zheng R-S, Sun X-D, Cui F-Z, Chen XB. Novel crosslinked alginate/hyaluronic acid hydrogels for nerve tissue engineering. Front Mater Sci 2013; 7: 269–284.
- 38Wang X, He J, Wang Y, Cui F-Z. Hyaluronic acid-based scaffold for central neural tissue engineering. Interface Focus 2012; 2: 278–291.
- 39Gumera C, Rauck B, Wang Y. Materials for central nervous system regeneration: Bioactive cues. J Mater Chem 2011; 21: 7033–7051.
- 40Hench LL, Polak JM. Third-generation biomedical materials. Science Feb 08 2002; 295: 1014–1017.
- 41James R, Nagarale RK, Sachan VK, Badalucco C, Bhattacharya PK, Kumbar SG. Synthesis and characterization of electrically conducting polymers for regenerative engineering applications: Sulfonated ionic membranes. Polym Adv Technol 2014; 25: 1439–1445.
- 42Willand MP, Nguyen MA, Borschel GH, Gordon T. Electrical stimulation to promote peripheral nerve regeneration. Neurorehabil Neural Repair 2016; 30: 490–496.
- 43Nagarale RK, Gohil GS, Shahi VK. Recent developments on ion-exchange membranes and electro-membrane processes. Adv Colloid Interface Sci 2006; 119: 97–130.
- 44Jiang X, Lim SH, Mao H-Q, Chew SY. Current applications and future perspectives of artificial nerve conduits. Exp Neurol 2010; 223: 86–101.
- 45Kumbar SG. Biodegradable polymeric compositions and methods of use in biomedical applications. US Patent 62671080, 2018.
- 46Kumbar HMD. Composite fibers and matrices thereof, PCT Patent Application No. PCT/US16/15104, 2016.
- 47James R, Kumbar SG, Laurencin CT, Balian G, Chhabra AB. Tendon tissue engineering: adipose-derived stem cell and GDF-5 mediated regeneration using electrospun matrix systems. Biomed Mater 2011; 6: 025011.
- 48James R, Toti US, Laurencin CT, Kumbar SG. Electrospun nanofibrous scaffolds for engineering soft connective tissues. Methods Mol Biol 2011; 726: 243–258.
- 49Kumbar SG, James R, Nukavarapu SP, Laurencin CT. Electrospun nanofiber scaffolds: Engineering soft tissues. Biomed Mater 2008; 3:034002.
- 50Lee P, Tran K, Chang W, Fang YL, Zhou G, Junka R, Shelke NB, Yu X, Kumbar SG. Bioactive polymeric scaffolds for osteochondral tissue engineering: in vitro evaluation of the effect of culture media on bone marrow stromal cells. Polym Adv Technol 2015; 26: 1476–1485.
- 51Lee P, Tran K, Chang W, Shelke NB, Kumbar SG, Yu X. Influence of chondroitin sulfate and hyaluronic acid presence in nanofibers and its alignment on the bone marrow stromal cells: Cartilage regeneration. J Biomed Nanotechnol 2014; 10: 1469–1479.
- 52Yeo MG, Kim GH. Preparation and characterization of 3D composite scaffolds based on rapid-prototyped PCL/β-TCP struts and electrospun PCL coated with collagen and HA for bone regeneration. Chem Mater 2011; 24: 903–913.
- 53Balasubramanian P, Roether JA, Schubert DW, Beier JP, Boccaccini AR. Bi-layered porous constructs of PCL-coated 45S5 bioactive glass and electrospun collagen-PCL fibers. J Porous Mat 2015; 22: 1215–1226.
- 54Lee P, Tran K, Zhou G, Bedi A, Shelke NB, Yu X, Kumbar SG. Guided differentiation of bone marrow stromal cells on co-cultured cartilage and bone scaffolds. Soft Matter 2015; 11: 7648–7655.
- 55Yu LM, Kazazian K, Shoichet MS. Peptide surface modification of methacrylamide chitosan for neural tissue engineering applications. J Biomed Mater Res A 2007; 82: 243–255.
- 56Junka R, Valmikinathan CM, Kalyon DM, Yu X. Laminin functionalized biomimetic nanofibers for nerve tissue engineering. J Biomater Tissue Eng 2013; 3: 494–502.
- 57Wei X-L, Wang Y, Long S, Bobeczko C, Epstein A. Synthesis and physical properties of highly sulfonated polyaniline. J Am Chem Soc 1996; 118: 2545–2555.
- 58Bellot M, Galandrin S, Boularan C, Matthies HJ, Despas F, Denis C, Javitch J, Mazères S, Sanni SJ, Pons V, Seguelas MH, Hansen JL, Pathak A, Galli A, Sénard JM, Galés C. Dual agonist occupancy of AT1-R-α2C-AR heterodimers results in atypical Gs-PKA signaling. Nat Chem Biol 2015; 11: 271–279.
- 59Shelke NB, Lee P, Anderson M, Mistry N, Nagarale RK, Ma X-M, Yu X, Kumbar SG. Neural tissue engineering: Nanofiber-hydrogel based composite scaffolds. Polym Adv Technol 2016; 27: 42–51.
- 60Kumbar SG, M. D. Harmon. Composite fibers and matrices thereof, Ed: Google Patents, 2018.
- 61Wei Y, Hudson S, Mayer J, Kaplan D. The crosslinking of chitosan fibers. J Polym Sci A Polym Chem 1992; 30: 2187–2193.
- 62Chen A-H, Liu S-C, Chen C-Y, Chen C-Y. Comparative adsorption of cu (II), Zn (II), and Pb (II) ions in aqueous solution on the crosslinked chitosan with epichlorohydrin. J Hazard Mater 2008; 154: 184–191.
- 63Shanahan M, Auriac Y. Water absorption and leaching effects in cellulose diacetate. Polymer 1998; 39: 1155–1164.
- 64Nada AA, James R, Shelke NB, Harmon MD, Awad HM, Nagarale RK, Kumbar SG. A smart methodology to fabricate electrospun chitosan nanofiber matrices for regenerative engineering applications. Polym Adv Technol 2014; 25: 507–515.
- 65Guo J, Berbano SS, Guo H, Baker AL, Lanagan MT, Randall CA. Cold sintering process of composites: Bridging the processing temperature gap of ceramic and polymer materials. Adv Funct Mater 2016; 26: 7115–7121.
- 66Kumbar SG, Nukavarapu SP, James R, Nair LS, Laurencin CT. Electrospun poly (lactic acid-co-glycolic acid) scaffolds for skin tissue engineering. Biomaterials 2008; 29: 4100–4107.
- 67Othman N, Piah M, Adzis Z, Ahmad H, Ahmad N. Simulation of voltage and electric-field distribution for contaminated glass insulator. Research and Development (SCOReD), 2013 I.E. Student Conference on 2013, 116–120.
- 68Soltani MH, Pichardo R, Song Z, Sangha N, Camacho F, Satyamoorthy K, Sangueza OP, Setaluri V. Microtubule-associated protein 2, a marker of neuronal differentiation, induces mitotic defects, inhibits growth of melanoma cells, and predicts metastatic potential of cutaneous melanoma. Am J Pathol 2005; 166: 1841–1850.
- 69Yuan A, Rao MV, Nixon RA. Neurofilaments at a glance, Ed: The Company of Biologists Ltd, 2012.
- 70Safaeijavan R, Soleimani M, Divsalar A, Eidi A, Ardeshirylajimi A. Biological behavior study of gelatin coated PCL nanofiberous electrospun scaffolds using fibroblasts. J Paramed Sci 2013; 5:67–73.
- 71Zavastin D, Cretescu I, Bezdadea M, Bourceanu M, Drăgan M, Lisa G, Mangalagiu I, Vasić V, Savić J. Preparation, characterization and applicability of cellulose acetate–polyurethane blend membrane in separation techniques. Colloids Surf A Physicochem Eng Asp 2010; 370: 120–128.
- 72Fernandes LL, Resende CX, Tavares DS, Soares GA, Castro LO, Granjeiro JM. Cytocompatibility of chitosan and collagen-chitosan scaffolds for tissue engineering. Polimeros 2011; 21: 1–6.
- 73Gregorio-Jauregui KM, Pineda MG, Rivera-Salinas JE, Hurtado G, Saade H, Martinez JL, et al. One-step method for preparation of magnetic nanoparticles coated with chitosan. J Nanomater 2012; 2012: 4.
- 74Mendes LC, Falco APS, Pinho MS, Marques PO. Sulfonated polyaniline: Influence of sulfonation routes on its thermal and structural characteristics. Mater Res 2011; 14: 466–471.
- 75Menczel JD, Prime RB. Thermal Analysis of Polymers: Fundamentals and Applications. Hoboken, New Jersey: John Wiley & Sons; 2014.
- 76Plueddemann EP. Interfaces in Polymer Matrix Composites: Composite Materials, Vol 6. Cambridge, MA, USA: Elsevier; 2016.
- 77Anseth KS, Bowman CN, Brannon-Peppas L. Mechanical properties of hydrogels and their experimental determination. Biomaterials 1996; 17: 1647–1657.
- 78Yim EK, Pang SW, Leong KW. Synthetic nanostructures inducing differentiation of human mesenchymal stem cells into neuronal lineage. Exp Cell Res 2007; 313: 1820–1829.
- 79Ge W, Ren C, Duan X, Geng D, Zhang C, Liu X, Chen H, Wan M, Geng R. Differentiation of mesenchymal stem cells into neural stem cells using cerebrospinal fluid. Cell Biochem Biophys 2015; 71: 449–455.
- 80Gunther K, Appelt-Menzel A, Kwok CK, Walles H, Metzger M. Rapid monolayer neural induction of induced pluripotent stem cells yields stably proliferating neural stem cells. J Stem Cell Res Ther 2016; 2016:1–6.
- 81Cheng Y-C, Tsao C-W, Chiang M-Z, Chung C-A, Chien C-C, Hu W-W, et al. Microfluidic platform for human placenta-derived multipotent stem cells culture and applied for enhanced neuronal differentiation. Microfluid Nanofluid 2015; 18: 587–598.
- 82Kai D, Jiang S, Low ZW, Loh XJ. Engineering highly stretchable lignin-based electrospun nanofibers for potential biomedical applications. J Mater Chem B 2015; 3: 6194–6204.
- 83Martínez-Campos E, Civantos A, Redondo JA, Guzmán R, Pérez-Perrino M, Gallardo A, et al. Cell adhesion and proliferation on sulfonated and non-modified chitosan films. AAPS PharmSciTech 2016;18:974–982.
- 84Bober P, Humpolíček P, Pacherník J, Stejskal J, Lindfors T. Conducting polyaniline based cell culture substrate for embryonic stem cells and embryoid bodies. RSC Adv 2015; 5: 50328–50335.
- 85Yang Z, Wang KK. Glial fibrillary acidic protein: From intermediate filament assembly and gliosis to neurobiomarker. Trends Neurosci 2015; 38: 364–374.
- 86Sensenbrenner M, Lucas M, Deloulme J-C. Expression of two neuronal markers, growth-associated protein 43 and neuron-specific enolase, in rat glial cells. J Mol Med 1997; 75: 653–663.
- 87Li Y, Cao J, Chen M, Li J, Sun Y, Zhang Y, Zhu Y, Wang L, Zhang C. Abnormal neural progenitor cells differentiated from induced pluripotent stem cells partially mimicked development of TSC2 neurological abnormalities. Stem Cell Rep 2017; 8: 883–893.
