J Korean Thyroid Assoc.  2015 May;8(1):88-97. 10.11106/cet.2015.8.1.88.

Regeneration of Recurrent Laryngeal Nerve using Polycaprolactone (PCL) Nerve Guide Conduit Coated with Conductive Materials

Affiliations
  • 1Department of Otorhinolaryngology-Head and Neck Surgery, School of Medicine, Inha University, Incheon, Korea. jylim@inha.ac.kr
  • 2Department of Chemical Engineering, School of Engineering, Inha University, Incheon, Korea.

Abstract

BACKGROUND AND OBJECTIVES
Recurrent laryngeal nerve (RLN) damage commonly occurs from a thyroid surgery and causes communication impairment, aspiration and dysphagia. The purpose of this study is to develop a polycaprolactone (PCL) nerve guide conduit (NGC) coated with conductive materials for facilitating regeneration from the RLN defects and to evaluate the usefulness of the PCL NGC coated with conductive materials in a rabbit model.
MATERIALS AND METHODS
The PCL NGCs coated with conductive materials were fabricated for this study. The types of conductive materials were single-walled carbon nanotubes (SWNTs) and poly (3,4-ethylenedioxythiophene): polystyrene sulfonate (PEDOT:PSS) which were coated on the PCL NGCs by layer-by-layer (LBL) assembly techniques. An 8-mm segment of left RLN was resected in 24 New Zealand white rabbits. Three different NGCs (PCL and PCL with two conductive materials) were interposed between both stumps and fixed with suture. For the assessment of functional regeneration, the vocal cord mobility was observed using endoscopic system after RLN stimulation, and the motion change was analyzed. The atrophies of thyroarytenoid muscle and nerve growth were evaluated by Hematoxylin-Eosin (H-E) and toluidine blue (T-B) staining, respectively. Immunohistochemical study using anti-neurofilament, S-100 staining was further performed to evaluate the nerve regeneration.
RESULTS
In endoscopic evaluation, the group with conductive PCL NGCs showed an improved tendency of vocal cord mobility compared to that of the other group. Nerve growth was observed with the time for 8 weeks in all groups and immunohistochemical staining revealed the expression of neurofilament and S-100 in regenerated nerve in all groups. The atrophies of thyroarytenoid muscle in the group with conductive PCL NGCs was also shown to be decreased compared to that of the nonconductive PCL NGC group.
CONCLUSION
The study shows that PCL NGC coated with conductive materials appears to be a good alternative option for the repair and regeneration of RNL damages.

Keyword

Recurrent laryngeal nerve; Regeneration; Nerve guide conduit

MeSH Terms

Atrophy
Deglutition Disorders
Laryngeal Muscles
Nanotubes, Carbon
Nerve Regeneration
Polystyrenes
Rabbits
Recurrent Laryngeal Nerve*
Regeneration*
Sutures
Thyroid Gland
Tolonium Chloride
Vocal Cords
Nanotubes, Carbon
Polystyrenes
Tolonium Chloride

Figure

  • Fig. 1. Fabrications of nerve guide conduits. (A) Micro- and nano- porous structured membrane template, (B) schematic illustration of coating process of conductive materials in membrane template, (C) application for nerve guide conduit with glue, (D) template NGC (left), conductive NGC coated with SWNTs (middle), conductive NGC coated with PEDOT:PSS (right).

  • Fig. 2. The animal operative procedure. (A) Careful dissection of left RLN, (B) 8 mm segmental resection of RLN, (C) NGC interposition, (D) schematic illustration of interposition of NGC between proximal and distal stumps. Asterisk: recurrent laryngeal nerve, arrow: resected nerve, arrowhead: interposed nerve guide conduit, respectively.

  • Fig. 3. Measurement of vocal fold movements. (A) Fully abducted position and fully adducted position were captured and measured the triangle area (angular points, abc), (B) The outcome of comparing the vocal cord movement among groups. The box and error bar denote means ± standard deviations. A: arytenoid, a: anterior commissure, b: fully abducted posterior commissure, c: fully adducted posterior commissure.

  • Fig. 4. Histologic evaluation of thyroarytenoid (TA) muscle atrophy. (A) The cross-sectional area of the TA muscles was measured by tracing outlines of the microscopic images, (B) the outcome of comparing the cross sectional area of the TA muscle among groups. The box and error bar denote means ± standard deviations. Arrowhead: atrophied TA muscle due to denervation of RLN, asterisk: compensated TA muscle due to reinnervation of RLN.

  • Fig. 5. Scanning electron microscope images of the template membrane and conductive materials coating membrane. (A) Micro-pore surface, (B) nano-pore surface, (C) cross section between inner and outer surface, (D) SWNT 3 layers membrane. (E) PEDOT:PSS 3 layers membrane.

  • Fig. 6. Histologic evaluation of regenerated RLN at 2, 4, 8 weeks after interposition. At 8 weeks, the regenerated nerve looked structurally normal in all groups. Arrows: regenerated RLNs.

  • Fig. 7. The expression of NF and S100 protein at 8 weeks. The NF and S100 protein in all groups were well expressed. DAPI (blue), NF, S100 protein (green).


Reference

References

1. Randolph GW. The importance of pre- and postoperative laryngeal examination for thyroid surgery. Thyroid. 2010; 20(5):453–8.
Article
2. Gardner GM, Smith MM, Yaremchuk KL, Peterson EL. The cost of vocal fold paralysis after thyroidectomy. Laryngoscope. 2013; 123(6):1455–63.
Article
3. Millesi H. Reappraisal of nerve repair. Surg Clin North Am. 1981; 61(2):321–40.
Article
4. Wang S, Cai Q, Hou J, Bei J, Zhang T, Yang J. et al. Acceleration effect of basic fibroblast growth factor on the regeneration of peripheral nerve through a 15-mm gap. J Biomed Mater Res A. 2003; 66(3):522–31.
5. Widmer MS, Gupta PK, Lu L, Meszlenyi RK, Evans GR, Brandt K. et al. Manufacture of porous biodegradable polymer conduits by an extrusion process for guided tissue regeneration. Biomaterials. 1998; 19(21):1945–55.
6. Kiyotani T, Teramachi M, Takimoto Y, Nakamura T, Shimizu Y, Endo K. Nerve regeneration across a 25-mm gap bridged by a polyglycolic acid-collagen tube: a histological and electrophysiological evaluation of regenerated nerves. Brain Res. 1996; 740(1-2):66–74.
Article
7. Pettersson J, McGrath A, Kalbermatten DF, Novikova LN, Wiberg M, Kingham PJ. et al. Muscle recovery after repair of short and long peripheral nerve gaps using fibrin conduits. Neurosci Lett. 2011; 500(1):41–6.
8. Keilhoff G, Stang F, Wolf G, Fansa H. Bio-compatibility of type I/III collagen matrix for peripheral nerve reconstruction. Biomaterials. 2003; 24(16):2779–87.
Article
9. Choi JS, Oh SH, An HY, Kim YM, Lee JH, Lim JY. Functional regeneration of recurrent laryngeal nerve injury during thyroid surgery using an asymmetrically porous nerve guide conduit in an animal model. Thyroid. 2014; 24(1):52–9.
Article
10. Oh SH, Kim JR, Kwon GB, Namgung U, Song KS, Lee JH. Effect of surface pore structure of nerve guide conduit on peripheral nerve regeneration. Tissue Eng Part C Methods. 2013; 19(3):233–43.
Article
11. Kim H, Shim BS. Stretchable conducting materials with multi-scale hierarchical structures for biomedical applications. Proceedings of SPIE 9172, Aug 27, 2014; Nanostructured Thin Films VII, 91720D. 2014; 9172(91720D):
12. Lee WT, Milstein C, Hicks D, Akst LM, Esclamado RM. Results of ansa to recurrent laryngeal nerve reinnervation. Otolaryngol Head Neck Surg. 2007; 136(3):450–4.
Article
13. Mu LC, Yang SL. Electromyographic study on end-to-end anastomosis of the recurrent laryngeal nerve in dogs. Laryngoscope. 1990; 100(9):1009–17.
Article
14. Nahm I, Shin T, Watanabe H, Maeyama T. Misdirected regeneration of injured recurrent laryngeal nerve in the cat. Am J Otolaryngol. 1993; 14(1):43–8.
Article
15. Crumley RL. Laryngeal synkinesis: its significance to the laryngologist. Ann Otol Rhinol Laryngol. 1989; 98(2):87–92.
Article
16. Nahm I, Shin T, Chiba T. Regeneration of the recurrent laryngeal nerve in the guinea pig: reorganization of motoneurons after freezing injury. Am J Otolaryngol. 1990; 11(2):90–8.
Article
17. de Ruiter GC, Malessy MJ, Yaszemski MJ, Windebank AJ, Spinner RJ. Designing ideal conduits for peripheral nerve repair. Neurosurg Focus. 2009; 26(2):E5.
Article
18. Azzam NA, Zalewski AA, Williams LR, Azzam RN. Nerve cables formed in silicone chambers reconstitute a perineurial but not a vascular endoneurial permeability barrier. J Comp Neurol. 1991; 314(4):807–19.
Article
19. Lee SK, Kim H, Shim BS. Graphene: an emerging material for biological tissue engineering. Carbon letters. 2013; 14(2):63–75.
Article
20. Malarkey EB, Fisher KA, Bekyarova E, Liu W, Haddon RC, Parpura V. Conductive single-walled carbon nanotube substrates modulate neuronal growth. Nano Lett. 2009; 9(1):264–8.
Article
21. Park SY, Park J, Sim SH, Sung MG, Kim KS, Hong BH. et al. Enhanced differentiation of human neural stem cells into neurons on graphene. Adv Mater. 2011; 23(36):H263–7.
22. del Valle LJ, Aradilla D, Oliver R, Sepulcre F, Gamez A, Armelin E. et al. Cellular adhesion and proliferation on poly(3,4-ethylenedioxythiophene): Benefits in the electroactivity of the conducting polymer. Eur Polym J. 2007; 43(6):2342–9.
23. Schmidt CE, Shastri VR, Vacanti JP, Langer R. Stimulation of neurite outgrowth using an electrically conducting polymer. Proc Natl Acad Sci U S A. 1997; 94(17):8948–53.
Article
24. Hwang JY, Shin US, Jang WC, Hyun JK, Wall IB, Kim HW. Biofunctionalized carbon nanotubes in neural regeneration: a mini-review. Nanoscale. 2013; 5(2):487–97.
Article
25. Jakubiec B, Marois Y, Zhang Z, Roy R, Sigot-Luizard MF, Dugre FJ. et al. In vitro cellular response to polypyrrole-coated woven polyester fabrics: potential benefits of electrical conductivity. J Biomed Mater Res. 1998; 41(4):519–26.
26. Iizuka T. Experimental studies on the nerve interception and atrophy of the intrinsic muscles of the larynx. Nihon Jibiinkoka Gakkai Kaiho. 1966; 69(2):176–95.
27. Pan YA, Misgeld T, Lichtman JW, Sanes JR. Effects of neurotoxic and neuroprotective agents on peripheral nerve regeneration assayed by time-lapse imaging in vivo. J Neurosci. 2003; 23(36):11479–88.
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