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Table of Contents    
Year : 2021  |  Volume : 69  |  Issue : 2  |  Page : 318-325

Efficacy of Silicone Conduit in the Rat Sciatic Nerve Repair Model: Journey of a Thousand Miles

1 Department of Neurosurgery, All India Institute of Medical Sciences, Raebareli, Uttar Pradesh, India
2 Department of Neurosurgery, Sanjay Gandhi Post Graduate Institute of Medical Sciences, Lucknow, Uttar Pradesh, India
3 Veterinary Scientist, Animal House, Sanjay Gandhi Post Graduate Institute of Medical Sciences, Lucknow, Uttar Pradesh, India
4 Department of Plastic and Reconstruction Surgery, Sanjay Gandhi Post Graduate Institute of Medical Sciences, Lucknow, Uttar Pradesh, India
5 Department of Pathology, Sanjay Gandhi Post Graduate Institute of Medical Sciences, Lucknow, Uttar Pradesh, India

Date of Submission22-May-2020
Date of Decision13-Aug-2020
Date of Acceptance25-Oct-2020
Date of Web Publication24-Apr-2021

Correspondence Address:
Arun Kumar Srivastava
Department of Neurosurgery, Sanjay Gandhi Post Graduate Institute of Medical Sciences, Lucknow, Uttar Pradesh
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Source of Support: None, Conflict of Interest: None

DOI: 10.4103/0028-3886.314576

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 » Abstract 

Background: A lot of options have been tried for bridging the two ends of the injured nerves. Researchers have used decellularized nerve grafts, artificial materials and even nerve growth factors to augment functional recovery. These materials are either costly or inaccessible in developing world.
Objective: The study aimed to evaluate the efficacy of the silicone conduit in a rat sciatic nerve injury model.
Materials and Methods: 24 healthy Sprague–Dawley (SD) rats (250-300 grams; 8-10 weeks) were used and right sciatic nerve was exposed; transected and re-anastomosed by two different methods in 16 rats. In control group, n = 8 (Group I) the sciatic nerve was untouched; Group II (reverse nerve anastomosis, n = 8): 1-centimeter of nerve was cut and re-anastomosed by using 10-0 monofilament suture; Group III (silicone conduit, n = 8) 1-centimeter nerve segment was cut, replaced by silicone conduit and supplemented by fibrin glue]. Evaluation of nerve recovery was done functionally (pain threshold and sciatic functional index) over 3 months and histologically and electron microscopically.
Results: Functional results showed a trend of clinical improvement in Group III and II but recovery was poor and never reached up to normal. Histopathological and electron microscopic results showed an incomplete axonal regeneration in Groups II and III. Psychological analyses showed that no outwards signs of stress were present and none of the rats showed paw biting and teeth chattering.
Conclusion: The silicone conduit graft may be an economical and effective alternative to presently available interposition grafts, however for short segments only.

Keywords: Anastomosis, conduit, electron microscopic, glue, rat nerve model, sciatic, sciatic functional index
Key Message: Silicone conduit may be used as cheaper alternative for short segment rat nerve injury models.

How to cite this article:
Singh S, Srivastava AK, Baranwal AK, Bhatnagar A, Das KK, Jaiswal S, Behari S. Efficacy of Silicone Conduit in the Rat Sciatic Nerve Repair Model: Journey of a Thousand Miles. Neurol India 2021;69:318-25

How to cite this URL:
Singh S, Srivastava AK, Baranwal AK, Bhatnagar A, Das KK, Jaiswal S, Behari S. Efficacy of Silicone Conduit in the Rat Sciatic Nerve Repair Model: Journey of a Thousand Miles. Neurol India [serial online] 2021 [cited 2021 Jun 18];69:318-25. Available from:

Traumatic peripheral nerve injury is a devastating event having a significant neurological, economical, psychological and functional impact. A lot of options, both autografts or isografts, have been tried to bridge the two ends of injured nerves.[1] Researchers have used decellularized nerve grafts, artificial materials and even nerve growth factors to augment functional recovery of damaged nerve.[2] Most of these materials are either costly or inaccessible in a developing country. The aim of our study was to provide an immunological safe, cost-effective, neuro-effective, and readily available alternative for the use of interposition graft. Silicone sheets are cheap, readily available and can be transformed into tubes easily.[3] In this article, we showed a comparative analysis of clinical, psychological and histopathological outcomes in a rat sciatic nerve injury model.

 » Methodology Top

Study design

We did a single-center, prospective study to analyze the efficacy of silicone conduit in rat nerve model (2018–2019). This animal experiment was approved by the Institute Animal Ethics Committee with Ethical number PGI/IMP/IAEC/17/27.08.2015. All the animals were kept in the institute vivarium (maintained at standard room temperature and humidity). Pellet feed and Reverse Osmosed (R. O.) treated water was offered ad libitum to animals.

Study design and rationale

Rats were operated in small animal operation theater of vivarium and observed for the next 15 weeks. The timeline of 15 weeks was taken from work of Waitayawinyu et al.[4] He proposed that the rodents' nerve have a higher regenerative capacity, and so, observations beyond 15-weeks may be confounded. Moreover, the macrophages and Schwann cells require nearly 3–6 weeks to reach the injury site and clear the debris.[5] Once the Schwann cells reach to their target, they organize and provide a scaffold for regenerating axons.

Study parameters

Mechanical pain threshold, Sciatic function test, and open field analyses were performed by a two-blinded observers (AKB, SS) unaware of animal grouping details. The efficacy of nerve regeneration was evaluated using the following standardized parameters

  1. Mechanical pain threshold testing [Von Frey hair (VFH) sensitivity test] - Rat's paw was stimulated by a stiff nylon monofilament and 'paw withdrawal' or 'flinch' was observed for. The intensity of pain threshold in incremented by using filaments of different thickness sequentially
  2. Bodyweight gain: Body weight of rats was recorded weekly. It as an indirect marker of altered feeding habits due to pain. As such, a rat in more pain or agony do not prefer to explore and eats less and thereby either do not gain weight or gain less weight as compared to normal age-matched rat
  3. Sciatic functional index (SFI): It was analyzed using footprint score method. Ipsilateral paw was dabbed gently on in-house made ink-pad and then rat was allowed to walk over open field with white paper floor [Figure 1]. The maximum score was 11, when foot-print strokes of paw was normal and score was zero in case of hind-limb paralysis. The SFI comprises values between –11 (normal functionality) to 0 (complete loss of functionality)
  4. Open field analyses and Toe spread pattern: Rats were allowed to walk in guarded open field to analyze animals' gait and development of any behavioral signs. The impression of rat's paw, smeared in ink, was taken in white sheet. The distorted prints were allowed to dry and then analyzed subsequently [Figure 2]
  5. Histopathological and Electron microscopy study by single-blinded pathologist (SJ).
Figure 1: The rat was allowed to walk white paper floor and ink-pad impression of the operated hind-paw was taken. (a) The eleven points were analyzed for an objective assessment; (b) The eleven points included impression of five toes, five foot-pads and one tarsus area. Representative photograph of rat (Group II) with right hind-paw (operated) showing the animal shifting his body weight to tarsus and internal meta tarsal foot pads absent. (c and d) This lead to hollow-prints impression white paper floor and ink-pad

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Figure 2: (a and b) Post-surgery, rats were allowed to walk and pattern of hind-paw observed. One can notice the hollow pattern in center; (c) Another rat shows dragging pattern of hind-limb (Group II); (d) Rats were kept in isolated in cages during initial post-operative weeks and observed for limb movement

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Surgical technique

The rats were operated under general anesthesia with intraperitoneal injection of xylazine 7 mg/kg and ketamine 70 mg/kg. Operation site was prepared by shaving of lateral side of hind limb and mid back area. Post shaving cetrimide solution and betadine scrub were applied to make site sterile. Rats were positioned in prone. The sciatic nerve was exposed at the dorso-caudal region. We give an incision was 0.5 cm lateral to midline, extending 3-centimeters towards the tibio-femoral joint. The incision was deepened to muscle and fascia and fascio-muscular layers were dissected bluntly in order to expose sciatic nerve. Once the sciatic nerve is isolated, further procedure depends on the group-wise predefined manipulations. In Group I, nothing further was done and surgical wound was closed. In Group II, sciatic nerve transection was performed at two sites 1-centimeters apart. The cut segment was reversed and re-sutured by epineural microsutures using 10-0 nylon. The sutured segment was supplemented by fibrin glue. In Group, III the nerve segment was discarded and a silicone conduit (made from autoclaved silicone sheet) was used as interposition graft, with two remaining ends of nerve being inserted inside the conduit and thereafter fibrin glue was applied for approximately 30-seconds [Figure 3]. In all the rats, the wound was closed in single layer using silk 2-0. The skin was painted with povidone iodine antiseptic solution. The rats were kept in recovery chambers till complete anesthetic reversal and finally shifted to their home cages over soft bedding with free access to water and food until complete recovery. Postoperative analgesia and antibiotic were given for four days.
Figure 3: Operative photographs showing (a) nerve was exposed; (b) 1-centimeter of nerve was cut; (c) the two cut ends were exposed; (d and e) both the ends were inserted inside silicone conduit supplemented with glue

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Rats were euthanized at 15 weeks using carbon dioxide asphyxiation. The sutured nerve (groups II and III) and intact nerve (group I) were harvested and preserved for histopathological and electron microscopy studies, respectively.

 » Results Top

Surgical outcome

A total of 24 rats were operated in three groups [8 rats each], and were observed for neurological, psychological for the next 15 weeks and histopathological changes (after euthanization). None of rats had surgical site infection or death.

Functional outcome

We found a poor recovery among rats of Group III and II both, and functional recovery in Group III was even poorer. Between 8th to 11th week, the rate of change in body weight remained similar in all three groups. After 11th–12th week, the rats in Group II catch-up weight to reach normal level at the end of 14th week, but rats in Group III showed a decline in growth curve [Figure 4]. The pain threshold response was better in Group II in comparison to group III but overall mechanical allodynia was present in Group II and III. A similar outcome was found for the sciatic function index.
Figure 4: (a) x-axis represents weeks and y-axis represents body-weight in grams. The blue line represents control group, orange line represents Group II and grey line represents Group III.; (b) y-axis represents pain threshold (in gram-force). Red line represents pain threshold in rats of Group II and green line shows Group III results; (c) y-axis represents sciatic function index. The initial rate of recovery in both Group II and Group III looks similar but after 5th week, recovery in Group III was lesser; (d) strength of hind-paw in strength/gram (rota-rod) were also analysed (Group II rats) showing an incremental response

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Psychological outcome

We found a unique pattern of behavioral changes among two groups, that, no outwards signs of stress were present and none of the rats showed teeth chattering and paw biting. This means rats were not considering the operated limb as 'Alien' and at-least rats were not irritated or depressed.

Toe pattern study (Out-field analysis)

We observed an interesting pattern of toe-print among rats of Group II and III. The toe print of operated limb in group II resembles a contracture posture, with fingermarks more prominent than center (we call it 'hollow paw sign') and there was a decrease stance stage, and limb dragging in a circumferential manner [Figure 5]. Contrarily, in the group III, the paw was a little outwardly placed, with medial finger marks more prominent than lateral fingermarks. The electrophysiological study would have added more insight but still, gross inference can be obtained that the regeneration in conduit group (group II) was complete, while that in group III was selective.
Figure 5: (a) Lateral twisting of paw among the rats in group III; (b) Corresponding inkpad photograph showing lateral twisting; (b) The right hind paw shows 'Paw dragging' while putting body-weight on the medial side; (c and d) rats in group III show decreased points with toe (marked with black arrow) and only lateral prints seen. M Medial, L Lateral

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Histopathological pattern

Histological results showed that after three months there was incomplete axonal regeneration in groups II and III [Figure 6]. The axonal regeneration in group III was less than that in group II with a mild degree of fibrosis.
Figure 6: H and E stained sections of regenerating nerve fibres; (a) transverse section; (b) longitudinal section; (c) Toludine blue stained transverse section shows both myelinated and unmyelinated fibres; (d) Transmission electron microscope image showing process of regeneration and degeneration containing myelinated and unmyelinated fibers

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 » Discussion Top

The estimated incidence of traumatic peripheral nerve injury is nearly one million per year worldwide. The peripheral nerve injury has a poor impact on patient's quality of life and economic burden.[6] The sensory and motor functional defects may result in complete paralysis of the affected limb, partial paralysis with functionally ineffective limb or neuropathic pain. Peripheral nerves have ability to regenerate, but the physiological process is often unsatisfactory or inadequate. There is an utmost need to find innovative therapies to supplement or augment the physiological process of repair. With the availability of new biomaterials and synthetic conduits materials, researchers are focusing on innovative cheaper alternatives.[1],[2]

The functional recovery or nerve regeneration depends on many factors like (a) age of the patient, (b) level of injury, (c) mechanism of injury, (d) duration from injury to repair, and (e) patient's compliance. Other physiological factors include the speed of axonal regeneration, amount of re-innervation and plasticity.[7],[8],[9] This regeneration process is often misaligned due to positional mismatch between regenerating axons and target.[10] The misalignment is prevented by using a conduit as it prevents the multi-directional growth of regenerating axons and ameliorates ill-effects of chronic flexure contracture. Therefore, we explored an economical, cost-effective conduit for interposition graft.

We used silicone sheets and made conduits in our own laboratory. For this study, we used the healthy Sprague–Dawley (SD) rats (250–300 g; 8–10 weeks) because rat nerves are comparatively larger and more resilient. The functional assessment and psychological tests is universally standardized for rat models and these tests can be performed easily. We used fibrin glue in both Groups II and III, and propose to use glue for nerve repair as reinforcement over suture anastomosis. Although the epineural suturing is the most popular method for peripheral nerve anastomosis, still the technique has inherent disadvantages of excessive handling, leading to trauma and inflammation.[11],[12] Some authors believe that the suture material, for endoneural anastomosis, may also cause hindrance in regeneration by hampering blood supply of fascicles. On the other hand, a fibrin glue is biocompatible, biodegradable, promotes angiogenesis and tissue growth and has less foreign body reactions. The fibrin glue induces less inflammation, tissue necrosis and fibrosis.[13],[14] We followed a middle path and suture nerve ends at two ends and then reinforced the anastomosis by fibrin glue. For making conduits, we used pre-autoclaved silicone sheets and roll the sheet to make silicon tube of desired size. The silicone conduits are cheap, easily available, easy to prepare tubes from silicone sheets, have no risk of viral transmission. Our results revealed that there was a decrease in the values of SFI and VHF intensity postoperatively in group III and then increased gradually but not reach to the preoperative values and this coincides with observations made by Haapaniemi et al. and Suri et al.[15],[16] However, it was observed that the axonal regeneration was more better in group III when compared with group II and also, group II showed more deposition of collagen fibers. There was no correlation between the morphological findings and the functional outcomes between group II and group III. Similar results has been shown in other studies.[17],[18],[19] Meng et al. used human amniotic membrane, as a conduit, wrapped over 10-mL transected defect but did not found any significant results.[20] We also transected same length of nerve and found similar results. We believe that the transected nerve segment, especially in rat model, should have been smaller (5–8 mL). The poor recovery pattern in Group III may be explained by (a) dissected length was more in proportionate to size of animal, (b) period of observation was less, (c) sample size was less, and (d) a possibility of technical failure. Herein, we want to highlight an interesting post-euthanization morphological finding, that majority of nerves among Group III showed good recovery [Figure 7]. It was encouraging to see morphologically aligned regenerating nerve, and a longer follow-up may provide better results. Similarly, other authors have found that subjects showed a less cold intolerance, in a median nerve injury models, using silicone conduits.[21] The digital nerves repaired by similar biodegradable polyglycolic acid tubes show good sensory recovery.[22] Other studies show less aberrant motor regeneration and good functional recovery after conduit repait.[23],[24],[25] A similar silicone conduit was described by Liang et al. and Merolli et al.[26],[27] None of our rats in group II or III showed any signs of stress and the surgical wound was satisfactory in all of them These facts, indirectly suggests that rats were in healing stage and they could still sense their hind limb (even when not responding to sensory tests). Rats with complete sensory and motor weakness usually show autophagy or at least surgical-site bite-marks. The proportional weight gain in all three groups, further adds to our inference.
Figure 7: (a-f) Post-euthanization photograph of rats showing morphological recovery in operated nerves of Group II and III. The nerves were thinned out and showed good realignment

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'Flexion contracture' or a 'unique pattern' suggesting recovery

The toe print of operated limb in group II resemble a contracture posture, with fingermarks more prominent than center (we call it 'hollow paw sign') and there was decrease stance stage, and limb dragging in a circumferential manner. Contrarily, in Group III, the paw was little outwardly placed, with medial finger marks more prominent than lateral fingermarks. The electrophysiological study would have added more insight but still gross inference can be obtained that the regeneration in conduit group (group II) was complete, while that in group III was selective.

Lack of autotomy of their fingers as a psychological sign of 'stress-relieved' or 'recovering sensory afferents'

There was no sign of infection or inflammatory reaction at the surgical site in any of the rats included. We found a unique pattern of behavioral changes among two groups, that, no outwards signs of stress were present and none of the rats showed paw biting or teeth chattering.[28] This means rats were not considering the operated limb as 'Alien' and at least rats were not irritated or depressed.

Translation of rat nerve research from desk to table

Although the rat model is most popular in 'PubMed' search for peripheral repair, some authors believe that the translation is notoriously unreliable. In Group II, we could have used suture alone or glue alone.[29] The ramp-loaded test showed inferiority of 'glue alone' with presence of more inflammation, slow absorption, and toxicity.[30] However, the issue remained under-discussed is that whether rat-nerve injury model is ideal and really translated to human periphery nerve injury surgery. It is believed that a transection of short gap may take 1–2 weeks to heal, and do better with empty conduits, compared to those conduits filled with fillers (due to viscosity of filler substance); whereas long gap takes nearly 6-weeks to heal, and may do better with a filler (axon regenerate better in Schwann cell-friendly atmosphere).[29],[30]

A 'critical nerve gap' is defined as a defect of size more than which functional recovery is not possible without an interposition graft. In rats, the critical nerve gap is nearly 1.5 cm, in rabbits it is 3 cm, and in pigs and humans, the critical nerve gap is 4-centimeters.[31],[32] [Table 1] shows a review of the literature of similar studies where conduits were used in rat regeneration models.[33],[34],[35],[36],[37],[38],[39],[40],[41] Majority of authors have used Wistar rats,[16],[34],[36],[37],[39] while Zhou et al. used SD rats.[40] In these studies, authors have shown promising electron microscopy and electrophysiological tests in the rats where conduits were used. The authors have used various growth factors; for example, seeded with Schwann cells, human bone marrow cells-seeded muscle-stuffing, brain-derived neurotrophic factor bone marrow stromal cells or even autologous nerve grafts inside the conduits. In our study, we did not use such growth factors and this may be a responsible factor for poor functional outcome. In the future, we have planned to conduct a study using growth factors stuffed silicone conduit in our animal laboratory. We are working on a larger-sized Rat model study and have planned to use several cost-effective materials as well. Our animal laboratory shall be upgraded with advanced armamentarium and readout methods such as ACTI-meter and nerve conduction study.
Table 1: Review of literature showing various studies on rat nerve regeneration models

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 » Conclusion Top

In conclusion, this study showed that silicone conduit is an inferior alternative to autologous nerve graft, but has good functional recovery in the late stages. Although the result did not show a significant improvement in outcome, but an improving clinical trend was seen. These effects could have therapeutic importance in clinical settings and silicone conduits may be used as economical and effective alternative for short segment gap. Further studies, with larger sample size and longer follow-up, are warranted to justify our results statistically.


I want to acknowledge Mr. Satish Kumar Gautam, Assistant Technician, Animal House, Sanjay Gandhi Post Graduate Institute of Medical Sciences, Lucknow for his invaluable contribution and help in animal handling.

Financial support and sponsorship

Intramural project funding number A-01-PGI/IMP/69/2016.

Conflicts of interest

There are no conflicts of interest.

 » References Top

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Muheremu A, Ao Q. Past, present, and future of nerve conduits in the treatment of peripheral nerve injury. Biomed Res Int 2015;2015:237507. doi: 10.1155/2015/237507.  Back to cited text no. 3
Waitayawinyu T, Parisi DM, Miller B, Luria S, Morton HJ, Chin SH, et al. A comparison of polyglycolic acid versus type 1 collagen bioabsorbable nerve conduits in a rat model: An alternative to autografting. J Hand Surg Am 2007;32:1521-9.  Back to cited text no. 4
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  [Figure 1], [Figure 2], [Figure 3], [Figure 4], [Figure 5], [Figure 6], [Figure 7]

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