Sciatic Nerve Repair In Rats Research Articles

Effects of Adipose-Derived Mesenchymal Stem Cells and Human Amniotic Membrane on Sciatic Nerve Repair in Rats

Background: Peripheral nerve injuries remain a great challenge for microsurgery despite the significant progress in recent decades. The current gold standard is autogenous nerve grafting with a success rate as low as 50% in long gaps. Current studies have focused on finding alternative methods for bridging nerve defects. Previous data have demonstrated the role of human amniotic membrane in stimulating neural regeneration. On the other hand, adipose-derived mesenchymal stem cells can differentiate into all three germ layers and could support nerve repair. The purpose of this study was to compare the role of the human amniotic membrane with and without adipose tissue stem cells in sciatic nerve injury with gap in rats. Objectives: We aimed to evaluate the effectiveness of the human amniotic membrane with and without adipose-derived mesenchymal stem cells in sciatic nerve injury with gap in rats. Methods: Twenty-four male Wistar rats in four random groups were used in our study. In the first group, the nerve gap was repaired using the inverse resected nerve segment (Control group), the second group was repaired with a human amniotic membrane (AM group), the third group was repaired with an amnion sheet with seeded adipose-derived mesenchymal stem cells (AM/ADMSCs group), and the last group was not repaired, and both stumps were sutured to muscles. Results: All the animals underwent the procedures and survived without complication. The sciatic function index and hot plate test results were significantly improved in the AM and AM/ADMSCs groups compared to the Control group (as a gold standard of care) (P>0.05). Based on histopathology findings, regenerative nerve fibers were seen in the implanted area of both AM and AM/ADMSCs groups; however, nerve fibers were surrounded by significant fibrosis (scar formation) in the AM/ADMSCs group. The axon count in the Control group was significantly higher than both experimental groups (P < 0.01). Conclusions: Our study showed the role of amniotic membrane in the promotion of nerve regeneration in sciatic nerve injury with a gap, but adding adipose-derived mesenchymal stem cells not only has no extra benefits, but also causes more tissue scar.

Effect of Oat (Avena Sativa L.) Extract on Experimental Sciatic Nerve Injury in Rats.

Peripheral nerve disorders are the most common neurological problems; therefore, it is important to intervene to treat or stop the resulting side effects. This study aimed to investigate the effect of oat extract on experimental sciatic nerve injury in rats. Totally, 50 adult male rats were divided into five groups (n=10). Group 1 was exposed to sham condition, and group 2 was regarded as the control group (nerve injury without treatment). Moreover, groups 3-5 were subjected to sciatic nerve injury, and they received oral gavages of the oat extract (100, 200, and 400 mg/kg), respectively. Subsequently, 2 and 4 weeks later, the rats were euthanized for pathological evaluation of nerve repair. The results showed an increase in the formation of the perineurium and epineurium dose in the oat-treated groups (100, 200, and 400 mg/kg), compared to the control group after 2 weeks (P<0.05). Furthermore, the presence of inflammatory cells in the oat extract-treated groups (100, 200, and 400 mg/kg) decreased, compared to that in the control group after 2 weeks (P<0.05). In addition, the swelling of the axon significantly decreased in the oat extract-treated groups (200 and 400 mg/kg), compared to the control group (P<0.05). However, the axon dose-dependently increased in oat-treated groups (100, 200, and 400 mg/kg), compared to that in the control group after 4 weeks (P<0.05). These results suggest that oat extract has positive effects on sciatic nerve repair in rats.

Research of a novel tissue-engineered nerve graft for sciatic nerve repair in rats

Objective To observe the effect of novel tissue engineered nerve grafts, which combined silk fibroin/collagen scaffold with a co-culture of Schwann cells (SCs) and adipose-derived stem cells (ADSCs) , to repair rat sciatic nerve defects. Methods From February, 2015 to August, 2016, the culture and purify SCs and ADSCs were isolated. And co-cultured at a ratio of 2∶1 and introduced into a silk fibroin (SF)/collagen scaffold to construct a tissue-engineered nerve conduit (TENC), which were transplanted to bridge 10 mm long sciatic nerve defects in rats. The experiment was divided randomly into 4 groups (10 rats/group): those bridged with plain SF/collagen scaffolds(Scaffold group), those bridged with TENCs (TENC group), those bridged with autografts (Autograft group) and those unoperated side (Normal group). The mechanical properties were examined using a universal testing machine(Instron 5865). Scanning electron microscopy was performed to observe the structure of the SF/collagen scaffold and the cells' growth. A series of electrophysiological examinations and morphological analyses were performed 12 weeks after surgery to evaluate the effect of the TENC on peripheral nerve regeneration. And One-way ANOVA was used to analyze the data. If the differences between groups were statistically significant, the Turkey's method was further applied for comparison. Results The plain SF/collagen scaffold showed appropriate pore size and good intercommunicating of holes. The cells were tightly attached to and partly coiled about the scaffold and exhibited either a spindle or a spherical shape. The results of the mechanical measurement revealed that the maximum and average Young’s moduli of the SF/collagen scaffold were (10.80 ± 0.30) MPa and (8.14 ± 0.20) MPa, respectively. The mechanical properties ensure that the scaffold could resist muscular contraction and maintain its shape unchanged for a considerable period of time after grafting. All rats in each group had achieved nerve defect regeneration in varying degrees. But in terms of the effect of the repaired nerve, those treated with TENC were similar to those with autologous nerve grafts but superior to those with plain SF/collagen scaffolds. Conclusion The TENC that combined silk fibroin/collagen scaffold with a co-culture of SCs and ADSCs had normal nerve-like structure, and can bridge sciatic nerve defect and promote nerve growth. Key words: Silk fibroin; Collagen; Schwann cells; Adipose-derived stem cells; Co-culture; Tissue engineered nerve

The Effect of Muscle Graft With Nerve Growth Factor and Laminin on Sciatic Nerve Repair in Rats.

Introduction:Peripheral nerve injury is one of the most common damages that lead to physical disability. Considering the similarity between the coatings of skeletal muscles and nerve fibers, we conducted this research to determine the effect of muscle graft with Nerve Growth Factor (NGF) and Laminin (L) on nerve repair.Methods:We cut a 10-mm length of the sciatic nerve from 42 female Wistar rats (Weight: 200±250 g) and equally divided the rats into three groups. In the muscle graft+NGF+laminin group, the degenerated skeletal muscle was sutured with proximal and distal ends of the transected sciatic nerve. Then, NGF (100 ng) and laminin (1.28 mg/mL) were injected into the muscle graft. In the muscle graft group, normal saline was injected into the muscle graft. In the control group, 10 mm of the sciatic nerve was removed without any treatment. Functional recovery was assessed based on Sciatic Functional Index (SFI). Also, tracing motor neurons and histological studies were performed to evaluate nerve repair. The obtained data were analyzed by ANOVA test.Results:The Mean±SD SFI value significantly increased in the muscle graft+NGF+laminin (−76.6±2.9) and muscle graft (−82.1±3.5) groups 60 days after the injury compared to the control group. The Mean±SD number of labeled motor neurons significantly increased in the muscle graft+NGF+laminin (78.6±3.1) and muscle graft (61.3±6.1) groups compared to the control group (P<0.001). The mean number of myelinated axons in the distal segments of the muscle graft+NGF+laminin increased significantly compared to the muscle graft group.Conclusion:These findings suggest that muscle graft followed by NGF and laminin administration have therapeutic effects on nerve repair.

Curcumin accelerates the repair of sciatic nerve injury in rats through reducing Schwann cells apoptosis and promoting myelinization

BackgroundSchwann cells (SCs) play an indispensable role in the repair and regeneration of injured peripheral nerve. Curcumin can reduce SCs apoptosis, and promote the regeneration and functional recovery of injured peripheral nerves. However, the corresponding mechanisms are not clear. ObjectiveThe article was aimed to explore the effect and corresponding mechanisms of curcumin on the repair of sciatic nerve injury in rats. MethodsAfter surgery induced sciatic nerve injury, the model rats were divided into three groups and treated with curcumin, curcumin+PD98059 and curcumin+IGF-1 respectively for 4days. The phosphorylation of Erk1/2 and Akt, and the expression of LC3-II, Beclin 1 and p62 were measured using western blotting. After treatment for 60days, myelination of the injured sciatic nerve was evaluated by MBP immunohistochemical staining and the expression of PMP22, Fibrin and S100 were determined using qRT-PCR and western blotting. In vitro, RSC96 cells were starved for 12h to induce autophagy, and received DMSO, curcumin, PD98059+curcumin, IGF–1+curcumin and BFA1 respectively. The phosphorylation of Erk1/2、Akt and the expression of LC3-II, Beclin 1, p62, PMP22, Fibrin and S100 were measured using western blotting, and the cell apoptosis was detected by flow cytometry. ResultsCurcumin could promote injury-induced cell autophagy, remyelination and axon regeneration in sciatic nerve of rats. In vitro, curcumin could accelerate cell autophagy through regulating autophagy related Erk1/2 and Akt pathway, prevent cell apoptosis and promote expression of PMP22 and S100, and reduced deposition of Fibrin in cultured RSC96 SCs. ConclusionsCurcumin could accelerate injured sciatic nerve repair in rats through reducing SCs apoptosis and promoting myelinization.

Expression of neutrophil gelatinase-associated lipocalin (NGAL) in peripheral nerve repair

Introduction: Trauma is one of the causes of peripheral nerve injuries. Free radicals increase after tissue damage. Free radicals are usually scavenged and detoxified by antioxidants. In this study, we assessed the antioxidative role of the NGAL molecule in sciatic nerve repair in rats. Materials and methods: The sciatic nerves of 40 rats were crushed and the total mRNA of samples from day 1 and 3 and week 1, 3, 5 post injury was extracted. The expression of the NGAL gene was confirmed by RT-PCR. For immunohistochemistry analysis, the samples were fixed in paraformaldehyde and cut in 20 micrometer slices by cryostat. Results: The expression of NGAL significantly upregulated in day 1, 3 and week 1 following the crushing of sciatic nerves in comparison with the intact nerves. Immunohistochemistry results also confirmed the protein expression of this gene. Discussion: The NGAL molecule showed upregulation in the degeneration process after nerve injury, so it may play an important role in nerve repair.

Use of adipose-derived stem cells to fabricate scaffoldless tissue-engineered neural conduits in vitro

Peripheral nerve injuries resulting from trauma or disease often necessitate surgical intervention. Although the gold standard for such repairs uses nerve autografts, alternatives that do not require invasive harvesting of autologous nerve tissues are currently being designed and evaluated. We previously established the use of scaffoldless engineered neural conduits (ENCs) fabricated from primary cells as one such alternative in sciatic nerve repair in rats [Baltich et al. (2010) In Vitro Cell Dev Biol Anim 46(5):438–444]. The present study establishes protocols for fabricating neural conduits from adipose-derived stem cells (ASCs) differentiated to either a fibroblast or neural lineage and co-cultured into a three-dimensional (3-D) scaffoldless tissue-ENC. Addition of ascorbic acid-2-phosphate and fibroblast growth factor (FGF)-2 to the medium induced and differentiated ASCs to a fibroblast lineage in more than 90% of the cell population, as confirmed by collagen I expression. ASC-differentiated fibroblasts formed monolayers, delaminated, and formed 3-D conduits. Neurospheres were formed by culturing ASCs on non-adherent surfaces in serum-free neurobasal medium with the addition of epidermal growth factor (EGF) and FGF-2. The addition of 10 ng EGF and 10 ng FGF-2 produced larger and more numerous neurospheres than treatments of lower EGF and FGF-2 concentrations. Subsequent differentiation to glial-like cells was confirmed by the expression of S100. ASC-derived fibroblast monolayers and neurospheres were co-cultured to fabricate a 3-D scaffoldless tissue-ENC. Their nerve-like structure and incorporation of glial-like cells, which would associate with regenerating axons, may make these novel, stem cell-derived neural conduits an efficacious technology for repairing critical gaps following peripheral nerve injury.

Effect of Hyaluronic Acid-Carboxymethylcellulose Solution on Perineural Scar Formation after Sciatic Nerve Repair in Rats

BackgroundScar tissue formation is the major cause of failure in peripheral nerve surgery. Use of a hyaluronic acid-carboxymethylcellulose (HA-CMC) membrane (Seprafilm) as a solid anti-adhesion barrier agent is one of the therapeutic approaches to reduce postoperative scar tissue formation. However, a solid membrane may not be suitable for repair of a weak peripheral nerve site. This study examined the effect of HA-CMC solution on perineural scar formation after peripheral nerve repair in rats.MethodsThe sciatic nerves of 40 rats were transected and then immediately repaired using 10-0 nylon. The nerves were divided randomly into two groups. Saline and HA-CMC solution were applied topically to the nerve repair sites in the control and experimental groups, respectively. Reoperation was performed at 3, 6, 9, and 12 weeks to assess scar tissue formation. The assessment included the quality of wound healing, presence of perinueral adhesion, cellular components of the scar tissue, thickness of the scar tissue and histomorphological organization of the repair site.ResultsTopical application of the HA-CMC solution significantly decreased the macroscopic nerve adherence score and the numbers of the cellular components such as fibroblasts and inflammatory cells (p < 0.05, Mann-Whitney U-test). The scar tissue formation index was significantly lower in the experimental group at 12 weeks than that in the control group (p < 0.05, Mann-Whitney U-test). The grading scores of the histomorphological axonal organization at the repair site were significantly higher in the experimental group than those in the control group at 12 weeks (p < 0.05, Mann-Whitney U-test). No evidence of wound dehiscence or inflammatory reactions against the HA-CMC solution was noted.ConclusionsTopical application of a HA-CMC solution is effective in reducing the perineural scar formation and adhesion after sciatic nerve repair in rats, and is effective in promoting peripheral nerve regeneration at the repair site.

Comparative Study of Scar Formation at the Site of Sciatic Nerve Repair in Rats

Purpose : To investigate the effectiveness and to compare the degree of axonal regeneration with the control after the topical application of dexamethasone, hyaluronic acid and amniotic fluid at the repair site of the peripheral nerve. Materials and Methods : The left sciatic nerves of 40 adult Sprague-Dawley rats were transected and neurorrhaphy was performed. The nerves were divided into four groups according to thesolution applied around the repair site: control group (group 1), 0.3 ml normal saline: experimental groups, 0.3 ml dexamethasone (group 2), 0.3 ml hyaluronic acid (group 3), and 0.3 ml amniotic fluid (group 4). The thickness of the perineural scar tissue, the count of the myelinated nerve fiber and axonal regeneration on the repair site were evaluated electromyographically and histologically 4 and 12 weeks after surgery. Results : The peak amplitude of the compound motor action potential was observed in group 2, at postoperative 4 weeks and group 3 at postoperative 12 weeks. The increase in the number of myelinated nerve fibers was most prominently in group 3 at 4 and 12 weeks after surgery. The degree of perineural scarring was lowest in group 2 at postoperative 4 weeks and in group 3 at postoperative 12 weeks. Conclusion : A single dose topical application of Dexamethasone, HA and Amniotic fluid is more effective in preventing perineural scar formation as well as in facilitating the nerve regeneration process in a rat hind-limb model than using saline.

Static footprint analysis: a time‐saving functional evaluation of nerve repair in rats

Walking track analysis is a widely accepted technique for functional evaluation after sciatic nerve repair in rats, but it is labour-intensive. In 2000, Bervar described a time-saving digitised static footprint analysis. In that study there were good correlations between the traditional sciatic function index (SFI) and the newly-developed static sciatic index (SSI) and static toe spread factor (TSF), respectively. Despite promising results, static footprint analysis is still not widely used. The present study was designed to validate it. After transection of the sciatic nerve, end-to-end repair was assessed using video recorded dynamic and static footprints in 45 Wistar rats. We found an even better correlation between the SFI and both the SSI and the static TSF. In conclusion, static footprint analysis is a time-saving and easy technique for accurate functional assessment of peripheral nerve regeneration in rats.