OBJECTIVE: To investigate the effect of folic acid coated-crosslinked urethane-doped polyester elastomer (fCUPE) nerve conduit in repairing long distance peripheral nerve injury. METHODS: Thirty-six 3-month-old male Sprague Dawley rats weighing 180-220 g were randomly assigned to 3 groups, each consisting of 12 rats: CUPE nerve conduit transplantation group (group A), fCUPE nerve conduit transplantation group (group B), and autologous nerve transplantation group (group C), the contralateral healthy limb of group C served as the control group (group D). A 20-mm-long sciatic nerve defect model was established in rats, and corresponding materials were used to repair the nerve defect according to the group. The sciatic function index (SFI) of groups A-C was calculated using the Bain formula at 1, 2, and 3 months after operation. The nerve conduction velocity (NCV) of the affected side in groups A-D was assessed using neuroelectrophysiological techniques. At 3 months after operation, the regenerated nerve tissue was collected from groups A-C for S-100 immunohistochemical staining and Schwann cell count in groups A and B to compare the level of nerve repair and regeneration in each group. RESULTS: At 3 months after operation, the nerve conduits in all groups partially degraded. There was no significant adhesion between the nerve and the conduit and the surrounding tissues, the conduit was well connected with the distal and proximal nerves, and the nerve-like tissues in the conduit could be observed when the nerve conduit stents were cut off. SFI in group A was significantly higher than that in group C at each time point after operation and was significantly higher than that in group B at 2 and 3 months after operation ( P<0.05). There was no significant difference in SFI between groups B and C at each time point after operation ( P>0.05). NCV in group A was significantly slower than that in the other 3 groups at each time point after operation ( P<0.05). The NCV of groups B and C were slower than that of group D, but the difference was significant only at 1 month after operation ( P<0.05). There was no significant difference between groups B and C at each time point after operation ( P>0.05). Immunohistochemical staining showed that the nerve tissue of group A had an abnormal cavo-like structure, light tissue staining, and many non-Schwann cells. In group B, a large quantity of normal neural structures was observed, the staining was deeper than that in group A, and the distribution of dedifferentiated Schwann cells was obvious. In group C, the nerve bundles were arranged neatly, and the tissue staining was the deepest. The number of Schwann cells in group B was (727.50±57.60) cells/mm ², which was significantly more than that in group A [(298.33±153.12) cells/mm ²] ( t=6.139, P<0.001). CONCLUSION The fCUPE nerve conduit is effective in repairing long-distance sciatic nerve defects and is comparable to autologous nerve grafts. It has the potential to be used as a substitute material for peripheral nerve defect transplantation.
Rats ; Animals ; Male ; Rats, Sprague-Dawley ; Polyesters ; Peripheral Nerve Injuries/surgery* ; Elastomers ; Urethane ; Sciatic Nerve/injuries* ; Carbamates ; Nerve Tissue ; Nerve Regeneration/physiology*

A previous study has indicated that Krüppel-like factor 7 (KLF7), a transcription factor that stimulates Schwann cell (SC) proliferation and axonal regeneration after peripheral nerve injury, is a promising therapeutic transcription factor in nerve injury. We aimed to identify whether inhibition of microRNA-146b (miR-146b) affected SC proliferation, migration, and myelinated axon regeneration following sciatic nerve injury by regulating its direct target KLF7. SCs were transfected with miRNA lentivirus, miRNA inhibitor lentivirus, or KLF7 siRNA lentivirus in vitro. The expression of miR146b and KLF7, as well as SC proliferation and migration, were subsequently evaluated. In vivo, an acellular nerve allograft (ANA) followed by injection of GFP control vector or a lentiviral vector encoding an miR-146b inhibitor was used to assess the repair potential in a model of sciatic nerve gap. miR-146b directly targeted KLF7 by binding to the 3'-UTR, suppressing KLF7. Up-regulation of miR-146b and KLF7 knockdown significantly reduced the proliferation and migration of SCs, whereas silencing miR-146b resulted in increased proliferation and migration. KLF7 protein was localized in SCs in which miR-146b was expressed in vivo. Similarly, 4 weeks after the ANA, anti-miR-146b increased KLF7 and its target gene nerve growth factor cascade, promoting axonal outgrowth. Closer analysis revealed improved nerve conduction and sciatic function index score, and enhanced expression of neurofilaments, P0 (anti-peripheral myelin), and myelinated axon regeneration. Our findings provide new insight into the regulation of KLF7 by miR-146b during peripheral nerve regeneration and suggest a potential therapeutic strategy for peripheral nerve injury.
Animals ; Cell Movement ; genetics ; Cell Proliferation ; genetics ; Disease Models, Animal ; Female ; Ganglia, Spinal ; cytology ; Gene Expression Regulation ; genetics ; physiology ; HEK293 Cells ; Humans ; Kruppel-Like Transcription Factors ; genetics ; metabolism ; Male ; MicroRNAs ; genetics ; metabolism ; Motor Endplate ; genetics ; Myelin P0 Protein ; metabolism ; Nerve Regeneration ; genetics ; physiology ; Nerve Tissue Proteins ; metabolism ; RNA, Small Interfering ; genetics ; metabolism ; Rats ; Rats, Sprague-Dawley ; Rats, Wistar ; Sciatic Neuropathy ; metabolism ; surgery ; therapy

Aged ; Aged, 80 and over ; Arthroplasty ; methods ; Hip Joint ; surgery ; Hip Prosthesis ; adverse effects ; Humans ; Male ; Nerve Compression Syndromes ; etiology ; Sciatic Neuropathy ; etiology

There have been many attempts for regeneration of peripheral nerve injury. In this study, we examined the in vivo effects of non-differentiated and neuronal differentiated adipose-derived stem cells (ADSCs) in inducing the neuronal regeneration in the Sprague-Dawley (SD) rats undergoing nerve defect bridged with the PCL nanotubes. Then, we performed immunohistochemical and histopathologic examinations, as well as the electromyography, in three groups: the control group (14 sciatic nerves transplanted with the PCL nanotube scaffold), the experimental group I (14 sciatic nerves with the non-differentiated ADSCs at a density of 7x105 cells/0.1 mL) and the experimental group II (14 sciatic nerves with the neuronal differentiated ADSCs at 7x105 cells/0.1 mL). Six weeks postoperatively, the degree of the neuronal induction and that of immunoreactivity to nestin, MAP-2 and GFAP was significantly higher in the experimental group I and II as compared with the control group. In addition, the nerve conduction velocity (NCV) was significantly higher in the experimental group I and II as compared with the control group (P=0.021 and P=0.020, respectively). On the other hand, there was no significant difference in the NCV between the two experimental groups (P>0.05). Thus, our results will contribute to treating patients with peripheral nerve defects using PCL nanotubes with ADSCs.
Adipose Tissue/cytology ; Animals ; Cell Differentiation ; Electromyography ; Male ; Nanotubes ; *Nerve Regeneration ; Nerve Tissue Proteins/immunology ; Nestin/immunology ; Neural Conduction/physiology ; Peripheral Nerve Injuries/*surgery ; Phosphoprotein Phosphatases/immunology ; Polyesters/*therapeutic use ; Rats ; Rats, Sprague-Dawley ; Sciatic Nerve/injuries/surgery ; Stem Cell Transplantation/*methods ; Stem Cells/*cytology ; Tissue Engineering/methods

mRNAs of alpha-adrenoceptor (α-AR) subtypes are found in neurons in dorsal root ganglion (DRG) and change after peripheral nerve injury. In this study, the distribution of α-AR subtype proteins was studied in L5 DRG of normal rats and rats with chronic constriction injury of sciatic nerve (CCI). Using immunofluorescence technique, it was found that α1A-, α1B-, and α2A-AR proteins were expressed in large, medium, and small size neurons in normal DRG, and significantly increased in all size neurons 14 days after CCI. α1D- and α2C-AR was also expressed in all size neurons in normal DRG. However, α1D-AR was significantly increased and α2C-AR was decreased in small size neurons 14 days post CCI. α2B-AR neurons were not detectable in normal and CCI DRG. Co-expression of α1A- and α2A-AR in the same neuron was observed in normal DRG and increased post CCI. Collectively, these results indicated that there is distinct distribution of α-AR subtypes in DRG neurons, and the distribution and levels of expression of α-AR subtypes change differently after CCI. The up-regulation of α-AR subtypes in DRG neurons may play an important role in the process of generating and transmitting neuropathic pain.
Animals ; Cell Size ; Chronic Disease ; Constriction, Pathologic ; Fluorescent Antibody Technique ; Ganglia, Spinal ; metabolism ; pathology ; Male ; Microscopy, Confocal ; Neurons ; metabolism ; pathology ; Pain Measurement ; methods ; Pain Threshold ; Protein Isoforms ; metabolism ; Rats ; Rats, Sprague-Dawley ; Receptors, Adrenergic, alpha-1 ; metabolism ; Receptors, Adrenergic, alpha-2 ; metabolism ; Sciatic Nerve ; injuries ; surgery

OBJECTIVEInjection of insulin-like growth factor-1 (IGF-1) can prevent bone loss in sciatic nerve transaction rats. We try to investigate the action mechanism of IGF-1 on bone formation.

METHODSA total of 40 adult male Spragne-Dawley rats were divided into two groups (experimental group and control group) with 20 animals in each. Sciatic neurectomy was performed to model disuse osteoporosis in all rats. IGF-1 was administered in experimental group with the dose of 100 microgramme/kilogram per day for 3 days. Meanwhile, the rats in control group were treated with saline. Bone mineral density was measured by dual-energy X-ray absorptiometry 4 and 6 weeks after neurectomy respectively. Expression of Osterix and Runx2 was determined by reverse transcription-polymerase chain reaction (RT-PCR) assay.

RESULTSThere was a significant increase in the bone mineral density of experimental group compared with control group. There was a significant decrease in the level of receptor activator of nuclear factor-kappaB-ligand but an increase in the level of osteoprotegerin 4 and 6 weeks after neurectomy in the experimental group compared with control one. The expression of Osterix and Runx2 was up-regulated in the bone marrow of experimental group compared with control group.

CONCLUSIONIGF-1 can increase bone formation by stimulation of osteoblast number and activity, and reduce bone resorption by restriction of differentiation of osteoclast, suggesting that IGF-1 may improve the therapeutic efficacy for disuse osteoporosis.

Animals ; Bone Density ; drug effects ; Bone Resorption ; prevention & control ; Cell Differentiation ; Core Binding Factor Alpha 1 Subunit ; metabolism ; Immunohistochemistry ; Injections ; Insulin-Like Growth Factor I ; administration & dosage ; Male ; Osteoblasts ; drug effects ; Rats ; Rats, Sprague-Dawley ; Sciatic Nerve ; surgery ; Transcription Factors ; metabolism ; Up-Regulation ; physiology

OBJECTIVETo evaluate the local effect of vascular endothelial growth factor (VEGF) on transected sciatic nerve regeneration.

METHODSSixty male white Wistar rats were divided into four experimental groups randomly (n equal to 15). In transected group the left sciatic nerve was transected and the stump was fixed to adjacent muscle. In treatment group the defect was bridged using a silicone graft filled with 10 microlitre VEGF. In silicone group the graft was filled with phosphate-buffered saline. In sham-operated group the sciatic nerve was exposed and manipulated. Each group was subdivided into three subgroups with five animals in each and nerve fibers were studied 4, 8 and 12 weeks after operation.

RESULTSBehavioral test, functional study of sciatic nerve, gastrocnemius muscle mass and morphometric indices confirmed a faster recovery of regenerated axons in VEGF group than in silicone group (P less than 0.05). In immunohistochemical assessment, reactions to S-100 in VEGF group were more positive than that in silicone group.

CONCLUSIONLocal administration of VEGF will improve functional recovery and morphometric indices of sciatic nerve.

Administration, Topical ; Animals ; Nerve Regeneration ; drug effects ; Rats ; Rats, Wistar ; Sciatic Nerve ; surgery ; Vascular Endothelial Growth Factor A

OBJECTIVETo assess local effect of celecoxib on nerve regeneration in a rat sciatic nerve transection model.

METHODSForty-five male healthy white Wistar rats were randomly divided into three experimental groups (n equal to 15 for each): sham-operation (SHAM), control (SIL) and celecoxib treated (SIL/CLX) groups. In SHAM group after anesthesia left sciatic nerve was exposed and after homeostasis muscle was sutured. In SIL group the left sciatic nerve was exposed in the same way and transected proximal to tibioperoneal bifurcation leaving a 10 mm gap. Proximal and distal stumps were each inserted into a silicone tube and filled with 10 microlitre phosphate buffered solution. In SIL/CLX group defect was bridged using a silicone tube filled with 10 microlitre celecoxib (0.1 g/L).

RESULTSFunctional study and gastrocnemius muscle mass confirmed faster and better recovery of regenerated axons in SIL/CLX than in SIL group (P less than 0.05). Morphometric indices of regenerated fibers showed number and diameter of the myelinated fibers in SIL/CLX were significantly greater than those in control group. In immunohistochemistry, location of reactions to S-100 in SIL/CLX was clearly more positive than that in SIL group.

CONCLUSIONResponse to local treatment of celecoxib demonstrates that it influences and improves functional recovery of peripheral nerve regeneration.

Animals ; Celecoxib ; Cyclooxygenase 2 Inhibitors ; pharmacology ; Male ; Nerve Regeneration ; drug effects ; Peripheral Nerves ; drug effects ; surgery ; Pyrazoles ; pharmacokinetics ; Random Allocation ; Rats ; Rats, Wistar ; Sciatic Nerve ; drug effects ; Silicones ; Sulfonamides ; pharmacokinetics

OBJECTIVEPresent study aimed at further comprehensive functional, histomorphometrical and immunohistochemical assessment of peripheral nerve regeneration using rat sciatic nerve transection model.

METHODSThe 10-mm rat sciatic nerve gap was created in rats. In control group nerve stumps were sutured to adjacent muscle and in treatment group the gap was bridged using an inside-out vein graft. In sham-operated group the nerve was manipulated and left intact. All animals underwent walking track analysis test 4, 8, and 12 weeks after surgery. Subsequently, muscle mass measurement was performed to assess reenervation, histological examination to observe the sciatic nerve regeneration morphologically and immunohistochemistry to detect Schwann cells using anti S-100. Results were analyzed using a factorial ANOVA with two between-subjects factors. Bonferroni test for pairwise comparisons was used to examine the effect of treatments.

RESULTSFunctional analysis of myelinated nerve fibers showed that nerve function improved significantly in the time course in treatment group. However, quantitative morphometrical analysis of myelinated nerve fibers showed that there was no significant difference between 8 and 12 weeks in treatment group. Muscle weight ratio was bigger and weight loss of the gastrocnemius muscle was ameliorated by inside-out vein grafting. The position of positive immunohistochemical reactions further implied that regenerated axons and Schwann cell-like cells existed after vein grafting was performed, and was accompanied by the process of myelination and structural recovery of regenerated nerves.

CONCLUSIONFunctional analysis of peripheral nerve repair is far more reliable than quantitative morphometrical analysis.

Animals ; Immunohistochemistry ; Male ; Nerve Regeneration ; Rats ; Recovery of Function ; S100 Proteins ; analysis ; Sciatic Nerve ; physiology ; surgery ; Veins ; transplantation

BACKGROUND: Scar 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. METHODS: The 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. RESULTS: Topical 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. CONCLUSIONS: Topical 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.
Animals ; Carboxymethylcellulose Sodium/therapeutic use ; Cicatrix/*prevention & control ; Drug Combinations ; Hyaluronic Acid/*therapeutic use ; *Membranes, Artificial ; Postoperative Complications/*prevention & control ; Rats ; Rats, Sprague-Dawley ; Sciatic Nerve/*surgery ; Solutions