Platelet-rich plasma promotes peripheral nerve regeneration after sciatic nerve injury

The effect of platelet-rich plasma on nerve regeneration remains controversial.In this study,we established a rabbit model of sciatic nerve small-gap defects with preserved epineurium and then filled the gaps with platelet-rich plasma.Twenty-eight rabbits were divided into the following groups(7 rabbits/group):model,low-concentrati on PRP(2.5-3.5-fold concentration of whole blood platelets),medium-concentration PRP(4.5-6.5-fold concentration of whole blood platelets),and high-concentration PRP(7.5-8.5-fold concentration of whole blood platelets).Electrophysiological and histomorphometrical assessments and proteomics analysis we re used to evaluate regeneration of the sciatic nerve.Our results showed that platelet-rich plasma containing 4.5-6.5-and 7.5-8.5-fold concentrations of whole blood platelets promoted repair of sciatic nerve injury.Proteomics analysis was performed to investigate the possible mechanism by which platelet-rich plasma promoted nerve regeneration.Proteomics analysis showed that after sciatic nerve injury,platelet-rich plasma increased the expression of integrin subunitβ-8(ITGB8),which participates in angiogenesis,and differentially expressed proteins were mainly enriched in focal adhesion pathways.Additionally,two key proteins,ribosomal protein S27 a(RSP27 a)and ubiquilin 1(UBQLN1),which were selected after protein-protein interaction analysis,are involved in the regulation of ubiquitin levels in vivo.These data suggest that platelet-rich plasma promotes peripheral nerve regeneration after sciatic nerve injury by affecting angiogenesis and intracellular ubiquitin levels.

Label-free quantitative proteomics analysis models in vivo and in vitro reveal key proteins and potential roles in sciatic nerve injury

Background:The underlying mechanism of sciatic nerve injury(SNI)is a common motor functional disorder,necessitates further research.Methods:A rat model of SNI was established,with the injury group subjected to compressive injury of the right sciatic nerve exposed at the midpoint of the thigh and the sham surgery group undergoing the same surgical procedure.An oxygen-glucose deprivation model was employed to simulate in vitro SNI in PC12 cells.Following data acquisition and quality control,differentially expressed proteins(DEPs)in each model were identified through differential analysis,and enrichment analysis was used to explore the potential functions and pathways of the DEPs.Venn diagrams were drawn,and DEPs from both in vivo and in vitro SNI models were imported into the STRING database to construct a protein-protein interaction network and screen for hub proteins.Results:After the peptide segments obtained from rat nerve blockade and PC12 cells met quality requirements,258 DEPs were identified in rat nerve samples,and 119 DEPs were screened in PC12 cells.Enrichment analysis revealed that DEPs in the rat model were predominantly concentrated in biological functions such as myogenic cell proliferation and signaling related to lipid and energy metabolism.DEPs in the in vitro model were mainly enriched in biological processes such as phagocytosis and were associated with lipid transport and metabolism.Two hub proteins,amyloid precursor protein(APP)and fibronectin 1(FN1),were identified through MCC,MCODE,and Degree scoring.Both PC12 cells and external validation sets showed relatively higher expression of APP and FN1 in injured samples.Results of gene set enrichment analysis indicated that these two proteins were associated with metabolic pathways,such as biosynthesis of glycosaminoglycan chondroitin sulfate and biosynthesis of unsaturated fatty acids.Conclusion:APP and FN1 are potential key molecules involved in SNI and are associated with various metabolic pathways in nerve repair.These findings provide a theoretical basis for the development of therapeutic targets for SNI.

Sciatic nerve injury repair: a visualized analysis of research fronts and development trends

A total of 3,446 publications regarding sciatic nerve injury repair and protection indexed by Web of Science during 2000-2004 were used for a detailed analysis of temporal-spatial distribu- tion characteristics. Reference co-citation networks of the 100 top-cited publications as per the number of total citations were created using the Web of Science database and the information visualization tool, CiteSpaceIIL The key words that showed high frequency in these publications were included for analyzing the research fronts and development trends for sciatic nerve injury repair and protection. Through word frequency trend analysis, studies on bone marrow mesen- chymal stem cells, adipose-derived stem cells, and skeletal muscle-derived multipotent stem cells combined with tissue-engineered scaffold material will become the forefronts in the field of sci- atic nerve injury repair and protection in the near future.

The effects of claudin 14 during early Wallerian degeneration after sciatic nerve injury

Claudin 14 has been shown to promote nerve repair and regeneration in the early stages of Wallerian degeneration （0-4 days） in rats with sciatic nerve injury, but the mechanism underlying this process remains poorly understood. This study reported the effects of claudin 14 on nerve degeneration and regeneration during early Wallerian degeneration. Claudin 14 expression was up-regulated in sciatic nerve 4 days after Wallerian degeneration. The altered expression of claudin 14 in Schwann cells resulted in expression changes of cytokines in vitro. Expression of claudin 14 affected c-Jun, but not Akt anal ERK1/2 patl^ways, l^urther studies reve^ed that enhanced expression of claudin 14 could promote Schwann cell proliferation and migration. Silencing of claudin 14 expression resulted in Schwann cell apoptosis and reduction in Schwann cell proliferation. Our data revealed the role of claudin 14 in early Wallerian degeneration, which may provide new insights into the molecular mechanisms of Wallerian degeneration.

Strain and stress variations in the human amniotic membrane and fresh corpse autologous sciatic nerve anastomosis in a model of sciatic nerve injury

A 10-mm long sciatic nerve injury model was established in fresh normal Chinese patient cadavers. Amniotic membrane was harvested from healthy maternal placentas and was prepared into multilayered,coiled,tubular specimens.Sciatic nerve injury models were respectively anastomosed using the autologous cadaveric sciatic nerve and human amniotic membrane.Tensile test results showed that maximal loading,maximal displacement,maximal stress,and maximal strain of sciatic nerve injury models anastomosed with human amniotic membrane were greater than those in the autologous nerve anastomosis group.The strain-stress curves of the human amniotic membrane and sciatic nerves indicated exponential change at the first phase,which became elastic deformation curves at the second and third phases,and displayed plastic deformation curves at the fourth phase,at which point the specimens lost their bearing capacity.Experimental findings suggested that human amniotic membranes and autologous sciatic nerves exhibit similar stress-strain curves, good elastic properties,and certain strain and stress capabilities in anastomosis of the injured sciatic nerve.

Differential gene expression in proximal and distal nerve segments of rats with sciatic nerve injury during Wallerian degeneration

Wallerian degeneration is a subject of major interest in neuroscience. A large number of genes are differentially regulated during the distinct stages of Wallerian degeneration： transcription factor activation, immune response, myelin cell differentiation and dedifferentiation. Although gene expression responses in the distal segment of the sciatic nerve after peripheral nerve injury are known, differences in gene expression between the proximal and distal segments remain unclear. In the present study in rats, we used microarrays to analyze changes in gene expression, biological processes and signaling pathways in the proximal and distal segments of sciatic nerves under- going Wallerian degeneration. More than 6,000 genes were differentially expressed and 20 types of expression tendencies were identified, mainly between proximal and distal segments at 7-14 days after injury. The differentially expressed genes were those involved in cell differentiation, cytokinesis, neuron differentiation, nerve development and axon regeneration. Furthermore, 11 biological processes were represented, related to responses to stimuli, cell apoptosis, inflammato- ry response, immune response, signal transduction, protein kinase activity, and cell proliferation. Using real-time quantitative PCR, western blot analysis and immunohistochemistry, microarray data were verified for four genes： aquaporin-4, interleukin 1 receptor-like 1, matrix metallopro- teinase-12 and periaxin. Our study identifies differential gene expression in the proximal and distal segments of a nerve during Wallerian degeneration, analyzes dynamic biological changes of these genes, and provides a useful platform for the detailed study of nerve injury and repair during Wallerian degeneration.

Differential expression of microRNAs in dorsal root ganglia after sciatic nerve injury

This study investigated the possible involvement of microRNAs in the regulation of genes that participate in peripheral neural regeneration. A microRNA microarray analysis was conducted and 23 microRNAs were identiifed whose expression was signiifcantly changed in rat dorsal root ganglia after sciatic nerve transection. The expression of one of the downregulated microRNAs, microRNA-214, was validated using quantitative reverse transcriptase-PCR. MicroRNA-214 was predicted to target the 3′-untranslated region of Slit-Robo GTPase-activating protein 3. In situ hybridization veriifed that microRNA-214 was located in the cytoplasm of dorsal root ganglia primary neurons and was downregulated following sciatic nerve transection. Moreover, a com-bination of in situ hybridization and immunohistochemistry revealed that microRNA-214 and Slit-Robo GTPase-activating protein 3 were co-localized in dorsal root ganglion primary neu-rons. Western blot analysis suggested that Slit-Robo GTPase-activating protein 3 was upregulated in dorsal root ganglion neurons after sciatic nerve transection. These data demonstrate that mi-croRNA-214 is located and differentially expressed in dorsal root ganglion primary neurons and may participate in regulating the gene expression of Slit-Robo GTPase-activating protein 3 after sciatic nerve transection.

Stress and strain analysis on the anastomosis site sutured with either epineurial or perineurial sutures after simulation of sciatic nerve injury

The magnitude of tensile stress and tensile strain at an anastomosis site under physiological stress is an important factor for the success of anastomosis following suturing in peripheral nerve injury treatment. Sciatic nerves from fresh adult cadavers were used to create models of sciatic nerve injury. The denervated specimens underwent epineurial and perineurial suturing. The elastic modulus （40.96 ＋ 2.59 MPa） and Poisson ratio （0.37 ＋ 0.02） of the normal sciatic nerve were measured by strain electrical measurement. A resistance strain gauge was pasted on the front, back left, and right of the edge of the anastomosis site after suturing. Strain electrical measurement results showed that the stress and strain values of the sciatic nerve following perineurial suturing were lower than those following epineurial suturing. Scanning electron microscopy revealed that the sciatic nerve fibers were disordered following epineurial compared with perineurial suturing. These results indicate that the effect of perineurial suturing in sciatic nerve injury repair is better than that of epineurial suturing.

Autologous nerve anastomosis versus human amniotic membrane anastomosisA rheological comparison following simulated sciatic nerve injury

The sciatic nerve is biological viscoelastic solid,with stress relaxation and creep characteristics.In this study,a comparative analysis of the stress relaxation and creep characteristics of the sciatic nerve was conducted after simulating sciatic nerve injury and anastomosing with autologous nerve or human amniotic membrane.The results demonstrate that,at the 7 200-second time point,both stress reduction and strain increase in the human amniotic membrane anastomosis group were significantly greater than in the autologous nerve anastomosis group.Our findings indicate that human amniotic membrane anastomosis for sciatic nerve injury has excellent rheological characteristics and is conducive to regeneration of the injured nerve.

Saikosaponin a increases interleukin-10 expression and inhibits scar formation after sciatic nerve injury

Nerve scarring after peripheral nerve injury can severely hamper nerve regeneration and functional recovery.Further,the anti-inflammatory cytokine,interleukin-10,can inhibit nerve scar formation.Saikosaponin a（SSa） is a monomer molecule extracted from the Chinese medicine,Bupleurum.SSa can exert anti-inflammatory effects in spinal cord injury and traumatic brain injury.However,it has not been shown whether SSa can play a role in peripheral nerve injury.In this study,rats were randomly assigned to three groups.In the sham group,the left sciatic nerve was directly sutured after exposure.In the sciatic nerve injury（SNI） ＋ SSa and SNI groups,the left sciatic nerve was sutured and continuously injected daily with SSa（10 mg/kg） or an equivalent volume of saline for 7 days.Enzyme linked immunosorbent assay results demonstrated that at 7 days after injury,interleukin-10 level was considerably higher in the SNI ＋ SSa group than in the SNI group.Masson staining and western blot assay demonstrated that at 8 weeks after injury,type I and III collagen content was lower and nerve scar formation was visibly less in the SNI ＋ SSa group compared with the SNI group.Simultaneously,sciatic functional index and nerve conduction velocity were improved in the SNI ＋ SSa group compared with the SNI group.These results confirm that SSa can increase the expression of the anti-inflammatory factor,interleukin-10,and reduce nerve scar formation to promote functional recovery of injured sciatic nerve.

Inhibitor of DNA binding 2 accelerates nerve regeneration after sciatic nerve injury in mice

Inhibitor of DNA binding 2(Id2)can promote axonal regeneration after injury of the central nervous system.However,whether Id2 can promote axonal regeneration and functional recovery after peripheral nerve injury is currently unknown.In this study,we established a mouse model of bilateral sciatic nerve crush injury.Two weeks before injury,AAV9-Id2-3×Flag-GFP was injected stereotaxically into the bilateral ventral horn of lumbar spinal cord.Our results showed that Id2 was successfully delivered into spinal cord motor neurons projecting to the sciatic nerve,and the number of regenerated motor axons in the sciatic nerve distal to the crush site was increased at 2 weeks after injury,arriving at the tibial nerve and reinnervating a few endplates in the gastrocnemius muscle.By 1 month after injury,extensive neuromuscular reinnervation occurred.In addition,the amplitude of compound muscle action potentials of the gastrocnemius muscle was markedly recovered,and their latency was shortened.These findings suggest that Id2 can accelerate axonal regeneration,promote neuromuscular reinnervation,and enhance functional improvement following sciatic nerve injury.Therefore,elevating the level of Id2 in adult neurons may present a promising strategy for peripheral nerve repair following injury.The study was approved by the Experimental Animal Ethics Committee of Jinan University(approval No.20160302003)on March 2,2016.

“Three Methods and Three Points” regulates p38 mitogen-activated protein kinase in the dorsal horn of the spinal cord in a rat model of sciatic nerve injury

Tuina is a traditional Chinese treatment for sensory disturbances caused by peripheral nerve injury and related diseases. Our previous studies showed that tuina regulates relevant regions and indices of the spinal dorsal horn using the Dian, Bo, and Rou method in Yinmen（BL37）, Yanglingquan（GB34）, and Weizhong（BL40）. Treatment prevents muscle atrophy, protects spinal cord neurons, and promotes sciatic nerve repair. The mechanisms of action of tuina for treating peripheral nerve injury remain poorly understood. This study established rat models of sciatic nerve injury using the crushing method. Rats received Chinese tuina in accordance with the principle of ＂Three Methods and Three Points,＂ once daily for 20 days. Tuina intervention reduced paw withdrawal latency and improved wet weight of the gastrocnemius muscle, as well as promoting morphological recovery of sciatic nerve fibers, Schwann cells, and axons. The protein expression levels of phospho-p38 mitogen-activated protein kinase, tumor necrosis factor-α, and interleukin-1β also decreased. These findings indicate that ＂Three Methods and Three Points＂ promoted morphological recovery and improved behavior of rats with peripheral nerve injury.

Bacterial melanin promotes recovery after sciatic nerve injury in rats

Bacterial melanin, obtained from the mutant strain of Bacillus Thuringiensis, has been shown to promote recovery after central nervous system injury. It is hypothesized, in this study, that bacterial melanin can promote structural and functional recovery after peripheral nerve injury. Rats subjected to sciatic nerve transection were intramuscularly administered bacterial melanin. The sciatic nerve transected rats that did not receive intramuscular administration of bacterial melanin served as controls. Behavior tests showed that compared to control rats, the time taken for instrumental conditioned reflex recovery was significantly shorter and the ability to keep the balance on the rotating bar was significantly better in bacterial melanin-treated rats. Histomor- phological tests showed that bacterial melanin promoted axon regeneration after sciatic nerve injury. These findings suggest that bacterial melanin exhibits neuroprotective effects on injured sciatic nerve, contributes to limb motor function recovery, and therefore can be used for rehabil- itation treatment of peripheral nerve injury.

Baculoviral inhibitor of apoptosis protein repeatcontaining protein 3 delays early Wallerian degeneration after sciatic nerve injury

Wallerian degeneration is a complex biological process that occurs after nerve injury,and involves nerve degeneration and regeneration.Schwann cells play a crucial role in the cellular and molecular events of Wallerian degeneration of the peripheral nervous system.However,Wallerian degeneration regulating nerve injury and repair remains largely unknown,especially the early response.We have previously reported some key regulators of Wallerian degeneration after sciatic nerve injury.Baculoviral inhibitor of apoptosis protein repeat-containing protein 3(BIRC3)is an important factor that regulates apoptosis-inhibiting protein.In this study,we established rat models of right sciatic nerve injury.In vitro Schwann cell models were also established and subjected to gene transfection to inhibit and overexpress BIRC3.The data indicated that BIRC3 expression was significantly up-regulated after sciatic nerve injury.Both BIRC3 upregulation and downregulation affected the migration,proliferation and apoptosis of Schwan cells and affected the expression of related factors through activating c-fos and ERK signal pathway.Inhibition of BIRC3 delayed early Wallerian degeneration through inhibiting the apoptosis of Schwann cells after sciatic nerve injury.These findings suggest that BIRC3 plays an important role in peripheral nerve injury repair and regeneration.The study was approved by the Institutional Animal Care and Use Committee of Nantong University,China(approval No.2019-nsfc004)on March 1,2019.

The longitudinal epineural incision and complete nerve transection method for modeling sciatic nerve injury

Injury severity, operative technique and nerve regeneration are important factors to consider when constructing a model of peripheral nerve injury. Here, we present a novel peripheral nerve injury model and compare it with the complete sciatic nerve transection method. In the experimental group, under a microscope, a 3-mm longitudinal incision was made in the epineurium of the sciatic nerve to reveal the nerve fibers, which were then transected. The small, longitudinal incision in the epineurium was then sutured closed, requiring no stump anastomosis. In the control group, the sciatic nerve was completely transected, and the epineurium was repaired by anastomosis. At 2 and 4 weeks after surgery, Wallerian degeneration was observed in both groups. In the experimental group, at 8 and 12 weeks after surgery, distinct medullary nerve fibers and axons were observed in the injured sciatic nerve. Regular, dense myelin sheaths were visible, as well as some scarring. By 12 weeks, the myelin sheaths were normal and intact, and a tight lamellar structure was observed. Functionally, limb movement and nerve conduction recovered in the injured region between 4 and 12 weeks. The present results demonstrate that longitudinal epineural incision with nerve transection can stably replicate a model of Sunderland grade IV peripheral nerve injury. Compared with the complete sciatic nerve transection model, our method reduced the difficulties of micromanipulation and surgery time, and resulted in good stump restoration, nerve regeneration, and functional recovery.

Guinea pigs as an animal model for sciatic nerve injury

The overwhelming use of rat models in nerve regeneration studies is likely to induce skewness in treatment outcomes.To address the problem,this study was conducted in 8 adult guinea pigs of either sex to investigate the suitability of guinea pig as an alternative model for nerve regeneration studies.A crush injury was inflicted to the sciatic nerve of the left limb,which led to significant decrease in the pain perception and neurorecovery up to the 4th weak.Lengthening of foot print and shortening of toe spread were observed in the paw after nerve injury.A 3.49 ± 0.35 fold increase in expression of neuropilin 1（NRP1） gene and 2.09 ± 0.51 fold increase in neuropilin 2（NRP2） gene were recorded 1 week after nerve injury as compared to the normal nerve.Ratios of gastrocnemius muscle weight and volume of the experimental limb to control limb showed more than 50% decrease on the 30 th day.Histopathologically,vacuolated appearance of the nerve was observed with presence of degenerated myelin debris in digestion chambers.Gastrocnemius muscle also showed degenerative changes.Scanning electron microscopy revealed loose and rough arrangement of connective tissue fibrils and presence of large spherical globules in crushed sciatic nerve.The findings suggest that guinea pigs could be used as an alternative animal model for nerve regeneration studies and might be preferred over rats due to their cooperative nature while recording different parameters.

Stem cell transplantation for repair of sciatic nerve injury

Three articles regarding transplantation of umbilical cord mesenchmal stem cells alone or in combination with Schwann cells and feridex and polylysine complex-labeled bone marrow stromal cell transplantation （MRI tracing） for repair of sciatic nerve injury were reported in Neural Regeneration

Tissue-engineered nerve for repair of sciatic nerve injury

Three articles regarding the use of nerve fragments bridging regeneration chambers, three-dimensional bionic nerve conduits and multiwalled carbon nanotubes for repair of sciatic nerve injury were reported in Neural Regeneration Research. We hope that our readers find these papers useful to their research.

Protective effect of sodium valproate on motor neurons in the spinal cord following sciatic nerve injury in rats

BACKGROUND: Sodium valproate (VPA) is used to be an effective anti-epileptic drug. VPA possesses the characteristics of penetrating rapidly through the blood-brain barrier (BBB) and increasing levels of Bcl-2 and growth cone-associated protein (GAP) 43 in spinal cord. OBJECTIVE: To observe the effect of VPA on Bcl-2 expression and motor neuronal apoptosis in spinal cord of rats following sciatic nerve transection. DESIGN: Randomized controlled experiment. SETTING: Department of Hand Surgery and Microsurgery, Wuhan Puai Hospital. MATERIALS: A total of 30 male healthy SD rats of clean grade and with the body mass of 180-220 g were provided by Experimental Animal Center of Medical College of Wuhan University. Sodium Valproate Tablets were purchases from Hengrui Pharmaceutical Factory, Jiangsu. METHODS: The experiment was performed in the Central Laboratory of Wuhan Puai Hospital and Medical College of Wuhan University from February to May 2006. Totally 30 rats were randomly divided into two groups: treatment group (n =15) and model group (n =15). Longitudinal incision along backside of right hind limbs of rats was made to expose sciatic nerves, which were sharply transected 1 cm distal to the inferior margin of piriform muscle after nerve liberation under operation microscope to establish sciatic nerve injury rat models. Sodium Valproate Tablets were pulverized and diluted into 50 g/L suspension with saline. On the day of operation, the rats in the treatment group received 6 mL/kg VPA suspension by gastric perfusion, once a day, whereas model group received 10 mL/kg saline by gastric perfusion, once a day. L4-6 spinal cords were obtained at days 1, 4, 7, 14 and 28 after operation, respectively. Terminal deoxyribonucleotidyl transferase (TdT)-mediated dUTP-biotin nick end labeling (TUNEL) technique and immunohistochemical method (SP method) were used to detect absorbance (A) of neurons with positive Bcl-2 expression. Apoptotic rate of cells (number of apoptotic cells/total number of cells×100%) was calculated. MAIN OUTCOME MEASURES: A value of neurons with positive Bcl-2 expression and apoptotic rate in spinal cord of rats in the two groups. RESULTS: A total of 30 SD rats were involved in the result analysis. ①expression of positive Bcl-2 neurons: A value of positive Bcl-2 neurons were 0.71±0.02, 0.86±0.04, 1.02±0.06 at days 4, 7 and 14, respectively after operation in the treatment group, which were obviously higher than those in the model group (0.62±0.03, 0.71±0.05, 0.89±0.04, t = 3.10-4.50, P < 0.05). ②apoptotic result of motor neurons: Apoptotic rate of motor neurons in spinal cord was (6.91±0.89)% and (15.12±2.34)% at days 7 and 14 in the treatment group, which was significantly lower than those in the model group [(9.45±1.61)%, (19.35±0.92)%, t = 2.39, 3.03. P < 0.05]. CONCLUSION: VPA can increase expression of Bcl-2 in spinal cord and reduce neuronal apoptosis in rats following sciatic nerve injury, and has protective effect on motor neuron in spinal cord of rats.

Microencapsulation improves inhibitory effects of transplanted olfactory ensheathing cells on pain after sciatic nerve injury

Olfactory bulb tissue transplantation inhibits P2X2/3 receptor-mediated neuropathic pain. However, the olfactory bulb has a complex cellular composition, and the mechanism underlying the action of purified transplanted olfactory ensheathing cells（OECs） remains unclear. In the present study, we microencapsulated OECs in alginic acid, and transplanted free and microencapsulated OECs into the region surrounding the injured sciatic nerve in rat models of chronic constriction injury. We assessed mechanical nociception in the rat models 7 and 14 days after surgery by measuring paw withdrawal threshold, and examined P2X2/3 receptor expression in L4–5 dorsal root ganglia using immunohistochemistry. Rats that received free and microencapsulated OEC transplants showed greater withdrawal thresholds than untreated model rats, and weaker P2X2/3 receptor immunoreactivity in dorsal root ganglia. At 14 days, paw withdrawal threshold was much higher in the microencapsulated OEC-treated animals. Our results confirm that microencapsulated OEC transplantation suppresses P2X2/3 receptor expression in L4–5 dorsal root ganglia in rat models of neuropathic pain and reduces allodynia, and also suggest that transplantation of microencapsulated OECs is more effective than transplantation of free OECs for the treatment of neuropathic pain.