Biological conduit small gap sleeve bridging method for peripheral nerve injury: regeneration law of nerve fibers in the conduit

The clinical effects of 2-mm small gap sleeve bridging of the biological conduit to repair peripheral nerve injury are better than in the traditional epineurium suture, so it is possible to replace the epineurium suture in the treatment of peripheral nerve injury. This study sought to identify the regeneration law of nerve fibers in the biological conduit. A nerve regeneration chamber was constructed in models of sciatic nerve injury using 2-mm small gap sleeve bridging of a biodegradable biological conduit. The results showed that the biological conduit had good histocompatibility. Tissue and cell apoptosis in the conduit apparently lessened, and regenerating nerve fibers were common. The degeneration regeneration law of Schwann cells and axons in the conduit was quite different from that in traditional epineurium suture. During the prime period for nerve fiber regeneration(2–8 weeks), the number of Schwann cells and nerve fibers was higher in both proximal and distal ends, and the effects of the small gap sleeve bridging method were better than those of the traditional epineurium suture. The above results provide an objective and reliable theoretical basis for the clinical application of the biological conduit small gap sleeve bridging method to repair 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. Histomorphological 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 rehabilitation treatment of peripheral nerve injury.     

Electrical stimulation of the vagus nerve protects against cerebral ischemic injury through an anti-infammatory mechanism

Vagus nerve stimulation exerts protective effects against ischemic brain injury; however, the underlying mechanisms remain unclear. In this study, a rat model of focal cerebral ischemia was established using the occlusion method, and the right vagus nerve was given electrical stimulation (constant current of 0.5 mA; pulse width, 0.5 ms; frequency, 20 Hz; duration, 30 seconds; every 5 minutes for a total of 60 minutes) 30 minutes, 12 hours, and 1, 2, 3, 7 and 14 days after surgery. Electrical stimulation of the vagus nerve substantially reduced infarct volume, improved neurological function, and decreased the expression levels of tumor necrosis factor-α and interleukin-6 in rats with focal cerebral ischemia. The experimental findings indicate that the neuroprotective effect of vagus nerve stimulation following cerebral ischemia may be associated with the inhibition of tumor necrosis factor-α and interleukin-6 expression.     

Blockade of transient receptor potential cation channel subfamily V member 1 promotes regeneration after sciatic nerve injury

The transient receptor potential cation channel subfamily V member 1(TRPV1) provides the sensation of pain(nociception). However, it remains unknown whether TRPV1 is activated after peripheral nerve injury, or whether activation of TRPV1 affects neural regeneration. In the present study, we established rat models of unilateral sciatic nerve crush injury, with or without pretreatment with AMG517(300 mg/kg), a TRPV1 antagonist, injected subcutaneously into the ipsilateral paw 60 minutes before injury. At 1 and 2 weeks after injury, we performed immunofluorescence staining of the sciatic nerve at the center of injury, at 0.3 cm proximal and distal to the injury site, and in the dorsal root ganglia. Our results showed that Wallerian degeneration occurred distal to the injury site, and neurite outgrowth and Schwann cell regeneration occurred proximal to the injury. The number of regenerating myelinated and unmyelinated nerve clusters was greater in the AMG517-pretreated rats than in the vehicle-treated group, most notably 2 weeks after injury. TRPV1 expression in the injured sciatic nerve and ipsilateral dorsal root ganglia was markedly greater than on the contralateral side. Pretreatment with AMG517 blocked this effect. These data indicate that TRPV1 is activated or overexpressed after sciatic nerve crush injury, and that blockade of TRPV1 may accelerate regeneration of the injured sciatic nerve.     

Neuroprotective effects of ultrasound-guided nerve growth factor injections after sciatic nerve injury

Nerve growth factor (NGF) plays an important role in promoting neuroregeneration after peripheral nerve injury. However, its effects are limited by its short half-life; it is therefore important to identify an effective mode of administration. High-frequency ultrasound (HFU) is increasingly used in the clinic for high-resolution visualization of tissues, and has been proposed as a method for identifying and evaluating peripheral nerve damage after injury. In addition, HFU is widely used for guiding needle placement when administering drugs to a specific site. We hypothesized that HFU guiding would optimize the neuroprotective effects of NGF on sciatic nerve injury in the rabbit. We performed behavioral, ultrasound, electrophysiological, histological, and immunohistochemical evaluation of HFU-guided NGF injections administered immediately after injury, or 14 days later, and compared this mode of administration with intramuscular NGF injections. Across all assessments, HFU-guided NGF injections gave consistently better outcomes than intramuscular NGF injections administered immediately or 14 days after injury, with immediate treatment also yielding better structural and functional results than when the treatment was delayed by 14 days. Our findings indicate that NGF should be administered as early as possible after peripheral nerve injury, and highlight the striking neuroprotective effects of HFU-guided NGF injections on peripheral nerve injury compared with intramuscular administration.     

Femoral nerve regeneration and its accuracy under different injury mechanisms

Surgical accuracy has greatly improved with the advent of microsurgical techniques. However, complete functional recovery after peripheral nerve injury has not been achieved to date. The mechanisms hindering accurate regeneration of damaged axons after peripheral nerve injury are in urgent need of exploration. The present study was designed to explore the mechanisms of peripheral nerve regeneration after different types of injury. Femoral nerves of rats were injured by crushing or freezing. At 2, 3, 6, and 12 weeks after injury, axons were retrogradely labeled using 1,1′-dioctadecyl-3,3,3′,3′-tetramethylindocarbocyanine perchlorate(Dil) and True Blue, and motor and sensory axons that had regenerated at the site of injury were counted. The number and percentage of Dil-labeled neurons in the anterior horn of the spinal cord increased over time. No significant differences were found in the number of labeled neurons between the freeze and crush injury groups at any time point. Our results confirmed that the accuracy of peripheral nerve regeneration increased with time, after both crush and freeze injury, and indicated that axonal regeneration accuracy was still satisfactory after freezing, despite the prolonged damage.     

Resuscitation therapy for traumatic brain injuryinduced coma in rats:mechanisms of median nerve electrical stimulation

In this study, rats were put into traumatic brain injury-induced coma and treated with median nerve electrical stimulation. We explored the wake-promoting effect, and possible mechanisms, of median nerve electrical stimulation. Electrical stimulation upregulated the expression levels of orexin-A and its receptor OX1 R in the rat prefrontal cortex. Orexin-A expression gradually increased with increasing stimulation, while OX1 R expression reached a peak at 12 hours and then decreased. In addition, after the OX1 R antagonist, SB334867, was injected into the brain of rats after traumatic brain injury, fewer rats were restored to consciousness, and orexin-A and OXIR expression in the prefrontal cortex was downregulated. Our findings indicate that median nerve electrical stimulation induced an up-regulation of orexin-A and OX1 R expression in the prefrontal cortex of traumatic brain injury-induced coma rats, which may be a potential mechanism involved in the wake-promoting effects of median nerve electrical stimulation.     

Polylactic-co-glycolic acid microspheres containing three neurotrophic factors promote sciatic nerve repair after injury

A variety of neurotrophic factors have been shown to repair the damaged peripheral nerve. However, in clinical practice, nerve growth factor, neurotrophin-3 and brain-derived neurotrophic factor are all peptides or proteins that may be rapidly deactivated at the focal injury site; their local effective concentration time following a single medication cannot meet the required time for spinal axons to regenerate and cross the glial scar. In this study, we produced polymer sustained-release microspheres based on the polylactic-co-glycolic acid copolymer; the microspheres at 300-μm diameter contained nerve growth factor, neurotrophin-3 and brain-derived neurotrophic factor. Six microspheres were longitudinally implanted into the sciatic nerve at the anastomosis site, serving as the experimental group; while the sciatic nerve in the control group was subjected to the end-to-end anastomosis using 10/0 suture thread. At 6 weeks after implantation, the lower limb activity, weight of triceps surae muscle, sciatic nerve conduction velocity and the maximum amplitude were obviously better in the experimental group than in the control group. Compared with the control group, more regenerating nerve fibers were observed and distributed in a dense and ordered manner with thicker myelin sheaths in the experimental group. More angiogenesis was also visible. Experimental findings indicate that polylactic-co-glycolic acid composite microspheres containing nerve growth factor, neurotrophin-3 and brain-derived neurotrophic factor can promote the restoration of sciatic nerve in rats after injury.     

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.     

Peripheral nerve regeneration with conduits:use of vein tubes

Treatment of peripheral nerve injuries remains a challenge to modern medicine due to the complexity of the neurobiological nerve regenerating process. There is a greater challenge when the transected nerve ends are not amenable to primary end-to-end tensionless neurorraphy. When facing a segmental nerve defect, great effort has been made to develop an alternative to the autologous nerve graft in order to circumvent morbidity at donor site, such as neuroma formation, scarring and permanent loss of function. Tubolization techniques have been developed to bridge nerve gaps and have been extensively studied in numerous experimental and clinical trials. The use of a conduit intends to act as a vehicle for moderation and modulation of the cellular and molecular ambience for nerve regeneration. Among several conduits, vein tubes were validated for clinical application with improving outcomes over the years. This article aims to address the investigation and treatment of segmental nerve injury and draw the current panorama on the use of vein tubes as an autogenous nerve conduit.     

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.     

Pulsed electrical stimulation protects neurons in the dorsal root and anterior horn of the spinal cord after peripheral nerve injury

Most studies on peripheral nerve injury have focused on repair at the site of injury, but very few have examined the effects of repair strategies on the more proximal neuronal cell bodies. In this study, an approximately 10-mm-long nerve segment from the ischial tuberosity in the rat was transected and its proximal and distal ends were inverted and sutured. The spinal cord was subjected to pulsed electrical stimulation at T10 and L3, at a current of 6.5 m A and a stimulation frequency of 15 Hz, 15 minutes per session, twice a day for 56 days. After pulsed electrical stimulation, the number of neurons in the dorsal root ganglion and anterior horn was increased in rats with sciatic nerve injury. The number of myelinated nerve fibers was increased in the sciatic nerve. The ultrastructure of neurons in the dorsal root ganglion and spinal cord was noticeably improved. Conduction velocity of the sciatic nerve was also increased. These results show that pulsed electrical stimulation protects sensory neurons in the dorsal root ganglia as well as motor neurons in the anterior horn of the spinal cord after peripheral nerve injury, and that it promotes the regeneration of peripheral nerve fibers.     

Changes in the blood-nerve barrier after sciatic nerve cold injury:indications supporting early treatment

Severe edema in the endoneurium can occur after non-freezing cold injury to the peripheral nerve, which suggests damage to the blood-nerve barrier. To determine the effects of cold injury on the blood-nerve barrier, the sciatic nerve on one side of Wistar rats was treated with low temperatures(3–5°C) for 2 hours. The contralateral sciatic nerve was used as a control. We assessed changes in the nerves using Evans blue as a fluid tracer and morphological methods. Excess fluid was found in the endoneurium 1 day after cold injury, though the tight junctions between cells remained closed. From 3 to 5 days after the cold injury, the fluid was still present, but the tight junctions were open. Less tracer leakage was found from 3 to 5 days after the cold injury compared with 1 day after injury. The cold injury resulted in a breakdown of the blood-nerve barrier function, which caused endoneurial edema. However, during the early period, the breakdown of the blood-nerve barrier did not include the opening of tight junctions, but was due to other factors. Excessive fluid volume produced a large increase in the endoneurial fluid pressure, prevented liquid penetration into the endoneurium from the microvasculature. These results suggest that drug treatment to patients with cold injuries should be administered during the early period after injury because it may be more difficult for the drug to reach the injury site through the microcirculation after the tissue fluid pressure becomes elevated.     

Allograft pretreatment for the repair of sciatic nerve defects: green tea polyphenols versus radiation

Pretreatment of nerve allografts by exposure to irradiation or green tea polyphenols can eliminate neuroimmunogenicity, inhibit early immunological rejection, encourage nerve regeneration and functional recovery, improve tissue preservation, and minimize postoperative infection. In the present study, we investigate which intervention achieves better results. We produced a 1.0 cm sciatic nerve defect in rats, and divided the rats into four treatment groups: autograft, fresh nerve allograft, green tea polyphenol-pretreated(1 mg/m L, 4°C) nerve allograft, and irradiation-pretreated nerve allograft(26.39 Gy/min for 12 hours; total 19 k Gy). The animals were observed, and sciatic nerve electrophysiology, histology, and transmission electron microscopy were carried out at 6 and 12 weeks after grafting. The circumference and structure of the transplanted nerve in rats that received autografts or green tea polyphenol-pretreated nerve allografts were similar to those of the host sciatic nerve. Compared with the groups that received fresh or irradiation-pretreated nerve allografts, motor nerve conduction velocity in the autograft and fresh nerve allograft groups was greater, more neurites grew into the allografts, Schwann cell proliferation was evident, and a large number of new blood vessels was observed; in addition, massive myelinated nerve fibers formed, and abundant microfilaments and microtubules were present in the axoplasm. Our findings indicate that nerve allografts pretreated by green tea polyphenols are equivalent to transplanting autologous nerves in the repair of sciatic nerve defects, and promote nerve regeneration. Pretreatment using green tea polyphenols is better than pretreatment with irradiation.     

Phrenic nerve transfer to the musculocutaneous nerve for the repair of brachial plexus injury: electrophysiological characteristics

Phrenic nerve transfer is a major dynamic treatment used to repair brachial plexus root avulsion. We analyzed 72 relevant articles on phrenic nerve transfer to repair injured brachial plexus that were indexed by Science Citation Index. The keywords searched were brachial plexus injury, phrenic nerve, repair, surgery, protection, nerve transfer, and nerve graft. In addition, we performed neurophysiological analysis of the preoperative condition and prognosis of 10 patients undergoing ipsilateral phrenic nerve transfer to the musculocutaneous nerve in our hospital from 2008 to 201 3 and observed the electromyograms of the biceps brachii and motor conduction function of the musculocutaneous nerve. Clinically, approximately 28% of patients had brachial plexus injury combined with phrenic nerve injury, and injured phrenic nerve cannot be used as a nerve graft. After phrenic nerve transfer to the musculocutaneous nerve, the regenerated potentials first appeared at 3 months. Recovery of motor unit action potential occurred 6 months later and became more apparent at 12 months. The percent of patients recovering ‘excellent' and ‘good' muscle strength in the biceps brachii was 80% after 18 months. At 12 months after surgery, motor nerve conduction potential appeared in the musculocutaneous nerve in seven cases. These data suggest that preoperative evaluation of phrenic nerve function may help identify the most appropriate nerve graft in patients with an injured brachial plexus. The functional recovery of a transplanted nerve can be dynamically observed after the surgery.     

Axon regeneration impediment: the role of paired immunoglobulin-like receptor B

Regenerative capacity is weak after central nervous system injury because of the absence of an enhancing microenvironment and presence of an inhibitory microenvironment for neuronal and axonal repair. In addition to the Nogo receptor(Ng R), the paired immunoglobulin-like receptor B(Pir B) is a recently discovered coreceptor of Nogo, myelin-associated glycoprotein, and myelin oligodendrocyte glycoprotein. Concurrent blocking of Ng R and Pir B almost completely eliminates the inhibitory effect of myelin-associated inhibitory molecules on axonal regeneration. Pir B participates in a key pathological process of the nervous system, specifically axonal regeneration inhibition. Pir B is an inhibitory receptor similar to Ng R, but their effects are not identical. This study summarizes the structure, distribution, relationship with common nervous system diseases, and known mechanisms of Pir B, and concludes that Pir B is also distributed in cells of the immune and hematopoietic systems. Further investigations are needed to determine if immunomodulation and blood cell migration involve inhibition of axonal regeneration.     

The role of exosomes in peripheral nerve regeneration

Peripheral nerve injuries remain problematic to treat, with poor functional recovery commonly observed. Injuries resulting in a nerve gap create specific difficulties for axonal regeneration. Approaches to address these difficulties include autologous nerve grafts (which are currently the gold standard treatment) and synthetic conduits, with the latter option being able to be impregnated with Schwann cells or stem cells which provide an appropriate micro-environment for neuronal regeneration to occur. Transplanting stem cells, however, infers additional risk of malignant transformation as well as manufacturing difficulties and ethical concerns, and the use of autologous nerve grafts and Schwann cells requires the sacrifice of a functioning nerve. A new approach utilizing exosomes, secreted extracellular vesicles, could avoid these complications. In this review, we summarize the current literature on exosomes, and suggest how they could help to improve axonal regeneration following peripheral nerve injury.     

Biological conduits combining bone marrow mesenchymal stem cells and extracellular matrix to treat long-segment sciatic nerve defects

The transplantation of polylactic glycolic acid conduits combining bone marrow mesenchymal stem cells and extracellular matrix gel for the repair of sciatic nerve injury is effective in some respects, but few data comparing the biomechanical factors related to the sciatic nerve are available. In the present study, rabbit models of 10-mm sciatic nerve defects were prepared. The rabbit models were repaired with autologous nerve, a polylactic glycolic acid conduit + bone marrow mesenchymal stem cells, or a polylactic glycolic acid conduit + bone marrow mesenchymal stem cells + extracellular matrix gel. After 24 weeks, mechanical testing was performed to determine the stress relaxation and creep parameters. Following sciatic nerve injury, the magnitudes of the stress decrease and strain increase at 7,200 seconds were largest in the polylactic glycolic acid conduit + bone marrow mesenchymal stem cells + extracellular matrix gel group, followed by the polylactic glycolic acid conduit + bone marrow mesenchymal stem cells group, and then the autologous nerve group. Hematoxylin-eosin staining demonstrated that compared with the polylactic glycolic acid conduit + bone marrow mesenchymal stem cells group and the autologous nerve group, a more complete sciatic nerve regeneration was found, including good myelination, regularly arranged nerve fibers, and a completely degraded and resorbed conduit, in the polylactic glycolic acid conduit + bone marrow mesenchymal stem cells + extracellular matrix gel group. These results indicate that bridging 10-mm sciatic nerve defects with a polylactic glycolic acid conduit + bone marrow mesenchymal stem cells + extracellular matrix gel construct increases the stress relaxation under a constant strain, reducing anastomotic tension. Large elongations under a constant physiological load can limit the anastomotic opening and shift, which is beneficial for the regeneration and functional reconstruction of sciatic nerve. Better regeneration was found with the polylactic glycolic acid conduit + bone marrow mesenchymal stem cells + extracellular matrix gel grafts than with the polylactic glycolic acid conduit + bone marrow mesenchymal stem cells grafts and the autologous nerve grafts.     

Lateral intracerebroventricular injection of Apelin-13 inhibits apoptosis after cerebral ischemia/reperfusion injury

Apelin-13 inhibits neuronal apoptosis caused by hydrogen peroxide, yet apoptosis following cerebral ischemia-reperfusion injury has rarely been studied. In this study, Apelin-13 (0.1 μg/g) was injected into the lateral ventricle of middle cerebral artery occlusion model rats. TTC, TUNEL, and immunohistochemical staining showed that compared with the cerebral ischemia/reperfusion group, infarct volume and apoptotic cell number at the ischemic penumbra region were decreased in the Apelin-13 treatment group. Additionally, Apelin-13 treatment increased Bcl-2 immunoreactivity and decreased caspase-3 immunoreactivity. Our findings suggest that Apelin-13 is neuroprotective against cerebral ischemia/reperfusion injury through inhibition of neuronal apoptosis.     

Electrospun and woven silk fibroin/poly(lactic-coglycolic acid) nerve guidance conduits for repairing peripheral nerve injury

We have designed a novel nerve guidance conduit(NGC) made from silk fibroin and poly(lactic-co-glycolic acid) through electrospinning and weaving(ESP-NGCs). Several physical and biological properties of the ESP-NGCs were assessed in order to evaluate their biocompatibility. The physical properties, including thickness, tensile stiffness, infrared spectroscopy, porosity, and water absorption were determined in vitro. To assess the biological properties, Schwann cells were cultured in ESP-NGC extracts and were assessed by morphological observation, the MTT assay, and immunohistochemistry. In addition, ESP-NGCs were subcutaneously implanted in the backs of rabbits to evaluate their biocompatibility in vivo. The results showed that ESP-NGCs have high porosity, strong hydrophilicity, and strong tensile stiffness. Schwann cells cultured in the ESP-NGC extract fluids showed no significant differences compared to control cells in their morphology or viability. Histological evaluation of the ESP-NGCs implanted in vivo indicated a mild inflammatory reaction and high biocompatibility. Together, these data suggest that these novel ESP-NGCs are biocompatible, and may thus provide a reliable scaffold for peripheral nerve repair in clinical application.