Knockdown of slit signaling during limb development leads to a reduction in humerus length.

BACKGROUND: Slits (1-3) and their Robo (1-3) receptors play multiple non-neuronal roles in development, including in development of muscle, heart and mammary gland. Previous work has demonstrated expression of Slit and Robo family members during limb development, where their functions are unclear. RESULTS: In situ hybridisation confirmed strong expression of Slit2, Slit3, Robo1, and Robo2 throughout mouse limb and joint development. No expression of Slit1 or Robo3 was detected. Analysis of Slit1/2 or Slit3 knockout mice revealed normal limb development. In contrast, locally blocking Slit signaling though grafting of cells expressing a dominant-negative Robo2 construct in the proximo-central region of developing chicken limb buds caused significant shortening of the humerus. CONCLUSIONS: These findings demonstrate an essential role for Slit/Robo signaling in regulating bone length during chicken limb development.

Migrating Schwann cells direct axon regeneration within the peripheral nerve bridge.

Schwann cells within the peripheral nervous system possess a remarkable regenerative potential. Current research shows that peripheral nerve-associated Schwann cells possess the capacity to promote repair of multiple tissues including peripheral nerve gap bridging, skin wound healing, digit tip repair as well as tooth regeneration. One of the key features of the specialized repair Schwann cells is that they become highly motile. They not only migrate into the area of damaged tissue and become a key component of regenerating tissue but also secrete signaling molecules to attract macrophages, support neuronal survival, promote axonal regrowth, activate local mesenchymal stem cells, and interact with other cell types. Currently, the importance of migratory Schwann cells in tissue regeneration is most evident in the case of a peripheral nerve transection injury. Following nerve transection, Schwann cells from both proximal and distal nerve stumps migrate into the nerve bridge and form Schwann cell cords to guide axon regeneration. The formation of Schwann cell cords in the nerve bridge is key to successful peripheral nerve repair following transection injury. In this review, we first examine nerve bridge formation and the behavior of Schwann cell migration in the nerve bridge, and then discuss how migrating Schwann cells direct regenerating axons into the distal nerve. We also review the current understanding of signals that could activate Schwann cell migration and signals that Schwann cells utilize to direct axon regeneration. Understanding the molecular mechanism of Schwann cell migration could potentially offer new therapeutic strategies for peripheral nerve repair.

Loc680254 regulates Schwann cell proliferation through Psrc1 and Ska1 as a microRNA sponge following sciatic nerve injury.

Peripheral nerve injury triggers sequential phenotype alterations in Schwann cells, which are critical for axonal regeneration. Long noncoding RNAs (lncRNAs) are long transcripts without obvious coding potential. It has been reported that lncRNAs participate in diverse biological processes and diseases. However, the role of lncRNA in Schwann cells and peripheral nerve regeneration is unclear. Here, we identified an lncRNA, loc680254, which is upregulated in rat sciatic nerve after peripheral nerve injury. The loc680254 knockdown inhibits Schwann cell proliferation, enhances apoptosis, and hinders cell cycle, while loc680254 overexpression has the opposite effect. Mechanically, we found that loc680254 might act as a microRNA sponge to regulate the expression of mitosis-related gene, spindle and kinetochore associated complex subunit 1 (Ska1) and proline/serine-rich coiled-coil 1 (Psrc1). Silencing of Psrc1 or Ska1 attenuates the effect of loc680254 overexpression on Schwann cell proliferation. Finally, we repaired the rat sciatic nerve gap with chitosan scaffolds loaded with loc680254-overexpressing Schwann cells and evaluated axon regeneration and functional recovery. Our results indicated that loc680254 is a new potential modulator for Schwann cell proliferation, which could be targeted to develop novel therapeutic strategies for peripheral nerve repair.

Sox2 expression in Schwann cells inhibits myelination in vivo and induces influx of macrophages to the nerve.

Correct myelination is crucial for the function of the peripheral nervous system. Both positive and negative regulators within the axon and Schwann cell function to ensure the correct onset and progression of myelination during both development and following peripheral nerve injury and repair. The Sox2 transcription factor is well known for its roles in the development and maintenance of progenitor and stem cell populations, but has also been proposed in vitro as a negative regulator of myelination in Schwann cells. We wished to test fully whether Sox2 regulates myelination in vivo and show here that, in mice, sustained Sox2 expression in vivo blocks myelination in the peripheral nerves and maintains Schwann cells in a proliferative non-differentiated state, which is also associated with increased inflammation within the nerve. The plasticity of Schwann cells allows them to re-myelinate regenerated axons following injury and we show that re-myelination is also blocked by Sox2 expression in Schwann cells. These findings identify Sox2 as a physiological regulator of Schwann cell myelination in vivo and its potential to play a role in disorders of myelination in the peripheral nervous system.

The role of p38alpha in Schwann cells in regulating peripheral nerve myelination and repair.

Myelination in the peripheral nervous system (PNS) is controlled by both positive and negative regulators within Schwann cells to ensure timely onset and correct myelin thickness for saltatory conduction by neurons. Transcription factors such as Sox10, octamer-binding transcription factor 6 (Oct6) and Krox20 form a positive regulatory network, whereas negative regulators such as cJun and Sox2 oppose myelination in Schwann cells. The role of the p38 MAPK pathway has been studied in PNS myelination, but its precise function remains unclear, with both positive and negative effects of p38 activity reported upon both myelination and processes of nerve repair. To clarify the role of p38 MAPK in the PNS, we have analysed mice with a Schwann cell-specific ablation of the major p38 isoform, p38alpha. In line with previous findings of an inhibitory role for p38 MAPK, we observe acceleration of post-natal myelination in p38alpha null nerves, a delay in myelin down-regulation following injury, together with a small increase in levels of re-myelination following injury. Finally we explored roles for p38alpha in controlling axonal regeneration and functional repair following PNS injury and observe that loss of p38alpha function in Schwann cells does not appear to affect these processes as previously reported. These studies therefore provide further proof for a role of p38 MAPK signalling in the control of myelination by Schwann cells in the PNS, but do not show an apparent role for signalling by this MAP kinase in Schwann cells controlling other elements of Wallerian degeneration and functional repair following injury. Cover Image for this issue: doi: 10.1111/jnc.13793.

Role of Netrin-1 Signaling in Nerve Regeneration.

Netrin-1 was the first axon guidance molecule to be discovered in vertebrates and has a strong chemotropic function for axonal guidance, cell migration, morphogenesis and angiogenesis. It is a secreted axon guidance cue that can trigger attraction by binding to its canonical receptors Deleted in Colorectal Cancer (DCC) and Neogenin or repulsion through binding the DCC/Uncoordinated (Unc5) A-D receptor complex. The crystal structures of Netrin-1/receptor complexes have recently been revealed. These studies have provided a structure based explanation of Netrin-1 bi-functionality. Netrin-1 and its receptor are continuously expressed in the adult nervous system and are differentially regulated after nerve injury. In the adult spinal cord and optic nerve, Netrin-1 has been considered as an inhibitor that contributes to axon regeneration failure after injury. In the peripheral nervous system, Netrin-1 receptors are expressed in Schwann cells, the cell bodies of sensory neurons and the axons of both motor and sensory neurons. Netrin-1 is expressed in Schwann cells and its expression is up-regulated after peripheral nerve transection injury. Recent studies indicated that Netrin-1 plays a positive role in promoting peripheral nerve regeneration, Schwann cell proliferation and migration. Targeting of the Netrin-1 signaling pathway could develop novel therapeutic strategies to promote peripheral nerve regeneration and functional recovery.

Loss of SOX10 function contributes to the phenotype of human Merlin-null schwannoma cells.

Loss of the Merlin tumour suppressor causes abnormal de-differentiation and proliferation of Schwann cells and formation of schwannoma tumours in patients with neurofibromatosis type 2. Within the mature peripheral nerve the normal development, differentiation and maintenance of myelinating and non-myelinating Schwann cells is regulated by a network of transcription factors that include SOX10, OCT6 (now known as POU3F1), NFATC4 and KROX20 (also known as Egr2). We have examined for the first time how their regulation of Schwann cell development is disrupted in primary human schwannoma cells. We find that induction of both KROX20 and OCT6 is impaired, whereas enforced expression of KROX20 drives both myelin gene expression and cell cycle arrest in Merlin-null cells. Importantly, we show that human schwannoma cells have reduced expression of SOX10 protein and messenger RNA. Analysis of mouse SOX10-null Schwann cells shows they display many of the characteristics of human schwannoma cells, including increased expression of platelet derived growth factor receptor beta (PDGFRB) messenger RNA and protein, enhanced proliferation, increased focal adhesions and schwannoma-like morphology. Correspondingly, reintroduction of SOX10 into human Merlin-null cells restores the ability of these cells to induce KROX20 and myelin protein zero (MPZ), localizes NFATC4 to the nucleus, reduces cell proliferation and suppresses PDGFRB expression. Thus, we propose that loss of the SOX10 protein, which is vital for normal Schwann cell development, is also key to the pathology of Merlin-null schwannoma tumours.

Drebrin controls neuronal migration through the formation and alignment of the leading process.

Formation of a functional nervous system requires neurons to migrate to the correct place within the developing brain. Tangentially migrating neurons are guided by a leading process which extends towards the target and is followed by the cell body. How environmental cues are coupled to specific cytoskeletal changes to produce and guide leading process growth is unknown. One such cytoskeletal modulator is drebrin, an actin-binding protein known to induce protrusions in many cell types and be important for regulating neuronal morphology. Using the migration of oculomotor neurons as a model, we have shown that drebrin is necessary for the generation and guidance of the leading process. In the absence of drebrin, leading processes are not formed and cells fail to migrate although axon growth and pathfinding appear grossly unaffected. Conversely, when levels of drebrin are elevated the leading processes turn away from their target and as a result the motor neuron cell bodies move along abnormal paths within the brain. The aberrant trajectories were highly reproducible suggesting that drebrin is required to interpret specific guidance cues. The axons and growth cones of these neurons display morphological changes, particularly increased branching and filopodial number but despite this they extend along normal developmental pathways. Collectively these results show that drebrin is initially necessary for the formation of a leading process and subsequently for this to respond to navigational signals and grow in the correct direction. Furthermore, we have shown that the actions of drebrin can be segregated within individual motor neurons to direct their migration independently of axon guidance.

Regulation of Schwann cell differentiation and proliferation by the Pax-3 transcription factor.

Pax-3 is a paired domain transcription factor that plays many roles during vertebrate development. In the Schwann cell lineage, Pax-3 is expressed at an early stage in Schwann cells precursors of the embryonic nerve, is maintained in the nonmyelinating cells of the adult nerve, and is upregulated in Schwann cells after peripheral nerve injury. Consistent with this expression pattern, Pax-3 has previously been shown to play a role in repressing the expression of the myelin basic protein gene in Schwann cells. We have studied the role of Pax-3 in Schwann cells and have found that it controls not only the regulation of cell differentiation but also the survival and proliferation of Schwann cells. Pax-3 expression blocks both the induction of Oct-6 and Krox-20 (K20) by cyclic AMP and completely inhibits the ability of K20, the physiological regulator of myelination in the peripheral nervous system, to induce myelin gene expression in Schwann cells. In contrast to other inhibitors of myelination, we find that Pax-3 represses myelin gene expression in a c-Jun-independent manner. In addition to this, we find that Pax-3 expression alone is sufficient to inhibit the induction of apoptosis by TGFß1 in Schwann cells. Expression of Pax-3 is also sufficient to induce the proliferation of Schwann cells in the absence of added growth factors and to reverse K20-induced exit from the cell cycle. These findings indicate new roles for the Pax-3 transcription factor in controlling the differentiation and proliferation of Schwann cells during development and after peripheral nerve injury.

Single Cell Transcriptome Data Analysis Defines the Heterogeneity of Peripheral Nerve Cells in Homeostasis and Regeneration.

The advances in single-cell RNA sequencing technologies and the development of bioinformatics pipelines enable us to more accurately define the heterogeneity of cell types in a selected tissue. In this report, we re-analyzed recently published single-cell RNA sequencing data sets and provide a rationale to redefine the heterogeneity of cells in both intact and injured mouse peripheral nerves. Our analysis showed that, in both intact and injured peripheral nerves, cells could be functionally classified into four categories: Schwann cells, nerve fibroblasts, immune cells, and cells associated with blood vessels. Nerve fibroblasts could be sub-clustered into epineurial, perineurial, and endoneurial fibroblasts. Identified immune cell clusters include macrophages, mast cells, natural killer cells, T and B lymphocytes as well as an unreported cluster of neutrophils. Cells associated with blood vessels include endothelial cells, vascular smooth muscle cells, and pericytes. We show that endothelial cells in the intact mouse sciatic nerve have three sub-types: epineurial, endoneurial, and lymphatic endothelial cells. Analysis of cell type-specific gene changes revealed that Schwann cells and endoneurial fibroblasts are the two most important cell types promoting peripheral nerve regeneration. Analysis of communication between these cells identified potential signals for early blood vessel regeneration, neutrophil recruitment of macrophages, and macrophages activating Schwann cells. Through this analysis, we also report appropriate marker genes for future single cell transcriptome data analysis to identify cell types in intact and injured peripheral nerves. The findings from our analysis could facilitate a better understanding of cell biology of peripheral nerves in homeostasis, regeneration, and disease.

Dynamic expression of Slit1-3 and Robo1-2 in the mouse peripheral nervous system after injury.

The Slit family of axon guidance cues act as repulsive molecules for precise axon pathfinding and neuronal migration during nervous system development through interactions with specific Robo receptors. Although we previously reported that Slit1-3 and their receptors Robo1 and Robo2 are highly expressed in the adult mouse peripheral nervous system, how this expression changes after injury has not been well studied. Herein, we constructed a peripheral nerve injury mouse model by transecting the right sciatic nerve. At 14 days after injury, quantitative real-time polymerase chain reaction was used to detect mRNA expression of Slit1-3 and Robo1-2 in L4-5 spinal cord and dorsal root ganglia, as well as the sciatic nerve. Immunohistochemical analysis was performed to examine Slit1-3, Robo1-2, neurofilament heavy chain, F4/80, and vimentin in L4-5 spinal cord, L4 dorsal root ganglia, and the sciatic nerve. Co-expression of Slit1-3 and Robo1-2 in L4 dorsal root ganglia was detected by in situ hybridization. In addition, Slit1-3 and Robo1-2 protein expression in L4-5 spinal cord, L4 dorsal root ganglia, and sciatic nerve were detected by western blot assay. The results showed no significant changes of Slit1-3 or Robo1-2 mRNA expression in the spinal cord within 14 days after injury. In the dorsal root ganglion, Slit1-3 and Robo1-2 mRNA expression were initially downregulated within 4 days after injury; however, Robo1-2 mRNA expression returned to the control level, while Slit1-3 mRNA expression remained upregulated during regeneration from 4-14 days after injury. In the sciatic nerve, Slit1-3 and their receptors Robo1-2 were all expressed in the proximal nerve stump; however, Slit1, Slit2, and Robo2 were barely detectable in the nerve bridge and distal nerve stump within 14 days after injury. Slit3 was highly ex-pressed in macrophages surrounding the nerve bridge and slightly downregulated in the distal nerve stump within 14 days after injury. Robo1 was upregulated in vimentin-positive cells and migrating Schwann cells inside the nerve bridge. Robo1 was also upregulated in Schwann cells of the distal nerve stump within 14 days after injury. Our findings indicate that Slit3 is the major ligand expressed in the nerve bridge and distal nerve stump during peripheral nerve regeneration, and Slit3/Robo signaling could play a key role in peripheral nerve repair after injury. This study was approved by Plymouth University Animal Welfare Ethical Review Board (approval No. 30/3203) on April 12, 2014.

Classic axon guidance molecules control correct nerve bridge tissue formation and precise axon regeneration.

The peripheral nervous system has an astonishing ability to regenerate following a compression or crush injury; however, the potential for full repair following a transection injury is much less. Currently, the major clinical challenge for peripheral nerve repair come from long gaps between the proximal and distal nerve stumps, which prevent regenerating axons reaching the distal nerve. Precise axon targeting during nervous system development is controlled by families of axon guidance molecules including Netrins, Slits, Ephrins and Semaphorins. Several recent studies have indicated key roles of Netrin1, Slit3 and EphrinB2 signalling in controlling the formation of new nerve bridge tissue and precise axon regeneration after peripheral nerve transection injury. Inside the nerve bridge, nerve fibroblasts express EphrinB2 while migrating Schwann cells express the receptor EphB2. EphrinB2/EphB2 signalling between nerve fibroblasts and migrating Schwann cells is required for Sox2 upregulation in Schwann cells and the formation of Schwann cell cords within the nerve bridge to allow directional axon growth to the distal nerve stump. Macrophages in the outermost layer of the nerve bridge express Slit3 while migrating Schwann cells and regenerating axons express the receptor Robo1; within Schwann cells, Robo1 expression is also Sox2-dependent. Slit3/Robo1 signalling is required to keep migrating Schwann cells and regenerating axons inside the nerve bridge. In addition to the Slit3/Robo1 signalling system, migrating Schwann cells also express Netrin1 and regenerating axons express the DCC receptor. It appears that migrating Schwann cells could also use Netrin1 as a guidance cue to direct regenerating axons across the peripheral nerve gap. Engineered neural tissues have been suggested as promising alternatives for the repair of large peripheral nerve gaps. Therefore, understanding the function of classic axon guidance molecules in nerve bridge formation and their roles in axon regeneration could be highly beneficial in developing engineered neural tissue for more effective peripheral nerve repair.

FGF5 Regulates Schwann Cell Migration and Adhesion.

The fibroblast growth factor (FGF) family polypeptides play key roles in promoting tissue regeneration and repair. FGF5 is strongly up-regulated in Schwann cells of the peripheral nervous system following injury; however, a role for FGF5 in peripheral nerve regeneration has not been shown up to now. In this report, we examined the expression of FGF5 and its receptors FGFR1-4 in Schwann cells of the mouse sciatic nerve following injury, and then measured the effects of FGF5 treatment upon cultured primary rat Schwann cells. By microarray and mRNA sequencing data analysis, RT-PCR, qPCR, western blotting and immunostaining, we show that FGF5 is highly up-regulated in Schwann cells of the mouse distal sciatic nerve following injury, and FGFR1 and FGFR2 are highly expressed in Schwann cells of the peripheral nerve both before and following injury. Using cultured primary rat Schwann cells, we show that FGF5 inhibits ERK1/2 MAP kinase activity but promotes rapid Schwann cell migration and adhesion via the upregulation of N-cadherin. Thus, FGF5 is an autocrine regulator of Schwann cells to regulate Schwann cell migration and adhesion.

Grape Seed Proanthocyanidin Extract Ameliorates Streptozotocin-induced Cognitive and Synaptic Plasticity Deficits by Inhibiting Oxidative Stress and Preserving AKT and ERK Activities.

Progressive memory loss and cognitive impairment are the main clinical manifestations of Alzheimer's disease (AD). Currently, there is no effective drug available for the treatment of AD. Previous studies have demonstrated that the cognitive impairment of AD is associated with oxidative stress and the inhibition of AKT and ERK phosphorylation. Grape seed proanthocyanidin extract (GSPE) has been shown to have strong antioxidant effect and can protect the nervous system from oxidative stress damage. This study aimed to investigate the protective effect of GSPE on the cognitive and synaptic impairments of AD using a sporadic AD rat model induced by intracerebroventricular (ICV) injection of streptozotocin (STZ) (ICV-STZ). Rats were treated with GSPE (50, 100, or 200 mg/kg every day) by intragastrical (ig.) administration for continuous 7 weeks, and ICV-STZ (3 mg/kg) was performed on the first day and third day of week 5. Learning and memory abilities were assessed by the Morris water maze (MWM) test at week 8. After behavioral test, hippocampal long-term potentiation (LTP) was recorded, and the levels of malondialdehyde (MDA), superoxide dismutases (SOD), glutathione (GSH) and the protein expression of AKT and ERK were measured in the hippocampus and cerebral cortex of rats. Our study revealed that ICV-STZ significantly impaired the working learning ability and hippocampal LTP of rats, significantly increased the levels of MDA, and decreased the activity of SOD and GSH in the hippocampus and cerebral cortex. In contrast, GSPE treatment prevented the impairment of cognitive function and hippocampal LTP induced by ICV-STZ, decreased the level of MDA, and increased the level of SOD and GSH. Furthermore, Western blot results showed that GSPE treatment could prevent the loss of AKT and ERK activities in the hippocampus and cerebral cortex induced by ICV-STZ. Our findings demonstrate that GSPE treatment could ameliorate the impairment of cognitive ability and hippocampal synaptic plasticity in a rat model of sporadic AD by inhibiting oxidative stress and preserving AKT and ERK activities. Therefore, GSPE may be an effective agent for the treatment of cognitive deficits associated with sporadic AD.

A preliminary study on DRGs and spinal cord of a galanin receptor 2-EGFP transgenic mouse.

The neuropeptide galanin functions via three G-protein coupled receptors, Gal1-3-R. Both Gal1-R and 2-R are involved in pain signaling at the spinal level. Here a Gal2-R-EGFP transgenic (TG) mouse was generated and studied in pain tests and by characterizing Gal2-R expression in both sensory ganglia and spinal cord. After peripheral spared nerve injury, mechanical allodynia developed and was ipsilaterally similar between wild type (WT) and TG mice. A Gal2-R-EGFP-positive signal was primarily observed in small and medium-sized dorsal root ganglion (DRG) neurons and in spinal interneurons and processes. No significant difference in size distribution of DRG neuronal profiles was found between TG and WT mice. Both percentage and fluorescence intensity of Gal2-R-EGFP-positive neuronal profiles were overall significantly upregulated in ipsilateral DRGs as compared to contralateral DRGs. There was an ipsilateral reduction in substance P-positive and calcitonin gene-related peptide (CGRP)-positive neuronal profiles, and this reduction was more pronounced in TG as compared to WT mice. Moreover, Gal2-R-EGFP partly co-localized with three pain-related neuropeptides, CGRP, neuropeptide Y and galanin, both in intact and injured DRGs, and with galanin also in local neurons in the superficial dorsal horn. Taken together, the present results provide novel information on the localization and phenotype of DRG and spinal neurons expressing the second galanin receptor, Gal2-R, and on phenotypic changes following peripheral nerve injury. Gal2-R may also be involved in autoreceptor signaling.

Analysis of Schwann Cell Migration and Axon Regeneration Following Nerve Injury in the Sciatic Nerve Bridge.

While it is proposed that interaction between Schwann cells and axons is key for successful nerve regeneration, the behavior of Schwann cells migrating into a nerve gap following a transection injury and how migrating Schwann cells interact with regenerating axons within the nerve bridge has not been studied in detail. In this study, we combine the use of our whole-mount sciatic nerve staining with the use of a proteolipid protein-green fluorescent protein (PLP-GFP) mouse model to mark Schwann cells and have examined the behavior of migrating Schwann cells and regenerating axons in the sciatic nerve gap following a nerve transection injury. We show here that Schwann cell migration from both nerve stumps starts later than the regrowth of axons from the proximal nerve stump. The first migrating Schwann cells are only observed 4 days following mouse sciatic nerve transection injury. Schwann cells migrating from the proximal nerve stump overtake regenerating axons on day 5 and form Schwann cell cords within the nerve bridge by 7 days post-transection injury. Regenerating axons begin to attach to migrating Schwann cells on day 6 and then follow their trajectory navigating across the nerve gap. We also observe that Schwann cell cords in the nerve bridge are not wide enough to guide all the regenerating axons across the nerve bridge, resulting in regenerating axons growing along the outside of both proximal and distal nerve stumps. From this analysis, we demonstrate that Schwann cells play a crucial role in controlling the directionality and speed of axon regeneration across the nerve gap. We also demonstrate that the use of the PLP-GFP mouse model labeling Schwann cells together with the whole sciatic nerve axon staining technique is a useful research model to study the process of peripheral nerve regeneration.

Distinct VIP and PACAP Functions in the Distal Nerve Stump During Peripheral Nerve Regeneration.

Vasoactive Intestinal Peptide (VIP) and Pituitary Adenylyl Cyclase Activating Peptide (PACAP) are regeneration-associated neuropeptides, which are up-regulated by neurons following peripheral nerve injury. So far, they have only been studied for their roles as autocrine signals for both neuronal survival and axon outgrowth during peripheral nerve regeneration. In this report, we examined VIP and PACAP's paracrine effects on Schwann cells and macrophages in the distal nerve stump during peripheral nerve regeneration. We show that VPAC1, VPAC2, and PAC1 are all up-regulated in the mouse distal nerve following peripheral nerve injury and are highly expressed in Schwann cells and macrophages within the distal sciatic nerve. We further investigated the effect of VIP and PACAP on cultured rat Schwann cells, and found that VIP and PACAP can not only promote myelin gene expression in Schwann cells but can also inhibit the release of pro-inflammatory cytokines by Schwann cells. Furthermore, we show that VIP and PACAP inhibit the release of pro-inflammatory cytokines and enhance anti-inflammatory cytokine expression in sciatic nerve explants. Our results provide evidence that VIP and PACAP could have important functions in the distal nerve stump following injury to promote remyelination and regulate the inflammatory response. Thus, VIP and PACAP receptors appear as important targets to promote peripheral nerve repair following injury.

Macrophage-Derived Slit3 Controls Cell Migration and Axon Pathfinding in the Peripheral Nerve Bridge.

Slit-Robo signaling has been characterized as a repulsive signal for precise axon pathfinding and cell migration during embryonic development. Here, we describe a role for Sox2 in the regulation of Robo1 in Schwann cells and for Slit3-Robo1 signaling in controlling axon guidance within the newly formed nerve bridge following peripheral nerve transection injury. In particular, we show that macrophages form the outermost layer of the nerve bridge and secrete high levels of Slit3, while migratory Schwann cells and fibroblasts inside the nerve bridge express the Robo1 receptor. In line with this pattern of Slit3 and Robo1 expression, we observed multiple axon regeneration and cell migration defects in the nerve bridge of Sox2-, Slit3-, and Robo1-mutant mice. Our findings have revealed important functions for macrophages in the peripheral nervous system, utilizing Slit3-Robo1 signaling to control correct peripheral nerve bridge formation and precise axon targeting to the distal nerve stump following injury.

The effect of pea albumin 1F on glucose metabolism in mice.

Pea albumin 1F (PA1F), a plant peptide isolated from pea seeds, can dramatically increase blood glucose concentration by subcutaneous injection with a dosage of 5 or 10 microg/g (body weight) in normal and type II diabetic mice (KK/upj-Ay). The voltage-dependent anion channel 1 (VDAC-1) has been identified as the PA1F binding protein from mice pancreatic cell membrane, which may be involved in the regulation of enhancing blood glucose in response to PA1F binding. The results clearly show that peptide-signaling molecules from plants can affect mammalian physiological functions, especially, in association with glucose metabolism.

Transection and Crush Models of Nerve Injury to Measure Repair and Remyelination in Peripheral Nerve.

Injury to the peripheral nervous system begins a well-characterized process within both neurons and Schwann cells to allow axonal regrowth, remyelination, and functional repair. Models of peripheral nerve injury have been widely used to study the behavior of Schwann cells, neurons, and other cell types such as macrophages as the events of Wallerian degeneration and regeneration take place. The most commonly used approaches in rodent models to model nerve injury in human patients are sciatic nerve transection and nerve crush, and both have well established time courses of demyelination, immune cell influx, axonal regrowth, and remyelination. We describe the techniques of sciatic nerve surgery for transection and crush injury, together with methods for the analysis of events within peripheral nerve repair in these two models.