In vivo knockdown of ErbB3 in mice inhibits Schwann cell precursor migration

The myelin sheath insulates neuronal axons and markedly increases the nerve conduction velocity. In the peripheral nervous system (PNS), Schwann cell precursors migrate along embryonic neuronal axons to their final destinations, where they eventually wrap around individual axons to form the myelin sheath after birth. ErbB2 and ErbB3 tyrosine kinase receptors form a heterodimer and are extensively expressed in Schwann lineage cells. ErbB2/3 is thought to be one of the primary regulators controlling the entire Schwann cell development. ErbB3 is the bona fide Schwann cell receptor for the neuronal ligand neuregulin-1. Although ErbB2/3 is well known to regulate both Schwann cell precursor migration and myelination by Schwann cells in fishes, it still remains unclear whether in mammals, ErbB2/3 actually regulates Schwann cell precursor migration. Here, we show that knockdown of ErbB3 using a Schwann cell-specific promoter in mice causes delayed migration of Schwann cell precursors. In contrast, littermate control mice display normal migration. Similar results are seen in an in vitro migration assay using reaggregated Schwann cell precursors. Also, ErbB3 knockdown in mice reduces myelin thickness in sciatic nerves, consistent with the established role of ErbB3 in myelination. Thus, ErbB3 plays a key role in migration, as well as in myelination, in mouse Schwann lineage cells, presenting a genetically conservative role of ErbB3 in Schwann cell precursor migration.     

Cell autonomy of the mouse claw paw mutation

Mice homozygous for the autosomal recessive mutation claw paw (clp) are characterized by limb posture abnormalities and congenital hypomyelination, with delayed onset of myelination of the peripheral nervous system but not the central nervous system. Although this combination of limb and peripheral nerve abnormalities in clp/clp mice might suggest a common neurogenic origin of the syndrome, it is not clear whether the clp gene acts primarily in the neurone, the Schwann cell or both. In the work described here, we address this question of cell autonomy of the clp mutation through reciprocal nerve grafting experiments between wild-type and clp/clp animals. Our results demonstrate that the clp mutation affects the Schwann cell compartment and possibly also the neuronal compartment. These data suggest that the clp gene product is expressed in Schwann cells as well as neurones and is likely to be involved in direct axon--Schwann cell interactions. Within the Schwann cell, clp affects a myelin-related signaling pathway that regulates periaxin and Krox-20 expression, but not Oct-6.     

Membrane-bound neuregulin1 type III actively promotes Schwann cell differentiation of multipotent Progenitor cells

Many steps of peripheral glia development appear to be regulated by neuregulin1 (NRG1) signaling but the exact roles of the different NRG1 isoforms in these processes remain to be determined. While glial growth factor 2 (GGF2), a NRG1 type II isoform, is able to induce a satellite glial fate in neural crest stem cells, targeted mutations in mice have revealed a prominent role of NRG1 type III isoforms in supporting survival of Schwann cells at early developmental stages. Here, we investigated the role of NRG1 isoforms in the differentiation of Schwann cells from neural crest-derived progenitor cells. In multipotent cells isolated from dorsal root ganglia, soluble NRG1 isoforms do not promote Schwann cell features, whereas signaling by membrane-associated NRG1 type III induces the expression of the Schwann cell markers Oct-6/SCIP and S100 in neighboring cells, independent of survival. Thus, axon-bound NRG1 might actively promote both Schwann cell survival and differentiation.     

A dual role of erbB2 in myelination and in expansion of the schwann cell precursor pool

Neuregulin-1 provides an important axonally derived signal for the survival and growth of developing Schwann cells, which is transmitted by the ErbB2/ErbB3 receptor tyrosine kinases. Null mutations of the neuregulin-1, erbB2, or erbB3 mouse genes cause severe deficits in early Schwann cell development. Here, we employ Cre-loxP technology to introduce erbB2 mutations late in Schwann cell development, using a Krox20-cre allele. Cre-mediated erbB2 ablation occurs perinatally in peripheral nerves, but already at E11 within spinal roots. The mutant mice exhibit a widespread peripheral neuropathy characterized by abnormally thin myelin sheaths, containing fewer myelin wraps. In addition, in spinal roots the Schwann cell precursor pool is not correctly established. Thus, the Neuregulin signaling system functions during multiple stages of Schwann cell development and is essential for correct myelination. The thickness of the myelin sheath is determined by the axon diameter, and we suggest that trophic signals provided by the nerve determine the number of times a Schwann cell wraps an axon.     

Autophagy Is Involved in the Reduction of Myelinating Schwann Cell Cytoplasm during Myelin Maturation of the Peripheral Nerve

Peripheral nerve myelination involves dynamic changes in Schwann cell morphology and membrane structure. Recent studies have demonstrated that autophagy regulates organelle biogenesis and plasma membrane dynamics. In the present study, we investigated the role of autophagy in the development and differentiation of myelinating Schwann cells during sciatic nerve myelination. Electron microscopy and biochemical assays have shown that Schwann cells remove excess cytoplasmic organelles during myelination through macroautophagy. Inhibition of autophagy via Schwann cell-specific removal of ATG7, an essential molecule for macroautophagy, using a conditional knockout strategy, resulted in abnormally enlarged abaxonal cytoplasm in myelinating Schwann cells that contained a large number of ribosomes and an atypically expanded endoplasmic reticulum. Small fiber hypermyelination and minor anomalous peripheral nerve functions are observed in this mutant. Rapamycin-induced suppression of mTOR activity during the early postnatal period enhanced not only autophagy but also developmental reduction of myelinating Schwann cells cytoplasm in vivo. Together, our findings suggest that autophagy is a regulatory mechanism of Schwann cells structural plasticity during myelination.     

CAP1 was associated with actin and involved in Schwann cell differentiation and motility after sciatic nerve injury

Adenylate cyclase-associated protein 1 (CAP1), a member of cyclase-associated proteins that regulating actin dynamics, was shown to regulate actin filaments, localize to dynamic actin structures and mediate such processes as establishment of cell polarity, motility, morphogenesis, receptor-mediated endocytosis and mRNA location. But little is known about the role of CAP1 during peripheral nervous system injury. Here, we found the spatiotemporal protein expression of CAP1 after sciatic nerve crush. After crush, CAP1 had an increased protein expression level, reached a peak at about day 5 and then returned to the normal level at 4 weeks, similar to Oct-6. Besides, in 5-day injured tissue, using double immunofluorescent staining we found CAP1 had a colocalization with S100 and Oct-6. In vitro, during the process of cAMP-induced Schwann cells differentiation, we observed enhanced expression of CAP1 and P0. Specially, CAP1-specific siRNA-tranfected SCs did not show significant actin structure which form cellure surface tension and protrusion shape after cAMP treatment. And we observed the interaction of CAP1 with actin and that CAP1-specific siRNA-transfected SCs had a decreased motility and migration. Together, all these data indicated that the change of CAP1 protein expression was associated with Schwann cells motility and differentiation after the crush of sciatic nerve.     

Conditional beta1-integrin gene deletion in neural crest cells causes severe developmental alterations of the peripheral nervous system

Integrins are transmembrane receptors that are known to interact with the extracellular matrix and to be required for migration, proliferation, differentiation and apoptosis. We have generated mice with a neural crest cell-specific deletion of the beta1-integrin gene to analyse the role of beta1-integrins in neural crest cell migration and differentiation. This targeted mutation caused death within a month of birth. The loss of beta1-integrins from the embryo delayed the migration of Schwann cells along axons and induced multiple defects in spinal nerve arborisation and morphology. There was an almost complete absence of Schwann cells and sensory axon segregation and defective maturation in neuromuscular synaptogenesis. Thus, beta1-integrins are important for the control of embryonic and postnatal peripheral nervous system development.     

Upregulated Expression of TRIM32 Is Involved in Schwann Cell Differentiation,Migration and Neurite Outgrowth After Sciatic Nerve Crush

Tripartite motif containing 32 (TRIM32), a member of the tripartite motif (TRIM) family, plays an indispensable role in myoblast proliferation. It also regulates neuron and skeletal muscle stem cell differentiation. Although it is of great importance, we know little about the roles of TRIM32 during peripheral nervous system injury. Here, we examined the dynamic changes of TRIM32 in acute sciatic nerve crush (SNC) model. After crush, TRIM32 rapidly increased and reached the climax at 1 week but then gradually declined to the normal level at 4 weeks post-injury. Meanwhile, we observed similar changes of Oct-6. What is more, we found co-localization of TRIM32 with S100 and Oct-6 in 1-week-injured tissues using double immunofluorescent staining. In further vitro experiments, enhancive expression of TRIM32 was detected during the process of cyclic adenosine monophosphate (cAMP)-induced Schwann cell differentiation and nerve growth factor (NGF)-induced PC12 cell neurite outgrowth. More interestingly, specific si-TRIM32-transfected RSC96 cells exhibited obvious reduction in the ability of migration. Taken together, we inferred that upregulated TRIM32 was not only involved in the differentiation and migration of Schwann cells but the neurite elongation after SNC.     

Growth Arrest and Spontaneous Differentiation Are Initiated Through an Autocrine Loop in Clonally Derived Schwann Cells by α1-Procollagen I C-Propeptide

Schwann cells cloned from rat sciatic nerve survive and display self-induced growth suppression, or undergo spontaneous apoptosis, on long-term serum-free subconfluent culture. Strain SCL4.1/F7 sustained the capacity to growth arrest for up to 40 generations. A soluble activity transmitted between neighbouring cells of this strain suppresses DNA synthesis within three cell cycles. Autocrine Schwann cell growth-inhibitory factor (SGIF) operates during the G1 phase of the cell cycle, overcomes the mitogenic action of Schwann cell/serum-associated (platelet-derived growth factor-BB) and axon-associated (axolemma-enriched fraction) stimuli in serum-free conditions, and suppresses DNA synthesis in sciatic nerve Schwann cell cultures in a stage-specific manner. A 35-kDa protein with N-terminal sequence and approximate molecular mass of the C-propeptide of rat alpha1-procollagen I makes a major contribution to SGIF. Growth suppression in the SCL4.1/F7 strain is mediated by the ras/extracellular signal-regulated kinase pathway, is accompanied by down-regulation of erbB2/erbB3 and of tetraethylammonium-sensitive K+ currents, and is followed by transition of cells within 5-10 days from O4+, p75 nerve growth factor receptor (p75NGF-R)+, glial fibrillary acidic protein (GFAP)+ to O4+, p75NGF-R-, GFAP-, periaxin+ phenotypes. Oct-6/SCIP mRNA is present in both proliferating and growth-arrested SCL4.1/F7 cells. These results demonstrate an autocrine/ paracrine loop for the growth arrest of clonally derived Schwann cells in the absence of axons linked in part to the metabolism of collagen. Schwann cells thus appear to self-regulate growth in a negative as well as a positive direction through characterized molecular mechanisms and signal pathways.     

Neuron-schwann cell interaction in basal lamina formation

The availability of tissue culture systems that allow the growth of nerve cells, Schwann cells, and fibroblasts separately or in various combinations now makes possible investigation of the role of cell interactions in the development of the peripheral nervous system. Using these systems it was earlier found that basal lamina is formed on the Schwann cell surface in cultures of sensory ganglion cells and Schwann cells without fibroblasts. It is here reported that the presence of nerve cells is required for the generation of basal lamina on the Schwann cell plasmalemma. Utilizing nerve cell-Schwann cell preparations devoid of fibroblasts, this was found in the following ways. (1) When nerve cells are removed from 3- to 5-week-old cultures, the basal lamina disappears from Schwann cells. (2) If nerve cells are added back to such Schwann cell populations, Schwann cell basal lamina reappears. (3) Removal of nerve cells from older (3–4 months) cultures does not lead to basal lamina loss; areas presumed not to have been coated with lamina before neurite degeneration remain so, suggesting that the lamina persists but is not reformed. (4) If basal lamina is removed with trypsin, it is reformed in neuron plus Schwann cell cultures but not in Schwann cell populations alone. Thus, the formation but not the persistence of Schwann cell basal lamina requires the presence of nerve cells.     

Conditional disruption of beta 1 integrin in Schwann cells impedes interactions with axons.

In dystrophic mice, a model of merosin-deficient congenital muscular dystrophy, laminin-2 mutations produce peripheral nerve dysmyelination and render Schwann cells unable to sort bundles of axons. The laminin receptor and the mechanism through which dysmyelination and impaired sorting occur are unknown. We describe mice in which Schwann cell-specific disruption of beta1 integrin, a component of laminin receptors, causes a severe neuropathy with impaired radial sorting of axons. beta 1-null Schwann cells populate nerves, proliferate, and survive normally, but do not extend or maintain normal processes around axons. Interestingly, some Schwann cells surpass this problem to form normal myelin, possibly due to the presence of other laminin receptors such as dystroglycan and alpha 6 beta 4 integrin. These data suggest that beta 1 integrin links laminin in the basal lamina to the cytoskeleton in order for Schwann cells to ensheath axons, and alteration of this linkage contributes to the peripheral neuropathy of congenital muscular dystrophy.     

Interleukin-6 induces proinflammatory signaling in Schwann cells: A high-throughput analysis

Interleukin-6 plays an important role in peripheral nerve regeneration. We recently reported that IL-6 targets Schwann cells in the peripheral nerve for its function. In this study, we analyzed genes whose expression is regulated by IL-6 in a cell line derived from Schwann cells, the peripheral glia, using the Illumina gene microarray. At measurements 3 and 12 h after IL-6 treatment, 35 genes were found to be upregulated by IL-6. Most upregulated genes were proinflammatory genes that are known to be induced in inflammatory conditions. Interestingly, the expression of immunoproteasome subunits was upregulated by IL-6 in Schwann cells. Treatment with forskolin, an agent that mimics axonal signaling, suppressed the expression of IL-6-inducible genes. Finally, we found for the first time that sciatic nerve injury induced immunoproteasome expression in vivo. These findings indicate that IL-6 is involved in peripheral nerve regeneration by regulating proinflammatory signaling in Schwann cells.     

Neuregulin, a factor with many functions in the life of a schwann cell

The signalling system comprising the ligand Neuregulin-1, and its receptors, ErbB2 and ErbB3, plays multiple and important roles in glial development. These include functions in early development of neural crest cells, in expansion of the Schwann cell precursor pool and in myelination. Neuregulin is one of the crucial axon-derived signals that influence development of Schwann cells. These are specialized cells that ensheath peripheral axons and provide electrical insulation. Schwann cells have also long been implicated in providing more than a simple ensheathing function. Compelling evidence for this has emerged from the analysis of mice lacking these cells, resulting from a non-functional or compromised Neuregulin signalling system. They serve as a model to study glia-nerve interactions in vivo and indicate that Schwann cells provide important neurotrophic signals, and also cues that regulate perineurium development and nerve fasciculation.