Regulation of Schwann cell proliferation in cultured segments of the adult rat sciatic nerve

Schwann cell proliferation was studied in cultured segments of the rat sciatic nerve by measurement of [³H] thymidine incorporation or through bromodeoxyuridine-(BrdU)-labelling and immunocytochemistry. The aim was to delineate mechanisms involved in the injury-induced proliferative response of Schwann cells. Removal of extracellular Ca²⁺ by addition of EGTA to the culture medium suppressed [³H] thymidine incorporation as did the calmodulin inhibitor 48/80. The Ca²⁺ ionophore A23187 increased incorporation. Staurosporin, an inhibitor of protein kinase C (PKC), suppressed [³H] thymidine incorporation while phorbol-12-myristate-13-acetate (PMA) enhanced incorporation. Manipulation of the cAMP system showed that increased cAMP levels inhibited proliferation. Inhibition of protein kinase A by HA 1004 increased the incorporation of [³H] thymidine. Immunostaining for BrdU and glial specific markers together with morphological evaluation of myelin association showed that proliferation occurred in Schwann cells. The results are consistent with a model in which Schwann cell proliferation is enhanced by Ca²⁺ through activation of calmodulin-dependent and/or PKCdependent mechanisms. Inhibition is achieved through the cAMP system. Together, these results show that Schwann cells regulate proliferation differently in an integrated environment, e.g. the nerve structure, than in isolation as primary monocultures. J. Neurosci. Res. 52:530–537, 1998. © 1998 Wiley-Liss, Inc.     

Activation of MAP kinases, Akt and PDGF receptors in injured peripheral nerves

A number of receptor tyrosine kinases (RTKs) and the downstream phosphatidylinositol-3-kinase (PI3K)/Akt and mitogen-activated protein (MAP) kinase signaling pathways have been critically involved in peripheral nerve regeneration. Here, we examined the activation of PI3K/Akt and MAP kinase pathways, and platelet-derived growth factor receptors (PDGFRs) in the distal segments of crushed rat sciatic nerve from 3 to 28 days after injury. In Western blot analyses, the phosphorylated forms of extracellular signal-regulated protein kinase (ERK) and c-Jun NH₂-terminal kinases (JNKs) were highly augmented on days 3 and 7 and on days 7 and 14 after injury, respectively. Phosphorylated Akt and p38 consistently increased from 3 to 28 days after injury. Phosphorylated PDGFR-α and -β were also increased from 3 to 14 days. In the immunohistological analyses, phosphorylated ERK and PDGFR-α were co-localized in many activated Schwann cells and regrowing axons 3 days after injury, while PDGFR-β was localized in a few spindle-shaped cells. The detected temporal profile of RTK signaling appears to be crucial for the regulation of Schwann cell proliferation and following redifferentiation. Furthermore, the immunohistological studies suggested a role of ERK and PDGFR-α in axon regeneration as well.     

Motor and sensory Schwann cell phenotype commitment is diminished by extracorporeal shockwave treatment in vitro

The gold standard for peripheral nerve regeneration uses a sensory autograft to bridge a motor/sensory defect site. For motor nerves to regenerate, Schwann cells (SC) myelinate the newly grown axon. Sensory SCs have a reduced ability to produce myelin, partially explaining low success rates of autografts. This issue is masked in pre‐clinical research by the excessive use of the rat sciatic nerve defect model, utilizing a mixed nerve with motor and sensory SCs. Aim of this study was to utilize extracorporeal shockwave treatment as a novel tool to influence SC phenotype. SCs were isolated from motor, sensory and mixed rat nerves and in vitro differences between them were assessed concerning initial cell number, proliferation rate, neurite outgrowth as well as ability to express myelin. We verified the inferior capacity of sensory SCs to promote neurite outgrowth and express myelin‐associated proteins. Motor Schwann cells demonstrated low proliferation rates, but strongly reacted to pro‐myelination stimuli. It is noteworthy for pre‐clinical research that sciatic SCs are a strongly mixed culture, not representing one or the other. Extracorporeal shockwave treatment (ESWT), induced in motor SCs an increased proliferation profile, while sensory SCs gained the ability to promote neurite outgrowth and express myelin‐associated markers. We demonstrate a strong phenotype commitment of sciatic, motor, and sensory SCs in vitro, proposing the experimental use of SCs from pure cultures to better mimic clinical situations. Furthermore we provide arguments for using ESWT on autografts to improve the regenerative capacity of sensory SCs.     

Insulin and the insulin-like growth factors I and II are mitogenic to cultured rat sciatic nerve segments and stimulate [3H]thymidine incorporation through their respective receptors

The factors that control proliferation of Schwann cells during peripheral nerve regeneration are not yet known. In this study we investigated the effects of insulin, insulin-like growth factor I and II (IGF-I and IGF-II), IGF-I analogues, and factors that interfere with their respective receptors, on [³H]thymidine incorporation into cultured nerve segments from the rat sciatic nerve. Segments cultured in nM (0.1–1.7 nM) concentrations of insulin, truncated IGF-I (tIGF-I), long R³IGF-I, or IGF-II exhibited an increase in [³H]thymidine incorporation compared with control segments. IGF-II was most potent. JB1, an IGF-I antagonist, counteracted the effects of tIGF-I and insulin. The results suggest that non-neuronal cells in the nerve segment, probably Schwann cells, possess distinct receptors for insulin, IGF-I, and IGF-II and that these receptors may be involved in the control of Schwann cell proliferation during peripheral nerve regeneration. © 1996 Wiley-Liss, Inc.     

MMP‐9 controls Schwann cell proliferation and phenotypic remodeling via IGF‐1 and ErbB receptor‐mediated activation of MEK/ERK pathway

Phenotypic remodeling of Schwann cells is required to ensure successful regeneration of damaged peripheral axons. After nerve damage, Schwann cells produce an over 100‐fold increase in metalloproteinase‐9 (MMP‐9), and therapy with an MMP inhibitor increases the number of resident (but not infiltrating) cells in injured nerve. Here, we demonstrate that MMP‐9 regulates proliferation and trophic signaling of Schwann cells. Using in vivo BrdU incorporation studies of axotomized sciatic nerves of MMP‐9?/? mice, we found increased Schwann cell mitosis in regenerating (proximal) stump relative to wild‐type mice. Treatment of cultured primary Schwann cells with recombinant MMP‐9 suppressed their growth, mitogenic activity, and produced a dose‐dependent, biphasic, and selective activation of ERK1/2, but not JNK and p38 MAPK. MMP‐9 induced ERK1/2 signaling in both undifferentiated and differentiated (using dbcAMP) Schwann cells. Using inhibitors to MEK and trophic tyrosine kinase receptors, we established that MMP‐9 regulates Ras/Raf/MEK—ERK pathways through IGF‐1, ErbB, and PDGF receptors. We also report on the early changes of MMP‐9 mRNA expression (within 24 h) after axotomy. These studies establish that MMP‐9 controls critical trophic signal transduction pathways and phenotypic remodeling of Schwann cells. © 2009 Wiley‐Liss, Inc.     

Electrical stimulation induces calcium‐dependent release of NGF from cultured Schwann cells

Production of nerve growth factor (NGF) from Schwann cells (SCs) progressively declines in the distal stump, if axonal regeneration is staggered across the suture site after peripheral nerve injuries. This may be an important factor limiting the outcome of nerve injury repair. Thus far, extensive efforts are devoted to modulating NGF production in cultured SCs, but little has been achieved. In the present in vitro study, electrical stimulation (ES) was attempted to stimulate cultured SCs to release NGF. Our data showed that ES was capable of enhancing NGF release from cultured SCs. An electrical field (1 Hz, 5 V/cm) caused a 4.1‐fold increase in NGF release from cultured SCs. The ES‐induced NGF release is calcium dependent. Depletion of extracellular or/and intracellular calcium partially/ completely abolished the ES‐induced NGF release. Further pharmacological interventions showed that ES induces calcium influx through T‐type voltage‐gated calcium channels and mobilizes calcium from 1, 4, 5‐trisphosphate‐sensitive stores and caffeine/ryanodine‐sensitive stores, both of which contributed to the enhanced NGF release induced by ES. In addition, a calcium‐triggered exocytosis mechanism was involved in the ES‐induced NGF release from cultured SCs. These findings show the feasibility of using ES in stimulating SCs to release NGF, which holds great potential in promoting nerve regeneration by enhancing survival and outgrowth of damaged nerves, and is of great significance in nerve injury repair and neuronal tissue engineering. © 2009 Wiley‐Liss, Inc.     

大鼠蛛网膜下腔出血后细胞外调节蛋白激酶1/2激活介导海马区神经细胞自噬

目的探讨大鼠蛛网膜下腔出血(subarachnoid hemorrhage,SAH)后海马区细胞外调节蛋白激酶1/2(extracellular regulated protein kinases,ERK1/2)激活与神经细胞自噬的关系。方法 120只雄性SD大鼠随机分成假手术组、SAH组、ERK1/2抑制剂U0126组、自噬诱导剂雷帕霉素(rapamycin,Rap)组。采用枕大池二次注血法制作SAH大鼠模型;U0126组和Rap组分别于造模前30min侧脑室注射U0126(5μg/μL)和Rap(10nmol/μL)。光镜观察海马区神经细胞形态结构;免疫组化法和实时荧光定量PCR法检测海马区磷酸化ERK1/2(p-ERK1/2)、ERK1/2 mRNA和自噬标志蛋白(Beclin-1和Beclin-1mRNA、LC3-Ⅱ和LC3mRNA)表达水平。结果与假手术组比较,SAH组神经细胞死亡率增加(14.9%±5.7%,28.3%±9.8%,44.2%±10.9%)(q值依次为27.56、35.65、44.81;均P0.05),ERK1/2 mRNA、Beclin-1 mRNA和LC3 mRNA水平增加(1.83±0.01,2.82±0.06,1.34±0.04;1.46±0.02,1.76±0.02,1.35±0.02;1.52±0.04,1.89±0.01,1.31±0.04)(q值依次为42.99、60.66、48.08,71.26、72.46、49.50,48.49、82.40、41.18;均P0.05),p-ERK1/2、Beclin-1和LC3–Ⅱ蛋白水平增加(均P0.05);与SAH组比较,U0126组神经细胞死亡率增加(19.6±6.5%,36.2±7.7%,58.2±12.7%)(q值依次为9.59、10.43、14.66;均P0.05),U0126组ERK1/2 mRNA、Beclin-1 mRNA和LC3 mRNA表达降低(1.23±0.02,1.40±0.02,1.12±0.02;1.22±0.04,1.48±0.06,1.24±0.03;1.34±0.04,1.33±0.02,1.14±0.04)(q值依次为75.66、65.35、31.11,37.18、26.70、15.56,16.79、51.85、22.58;P0.05),p-ERK1/2、Beclin-1和LC3–Ⅱ蛋白水平降低(P0.05);与SAH组比较,Rap组神经细胞死亡率降低(9.1%±4.6%,18.8%±8.6%,28.21%±9.2%)(q值依次为11.86、12.54、16.74;均P0.05),Rap组Beclin-1mRNA和LC3mRNA增加(1.78±0.02,2.27±0.05,1.86±0.04;1.97±0.06,2.31±0.08,1.85±0.00)(q值依次为49.57、48.63、72.12、41.96、38.88、71.73;P0.05),Beclin-1和LC3-Ⅱ蛋白增加(P0.05),ERK1/2 mRNA变化差异无统计学意义(q值依次为2.63、2.65、2.83,P0.05),p-ERK1/2蛋白变化差异无统计学意义(P0.05)。结论 SAH后激活的ERK1/2激活,可促进Beclin-1和LC3-Ⅱ表达介导神经细胞丢失。     

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 and ERK1/2 pathways. Further studies revealed 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.     

Time-dependent effects of insulin on Schwann cell proliferation in the in vitro regenerating adult frog sciatic nerve

The present study showed that insulin (0.01 μg/ml, ≈? 2 nM) inhibited [³H]-thymidine incorporation in support cells, most likely Schwann cells, of the cultured frog sciatic nerve. A 25–35% inhibition took place in regenerating nerve preparations as well as in preparations devoid of neuronal protein synthesis, i.e., in isolated 5 mm nerve segments and in gangliectomized nerves, suggesting that the effect was direct and not mediated via the neuronal cells. The inhibition by insulin was time-dependent in that an effect was seen after 4 days but not at shorter or at longer periods of culturing. In separate experiments biotinylated insulin was shown to be taken up by Schwann cells in the regenerating nerve. Addition of serum increased the [³H]-thymidine incorporation severalfold and abolished the inhibitory action of insulin. Our results suggest that insulin, at a certain stage of the regeneration programme, exerts a direct, inhibitory effect on the proliferation of the Schwann cells in the cultured frog sciatic nerve. © 1993 Wiley-Liss, Inc.     

Increase of MCP-1 (CCL2) in myelin mutant Schwann cells is mediated by MEK-ERK signaling pathway

Macrophages are critically involved in the pathogenesis of genetically caused demyelination, as it occurs in inherited demyelinating neuropathies. On the basis of the observation that upregulation of the Schwann cell-derived chemokine MCP-1 (CCL2) is a pathologically relevant mechanism for macrophage activation in mice heterozygously deficient for the myelin component P0 (P0+/-), we posed the question of the intracellular signaling cascade involved. By using western blot analysis of peripheral nerve lysates the MAP-kinases extracellular signal-regulated kinase 1/2 (ERK1/2) and MAP kinase/ERK kinase 1/2 (MEK1/2) showed an early and constantly increasing activation in P0 mutants. Furthermore, in nerve fibers from the P0+/- mutants, Schwann cell nuclei were much more often positive for phosphorylated ERK1/2 than in nerve fibers from wild type mice. In vitro experiments using the MEK1/2-inhibitor CI-1040 decreased ERK1/2-phosphorylation and MCP-1 expression in a Schwann cell-derived cell line. Finally, systemic application of CI-1040 lead to a decreased ERK1/2-phosphorylation and substantially reduced MCP-1-production in peripheral nerves of P0+/- mutant mice. Our study identifies MEK1/2-ERK1/2 signaling as an important intracellular pathway that connects the Schwann cell mutation with the activation of pathogenetically relevant macrophages in the peripheral nerves. These findings may have important implications for the treatment of inherited peripheral neuropathies in humans.     

Pre-degenerated peripheral nerves co-cultured with bone marrow-derived cells：a new technique for harvesting high-purity Schwann cells

Schwann cells play an important role in the peripheral nervous system, especially in nerve repair following injury, so artificial nerve regeneration requires an effective technique for obtaining purified Schwann cells. In vivo and in vitro pre-degeneration of peripheral nerves have been shown to obtain high-purity Schwann cells. We believed that in vitro pre-degeneration was simple and controllable, and available for the clinic. Thus, we co-cultured the crushed sciatic nerves with bone marrow-derived cells in vitro. Results demonstrated that, 3 hours after injury, a large number of mononuclear cells moved to the crushed nerves and a large number of bone marrow-derived cells infiltrated the nerve segments. These changes promoted the degradation of the nerve segments, and the dedifferentiation and proliferation of Schwann cells. Neural cell adhesion molecule and glial fibrillary acidic protein expression were detected in the crushed nerves. Schwann cell yield was 9.08 ± 2.01 × 104/mg. The purity of primary cultured Schwann cells was 88.4 ± 5.79%. These indicate a successful new method for obtaining Schwann cells of high purity and yield from adult crushed sciatic nerve using bone marrow-derived cells.     

Role of intracellular Ca2+ and calmodulin/MAP kinase kinase/extracellular signal-regulated protein kinase signalling pathway in the mitogenic and antimitogenic effect of nitric oxide in glia- and neurone-derived cell lines

To elucidate the mechanism of cell growth regulation by nitric oxide (NO) and the role played in it by Ca2+, we studied the relationship among intracellular Ca2+ concentration ([Ca2+]i), mitogen-activated protein kinases [extracellular signal-regulated protein kinase (ERK)] and proliferation in cell lines exposed to different levels of NO. Data showed that NO released by low [(z)-1-[2-aminiethyl]-N-[2-ammonioethyl]amino]diazen-1-ium-1,2diolate (DETA/NO) concentrations (10 microm) determined a gradual, moderate elevation in [Ca2+]i (46.8 +/- 7.2% over controls) which paralleled activation of ERK and potentiation of cell division. Functionally blocking Ca2+ or inhibiting calmodulin or MAP kinase kinase activities prevented ERK activation and antagonized the mitogenic effect of NO. Experimental conditions favouring Ca2+ entry into cells led to increased [Ca2+]i (189.5 +/- 4.8%), ERK activation and cell division. NO potentiated the Ca2+ elevation (358 +/- 16.8%) and ERK activation leading to expression of p21Cip1 and inhibition of cell proliferation. Furthermore, functionally blocking Ca2+ down-regulated ERK activation and reversed the antiproliferative effect of NO. Both the mitogenic and antimitogenic responses induced by NO were mimicked by a cGMP analogue whereas they were completely antagonized by selective cGMP inhibitors. These results demonstrate for the first time that regulation of cell proliferation by low NO levels is cGMP dependent and occurs via the Ca2+/calmodulin/MAP kinase kinase/ERK pathway. In this effect the amplitude of Ca2+ signalling determines the specificity of the proliferative response to NO possibly by modulating the strength of ERK activation. In contrast to the low level, the high levels (50-300 microm) of DETA/NO negatively regulated cell proliferation via a Ca2+-independent mechanism.     

Differential gene expression in motor and sensory Schwann cells in the rat femoral nerve

Phenotypic differences in Schwann cells (SCs) may help to guide axonal regeneration down motor or sensory specific pathways following peripheral nerve injury. The goal of this study was to identify phenotypic markers for SCs harvested from the cutaneous (sensory) and quadriceps (motor) branches of the rat femoral nerve and to study the effects of expansion culture on the expression patterns of these motor or sensory phenotypic markers. RNA was extracted from SCs harvested from the motor and sensory branches of the rat femoral nerve and analyzed using Affymetrix Gene Chips (Rat Genome 230 v2.0 Array A). Genes that were upregulated in motor SCs compared with the sensory SCs or vice versa were identified, and the results were verified for a subset of genes using quantitative real-time polymerase chain reaction (qRT-PCR). The expression levels of the "phenotype-specific" genes were then evaluated in SC expansion cultures at various time points over 30 days by qRT-PCR to determine the effect of expansion on SC phenotype. Expression levels of the phenotype-specific genes were significantly altered after expansion culture for both the motor and the sensory markers compared with fresh nerve tissue. These results indicate that both motor and sensory SC gene expression patterns are disrupted during expansion in vitro and may affect the ability of SCs to express phenotype-specific genes after transplantation.     

Fibroblast‐derived tenascin‐C promotes Schwann cell migration through β1‐integrin dependent pathway during peripheral nerve regeneration

Peripheral nerve regeneration requires precise coordination and dynamic interaction among various types of cells in the tissue. It remains unclear, however, whether the cellular crosstalk between fibroblasts and Schwann cells (SCs) is related to phenotype modulation of SCs, a critical cellular process after peripheral nerve injury. In this study, microarray analysis revealed that a total of 6,046 genes were differentially expressed in the proximal nerve segment after sciatic nerve transection in rats, and bioinformatics analysis further identified tenascin‐C (TNC), an extracellular matrix (ECM) protein, as a key gene regulator. TNC was abundantly produced by nerve fibroblasts accumulating at the lesion site, rather than by SCs as usually expected. TNC significantly promoted SC migration without effects on SC proliferation in primary culture. In co‐culture of fibroblasts and SCs, inhibition of TNC expression either by siRNA transfection or antibody blockade could suppress SC migration, while TNC‐stimulated SC migration was mediated by TNC binding to β1‐integrin receptor in SCs through activation of Rac1 effectors. The in vivo evidence showed that exogenous TNC protein enhanced SC migration and axonal regrowth. Our results highlight that TNC‐mediated cellular interaction between fibroblasts and SCs may regulate SC migration through β1‐integrin‐dependent pathway during peripheral nerve regeneration. GLIA 2016;64:374–385     

Electrospun silk fibroin nanofibers promote Schwann cell adhesion, growth and proliferation

In this study, Schwann cells, at a density of 1 × 105 cells/well, were cultured on regenerated silk fibroin nanofibers (305 ± 84 nm) prepared using the electrospinning method. Schwann cells cultured on the silk fibroin nanofibers appeared more ordered, their processes extended further, and they formed more extensive and complex interconnections. In addition, the silk fibroin nanofibers had no impact on the proliferation of Schwann cells or on the secretion of ciliary neurotrophic factor, brain-derived neurotrophic factor or nerve growth factor. These findings indicate that regenerated electrospun silk fibroin nanofibers can promote Schwann cell adhesion, growth and proliferation, and have excellent biocompatibility.     

Nerve guides seeded with autologous schwann cells improve nerve regeneration

This study evaluates the ability of Schwann cells (SCs) transplanted into a nerve guide to improve regeneration and reinnervation after sciatic nerve resection and repair, leaving a 6-mm gap, in the mouse. SCs were isolated from predegenerated adult sciatic nerves and expanded in culture using a chemically defined medium. Syngeneic, isogeneic, and autologous SCs were suspended in Matrigel and seeded in resorbable, permeable poly(l-lactide-co-epsilon-caprolactone) guides at 150,000 cells/tube. Guides containing SCs were compared to guides filled with Matrigel alone and with peroneal nerve autografts. Functional reinnervation was assessed by noninvasive methods to determine recovery of sweating, nociceptive, sensory, and motor functions in the hindpaw during 4 months postoperation. Morphological analysis of the regenerated nerves was performed at the end of follow-up. The group with an autograft achieved faster and higher levels of reinnervation and higher number of regenerated myelinated fibers than groups repaired by tubulization. The immunogenicity of transplanted SCs influenced the outcome of nerve regeneration. Transplants of autologous SCs resulted in slightly lower levels of reinnervation than autografts, but higher recovery and number of regenerated fibers reaching the distal nerve than transplants of isologous and syngeneic SCs, although most of the differences were not statistically significant. Syngeneic SCs did not improve regeneration with respect to acellular guides. Prelabeled transplanted SCs were found to survive into the guide 1-3 months after implantation, to a larger number when they were autologous than syngeneic. Cellular prostheses composed of a resorbable guide seeded with autologous SCs appear as an alternative for repairing long gaps in injured nerves, approaching the success of autografts.     

Injury-induced activation of ERK 1/2 in the sciatic nerve of healthy and diabetic rats

Phosphorylation of extracellular-signal-regulated kinase 1/2 (p-ERK 1/2) was investigated by immunohistochemistry at 30 min, 1 h, and 48 h after nerve transection in the sciatic nerve of healthy and diabetic [streptozotocin (STZ)-induced diabetes mellitus and BioBreeding (BB; i.e. DR.lyp/lyp or BBDP)] rats. Transection injury increased the intensity of p-ERK 1/2 in nerve stumps at all time points. Staining was confined to Schwann cells with occasional faint staining in single axons. In diabetic rats, a lower intensity of p-ERK 1/2 was found at 1 and 48 h in the distal and proximal nerve stumps compared with healthy rats. STZ-induced diabetic rats were not different from BB rats. p-ERK 1/2 is activated differentially in Schwann cells after nerve injury in diabetic rats, whereas activation in STZ-induced diabetic rats did not differ from BB rats.     

Lithium promotes proliferation and suppresses migration of Schwann cells

Schwann cell proliferation,migration and remyelination of regenerating axons contribute to regeneration after peripheral nervous system injury.Lithium promotes remyelination by Schwann cells and improves peripheral nerve regeneration.However,whether lithium modulates other phenotypes of Schwann cells,especially their proliferation and migration remains elusive.In the current study,primary Schwann cells from rat sciatic nerve stumps were cultured and exposed to 0,5,10,15,or 30 mM lithium chloride(LiCl)for 24 hours.The effects of LiCl on Schwann cell proliferation and migration were examined using the Cell Counting Kit-8,5-ethynyl-2′-deoxyuridine,Transwell and wound healing assays.Cell Counting Kit-8 and 5-ethynyl-2′-deoxyuridine assays showed that 5,10,15,and 30 mM LiCl significantly increased the viability and proliferation rate of Schwann cells.Transwell-based migration assays and wound healing assays showed that 10,15,and 30 mM LiCl suppressed the migratory ability of Schwann cells.Furthermore,the effects of LiCl on the proliferation and migration phenotypes of Schwann cells were mostly dose-dependent.These data indicate that lithium treatment significantly promotes the proliferation and inhibits the migratory ability of Schwann cells.This conclusion will inform strategies to promote the repair and regeneration of peripheral nerves.All of the animal experiments in this study were ethically approved by the Administration Committee of Experimental Animal Center of Nantong University,China(approval No.20170320-017)on March 2,2017.     

In vitro biocompatibility of three chitosan/polycation composite materials for nerve regeneration

BACKGROUND:It has been reported that chitosan nerve conduits could support axon elongation and improve relevant function during in vivo nerve regeneration. OBJECTIVE: To investigate in vitro biocompatibility of three novel, chitosan/polycation composite materials for nerve regeneration in cultured mouse Schwann cells and PC12 cells. DESIGN, TIME AND SETTING: The observational, control experiments for nerve tissue engineering were performed at the Department of Biological Sciences and Biotechnology of Tsingh...     

Cyclic AMP synergistically enhances neuregulin-dependent ERK and Akt activation and cell cycle progression in Schwann cells

The elevation of intracellular cAMP synergistically enhances the neuregulin-dependent proliferation of cultured Schwann cells (SCs); however, the mechanism by which this occurs has not been completely defined. To better understand this mechanism, we investigated the effect of cAMP on the activation of the extracellular signal-regulated kinase (ERK) and phosphoinositide 3-kinase (PI3-K)-Akt (PKB) pathways by heregulin, a member of the neuregulin family. Using primary cultures of adult SCs, we demonstrated that the adenylyl cyclase activator, forskolin, enhanced heregulin-dependent SC proliferation by reducing the time required for S-phase entry. When cAMP levels were increased, using either forskolin or a cell permeable analogue of cAMP, the heregulin-induced phosphorylation of ERK was converted from transient to sustained and the heregulin-induced phosphorylation of Akt was synergistically increased. Consistent with these observations, studies in which inhibitors of MEK, the upstream stimulating ERK kinase, and PI3-K were administered at different times following the onset of stimulation indicated that sustained high levels of both MEK/ERK and PI3-K/Akt activity before S-phase initiation were essential for S-phase entry. Overall, these novel results indicate that in neuregulin-stimulated SCs the activation of cAMP-mediated pathways accelerates G1-S progression by prolonging ERK activation and concurrently enhancing Akt activation.