Elsevier

Neuroscience Research

Volume 70, Issue 4, August 2011, Pages 383-390
Neuroscience Research

The mood stabilizer valproic acid induces proliferation and myelination of rat Schwann cells

https://doi.org/10.1016/j.neures.2011.04.002Get rights and content

Abstract

Schwann cells (SCs) within peripheral nerve respond robustly after exposure to neurotrophic factors. Recent results have revealed that valproic acid (VPA), at a clinically relevant therapeutic concentration, produces effects similar to neurotrophic factors, and promotes neurite growth and cell survival. We hypothesized that VPA could also induce Schwann cell response. In this study, we sought to determine how pure Schwann cells responded to VPA by evaluating for proliferation, expression of S-100, growth cone-associated protein 43 (GAP-43), myelin-associated glycoprotein (MAG), and myelin basic protein (MBP). Immunohistochemistry demonstrated that the Schwann cells were positive for S-100, GAP-43, MAG, and MBP greater than 99% of the experimental cells. The rate of proliferation was increased in experimental cells from MTT assay and Bromodeoxyuridine/DAPI double staining. Furthermore, Western blot showed an up-regulation in GAP-43, MAG and MBP protein expression in experimental cells, respectively. We also found that mitogen-activated protein kinase (MAPK)/extracellular signal-regulated kinase (ERK) 1/2 pathway was involved in the enhanced cell proliferation of Schwann cells evoked by VPA. This study provides novel information regarding Schwann cell response to VPA, which might help the understanding of VPA-based treatment for peripheral nerve injury.

Highlights

► This is an original research about the neuroprotective effects by valproic acid. ► It reveals how Schwann cells response to valproic acid in vitro. ► MAPK/ERK1/2 is required for VPA-induced proliferation of Schwann cells.

Introduction

The mood-stabilizing agent, valproic acid (VPA), is widely prescribed in the treatment of manic-depressive disorders (Bowden, 1995). Recent evidence also indicated that locally or systemically applied VPA can enhance neurite outgrowth and recovery of motor function in adult rats (Cui et al., 2003, Wu et al., 2008). However, the precise mechanisms by which VPA brings about these effects are currently unknown.

It is widely accepted that Schwann cells (SCs) play a crucial role in neurite growth (Mirsky et al., 2002, Bhatheja and Field, 2006). SCs play a dual role: they serve as scaffolds of regenerating axons by expressing adhesion molecules on the surface plasma membrane, and produce various trophic factors for regenerating axons (Mirsky and Jessen, 1996, D’Antonio et al., 2006, Santos et al., 2000). VPA has been previously shown to protect cultured rat hippocampal neurons against amyloid and glutamate neurotoxicity and can protect cultured cerebellar granule cells against low-K+-induced apoptosis by acting on the phosphatidylinositol 3-kinase pathway (Plant et al., 2009, Fatemi et al., 2009). Yuan et al. (2001) have further reported that VPA, at a clinically relevant therapeutic concentration, increases the expression of the cytoprotective protein growth cone-associated protein 43 (GAP-43) and bcl-2 in the central nervous system (CNS) and human neuroblastoma SH-SY5Y cells, namely, the activation of the extracellular signal regulated kinase (ERK) pathway and promotion of neurite growth and cell survival. Although this neuroprotective effects exerted by VPA on cultured cells in CNS are well characterized, there remains limited information as to how SCs would directly respond to VPA.

Damage to a peripheral nerve may produce an axonal injury which may result in Wallerian degeneration in the site of injury, and subsequently triggers a cascade of glial cell responses including marked SCs proliferation and myelination. Both myelin-associated glycoprotein (MAG) and myelin basic protein (MBP) are key myelin specific protein constituents in the peripheral nervous system (PNS) (Ryu et al., 2008). MAG is known as a transmembrane protein of the immunoglobulin superfamily, and has been recognized for its bifunctionality whereby it plays a key role in the early phases of myelination as well as in the maintenance of stable axon–myelin interactions (Ryu et al., 2008, Filbin, 1995, Shim and Ming, 2009). Furthermore, previous data support the central importance of MAG as a promyelinogenic marker in SCs. MBP has been shown to be vital to the myelination process. Existing in at least six functionally similar isoforms, MBP is essential for proper formation of myelin thickness and compactness in the CNS and PNS (Ryu et al., 2008, Zhang et al., 2007). In addition, this pro-myelinating protein is also an important component of the major dense line (MDL) in the myelin sheath (Zhang et al., 2007).

In order to help the understanding of VPA treatment for peripheral nerve injury, the present study focus on investigating the influences of VPA on primary cultures of rat SCs, by a set of immunohistochemical, biochemical and molecular biological assessments. We hope to provide further evidence about the neuroprotective functions of VPA on peripheral nerve regeneration.

Section snippets

Subjects

The subjects were SCs obtained from 3-day-old Sprague–Dawley rats (SPF grade, license No. 2009–2010), provided by Experimental Animal Center at the Medical School of Wuhan University. VPA was purchased from the Sigma–Aldrich Company. All other chemicals used in the present experiment were of analytical reagent grade or better, and were obtained from the usual commercial sources.

All procedures were carried out in accordance with NIH Guidelines for the Care and Use of Laboratory Animals, and were

VPA treatment increases the cellular viability of SCs

To determine whether VPA at therapeutic concentration influenced the cell viability of SCs, we treated SCs with different medium for 12, 24, 36, 48 and 72 h. MTT assay indicated that the cellular viability of SCs treated with VPA was significantly increased compared to that treated with plain medium for 24, 36 and 48 h culture (Fig. 1).

VPA treatment enhances the cell proliferation of SCs

Control SCs stained positive for the panspecific SCs marker S-100 in greater than 99% of the evaluated cells. SCs subjected to VPA at therapeutic concentration for

Discussion

Recently, Cui et al. (2003) reported that VPA could facilitate neurite regeneration and rehabilitation of motor function, based on the research that rats whose sciatic nerves transected were systemically administered VPA orally. We have furtherly developed an in vivo animal model for peripheral nerve injury where a silicon tube is bridged a rat sciatic nerve deficit (Wu et al., 2008). Data from this model indicated that locally applied VPA promotes sciatic nerve regeneration in the defect nerve.

Acknowledgements

The authors gratefully acknowledge Fang Yu and Kai Guo for expert technical assistance and Qingsong Zhang for help in editing the manuscript.

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