Evidence that spinal astrocytes but not microglia contribute to the pathogenesis of Paclitaxel-induced painful neuropathy - PubMed (original) (raw)

Evidence that spinal astrocytes but not microglia contribute to the pathogenesis of Paclitaxel-induced painful neuropathy

Haijun Zhang et al. J Pain. 2012 Mar.

Abstract

Paclitaxel often induces persistent painful neuropathy as its most common treatment-limiting side effect. Little is known concerning the underlying mechanisms. Given the prominent role of glial cells in many types of neuropathic pain, we investigated here the morphological and functional changes of spinal astrocytes and microglia in a rat model of paclitaxel-induced neuropathy. Immunohistochemistry, western blotting, and real-time polymerase chain reaction were performed with samples from 109 rats up to 28 days after paclitaxel treatment. Paclitaxel (2 mg/kg, i.p.) induced a rapid and persistent activation of spinal astrocytes assessed using glial fibrillary acidic protein, but not apparent activation of microglia assessed using OX42, Iba-1, and phosphorylated p38. In the context of astocyte activation, there was a significant downregulation of glial glutamate transporters GLAST and GLT-1 in spinal dorsal horn. The activation of spinal astrocytes by paclitaxel was not associated with expression of pro-inflammatory cytokines including tumor necrosis factor-α, interleukin-1β, or interleukin-6 in spinal dorsal horn. Systemic treatment with minocycline (50 mg/kg, i.p.) prevented activation of astrocytes and downregulation of glial glutamate transporters in spinal dorsal horn induced by paclitaxel. These data suggest the involvement of spinal astrocytes but not microglia in the pathogenesis of paclitaxel-induced neuropathy.

Perspective: Spinal astrocytes and/or glial glutamate transporters could be new therapeutic targets for paclitaxel-induced painful neuropathy.

Copyright © 2012 American Pain Society. Published by Elsevier Inc. All rights reserved.

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Conflict of interest statement

DISCLOSURES

All authors declare no conflict of interest.

Figures

Figure 1

Figure 1

Illustration of quantifying immunohistochemcal stainings. A, A spinal dorsal horn was viewed under differential interference contrast (DIC) and the area of spinal lamina II was determined. B, The same slice was viewed under fluorescence without changing the position. C, Background fluorescence was measured first by defining 3 circular regions (1–3) adjacent to immune-positive neurons in spinal lamina II and averaged to obtain a mean background value. Then the mean fluorescent intensity subtracted by background for the whole of spinal lamina II of each section was determined. Scale: 50 um.

Figure 2

Figure 2

Expression of GFAP in spinal cord after paclitaxel or vehicle treatment. (A) Representative spinal cord slices of GFAP staining 4 Hrs (a, b), 7 days (c, d) and 28 days (e, f) after treatment. (B) Quantification of GFAP in spinal lamina II in paclitaxel- and vehicle-treated groups. The number of animals in each group was presented in bar graphs. * p < 0.05, ** p < 0.01 for paclitaxel vs. vehicle, two-way ANOVA with Bonferroni post hoc test. Scale: 500 um.

Figure 3

Figure 3

Expression of OX42 in spinal dorsal horn after L5 spinal nerve ligation (SNL) and paclitaxel treatment. (A) A robust increase in the expression of OX42 was detected in ipsilateral (ipsi) spinal dorsal horn versus contralateral (contra) side 7 days after SNL. Lower panels show the enlargement views of ipsilateral and contralateral dorsal horns separately. (B) Paclitaxel did not induce any increase in the expression of OX42 in spinal dorsal horn. (C) Statistical analysis revealed a significant increase in the expression of OX42 induced by L5-SNL but not paclitaxel. N=4 for each group. * p < 0.01, ipsilateral vs. contralateral, paired _t_-test. Scale: 500 um.

Figure 4

Figure 4

Expression of Iba-1 and p-p38 in spinal dorsal horn. (A) Spinal nerve ligation (SNL) induced a significant increase in the expression of Iba-1 in spinal dorsal horn than sham or naïve animals. N=4 for each group. (B) Paclitaxel (Pac) did not induce any increase in the expression of Iba-1 compared to vehicle (Veh) or naïve groups. N=4 for each group. (C) SNL induced a significant increase in the expression of p-p38 in spinal dorsal horn than sham or naïve animals. N=4 for each group. (D) Paclitaxel did not induce any increase in the expression of p-p38 compared to vehicle or naïve groups. N=4 for each group. * p < 0.05, ** p < 0.01, one-way ANOVA with Student Newman-Keuls post hoc test.

Figure 5

Figure 5

Expression of GLAST and GLT-1 in spinal dorsal horn. (A, B) Paclitaxel (Pac) but not vehicle (Veh) induced a persistent downregulation of GLAST in spinal dorsal horn. (C,D) Paclitaxel but not vehicle induced a rapid downregulation of GLT-1 in spinal dorsal horn. N=4 for each group. * p < 0.05, ** p < 0.01 for paclitaxel vs. vehicle; ^ p < 0.05, ^^ p < 0.01 for paclitaxel vs. naïve, two-way ANOVA with Bonferroni post hoc test. (E) Quantification of pro-inflammatory cytokines in spinal dorsal horn by real-time PCR. Paclitaxel did not induce any increase in the expression of TNFα, IL-1β or IL-6 in spinal dorsal horn compared to vehicle or naïve animals. N=4 for each group.

Figure 6

Figure 6

Systemic treatment of minocycline prevented the increased expression of GFAP and downregulation of GLAST and GLT-1 in spinal dorsal horn induced by paclitaxel. The number of animals in each group was presented in bar graphs. Veh: vehicle; Pac: paclitaxel; Mino: minocycline. * p < 0.01, one-way ANOVA with Student Newman-Keuls post hoc test. Scale: 500 um.

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