Shaofeng Pu

The effect of intrathecal administration of glial activation inhibitors on dorsal horn BDNF overexpression and hind paw mechanical allodynia in spinal nerve ligated rats.

Recent studies have suggested that activated glia in the spinal cord may play a vital role at different times during spinal nerve ligation (SNL)-induced neuropathic pain; therefore, glial activation inhibitors have been used as effective painkillers. Brain-derived neurotrophic factor (BDNF) is also known to be a powerful pain modulator, but it remains unclear how it contributes to the glial activation inhibitor-based treatment. This study revealed the following results: (1) intrathecal administration of minocycline (a microglial activation inhibitor) could prevent mechanical allodynia during the initiation of SNL-induced neuropathic pain, and its action was associated with the elimination of BDNF overexpression in the dorsal horn; (2) the spinal injection of fluorocitrate (an astrocytic activation inhibitor) but not minocycline could reverse mechanical allodynia during the maintenance phase of SNL-induced pain, and its action was also related to a decrease in BDNF overexpression in the dorsal horn; and (3) treatment with TrkB/Fc (a BDNF-sequestering protein) had a similar effect during both the early development and maintenance periods. These results led to the following conclusions: (1) elevated BDNF expression in the dorsal horn was required to develop and maintain neuropathic pain; (2) minocycline could only prevent mechanical allodynia in the early stages, possibly by inhibiting BDNF release from microglia; and (3) fluorocitrate could reverse existing mechanical allodynia, and its action was associated with the inhibition of BDNF upregulation induced by astrocytic activation.

Brain-derived neurotrophic factor–activated astrocytes produce mechanical allodynia in neuropathic pain

Neuropathic pain management is challenging for physicians and a vexing problem for basic researchers. Recent studies reveal that activated spinal astrocytes may play a vital role in nerve injury-induced neuropathic pain, although the mechanisms are not fully understood. We have found increased glial fibrillary acidic protein (GFAP) expression, a hallmark of reactive gliosis, and elevated brain-derived neurotrophic factor (BDNF) expression in the dorsal horn in a rat model of allodynia induced by spinal nerve ligation (SNL). The high GFAP expression and mechanical allodynia that SNL induces were prevented by the intrathecal injection of the BDNF-sequestering fusion protein TrkB/Fc. Additionally, mechanical allodynia and GFAP overexpression was induced by the spinal administration of exogenous BDNF to naive rats, and exogenous BDNF given together with fluorocitrate, an astrocytic metabolism inhibitor, inhibited allodynia and GFAP upregulation. Exogenous BDNF also activated the astrocytes directly when tested in vitro. Furthermore, intrathecal administration of BDNF-stimulated astrocytes also induced mechanical allodynia in naive rats. All of these results indicate that astrocytes activated by BDNF might contribute to mechanical allodynia development in neuropathic pain in rats.

Positive feedback loop of autocrine BDNF from microglia causes prolonged microglia activation.

Background/Aims: Microglia, which represent the immune cells of the central nervous system (CNS), have long been a subject of study in CNS disease research. Substantial evidence indicates that microglial activation functions as a strong neuro-inflammatory response in neuropathic pain, promoting the release of pro-inflammatory cytokines, such as tumor necrosis factor (TNF)-α. In addition, activated microglia release brain-derived neurotrophic factor (BDNF), which acts as a powerful cytokine. In this study, we performed a series of in vitro experiments to examine whether a positive autocrine feedback loop existed between microglia-derived BDNF and subsequent microglial activation as well as the mechanisms underlying this positive feedback loop. Methods: Because ATP is a classic inducer of microglial activation, firstly, we examined ATP-activated microglia in the present study. Secondly, we used TrkB/Fc, the BDNF sequester, to eliminate the effects of endogenous BDNF. ATP-stimulated microglia without BDNF was examined. Finally, we used exogenous BDNF to further determine whether BDNF could directly activate BV2 microglia. In all experiments, to quantify BV2 microglia activation, the protein levels of CD11b, a microglial activation marker, were measured by western blot. A Transwell migration assay was used to examine microglial migration. To assess the synthesis and release of proinflammatory cytokines, western blot was used to measure BDNF synthesis, and ELISA was used to quantify TNF-α release. Results: In our present research, we have observed that ATP dramatically activates microglia, enhancing microglial migration, increasing the synthesis of BDNF and up-regulating the release of TNF-α. Microglial activation is inhibited following the sequestration of endogenous BDNF, resulting in impaired microglial migration and decreased TNF-α release. Furthermore, exogenous BDNF can also activate microglia to subsequently enhance migration and increase TNF-α release. Conclusion: Therefore, we suggest that microglial activation increases the synthesis of BDNF and that the release of BDNF can, in turn, activate microglia. A positive autocrine BDNF feedback loop from microglia may contribute to prolonged microglial activation.

p38/MAPK inhibitor modulates the expression of dorsal horn GABA(B) receptors in the spinal nerve ligation model of neuropathic pain.

Background: Neuropathic pain is one of the most challenging clinical problems due to a lack of understanding the mechanisms. Recent studies have suggested that activated microglia in spinal cord may play a vital role in nerve injury-induced neuropathic pain, but the exact mechanisms have not been fully determined. Methods: First, we investigated the changes of dorsal horn GABA(B) receptor 1 (R1) expression in spinal nerve ligation rats. Second, we explored whether activated microglia contributed to such neuron changes by intrathecal administration of the p38 inhibitor, SB203580. Results: In this study, we found a dynamic change of GABA(B)R1a protein expression after spinal nerve ligation, and the peripheral nerve injury-induced downregulation of GABA(B)R1a expression in the spinal dorsal horn could be prevented by intrathecal administration of a p38/MAPK inhibitor SB203580. Conclusions: Our results provide valuable information for a better understanding of neuropathic pain and may contribute to developing effective treatments in future studies.

Circulating microRNA expression profile: a novel potential predictor for chronic nervous lesions

The mechanisms of chronic neuropathic pain are not clear. Serum microRNAs (miRNAs) might show a special feature for chronic nervous lesions. However, little is known about the changes in circulating miRNAs for the neuropathic pain. Therefore, changes in the circulating miRNAs expression profile for the neuropathic pain were investigated. Serum was collected from rats before and after spinal nerve ligation (SNL) surgery, and a microarray analysis was performed to determine the changes in miRNA expression profile. The expression of inflammatory cytokines in serum from the same individuals, including interleukin-6 (IL-6), tumor necrosis factor-α (TNF-α), and monocyte chemotactic protein-1 (MCP-1), was also measured. The results showed that the expression levels of IL-6, TNF-α, and MCP-1 were significantly elevated in SNL rats which were significantly correlated with pain levels. Nine miRNAs with significantly different expression levels before and after SNL surgery were identified by microarray analysis, which were further validated by quantitative real-time polymerase chain reaction analyses. Compared with naive rats without SNL surgery, the expression of five miRNAs (hsa-miR-221, hsa-miR-34c, hsa-miR-21, hsa-miR-30a-5p, and hsa-miR-206) in the serum of rats after SNL surgery was decreased and four miRNAs (hsa-miR-31-5p, hsa-miR-133b, hsa-miR-22, and hsa-miRPlus-A1087) were increased, suggesting that miRNA changes may involve in the regulation of neuropathic pain. TargetScan was used to predict mRNA targets for these miRNAs, and the results showed that the transcripts with multiple predicted target sites belonged to neurologically important pathways. Bioinformatics analysis revealed that several target genes are related to the activation of cell signaling associated with nervous lesions. In this study, the changes to miRNA profiles in serum under neuropathic pain conditions were shown for the first time, suggesting that circulating miRNAs profile in serum is a potential predictor for neuropathic pain.