Molecular pharmacological study on the roles of chemokines in neuropathic pain

单核细胞趋化蛋白-1（MCP-1，CCL 2）是一种明确的CC趋化因子，与各种类型的脑损伤（如缺血性和创伤性损伤）的病理学有关。以前，我们证明了MCP-1的生产上调，在背根神经节（DRG）的神经病理性疼痛模型大鼠。本研究采用MCP-1和转录激活因子3（activating transcription factor-3，ATF 3）双重免疫荧光染色技术，观察大鼠背根神经节（DRG）神经元在结扎后是否产生MCP-1。在较小的神经元（胞体面积<600 μm^2）中，MCP-1免疫反应（ir）不仅在损伤的神经元中观察到，而且在未损伤的神经元中也观察到。另一方面，在大尺寸神经元（>1200μm^2）中，几乎所有MCP-1免疫阳性神经元均为ATF 3免疫阳性。本实验进一步研究了腺苷5 '-O-（3-thiotriphosphate）（ATPγS）对大鼠皮层纹状体脑片培养的DRG小神经元MCP-1表达的影响。ATPγS可诱导星形胶质细胞MCP-1 mRNA表达和蛋白生成。通过使用几种MAP激酶抑制剂来检查MAP激酶在这种诱导中的参与。MEK抑制剂PD 98059和U 0126显著抑制ATPγ S诱导的MCP-1 mRNA表达和蛋白质生成。SP 600125对JNK的抑制可部分抑制它们。而p38 MAP激酶抑制剂SB 203580则能显著增强ATPγ S诱导的MCP-1生成。进一步的研究表明，SB 203580延长了ATPγS诱导的MCP-1 mRNA表达的持续时间，从而增加了MCP-1的净产量。这些结果表明，在星形胶质细胞中，ERK和p38 MAP激酶对ATPγ S诱导的MCP-1产生具有相互调节作用。

Monocyte chemoattractant protein-1(MCP-1,CCL2) is a well-defined CC chemokine implicated in the pathology of various types of brain injuries, such as ischemic and traumatic injuries. Previously, we demonstrated that MCP-1 production was upregulated in the dorsal root ganglia (DRG) of neuropathic pain model rats. In this study, we carried out the double immunofluorescent staining between MCP-1 and activating transcription factor-3(ATF3) to examine whether MCP-1 was produced in the injured or uninjured DRG neurons after the nerve ligation. In the small-sized neurons (cell body area <600 μm^2), MCP-1-immunoreactivity (ir) was observed not only in the injured neurons, but also in the uninjured ones. On the other hand, in the large-sized neurons (>1200μm^2), almost all of the MCP-1-ir neurons were ATF3-ir-positive. This result suggests that intercellular interaction between the injured and uninjured neurons is involved in the MCP-1 upregulation in the small-sized DRG neurons.Furthermore, we investigated the effects of adenosine 5'-O-(3-thiotriphosphate)(ATPγS) on MCP-1 production using the rat cortico-striatal slice culture. ATPγS induced MCP-1 mRNA expression and protein production in astrocytes. The involvement of MAP kinases in this induction was examined by using several kinds of MAP kinase inhibitors. PD98059 and U0126, MEK inhibitors, significantly suppressed ATPγS-induced MCP-1 mRNA expression and protein production. Inhibition of JNK by SP600125 resulted in a partial suppression of them. On the other hand, SB203580, a p38 MAP kinase inhibitor, significantly enhanced ATPγS-induced MCP-1 productfon. Further investigation revealed that SB203580 extended the duration of MCP-1 mRNA expression induced by ATPγS and thereby increased the net production of MCP-1. These results demonstrate the reciprocal regulation of ATPγS-induced MCP-1 production by ERK and p38 MAP kinases in astrocytes.