Molecular pharmacological study on the roles of chemokines in neuropathic pain

单核细胞趋化蛋白-1(MCP-1，CCL2)是一种定义明确的CC趋化因子，参与脑损伤的病理过程，如脑缺血和创伤。在此之前，我们证明了神经病理性疼痛模型大鼠背根神经节(DRG)中MCP-1的产生上调。在本研究中，我们进行了MCP-1和激活转录因子-3(ATF3)的双重免疫荧光染色，以检测神经结扎后损伤和未损伤的DRG神经元是否产生MCP-1。在胞体面积为600μm^2的小型神经元中，损伤神经元和未损伤神经元均可见单核细胞趋化蛋白-1免疫反应阳性(Ir)。另一方面，在大型神经元(&gt；1200μm^2)中，几乎所有的单核细胞趋化蛋白-1免疫反应阳性神经元都是ATF3阳性的。这一结果提示损伤和未损伤神经元之间的细胞间相互作用参与了小型背根节神经元单核细胞趋化蛋白-1的上调。此外，我们利用大鼠皮质-纹状体脑片培养方法观察了腺苷5‘-O-(3-硫代三磷酸)(γS)对单核细胞趋化蛋白-1表达的影响。三磷酸腺苷γS诱导星形胶质细胞单核细胞趋化蛋白-1mRNA表达和蛋白合成。用几种MAP激酶抑制剂检测MAP激酶在这种诱导过程中的作用。酪氨酸激酶抑制剂PD98059和U0126可显著抑制γS诱导的单核细胞趋化蛋白-1mRNA表达和蛋白合成。SP600125对JNK的抑制作用可部分抑制它们的表达。另一方面，p38MAPK抑制剂SB203580可显著增强γS诱导的单核细胞趋化蛋白-1合成。进一步的研究表明，SB203580延长了三磷酸腺苷γS诱导的单核细胞趋化蛋白-1mRNA表达的持续时间，从而增加了单核细胞趋化蛋白-1的净产量。这些结果表明，三磷酸腺苷-γ-S诱导星形胶质细胞产生单核细胞趋化蛋白-1的相互调节是通过ERK和p38MAP激酶实现的。

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.