Jin Xiao-Gao, He Song-Qing, Yan Xue-Tao, Zhang Guangxiong, Wan Li, Wang Jintao, Li Yawen, Tian Xuebi, Tian Yuke, Luo Ailin
Department of Anesthesiology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei Province, China.
J Pain. 2009 Jan;10(1):80-9. doi: 10.1016/j.jpain.2008.07.009. Epub 2008 Oct 31.
Neuropathic pain due to nerve injury is associated with overactivity of spinal N-methyl-D-aspartate (NMDA) receptors and nitric oxide synthases (NOS). Spinal NOS and NMDA receptors could act in a concerted manner to excite each other in nociceptive signaling. Among the 3 major NOS isoforms, neuronal NOS (nNOS) has the most functional relationship with NMDA receptors through a PDZ-PDZ (PSD-95, Dlg, ZO-1 homology) postsynapse interaction. However, some nNOS variants lack the PDZ domain, which may result in the changes in the interaction with the NMDA receptor and subsequent localization and enzymatic activity. The aim of this study was to determine which nNOS variants are expressed in the spinal cord in neuropathic rats and deduce their role in neuropathic pain by testing the effects of these kinds of nNOS on nuclear factor-kappaB (NF-kappaB) activity in PC12 cells. Western blot analysis revealed that there were at least 3 bands of nNOS (155, 135, and 125 kDa) in the spinal cord and, moreover, that nNOS at 135 kDa decreased significantly after development of neuropathic pain. 5'-RACE-PCR and Southern blots determined that the nNOS at 155 and 135 kDa corresponded to nNOSalpha and nNOSbeta, respectively, which was confirmed by RT-PCR. PC12 cells transfected with the nNOSalpha gene had no effect on NF-kappaB activity, but nNOSbeta without the PDZ domain significantly decreased that in PC12 cells. Considering the importance of spinal NF-kappaB signaling in neuropathic rat, it could be concluded that changes in spinal nNOS variants and quantity after peripheral nerve injury implicate nNOS in the generation of neuropathic pain.
This article presents data demonstrating that nNOS variants change in the spinal cord of the rats after neuropathic pain and result in differential effects on NF-kappaB activity in PC12 cells. These changes in nNOS variants and their different characteristics may account for the spinal NO paradox role in neuropathic pain. Furthermore, these data suggest that nNOSbeta may represent a new therapeutic target for the treatment of chronic neuropathic pain.
神经损伤所致的神经性疼痛与脊髓N-甲基-D-天冬氨酸（NMDA）受体和一氧化氮合酶（NOS）的过度活跃有关。脊髓NOS和NMDA受体可能在伤害性信号传导中协同作用以相互兴奋。在3种主要的NOS同工型中，神经元型NOS（nNOS）通过PDZ-PDZ（PSD-95、Dlg、ZO-1同源性）突触后相互作用与NMDA受体具有最密切的功能关系。然而，一些nNOS变体缺乏PDZ结构域，这可能导致与NMDA受体相互作用以及随后的定位和酶活性发生变化。本研究的目的是确定哪些nNOS变体在神经性大鼠的脊髓中表达，并通过测试这些类型的nNOS对PC12细胞中核因子-κB（NF-κB）活性的影响来推断它们在神经性疼痛中的作用。蛋白质印迹分析显示脊髓中至少有3条nNOS条带（155、135和125 kDa），此外，神经性疼痛形成后135 kDa的nNOS显著减少。通过5'-RACE-PCR和Southern印迹确定155和135 kDa的nNOS分别对应于nNOSα和nNOSβ，这通过逆转录聚合酶链反应（RT-PCR）得到证实。用nNOSα基因转染的PC12细胞对NF-κB活性没有影响，但没有PDZ结构域的nNOSβ显著降低了PC12细胞中的NF-κB活性。考虑到脊髓NF-κB信号传导在神经性大鼠中的重要性，可以得出结论，外周神经损伤后脊髓nNOS变体和数量的变化表明nNOS参与了神经性疼痛的产生。
本文提供的数据表明，神经性疼痛后大鼠脊髓中的nNOS变体发生变化，并对PC12细胞中的NF-κB活性产生不同影响。nNOS变体的这些变化及其不同特征可能解释了脊髓NO在神经性疼痛中的矛盾作用。此外，这些数据表明nNOSβ可能代表治疗慢性神经性疼痛的一个新的治疗靶点。
