Department of Zoology, Biochemistry and Molecular Biology Laboratory, Institute of Science, Banaras Hindu University, Varanasi, 221005, India.
Neurochem Int. 2020 Mar;134:104652. doi: 10.1016/j.neuint.2019.104652. Epub 2019 Dec 28.
Role of NADPH oxidase1 in the development of inflammatory pain has been demonstrated by gene knockout studies. Nevertheless, pharmacological inhibition of NOX1 is a requisite approach for therapeutic utility. Recently, we have reported the anti-nociceptive effect of newly identified NOX1 specific inhibitor ML171 (2-acetylphenothiazine). Inhibition of NOX1 resulted in attenuation of nociceptive sensitization during acute inflammatory pain via inhibition of ROS generation and its downstream ERK1/2 activation. However, glial activation accompanying inflammation is closely related to the initiation and maintenance of pain. Peripheral nociceptive inputs activate the primary afferents via release of various chemical mediators which are potentially capable of mediating signals from neuron to glia in DRG and subsequently in spinal cord dorsal horn. The subsequent interactions between neuron and glia contribute to pain hypersensitivity. Thus, the present study was focused to investigate the effect of ML171 on ERK1/2 signaling, glial activation, and crosstalk between neuron and glia in a mouse model of formalin induced acute nociception. Thus, the present study was focused to investigate the effect of ML171 on ERK1/2 signaling, glial activation, and crosstalk between neuron and glia in DRG and dorsal horn of the spinal cord of lumbar region (L3-L5) in a mouse model of formalin induced acute nociception. Intraperitoneal administration of ML171 decreased nociceptive behavioral responses, i.e. the flinch and lick counts, in formalin induced nociceptive mice. Immunofluorescence and Western blot analysis demonstrated decreased levels of nociceptive mediators like p-ERK1/2, p-NFκB p65, Iba1 and GFAP in DRG as well as in spinal cord dorsal horn; supporting anti-nociceptive potential of ML171. Further, co-localization studies showed the neuron-glia crosstalk in tissue dependent manner. ERK1/2 was found to be activated in glia and NFκB in neurons in DRG; whereas in case of spinal cord ERK1/2 was activated in neurons and NFκB in astrocytes. Decrease in nociceptive behavioral response and activation of nociceptive mediators after intraperitoneal administration of ML171 strongly advocate anti-nociceptive potential of ML171. This is the first report demonstrating modulation of ERK1/2-NFκB signaling pathway, glial activation and regulation of neuron-glia crosstalk by NADPH oxidase1 inhibition towards its anti-nociceptive action.
NADPH 氧化酶 1（NOX1）在炎症性疼痛发展中的作用已经通过基因敲除研究得到了证明。然而，NOX1 的药理学抑制是治疗应用的必要方法。最近，我们报道了新鉴定的 NOX1 特异性抑制剂 ML171（2-乙酰吩噻嗪）的镇痛作用。NOX1 的抑制通过抑制 ROS 的产生及其下游 ERK1/2 的激活，导致急性炎症性疼痛时的伤害感受敏化减弱。然而，伴随炎症的神经胶质激活与疼痛的起始和维持密切相关。外周伤害性传入通过释放各种化学介质激活初级传入纤维，这些化学介质有可能介导 DRG 中神经元到神经胶质的信号，并随后在脊髓背角中进行介导。神经元和神经胶质之间的后续相互作用有助于疼痛敏感性。因此，本研究的重点是研究 ML171 对 ERK1/2 信号、神经胶质激活以及在福尔马林诱导的急性疼痛模型中 DRG 和脊髓背角神经元与神经胶质之间的串扰的影响。因此，本研究的重点是研究 ML171 对 ERK1/2 信号、神经胶质激活以及在福尔马林诱导的急性疼痛模型中 DRG 和脊髓背角神经元与神经胶质之间的串扰的影响。福尔马林诱导的急性疼痛小鼠模型中，腹腔内给予 ML171 可减少伤害性行为反应，即退缩和舔舐次数。免疫荧光和 Western blot 分析表明，在 DRG 以及脊髓背角中，伤害性介质如 p-ERK1/2、p-NFκB p65、Iba1 和 GFAP 的水平降低，支持 ML171 的镇痛潜力。此外，共定位研究表明，神经元-神经胶质串扰以组织依赖的方式发生。在 DRG 中，ERK1/2 在神经胶质中被激活，NFκB 在神经元中被激活；而在脊髓中，ERK1/2 在神经元中被激活，NFκB 在星形胶质细胞中被激活。腹腔内给予 ML171 后，伤害性行为反应的减少和伤害性介质的激活强烈支持 ML171 的镇痛潜力。这是第一项表明 NADPH 氧化酶 1（NOX1）抑制通过调节 ERK1/2-NFκB 信号通路、神经胶质激活和调节神经元-神经胶质串扰来发挥其镇痛作用的报告。
