Department of Pharmacology and Toxicology, Virginia Commonwealth University School of Medicine, Richmond, VA 23298-0613, USA.
Mol Neurodegener. 2011 Nov 17;6:78. doi: 10.1186/1750-1326-6-78.
Fractalkine/CX3CL1 and its cognate receptor CX3CR1 are abundantly expressed in the CNS. Fractalkine is an unusual C-X3-C motif chemokine that is important in neuron-microglial communication, a co-receptor for HIV infection, and can be neuroprotective. To assess the effects of fractalkine on opiate-HIV interactive neurotoxicity, wild-type murine striatal neurons were co-cultured with mixed glia from the striata of wild-type or Cx3cr1 knockout mice ± HIV-1 Tat and/or morphine. Time-lapse digital images were continuously recorded at 20 min intervals for up to 72 h using computer-aided microscopy to track the same cells repeatedly.
Co-exposure to Tat and morphine caused synergistic increases in neuron death, dendritic pruning, and microglial motility as previously reported. Exogenous fractalkine prevented synergistic Tat and morphine-induced dendritic losses and neuron death even though the inflammatory mediator TNF-α remained significantly elevated. Antibody blockade of CX3CR1 mimicked the toxic effects of morphine plus Tat, but did not add to their toxicity; while fractalkine failed to protect wild-type neurons co-cultured with Cx3cr1-/--null glia against morphine and Tat toxicity. Exogenous fractalkine also normalized microglial motility, which is elevated by Tat and morphine co-exposure, presumably limiting microglial surveillance that may lead to toxic effects on neurons. Fractalkine immunofluorescence was expressed in neurons and to a lesser extent by other cell types, whereas CX3CR1 immunoreactivity or GFP fluorescence in cells cultured from the striatum of Cx3cr1-/- (Cx3cr1GFP/GFP) mice were associated with microglia. Immunoblotting shows that fractalkine levels were unchanged following Tat and/or morphine exposure and there was no increase in released fractalkine as determined by ELISA. By contrast, CX3CR1 protein levels were markedly downregulated.
The results suggest that deficits in fractalkine-CX3CR1 signaling contribute to the synergistic neurotoxic effects of opioids and Tat. Importantly, exogenous fractalkine can selectively protect neurons from the injurious effects of chronic opioid-HIV-1 Tat co-exposure, and this suggests a potential therapeutic course for neuroAIDS. Although the cellular mechanisms underlying neuroprotection are not certain, findings that exogenous fractalkine reduces microglial motility and fails to protect neurons co-cultured with Cx3cr1-/- mixed glia suggest that fractalkine may act by interfering with toxic microglial-neuron interactions.
趋化因子 fractalkine/CX3CL1 及其同源受体 CX3CR1 在中枢神经系统中大量表达。趋化因子 fractalkine 是一种不寻常的 C-X3-C 基序趋化因子，在神经元-小胶质细胞通讯中起重要作用，是 HIV 感染的共受体，具有神经保护作用。为了评估 fractalkine 对阿片类药物-HIV 相互作用的神经毒性的影响，将野生型鼠纹状体神经元与野生型或 Cx3cr1 敲除小鼠纹状体的混合神经胶质共培养，同时存在 HIV-1 Tat 和/或吗啡。使用计算机辅助显微镜以 20 分钟的间隔连续记录长达 72 小时的时间推移数字图像，以反复跟踪相同的细胞。
先前报道，Tat 和吗啡的共同暴露会协同增加神经元死亡、树突修剪和小胶质细胞运动。外源性 fractalkine 可防止 Tat 和吗啡协同诱导的树突丢失和神经元死亡，尽管炎症介质 TNF-α仍显著升高。CX3CR1 抗体阻断模拟了吗啡加 Tat 的毒性作用，但没有增加其毒性；而 fractalkine 未能保护与 Cx3cr1-/--null 神经胶质共培养的野生型神经元免受吗啡和 Tat 毒性的影响。外源性 fractalkine 还使 Tat 和吗啡共同暴露引起的小胶质细胞运动正常化，这可能限制了小胶质细胞的监视，从而导致对神经元的毒性作用。Fractalkine 免疫荧光在神经元中表达，在其他细胞类型中表达较少，而在 Cx3cr1-/-（Cx3cr1GFP/GFP）小鼠纹状体培养的细胞中，CX3CR1 免疫反应性或 GFP 荧光与小胶质细胞有关。免疫印迹显示，Tat 和/或吗啡暴露后 fractalkine 水平没有变化，并且通过 ELISA 确定没有增加释放的 fractalkine。相比之下，CX3CR1 蛋白水平明显下调。
研究结果表明，fractalkine-CX3CR1 信号通路的缺陷导致阿片类药物和 Tat 的协同神经毒性作用。重要的是，外源性 fractalkine 可以选择性地保护神经元免受慢性阿片类药物-HIV-1 Tat 共同暴露的损伤作用，这表明神经艾滋病有潜在的治疗方法。虽然神经保护的细胞机制尚不确定，但发现外源性 fractalkine 降低小胶质细胞运动，并且不能保护与 Cx3cr1-/-混合神经胶质共培养的神经元，表明 fractalkine 可能通过干扰毒性小胶质细胞-神经元相互作用来发挥作用。
