Department of Anesthesiology and Critical Care Medicine, Johns Hopkins University School of Medicine, Baltimore, MD, 21205, USA.
Center for Nanomedicine, Department of Ophthalmology, Wilmer Eye Institute, Johns Hopkins University School of Medicine, Baltimore, MD, 21231, USA.
J Neuroinflammation. 2017 Dec 19;14(1):252. doi: 10.1186/s12974-017-1004-5.
Rett syndrome (RTT) is a pervasive developmental disorder that is progressive and has no effective cure. Immune dysregulation, oxidative stress, and excess glutamate in the brain mediated by glial dysfunction have been implicated in the pathogenesis and worsening of symptoms of RTT. In this study, we investigated a new nanotherapeutic approach to target glia for attenuation of brain inflammation/injury both in vitro and in vivo using a Mecp2-null mouse model of Rett syndrome.
To determine whether inflammation and immune dysregulation were potential targets for dendrimer-based therapeutics in RTT, we assessed the immune response of primary glial cells from Mecp2-null and wild-type (WT) mice to LPS. Using dendrimers that intrinsically target activated microglia and astrocytes, we studied N-acetyl cysteine (NAC) and dendrimer-conjugated N-acetyl cysteine (D-NAC) effects on inflammatory cytokines by PCR and multiplex assay in WT vs Mecp2-null glia. Since the cysteine-glutamate antiporter (Xc) is upregulated in Mecp2-null glia when compared to WT, the role of Xc in the uptake of NAC and L-cysteine into the cell was compared to that of D-NAC using BV2 cells in vitro. We then assessed the ability of D-NAC given systemically twice weekly to Mecp2-null mice to improve behavioral phenotype and lifespan.
We demonstrated that the mixed glia derived from Mecp2-null mice have an exaggerated inflammatory and oxidative stress response to LPS stimulation when compared to WT glia. Expression of Xc was significantly upregulated in the Mecp2-null glia when compared to WT and was further increased in the presence of LPS stimulation. Unlike NAC, D-NAC bypasses the Xc for cell uptake, increasing intracellular GSH levels while preventing extracellular glutamate release and excitotoxicity. Systemically administered dendrimers were localized in microglia in Mecp2-null mice, but not in age-matched WT littermates. Treatment with D-NAC significantly improved behavioral outcomes in Mecp2-null mice, but not survival.
These results suggest that delivery of drugs using dendrimer nanodevices offers a potential strategy for targeting glia and modulating oxidative stress and immune responses in RTT.
雷特综合征（RTT）是一种进行性的普遍发育障碍，目前尚无有效的治疗方法。神经胶质功能障碍导致的脑内免疫失调、氧化应激和谷氨酸过量，与 RTT 的发病机制和症状恶化有关。在这项研究中，我们使用 Mecp2 基因敲除的 RTT 小鼠模型，研究了一种针对神经胶质细胞的新型纳米治疗方法，以减轻脑内炎症/损伤。
为了确定炎症和免疫失调是否是 RTT 中基于树突状聚合物治疗的潜在靶点，我们评估了来自 Mecp2 基因敲除和野生型（WT）小鼠的原代神经胶质细胞对 LPS 的免疫反应。我们使用内在靶向激活的小胶质细胞和星形胶质细胞的树突状聚合物，通过 PCR 和多重分析研究了 N-乙酰半胱氨酸（NAC）和树突状聚合物共轭 N-乙酰半胱氨酸（D-NAC）对 WT 与 Mecp2 基因敲除神经胶质细胞中炎症细胞因子的影响。由于胱氨酸-谷氨酸反向转运体（Xc）在 Mecp2 基因敲除神经胶质细胞中的表达上调，我们比较了 Xc 与 D-NAC 在 BV2 细胞内摄取 NAC 和 L-半胱氨酸的作用。然后，我们评估了每周两次系统给予 D-NAC 对 Mecp2 基因敲除小鼠改善行为表型和寿命的能力。
我们发现，与 WT 神经胶质细胞相比，源自 Mecp2 基因敲除小鼠的混合神经胶质细胞对 LPS 刺激的炎症和氧化应激反应更为剧烈。与 WT 相比，Mecp2 基因敲除神经胶质细胞中的 Xc 表达显著上调，而在 LPS 刺激下进一步增加。与 NAC 不同，D-NAC 绕过 Xc 进行细胞摄取，增加细胞内 GSH 水平，同时防止细胞外谷氨酸释放和兴奋性毒性。系统给予的树突状聚合物在 Mecp2 基因敲除小鼠的小胶质细胞中定位，但不在年龄匹配的 WT 同窝仔鼠中定位。D-NAC 治疗显著改善了 Mecp2 基因敲除小鼠的行为结果，但不能提高其生存率。
这些结果表明，使用树突状聚合物纳米器件递送药物为靶向神经胶质细胞和调节 RTT 中的氧化应激和免疫反应提供了一种潜在策略。
