Department of Pathology and Laboratory Medicine, University of British Columbia, G227-2211 Wesbrook Mall, Vancouver, BC, V6T 2B5, Canada.
J Neuroinflammation. 2018 Sep 19;15(1):270. doi: 10.1186/s12974-018-1290-6.
Axonal degeneration and neuronal loss have been described as the major causes of irreversible clinical disability in multiple sclerosis (MS). The aryl-hydrocarbon receptor nuclear translocator 2 (ARNT2) protein has been associated with neuroprotection in models of ischemia and neuronal responses to stressors.
To characterize its potential to influence inflammatory neurodegeneration, we examined ARNT2 expression in the experimental autoimmune encephalomyelitis (EAE) model of MS and characterized mediators that influence ARNT2 expression as well as plausible partners and targets.
Arnt2 message and protein levels dropped significantly in EAE spinal cords as disease developed and were lowest at peak disability. ARNT2 expression is prominent in neuronal cell bodies within the gray matter with some staining in glial fibrillary acidic protein (GFAP) astrocytes in healthy animals. At peak disease, ARNT2 expression is reduced by 20-50% in gray matter neurons compared to healthy controls. ARNT2 intensity in neurons throughout the EAE spinal cord correlated inversely with the degree of immune cell infiltration (r = - 0.5085, p < 0.01) and axonal damage identified with SMI32 staining (r = - 0.376, p = 0.032). To understand the relationship between ARNT2 expression and neuronal health, we exposed enriched cortical cultures of neurons to hydrogen peroxide (HO) to mimic oxidative stress. HO at lower doses rapidly increased ARNT2 protein levels which returned to baseline within 3-4 h. Exposure to higher doses of HO) dropped ARNT2 levels below baseline, preceding cytotoxicity measured by morphological changes and lactate dehydrogenase release from cells. Decreases in ARNT2 secondary to staurosporine and HO preceded increases in cleaved caspase 3 and associated apoptosis. We also examined expression of neuronal pas 4 (Npas4), whose heterodimerization with ARNT2 drives expression of the neurotrophic factor brain-derived neurotrophic factor (Bdnf). Like ARNT2, Npas4 levels also decline at the onset of EAE and are linked to decreases in Bdnf. In vitro, HO exposure drives Npas4 expression that is tied to increases in Bdnf.
Our data support ARNT2 as a neuronal transcription factor whose sustained expression is linked to neuronal and axonal health, protection that may primarily be driven through its partnering with Npas4 to influence BDNF expression.
轴突变性和神经元丢失已被描述为多发性硬化症（MS）中不可逆临床残疾的主要原因。芳香烃受体核转位蛋白 2（ARNT2）与缺血模型中的神经保护以及神经元对应激源的反应有关。
为了表征其影响炎症性神经退行性变的潜力，我们检查了 MS 的实验性自身免疫性脑脊髓炎（EAE）模型中的 ARNT2 表达，并描述了影响 ARNT2 表达的介质以及可能的伴侣和靶标。
随着疾病的发展，Arnt2 信使和蛋白质水平在 EAE 脊髓中显着下降，在疾病高峰期时最低。ARNT2 表达在灰质内的神经元细胞体中最为明显，在健康动物的胶质纤维酸性蛋白（GFAP）星形胶质细胞中有一些染色。在疾病高峰期，与健康对照组相比，灰质神经元中的 ARNT2 表达减少了 20-50%。EAE 脊髓中神经元的 ARNT2 强度与免疫细胞浸润程度呈负相关（r=-0.5085，p<0.01），与 SMI32 染色鉴定的轴突损伤呈负相关（r=-0.376，p=0.032）。为了了解 ARNT2 表达与神经元健康之间的关系，我们用过氧化氢（HO）暴露于富含皮质的神经元培养物中，以模拟氧化应激。较低剂量的 HO 迅速增加 ARNT2 蛋白水平，3-4 小时内恢复基线。暴露于更高剂量的 HO 会使 ARNT2 水平降至基线以下，这先于细胞形态变化和乳酸脱氢酶从细胞释放所测量的细胞毒性。由于 staurosporine 和 HO 导致的 ARNT2 减少先于 cleaved caspase 3 的增加和相关的细胞凋亡。我们还检查了神经元 pas 4（Npas4）的表达，其与 ARNT2 的异二聚化驱动神经营养因子脑源性神经营养因子（Bdnf）的表达。与 ARNT2 一样，Npas4 水平也在 EAE 发作时下降，并与 Bdnf 减少有关。在体外，HO 暴露会驱动 Npas4 表达，这与 Bdnf 的增加有关。
我们的数据支持 ARNT2 作为神经元转录因子，其持续表达与神经元和轴突健康有关，这种保护主要可能通过其与 Npas4 的结合来影响 BDNF 的表达。
