Fellner Avi, Barhum Yael, Angel Ariel, Perets Nisim, Steiner Israel, Offen Daniel, Lev Nirit
Department of Neurology, Rabin Medical Center, Beilinson Hospital, 39 Jabotinski St., Petah Tikva 49100, Israel.
Sackler Faculty of Medicine, Tel-Aviv University, Tel Aviv 69978, Israel.
Int J Mol Sci. 2017 Aug 1;18(8):1666. doi: 10.3390/ijms18081666.
Neuroinflammation contributes to amyotrophic lateral sclerosis (ALS) progression. TLR4, a transmembrane protein that plays a central role in activation of the innate immune system, has been shown to induce microglial activation in ALS models. TLR4 is up-regulated in the spinal cords of hSOD1 mice. We aimed to examine the effects of specific TLR4 inhibition on disease progression and survival in the hSOD1 mouse model of ALS. Immunologic effect of TLR4 inhibition in vitro was measured by the effect of TAK-242 treatment on LPS-induced splenocytes proliferation. hSOD1 transgenic mice were treated with TAK-242, a selective TLR4 inhibitor, or vehicle. Survival, body weight, and motor behavior were monitored. To evaluate in vivo immunologic modifications associated with TAK-242 treatment, we measured serum IL-1β in the plasma, as well as IL-1β and TNF-α mRNAs in the spinal cord in wild-type mice and in TAK-242-treated and vehicle-treated early symptomatic hSOD1 mice. Immunohistochemical analysis of motor neurons, astrocytes, and microglial reactivity in the spinal cords were performed on symptomatic (100 days old) TAK-242-treated and vehicle-treated hSOD1 mice. In vitro, splenocytes taken from 100 days old hSOD1 mice showed significantly increased proliferation when exposed to LPS ( = 0.0002), a phenomenon that was reduced by TAK-242 ( = 0.0179). TAK-242 treatment did not attenuate body weight loss or significantly affect survival. However, TAK-242-treated hSOD1 mice showed temporary clinical delay in disease progression evident in the ladder test and hindlimb reflex measurements. Plasma IL-1β levels were significantly reduced in TAK-242-treated compared to vehicle-treated hSOD1 mice ( = 0.0023). TAK-242 treatment reduced spinal cord astrogliosis and microglial activation and significantly attenuated spinal cord motor neuron loss at early disease stage ( = 0.0259). Compared to wild-type animals, both IL-1β and TNF-α mRNAs were significantly upregulated in the spinal cords of hSOD1 mice. Spinal cord analysis in TAK-242-treated hSOD1 mice revealed significant attenuation of TNF-α mRNA ( = 0.0431), but no change in IL-1β mRNA. TLR4 inhibition delayed disease progression, attenuated spinal cord astroglial and microglial reaction, and reduced spinal motor neuron loss in the ALS hSOD1 mouse model. However, this effect did not result in increased survival. To our knowledge, this is the first report on TAK-242 treatment in a neurodegenerative disease model. Further studies are warranted to assess TLR4 as a therapeutic target in ALS.
神经炎症促进肌萎缩侧索硬化症（ALS）的进展。Toll样受体4（TLR4）是一种跨膜蛋白，在先天免疫系统的激活中起核心作用，已证实在ALS模型中可诱导小胶质细胞活化。TLR4在hSOD1小鼠的脊髓中上调。我们旨在研究特异性抑制TLR4对ALS的hSOD1小鼠模型疾病进展和存活的影响。通过TAK - 242处理对脂多糖（LPS）诱导的脾细胞增殖的影响来测定体外抑制TLR4的免疫效应。hSOD1转基因小鼠用选择性TLR4抑制剂TAK - 242或赋形剂处理。监测存活情况、体重和运动行为。为了评估与TAK - 242处理相关的体内免疫改变，我们测量了野生型小鼠以及TAK - 242处理和赋形剂处理的早期有症状hSOD1小鼠血浆中的血清白细胞介素 - 1β（IL - 1β），以及脊髓中的IL - 1β和肿瘤坏死因子 - α（TNF - α）mRNA。对有症状的（100日龄）TAK - 242处理和赋形剂处理的hSOD1小鼠的脊髓运动神经元、星形胶质细胞和小胶质细胞反应性进行免疫组织化学分析。在体外，取自100日龄hSOD1小鼠的脾细胞在暴露于LPS时增殖显著增加（P = 0.0002），这一现象被TAK - 242降低（P = 0.0179）。TAK - 242处理并未减轻体重减轻或显著影响存活。然而，TAK - 242处理的hSOD1小鼠在阶梯试验和后肢反射测量中显示出疾病进展的暂时临床延迟。与赋形剂处理的hSOD1小鼠相比，TAK - 242处理的小鼠血浆IL - 1β水平显著降低（P = 0.0023）。TAK - 242处理减少了脊髓星形胶质细胞增生和小胶质细胞活化，并在疾病早期显著减轻了脊髓运动神经元损失（P = 0.0259）。与野生型动物相比，hSOD1小鼠脊髓中的IL - 1β和TNF - α mRNA均显著上调。TAK - 242处理的hSOD1小鼠的脊髓分析显示TNF - α mRNA显著降低（P = 0.0431），但IL - 1β mRNA无变化。在ALS的hSOD1小鼠模型中，抑制TLR4可延迟疾病进展，减轻脊髓星形胶质细胞和小胶质细胞反应，并减少脊髓运动神经元损失。然而，这种作用并未导致存活率增加。据我们所知，这是关于TAK - 242在神经退行性疾病模型中治疗的首次报告。有必要进一步研究评估TLR4作为ALS的治疗靶点。
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