Van Dyke Jonathan M, Smit-Oistad Ivy M, Macrander Corey, Krakora Dan, Meyer Michael G, Suzuki Masatoshi
Department of Comparative Biosciences and The Stem Cell and Regenerative Medicine Center, University of Wisconsin-Madison, Madison, WI, USA.
Department of Comparative Biosciences and The Stem Cell and Regenerative Medicine Center, University of Wisconsin-Madison, Madison, WI, USA.
Exp Neurol. 2016 Mar;277:275-282. doi: 10.1016/j.expneurol.2016.01.008. Epub 2016 Jan 13.
Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease characterized by progressive motor dysfunction and loss of large motor neurons in the spinal cord and brain stem. While much research has focused on mechanisms of motor neuron cell death in the spinal cord, degenerative processes in skeletal muscle and neuromuscular junctions (NMJs) are also observed early in disease development. Although recent studies support the potential therapeutic benefits of targeting the skeletal muscle in ALS, relatively little is known about inflammation and glial responses in skeletal muscle and near NMJs, or how these responses contribute to motor neuron survival, neuromuscular innervation, or motor dysfunction in ALS. We recently showed that human mesenchymal stem cells modified to release glial cell line-derived neurotrophic factor (hMSC-GDNF) extend survival and protect NMJs and motor neurons in SOD1(G93A) rats when delivered to limb muscles. In this study, we evaluate inflammatory and glial responses near NMJs in the limb muscle collected from a rat model of familial ALS (SOD1(G93A) transgenic rats) during disease progression and following hMSC-GDNF transplantation. Muscle samples were collected from pre-symptomatic, symptomatic, and end-stage animals. A significant increase in the expression of microglial inflammatory markers (CD11b and CD68) occurred in the skeletal muscle of symptomatic and end-stage SOD1(G93A) rats. Inflammation was confirmed by ELISA for inflammatory cytokines interleukin-1 β (IL-1β) and tumor necrosis factor-α (TNF-α) in muscle homogenates of SOD1(G93A) rats. Next, we observed active glial responses in the muscle of SOD1(G93A) rats, specifically near intramuscular axons and NMJs. Interestingly, strong expression of activated glial markers, glial fibrillary acidic protein (GFAP) and nestin, was observed in the areas adjacent to NMJs. Finally, we determined whether ex vivo trophic factor delivery influences inflammation and terminal Schwann cell (TSC) response during ALS. We found that intramuscular transplantation of hMSC-GDNF tended to exhibit less inflammation and significantly maintained TSC association with NMJs. Understanding cellular responses near NMJs is important to identify suitable cellular and molecular targets for novel treatment of ALS and other neuromuscular diseases.
肌萎缩侧索硬化症（ALS）是一种致命的神经退行性疾病，其特征为进行性运动功能障碍以及脊髓和脑干中大型运动神经元的丧失。尽管许多研究聚焦于脊髓中运动神经元细胞死亡的机制，但在疾病发展早期也观察到骨骼肌和神经肌肉接头（NMJ）的退行性变化。虽然最近的研究支持针对ALS患者骨骼肌进行治疗可能带来的益处，但对于骨骼肌和NMJ附近的炎症及神经胶质反应，或者这些反应如何影响ALS患者运动神经元的存活、神经肌肉支配或运动功能障碍，我们了解得还相对较少。我们最近发现，经过修饰能释放胶质细胞源性神经营养因子（hMSC-GDNF）的人间充质干细胞，在被递送至肢体肌肉时，可延长SOD1(G93A)大鼠的生存期，并保护其NMJ和运动神经元。在本研究中，我们评估了在疾病进展过程中以及hMSC-GDNF移植后，取自家族性ALS大鼠模型（SOD1(G93A)转基因大鼠）的肢体肌肉中NMJ附近的炎症和神经胶质反应。肌肉样本取自症状前期、症状期和终末期动物。在症状期和终末期SOD1(G93A)大鼠的骨骼肌中，小胶质细胞炎症标志物（CD11b和CD68）的表达显著增加。通过ELISA检测SOD1(G93A)大鼠肌肉匀浆中的炎性细胞因子白细胞介素-1β（IL-1β）和肿瘤坏死因子-α（TNF-α），证实了炎症的存在。接下来，我们观察到SOD1(G93A)大鼠肌肉中存在活跃的神经胶质反应，特别是在肌内轴突和NMJ附近。有趣的是，在与NMJ相邻的区域观察到活化神经胶质标志物胶质纤维酸性蛋白（GFAP）和巢蛋白的强烈表达。最后，我们确定了离体营养因子递送是否会影响ALS期间的炎症和终末雪旺细胞（TSC）反应。我们发现，hMSC-GDNF的肌内移植往往炎症较少，并且能显著维持TSC与NMJ的关联。了解NMJ附近的细胞反应对于确定针对ALS和其他神经肌肉疾病的新型治疗方法的合适细胞和分子靶点至关重要。
