BMC Complement Altern Med. 2013 Jul 18;13:179. doi: 10.1186/1472-6882-13-179.
Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease that results from a progressive loss of motor neurons. Familial ALS (fALS) is caused by missense mutations in Cu, Zn-superoxide dismutase 1 (SOD1) that frequently result in the accumulation of mutant protein aggregates that are associated with impairments in the ubiquitin-proteasome system (UPS). UPS impairment has been implicated in many neurological disorders. Bee venom (BV) extracted from honey bees has been used as a traditional medicine for treating inflammatory diseases and has been shown to attenuate the neuroinflammatory events that occur in a symptomatic ALS animal model.
NSC34 cells were transiently transfected with a WT or G85R hSOD1-GFP construct for 24 hrs and then stimulated with 2.5 μg/ml BV for 24 hrs. To determine whether a SOD1 mutation affects UPS function in NSC34 cells, we examined proteasome activity and performed western blotting and immunofluorescence using specific antibodies, such as anti-misfolded SOD1, anti-ubiquitin, anti-GRP78, anti-LC3, and anti-ISG15 antibodies.
We found that GFP-hSOD1G85R overexpression induced SOD1 inclusions and reduced proteasome activity compared with the overexpression of GFP alone in NSC34 motor neuronal cells. In addition, we also observed that BV treatment restored proteasome activity and reduced the accumulation of ubiquitinated and misfolded SOD1 in GFP-hSOD1G85R-overexpressing NSC34 motor neuronal cells. However, BV treatment did not activate the autophagic pathway in these cells.
Our findings suggest that BV may rescue the impairment of the UPS in ALS models.
肌萎缩侧索硬化症（ALS）是一种神经退行性疾病，由运动神经元的进行性丧失引起。家族性 ALS（fALS）是由铜、锌-超氧化物歧化酶 1（SOD1）的错义突变引起的，这些突变通常导致突变蛋白聚集体的积累，与泛素-蛋白酶体系统（UPS）的损伤有关。UPS 损伤与许多神经退行性疾病有关。从蜜蜂中提取的蜂毒已被用作治疗炎症性疾病的传统药物，并已被证明可以减轻在有症状的 ALS 动物模型中发生的神经炎症事件。
将 WT 或 G85R hSOD1-GFP 构建体瞬时转染到 NSC34 细胞中 24 小时，然后用 2.5μg/ml BV 刺激 24 小时。为了确定 SOD1 突变是否影响 NSC34 细胞中的 UPS 功能，我们使用特定的抗体（如抗错误折叠的 SOD1、抗泛素、抗 GRP78、抗 LC3 和抗 ISG15 抗体）检查蛋白酶体活性并进行 Western blot 和免疫荧光。
我们发现 GFP-hSOD1G85R 过表达诱导 SOD1 包涵体并降低蛋白酶体活性，与 GFP 单独过表达相比，在 NSC34 运动神经元细胞中。此外，我们还观察到 BV 处理可恢复蛋白酶体活性并减少 GFP-hSOD1G85R 过表达的 NSC34 运动神经元细胞中泛素化和错误折叠的 SOD1 的积累。然而，BV 处理并未激活这些细胞中的自噬途径。
我们的研究结果表明，BV 可能挽救 ALS 模型中 UPS 的损伤。
