Harmol hydrochloride dihydrate (HHD) is a novel alkaloid salt of the natural β-carboline harmol, which is isolated from Peganum harmala L. Here, we studied whether HHD could induce autophagy in neuro cells and investigated the underlying molecular mechanism. After incubation with HHD, the number of GFP-LC3 puncta in cells was measured using confocal microscopy. The distribution and colocalization of autophagosomes and autolysosomes in the cells were also detected. LC3 was gathered and cultured in a medium containing HHD. Compared with control cells and cells starved for 2 h, the number of GFP-LC3 puncta and the LC3-II expression level were significantly increased in HHD-treated cells (p < .05). The number of autophagosome (red) was increased and most of them were colocalized with lysosomes (green). Moreover, HHD induced the formation of puncta with Lysotracker Red positive in the L3 fat bodies (p < .05). When treated HEK cells with HHD, the protein expression level of LC3-II was markedly increased, and the protein expression level of α-Syn was significantly decreased (p < .05). HHD could induce the increased autophagosome in neuro cells by induction of autophagy. Moreover, HHD may promote the degradation of α-Syn protein to protect neuro cells by inducing autophagy.

中文翻译：

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Harmol 盐酸盐二水合物诱导神经细胞自噬并通过 Atg5/Atg12 依赖途径促进 α-Syn 的降解

Harmol hydrochloride dihydrate (HHD) 是天然 β-咔啉 harmol 的新型生物碱盐，它是从Peganum harmala L 中分离出来的。在这里，我们研究了 HHD 是否可以诱导神经细胞自噬并研究其潜在的分子机制。与 HHD 孵育后，使用共聚焦显微镜测量细胞中 GFP-LC3 斑点的数量。还检测了细胞中自噬体和自溶酶体的分布和共定位。收集 LC3 并在含有 HHD 的培养基中培养。与对照细胞和饥饿 2 h 的细胞相比，HHD 处理的细胞中 GFP-LC3 斑点数量和 LC3-II 表达水平显着增加（p < .05)。自噬体（红色）的数量增加，其中大部分与溶酶体（绿色）共定位。此外，HHD 诱导了 L3 脂肪体中 Lysotracker Red 阳性斑点的形成 ( p  < .05)。当用 HHD 处理 HEK 细胞时，LC3-II 的蛋白表达水平显着升高，而 α-Syn 的蛋白表达水平显着降低 ( p  < .05)。HHD可通过诱导自噬诱导神经细胞自噬体增加。此外，HHD 可能通过诱导自噬促进 α-Syn 蛋白的降解以保护神经细胞。