Supplementary Material for: Curcumin Protects an SH-SY5Y Cell Model of Parkinson’s Disease Against Toxic Injury by Regulating HSP90

<b><i>Background/Aims:</i></b> We aimed to explore the protective role of curcumin (Cur) in a cell model of Parkinson’s disease (PD) and its underlying mechanism. <b><i>Methods:</i></b> In this study, genes concerned with PD-related keywords were screened within DiGSeE database. The association network between Cur and selected genes was downloaded from STITCH, with the interactions analyzed by STRING. We built a mitochondrial toxin 1-methyl-4-phenylpyridinium (MPP⁺)-induced SH-SY5Y cell model of PD. Cell morphology was observed under an electron microscope. MTT assay was applied to detect cell proliferation rate. Western blot assay was conducted to determine the level of apoptotic markers, including cleaved caspase 3, Bcl-2-associated X protein (Bax) and B-cell lymphoma-extra-large (Bcl-xl). Tyrosine hydroxylase (TH), dopamine transporter (DAT) protein levels and dopamine (DA) concentration were identified as dopaminergic neuron markers and measured by western blotting or Enzyme-linked immunosorbent assay (ELISA). <b><i>Results</i></b>: Cur rescued the toxicity effects of MPP⁺ on SH-SY5Y cells, by controlling morphological change, promoting cell proliferation and inhibiting apoptosis. Of all PD-related genes, <i>HSP90</i> played an important role in Cur-gene network. HSP90 protein level was elevated by MPP⁺, whereas Cur could reverse this effect. Silencing of <i>HSP90</i> significantly attenuated the curative effect introduced by Cur, while <i>HSP90</i> overexpression enhanced the impact of Cur on PD. <b><i>Conclusion:</i></b> Cur can effectively inhibit the toxic effect of MPP⁺ on SH-SY5Y cells and significantly reduce the adverse effects of MPP⁺ on dopaminergic neurons via up-regulation of <i>HSP90</i>.

**背景与目的：** 本研究旨在探讨姜黄素（Cur）在帕金森病（PD）细胞模型中的保护作用及其潜在机制。**方法：** 本研究通过DiGSeE数据库筛选与帕金森病相关关键词关联的基因；从STITCH数据库下载姜黄素与筛选所得基因的关联网络，并通过STRING数据库分析其相互作用。本研究构建了线粒体毒素1-甲基-4-苯基吡啶鎓（MPP⁺）诱导的SH-SY5Y帕金森病细胞模型。通过电子显微镜观察细胞形态；采用MTT实验检测细胞增殖率；通过蛋白质印迹实验检测包括裂解型半胱氨酸天冬氨酸蛋白酶3、Bcl-2相关X蛋白（Bax）及B细胞淋巴瘤-XL（Bcl-xl）在内的细胞凋亡标志物水平。以酪氨酸羟化酶（TH）、多巴胺转运体（DAT）蛋白水平及多巴胺（DA）浓度作为多巴胺能神经元标志物，分别通过蛋白质印迹或酶联免疫吸附试验（ELISA）进行检测。**结果：** 姜黄素可通过抑制细胞形态改变、促进细胞增殖及抑制细胞凋亡，逆转MPP⁺对SH-SY5Y细胞的毒性作用。在所有帕金森病相关基因中，热休克蛋白90（HSP90）在姜黄素-基因网络中发挥重要作用。MPP⁺可升高HSP90蛋白水平，而姜黄素能够逆转这一效应。沉默HSP90可显著减弱姜黄素的治疗效果，而过表达HSP90则可增强姜黄素对帕金森病的作用。**结论：** 姜黄素可有效抑制MPP⁺对SH-SY5Y细胞的毒性作用，并通过上调HSP90显著减轻MPP⁺对多巴胺能神经元的损伤。