帕金森病（PD）的发病率随着年龄的增长而迅速上升。随着全球老龄化的到来，PD患者数量随着老年人口的增加而不断增加，特别是在中国。此前，我们根据临床经验发现益肾除颤汤（YCD）具有缓解症状、延缓病情进展、控制PD发展的潜力。然而，其潜在机制仍有待探讨。本研究通过网络药理学和实验验证的系统方法，探讨了YCD在缓解PD方面可能的治疗效果，旨在为中医治疗PD提供新的认识。 YCD的化学结构和性质取自中药系统药理学数据库(TCMSP)、SwissADME、PubChem和PubMed。使用 Swiss Target Prediction、GeneCard、PubChem 和 UniProt 确定了 YCD 和 PD 的潜在目标。草药成分目标网络是通过 Cytoscape 软件创建的。此外，通过使用STRING数据库，筛选了蛋白质-蛋白质相互作用（PPI）网络。通过Metascape数据库进行基因功能GO和KEGG通路富集分析。制备来自 Sprague-Dawley (SD) 大鼠的 YCD 药物大鼠血清，并用鱼藤酮预处理 SH-SY5Y 细胞以建立 PD 模型。为了检查 YCD 对这些细胞的影响并探索 p38 丝裂原激活蛋白激酶 (MAPK) 途径的机制作用，用血清或 p38 MAPK 途径抑制剂预处理细胞。本研究采用细胞计数试剂盒 (CCK)-8 测定和 Hoechst 33,342 染色来评估 YCD 含药大鼠血清对鱼藤酮处理的 SH-SY5Y 细胞诱导的活力和形态变化。通过流式细胞术评估细胞凋亡。免疫荧光染色评估微管相关蛋白 2 (MAP2) 水平。采用酶联免疫吸附测定（ELISA）来定量炎症介质白细胞介素-1β（IL-1β）、白细胞介素-6（IL-6）和肿瘤坏死因子-α（TNF-α）的浓度。此外，还测定了活性氧（ROS）和超氧化物歧化酶（SOD）水平。 Western Blotting 测量了总 p38 MAPK (p-p38) 和磷酸化 p38 MAPK (p-p38) 的表达。本研究鉴定了 YCD 中的 65 种活性成分，这些成分被发现针对 801 个特定基因。通过筛选，从 PD 和 YCD 之间的 172 个重叠靶标中识别出 63 个潜在核心靶标。通过 GO 和 KEGG 分析对这些靶标进行了检查，揭示了它们与 MAPK、PI3K-Akt 信号通路、正向控制蛋白磷酸化和神经退行性疾病通路的显着相关性。 SH-SY5Y细胞用2μM鱼藤酮处理48小时，细胞活力降低至50%，并降低MAP2表达，增加细胞凋亡、氧化应激、炎症和p-p38表达的发生率。 YCD含药大鼠血清显着提高存活率，降低细胞凋亡率，并增加MAP2表达。 YCD 含药血清增加 SOD，减少 ROS，并抑制 IL-6、IL-1β 和 TNF-α 水平，从而抑制鱼藤酮处理的 SH-SY5Y 细胞的氧化应激和炎症。此外，YCD 含药血清显着降低了鱼藤酮诱导的 p-p38 表达。 SB203580是p38 MAPK的特异性抑制剂，还可以抑制p-p38表达、凋亡，恢复细胞形态损伤，改善炎症和氧化应激。YCD在体外增强细胞活力，降低凋亡率、炎症和氧化应激。这些有益作用可能涉及抑制 p38 通路和抑制 p38 MAPK 的磷酸化。© 2024 作者。

The incidence of Parkinson's disease (PD) rises rapidly with the increase of age. With the advent of global aging, the number of patients with PD is rising along with the elderly population, especially in China. Previously, we found that Yishen chuchan decoction (YCD), prescribed based on clinical experience, has the potential of alleviating symptoms, delaying the progression, and controlling the development of PD. Nonetheless, the underlying mechanistic role is yet to be explored.This research examined the possible therapeutic effects of YCD in alleviating PD via a systematic approach with network pharmacology and experimental validation, aiming at providing a new understanding of traditional Chinese medicine management regarding PD.The chemical structure and properties of YCD were adopted from Traditional Chinese Medicine System Pharmacology Database (TCMSP), SwissADME, PubChem, and PubMed. The potential targets for YCD and PD were identified using Swiss Target Prediction, GeneCard, PubChem, and UniProt. The herbal-component-target network was created via the Cytoscape software. Moreover, by using the STRING database, the protein-protein interaction (PPI) network was screened. Gene function GO and KEGG pathway enrichment analyses were performed via the Metascape database. YCD-medicated Rat Serum from Sprague-Dawley (SD) Rats was prepared, and SH-SY5Y cells were preconditioned with rotenone to develop the PD model. To examine the impact of YCD on these cells and explore the mechanistic role of the p38 mitogen-activated protein kinase (MAPK) pathway, the cells were pretreated with either serum or a p38 MAPK pathway inhibitor. This study employed the Cell Counting Kit (CCK)-8 assay and Hoechst 33,342 staining to evaluate the viability and morphological changes induced by the YCD-medicated rat serum on rotenone-treated SH-SY5Y cells. Apoptosis was assessed by Flow cytometry. Immunofluorescence staining assessed the microtubule-associated protein 2 (MAP2) level. Enzyme-linked immunosorbent assay (ELISA) was employed to quantify the concentrations of inflammatory mediators interleukin-1β (IL-1β), interleukin-6 (IL-6) and tumor necrosis factor-alpha (TNF-α). Also, reactive oxygen species (ROS) and superoxide dismutase (SOD) levels were determined. Western Blotting measured the expression of total and phospho-p38 MAPK (p-p38).This study identified 65 active components in YCD, which were found to target 801 specific genes. By screening, 63 potential core targets were identified from a pool of 172 overlapping targets between PD and YCD. These targets were examined by GO and KEGG analyses revealing their substantial correlation to MAPK, PI3K-Akt signaling pathways, positively controlling protein phosphorylation, and pathways of neurodegenerative diseases. SH-SY5Y cells were treated with 2 μM rotenone for 48 h, which reduced cell viability to 50 %, and reduced MAP2 expression, increased the rate of apoptosis, oxidative stress, inflammation, and p-p38 expressions. YCD-medicated rat serum significantly improved the viability, reduced the apoptosis rate, and increased the MAP2 expression. YCD-medicated serum increased SOD, reduced ROS and suppressed IL-6, IL-1β and TNF-α levels, thus inhibiting oxidative stress and inflammation in rotenone-treated SH-SY5Y cells. Moreover, YCD-medicated serum substantially lowered the p-p38 expression induced by rotenone. SB203580, a specific inhibitor of p38 MAPK, could also inhibit the p-p38 expression, apoptosis, and restore morphological damage of cells, also improve inflammation and oxidative stress.YCD enhanced cell viability and reduced apoptosis rate, inflammation, and oxidative stress in vitro. These beneficial effects could potentially involve the suppression of p38 pathway and suppressed the phosphorylation of p38 MAPK.© 2024 The Authors.