Table9_Unraveling the Catha edulis Extract Effects on the Cellular and Molecular Signaling in SKOV3 Cells.xlsx

Khat (Catha edulis (Vahl) Endl.) is an evergreen flowering shrub used as a stimulant in many regions worldwide including East Africa, the Arabian Peninsula, Europe, and the United States. Chewing leaves of khat induces excitement and euphoria, which are primarily attributed to two major constituents, cathinone and cathine. Khat also contains other important constituents such as cathedulins. A considerable number of studies reported side effects induced by the khat extracts to both embryos and adults. These include teratogenicity and developmental retardation, oral cancer and ulcers, high blood pressure, and myocardial infarction. So far, little attention has been paid to the effects of khat extracts on the molecular signaling interactions. We aimed in this study to investigate this through evaluating the effects of khat extracts on SKOV3, a human ovarian adenocarcinoma cell line. We show, by in vitro assays, that khat induces several cellular defects including reduced cell size, cell membrane damage, and apoptosis. At high khat extract concentrations, the cell metabolic activity, cell cycle, and cellular proliferation were affected. RT-qPCR analysis showed an increase in the gene expression of the apoptotic marker BAX, the tumor suppressor p53, and the inflammatory cytokine IL-6. Protein expression analysis by immunostaining showed downregulation of β-catenin, E-cadherin, and Ki-67 and upregulation of FZD8 and SPRY2, suggesting that Wnt and FGF signaling were implicated. SwissTargetPrediction in silico analysis showed that khat constituents cathine, cathinone, catheduline K2, and catheduline E5 bind to family A G-protein-coupled receptor, cause many neurological diseases and disorders such as Alzheimer's, schizophrenia, depression, and anxiety, and induce many ovarian cancer-related diseases. The analysis also showed that important signaling pathways such as CREB, Wnt, FGF, IL-6, and ERK/MAPK, and that of the endometrial cancer, and cell cycle were implicated. Upstream regulators of cathine and cathinone were found to potentially target several molecules including interleukin-8, MMP2, PLAU, and micro-RNAs. In conclusion, khat induces significant cellular and molecular changes that could potentially cause a wide range of serious diseases and syndromes. Such an impact could have a heavy burden on the health care system in the countries where khat is consumed.

阿拉伯茶（Catha edulis (Vahl) Endl.）是一种常绿开花灌木，在东非、阿拉伯半岛、欧洲及美国等全球诸多地区被用作精神兴奋剂。咀嚼其叶片可引发兴奋与欣快感，该效应主要归因于其两种主要活性成分——卡西酮（cathinone）与卡辛（cathine）。阿拉伯茶还含有卡杜林类（cathedulins）等其他重要活性成分。已有大量研究报道了阿拉伯茶提取物对胚胎与成人的不良反应，包括致畸性与发育迟缓、口腔癌与溃疡、高血压及心肌梗死。迄今为止，学界对阿拉伯茶提取物在分子信号交互通路中的影响尚未受到足够关注。本研究旨在通过评估阿拉伯茶提取物对人卵巢腺癌细胞系SKOV3的效应，探究该类影响。本研究通过体外实验证实，阿拉伯茶可诱发多种细胞损伤，包括细胞体积缩小、细胞膜损伤及细胞凋亡。在高浓度阿拉伯茶提取物处理下，细胞代谢活性、细胞周期及细胞增殖均受到显著抑制。实时定量聚合酶链反应（RT-qPCR）分析结果显示，凋亡标志物BAX、抑癌基因p53及炎症因子IL-6的基因表达水平均显著上调。采用免疫染色法开展的蛋白表达分析显示，β-连环蛋白（β-catenin）、上皮钙黏蛋白（E-cadherin）及Ki-67的表达水平下调，而FZD8与SPRY2的表达水平上调，提示Wnt信号通路与成纤维细胞生长因子（FGF）信号通路可能参与了该调控过程。基于SwissTargetPrediction平台的计算机模拟（in silico）分析显示，阿拉伯茶中的卡辛（cathine）、卡西酮（cathinone）、卡杜林K2（catheduline K2）及卡杜林E5（catheduline E5）可与G蛋白偶联受体A家族结合，进而引发阿尔茨海默病、精神分裂症、抑郁症及焦虑症等多种神经系统疾病与病症，并可诱发多种卵巢癌相关疾病。该分析还显示，CREB、Wnt、FGF、IL-6及ERK/MAPK等重要信号通路，以及子宫内膜癌、细胞周期相关通路均被牵涉其中。研究还发现，卡辛与卡西酮的上游调控因子可靶向包括白细胞介素-8（IL-8）、基质金属蛋白酶2（MMP2）、纤溶酶原激活物（PLAU）及微小RNA（micro-RNAs）在内的多种分子。综上所述，阿拉伯茶可引发显著的细胞与分子水平改变，潜在诱发多种严重疾病与综合征。此类影响可能给阿拉伯茶消费国的医疗保健系统带来沉重负担。