Abstract：
BACKGROUND: Our previous study has revealed that asiaticoside (AC) promotes endoplasmic reticulum stress and antagonizes proliferation and migration of gastric cancer (GC) via miR-635/HMGA1 axis. However, the effect and mechanism of AC on other progressions of GC, such as ferroptosis and immune escape, are still unknown.
METHODS: AGS and HGC27 cells were incubated with 1, 2 and 4 μM of AC for 24 h. Mice xenografted with AGS cells were intragastrically injected with AC. The effect and mechanism of AC on GC were determined by the measurement of the ferrous iron level, the ROS level and the glutathione peroxidase (GSH) content, flow cytometry, enzyme-linked immunosorbent assay (ELISA), immunohistochemistry and western blotting assays.
RESULTS: AC increased the Fe2+ level and the ROS level, but decreased the expression of GPX4 and SLC7A11 and the GSH level. Besides, AC enhanced the percent of CD8+ T cells and the IFN-γ concentration, but reduced the PD-L1 expression and the IL-10 level. Mechanically, AC downregulated the relative levels of β-catenin, active-β-catenin, p-GSK3β/GSK3β, cyclin D1 and c-Myc in GC cells, which were rescued with the application of LiCl (an activator of Wnt/β-catenin pathway) in AGS cells. Moreover, activation of Wnt/β-catenin pathway by LiCl or the β-catenin overexpression inverted the effect of AC on ferroptosis and immune escape in GC cells. In vivo, AC treatment declined the tumor size and weight, the level of GPX4, SLC7A11, PD-L1 and IFN-γ, and the expression of Wnt/β-catenin pathway.
CONCLUSIONS: AC enhanced ferroptosis and repressed immune escape by downregulating the Wnt/β-catenin signaling in GC.
METHODS: AGS and HGC27 cells were incubated with 1, 2 and 4 μM of AC for 24 h. Mice xenografted with AGS cells were intragastrically injected with AC. The effect and mechanism of AC on GC were determined by the measurement of the ferrous iron level, the ROS level and the glutathione peroxidase (GSH) content, flow cytometry, enzyme-linked immunosorbent assay (ELISA), immunohistochemistry and western blotting assays.
RESULTS: AC increased the Fe2+ level and the ROS level, but decreased the expression of GPX4 and SLC7A11 and the GSH level. Besides, AC enhanced the percent of CD8+ T cells and the IFN-γ concentration, but reduced the PD-L1 expression and the IL-10 level. Mechanically, AC downregulated the relative levels of β-catenin, active-β-catenin, p-GSK3β/GSK3β, cyclin D1 and c-Myc in GC cells, which were rescued with the application of LiCl (an activator of Wnt/β-catenin pathway) in AGS cells. Moreover, activation of Wnt/β-catenin pathway by LiCl or the β-catenin overexpression inverted the effect of AC on ferroptosis and immune escape in GC cells. In vivo, AC treatment declined the tumor size and weight, the level of GPX4, SLC7A11, PD-L1 and IFN-γ, and the expression of Wnt/β-catenin pathway.
CONCLUSIONS: AC enhanced ferroptosis and repressed immune escape by downregulating the Wnt/β-catenin signaling in GC.
摘要：
背景：我们之前的研究表明积雪草苷（AC）通过miR-635/HMGA1轴促进内质网应激并拮抗胃癌（GC）的增殖和迁移。然而,AC对GC其他进展的影响和机制，比如铁性凋亡和免疫逃逸，仍然未知。
方法：将AGS和HGC27细胞与1、2和4μM的AC一起孵育24小时。将用AGS细胞异种移植的小鼠灌胃注射AC。通过亚铁含量的测定，确定了AC对GC的影响和机理。ROS水平和谷胱甘肽过氧化物酶（GSH）含量，流式细胞术,酶联免疫吸附测定(ELISA),免疫组织化学和蛋白质印迹分析。
结果：AC提高了Fe2+水平和ROS水平，但降低了GPX4和SLC7A11的表达和GSH水平。此外,AC提高了CD8+T细胞的百分比和IFN-γ浓度，但降低PD-L1表达和IL-10水平。机械上,AC下调β-连环蛋白的相对水平，活性β-连环蛋白，p-GSK3β/GSK3β,GC细胞中的细胞周期蛋白D1和c-Myc，通过在AGS细胞中应用LiCl（Wnt/β-catenin途径的激活剂）来挽救它们。此外，LiCl或β-catenin过表达对Wnt/β-catenin途径的激活逆转了AC对GC细胞铁凋亡和免疫逃逸的影响。在体内,AC治疗降低了肿瘤的大小和重量，GPX4、SLC7A11、PD-L1和IFN-γ的水平，Wnt/β-catenin通路的表达。
结论：AC通过下调GC中的Wnt/β-catenin信号传导来增强铁凋亡并抑制免疫逃逸。
方法：将AGS和HGC27细胞与1、2和4μM的AC一起孵育24小时。将用AGS细胞异种移植的小鼠灌胃注射AC。通过亚铁含量的测定，确定了AC对GC的影响和机理。ROS水平和谷胱甘肽过氧化物酶（GSH）含量，流式细胞术,酶联免疫吸附测定(ELISA),免疫组织化学和蛋白质印迹分析。
结果：AC提高了Fe2+水平和ROS水平，但降低了GPX4和SLC7A11的表达和GSH水平。此外,AC提高了CD8+T细胞的百分比和IFN-γ浓度，但降低PD-L1表达和IL-10水平。机械上,AC下调β-连环蛋白的相对水平，活性β-连环蛋白，p-GSK3β/GSK3β,GC细胞中的细胞周期蛋白D1和c-Myc，通过在AGS细胞中应用LiCl（Wnt/β-catenin途径的激活剂）来挽救它们。此外，LiCl或β-catenin过表达对Wnt/β-catenin途径的激活逆转了AC对GC细胞铁凋亡和免疫逃逸的影响。在体内,AC治疗降低了肿瘤的大小和重量，GPX4、SLC7A11、PD-L1和IFN-γ的水平，Wnt/β-catenin通路的表达。
结论：AC通过下调GC中的Wnt/β-catenin信号传导来增强铁凋亡并抑制免疫逃逸。
