Background. Ganoderma lucidum has certain components with known pharmacological effects, including strengthening immunity and anti-inflammatory activity. G. lucidum seeds inherit all its biological characteristics. G. lucidum spore polysaccharide (GLSP) is the main active ingredient to enhance these effects. However, its specific biological mechanisms are not exact. Our research is aimed at revealing the specific biological mechanism of GLSP to enhance immunity and inhibit the growth of H22 hepatocellular carcinoma cells. Methods. We extracted primary macrophages (Mø) from BALB/c mice and treated them with GLSP (800 μg/mL, 400 μg/mL, and 200 μg/mL) to observe its effects on macrophage polarization and cytokine secretion. We used GLSP and GLSP-intervened macrophage supernatant to treat H22 tumor cells and observed their effects using MTT and flow cytometry. Moreover, real-time fluorescent quantitative PCR and western blotting were used to observe the effect of GLSP-intervened macrophage supernatant on the PI3K/AKT and mitochondrial apoptosis pathways. Results. In this study, GLSP promoted the polarization of primary macrophages to M1 type and the upregulation of some cytokines such as TNF-α, IL-1β, IL-6, and TGF-β1. The MTT assay revealed that GLSP+Mø at 400 μg/mL and 800 μg/mL significantly inhibited H22 cell proliferation in a dose-dependent manner. Flow cytometry analysis revealed that GLSP+Mø induced apoptosis and cell cycle arrest at the G2/M phase, associated with the expression of critical genes and proteins (PI3K, p-AKT, BCL-2, BAX, and caspase-9) that regulate the PI3K/AKT pathway and apoptosis. GLSP reshapes the tumor microenvironment by activating macrophages, promotes the polarization of primary macrophages to M1 type, and promotes the secretion of various inflammatory factors and cytokines. Conclusion. Therefore, as a natural nutrient, GLSP is a potential agent in hepatocellular carcinoma cell treatment and induction of apoptosis.

中文翻译：

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灵芝孢子多糖通过改变巨噬细胞极性和诱导细胞凋亡抑制肝癌细胞的生长

背景。灵芝含有某些具有已知药理作用的成分，包括增强免疫力和抗炎活性。G. lucidum种子继承了它的所有生物学特性。灵芝孢子多糖（GLSP）是增强这些作用的主要活性成分。然而，其具体的生物学机制并不确切。我们的研究旨在揭示GLSP增强免疫力和抑制H22肝癌细胞生长的具体生物学机制。方法。我们从 BALB/c 小鼠中提取原代巨噬细胞 ( Mø ) 并用 GLSP（800  μg /mL、400  μg /mL 和 200 μ g/mL) 观察其对巨噬细胞极化和细胞因子分泌的影响。我们使用 GLSP 和 GLSP 干预的巨噬细胞上清液治疗 H22 肿瘤细胞，并使用 MTT 和流式细胞术观察它们的作用。此外，实时荧光定量PCR和蛋白质印迹用于观察GLSP干预巨噬细胞上清液对PI3K/AKT和线粒体凋亡通路的影响。结果。在本研究中，GLSP 促进原代巨噬细胞向 M1 型极化和一些细胞因子如 TNF - α、IL- 1β、IL-6 和 TGF- β1的上调。MTT 测定显示 GLSP+M ø在 400  μ g/mL 和 800  μg/mL以剂量依赖性方式显着抑制H22细胞增殖。流式细胞术分析显示，GLSP+Mø 在 G2/M 期诱导细胞凋亡和细胞周期停滞，这与调节调节的关键基因和蛋白质（PI3K、p-AKT、BCL-2、BAX 和 caspase-9）的表达有关PI3K/AKT 通路和细胞凋亡。GLSP通过激活巨噬细胞重塑肿瘤微环境，促进原代巨噬细胞极化为M1型，促进各种炎症因子和细胞因子的分泌。结论。因此，作为一种天然营养素，GLSP是一种潜在的治疗肝癌细胞和诱导细胞凋亡的药物。