目的 探索中药淫羊藿提取物阿可拉定通过肝细胞癌(HCC)外泌体调控信号转导与转录激活因子 3(STAT3)介导 的巨噬细胞极化的药理作用。 方法 人肝癌细胞株 HepG2 细胞分别给予阿可拉定(10 μmol / L)或二甲基亚砜(DMSO)体外干 预后,将其与 THP-1 衍生的巨噬细胞培养于 Transwell 共培养系统中,ELISA 法检测巨噬细胞白细胞介素-6(IL-6)和转化生 长因子-β(TGF-β)分泌水平,流式细胞仪检测巨噬细胞 M2 型极化标记 CD206 的表达。 提取、鉴定经阿可拉定或 DMSO 干预 HepG2 细胞株外泌体,并利用外泌体干预 THP-1 衍生的巨噬细胞,再利用酶联免疫吸附法(ELISA)检测巨噬细胞分泌 IL-6 和 TGF-β 水平,蛋白质印迹法检测外泌体极化蛋白 STAT3 的表达和磷酸化。 结果 与阿可拉定诱导的 HepG2 细胞共培养 后,巨噬细胞 IL-6 和 TGF-β 的分泌和 CD206 的表达明显降低;阿可拉定诱导的 HepG2 外泌体也可抑制巨噬细胞分泌 IL-6 和 TGF-β,且能显著下调巨噬细胞信号转导与转录激活因子 3(STAT3)的磷酸化。 结论 阿可拉定可抑制 HCC 相关巨噬细胞 向 M2 极化,其机制与其诱导的 HCC 细胞外泌体相关。 阿可拉定是一种免疫调节剂,可影响 HCC 肿瘤微环境。

Objective To explore the pharmacological effect of acodarin an extract from the Chinese herbal medicine Epimedium in regulating signal transducer and activator of transcription 3 STAT3 -mediated macrophage polarization via exosomes derived from hepatocellular carcinoma HCC cells. Method HepG2 cell line was administrated by Icaritin 10 μmol / L or DMSO in vitro and then was cocultured with THP-1 cell-derived macrophage in a Transwell coculture system. The levels of IL-6 and TGF-β secreted by the macrophage were detected by ELISA assay. Flow cytometry was utilized to examine CD206 expression which is a biomarker of M2 macrophage. We extracted and identified the exosomes derived from DMSO or Icaritin-treated HepG2 cells. ELISA assay was utilized to detect IL-6 and TGF-β expressed in macrophage after administrated with exosomes isolated from HCC cells with or without Icaritin treatment. The expression and activity of STAT3 were assessed by Western blot assay. Result the expression of IL-6 TGF-β and CD206 were increased after cocultured with HCC cells but decreased significantly after cocultured with Icaritin - induced HepG2 cells. Meanwhile Icaritin-induced exosome also inhibited the expression of IL-6 and TGF-β as well as STAT3 activity. Conclusion Icaritin can inhibit macrophage polarize to M2 via HCC cells-derived exosomes. These findings indicate that Icaritin was an immune regulator in HCC microenvironment.