齐墩果酸通过靶向miR-130b-3p-PTEN-PI3K-Akt信号通路和糖酵解抑制M2巨噬细胞极化并增强肝细胞癌的抗PD-1治疗效果。

Oleanolic acid inhibits M2 macrophage polarization and potentiates anti-PD-1 therapy in hepatocellular carcinoma by targeting miR-130b-3p-PTEN-PI3K-Akt signaling and glycolysis.

作者信息

Tu Xiaoyu, Lin Wanfu, Zhai Xiaofeng, Liang Shufang, Huang Guokai, Wang Jingfang, Jia Wentao, Li Shu, Li Bai, Cheng Binbin

机构信息

Faculty of Traditional Chinese Medicine, Naval Medical University (Second Military Medical University), Shanghai 200433, China; Department of Rehabilitation Medicine and Physiotherapy, the First Affiliated Hospital of Naval Medical University (Second Military Medical University), Shanghai 200433, China.

Faculty of Traditional Chinese Medicine, Naval Medical University (Second Military Medical University), Shanghai 200433, China; Oncology Department of Traditional Chinese Medicine, the First Affiliated Hospital of Naval Medical University (Second Military Medical University), Shanghai 200433, China.

出版信息

Phytomedicine. 2025 Jun;141:156750. doi: 10.1016/j.phymed.2025.156750. Epub 2025 Apr 9.

DOI:10.1016/j.phymed.2025.156750

PMID:40250003

Abstract

BACKGROUND

Hypoxia promotes M2 polarization of macrophages and the formation of the immunosuppressive tumor microenvironment (TME) in hepatocellular carcinoma (HCC). Oleanolic acid (OA) has shown great potential in the treatment of HCC. However, the mechanisms of macrophage M2 polarization in hypoxic tumor TME and the regulating effect of OA is still unclear.

OBJECTIVE

To investigate the mechanisms of macrophage M2 polarization induced by hypoxic HCC cells-derived exosomes and examine the efficacy of OA in remedying the immunosuppressive TME and the anti-PD1 therapy potential.

METHODS

Hypoxic and normoxic HCC-derived exosomes (H-Exo and N-Exo) were collected by centrifugation. The microRNAs (miRNA) carried by the exosomes were sequenced and then screened to identify the functional miRNA. THP-1-induced macrophages were treated with exosomes or miRNAs to induce the M2 polarization of macrophages. Real-time RT-PCR and Western blotting were used to identify the direct target of miR-130b-3p and its downstream molecules. Hepa1-6 hepatoma-bearing mice were subjected to determine the efficacy of OA in regulating the TME and the anti-PD1 therapy potential.

RESULTS

H-Exo promotes macrophage M2 polarization, and thereby accelerates the migration and epithelial-mesenchymal transition (EMT) of HCC cells. Exosomal miRNA sequencing and subsequent functional validation showed that miR-130b-3p was the mediator of H-Exo-induced macrophage M2 polarization. PTEN was identified as the target of miR-130b-3p, and downregulation of PTEN by miR-130b-3p led to the activation of PI3K/Akt signaling and macrophage M2 polarization. In addition, miR-130b-3p also enhanced the glycolysis. OA suppressed H-Exo and miR-130b-3p-induced macrophage M2 polarization, also inhibited miR-130b-3p-induced glycolysis. In vivo, OA treatment enhanced the efficacy of anti-PD1 antibody by decreasing the number of M2 macrophages and increasing the number of CD8+ T cells.

CONCLUSION

Our findings uncover a new mechanism of hypoxic HCC cells-induced M2 polarization of macrophages through exosomal miR-130b-3p-PTEN-PI3K-Akt signaling. The combination therapy of OA with anti-PD1 antibody may lead to substantial improvements of the immunotherapy efficacy and expand the beneficiaries.

摘要

背景

缺氧促进巨噬细胞的M2极化以及肝细胞癌（HCC）中免疫抑制性肿瘤微环境（TME）的形成。齐墩果酸（OA）在HCC治疗中显示出巨大潜力。然而，缺氧肿瘤TME中巨噬细胞M2极化的机制以及OA的调节作用仍不清楚。

目的

探讨缺氧HCC细胞来源的外泌体诱导巨噬细胞M2极化的机制，并研究OA纠正免疫抑制性TME的疗效及抗PD1治疗潜力。

方法

通过离心收集缺氧和常氧HCC来源的外泌体（H-Exo和N-Exo）。对外泌体携带的微小RNA（miRNA）进行测序，然后筛选以鉴定功能性miRNA。用外泌体或miRNA处理THP-1诱导的巨噬细胞以诱导巨噬细胞的M2极化。采用实时RT-PCR和蛋白质印迹法鉴定miR-130b-3p的直接靶标及其下游分子。对荷Hepa1-6肝癌小鼠进行实验以确定OA调节TME的疗效及抗PD1治疗潜力。

结果

H-Exo促进巨噬细胞M2极化，从而加速HCC细胞的迁移和上皮-间质转化（EMT）。外泌体miRNA测序及后续功能验证表明，miR-130b-3p是H-Exo诱导巨噬细胞M2极化的介质。PTEN被鉴定为miR-130b-3p的靶标，miR-130b-3p下调PTEN导致PI3K/Akt信号通路激活和巨噬细胞M2极化。此外，miR-130b-3p还增强了糖酵解。OA抑制H-Exo和miR-130b-3p诱导的巨噬细胞M2极化，也抑制miR-130b-3p诱导的糖酵解。在体内，OA治疗通过减少M2巨噬细胞数量和增加CD8+T细胞数量增强了抗PD1抗体的疗效。