Dual targeting by Artemisia rupestris L. disrupting TLR4/NF-κB signaling and macrophage driven immune evasion in hepatocellular carcinoma
Sirong Zhang1, Jie Li2, Man Ding3
1School of Biological and Food Engineering, Engineering Research Center for Development and High Value Utilization of Genuine Medicinal Materials in North Anhui Province, Suzhou University, Suzhou, Anhui, 234000, China; Xinjiang Institute of Materia Medica, Key Laboratory of Xinjiang Uygur Medicine, Urumqi, 830010, China; College of Pharmacy, Xinjiang Medical University, Urumqi, 830011, China.
Ethnopharmacological Relevance:
Hepatocellular carcinoma (HCC) is a malignant tumor with a high mortality rate in China, and its progression is closely related to the inflammatory microenvironment. The role and mechanism of Artemisia rupestris L. (ARL) in regulating immune response and exerting anti-HCC effects remains unclear.
Aim Of The Study:
This study aimed to evaluate the therapeutic potential of ARL against HCC by targeting inflammatory pathways and macrophage polarization.
Materials And Methods:
Co-culture models of HepG2, Hep3B, and THP-1 cells under inflammatory conditions were treated with cisplatin (DDP) or ARL low, medium and high concentration of 25, 50 and 100 μg/mL, respectively. Cytotoxicity was assessed via MTT assay. Cytokines such as IL-1β, IL-6, TNF-α were detected by ELISA while cell migration and invasion were studied using scratch and Transwell assays. The protein expressions of TLR4/MyD88/NF-κB pathway were analyzed by Western blot and immunofluorescence assay. CD86 expression in M1 macrophages was quantified using flow cytometry.
Results:
ARL significantly inhibited HCC cell proliferation at 100 μg/mL, reduced migration by 31.9-49.3 %, and decreased cell invasion up to 3.2 folds. Dose dependent suppressions of IL-1β, IL-6, and TNF-α were corelated significantly with reduced TLR4, MyD88, NF-κB, and phosphorylated IκB levels. ARL-H demonstrated efficacy comparable to the NF-κB inhibitor PDTC, blocking NF-κB nuclear translocation. Flow cytometry demonstrated that in the HCC cell co-culture system, ARL treatment downregulated CD86 expression, suggesting impaired M1 macrophage activation and ARL's immunomodulatory role in inflammation.
Conclusion:
ARL suppresses HCC progression by dual mechanisms that include the interference in TLR4/MyD88/NF-κB cascade to reduce proinflammatory cytokine production, and reprogramming macrophage polarization to attenuate immune driven tumor growth. These findings make ARL a promising natural therapeutic agent and provide an experimental basis for further in-depth subsequent exploration of the application of ARL in hepatocellular carcinoma.
