Authors
De-Guo Liu, Zi-Rong Li, Yi Shu, Jin-Qiu Li, Wan-Ting Dong, Qian Liu
Published in
Zhongguo Zhong yao za zhi = Zhongguo zhongyao zazhi = China journal of Chinese materia medica. Volume 51. Issue 12. Pages 3510-3521.
Abstract
This study investigated the effect of gypenosides(GPs) on anoikis resistance(AR) in human triple-negative breast cancer(TNBC) MDA-MB-231 cells and explored the underlying mechanism involving the transforming growth factor β1(TGF-β1)/forkhead box protein P3(Foxp3)/retinoic acid-related orphan receptor γt(RORγt) signaling axis. An anoikis model was established using poly-HEMA coated plates, and cells were divided into normal adherent, anoikis model, GPs(100 μg·mL~(-1)), TGF-β1 inhibitor(LY, 10 μmol·L~(-1)), and GPs + LY groups. Cell viability, apoptosis, invasion and migration were assessed by CCK-8 assay, flow cytometry with AnnexinV-FITC/PI staining, and Transwell assay, respectively. The mRNA and protein expression levels of TGF-β1, Foxp3, RORγt, B-cell Lymphoma 2(Bcl-2), Bcl-2 associated X protein(Bax), cysteine aspartic acid specific protease-3(caspase-3), and Smad2/3 phosphorylation levels were measured by quantitative real-time PCR(qRT-PCR) and Western blot. Nuclear localization of Foxp3 and RORγt was examined by immunofluorescence staining. A breast cancer bone metastasis model was established in nude mice by tail vein injection. After intervention, the circulating tumor cells(CTCs) in peripheral blood and the formation of bone metastasis foci were evaluated. The mRNA and protein expression levels of TGF-β1, Foxp3, RORγt in bone metastasis foci of each nude mouse breast cancer bone metastasis model were detected using qRT-PCR and Western blot. In vitro experimental results showed that GPs inhibited the viability of human breast cancer MDA-MB-231 cells, promoted apoptosis, and reduced the number of invasive and migratory cells. GPs downregulated the mRNA and protein expression levels of TGF-β1, Foxp3, and Bcl-2, upregulated the expression levels of RORγt, Bax, and caspase-3, and inhibited Smad2/3 phosphorylation. Furthermore, GPs decreased Foxp3 nuclear accumulation and enhanced RORγt nuclear localization, and exhibited synergistic effects with LY. In vivo experimental results indicated that GPs reduced the number of CTCs in peripheral blood and the number of bone metastases in nude mice. The changes in key mRNA and protein expression levels of the TGF-β1/Foxp3/RORγt signaling axis in bone tissue were consistent with those observed in in vitro experiments. In summary, GPs inhibit the TGF-β1/Smad signaling axis, downregulate Foxp3 expression and its nuclear localization, thereby relieving its negative regulation on RORγt. This leads to upregulation of pro-apoptotic proteins Bax and caspase-3, downregulation of anti-apoptotic protein Bcl-2, disruption of anoikis resistance, and reduction of metastatic potential in MDA-MB-231 cells. These findings reveal that the TGF-β1/Foxp3/RORγt signaling axis is a key target of GPs against metastatic triple-negative breast cancer(TNBC), providing new experimental evidence and clinical strategies for natural product-based therapy in TNBC.
PMID:
42543310
Bibliographic data and abstract were imported from PubMed on 03 Aug 2026.
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