Ze-Qi-Tang Formula Induces Granulocytic Myeloid-Derived Suppressor Cell Apoptosis via STAT3/S100A9/Bcl-2/Caspase-3 Signaling to Prolong the Survival of Mice with Orthotopic Lung Cancer

Figure 6

ZQT selectively induced G-MDSC apoptosis by activating the STAT3/S100A9/Bcl-2/caspase-3 signaling pathway. (a) MDSCs from the tumor tissue of tumor-bearing mice treated with normal saline, ZQT, STAT3 inhibitor (S3I-201), or ZQT combined with S3I-201 were isolated using the MojoSort Mouse Isolation Kit. The protein expression of Ki67, STAT1, p-STAT1, STAT3, p-STAT3, S100A9, Arg-1, Bcl-2, Bax, cl-PARP, cyt c, and cl-caspase-3 in MDSCs from both normal saline and ZQT-treated mice was confirmed by Western blotting. (b) Lung tumor tissues from normal saline or ZQT-treated tumor-bearing mice were stained using S100A9, Ly6G, Ly6C, CD4, and CD8 antibodies for IHC assay (scale bar: 50 μM). (c) T-lymphocyte proliferation assay was used to show dose-dependent suppression of T cell proliferation by G-MDSCs isolated from tumor tissue of tumor-bearing mice treated with normal saline or ZQT. CD3⁺ T cells isolated from tumor stained with 5 μM CFSE were incubated with G-MDSCs for 72 h. Meanwhile, CD3 and CD28 antibodies were added for CD3⁺ T cell stimulation. Flow cytometry was used to quantify 72 h CFSE dilution. (d, e) G-MDSCs from tumor tissue of tumor-bearing mice treated with normal saline or ZQT were isolated using flow cytometry. The purity (CD11b⁺ Ly6G⁺) was >90% as assessed, and the protein expression of STAT3 and cl-caspase-3 was determined by IF (scale bar: 20 μM). . Data are expressed as the . n/s: nonstatistical significance. ; ; .

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