Akkermansia muciniphila Enhances the Antitumour Efficacy of αPD1 Therapy in Gastric Cancer by Remodelling the Tumour Immune Microenvironment
Description
The probiotic Akkermansia muciniphila (A. muciniphila) enhanced the efficacy of αPD1 therapy for gastric cancer by promoting the infiltration and activation of CD8⁺ T cells and altering the composition of the gut microbiota.
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Animal Modelling and Treatment Established a subcutaneous gastric cancer mouse model TUNEL and Ki67 Staining The tumour tissues were fixed, paraffin-embedded, and sectioned. The sections were deparaffinized, rehydrated, and incubated with a CK8 antibody to label the cancer cells. The TUNEL reaction mixture was applied and incubated at 37 °C for 1 h, followed by incubation with a Ki67 antibody. The sections were then imaged under a fluorescence microscope. Flow Cytometry Assay For surface staining, the cells were incubated with fluorochrome-conjugated antibodies against CD45, CD3, CD4, and CD8 for 30 min at 4 °C. For intracellular staining, the cells were fixed, permeabilized, and incubated with antibodies against FoxP3 and IFN-γ for 30 min at 4 °C. Immunohistochemistry The sections were incubated overnight at 4 °C with primary antibodies against CD4 and CD8. Enzyme-Linked Immunosorbent Assay (ELISA) Tumour tissues were homogenized in PBS and stored at −20 °C after centrifugation. Cytokine levels were quantified using ELISA kits according to the manufacturer’s instructions. 16S rRNA Sequencing All the faecal DNA samples were processed in a single batch and sequenced using the Illumina HiSeq platform for shotgun metagenomic analysis.