HKU Data Repository
Browse

File(s) under embargo

Supporting data for "Differential effect of saturated and unsaturated fatty acids on γδ-T cell-mediated antitumor immunity"

dataset
posted on 2024-07-18, 01:54 authored by Yanmei ZhangYanmei Zhang
huPBMCs were cultured with BSA, PA, OA, or PA+OA conditions for 14 days. Then BSA-, PA-, OA-, or PA+OA-treated Vγ9Vδ2-T cells were negatively isolated using the TCR γ/δ+ T cell isolation reagent. Purified Vγ9Vδ2-T cells were co-cultured with target cancer cells for 6 hours. The supernatant was collected for further analyzing the secretions of their effector cytokines and lytic granules. The dead cancer cells were examined by flow cytometry. To explore the effect of PA or OA on the antitumor ability of Vγ9Vδ2-T cells in vivo, human lung cancer A549 cells were injected into Rag2–/–γc–/– mice conditioning different diets. To examine the changes of glycolysis and OXPHOS of BSA-, PA-, OA-, or PA+OA- Vγ9Vδ2-T cells, Vγ9Vδ2-T cells were isolated and the corresponding ECAR and OCR were detected using seahorse assay. To determine the role of pyroptosis on the impaired antitumor activity induced by PA, after treated Vγ9Vδ2-T cells with BSA, PA, OA, or PA+OA for 12 days, the inhibitor of pyroptosis DMF was used to treat Vγ9Vδ2-T cells overnight. Then the level of LDH release and the expression of pyroptosis markers were determined to verify the inhibitory effect of DMF on pyroptosis. To explore the mechanisms underlying distinct antitumor activity of PA and OA, purified PA or OA treated Vγ9Vδ2-T cells were performed proteomics analysis compared to BSA-treated Vγ9Vδ2-T cells. The Venn program was used to find the distinguished proteins and pathway analysis was used to find the top expressed pathway. After proteomics analysis, the critical pathways or molecules were verified in the BSA, PA, OA, or PA+OA-treated Vγ9Vδ2-T cells. To determine the effect of critical molecule IFNγ on pyroptosis in Vγ9Vδ2-T cells, anti-IFNγ Ab or recombinant IFNγ (rhIFNγ) was used to treat BSA, PA, OA, or PA+OA-treated Vγ9Vδ2-T cells. To determine the role of IFNγ in the antitumor activity of Vγ9Vδ2-T cells, PBMCs were cultured under BSA, PA, OA, or PA+OA conditions for 14 days. Then anti-IFNγ Ab or recombinant IFNγ (rhIFNγ) was used to establish the model of blockade or supplementation of IFNγ. Then the antitumor activity, the secretions of lytic granules, glycolysis, and OXPHOS were detected. We further verified the effect of IFNγ on the antitumor function of Vγ9Vδ2-T cells in vivo. Rag2–/–γc–/– mice were fed with LFD, palm oil HFD, or olive oil HFD for 30 days. Then GFP+ A549 tumor cells were subcutaneously inoculated into these mice. On day 7, 12, 17, and 22 following A549 cells inoculation, human anti-IFNγ neutralizing mAb or rhIFNγ was injected into mice intraperitoneally. After 12 hours, the tumor-bearing mice were intravenously injected with BSA-, PA-, or OA-treated Vγ9Vδ2-T cells, respectively. The tumor weight, volume, and survival time were monitored during the experiment.


History

Usage metrics

    Research Postgraduates

    Exports

    RefWorks
    BibTeX
    Ref. manager
    Endnote
    DataCite
    NLM
    DC