Establishment of a CD8+T cell exhaustion model in vitro
10.7644/j.issn.1674-9960.2025.04.004
- VernacularTitle:CD8+T细胞体外耗竭模型的建立
- Author:
Lingmin ZENG
1
;
Dingyi LU
;
Jiayi CHEN
;
Haoqian ZHANG
;
Jun GAO
;
Qiuying HAN
;
Xin PAN
Author Information
1. 军事科学院军事医学研究院,北京 100850
- Keywords:
CD8+T cells;
T-cell exhaustion;
exhaustion system;
immunotherapy
- From:
Military Medical Sciences
2025;49(4):265-272
- CountryChina
- Language:Chinese
-
Abstract:
Objective To establish a stable in vitro model of CD8+T cell exhaustion.Methods CD8+T cells were isolated and purified from the spleens of ovalbumin-specific CD8+T cell receptor(OT-I)transgenic mice and subjected to chronic antigen stimulation to induce exhaustion in vitro.Flow cytometry was employed to evaluate the expressions of exhaustion markers,secretion of effector cytokines,and transcription factor profiles in CD8+T cells.Exhausted and effector(non-exhausted)CD8+T cells were co-cultured with tumor cells before tumor cell viability was measured to assess the cytotoxic potential of CD8+T cells.Additionally,N-acetyl-L-cysteine(N-AC)was used as a positive control during exhaustion induction to validate the model.Results Chronic stimulation resulted in a significant upregulation of inhibitory receptors,including programmed cell death protein 1(PD-1),T cell immunoglobulin and mucin domain-containing protein 3(TIM-3),T-cell immunoglobulin and immunoreceptor tyrosine-based inhibitory motifdomain(TIGIT),and lymphocyte activation gene 3(LAG-3).Concurrently,the secretion of key effector cytokines such as tumor necrosis factor-α(TNF-α)and interferon-γ(IFN-γ)was markedly reduced.Exhausted CD8+T cells exhibited diminished cytotoxicity against tumor cells compared to effector CD8+T cells.Notably,treatment with N-AC effectively restored the function of exhausted CD8+T cells and enhanced their anti-tumor activity.Conclusion This study has established an effective in vitro model for CD8+T cell exhaustion.The use of N-AC demonstrates its potential to restore functionality in exhausted CD8+T cells,underscoring the reliability and utility of this model for investigating the anti-tumor potential of exhausted T cells.
