精准医学杂志2025，Vol.40Issue(5)：384-390,397,8.DOI:10.13362/j.jpmed.202540094
CCR2过表达小鼠CD8+TILs活化对小鼠脑胶质瘤细胞增殖与凋亡的影响
Effect of activation of CC chemokine receptor 2-overexpressing mouse CD8+tumor-infiltrating lym-phocytes on the proliferation and apoptosis of mouse glioma cells
摘要
Abstract
Objective To construct CC chemokine receptor 2(CCR2)-overexpressing mouse CD8+tumor-infiltrating lymphocytes(TILs),and to investigate the effect of the activation of CCR2-overexpressing mouse CD8+TILs on the proliferation and apoptosis of mouse glioma cells.Methods CD8+TILs were isolated from lung adenocarcinoma tissue of C57BL/6J mice.A lentiviral vector with CCR2 overexpression was constructed and packaged to obtain the lentivirus,which was used to transfect mouse CD8+TILs.Mouse CD8+TILs were divided into CD8+T group(without transfection),CD8+T LV-Empty group(trans-fected with empty vector),and CD8+T CCR2OE group(transfected with CCR2-overexpressing virus),and flow cytometry was used to measure the proportion of CCR2-positive cells in each group.GL261 cells were divided into GL261 group(routine culture),group A(GL261 cells co-cultured with mouse CD8+TILs),group B(GL261 cells co-cultured with mouse CD8+TILs with the ad-dition of PD-1 antibody),group C(GL261 cells co-cultured with CCR2-overexpressing mouse CD8+TILs),and group D(GL261 cells co-cultured with CCR2-overexpressing mouse CD8+TILs with the addition of PD-1 antibody).Flow cytometry was used to measure the proportions of CD69+,CD107A+,IFN-γ-positive,and Granzyme B-positive cells in mouse CD8+TILs,and Annexin V-FITC/PI staining was used to measure the apoptosis rate of GL261 cells in each group of A-D.GL261 cells were divided into GL261 group(routine culture),group E(GL261 cells co-cultured with mouse CD8+TILs),group F(GL261 cells co-cultured with mouse CD8+TILs with the addition of PD-1 antibody),group G(GL261 cells co-cultured with CCR2-overexpressing mouse CD8+TILs),and group H(GL261 cells co-cultured with CCR2-over-expressing mouse CD8+TILs with the addition of PD-1 antibody),and CCK-8 assay was used to measure the proliferative capacity of GL261 in each group.Mouse CD8+TILs were divided into group a(routine culture),group b(treated with CCL2),group c(treated with CD3 antibody),and group d(treated with CD3 antibody and CCL2),and flow cytometry was used to measure the proportions of CD69+and CD107A+cells in CCR2-overexpressing mouse CD8+TILs in groups a-d.Results Flow cytometry showed that the proportion of CCR2-positive cells in the CD8+T CCR2OE group was significantly higher than that in the CD8+T LV-Empty group(F=202.80,P＜0.05).In the co-culture activation experiment of GL261 and mouse CD8+TILs,group C had significantly higher proportions of CD69+,CD107A+,IFN-γ-positive,and Granzyme B-positive cells than group A,and group D had significantly higher proportions than group B(F=17.47-146.70,P＜0.05).The apoptosis analysis of GL261 cells showed that group C had a significantly higher apoptosis rate than group A,and group D had a significantly higher apoptosis rate than group B(F=88.27,P＜0.05).The proliferation assay of GL261 cells showed that group G had a significantly lower proliferative capacity than group E,and group H had a significantly lower proliferative capacity than group F(F=40.85,P＜0.05).The CD3 and CCL2 stimulation experiment showed that group c had significantly higher proportions of CD69+and CD107A+cells than group a,and group d had significantly higher proportions than group c(F=108.80,204.06,P＜0.05).Conclusion CCR2 over-expression in mouse CD8+TILs can significantly enhance the activation of mouse CD8+TILs,thereby promoting the apoptosis of glioma cells and inhibiting their proliferation.关键词
神经胶质瘤/受体,CCR2/CD8阳性T淋巴细胞/肿瘤浸润/淋巴细胞活化/细胞增殖/细胞凋亡Key words
Glioma/Receptors,CCR2/CD8-positive T-lymphocytes/Tumor infiltrating/Lymphocyte activation/Cell proliferation/Apoptosis分类
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邱旭,于萌萌,王震,张强,范天宇,王斌,张丽..CCR2过表达小鼠CD8+TILs活化对小鼠脑胶质瘤细胞增殖与凋亡的影响[J].精准医学杂志,2025,40(5):384-390,397,8.基金项目
山东省自然科学基金面上项目(ZR2021M-H138) （ZR2021M-H138）
