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目的探讨以腺相关病毒(AAV)为载体,前列腺特异性抗原(PSA)基因转染树突状细胞(DC)诱导前列腺癌患者外周血T细胞亚群变化特点及临床意义。方法抽取30例前列腺癌患者外周血,采用密度梯度离心法分离外周血单个核细胞,以rAAV/PSA感染DC前体细胞,采用系列细胞因子诱导DC前体细胞成熟。第6天收集成熟DC并与T细胞按比例混合培养,诱导细胞毒性T淋巴细胞(CTL)。分别于DC与T细胞混合培养前后应用流式细胞术分析外周血T细胞亚群及调节性T细胞(CD4+CD25+FoxP3+Treg)的表达水平。结果 PSA基因转染DC刺激T淋巴细胞爆发增殖,与培养前比较,混合培养6d后CD8+、CD8+CD69+、CD8+CD28+T细胞的比例和CD8+/CD4+比值均明显增高,差异有统计学意义(P<0.01);而CD8+CD28-T细胞和Treg细胞的比例均显著降低,差异有统计学意义(P<0.01)。CD4+T细胞比例较前略有升高,但差异无统计学意义(P>0.05)。结论 PSA基因转染DC能够有效地激活CD8+抗原特异性CTL,下调免疫抑制性T细胞,提高患者的细胞免疫功能,为前列腺癌的免疫治疗提供新的有效策略。
Objective To investigate the characteristics and clinical significance of T cell subsets in peripheral blood of patients with prostate cancer induced by adeno-associated virus (AAV) vector and prostate-specific antigen (PSA) gene transfected dendritic cells (DCs). Methods Peripheral blood samples from 30 patients with prostate cancer were collected. Peripheral blood mononuclear cells were isolated by density gradient centrifugation. DC precursor cells were infected with rAAV / PSA. Serial cytokines were used to induce DC precursor cell maturation. On the 6th day, mature DCs were harvested and mixed with T cells in proportion to induce cytotoxic T lymphocytes (CTLs). The levels of T lymphocyte subsets and regulatory T cells (CD4 + CD25 + FoxP3 + Treg) in peripheral blood were analyzed by flow cytometry before and after mixed culture of DC and T cells respectively. Results The PSA gene transfected DC stimulated the proliferation of T lymphocytes. Compared with the pre-culture, the ratio of CD8 +, CD8 + CD69 +, CD8 + CD28 + T cells and the ratio of CD8 + / CD4 were significantly increased after 6 d (P <0.01). However, the proportion of CD8 + CD28-T cells and Treg cells were significantly decreased, the difference was statistically significant (P <0.01). The proportion of CD4 + T cells was slightly higher than before, but the difference was not statistically significant (P> 0.05). Conclusion DCs transfected with PSA can effectively activate CD8 + antigen - specific CTLs, down - regulate immunosuppressive T cells, and improve cellular immune function in patients with prostate cancer, providing a novel and effective strategy for immunotherapy of prostate cancer.