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目的:构建前列腺癌PSA特异性树突状细胞(PSA-DC)瘤苗,并观察其体外免疫活性,为后续研究奠定基础。方法:分离骨髓前体细胞,大量制备骨髓DCs(bone marrowd erived DC,BMDC);分别以PSA、癌细胞裂解产物(lysate,Lys)、无关蛋白卵清蛋白(Ova)冲击DC制备PSA-DC、Lys-DC、Ova-DC瘤苗。ELISA法检测PSA-DC瘤苗培养上清中细胞因子(IL-12p70和IL-1β)水平的变化;观察PSA-DC瘤苗刺激抗原特异性T细胞增殖活性和诱导抗原特异性CTL杀伤活性,并与Lys-DC、Ova-DC瘤苗及未加入抗原冲击的DC(Non-DC)组作比较。结果:成功获得成熟DC,纯度可以达到>95%。ELISA法检测结果表明,PSA-DC、Lys-DC和Ova-DC组培养上清中IL-12p70和IL-1β水平均较Non-DC组明显升高(P<0.05)。混合淋巴细胞培养(MLR)显示:PSA-DC、Lys-DC组DCs刺激CD4+T细胞增殖的能力明显优于Ova-DC、Non-DC组(P<0.01);PSA-DC、Lys-DC组培养上清中IL-2、IFN-γ水平明显高于后两者(P<0.01),而IL-10和IL-4水平下调(P<0.05)。PSA-DC、Lys-DC组可产生针对PSA和肿瘤裂解物的DTH反应,而对无关抗原无效(P<0.05)。与OVA-DC、Non-DC组相比,Lys-DC、PSA-DC组体外诱导的CTL细胞对LNCaP细胞的杀伤活性较强(P<0.05),且具有抗原特异性。结论:利用PSA蛋白冲击DC可成功制备PSA特异性DC瘤苗,该瘤苗体外具有较强的免疫活性,能有效杀伤LNCaP细胞。
Objective: To construct prostate cancer PSA-specific dendritic cell (PSA-DC) tumor vaccine and observe its immunological activity in vitro, which will lay the foundation for further research. METHODS: Bone marrow progenitor cells (DCs) were isolated and bone marrow DCs (BMDCs) were prepared in large quantities. PSA-DCs were prepared by shocking DCs with PSA, lysate (Lys), and unrelated protein ova (Ova) Lys-DC, Ova-DC tumor vaccine. The changes of cytokines (IL-12p70 and IL-1β) in the culture supernatant of PSA-DC vaccine were detected by ELISA. The proliferation of PSA-DCs stimulated by PSA-DC vaccine and the induction of antigen-specific CTLs killing activity were observed. And compared with Lys-DC, Ova-DC tumor vaccine and non-DC (non-DC) group. Results: The successful acquisition of mature DC, purity can reach> 95%. The results of ELISA showed that the levels of IL-12p70 and IL-1β in the supernatant of PSA-DC, Lys-DC and Ova-DC group were significantly higher than those in Non-DC group (P <0.05). In mixed lymphocyte culture (MLR), the proliferation of CD4 + T cells in PSA-DC and Lys-DC groups was significantly better than that in Ova-DC and Non-DC groups (P <0.01) The levels of IL-2 and IFN-γ in the supernatants were significantly higher than those in the latter two groups (P <0.01), while the levels of IL-10 and IL-4 in the supernatants were decreased (P <0.05). The PSA-DC and Lys-DC groups produced DTH responses to PSA and tumor lysates, but not to irrelevant antigens (P <0.05). Compared with OVA-DC and Non-DC groups, CTL cells induced by Lys-DC and PSA-DC had stronger cytotoxicity against LNCaP cells (P <0.05) and antigen-specific. CONCLUSION: PSA-specific DC vaccine can be successfully prepared by using PSA protein to impact DCs. The vaccine has strong immunocompetence in vitro and can effectively kill LNCaP cells.