Krawczyk Paweł, Wojas Kamila, Milanowski Janusz, Roliński Jacek
Department of Pneumology, Oncology and Allergology, Medical University of Lublin, Jaczewskiego 8, 20-950, Lublin, Poland.
Arch Immunol Ther Exp (Warsz). 2007 Nov-Dec;55(6):405-15. doi: 10.1007/s00005-007-0046-2. Epub 2007 Dec 3.
Monocyte-derived dendritic cells (DCs) are currently under extensive evaluation as cell vaccines for cancer treatment. Many protocols regarding DCs generation in vitro with different protein components, especially autologous proteins, have been described. On the other hand, active tumor-derived factors in patients' serum could impair monocytes, which might result in their abrogated differentiation into DCs in vitro.
Autologous DCs from non-small-cell lung cancer (NSCLC)-bearing patients were generated in different culture microenvironments. Peripheral blood mononuclear cells (PBMCs) were cultured in the presence of interleukin-4 and granulocyte-monocyte-stimulating factor with supplementation of 10% autologous serum, 10% allogenic serum, or 2% human albumin. The course of apoptosis, phagocytic ability, and the immunophenotype of the generated DCs were analyzed using flow cytometric methods.
After 48 h of culture, we found a lower percentage of CD1a+/CD14+ and a higher percentage of CD1a+/CD14(-) cells in the culture supplemented with human albumin than in the cultures supplemented with serums. The lowest CD14 antigen expression was found in the human albumin-supplemented 48-h cultures. After 48 h in the cultures carried out with human albumin we found significantly higher percentages of AV+/PI+ cells and AV(-)/PI+ cells than in cultures supplemented with autologous or allogenic serum. We also noted that the expression of FITC-dextran after 4 and 24 h of incubation was significantly higher in the cultures supplemented with both serums than in the HA-SC. The percentage of semi-mature DCs and of CD83 expression was lowest in the culture supplemented with 2% human albumin.
The kind of culture supplementation had a great impact on the apoptosis of cultured PBMCs. It could also influence the yield of monocyte-derived DCs. It was also confirmed that autologous and allogenic serums provide suitable microenvironments for the generation of autologous DCs from NSCLC patients. The choice of culture supplementation for DC generation is still unsolved and further studies should be undertaken.
单核细胞衍生的树突状细胞（DCs）目前作为癌症治疗的细胞疫苗正在接受广泛评估。已经描述了许多关于在体外使用不同蛋白质成分，特别是自体蛋白质生成DCs的方案。另一方面，患者血清中的活性肿瘤衍生因子可能会损害单核细胞，这可能导致它们在体外无法分化为DCs。
在不同的培养微环境中生成来自非小细胞肺癌（NSCLC）患者的自体DCs。外周血单核细胞（PBMCs）在白细胞介素-4和粒细胞-单核细胞刺激因子存在下培养，并补充10%自体血清、10%异体血清或2%人白蛋白。使用流式细胞术方法分析所生成DCs的凋亡过程、吞噬能力和免疫表型。
培养48小时后，我们发现补充人白蛋白的培养物中CD1a+/CD14+细胞的百分比低于补充血清的培养物，而CD1a+/CD14(-)细胞的百分比更高。在补充人白蛋白的48小时培养物中发现CD14抗原表达最低。在用人白蛋白进行的培养中培养48小时后，我们发现AV+/PI+细胞和AV(-)/PI+细胞的百分比显著高于补充自体或异体血清的培养物。我们还注意到，在孵育4小时和24小时后，补充两种血清的培养物中FITC-葡聚糖的表达明显高于人白蛋白补充的培养物（HA-SC）。在补充2%人白蛋白的培养物中，半成熟DCs的百分比和CD83表达最低。
培养补充物的种类对培养的PBMCs的凋亡有很大影响。它也可能影响单核细胞衍生DCs的产量。还证实自体和异体血清为从NSCLC患者生成自体DCs提供了合适的微环境。用于DC生成的培养补充物的选择仍未解决，应进行进一步研究。
