肿瘤细胞对CD4^+CD25^+Treg细胞数量和功能的影响

Influence of tumor cells in number and function of CD4^+CD25^+Treg cells

目的:探讨肿瘤细胞与免疫细胞相互作用对CD4+CD25+Treg细胞数量和功能的影响。方法:建立Lewis肺癌细胞与小鼠脾淋巴细胞共培养体系。Lewis肺癌细胞与不同浓度小鼠脾淋巴细胞共培养,分为4组:实验组Ⅰ(5×105个Lewis肺癌细胞与1×106个淋巴细胞共培养)、对照组Ⅰ(1×106个淋巴细胞单独培养)、实验组Ⅱ(5×105个Lewis肺癌细胞与2×106个淋巴细胞共培养)、对照组Ⅱ(2×106个淋巴细胞单独培养);Lewis肺癌细胞与淋巴细胞共培养不同时间,采用3个时间点:24、48和72h;Lewis肺癌细胞培养上清与淋巴细胞共培养,选择培养上清浓度为20%和50%。采用流式细胞术检测了Lewis肺癌细胞与脾淋巴细胞共培养系统中CD4+CD25+Treg细胞数量变化,通过RT-PCR方法检测了共培养对Foxp3 mRNA表达的影响。结果:与对照组比较,实验组Ⅰ中CD4+CD25+Treg细胞数量和Foxp3 mRNA表达明显增强(P<0.05),实验组Ⅱ中CD4+CD25+Treg细胞数量和Foxp3 mRNA表达无明显变化(P>0.05);Lewis肺癌细胞与淋巴细胞培养24及48h可见CD4+CD25+Treg细胞数量及Foxp3 mRNA表达明显升高(P<0.05),而72h后变化不明显;20%和50%Lewis肺癌细胞培养上清均可明显提高CD4+CD25+Treg数量及Foxp3 mRNA表达(P<0.05)。结论:肿瘤细胞及其培养上清可诱导CD4+CD25+Treg细胞数量增加、功能增强,由肿瘤细胞所引起的CD4+CD25+Treg细胞产生及功能增强可能是肿瘤逃避免疫监视机制之一。

Objective To explore the influence of tumor cells in the number and function of CD4^＋CD25^＋Treg cells.Methods Lewis lung cancer cells and mouse spleen lymphocytes co-culture system was established.Lewis lung cancer cells with different concentrations of lymphocytes co-cultured were divided into 4 groups：experimental group Ⅰ（5×10^5 Lewis lung cancer cells and 1×10^6 lymphocytes co-culture）,control group Ⅰ（1×10^6 lymphocytes culture）,experimental group Ⅱ（5×10^5 Lewis lung cancer cells and 2×10^6 lymphocytes co-culture）,control group Ⅱ（2×10^6 lymphocytes culture）;Lewis lung cancer cells were co-cultivated with lymphocytes for different time,24,48,and 72 h three time points were selected.Lewis lung cancer cells culture supernatant was co-cultivated with lymphocytes,the concentrations of culture supernatant were 20% and 50%.The number changes of CD4^＋CD25^＋Treg cells in the co-culture system of Lewis lung cancer cells and splenic lymphocytes were detected by flow cytometry;the expression of Foxp3 mRNA after co-culture was detected by RT-PCR method.Results Compared with control group,the number of CD4＋CD25＋Treg cells and the expression of Foxp3 mRNA were significantly increased in experimental group Ⅰ （P〈0.05）,and there was no significant difference in experimental group Ⅱ （P〉0.05）;24 and 48 h after co-culture of Lewis lung cancer cells and lymphocytes,the number of CD4^＋CD25^＋Treg cells and the expression of Foxp3 mRNA were significantly increased （P〈0.05）,and there was no significant changes at 72 h;20% and 50% Lewis lung cancer cells supernatant could significantly increase the number of CD4＋CD25＋Treg cells and Foxp3 mRNA expression （P〈0.05）.Conclusion Tumor cells and their supernatants could induce the increase of the number of CD4^＋CD25^＋Treg cells and their function,this might be one of mechanisms of tumor-induced immune tolerance.