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目的初步探讨CD4+CD25+调节性T细胞(CD4+CD25+ regulatory T cells,CD4+CD25+Treg)在急性淋巴细胞白血病(acute lymphocytic leukemia,ALL)患者化疗前及化疗缓解后外周血中的表达水平,并研究患者血清能否诱导外周血CD4+CD25-T细胞转化为CD4+CD25+Treg。方法①采用流式细胞术分别检测ALL初诊组、化疗完全缓解或部分缓解组及正常对照组外周血中CD4+CD25+T细胞所占比例,然后通过荧光定量RT-PCR检测各组外周血中转录因子Foxp3mRNA的表达水平,并逐层分析比较。②采集正常人外周血单个核细胞后,对照组用正常人血清,实验组用患者血清并分别设浓度梯度进行培养,72h后采用流式细胞术、荧光定量RT-PCR分别检测CD4+CD25+T细胞和Foxp3mRNA表达。结果ALL化疗缓解组CD4+CD25+T细胞及Foxp3mRNA表达水平均明显高于ALL初诊组和正常对照组(P<0.05),后两者之间CD4+CD25+T细胞水平无统计学差异(P>0.05),但ALL初诊组Foxp3mRNA含量较正常对照组明显升高(P<0.01),差异具有统计学意义;并且血清培养对照组CD4+CD25+T细胞水平及Foxp3mRNA含量均明显低于实验组(P<0.05),且其表达并不随血清浓度的增加而升高。结论CD4+CD25+Foxp3+Treg在ALL初诊组及化疗缓解组患者外周血中比例明显升高,且初步表明患者血清中的可溶性物质可诱导外周血CD4+CD25+T细胞转化为CD4+CD25+Treg,提示CD4+CD25+Treg可能是ALL免疫抑制的一个重要原因。
Objective To investigate the expression of CD4 + CD25 + regulatory T cells (CD4 + CD25 + regulatory T cells) in peripheral blood of patients with acute lymphocytic leukemia (ALL) before chemotherapy and after remission, And study the patient’s serum can induce peripheral blood CD4 + CD25-T cells into CD4 + CD25 + Treg. Methods ① Flow cytometry was used to detect the proportion of CD4 + CD25 + T cells in the newly diagnosed group, complete remission chemotherapy or partial remission group and normal control group, respectively, and then detected by peripheral blood fluorescence quantitative RT-PCR The expression level of transcription factor Foxp3mRNA was analyzed by layer by layer. (2) After collecting normal human peripheral blood mononuclear cells, the control group was treated with normal human serum, and the experimental group was incubated with serum and concentration gradient respectively. After 72 hours, the expression of CD4 + CD25 + T cells was detected by flow cytometry and fluorescence quantitative RT- T cells and Foxp3 mRNA expression. Results The expressions of CD4 + CD25 + T cells and Foxp3 mRNA in ALL group were significantly higher than those in the newly diagnosed group and the normal control group (P <0.05). There was no significant difference in CD4 + CD25 + T cell level between the two groups > 0.05). However, the levels of Foxp3mRNA in newly diagnosed ALL patients were significantly higher than those in normal controls (P <0.01), and the difference was statistically significant. The levels of CD4 + CD25 + T cells and Foxp3mRNA in serum-cultured control group were significantly lower than those in experimental group (P <0.05), and its expression does not increase with the increase of serum concentration. Conclusion The proportion of CD4 + CD25 + Foxp3 + Treg in peripheral blood of newly diagnosed group and chemotherapy-remission group was significantly higher than that of the control group, and it was preliminarily indicated that the soluble substance in serum could induce the conversion of CD4 + CD25 + T cells into CD4 + CD25 + Treg, suggesting that CD4 + CD25 + Treg may be an important cause of ALL immunosuppression.