CD4^+CD25^+FOXP3^+调节性T淋巴细胞数量及功能变化与子宫内膜异位症患者发病相关性研究

The function and distribution of CD4^+CD25^+FOXP3^+ regulatory T lymphocytes in endometriosis

目的:分析子宫内膜异位症(EMs)患者在位、异位内膜及外周血内CD4^+CD25^+FOXP3^+调节性T淋巴细胞(Treg细胞)的数量、分布及功能特性,探讨Treg细胞与EMs发病之间的关系。方法:收集EMs患者在位、异位内膜组织及外周血标本,并以非内异症患者作为对照,免疫组化分析在位、异位内膜内叉头状/翅膀状螺旋转录因子3(FOXP3)阳性细胞的数量及分布变化情况,并与EMs患者的r AFS临床分期进行相关性分析。流式细胞术检测Treg细胞占外周血CD4^+T淋巴细胞的百分比,及磁珠分选外周血内CD4^+CD25^+T细胞及CD4^+CD25-T细胞,3H-thymidine法测定CD4^+CD25^+T细胞免疫抑制功能变化情况。结果:EMs、非EMs患者在位内膜内平均FOXP3阳性细胞密度并无显著差异[(76.44±62.14)/mm^2vs(50.59±20.79)/mm^2;WU=152.0,P=0.20]。进一步按月经周期行亚组分析:EMs分泌期内膜内FOXP3阳性细胞密度显著高于非EMs患者[(94.84±53.91)/mm^2vs(31.37±19.02)/mm^2;MU=43.00,P=0.03]。不同r AFS期别患者卵巢异位症病灶内FOXP3阳性细胞密度并无显著差异[I^II期:(123.00±115.00)/mm^2vs III^IV期:(111.00±108.00)/mm^2;MU=139.5,P>0.05]。EMs患者外周血内Treg细胞比例显著低于非EMs患者[(0.58±0.21)%vs(1.35±0.38)%,P<0.001],但EMs患者外周血CD4^+CD25^+T淋巴细胞对CD4^+CD25-T淋巴细胞增殖抑制率,与非EMs相比并无显著变化[(68.43±18.15)%vs(66.37±17.78)%,P>0.05]。结论:EMs患者在位内膜内Treg细胞失去正常的周期波动性,EMs患者分泌期内Treg细胞数目增加可能与内异症发病相关。EMs患者外周血内Treg细胞比例下降,但其免疫抑制功能并无明显改变。

Objective： To investigate the distribution and function of Treg cells in endometriosis and non-endometriosis patients,and to determine the correlation CD4~＋CD25~＋FOXP3~＋regulatory T lymphocytes（Treg cells） with the pathogenesis of endometriosis.Methods： Eutopic endometrial specimens were obtained from 80 patients（40 with endometriosis and 40 without endometriosis） and ovarian endometriosis specimens were from 35 endometriosis patients.The quantity and distribution of the FOXP3 positive cells in the samples were detected by immunohistochemical analysis.Peripheral blood mononuclear cells（PBMC） were isolated from endometriosis and non-endometriosis patients.The percentage of Treg cells in CD4~＋T lymphocytes was analyzed by flow cytometry.Magnetic beads were applied to separate and collect CD4~＋CD25~＋T cells and CD4~＋CD25-T cells from PBMC,and then the immunosuppressive function of CD4~＋CD25~＋T cells was measured by 3H-thymidine method.Results： The FOXP3~＋regulatory T cells can be observed in all eutopic endometrial samples.The mean density of Foxp3~＋cells were（76.44±62.14）/mm^2and（50.59 ±20.79）/mm^2 for women with and without endometriosis,respectively（WU = 152.00; P = 0.20）.No significant difference was detected between women with[（55.33 ± 32.21）/mm^2]and without[（58.05 ±60.14） /mm^2]endometriosis during the proliferative phase too（WU = 54.00,P = 0.72）.However,highly significant increase in Foxp3~＋cell density was observed in women with endometriosis[（94.84 ±53.91）/mm^2]compared with those women without endometriosis[（31.37 ±19.02）/mm^2],during the secretory phase（WU=43.00,P = 0.03）.The density of FOXP3~＋cell was（123.00 ±115.00）/mm^2 in mild endometriosis（stage I ~ II）,and（111.00 ±108.00）/mm^2 in advanced endometriosis（stage III ~ IV）.There was no statistically significant differences between the two groups（WU = 27.00,P 〉0.05）.Compared with non-endometriosis patients,endometriosis patients had a significantly lower Treg： CD4~＋T cells ratio in the peripheral blood[（0.58 ±0.21） % vs（1.35 ±0.38） %,P〈0.001].The inhibitory rate of cell proliferation of CD4~＋CD25~＋T was（68.43 ±18.15） % in endometriosis patients,and（66.37 ± 17.78） % in non-endometriosis patients,and the differences were not significant（P〉0.05）.Conclusion： The quantity of Treg cells lose the periodic fluctuation in eutopic endometrium of endometriosis patients across the menstrual cycle.During the secretory phase,the density of Treg cell still significantly increased,which may be associated with the incidence of endometriosis.The percentage of Treg cell in peripheral blood decreased in endometriosis patients,but the immunosuppressive function of Treg cells group had no obvious change.