人白细胞抗原-G在子宫腺肌症患者子宫内膜腺细胞中通过白细胞介素-10诱导后对自然杀伤细胞的影响

Effect of HLA- G on NK cells in endometrial cells extracted from adenomyosis patients induced by IL- 10

目的:从临床提取子宫腺肌症患者子宫内膜组织与同时提取的相应患者的外周血自然杀伤(natural killer,NK)细胞共培养,加入白细胞介素-10(interleukin-10,IL-10)干预,从而产生人白细胞抗原-G(human leucocyte antigen-G,HLA-G),观察前者对NK细胞的影响。方法:首先从临床先后提取20例子宫腺肌症患者的子宫内膜组织,将组织细胞分离,弃去间质细胞,剩余的腺细胞分为A、B、C3组,同时每一组分为2个部分,即干预组和非干预组,用含有10%胎牛血清和1%的双抗DMEM F12培养基体外培养以上3组细胞。同时干预组加入20 ng/m L的IL-10,非干预组添加等量的培养基。24 h后,A组细胞利用酶联免疫吸附试验(enzymelinkedimmunosorbentassay,ELISA)分别测定干预组与非干预组的HLA-G的表达程度。随后应用免疫磁珠法从人外周血单个核细胞中分选出CD56+单核细胞,即NK细胞,将NK细胞与B组、C组细胞共培养,48 h后,B组用流式细胞仪检测干预组与非干预组的NK细胞凋亡率。C组用单纯计数法计数干预组与非干预组的NK细胞数量。结果:A组中,干预组HLA-G浓度平均为11.2 ng/m L,非干预组HLA-G浓度平均为3.2 ng/m L。B组中,加入IL-10的干预组与非干预组的NK细胞凋亡率分别平均为8.5%与4.2%。C组中,干预组与非干预组的NK细胞计数分别平均为2.5×105个/m L与1.7×105个/m L,48 h后干预组与非干预组的NK细胞计数分别平均为1.5×105个/m L与1.6×105个/m L。结论:IL-10增加了HLA-G表达,同时HLAG的表达对NK细胞的凋亡影响并不明显,但NK细胞数量减少,说明HLA-G对NK细胞有一定的抑制作用。

Objective： To observe the effects of human leucocyte antigen- G（ HLA- G） on natural killer（ NK） cells when endometrial tissue extracted from adenomyosis patients and peripheral blood NK cell are added with interleukin- 10（ IL- 10） to produce HLA- G. Method： 20 cases of uterine adenomyosis patients were selected to extract endometrial tissue successively. With tissue cells separated and interstitial cell abandoned,the remaining gland cells were divided into group A,B and C,with two parts in each group,the intervention group and non- intervention group.Cells in the three groups were cultured in vitro with 10 % fetal bovine serum and 1 % of the double resistance DMEM F12 medium. IL- 10（ 20 ng/m L） was added into the intervention group and the same amount of medium into the non- intervention group. 24 hours later,enzyme linked immunosorbent assay（ ELISA） method was adopted to detect the expressions HLA- G in the two groups. The immunomagnetic sand method was then used to sort out CD 56 ＋ mononuclear cells,namely the NK cells,from human peripheral blood mononuclear cells. The sorted NK cells and cells in group B and C were co- cultured. 48 hours later,flow cytometry was performed to analyze the NK cells apoptosis ratio in group B.Counting method was used to count cells in group C. Results： In group A,HLA- G concentration averaged 11. 2 ng / ml in the intervention group and 3. 2 ng / ml in the non- intervention group. In group B,the NK cells apoptosis rates averaged 8. 5 % and 4. 2 % respectively in the intervention group and in the non- intervention group. In group C,NK cells counting in the intervention group and in the non- intervention group averaged2. 5 × 105/ m L and 1. 7 × 105/ m L,with an average of 1. 5 × 105/ m L and 1. 6 × 105/ m L 48 hours later. Conclusion： IL- 10 significantly increased the expression of HLA- G with no obvious effects on NK cells apoptosis,while the number of NK cells decreasing significantly showed HLA- G has certain inhibitory effect on NK cells.