CAJ | 학술논문

目的：研究HLA-G对育龄女性外周血中淋巴细胞（ PBLC）增殖及Treg细胞亚群的影响，探讨其在妊娠免疫耐受中的作用机制。方法：高表达HLA-G的绒癌细胞系JEG-3细胞与健康育龄女性外周血淋巴细胞（ PBLC）分组共培养，利用HLA-G中和抗体（87G）和重组人Ⅱ型肿瘤坏死因子受体-抗体融合蛋白（rhTNFR∶Fc）进行干预，实验分为7组：①JEG-3＋PBLC培养组；②JEG-3＋PBLC＋87G培养组；③JEG-3＋PBLC非接触培养组；④JEG-3＋PBLC＋87G非接触培养组；⑤对照组（单纯PBLC组）；⑥JEG-3＋PBLC＋rhTNFR：Fc培养组；⑦JEG-3＋PBLC＋87G ＋rhTNFR：Fc培养组。①～⑤每组CCK-8法检测各组PBLC增殖抑制情况，RT-PCR方法检测TNF-αmRNA的表达，①～⑦每组流式细胞术检测Treg细胞的比例。结果：CCK-8法显示，JEG-3＋PBLC培养组、JEG-3＋PBLC＋87G培养组、JEG-3＋PBLC非接触培养组、JEG-3＋PBLC＋87G非接触培养组PBLC增殖抑制率分别为（48.00±5.56）％、（14.67±4.04）％、（37.67±2.31）％、（8.33±3.21）％，各组间比较差异均有统计学意义，P值均＜0.05；RT-PCR结果显示JEG-3＋PBLC培养组、JEG-3＋PBLC非接触培养组TNF-αmRNA的表达量明显低于对照组，加入87G干预后，TNF-αmRNA的表达量较相应无87G组明显上升，P值均＜0.05。流式细胞检测显示，与对照组相比，JEG-3＋PBLC培养组Treg 细胞比例明显上升（P＜0.05），而JEG-3＋PBLC非接触培养组差异无统计学意义（P＞0.05），JEG-3＋PBLC＋87G培养组Treg细胞比例较JEG-3＋PBLC培养组明显下降（P＜0.05）；JEG-3＋PBLC ＋rhTNFR：Fc培养组与JEG-3＋PBLC培养组相比，Treg 细胞比例明显上升（P＜0.05）；以JEG-3＋PBLC ＋rhTNFR：Fc培养组为对照，JEG-3＋PBLC＋rhTNF：FC＋87G培养组Treg细胞比例明显下降，但明显高于JEG-3＋PBLC＋87G培养组，各组间比较差异均有统计学意义，P值均＜0.05。结论：HLA-G可以抑制育龄女性外周血淋巴细胞的增殖及抑制TNF-α的表达，上调Treg细胞的比例。目的：研究HLA-G對育齡女性外週血中淋巴細胞（ PBLC）增殖及Treg細胞亞群的影響，探討其在妊娠免疫耐受中的作用機製。方法：高錶達HLA-G的絨癌細胞繫JEG-3細胞與健康育齡女性外週血淋巴細胞（ PBLC）分組共培養，利用HLA-G中和抗體（87G）和重組人Ⅱ型腫瘤壞死因子受體-抗體融閤蛋白（rhTNFR∶Fc）進行榦預，實驗分為7組：①JEG-3＋PBLC培養組；②JEG-3＋PBLC＋87G培養組；③JEG-3＋PBLC非接觸培養組；④JEG-3＋PBLC＋87G非接觸培養組；⑤對照組（單純PBLC組）；⑥JEG-3＋PBLC＋rhTNFR：Fc培養組；⑦JEG-3＋PBLC＋87G ＋rhTNFR：Fc培養組。①～⑤每組CCK-8法檢測各組PBLC增殖抑製情況，RT-PCR方法檢測TNF-αmRNA的錶達，①～⑦每組流式細胞術檢測Treg細胞的比例。結果：CCK-8法顯示，JEG-3＋PBLC培養組、JEG-3＋PBLC＋87G培養組、JEG-3＋PBLC非接觸培養組、JEG-3＋PBLC＋87G非接觸培養組PBLC增殖抑製率分彆為（48.00±5.56）％、（14.67±4.04）％、（37.67±2.31）％、（8.33±3.21）％，各組間比較差異均有統計學意義，P值均＜0.05；RT-PCR結果顯示JEG-3＋PBLC培養組、JEG-3＋PBLC非接觸培養組TNF-αmRNA的錶達量明顯低于對照組，加入87G榦預後，TNF-αmRNA的錶達量較相應無87G組明顯上升，P值均＜0.05。流式細胞檢測顯示，與對照組相比，JEG-3＋PBLC培養組Treg 細胞比例明顯上升（P＜0.05），而JEG-3＋PBLC非接觸培養組差異無統計學意義（P＞0.05），JEG-3＋PBLC＋87G培養組Treg細胞比例較JEG-3＋PBLC培養組明顯下降（P＜0.05）；JEG-3＋PBLC ＋rhTNFR：Fc培養組與JEG-3＋PBLC培養組相比，Treg 細胞比例明顯上升（P＜0.05）；以JEG-3＋PBLC ＋rhTNFR：Fc培養組為對照，JEG-3＋PBLC＋rhTNF：FC＋87G培養組Treg細胞比例明顯下降，但明顯高于JEG-3＋PBLC＋87G培養組，各組間比較差異均有統計學意義，P值均＜0.05。結論：HLA-G可以抑製育齡女性外週血淋巴細胞的增殖及抑製TNF-α的錶達，上調Treg細胞的比例。
목적：연구HLA-G대육령녀성외주혈중림파세포（ PBLC）증식급Treg세포아군적영향，탐토기재임신면역내수중적작용궤제。방법：고표체HLA-G적융암세포계JEG-3세포여건강육령녀성외주혈림파세포（ PBLC）분조공배양，이용HLA-G중화항체（87G）화중조인Ⅱ형종류배사인자수체-항체융합단백（rhTNFR∶Fc）진행간예，실험분위7조：①JEG-3＋PBLC배양조；②JEG-3＋PBLC＋87G배양조；③JEG-3＋PBLC비접촉배양조；④JEG-3＋PBLC＋87G비접촉배양조；⑤대조조（단순PBLC조）；⑥JEG-3＋PBLC＋rhTNFR：Fc배양조；⑦JEG-3＋PBLC＋87G ＋rhTNFR：Fc배양조。①～⑤매조CCK-8법검측각조PBLC증식억제정황，RT-PCR방법검측TNF-αmRNA적표체，①～⑦매조류식세포술검측Treg세포적비례。결과：CCK-8법현시，JEG-3＋PBLC배양조、JEG-3＋PBLC＋87G배양조、JEG-3＋PBLC비접촉배양조、JEG-3＋PBLC＋87G비접촉배양조PBLC증식억제솔분별위（48.00±5.56）％、（14.67±4.04）％、（37.67±2.31）％、（8.33±3.21）％，각조간비교차이균유통계학의의，P치균＜0.05；RT-PCR결과현시JEG-3＋PBLC배양조、JEG-3＋PBLC비접촉배양조TNF-αmRNA적표체량명현저우대조조，가입87G간예후，TNF-αmRNA적표체량교상응무87G조명현상승，P치균＜0.05。류식세포검측현시，여대조조상비，JEG-3＋PBLC배양조Treg 세포비례명현상승（P＜0.05），이JEG-3＋PBLC비접촉배양조차이무통계학의의（P＞0.05），JEG-3＋PBLC＋87G배양조Treg세포비례교JEG-3＋PBLC배양조명현하강（P＜0.05）；JEG-3＋PBLC ＋rhTNFR：Fc배양조여JEG-3＋PBLC배양조상비，Treg 세포비례명현상승（P＜0.05）；이JEG-3＋PBLC ＋rhTNFR：Fc배양조위대조，JEG-3＋PBLC＋rhTNF：FC＋87G배양조Treg세포비례명현하강，단명현고우JEG-3＋PBLC＋87G배양조，각조간비교차이균유통계학의의，P치균＜0.05。결론：HLA-G가이억제육령녀성외주혈림파세포적증식급억제TNF-α적표체，상조Treg세포적비례。
Objective:To study the effect of HLA-Gon proliferation in peripheral blood lymphocytes of childbearing age women and Treg cell subsets,and investigate the mechanisms of immune tolerance in pregnancy.Methods:The high expression of HLA-G cho-riocarcinoma cell lines JEG-3 cells with peripheral blood lymphocytes( PBLC) of healthy childbearing age women co-culture,using the HLA-G neutralizing antibodies(87G)and recombinant human tumor necrosis factor receptor typeⅡ-antibody fusion protein(rhTNFR:Fc)to intervene.Experiments were divided into seven groups:①JEG-3+PBLC culture group;②JEG-3+PBLC+87G culture group;③JEG-3+PBLC non-contact culture group;④JEG-3+PBLC+87G non-contact culture group;⑤The control group(PBLC group);⑥JEG-3+PBLC+rhTNFR:Fc culture group;⑦JEG-3+PBLC+87G+rhTNFR:Fc culture group.Detected the PBLC proliferation inhibition by CCK-8 method and the expression of TNF-αmRNA by RT-PCR in①-⑤groups.The proportion of Treg cells were detected by flow cytometry in①-⑦groups.Results:The assay of CCK-8 showed that the PBLC proliferation inhibition rate of JEG-3+PBLC culture group,JEG-3 +PBLC+87G culture group,JEG-3+PBLC non-contact culture group,and JEG-3+PBLC+87G non-contact culture group were(48.00±5.56)%,(14.67±4.04)%,(37.67±2.31)% and(8.33±3.21)%,there was a statistically significant difference on each group ( P<0.05 ).The result of RT-PCR showed that the expression of TNF-αmRNA in JEG-3+PBLC culture group and JEG-3+PBLC non-contact culture group were significantly lower than the control group.Compared with the corresponding non-87G group,the expression of TNF-αmRNA increased significantly after the intervention of 87G,P<0.05.Compared with the control group, the proportion of Treg cells of JEG-3+PBLC culture group detected by flow cytometry was significantly increased(P<0.05).There was no significant difference between JEG-3+PBLC non-contact culture group and control group ( P>0.05 ).Compared with the JEG-3+PBLC culture group,the proportion of Treg cells of JEG-3+PBLC+87G culture group was significantly decreased(P<0.05).Compared with JEG-3+PBLC culture group,the proportion of Treg cells of JEG-3+PBLC+rhTNFR:Fc culture group was significantly increased( P<0.05).Set JEG-3+PBLC+rhTNFR:Fc culture group as control,the proportion of Treg cells of JEG-3+PBLC+rh TNF:FC+87G culture was significantly decreased,but obviously higher than JEG-3+PBLC+87G culture group,there was a statistically significant difference on each group(P<0.05).Conclusion:HLA-G can inhibit peripheral blood lymphocyte proliferation of childbearing age women and inhibit the expression of TNF-α,and up-regulate the proportion of Treg cells.