周围血中CD3^+ TCRvα24^+与CD3^+ TCRvβ11^+ NKT细胞之间的差异

Differences between CD3^+ TCRvα24^+ NKT cell and CD3^+ TCRvβ11^+ NKT cell in PBMC

目的:比较正常人周围血中CD3+TCRvβ11+NKT细胞和CD3+TCRvα24+NKT细胞在频率、亚群、表型特征及功能方面的异同,以进一步了解NKT细胞在免疫应答中的作用。方法:分离正常成年人PBMCs,利用流式细胞术(FCM)检测TCRvα24、TCRvβ11、CD4、CD8、CD45RA、CD62L和CCR7表面分子的表达;PMA+Ionomyc in刺激PBMCs后,检测CD3+TCRvα24+、CD3+TCRvβ11+NKT细胞产生细胞因子IL-4和IFN-γ的情况。结果:PBMCs中CD3+TCRvα24+和CD3+TCRvβ11+NKT细胞的平均频率分别为0.63%和0.43%,NKT细胞频率的个体差异较大,少数细胞同时表达TCRvα24和TCRvβ11;根据CD4和CD8分子的表达,PBMCs中CD3+TCRvα24+NKT细胞可分为CD4+、CD8+、CD4-CD8-3个亚群,平均频率分别为64.35%、19.04%、17.18%,CD3+TCRvβ11+NKT细胞同样可分为CD4+、CD8+、CD4-CD8-3个亚群,其平均频率分别为53.69%、18.99%、29.74%,相应各亚群之间无显著差异;CD45RA+CD3+TCRvβ11+NKT细胞的频率(71.14%)要高于CD45RA+CD3+TCRvα24+NKT细胞的频率(46.55%),二者之间差异有显著性,CD62L+CD3+TCRvα24+NKT(46.26%)对CD62L+CD3+TCRvβ11+NKT(42.36%)以及CCR7+CD3+TCRvα24+NKT(9.24%)对CCR7+CD3+TCRvβ11+NKT(8.22%)之间的差异均无统计学意义;细胞因子检测的结果表明CD3+TCRvα24+NKT细胞和CD3+TCRvβ11+NKT细胞产生的IL-4(13.01%对6.62%)和IFN-γ(38.12%对26.95%)的总体水平间无显著性差异,但是IFN-γ+IL-4+CD3+TCRvα24+NKT细胞的平均频率(12.65%)要高于IFN-γ+IL-4+CD3+TCRvβ11+NKT细胞的平均频率(3.02%),且二者之间的差异有统计学意义。结论:正常人周围血中CD3+TCRvα24+NKT细胞和CD3+TCRvβ11+NKT细胞在频率、表型及产生细胞因子方面均有一定差异,总体来看,二者频率虽小但表型复杂,产生细胞因子IFN-γ和IL-4的水平高,参与免疫调节及免疫应答的过程。

AIM： Clarified the differences between CD3＋TCRvα24＋ NKT cells and CD3＋TCRvβ11＋ NKT cells in their frequencies,subpopulations,phenotypes and biological functions,so as to fully understand the effects of NKT cells in immune responses.METHODS： PBMCs from blood donors were isolated and cell surface markers（CD3,TCRvα24,TCRvβ11,CD4,CD8,CD45RA,CD62L,CCR7） and intracellular cytokines（IL-4,IFN-γ） were detected by flow cytometry directly or after stimulation with PMA plus Ionomycin.RESULTS： The mean frequencies of CD3＋TCRvα24＋ NKT cells and CD3＋TCRvβ11＋ NKT cells in PBMCs were 0.63% and 0.43% and they varied according to individuals.A small population of NKT cells coexpressed TCRvα24 and TCRvβ11.The subpopulations of CD4＋ NKT 64.35%,CD8＋ NKT 19.04%,CD4-CD8-NKT 17.18% in human CD3＋TCRvα24＋ NKT cells and CD4＋ NKT 53.69%,CD8＋ NKT 18.99%,CD4-CD8-NKT 29.74% in CD3＋TCRvβ11＋ NKT cells could be identified based upon the expressions of CD4 and CD8 molecules.There were no significant differences between relative subtypes.The frequency of CD45RA＋CD3＋TCRvβ11＋ NKT cells（71.14%） was higher than the frequency of CD45RA＋CD3＋TCRvα24＋ NKT cells and the differences between them were significant.The differences between the frequencies of CD62L＋CD3＋TCRvα24＋ NKT cells（46.26%） and CD62L＋CD3＋TCRvβ11＋ NKT cells（42.36%）,the frequencies of CCR7＋CD3＋TCRvα24＋ NKT cells（9.24%） and CCR7＋CD3＋TCRvβ11＋ NKT cells（8.22%） were not significant.There were no significant differences in the secretions of IL-4 by CD3＋TCRvα24＋ NKT cells（13.01%） and CD3＋TCRvβ11＋ NKT cells（6.62%）,and IFN-γ by CD3＋TCRvα24＋ NKT cells（38.12%） and CD3＋TCRvβ11＋ NKT cells（26.95%）.However,there were significant differences between the mean frequency of IFN-γ＋IL-4＋CD3＋TCRvα24＋ NKT cells（12.65%） and that of IFN-γ＋IL-4＋CD3＋TCRvβ11＋ NKT cells（3.02%）.CONCLUSION： There were some differences between CD3＋TCRvα24＋ NKT cells and CD3＋TCRvβ11＋ NKT cells in their frequencies,phenotypes and productions of cytokines.In all,although their frequencies were low,the complicated phenotypes and high secretions of cytokines（IL-4 and IFN-γ） assigned NKT cells immunoregulatory effects.