摘要目的 探讨CD160分子在荷瘤小鼠CD8+T淋巴细胞上的表达及其对CD8+T细胞活性和免疫功能的影响.方法 C57BL/6小鼠接种TC-1宫颈癌细胞建立小鼠荷瘤模型,免疫磁珠分选小鼠脾脏细胞获得CD8+T细胞,流式细胞术检测CD160在CD8+T细胞上的表达;流式细胞仪(FACS)分选CD8+T细胞获得CD160+ CD8+T细胞群和CD160-CD8+T细胞群,将两群细胞分别与肿瘤细胞抗原肽热休克蛋白70 (HSP70)-TC-1复合物和去除CD8+T的脾细胞混合,两群混合细胞各分为两组,其中一组同时加入CD160高亲和力配体单纯疱疹病毒侵入介导子(HVEM)的抗体,刺激培养6d,羧基荧光素二醋酸盐琥珀酰亚胺脂(CFSE)标记法检测CD8+T细胞增殖,流式细胞术检测各组细胞胞内穿孔素和γ-干扰素(IFN-γ)的表达.结果 荷瘤3周,小鼠CD8+T细胞上CD160分子的阳性率为(18.050 ±7.944)％,显著高于对照小鼠的阳性率[(8.663 ±3.921)％,P＜0.01];刺激培养后,CD160+ CD8+T细胞群中增殖细胞比例为(3.425 ±2.352)％,显著低于CD160-CD8+T细胞群的(15.440±6.673)％ (P＜0.01),抗HVEM抗体阻断CD160与HVEM的相互作用后,CD160+ CD8+T细胞群中增殖细胞比例显著增高达到(12.000 ±5.286)％ (P ＜0.01);同时,CD160+CD8+T细胞群中表达穿孔素和IFN-γ的阳性细胞率分别为(5.263±2.752)％和(4.150±2.499)％,显著低于CD160-CD8+T细胞群的(17.600±7.203)％和(11.480 ±4.790)％ (P＜0.01),抗HVEM抗体的作用使CD160+ CD8+T细胞群中表达穿孔素和IFN-γ的阳性细胞率显著增高,分别达到(12.380±4.922)％和(11.280 ±4.037)％ (P ＜0.01).结论 CD160在荷瘤小鼠的脾CD8+T细胞上表达升高且与其免疫活性功能的下降有关;阻断CD160和HVEM的相互作用,部分逆转了CD8+T细胞的免疫活性的抑制.

abstractsObjective To investigate the expression level of CD160 on CD8 + T cells and the effects of CD160 on the immunological activity of CD8 + T cells from the tumor-bearing mice.Methods CD8 +T cells were separated from C57BL/6 mice splenocytes with immunomagnetic beads and CD160 was detected by flow cytometry after mice were inoculated with TC-1 cervical cancer cells.The purified CD160 + CD8 + T cells population and CD160-CD8 + T cells population were obtained by fluorescence activated cell sorting (FACS),and then stimulated separately in vitro with heat shock protein(HSP)70-TC-1 peptide complexes in the context of CD8 + T-depleted splenocytes in the presence or absence of anti-herpesvirus entry mediator (HVEM,the high affinity ligand for CD160) antibody for 6 days.Flow cytometry was performed to measure the proliferation of carboxyfluorescein succinimidyl amino ester (CFSE) labeling-CD8 + T cells and the intracellular expression of perforin and γ-interferon (IFN-γ) in CD8 + T cells.Results On the week 3 after the inoculation with TC-1 cells,the positive percentageof CD160 + CD8 +T cells of tumor-bearing mice was significantly higher than that of control mice [(18.050 ± 7.944) ％ vs.(8.663 ± 3.921) ％,P ＜ 0.01].The proliferation percentage of CD160 + CD8 + T cells population was significantly lower than that of the CD160-CD8 + T cells population [(3.425 ± 2.352) ％ vs.(15.440 ± 6.673) ％,P ＜ 0.01] after the stimulation culture,and had a significant increase to (12.000 ± 5.286)％ (P ＜ 0.01) by blocking the interaction of CD160 and HVEM with anti-HVEM Ab.Meanwhile,the positive percentage of perforin and IFN-γ in CD160 + CD8 + T cells population was respectively lower than that of the CD160-CD8 + T cells population [(5.263 ± 2.752) ％ vs.(17.600 ± 7.203) ％,P ＜0.01,and (4.150 ±2.499)％ vs.(11.480 ±4.790)％,P ＜0.01],and increased significantly to (12.380 ± 4.922) ％ (P ＜ 0.01) and (11.280 ± 4.037) ％ (P ＜ 0.01) by anti-HVEM Ab.Conclusion The expression of CD160 is increased on CD8 + T cells from the tumor-bearing mice and related to the declined immunological activity of CD8 + T cells.The blockade of the interaction between CD160 and HVEM can reverse partly the inhibited activity and function of CD8 + T cells.