自身肽结合于HLA-A2分子上能够在体外诱导抗原肽特异性的同种T细胞

在同种反应性T细胞(同种T细胞)识别的配体中,抗原肽的作用是免疫学长期争论的问题,即同种T细胞识别是否具有抗原肽特异性.为了证实通过长期混合淋巴细胞培养(LTMLC)方法能够诱生抗原肽/MHC复合物(pMHC)特异性的同种T细胞,本研究利用仅表达HLA-A2,TAP缺陷的T2细胞,将酪氨酸激酶来源的自身抗原肽(Tyr369-377)和EB病毒来源的病毒抗原肽(LMP2A426-434)分别加载到T2细胞上,使T2细胞提呈单一的T细胞抗原识别表位,并且选择4个HLA-A2阳性(HLA-A2+ve)与4个HLA-A2阴性(HLA-A2-ve)个体的PBL样本,与加载上述抗原肽的T2细胞混合培养.在此实验系统中,HLA-A2+ve PBL与加载病毒抗原肽的T2细胞(T2/LMP)混合培养代表T细胞对普通抗原的反应,而HLA-A2-ve PBL与加载自身抗原肽的T2细胞(T2/Tyr)混合培养则为T细胞对同种抗原的反应.利用特异性pMHC四聚体染色与特异性细胞毒试验检测LTMLC诱生CTL的特异性,其中利用HIV抗原肽(Gag77-85)作为对照.结果显示:(ⅰ)T2/LMP与HLA-A2+ve个体的PBL混合培养产生CTL(CTL-T2/LMP),CTL-T2/LMP对T2/LMP的杀伤显著高于对照T2/HIV的杀伤(26.52%±3.72%vs7.01%±0.87%,P<0.001);LMP四聚体对CTL-T2/LMP染色的阳性细胞数显著高于对照HIV四聚体(0.98%±0.33%vs0.05%±0.01%,P=0.0014);(ⅱ)加载自身抗原肽的T2细胞(T2/Tyr)可诱导HLA-A2-ve个体的PBL产生CTL(CTL-T2/Tyr),CTL-T2/Tyr对T2/Tyr的杀伤显著高于对T2/HIV的杀伤(28.07%±2.58%vs6.87±1.01%,P<0.001);Tyr四聚体对CTL-T2/Tyr染色的阳性细胞数显著高于HIV四聚体(0.88%±0.3%vs0.06±0.03%,P=0.0018).结果说明:结合于自身MHC分子上的病毒抗原肽与结合于同种MHC分子上的自身抗原肽都能诱导产生抗原肽特异性的CTL;在LTMLC诱生的同种CTL中,有相当数量的CTL具有pMHC特异性,这些同种CTL的识别机制与普通抗原反应性CTL一样,识别的对象也是特异性的pMHC.支持了同种抗原的pMHC种类繁多造成同种T细胞反应强度极高的假说.利用LTMLC诱生抗原肽特异性同种T细胞方法对于T细胞过继治疗具有潜在的应用价值.

The cross-reactivity of I-A^(g7) and I-A^d tetramers

OBJECTIVE: To investigate the cross-reactivity between glutamic acid decarboxylase (GAD)-I-A(g7) and I-A(d) tetramer in diabetes-prone non-obese diabetic (NOD) mice (I-A(g7)) and diabetes-free Balb/c mice (I-A(d)). METHODS: Two GAD peptide I-A(g7) and I-A(d) tetramers were generated and compared for phenotype and function of sorted GAD peptide I-A(g7) and I-A(d) tetramer-positive (tet+) T cells. RESULTS: The cross-reactivity is shown in either tetramer positive percentage or tetramer staining intensity. The NOD and Balb/c derived-tet+ T cells were able to be cross-stained by GAD peptide I-A(g7) and I-A(d) tetramers, and responded to both irradiated NOD and Balb/c splenotyes under stimulation by synthetic and recombinant GAD peptides. CONCLUSION: Although I-A(g7) and I-A(d) are closely related in biochemical and biological aspects, their most notable difference is the presence or absence of a negatively charged residue at position beta57 that links to insulin-dependent diabetes mellitus.

The cross-reactivity of I-Ag7 and I-Ad tetramers

To investigate the cross reactivity between glutamic acid decarboxylase (GAD) I A g7 and I A d tetramer in diabetes prone non obese diabetic (NOD) mice (I A g7 ) and diabetes free Balb/c mice (I A d) Methods Two GAD peptide I A g7 and I A d tetramers were generated and compared for phenotype and function of sorted GAD peptide I A g7 and I A d tetramer positive (tet +) T cells Results The cross reactivity is shown in either tetramer positive percentage or tetramer staining intensity The NOD and Balb/c derived tet + T cells were able to be cross stained by GAD peptide I A g7 and I A d tetramers, and responded to both irradiated NOD and Balb/c splenotyes under stimulation by synthetic and recombinant GAD peptides Conclusion Although I A g7 and I A d are closely related in biochemical and biological aspects, their most notable difference is the presence or absence of a negatively charged residue at position β57 that links to insulin dependent diabetes mellitus