【作者】 李廷；

【导师】 张学光；

【作者基本信息】 苏州大学 ， 免疫学， 2004， 硕士

【摘要】 目的：初步探讨Graves病(GD)患者外周血CD4~+CD28~-和CD4~+CD28~+T细胞亚群TCR Vβ基因是否存在限制性选用及其相应的生物学特性。方法：运用real-time quantitative PCR(RQ-PCR)定量测定10例GD患者外周血CD4~+CD28~-和CD4~+CD28~+T细胞TCR Vβ的表达。运用流式细胞仪(FCM)检测CD4~+CD28~-和CD4~+CD28~+短期T细胞系胞内IFN-γ和IL-4的表达。运用~3H-TdR掺入试验观测激发型CD40单抗、阻断型CD40L单抗和阻断型B7-1单抗对CD4~+CD28~-和CD4~+CD28~+短期T细胞系与非毒性甲状腺肿(NTG)来源的甲状腺上皮细胞的增殖影响。结果：在10例GD患者CD4~+CD28~-T细胞亚群中，共出现12次Vβ基因的过高表达，而在8例GD患者(2例未测)CD4~+CD28~+T亚群中，共出现6次Vβ基因的过高表达。CD4~+CD28~-与CD4~+CD28~+亚群所选用的Vβ基因有部分不同，Vβ7和Vβ17在CD4~+CD28~+亚群中的表达水平高于CD4~+CD28~-亚群(分别为P＜0.05和P＜0.01)，而Vβ21、Vβ23和vβ25在CD4~+CD28~-亚群的表达水平高于CD4~+CD28~+亚群(分别为P＜0.05，P＜0.05和P＜0.01)。CD4~+CD28~-短期T细胞系产生的IFN-γ(27.9±15.9％)显著高于其自体CD4~+CD28~+短期T细胞系(10.3±12.1％，P＜0.001)；而IL-4的产生在CD4~+CD28~-和CD4~+CD28~+短期T细胞系中无明显统计学差异(分别为7.8±9.2％和13.5±18.7％，P＞0.05)。激发型CD40单抗能明显刺激CD4~+CD28~+T细胞系的增殖(P＜0.05)，但不影响CD4~+CD28~-T细胞系的增殖(P＞0.05)；阻断型CD40L单抗对两种T细胞系均有不同程度的阻断作用，以对CD4~+CD28~+T细胞系的阻断作用更明显(分别为P＜0.05和P＜0.01)；阻断型B7-1单抗仅能阻断CD4~+CD28~+T细胞系的增殖(P＜0.05)，而对CD4~+CD28~-T细胞系无明显影响(P＞0.05)。结论：TCR Vβ基因的限制性选用反映了两类T细胞亚群均存在寡克隆性增殖，其中CD4~+CD28~-T细胞亚群可能为Th1型T细胞，二者在GD的发病和活动中可能分别起不同的作用。激发型CD40单抗、阻断型CD40L单抗和阻断型B7-1单抗对两类T细胞亚群的增殖均有一定的调节作用。更多还原

【Abstract】 Objective: To evaluate if there is a restricted usage of TCR VJ3 gene segment in CD4+ CD28+ and CD4+CD28~ T cell subsets from peripheral blood with Graves disease, and the biological function of these two subsets. Methods: We detected the expression level of TCR Vp in CD4+CD28 + and CD4+CD28~T subset with real-time quantitative PCR(RQ-PCR) in ten Graves disease patients, the intracytoplasmic level of IFN-y and IL-4 in the two short-term cell lines using Flow Cytometer. Furthermore, the function of agonistic anti-CD40mAb, antagonistic anti-CD40LmAb and antagonistic anti-B7-lmAb to the interaction of thyrocyte with these two subsets also be primarily assessed by 3H-TdR incorporation. Results: There were overexpression of CD4+CD28-T cell subset in these 10 patients for 12 times, and only 6 times of CD4+CD28+T cell subset in these 8 patients(two patients not detected). The usage of V(3 in these two subsets is partially different. The intracytoplasmic level of IFN-y( 27.9 15.9% )in CD4+CD28-T cell subset show marked higher than that in CD4+CD28+T cell subset (10.3 + 12.1%, P<0.001) of the same patient, while the intracytoplasmic level of IL-4 in these two subset has no different significance (7.8 + 9.2% and 13.5 + 18.7% respectively, P>0.05) . The promoting proliferation effect of agonistic CD40mAb could be found in the CD4+CD28+T cell group rather than CD4+CD28-; the proliferation of both groups could be inhibited by antagonistic CD40LmAb; the antagonistic B7-lmAb could only restrain the proliferation of CD4+CD28+T cell group, but not CD4+CD28- group. Conclusion: The restricted usage of TCR Vp gene segment suggests that oligoclonal proliferation happened in both subsets, CD4+CD28-T cell subset maybe belong to Th1, and both of them probably playdifferent role in pathogenesis and activation of Graves disease. All of agonistic CD40mAb, antagonistic CD40LmAb and antagonistic B7-lmAb have relavative effects on these two T cell subsets.更多还原