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目的观察促甲状腺素受体A亚单位上的第52~71位(PA)及第67~86位(ID)氨基酸多肽对格雷夫斯病(GD)患者外周血单个核细胞(PBMC)增殖活性及细胞因子分泌的影响。方法选取GD患者13例、健康志愿者8例为研究对象,取其静脉血,分离PBMC。PBMC分为空白对照组(无细胞及抗原)、阴性对照组(不加抗原刺激)、阳性对照组(刀豆蛋白A 5μg·m L-1)、PA组(PA多肽20μg·m L-1)、ID组(ID多肽20μg·m L-1),均培养3 d。酶联免疫吸附法检测各组细胞上清液中干扰素γ(IFN-γ)及白细胞介素4(IL-4)水平,并计算各组增殖指数。结果健康志愿者和GD患者的阳性对照组IFN-γ、IL-4水平均显著高于阴性对照组(P<0.01),而PA组和ID组的IFN-γ、IL-4水平与阴性对照组比较无显著差异(P>0.05)。健康志愿者和GD患者的阳性对照组增殖指数均显著高于阴性对照组(P<0.01)。健康志愿者PA组和ID组增殖指数和阴性对照组相比无显著差异(P>0.05)。GD患者PA组增殖指数高于阴性对照组(P<0.01)和ID组(P<0.05),ID组增殖指数与阴性对照组比较无显著差异(P>0.05)。结论 PA多肽及ID多肽均为具有免疫活性的抗原肽,且PA多肽的激活作用更强,可作为肽疫苗的备选多肽。
Objective To observe the proliferation activity of peripheral blood mononuclear cells (PBMCs) in patients with Graves’ disease (GD) from the 52th to the 71st (PA) and the 67th to the 86th (ID) amino acids of the thyroid stimulating hormone receptor A subunit And cytokine secretion. Methods Thirteen GD patients and eight healthy volunteers were selected as the research object. The venous blood was collected and PBMCs were isolated. PBMCs were divided into blank control group (no cell and antigen), negative control group (without antigen stimulation), positive control group (concanavalin A 5μg · m L-1), PA group (PA polypeptide 20μg · m L-1 ), ID group (20μg · m L-1 ID polypeptide) were cultured for 3 days. The levels of interferon γ (IFN-γ) and interleukin-4 (IL-4) in supernatant of each group were detected by enzyme-linked immunosorbent assay. Proliferation index of each group was calculated. Results The levels of IFN-γ and IL-4 in the positive control group were significantly higher than those in the negative control group (P <0.01), while the levels of IFN-γ and IL-4 in the PA group and ID group were significantly lower than those in the negative control group There was no significant difference between the two groups (P> 0.05). The proliferation index of positive control group was significantly higher than that of negative control group (P <0.01) in healthy volunteers and GD patients. There was no significant difference between the proliferation index of PA group and ID group in healthy volunteers and negative control group (P> 0.05). The proliferation index of PA patients in GD group was higher than that in negative control group and ID group (P <0.05). There was no significant difference in proliferation index between ID group and negative control group (P> 0.05). Conclusion PA polypeptide and ID polypeptide are both immunocompetent antigen peptide, and the activation of PA polypeptide is stronger, which can be used as peptide vaccine candidate polypeptide.