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目的研究多肽类抗原-超抗原金黄色葡萄球菌肠毒素B(staphylococcal enterotoxin B,SEB)活化的NKT细胞亚群及耐受特征。方法小鼠脾细胞分别与SEB和ConA体外培养,MTT方法测定细胞增殖;特异性耐受特征的研究使用体外活化第3天的细胞,吸出上清后添加二次抗原ConA、LPS、IL-2,继续培养3 d,MTT方法测定细胞对二次抗原的应答反应能力。用流式细胞测定法解析SEB和ConA体外活化的淋巴细胞在第0、5、10和15天时的T和NKT淋巴细胞亚群。结果SEB和ConA均诱导了小鼠淋巴细胞在体外增殖;SEB活化的淋巴细胞对ConA、LPS和IL-2的二次应答反应能力消失,细胞增殖的OD570nm值由一次应答反应的0.433±0.07分别下降到0.19±0.01、0.14±0.02和0.15±0.04(P<0.01)。ConA活化的淋巴细胞的二次应答反应依然存在并依赖IL-2。SEB活化的淋巴细胞是CD4+NK1.1+和CD8+NK1.1+NKT细胞,而不是CD4-CD8-NKT细胞。ConA活化的淋巴细胞是CD3+、CD4+和CD8+T淋巴细胞并包括了CD4-CD8-/CD3+NK1.1+NKT细胞。结论除了α-Calcer脂多糖类抗原能够活化NKT细胞,多肽类超抗原SEB也能活化NKT细胞并具有特异性免疫耐受功能。能够介导免疫耐受的NKT细胞亚群可能是SEB活化的CD4+、CD8+NKT细胞而不是CD4-CD8-/CD3+NK1.1+NKT细胞。
Objective To study the subsets and tolerance of NKT cells activated by peptide antigen - superantigen staphylococcal enterotoxin B (SEB). Methods The spleen cells of mice were cultured in vitro with SEB and ConA respectively. The cell proliferation was measured by MTT assay. The cells were cultured in vitro for 3 days. After the supernatant was aspirated, the secondary antigens ConA, LPS and IL-2 , Continue to cultivate 3 d, MTT method to determine the cell response to secondary antigen response ability. Flow cytometry was used to resolve T and NKT lymphocyte subsets on days 0, 5, 10, and 15 of SEB and ConA activated lymphocytes in vitro. Results Both SEB and ConA induced the proliferation of mouse lymphocytes in vitro. The ability of SEB-activated lymphocytes to respond to ConA, LPS and IL-2 disappeared. The OD570nm value of cell proliferation increased from 0.433 ± 0.07 Decreased to 0.19 ± 0.01, 0.14 ± 0.02 and 0.15 ± 0.04 (P <0.01). Secondary response to ConA-activated lymphocytes is still present and dependent on IL-2. SEB-activated lymphocytes are CD4 + NK1.1 + and CD8 + NK1.1 + NKT cells, but not CD4-CD8-NKT cells. ConA activated lymphocytes are CD3 +, CD4 + and CD8 + T lymphocytes and include CD4-CD8- / CD3 + NK1.1 + NKT cells. Conclusion Except that α-Calcer lipopolysaccharide antigen can activate NKT cells, SEB can activate NKT cells and has specific immune tolerance function. The subset of NKT cells capable of mediating immune tolerance may be SEB-activated CD4 +, CD8 + NKT cells but not CD4-CD8- / CD3 + NK1.1 + NKT cells.