目的: 研究淫羊藿水溶性提取物对刀豆蛋白A(ConA)刺激的小鼠T淋巴细胞体外早期活化和增殖的影响。方法: 无菌分离小鼠各部位淋巴结, 制备单细胞悬液, 加入不同终质量浓度(5、25、50mg/L)的淫羊藿提取物, 以MTT法检测药物对T细胞活性的影响; 利用流式细胞术(FCM)结合双色荧光抗体染色技术检测早期活化标志CD69分子的表达; 运用流式细胞术结合活体染料CFDA-SE染色技术和荧光抗体染色技术检测T细胞增殖。结果: MTT法检测淫羊藿提取物在50mg/L时细胞存活率达52.36%。 ConA作用6h后, 对照组CD69⁺表达率为(4.61±0.85)%, ConA组CD69+T细胞的比率上升至(38.30±2.54)%, 淫羊藿提取物在终质量浓度5、25、50mg/L时均抑制ConA诱导的CD3⁺T淋巴细胞CD69的表达(P<0.01), 其表达率分别下调为(36.16±2.14)%, (30.5±1.72)%, (15.99±0.87)%。培养48、72后, 对照组增殖指数(PI)分别为1.01±0.01和1.05±0.01, ConA组PI分别达到1.42±0.01、2.32±0.01, 淫羊藿提取物在终质量浓度5、25、50mg/L均抑制T淋巴细胞增殖(P<0.01), 48h时PI分别为1.28±0.01、1.26±0.01、1.21±0.01;72h时PI分别为2.30±0.03、2.13±0.02、2.10±0.01。结论: 淫羊藿提取物能明显抑制ConA刺激的小鼠T淋巴细胞的早期活化和增殖, 并呈剂量依赖性。

Objective: To study the effect of water extract from Herba Emidemii on the early activation and proliferation of mouse T lymphocytes stimulated by concanvalin A(Con A)in vitro. Methods: Single cell suspension was prepared from mouse lymphoid nodes in germ free condition. The drug toxicity of water extract from Herba Emidemii(5, 25, and 50 mg/L)on T lymphocytes was measured by MTT. The expression of CD69, the marker of early activation of T lymphocytes was measured by flow cytometry (FCM)combined with two-colour immunofluorescent staining of eell surface antigen. The proliferation of T lymphocytes was measured by FCM combined with carboxyl fluorescin diacetate succinmidyl ester(CFDA-SE)staining and immunofluorescent staining. Results: The survival ratio of T lymphocytes in 50 mg/L was 52.36%. After 6h culture, the expression rate of CD69⁺ Tcellin control group was(4.61士0.85)%, that in Con A group was(38.30士2.54)%, water extrat from Herba Emidmii(5, 25, and 50 mg/L)inhibited the expression of CD69 in CD3+T lymphocytes stimulated by Con A(P<0.01), the expression rates were(36.16±2.14), % (30.5士1.72)%, and(15.99士O.87)%, respectively. After 48 and 72 h culture, the proliferation index(PI)of control group were 1.01±O.01 and 1.05士O.01, of Con A group were 1.42士O.01 and 2.32±0.01, respectively. Water extract from Herba Emidmii(5, 25, and 50 mg/L)inhibited the proliferation of T lymphocytes(P<0.01), the PI of 48h were 1.28土O.01, 1.26土O.01, 1-21±O.01；the PI of 72h were 2.30士O.03, 2.13士0.02, 2.10土O.01, respectively. Conclusion: Water extract from Herba Emidmii could significantly inhibit Con A-induced early activation and proliferation of mouse T lymphocytes in vitro in a dose dependent manner.