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目的探讨基于CD4立体构型设计的新型免疫抑制剂(J2)在小鼠体内的免疫抑制作用以及其作用机制。方法将羧基荧光素二醋酸琥珀酰亚胺酯(CFSE)标记的C57BL/6(H-2b)小鼠的淋巴细胞4×10~6～6×10~6经尾静脉注入经~(60)钴照射(900拉德)后的DBA/2(H-2d)小鼠体内构建模型。将DBA/2小鼠随机分为5组,每组5只,模型建立后即刻腹腔注射J2,每天1次。对照组注射生理盐水、CsA组注射环孢素A(10mg/kg体重);实验组J2A组(1mg/kg体重)、J2B组(4 mg/kg体重)、J2C组(8mg/kg体重)。给药3d后分组处死DBA/2小鼠取脾制备淋巴细胞,用CD3、CD4、CD8单抗分别标记受检T细胞,FACS流式细胞仪检测CD4~+T细胞和CD8~+T细胞在小鼠体内分裂增殖的情况,并检测CD4~+T细胞的凋亡情况。结果CFSE标记C57BL/6小鼠的脾淋巴细胞着染率大于99%。对照组分裂的CD4~+ T细胞(75.4±1.58)%、分裂的CD8~+ T细胞(83.48±1.25)%明显多于CsA组和实验组(P<0.01)。J2B组和J2C组分裂的细胞数分别与CsA组比较差异无统计学意义(P>0.05),但J2A组多于CsA组(P<0.05)。对照组CD4~+ T细胞早期凋亡的比例(41.1±3.4)%明显高于其他各组(P<0.01)。J2A组(35.6±4.1)%、J2B组(24.0±3.7)%和J2C组(13.6±2.3)%CD4~+ T细胞早期凋亡的比例显著高于CsA组(7.4±1.9)%(P<0.05)。结论J2可能抑制T细胞亚群CD4~+ T细胞的活化,从而进一步抑制CD8~+ T细胞的分裂增殖而具有免疫抑制作用。
Objective To investigate the immunosuppressive effects of a novel immunosuppressive agent (J2) based on the CD4 stereoconfiguration in mice and its mechanism of action. Methods The lymphocytes from 4 × 10 ~ 6 ~ 6 × 10 ~ 6 of CFSE-labeled C57BL / 6 (H-2b) DBA / 2 (H-2d) mice were irradiated with cobalt (900 rad) in vivo. DBA / 2 mice were randomly divided into 5 groups with 5 mice in each group. J2 was injected intraperitoneally immediately after the model was established. The rats in the control group were injected with normal saline. The CsA group was injected cyclosporine A (10mg / kg body weight); the J2A group (1mg / kg body weight), the J2B group (4mg / kg body weight) and the J2C group (8mg / kg body weight). After 3 days of administration, the lymphocytes were prepared from the spleens of DBA / 2 mice and the T cells were labeled with CD3, CD4 and CD8 monoclonal antibodies respectively. The percentage of CD4 ~ + T cells and CD8 ~ + T cells were detected by FACS Mice in vivo proliferation and proliferation, and detect CD4 ~ + T cell apoptosis. Results CFSE-labeled C57BL / 6 mice had lymphocyte staining rates> 99%. In the control group, CD4 ~ + T cells (75.4 ± 1.58)% and CD8 ~ + T cells (83.48 ± 1.25%) were significantly more than CsA group and experimental group (P <0.01). There were no significant differences in the number of cells split between J2B and J2C groups compared with CsA group (P> 0.05), but more in J2A group than CsA group (P <0.05). The percentage of early apoptotic CD4 ~ + T cells in control group (41.1 ± 3.4%) was significantly higher than that in other groups (P <0.01). The percentage of early apoptosis in J2A group (24.6 +/- 3.7)% and J2C group (13.6 +/- 2.3)% CD4 + T cells was significantly higher than that in CsA group (7.4 +/- 1.9)% (P < 0.05). Conclusions J2 may inhibit the activation of CD4 ~ + T cells in T cell subsets and further inhibit the proliferation and proliferation of CD8 ~ + T cells.