术前输注供者CD80^(low)/CD86^(low)树突状细胞,延长小鼠同种异体移植心脏存活

Donor dendritic cells treated with B7-1,B7-2 antisense oligonucleotide prolonged mouse cardiac allograft survival

目的 :应用B7- 1和B7- 2反义寡核苷酸 (ASB7- 1/ASB7- 2oligo)抑制CD80 (B7- 1)、CD86 (B7- 2 )在供体小鼠骨髓树突状细胞 (DC)上的表达 ,观察这类DC对同种异体小鼠心脏移植存活时间的影响并探讨其机理。方法 :小鼠B7- 1和B7- 2反义寡核苷酸在lipofectamine协助下分别转染供体鼠C5 7BL/10J(B10 )小鼠骨髓DC ,流式细胞仪检测其CD80 /CD86的表达 ,证实为CD80 low/CD86 low。将各组DC经尾静脉输注到受体小鼠C3H/HeJ(C3H)体内 ,1周后进行心脏移植术 ,观察存活时间 ;体外实验观察各组DC对同种异体T细胞的激活作用 ,包括混合淋巴细胞反应、细胞毒性效应、及IL - 2的产生。结果 :ASB7- 1和ASB7- 2分别显著抑制DC表达CD80 /CD86 ;转输这些CD80 low/CD86 lowDC可使小鼠心脏移植物存活时间显著延长 ,分别为 (18.6± 0 .89)d和 (2 3.6 7± 10 .73)d ,与转输成熟骨髓DC(IL - 4DC)组和生理盐水注射组(6 .2 2± 0 .97)d、(11.17± 1.72 )d比较 ,均有显著差异 (P <0 0 1) ;CD80 low或CD86 lowDC对异体T细胞激活作用较弱 ,表现为T细胞增殖能力、IL - 2产生及细胞毒杀伤均明显低。结论 :应用反义寡聚核苷酸转染供者DC ,降低其CD80或CD86的表达 ,可以抑制供者特异性的免疫应答 ,延长移植物存活时间。

AIM:To investigate the effect of donor bone marrow derived dentritic cell (DC) treated with B7-1, B7-2 antisense oligonucleotide on mouse heart allografe survival time and its mechanism. METHODS: There were 7 groups of C57BL/10J (B10) mouse bone marrow DCs which were treated by 400 nM antisense oligonucleotide target to B7-1, B7-2 mRNA (AS B7-1/2), B7-1 mismatch oligo control ,B7-2 mismatch control(mASB7-1/2), lipofectamine only and non-treatment, respectively. Each group of DC were named as ASB7-1 DC, ASB7-2 DC, mASB7-1DC, mAS B7-2DC, and Lipo DC, respectively.RESULTS: Flow cytometer results shown that AS B7-1/2 can inhibit B7-1(CD80)and B7-2 (CD86) molecule express on DC surface, while control groups have no effects. To observe their tolerogenicity in mouse cardiac allograft model, B10→C3H heterotopic heart transplantation were performed. Recepients were received 2×106 of DC injection 7 days before transplantation. Results showed that both AS B7-1DC and AS B7-2 DC can prolong mouse cardiac allograft survival time to (18.6±0.89) days and (23.67±10.73) days, respectively, compared with IL-4 DC [(6.22 ±0.97) days(P<0.01)].Two mismatch control groups can slightly prolong while oligo DC has no effect. For understanding its mechanism, each group of DC was used as stimulator to stimulated C3H spleen T cell. Results suggested that AS B7-1DC and AS B7-2 DC had less allo-stimulate function, including MLR and generation CTL and IL-2 production than IL-4 DC but control groups have no effect.CONCLUSION: Donor bone marrow derived DC treated with AS B7-1 oligo and AS B7-2 oligo expressed lower level of CD80 and CD86, respectively. These cells can induce allogeneic T cells anergy in vitro and markedly prolong mouse heart allograft survival time in vivo.