血红素氧化酶-1在鼠异种肝移植中的抗排斥作用及其机制研究

目的探讨血红素氧化酶1(HO1)在异种肝移植中的作用及其机制方法采用豚鼠对大鼠肝移植模型,实验动物分为3组,每组10对鼠。A组供体鼠术前24h腹腔注射5ml/kg生理盐水作为对照;B组供体术前24h腹腔注射HO1诱导剂钴原卟啉(CoPP),C组供体注射CoPP的同时给予HO1抑制剂锌原卟啉(ZnPP)。每组受体均接受眼镜蛇毒因子(CVF)。分别以RTPCR和western印迹法观察HO1mRNA及蛋白表达。通过凝胶电泳迁移率改变法(EMSA)检测移植肝核转录因子(NF)κB的活化,免疫组化观察E选择素表达,来评价HO1对内皮细胞活化的抑制作用。并比较各组受体鼠存活时间。结果各组均未发生超急性排斥反应。CoPP预处理可以诱导HO1mRNA和蛋白过度表达,进而显著抑制移植肝内NFκB的活化和内皮细胞E选择素表达;B组与(0.112±0.039)A组(0.211±0.030)、C组(0.283±0.075)相比差异有统计学意义(P<0.05)。HO1过度表达组与其它组相比,移植肝内皮细胞肿胀减轻,浸润细胞数减少,肝脏功能改善,而且受体鼠存活时间显著延长;B组(15.5h±3.8h)比A组(7.3h±2.1h)差异有统计学意义(P<0.01)。这种保护效应可被同时给予HO1抑制剂ZnPP所取消;B组比C组(6.7h±2.9h)差异有统计学意义(P<0.01)。结论HO1通过抑制内皮细胞活化,延缓异种移植肝排斥反应发生。

Effect of heme oxugenase-1 on delayed xenograft rejection: experiment of guinea pig-to-rat liver xenotransplantation

OBJECTIVE: To investigate the effects and mechanism of heme oxygenase-1 (HO-1) in liver xenotransplantation and mechanism thereof. METHODS: Thirty male guinea-pigs used as donors were injected intravenously with cobra venom factor (CVF) and then randomly divided into 3 groups 24 hours after: Group A injected intraperineally with NaCl, Group B injected intraperineally with cobalt-protoporphyrin (CoPP), heme oxygenase-1 inducer, and Group C injected intraperineally with CoPP and zinc protoporphyrin (ZnPP), HO-1 inhibitor zinc before their livers were harvested. Thirty male SD rats used as recipients underwent the above-mentioned treatment 24 hours before receiving the xenografts. Five pairs of guinea pigs and rats in each group underwent collection of blood and liver tissues 3 hours after the recovery of blood perfusion in the transplanted livers for detection of serum enzymes by biochemical methods and expression of HO-1 mRNA and protein in the transplanted livers by RT-PCR and Western blotting respectively. The other 5 pairs in each group were used to observe the survival time. RESULTS: The survival time of Group B was 15.5 h +/- 3.8 h, significantly longer than those of Group A (7.3 h +/- 2.1 h) and Group C (6.7 h +/- 2.9 h, both P < 0.01). The values of ALT and LDH of Group B were significantly lower than those of Group A and C (all P < 0.05). HOI-1 mRNA expression was not detected or only expressed in trace amount in the livers of normal guinea pigs, expressed in a small amount in the transplanted livers of Group A. The expression of HO-1 mRNA and that of HO-1 protein in the transplanted livers of Group B were significantly higher than those of Group A (both P < 0.01), and the expression of HO-1 mRNA and that of HO-1 protein in the transplanted livers of Group C were not significantly different from those of Group A (both P > 0.05). Remarkable NF-kB band was detected in Groups A and C, and only weak NF-kB band was seen in Group B. The E-selectin expression was significantly lower in the transplanted livers of Group B than in those of Group A and C (both P < 0.05). CONCLUSION: HO-1 delays the occurrence of delayed xenograft rejection in liver xenotransplantation. This effect depends, at least in part, on HO-1-mediated inhibition of endothelium activation in xenografts.