Mechanism of liver ischemia-reperfusion injury with monoclonal antibodies of adhesin molecules and its application to organ transplantation

为探讨中性粒细胞(PMN)在肝脏缺血再灌流和内毒素血症中是否参与细胞损伤，采用Wistar大鼠肝脏热缺血再灌流模型，用动脉夹闭供肝正中叶和左外叶的血管90min。阻断前5min静脉注射抗ICAM-1(IA29)、CD11a(WT-1)、CD18(WT-3)的单抗或PBS作为安慰剂。再灌流后观察肝脏中性粒细胞的浸润及ICAM-1的表达。以肝组织中三磷酸腺苷(ATP)和丙二醛(MDA)作为肝细胞损伤的指标，观察单抗治疗的效果。至再灌流后24小时，PMN积累量持续增加。肝组织ICAM-1表达在再灌流后4周明显增强。单抗治疗可抑制PM…的渗入再灌流后6小时增加生理盐水，6、12和24小时恢复ATP合成，12小时降低肝组织丙二醛水平。ICR小鼠腹腔注射内毒素血症脂多糖，同时静脉注射1 mg/kg抗白细胞黏附分子单抗(抗CD11a：KAB，抗CD11b：MI/70，抗CD18；C17/16，抗ICAM-1：KAT-1，抗LECAM-1：MEL-14)，30 mg/kg甲基强的松龙(MP)作为常规休克药物，或PBS作为安慰剂。安慰剂组给药后48周存活率为36%(20/55)。经抗CD11a、抗CD18、抗LECAM-1和MP治疗后，存活率分别提高到70(7/10)、62(8/13)、(7/11)和100%(11/11)。而抗CD11b和抗ICAM-1对生存率无明显影响。用抗CD18单抗进行流式细胞仪检测发现，在30min内，粒细胞表面CD18的表达增加了12倍，而MP对其的抑制作用持续到3h。4h后肝组织丙二醛含量由未处理组的0.50升至2.46nmol/mg蛋白，抗CD18单抗和MP分别抑制其含量至1.80和1.41nmol/mg蛋白。提示白细胞在肝缺血再灌注和内毒素血症下的氧化损伤中起重要作用。这些单抗可以是预防这种损伤的治疗剂。较少

The present study is to determine whether neutrophils (PMNs) contribute to the cellular injuries under hepatic ischermia/reperfusion and endotoxemia.Hepatic warm ischemia/reperfusion-Mate Wistar rats were used and the vessels supplying the median and the left lateral hepatic lobes were occuluded with arterial clamp for 90 min. Five minutes before the occulusion, monoclonal anitibody (mAb) against ICAM-1 (IA29), CD11a (WT-1), CD18 (WT-3) or PBS as the placebo was administered intravenously. After reperfusion, the infiltration of PMNs and the expression of ICAM-1 in the liver were examined histologically. To examine the effect of mAb treatment, hepatic adenosine triphosphate (ATP) and malondialdehyde (MDA) levels were determined as indices of hepatic cellular injury. The number of accumulated PMNs increased continuously up to 24 hours after reperfusion. The expression of ICAM-1 in the liver was enhanced 4 houres after reperfusion. Treatment with the mAbs suppressed the infiltration of PM … More NS by 6 hours, ATP resynthesis by 6,12 and 24 hours, and reduced hepatic MDA level by 12 hours after reperfusion. Then, these mAbs increased the survival rate of rats receiving total hepatic ischemin (90min) / reperfusion.Endotoxemia-Lipopolysaccharide (LPS : E.coli) was administered to ICR mice intraperitoneally, and lmg/kg antileukocyte adhesion molecule mAb (anti-CD11a : KAB,anti-CD11b : MI/70, anti-CD18 ; C17/16, anti-ICAM-1 : KAT-1, anti-LECAM-1 : MEL-14), 30mg/kg methylprednisolone (MP) as a conventional agent for shock, or PBS as the placebo was administered intravenously, simultancously. In the placebo group, the survival rate of mice 48 houres after LPS administration was 36% (20/55). Treatment with anti-CD11a, antiCD18, anti-LECAM-1 and MP increased the survival rate to 70 (7/10), 62 (8/13), 64 (7/11) and 100% (11/11), respectively. ON the other hand, anti-CD11b and anti-ICAM-1 had no significant effect on the survial rate. FACS analysis using the anti-CD18 mAb revealed that the expression of CD18 on the surfaces of granulocytes increased 12 folds within 30 min, and MP suppressed it significantly for up to 3 hours after LPS administration. The hepatic MDA content increased from 0.50 (untreated mice) to 2.46 nmol/mg protein 4 hours after LPS administration, and anti-CD18 mAb and MP suppressed it to 1.80 and 1.41 nmol/mg protein, respectively.Leukocytes are concluded to mediate significant roles for oxidative injury causing hapatic cellular damages under hepatic ischemia/reparfuion and endotoxemia. These mAbs can be therapeutic agents preventing such injuries. Less