大环内酯类抗生素对大鼠肝移植后缺血再灌注损

背景：肝移植是终末期肝病和肝衰竭的标准治疗方法，而缺血再灌注损伤能影响肝移植成功率。当有限的肝脏供体可供移植时，如何降低肝缺血再灌注损伤成了肝移植的首要问题。

目的：探究大环内酯类抗生素对大鼠肝移植后肝缺血再灌注损伤的作用及机制。

方法：构建自体原位肝移植大鼠模型，将Wistar 大鼠随机分为大环内酯类抗生素组和对照组。大环内酯类抗生素组大鼠在肝切除前30 min，采用大环内酯类抗生素(含60 mg/kg 罗红霉素、20 mg/kg 克拉霉素和40 mg/kg 红霉素)混合液预处理供体肝脏，在原位肝移植时向门静脉中推注上述大环内酯类抗生素混合液；对照组大鼠在肝切除前30 min，同体积生理盐水预处理供体肝脏，在原位肝移植时向门静脉中推注同体积的生理盐水。肝移植后连续7 d 观察大鼠存活率；检测肝移植后48，72 h 大鼠血清天冬氨酸氨基转移酶和丙氨酸氨基转移酶活性；苏木精-伊红染色和免疫组化检测大鼠肝组织形态学改变和Ki-67 阳性细胞数；TUNEL 法和蛋白质免疫印迹分别检测大鼠肝细胞凋亡数量和caspase-3、cleaved caspase-3 蛋白表达水平；免疫荧光和ELISA 法分别检测肝移植后大鼠肝组织中Kupffer 细胞数量改变和白细胞介素6、肿瘤坏死因子α和白细胞介素1β 水平。

结果与结论：大环内酯类抗生素提高了肝移植大鼠的整体存活率，改善了移植肝脏的功能失调，减轻了肝移植后缺血再灌注的损伤程度，提高了移植肝脏的再生能力，降低了移植肝组织中凋亡细胞数和凋亡蛋白cleaved caspase-3/ caspase-3 比值，降低了移植肝组织中Kupffer 细胞数量和白细胞介素6、肿瘤坏死因子α和白细胞介素1β 水平。以上结果提示，大环内酯类抗生素对大鼠肝移植后的肝缺血再灌注损伤起保护作用。

海南省自然科学基金资助项目(SQ2015ZRJJ0364)，项目负责人：常顺伍

BACKGROUND:Liver transplantation is the standard treatment for end-stage liver disease and liver ,ischemia-reperfusion injury can reduce the success rate of liver a limited number of liver donors are available for transplantation,how to reduce liver ischemia-reperfusion injury has become the primary issue in liver transplantation.

OBJECTIVE:To evaluate the effect of macrolide antibiotics on ischemia-reperfusion injury after liver transplantation in rats.

METHODS:Rat autologous orthotopic liver transplantation model was rats were randomly divided into macrolide antibiotics group and control group.In the macrolide antibiotics group,the donor liver was treated with macrolide antibiotics (60 mg/kg roxithromycin,20 mg/kg clarithromycin and 40 mg/kg erythromycin) 30 minutes before hepatectomy,and the above macrolide antibiotic mixture was injected into the portal vein immediately after orthotopic liver the control group,rats were pretreated with the same volume of saline for 30 minutes before hepatectomy,and the same volume of saline was injected into the portal vein immediately after orthotopic liver survival rate of the rats was observed within 7 days after liver serum aspartate aminotransferase and alanine aminotransferase activities were detected by automatic biochemical analyzer at 48 and 72 hours after liver staining and immunohistochemistry assay were used to detect the morphological changes of liver tissues and the number of Ki-67 positive cells in liver transplantation and western blot assay were used to detect the number of apoptotic hepatocytes and the expression of caspase-3 and cleaved caspase-3 proteins in liver transplantation rats, Kupffer cell number changes and levels of interleukin-6,interleukin-1β,and tumor necrosis factor-α in rat liver tissues after liver transplantation were detected by immunofluorescence and ELISA,respectively.

RESULTS AND CONCLUSION:Macrolide antibiotics increased the overall survival rate of liver transplanted rats,improved the dysfunction of transplanted liver,reduced the severity of ischemia-reperfusion injury after liver transplantation,increased the regenerative capacity of transplanted liver,reduced the number of apoptotic cells and the ratio of cleaved caspase-3/caspase-3 in the transplanted liver tissue,and decreased the number of Kupffer cells and the levels of interleukin-6,interleukin-1β and tumor necrosis factor-α in the transplanted the results indicate that macrolide antibiotics protect against ischemia-reperfusion injury in rats undergoing liver transplantation.

0 引言 Introduction

尽管为寻找新的治疗肝衰竭的方法做出了巨大努力，但肝移植仍然是终末期肝病和急性肝衰竭的标准治疗方 法[1]，而缺血再灌注损伤(ischemia/reperfusion injury，IRI)的存在直接降低了肝移植的成功率。当有限的肝脏供体可供移植时，如何解决或降低肝IRI成了肝移植的首要的问 题[2]。IRI级联是多个步骤的结果：最初，缺血会造成肝供体线粒体损伤并导致氧化应激和细胞损伤[3]；植入后，再灌注会诱导肝供体促炎细胞因子的释放和活性氧的产生，这会进一步损害肝细胞[4]；这些因素的组合可导致肝脏中微循环障碍和血栓形成，并最终导致肝组织坏死[4-6]；此外，由于边缘供体移植物对缺血性损伤的敏感性较高，IRI导致缺乏适合移植的肝脏[7]。因此，减少IRI的疗法可能会增加合适供体肝脏的储备并降低等待肝移植的患者的死亡率。在肝移植手术中，冷藏后缺血性损伤和再灌注损伤仍然是移植肝功能受损和移植肝脏再衰竭的主要危险因素[8-9]。虽然，研究人员进行了大量关于限制肝供体冷缺血时间方面的研究工作[10]，然而，随着可用移植器官的需求增加，冷缺血期通常会延长，这通常导致移植时肝脏质量降低。临床已经进行了各种研究发掘最小化IRI的药物[11]，但该问题仍未得到妥善解决。因此，关于该问题的改进仍需要进一步探究。

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