肝窦内皮细胞HO-1表达促进小鼠肝脏缺血再灌注损伤后再生重建的机制研究

摘要/Abstract

摘要： 目的探讨肝窦内皮细胞(LSEC)血红素加氧酶-1(HO-1)表达促进小鼠肝脏缺血再灌注损伤后再生重建的机制。方法 60只小鼠分为假手术组、模型组、空载组、HO-1转染组,每组15只,除假手术组仅做麻醉开腹操作外,其余3组均使用经典肝脏70%热缺血方法构建小鼠肝脏缺血再灌注损伤模型,再灌注后空载组经门静脉注射200 μL空载体病毒转染的LSEC,HO-1转染组经门静脉注射200 μL CD31标记的2×10⁶个/mL HO-1转染的LSEC。比较各组小鼠肝功能、血清及肝组织炎症因子、氧化应激因子水平、肝组织内肝细胞生长因子(HGF)、凋亡蛋白、Nrf2/HO-1信号通路蛋白表达量变化及肝细胞凋亡情况。结果携带HO-1目的基因的慢病毒的HO-1 mRNA表达量显著高于空载组(P<0.05)。模型组、空载组和HO-1转染组的ALT和AST水平高于假手术组,HO-1转染组的ALT和AST水平低于模型组和空载组(P<0.05);模型组和空载组比较差异无统计学意义(P>0.05)。模型组、空载组和HO-1转染组血清及肝组织的TNF-α、IL-1β、IL-6和MDA水平高于假手术组,SOD水平降低(P<0.05);HO-1转染组血清及肝组织的TNF-α、IL-1β、IL-6和MDA水平低于模型组和空载组,SOD升高(P<0.05);模型组和空载组比较差异无统计学意义(P>0.05)。模型组、空载组和HO-1转染组肝细胞病理变化程度和凋亡数高于假手术组,HO-1转染组低于模型组和空载组(P<0.05);模型组和空载组比较差异无统计学意义(P>0.05)。模型组、空载组和HO-1转染组肝组织的Bax和Caspase-3水平高于假手术组,HGF和Bcl-2降低(P<0.05);HO-1转染组肝组织的Bax和Caspase-3水平低于模型组和空载组,HGF和Bcl-2升高(P<0.05);模型组和空载组比较差异无统计学意义(P>0.05)。模型组、空载组和HO-1转染组肝组织的Nrf2和HO-1水平低于假手术组(P<0.05),HO-1转染组肝组织的Nrf2和HO-1水平高于于模型组和空载组(P<0.05);模型组和空载组比较差异无统计学意义(P>0.05)。结论肝脏缺血再灌注损伤移植HO-1转染的LSEC可减轻肝脏氧化应激、炎症反应,减少肝细胞凋亡,促进肝再生重建,恢复肝功能,其机制可能与Nrf2/HO-1信号通路激活有关。

关键词: 肝脏缺血再灌注损伤, 肝窦内皮细胞, 血红素加氧酶-1, 再生重建

Abstract: Objective To investigate the mechanism of heme oxygenase-1 (HO-1) expression in hepatic sinusoidal endothelial cells promoting liver regeneration and reconstruction after ischemia-reperfusion injury in mice. Methods Sixty male mice were randomly divided into a sham surgery group, a model group, an empty vector group, and a HO-1 transfection group of 15 mice each. Except for the sham surgery group, which only underwent anesthesia and open surgery, the other three groups used the classic liver 70% warm ischemia method to establish a mouse model of liver ischemia-reperfusion injury. After reperfusion, the empty vector group was injected with 200 μL of empty vector virus transfected LSECs via the portal vein, while the HO-1 transfection group was injected with 200 μL of CD31 labeled 2×10⁶/mL HO-1 transfected LSECs via the portal vein. Evaluate liver function indicators [alanine aminotransferase (ALT), aspartate aminotransferase (AST)], serum and liver tissue inflammatory factors [tumor necrosis factor] among different groups of mice- α (TNF- α), Interleukin-1 β (IL-1β), changes in levels of interleukin-6 (IL-6), oxidative stress factors (superoxide dismutase (SOD), malondialdehyde (MDA), hepatocyte growth factor (HGF), apoptotic proteins Bax, Bcl-2, Caspase-3, and nuclear factor E2 related factor 2 (Nrf2) and HO-1 protein expression in the Nrf2/HO-1 signaling pathway were observed in liver tissue. Pathological changes in liver tissue and liver cell apoptosis rate were also observed. Methods RT-PCR assay showed that the mRNA expression of HO-1 in lentivirus carrying HO-1 target gene was significantly higher than that in no-load group (P<0.05). ALT and AST levels in model group, no-load group and HO-1 transfection group were higher than those in sham operation group, while ALT and AST levels in HO-1 transfection group were lower than those in model group and no-load group (P<0.05). There was no difference between model group and no-load group (P>0.05). The levels of TNF-α, IL-1β, IL-6 and MDA in serum and liver tissue of model group, no-load group, and HO-1 transfection group were higher than those of sham operation group, and the level of SOD was decreased (P<0.05). The levels of TNF-α, IL-1β, IL-6 and MDA in serum and liver tissue of HO-1 transfection group were lower than those of model group and no-load group, and SOD was increased (P<0.05). There was no difference between model group and no-load group (P>0.05). The pathological changes and apoptosis rate of hepatocytes in model group, no-load group and HO-1 transfection group were higher than those in sham operation group, while those in HO-1 transfection group were lower than those in model group and no-load group (P<0.05). There was no difference between model group and no-load group (P>0.05). The levels of Bax and Caspase-3 in liver tissues of model group, no-load group, and HO-1 transfection group were higher than those of sham operation group, while HGF and Bcl-2 were decreased (P<0.05). The levels of Bax and Caspase-3 in liver tissue of HO-1 transfection group were lower than those of model group and no-load group, while HGF and Bcl-2 were increased (P<0.05). There was no difference between model group and no-load group (P>0.05). The levels of Nrf2 and HO-1 in liver tissue of model group, no-load group and HO-1 transfection group were lower than those of sham operation group (P<0.05), and the levels of Nrf2 and HO-1 in liver tissue of HO-1 transfection group were higher than those of model group and no-load group (P<0.05). There was no difference between model group and no-load group (P>0.05). Conclusion The transplanted HO-1-transfected LSECs with liver ischemia and reperfusion injury can reduce liver oxidative stress and inflammation, reduce hepatocyte apoptosis, promote liver regeneration and reconstruction, and restore liver function, and the mechanism may be related to the activation of Nrf 2/HO-1 signaling pathway.

Key words: Liver ischemia-reperfusion injury, Liver sinusoidal endothelial cells, Heme oxygenase-1, Liver regeneration

李鹏程, 王路兵, 朱荣华, 龚庆豪. 肝窦内皮细胞HO-1表达促进小鼠肝脏缺血再灌注损伤后再生重建的机制研究[J]. 肝脏, 2025, 30(1): 122-127.

LI Peng-cheng, WANG Lu-bing, ZHU Rong-hua, GONG Qing-hao. Mechanism of HO-1 expression in hepatic sinusoidal endothelial cells promoting liver regeneration and reconstruction after ischemia-reperfusion injury in mice[J]. Chinese Hepatolgy, 2025, 30(1): 122-127.

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参考文献

[1] ItoY, Hosono K, Amano H. Responses of hepatic sinusoidal cells to liver ischemia-reperfusion injury[J]. Front Cell Dev Biol, 2023,11(1):1171317.
[2] MachadoIF, Palmeira CM, Rolo AP. Preservation of mitochondrial health in liver ischemia/reperfusion injury[J]. Biomedicines, 2023,11(3):948.
[3] 钱子冰,张琦,曾佩芸,等. 肝窦内皮细胞-肝脏的守门员[J]. 海南医学院学报,2022,28(4):315-320.
[4] 姜婷婷,胡媛,曹建平. 肝窦内皮细胞在肝损伤中的功能研究进展[J]. 中国血吸虫病防治杂志,2023,35(1):92-97.
[5] WangT, Fang Y, Zhang X, et al. Heme oxygenase-1 alleviates ischemia-reperfusion injury by inhibiting hepatocyte pyroptosis after liver transplantation in rats[J]. Front Biosci (Landmark Ed), 2023,28(10):275.
[6] TianX, Wang Y, Yuan M, et al. Heme Oxygenase-1-Modified BMMSC activate AMPK-Nrf2-FTH1 to reduce severe steatotic liver ischemia-reperfusion injury[J]. Dig Dis Sci, 2023,68(11):4196-4211.
[7] 戴伟明,陆伦根,蔡晓波. 肝窦内皮细胞与肝纤维化的关系[J]. 临床肝胆病杂志,2023,39(2):419-423.
[8] 房志强,阮柏,刘晶晶,等. 肝血窦内皮细胞介导的他莫昔芬对非酒精性脂肪性肝炎治疗作用的转录组学分析[J]. 现代生物医学进展,2022,22(9):1609-1613.
[9] 罗昕,蔡晓波,陆伦根. 肝窦内皮细胞介导的细胞串扰在肝纤维化中的作用[J]. 肝脏,2022,27(12):1263-1265.
[10] 孙睿,李涛,任万华. 肝窦内皮细胞损伤在肝窦阻塞综合征发生进程中的作用:聚焦免疫炎性机制的研究进展[J]. 中华肝脏病杂志,2024,32(3):279-283.
[11] LiX, Wu L, Tian X, et al. miR-29a-3p in exosomes from heme oxygenase-1 modified bone marrow mesenchymal stem cells alleviates steatotic liver ischemia-reperfusion injury in rats by suppressing ferroptosis via iron responsive element binding protein 2[J]. Oxid Med Cell Longev, 2022,22(1):6520789.
[12] LiJJ, Dai WQ, Mo WH, et al. Fucoidan ameliorates ferroptosis in ischemia-reperfusion-induced liver injury through Nrf2/HO-1/GPX4 activation[J]. J Clin Transl Hepatol, 2023,11(6):1341-1354.
[13] ZuoH, Wang Y, Yuan M, et al.Small extracellular vesicles from HO-1-modified bone marrow-derived mesenchymal stem cells attenuate ischemia-reperfusion injury after steatotic liver transplantation by suppressing ferroptosis via miR-214-3p[J]. Cell Signal, 2023,109(1):110793.
[14] 袁博,明廷凤,瞿思铭,等. 血红素加氧酶-1对肝窦内皮细胞增殖及促肝再生表型的影响[J]. 中华肝胆外科杂志,2022,28(7):536-541.
[15] 马晓飞,焦彦如,张俊杰. 栀子苷对肝脏缺血再灌注损伤大鼠炎症反应、氧化应激和PI3K/Akt信号通路的影响[J]. 现代药物与临床,2023,38(1):29-36.
[16] 邓银芝,倪利华. 肝脏缺血再灌注损伤的机制及防治策略的研究进展[J]. 微循环学杂志,2023,33(2):109-115.
[17] XiaoX, Liu D, Chen S, et al. Sevoflurane preconditioning activates HGF/Met-mediated autophagy to attenuate hepatic ischemia-reperfusion injury in mice[J]. Cell Signal, 2021,82(1):109966.
[18] SalehH, El-Shorbagy HM. Chitosan protects liver against ischemia-reperfusion injury via regulating Bcl-2/Bax, TNF-α and TGF-β expression[J]. Int J Biol Macromol, 2020,164(1):1565-1574.
[19] MorsyMA, Ibrahim YF, Abdel Hafez SMN, et al. Paeonol attenuates hepatic ischemia/reperfusion injury by modulating the Nrf2/HO-1 and TLR4/MYD88/NF-κB signaling pathways[J]. Antioxidants (Basel), 2022,11(9):1687.
[20] YounisNS, Abdelnaby RM, Mohamed ME. Hepatoprotective effects of linalool against liver ischemia-reperfusion: the role of Nrf2/HO-1/NQO1 and TLR4/RAGE/NFκB pathways[J]. Eur Rev Med Pharmacol Sci, 2023,27(20):10094-10111.
[21] TaoT, Ye B, Xu Y, et al. β-Patchoulene preconditioning protects mice against hepatic ischemia-reperfusion injury by regulating Nrf2/HO-1 signaling pathway[J]. J Surg Res, 2022,275(1):161-171.
[22] WangS, Zeng X, Yang Y, et al. Hypothermic oxygenated perfusion ameliorates ischemia-reperfusion injury of fatty liver in mice via Brg1/Nrf2/HO-1 axis[J]. Artif Organs, 2022,46(2):229-238.
[23] AliFF, Mohammed MM, Hussein Y, et al. Targeting PI3K/p-Akt/eNOS, Nrf2/HO-1, and NF-κB/p53 signaling pathways by angiotensin 1-7 protects against liver injury induced by ischemia-reperfusion in rats[J]. Cell Biochem Funct, 2024,42(1):e3938.