Ischemia-reperfusion of small liver remnant promotes liver tumor growth and metastases--activation of cell invasion and migration pathways.

Elucidating the mechanism of liver tumor growth and metastasis after hepatic ischemia-reperfusion (I/R) injury of a small liver remnant will lay the foundation for the development of therapeutic strategies to target small liver remnant injury, and will reduce the likelihood of tumor recurrence after major hepatectomy or liver transplantation for liver cancer patients. In the current study, we aimed to investigate the effect of hepatic I/R injury of a small liver remnant on liver tumor development and metastases, and to explore the precise molecular mechanisms. A rat liver tumor model that underwent partial hepatic I/R injury with or without major hepatectomy was investigated. Liver tumor growth and metastases were compared among the groups with different surgical stress. An orthotopic liver tumor nude mice model was used to further confirm the invasiveness of the tumor cells from the above rat liver tumor model. Significant tumor growth and intrahepatic metastasis (5 of 6 vs. 0 of 6, P=0.015), and lung metastasis (5 of 6 vs. 0 of 6, P=0.015) were found in rats undergoing I/R and major hepatectomy compared with the control group, and was accompanied by upregulation of mRNA levels for Cdc42, ROCK (Rho kinase), and vascular endothelial growth factor, as well as activation of hepatic stellate cells. Most of the nude mice implanted with liver tumor from rats under I/R injury and major hepatectomy developed intrahepatic and lung metastases. In conclusion, hepatic I/R injury of a small liver remnant exacerbated liver tumor growth and metastasis by marked activation of cell adhesion, invasion, and angiogenesis pathways.     

The role of toll-like receptor-4 in the development of multi-organ failure following traumatic haemorrhagic shock and resuscitation

Haemorrhagic shock and resuscitation (HS/R) following major trauma results in a global ischaemia and reperfusion injury that may lead to multiple organ dysfunction syndrome (MODS). Systemic activation of the immune system is fundamental to the development of MODS in this context, and shares many features in common with the systemic inflammatory response syndrome (SIRS) that complicates sepsis. An important advancement in the understanding of the innate response to infection involved the identification of mammalian toll-like receptors (TLRs) expressed on cells of the immune system. Ten TLR homologues have been identified in humans and toll-like receptor-4 (TLR4) has been studied most intensively. Initially found to recognise bacterial lipopolysaccharide (LPS), it has also recently been discovered that TLR4 is capable of activation by endogenous 'danger signal' molecules released following cellular injury; this has since implicated TLR4 in several non-infectious pathophysiologic processes, including HS/R. The exact events leading to multi-organ dysfunction following HS/R have not yet been clearly defined, although TLR4 is believed to play a central role as has been shown to be expressed at sites including the liver, lungs and myocardium following HS/R. Multi-organ dysfunction syndrome remains an important cause of morbidity and mortality in trauma patients, and current therapy is based on supportive care. Understanding the pathophysiology of HS/R will allow for the development of targeted therapeutic strategies aimed at minimising organ dysfunction and improving patient outcomes following traumatic haemorrhage. A review of the pathogenesis of haemorrhagic shock is presented, and the complex, yet critical role of TLR4 as both a key mediator and therapeutic target is discussed.     

Cellular and molecular functions of hepatic stellate cells in inflammatory responses and liver immunology

The liver is a central immunological organ. Liver resident macrophages, Kupffer cells (KC), but also sinusoidal endothelial cells, dendritic cells (DC) and other immune cells are involved in balancing immunity and tolerance against pathogens, commensals or food antigens. Hepatic stellate cells (HSCs) have been primarily characterized as the main effector cells in liver fibrosis, due to their capacity to transdifferentiate into collagen-producing myofibroblasts (MFB). More recent studies elucidated the fundamental role of HSC in liver immunology. HSC are not only the major storage site for dietary vitamin A (Vit A) (retinol, retinoic acid), which is essential for proper function of the immune system. This pericyte further represents a versatile source of many soluble immunological active factors including cytokines [e.g., interleukin 17 (IL-17)] and chemokines [C-C motif chemokine (ligand) 2 (CCL2)], may act as an antigen presenting cell (APC), and has autophagy activity. Additionally, it responds to many immunological triggers via toll-like receptors (TLR) (e.g., TLR4, TLR9) and transduces signals through pathways and mediators traditionally found in immune cells, including the Hedgehog (Hh) pathway or inflammasome activation. Overall, HSC promote rather immune-suppressive responses in homeostasis, like induction of regulatory T cells (Treg), T cell apoptosis (via B7-H1, PDL-1) or inhibition of cytotoxic CD8 T cells. In conditions of liver injury, HSC are important sensors of altered tissue integrity and initiators of innate immune cell activation. Vice versa, several immune cell subtypes interact directly or via soluble mediators with HSC. Such interactions include the mutual activation of HSC (towards MFB) and macrophages or pro-apoptotic signals from natural killer (NK), natural killer T (NKT) and gamma-delta T cells (γδ T-cells) on activated HSC. Current directions of research investigate the immune-modulating functions of HSC in the environment of liver tumors, cellular heterogeneity or interactions promoting HSC deactivation during resolution of liver fibrosis. Understanding the role of HSC as central regulators of liver immunology may lead to novel therapeutic strategies for chronic liver diseases.     

Bromelain ameliorates hepatic microcirculation after warm ischemia

BACKGROUND: Because of its immunomodulatory action, the protease bromelain represents a novel strategy for the treatment of hepatic ischemia/reperfusion (I/R) injury. A dose-response study was performed to investigate the effect of bromelain on liver function, microcirculation, and leukocyte-endothelium interactions in hepatic I/R injury. MATERIALS AND METHODS: One hundred forty rats were randomized to 8 short-term or 12 long-term groups (n=7 each). A 30 min normothermic hepatic ischemia was induced by Pringle maneuver with a portocaval shunt. Animals were treated 60 min prior to ischemia with either no therapy, 0.1, 1.0, or 10 mg/kg b.w. bromelain i.v. In the short-term experiments, microcirculation was investigated 30 min after sham operation or ischemia using intravital microscopy. In the long-term experiments AST, ALT, and bradykinin levels were determined for 14 d after central venous catheter (CVC) placement only, sham operation, or ischemia. Additionally, apoptosis rate, Kupffer cell activation, endothelial cell damage, and eNOS expression were analyzed. RESULTS: In sham-operated animals, treatment with 10 mg/kg b.w. bromelain led to a disturbed microcirculation with increased leukocyte adherence, apoptosis rate, Kupffer cell activation, and endothelial cell damage. Six h after CVC placement and administration of 10 mg/kg b.w. bromelain, AST and ALT levels were significantly increased. After I/R, rats treated with 0.1 mg/kg b.w. bromelain showed an improved microcirculation, reduction in leukocyte adhesion, apoptosis rates, Kupffer cell activation and endothelial cell damage, increased eNOS expression, and significantly lower AST levels compared with untreated animals. CONCLUSION: Bromelain represents a novel approach to the treatment of hepatic I/R injury with a limited therapeutic window.     

HEGPOL: Randomized,placebo controlled,multicenter, double-blind clinical trial to investigate hepatoprotective effects of glycine in the postoperative phase of liver transplantation [ISRCTN69350312]

Background  

Kupffer cell-dependent ischemia / reperfusion (I/R) injury after liver transplantation is still of high clinical relevance, as it is strongly associated with primary dysfunction and primary nonfunction of the graft. Glycine, a non-toxic, non-essential amino acid has been conclusively shown in various experiments to prevent both activation of Kupffer cells and reperfusion injury. Based on both experimental and preliminary clinical data this study protocol was designed to further evaluate the early effect of glycine after liver transplantation.     

移植肝再灌注损伤的发生机制

目的：介绍有关移植肝再灌注损伤发生机制的研究动向,方法：复习有关文献并进行综述性报告。结果：移植肝冷保存后的再灌注损伤的发生机制主要有：（1）内皮细胞损伤和Kupffer细胞激活,导致一系列细胞因子的产生,引起移植肝损伤,并引发全身炎症反应综合征。（2）白细胞,血小板与肝血窦壁的粘附而损害肝细胞,并可阻塞肝血窦造成“无复流”现象;（3）pH值的变化,再灌注后移植肝的代谢恢复正常后,组织内pH值的改变可引起肝细胞损伤,并可造成线粒体的肿胀,使肝细胞的功能降低,（4）复氧损伤,主要与白细胞释放活性氧（ROS）有关,结论：移植肝再灌注损伤是多种因素综合作用的结果,在再灌注前后提高肝细胞和内皮细胞的活性,抑制Kupffer细胞的激活,减少ROS及肿瘤坏死因子（TNF）的产生将是今后预防移植肝再灌注损伤研究的关键。     

Liver Ischemia and Reperfusion Injury: New Insights into Mechanisms of Innate—Adaptive Immune‐Mediated Tissue Inflammation

Ischemia and reperfusion injury (IRI) is a dynamic process that involves two distinctive yet interrelated phases of ischemic organ damage and inflammation‐mediated reperfusion injury. Although multiple cellular and molecular pathways contribute and regulate tissue/organ damage, integration of different players into a unified mechanism is warranted. The crosstalk between innate and adaptive immune systems plays a significant role in the pathogenesis of liver IRI. In this review, we focus on recent progress in the mechanism of liver innate immune activation by IR. Kupffer cells (KC), DCs, NK, as well as T cells initiate local inflammation response, the hallmark of IRI, by utilizing distinctive immune receptors to recognize and/or trigger various molecules, both endogenous and exogenous. The interlocked molecular signaling pathways in the context of multiple liver cell types, the IRI kinetics and positive versus negative regulatory loops in the innate immune activation process are discussed. Better appreciation of molecular interactions that mediate these intricate cascades, should allow for the development of novel therapeutic approached against IRI in liver transplant recipients.     

Toll-Like Receptor Signaling in Liver Ischemia and Reperfusion

ABSTRACT

Liver ischemia–reperfusion (I/R) injuries are significant clinical challenges implicated in various hepatic surgical procedures and transplantations. Associated with varying degrees of insult, the hallmark of I/R is the excessive inflammatory response potentiated by the host immune system. Toll-like receptors (TLRs), known to play an important role in pathogen-derived inflammation, are now thought to participate in I/R injury-derived inflammation signaling pathways. Endogenous particles (proteins, cytokines, nucleic acids) that are released from damaged host cells bind to TLR2, TLR4, and TLR9, resulting in even further injury by subsequent inflammatory reactions and activation of the innate immune system. This review aims to systematically examine the current literature about TLR signaling mechanisms, allowing for a greater understanding of the precise role of TLRs in hepatic I/R injuries.     

Kupffer细胞在肝移植缺血再灌注损伤中的双重作用

Kupffer细胞足定居于肝内的巨细胞,在月十移植缺血再灌注损伤中发挥着重要的作用,门静脉恢复血流后刺激Kupffer细胞激活,释放活性氧族、多种炎性介质和细胞因子,对肝脏造成损伤.另一方面又可上调HO-1的表达,保护肝脏缺血再灌注损伤,因此,Kupffer细胞在肝移植缺血再灌注损伤中发挥着双重效应.     

Long-term effects of acute ischemia and reperfusion injury

Ischemia reperfusion (I/R) injury plays a major role in delayed graft function and long-term changes after kidney transplantation. By using different therapeutic strategies to prevent I/R injury in rat models of kidney transplantation we studied relationships between inflammatory cell arrival and adhesion molecule expression. In other rat models for acute renal failure we investigated the effect of up-regulation of protective genes such as heme oxygenase-1 (HO-1) on infiltrating cells, showing that infiltrating cells also contribute to beneficial effects. In order to gain more insight into the complex mechanisms of long-term changes after kidney transplantation, we started a protocol biopsy program to study histologic changes 6, 12, and 26 weeks after transplantation. The following article clarifies some of the complex mechanisms contributing to long-term changes caused by I/R injury.     

血红素氧合酶-1减轻大鼠移植肝缺血再灌注损伤的作用及机制

目的 探讨血红素氧合酶-1(HO-1)减轻大鼠移植肝缺血再灌注损伤的作用及其机制.方法 选择近交系雄性SD大鼠作为肝移植的供、受者;采用单纯随机方法将48只SD大鼠随机分为对照组、抑制组和诱导组(每组供、受者各8只).对照组:供肝不用任何药物处理;抑制组:在获取供肝前24 h,经供者腹腔注射HO-1抑制剂锌原卟啉20 mg/kg进行预处理;诱导组:在获取供肝前24 h,经供者腹腔注射HO-1诱导剂钴原卟啉5 mg/kg进行预处理.获取供肝后,在4℃UW液中冷保存24 h.肝移植前检测供肝HO-1的表达水平;肝移植后6 h采血并获取移植肝标本,分离培养枯否细胞;检测受者的肝功能;检测枯否细胞培养上清中肿瘤坏死因子α(TNF-α)和白细胞介素6(IL-6)的含量;观察移植肝组织病理学表现以及枯否细胞CD14 mRNA的表达水平和蛋白含量测定.结果 移植前诱导组供肝HO-1的表达水平明显增高.移植后诱导组血清丙氨酸转氨酶(ALT)和天冬氨酸转氨酶(AST)含量明显降低;移植肝组织病理学损伤减轻;枯否细胞培养上清中TNF-α和IL-6含量减少;而且枯否细胞上的CD14 mRNA表达水平和蛋白含量也明显低于抑制组.结论 诱导供肝HO-1表达上调可能抑制了枯否细胞的激活,从而减轻大鼠移植肝缺血再灌注损伤.     

Hyperglycemia and Liver Ischemia Reperfusion Injury: A Role for the Advanced Glycation Endproduct and Its Receptor Pathway

Although pretransplant diabetes is a risk factor for mortality post–liver transplant, the underlying mechanism has not been fully defined. In a murine liver partial warm ischemia model, we addressed the question of how diabetes/hyperglycemia impacted tissue inflammatory injuries against ischemia reperfusion (IR), focusing on the advanced glycation endproduct (AGE) and its receptor (RAGE) pathway. Our results showed that hepatocellular injury was exacerbated in streptozotocin‐induced diabetic mice against IR, in association with hyper‐inflammatory immune activation in livers. Serum levels of AGEs, but not HMGB1, were increased in diabetic mice in response to liver IR. Both RAGE antagonist peptides and small interfering RNA alleviated liver injuries and inhibited inflammatory immune activation against IR in diabetic, but not normal, mice. Kupffer cells (KCs)/macrophages, but not hepatocytes, from diabetic mice expressed significantly higher levels of RAGE, leading to their hyper‐inflammatory responsiveness to both TLR ligands and AGEs. In vitro, hyperglycemia increased macrophage RAGE expression and enhanced their TLR responses. Our results demonstrated that activation of the AGE–RAGE signaling pathway in KCs was responsible for hyper‐inflammatory immune responses and exacerbated hepatocellular injuries in diabetic/hyperglycemic hosts against liver IR.     

Kupffer Cell Function in Ischemic and Nonischemic Livers After Hepatic Partial Ischemia/Reperfusion

Hepatic partial ischemic/reperfusion (I/R) injury, in which ischemic and nonischemic areas of the liver are likely to respond to each other after reperfusion, often occurs following hepatobiliary surgical procedures. Kupffer cells (KCs) are considered to play a major role in hepatic I/R injury. To study the activation of KCs in ischemic and nonischemic liver tissues following hepatic I/R, we investigated the superoxide generation and proinflammatory cytokine production of KCs in both liver parts in a rat model of partial hepatic I/R injury. KC superoxide generation in the ischemic and nonischemic lobes was upregulated 6 and 24 h after reperfusion, respectively, and then accelerated. The production of interleukin-1β (IL-1β) by KCs in the ischemic lobes increased during the early and late phases, 6 h and 48–72 h after reperfusion, respectively. A late increase in IL-1β production was also observed in the nonischemic lobes. Production of tumor necrosis factor-α (TNF-α) increased 6–24 h after reperfusion in both lobes. Upregulation of IL-1β mRNA in the ischemic lobes preceded the upregulation of TNF-α mRNA in both lobes. The hepatic partial I/R process results in activation of KCs in ischemic and nonischemic areas of the liver. The KCs are activated during the early phase after reperfusion in the ischemic areas, followed by activation in both the ischemic and nonischemic areas. This could be a cause of liver dysfunction after partial hepatic I/R during surgery. Received: September 9, 1999 / Accepted: September 26, 2000     

肝脏缺血再灌注损伤的研究进展

肝脏缺血再灌注损伤是肝脏外科常见的病理生理过程,是集肝窦血管内皮损伤、氧化应激、炎症反应、细胞凋亡与自噬等多种机制共同作用的结果,对于肝切除、肝移植等肝脏外科手术后肝功能恢复和围手术期安全具有显著影响.然而,迄今为止,其具体的损伤机制尚未完全阐明,在临床上也始终缺乏有效的干预手段.因此,对于肝脏缺血再灌注损伤发病机制以及防御举措的深入研究具有重要的临床意义.笔者结合近年来最新文献报道,对该领域的实验研究进展予以简要综述.     

Mycophenolate mofetil preconditioning protects mouse liver against ischemia/reperfusion injury in wild type and toll-like receptor 4 knockout mice

BackgroundMycophenolate mofetil (MMF), an immunosuppressive drug, exerts anti-inflammatory effects on organs during ischemia/reperfusion (I/R) injury. However, the exact function of MMF in hepatic I/R injury remains largely unknown. The purpose of this study was to explore the role and potential mechanism of MMF protection in hepatic I/R injury.MethodsMale wild type (WT) and TLR4 knockout (KO) mice were injected intraperitoneally with MMF or normal saline. Animals underwent 90 min of partial hepatic ischemia, followed by 1, 6, or 24 h of reperfusion. Hepatic histology, serum amiotransferase, inflammatory cytokines, hepatocyte apoptosis, and hepatocyte autophagy were examined to assess liver injury.ResultsTreatment with MMF significantly decreased hepatic I/R injury as indicated by a reduction in serum aminotransferase levels, Suzuki scores, and the overall degree of necrosis. MMF treatment inhibited TLR4 activation dramatically. MMF administration also significantly inhibited the activation of the NF-κB pathway and the expression of pro-inflammatory cytokines. In TLR4 KO mice, MMF still exerted protection from hepatic I/R injury. MMF treatment inhibited hepatocyte apoptosis, as indicated by reduced TUNEL staining, and reduced the accumulation of cleaved caspase-3. In addition, MMF may induce autophagy and increase autophagic flux before and after hepatic reperfusion by augmenting the expression of LC3-II, P62, and Beclin-1. The induction of autophagy by MMF treatment may be related to TLR4 activation.ConclusionsOur results indicate that MMF treatment ameliorates hepatic I/R injury. The mechanism of action likely involves the ability of MMF to decrease apoptosis and the inflammatory response while inducing autophagy.