Effects of ischemic preconditioning on regenerative capacity of hepatocyte in the ischemically damaged rat livers

BACKGROUND: Liver regeneration after partial hepatectomy is regulated by several factors that activate or inhibit hepatocyte proliferation. A short period of ischemia-reperfusion (IR), called ischemic preconditioning (IPC), protects the liver against subsequent sustained ischemic insults. The present study investigated the effects of IPC on liver regeneration after partial hepatectomy under IR in rats. MATERIALS AND METHODS: Male Wistar rats were subjected to 45 min of total hepatic ischemia, and 70% hepatectomy was performed just before reperfusion. Animals were pre-treated with either IPC (10/15 min) (IPC + PHx group) or not (ischemia + PHx). The survival rate, serum transaminases, tumor necrosis factor (TNF)-alpha, and interleukin (IL)-6 levels, hepatocyte proliferation and histological change of the remnant liver were measured in both groups and compared with non-ischemic controls subjected to 70% hepatectomy alone (PHx group). RESULTS: The survival rate was significantly better in the IPC + PHx group than in the ischemia + PHx group. Furthermore, IPC reduced liver injury determined by liver histology and serum transaminases. There was an early rise in serum TNF-alpha and IL-6 levels in the ischemia + PHx group. Compared with non-ischemic controls, IPC significantly decreased TNF-alpha, but not IL-6 during the late (24 and 48 h) phases of reperfusion. Rats subjected to 70% hepatectomy and 45 min of hepatic ischemia showed significantly reduced hepatocyte proliferation (mitotic index, proliferating cell nuclear antigen, and relative liver weight) when compared with animals subjected to hepatectomy alone. However, hepatocyte proliferation was markedly increased in rats pretreatment with IPC when compared with ischemic controls. CONCLUSION: These results suggest that ischemic pre-conditioning ameliorates the hepatic injury associated with ischemia-reperfusion and has a stimulatory effect on liver cell regeneration that may make it valuable as a hepatoprotective modality. Il-6 appears to be key mediator in promoting regeneration after combined ischemia and hepatic resection.     

Modulation of Remote Ischemic Preconditioning by Proinflammatory Cytokines in Renal Transplant Recipients

Aim: Remote ischemic preconditioning (RIPC) has been used as a strategy to reduce acute renal injury and ischemia-reperfusion injury (IRI) in renal transplantation (RT) with controversial results. Objective: To determine if RIPC modifies IRI in cadaveric RT recipients through inflammatory mediators and graft function. Methods: Twenty-nine RT recipients were studied, 12 in the control group (CG) and 17 in the RIPC group. RIPC which was performed on donors using a pneumatic tourniquet placed on both thighs for 10 min followed by the determination of IL-1, IL-6, TNF-α, VEGF, and ICAM-1, and hematological and biochemical parameters in different phases of RT. Results: Serum creatinine levels were significantly lower in the RIPC group versus the CG at 15 and 30 days; however, the estimated glomerular filtration rate (eGFR) showed no significant difference in any phase between either group, only TNF-α showed significantly higher values in the RIPC group versus the CG in almost all phases of the study, meanwhile IL6 was increased at 72 hours (hr) and 30 days, IL1 at 72 hr and 15 days and ICAM-1 post reperfusion, contrary to this VEGF showed a decrease at 7 and 15 days. Conclusion: RIPC did not improve eGFR or serum creatinine; however, it modifies the inflammatory response in RT recipients.     

Intracellular oxygenation and cytochrome oxidase C activity in ischemic preconditioning of steatotic rabbit liver

Background

Mild to moderate steatotic livers are used as marginal donors in liver transplantation. Very little is known about the mechanisms of ischemia reperfusion (IR) injury (IRI) in fatty liver. This study aimed to establish whether cytochrome oxidase C (COX) activity is compromised by IRI in fatty liver and whether ischemic preconditioning (IPC) can protect COX activity.

Methods

New Zealand rabbits were fed on a high-cholesterol diet for 8 weeks to induce moderate hepatic steatosis. Three groups were tested. The IR group underwent 60 minutes of ischemia, followed by 7 hours of reperfusion. The IPC group (IPC + IR) underwent 5 minutes of ischemia, followed by 10 minutes of reperfusion and then 60 minutes of ischemia and 7 hours of reperfusion. The control group (sham) underwent the same surgical procedure, but ischemia was not induced. Deoxyhemoglobin, oxyhemoglobin, and change in the redox state of COX was continuously monitored in vivo by near-infrared spectroscopy. COX and citrate synthase (CS) activity assays were carried out on liver biopsy specimens in vitro. Bile was collected continuously during the procedure and analyzed using proton nuclear magnetic resonance spectroscopy.

Results

The IR group had decreased COX activity and tissue oxygenation represented by deoxyhemoglobin, oxyhemoglobin, COX, and elevated redox ratios of lactate/pyruvate and β-hydroxybutarate/acetoacetate in vivo and a decrease in COX and CS activity in vitro. The IPC + IR group showed higher levels of all measured parameters in vivo and showed a smaller decrease in COX and CS activity in vitro.

Conclusion

This study shows that IRI affects COX activity in fatty livers. This is attenuated by IPC.     

缺血预处理对大鼠肝脏部分切除术后残留肝脏的保护作用及其机制的研究

目的　探讨缺血预处理 (ischemicpreconditioning ,IPC)对大鼠肝脏部分切除术后残留肝脏的保护作用及其机制。方法　 5 0只雄性SD大鼠随机均分为假手术 (Sham)、单纯热缺血 (warmis chemia ,WI)、IPC、IPC L arginine(NO供体 )和WI NAME(NO合成酶抑制剂 ) 5组。术前、术后 1、2、3d检测血清AST和ALT ,术前、IPC后、热缺血后 0 5、1、2、3h检测肝脏组织NO浓度。AST和ALT用自动生化分析仪检测 ,NO用硝酸还原法检测。结果　WI、IPC、IPC L arginine及WI NAME组术后AST和ALT均高于Sham组 (P <0 0 5或P <0 0 1) ,IPC和WI NAME组术后AST和ALT低于WI和IPC L arginine组 (P <0 0 5 )。WI、IPC、IPC L arginine及WI NAME组术后 0 5h肝脏组织NO浓度开始升高(P <0 0 5或P <0 0 1) ,IPC和WI NAME组术后肝脏组织NO浓度低于WI和IPC L arginine组 (P <0 0 5 )。结论　IPC对大鼠肝脏部分切除术后残留肝脏的缺血再灌注损伤有保护作用。IPC通过抑制大鼠热缺血再灌注肝脏产生NO ,减少NO所诱发的肝脏组织细胞的凋亡或坏死 ,保护肝脏功能。     

Relevance of epidermal growth factor to improve steatotic liver preservation in IGL-1 solution

Aim

Static preservation solution is critical for liver graft outcomes, especially when steatosis is present. Institut Georges Lopez (IGL)-1 solution protects fatty livers effectively against cold ischemia reperfusion injury. Its benefits are mediated by nitric oxide and prevention of oxidative stress. The supplementation of IGL-1 with epidermal growth factor (EGF) enhances steatotic graft preservation by increasing adenosine triphosphate content, thereby mitigating oxidative stress and mitochondrial damage.

Methods

After steatotic livers were preserved for 24 hours in IGL-1 solution with or without EGF supplements, they were perfused ex vivo for 2 hours at 37°C. The benefits of EGF were assessed by evidences of hepatic damage and function—transaminases, bile production, and flow rate—as well as by other factors presumably associated with the poor tolerance of fatty livers toward cold ischemia-reperfusion injury (IRI)—energy metabolism, mitochondrial damage, oxidative stress, eNOS activity and proinflammatory interleukin (IL) beta content.

Results

Steatotic livers preserved in IGL-1 solutions supplemented with EGF (10 μg/L) showed lower transaminase levels, greater bile production, and ameliorated flow rates when compared to IGL-1 alone. In addition, energy metabolism deterioration, mitochondrial damage, oxidative stress, and cytokine IL-1 beta release were prevented.

Conclusion

EGF addition to IGL-1 increased fatty liver graft preservation, thereby reducing steatotic liver damage against cold IRI.     

Severe steatosis increases hepatocellular injury and impairs liver regeneration in a rat model of partial hepatectomy

OBJECTIVE: The aim of this study was to assess the influence of severe steatosis with inflammation on hepatocellular recovery after 70% hepatectomy in a rat model of diet-induced steatosis. BACKGROUND: Patients with steatosis have an increased risk of inflammatory complications after liver resection. This might be attributable to Kupffer cell-mediated inflammation in steatotic livers causing progressive injury. METHODS: Male Wistar rats were fed a standard methionine- and choline-deficient diet for 1 or 5 weeks. A 70% partial hepatectomy (PH) was performed, after which rats were killed at 24, 48, or 72 hours. The extent of steatosis and inflammation was determined by assessment of hepatic triglycerides, cytokine content, and histopathology. Outcome parameters were: liver regeneration (MIB-5 proliferation rate, mitotic index, and regenerating liver mass), hepatocellular injury (plasma aminotransferases, lipid peroxidation, histopathology, and apoptosis), Kupffer cell-mediated proinflammatory response (TNF-alpha, IL-1beta, IL-6, IL-10 in plasma and liver) and antioxidant content (total glutathione). RESULTS: Methionine- and choline-deficient diet induced uncomplicated steatosis after 1 week (<30% hepatocytes affected without inflammation) and severe steatosis after 5 weeks (>60% hepatocytes affected, including prominent inflammation) as confirmed by histopathology. After PH, liver regeneration was impaired at all time points in the severe steatosis group as compared with the mild and control groups (P < 0.05). Hepatocellular injury was significantly increased in the severe steatosis group at all time points (P < 0.05). Kupffer cell-mediated inflammatory responses were aggravated in the severe steatosis group along with decreased antioxidant content (P < 0.05). Necrosis was the main type of cell death in severe steatotic livers compared with mainly apoptotic cell death in mild steatotic and normal livers. CONCLUSION: Steatosis with prominent inflammation impaired liver regeneration probably because of increased hepatocellular lipid peroxidation and damage in concert with Kupffer cell-mediated proinflammatory responses. These results suggest an increased risk of performing extensive liver resection in the presence of severe steatosis.     

肝切除及肝移植术中缺血预处理的临床研究进展

缺血再灌注损伤一直是肝切除及肝移植术中存在的问题,是术后肝功能衰竭和移植物无功能的主要原因,而缺血预处理能减轻这种损伤。文中综述了近年来缺血预处理及其在临床应用中的研究进展,探讨了该技术在临床上的实用价值。     

Cross-Talk Between Sirtuin 1 and High-Mobility Box 1 in Steatotic Liver Graft Preservation

Background

Sirtuin 1 (SIRT1) is a nicotinamide adenine dinucleotide +–dependent histone deacetylase that regulates various pathways involved in ischemia-reperfusion injury (IRI). Moreover, high-mobility group box 1 protein (HMGB1) has also been involved in inflammatory processes during IRI. However, the roles of both SIRT1 and HMGB1 in liver preservation is poorly understood. In this communication, we evaluated the potential relationship between SIRT1 and HMGB1 in steatotic and non-steatotic liver grafts preserved in Institute Georges Lopez solution (IGL-1) preservation solution enriched or not enriched with trimetazidine (TMZ).

Cytoprotective Effects of a Cyclic RGD Peptide in Steatotic Liver Cold Ischemia and Reperfusion Injury

The serious need for expanding the donor population has attracted attention to the use of steatotic donor livers in orthotopic liver transplantation (OLT). However, steatotic livers are highly susceptible to hepatic ischemia–reperfusion injury (IRI). Expression of fibronectin (FN) by endothelial cells is an important feature of hepatic response to injury. We report the effect of a cyclic RGD peptide with high affinity for the α5β1, the FN integrin receptor, in a rat model of steatotic liver cold ischemia, followed by transplantation. RGD peptide therapy ameliorated steatotic IRI and improved the recipient survival rate. It significantly inhibited the recruitment of monocyte/macrophages and neutrophils, and depressed the expression of pro-inflammatory mediators, such as inducible nitric oxide synthase (iNOS) and interferon (IFN)-γ. Moreover, it resulted in profound inhibition of metalloproteinase-9 (MMP-9) expression, a gelatinase implied in leukocyte migration in damaged livers. Finally, we show that RGD peptide therapy reduced the expression of the 17-kDa active caspase-3 and the number of apoptotic cells in steatotic OLTs. The observed protection against steatotic liver IRI by the cyclic RGD peptides with high affinity for the α5β1 integrin suggests that this integrin is a potential therapeutic target to allow the successful utilization of marginal steatotic livers in transplantation.     

Protective effect of ischemic preconditioning against intermittent warm-ischemia-induced liver injury

BACKGROUND: Although ischemic preconditioning (IPC) has been reported to protect the liver from injury when subjected to continuous hepatic ischemia, whether IPC protects rat livers against ischemia-reperfusion (I/R) injury after intermittent ischemia has not been elucidated. MATERIALS AND METHODS: Five groups of Wistar rats were subjected to intermittent hepatic ischemia (I) comprising 15-min ischemia and 5-min reperfusion three times with or without prior IPC (10-min ischemia and 10-min reperfusion), 45-min continuous ischemia (C) with or without IPC, and sham operation. Serum transaminase and lactic acid levels, hepatic tissue energy charges, and hepatic blood perfusion were measured after reperfusion. Plasma tumor necrosis factor-alpha (TNF-alpha) levels were determined after reperfusion for 120 min. Histological and apoptotic findings were evaluated after reperfusion for 180 min. RESULTS: IPC significantly reduced serum transaminase levels after continuous and intermittent ischemia (IPC + C, 1107 vs C, 2684 IU/l; IPC + I, 708 vs I, 1859 IU/l). After hepatic ischemia without IPC, apoptosis and necrosis with increased plasma TNF-alpha levels were observed. IPC protected livers from injury by interfering with the increase in plasma TNF-alpha (IPC + I, 27.6 vs I, 64.8 pg/ml; IPC + C, 21.6 vs C, 49.3 pg/ml). This resulted in the attenuation of hepatic necrosis after continuous ischemia and significantly reduced necrosis and apoptosis after intermittent ischemia. CONCLUSIONS: IPC exerts a greater protective effect against hepatic I/R injury after intermittent hepatic ischemia than after continuous hepatic ischemia.     

Meloxicam increases epidermal growth factor receptor expression improving survival after hepatic resection in diet-induced obese mice

Background

Patients with fatty liver have delayed regenerative responses, increased hepatocellular injury, and increased risk for perioperative mortality. Currently, no clinical therapy exists to prevent liver failure or improve regeneration in patients with fatty liver. Previously we demonstrated that obese mice have markedly reduced levels of epidermal growth factor receptor in liver. We sought to identify pharmacologic agents to increase epidermal growth factor receptor expression to improve hepatic regeneration in the setting of fatty liver resection.

Human C1 inhibitor attenuates liver ischemia-reperfusion injury and promotes liver regeneration

Liver ischemia-reperfusion injury (IRI) is a well-known cause of morbidity and mortality after liver transplantation (LT). Activation of the complement system contributes to the pathogenesis of IRI. Effective treatment strategies aimed at reducing hepatic IRI and accelerating liver regeneration could offer major benefits in LT. Herein, we investigated the effect of C1-esterase inhibitor (human) [C1-INH] on IRI and liver regeneration. Mice were subjected to 60-min partial IRI, with or without 70% partial hepatectomy, or CCl₄-induced acute liver failure. Before liver injury, the animals were pretreated with intravenous C1-INH or normal saline. Liver IRI was evaluated using serum levels of alanine aminotransferase, serum interleukin-6, and histopathology. Liver samples were stained for specific markers of regeneration (5-bromo-2'-deoxyuridine [BrdU] staining and proliferating cell nuclear antigen [PCNA]). Histology, serum interleukin-6, and alanine aminotransferase release revealed that C1-INH treatment attenuated liver injury compared with controls. Improved animal survival and increased number of BrdU- and PCNA-positive cells were observed in C1-INH–treated animals which underwent IRI + partial hepatectomy or CCl₄ injection compared with control group. These data indicate that complement plays a key role in IRI and liver regeneration. C1-INH represents a potential therapeutic strategy to reduce IRI and promote regeneration in LT.     

不同时间的缺血预处理对肝硬化大鼠缺血再灌注损伤的保护作用

目的 探讨缺血预处理(IPC)对肝硬化大鼠缺血再灌注损伤(I/R)的保护作用,并寻找对肝硬化I/R保护作用的最佳有效时间窗和理想方案.方法 通过构建正常大鼠及肝硬化大鼠模型,以正常肝脏I/R(A组)和正常肝脏10-10 min-IPC方案(B组)为对照组,肝硬化组则分别施行单纯I/R(C组)、5-10 min(D组)、8-10min(E组)、10-10 min(F组)及15-10 min(G组)的IPC方案,每组18只,分别在术后1 h、4 h及24 h三个时间点采静脉血,检测血清ALT、AST、LDH及肝组织中MDA、MPO、NO、SOD水平,评价肝功能及肝脏组织炎性浸润及抗损伤程度.结果 肝脏缺血30 min后肝脏功能受损明显,表现为各组的ALT、AST、LDH水平升高,尤以再灌注4 h时显著(P<0.05),但经过缺血预处理后,各IPC组中的NO、MDA、MPO及SOD水平亦显著高于其对应的I/R组,以E组的差别有显著意义(P<0.05).结论 10-10 min的IPC方案对于正常肝脏I/R确有保护作用,而8-10 min的IPC方案能最有效地启动对肝硬化大鼠肝脏I/R的保护作用.     

Liver and lung late alterations following hepatic reperfusion associated to ischemic preconditioning or N-acetylcysteine

This study aimed the effect of n-acetylcysteine or ischemic preconditioning in hepatic and pulmonary damage after liver ischemia-reperfusion injury. Twenty-four male Wistar-EPM rats were assigned into four groups: (IR) Hepatic ischemia-reperfusion; (IPC) IPC achieved before hepatic ischemia; (NAC) Animals received NAC pretreatment; and Sham operated group. After 24 h of hepatic reperfusion, blood, liver, and pulmonary samples were evaluated. Nonparametric tests were used (P 相似文献   

Ischemic Preconditioning to Prevent Lethal Ischemic Spinal Cord Injury in a Swine Model

Objective: Paraplegia is a serious complication of thoracic and thoracoabdominal aortic operations and is the result of ischemic spinal cord injury induced by low perfusion pressure during cross-clamping of the aorta. Ischemic preconditioning (IPC) of the heart or brain with reversible sublethal ischemic injury induces resistance to subsequent lethal ischemia. The aim of this study is to investigate whether ischemic tolerance can be induced by IPC of the spinal cord in a swine model. Study Design: The animals were randomly divided into three groups: the sham group (n = 3), control group (n = 6) and IPC group (n = 8). In the sham group, we performed a left thoracotomy without any ischemic injury. In the IPC group, the swine received a reversible ischemic spinal cord injury by aortic clamping for 20 min, whereas in the control group, no aortic cross-clamping was performed. Forty-eight hours later, the animals in both the IPC and control groups underwent aortic clamping for 30 min. Neurological examination was done 24 h later, and then the animals were euthanized for histopathology and a malonedialdehyde spectrophotometry assay of the spinal cord tissue. Results: A statistically significant difference in neurological outcome was observed between the control and IPC groups at 24 h after ischemic injury. The incidence of paraplegia and severe paresis was 100% in the control group and 62.5% in the IPC group (p =. 028). Between control and IPC groups, there was no statistically significant difference in histopathology and only a borderline statistical difference in the malonedialdehyde assay of the ischemic spinal cord (p =. 0745). Conclusion: In this study, IPC induced protection against a 30-min ischemic insult of the spinal cord, although complete recovery was not achieved (standing up or walking). We expect that combining this IPC with other existing protective methods might lead to a synergistic effect, which warrants further investigation.     

Recombinant Human Soluble Thrombomodulin Attenuates Hepatic Ischemia and/or Reperfusion Injury by Inhibiting Leukocyte Accumulation in Mice With Normal and Fatty Liver

In an attempt to increase the number of donor livers, there has been an increased use of marginal donor livers, such as steatotic (fatty) livers that increase susceptibility to ischemia and reperfusion injury (IRI). Inflammatory cell accumulation has a greater role in IRI in steatotic liver than in normal liver. Although the recombinant human soluble thrombomodulin (rhsTM) attracts attention as a new treatment for disseminated intravascular coagulation, the therapeutic efficacy of rhsTM in hepatic IRI remains uncertain, especially in fatty livers. We aimed to demonstrate the effect of rhsTM on hepatic IRI using well-established in vivo experimental models with steatotic liver.

Liver-Wrapping,Nitric Oxide-Releasing Nanofiber Downregulates Cleaved Caspase-3 and Bax Expression on Rat Hepatic Ischemia-Reperfusion Injury

Background

Hepatic ischemia-reperfusion injury (IRI) is an important determinant of the outcome of hepatic surgery, including re-section and transplantation. Previous studies have shown that nitric oxide (NO) has a protective effect against IRI. Therefore, many studies have examined methods for supplying NO. In this study, we investigated the effect of NO-releasing nanofibers on hepatic IRI in a rat model.

阿霉素预处理替代缺血预处理提供鼠肝缺血耐受力的实验研究

目的 探讨缺血预处理（IPC）延迟保护作用的发生机制以及应用阿霉素预处理（DPC）是否可以模拟IPC的延迟保护作用。方法 建立大鼠部分肝脏热缺血再灌注模型。IPC组采用肝脏缺血10min，再灌注10in，DPC组经静脉注射阿霉素（1mg/kg体重），对照组等量生理盐水注射。肝组织HSP70和HO－1蛋白和血清TNF－α、IL－10浓度分别采用Western blot法和ELISA法测定。结果 IPC后HO－1和HSP70含量分别于12h和24h达到高峰；IPC和DPC后24h诱导HSP70、HO－1的量无显著差异（P＞0.05）。对照组缺血再灌注后3h血清中TNF－α、AST、ALT、LDH及W／D（湿重／干重）的水平明显升高，而IL－10的含量降低，和假手术组相比差异显著（P＜0.01）；IPC或DPC后降低了TNF－α的释放和AST、ALT、LDH及W／D的水平，提高了IL－10的含量，和对照组相比差异显著（P＜0.01）。结论 IPC的延迟保护作用与HSP70和HO－1的诱导生成有关，DPC可以模拟IPC的延尺性保护作用，诱导HSP70和HO－1的产生。     

Expansion of hepatic progenitor cell in fatty liver graft after living donor liver transplantation

Although it is known that steatotic livers have a reduced ability to regenerate, most individuals with steatosis show generally benign prognosis. We hypothesized that a proliferative blockade in steatotic hepatocytes results in the compensatory expansion of hepatic progenitor cells (HPC) during fatty liver regeneration. Fifty‐four cases of living donor liver transplantation (LDLT) with a liver biopsy performed at the postoperative 10th day were examined. HPC were counted by immunofluorescence histochemical dual‐staining technique using cytokeratin 7 and Ki‐67, and the replicative arrest of hepatocytes was assessed by p21 immunohistochemistry. The degree of ductular proliferation during regeneration 10 days after LDLT correlated both with the degree of steatosis and the number of HPC (P < 0.001). There was no difference in the average number of HPC and the replicative arrest index between donors with or without steatosis before LDLT (P = 0.111 and P = 0.062). However, degree of steatosis correlated with both the expansion of HPC and the replicative arrest index during liver regeneration 10 days after LDLT (P < 0.001 and P < 0.001, respectively). Moreover, increased replicative arrest was strongly associated with HPC expansion (P < 0.001). In conclusion, the compensatory expansion of HPC as a result of impaired hepatocyte replication occurred during steatotic liver regeneration after LDLT.     

Pilot study of a noninvasive real-time optical backscatter probe in liver transplantation

Transplantation of severely steatotic donor livers is associated with early allograft dysfunction and poorer graft survival. Histology remains the gold standard diagnostic of donor steatosis despite the lack of consensus definition and its subjective nature. In this prospective observational study of liver transplant patients, we demonstrate the feasibility of using a handheld optical backscatter probe to assess the degree of hepatic steatosis and correlate the backscatter readings with clinical outcomes. The probe is placed on the surface of the liver and emits red and near infrared light from the tip of the device and measures the amount of backscatter of light from liver tissue via two photodiodes. Measurement of optical backscatter (Mantel–Cox P < 0.0001) and histopathological scoring of macrovesicular steatosis (Mantel–Cox P = 0.046) were predictive of 5-year graft survival. Recipients with early allograft dysfunction defined according to both Olthoff (P = 0.0067) and MEAF score (P = 0.0097) had significantly higher backscatter levels from the donor organ. Backscatter was predictive of graft loss (AUC 0.75, P = 0.0045). This study demonstrates the feasibility of real-time measurement of optical backscatter in donor livers. Early results indicate readings correlate with steatosis and may give insight to graft outcomes such as early allograft dysfunction and graft loss.