Induction of stress response proteins and experimental renal ischemia/reperfusion

BACKGROUND: The induction of stress response (heat shock) proteins (HSPs) is a highly conserved response that protects many cell types from diverse physiological and environmental stressors. We tested the hypothesis that the induction of HSPs is protective in experimental renal ischemia/reperfusion injury. METHODS: The effect of prior heat stress was examined in a rat model of renal ischemia. Postischemic renal function, histopathology, myeloperoxidase activity, and mortality were determined in hyperthermia and sham hyperthermia groups. RESULTS: HSP84, HSP70, and HSP22 mRNA were increased after eight minutes but not four minutes of hyperthermia. The induction of HSP84 and HSP70 was blocked by pretreatment with quercetin. Improvement in renal function, mortality, and histologic abnormalities was seen with eight minutes of hyperthermia six hours before ischemia. Protection was dependent on the timing of ischemia relative to heat stress and was not observed when HSPs were not induced. Postischemic increases in renal myeloperoxidase activity were markedly attenuated in the hyperthermia compared with the sham hyperthermia group. CONCLUSION: Endogenous protective mechanisms may be important in renal ischemia/reperfusion injury.     

Comparison of ischemic preconditioning and intermittent and continuous inflow occlusion in the murine liver

OBJECTIVE: To compare protection of the liver by ischemic preconditioning and intermittent inflow occlusion in a mouse model of prolonged periods of ischemia. SUMMARY BACKGROUND DATA: Preconditioning (short ischemic stress prior to a prolonged period of ischemia) and intermittent inflow occlusion protect the liver against reperfusion injury. This is the first study comparing these two modalities with continuous inflow occlusion (control). METHODS: Mice were subjected to 75 or 120 minutes of 70% hepatic ischemia and 3 hours of reperfusion. Each ischemic period was evaluated using three different protocols: continuous ischemia (control), preconditioning (10 minutes ischemia and 15 minutes reperfusion) prior to the prolonged ischemic insult, and intermittent clamping (cycles of 15 minutes ischemia and 5 minutes reperfusion). Organ injury was evaluated using serum levels of aspartate aminotransferase (AST), hematoxylin and eosin staining, and specific markers of apoptosis (cytochrome C release, caspase 3 activity, and TUNEL staining). Animal survival was determined using a model of total hepatic ischemia. RESULTS: Intermittent inflow occlusion and ischemic preconditioning were both protective against ischemic insults of 75 and 120 minutes compared with controls (continuous ischemia only). Protection against 75 minutes of ischemia was comparable in the intermittent clamping and the ischemic preconditioning group, whereas intermittent clamping was superior at 120 minutes of ischemia. One hundred percent animal survival was observed after 75 minutes of total hepatic ischemia using both protective protocols, whereas all animals subjected to continuous ischemia died after surgery. After 120 minutes of ischemia, intermittent inflow occlusion was associated with better animal survival (71%) compared with preconditioning (14%). CONCLUSIONS: Preconditioning and intermittent clamping are both protective against prolonged periods of ischemia. In the clinical setting, preconditioning is superior for ischemic periods of up to 75 minutes because it is not associated with blood loss during transection of the liver. However, for prolonged ischemic insults exceeding 75 minutes, intermittent clamping is superior to preconditioning.     

缺血预处理和Caspase抑制剂对缺血再灌注损伤保护作用的比较

目的　比较缺血预处理和Caspase抑制剂治疗对大鼠肝缺血再灌注的保护作用及其机制。方法　SD大鼠随机分为 6组 :( 1)缺血再灌注1 (IR1 )组 ;( 2 )IR2 组 ;( 3 )缺血预处理1 (IP1 )组 ;( 4 )IP2 组 ;( 5 )Caspase抑制剂治疗1 (T1 )组 ;( 6)T2 组。比较各组大鼠 70 %肝脏 60min或 12 0min缺血 ,在再灌注 3h时的肝组织Caspase 3活性 ,肝细胞凋亡率和血清AST和ALT水平 ,及实验动物 7d存活率。结果　在 60min缺血及 3h再灌注时间 ,IP1 组和T1 组的保护作用相同 ,在 12 0min缺血及 3h再灌注时 ,T2 组对Caspase活性和肝细胞凋亡的抑制优于IP2 组 (P <0 .0 1) ,但两者的AST和ALT水平及动物 7d存活率均无显著性差异。结论　缺血预处理和Caspase抑制剂治疗对鼠肝缺血再灌注损伤都有保护作用 ,两者的保护效果无显著性差异。缺血预处理对缺血再灌注损伤的保护更简便、经济、安全 ,临床应用前景十分广阔。     

Increased ischemic injury in old mouse liver: an ATP-dependent mechanism.

Although livers exhibit only minimal morphologic changes with age, how older livers tolerate pathologic conditions such as normothermic ischemia is unknown. Young 6-week-old mice and old 60-week-old mice underwent 60 minutes of hepatic ischemia and various periods of reperfusion. Markers of hepatocyte injury, hepatic energy content, and mitochondrial function were determined. Ischemic preconditioning and glucose injection were evaluated as protective strategies against reperfusion injury in old mice. Reperfusion injury was far worse in old mice compared with mice in the young control group. Ischemic preconditioning was highly protective against reperfusion injury in young but not in old mice. Older livers had dramatically reduced adenosine triphosphate (ATP) levels and glycogen contents. The low intrahepatic energy level in old mice was associated with a reduced mitochondrial ATP production. Preoperative injection of glucose restored the intrahepatic ATP content and protected against reperfusion injury. Furthermore, glucose injection restored the protective effect of ischemic preconditioning, resulting in additive protection when both strategies were combined. Aging of the liver is associated with mitochondrial dysfunction and decreased intrahepatic energy content, resulting in poorer tolerance against ischemic injury. Improving intrahepatic ATP levels in old livers by glucose injection protects the old liver against ischemic injury and restores the protective effects of ischemic preconditioning.     

Preconditioning protects against ischemia/reperfusion injury of the liver

Ischemic preconditioning (IPC) of an organ may induce protection against the injury caused by longer duration of ischemia and subsequent reperfusion. In a standardized model of such injury in the rat liver, we used the following protocol to investigate whether adenosine played a role in IPC by preventing its enzymatic degradation by dipyridamole pretreatment according to the following protocol: group 1, non-ischemic control rats; group 2, ischemic control rats subjected to 60 minutes of ischemia by clamping of the common hepatic artery followed by 60 minutes of reperfusion; group 3, IPC with 10 minutes of ischemia followed by 15 minutes of reperfusion, prior to the ischemia/reperfusion period as in group 2; group 4, pharmacologic preconditioning with administration of dipyridamole prior to the ischemia/reperfusion period as in group 2. Peripheral liver blood flow was significantly reduced during clamping (groups 2 to 4). After unclamping, blood flow was still reduced in the ischemic rats (group 2) but had returned to preclamp values in the animals that had been subjected to ischemic (group 3) or pharmacologic (group 4) preconditioning. Liver cell injury was significantly increased in the ischemia group (group 2) only. In our experimental model of ischemia/reperfusion injury in the rat liver, we found an equally beneficial effect with ischemic and pharmacologic preconditioning. Adenosine appears to be a crucial factor in IPC.     

Role of EP4 prostaglandin E2 receptor in the ischemic liver

Prostaglandin E(2) (PGE(2)) mediates a variety of both innate and adaptive immunity responses through 4 distinct receptors, EP1-4. Recent studies have suggested the physiological and pathological role of EP4 in various inflammatory diseases. In this study, we investigated the importance of the EP4 receptor, and the efficacy of a selective EP4 agonist to alter hepatic ischemia/reperfusion (I/R) injury, an important cause of damage in liver resection and transplantation. We used an established murine I/R injury model, 70% partial hepatic ischemia for 90 minutes in male C57BL/6 mice. The local expression of EP4 messenger RNA (mRNA) in the naive and the ischemic liver at 2 hours after reperfusion was examined using RT-PCR analysis. Some mice received the EP4 selective agonist during I/R. Serum aspartate aminotransferase (AST) and alanine aminotransferase (ALT) were measured as markers of hepatic injury. EP4 expression in the liver was significantly up-regulated at 2 hours after reperfusion. Furthermore, treatment with EP4 agonist significantly inhibited hepatic injury at 6 hours after reperfusion. Our data suggest an inhibitory role of EP4 PGE(2) receptor in hepatic I/R injury and the therapeutic efficacy of a selective EP4 agonist for liver protection.     

Efficacy of HSP72 induction in rat liver by orally administered geranylgeranylacetone

Abstract It is well known that heat‐shock proteins (HSPs) have a cyto‐protective function as “molecular chaperones” when cells are exposed to several stress conditions. Geranylgeranylacetone (GGA) is an antiulcer drug that was developed in Japan and it has recently been reported to induce HSP72 in rat gastric mucosa. In this experiment, we investigated the induction of HSP72 in rat liver in response to oral administration of GGA and assessed its ability to induce tolerance to warm ischemic injury by this approach. We prepared donor rats by orally administering GGA to them and compared HSP72 expression in graft liver, survival rates, and serum TNF‐α concentrations after liver transplantation with the findings in controls. The survival rates were significantly increased when the livers were obtained from donor rats given GGA. Western blotting revealed expression of HSP72 in graft livers given GGA, and the serum TNF‐α levels were significantly suppressed in the rats given GGA. Oral administration of GGA induced HSP72 in graft livers, and they were better able to tolerate warm ischemic injury. Oral administration of GGA appears to provide a promising new strategy for preventing ischemia‐reperfusion injury.     

Interleukin-6 and STAT3 protect the liver from hepatic ischemia and reperfusion injury during ischemic preconditioning

BACKGROUND: Ischemic preconditioning has been shown to protect the liver from ischemia/reperfusion injury. We hypothesized that IL-6 directly modulates the protective effects of ischemic preconditioning. METHODS: Three weeks after undergoing splenic transposition, wild-type C57BL/6 and IL-6 null mice underwent 75 minutes of total hepatic ischemia with or without prior ischemic preconditioning (10 minutes of ischemia followed by 15 minutes of reperfusion). After reperfusion, serum ALT, serum IL-6, hepatic IL-6 mRNA, hepatic pSTAT3, and liver histology were evaluated. RESULTS: In wild-type mice, survival at 24 hours was greater in the preconditioned group compared with the non-preconditioned group (75% vs 40%, P<.05). In IL-6 null mice, however, ischemic preconditioning did not improve survival when compared with the non-preconditioned group. Preconditioning significantly reduced hepatocellular injury in wild-type mice (P<.05) when compared with IL-6 null animals. This protection was associated with significant increases in serum IL-6, hepatic IL-6 mRNA, and hepatic pSTAT3 levels (P<.05). The protective effects of ischemic preconditioning that correlated with significant increases in systemic IL-6, hepatic IL-6 mRNA abundance, and pSTAT3 levels, were not observed in IL-6 null mice. CONCLUSIONS: The protective effects of ischemic preconditioning during total hepatic ischemia/reperfusion injury are dependent on IL-6 signaling and are associated with increased phosphorylation of hepatic STAT3.     

Protective effects of ischemic preconditioning for liver resection performed under inflow occlusion in humans

OBJECTIVE: To determine whether ischemic preconditioning protects the human liver against a subsequent period of ischemia in patients undergoing hemihepatectomy, and to identify possible underlying protective mechanisms of ischemic preconditioning, such as inhibition of hepatocellular apoptosis. SUMMARY BACKGROUND DATA: Ischemic preconditioning is a short period of ischemia followed by a brief period of reperfusion before a sustained ischemic insult. Recent studies in rodents suggest that ischemic preconditioning is a simple and powerful protective modality against ischemic injury of the liver. The underlying mechanisms are thought to be related to downregulation of the apoptotic pathway. METHODS: Twenty-four patients undergoing hemihepatectomy for various reasons alternatively received ischemic preconditioning (10 minutes of ischemia and 10 minutes of reperfusion) before transection of the liver performed under inflow occlusion for exactly 30 minutes. Liver wedge and Tru-cut biopsy samples were obtained at the opening of the abdomen and 30 minutes after the end of the hepatectomy. Serum levels of aspartate transferase, alanine transferase, bilirubin and prothrombin time were determined daily until discharge. Hepatocellular apoptosis was evaluated by in situ terminal deoxynucleotidyl transferase mediated d-UTP nick end-labeling (TUNEL) assay and electron microscopy. Caspase 3 and 8 activities were measured in tissue using specific fluorometric assays. RESULTS: Serum levels of aspartate transferase and alanine transferase were reduced by more than twofold in patients subjected to ischemic preconditioning versus controls. The analysis of a subgroup of patients with mild to moderate steatosis indicated possible increased protective effects of ischemic preconditioning. In situ TUNEL staining demonstrated a dramatic reduction in the number of apoptotic sinusoidal lining cells in the ischemic preconditioning group. Electron microscopy confirmed features of apoptosis present in control but not in ischemic preconditioning patients. There was no significant difference in caspase 3 and 8 activity when patients with ischemic preconditioning were compared with controls. CONCLUSIONS: Ischemic preconditioning is a simple and effective modality protecting the liver against subsequent prolonged periods of ischemia. This strategy may be a more attractive technique than intermittent inflow occlusion, which is associated with increased blood loss during each period of reperfusion.     

Clinical Aspect of Cytokines and Chemokines in Liver Transplant

BackgroundThe objective of this prospective observational study was to determine whether a transplanted liver graft releases proinflammatory cytokines and chemokines on reperfusion and to determine the relationship between these molecules and subsequent ischemic reperfusion injury and acute kidney injury.MethodsBlood samples from 66 consecutive patients undergoing liver transplant were analyzed for cyto- and chemokines at different time frames before and after liver transplant. Ischemic reperfusion injury was defined based on the peak levels of arginine transaminase and divided into 3 groups: mild, moderate, and severe. Acute kidney injury was defined according to the latest Kidney Disease: Improving Global Outcomes classification.ResultsFor more than 40 different cyto/chemokines and growth factors, a certain pattern of expression was observed in all patients with ischemic reperfusion injury. G-SCF, IP10, and HSP90a were significantly elevated in all patients with ischemic reperfusion injury. On the other hand, eotaxin and MCP1 levels were markedly elevated in patients without ischemic reperfusion, suggesting a possible cytoprotective effect. We identified cold ischemia, macrosteatosis > 30%, postreperfusion syndrome, and postoperative use of 2 or more vasoactive agents as independent risk factors for ischemic reperfusion injury.ConclusionsIschemia reperfusion injury is accompanied by distinct innate and adaptive immune cytokine signatures before and after transplant. It can be used for therapeutic intervention with goal to improve post-transplant graft outcomes.     

Ergothioneine pretreatment protects the liver from ischemia-reperfusion injury caused by increasing hepatic heat shock protein 70

BACKGROUND: Reperfusion of the liver after ischemia induces the expression of the heat shock genes and the synthesis of the heat shock proteins (HSP). We studied the effects of the natural antioxidant ergothioneine (EGT) treatment on the expression of HSP70 in ischemic-reperfused (IR) liver. METHODS: Adult male Wistar rats were randomly divided into three groups: Sham group given standard laboratory chow and water for 3 weeks followed by sham operation; Control group given standard laboratory chow and water for 3 weeks followed by liver IR injury; EGT group given standard laboratory chow supplementation l-ergothioneine (1.2 mg/kg/d body weight) administered by gavage and water for 3 weeks followed by liver IR injury. Ten rats from each group were killed to determine serum aspartate aminotransferase (AST), alanine aminotransferase (ALT), lactic dehydrogenase (LDH), tissue malondialdehyde (MDA), HSP70 levels, and histologic changes at 30, 60, and 120 min of reperfusion, respectively. Survival was followed for 1 week. RESULTS: IR caused significant increase in serum AST, ALT, LDH, and tissue MDA levels. As compared with the control group, animals treated with EGT experienced a significant decrease in serum AST, ALT, and LDH levels in all reperfusion periods. Tissue MDA levels in animals receiving EGT were significantly reduced as compared with control group at 30 min and 60 min after reperfusion. After ischemia, reperfusion caused a remarkable production of HSP70 in the control group. When the rats were pretreated with EGT, the levels of HSP70 increased significantly in their livers after reperfusion compared with the control group. Liver injury in the EGT-treated animals was lower to that in the control group. The 7-day survival rate was significantly improved (from 50% to 80%) by EGT pretreatment. CONCLUSION: HSP70 has been shown to induce tolerance against warm IR injury in rat livers. EGT pretreatment protects the liver from IR injury by over-expression of HSP and the subsequent suppression of lipid peroxidation.     

Ischemic preconditioning and liver tolerance to warm or cold ischemia: experimental studies in large animals

BACKGROUND: In the rodent, ischemic preconditioning (IPC) has been shown to improve the tolerance of the liver to ischemia-reperfusion under normothermic or hypothermic conditions. The aim of the present study was to test this hypothesis in a dog model, which may be more relevant to the human. METHODS: Beagle dogs were used in two distinct animal models of hepatic warm ischemia and orthotopic liver transplantation (hypothermic ischemia). IPC consisted of 10 minutes of ischemia followed by 10 minutes of reperfusion. In the first model, livers were exposed to 55 minutes prolonged warm ischemia and reperfused for 3 days (n = 6). In the second model, livers were retrieved and preserved for 48 hours at 4 degrees C in University of Wisconsin solution, transplanted, and reperfused without immunosuppression for 7 days (n = 5). In each model, nonpreconditioned animals served as controls (n = 5 in each group). Also, isolated dog hepatocytes were subjected to warm and cold storage ischemia-reperfusion to model the animal transplant studies using IPC. RESULTS: In the first model (warm ischemia), IPC significantly decreased serum aminotransferase activity at 6 and 24 hours post-reperfusion. After 1 hour of reperfusion, preconditioned livers contained more adenosine triphosphate and produced more bile and less myeloperoxidase activity (neutrophils) relative to controls. In the second model (hypothermic preservation), IPC was not protective. Finally, IPC significantly attenuated hepatocyte cell death after cold storage and warm reperfusion in vitro. CONCLUSIONS: IPC is effective in large animals for protecting the liver against warm ischemia-reperfusion injury but not injury associated with cold ischemia and reperfusion (preservation injury). However, the IPC effect observed in isolated hepatocytes suggests that preconditioning for preservation is theoretically possible.     

Protective Effect of Citric Acid Against Hepatic Ischemia Reperfusion Injury in Sprague-Dawley Rats

ObjectiveHepatic ischemia reperfusion (I/R) injury is regarded as a serious concern in clinical practice. Citric acid reduces oxidative stress and inflammation during hypoxia and reoxygenation. Our objective was to investigate the protective effect of citric acid against hepatic I/R injury in rats.MethodsWe fed Sprague-Dawley rats either citric acid (100 mg/kg/d) or saline. One week later, ischemia was induced by clamping the rats’ common hepatic artery and portal vein for 30 minutes. The rats were randomly divided into 3 major groups that were treated as follows: 1. the sham operated group; 2. the I/R group; and 3. the I/R-citric acid group.ResultsCompared to the sham group, the I/R group had higher expression of aspartate aminotransferase and alanine aminotransferase and lower expression of catalase, superoxide dismutase, glutathione peroxidase, antioxidant, nitric oxide, and albumin. Compared to the I/R group, the I/R-citric acid group had higher expression of catalase, superoxide dismutase, antioxidants, and nitric oxide, and lower expression of aspartate aminotransferase and alanine aminotransferase.ConclusionsThese results suggest that citric acid therapy has significant therapeutic potential in ischemic liver injury.     

Effect of prostaglandin E1 derivative on labilization of liver lysosomal membrane in partial liver ischemia]

The change of liver lysosomal enzymes in tissue and serum during a reperfusion period was studied in partial liver ischemic model in rats and effect of Prostaglandin E1 (PGE1) derivative on partial liver ischemia was investigated. Partial liver ischemia was induced by clamping the branches of the vessels to the right and caudate lobes of rat liver. The clamp was released after 30 minutes of ischemia. Ischemic and nonischemic lobes of the liver were separately removed and the serum was also collected immediately and two hours after the release of the clamp. Lysosomal enzyme activities from free and bound lysosomal fraction were measured separately and the fragility index (F.I.) was calculated. PGE1 derivative was administered intraperitoneally 24, 6, 0.5 hours prior to the induction of ischemia at each dose of 0.05 microgram/kg. Pretreatment with PGE1 derivative prevented lysosomal labilization in ischemic lobe, since there was a significant decrease in F.I. of cathepsin D in the PGE1-pretreated group (preischemia; 28.3 +/- 2.4%, immediately after reperfusion; 30.3 +/- 2.5%, two hours after reperfusion; 30.3 +/- 2.5%) compared to the placebo group (immediately after reperfusion; 40.9 +/- 3.4%, two hours after reperfusion; 41.7 +/- 3.4%, p less than 0.05, p less than 0.05, p less than 0.01, respectively). Pretreatment with PGE1 derivative also significantly suppressed the increase of serum lysosomal enzyme activity. These results showed that PGE1 derivative improved liver lysosomal labilization in partial liver ischemia.     

Experimental study of the effect of intraportal prostaglandin E1 on hepatic blood flow during reperfusion after ischaemia and hepatectomy.

BACKGROUND: Prostaglandin E1 (PGE1) has protective effects experimentally and clinically in individual models of hepatic ischaemia-reperfusion injury and of partial hepatectomy. The present study investigated the effects of intraportal administration of PGE1 on hepatic blood flow, systemic arterial pressure and long-term animal survival after 60 min of total liver ischaemia followed by 70 per cent partial hepatectomy in rats. METHODS: Total liver ischaemia was induced by occluding the hepatoduodenal ligament for 60 min. PGE1 0.5 microg per kg per min was infused intraportally for 15 min before inducing ischaemia and for 120 min after ischaemia in the treatment group. Normal saline was infused in the control group. During ischaemia 70 per cent partial hepatectomy was performed. Portal venous flow (PVF), peripheral tissue blood flow (PTBF) and hepatic artery flow were measured before and after ischaemia. Serum biochemical analysis was carried out at 1, 3 and 24 h, and 7 and 14 days; and liver histology at 1 and 24 h, and 7 days after reperfusion. Survival was followed for 1 year. RESULTS: Intraportal infusion of PGE1 significantly improved PVF and PTBF without affecting the systemic arterial pressure. Long-term survival was significantly higher in the PGE1 group. Serum aspartate aminotransferase, alanine aminotransferase and alkaline phosphatase levels decreased significantly, and 2-h bile flow was significantly improved, in the PGE1 group. Histological examination revealed significant portal venous congestion, sinusoidal congestion, fatty degeneration and tissue necrosis 24 h and 7 days after reperfusion in the control group. CONCLUSION: PGE1 has a protective effect against liver damage when the liver is injured by warm ischaemia and reperfusion followed by partial resection.     

热休克蛋白70对肝硬化大鼠肝脏缺血再灌注损伤保护作用的研究

目的 探讨热休克蛋白70（HSP70）对肝硬化大鼠肝脏缺血再灌注损伤的保护作用。方法Wistar雄性大鼠用四氯化碳皮下注射制成肝硬化模型。分两组:IP组（缺血预处理组）,用 Pringle’s法阻断肝门15min后,恢复血供,关腹;C组（对照组）,只予以开、关腹。48h后,再次阻断肝门30min,恢复血供。用Western blotting法检测IP后6、24、48h肝组织中HSP70的表达水平;测再灌注1h血清生化酶（ALT、AST、LDH）水平;计算术后一周生存率,并行肝脏病理组织学检查。结果 IP组在缺血预处理后24-48hHSP70表达显著增强,呈高水平;而C组中在各时点HSP70均无表达增强。再灌注1h,IP组的ALT、AST、LDH水平显著低于C组（P<0.01或P<0.05,n=7）。术后一周生存率IP组（93.10％,n=29）明显高于C组（73.33％,n=30）（P<0.05）。缺血再灌注后1h,IP组的肝细胞损伤明显轻于C组。结论 HSP70能够减轻肝硬化大鼠肝脏缺血再灌注损伤,提高术后生存率。     

不同预处理对肝硬化兔缺血再灌注肝组织HSP70及ET-1表达的影响

目的:探讨两种预处理方式,即经典缺血预处理(IPC)与肢体缺血预处理(LIPC),对肝硬化兔肝缺血再灌注(I/R)损伤的保护作用及可能的作用机制。方法:皮下注射CCl4-橄榄油溶液制备兔肝硬化模型,随后将模型兔随机分为假手术组,肝I/R组(I/R组),IPC+肝I/R组(IPC组),LIPC+肝I/R组(LIPC组),每组7只。肝I/R模型制作方法:阻断入肝血流30 min,再灌注2 h;IPC诱导方法:在行肝I/R处理前阻断入肝血流10 min,开放10 min;LIPC诱导方法:在行肝I/R处理前24 h,采用止血带捆扎兔单侧后肢5 min,再开放5 min,重复3次。各组于再灌注2 h后切取肝组织,行组织形态学观察,用ELISA法测定内皮素1(ET-1)含量及Western blot法检测热休克蛋白(HSP70)的表达。结果:与假手术组比较,其余各组在肝硬化病变的基础上均出现不同程度的变性、水肿和炎性细胞浸润,但IPC组与LIPC组明显轻于I/R组,而LIPC组及IPC组间病变程度无明显差异;与假手术组比较,其余各组肝组织ET-1含量和HSP70的表达均明显增加(均P<0.05),但IPC组与LIPC组肝组织ET-1含量低于I/R组,HSP70的表达高于I/R组(均P<0.05),而上述2项指标在LIPC及IPC组间均无统计学差异(均P>0.05)。结论:LIPC和IPC均能对肝硬化肝I/R损伤有保护作用,且保护强度相似,其机制可能均与抑制ET-1的释放及增加HSP70的表达有关;LIPC具有无创性,可能具有更大的临床应用前景。     

Heme oxygenase-1 attenuates ischemia/reperfusion-induced apoptosis and improves survival in rat renal allografts

BACKGROUND: Kidneys can be preserved only for a limited time without jeopardizing graft function and survival. Induction of heat shock proteins (HSPs) can protect against ischemia/reperfusion (I/R) injury. Therefore, we investigated whether the induction of the HSP, heme oxygenase-1 (HO-1), improves outcome following isotransplantation after an extended period of cold storage. METHODS: Rats were subjected to heat preconditioning (HP; 42 degrees C for 20 minutes). Kidneys harvested after 24 hours, were preserved in cold University of Wisconsin (UW) solution at 4 degrees C for 45 hours and transplanted into bilateral nephrectomized rats. Cobalt protoporphyrin (CoPP) was administered in another group of animals in order to induce HO-1 pharmacologically, while other groups of animals received the HO-1 inhibitor, tin protophorphyrine (SnPP), following HP or CoPP. RESULTS: Cold ischemia caused a complete attenuation of graft function within 3 days following transplantation and subsequent death of all animals, whereas HP protected graft function and five of nine rats survived for 3 weeks. HP inhibited the induction of osteopontin and induced the expression of HO-1, HSP 70 and 90, and the antiapoptotic factor Bcl-XL. Grafts exposed to HP were protected against structural I/R injuries as revealed by histologic assessment using a semiquantitative score. Furthermore, induction of apoptosis was attenuated and activation of caspase-3 was inhibited. Comparable results were observed after administration of CoPP, whereas SnPP inhibited the effects of HP and CoPP. CONCLUSION: HP or administration of CoPP induced both HO-1, preserved kidney graft function, and prevented postreperfusion apoptosis after cold preservation.     

Prolonged survival and decreased mucosal injury after low-dose enteral allopurinol prophylaxis in mesenteric ischemia

Previous studies demonstrating protective effects of allopurinol in intestinal ischemia have evaluated intravenous allopurinol (presently unavailable for human use) or enteral allopurinol at supranormal doses and, therefore, have questionable clinical relevance. To address this problem, we evaluated the protective effects of clinically used doses of enteral allopurinol in rats with intestinal ischemia. Forty male Sprague-Dawley rats (weighing 300 to 400 g) received enteral allopurinol (10 mg/kg) or water daily for 1 week. Rats were then subjected to superior mesenteric artery occlusion with interruption of collateral flow for 20 minutes to produce ischemic injury to the intestine. Segmental small bowel resections were performed in 10 control rats and 10 allopurinol-treated rats before and after reperfusion to identify histopathologic evidence of reperfusion injury. Mucosal injury was quantitated using a grading scale of 0 to 5 (5 being most severe). The remaining 20 rats (10 in each group) were observed for mortality (death within 7 days) after reperfusion. Mucosal injury after reperfusion was graded at 4.4 +/- 0.20 in controls versus 2.3 +/- 0.23 in the treated group (P less than .001). In addition, there was a significant increase in mucosal damage in the control group when postreperfusion specimens were compared with specimens taken before reperfusion (2.8 +/- 0.19 before and 4.4 +/- 0.20 after reperfusion, P less than .001). Injury score for the allopurinol-treated group did not significantly increase after reperfusion. Survival was 50% in the water-fed control group compared with 100% survival in allopurinol-treated rats (P = .016). We conclude that enteral allopurinol in the presently available form and dose is effective in reducing mesenteric reperfusion injury.