Plasma D(-)-lactate as a new marker for diagnosis of acute intestinal injury following ischemia-reperfusion

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Protective effect of pyrrolidine dithiocarbamate on liver injury induced by intestinal ischemia-reperfusion in rats

BACKGROUND: The nuclear translocation of transcription factors may be a critical factor in the intracellular pathway involved in ischemia/reperfusion(I/R) injury. The aim of the study was to evaluate the role of nuclear factor-kappa B (NF-κB) in the pathogenesis of liver injury induced by intestinal ischemia/reperfusion (IIR) and to investigate the effect of pyrrolidine dithiocarbamate (PDTC) on this liver injury. METHODS: Male Wistar rats were divided randomly into three experimental groups (8 rats in each): sham operation group (control group); intestinal/reperfusion group(I/R group): animals received 1-hour of intestinal ischemia and 2-hour reperfusion; and PDTC treatment group (PDTC group): animals that received I/R subject to PDTC treatment (100 mg/kg). The histological changes in the liver and intestine were observed, and the serum levels of tumor necrosis factor-α (TNF-α), alanine aminotransferase (ALT), aspartate aminotransferase (AST), liver superoxide dismutase (SOD), and nitrite/nitrate (NO) were measured. The immunohistochemical expression and Western blot analysis of liver NF-κB and intercellular adhesion molecule-1(ICAM-1) were observed. RESULTS: IIR induced liver injury characterized by the histological changes of liver edema, hemorrhage, polymorphonuclear neutrophil (PMN) infiltration, and elevated serum levels of AST and ALT. The serum TNF-α level was significantly higher than that of the control group(P<0.01) and a high level of liver oxidant product was observed (P<0.01). These changes were parallel to the positive expression of NF-κB and ICAM-1. After the administration of PDTC, the histological changes after liver injury were improved; the levels of SOD and NO in the liver were elevated and reduced, respectively (P<0.01). The expressions of ICAM-1 and NF-κB in the liver were weakened (P<0.01). CONCLUSION: NF-κB plays an important role in the pathogenesis of liver injury induced by HR. PDTC, an agent known to inhibit the activation of NF-κB, can reduce and prevent this injury.

Attenuation of graft ischemia-reperfusion injury by urinary trypsin inhibitor in mouse intestinal transplantation

AIM: Ischemia/reperfusion (I/R) injury is one of the major obstacles for intestinal transplantation (ITx). Urinary trypsin inhibitor (Ulinastatin, UTI) suppresses proteases and stabilizes lysosomal membranes. We supposed that Ulinastatin would diminish I/R injury of intestinal graft.METHODS: UTI- treated group and untreated control group were investigated by histological assessment at 1.5, 4, 24, and 72 h after ITx. Myeloperoxidase (MPO)activity was used as the activity of neutrophils, and malondialdehyde (MDA) was used as an index of lipid peroxidation. TNFα and i-NOS mRNA expression in graft tissue were measured by semi-quantitative RT-PCR.CD11b+ Gr1+ cells in graft lamina propria were analyzed by flow cytometry.RESULTS: Histological scores of the graft showed that the tissue injury was markedly attenuated by UTI treatment at different time points after ITx, with reduced MPO and MDA value in the grafts. The expression of TNFα and i-NOS mRNA was profoundly inhibited, while the infiltration of CD11b+ Gr1+ cells into the intestinal graft was decreased in UTI group.CONCLUSION: Urinary trypsin inhibitor attenuates I/R injury in mouse intestinal transplantation by reducing monocytes infiltration and down-regulation of TNFα and i-NOS mRNA expression.

Intestinal ischemia-reperfusion of macaques triggers a strong innate immune response

AIM:To investigate inflammatory injury in the intestinal mucosa after intestinal ischemia-reperfusion(ⅡR) with Toll-like receptor(TLR)-mediated innate immunity.METHODS:Ten macaques were randomized into control and ⅡR groups.The distribution and expression level of TLR2,TLR4,MD2,nuclear factor(NF)-κB p65 and interferon(IFN)-γ were measured by immunohistochemical stain and western blotting.The mRNA expression of TLR4,TLR2,MD2,interleukin(IL)-1β and tumor necrosis factor(TNF)-α were measured by reverse transcriptase-polymerase chain reaction.The cytokine levels in blood and intestinal tissues were measured by ELISA.RESULTS:Obvious hemorrhage and erosion of mucosae were seen in the ⅡR group.Expression of TLR2,TLR4,MD2,NF-κB p65 and IFN-γ was significantly higher in the ⅡR group than in the control group(0.13 ± 0.04,0.22 ± 0.04,0.16 ± 0.06,0.65 ± 0.12,0.38 ± 0.10 vs 0.07 ± 0.04,0.08 ± 0.03,0.04 ± 0.02,0.19 ± 0.06,0.14 ± 0.05,P < 0.05).In addition,the expression of TLR2,TLR4,MD2,IL-1β and TNF-α mRNA in the ⅡR group were significantly higher than those of control group(1.52 ± 0.15,1.39 ± 0.06,1.94 ± 0.12,1.48 ± 0.15,0.66 ± 0.08 vs 0.31 ± 0.05,0.5 ± 0.04,0.77 ± 0.05,0.35 ± 0.08,0.18 ± 0.04,P < 0.05).Furthermore,IL-1β,IL-6 and TNF-α levels in the macaques ileum and plasma were significantly higher than in the control group(plasma:86.3 ± 15.2,1129 ± 248.3,77.8 ± 16.2 vs 29.5 ± 7.3,19.8 ± 8.2,5.6 ± 1.7; ileum:273.4.± 44.7,1636 ± 168.0,205.5 ± 30.7 vs 76.8 ± 20.5,663.4 ± 186.9,49.0 ± 9.4; P < 0.05).CONCLUSION:After ⅡR,general inflammatory injury in the intestinal mucosa is correlated with a strong innate immune response,mediated by activation of the TLR-NF-κB-cytokine pathway.

Effect of intestinal ischemia-reperfusion injury on protein levels of leptin and orexin-A in peripheral blood and central secretory tissues

AIM: To explore the effect of intestinal ischemia-reperfusion injury on protein levels of leptin and orexin-A in peripheral blood and their central secretory tissues and to find out the role leptin and orexin-A play in acute inflammatory responses.METHODS: An intestinal ischemia-reperfusion (I/R)injury model of rats was established and rats were divided randomly into six groups: sham-operation group, 60 min ischemia/30 min reperfusion group (I60'R30'), I60'R90',I60'R150', I60'R240' and I60'R360', 9 rats each group.Two highly-sensitive radioimmunoassays for leptin and orexin-A were established and used to check the change of their concentrations in peripheral blood and central secretory tissues before and after intestinal I/R injury.RESULTS: Compared with the serum leptin level before injury, it decreased significantly in I60'R30' group and increased significantly in I60'R360' group; compared to sham-operation group after injury, serum leptin level increased significantly in I60'R360' group; compared to sham-operation group after injury, adipose leptin levels decreased significantly in I60'R30' and I60'R90' groups,while increased significantly in I60'R360' group. There was no significant difference between the expression levels of orexin-A before and after I/R injury.CONCLUSION: Leptin has a time-dependent response and orexin-A has a delayed response to acute inflammatory stimuli such as intestinal I/R injury and they may participate in metabolic disorders in injury as inflammatory cytokines.

Effects of penehyclidine hydrochloride in small intestinal damage caused by limb ischemia-reperfusion

AIM:To investigate the protective effect of penehyclidine hydrochloride post-conditioning in the damage to the barrier function of the small intestinal mucosa caused by limb ischemia-reperfusion(LIR) injury. METHODS:Male Wistar rats were randomly divided into three groups(36 rats each) :the sham-operation group(group S) ,lower limb ischemia-reperfusion group(group LIR) ,and penehyclidine hydrochloride postconditioning group(group PHC) .Each group was divided into subgroups(n=6 in each group) according to ischemic-reperfusion time,i.e.immediately 0 h(T1) ,1 h(T2) ,3 h(T3) ,6 h(T4) ,12 h(T5) ,and 24 h(T6) .Bilateral hind-limb ischemia was induced by rubber band application proximal to the level of the greater trochanter for 3 h.In group PHC,0.15 mg/kg of penehyclidine hydrochloride was injected into the tail vein immediately after 3 h of bilateral hind-limb ischemia.The designated rats were sacrificed at different time-points of reperfusion;diamine oxidase(DAO) ,superoxide dismutase(SOD) activity,myeloperoxidase(MPO) of small intestinal tissue,plasma endotoxin,DAO,tumor necrosis factor-α(TNF-α) ,and interleukin(IL) -10 in serum were detected in the rats. RESULTS:The pathological changes in the small intestine were observed under light microscope.The levels of MPO,endotoxin,serum DAO,and IL-10 at T1-T6,and TNF-αlevel at T1-T4 increased in groups LIR and PHC(P<0.05) compared with those in group S,but tissue DAO and SOD activity at T1-T6 decreased(P<0.05) .In group PHC,the tissue DAO and SOD activity at T2-T6,and IL-10 at T2-T5 increased to higher levels than those in group LIR(P<0.05) ;however,the levels of MPO,endotoxin,and DAO in the blood at T2-T6,and TNF-αat T2 and T4 decreased(P<0.05) . CONCLUSION:Penehyclidine hydrochloride post-conditioning may reduce the permeability of the small intestines after LIR.Its protection mechanisms may be related to inhibiting oxygen free radicals and inflammatory cytokines for organ damage.

Life and death at the mucosal-luminal interface: New perspectives on human intestinal ischemia-reperfusion

Intestinal ischemia is a frequently observed phenomenon. Morbidity and mortality rates are extraordinarily high and did not improve over the past decades. This is in part attributable to limited knowledge on the pathophysiology of intestinal ischemia-reperfusion(IR) in man, the paucity in preventive and/or therapeutic options and the lack of early diagnostic markers for intestinal ischemia. To improve our knowledge and solve clinically important questions regarding intestinal IR, we developed a human experimental intestinal IR model. With this model, we were able to gain insight into the mechanisms that allow the human gut to withstand short periods of IR without the development of severe inflammatory responses. The purpose of this review is to overview the most relevant recent advances in our understanding of the pathophysiology of human intestinal IR, as well as the(potential) future clinical implications.

Bone-marrow mesenchymal stem cells reduce rat intestinal ischemia-reperfusion injury, ZO-1 downregulation and tight junction disruption via a TNF-α-regulated mechanism

AIM: To investigate the effect of bone-marrow mesenchymal stem cells (BM MSCs) on the intestinal mucosa barrier in ischemia/reperfusion (I/R) injury. METHODS: BM MSCs were isolated from male Sprague-Dawley rats by density gradient centrifugation, cultured, and analyzed by flow cytometry. I/R injury was induced by occlusion of the superior mesenteric artery for 30 min. Rats were treated with saline, BM MSCs (via intramucosal injection) or tumor necrosis factor (TNF)-α blocking antibodies (via the tail vein). I/R injury was assessed using transmission electron microscopy, hematoxylin and eosin (HE) staining, immunohistochemistry, western blotting and enzyme linked immunosorbent assay.RESULTS: Intestinal permeability increased, tight junctions (TJs) were disrupted, and zona occludens 1 (ZO-1) was downregulated after I/R injury. BM MSCs reduced intestinal mucosal barrier destruction, ZO-1 downregulation, and TJ disruption. The morphological abnormalities after intestinal I/R injury positively correlated with serum TNF-α levels. Administration of anti-TNF-α IgG or anti-TNF-α receptor 1 antibodies attenuated the intestinal ultrastructural changes, ZO-1 downregulation, and TJ disruption. CONCLUSION: Altered serum TNF-α levels play an important role in the ability of BM MSCs to protect against intestinal I/R injury.

Acid fibroblast growth factor reduces rat intestinal mucosal damage caused by ischemia-reperfusion insult

AIM: To detect the effects of acid fibroblast growth factor (aFGF) on apoptosis and proliferation of intestinal epithelial cells in differentiation or proliferation status to explore the protective mechanisms of aFGF.METHODS: Wistar rats were randomly divided into sham-operated control group (C, n = 6), intestinal ischemia group (I,n = 6), aFGF treatment group (A,n =48) and intestinal ischemia-reperfusion group (R, n = 48). Apoptosis of intestinal mucosal cells was determined with terminal deoxynucleotidyl transferasemediated dUTP-biotin nick-end labeling (TUNEL)technique. Proliferating cell nuclear. antigen (PCNA)protein expression and distribution were detected with immunohistochemical method. Plasma levels of D-lactate were determined with modified Brandts method.RESULTS: In A group, administration of exogenous aFGF could improve intestinal histological structure and decrease plasma D-lactate levels at 2-12 h after the reperfusion compared with R group. The apoptotic rates and PCNA protein expressions were not increased until 2 h after reperfusion and were maximal at 12 h. After reperfusion for 2-12 h, the apoptotic rates were gradually augmented along the length of jejunal crypt-villus units.Administration of aFGF could significantly reduce the apoptotic response at 2-12 h after reperfusion (P＜0.05).Apoptosis rates in villus and crypt epithelial cells in A group at 12 h after reperfusion were (62.5±5.5)% and (73.2±18.6)% of those in R group, respectively.Treatment of aFGF could apparently induce protein expression of PCNA in intestinal mucosal cells of A group compared with R group during 2-12 h after reperfusion (P＜0.05). There were approximately 1.3- and 1.5-times increments of PCNA expression levels in villus and crypt cells in A group at 12 h after reperfusion compared with R group, respectively.CONCLUSION: Intestinal I/R insult could lead to histological structure change and apoptotic rate increment. The protective effects of aFGF against ischemia/reperfusion in rat intestinal mucosa might be partially due to its ability to inhibit ischemia/reperfusioninduced apoptosis and to promote cell proliferation of crypt cells and villus epithelial cells.

Rosuvastatin reduces rat intestinal ischemia-reperfusion injury associated with the preservation of endothelial nitric oxide synthase protein

瞄准：为了在 ischemia-reperfusion (红外) 上调查 rosuvastatin 的保护的效果，并且在内皮的氮的氧化物 synthase (eNOS ) 的表示上决定这个代理人的效果，在老鼠导致了小肠的损害和发炎蛋白质。方法：肠的损坏被为 30 min 夹钳优异 mes 伤寒动脉和腹的箱子在男 Sprague-Dawley 老鼠导致，为 60 min 由灌注列在后面。在生理盐水溶解的 Rosuvastatin intraperitoneally 被管理在局部缺血前的 60 min。肠的粘膜损害和发炎的严厉被几个生物化学的标记，以及由组织检查所见评估。eNOS 的蛋白质层次被西方的污点决定。结果：当粘膜的索引损坏，管腔内血红素和蛋白质的层次显著地在假冒操作组与那些相比在红外组被增加。然而，这些增加被处理显著地以一种剂量依赖者方式与 rosuvastatin 禁止。rosuvastatin 的保护的效果被组织检查所见也证实。到红外的小肠的暴露导致了 thiobarbituric 的重要增加描绘的粘膜发炎酸反应的物质，联系织物的 myeloperoxidase 活动，和老鼠的粘膜内容导致 cytokine 的嗜中性的 chemoattractant-1 (CINC-1 ) 和肿瘤坏死 factor-alpha (TNF-alpha ) 。在红外以后的煽动性的参数的这些增加被预告的处理显著地在 10 mg/kg 的剂量与 rosuvastatin 禁止。而且， CINC-1 和 TNF-alpha 的 mRNA 表示在红外以后被增加，并且这增加被 rosuvastatin 也禁止。eNOS 的粘膜蛋白质层次在红外期间减少了，但是在与 rosuvastatin 对待的老鼠被保存。结论：Rosuvastatin 禁止老鼠红外导致的肠的损害和发炎，和它的保护与 eNOS 的保藏被联系蛋白质。

Lansoprazole ameliorates intestinal mucosal damage induced by ischemia-reperfusion in rats

AIM: To investigate the protective effect of lansoprazoleon ischemia and reperfusion (I/R)-induced rat intestinalmucosal injury in vivo.METHODS: Intestinal damage was induced by clampingboth the superior mesenteric artery and the celiac trunkfor 30 rain followed by reperfusion in male Sprague-Dawleyrats. lansoprazole was given to rats intraperitoneally 1 hbefore vascular clamping.RESULTS: Both the intraluminal hemoglobin and proteinlevels, as indices of mucosal damage, significantlyincreased in I/R-groups comparion with those of sham-operation groups. These increases in intraluminal hemoglobinand protein levels were significantly inhibited by the treatmentwith lansoprazole at a dose of 1 mg/kg. Small intestineexposed to I/R resulted in mucosal inflammation that wascharacterized by significant increases in thiobarbituric acid-reactive substances (TBARS), tissue-associatedmyeloperoxidase activity (MPO), and mucosal content of ratcytokine-induced neutrophil chemoattractant-1 (CINC-1).These increases in TBARS, MPO activities and CINC-1 contentin the intestinal mucosa after I/R were all inhibited bypretreatment with lansoprazole at a dose of 1 mg/kg.Furthermore, the CINC-1 mRNA expression was increasedduring intestinal I/R, and this increase in mRNA expressionwas inhibited by treatment with lansoprazole.CONCLUSION: Lansoprazole inhibits lipid peroxidation andreduces development of intestinal mucosal inflammationinduced by I/R in rats, suggesting that lansoprazole mayhave a therapeutic potential for I/R injury.

Protective effect and mechanism of clemastine fumarate on acute lung injury in mice with intestinal ischemia-reperfusion

Objective:To evaluate the protective effect and mechanism of clemastine fumarate(CLE)on acute lung injury(ALI)in intestinal ischemia-reperfusion(I/R)mice.Methods:Twenty-four SPF Balb/c mice were randomly divided into sham operation group(sham group),ischemia-reperfusion group(I/R group),and clemastine fumarate pretreatment group(I/R+C group).In the I/R group,an intestinal ischemia-reperfusion model was established(ischemia for 40 minutes,reperfusion for 2 hours).In the I/R+C group,CLE 5 mg/kg was intraperitoneally injected before the operation.Lung tissue morphology was observed and scored by HE staining;and the ratios of wet weight to dry weight(W/D)were recorded.the levels of MDA,SOD,GSH-px,NF-κB and TNF-αin lung tissue of each group were determined by ELISA;Western blot method was used to determine the expression of TLR4 protein in lung tissue.Results:Compared with the Sham group,the I/R group had significantly higher lung tissue injury score and wet/dry ratio(P<0.05),increased lung tissue MDA level(P<0.05),decreased SOD and GSH-px levels(P<0.05),and increased NF-κB and TNF-αlevels,the expression of TLR4 protein in lung tissue increased(P<0.05);compared with the I/R group,the lung tissue injury score and wet/dry ratio of the I/R+C group decreased(P<0.05),the level of MDA in lung tissue decreased(P<0.05),the levels of SOD and GSH-px increased(P<0.05),and the levels of NF-κB and TNF-毩decreased(P<0.05),the expression of TLR4 protein in lung tissue decreased(P<0.05).Conclusion:Clemastine fumarate can alleviate acute lung injury after intestinal ischemia-reperfusion in mice,and the mechanism may be related to the inhibition of oxidative stress and inflammatory response in lung tissue.

EFFECT OF PREOPERATIVE GLUTAMINE ADMINISTRATION ON ICAM-1 EXPRESSION IN RAT LUNG INDUCED BY INTESTINAL ISCHEMIA-REPERFUSION

Objective To evaluate the effect of preoperative glutamine administration on intracellular adhesion molecule-1 (ICAM-1) expression in rat lung induced by intestinal ischemia-reperfusion(I/R). MethodsSprague-Dawley rats (n=25) were randomly divided into 5 groups: sham group (sham surgery), glutamine groups (three different doses) and control group. All groups except sham were subjected to intestinal I/R injury, and superior mesenteric artery (SMA) occluded for 60 min followed by 90 min of reperfusion. Lung injury was evaluated with Evans blue dye concentration and histopathologic examination. The immunohistochemical expression and mRNA expression of ICAM-1 were measured with immunohistochemical staining and RT-PCR method respectively. The level of myeloperoxidase (MPO) was also measured with biochemistry method. Results Intestinal I/R resulted in lung injury characterized by an increase in Evans blue dye concentration, neutrophil sequestration, and obvious staining for expression of pulmonary ICAM-1, compared with sham group. The expression of ICAM-1 and the level of MPO in rat lung were lower in glutamine groups compared with control group. Conclusion I-R injury increases the expression of ICAM-1 within the lung. This may contribute to the migration, accumulation and activation of polymorphonuclear neutrophils (PMNs) after such injury. Preoperative glutamine administration attenuates rat lung injury induced by intestinal I-R, and inhibiting ICAM-1 expression maybe one of the potential mechanisms.

Sulforaphane protects liver injury induced by intestinal ischemia reperfusion through Nrf2-ARE pathway

AIM: To investigate the effect of sulforaphane (SFN) on regulation of NF-E2-related factor-2 (Nrf2)-antiox-idant response element (ARE) pathway in liver injury induced by intestinal ischemia/reperfusion (I/R). METHODS: Rats were divided randomly into four ex-perimental groups: control, SFN control, intestinal I/R and SFN pretreatment groups (n = 8 in each group). The intestinal I/R model was established by clamping the superior mesenteric artery for 1 h and 2 h reperfu-sion. In the SFN pretreatment group, surgery was performed as in the intestinal I/R group, with intraperitoneal administration of 3 mg/kg SFN 1 h before the op-eration. Intestine and liver histology was investigated. Serum levels of aspartate aminotransferase (AST), and alanine aminotransferase (ALT) were measured. Liver tissue superoxide dismutase (SOD), myeloperoxidase (MPO), glutathione (GSH) and glutathione peroxidase (GSH-Px) activity were assayed. The liver transcription factor Nrf2 and heme oxygenase-1 (HO-1) were determined by immunohistochemical analysis and Western blotting analysis.RESULTS: Intestinal I/R induced intestinal and liver injury, characterized by histological changes as well as a signif icant increase in serum AST and ALT levels (AST: 260.13 ± 40.17 U/L vs 186.00 ± 24.21 U/L, P < 0.01; ALT: 139.63 ± 11.35 U/L vs 48.38 ± 10.73 U/L, P < 0.01), all of which were reduced by pretreatment with SFN, respectively (AST: 260.13 ± 40.17 U/L vs 216.63 ± 22.65 U/L, P < 0.05; ALT: 139.63 ± 11.35 U/L vs 97.63 ± 15.56 U/L, P < 0.01). The activity of SOD in the liver tissue decreased after intestinal I/R (P < 0.01), which was enhanced by SFN pretreatment (P < 0.05). In ad-dition, compared with the control group, SFN markedly reduced liver tissue MPO activity (P < 0.05) and elevat-ed liver tissue GSH and GSH-Px activity (P < 0.05, P < 0.05), which was in parallel with the increased level of liver Nrf2 and HO-1 expression.CONCLUSION: SFN pretreatment attenuates liver injury induced by intestinal I/R in rats, attributable to the antioxidant effect through Nrf2-ARE pathway.

Fish oil alleviates liver injury induced by intestinal ischemia/reperfusion via AMPK/SIRT-1/autophagy pathway

AIM To evaluate whether fish oil(FO) can protect liver injury induced by intestinal ischemia/reperfusion(I/R) via the AMPK/SIRT-1/autophagy pathway.METHODS Ischemia in wistar rats was induced by superior mesenteric artery occlusion for 60 min and reperfusion for 240 min. One milliliter per day of FO emulsion or normal saline was administered by intraperitoneal injection for 5 consecutive days to each animal. Animals were sacrificed at the end of reperfusion. Blood andtissue samples were collected for analyses. AMPK, SIRT-1, and Beclin-1 expression was determined in lipopolysaccharide(LPS)-stimulated HepG2 cells with or without FO emulsion treatment.RESULTS Intestinal I/R induced significant liver morphological changes and increased serum alanine aminotransferase and aspartate aminotransferase levels. Expression of p-AMPK/AMPK, SIRT-1, and autophagy markers was decreased whereas tumor necrosis factor-α(TNF-α) and malonaldehyde(MDA) were increased. FO emulsion blocked the changes of the above indicators effectively. Besides, in LPS-stimulated HepG2 cells, small interfering RNA(siRNA) targeting AMPK impaired the FO induced increase of p-AMPK, SIRT-1, and Beclin-1 and decrease of TNF-α and MDA. SIRT-1 siRNA impaired the increase of SIRT-1 and Beclin-1 and the decrease of TNF-α and MDA.CONCLUSION Our study indicates that FO may protect the liver against intestinal I/R induced injury through the AMPK/SIRT-1/autophagy pathway.

Protective role of adiponectin in a rat model of intestinal ischemia reperfusion injury

AIM: To determine the potential protective role of adiponectin in intestinal ischemia reperfusion(I/R) injury.METHODS: A rat model of intestinal I/R injury was established. The serum level of adiponectin in rats with intestinal I/R injury was determined by enzymelinked immunosorbent assay(ELISA). The serum levels of interleukin(IL)-1β, IL-6, and tumor necrosis factor(TNF)-α were also measured by ELISA. Apoptosis of intestinal cells was detected using the terminal deoxynucleotidyl transferase d UTP nick end labeling assay. The production of malondialdehyde(MDA) and superoxide dismutase(SOD) and villous injury scores were also measured.RESULTS: Adiponectin was downregulated in the serum of rats with intestinal I/R injury compared with sham rats. No significant changes in the expression of adiponectin receptor 1 and adiponectin receptor 2 were found between sham and I/R rats. Pre-treatment with recombinant adiponectin attenuated intestinal I/R injury. The production of pro-inflammatory cytokines,including IL-6, IL-1β, and TNF-α, in rats with intestinal I/R injury was reduced by adiponectin pre-treatment. The production of MDA was inhibited, and the release of SOD was restored by adiponectin pre-treatment in rats with intestinal I/R injury. Adiponectin pre-treatment also inhibited cell apoptosis in these rats. Treatment with the AMP-activated protein kinase(AMPK) signaling pathway inhibitor, compound C, or the heme oxygenase 1(HO-1) inhibitor, Snpp, attenuated the protective effects of adiponectin against intestinal I/R injury. CONCLUSION: Adiponectin exhibits protective effects against intestinal I/R injury, which may involve the AMPK/HO-1 pathway.

Ischemic post-conditioning to counteract intestinal ischemia/reperfusion injury

Intestinal ischemia is a severe disorder with a variety of causes.Reperfusion is a common occurrence during treatment of acute intestinal ischemia but the injury resulting from ischemia/reperfusion(IR)may lead toeven more serious complications from intestinal atrophy to multiple organ failure and death.The susceptibility of the intestine to IR-induced injury(IRI)appears from various experimental studies and clinical settings such as cardiac and major vascular surgery and organ transplantation.Where as oxygen free radicals,activation of leukocytes,failure of microvascular perfusion,cellular acidosis and disturbance of intracellular homeo-stasis have been implicated as important factors inthe pathogenesis of intestinal IRI,the mechanisms underlying this disorder are not well known.To date,increasing attention is being paid in animal studies to potential pre-and post-ischemia treatments that protect against intestinal IRI such as drug interference with IR-induced apoptosis and inflammation processes and ischemic pre-conditioning.However,better insight is needed into the molecular and cellular events associated with reperfusion-induced damage to develop effective clinical protection protocols to combat this disorder.In this respect,the use of ischemic post-conditioning in combination with experimentally prolonged acidosis blocking deleterious reperfusion actions may turn out to have particular clinical relevance.

Recombinant angiopoietin-like protein 4 attenuates intestinal barrier structure and function injury after ischemia/reperfusion

BACKGROUND Intestinal barrier breakdown,a frequent complication of intestinal ischemiareperfusion(I/R)including dysfunction and the structure changes of the intestine,is characterized by a loss of tight junction and enhanced permeability of the intestinal barrier and increased mortality.To develop effective and novel therapeutics is important for the improvement of outcome of patients with intestinal barrier deterioration.Recombinant human angiopoietin-like protein 4(rhANGPTL4)is reported to protect the blood-brain barrier when administered exogenously,and endogenous ANGPTL4 deficiency deteriorates radiationinduced intestinal injury.AIM To identify whether rhANGPTL4 may protect intestinal barrier breakdown induced by I/R.METHODS Intestinal I/R injury was elicited through clamping the superior mesenteric artery for 60 min followed by 240 min reperfusion.Intestinal epithelial(Caco-2)cells and human umbilical vein endothelial cells were challenged by hypoxia/reoxygenation to mimic I/R in vitro.RESULTS Indicators including fluorescein isothiocyanate-conjugated dextran(4 kilodaltons;FD-4)clearance,ratio of phosphorylated myosin light chain/total myosin light chain,myosin light chain kinase and loss of zonula occludens-1,claudin-2 and VE-cadherin were significantly increased after intestinal I/R or cell hypoxia/reoxygenation.rhANGPTL4 treatment significantly reversed these indicators,which were associated with inhibiting the inflammatory and oxidative cascade,excessive activation of cellular autophagy and apoptosis and improvement of survival rate.Similar results were observed in vitro when cells were challenged by hypoxia/reoxygenation,whereas rhANGPTL4 reversed the indicators close to normal level in Caco-2 cells and human umbilical vein endothelial cells significantly.CONCLUSION rhANGPTL4 can function as a protective agent against intestinal injury induced by intestinal I/R and improve survival via maintenance of intestinal barrier structure and functions.

Protective Effects of Epigallocatechin-3-gallate on Intestinal Ischemia Reperfusion Injury through Enhanced Activation of PI3K/Akt Pathway in Rats

Inflammation plays a critical role in intestinal ischemia reperfusion injury（IRI）. Epigallocatechin-3-gallate（EGCG） has been demonstrated to possess anti-inflammatory effect. This study examined the effect of EGCG on intestinal IRI and explored the possible mechanisms. Male Wistar rats were randomly divided into three groups： sham-operated group（Sham）, IRI control group（IRI） and IRI-EGCG group（EGCG）. Rats in IRI-EGCG group were administered dissolved EGCG in drinking water（0.4 mg/m L） for 14 days prior to IRI induction. A rat model of intestinal IRI was established by ligating the superior mesenteric artery（SMA） for 30 min, followed by reperfusion for 1 h. Intestinal histology, pro-inflammatory cytokines and mediators were examined and the effect of EGCG on PI3K/Akt signalling was assessed. EGCG significantly alleviated the pathological changes of the intestine and suppressed the IRI-induced up-regulation of TNF-α, IL-1 and IL-6 m RNA and protein expression in the serum and intestine. The mechanism might be that EGCG enhanced the activation of PI3K/Akt signalling pathway. In conclusion, the administration of EGCG can significantly mitigate the acute intestinal IRI in rats by enhancing the activation of PI3K/Akt signalling pathway to suppress inflammatory response and might be a promising alternative for the prevention or treatment of intestinal IRI in the clinical practice.

PTEN-induced kinase 1-induced dynamin-related protein 1 Ser637 phosphorylation reduces mitochondrial fission and protects against intestinal ischemia reperfusion injury

BACKGROUND Intestinal ischemia reperfusion(I/R)occurs in various diseases,such as trauma and intestinal transplantation.Excessive reactive oxygen species(ROS)accumulation and subsequent apoptotic cell death in intestinal epithelia are important causes of I/R injury.PTEN-induced putative kinase 1(PINK1)and phosphorylation of dynamin-related protein 1(DRP1)are critical regulators of ROS and apoptosis.However,the correlation of PINK1 and DRP1 and their function in intestinal I/R injury have not been investigated.Thus,examining the PINK1/DRP1 pathway may help to identify a protective strategy and improve the patient prognosis.AIM To clarify the mechanism of the PINK1/DRP1 pathway in intestinal I/R injury.METHODS Male C57BL/6 mice were used to generate an intestinal I/R model via superior mesenteric artery occlusion followed by reperfusion.Chiu’s score was used to evaluate intestinal mucosa damage.The mitochondrial fission inhibitor mdivi-1 was administered by intraperitoneal injection.Caco-2 cells were incubated in vitro in hypoxia/reoxygenation conditions.Small interfering RNAs and overexpression plasmids were transfected to regulate PINK1 expression.The protein expression levels of PINK1,DRP1,p-DRP1 and cleaved caspase 3 were measured by Western blotting.Cell viability was evaluated using a Cell Counting Kit-8 assay and cell apoptosis was analyzed by TUNEL staining.Mitochondrial fission and ROS were tested by MitoTracker and MitoSOX respectively.RESULTS Intestinal I/R and Caco-2 cell hypoxia/reoxygenation decreased the expression of PINK1 and p-DRP1 Ser637.Pretreatment with mdivi-1 inhibited mitochondrial fission,ROS generation,and apoptosis and ameliorated cell injury in intestinal I/R.Upon PINK1 knockdown or overexpression in vitro,we found that p-DRP1 Ser637 expression and DRP1 recruitment to the mitochondria were associated with PINK1.Furthermore,we verified the physical combination of PINK1 and p-DRP1 Ser637.CONCLUSION PINK1 is correlated with mitochondrial fission and apoptosis by regulating DRP1 phosphorylation in intestinal I/R.These results suggest that the PINK1/DRP1 pathway is involved in intestinal I/R injury,and provide a new approach for prevention and treatment.