Involvement of α2-Adrenoceptors in the Gastric Protective Effect of Nitroglycerin Against Acidified Ethanol-Induced Mucosal Injury

Clinical observations reveal that alcohol intake is associated with an increase in upper gastrointestinal hemorrhage requiring hospitalization and that nitroglycerin or long-acting nitrates lower this risk. Nitroglycerin, a gastric vasodilator that can increase gastric fluid volume, protects the rodent stomach against damage, including that caused by 70% ethanol. Blockade of α₂-adrenoceoptors attenuates gastric protection by intragastric nicotine against 40% ethanol. We tested the hypothesis that the protective effect of nitroglycerin is mediated by an increase in gastric fluid volume and α₂-adrenoceoptors. Nitroglycerin, 5 mg/kg, vehicle, or acidified ethanol was administered intragastrically. In study 1 acidified ethanol-induced mucosal injury was measured. In study 2 the effect of increasing gastric volume (1 ml/kg) on mucosal injury was assessed. In study 3 the effect of yohimbine (α₂-adrenoceoptor antagonist), 5 mg/kg subcutaneously, on the nitroglycerein-mediated protective effect was determined. Results showed that nitroglycerin significantly attenuated the number and length of mucosal lesions induced by acidified ethanol. Increase in gastric fluid volume by exogenously administered saline did not alter the protective effect. Yohimbine blocked the nitroglycerin-mediated protection. These experimental data are consistent with the observation that nitrates lower the risk of ethanol-induced gastrointestinal complications. α₂-Adrenoceoptors are responsible in part for the protective effect of nitroglycerin.     

Gastric mucosal protection with β-phenylethylamine (PEA) and other sympathomimetic amines against absolute ethanol in the rat

The purpose of this study was to investigate the potential of -phenylethylamine (PEA), an amphetamine-like compound present in the blood during high stress situations, to protect rat gastric mucosa against absolute ethanol. F-344 rats were pretreated with PEA in saline at several dose levels and at various times prior to oral administration of 1 ml absolute ethanol. PEA at dose levels of 50 and 100 mg/kg significantly reduced the severity of alcohol-induced lesions following oral, but not parenteral, treatment. The duration of protection with PEA was approximately 90 min, with maximum protection observed when PEA was administered 15–30 min before alcohol. Pretreatment with indomethacin did not prevent or reduce the protection induced by PEA. Other sympathomimetic amines such as isoproterenol and ephedrin were similarly cytoprotective against absolute ethanol while amphetamine, phenylephrine, and epinephrine proved ineffective. These results add further support to the role of the sympathetic nervous system in regulating gastric mucosal protection in the rat.     

β-catenin is essential for ethanol metabolism and protection against alcohol-mediated liver steatosis in mice

The liver plays a central role in ethanol metabolism, and oxidative stress is implicated in alcohol-mediated liver injury. β-Catenin regulates hepatic metabolic zonation and adaptive response to oxidative stress. We hypothesized that β-catenin regulates the hepatic response to ethanol ingestion. Female liver-specific β-catenin knockout (KO) mice and wild-type (WT) littermates were fed the Lieber-Decarli liquid diet (5% ethanol) in a pairwise fashion. Liver histology, biochemistry, and gene-expression studies were performed. Plasma alcohol and ammonia levels were measured using standard assays. Ethanol-fed (EtOH) KO mice exhibited systemic toxicity and early mortality. KO mice exhibited severe macrovesicular steatosis and 5 to 6-fold higher serum alanine aminotransferase and aspartate aminotransferase levels. KO mice had a modest increase in hepatic oxidative stress, lower expression of mitochondrial superoxide dismutase (SOD2), and lower citrate synthase activity, the first step in the tricarboxylic acid cycle. N-Acetylcysteine did not prevent ethanol-induced mortality in KO mice. In WT livers, β-catenin was found to coprecipitate with forkhead box O3, the upstream regulator of SOD2. Hepatic alcohol dehydrogenase and aldehyde dehydrogenase activities and expression were lower in KO mice. Hepatic cytochrome P450 2E1 protein levels were up-regulated in EtOH WT mice, but were nearly undetectable in KO mice. These changes in ethanol-metabolizing enzymes were associated with 30-fold higher blood alcohol levels in KO mice. CONCLUSION: β-Catenin is essential for hepatic ethanol metabolism and plays a protective role in alcohol-mediated liver steatosis. Our results strongly suggest that integration of these functions by β-catenin is critical for adaptation to ethanol ingestion in vivo.     

Involvement of α5 integrin in survivin-mediated osteosarcoma metastasis

Objective: To investigate the role of survivin in osteosarcoma metastasis. Methods: Small interfering RNA(si RNA) was used to knockdown the expression of survivin and α5 integrin in the human osteosarcoma cell line MG63. Western blotting and immunostaining methods was used to assessed the effect of survivin knockdown on the expression of α5 integrin through flow cytometry and fluorescence microscopy detection. Meanwhile, the invasion and migration of transfected cells in Transwell and wound healing assays were probed, and the growth situation of these cells transplanted into nude mice was monitored. Results: Knockdown of survivin expression could inhibit the invasion and migration of osteosarcoma MG64 cells in vitro and the expression of α5 integrin on osteosarcoma MG64 cell surface, suggesting that survivin can inhibit the invasion and migration of osteosarcoma cells through downregulation of α5 integrin. Anti-α5 integrin antibody could also markedly decrease the capability of invasion and migration of osteosarcoma MG64 cells. Additionally, knockdown of survivin expression could slow the growth of osteosarcoma MG63 cells transplanted into nude mice. Conclusions: Survivin-directed anti-tumor strategies might be an effective method in the treatment of osteosarcoma.     

The role of nicotinic acetylcholine receptor (nAChR) α7 subtype in the functional interaction between nicotine and ethanol in mouse cerebellum

Background: Many epidemiological studies report that alcoholics overwhelmingly smoke tobacco and vice versa, which suggests a possible functional interaction between ethanol and nicotine. Although nicotine–ethanol interaction is well documented within the central nervous system, the mechanism is not well understood. Therefore, it is important from a public health standpoint to understand the mechanisms involved in nicotine and ethanol functional interaction. The intracerebellar (ICB) administration of nicotine significantly attenuates ethanol ataxia through nicotinic acetylcholine receptor (nAChR) α₄β₂ subtype. This study, an extension of earlier work, was intended to investigate the possible role of nAChR subtype α₇ in mitigating ethanol ataxia. Methods: The effect of ICB injection of PNU‐282987 (α₇ agonist; 25 ng to 2.5 μg) and the antagonist methyllycaconitine was evaluated on ethanol (2 g/kg; i.p.)‐induced ataxia with a Rotorod. Cerebellar nitric oxide was determined fluorometrically in the presence of ethanol and/or PNU‐282987. Results: Attenuation of ethanol‐induced ataxia following PNU‐282987 microinfusion was dose‐dependent suggesting the participation of α₇ subtype in nicotine and ethanol interaction. Intracerebellar pretreatment with methyllycaconitine (α₇‐selective antagonist; 6 ng) virtually abolished the attenuating effect of PNU‐282987 as well as the effect of nicotine, but not of RJR‐2403 (α₄β₂‐selective agonist; 125 ng) on ethanol‐induced ataxia. Finally, ethanol administration significantly decreased cerebellar NO_x, whereas ICB PNU‐282987 significantly increased and/or opposed ethanol‐induced decrease in NO_x. These results were functionally in agreement with our Rotorod data. Conclusions: These observations confirmed the following: (i) α₇ participation in nicotine–ethanol interaction and (ii) α₇ selectivity of methyllycaconitine. Overall, the results demonstrate the role of cerebellar nAChR α₇ subtype in nicotine‐induced attenuation of ethanol‐induced ataxia in cerebellar NO_x‐sensitive manner.     

Increased glutathione conjugate transport: a possible compensatory protection mechanism against oxidative stress in obesity?

OBJECTIVE: To compare glutathione S-conjugate transport in obese and nonobese persons, and how glutathione S-conjugates are involved in the antioxidant status in obesity. MATERIALS AND METHODS: The efflux of glutathione conjugates and malondialdehyde (MDA) levels were measured in erythrocytes of obese (N = 33) and nonobese (N = 28) persons at every 30 min during a 120 min incubation time in vitro. 2,4-dinitrophenyl-S-glutathione (DNP-SG) represented the glutathione S-conjugate. RESULTS: The efflux of conjugate in erythrocytes from obese subjects (708 +/- 147 DNP-SG efflux nmol/ml erythrocytes/h) was significantly higher than that of control group (490 +/- 105 DNP-SG efflux nmol/ml erythrocytes/h) (P < 0.05). At all time points measured (30-120 min), there was an increase in DNP-SG efflux in obese group (P < 0.05). This is manifested by a decrease in cellular DNP-SG levels. The susceptibility of erythrocytes to in vitro 1-chloro-2,4-dinitrobenzene (CDNB)-induced oxidative stress were greater for cells of control group (P < 0.05), although hemolysis sensitivity of these cells are not different between both groups (P > 0.05). Following CDNB pretreatment, incubation of erythrocyte with vanadate, a DNP-SG transport inhibitor, resulted in an increase of MDA in both groups. However, in this case, the difference in susceptibility was not related to obesity. On the other hand, while erythrocyte glutathione level was lower in obese subjects (79% of control) than in controls (P < 0.05), the adenosine 5'-triphosphate (ATP) levels, the enzyme activities of glutathione S-transferase (GST) and the conjugation capacities of the erythrocytes were not different between groups (P>0.05). CONCLUSION: Obesity may increase erythrocyte glutathione conjugate transport independent from ATP and GST activity that may protect against MDA formation in vitro.     

Effects of L-arginine during ischemia-reperfusion injury in rat orthotopic liver transplantation

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Do chilling injury and heat stress share the same mechanism of injury in oocytes?

Chilling injury is the major limiting factor for successful cryopreservation of both human and animal oocytes. Maternal hyperthermia is the main reason for summer infertility in dairy cows. In this paper, we will present evidence for the involvement of membrane lipid composition and its thermotrophic behavior and the mechanism by which chilling injury and/or heat stress disrupt oocytes' developmental competence. Although oocyte and single zygote are similar in shape and size, the oocyte is known to be very sensitive to cryopreservation, whereas, the zygote is successfully cryopreserved. Recently, we have shown that the lipid-phase transition (LPT) in human MII oocytes occurs at about 20 degrees C, while the LPT temperature of zygotes is lower by 10 degrees C. Similarly, the LPT in oocytes collected from dairy cows was found to be elevated by 10 degrees C in the summer vs. the winter. This feature was associated with alterations in membrane lipid composition. In particular, during the winter, the oocyte membrane is composed mainly of mono- and polyunsaturated fatty acids while in the hot summer, it is composed of more saturated fatty acids. In another study, we showed that exposure of bovine oocytes to physiologically relevant heat shock increases the proportion of oocytes that undergo apoptosis, presumably via sphingomyelin hydrolysis and ceramide formation (i.e. the sphingomyelin apoptosis pathway). Using a mouse model, we have recently shown that hyperthermia of 1.5 degrees C affects the follicle enclosed oocyte as determined by lower developmental competence. Given the importance of the membrane's composition and integrity, it appears that alterations in the oocyte-membrane underlie the disruption of developmental competence in mammalian oocytes exposed to thermal stress (i.e. chilling injury or heat shock).     

Antioxidative properties of curcumin in the protection of blood platelets against oxidative stress in vitro

The present in?vitro study was designed to estimate the antioxidative activity of curcumin in the protection of human blood platelets and plasma against peroxynitrite (ONOO(-))-induced oxidative stress. The effects of curcumin (12.5-50?μg/ml) on ONOO(-)-induced damage of proteins and lipids were determined by the estimation of protein carbonyl groups, 3-nitrotyrosine formation, and thiobarbituric acid reactive substance (TBARS) generation. Exposure of blood platelets and plasma to 100?μM ONOO(-) resulted in an increased level of carbonyl groups, nitration of protein tyrosine residues, and enhanced lipid peroxidation. Curcumin inhibited carbonyl group formation in plasma and in platelet proteins. The highest dose of curcumin (50?μg/ml) reduced blood platelet protein carbonylation by approximately 40%. In the protection of blood plasma protein, the lower doses of curcumin (12.5 and 25?μg/ml) were more effective. Curcumin partially prevented 3-nitrotyrosine formation in plasma proteins; the effect of curcumin was only statistically significant in blood platelets at the highest dose (50?μg/ml). The antioxidative action of curcumin in the protection against lipid peroxidation caused by ONOO(-) was also observed. Curcumin suppressed the formation of TBARS both in blood platelets and in plasma samples. The highest concentration of curcumin (50?μg/ml) decreased the TBARS level by approximately 35% in both blood platelets and plasma samples. In conclusion, the present study demonstrates the antioxidative properties of curcumin and its protective effects against oxidative/nitrative changes of blood platelets and plasma components, especially proteins and lipids.     

Exogenous nitric oxide directly inhibits antral circular muscle motility of rat stomach in vitro

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Is prenatal myo-inositol deficiency a mechanism of CNS injury in galactosemia?

Classic Galactosemia due to galactose-1-phosphate uridyltransferase (GALT) deficiency is associated with apparent diet-independent complications including cognitive impairment, learning problems and speech defects. As both galactose-1-phosphate and galactitol may be elevated in cord blood erythrocytes and amniotic fluid despite a maternal lactose-free diet, endogenous production of galactose may be responsible for the elevated fetal galactose metabolites, as well as postnatal CNS complications. A prenatal deficiency of myo-inositol due to an accumulation of both galactose-1- phosphate and galactitol may play a role in the production of the postnatal CNS dysfunction. Two independent mechanisms may result in fetal myo-inositol deficiency: competitive inhibition of the inositol monophosphatase1 (IMPA1)-mediated hydrolysis of inositol monophosphate by high galactose-1- phosphate levels leading to a sequestration of cellular myo-inositol as inositol monophosphate and galactitol-induced reduction in SMIT1-mediated myo-inositol transport. The subsequent reduction of myo-inositol within fetal brain cells could lead to inositide deficiencies with resultant perturbations in calcium and protein kinase C signaling, the AKT/mTOR/ cell growth and development pathway, cell migration, insulin sensitivity, vescular trafficking, endocytosis and exocytosis, actin cytoskeletal remodeling, nuclear metabolism, mRNA export and nuclear pore complex regulation, phosphatidylinositol-anchored proteins, protein phosphorylation and/or endogenous iron "chelation". Using a knockout animal model we have shown that a marked deficiency of myo-inositol in utero is lethal but the phenotype can be rescued by supplementing the drinking water of the pregnant mouse. If myo-inositol deficiency is found to exist in the GALT-deficient fetal brain, then the use of myo-inositol to treat the fetus via oral supplementation of the pregnant female may warrant consideration.     

Remote ischemic preconditioning confers late protection against myocardial ischemia–reperfusion injury in mice by upregulating interleukin-10

Remote ischemic preconditioning (RIPC) induces a prolonged late phase of multi-organ protection against ischemia–reperfusion (IR) injury. In the present study, we tested the hypothesis that RIPC confers late protection against myocardial IR injury by upregulating expression of interleukin (IL)-10. Mice were exposed to lower limb RIPC or sham ischemia. After 24?h, mice with RIPC demonstrated decreased myocardial infarct size and improved cardiac contractility following 30-min ischemia and 120-min reperfusion (I-30/R-120). These effects of RIPC were completely blocked by anti-IL-10 receptor antibodies. In IL-10 knockout mice, RIPC cardioprotection was lost, but it was mimicked by exogenous IL-10. Administration of IL-10 to isolated perfused hearts increased phosphorylation of the protein kinase Akt and limited infarct size after I-30/R-120. In wild-type mice, RIPC increased plasma and cardiac IL-10 protein levels and caused activation of Akt and endothelial nitric oxide synthase in the heart at 24?h, which was also blocked by anti-IL-10 receptor antibodies. In the gastrocnemius muscle, RIPC resulted in immediate inactivation of the phosphatase PTEN and activation of Stat3, with increased IL-10 expression?24?h later. Myocyte-specific PTEN inactivation led to increased Stat3 phosphorylation and IL-10 protein expression in the gastrocnemius muscle. Taken together, these results suggest that RIPC induces late protection against myocardial IR injury by increasing expression of IL-10 in the remote muscle, followed by release of IL-10 into the circulation, and activation of protective signaling pathways in the heart. This study provides a scientific basis for the use of RIPC to confer systemic protection against IR injury.     

Trimetazidine protects isolated rat hearts against ischemia–reperfusion injury in an experimental timing – dependent manner

The present study investigated the tolerance of the isolated rat heart to ischemia–reperfusion after administration of trimetazidine (TMZ) at different experimental phases, as well as the possible involvement of p38 MAPK and JNKs in this response. Isolated rat hearts were perfused in Langendorff mode. Untreated hearts after stabilization (S) were subjected to 20 min of zero-flow global ischemia (I) and 45 min of reperfusion (R), (NORM), n = 9. TMZ (10^–5 M) was administered (in the perfusate): a) only at S phase, (TMZ–STAB), n = 8, b) only at R, (TMZ–REP), n = 8 and c) during both S and R, (TMZ–STAB+REP), n = 8. Recovery of left ventricular developed pressure at 45 min of R (Rec) was significantly higher in TMZ–STAB and TMZ–STAB+REP and LDH release was lower in TMZ–STAB+REP and TMZ–STAB than NORM, [1153.2 (121.0) and 1152.1 (86.8) vs 1573.5 (138.2), P < 0.05]. TMZ induced cardioprotection did not involve p38 MAPK and JNKs. Phospho–p38 MAPK and JNKs levels after I/R were not changed with TMZ treatment. In TMZ–REP, Rec and LDH release were similar to NORM, but the rate of functional recovery (ratio of Rec at 10 min of R to Rec) was 86.7% (13.3) for TMZ–REP vs 53.8% (7.7) for NORM, P < 0.05. This effect was associated with decreased myocardial lactate content early at reperfusion. In conclusion, preischemic administration of TMZ protects against I/R injury while TMZ given only at reperfusion accelerates recovery of function without reducing the extent of injury.     

Intravenous administration of glutathione protects parenchymal and non-parenchymal liver cells against reperfusion injury following rat liver transplantation

AIM:To investigate the effects of intravenous administrationof the antioxidant glutathione (GSH) on reperfusion injuryfollowing liver transplantation.METHODS:Livers of male Lewis rats were transplantedafter 24 h of hypothermic preservation in University ofWisconsin solution in a syngeneic setting.During a 2-hreperfusion period either saline (controls,n=8) or GSH(50 or 100 μmol/(h·kg),n=5 each) was continuouslyadministered via the jugular vein.RESULTS:Two hours after starting reperfusion plasmaALT increased to 1 457±281 U/L (mean±SE) in controlsbut to only 908±187 U/L (P<0.05) in animals treated with100 μmol GSH/(h·kg).No protection was conveyed by50μmol GSH/(h·kg).Cytoprotection was confirmed bymorphological findings on electron microscopy:GSHtreatment prevented detachment of sinusoidal endothelialcells (SECs) as well as loss of microvilli and mitochondrialswelling of hepatocytes.Accordingly,postischemic bile flowincreased 2-fold.Intravital fluorescence microscopy revealeda nearly complete restoration of sinusoidal blood flow and asignificant reduction of leukocyte adherence to sinusoidsand postsinusoidal venules.Following infusion of 50μmol and100 μmol GSH/(h·kg),plasma GSH increased to 65±7 mol/Land 97±18 mol/L,but to only 20±3 mol/L in untreated recipients.Furthermore,plasma glutathione disulfide (GSSG) increasedto 7.5±1.0 mol/L in animals treated with 100μmol/(h·kg) GSHbut infusion of 50μmol GSH/(h·kg) did not raise levels ofuntreated controls (1.8±0.5 mol/L vs 2.2±0.2 mol/L).CONCLUSION:Plasma GSH levels above a critical level mayact as a “sink” for ROS produced in the hepatic vasculature during reperfusion of liver grafts.Therefore,GSH can beconsidered a candidate antioxidant for the Drevention ofreperfusion injury after liver transplantation,in particularsince it has a low toxicity in humans.     

Ligustrazine alleviates acute lung injury in a rat model of acute necrotizing pancreatitis

Introduction Acute pancreatitis is often complicated by lung injury. However, its pathogenesis remains unclear. It is known that AP involves a complex array of mediators that can initiate and amplify the systemic inflammatory response. This can lead to the failure of distant organ systems, such as the lungs, heart and kidneys,[1-3] among which, pulmonary failure is the most common.[4-7] Recent studies have indicated that during the pathogenesis of acute necrotizing pancreatitis (ANP), chang…     

Ligustrazine alleviates gastric mucosal injury in a rat model of acute necrotizing pancreatitis

BACKGROUND: Acute necrotizing pancreatitis (ANP) leads to a systemic inflammatory response characterized by widespread leukocyte activation and, as a consequence, distant organ injury. The aim of this study was to explore the relationship between gastric microcirculatory impairment and inflammatory mediators released in rats and to evaluate the therapeutic effect of ligustrazine extracted from Rhizoma ligusticum wallichii on gastric mucosa injury in a rat model of ANP. METHODS: Ninety-six Sprague-Dawley rats were randomly divided into three groups: normal control (group C); ANP without treatment (group P); and ANP treated with ligustrazine (group T). The ANP model was induced by injection of 50 g/L sodium taurocholate under the pancreatic membrane (4 ml/kg). Group C was given isovolumetric injection of 9 g/L physiological saline by the same route. Group T was injected with ligustrazine (10 ml/kg) via the portal vein. The radioactive biomicrosphere technique was used to measure the blood flow 2 and 12 hours after the induction of ANP. Samples of the pancreas and stomach were taken to assess pathological changes by a validated histology score; meanwhile, the levels of serum interleukin-1β (IL-1β) were determined. Gastric tissues were also used to measure the level of myeloperoxidase (MPO), which is expressed intracellularly in the azurophilic granules of neutrophils. RESULTS: Blood flow in group P was significantly lower than that in group C (P<0.01). Pathological changes were significantly aggravated in group P. The gastric MPO activity in group P was significantly higher than that in group C (P<0.01). The level of serum IL-1β in group P increased more significantly than that in group C (P<0.01). Blood flow of the stomach in group T was significantlyhigher than that in group P after 2 hours (P<0.01). The pathological changes were significantly alleviated in group T. The MPO activity of group T was significantly lower than that of group P (P<0.01). Although serum IL-1β level of group T, was higher than of group C (P<0.01), it was lower than that of group P (P<0.01). There was a negative correlation between gastric blood flow and MPO activity (r=-0.983, P<0.01), and between gastric blood flow and pathological score (r=-0.917, P<0.05). CONCLUSIONS: Decreased gastric blood flow and increased inflammatory mediators can be seen early in ANP, and both are important factors for gastric and mucosal injury. Ligustrazine can ameliorate microcirculatory disorder and alleviate the damage to the pancreas and stomach.