四氯化碳诱导肝损伤大鼠自由基及微量元素含量的动态变化(英文)

为探讨微量元素与自由基代谢的关系,本实验采用四氯化碳诱导肝损伤的大鼠模型,研究了中毒大鼠体内自由基和微量元素含量的动态变化。结果大鼠经四氯化碳致毒后,肝组织自由基浓度增加,肝脏中Cu,Zn-SOD含量增加,存在着自由基底物对超氧化物歧化酶的诱导合成作用。肝脏出现明显的脂质过氧化增强现象,MDA浓度增加,GSH降低和GSSG升高。实验还观察到脑组织亦发生轻度脂质过氧化增强。在上述变化的同时,与自由基代谢有关的微量元素Zn、Cu、Fe、Mn和Se也发生了明显的改变,其中肝脏中Zn、Cu和Se含量显著降低,而肝Fe却明显增高,在脑组织中表现为Zn的降低和Se升高。上述结果说明微量元素改变在肝损伤时的自由基代谢中具有一定的作用。     

Baicalin prevents LPS-induced activation of TLR4/NF-κB p65 pathway and inflammation in mice via inhibiting the expression of CD14

Previous studies have shown that baicalin,an active ingredient of the Chinese traditional medicine Huangqin,attenuates LPS-induced inflammation by inhibiting the activation of TLR4/NF-κBp65 pathway,but how it affects this pathway is unknown.It has been shown that CD14 binds directly to LPS and plays an important role in sensitizing the cells to minute quantities of LPS via chaperoning LPS molecules to the TLR4/MD-2 signaling complex.In the present study we investigated the role of CD14 in the anti-inflammatory effects of baicalin in vitro and in vivo.Exposure to LPS(1μg/mL)induced inflammatory responses in RAW264.7 cells,evidenced by marked increases in the expression of MHC II molecules and the secretion of NO and IL-6,and by activation of MyD88/NF-κB p65 signaling pathway,as well as the expression of CD14 and TLR4.These changes were dose-dependently attenuated by pretreatment baicalin(12.5–50μM),but not by baicalin post-treatment.In RAW264.7 cells without LPS stimulation,baicalin dose-dependently inhibit the protein and mRNA expression of CD14,but not TLR4.In RAW264.7 cells with CD14 knockdown,baicalin pretreatment did not prevent inflammatory responses and activation of MyD88/NF-κB p65 pathway induced by high concentrations(1000μg/mL)of LPS.Furthermore,baicalin pretreatment also inhibited the expression of CD14 and activation of MyD88/NF-κB p65 pathway in LPS-induced hepatocyte-derived HepG2 cells and intestinal epithelial-derived HT-29 cells.In mice with intraperitoneal injection of LPS and in DSS-induced UC mice,oral administration of baicalin exerted protective effects by inhibition of CD14 expression and inflammation.Taken together,we demonstrate that baicalin pretreatment prevents LPS-induced inflammation in RAW264.7 cells in CD14-dependent manner.This study supports the therapeutic use of baicalin in preventing the progression of LPS-induced inflammatory diseases.     

Tunisian radish extract (Raphanus sativus) enhances the antioxidant status and protects against oxidative stress induced by zearalenone in Balb/c mice

Radish (Raphanus sativus) is a food plant known worldwide. From antiquity it has been used in folk medicine as a natural drug against many toxicants. Zearalenone (zen) is a non-steroidal estrogenic mycotoxin present in corn and food mixture for farm animals and it is hepatotoxic, hematotoxic, immunotoxic, nephrotoxic and genotoxic. The objectives of the present study were to assess the biological activity of radish extract and to evaluate the protective role of radish extract against the toxicity of zen in female Balb/c mice. Animals were divided into seven groups and treated orally for 10 days as follows: a control, an olive oil group, groups treated with radish extract alone (5, 10 and 15 mg kg(-1) b.w.), a group treated with zen (40 mg kg(-1) b.w.) and a group treated with zen plus the lowest dose of radish extract. The results indicate that radish extract improved the antioxidant status and had no significant effects on hematological and biochemical parameters tested or histology of the liver and kidney. Treatment with zen results in a significant increase in ALT, AST, ALP, BILT, BILD, CRE accompanied with significant changes in most of hematological parameters and the antioxidant enzyme activities, co-treatment of zen and the radish extract results in a significant reestablishment of hematological, serum biochemical parameters, and the histology of the liver and kidney. These findings suggest that radish extract is safe and can be overcome or, at least, significantly diminish zen effects.     

Potentiating effect of acetaminophen and carbon tetrachloride-induced hepatotoxicity is mediated by activation of receptor interaction protein in mice

When multiple drugs or chemicals are used in combination, it is important to understand the risk of their interactions and predict potential additive effects. The aim of the current study was to investigate the molecular mechanism(s) accounting for the additive/synergistic effect of combination treatment with acetaminophen (APAP) and carbon tetrachloride (CCl₄). Mice were intraperitoneally administered vehicle or 100?mg/kg (5?mL/kg) APAP and 30?min after vehicle or 15?mg/kg (5?mL/kg) CCl₄. Sixteen hours after treatment, mice from each group were sacrificed and the livers were removed. CCl₄ administration caused slight glycogen depletion; this effect was more pronounced following co-administration of APAP and CCl₄. ATP and NADPH levels showed the same trend as glycogen levels. The levels of receptor interacting protein 1 and 3 increased following combination treatment with APAP and CCl₄. In contrast, levels of the glutamate cysteine ligase catalytic subunit and glutamate cysteine ligase modifier subunits were not significantly affected by combination treatment. APAP and CCl₄ co-administration potentiated the phosphorylation of c-Jun N-terminal kinase and p38 kinases, although phosphorylated activation of extracellular signal-regulated kinase was not changed. Our results suggest that APAP and CCl₄ co-administration potentiates hepatotoxicity in an additive/synergistic manner via receptor interacting protein activation.     

The natural polyamines spermidine and spermine prevent bone loss through preferential disruption of osteoclastic activation in ovariectomized mice

BACKGROUND AND PURPOSE

Although naturally occurring polyamines are indispensable for a variety of cellular events in eukaryotic cells, little attention has been paid to their physiological and pathological significance in bone remodelling to date. In this study, we evaluated the pharmacological properties of several natural polyamines on the functionality and integrity of bone in both in vitro and in vivo experiments.

EXPERIMENTAL APPROACH

Mice were subjected to ovariectomy (OVX) and subsequent oral supplementation with either spermidine or spermine for determination of the bone volume together with different parameters regarding bone formation and resorption by histomorphometric analyses in vivo. Pre-osteoclasts were cultured with receptor activator of NF-κB ligand (RANKL), with or without spermidine and spermine to determine cellular maturation by tartrate-resistant acid phosphatase (TRAP) staining and cellular viability by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide reduction in vitro.

KEY RESULTS

Spermidine or spermine, given in drinking water for 28 days, significantly prevented the increased osteoclast surface/bone surface ratio and the reduced bone volume following OVX in mice. Either spermidine or spermine significantly inhibited the increased number of multinucleated TRAP-positive cells in osteoclasts cultured with RANKL in a concentration-dependent manner without affecting cell survival.

CONCLUSIONS AND IMPLICATIONS

The natural polyamines spermidine and spermine prevented OVX-induced bone loss through the disruption of differentiation and maturation of osteoclasts, rather than affecting osteoblasts. The supplementation with these natural polyamines could be beneficial for the prophylaxis as well as therapy of metabolic bone diseases such as post-menopausal osteoporosis.     

黄芩素对四氯化碳诱导小鼠急性肝损伤的保护作用

<正>黄芩素是一种天然类黄酮物质,提取于唇形科植物高黄芩(Scutellaria altissima L.)的全草中,因其抗炎、解热作用[1]显著、毒副作用小越来越多的受到人们的重视。本实验室首次通过灌胃黄芩素并采用四氯化碳诱导急性肝损伤小鼠模型,观察黄芩素对肝脏的影响并讨论了其可能的作用机制。1材料与方法1.1材料及仪器健康昆明小鼠(♂),体质量18～22 g,清洁级,由第四军医大学动物实验中心提供。黄芩素(中国食     

Hepatoprotective effect of ginsenoside Rg1 from Panax ginseng on carbon tetrachloride‐induced acute liver injury by activating Nrf2 signaling pathway in mice

Oxidative stress and inflammatory response are well known to be involved in the pathogenesis of acute liver injury. This study was performed to examine the hepatoprotective effect of ginsenoside Rg1 (Rg1) against CCl₄‐induced acute liver injury, and further to elucidate the involvement of Nrf2 signaling pathway in vivo and in vitro. Mice were orally administered Rg1 (15, 30, and 60 mg/kg) or sulforaphane (SFN) once daily for 1 week prior to 750 μL/kg CCl₄ injection. The results showed that Rg1 markedly altered relative liver weights, promoted liver repair, increased the serum level of TP and decreased the serum levels of ALT, AST and ALP. Hepatic oxidative stress was inhibited by Rg1, as evidenced by the decrease in MDA, and increases in GSH, SOD, and CAT in the liver. Further research demonstrated that Rg1 suppressed liver inflammation response through repressing the expression levels of inflammation‐related genes including TNF‐α, IL‐1β, IL‐6, COX‐2, and iNOS. In addition, Rg1 enhanced antioxidative stress and liver detoxification abilities by up‐regulating Nrf2 and its target‐genes such as GCLC, GCLM, HO‐1, NQO1, Besp, Mrp2, Mrp3, Mrp4, and down‐regulating Cyp2e1. However, the changes in Nrf2 target‐genes, as well as ameliorative liver histology induced by Rg1 were abrogated by Nrf2 antagonist all‐transretinoic acid in vivo and Nrf2 siRNA in vitro. Overall, the findings indicated that Rg1 might be an effective approach for the prevention against acute liver injury by activating Nrf2 signaling pathway.     

A high‐fat diet and multiple administration of carbon tetrachloride induces liver injury and pathological features associated with non‐alcoholic steatohepatitis in mice

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Molecular Mechanisms of Lipoic Acid Protection against Aflatoxin B1-Induced Liver Oxidative Damage and Inflammatory Responses in Broilers

Alpha-lipoic acid (α-LA) was evaluated in this study for its molecular mechanisms against liver oxidative damage and inflammatory responses induced by aflatoxin B₁ (AFB₁). Birds were randomly allocated into four groups with different diets for three weeks: a basal diet, a 300 mg/kg α-LA supplementation in a basal diet, a diet containing 74 μg/kg AFB₁, and 300 mg/kg α-LA supplementation in a diet containing 74 μg/kg AFB₁. In the AFB₁ group, the expression of GSH-P_X mRNA was down-regulated (p < 0.05), and the levels of lipid peroxide and nitric oxide were increased (p < 0.05) in the chicken livers compared to those of the control group. Additionally, the mRNA level of the pro-inflammatory factor interleukin-6 was up-regulated significantly (p < 0.05), the protein expressions of both the nuclear factor kappa B (NF-κB) p65 and the inducible nitric oxide synthase were enhanced significantly (p < 0.05) in the AFB₁ group. All of these negative effects were inhibited by α-LA. These results indicate that α-LA may be effective in preventing hepatic oxidative stress, down-regulating the expression of hepatic pro-inflammatory cytokines, as well as inhibiting NF-κB expression.     

Protective effects of Mangifera indica L. extract, mangiferin and selected antioxidants against TPA-induced biomolecules oxidation and peritoneal macrophage activation in mice

We compared the protective abilities of Mangifera indica L. stem bark extract (Vimang) 50-250 mgkg(-1), mangiferin 50 mgkg(-1), vitamin C 100 mgkg(-1), vitamin E 100 mgkg(-1)and beta -carotene 50 mgkg(-1)against the 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced oxidative damage in serum, liver, brain as well as in the hyper-production of reactive oxygen species (ROS) by peritoneal macrophages. The treatment of mice with Vimang, vitamin E and mangiferin reduced the TPA-induced production of ROS by the peritoneal macrophages by 70, 17 and 44%, respectively. Similarly, the H(2)O(2)levels were reduced by 55-73, 37 and 40%, respectively, when compared to the control group. The TPA-induced sulfhydryl group loss in liver homogenates was attenuated by all the tested antioxidants. Vimang, mangiferin, vitamin C plus E and beta -carotene decreased TPA-induced DNA fragmentation by 46-52, 35, 42 and 17%, respectively, in hepatic tissues, and by 29-34, 22, 41 and 17%, in brain tissues. Similar results were observed in respect to lipid peroxidation in serum, in hepatic mitochondria and microsomes, and in brain homogenate supernatants. Vimang exhibited a dose-dependent inhibition of TPA-induced biomolecule oxidation and of H(2)O(2)production by peritoneal macrophages. Even if Vimang, as well as other antioxidants, provided significant protection against TPA-induced oxidative damage, the former lead to better protection when compared with the other antioxidants at the used doses. Furthermore, the results indicated that Vimang is bioavailable for some vital target organs, including liver and brain tissues, peritoneal exudate cells and serum. Therefore, we conclude that Vimang could be useful to prevent the production of ROS and the oxidative tissue damages in vivo.     

Differential hepatoprotective mechanisms of rutin and quercetin in CCl4-intoxicated BALB/cN mice

Aim:

To investigate the mechanisms underlying the protective effects of quercetin-rutinoside (rutin) and its aglycone quercetin against CCl₄-induced liver damage in mice.

Methods:

BALB/cN mice were intraperitoneally administered rutin (10, 50, and 150 mg/kg) or quercetin (50 mg/kg) once daily for 5 consecutive days, followed by the intraperitoneal injection of CCl₄ in olive oil (2 mL/kg, 10% v/v). The animals were sacrificed 24 h later. Blood was collected for measuring the activities of ALT and AST, and the liver was excised for assessing Cu/Zn superoxide dismutase (SOD) activity, GSH and protein concentrations and also for immunoblotting. Portions of the livers were used for histology and immunohistochemistry.

Results:

Pretreatment with rutin and, to a lesser extent, with quercetin significantly reduced the activity of plasma transaminases and improved the histological signs of acute liver damage in CCl₄-intoxicated mice. Quercetin prevented the decrease in Cu/Zn SOD activity in CCl₄-intoxicated mice more potently than rutin. However, it was less effective in the suppression of nitrotyrosine formation. Quercetin and, to a lesser extent, rutin attenuated the inflammation in the liver by down-regulating the CCl₄-induced activation of nuclear factor-kappa B (NF-κB), tumor necrosis factor-α (TNF-α) and cyclooxygenase (COX-2). The expression of inducible nitric oxide synthase (iNOS) was more potently suppressed by rutin than by quercetin. Treatment with both flavonoids significantly increased NF-E2-related factor 2 (Nrf2) and heme oxygenase (HO-1) expression in injured livers, although quercetin was less effective than rutin at an equivalent dose. Quercetin more potently suppressed the expression of transforming growth factor-β1 (TGF-β1) than rutin.

Conclusion:

Rutin exerts stronger protection against nitrosative stress and hepatocellular damage but has weaker antioxidant and anti-inflammatory activities and antifibrotic potential than quercetin, which may be attributed to the presence of a rutinoside moiety in position 3 of the C ring.     

Hepatoprotective activity of petroleum ether, diethyl ether, and methanol extract of Scoparia dulcis L. against CCl4-induced acute liver injury in mice

Objectives:

The present study was aimed at assessing the hepatoprotective activity of 1:1:1 petroleum ether, diethyl ether, and methanol (PDM) extract of Scoparia dulcis L. against carbon tetrachloride-induced acute liver injury in mice.

Materials and Methods:

The PDM extract (50, 200, and 800 mg/kg, p.o.) and standard, silymarin (100 mg/kg, p.o) were tested for their antihepatotoxic activity against CCl4-induced acute liver injury in mice. The hepatoprotective activity was evaluated by measuring aspartate aminotransferase, alanine aminotransferase, alkaline phosphatase, and total proteins in serum, glycogen, lipid peroxides, superoxide dismutase, and glutathione reductase levels in liver homogenate and by histopathological analysis of the liver tissue. In addition, the extract was also evaluated for its in vitro antioxidant activity using 1, 1-Diphenyl-2-picrylhydrazyl scavenging assay.

Results:

The extract at the dose of 800 mg/kg, p.o., significantly prevented CCl4-induced changes in the serum and liver biochemistry (P < 0.05) and changes in liver histopathology. The above results are comparable to standard, silymarin (100 mg/kg, p.o.). In the in vitro 1, 1-diphenyl-2-picrylhydrazyl scavenging assay, the extract showed good free radical scavenging potential (IC 50 38.9 ± 1.0 μg/ml).

Conclusions:

The results of the study indicate that the PDM extract of Scoparia dulcis L. possesses potential hepatoprotective activity, which may be attributed to its free radical scavenging potential, due to the terpenoid constituents.     

Curcumin attenuates brain edema in mice with intracerebral hemorrhage through inhibition of AQP4 and AQP9 expression

Aim:

Aquaporins (AQPs) are the water-channels that play important roles in brain water homeostasis and in cerebral edema induced by brain injury. In this study we investigated the relationship between AQPs and a neuroprotective agent curcumin that was effective in the treatment of brain edema in mice with intracerebral hemorrhage (ICH).

Methods:

ICH was induced in mice by autologous blood infusion. The mice immediately received curcumin (75, 150, 300 mg/kg, ip). The Rotarod test scores, brain water content and brain expression of AQPs were measured post ICH. Cultured primary mouse astrocytes were used for in vitro experiments. The expression of AQP1, AQP4 and AQP9 and NF-κB p65 were detected using Western blotting or immunochemistry staining.

Results:

Curcumin administration dose-dependently reduced the cerebral edema at d 3 post ICH, and significantly attenuated the neurological deficits at d 5 post ICH. Furthermore, curcumin dose-dependently decreased the gene and protein expression of AQP4 and AQP9, but not AQP1 post ICH. Treatment of the cultured astrocytes with Fe²⁺ (10–100 μmol/L) dose-dependently increased the expression and nuclear translocation of NF-κB p65 and the expression of AQP4 and AQP9, which were partly blocked by co-treatment with curcumin (20 μmol/L) or the NF-κB inhibitor PDTC (10 μmol/L).

Conclusion:

Curcumin effectively attenuates brain edema in mice with ICH through inhibition of the NF-κB pathway and subsequently the expression of AQP4 and AQP9. Curcumin may serve as a potential therapeutic agent for ICH.     

iTRAQ-based proteomic analysis of combination therapy with taurine,epigallocatechin gallate,and genistein on carbon tetrachloride-induced liver fibrosis in rats

Combination therapy with taurine, epigallocatechin gallate, and genistein was effective in alleviating the progression of liver fibrosis in our previous study. To better understand the anti-fibrotic mechanisms of combination therapy, an iTRAQ-based proteomics approach was used to study the expression profiles of proteins in carbon tetrachloride-induced liver fibrosis rats following combination therapy. The anti-fibrotic effects of combination therapy were assessed directly by liver histology, and indirectly by measurement of serum biochemical markers and antioxidant enzymes. The results showed that combination therapy could significantly improve the liver function, as indicated by decreasing levels of alanine aminotransferase (ALT), aspartate transaminase (AST), transforming growth factor-β1 (TGF-β1), and collagen I, increasing levels of total antioxidative capacity (T-AOC), superoxide dismutase (SOD), and glutathione peroxidase (GSH-Px), and reducing the pathological tissue damage. A total of 89 differential expressed proteins in response to combination therapy were identified by iTRAQ, which were interacted with each other and involved in different biological processes and pathways. Four differentially expressed proteins (Tpi1, Txn1, Fgb, and F7) involved in antioxidant defense system, glycolysis pathway and coagulation cascade pathway were validated by enzyme-linked immunosorbent assay. Our work provided valuable insights into the molecular mechanism of combination therapy against liver fibrosis, and the identified targets may be useful for treatment of liver fibrosis in future.     

Role of H(2)O(2) in hypertension, renin-angiotensin system activation and renal medullary disfunction caused by angiotensin II

BACKGROUND AND PURPOSE

Activation of the intrarenal renin-angiotensin system (RAS) and increased renal medullary hydrogen peroxide (H₂O₂) contribute to hypertension. We examined whether H₂O₂ mediated hypertension and intrarenal RAS activation induced by angiotensin II (Ang II).

EXPERIMENTAL APPROACH

Ang II (200 ng·kg⁻¹·min⁻¹) or saline were infused in Sprague Dawley rats from day 0 to day 14. Polyethylene glycol (PEG)-catalase (10 000 U·kg⁻¹·day⁻¹) was given to Ang II-treated rats, from day 7 to day 14. Systolic blood pressure was measured throughout the study. H₂O₂, angiotensin AT₁ receptor and Nox4 expression and nuclear factor-κB (NF-κB) activation were evaluated in the kidney. Plasma and urinary H₂O₂ and angiotensinogen were also measured.

KEY RESULTS

Ang II increased H₂O₂, AT₁ receptor and Nox4 expression and NF-κB activation in the renal medulla, but not in the cortex. Ang II raised plasma and urinary H₂O₂ levels, increased urinary angiotensinogen but reduced plasma angiotensinogen. PEG-catalase had a short-term antihypertensive effect and transiently suppressed urinary angiotensinogen. PEG-catalase decreased renal medullary expression of AT₁ receptors and Nox4 in Ang II-infused rats. Renal medullary NF-κB activation was correlated with local H₂O₂ levels and urinary angiotensinogen excretion. Loss of antihypertensive efficacy was associated with an eightfold increase of plasma angiotensinogen.

CONCLUSIONS AND IMPLICATIONS

The renal medulla is a major target for Ang II-induced redox dysfunction. H₂O₂ appears to be the key mediator enhancing intrarenal RAS activation and decreasing systemic RAS activity. The specific control of renal medullary H₂O₂ levels may provide future grounds for the treatment of hypertension.