Hepatic Caspase-3 Activity Research Articles

Pharmacological Evaluation of Novel 1,2,4-triazine Derivatives Containing Thiazole Ring against Hepatocellular Carcinoma

Background: New 6-hydroxy-5-(p-hydroxybenzylidene)-3-phenyl-2- [(5-pchlorophenyl)- 1,3-thiazol-2-yl]-1, 2, 4-triazine derivatives containing a thiazole ring were synthesised as potential antitumor agents. Methods: Cytotoxicity of compounds (3) and (4) was evaluated in human hepatocellular carcinoma (HCC) cell lines (HepG2); compound (3) showed more cytotoxicity (IC50=9.0μg/ml) than compound (4) (IC50=18.40μg/ml) using doxorubicin as standard. The degree of toxicity of compound (3) was assessed by the LD50 with its anticancer performance by suppressing tumor angiogenesis against diethylnitrosamine (DENA) induced hepatocellular carcinoma (HCC) in male rat model. Results : Carcinogenic rats showed a significant increase in markers of angiogenesis, tumour growth, and liver function tests and malondialdehyde level coupled with reduced hepatic glutathione level and caspase-3 activity. The distribution of compound (3) to animals after the development of HCC improved biochemical alterations from a DENA chemical carcinogen that is confirmed by hepatic histopathology. Conclusion: Compound 3 perhaps utilized as a strong applicant for newly therapeutic protocols against hepatocarcinogenesis by controlling tumor angiogenesis and renovating the activity of hepatic marker enzymes in addition to reversing the oxidant-antioxidant imbalance in corporation with amelioration of histopathology. While the trial supports the use of compound 3 for improved HCC outcome and the toxicity and side effects should be considered.

Ziziphus spina-christi (L.) fortified with Camellia sinensis mediates apoptosis, Notch-1 signaling, and mitigates obesity-induced non-alcoholic fatty liver.

The habit of drinking tea is highly prevalent and could be utilized to introduce more health benefits through fortification with medicinal plants. The purpose of this analysis was to assess the nutritional quality and health benefits of fortified Ziziphus tea (ZT) with green tea (GT) against obesity and non-alcoholic fatty liver disease (NAFLD). Proximate analysis and sensory evaluation were carried out on the fortified tea. In the in vivo study, 15 SD rats were used for each group. Flow cytometry was utilized for caspase 3 analysis. ELISA was used for the detection of tumor necrosis factor-alpha and adiponectin levels. Real-time PCR was used to detect Notch-1 and Hes-1 gene expression. The composition of fortified (GT+ZT) showed a significant improvement in the nutritional value represented by the increase in overall protein, crude fat, crude fiber, ash, carbohydrate, mineral contents, and antioxidant capacity. Treatment with GT+ZT restored the disturbance in body weight, lipid profile, liver function, glucose, insulin sensitivity index, and oxidative status. It reversed the changes in TNF-α and adiponectin levels. Their protective effects against NAFLD were indicated by the inhibition of hepatic caspase-3 activity, suppression of Notch-1, and Hes-1 gene expression and amelioration of high-fat diet (HFD)-induced histological alterations. Collectively, our findings, elucidate the precise mechanism of fortified ZT+GT for the attenuation of obesity-induced metabolic disorders and NAFLD via regulating lipolysis, TNF-α, adiponectin, apoptosis, and Notch-1 signaling pathways, and provide a foundation for an easily implemented healthy habit of drinking. PRACTICAL APPLICATIONS: The incorporation of bioactive compounds into functional foods is a growing market. Consumer attention in well-being has increased rapidly toward a fortified diet that provides additional health effects. The fortified (GT+ZT) tea may potentially serve as an easily implemented healthy drinking habit to prevent and manage obesity and NAFLD and reduce the risk of other diseases. Fortification with ZT improved the health-promoting functionality of GT through the enhancement of total protein, carbohydrates, antioxidant, and mineral contents. This was reflected by their synergetic therapeutic activity in ameliorating the disturbance in obesity-related disorders and NAFLD via regulating lipolysis, inflammation, oxidative stress, apoptosis, and Notch-1 signaling pathways. Therefore, (GT+ZT) could be considered functional foods which attribute to functional improvement and reduction in disease risk.

Exacerbated LPS/GalN-Induced Liver Injury in the Stress-Sensitive Wistar Kyoto Rat Is Associated with Changes in the Endocannabinoid System.

Acute liver injury (ALI) is a highly destructive and potentially life-threatening condition, exacerbated by physical and psychological stress. The endocannabinoid system plays a key role in modulating stress and hepatic function. The aim of this study was to examine the development of acute liver injury in the genetically susceptible stress-sensitive Wistar-Kyoto (WKY) rat compared with normo-stress-sensitive Sprague Dawley (SD) rats, and associated changes in the endocannabinoid system. Administration of the hepatotoxin lipopolysaccharide/D-Galactosamine (LPS/GalN) resulted in marked liver injury in WKY, but not SD rats, with increased alanine aminotransferase (ALT), aspartate aminotransferase (AST) and glutamate dehydrogenase (GLDH) plasma levels, significant histopathological changes, increased hepatic pro-inflammatory cytokine expression and caspase-3 activity and expression and reduced Glutathione (GSH) activity. Furthermore, compared to SD controls, WKY rats display increased anandamide and 2-Arachidonoylglycerol levels concurrent with decreased expression of their metabolic enzymes and a decrease in cannabinoid (CB)1 receptor expression following LPS/GalN. CB1 antagonism with AM6545 or CB2 agonism with JWH133 did not alter LPS/GalN-induced liver injury in SD or WKY rats. These findings demonstrate exacerbation of acute liver injury induced by LPS/GalN in a stress-sensitive rat strain, with effects associated with alterations in the hepatic endocannabinoid system. Further studies are required to determine if the endocannabinoid system mediates or modulates the exacerbation of liver injury in this stress-sensitive rat strain.

Sleep deprivation aggravated lipopolysaccharide/D-galactosamine-induced acute liver injury by suppressing melatonin production.

Sleep loss is common in patients with liver injury, but the effects of sleep deprivation (SD) on liver injury remain unclear. In the present study, the potential effects of SD on acute liver injury and the underlying mechanisms have been investigated. The sleep of male BALB/c mice has been deprived by using a modified multiple platform water bath for 3 days and acute liver injury was induced by intraperitoneal injection of lipopolysaccharide (LPS) and D-galactosamine (D-Gal). The degree of liver injury was detected by aminotransferase determination, histopathology and survival rate analysis. Inflammatory response and melatonin (MT) were measured by enzyme-linked immunosorbent assay (ELISA). In addition, hepatocyte apoptosis was determined by caspase activity measurement and terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay. We observed that SD increased plasma aminotransferases, TUNEL-positive hepatocytes, histological abnormalities and mortality rates in mice with LPS/D-Gal treatment. SD also promoted LPS/D-Gal-induced production of TNF-α and upregulated hepatic caspase-8, caspase-9, and caspase-3 activities in LPS/D-Gal-exposed mice. In addition, SD significantly decreased MT contents in plasma of mice with acute liver injury, but supplementation with MT reversed these SD-promoted changes. Our data suggested that SD exacerbated LPS/D-Gal-induced liver injury via decreasing melatonin production.

Olive oil combined with Lycium barbarum polysaccharides attenuates liver apoptosis and inflammation induced by carbon tetrachloride in rats

Olive oil and Lycium barbarum polysaccharides were considered to be hepatoprotective for liver fibrosis. This study investigated anti-apoptotic, anti-inflammatory and anti-fibrotic effects of olive oil and/or Lycium barbarum polysaccharides (LBP) on CCl4-induced liver fibrosis in rats. The results showed that CCl4 caused liver fatty change, cell death, inflammation and collagen accumulation. The olive oil-treated groups reduced hepatic transforming growth factor (TGF)-β1 and tissue inhibitors of metalloproteinase (TIMP)-1 levels. The LBP-treated groups inhibited hepatic caspase-9 and caspase-3 activities, decreased hepatic tumor necrosis factor (TNF)-α levels, increased hepatic interleukin (IL)-10 levels and IL-10/TNF-α ratios, and reduced hepatic TGF-β1 and TIMP-1 levels. Olive oil combined with LBP suppressed liver apoptotic markers, inhibited liver inflammatory markers, and attenuated hepatic TGF-β1 levels. Therefore, LBP improves liver apoptotic, inflammatory and fibrotic markers, while olive oil combined with LBP has better effects on anti-apoptosis and anti-inflammation compared with olive oil treatment in rats with CCl4-induced liver fibrosis.

Dopamine alleviated acute liver injury induced by lipopolysaccharide/d-galactosamine in mice

Dopamine (DA), a crucial neurotransmitter, not only functions in the central nervous system but also plays important roles in the modulation of inflammation. Several studies suggest that DA might suppress the inflammatory response both in vitro and in vivo. In the present study, the potential effects of DA in a mouse model with lipopolysaccharide (LPS)/d-galactosamine (D-Gal)-induced acute liver injury were investigated. The results show that DA-treated LPS/D-Gal-exposed mice had reduced incidence of histologic lesions, lower plasma aminotransferases and improved the survival rates compared to LPS/D-Gal-exposed mice. Treatment with DA also suppressed LPS/D-Gal-induced production of TNF-α, phosphorylation of c-jun-N-terminal kinase (JNK), cleavage of caspase-3, up-regulation of hepatic caspase-3, caspase-8, and caspase-9 activities and reduced the count of TUNEL-positive hepatocytes. These data indicate that DA attenuated LPS/D-Gal-induced fulminant liver injury in mice, which implies that DA might have value for the prevention of inflammatory liver disease.

Tributyltin promoted hepatic steatosis in zebrafish (Danio rerio) and the molecular pathogenesis involved

Endocrine disruptor effects of tributyltin (TBT) are well established in fish. However, the adverse effects on lipid metabolism are less well understood. Since the liver is the predominant site of de novo synthesis of lipids, the present study uses zebrafish (Danio rerio) to examine lipid accumulation in the livers and hepatic gene expression associated with lipid metabolism pathways. After exposure for 90 days, we found that the livers in fish exposed to TBT were yellowish in appearance and with accumulation of lipid droplet, which is consistent with the specific pathological features of steatosis. Molecular analysis revealed that TBT induced hepatic steatosis by increasing the gene expression associated with lipid transport, lipid storage, lipiogenic enzymes and lipiogenic factors in the livers. Moreover, TBT enhanced hepatic caspase-3 activity and up-regulated genes related to apoptosis and cell-death, which indicated steatotic livers of fish exposed to TBT and the subsequent liver damage were likely due to accelerated hepatocyte apoptosis or cell stress. In short, TBT can produce multiple and complex alterations in transcriptional activity of lipid metabolism and cell damage, which provides potential molecular evidence of TBT on hepatic steatosis.

Rotenone, a mitochondrial respiratory complex I inhibitor, ameliorates lipopolysaccharide/D-galactosamine-induced fulminant hepatitis in mice.

The syntheses of inflammatory mediators are energy-intensive processes and the mitochondria play pivotal roles in supporting these energy-requiring molecular responses. In the present studies, a mitochondrial respiratory complex I inhibitor rotenone was administrated in mice with lipopolysaccharide/D-galactosamine (LPS/D-Gal)-induced fulminant liver injury and the prophylactic and therapeutic effects on tissue injury were evaluated. We found that pretreatment with rotenone suppressed the elevation of plasma aminotransferases, alleviated the histopathological abnormalities and improved the survival rate of LPS/D-Gal-challenged mice. Pretreatment with rotenone has no obvious effects on hepatic malondialdehyde (MDA) contents but it significantly inhibited the up-regulation of both hepatic mRNA level and plasma protein level of TNF-α and IL-6. In the rotenone-pretreated group, the elevation of hepatic caspase-3, caspase-8 and caspase-9 activities induced by LPS/D-Gal decreased and rotenone reduced the count of TUNEL-positive apoptotic hepatocytes. In addition, posttreatment with rotenone at 1h after LPS/D-Gal challenge also suppressed the elevation of plasma aminotransferases. These data suggest that mitochondrial complex I inhibition might be a potential approach for the control of inflammation.

Effect ofPunica Granatum(Pomegranate) Juice Extract on Healthy Liver and Hepatotoxicity Induced by Diethylnitrosamine and Phenobarbital in Male Rats

The effect of pomegranate juice (PJ) on hepatic antioxidant enzyme activities, lipid peroxidation, DNA fragmentation (DNAF), and caspase-3 activity in rats both treated and not treated with diethylnitrosamine (DEN) and phenobarbital (PB) was studied. Administration of rats with DEN and PB caused an elevation in the levels of malondialde-hyde (MDA), DNAF, and activities of glutathione reductase (GSR) and caspase-3, while the activities of superoxide dismutase, glutathione S-transferase, total glutathione peroxidase (t-GPx), and glutathione (GSH) level were decreased in hepatocytes compared to the control. Treatment of rats with PJ pre, during, and post DEN and PB administration improved liver function and decreased the levels of MDA, DNAF, t-GPx, GSR, and caspase-3 activities, but the GSH level did not change compared to the D-P group. This indicates that PJ reduced the oxidative stress and apoptosis induced by DEN and PB. Administration of healthy rats with PJ only for a long period induced oxidative stress and apoptosis for hepatocytes.

Arsenic-induced hepatic mitochondrial toxicity in rats and its amelioration by diallyl trisulfide

The present investigation was aimed to investigate the possible protective role of diallyl trisulfide (DATS) against arsenic (As)-induced hepatic mitochondrial toxicity in rats. Mitochondria were isolated from the liver tissue of rats from all the groups. Lipid profile, lipid peroxidation, antioxidant enzyme activities, hepatic function enzymes, mitochondrial swelling, cytochrome c oxidase activity, mitochondrial Ca+-ATPase and Na+/K+-ATPase activity, mitochondrial calcium content and mitochondrial enzyme activities were measured. Short-term As exposure (5 mg/kg bw/d for 28 d) caused liver damage as evidenced by changes in activities of liver enzymes. The effects of As were coupled with enhanced reactive oxygen species generation, mitochondrial swelling, inhibition of cytochrome c oxidase, complex I-mediated electron transfer, decreased Ca2+-ATPase and Na+/K+-ATPase activity, a reduction in mitochondrial calcium content, changes in indices of hepatic mitochondrial oxidative stress, significant increase in mitochondrial lipid peroxidation products and alterations in mitochondrial lipid profile. Significant decreases in mitochondrial antioxidants and tricarboxylic acid cycle enzymes were also found in the liver mitochondria of As-induced hepatic mitochondrial toxicity in rats. As also increased hepatic caspase-3 activity and DNA fragmentation. All these apoptosis-related molecular changes caused by As could be alleviated by supplementation with DATS, which likely suggests a protective role against As-induced hepatotoxic changes and hepatic mitochondrial toxicity. The protective effect of DATS on the liver mitochondria was evidenced by altering all the changes induced by As. Free radical scavenging and metal chelating activities of DATS may be the mechanism, responsible for the protective action against As-induced mitochondrial damage in liver.

Amelioration of Liver Injury by Continuously Targeted Intervention against TNFRp55 in Rats with Acute-on-Chronic Liver Failure

BackgroundAcute-on-chronic liver failure (ACLF) is an acute deterioration of established liver disease. Blocking the TNF (tumor necrosis factor)/TNFR (tumor necrosis factor receptor) 1 pathway may reduce hepatocyte apoptosis/necrosis, and subsequently decrease mortality during development of ACLF. We demonstrated that a long-acting TNF antagonist (soluble TNF receptor: IgG Fc [sTNFR:IgG-Fc]) prevented/reduced development of acute liver failure by blocking the TNF/TNFR1 (TNFRp55) pathway. However, it is still unclear if sTNFR:IgG-Fc can inhibit hepatocyte damage during development of ACLF.MethodologyChronic liver disease (liver fibrosis/cirrhosis) was induced in Wistar rats by repeatedly challenging with human serum albumin (HSA), and confirmed by histopathology. ACLF was induced with D-galactosamine (D-GalN)/lipopolysaccharide (LPS) i.p. in the rats with chronic liver disease. Serum and liver were collected for biochemical, pathological and molecular biological examinations.Principal FindingsReduced mortality was observed in sTNFR:IgG-Fc treated ACLF rats, consistent with reduced interleukin (IL)-6 levels in serum and liver, as well as reduced hepatic caspase-3 activity, compared to that of mock treated group. Reduced hepatic damage was confirmed with histopathology in the sTNFR:IgG-Fc treated group, which is consistent with reduced Bcl-2 and Bax, at mRNA and protein levels, but increased hepatocyte proliferation (PCNA). This is also supported by the findings that caspase-3 production was up-regulated significantly in ACLF group compared to the mock treated group. Moreover, up-regulated caspase-3 was inhibited following sTNFR:IgG-Fc treatment. Finally, there was up-regulation of hepatic IL-22R in sTNFR:IgG-Fc treated ACLF rats.ConclusionssTNFR:IgG-Fc improved survival rate during development of ACLF via ameliorating liver injury with a potential therapeutic value.

Deep hypothermia protects against acute hypoxia in vivo in rats: a mechanism related to the attenuation of oxidative stress

There is growing interest in using hypothermia to prevent hypoxic damage in clinical and experimental models, although the mechanisms regulated by hypothermia are still unclear. As reactive oxygen and nitrogen species are the main factors causing cellular damage, our objective was to study the scope of hypothermia in preventing hypoxia-induced oxidative damage. We analysed systemic and hepatic indicators of oxidative stress after an acute hypoxic insult (10% oxygen in breathing air) in normothermic (37°C body temperature) and hypothermic conditions (22°C) in rats. Exposure to hypoxia resulted in tissue damage (aspartate aminotransferase increased from 54.6 ± 6.9 U l(-1) in control animals to 116 ± 1.9 U l(-1) in hypoxia, and alanine aminotransferase increased from 19 ± 0.8 to 34 ± 2.9 U l(-1)), oxidative stress (nitric oxide metabolites increased from 10.8 ± 0.4 μM in control rats to 23 ± 2.7 μM in hypoxia, and thiobarbituric reactive substances increased from 3.3 ± 0.2 to 5.9 ± 0.4 nm) and antioxidant consumption (reduced/oxidized glutathione ratio changed from 9.8 ± 0.3 to 6.8 ± 0.3). In contrast, when hypothermia was applied prior to hypoxia, the situation was reversed, with a reduction in aspartate aminotransferase (from 116 ± 1.9 in hypoxic animals to 63 ± 7.8 U l(-1) in animals exposed to hypothermia followed by hypoxia), alanine aminotransferase (from 34 ± 2.9 to 19 ± 0.9 U l(-1)), oxidative stress (nitric oxide metabolites decreased from 23 ± 2.7 to 17.8 ± 1.9 μM and thiobarbituric acid-reactive substances decreased from 5.9 ± 0.4 to 4.3 ± 0.2 nm) and antioxidant preservation (reduced/oxidized glutathione ratio changed from 6.8 ± 0.3 to 11.1 ± 0.1). Hypoxia induced a decrease in liver enzymatic antioxidant activities even during hypothermia. Both treatments, hypoxia and hypothermia, produced a similar increase in hepatic caspase-3 activity. In conclusion, hypothermia prevented the tissue damage and oxidative stress elicited by hypoxia. Our results provide new evidence concerning the protective mechanism of hypothermia in vivo.

Dual cytoprotective effects of splenectomy for small-for-size liver transplantation in rats

The problems associated with small-for-size liver grafts (ie, high mortality rates, postoperative complications, and acute rejection) remain critical issues in partial orthotopic liver transplantation (OLT). In association with partial OLT, splenectomy (SP) is a procedure used to reduce the portal pressure. However, the precise effects of SP on partial OLT have been unclear. In this study, using small-for-size liver grafts in rats, we examined the cytoprotective effects of SP on OLT. Liver grafts were assigned to 2 groups: a control group (OLT alone) and an SP group (OLT after SP). SP significantly increased animal survival and decreased liver damage. SP exerted the following cytoprotective effects: (1) it improved hepatic microcirculation and prevented increases in the portal pressure after OLT, (2) it suppressed the hepatic infiltration of neutrophils and macrophages through the direct elimination of splenic inflammatory cells before OLT, (3) it decreased the hepatic expression of tumor necrosis factor α and interleukin-6, (4) it attenuated sinusoidal endothelial injury, (5) it decreased plasma endothelin 1 levels and increased hepatic heme oxygenase 1 expression, (6) it suppressed hepatocellular apoptosis through the down-regulation of hepatic caspase-3 and caspase-8 activity, and (7) it increased hepatic regeneration. In conclusion, SP for small-for-size grafts exerts dual cytoprotective effects by preventing excessive portal vein hepatic inflow and eliminating splenic inflammatory cell recruitment into the liver; this in turn inhibits hepatocellular apoptosis and improves liver regeneration.

Hepatoprotective effects of IL-22 on fulminant hepatic failure induced by d-galactosamine and lipopolysaccharide in mice

Interleukin-22 (IL-22), a member of the IL-10 cytokine family that is produced by activated Th22, Th1 and Th17 cells as well as natural killer cells, plays an important role in increase of innate immunity, protection from damage and enhancement of regeneration. Here, we examined the effects of IL-22 on acute liver failure model induced by d-galactosamine (GalN) and lipopolysaccharide (LPS). Administration of recombinant human IL-22 (rhIL-22) reduced the death rate markedly and prevented mice from severe hepatic injury, as evidenced by decreased serum alanine aminotransferase (ALT) and total bilirubin (T.Bil) activity as well as improved histological signs in liver. Furthermore, IL-22 treatment decreased the hepatic malondialdehyde (MDA) contents and increased the reduced glutathione levels. Serum tumor necrosis factor α (TNF-α) level and hepatic caspase-3 activity were significantly lower in mice administrated with IL-22. Moreover, IL-22 treatment significantly enhanced activation of STAT3 and up-regulated the expression of Bcl-xL, heme oxygenase-1 (HO-1) and redox factor-1 (Ref-1) in the liver injury induced by GalN/LPS. Collectively, these data indicate that IL-22 can provide critical protection against GalN/LPS-induced liver injury through anti-apoptotic, anti-oxidant and anti-inflammatory actions.

Hepatic and HSC-specific sorafenib effects in rats with established secondary biliary cirrhosis

Portal hypertension in cirrhosis depends on increased intrahepatic vascular resistance, which is explained by fibrosis and intrahepatic hyperresponsiveness to vasoconstrictors. Both are caused by activation and proliferation of hepatic stellate cells (HSCs). Portal hypertension of cirrhotic rats can be reduced by the multikinase inhibitor sorafenib, due to a reduction of intrahepatic vascular resistance. Therefore, the hepatic effects of sorafenib require further understanding. Here, we investigated hepatic and HSC-specific sorafenib effects in cirrhotic rats. Animal models of bile duct ligation-induced secondary biliary cirrhosis in rats were studied. The rats were treated with sorafenib (60 mg/kg/day) for 1 week, starting after established cirrhosis. Histological evaluation was carried out using hemalaun and eosin (HE) staining. Apoptosis was studied by PARP cleavage, colorimetric caspase-3 assay, and electrophoretic DNA detection. HSC activation was studied by hepatic Sirius red and immunohistochemical αSMA (α-smooth muscle actin) staining, and by in vitro experiments with culture-activated primary HSCs. Biochemical serum parameters suggested the occurrence of sorafenib-induced liver damage. HE staining revealed histological changes in livers of sham-operated and bile duct-ligated (BDL) rats in response to sorafenib, which were different in both groups. In BDL rats and isolated HSCs, the treatment with sorafenib reduced hepatic αSMA and procollagen-1α mRNA expression. As shown by immunohistochemical staining, perisinusoidal αSMA expression was reduced by sorafenib in BDL rats. This was associated with reduced perisinusoidal deposition of extracellular matrix, as revealed by Sirius red staining. Although no change in PARP cleavage and only a minor increase in hepatic caspase-3 activity were detected in BDL rats in response to sorafenib, livers of sorafenib-treated BDL rats contained small DNA fragments, which were not observed in untreated BDL rats. In conclusion, sorafenib treatment reduces the number of activated HSCs in cirrhotic livers. This leads to the decrease in intrahepatic vascular resistance, but also to liver damage in the dosage we used. Therefore, any translation to portal hypertensive patients who may profit from sorafenib should be done with particular care.

Protective effects of Asiaticoside on acute liver injury induced by lipopolysaccharide/D-galactosamine in mice

Asiaticoside (AS), a triterpenoid product isolated from Centella asiatica, has been described to exhibit anti-in fl ammatory activities in several inflammatory models. However, the effects of AS on liver injury are poorly understood. The present study was undertaken to investigate whether AS is efficacious against Lipopolysaccharide (LPS) /D-galactosamine (D-GalN)-induced acute liver injury in mice and its potential mechanisms. AS (5, 10 and 20 mg/kg/d) was pretreated orally once daily for 3 days before LPS/D-GalN injected in mice. The mortality, hepatic tissue histology, plasma levels of Tumor necrosis factor-alpha (TNF-alpha) and alanine aminotransferase (ALT) and aspartate aminotransferase (AST), hepatic tissue TNF-alpha and caspase-3 activity were measured. Besides, western blotting analysis of phospho-p38 mitogen-activated protein kinase (phospho-p38 MAPK), phospho-c-jun N-terminal kinase (phospho-JNK) and phospho-extracellular signal regulated kinase (phospho-ERK) were determined. As a result, AS showed significant protection as evidenced by the decrease of elevated aminotransferases, hepatocytes apoptosis and caspase-3, alleviation of mortality and improvement of liver pathological injury in a dose-dependent manner. Further, we found that AS dose-dependently reduced the elevation of phospho-p38 MAPK, phospho-JNK, phospho-ERK protein and TNF-alpha mRNA expression in liver tissues and plasma TNF-alpha. These results suggest that AS has remarkable hepatoprotective effects on LPS/D-GalN-induced liver injury and the possible mechanism is related to inhibition of TNF-alpha and MAPKs.

Dual effect of erythropoietin on liver protection and regeneration after subtotal hepatectomy in rats

The only currently offered curative option for many patients with primary or secondary liver tumors is the resection of hepatic tumors. The aim of this study was to evaluate the role of recombinant human erythropoietin (rhEPO) in liver protection and regeneration after subtotal hepatectomy in rats. Rats undergoing 70% hepatectomy received an intraperitoneal injection of saline (control) or rhEPO (4 U/g) 30 minutes prior to resection. Liver function was assessed by the measurement of the international normalized ratio (INR) levels, and hepatic injury was assessed by serum alanine aminotransferase and aspartate aminotransferase levels. Hepatic apoptosis was assessed by intrahepatic caspase-3 activity and morphological criteria. The regeneration capacity of remnant livers was assessed over 7 days with the regenerated liver/body weight ratio, immunohistochemistry markers of cell proliferation (Ki-67) and angiogenesis (von Willebrand factor), and phosphorylated extracellular signal-regulated kinase signaling. Two and 4 days after subtotal hepatectomy, the regenerated liver/body weight ratio was significantly higher in animals treated with rhEPO versus the control group (P < 0.005). Serum liver enzymes and INR levels on days 2 and 4 post-hepatectomy were significantly lower in animals pretreated with rhEPO in comparison with the control group (P < 0.005). No statistically significant difference was noted in intrahepatic hepatic caspase-3 activity, immunohistochemistry for caspase-3, or a terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling assay between the hepatectomized groups. In the rhEPO-pretreated group, the mitotic index, Ki-67 and von Willebrand factor expression, and extracellular signal-regulated kinase activity were significantly higher on day 2 post-hepatectomy (P < 0.05) in comparison with the control group. In conclusion, rhEPO treatment may offer a unique beneficial dual-function strategy for hepatic protection and regeneration immediately after subtotal hepatectomy in rats.

Protective role of carnosine in mice with cadmium-induced acute hepatotoxicity

The hepatoprotective effect of carnosine was investigated against cadmium-induced acute liver injury in mice. Hepatotoxicity was induced by a single i.p. injection of cadmium chloride (6.5 mg/kg). Carnosine treatment (10 mg/kg/day, i.p.) was applied for three consecutive days, starting one day before cadmium administration. Carnosine significantly decreased the cadmium-induced elevations in serum aminotransferases. Carnosine suppressed lipid peroxidation and restored the deficits in the antioxidant defense mechanisms (reduced glutathione level, and catalase and superoxide dismutase activities) in liver tissue resulted from cadmium administration. Also, the reductions in hepatic nitric oxide and zinc ion levels, and the increases in hepatic cadmium ion concentration, and myeloperoxidase and caspase-3 activities following cadmium exposure were significantly attenuated by carnosine treatment. In addition, carnosine markedly ameliorated cadmium-induced liver tissue damage as evidenced by light and electron microscopic examinations. It was concluded that carnosine can be considered a potential candidate to protect the liver against the deleterious effect of acute cadmium intoxication.

The BH3-Only Protein Bid Does Not Mediate Death-Receptor-Induced Liver Injury in Obstructive Cholestasis

The accumulation of bile acids during obstructive cholestasis causes liver injury and fibrosis, which is at least partly mediated by the death receptors Tumor necrosis factor-related apoptosis-inducing ligand, Tumor necrosis factor-alpha, and Fas. The BH3-interacting domain death agonist Bid is a critical mediator of death receptor-induced apoptosis in hepatocytes. Our aim for this study was, therefore, to elucidate whether Bid also mediates death receptor-induced liver injury in obstructive cholestasis. Overall, survival and various aspects of liver injury were analyzed in wild-type and Bid(-/-) mice after bile duct ligation (BDL), a commonly used model to study obstructive cholestasis in mice. Liver injury was examined at 3, 7, and 14 days after BDL. Loss of Bid did not affect the number of bile infarcts, serum aspartate aminotransferase values, or animal survival. Processing of procaspase-3 and procaspase-9, and caspase-3 enzyme activities, were not detectable in either group, and Bid(-/-) mice displayed the same pattern of terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling positive hepatocytes as wild-type controls following BDL. In contrast to Fas-receptor deficient lpr mice, hepatic fibrosis and the inflammatory response was not affected by loss of Bid. Together, these data suggest that Bid is not a downstream target of death receptors in obstructive cholestasis and does not significantly contribute to bile acid induced liver injury and fibrosis.

Mechanism of the suppression against d-galactosamine-induced hepatic injury by dietary amino acids in rats

To elucidate the mechanism by which dietary amino acids suppress the D-galactosamine (D-GalN)-induced hepatitis, we examined the involvement of Kupffer cells, tumor necrosis factor-alpha (TNF-alpha) and apoptosis in the mechanism. In experiment 1, the rats were fed with 10% L-glutamine or 5% glycine diet injected with D-GalN with or without gadolinium chloride (GdCl3)-pretreatment. The results indicated that these amino acids suppressed the D-GalN-induced elevation of serum transaminase activities, irrespective of GdCl3-pretreatment. In experiment 2, rats were fed with 10% of L-glutamine, L-serine, L-alanine or L-glutamic acid diets injected with D-GalN. The results demonstrated that all these amino acids suppressed the D-GalN-induced elevation of serum transaminase activities, but that serum TNF-alpha concentrations and hepatic caspase-3 activities in the rats were not appreciably changed. In conclusion, the suppressive effects of amino acids on D-GalN-induced hepatitis were suggested not to be always mediated by the inhibition of Kupffer cells --> TNF-alpha --> apoptosis pathway.