BACKGROUND: Duration of action varies in conditions with reduced plasma cholinesterase activity. To evaluate the action duration and recovery of mivacurium under the experimental hepatic failure and cholestasis, the pharmacodynamic studies were done. METHODS: The pharmacodynamic studies were done using a common peroneal nerve-anterior tibialis muscle preparation in 18, either sex, adult cats (weight, 2.0~4.0 kg). For the hepatic failure, galactosamine chloride (4.25 mmol/kg) was given 16 hours prior to the pharmacodynamic study. The cholestasis was made by the ligation of CBD and cystic duct 8 days prior to the pharmacodynamic study. All the cat had 4XED95 of mivacurium. The action durations and recovery indices were measured. And plasma cholinesterase activities were checked. RESULTS: The duration of mivacurium was prolonged significantly with either hepatic failure (14.96 4.44 min.) or cholestasis (11.21+/- 5.11 min.) group compared to control group (5.27 +/-0.67 min.) and also the recovery indices were significantly increased in the hepatic failure (4.58+/- 1.40 min.) and cholestasis (3.21+/- 1.00 min.) groups, as compaired with the control group (1.57+/- 0.40 min.). CONCLUSION: The mivacurium-induced neuromuscular blockade is prolonged by the experimental hepatic failure and cholestasis, and the effects may be caused by the hepatic dysfunction, impairment of direct biliary excretion.
Adult ; Animals ; Cats* ; Cholestasis* ; Cholinesterases ; Cystic Duct ; Galactosamine ; Humans ; Ligation ; Liver Failure* ; Neuromuscular Blockade* ; Plasma

Animals ; Disease Models, Animal ; Ephedra ; chemistry ; Galactosamine ; Lipopolysaccharides ; Liver Failure, Acute ; chemically induced ; drug therapy ; Protective Agents ; pharmacology ; Rats

BACKGROUND: Though the mivacurium is the short acting nondepolarizing neuromuscular blocking agent, the action duration of it is not prospected in condition of hepatic failure owing to its being metabolized by serum cholinesterase and other esterase produced in liver. The purpose of this study is to evaluate neuromuscular effect of the mivacurium in cats with acute hepatic failure. METHODS: Six cats administrated only mivacurium are the control group, and six cats with acute hepatic failure by galactosamine hydrochloride are the experimental group. The force of the anterior tibialis muscle in response to supramaximal common peroneal nerve stimulations were recorded, the time intervals from mivacurium administration to attain 100% twitch depression (onset time), from mivacurium administration to recovery of 25% twitch tension (duration) and from 25% to 75% of twitch recovery (recovery index) were compared between the control group and the experimental group. RESULTS: In experimental group, SGOT and SGPT prior to administration of galatosamine were 28.8+/-5.6 (IU/L) and 43.0+/-7.9 (IU/L), respectively, SGOT and SGPT in acute hepatic failure were 5004.0+/-8113.2 (IU/L) and 3763.0+/-5416.4 (IU/L), respectively, and there were significant differences between the control group and the experimental group. The action duration{47.6+/-18.0 (min)} and the recovery time{7.7+/-3.7 (min)} of mivacurium in the experimental group were more prolonged than the action duration{21.9+/-5.0 (min)} and the recovery time{4.4+/-0.7 (min)} of mivacurium in the control group. CONCLUSIONS: These results indicate that the hepatic failure can prolong the action duration and the recovery index of mivacurium, but the other factors affecting the action of mivacurium must be studied.
Alanine Transaminase ; Animals ; Aspartate Aminotransferases ; Cats* ; Cholinesterases ; Depression ; Galactosamine* ; Liver ; Liver Failure ; Liver Failure, Acute* ; Neuromuscular Agents ; Neuromuscular Blockade* ; Peroneal Nerve

OBJECTIVETo study the intestinal expression of defensin-5 (RD-5), soluble phospholipase A2 (sPLA2) and lysozyme in acute liver failure (ALF) using rat models, and to determine the relation of these expressions to intestinal bacterial translocation.

METHODSForty-eight healthy male Sprague-Dawley rats were divided into a control group (n=8) and a model group (n=40; intraperitoneal injection of 10% D-galactosamine). The model group was further divided into five subgroups according to the time lapse after model establishment (8, 16, 24, 48, and 72 hours). At the end of the experiments, homogenates of mesenteric lymph nodes, liver and spleen were cultured in agar for bacterial outgrowth.Hematoxylin-eosin stained sections of liver and terminal ileum were examined under an optical microscope to assess pathological changes. mRNA expression of RD-5, sPLA2 and lysozyme in the terminal ileum was determined by reverse transcription-polymerase reaction (RT-PCR), and protein expression of sPLA2 and lysozyme from the same anatomic location was determined by western blotting and immunohistochemistry. Means between groups were compared with one-way analysis of variance.

RESULTSALF was successfully induced in the D-galactosamine injected rats. No bacteria grew in the organ cultures from the control group, while 8.3%, 37.5% and 58.3% of the rats in the 24-, 48-and 72-hour model groups showed positive cultures. Despite this, the structure of the terminal ileum from the rats in the 72-hour model group was nearly intact, without obvious necrosis of mucosal epithelial cells. Expression of RD-5 and sPLA2 mRNA in the model groups gradually increased at early time points and peaked at 16 hours after induction of ALF (1.291+/-0.153 and 1.131+/-0.128), which was significantly higher than that detected in the control group (0.725+/-0.116 and 0.722+/-0.112, t=69.25, 95.71, all P<0.01). After that, the expression of RD-5 and sPLA2 mRNA progressively decreased, and by 72 hours after the induction of ALF, the expression (0.415+/-0.104 and 0.425+/-0.076) was significantly lower than that of the control group (t=31.55 and 44.98, all P<0.01). Lysozyme mRNA expression in the model group peaked at 8 hours after ALF induction (1.211+/-0.107), which was higher than that of the control group at this time point (0.853+/-0.093), and by 72 hours after ALF induction it declined to 0.704+/-0.103, which was significantly lower than that of the control group (t=9.224; all P=0.009). In addition, at 72 hours after ALF induction the protein expression of both lysozyme and sPLA2 was significantly lower in the model group (0.327+/-0.086 and 0.382+/-0.057) than in the control group (0.583+/-0.121 and 0.650+/-0.093, t=12.28 and 15.83, P=0.004 and 0.001). Similar results were obtained with immunohistochemical staining.

CONCLUSIONThe function of the ileal mucosal immune barrier in the rat model of acute liver failure decreased, along with decreases in expression of RD-5, sPLA2 and lysozyme in the Paneth cells.At the same time, the rate of organ bacterial translocation increased without obvious injury to the intestinal mucosa structure.

Animals ; Bacterial Translocation ; Defensins ; Disease Models, Animal ; Galactosamine ; Injections, Intraperitoneal ; Intestines ; Liver Failure, Acute ; Male ; Muramidase ; Phospholipases A2 ; Protein Precursors ; RNA, Messenger ; Rats ; Rats, Sprague-Dawley

OBJECTIVETo investigate the role of the Notch signaling pathway, and the underlying mechanism, in development of acute liver failure (ALF) in a mouse model.

METHODSFor in vivo analysis of the role of Notch signaling in ALF, a mouse model of ALF was generated by intraperitoneal injection of 3.0 g/kg D-galactosamine. Histological specimens were stained by hematoxylin-eosin, and then studied microscopically.Expression level of Jaggedl, Notchl, NICD, and Hes5 was measured by western blotting (for protein) and real time-PCR (for mRNA). The level of CD68 protein was detected by immunohistochemical staining. Serum levels of alanine aminotransferase (ALT), aspartate aminotransferase (AST), IL-10, high mobility group box 1 (HMGB1) chromatin protein, and lipopolysaccharide (LPS) were measured by standard methods. For in vitro analysis of the molecular mechanism, the RAW264.7 macrophage cell line was cultured with LPS in the absence or presence of the Notch inhibitor DAPT, and the intracellular levels of Notch1, NICD, and Hes5 were measured by western blotting and real time-PCR and the extracellular levels of IL-10 and HMGB1 were detected in the supematant.

RESULTSCompared with unmodeled (normal control) mice, the ALF modeled mice showed higher levels of serum ALT (848.40+/-94.83 U/L vs. 38.99+/-9.63 U/L), AST (911.49+/-67.65 U/L vs. 55.28+/-7.50 U/L), HMGB1 (101.91+/-12.43 µg/L vs. 20.73+/-5.37 µg/L), 1L-10 (4 627.88+/-842.45 pg/mL vs. 1 064.92+/-238.46 pg/mL) and LPS (11.80+/-0.89 EU/mL vs. 0.58+/-0.12 EU/mL), as well as higher expression of Jagged1 (mRNA: 7.63+/-1.41 vs. 1.00+/-0.00; protein: 0.71+/-0.07 vs. 0.34+/-0.07), Notch1 (mRNA: 7.10+/-0.66 vs. 1.00+/-0.00; protein: 0.66+/-0.11 vs. 0.27+/-0.08), NICD (protein: 0.76+/-0.08 vs. 0.27+/-0.08), Hes5 (mRNA: 7.95+/-0.71 vs. 1.00+/-0.00; protein: 1.20+/-0.07 vs. 0.76+/-0.07), and CD68 (protein: 7 685.05+/-417.34 vs. 2 294.01+/-392.93) (all P<0.01). In vitro, LPS increased the extracellular levels of HMGB1 (7.44+/-0.63 vs. 0.21+/-0.05), IL-10 (315.19+/-79.13 vs. 59.19+/-23.30) and the intracellular expression of Notch1 (mRNA: 6.49+/-0.73 vs. 1.00+/-0.00), NICD (protein: 0.65+/-0.10 vs. 0.23+/-0.07), and Hes5 (mRNA: 7.30+/-0.85 vs. 1.00+/-0.00; protein: 0.96+/-0.10 vs. 0.54+/-0.07) (all P<0.01). DAPT treatment led to a decrease above the index serum levels of HMGB1 (6.22+/-0.71) and IL-10 (252.06+/-57.63), and of expression of Notch 1 (mRNA: 3.20+/-0.68), NICD (protein: 0.42+/-0.05), and Hes5 (mRNA: 4.72+/-0.67; protein: 0.84+/-0.09) (P<0.01 or <0.05).

CONCLUSIONThe Notch signaling pathway may plan an important role in the development of ALF upon activation of the pathway in macrophages by LPS and leading to promoted secretion of HMGB 1 and IL-10, with a greater effect on the former.

Alanine Transaminase ; Animals ; Aspartate Aminotransferases ; Disease Models, Animal ; Galactosamine ; HMGB1 Protein ; Lipopolysaccharides ; Liver Failure, Acute ; Mice ; RAW 264.7 Cells ; RNA, Messenger ; Receptors, Notch ; metabolism ; Signal Transduction

OBJECTIVETo apply an orthogonal design optimization strategy to a mouse model of acute liver failure induced by D-galactosamine (D-GalN) and lipopolysaccharide (LPS) exposure.

METHODSA four-level orthogonal array design (L16(45)) was constructed to test factors with potential impact on successful establishment of the model (D-GalN and LPS dosages, and dilution rate of the D-GalN/LPS mixture). The mortality rate of mice within 24 hours of D-GalN/LPS administration was determined by the Kaplan-Meier method. The model outcome was verified by changes in serum alanine transferase level, liver histology, and hepatocyte apoptosis.

RESULTSThe orthogonal array identified the optimal model technique as intraperitoneal injection of a combination of D-GalN and LPS at dosages of 350 mg/kg and 30 mug/kg, respectively, and using a dilution rate of 3. The dosages tested had no effect on survival. The typical signs of liver failure appeared at 6 hrs after administration of the D-GalN/LPS combination.

CONCLUSIONThe orthogonal design optimization strategy provided a procedure for establishing a mouse model of acute liver failure induced by D-GalN and LPS that showed appropriate disease outcome and survival, and which will serve to improve future experimental research of acute liver failure.

Animals ; Apoptosis ; Disease Models, Animal ; Galactosamine ; adverse effects ; Lipopolysaccharides ; adverse effects ; Liver Failure, Acute ; chemically induced ; Male ; Mice ; Mice, Inbred C57BL

OBJECTIVETo reproduce a Kunming murine endotoxin shock model suitable for the anti-endotoxin pharmaceutical research.

METHODSKunming mice were challenged with an intraperitoneal (i. p.) injection of different doses of D-galactosamine (D-Gal) and endotoxin (LPS) and divided into 10 groups: i.e, group 1 [with injection of isotonic saline solution (NS) and LPS]; group 2 (with injection of NS and 90mg/kg LPS), group 3 (with injection of NS and 500mg/kg D-Gal), group 4 (with injection of 500mg/kgD-Gal and 25 microg /kg LPS), group 5 (with injection of 500mg/kg D-Gal and 50 microg/kg LPS), group 6(with injection of 500mg/kg D-Gal and 250 microg/kg LPS), group 7( with injection of NS and 600mg/ kg D-Gal), group 8 (with injection of 600mg/kg D-Gal and 10 microg/kg LPS), group 9( with injection of 600mg/kg D-Gal and 25 microg/kg LPS), group 10 (with injection of 600mg/kg D-Gal and 50 microg/kg LPS). The death of the mice were observed and the mortality rate was recorded at 48 post-injection hour (PIH). The dose of D-Gal and LPS which caused 100% lethality was chosen for the subsequent experiment to serve as control group (with injection of NS and 600mg/kg D-Gal), LPS group (with injection of 600mg/kg D-Gal and 580mg/kg LPS for later experiment). The venous blood of the mice were collected for the detection of serum content of TNF-alpha with ELISA method at 30, 75 and 120 post-injection minutes (PIM). The tissues of lung, liver, intestine were also harvested at 5 PIH for the pathological examination.

RESULTSThe lethality of mice was 100% in the groups 2, 6 and 10 (P < 0.01). The serum content of TNF-alpha was maintained in a low level in control group, but it increased remarkably in LPS group, and it reached peak at 75 PIM (6365 +/- 2087ng/L, P < 0.01). Obvious inflammatory reaction was observed in the lung, liver and intestine in LPS group, while only mild inflammatory reaction was observed in liver in control group.

CONCLUSIONThe Kunming mice showed signs of endotoxin shock after D-galactosamine presensitizing and endotoxin challenge, and it is suitable for anti-endotoxin pharmaceutical research.

Animals ; Disease Models, Animal ; Female ; Galactosamine ; adverse effects ; Male ; Mice ; Mice, Inbred Strains ; Serum ; chemistry ; Shock, Septic ; chemically induced ; Tumor Necrosis Factor-alpha ; metabolism

OBJECTIVETo compare the effects of different approaches to establishing rat models of acute liver failure (ALF).

METHODSSixty-eight Sprague-Dawley rats were randomly divided into 3 groups for establishing ALF models using 3 different approaches, namely conventional hepatectomy for resecting 90% liver tissue as described by Higgins and Anderson, modified bloodless hepatectomy for resecting 90% liver tissue, and intraperitoneal injections of 700 mg/kg D-galactosamine (D-gal) and 5 µg/kg lipopolysaccharide (LPS). The mortality of the rats due to postoperative bleeding and survival rate at 7 days after the surgery were recorded. The levels of alanine aminotransferase (ALT), total bilimbin (Tbil), albumin (ALB), NH3, glucose (Glu) and prothrombin time (PT) were monitored, and histopathologies of the liver were examined at 24 and 72 h after the surgery.

RESULTSThe mortality rate due to postoperative bleeding was higher in conventional hepatectomy group than in the modified surgical group (15% vs 0). The survival rate at 7 days was 25%, 0%, 15% in conventional surgical group, modified surgical group and drug injection group, respectively. In the latter two groups, significant changes of ALT, Tbil, ALB, NH3, Glu, and PT were recorded at 24 and 72 h after the modeling (P<0.05), and these changes were the most obvious at 24 h in modified surgical group and at 72 h in the drug injection group; ALB in both groups declined to the lowest at 7 days and then increased gradually. Liver cell degeneration and necrosis were found in modified surgical group and drug injection group at 24 h and 72 h after the modeling.

CONCLUSIONBoth the modified 90% bloodless hepatectomy and injections of D-gal and LPS can be used to establish ideal rat models of ALF to suit different ALF-related researches.

Animals ; Disease Models, Animal ; Galactosamine ; adverse effects ; Hepatectomy ; Injections, Intraperitoneal ; Lipopolysaccharides ; adverse effects ; Liver Failure, Acute ; chemically induced ; pathology ; Rats ; Rats, Sprague-Dawley

OBJECTIVETo study the role of endoplasmic reticulum stress (ERS) in acute liver failure (ALF) using a mouse model of D-Galactosamine/lipopolysaccharide (D-GalN/LPS)-induced ALF.

METHODSThe ALF model was established by administering intraperitoneal (i.p.) injections of D-Ga1N (700 mg/kg) and LPS (10 mug/kg) to six C57BL/6 mice. Three of the modeled mice were also administered 4-phenylbutyrate (4-PBA; 100 mg/kg i.p.) at 6 hours before the onset of ALF and served as the intervention group. Non-modeled mice served as controls. All mice were analyzed by western blotting and qRT-PCR to determine the expression levels of ERS-related proteins in liver tissue. Liver function was assessed by measuring levels of alanine aminotransferase (ALT) and aspartate aminotransferase (AST) in serum. Extent of injury to the liver tissue was assessed by hematoxylin-eosin staining and histological analysis. qRT-PCR was also used to detect differences in expression of inflammation-related genes, and western blotting was also used to detect differences in expression of the apoptosis related protein Caspase-3.The extent of apoptosis in liver tissue was assessed by TUNEL assay.

RESULTSThe ERS markers GRP78 and GRP94 showed increased expression at both the gene and protein levels which followed progression of ALF. The ERS effector proteins XBP-1, ATF-6 and IRE 1 a involved in the unfolded protein response were activated in the early stages of ALF, and the ERS-induced apoptosis regulators Caspase-12 and CHOP were activated in the late stage of ALF. Inhibition of ERS by 4-PBA intervention protected against injury to liver tissue and function, as evidenced by significantly lower levels of serum ALT and AST and a remarkably decreased extent of histological alterations. Furthermore, the inhibition of ERS suppressed expression of the proinflammatory cytokines TNFa, IL-6 and IL-1 β, and reduced the extent of hepatocyte apoptosis.

CONCLUSIONERS is activated in the mouse model of D-GalN/LPS-induced ALF. Inhibition of ERS may be protective against liver injury and the mechanism of action may involve reductions in inflammatory and apoptotic factors and/or signaling. Therefore, inhibiting ERS may represent a novel therapeutic approach for treating ALF.

Animals ; Apoptosis ; Disease Models, Animal ; Endoplasmic Reticulum Stress ; Galactosamine ; adverse effects ; Lipopolysaccharides ; adverse effects ; Liver Failure, Acute ; chemically induced ; metabolism ; pathology ; Male ; Mice ; Mice, Inbred C57BL

OBJECTIVETo establish a practical and reproducible animal model of human acute-on-chronic liver failure for further study of the pathophysiological mechanism of acute-on-chronic liver failure and for drug screening and evaluation in its treatment.

METHODSImmunological hepatic fibrosis was induced by human serum albumin in Wistar rats. In rats with early-stage cirrhosis (fibrosis stage IV), D-galactosamine and lipopolysaccharide were administered. Mortality and survival time were recorded in 20 rats. Ten rats were sacrificed at 4, 8, and 12 hours. Liver function tests and plasma cytokine levels were measured after D-galactosamine/lipopolysaccharide administration and liver pathology was studied. Cell apoptosis was detected by terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling assay.

RESULTSMost of the rats treated with human albumin developed cirrhosis and fibrosis, and 90% of them died from acute liver failure after administration of D-galactosamine/lipopolysaccharide, with a mean survival time of (16.1+/-3.7) hours. Liver histopathology showed massive or submassive necrosis of the regenerated nodules, while fibrosis septa were intact. Liver function tests were compatible with massive necrosis of hepatocytes. Plasma level of TNFalpha increased significantly, parallel with the degree of the hepatocytes apoptosis. Plasma IL-10 levels increased similarly as seen in patients with acute-on-chronic liver failure.

CONCLUSIONWe established an animal model of acute-on-chronic liver failure by treating rats with human serum albumin and later with D-galactosamine and lipopolysaccharide. TNFalpha-mediated liver cell apoptoses plays a very important role in the pathogenesis of acute liver failure.

Animals ; Disease Models, Animal ; Female ; Galactosamine ; adverse effects ; Humans ; Lipopolysaccharides ; adverse effects ; Liver Failure, Acute ; chemically induced ; Rats ; Rats, Wistar ; Serum Albumin ; adverse effects