The experimental trial was conducted on bull white mice, ddY strain, weight of 18-20g, were randomly assigned to different lots, with 8-15 mice in each lot. Experimental product was total dry lingzhi extract produced by OPC Company which reached standards at basic level. Lingzhi dry powder was prepared with distilled water to make liquid extract with concentration of 15% (15g dry extract/100ml water), 30% and 75%. These liquid extracts were used to test hepatoprotective effect effect of Lingzhi extract on chronic hepatic toxication by DEN and CCl4. The results showed that lingzhi extract did not show microprotective effects. However, lingzhi extract can inhibit hyperplastic nodule formation. On the other hand, lingzhi extract also reduced abnormal cells formation
Drugs, Chinese Herbal ; Liver ; Diethylnitrosamine ; Carbon Tetrachloride

No abstract available.
Animals ; Autoradiography ; Carcinogenesis* ; Diethylnitrosamine ; gamma-Glutamyltransferase* ; Liver* ; Rats*

No abstract available.
Animals ; Autoradiography ; Carcinogenesis* ; Diethylnitrosamine ; gamma-Glutamyltransferase* ; Liver* ; Rats*

OBJECTIVETo establish a zebrafish model of liver fibrosis via diethylnitrosamine (DEN)-induced liver injury.

METHODSA total of 120 wild-type 3-month-old zebrafish were randomly divided into DEN-treated group and control group. The survival rate and behavioral changes of each group were observed. After treatment with DEN for 2, 4, and 6 weeks, liver index was measured, and liver fibrosis was evaluated with HE staining, Gomori staining and Sirius red staining.

RESULTSNo obvious behavioral change was observed in DEN-treated group during the experiment. Compared with that in control group, the liver index of zebrafish in DEN-treated group showed no significantly changes at the time points of observation. Proliferation of reticulate fibers was found in 30% of zebrafish treated with DEN for 4 weeks, and the rate increased to 80% at 6 weeks when reticulate fibers and collagen fibers actively proliferated to result in fiber collapse and formation of fibrotic nodules.

CONCLUSIONA stable zebrafish liver fibrosis model was successfully established by inducing liver damage to facilitate studies of the pathogenesis of liver fibrosis and screening therapeutic drugs.

Animals ; Diethylnitrosamine/toxicity* ; Liver ; Liver Neoplasms/chemically induced* ; Rats

OBJECTIVETo investigate the expression and role of B-cell translocation gene 2(BTG2) in the carcinogenesis of hepatocellular carcinoma (HCC).

METHODSModified Diethylnitrosamine (DEN)-induced primary hepatocellular carcinoma rat model was established. The expression of BTG2, p53 and cyclinD1 was detected by RT-PCR, western blot and immunohistochemistry.

RESULTSThe BTG2 protein was predominantly localized in the nucleus, with faint cytoplasmic staining in normal liver cells; however, it is mainly a cytoplasmic protein in HCC cells. BTG2 was over-expressed during the early stage after DEN treatment, the expression level peaked at 5 weeks and then it gradually decreased to the normal level after 16 weeks. The expression of cyclin D1 and cyclin E was increased gradually after DEN treatment, and peaked at 16 weeks and 5 weeks respectively. A significant increase in p53 was not observed until 5 weeks after DEN treatment, and it gradually decreased after 16 weeks.

CONCLUSIONSDecreased expression of BTG2 may be an important step in carcinogenesis of HCC. BTG2 may positively regulate p53 expression and negatively regulate cyclin D1 expression in the carcinogenesis of HCC.

OBJECTIVE: To study the inhibitory effect of pills (BJJ) agaisnt diethylnitrosamine (DEN)-induced hepatocarcinogenesis and explore the relation between this effect and the inflammasome signaling pathway. METHODS: Sixty-five male SD rats were randomly divided into control group, DEN model group, and 3 BJJ treatment groups at low, medium and high dose (with daily dose of 0.55, 1.1 and 2.2 g/kg, respectively, for 12 consecutive weeks starting from the 5th week after modeling). The pathological changes of the liver tissue were observed with HE and Masson staining, and serum levels of alanine transaminase (ALT), glutamic oxaloacetic transaminase (AST), alkaline phosphatase (ALP) and total bilirubin (TBIL) of the rats were detected using ELISA. Oxidation stress in the liver tissue was assessed with ELISA, and Western blotting and ELISA were used to detect the molecular expressions of inflammasome-related pathway. RESULTS: BJJ significantly inhibited tumor growth in the liver of the rats. HE and Masson staining showed that BJJ treatment obviously ameliorated liver fibrosis and reduced cancer cell and inflammatory cell infiltration in the liver. BJJ significantly reduced elevations of serum ALT, AST, ALP and TBIL levels, increased the contents of superoxide dismutase, catalase and glutathione peroxidase in the liver and suppressed malondialdehyde in Den-treated rats. BJJ also dose-dependently decreased the expressions of NLRP3, apoptosis-associated speck-like protein (ASC), caspase-1, pro-IL-1β, pro-IL-18, IL-1β and IL-18 in the liver of Den-treated rats. CONCLUSIONS BJJ treatment can dose-dependently inhibit DEN-induced hepatocarcinogenesis by enhancing antioxidant capacity and down-regulating inflammatory-related pathways in rats.
Animals ; Aspartate Aminotransferases ; Diethylnitrosamine ; Liver ; Liver Neoplasms ; Male ; Rats ; Rats, Sprague-Dawley

The expression of major human apurinic/apyrimidinic DNA endonuclease (APEX) from its cDNA in E. coli (DH5 alpha) was attempted in order to obtain a biologically active recombinant APEX. E. coli cells were transformed by a prokaryotic translation vector (pGEX-4T-3) harboring APEX cDNA. GST-APEX fusion protein with a molecular weight of 6.3 KDa was induced by IPTG (1.0 mM) treatment. Western blot immunodetection identified the induced protein as the GST-APEX fusion protein. The survival rate of E. coli cells (DH5 alpha) transformed with pGEX-4T-3-APEX increased when the cells were treated with N-diethyl-N-nitrosamine (DENA) or 3'-methyl-4-monomethylaminoazobenzene (3'-MeMAB), indicating that APEX expression had a protective effect on the cytotoxicity of these carcinogens. The fusion protein extracted from E. coli cells and purified by GSH-agarose gel affinity chromatography exhibited APEX activity. Treatment of thrombin to the GST-APEX fusion protein and affinity purification followed by Sephacryl S-100 gel filtration resulted in APEX peptide with MW 36 KDa, which exhibited AP DNA repair activity (8,7000 EU/mg protein). N-ethylmaleimide (0.1 mM) or AMP (0.98 mM) inhibited APEX activity by 50% and kinetic analysis indicated that the recombinant APEX (rAPEX) had a Km value of 0.022 microM (AP sites for AP DNA) and the Ki value was 0.48 mM for AMP. These results indicated that E. coli cells expressing biologically active GST-APEX were resistant to the cell damage caused by chemical carcinogens and that rAPEX purified from E. coli cells transformed with APEX cDNA-inserted translation vector was similar to native APEX in some properties.
Carbon-Oxygen Lyases/biosynthesis* ; Diethylnitrosamine/pharmacology ; Escherichia coli/genetics ; Escherichia coli/drug effects ; Human ; Recombinant Fusion Proteins/biosynthesis*

Vitamin C (ascorbic acid) is an essential nutrient of most living tissues. We established a strain of Gulo-/- mice with known deficiency, in which vitamin C intake can be controlled by diet, like humans, and investigated the differentially expressed proteins following treatments with Helicobacter pylori and diethylnitrosamine (DENA) in the liver of Gulo-/- mice using a proteomic approach. Expression of p53, 14-3-3epsilon and 14-3-3delta in Gulo-/- mice liver tissue was analyzed by immunohistochemistry. 2-DE maps constructed from Gulo-/- mice liver and differentially expressed proteins in liver tissue were identified by matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF/MS). In Gulo-/- mice after H. Pylori infection, followed by treatment with DENA, no differences in p53, 14-3-3epsilon and 14-3-3delta were observed by immunohistochemistry. Proteome analyses using MALDI-TOF/MS resulted in successful identification of 12 proteins (nine proteins were up-regulated and three were down-regulated). Specifically, peroxiredoxin-6 and Alpha-1-antitrypsin 1-4 were up-regulated in liver after H. Pylori infection followed by treatment with DENA. These results indicated that oral supplementation with vitamin C led to rescue of Gulo-/- mice from vitamin deficiency, and protected the liver from H.pylori infection and/or DENA effect, and vitamin C also protected the liver against oxidative stress.
Animals ; Ascorbic Acid ; Avitaminosis ; Diet ; Diethylnitrosamine* ; Helicobacter pylori* ; Helicobacter* ; Humans ; Immunohistochemistry ; Liver* ; Mice* ; Oxidative Stress ; Proteins* ; Proteome

PURPOSE: Tc-99m labeled diethylenetriaminepentaacetic acid (DTPA) -coupled galactosylated human serum albumin (GSA) is a currently used imaging agent for asialoglycoprotein receptor (ASGPR) of the liver, but, it has several shortcomings. Recently a new ASGPR imaging agent, (99m) Tc-lactosylated human serum albumin (LSA), with simple labeling procedure, high labeling efficiency, high stability was developed. In order to assess the feasibility of the (99m) Tc-LSA as a ASGPR imaging radiopharmaceuticals, we performed biodistribution study of the tracer in liver injured mice model and the results were compared with histolgic data. MATERIALS AND METHODS: To induce hepatic damage in ICR mice, diethylnitrosamine (DEN) (60 mg/kg/week X 5 time, low dose or 180 mg/kg/week X 2 times, high dose) and thioacetamide (TAA) (50 mg/kg X 1 time) were administrated intraperitoneally. Degree of liver damage was evaluated by tissue hematoxilin-eosin stain, and expression of asialoglycoprotein receptor (ASGPR) was assessed by immunohistochemistry using ASGPR antibody. (99m) Tc-LSA was intravenously administrated via tail vein in DEN or TAA treated mice, and biodistribution study of the tracer was also performed. RESULTS: DEN treated mice showed ballooning of hepatocyte and inflammatory cell infiltration in low dose group and severe hapatocyte necrosis in high dose group, and low dose group showed higher ASGPR staining than control mice in immunohistochemical staining. TAA treated mice showed severe hepatic necrosis. (99m) Tc-LSA Biodistribution study showed that mice with hepatic necrosis induced by high dose DEN or TAA revealed higher blood activity and lower liver activity than control mice, due to slow clearance of the tracer by the liver. The degree of liver uptake was inversely correlated with the degree of histologic liver damage. But low dose DEN treated mice with mild hepatic injury showed normal blood clearance and hepatic activity, partly due to overexpression of ASGPR in mice with mild degree hepatic injury. CONCLUSION: Liver uptake of (99m) Tc-LSA was inversely correlated with degree of histologic hepatic injury in DEN and TAA treated mice. These results support that (99m) Tc-LSA can be used to evaluate the liver status in liver disease patients.
Animals ; Asialoglycoprotein Receptor ; Diethylnitrosamine* ; Hepatocytes ; Humans ; Immunohistochemistry ; Liver Diseases ; Liver* ; Mice* ; Mice, Inbred ICR ; Necrosis ; Radiopharmaceuticals ; Serum Albumin* ; Thioacetamide* ; Veins