metallothionein and Liver-Diseases

Zinc (Zn) is an essential and the second most abundant trace element after Iron. It can apply antioxidant, anti-inflammatory, and anti-apoptotic activity. It is assumed to be indispensable for cell division, cellular differentiation and cell signalling. Zinc is essential for proper liver function which is also the site of its metabolism. Depleted Zn concentrations have been observed in both acute and chronic hepatic diseases. It is reported that Zn deficiency or abnormal Zn metabolism during majority of liver diseases is attributed to deficient dietary intake of Zn, augmented disposal of Zn in the urine, activation of certain Zn transporters, and expression of hepatic metallothionein. Undoubtedly, Zn is involved in generating many diseases but how and whether it plays role from acute to fulminant stage of all chronic liver diseases remains to be cleared. Here, we will discuss the role of Zn in development of different diseases specifically the involvement of Zn to understand the aetiology and intricate mechanism of dynamic liver diseases.

Topics: Humans; Liver; Liver Diseases; Metallothionein; Minerals; Trace Elements; Zinc

Hepatic metallothionein (MT) expression, with various isoforms, and varying cellular localizations is a useful marker for clinico-pathogenesis of liver diseases. In acute liver toxicity caused by cadmium, carbon tetrachloride, or acetaminophen, MT plays a protective role, via the scavenging of radical species. In chronic hepatitis C patients, hepatic MT levels appear to be a biological factor associated with the severity of HCV infection, and are associated with a better response to IFN therapy. Transgenic mice that express HBsAg in the liver show hepatocellular damage, inflammation, regeneration, hyperplasia, and, eventually, neoplasia. The MT isoform, MT-1 help mitigate HBV-induced hepatitis. Analysis of MT gene expression in the livers of chronic hepatitis B patients is useful for understanding the features of distinct liver diseases and for judging disease progression. A profound down-regulation of isoform MT-1G in hepatocellular carcinoma was observed in 63% of tumors relative to the adjacent nonmalignant liver. MT has been implicated in the control of p53 folding with zinc exchange. Therefore, it appears MT may play a role in the pathogenesis of hepatocellular carcinoma. Overall MT is linked to a variety of liver diseases.

Topics: Animals; Carcinoma, Hepatocellular; Gene Expression; Humans; Liver; Liver Diseases; Liver Neoplasms; Metallothionein

A review is presented of various aspects of copper (Cu) metabolism. The Cu absorption from the gastrointestinal tract in monogastric animals differs from that in ruminants. This is influenced by Cu binding compounds, sulphide production in the rumen, and molybdenum and zinc concentrations of the diet. Moreover, the valence of the Cu ions may influence the availability of Cu in the intestine. Metallothionein and lysosomes are involved in the accumulation of copper in the liver. The different findings in various Cu storage diseases may reflect different mechanisms of disease. Cu-induced liver cell damage and haemolysis may be the result of lipid peroxidation.

Topics: Animals; Cattle; Cattle Diseases; Ceruloplasmin; Chemical and Drug Induced Liver Injury; Copper; Digestive System; Dog Diseases; Dogs; Female; Hemolysis; Intestinal Absorption; Kinetics; Liver; Liver Diseases; Lysosomes; Metal Metabolism, Inborn Errors; Metallothionein; Mice; Rats; Rumen; Sheep; Sheep Diseases

Zinc is an essential trace element that is often reduced under the type 1 diabetic condition. Previous studies demonstrated that zinc deficiency enhanced type 1 diabetes-induced liver injury and that zinc supplementation significantly helped to prevent this. Due to the differences in pathogenesis between type 1 and type 2 diabetes, it is unknown whether zinc supplementation can induce a beneficial effect on type 2 diabetes-induced liver injury. This possible protective mechanism was investigated in the present study. A high-fat diet, along with a one-time dose of streptozotocin, was applied to metallothionein (MT) knockout mice, nuclear factor-erythroid 2-related factor (Nrf) 2 knockout mice, and age-matched wild-type (WT) control mice, in order to induce type 2 diabetes. This was followed by zinc treatment at 5 mg/kg body weight given every other day for 3 months. Global metabolic disorders of both glucose and lipids were unaffected by zinc supplementation. This induced preventive effects on conditions caused by type 2 diabetes like oxidative stress, apoptosis, the subsequent hepatic inflammatory response, fibrosis, hypertrophy, and hepatic dysfunction. Additionally, we also observed that type 2 diabetes reduced hepatic MT expression, while zinc supplementation induced hepatic MT expression. This is a crucial antioxidant. A mechanistic study showed that MT deficiency blocked zinc supplementation-induced hepatic protection under the condition of type 2 diabetes. This suggested that endogenous MT is involved in the hepatic protection of zinc supplementation in type 2 diabetic mice. Furthermore, zinc supplementation-induced hepatic MT increase was unobserved once Nrf2 was deficient, indicating that Nrf2 mediated the upregulation of hepatic MT in response to zinc supplementation. Results of this study indicated that zinc supplementation prevented type 2 diabetes-induced liver injury through the activation of the Nrf2-MT-mediated antioxidative pathway.

Topics: Animals; Diabetes Complications; Diabetes Mellitus, Experimental; Diabetes Mellitus, Type 2; Dietary Supplements; Endoplasmic Reticulum Stress; Lipid Metabolism; Liver Diseases; Male; Metallothionein; Mice; NF-E2-Related Factor 2; Oxidative Stress; Streptozocin; Zinc

Background The aim of this study was to investigate the effects of selenium, zinc, insulin, and metallothionein on oxidative damage and metallothionein (MT) gene expression levels in streptozotocin (STZ)-induced type 1 diabetic rats exposed to Cd. Methods Rats were categorized under eight groups (control, STZ, Cd, STZ + Cd, Group 5, Group 6, Group 7, and STZ + Cd + MT [n:8/group]) were used. After diabetes was induced by STZ (55 mg/kg, i.p.), Cd was administered (1 mg/kg CdCl, orally) for 4 weeks. In cadmium-treated groups selenium (Na2SeO3 1.5 mg/kg, i.p.), zinc (ZnSO4 10 mg/kg via oral gavage), insulin (insulin glargine, 2U/day, s.c.), and MT (1mg/kg, every other 10 days, s.c.) were administered. MT gene expression levels, MDA levels, GPx, SOD, and CAT activity levels were determined in liver and kidney tissues. Results MT gene expression and MDA levels increased (p < 0.05) while GPx and SOD activity levels decreased (p < 0.05) in STZ, Cd, and STZ + Cd groups. In Group 5, Group 6, Group 7, and Group 8 groups MT gene expression and MDA levels were decreased while GPx and SOD activity levels were increased (p < 0.05). CAT activity significantly increased (p < 0.05) in STZ + Cd group while there were no significance in other groups (p > 0.05). Compared to the control, Group 5, Group 6, Group 7, and Group 8 groups provided no difference for alanine aminotransferase, aspartate aminotransferase, blood urea nitrogen and creatinine levels (p > 0.05). Conclusions Our results suggest that Se, insulin, Zn and MT may have protective effects against hepatotoxicity and nephrotoxicity caused by Cd exposure in diabetic rats by reducing oxidative stress and MT gene expression levels.

Topics: Animals; Cadmium; Diabetes Mellitus, Experimental; Diabetes Mellitus, Type 1; Gene Expression Regulation; Insulin; Kidney Diseases; Liver Diseases; Male; Metallothionein; Oxidative Stress; Rats; Rats, Wistar; Selenium; Streptozocin; Zinc

The chronic form of primary hepatitis occurs commonly in dogs, and the etiology is rarely found. Metallothionein (MT) is a heavy metal-binding protein found in many organs, including the liver. MT was recently shown to enhance liver regeneration and decrease hepatic fibrosis in human beings. This study examined the expression of MT in 24 cases of chronic hepatitis in dogs using immunohistochemistry. To understand the role of MT as a determinant of hepatic inflammation, fibrosis, bile duct proliferation, and regeneration, we correlated its expression with histologic lesions of chronic hepatitis, such as hepatic inflammation, fibrosis, and bile duct proliferation, as well as hepatocellular growth fraction as measured by Ki67 immunolabeling. Hepatocellular growth fraction was used as a measure of hepatic regeneration. Regression analysis revealed a significant positive correlation between MT labeling intensity and growth fraction (r(2) = 0.29, P < .05). The percentage of MT-positive cells and the overall MT expression were both positively correlated with growth fraction (r(2) = 0.25 and 0.26, respectively; P < .05). A negative correlation was found between the overall MT labeling and fibrosis (r(2) = 0.18, P < .05). A similar trend of negative correlation was also found between the percentage of MT-positive cells and fibrosis, but the P value was not statistically significant (r(2) = 0.14, P = .0684). These findings suggest a protective role of MT in dogs affected by chronic hepatitis, similar to its role in human beings. These dogs may respond to treatment modules focusing on enhancing the expression of MT.

Topics: Animals; Bile Ducts; Cell Proliferation; Dogs; Gene Expression Regulation; Hepatitis, Chronic; Immunohistochemistry; Inflammation; Ki-67 Antigen; Liver; Liver Cirrhosis; Liver Diseases; Liver Regeneration; Metallothionein

This study was aimed to examine circadian variations of hepatic antioxidant components, including the Nrf2- pathway, the glutathione (GSH) system, antioxidant enzymes and metallothionein in mouse liver.. Adult mice were housed in light- and temperature-controlled facilities for 2 weeks, and livers were collected every 4 h during the 24 h period. Total RNA was isolated, purified, and subjected to real-time RT-PCR analysis. Hepatic mRNA levels of Nrf2, Keap1, Nqo1 and Gclc were higher in the light-phase than the dark-phase, and were female-predominant. Hepatic GSH presented marked circadian fluctuations, along with glutathione S-transferases (GST-α1, GST-µ, GST-π) and glutathione peroxidase (GPx1). The expressions of GPx1, GST-µ and GST-π mRNA were also higher in females. Antioxidant enzymes Cu/Zn superoxide dismutase (Sod1), catalase (CAT), cyclooxygenase-2 (Cox-2) and heme oxygenase-1 (Ho-1) showed circadian rhythms, with higher expressions of Cox-2 and CAT in females. Metallothionein, a small non-enzymatic antioxidant protein, showed dramatic circadian variation in males, but higher expression in females. The circadian variations of the clock gene Brain and Muscle Arnt-like Protein-1(Bmal1), albumin site D-binding protein (Dbp), nuclear receptor Rev-Erbα (Nr1d1), period protein (Per1 and Per2) and cryptochrome 1(Cry1) were in agreement with the literature. Furthermore, acetaminophen hepatotoxicity is more severe when administered in the afternoon when hepatic GSH was lowest.. Circadian variations and gender differences in transcript levels of antioxidant genes exist in mouse liver, which could affect body responses to oxidative stress at different times of the day.

Topics: Acetaminophen; Aging; Animals; Antioxidants; Circadian Rhythm; CLOCK Proteins; Female; Gene Expression Regulation; Glutathione; Liver; Liver Diseases; Male; Metallothionein; Mice; NF-E2-Related Factor 2; RNA, Messenger

The biochemical mechanisms of Chlorella vulgaris protection against cadmium (Cd)-induced liver toxicity were investigated in male Sprague-Dawley rats (5 weeks of age, weighing 90-110 g). Forty rats were randomly divided into one control and three groups treated with 10 ppm Cd: one Cd without Chlorella (Cd-0C), one Cd with 5% Chlorella (Cd-5C), and one Cd with 10% Chlorella (Cd-10C) groups. The rats had free access to water and diet for 8 weeks. Body weight gain and relative liver weight were significantly lower in the Cd-0C group than in Cd-5C and Cd-10C groups. Rats in the Cd-0C group had significantly higher hepatic concentrations of Cd and metallothioneins (MTs) than in the Cd-5C or Cd-10C group. The hepatic MT I/II mRNA was expressed in all experimental rats. MT II was more expressed in the Cd-5C and Cd-10C groups than in the Cd-0C group. Morphologically, a higher level of congestion and vacuolation was observed in the livers of the Cd-0C group compared to those of the Cd-5C and Cd-10C groups. Therefore, this study suggests that C. vulgaris has a protective effect against Cd-induced liver damage by reducing Cd accumulation and stimulating the expression of MT II in liver. However, the details of the mechanism of C. vulgaris on liver toxicity remains to be clarified by further studies.

Topics: Animals; Body Weight; Cadmium; Cadmium Poisoning; Chemical and Drug Induced Liver Injury; Chlorella vulgaris; Dietary Supplements; Hepatocytes; Hyperemia; Liver; Liver Diseases; Male; Metallothionein; Organ Size; Powders; Rats

Topics: Animals; Blood Coagulation; Chemical and Drug Induced Liver Injury; Disease Models, Animal; Disseminated Intravascular Coagulation; Fibrinogen; Galactosamine; Lipopolysaccharides; Liver; Liver Diseases; Metallothionein; Mice; Mice, Inbred C57BL; Mice, Knockout; Platelet Count; Time Factors

Cadmium (Cd), a widely distributed toxic trace metal, has been shown to accumulate in liver after long- and short-term exposure. Cd (2 mg/kg/day CdCl2) was intraperitoneally given to rats for eight days. Vitamin C (250 mg/kg/day) + vitamin E (250 mg/kg/day) + sodium selenate (0.25 mg/kg/day) were given to rats by oral means. The animals were treated by anti-oxidants one hour prior to treatment with Cd every day. The degenerative changes were observed in the groups given only Cd and anti-oxidants + Cd. Metallothionein (MT) immunoreactivity increased in cytoplasm of hepatocytes of the rats given Cd when compared with controls. In a number of cells with Cd and anti-oxidants treatment, immunoreactivity increase was more than in the group given Cd only and nuclear MT expression was also detected. Cell proliferation was assessed with proliferating cell nuclear antigen (PCNA) immunohistochemistry. PCNA expressions increased in all groups more than in the controls. Anti-oxidants treatment increased cell proliferation. In the animals administered with Cd, an increase in serum aspartate (AST) and alanine (ALT) aminotransferases, liver glutathione (GSH) and lipid peroxidation (LPO) levels were observed. On the other hand, in the rats treated with anti-oxidants and Cd, serum AST and ALT, liver glutathione and LPO levels decreased. As a result, these results suggest that combined anti-oxidants treatment might be useful in protection of liver against Cd toxicity.

Topics: Animals; Antioxidants; Ascorbic Acid; Cadmium; Cadmium Poisoning; Chemical and Drug Induced Liver Injury; Hepatocytes; Lipid Peroxidation; Liver Diseases; Male; Metallothionein; Proliferating Cell Nuclear Antigen; Rats; Rats, Sprague-Dawley; Selenic Acid; Selenium Compounds; Vitamin E

It is desirable to diagnose hepatocellular carcinoma (HCC) in the early stages during its development since its treatment is usually difficult. We previously proposed a new diagnostic method that made use of the total metallothionein (MT), zinc (Zn), and copper (Cu) concentrations in the liver of the HCC patients. We recently found that MT-1 is involved in the metabolism or detoxification of toxic metals, such as cadmium; on the other hand, MT-2 is responsible for the homeostasis of essential metals such as copper, in experimental models such as Long Evans Cinnamon (LEC) rats. In order to device a better diagnostic method than the one we proposed previously, in this study, we newly propose an improved method that includes the discriminative determination data regarding the MT isomers, namely, MT-1 and MT-2, in the liver of patients with or without HCC as compared with the total MT level. The total MT and Zn concentrations in the HCC patients were confirmed to be significantly lower than those in patients without hepatic disorders (Ctrl). In contrast, Cu concentrations of the HCC patients were higher than those of the Ctrl patients. In addition, in the juxta-tumor portion with HCC, MT-1 concentrations were significantly higher than those of MT-2. In contrast, the MT-1 concentrations in the tumor portion were significantly lower than that in the juxta-tumor portion. In addition, MT-1/MT-2 ratio in the tumor portion was significantly lower than that of the juxta-tumor portion. By using parameters such as concentrations of Cu, Zn, total MT, and MT isomers, we performed the multivariate discriminative analysis (MDA). The results suggest that the concentrations of MT isomers change depending on the progress of the tumor, and information on MT isomers and trace elements is very useful in determining the stage of the chronic hepatic disorder.

Topics: Adult; Aged; Aged, 80 and over; Animals; Carcinoma, Hepatocellular; Chronic Disease; Cytosol; Electrophoresis, Capillary; Female; Humans; Indicators and Reagents; Isomerism; Liver; Liver Diseases; Liver Neoplasms; Male; Mass Spectrometry; Metallothionein; Metals; Middle Aged; Multivariate Analysis; Rats; Rats, Long-Evans

Cyclosporine A (CsA) is the most widely used immunosuppressive drug for preventing graft rejection and autoimmune disease. However, the therapeutic treatment induces several side effects such as nephrotoxicity, cardiotoxicity, hypertension and hepatotoxicity. Among possible mechanisms of CsA-induced hepatic damage, oxidative stress has been suggested. Melatonin (Mel) has been successfully used as a potent antioxidant against many pathophysiological states. This experimental study was performed to test, during CsA treatment, the alterations of some heat shock proteins (HSP) and the Mel antioxidant properties against CsA-induced injury. Rats were divided into four groups, which were treated respectively with olive oil, Mel alone, CsA and CsA plus Mel for 30 days. At the end of the treatments, the animals were killed and hepatic tissue was treated for morphological (haematoxylin-eosin), biochemical (reduced glutathione, GSH and malondialdehyde, MDA) and immunohistochemical (HSP60, HSP72, GRP75 and MT) analyses. The results indicate that CsA-induced hepatotoxicity was characterised by morphological alterations in tissue architecture, changes in GSH and MDA levels and increase in stress protein expression. In conclusion, our data suggest that the imbalance between production of free oxygen radicals and antioxidant defence systems, due to CsA administration, is a mechanism responsible for oxidative stress. Moreover, we show that Mel plays a protective action against CsA-induced oxidative stress, as supported by biochemical and immunohistochemical results.

Topics: Animals; Antioxidants; Chaperonin 60; Chemical and Drug Induced Liver Injury; Cyclosporine; Glutathione; Heat-Shock Proteins; HSP70 Heat-Shock Proteins; HSP72 Heat-Shock Proteins; Immunosuppressive Agents; Liver; Liver Diseases; Male; Malondialdehyde; Melatonin; Membrane Proteins; Metallothionein; Oxidative Stress; Rats; Rats, Wistar

Metallothionein (MT) is induced in the liver not only by heavy metals, but also by stress such as starvation. However, the meaning of the induced MT during starvation has never been clear. In this study, we investigated the relationship between changes in hepatic MT synthesis and the hepatic damage that occurs during starvation. MT synthesis was assessed by measuring MT contents and the expression of the MT gene in the liver. The hepatic damage was assessed by measuring glutamic pyruvic transaminase (GPT) and glutamic oxaloacetic transaminase (GOT) activities in the serum. MT synthesis in the liver increased over the normal level by starvation, but decreased under the normal level by refeeding after starvation. Both GPT and GOT activities of the refeeding group were higher than those of the control group. However, MT synthesis increased by a subcutaneous injection with CdCl(2) (1 mg Cd /kg) at the same time as refeeding after starvation. At this point, GOT activity decreased until it reached the normal level. MT synthesis decreased by refeeding after starvation, and from the results found in this study, we proposed the hypothesis that the liver damage caused by refeeding after starvation might be due to the decrease in the synthesis of a sufficient amount of MT induced by metals.

Topics: Alanine Transaminase; Animals; Aspartate Aminotransferases; Cadmium Chloride; Food; Gene Expression Regulation; Injections, Subcutaneous; Liver; Liver Diseases; Male; Metallothionein; Mice; Mice, Inbred Strains; RNA, Messenger; Spain; Starvation; Time Factors

Metallothionein and zinc have been implicated in cellular defense against a number of cytotoxic agents. With respect to the free radical-generating hepatotoxin carbon tetrachloride, conclusions about a defensive role were reached from in vitro studies, in vivo studies using inducers of metallothionein and studies using injections of pharmacological amounts of zinc. Metallothionein knockout (null) and metallothionein transgenic mice are more direct models to examine the effects of metallothionein expression on induced cytotoxicity. Similarly, zinc presented via the diet is a more physiological model than that presented via injection. We examined whether metallothionein-overexpressing mice or metallothionein knockout mice had altered sensitivity to carbon tetrachloride and whether supplemental dietary zinc reduced sensitivity to carbon tetrachloride in these genotypes. Metallothionein knockout mice produced no metallothionein and were unable to sequester additional hepatic zinc in response to elevated dietary zinc. Hepatotoxicity, as measured by serum alanine aminotransferase activity, histological analyses and hepatic thiol levels, was greater in the knockout mice than in controls 12 h after carbon tetrachloride treatment but not at later time points (up to 48 h). In contrast, metallothionein-overexpressing mice produced more metallothionein and sequestered more liver zinc than control mice, but hepatotoxicity was similar between genotypes. Supplemental dietary zinc had no effect on hepatotoxicity with either genotype. These data suggest metallothionein null mice were more susceptible to carbon tetrachloride-induced hepatotoxicity than were control mice. However, neither metallothionein overexpression nor supplemental dietary zinc provided further protection.

Topics: Alanine Transaminase; Animals; Carbon Tetrachloride; Chemical and Drug Induced Liver Injury; Dietary Supplements; Female; Gene Expression Regulation; Liver; Liver Diseases; Male; Metallothionein; Mice; Mice, Inbred C57BL; Mice, Knockout; Mice, Transgenic; Microscopy; Oxidative Stress; Time Factors; Zinc

Inhibins and activins are dimeric proteins belonging to the transforming growth factor-beta superfamily. Follistatin is an activin-binding protein that antagonizes the function of activin via binding to its beta-subunits. Previously, we demonstrated that mice deficient in inhibin develop ovarian and testicular sex cord-stromal tumors of granulosa and Sertoli cell origin, with 100% penetrance as early as 4 weeks of age. Overproduction of activins in the serum directly causes a cachexia-like wasting syndrome that results in lethality of these mice at an early stage after the onset of the tumors. In an independent set of studies, overexpression of mouse follistatin using the mouse metallothionein I promoter in transgenic mice led to gonadal defects and eventual infertility, primarily due to local effects of follistatin in these tissues. Activin has a positive growth effect on gonadal tumor cells in culture and directly causes the cancer cachexia-like syndrome in inhibin-deficient mice via interaction with activin receptor type IIA in livers and stomachs. We therefore hypothesized that an activin antagonist such as follistatin can act as a physiological modifier, either locally or via the serum, to block the activin-mediated cancer cachexia-like syndrome in inhibin-deficient mice and/or slow the progression of gonadal cancers in these mice. To test this hypothesis, we generated mice that are homozygous mutant for the inhibin alpha null allele (i.e. inham1/inham1) and carry the mouse metallothionein I follistatin (MT-FS) transgene. Our results show that gonadal tumors that are histologically similar in most, but not all, cases to the tumors in inhibin-deficient mice develop in these inham1/inham1, MT-FS+ mice. However, inham1/inham1, MT-FS+ mice exhibit a less severe wasting syndrome, lower serum activin levels, and a statistically significant prolonged survival in a number of cases compared with mice deficient in inhibin alone. Thus, follistatin can act as a modulator of tumor growth and the activin-induced cancer cachexia-like syndrome in inhibin-deficient mice.

Topics: Activins; Animals; Disease Progression; Female; Follistatin; Glycoproteins; Inhibins; Liver Diseases; Male; Metallothionein; Mice; Mice, Transgenic; Ovarian Neoplasms; Peptides; Stomach Diseases; Survival Analysis; Testicular Neoplasms; Wasting Syndrome

The objective of this study was to correlate hepatic and renal cadmium (Cd) accumulation, Cd-binding capacity of metallothionein (MT) and lipid peroxidation with the tissue injury in the male bank voles raised under short (8 h light/16 h dark) and long (16 h light/8 h dark) photoperiods that affect differently Cd accumulation and MT induction in these rodents. The animals were exposed to dietary Cd (0, 40 and 80 microg/g) for 6 weeks. The accumulation of Cd in the liver and kidneys appeared to be dose-dependent in bank voles from the two photoperiod groups; however, the short-photoperiod animals exhibited significantly higher concentrations of Cd in both organs than the long-photoperiod bank voles. Cd-Binding capacity of MT in the liver and kidneys of bank voles from the long photoperiod was sufficiently high to bind and detoxify all Cd ions, while in the animals fed 80 microg Cd/g under the short photoperiod, the concentrations of Cd in both organs exceeded (by about 10 microg/g) the MT capacity. However, similar histopathological changes in the liver (a focal hepatocyte swelling and granuloma) and kidneys (a focal degeneration of proximal tubules) occurred in Cd-80 bank voles from the two photoperiods. Likewise, in either photoperiod group, dietary Cd brought about a similar, dose-dependent decrease in the hepatic and renal lipid peroxidation, which paralleled closely that of the iron (Fe) concentrations. These data indicate that: (1) MT does not protect the liver and kidneys against Cd-induced injury in the bank vole exposed to the higher level of dietary Cd; and (2) lipid peroxidation cannot be responsible for the tissue damage. It is hypothesized that dietary Cd produces histopathological changes indirectly, through depressing the tissue Fe and Fe-dependent oxidative processes.

Topics: Animals; Arvicolinae; Body Weight; Cadmium; Chemical and Drug Induced Liver Injury; Copper; Diet; Iron; Kidney; Kidney Diseases; Light; Lipid Peroxidation; Liver; Liver Diseases; Male; Metallothionein; Organ Size; Photoperiod; Zinc

The influence of aging on the sensitivity of the liver to the acute toxicity of cadmium has not been studied previously in adult rats. In this study hepatotoxicity caused by a single sc injection of CdCl(2) was compared in 5-, 18-, and 28-month-old male Fischer 344 rats. Doses of Cd were adjusted on the basis of the mean lean body mass for each age group of rats, and liver injury was evaluated 24 h after treatment. Cd treatment produced substantial increases in serum alanine aminotransferase (ALT) and sorbitol dehydrogenase (SDH) activities in 5- and 18-month-old rats, whereas no significant increases were observed in 28-month-old rats. Histologic examination of representative livers from each age group confirmed the findings for serum enzyme activity; hepatocellular necrosis was observed only in livers from 5- and 18-month-old rats. The attenuation of Cd hepatotoxicity in senescent rats did not appear to be related to pretreatment levels of metallothionein or glutathione. Likewise, resistance to Cd could not be explained on the basis of metallothionein induction, which decreased as a function of aging. Thus, the mechanisms that account for the postmaturational decline in sensitivity to Cd do not appear to be associated with alterations in levels of the major factors that protect against Cd-induced hepatotoxicity.

Topics: Aging; Alanine Transaminase; Animals; Cadmium; Chemical and Drug Induced Liver Injury; Glutathione; L-Iditol 2-Dehydrogenase; Liver; Liver Diseases; Male; Metallothionein; Rats; Rats, Inbred F344

cis-Diamminedichloroplatinum (cisplatin) is an important anticancer drug used to treat solid tumors. The nephrotoxicity of cisplatin is recognized as the most important dose-limiting factor, but high doses of cisplatin also produce hepatotoxicity. However, little is known about cisplatin-induced liver injury and the role of metallothionein, a cysteine-rich, metal-binding protein, in modulating its hepatotoxicity. This study was designed to examine cisplatin hepatotoxicity in control and metallothionein-I/II knockout (MT-null) mice. Animals were given a single injection of cisplatin (50-200 mumol/kg i.p.), and liver injury was evaluated 3-16 h later. Cisplatin produced dose- and time-dependent liver injury, as evidenced by increased serum activity of alanine aminotransferase (ALT), as well as by histopathology. Apoptosis, rather than necrosis, predominates in cisplatin-induced liver injury, as indicated by increased numbers of apoptotic cells (hematoxylin and eosin staining), in situ apoptotic DNA detection, and DNA fragmentation on agarose gel electrophoresis. MT-null mice were more sensitive than controls to cisplatin-induced hepatotoxicity. Cisplatin (200 mumol/kg) was lethal to 12% of control mice, but 60% of MT-null mice died within 16 h. At the dose of 150 mumol/kg, serum ALT activities were increased 2-fold in control mice compared to 6.5-fold in MT-null mice. Apoptotic lesions were more pronounced in MT-null than in control mice. MT-null mice were also more susceptible than controls to cisplatin-induced nephrotoxicity, as evidenced by having higher blood urea nitrogen concentrations. Furthermore, cultured MT-null hepatocytes were more sensitive than control cells to the cytotoxicity of cisplatin (50-200 microM), as indicated by lactate dehydrogenase leakage into the medium. These results demonstrate that (1) high doses of cisplatin produce hepatotoxicity, with apoptosis as the major lesion, and (2) MT protects against cisplatin-induced liver injury.

Topics: Animals; Antineoplastic Agents; Apoptosis; Cadmium; Cells, Cultured; Chemical and Drug Induced Liver Injury; Cisplatin; Liver; Liver Diseases; Metallothionein; Mice; Mice, Knockout

The hepatotoxicity of cadmium was studied in 1-, 2-, and 6-month-old male Wistar rats. Liver damage, indicated by the increase in serum alanine aminotransferase activity 24 h after sc administration of 3 and 6 mg/kg cadmium, was observed only in 6-month-old rats. Dose-dependent increases in the cadmium content of the liver were similar for all three age groups. Basal and induced metallothionein contents were higher in livers of 1-month-old rats than in those of 2- and 6-month-old rats. In contrast, the basal glutathione content of the liver was higher in 6-month-old rats than in 1- and 2-month-old rats, and glutathione content increased slightly in all three age groups after cadmium administration. Thus, the higher susceptibility to cadmium-induced hepatotoxicity in 6-month-old rats seemed not to be explained by differences in cadmium uptake or by the metallothionein and glutathione contents of the liver. Inactivation of Kupffer cells with gadolinium chloride or depletion of neutrophils with cyclophosphamide relieved cadmium hepatotoxicity only in 6-month-old rats. In addition, 6-month-old rats were more susceptible than 2-month-old rats to lipopolysaccharide-induced hepatotoxicity. The results suggest that age-associated changes in Kupffer cell function and infiltration of neutrophils are important determinants of cadmium-induced hepatotoxicity in rats.

Topics: Age Factors; Alanine Transaminase; Alkylating Agents; Animals; Anti-Inflammatory Agents; Cadmium; Chemical and Drug Induced Liver Injury; Cyclophosphamide; Dose-Response Relationship, Drug; Escherichia coli; Gadolinium; Glutathione; Kupffer Cells; Lipopolysaccharides; Liver Diseases; Male; Metallothionein; Neutrophils; Rats; Rats, Wistar

Previous work has indicated that testosterone pretreatment protects against cadmium-induced toxicity in male rats while other data indicate that pretreatment of mice with testosterone offers no such protection against cadmium. Since cadmium toxicity may vary widely with species and strain, we examined the effect of testosterone pretreatment on cadmium toxicity in two strains of mice, one that is sensitive (C3H) and one that is resistant (C57) to cadmium toxicity. A single sc injection of 20 micromol CdCl2/kg to C3H mice or 45 micromol CdCl2/kg to C57 mice proved very toxic, causing 50%, and 44% mortalities, respectively. However, when C57 mice were pretreated with testosterone (5 mg/kg, s.c., at - 48, - 24, and 0 h) prior to cadmium (45 micromol/kg), complete resistance to cadmium-induced lethality developed. Testosterone had no effect on cadmium-induced lethality in C3H mice. Testosterone prevented extensive hepatocellular damage caused by cadmium in C57 mice and also significantly reduced cadmium-induced elevations in serum lactate dehydrogenase (LDH) activity and blood urea nitrogen (BUN), which are indicators of hepatic and renal function, respectively. The toxicokinetics of cadmium were apparently not affected by testosterone pretreatment, as the distribution of cadmium to liver in either strain was unchanged by the steroid. Cadmium-induced metallothionein (MT) levels in liver and kidney of C57 mice were increased in testosterone-pretreated mice given the higher doses of metal but no such enhancement of MT synthesis occurred in C3H mice. This increase in MT may provide some level of protection against cadmium toxicity in the C57 mice. These results indicate that testosterone pretreatment prevents toxicity of cadmium in male C57 mice, possibly through enhancement of MT synthesis, but has no effect in male C3H mice.

Topics: Animals; Antidotes; Cadmium; Cadmium Poisoning; Chemical and Drug Induced Liver Injury; Kidney; L-Lactate Dehydrogenase; Liver; Liver Diseases; Male; Metallothionein; Mice; Mice, Inbred C3H; Mice, Inbred C57BL; Species Specificity; Testosterone; Urea

Recently, it has been shown that large doses of all-trans-retinol (vitamin A) can potentiate the hepatotoxicity of several organic chemicals in the rat. Whether retinol pretreatment can alter the acute hepatotoxicity of an inorganic chemical, such as cadmium, is unknown. Therefore, the objective of this study was to determine how retinol might affect the acute toxicity of cadmium chloride (CdCl2) and to elucidate possible mechanisms. Cadmium exposure can induce acute, lethal hepatocellular necrosis in rodents, as well as lesions in the lung, kidney, testis, and gastrointestinal tract. In the present studies, male Sprague-Dawley rats were pretreated with retinol (75 mg/kg/day, po) for 7 consecutive days. One day after the last dose of retinol, animals were given a single injection of CdCl2 (2.5 to 4.0 mg/kg, iv). Cadmium chloride administration to unpretreated control rats caused extensive hepatic, renal, pulmonary, and testicular toxicity at 6, 24, and 48 hr postdosing as evaluated by plasma enzymes and/or histopathology. In retinol-pretreated rats, a significant attenuation of CdCl2-induced tissue injury was observed. Since the inducible cadmium-binding protein metallothionein (MT) is often an essential aspect of cadmium tolerance, its content in tissue was assessed using the cadmium-hemoglobin assay. Interestingly, retinol pretreatment significantly increased MT in the liver by sevenfold, but had no effect on lung, kidney, testicular, or pancreatic MT content. Although this increase in hepatic MT was much less than that induced by CdCl2, it was additive to the induction of CdCl2. Furthermore, the tissue distribution of cadmium was significantly altered by retinol pretreatment. The liver accumulated more cadmium, while less cadmium was found in the lung, kidney, and testis in retinol-pretreated rats than in controls. In monolayers of primary isolated hepatocytes, CdCl2-induced toxicity was significantly reduced in cells isolated from retinol-pretreated rats compared to those isolated from control rats. The dose response was shifted to the right and the in vitro cadmium LC50 was increased by in vivo retinol exposure from 1.1 +/- 0.1 to 2.4 +/- 0.04 microM. From these data it is concluded that the induction of hepatic MT is an essential aspect of retinol-induced tolerance to CdCl2 hepatotoxicity, as well as toxicity in other tissues.

Topics: Animals; Cadmium; Cadmium Chloride; Cells, Cultured; Chemical and Drug Induced Liver Injury; Drug Administration Schedule; Drug Tolerance; Injections, Intravenous; L-Lactate Dehydrogenase; Liver; Liver Diseases; Male; Metallothionein; Rats; Rats, Sprague-Dawley; Testis; Tissue Distribution; Vitamin A

Exposure of humans to toxic metals and metalloids is a major environmental problem. Many metals, such as cadmium, can be hepatotoxic. However, the mechanisms by which metals cause acute hepatic injury are in many cases unknown. Previous reports suggest a major role for inflammation in acute cadmium induced hepatotoxicity. In initial experiments we found that a non-hepatotoxic dose of cadmium chloride (CdCl2; 2.0 mg/kg, i.v.) markedly increased the clearance rate of colloidal carbon from the blood, which is indicative of enhanced phagocytic activity by Kupffer cells (resident hepatic macrophages). Thus. the objective these studies was to determine the involvement of Kupffer cells in cadmium induced liver injury by inhibiting their function with gadolinium chloride (GdCl3). Male Sprague-Dawley rats were administered GdCl3 (10 mg/kg, i.v.) followed 24 h later by a single dose of CdCl2 (3.0 and 4.0 mg/kg, i.v.). Twenty four hours after CdCl2 administration animals were killed and the degree of liver toxicity was assessed using plasma alanine aminotransferase (ALT), as well as light microscopy. Cadmium chloride administration produced multifocal hepatocellular necrosis and increased plasma ALT activity. Pretreatment with GdCl3 significantly reduced both the morphological changes and hepatic ALT release caused by CdCl2. However, the protection was specific to the liver, and did not alter CdCl2 induced testicular injury, as determined by histopathological damage. In many cases, the inducible cadmium-binding protein, metallothionein (MT) is often an essential aspect of the acquisition of cadmium tolerance in the liver. Although cadmium caused a dramatic induction of hepatic MT (32-fold), GdCl3 caused only a minor increase (2-fold). Combined CdCl2 and GdCl3 treatment did not induce levels to an extent greater than CdCl2 alone. As expected, GdCl3 also caused a slight increase in the amount of cadmium associated with the liver. In cultured hepatocytes isolated from GdCl3 pretreated rats, CdCl2 induced cytotoxicity was not significantly altered compared to control hepatocytes, indicating that the mechanism of tolerance required the presence of other cell types. Thus, GdCl3 attenuation of CdCl2 induced hepatotoxicity does not appear to be caused by increased tissue MT content or a decreased susceptibility of hepatocytes to cadmium. From these data, we concluded that tolerance to cadmium induced hepatotoxicity involves the inhibition of Kupffer cell function which resu

Topics: Alanine Transaminase; Animals; Cadmium Chloride; Carcinogens; Cells, Cultured; Chemical and Drug Induced Liver Injury; Disease Models, Animal; Dose-Response Relationship, Drug; Gadolinium; Gout Suppressants; Injections, Intravenous; Kupffer Cells; Lethal Dose 50; Liver; Liver Diseases; Male; Metallothionein; Necrosis; Phagocytosis; Rats; Rats, Sprague-Dawley; Testis; Tissue Distribution

To date, numerous correlative studies have implicated metallothionein in the detoxification of heavy metals and in the regulation of metal distribution within an organism. In the present study cadmium-binding proteins (metallothionein equivalents), cadmium acute toxicity, and cadmium distribution in tissues and subcellular fractions were compared in metallothionein-I and -II deficient (MT-/-) mice and the parental strain carrying intact metallothionein genes (MT+/+) to determine if the absence of metallothionein altered any of these parameters. In an uninduced state, MT-/- mice expressed lower levels of cadmium-binding proteins relative to MT+/+ mice in several tissues. Administration of zinc enhanced the levels of cadmium-binding proteins in liver, small intestine, kidney, pancreas, and male sex organs, but not in cecum or brain of MT+/+ mice compared to zinc pretreated MT-/- mice. The cadmium LD50 was similar for MT-/-, MT+/+, and zinc-pretreated MT-/- mice (15-17 mumol CdCl2/kg body weight delivered i.p.). However, zinc-pretreated MT+/+ mice had a cadmium LD50 of 58-63 mumol CdCl2/kg body weight. Over two-thirds of cadmium was found in liver, cecum, small intestine, and kidney in both MT+/+ and MT-/- mice; therefore, metallothionein levels do not appear to play a major role in the tissue distribution of cadmium. However, after zinc pretreatment, MT+/+ mice accumulated more cadmium in the liver and less in other tissues, whereas the amount of cadmium in the liver was not altered by zinc pretreatment in MT-/- mice. In general, the cytosolic/particulate ratio of cadmium was significantly higher in tissues of noninduced MT+/+ mice relative to MT-/- mice. This difference was accentuated after zinc pretreatment. Together these results indicate that basal levels of metallothionein do not protect from the acute toxicity of a single i.p. cadmium challenge. Furthermore, it does not appear that the cytosolic compartmentalization of cadmium is correlated with reduced toxicity.

Topics: Animals; Cadmium; Chemical and Drug Induced Liver Injury; Female; Lethal Dose 50; Liver; Liver Diseases; Male; Metallothionein; Mice; Mice, Transgenic; Oxidoreductases; Subcellular Fractions; Tissue Distribution; Zinc

To explore the hypothesis that a second xenobiotic agent is required with excess copper to produce Indian Childhood Cirrhosis, this study investigated the effect of the pyrrolizidine alkaloid retrorsine fed to the mother during the suckling period upon the serial changes in neonatal copper status.. Female Wistar rats with new-born litters were fed either a control or a retrorsine (50 mg/kg) diet. At 0, 4, 8, 11, 15, 18 and 21 days, pups from each litter were weighed, sacrificed and their livers removed for copper, DNA and metallothionein analysis. Serum samples were assayed for caeruloplasmin oxidase activity and albumin.. 1) Higher than adult level of hepatic copper in normal rats which rose post-natally before declining from day 11 after birth, 2) raised hepatic copper concentrations and total copper in the retrorsine group from day 15; levels were higher than adult at birth, 3) reduced serum caeruloplasmin oxidase activity and albumin levels in retrorsine group, but both groups lower than adult, 4) lower hepatic metallothionein levels in retrorsine group, but both groups higher than adult, and 5) reduced liver DNA in the retrorsine group when expressed as total DNA and per gram of tissue. These changes were not secondary to under-nutrition as a small study on under-nourished rat neonates showed that copper handling is not significantly altered when compared to well-nourished rats.. Retrorsine passing to rat neonates via breast milk causes: 1) the accumulation of hepatic copper, 2) impairment of the rise in serum caeruloplasmin, which could indicate a decline in synthesis or failure of copper incorporation into the apo-protein, 3) a decrease in hepatic metallothionein and serum albumin levels, again suggesting diminished protein synthesis, and 4) reduced hepatic DNA indicative of decreased cell number but increased cell size. Accumulation of liver copper but reduction of copper-binding proteins could result in free copper and explain the synergistic hepatotoxicity of copper and retrorsine.

Topics: Animals; Animals, Suckling; Body Weight; Ceruloplasmin; Chemical and Drug Induced Liver Injury; Copper; DNA; Drug Evaluation, Preclinical; Female; Liver; Liver Diseases; Metallothionein; Milk; Nutrition Disorders; Organ Size; Pyrrolizidine Alkaloids; Rats; Rats, Wistar; Serum Albumin

The LEC rat spontaneously develops liver cancer after suffering chronic liver injury caused by abnormal copper accumulation in the liver, but the role of copper accumulation in the induction of liver cancer remains obscure. We histochemically and biochemically examined the content of copper and metallothionein (MT), a cytoplasmic copper binding protein, in spontaneously developed preneoplastic and neoplastic liver lesions and compared them with those in the surrounding liver tissues. Histochemically, the majority of the preneoplastic liver lesions (68%) and liver cancers (59%) showed lower copper contents than the surrounding liver tissues and no lesions were shown to accumulate more copper than the surrounding tissues. A marked heterogeneity in copper staining was observed in cancer tissues. In contrast, these lesions showed an equal to higher MT content than their surroundings. Biochemical measurements of copper and MT in cancer tissues supported the histochemical findings. The bromodeoxyuridine (BrdU) labeling index was high in all cancer tissues and some of the preneoplastic liver lesions. Parts of the cancer tissues with negative or weak staining for copper were highly labeled with BrdU. Taking these results together, copper accumulation may exert a growth inhibitory effect on surrounding hepatocytes, whereas the hepatocytes in the liver lesions could proliferate, escaping from the effect of copper toxicity by increasing their MT induction and lowering copper accumulation. Thus, accumulation of copper may act as a promoting factor for the development of liver cancer in LEC rats by creating a selective growth environment.

Topics: Animals; Copper; Liver Diseases; Liver Neoplasms, Experimental; Male; Metallothionein; Precancerous Conditions; Rats; Rats, Mutant Strains; Staining and Labeling

Oleanolic acid (OA) is a triterpenoid compound that has been shown to protect against some hepatotoxicants and is used in China to treat hepatitis. This study was conducted to examine the protective effects of OA against cadmium (Cd)-induced liver injury in mice and the mechanism of protection. OA (100 mg/kg x 3 days) pretreatment dramatically decreased Cd (3.7 mg/kg i.v.)-induced liver injury as indicated by decreased serum activities of alanine aminotransferase and sorbitol dehydrogenase, as well as by histopathological observation. To examine the mechanism of protection, the distribution of Cd to major organs and the hepatic subcellular distribution of Cd were determined 2 hr after 109Cd injection (3.5 mg/kg of Cd and 10 microCi/mg of Cd i.v.). OA did not reduce the amount of Cd in liver, but significantly altered the hepatic subcellular distribution of Cd, with more Cd in hepatic cytosol bound to metallothionein (MT), and with less Cd in other organelles and proteins. OA produced an approximately 30-fold increase in hepatic MT, but had no appreciable effects on MT levels of five other organs. Furthermore, OA increased both hepatic MT-I and MT-II levels, as determined by high-performance liquid chromatography/atomic absorption spectrophotometry. Northern blot analysis revealed that OA increases MT mRNA expression. In summary, OA pretreatment protects against Cd-induced hepatotoxicity by inducing MT. MT bound Cd in the cytosol, and thus decreased the amount of Cd in other critical organelles and proteins. OA is a hepatic MT inducer for both MT-I and MT-II isoforms, and this effect is due, at least in part, to an increased MT mRNA accumulation.

Topics: Alanine Transaminase; Animals; Blotting, Northern; Cadmium; Chemical and Drug Induced Liver Injury; Cytosol; Dose-Response Relationship, Drug; Isomerism; L-Iditol 2-Dehydrogenase; Liver; Liver Diseases; Male; Metallothionein; Mice; Mice, Inbred Strains; Oleanolic Acid; Organ Size; RNA Probes; RNA, Messenger; Subcellular Fractions; Tissue Distribution

Metallothionein (MT) is a sulfhydryl-rich protein whose levels are increased by administration of a variety of agents including metals, cytokines, and oxidative stress agents. Recent studies have suggested that MT is involved in protecting against various forms of oxidative stress, but little is known about the induction of MT by oxidative stress agents. Diethyl maleate (DEM) causes oxidative stress by depleting glutathione levels and is quite effective at increasing hepatic concentrations of MT. The purpose of the current study was to learn more about the relationship between induction of MT and oxidative stress by characterizing this increase in hepatic MT levels produced by DEM. Administration of DEM (3 to 9 mmol/kg, sc) increased hepatic MT concentration in mice as much as 37-fold to 213 micrograms MT/g liver, which is similar to the hepatic MT level seen after administration of other effective MT inducers, such as Cd. The maximal increase of hepatic MT took place 12 to 24 hr after administration of 5 mmol DEM/kg. This rise in MT was preceded by a 60% depletion of hepatic glutathione 3 hr after DEM and increases in both MT-I and MT-II mRNA, which reached a peak 6 to 9 hr after DEM. Administration of DEM (3-5 mmol/kg, sc) also increased MT levels in Sprague-Dawley rats. Pretreatment with DEM protected against Cd-induced hepatotoxicity in a fashion which suggested that a functional MT was being synthesized. In summary, DEM is a highly effective inducer of MT which increases MT at the mRNA level.

Topics: Animals; Base Sequence; Blotting, Northern; Cadmium; Chemical and Drug Induced Liver Injury; Cytosol; Dose-Response Relationship, Drug; Glutathione; Immunoblotting; Liver; Liver Diseases; Male; Maleates; Metallothionein; Mice; Mice, Inbred Strains; Molecular Sequence Data; Rats; Rats, Inbred Strains; RNA, Messenger; Subcellular Fractions; Time Factors; Transcription, Genetic

Female Wistar rats received an oral dose of 700 mg salicylic acid/kg body wt., given as sodium salicylate. Some of the salicylate-treated rats received two subcutaneous injections of 100 mumol kg-1 ZnCl2 (24 h before and simultaneously with the salicylate administration). Other animals were given one subcutaneous injection of 100 mumol kg-1 ZnCl2 simultaneously with the salicylate treatment. Control rats were similarly injected with ZnCl2. Twenty four hours after salicylate treatment, serum and livers were taken for histochemical and biochemical analysis. The most remarkable effects of the treatment were enrichment of lipid droplets and iron and a reduction of glycogen, particularly in the periportal hepatocytes. The effects of salicylate were partially prevented by two ZnCl2 injections. The protective effects of ZnCl2 may be due to lower iron uptake into hepatocytes and by the induction of zinc metallothionein, which can serve as a scavenger for oxygen radicals.

Topics: Alanine Transaminase; Alkaline Phosphatase; Animals; Cations; Chemical and Drug Induced Liver Injury; Female; Histocytochemistry; Iron; Lipids; Liver Diseases; Liver Glycogen; Metallothionein; Rats; Rats, Inbred Strains; Salicylates; Salicylic Acid; Zinc

The plasma metallothionein concentration was evaluated in healthy subjects and in patients with several types of liver disorders. Plasma metallothionein concentrations in controls varied between 2.4 and 4.8 ng/ml. Patients with disorders associated with increased liver copper concentrations (i.e., primary biliary cirrhosis and primary sclerosing cholangitis) had significantly (both p less than 0.002) elevated plasma metallothionein concentrations (range = 1.8 to 52.2 ng/ml), and a considerable number of these were above the maximum control level (21 of 41 patients). In contrast, patients with liver disorders not associated with increased liver copper concentrations (alcoholic and cryptogenic cirrhosis, and acute viral and chronic active hepatitis) generally had normal plasma metallothionein concentrations and only a few were above the maximum control level (11 of 64 patients, maximum = 8.8 ng/ml). The metallothionein concentrations in plasma samples from patients in stage I or II primary biliary cirrhosis were within or slightly above the control range, whereas most patients in stage III had elevated levels (p less than 0.002), and almost all patients in stage IV had clearly elevated (p less than 0.0001) concentrations. In primary biliary cirrhosis the plasma metallothionein concentration tended to increase during the evolution of the disorder, and the concentration correlated significantly with the serum total bilirubin concentration. In conclusion, the plasma metallothionein concentration is significantly elevated in patients with primary biliary cirrhosis and in patients with primary sclerosing cholangitis. Although related to the histological stage of primary biliary cirrhosis, the measurement of plasma metallothionein concentrations contributes little to the diagnosis or the evaluation of the severity of these disorders.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Humans; Liver Cirrhosis, Biliary; Liver Diseases; Metallothionein; Osmolar Concentration; Radioimmunoassay; Regression Analysis

Patients with Wilson's disease often have a further increase in hepatic copper when given zinc as an initial treatment, although there is no associated clinical deterioration. To better understand this situation an animal model was developed in which copper-loaded rats are treated with zinc administered subcutaneously. In the presence of equal amounts of copper loading in liver, control rats show hepatic damage but zinc-treated rats do not. Zinc-treated rats have much higher levels of hepatic metallothionein. Gel filtration studies reveal that much of the hepatic copper in zinc-treated rats is in this metallothionein fraction, whereas the copper in control animals is primarily associated with fractions of high or low molecular weight. Subcutaneous zinc therapy also induces intestinal, but not brain, metallothionein. We interpret these findings to indicate that zinc therapy protects against copper toxicity in liver by induction of hepatic metallothionein, which sequesters copper in a nontoxic form.

Topics: Alanine Transaminase; Animals; Chromatography, Gel; Copper; Cytosol; Delayed-Action Preparations; Diet; Hepatolenticular Degeneration; Liver; Liver Diseases; Male; Metallothionein; Rats; Zinc

We have examined the relationship between (a) histochemically demonstrable copper using rubeanic acid, (b) copper-associated protein (CAP) using orcein, and (c) immunoreactive metallothionein (MT) using DNP hapten sandwich staining and have correlated these with histological lesions in 95 liver biopsies from patients with conditions associated with hepatic copper retention, 4 fetal livers, and 25 histologically normal adult controls. No copper or CAP was present in normal adult liver but periportal CAP was present in fetal liver. MT was present in hepatocytes of normal livers with a predominantly perivenular (centrilobular) cytoplasmic distribution varying in staining intensity; all fetal hepatocytes stained strongly for MT. Fifty-two of 95 (55 per cent) abnormal livers contained CAP and 42 (44 per cent) contained both CAP and copper. In CAP-positive livers, the commonest histological lesions were piecemeal necrosis and cholestasis. CAP was present in (a) 15/15 cases of primary biliary cirrhosis including early cases with minimal pathology; and (b) 5/5 cases of Wilson's disease, 6/6 cases of biliary atresia, and 3/9 cases of sclerosing cholangitis. In other conditions, it was present in 25-50 per cent of cases. MT distribution was abnormal in most CAP-positive livers. Necrotic hepatocytes were intensely MT-positive and in Wilson's disease had a characteristic appearance.

Topics: Copper; Fetus; Humans; Liver; Liver Diseases; Metallothionein

Topics: Adult; Ceruloplasmin; Copper; Histocytochemistry; Humans; Immunoenzyme Techniques; Liver Diseases; Metallothionein

To study the influence of hepatic metallothionein (MT) on the hepatotoxic response to carbon tetrachloride (CCl4), adult male rats were pretreated with a 10 mg X kg-1 dose of zinc (Zn) 24 h prior to CCl4 (i.p., l mL X kg-1) treatment. Zn pretreatment increased the hepatic MT concentrations markedly and reduced the magnitudes of the CCl4-induced reduction of cytochrome P450 concentration as well as elevation of serum alanine aminotransferase and aspartate aminotransferase activities when determined at 4 or 24 h following CCl4 treatment. Treatment of Zn-exposed animals with CCl4 also resulted in significant reduction of the concentrations of hepatic MT (as determined by the cadmium-saturation method) as well as cytosolic Zn. Sephadex G-75 chromatographic study of hepatic cytosols showed that MT-bound Zn was selectively depleted by CCl4 exposure. Moreover, it was demonstrated that CCl4, after metabolic activation, reduced the cadmium binding capacity of Zn-induced hepatic MT in vitro. To examine the possible protective effect of Zn independent of induction of MT synthesis, CCl4 was administered 2 h following Zn pretreatment and the hepatotoxic response was examined 4 h later. This study revealed limited protection by Zn prior to the induction of MT synthesis. These data further support a role of MT in the modulation of CCl4 hepatotoxicity.

Topics: Animals; Cadmium; Carbon Tetrachloride Poisoning; Chemical and Drug Induced Liver Injury; Cytosol; In Vitro Techniques; Liver Diseases; Male; Metallothionein; Protein Binding; Rats; Rats, Inbred Strains; Zinc

Three different histochemical methods for copper detection were compared. Atomic absorption analysis was used to substantiate the tissue stains. There was good correlation between rhodanine staining and rubeanic acid-stained tissue sections. The orcein reaction for copper-associated protein did not consistently correlate with the methods demonstrating copper. Prolonged staining (72 hours) with rubeanic acid more consistently and clearly detected increased copper in canine livers than did staining with rhodanine. Seventy-two hour staining with rubeanic acid is the method of choice for histochemical detection of copper in canine liver.

Topics: Animals; Copper; Dog Diseases; Dogs; Female; Liver; Liver Diseases; Metallothionein; Oxazines; Rhodanine; Spectrophotometry, Atomic; Staining and Labeling; Thioamides

Rats were injected sc with 0.5 mg Cd/kg, 6 days/week, for up to 26 weeks. Hepatic and renal function and tissue Cd and metallothionein (MT) content were determined in tissues and plasma at various times after Cd injection. Cd in liver and kidney increased linearly for the first 10 weeks of treatment, but thereafter hepatic concentrations of Cd decreased by 33% whereas the content of Cd in kidney remained constant. MT in liver and kidney increased linearly during the first 12 weeks of Cd treatment to 4400 and 2300 micrograms MT/g, respectively, but rose only slightly thereafter. Circulating concentrations of MT progressively increased beginning 2 weeks after Cd treatment and were approximately 10 times control values in rats dosed with Cd for 12 or more weeks. Plasma activities of alanine and aspartate aminotransferase exhibited a time course similar to that observed with MT, and were elevated as early as the sixth week of Cd exposure. Sharp increases in activities of these enzymes also occurred after 10 to 12 weeks of dosing. Hepatic microsomal metabolism of benzo[a]pyrene and ethylmorphine was severely attenuated beginning 4 weeks after Cd. Renal injury occurred after hepatic damage, as evidenced by decreased in vitro p-aminohippuric acid uptake beginning 8 weeks after exposure. Urine outflow increased threefold 11 weeks after Cd exposure began, while urinary protein and Cd excretion increased beginning at Week 9. These data indicate the liver is a major target organ of chronic Cd poisoning, and suggest that Cd-induced hepatic injury, via release of Cd-MT, may play an important role in the nephrotoxicity observed in response to long-term exposure to Cd.

Topics: Animals; Body Weight; Cadmium; Cadmium Poisoning; Chemical and Drug Induced Liver Injury; Kidney; Kidney Diseases; Liver; Liver Diseases; Male; Metallothionein; Organ Size; Rats; Rats, Inbred Strains; Tissue Distribution

Of 1361 consecutive liver biopsy specimens, 24% contained orcein-positive granules. The highest incidence of positivity was found in biliary disease (90.9%), long before cirrhosis had developed, whereas in chronic non-primarily biliary disease, positive results were almost exclusively in patients with well established cirrhosis. Orcein-positive granules were never found in acute liver disease. These granules were also demonstrated in tumour cells of primary hepatocellular tumours (benign 4 of 4 cases; malignant 9 of 37 cases), while all the secondary tumour deposits were negative. In our view the additional information obtained by this technique warrants its adoption as a routine procedure.

Topics: Carcinoma, Hepatocellular; Coloring Agents; Cytoplasmic Granules; Humans; Liver; Liver Cirrhosis, Alcoholic; Liver Cirrhosis, Biliary; Liver Diseases; Liver Neoplasms; Male; Metalloproteins; Metallothionein; Oxazines; Staining and Labeling

In order to test the possibility of metallothionein (MT) transfer from liver to kidney, experimental hepatic disorders produced by hepatotoxins were examined to study the release of MT from liver. 109Cd exposed rats were treated with carbon tetrachloride (CCl4) and the distribution of cadmium (Cd) in the body was studied. Hepatic Cd was significantly decreased corresponding to the dose of CCl4. Cd in plasma, kidney, and urine was increased remarkably in contrast with the decrease of hepatic Cd. No remarkable changes in Cd of other tissues and feces were observed. These phenomena were produced by other hepatotoxins like galactosamine and ethionine, and long-term administration of Cd, too. In every case that plasma Cd increased markedly, plasma levels of glutamate oxaloacetate transaminase (GOT), glutamate pyruvate transaminase (GPT), and lactate dehydrogenase (LDH) rose simultaneously, and a significant, positive correlation between Cd concentration and each of enzyme activities in plasma was observed. Cd in hepatic supernatant of CCl4 treated rats was bound mostly to MT fraction, and in kidney, plasma or urine, Cd was also in the form of MT. These results suggest that hepatic MT can be released into blood in the same manner as hepatic enzymes and transported to kidney and urine in some types of hepatic disorders.

Topics: Animals; Cadmium; Cadmium Poisoning; Carbon Tetrachloride Poisoning; Dose-Response Relationship, Drug; Female; Kidney; Liver; Liver Diseases; Male; Metallothionein; Rats; Rats, Inbred Strains; Time Factors; Tissue Distribution