bromochloroacetic-acid and Chemical-and-Drug-Induced-Liver-Injury

Liver penetration is a rare but serious complication of peptic ulcer disease. We report a case of a 33-year-old woman who took large doses of nonsteroidal antiinflammatory drugs and developed a giant duodenal ulcer that penetrated into her liver. The diagnosis was based on histologic examination of endoscopic biopsies. She was initially treated with a proton pump inhibitor, but, within 5 weeks, she developed a symptomatic postbulbar stricture that required surgical correction. We also review 11 other reported cases of endoscopically and histologically diagnosed peptic ulcer penetration into the liver.

Topics: Adult; Anti-Inflammatory Agents, Non-Steroidal; Chemical and Drug Induced Liver Injury; Diclofenac; Duodenal Ulcer; Duodenoscopy; Duodenum; Female; Humans; Immunoenzyme Techniques; Intervertebral Disc Displacement; Keratins; Liver; Liver Diseases; Liver Function Tests; Peptic Ulcer Hemorrhage; Peptic Ulcer Perforation

Mallory bodies are a morphological key feature of severe alcoholic liver cell injury (alcoholic hepatitis) and the morphological expression of dysregulation and derangement of the intermediate filament cytoskeleton of the hepatocyte. Their pathogenesis is still unclear. Studies on Mallory body formation may not only help to elucidate the mechanisms of liver cell injury associated with alcoholic hepatitis, but may also contribute to our understanding of the regulation and function of the intermediate filament cytoskeleton.

Topics: Animals; Chemical and Drug Induced Liver Injury; Colchicine; Cytoskeleton; Electrophoresis, Polyacrylamide Gel; Endoplasmic Reticulum; Fluorescent Antibody Technique; Griseofulvin; Hepatitis, Alcoholic; Humans; Isoantibodies; Isoelectric Focusing; Keratins; Liver; Liver Cirrhosis, Alcoholic; Mice; Microscopy, Electron; Rats

The formation of protein inclusions is frequently associated with chronic metabolic diseases. In mice, short-term intoxication with 3,5-diethoxycarbonyl-1,4-dihydrocollidine (DDC) leads to hepatocellular damage indicated by elevated serum liver enzyme activities, whereas only minor morphological changes are observed. Conversely, chronic administration of DDC for several weeks results in severe morphological damage, characterized by hepatocellular ballooning, disruption of the intermediate filament cytoskeleton, and formation of Mallory-Denk bodies consisting predominantly of misfolded keratins, Sqstm1/p62, and heat shock proteins. To evaluate the mechanistic underpinnings for this dichotomy we dissected the time-course of DDC intoxication for up to 10 weeks. We determined body weight change, serum liver enzyme activities, morphologic alterations, induction of antioxidant response (heme oxygenase-1, HO-1), oxidative damage and ATP content in livers as well as respiration, oxidative damage and the presence and activity of HO-1 in endoplasmic reticulum and mitochondria (mtHO-1). Elevated serum liver enzyme activity and oxidative liver damage were already present at early intoxication stages without further subsequent increase. After 2 weeks of intoxication, mice had transiently lost 9% of their body weight, liver ATP-content was reduced to 58% of controls, succinate-driven respiration was uncoupled from ATP-production and antioxidant response was associated with the appearance of catalytically active mtHO-1. Oxidative damage was associated with both acute and chronic DDC toxicity whereas the onset of chronic intoxication was specifically associated with mitochondrial dysfunction which was maximal after 2 weeks of intoxication. At this transition stage, adaptive responses involving mtHO-1 were induced, indirectly leading to improved respiration and preventing further drop of ATP levels. Our observations clearly demonstrate principally different mechanisms for acute and chronic toxic damage.

Topics: Acute Disease; Adaptor Proteins, Signal Transducing; Adenosine Triphosphate; Animals; Body Weight; Chemical and Drug Induced Liver Injury; Chemical and Drug Induced Liver Injury, Chronic; Energy Metabolism; Enzyme Induction; Gene Expression; Heat-Shock Proteins; Heme Oxygenase-1; Hepatocytes; Keratins; Liver; Male; Mallory Bodies; Mice; Mitochondria; Oxidative Stress; Protein Folding; Pyridines; Sequestosome-1 Protein; Time Factors

Liver failure represents a serious challenge for cell based therapies. Mesenchymal stem cells (MSCs) possess potential for regeneration of fibrotic liver; however, there is a dire need to improve their hepatic differentiation. This study examines a pretreatment strategy to augment the differentiation potential of MSCs towards hepatic lineage. MSCs were isolated from C57BL/6 wild type mice and were characterized by flow cytometry for CD44 (92.4%), CD90 (96.6%), CD105 (94.7%), CD45 (0.8%) and CD34 (1.4%) markers. To improve the differentiation potential of MSCs towards hepatic lineage, cells were pretreated with injured liver tissue in an in-vitro model, which resulted in high expression of albumin, cytokeratin 8, 18, TAT and HNF1α as compared to untreated MSCs. The efficacy of pretreated MSCs was evaluated by preparing in-vivo mouse model with liver fibrosis by intraperitoneal administration of CCl(4). Pretreated MSCs were transplanted in the left lateral lobe of mice with liver fibrosis and showed enhanced localization and differentiation abilities after 1 month. The expression for cytokeratin 8, 18, albumin and Bcl-xl was up-regulated and that of HGF, Bax and Caspase- 3 was down-regulated in animals transplanted with pretreated MSCs. Sirus red staining also confirmed a significant reduction in the fibrotic area in liver tissue transplanted with pretreated MSCs as compared to untreated MSCs and was concomitant with improved serum levels of bilirubin and alkaline phosphatase (ALP). Therefore, it was concluded that pretreatment with injured liver tissue augment homing and hepatic differentiation abilities of MSCs and provides an improved procedure for the treatment of liver fibrosis.

Topics: Albumins; Animals; Carbon Tetrachloride; Cell Differentiation; Cells, Cultured; Chemical and Drug Induced Liver Injury; Flow Cytometry; Gene Products, tat; Hepatocyte Nuclear Factor 1-alpha; Hepatocytes; Keratins; Liver; Liver Cirrhosis; Mesenchymal Stem Cell Transplantation; Mesenchymal Stem Cells; Mice; Mice, Inbred C57BL; Receptors, Cell Surface

Keratins 8 and 18 (K8 and K18) are heteropolymeric intermediate filament phosphoglycoproteins of simple-type epithelia. Mutations in K8 and K18 predispose the affected individual to liver disease as they protect hepatocytes from apoptosis. K18 undergoes dynamic O-linked N-acetylglucosamine glycosylation at Ser 30, 31 and 49. We investigated the function of K18 glycosylation by generating mice that overexpress human K18 S30/31/49A substitution mutants that cannot be glycosylated (K18-Gly(-)), and compared the susceptibility of these mice to injury with wild-type and other keratin-mutant mice. K18-Gly(-) mice are more susceptible to liver and pancreatic injury and apoptosis induced by streptozotocin or to liver injury by combined N-acetyl-D-glucosaminidase inhibition and Fas administration. The enhanced apoptosis in the livers of mice that express K18-Gly(-) involves the inactivation of Akt1 and protein kinase Ctheta as a result of their site-specific hypophosphorylation. Akt1 binds to K8, which probably contributes to the reciprocal hyperglycosylation and hypophosphorylation of Akt1 that occurs on K18 hypoglycosylation, and leads to decreased Akt1 kinase activity. Therefore, K18 glycosylation provides a unique protective role in epithelial injury by promoting the phosphorylation and activation of cell-survival kinases.

Topics: Acetylglucosamine; Amino Acid Substitution; Animals; Antibodies, Monoclonal; Apoptosis; beta-N-Acetylhexosaminidases; Caspase 3; Chemical and Drug Induced Liver Injury; Cytoskeleton; Cytosol; Enzyme Inhibitors; fas Receptor; Glycogen Synthase Kinase 3; Glycosylation; Hepatocytes; HSP70 Heat-Shock Proteins; Humans; Keratin-18; Keratin-8; Keratins; Liver; Mice; Mice, Inbred Strains; Mice, Knockout; Mice, Transgenic; Models, Biological; Oximes; Pancreas; Phenylcarbamates; Phosphorylation; Protein Binding; Protein Kinase C-delta; Protein Serine-Threonine Kinases; Proto-Oncogene Proteins c-akt; Pyruvate Dehydrogenase Acetyl-Transferring Kinase; Streptozocin

Mounting evidence suggests that keratin post-translational modifications are crucial for many cellular processes. Now, keratin 18 modified by the addition of an O-linked N-acetylglucosamine residue is shown to be as a critical effector of stress-responsive Akt signalling, providing an important link between keratin glycosylation and cell survival.

Topics: Acetylglucosamine; Animals; Apoptosis; beta-N-Acetylhexosaminidases; Cell Survival; Chemical and Drug Induced Liver Injury; fas Receptor; Glycosylation; Humans; Keratin-18; Keratin-8; Keratins; Models, Biological; Oximes; Phenylcarbamates; Phosphorylation; Protein Processing, Post-Translational; Proto-Oncogene Proteins c-akt; Streptozocin

Mallory bodies (MBs) and intracellular hyaline bodies (IHBs) are cytoplasmic hepatocellular inclusions that consist of aggregated proteins. MBs are characteristically associated with alcoholic and non-alcoholic steatohepatitis, but may also be found in chronic cholestatic and metabolic (eg copper intoxication) diseases and hepatocellular neoplasms, particularly hepatocellular carcinomas. IHBs have hitherto only been described in hepatocellular carcinoma cells. In the present study hepatocellular carcinomas (HCCs) and a case of idiopathic copper toxicosis were evaluated with respect to the presence and mutual relationship of MBs and IHBs. IHBs alone were present in 8.6%, MBs alone in 16.1% and both types of inclusion in 7.5% of HCCs. It is shown that IHBs may also occur in non-neoplastic hepatocytes in association with idiopathic copper toxicosis, together with MBs. In HCCs and idiopathic copper toxicosis, MBs and IHBs may be present within the same cell. Moreover, hybrid inclusions holding an intermediate position between MBs and IHBs regarding light microscopy, ultrastructure and composition exist. MBs and IHBs contain p62, a stress-inducible adapter protein, as the major constituent. In MBs p62 is associated with keratins, whereas classical IHBs lack keratins. Light microscopic, electron microscopic and immunohistochemical data suggest a close pathogenetic relationship between MBs and IHBs. Both types of inclusion are the result of over-expression and accumulation of the stress protein p62. If p62 is induced alone, or at least prevails, IHBs may arise by aggregation. However, if abnormal keratins are present in addition to p62, p62 associates and co-aggregates with keratins, finally leading to classical MBs.

Topics: Adaptor Proteins, Signal Transducing; Carcinoma, Hepatocellular; Chemical and Drug Induced Liver Injury; Copper; Hepatocytes; Humans; Hyalin; Inclusion Bodies; Keratins; Liver Diseases; Liver Neoplasms; Microscopy, Electron; Microscopy, Immunoelectron; Neoplasm Proteins; Sequestosome-1 Protein

Mallory bodies (MBs) are characteristic of several liver disorders, and consist primarily of keratins with transglutaminase-generated keratin crosslinks. We tested the effect of the transglutaminase-2 (TG2) inhibitor KCC009 on MB formation in a mouse model fed 3,5-diethoxycarbonyl-1,4-dihydrocollidine (DDC). KCC009 decreased DDC-induced liver enlargement without affecting MB formation or extent of liver injury. TG2 protein and activity increased after DDC feeding and localized within and outside hepatocytes. KCC009 inhibited DDC-induced hepatomegaly by affecting hepatocyte cell size rather than proliferation. Hence, TG2 is a potential mediator of injury-induced hepatomegaly via modulation of hepatocyte hypertrophy, and KCC009-mediated TG2 inhibition does not affect mouse MB formation.

Topics: Animals; Cell Size; Chemical and Drug Induced Liver Injury; Dicarbethoxydihydrocollidine; Enzyme Inhibitors; GTP-Binding Proteins; Hepatomegaly; Humans; Inclusion Bodies; Isoxazoles; Keratins; Liver Diseases; Mice; Mice, Inbred C3H; Protein Glutamine gamma Glutamyltransferase 2; Proteins; Transglutaminases

Keratin 8 (K8) variants predispose to human liver injury via poorly understood mechanisms. We generated transgenic mice that overexpress the human disease-associated K8 Gly61-to-Cys (G61C) variant and showed that G61C predisposes to liver injury and apoptosis and dramatically inhibits K8 phosphorylation at serine 73 (S73) via stress-activated kinases. This led us to generate mice that overexpress K8 S73-to-Ala (S73A), which mimicked the susceptibility of K8 G61C mice to injury, thereby providing a molecular link between K8 phosphorylation and disease-associated mutation. Upon apoptotic stimulation, G61C and S73A hepatocytes have persistent and increased nonkeratin proapoptotic substrate phosphorylation by stress-activated kinases, compared with wild-type hepatocytes, in association with an inability to phosphorylate K8 S73. Our findings provide the first direct link between patient-related human keratin variants and liver disease predisposition. The highly abundant cytoskeletal protein K8, and possibly other keratins with the conserved S73-containing phosphoepitope, can protect tissue from injury by serving as a phosphate "sponge" for stress-activated kinases and thereby provide a novel nonmechanical function for intermediate filament proteins.

Topics: Animals; Apoptosis; Chemical and Drug Induced Liver Injury; Disease Models, Animal; Fas Ligand Protein; Genetic Predisposition to Disease; Genetic Variation; Hepatocytes; Humans; Intermediate Filament Proteins; Keratin-8; Keratins; Liver Diseases; Liver Function Tests; Marine Toxins; Membrane Glycoproteins; Mice; Mice, Knockout; Mice, Transgenic; Microcystins; Mitogen-Activated Protein Kinases; Mutation; Peptides, Cyclic; Phosphorylation; Tumor Necrosis Factors

We examined the response of murine oval cells, that is, the putative liver progenitor cells, to acetaminophen. Female C57BL/6J mice were injected intraperitoneally with varying doses of N-acetyl-paraaminophen (APAP) (250, 500, 750, and 1,000 mg/kg of weight) and sacrificed at 3, 6, 9, 24, and 48 hours. In preliminary studies, we showed that anticytokeratin antibodies detected A6-positive cells with a sensitivity and specificity of greater than 99%. The oval cell reaction was quantified, on immunostaining for biliary-type cytokeratins, as both number and density of oval cells per portal tract, analyzed by size of portal tract. Acetaminophen injury was followed by periportal oval cell accumulation displaying a moderate degree of morphological homogeneity. Oval cell response was biphasic, not temporally correlating with the single wave of injury seen histologically. Increases in oval cells were largely confined to the smallest portal tracts, in keeping with their primary derivation from the canals of Hering, and increased in a dose-dependent fashion. The timing of the two peaks of the oval cell reaction also changed with increasing dose, the first becoming earlier and the second later. In conclusion, our studies indicate a marked oval cell activation during the height of hepatic injury. Oval cells appear to be resistant to acetaminophen injury. The close fidelity of mechanism and histology of acetaminophen injury between mouse and human livers makes it a useful model for investigating liver regeneration and the participation of stem/progenitor cells in that process.

Topics: Acetaminophen; Animals; Chemical and Drug Induced Liver Injury; Dose-Response Relationship, Drug; Female; Immunologic Techniques; Keratins; Liver; Liver Diseases; Mice; Mice, Inbred C57BL; Stem Cells; Time Factors

Several studies have demonstrated that bone marrow contains a subpopulation of stem cells capable of participating in the hepatic regenerative process, even if some reports indicate quite a low level of liver repopulation by human stem cells in the normal and transiently injured liver.. In order to overcome the low engraftment levels seen in previous models, we tried the direct intraperitoneal administration of human cord blood stem cells, using a model of hepatic damage induced by allyl alcohol in NOD/SCID mice.. We designed a protocol based on stem cell infusion following liver damage in the absence of irradiation. Flow cytometry, histology, immunohistochemistry and RT-PCR for human hepatic markers were performed to monitor human cell engraftment.. Human stem cells were able to transdifferentiate into hepatocytes, to improve liver regeneration after damage and to reduce the mortality rate both in both protocols, even if with qualitative and quantitative differences in the transdifferentiation process.. We demonstrated for the first time that the intraperitoneal administration of stem cells can guarantee a rapid liver engraftment. Moreover, the new protocol based on stem cell infusion following liver damage in the absence of irradiation may represent a step forward for the clinical application of stem cell transplantation.

Topics: Animals; Chemical and Drug Induced Liver Injury; Cord Blood Stem Cell Transplantation; Disease Models, Animal; Flow Cytometry; Glyceraldehyde-3-Phosphate Dehydrogenases; Humans; Immunohistochemistry; Keratin-7; Keratins; Liver; Mice; Mice, Inbred NOD; Mice, SCID; Propanols; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Survival Rate; Transplantation, Heterologous; Treatment Outcome

Mallory bodies (MBs) consist of abnormal keratins, ubiquitin, heat shock proteins, and the protein p62. p62 is encoded by an immediate-early response gene that rapidly responds to a variety of extracellular signals involved in cell proliferation, differentiation, and particularly oxidative stress. It acts as an adapter in signal transduction and binds noncovalently to ubiquitin, possibly being involved in the regulation of the fate of ubiquitinated proteins by segregation (i.e., sequestosome or aggresome formation). The presence of p62 together with ubiquitinated abnormal keratins in the MB characterizes MBs as a disease-associated type of sequestosome. A detailed study on the expression of p62 and its relationship to MB formation in the 3,5-diethoxycarbonyl-1,4-dihydrocollidine (DDC)-treated mouse liver is reported based on immunohistochemical, immunoblot, and Northern blot analyses. The results indicate that p62 is rapidly induced in hepatocytes of intoxicated animals preceding MB formation. As suggested by experiments with short-term DDC-treated naive mice and mice refed DDC after recovery from long-term DDC treatment (primed mice), p62 does not exert an initiating effect on MB formation but the appearance of MBs requires the presence of abnormal keratins, which associate with p62 after ubiquitination. The rapid induction of p62 and its association with MBs further support the role of oxidative stress in MB formation. In conclusion, the constant presence of p62 in MBs suggests that binding of p62 to abnormal keratins may allow hepatocytes to dispose potentially harmful proteins in a biologically inert manner.

Topics: Adaptor Proteins, Signal Transducing; Animals; Antibodies; Biopsy; Blotting, Northern; Blotting, Western; Carrier Proteins; Chemical and Drug Induced Liver Injury; Dicarbethoxydihydrocollidine; Humans; Immediate-Early Proteins; Inclusion Bodies; Keratins; Liver; Liver Diseases; Male; Mice; Microscopy, Immunoelectron; Proteins; RNA, Messenger; Sequestosome-1 Protein; Transcription Factor TFIIH; Transcription Factors; Ubiquitin

Tumor necrosis factor (TNF) is a cytokine produced by macrophages and T lymphocytes that acts through two distinct receptors, TNFR1 (60 kD, CD120a) and TNFR2 (80 kD, CD120b), to affect cellular proliferation, differentiation, survival, and cell death. In addition to its proinflammatory actions in mucosal tissue, TNF is important for liver regeneration. Keratin 8 (K8) and keratin 18 (K18) form intermediate filaments characteristic of liver and other single cell layered, internal epithelia and their derivative cancers. K8-deficient (K8(-)) mice, which escape embryonic lethality, develop inflammatory colorectal hyperplasia, mild liver abnormalities, and tolerate hepatectomy poorly. We show that normal and malignant epithelial cells deficient in K8 and K18 are approximately 100 times more sensitive to TNF-induced death. K8 and K18 both bind the cytoplasmic domain of TNFR2 and moderate TNF-induced, Jun NH(2)-terminal kinase (JNK) intracellular signaling and NFkappaB activation. Furthermore, K8(-) and K18(-) mice are much more sensitive to TNF dependent, apoptotic liver damage induced by the injection of concanavalin A. This moderation of the effects of TNF may be the fundamental function of K8 and K18 common to liver regeneration, inflammatory bowel disease, hepatotoxin sensitivity, and the diagnostic, persistent expression of these keratins in many carcinomas.

Topics: Animals; Apoptosis; Cell Line; Chemical and Drug Induced Liver Injury; Concanavalin A; Epithelial Cells; Humans; JNK Mitogen-Activated Protein Kinases; Keratins; Liver; Mice; Mice, Knockout; Mitogen-Activated Protein Kinases; NF-kappa B; Protein Binding; Receptors, Tumor Necrosis Factor; Signal Transduction; Transfection; Tumor Cells, Cultured; Tumor Necrosis Factor-alpha

In alcoholic hepatitis, a severe form of alcohol-induced toxic liver injury, as well as in experimental intoxication of mice with the porphyrinogenic drugs griseofulvin and 3,5-diethoxycarbonyl-1, 4-dihydrocollidine, hepatocytes form cytoplasmic protein aggregates (Mallory bodies; MBs) containing cytokeratins (CKs) and non-CK components. Here we report that mice lacking the CK8 gene and hence CK intermediate filaments in hepatocytes, but still expressing the type I partner, ie, the CK18 gene, do not form MBs but suffer from extensive porphyria and progressive toxic liver damage, leading to the death of a considerable number of animals (7 of 12 during 12 weeks of intoxication). Our observations show that 1) in the absence of CK8 as well as in the situation of a relative excess of CK18 over CK8 no MBs are formed; 2) the loss of CK8 is not compensated by other type II CKs; and 3) porphyria and toxic liver damage are drastically enhanced in the absence of CK8. Our results point to a protective role of CKs in certain types of toxic liver injury and suggest that MBs by themselves are not harmful to hepatocytes but may be considered as a product of a novel defense mechanism in hepatocytes.

Topics: Animals; Bile Ducts; Chemical and Drug Induced Liver Injury; Cytoplasm; Cytoskeleton; Dicarbethoxydihydrocollidine; Epithelium; Keratin-7; Keratins; Liver; Liver Diseases; Mice; Mice, Inbred Strains; Mice, Knockout

Simple epithelia express keratins 8 (K8) and 18 (K18) as their major intermediate filament (IF) proteins. One important physiologic function of K8/18 is to protect hepatocytes from drug-induced liver injury. Although the mechanism of this protection is unknown, marked K8/18 hyperphosphorylation occurs in association with a variety of cell stresses and during mitosis. This increase in keratin phosphorylation involves multiple sites including human K18 serine-(ser)52, which is a major K18 phosphorylation site. We studied the significance of keratin hyperphosphorylation and focused on K18 ser52 by generating transgenic mice that overexpress a human genomic K18 ser52--> ala mutant (S52A) and compared them with mice that overexpress, at similar levels, wild-type (WT) human K18. Abrogation of K18 ser52 phosphorylation did not affect filament organization after partial hepatectomy nor the ability of mouse livers to regenerate. However, exposure of S52A-expressing mice to the hepatotoxins, griseofulvin or microcystin, which are associated with K18 ser52 and other keratin phosphorylation changes, resulted in more dramatic hepatotoxicity as compared with WT K18-expressing mice. Our results demonstrate that K18 ser52 phosphorylation plays a physiologic role in protecting hepatocytes from stress-induced liver injury. Since hepatotoxins are associated with increased keratin phosphorylation at multiple sites, it is likely that unique sites aside from K18 ser52, and phosphorylation sites on other IF proteins, also participate in protection from cell stress.

Topics: 3T3 Cells; Actin Cytoskeleton; Amino Acid Substitution; Animals; Chemical and Drug Induced Liver Injury; Genetic Predisposition to Disease; Griseofulvin; Hepatectomy; Humans; Intermediate Filaments; Keratins; Liver Regeneration; Mice; Mice, Transgenic; Microcystins; Okadaic Acid; Peptides, Cyclic; Phosphorylation; Point Mutation; Protein Processing, Post-Translational

A cytokeratin immunohistochemical study was performed on 38 liver biopsies from cases of primary biliary cirrhosis, primary sclerosing cholangitis, extrahepatic biliary obstruction or drug-induced liver disease in order to analyse the cytoskeletal changes in detail. On paraffin sections of 27 cases, a variable number of hepatocytes were reactive with a polyclonal anti-cytokeratin antiserum that, in the normal liver, stains bile duct cells only. On cryostat sections of 23 cases, a variable number of hepatocytes were immunoreactive with a monoclonal antibody specifically directed against cytokeratin no. 7 and were most numerous in cases of long-standing cholestasis irrespective of the aetiology. In three cases of primary sclerosing cholangitis and two cases of primary biliary cirrhosis a few hepatocytes were also weakly positive with a monoclonal antibody specific for cytokeratin no. 19. Since cytokeratins no. 7 and no. 19 are, in the normal liver, restricted to bile duct cells, these results further support the concept of 'ductular metaplasia' of hepatocytes, the mechanism of which remains unclear.

Topics: Antibodies, Monoclonal; Autopsy; Biopsy; Chemical and Drug Induced Liver Injury; Cholangitis, Sclerosing; Cholestasis, Intrahepatic; Cytoskeleton; Frozen Sections; Humans; Keratins; Liver; Liver Cirrhosis, Biliary; Liver Diseases; Metaplasia; Paraffin

A monoclonal murine antibody (KM 54-5) was produced against Mallory body (MB) material isolated from liver tissue of griseofulvin treated mice. The antigen was identified by positive immunofluroescence microscopy of MBs and by the immunoblotting technique on polypeptides separated by sodium dodecyl sulfate polyacrylamide gel electrophoresis. In immunoblotting experiments, antibody KM 54-5 reacted with cytokeratins A (human no. 8) and D (human no. 18) of murine, bovine, and human hepatocytes as well as with cytokeratin A (no. 8) and its degradation products present in isolated murine MB. In immunofluorescence microscopy the antibody did not react with cytokeratin filaments of normal liver but showed a positive reaction with MBs after a certain stage in MB development had been reached. In a dot blot assay, using individual cytokeratin polypeptides isolated from murine liver and purified by ion exchange chromatography in pH 8 buffer containing 8 M urea, the antibody reacted with the individual polypeptides A (no. 8) and D (no. 18) but not with the heterotypic tetramer (A2D2) reconstituted from these polypeptides in 4 M urea. These findings confirm the cytokeratin nature of MB filaments. In addition, they show that the pathologic process of MB formation involves changes in cytokeratin organization and conformation, resulting in the accessibility of a specific antigenic determinant which is inaccessible ("masked") in the heterotypic tetramer subunit and in the cytokeratin filaments of normal cells. Hence this study presents an example of a pathological change of cytokeratin filaments and illustrates the value of monoclonal antibodies in the detection of such changes.

Topics: Animals; Antibodies, Monoclonal; Cattle; Cell Line; Chemical and Drug Induced Liver Injury; Electrophoresis, Polyacrylamide Gel; Fluorescent Antibody Technique; Griseofulvin; Humans; Immunologic Techniques; Inclusion Bodies; Keratins; Liver; Liver Diseases; Macromolecular Substances; Male; Mice; Rats