PubMed Health⌕ Search

Biomedical subjects

K Zatloukal

Publications and source records attributed to K Zatloukal.

At least 73 records · Page 4Linked to original sources
PubMed1992

Common epitopes of human and murine Mallory bodies and Lewy bodies as revealed by a neurofilament antibody.

The antibody SMI 31, which is directed against a phosphorylated epitope, associated with neurofilaments and recognizes Lewy bodies in brains of patients with Parkinson's disease (Bancher C, Lassmann H, Budka H, Jellinger K, Grundgke-Iqbal I, Iqbal K, Wiche G, Seitelberger F, Wisniewski H: J Neuropathol Exp Neurol 1:81, 1989), decorated in immunofluorescence microscopy Mallory bodies (MBs) present in livers of mice chronically treated with griseofulvin and 3,5-diethoxycarbonyl-1,4-dihydrocollidine. In immunoblots it recognized very acidic MB components in a molecular weight range between 55 and 69.5 kilodaltons in addition to poorly soluble high molecular weight material. Moreover, an antibody to tau protein showed similar reactivities in immunofluorescence microscopy and immunoblotting experiments. Both antibodies also stained MBs in human liver with alcoholic hepatitis. These observations support and extend earlier findings which indicate that several intermediate filament-related cellular inclusion bodies, including MBs, share a variety of morphologic, structural and antigenic features. They also suggest the involvement of tau or tau-like proteins in MB formation.

Animals↗
PubMed1992

Alcoholic liver disease. Parenchyma to stroma relationship in fibrosis and cirrhosis as revealed by three-dimensional reconstruction and immunohistochemistry.

Severe ethanol-induced liver damage is characterized by fibrous dissociation of liver cell plates leading to many apparently isolated hepatocytes. Three-dimensional reconstruction, however, revealed hepatocytes that were surrounded by connective tissue as endpoints of "parenchymal pillars" or in association with liver cell plates and bile ductules. Double immunofluorescence studies displayed the expression of cytokeratin (CK) 7 in bile ducts, including bile ductules, but also in some hepatocytes still organized in liver cell plates. The other bile duct, typical CK, namely CK 19, was only detectable in few hepatocytes. However, the expression of CK 7 and/or CK 19 was less frequent in hepatocytes that were closely associated with bile ductules. CK 7 and CK 19 were also found in some, but not all, Mallory bodies. These observations indicate that the expression of these two CKs is neither related to a transformation of hepatocytes to bile duct-like structures ("ductal metaplasia") nor to the formation of Mallory bodies. Furthermore, double immunofluorescence studies revealed small groups of hepatocytes and bile ductules that were encircled by basement membrane material, thus suggesting the formation of "secretory units."

Aged↗
PubMed1992

Modulation of protein composition of nuclear lamina. Reduction of lamins B1 and B2 in livers of griseofulvin-treated mice.

Chronic griseofulvin (GF) intoxication of mice leads to severe alterations of the hepatocytic intermediate filament cytoskeleton similar to that found in alcoholic hepatitis in humans (i.e., derangement and diminution of the keratin filament network and appearance of cytoplasmic aggregates of keratin-containing material, termed Mallory bodies). To investigate eventual alterations of nuclear lamins under these pathologic situations monoclonal antibodies were produced. One of these, GL-35, was directed to lamin B1 and lamin B2. Immunofluorescence microscopy revealed in GF-treated livers, in comparison to normal mouse livers, a highly reduced immunoreaction for lamins B1 and B2, whereas the staining for lamins A and C was unchanged. This reduction of both B-type lamins occurred before detectable alterations of keratin filaments and was reversible after cessation of GF intoxication. A diminished content of both B-type lamins in relation to lamins A and C in GF-intoxicated livers was also revealed by analysis of isolated nuclear envelopes on two-dimensional gels. Moreover, there was a clear predominance of more acidic isoelectric variants of lamins B1 and B2. In contrast to the reduced amount of B-type lamin proteins no reduction in the concentration of the mRNA for lamin B1 was found. For lamin B2 even an increase of mRNA was detected in GF-treated livers. These results indicate that GF not only interferes with expression of the keratin intermediate filament skeleton of hepatocytes but also leads to selective alterations of the nuclear lamins, most likely by posttranslational modifications of intermediate filament proteins.

Animals↗
PubMed1991

Co-expression of cytokeratin and vimentin filaments in rete testis and epididymis. An immunohistochemical study.

In 11 testes of different developmental stages (from 10-week-old embryos to adult) the cytokeratin and vimentin expression patterns of rete testis and epididymis were investigated immunohistochemically in formaldehyde-fixed paraffin-embedded material. In addition, immunofluorescence microscopy including double immunofluorescence was performed on frozen sections of 3 of these 11 cases. Rete testis and epididymis cells displayed a heterogeneous co-expression of cytokeratin and vimentin. In double immunohistochemistry, differences in distribution of keratin and vimentin intermediate filaments with predominance of cytokeratins in the apical cytoplasmic regions and of vimentin filaments in the basal portions of the cells were found. Cytokeratin expression preceded the appearance of vimentin: cytokeratin was already detectable in 10-week-old embryos, while weak vimentin immunoreactivity was first seen in 12-week-old embryos and became conspicuous in testes around the perinatal period. In testes of children up to 2 years of age the cytoplasmic distribution of cytokeratin and vimentin was more homogeneous. Predominance of the basal cell portions for vimentin and the apical regions for cytokeratin staining were less pronounced than in adult testes. In the proximal and distal parts of the epididymis a different intermediate filament expression pattern was found with a clear predominance of cytokeratin near the rete.

Adult↗
PubMed1991

Enzyme-histochemical studies of griseofulvin-intoxicated mouse livers.

Enzyme-histochemical studies were conducted on livers of mice chronically fed griseofulvin (GF) in order to produce Mallory bodies (MBs) in hepatocytes. The development of MBs is associated with derangement of the immunohistochemically detectable intermediate filament (IF) cytoskeleton of the cytokeratin (CK) type, although no strict correlation between appearance or involution of MBs and the cytoskeletal alterations exists. Since the function of the IF cytoskeleton and the relationship of its disturbance to cell injury is unknown, the aim of the present study was to correlate the activities of several key enzymes of cellular metabolic pathways with the disturbance of the cytoskeleton architecture. For that purpose enzyme-histochemistry in combination with immunohistochemical CK-IF stainings were performed on identical sections. In GF-intoxicated mouse livers the normal topography of enzyme activities was disturbed, but no strict colocalization of enzymatic and cytoskeletal changes was found. Glucose-6-phosphatase, a microsomal enzyme involved in glucose output and gluconeogenesis, showed elevated activity in MB-free hepatocytes with diminished immunostainable CK-IF cytoskeleton refuting the concept of a disability of those cells to export glucose. It could indeed indicate that those cells without MBs are in the state of recovery. However, these cells could also resemble "hyperactive foci". Glycogen was decreased in MB-containing hepatocytes with disturbed cytoskeleton, and this feature favours the assumption of cell degeneration. On the other hand, the mitochondrial marker enzymes, i.e. succinate dehydrogenase, cytochrome-c-oxidase and 3-hydroxybutyrate dehydrogenase, remained unchanged in altered hepatocytes. Alkaline phosphatase activity at the canalicular pole of GF-intoxicated hepatocytes was elevated, indicating cholestatic features associated with this disorder. However, since altered hepatocytes did not show impairment of oxido-reductase activities, a severe impairment of bile secretion as a consequence of cell damage is unlikely. Unchanged or even increased ATPase activity of altered hepatocytes also indicated their sustained metabolic abilities. The results presented provide indirect evidence that hepatocytes with disturbed IF cytoskeleton do not significantly differ from normal cells with respect to oxidative metabolism, fatty acid synthesis and gluconeogenesis. This suggests that alterations of the IF cytoskeleton associated with GF intoxication and MB formation have no significant adverse influence on the metabolic functions of liver cells, as far as can be assessed by evaluation by enzyme-histochemical staining of several key enzymes.

Animals↗
PubMed1991

Changes of cytokeratin filament organization in human and murine Mallory body-containing livers as revealed by a panel of monoclonal antibodies.

Mallory bodies (MBs) are characteristics morphologic features of alcoholic hepatitis and can be produced in mouse hepatocytes by chronic griseofulvin (GF) intoxication. The formation of MBs, which share some immunological, biochemical, and ultrastructural features with cytokeratin (CK) filaments of normal liver, is accompanied by derangement and even loss of the CK cytoskeleton of hepatocytes ("empty cells") as revealed by immunofluorescence microscopy. To clarify whether this diminution or lack of CK-related staining of MB-containing hepatocytes was due to loss of CK filaments or changes in antigenicity or accessibility of antigenic determinants immunohistochemical studies using a battery of monoclonal and polyclonal CK antibodies were performed. It could be shown that all these antibodies directed against different CK polypeptide components and antigenic determinants of CKs revealed a highly reduced or even undetectable cytoplasmic CK meshwork in most cells with fully developed large MBs. In the light of our present knowledge of the organization of CK intermediate filaments, these results indicate that the phenomenon of the "empty cells" reflects a diminution of CK meshwork rather than altered antigenic determinants.

Animals↗
PubMed1991

High molecular weight components are main constituents of Mallory bodies isolated with a fluorescence activated cell sorter.

Mallory bodies (MBs) are cytoplasmic filamentous aggregates containing cytokeratin (CK) material. They occur in hepatocytes of patients with alcoholic liver disease (i.e., alcoholic hepatitis) and can also be induced experimentally in mice by chronic griseofulvin intoxication. To further investigate components and mechanisms involved in MB formation, a new method for MB purification was established. MBs present in a liver homogenate of griseofulvin-fed mice were labeled with a murine monoclonal antibody specific for MBs and a second fluorescein isothiocyanate-conjugated (anti-mouse IgG and IgM) antibody and subsequently isolated by two sequential sorting procedures using a fluorescence activated cell sorter (FACS). Purity of MB isolates was over 90% as revealed by computer analysis of sorting signals and fluorescence and electron microscopy. Electrophoretic separation on sodium dodecyl sulfate-polyacrylamide gels revealed three MB-related polypeptides with apparent molecular masses of 48, 55, and 65 kilodaltons but most of the highly purified MB material did not enter the gel or remained at the interphase between stacking and resolving gels. Western blotting with CK-specific antibodies showed the presence of CK epitopes in the high molecular weight MB material, which has a similar amino acid composition as normal liver CKs. These results establish that very high molecular weight material is the main constituent of MBs and suggest that a post-translational modification of CKs by covalent crosslinks is a principal mechanism of MB pathogenesis.

Amino Acids↗
PubMed1990

Fate of Mallory body-containing hepatocytes: disappearance of Mallory bodies and restoration of the hepatocytic intermediate filament cytoskeleton after drug withdrawal in the griseofulvin-treated mouse.

Mallory bodies are characteristic morphological features of alcoholic hepatitis in man and can be produced in the mouse by chronic griseofulvin intoxication. The appearance of Mallory bodies in hepatocytes is associated with derangement of the cytokeratin intermediate filament cytoskeleton, at least as revealed by immunofluorescence and suggested by immunoelectron microscopy. Immunohistochemical studies were performed to answer the question whether Mallory body formation and cytoskeleton alterations finally lead to cell death or are reversible phenomena. Chronically griseofulvin-intoxicated mice killed at different stages of recovery on a normal diet served as experimental animals. It could be shown that (a) Mallory bodies are very durable structures and are found for up to 6 mo after griseofulvin withdrawal as a result of persistence and neoformation; (b) new Mallory bodies can appear even several months after cessation of griseofulvin feeding; (c) Mallory body formation and cytoskeletal changes by themselves do not lead to irreversible cell damage; (d) the cytoskeletal changes are reversible within 7 mo after griseofulvin withdrawal; (e) a dissociation between disappearance of Mallory bodies and restoration of a regularly immunostained cytoplasmic cytokeratin meshwork is observed.

Animals↗
PubMed1990

Characterization of a second highly conserved B-type lamin present in cells previously thought to contain only a single B-type lamin.

Previous analyses of the nuclear lamina of mammalian cells have revealed three major protein components (lamins A, B and C) that have been identified by protein sequence homology as members of the intermediate filament (IF) protein family. It has been claimed that mammalian cells contain either all three lamins or lamin B alone. Using monoclonal antibodies specific for B-type lamins and cDNA cloning we identified a second major mammalian B-type lamin (murine lamin B2), thus showing that lamin composition in mammals is more complex than previously thought. Lamin B2 is coexpressed with lamin B1 (formerly termed lamin B) in all somatic cells and mammalian species that we analysed, including a variety of cells currently believed to contain only a single lamin. This suggests that two B-type lamins are necessary to form a functional lamina in mammalian somatic cells. By cDNA cloning we found that Xenopus laevis lamin LII is the amphibian homolog of mammalian lamin B2. Lamin expression during embryogenesis of amphibians and mammals shows striking similarities. The first lamins expressed in the early embryo are the two B-type lamins, while A-type lamins are only detected much later in development. These findings indicate that the genomic differentiation into two B-type lamins occurred early in vertebrate evolution and has been maintained in both their primary structure and pattern of expression.

Amino Acid Sequence↗
PubMed1990

High molecular weight component of Mallory bodies detected by a monoclonal antibody.

To identify Mallory body (MB) constituents, monoclonal antibodies to murine MBs induced by long-term griseofulvin (GF) feeding were produced. One of these, antibody MM 120-1, specifically reacted in immunofluorescence microscopy with MBs in all developmental stages but not with other cell structures of human and mouse liver and other organs. The MM 120-1 antigen was present in murine MBs induced by griseofulvin or 3,5-diethoxycarbonyl-1,4-dihydrocollidine feeding and also in human MBs in livers of patients with alcoholic hepatitis. In immunoblots, the MM 120-1 antigen was detectable in the high molecular weight fraction of MB preparations, most of which remained in the well and at the interphase between stacking and resolving gel. No reactivity with cytokeratin polypeptides of different conformational states (i.e., isolated cytoker atin components A and D, heterotetramers A2D2, reconstituted intermediate filaments) was found. It is concluded that the antibody MM 120-1 is a highly specific and sensitive marker for murine and human MBs recognizing a high molecular weight nonkeratin component. This component could play a central role in the pathogenesis of MBs.

Animals↗
PubMed1990

[Cytoskeleton--function and pathology: studies on the pathology of the intermediate filament cytoskeleton of liver cells].

Microfilaments, microtubules and intermediate filaments (IF) are major filamentous components of the cytoskeleton and play a role in the modulation of cell shape, in cellular movements, cellular stability, intracellular organisation as well as cell-to-cell and cell-to-stroma interactions. Particular interest was concentrated in the last few years on IF because of their cell type-specificity and, consequently, their suitability as cell markers in diagnostic pathology. Despite their apparent stability, IF are dynamic structures which may be modified under pathologic conditions. In recent years, pathologic alterations related to the IF cytoskeleton have been described in a diversity of chronic and degenerative disorders, including alcoholic hepatitis and neurologic diseases (e.g. M. Alzheimer, M. Parkinson). Our studies were particularly devoted to the elucidation of the pathogenesis of severe alcoholic liver injury (alcoholic hepatitis), which is associated with inflammation, liver cell degeneration and necrosis and morphologically characterized by the appearance of cytoplasmic hyaline inclusions (i.e., Mallory bodies). In the present review morphologic, immunologic and biochemical studies on nature and pathogenesis of Mallory bodies are summarized. Moreover, similarities between Mallory bodies and other cytoskeleton-related inclusion bodies suggest common routes of pathogenesis. Consequently, studies along these lines may not only lead to the understanding of mechanisms involved in alcoholic injury but may also provide information on general principles of cell damage as well as on regulation and function of the IF cytoskeleton.

Animals↗
PubMed1989

Hepatocellular cytokeratins as substrates of transglutaminases.

To examine whether hepatocellular cytokeratins can serve as substrates of transglutaminases (TG) TG-catalyzed incorporation of [3H]putrescine into, as well as cross-linking of, cytokeratins was studied. Purified guinea pig liver TG and mouse liver TG present in 105,000 x g supernatants were used as enzymes. Isolated mouse liver cytokeratin filaments, heterotypic tetramers (A2D2), as well as cytokeratin filaments reconstituted from isolated and column-purified liver cytokeratin polypeptides A and D served as substrates. Moreover, to more closely mimic the situation within the cell, mouse liver homogenate containing cytokeratins was also used. Cross-linked proteins were identified as cytokeratins by immunoblotting after sodium dodecyl sulfate-polyacrylamide gel electrophoresis using cytokeratin antibodies. The results indicated that mouse liver cytokeratins can serve as substrates of homologous and heterologous TG. However, liver cytokeratin components A and D differed in this respect. Depending on the experimental conditions either components D or components A were better substrates of TG-mediated cross-linking as revealed by increased high molecular weight aggregates, which failed to enter the gels, concomitant with a decrease of the monomer band suggesting a more intimate relationship between homologous cytokeratin polypeptides within the filament. The results presented provide the basis for studies of TG-induced cross-linking of cytoskeletal components in hepatocytes that may occur during liver cell injury associated with increased intracellular Ca2+ concentrations.

Animals↗
PubMed1989

[Immunohistochemical analysis of 42 renal cell carcinomas and one oncocytoma with mono- and polyclonal antibodies against vimentin and cytokeratin].

42 renal cell carcinomas and 1 oncocytoma were investigated by means of immunofluorescence (including double immunofluorescence) using a panel of mono- and polyclonal antibodies to vimentin and cytokeratins. In all tumors except chromophobe cell renal carcinoma (CCRC) and oncocytoma generally a coexpression of vimentin and cytokeratins could be demonstrated; however, the intermediate filament expression was often very heterogeneous with regard to the distribution of vimentin and cytokeratins in general, depending on the mono- and polyclonality of the antibodies and on the areas of a tumor investigated. In CCRC and oncocytoma all tumor cells contained cytokeratin filaments. In addition, as revealed by double immunofluorescence, in only occasional tumor cells we could demonstrate vimentin.

Adenocarcinoma↗
PubMed1988

A new technical of liver biopsy with plugging of the needle track using a double channel biopsy device. An experimental study.

A new liver biopsy technique with plugging of the intrahepatic needle track in one procedure using a double channel biopsy device (Austrian patent No 384,165 invented by K. Zatloukal and H.P. Dinges) is presented in an experimental study. This technique is similar to that described by Menghini, but is combined with a fibrin sealing system (Tissucol, Tisseel). Biopsy and sealing of the biopsy site is performed within less than two seconds. 15 liver biopsies were performed on 6 heparinized dogs after laparotomy. Heparinization was maintained for 6 hours and the efficiency of the sealing in preventing biopsy-related bleeding was checked before the abdominal wound was closed. No bleeding occurred after biopsy with the new needle system and occlusion of the needle track with fibrin sealant. The livers with the sealed needle tracks and all biopsies obtained were examined histologically. It was found that the needle tracks were filled with the fibrin sealant which was almost completely replaced by granulation tissue after 8 days. Biopsies performed with a conventional Menghini needle led to continuous bleeding from the needle track. The results show that this new biopsy device efficiently prevents bleeding from the biopsy site in animals with severely impaired blood coagulation. Therefore, application of this biopsy technique in humans might reduce bleeding complications particularly in patients with severe liver diseases and disturbed blood coagulation.

Animals↗
PubMed1987

Turnover of cytokeratin polypeptides in mouse hepatocytes.

The turnover of cytokeratin polypeptides A (equivalent to No. 8 of the human cytokeratin catalog) and D (equivalent to human cytokeratin No. 18) of mouse hepatocytes was studied by pulse-labeling of mouse liver proteins after intraperitoneal injection of L-[guanido-14C]arginine and [14C]sodium bicarbonate. At various times after injection cytoskeletal proteins were prepared and separated by SDS-polyacrylamide gel electrophoresis, and the specific radioactivities of polypeptides recovered from excised gel slices were determined. With L-[guanido-14C]arginine a rapid increase in the specific radioactivity of both cytokeratins was observed which reached a plateau between 12 and 24 h. With [14C]sodium bicarbonate maximal specific radioactivity was obtained at 6 h followed by a rapid decrease to half maximum values within the subsequent 6 h and then a slower decrease. Half-lives were determined from the decrease of specific radioactivities after pulse-labeling by least-squares plots and found to be 84 h (for cytokeratin component A) and 104 h (component D) for arginine labeling. Values obtained after bicarbonate labeling were similar (95 h for A and 98 h for D). These results show that liver cytokeratins are relatively stable proteins and suggest that components A and D are synthesized and degraded at similar rates, probably in a coordinate way.

Animals↗
PubMedDate not supplied

Induction of heat shock proteins in short-term cultured hepatocytes derived from normal and chronically griseofulvin-treated mice.

Freshly isolated mouse hepatocytes were tested with respect to the induction of heat shock (stress) proteins by elevated temperature, sodium arsenite and ethanol treatment. With heat, arsenite and ethanol treatments, the synthesis of a protein with a molecular weight of 68 kD (heat shock protein 68) was predominantly elevated; arsenite and ethanol exerted their effects on heat shock protein synthesis in a dose-dependent manner. Hepatocytes derived from livers of chronically griseofulvin-pretreated mice differed in their response from normal hepatocytes in that ethanol was ineffective in these cells. These results indicate that different modes and pathways of the stress response exist, depending on the nature of the inducing agent but also on pretreatment conditions. In vivo, pathologic alterations of cells and organs (e.g., in the course of chronic diseases) can, therefore, be expected to modulate the stress response.

Animals↗