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PubMed1986

Neoplastic angioendotheliomatosis: immunopathologic and morphologic evidence for intravascular malignant lymphomatosis.

Neoplastic angioendotheliomatosis (NAE) is a rare entity characterized by multifocal, intravascular proliferation of large pleomorphic cells within small vessels of most organs, with a particular affinity for the central nervous system. Clinically, patients with NAE present with focal neurologic signs and a progressive decline in mental status, followed by death in a few months. The histogenesis of NAE is controversial but has been previously thought to represent a malignant proliferation of endothelial cells. Three autopsy cases with clinical and histologic features of NAE were investigated by electron microscopic, standard histochemical, and immunohistochemical technics that included the use of three panleukocyte monoclonal antibodies (PLA), the endothelial-cell-specific reagents, FVIII-RAG anti-sera and Ulex europaeus agglutinin (UEA), and muramidase. The NAE cells in all three cases were stained positively by the PLA, whereas the adjacent endothelial cells and not the NAE cells were stained by FVIII-RAG and UEA. Muramidase by immunoperoxidase technic and nonspecific esterase (chloracetate) were not present in NAE cells. These results indicate that NAE is a leukocyte-derived neoplasm and not a malignant endothelial cell neoplasm. Based on these findings and on a review of the literature, it is proposed that NAE represents intravascular malignant lymphomatosis (IML). IML appears to be a primary manifestation and/or a major secondary form of disseminated malignant lymphoma. This would explain the spectrum of findings in previously reported cases.

Adult↗
PubMed2003

Characterization of AcmB, an N-acetylglucosaminidase autolysin from Lactococcus lactis.

A gene encoding a putative peptidoglycan hydrolase, named acmB, which is a paralogue of the major autolysin acmA gene, was identified in the Lactococcus lactis genome sequence. The acmB gene is transcribed in L. lactis MG1363 and its expression is modulated during cellular growth. The encoded AcmB protein has a modular structure with three domains: an N-terminal domain, especially rich in Ser, Thr, Pro and Asn residues, resembling a cell-wall-associated domain; a central domain homologous to the Enterococcus hirae muramidase catalytic domain; and a C-terminal domain of unknown function. A recombinant AcmB derivative, devoid of its N-terminal domain, was expressed in Escherichia coli. It exhibited hydrolysing activity on the peptidoglycan of several Gram-positive bacteria, including L. lactis. Though showing sequence similarity with enterococcal muramidase, AcmB has N-acetylglucosaminidase specificity. The acmB gene was inactivated in order to evaluate the role of the enzyme. AcmB does not appear to be involved in cell separation but contributes to cellular autolysis.

Acetylglucosaminidase↗
PubMed1993

The use of monoclonal antibodies for studying the biological properties of Staphylococcus aureus endo-beta-N-acetylglucosaminidase.

Staphylococcus aureus endo-beta-N-acetylglucosaminidase (SaG) has been suggested to function as a virulence determinant which interferes with the host cellular immune response. To further characterize the biological properties of SaG, monoclonal antibodies (mAbs) were raised against purified SaG. Four IgG1 subclass mAbs were obtained, none of which reacted with the reduced, sodium dodecyl sulphate pretreated or boiled enzyme. The ability of the mAbs to react with the enzymes present in supernatants obtained from 197 S. aureus strains indicated that they recognized epitopes which are highly conserved; bacteriolytic enzymes produced by staphylococci other than S. aureus did not show any cross-reactivity. After pretreatment of SaG with mAbs (mAb-SaG molar ratios varying from 1 to 20), it was shown that all selected mAbs caused, at a mAb:SaG molar ratio of 10, a 90% inhibition of SaG bacteriolytic activity and a statistically significant reduction of its ability to interfere with phagocytosis by human polymorphonuclear leukocytes. All selected mAbs reacted with several commercially available exo-beta-N-acetylglucosaminidases; mAb C1/10-11 also reacted with chicken and turkey egg muramidases and, at a mAb:SaG molar ratio of 10, inhibited their bacteriolytic activity by 97%. This suggests that one or more epitopes present in the above exo-glucosaminidases and muramidases share some degree of homology with others present in SaG.

Acetylglucosaminidase↗
PubMed1981

Effect of fructose and other carbohydrates on the surface properties, lipoteichoic acid production, and extracellular proteins of Streptococcus mutans Ingbritt grown in continuous culture.

Streptococcus mutans Ingbritt was grown in a chemostat at destined dilution rates in either 0.5% fructose or 0.5% sorbitol and at destined pH values in 0.5% fructose. The yield of cells was affected by the carbohydrate source, as well as by the pH, with the lowest yield being at pH 5.5 in 0.5% fructose. Fructose-grown cells showed greater susceptibility to lysis by a muramidase than the corresponding glucose-grown cells, but there were no marked differences in the lytic susceptibilities of the corresponding cell wall preparations or in the serological reactivities of wall lysates with antiserum to S. mutans Ingbritt. The greatest amounts of cellular lipoteichoic acid were obtained at high dilution rates in both fructose and sorbitol, as well as at high pH values in fructose. The greatest amounts of extracellular lipoteichoic acid were found at low dilution rates, as estimated by rocket immunoelectrophoresis and also by hemagglutination. Three major extracellular protein components were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and the effects of growth conditions on these components were determined. Results for batch-grown cultures showed that there was genotypic variation in the susceptibility of cells to lysis by a muramidase. The enhancement of lipoteichoic acid production by fructose and sorbitol in batch cultures was not identical in representative strains of S. mutans serotype c, nor was the effect of fructose found uniformly in representative strains of the different S. mutans serotypes.

Bacterial Proteins↗
PubMed1991

Analysis of murein and murein precursors during antibiotic-induced lysis of Escherichia coli.

Lysis of Escherichia coli induced by either D-cycloserine, moenomycin, or penicillin G was monitored by studying murein metabolism. The levels of the soluble murein precursor UDP-N-acetylmuramyl-L-alanyl-D-glutamyl-m-diaminopimelyl-D-alanyl- D-alanine (UDP-MurNAc-pentapeptide) and the carrier-linked MurNAc-(pentapeptide)-pyrophosphoryl-undecaprenol as well as N-acetylglucosamine-beta-1,4-MurNAc-(pentapeptide)-pyrophosphoryl- undecaprenol varied in a specific way. In the presence of penicillin, which is known to interfere with the cross-linking of murein, the concentration of the lipid-linked precursors unexpectedly decreased before the onset of lysis, although the level of UDP-MurNAc-pentapeptide remained normal. In the case of moenomycin, which specifically blocks the formation of the murein polysaccharide strands, the lipid-linked precursors as well as UDP-MurNAc-pentapeptide accumulated as was expected. D-Cycloserine, which inhibits the biosynthesis of UDP-MurNAc-pentapeptide, consequently caused a decrease in all three precursors. The muropeptide composition of the murein showed general changes such as an increase in the unusual DL-cross bridge between two neighboring meso-diaminopimelic acid residues and, as a result of uncontrolled DL- and DD-carboxypeptidase activity, an increase in tripeptidyl and a decrease in tetrapeptidyl and pentapeptidyl moieties. The average length of the glycan strands decreased. When the glycan strands were fractionated according to length, a dramatic increase in the amount of single disaccharide units was observed not only in the presence of penicillin but also in the presence of moenomycin. This result is explained by the action of an exo-muramidase, such as the lytic transglycosylases present in E. coli. It is proposed that antibiotic-induced bacteriolysis is the result of a zipperlike splitting of the murein net by exo-muramidases locally restricted to the equatorial zone of the cell.

Bacteriolysis↗
PubMed1985

Meningeal tumors of infancy and childhood.

Seventeen meningeal tumors were examined for pathology with electron microscopy and immunohistochemistry including glial fibrillary acidic protein (GFAP), S-100 protein, muramidase, and factor VIII. These tumors included seven meningiomas, one hemangiopericytoma, three meningeal sarcomas (1 pleomorphic-cell type and 2 myxofibrosarcomas), two fibrous histiocytomas, and four malignant melanomas. Two of seven children with meningioma had a poor outcome despite the benign histological features of the tumor. S-100 protein was present in the two tumors. All three children with meningeal sarcoma had a rapid downhill clinical course, although the myxofibrosarcoma initially had a relatively benign histological appearance. The two children with fibrous histiocytoma did well despite the aggressive histological features. Muramidase was a good marker of such tumors. Because of the morphological difficulties associated with childhood meningeal tumors, both electron microscopy and immunohistochemistry can be of diagnostic benefit.

Adolescent↗
PubMed1984

Sural nerve biopsy in leprosy patients after varying periods of treatment: histopathological and bacteriological findings on light microscopy.

As part of a larger study of nerve biopsies from leprosy patients in Ethiopia for the presence of muramidase (lysozyme), sections were also examined by light microscopy after staining with hematoxylin and eosin for cellular infiltrate and a modification of the Ziehl-Neelsen stain for leprosy bacilli. The muramidase findings will be reported separately. This paper describes the infiltrative and bacterial findings in a group of 18 patients, including four with nonlepromatous forms of leprosy who were suffering from delayed hypersensitivity reaction at the time of biopsy. The findings were unexpectedly interesting and revealing. Lepromatous and borderline-lepromatous patients all showed endoneurial and perineurial infiltration of considerable extent and, in several instances, bacilli were wide-spread from one end of the biopsy to the other; in two patients, solid-staining bacilli and globi were found, indicating relapse. In all except two of the nonlepromatous patients (mainly borderline-tuberculoid) there was an extensive and severe granulomatous infiltration, and in one case there was marked caseation in the endoneurial zone. Within the limits of the present study, the findings indicate that biopsy of a peripheral nerve, even when it is not obviously associated with a skin lesion, may reveal pathological changes which are greater in degree than those suggested by skin biopsy or clinical examination. These observations in a somewhat heterogeneous group of patients treated for varying periods of time, and in a study which was not prospectively planned, suggest that similar observations in a larger group of untreated and treated patients, including those who have relapsed, may be of value.

Adolescent↗
PubMed1997

Reevaluation of the effect of lysoyzme on Escherichia coli employing ultrarapid freezing followed by cryoelectronmicroscopy or freeze substitution.

Lysozyme is able to lyse Gram-positive bacteria acting as muramidase on the peptidoglycan polymer. Gram-negative bacteria in vitro are not lysed by lysozyme. It was assumed that the peptido-glycan is protected by the outer membrane and thus that Gram-negative bacteria are not affected by lysozyme without the aid of other factors such as EDTA or complement which enable lysozyme to penetrate the outer membrane. Accidentally, Pellegrini et al. [(1992) J. Appl. Bacteriol., 72:180-187] found that lysozyme per se is able to kill some Gram-negative bacteria. On the basis of morphological and immunocytochemical findings obtained from chemically fixed bacteria, it was concluded that lysozyme does not lyse Gram-negative bacteria but affects the cytoplasm of for example, Escherichia coli, leading to its disintegration, whilst the membranes do not break down. In an attempt to clarify the action of lysozyme on E. coli, we employed cryotechniques including ultrarapid freezing, cryomicroscopy and freeze substitution, and immunolabeling. Bacteria that were immediately frozen after exposure to lysozyme remained morphologically intact. Individual bacteria plated on agar after exposure to lysozyme were mostly intact when frozen within a few seconds. However, inner and outer membranes of 80% of the bacteria were disrupted, whereas the cytoplasm of only a few bacteria showed signs of disintegration when bacteria were frozen with a delay of only 5 min of plating onto pure agar or agar containing growth medium. After a period of time of 15 min between plating onto agar and freezing, about 97% of the bacteria showed changes of disintegration of various extent. Immunolabeling showed that lysozyme binds to the outer cell membrane and may penetrate the membrane, reaching the periplasmic space and possibly the inner cell membrane. The ultrastructural findings and the results of antibacterial assays suggest that lysozyme is bactericidal for E. coli but is not able to induce disintegration. Disintegration is accomplished by changes of the environment starting at the cell membranes. The mechanism by which lysozyme penetrates the membrane, the way it acts to be bactericidal, and the way disintegration is initiated remain to be clarified.

Escherichia coli↗
PubMed1981

Neoplastic Paneth cells in adenocarcinoma of the urinary bladder: a first case report.

A case of a highly differentiated, primary, nonurachal adenocarcinoma of intestinal type, originating from the urinary bladder mucosa, is reported. The tumor contained Paneth cells as an integrated part, and their identity on the light microscopic level was confirmed by histochemical stains including immunohistochemical stain for muramidase. A gradual change from Paneth cell types to mucous cells was seen. Argentaffin cells and goblet cells appeared in some parts of the tumor but no detectable transitional forms were found. Only a few carcinomas containing malignant Paneth cells have been recorded, and this case is believed to be the first observed in an extraintestinal site.

Adenocarcinoma↗
PubMed1983

Immunohistochemistry of Hodgkin's disease. A study of 20 cases.

An immunoperoxidase study of 20 cases of Hodgkin's disease demonstrated universal staining of Reed Sternberg cells and their mononuclear variants for both kappa and lambda light chains and, in all but one case, for IgG. Staining for IgA and albumin was variable and for IgD and IgM uniformly negative. A double staining procedure using two different chromogens produced the paradoxical finding of both light chain types within the same cell, but these could only be demonstrated sequentially and not simultaneously, suggesting a blocking phenomenon. The above findings coupled with the demonstration of muramidase and/or alpha-1-antitrypsin in Reed-Sternberg cells and their mononuclear variants in all but two cases studied favor a histiocytic origin for these cells. This characteristic profile of results is also very helpful in distinguishing Hodgkin's disease from other neoplasms which mimic Hodgkin's disease because of the presence of Reed-Sternberg-like cells.

Adolescent↗
PubMed1990

An immunohistochemical and ultrastructural study of the sinusoids of hepatocellular carcinoma.

The sinusoids of 30 human hepatocellular carcinomas of various types were examined by electron microscopy and histochemically for binding to the Ulex europaeus lectin (UEA1). A population of sinusoidal macrophages was identified with an antibody to lysozyme (muramidase). The UEA1 binding was negative in normal sinusoids but positive in the tumor vessels. Macrophages resembling Kupffer cells were found within the tumor vessels but in smaller numbers than in either normal or cirrhotic liver tissue. Fibrolamellar and sclerosing carcinomas contained the smallest numbers. Ultrastructurally, endothelial cells of tumor vessels were thicker than normal, with fewer fenestrations. They contained bundles of microfilaments and showed basement membrane formation. Subendothelial myoid cells were found. These findings indicate that the sinusoidal vessels of hepatocellular carcinomas show features of true capillaries and precapillary blood vessels. The degree of this difference from normal hepatic sinusoids may reflect the relative immaturity of the cancer cells.

Adolescent↗
PubMed1993

Structural characterization of the cyanelle peptidoglycan of Cyanophora paradoxa by 252Cf plasma desorption mass spectrometry and fast atom bombardment/tandem mass spectrometry.

A strategy for the structural characterization of the four major NaBH4-reduced peptidoglycan monomers derived from muramidase-digested peptidoglycan from the cyanelles of the flagellate Cyanophora paradoxa Korschikoff is described. Initial molecular weight determination of these glycopeptides was performed by positive and negative ion plasma desorption mass spectrometry. Due to the presence of two pairs of disaccharide tripeptide and disaccharide tetrapeptide monomers differing in mass by 112 units, respectively, an as yet unknown peptidoglycan modification either at the carbohydrate or at the peptide moiety was assumed. beta-Elimination of the disaccharide unit from the unreduced peptidoglycan monomers yielded the corresponding (modified) N1-lactyltripeptides and -tetrapeptides, respectively. These peptides, N-terminally blocked with lactic acid, unambiguously showed the modification to be located on the peptide moiety. By positive ion fast atom bombardment/hybrid tandem mass spectrometry of the reduced peptidoglycan monomers as well as of the corresponding deglycosylated monomers (= N1-lactylpeptides) the modification was determined to be linked to the glutamic acid moiety. Based on combined data from plasma desorption mass spectrometry, tandem mass spectrometry, accurate mass measurement and amino acid analysis of the acid hydrolysate after derivatization with o-phthaldialdehyde by high-performance liquid chromatography we could establish the structure of the modification as N-acetylputrescine. Finally, the confirmation of the linkage of the glutamic acid to diaminopimelic acid via the gamma-COOH was based on the presence of a-type peptide backbone fragment ions in the positive ion plasma desorption mass spectra of the modified N1-lactylpeptides.

Amino Acid Sequence↗
PubMedDate not supplied

An immunohistochemical and clinicopathological study of granulocytic sarcoma ('chloroma').

A small series of granulocytic sarcomas (GS) or 'chloromas' has been studied by means of conventional histology and immunohistochemistry. The latter was found to be most useful in the diagnosis and characterization of these neoplasms, which are rare in Great Britain. Polytypic antisera to leukocyte elastase and cathepsin G have proved to be most useful, giving intense staining in all cases. These two markers are more specific than muramidase (lysozyme) and alpha 1-antitrypsin, since, unlike the latter two, they do not stain histiocytic cells. The results of staining with two monoclonal antisera, AGF 4.48 and AGF 4.36, are also described. In addition, the clinicopathological details of the seven cases are summarized and the literature is briefly surveyed.

Adult↗
PubMed1981

The enzyme histochemistry of lymphoid and non-lymphoid cells of the human palatine tonsil: a basis for the study of lymphomas.

The distribution of various hydrolytic enzymes has been determined in 27 human palatine tonsils by means of conventional enzyme histochemical techniques, and lysozyme (muramidase) activity has been localised in eight tonsils by the unlabelled antibody peroxidase-antiperoxidase complex (PAP) method. The enzyme activities of various cell types are compared and the effect of various methods of fixation and processing discussed. The results suggest that arrangement of various histiocytic cells types within the tonsillar follicles and crypt epithelium is related to the processing of antigen. The PAP method for lysozyme demonstrates a smaller population of cells than is demonstrated by the alpha-naphthyl acetate esterase (ANAE) method. T-cells are demonstrated by the presence of dot-like ANAE activity in their cytoplasm. Large numbers of the lymphocytes of this type were located in the T-dependent areas of the tonsil, and are frequent beneath the crypt epithelium. The efferent lymphatic vessels appeared to contain an almost pure population of T-cells. The immunohistochemical method for lysozyme did not differentiate between T- and B-cell areas, dot-like activity being absent. As in other workers' studies on non-lymphoid cells of the murine spleen, several types of glass-adherent cells have been identified in short-term cell cultures from the human tonsil. True dendritic cells and branching macrophages differ in several ways (as in the mouse spleen). The tonsil is considered to be a useful control "reactive" lymphoid organ, to act as a baseline tissue in an extended study of morphology and enzyme histochemistry in the lymphomas.

Adolescent↗
PubMed2000

Assay for lytic transglycosylases: a family of peptidoglycan lyases.

An assay has been developed to monitor the activity of the lytic transglycosylases which does not involve the use of radiolabel. Samples of lytic transglycosylase were incubated with isolated and purified insoluble peptidoglycan as substrate for varying lengths of time. Residual insoluble material was removed by ultracentrifugation in a microfuge and the solubilized components were treated with sodium borohydride prior to acid hydrolysis. The optimal conditions for this acid hydrolysis were established to be incubation at 96 degrees C for 1 h in 6 M HCl, in vacuo. The hydrolyzed samples were subjected to amino acid/sugar analysis by cation-exchange chromatography on a Beckman System Gold amino acid analyzer. To effect a clear resolution of muramic acid from serine and glutamic acid, the equilibration buffer was modified to be composed of 33 mM sodium citrate, pH 3.12. The product of the lyase reaction of the lytic transglycosylases are 1,6-anhydromuramyl residues, which are not reduced by the sodium borohydride treatment. On the other hand, the muramyl residues arising at the reducing ends of peptidoglycan after treatment with muramidases (hydrolyases) are reduced to muramitol residues, which elute from the amino acid analyzer prior to aspartic acid. This assay thus distinguishes the activity of the two enzymes and was applied to determine the initial activities of increasing concentrations of a soluble derivative of lytic transglycosylase B from the opportunistic pathogen Pseudomonas aeruginosa.

Amino Acids↗
PubMed1991

Increased yield of a lysozyme after self-cloning of the gene in Streptomyces coelicolor "Müller".

Streptomyces coelicolor "Müller" DSM3030 excretes a lysozyme comprising both beta-1,4-N-acetyl- and beta-1,4-N,6-O-diacetyl muramidase activities. The lysozyme is named Cellosyl. Gene libraries have been established using genomic DNA from the wild-type strain, S. coelicolor DSM3030, and from an overproducing mutant, S. coelicolor HP1, which exhibits about a twofold increase in lysozyme production. The lysozyme-encoding genes (cel) from both strains were detected by oligodeoxynucleotide hybridization. The nucleotide sequence of the cel genes isolated from both strains was shown to be identical. The different levels of lysozyme production could not be correlated with any mutations at the cel gene locus. The cel gene isolated from the wild-type strain could not be expressed in some other species of Streptomyces. However, self-cloning of the cel gene into S. coelicolor DSM3030 and HP1 resulted in a 2.5-fold increase in lysozyme production.

Amino Acid Sequence↗
PubMed1990

Macrophage histology in paraffin-embedded multiple sclerosis plaques is demonstrated by the monoclonal pan-macrophage marker HAM-56: correlation with chronicity of the lesion.

Using the monoclonal antibody HAM-56 with the avidin-biotin method on recent or archival paraffin-embedded tissue from multiple sclerosis brains, we have been able to distinguish between acute, chronic active and inactive lesions. HAM-56 stains all macrophages, monocytes and at least some microglia; it is the only pan-macrophage marker to our knowledge that can be satisfactorily used on conventional paraffin sections. A much narrower range of mainly perivascular macrophages in acute plaques of multiple sclerosis is stained with MAC-387, anti-muramidase and anti-alpha1-anti-trypsin. The acute plaques show HAM-56-stained macrophages throughout the lesion, and these macrophages exhibit profiles of phospholipid-rich myelinic bodies, which are also usually stainable with Luxol fast blue. Active ongoing lesions show a rim of macrophages at the edge of the lesion. These macrophages show profiles of large vacuoles, thought to represent the sudanophilic esterified cholesterol formed during demyelination. Inactive cases show none of these features; the few perivascular macrophages present often contain the end product of lipid peroxidation, ceroidlipofuscin.

Antibodies, Monoclonal↗
PubMed1985

Immunohistochemical findings in rheumatoid nodules.

Eighteen nodules from patients with rheumatoid disease were studied histologically and immunohistochemically. A continuum of microscopic changes was observed with varying degrees of fibrinoid necrosis, mononuclear cell infiltration and healing by fibrous scarring. In two cases there was focal evidence of arteritis. Fibrin was plentiful in the necrotic areas of nodules. Small amounts of immunoglobulin were identified in plasma cells and as irregular extracellular deposits in and around areas of necrosis. In a single small vein abnormal IgG was detected. Mononuclear cells surrounding areas of necrosis stained strongly with antisera to ferritin and a cytoplasmic macrophage antigen, stained variably with muramidase (lysozyme) and negatively with alpha-1 antitrypsin antibodies. Perls' stain for ferric iron was almost entirely negative and ultrastructural x-ray microanalysis indicated that the cytoplasm of these cells were entirely free of iron. These findings confirm the chronic inflammatory nature of rheumatoid nodules but provide no support for the view that they originate in areas of vasculitis. A relative lack of cytoplasmic antiprotease along with a strong expression of ferritin appears to be a characteristic feature of macrophages in rheumatoid tissue.

Adult↗