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H J Gabius

Publications and source records attributed to H J Gabius.

At least 253 records · Page 14Linked to original sources
PubMed1990

Sugar receptors of the stromal cell layer in human long-term bone marrow cultures: their presence, modulatory responses to changes in the microenvironment and potential role in cellular adhesion.

Intimate cellular contacts and coordinated supply of regulatory factors are required to maintain the still inexplicable dynamic equilibrium of hemopoiesis. To infer the potential participation of protein-carbohydrate interaction in this complex process, human long-term bone marrow cultures were initiated from eleven donors, and the adherent cell layer was characterized enzyme- and immunohistochemically. Utilizing an array of carrier-immobilized carbohydrate ligands and sulfated polysaccharides as probes, specific binding of various constituents of the carbohydrate chains of cellular glycoconjugates to the stromal cells was unmistakably disclosed. Biochemical analysis, employing glycocytologically effective ligands in affinity chromatography, corroborated this result. The extent of binding was markedly lower in the two samples, derived from leukemia patients. Pronounced adaptive responses for this characteristic followed changes in the culture microenvironment that are known to influence qualitative and quantitative aspects of hemopoiesis in vitro, namely omission of hydrocortisone and horse serum or addition of cytokines. Similarly, such adaptive modulation occurred on the level of accessible cell surface receptors, monitored by neoglycoenzymes. These binding sites can be involved in mediation of cellular interactions, as revealed in a model system by the interference of N-acetyl-D-galactosamine in cell adhesion. Overall, the results support the idea that glycobiological recognition may contribute to the functional integrity of the stromal cell layer as well as provide the basis for further analysis.

Bone Marrow↗
PubMed1990

Are there characteristic alterations in lung tissue associated with Crohn's disease?

Two male patients aged 12 and 31 years suffered from Crohn's disease for more than six years and were treated with Cortison for more than four years. Surgical excision of parts of the terminal ileum was performed in both patients. They suffered from pulmonary symptoms as dyspnoea, shortness of breath and ventilation disturbances two years after operation. Wedge biopsies of the lungs revealed the following histomorphological findings: 1. Granulomatous interstitial lymphocyte infiltrates 2. Acute alveolitis with severe dysplasia of pneumocytes 3. Moderate interstitial fibrosis. Immunohistology performed in one case showed predominantly lambda chains expressed by lymphocytes associated with IgA and IgM. IgG was missing, furthermore kappa chains could not be detected. Macrophages contained endogenous lectins (sugar receptors) for fucose, maltose, and N-acetyl-D-glucosamine (glcNAc). No receptors specific for mannose, lactose, and heparin could be found. Pneumocytes did not bind the neoglycoproteins but were found to express HLA-DR receptors detectable by the monoclonal antibody LN 3 in dysplastic pneumocytes only. The histomorphological and immunohistochemical findings suggest that the analyzed alterations of lung tissue are related to the underlying disease of enteritis regionalis.

Adolescent↗
PubMed1990

Reduced expression of mannose-specific receptors on murine peripheral blood polymorphonuclear leukocytes following prolonged anaesthesia with different inhalation agents.

Inhalation anaesthetic agents are known to depress phagocytic functions such as mobilization, attachment, chemotactic motility, engulfment and intracellular killing. Mannose-specific sugar receptors on the surface of leukocytes are involved in a series of phagocytosis-related activities. To investigate the effect of anaesthesia on the expression of this type of sugar receptor, mice were anaesthetized with halothane, enflurane and isoflurane. The presence of mannose-binding receptors on peripheral blood polymorphonuclear leukocytes was examined glycocytochemically using the biotinylated neoglycoprotein mannosylated bovine serum albumin. Prolonged administration of inhalation anaesthetic agents, especially halothane, markedly depressed expression of mannose-specific receptors. This reduction may possibly contribute to postoperative immunodepression, resulting from the impaired cellular interaction which is involved in the phagocytic function of granulocytes.

Bupivacaine↗
PubMed1990

Binding of T-antigen-bearing neoglycoprotein and peanut agglutinin to cultured tumor cells and breast carcinomas.

Chemical conjugation of appropriate carbohydrate ligands to an inert labeled carrier renders probes available to screen for the presence of respective binding sites. A set with a certain plant lectin and a suitable neoglycoprotein can thus determine complementary parts of a potentially relevant glycobiological interaction system. Owing to the interest in the peanut agglutinin-reactive T-antigen, we performed chemical synthesis of the respective disaccharide structure to serve as glycohistochemical ligand and established refinements of the synthetic patway. Coupling of the derivatized monomers had to be performed in the presence of sodium sulfate for optimal results. Complete removal of the protective groups from the p-nitrophenyl derivative of the N-acetylgalactosamine moiety was achieved under mild conditions with 2,3-dichloro-5,6-dicyanobenzoquinone without affecting any other functional groups. Specific binding sites for the synthetic neoglycoprotein as well as for the plant lectin were demonstrated in cell lines of human breast carcinoma colon adenocarcinoma, and erythroleukemia. ABC reagents in conjunction with DAB as peroxidase substrate were used to visualize specific binding sites. Binding complied with the accepted criteria for specificity. Moreover, carbohydrate-specific binding sites were detected in sections of nine out of 14 cases with malignant breast lesions. The percentage of positive tumor cells with both neoglycoprotein and lectin was similar in each of the individual sections, regardless of quantitative variations between cases, lectin staining intensity often being more pronounced. The reactivity pattern in sections of primary and metastatic lesions was not significantly correlated with the lymph node status. This study emphasized that custom synthesis of saccharides and histochemical application of the resulting neoglycoprotein has a remarkable potential for complementary assessment of endogenous binding sites for carbohydrate structures, localized by external tools such as plant lectins, as a step to elucidate the importance of a putative proteincarbohydrate interaction.

Adenocarcinoma↗
PubMed1990

Tissue distribution of radioiodinated neoglycoproteins and mammalian lectins.

Quantitation of tissue distribution of radioiodinated neoglycoproteins 1 h after intravenous injection into mice allowed to evaluate their suitability to uncover potential selectivity in tracer retention. Variations within the panel of neoglycoproteins were introduced to the carbohydrate determinant, its density and linkage to the carrier. Five arrays of neoglycoproteins, encompassing up to twelve different carbohydrate moieties were used. The individual response on the level of organ content showed differences, accounted for by carbohydrate structure and density. However, increase in sugar density eventually caused general decrease in tissue retention, emphasizing the importance of synthetic parameters. Attachment of sugar residues to the spacer via primarily the C-6 group of monosaccharides led to rather prolonged survival in circulation of the resulting neoglycoprotein compared to the application of neoglycoproteins with p-aminophenyl glycosides as derivatives for coupling. Besides applying neoglycoproteins tissue uptake was also measured for several organs, when four mammalian lectins were employed as radiotracers. These lectins bind to cellular carbohydrate ligands, namely beta-galactosides, alpha-fucosides or heparin. Differences were measured for retention in liver, kidneys, spleen, stomach, thymus and bone marrow. The distinct properties of different tissues with respect to binding of neoglycoproteins as well as to endogenous lectins, exhibiting a certain degree of selectivity, are a step within the framework to attempt to therapeutically exploit the carrier potential of probes by recognitive protein-carbohydrate interactions.

Animals↗
PubMed1990

Endogenous lectins with specificity to beta-galactosides and alpha- or beta-N-acetyl-galactosaminides in human breast cancer. Their glycohistochemical detection in tissue sections by synthetically different types of neoglycoproteins, their quantitation on cultured cells by neoglycoenzymes and their usefulness as targets in lectin-mediated phototherapy in vitro.

Endogenous lectins may augment the panel of tumor markers. Specific protein-carbohydrate interactions especially involve carbohydrate moieties that are located at sequence termini, e.g. D-galactose and N-acetyl-D-galactosamine. Respective endogenous lectins can be detected by suitably constructed neoglycoproteins. In order to evaluate the influence of sugar and label density as well as coupling mode of the carbohydrate moiety to the carrier protein for lectin localization in histopathology, four different types of neoglycoproteins, carrying beta-galactosides or alpha- and beta-anomers of N-acetyl-D-galactosamine were employed to reveal the presence of specific receptors in invasive ductal mammary carcinomas with propensity for metastasis formation. Staining of tumor cells was more intense than staining of normal cell types. Coupling of the diazo derivatives of p-aminophenyl glycosides led in most cases to the relatively highest extent of staining in terms of number of stained cells and staining intensity. Classified next according to these categories attachment of sugars via p-isothiocyanato derivatives or via an aliphatic linker after his reaction with the C6-hydroxyl group of the sugar moiety was rather equally well effective, whereas reductive amination with concomitant ring opening at the reducing end of the disaccharide lactose resulted in neoglycoproteins, yielding the lowest extent of staining. The alpha-anomer is preferred as a ligand to endogenous lectins of tumor cells to the beta-anomer of N-acetyl-D-galactosamine. To reduce the number of steps in glycohistochemical processing, glycosylated enzymes were successfully employed. They also allowed to measure the lectin density on breast carcinoma cells, leading to rational selection for demonstrated lectin-mediated targeting of neoglycoprotein-hematoporphyrin conjugates. Immobilization of ligands as an approach to prepare histochemically valuable reagents to localize respective receptors is not confined to tumor lectinology, as emphasized by additional application of hormone-protein conjugates, termed neohormoproteins.

Acetylgalactosamine↗
PubMed1990

Differentiation-related expression of epidermal growth factor receptors in human lung carcinoma demonstrated histochemically by biotinylated epidermal growth factor.

Biotin-labeled epidermal growth factor (EGF) was applied to routinely processed sections of 64 cases of human lung carcinoma as a histochemical tool for demonstrating EGF-specific receptors. Formalin-fixed, paraffin-embedded tissue sections were deparaffinized and incubated with the labeled EGF (10 micrograms/ml) for 60 min at room temperature. The specific binding of the growth factor to its receptor was visualized by the avidin-biotin complex (ABC) technique. Positive binding capacities were obtained for the following cases: 15/16 epidermoid carcinoma; 13/15 adenocarcinoma; 2/11 large cell anaplastic carcinoma; 12/20 small cell anaplastic carcinoma; 0/11 normal lung tissue; 0/6 main bronchi; 0/1 hamartoma; 0/1 primary fibrosarcoma of lung. In addition, a strong positive reaction was seen for neutrophilic granulocytes present within the tumorous tissue. Data indicate that EGF receptors are frequently expressed in more differentiated carcinoma in comparison with anaplastic carcinoma of lung.

Aged↗
PubMed1990

Detection of binding sites for biotinylated neoglycoproteins and heparin (endogenous lectins) during cerebellar ontogenesis in the rat.

Endogenous carbohydrate-binding sites were studied during rat cerebellar development on sections of fixed tissue using synthetic tools, biotinylated neoglycoproteins, in conjunction with subsequent avidinperoxidase staining. Neoglycoproteins were constructed by chemically coupling the histochemically pivotal carbohydrate moieties to an inert carrier protein. The sugar part of the neoglycoproteins included common constituents of the carbohydrate part of cellular glycoconjugates, namely mannose, galactose, fucose, N-acetyl-glucosamine, N-acetylgalactosamine and N-acetyl-neuraminic acid to probe for the presence of respective endogenous receptors. Heparin was biotinylated after mild cyanogen bromide activation and aminoalkylation. Specific positive reactions were obtained for all neoglycoproteins and heparin. The staining pattern with the individual probes disclosed variable developmental regulation. Consequently, these results suggest that recognition processes during cerebellar development may include several types of carbohydrate determinants. In two instances, the binding of neoglycoproteins could be compared to endogenous lectin-specific antibodies. Despite a significant extent of accordance the comparison revealed notable differences. These differences were attributed primarily to fixation and the presence of physiological ligands that can mask the active endogenous carbohydrate-binding proteins. In any case, histochemical application of labeled neoglycoproteins is valuable to discern the presence, localization and developmental pattern of binding sites for the carbohydrate part of glycoconjugates, on which further biochemical and cell biological studies can consequently be based.

Acetylgalactosamine↗
PubMed1990

[An evaluation of endogenous lectins in the intrapulmonary metastases of osteogenic sarcoma].

Assessment is made of the presence or absence of specific sugar receptors (lectins) in patients operated for intrapulmonary metastases of osteogenic sarcoma. Investigations were carried out on histochemically stained paraffin sections for the separate types of sugars. Highest was the presence of specific receptors for sialic acid and for N-acetylglucosamine, followed by those for ramnose and heparin and least for mannose and maltose. Lectin determination is a new branch in morphological science, assisting clinical practice, special surgical activity by the attempt for quantitative grading of cell populations in norm and in their neoplastic development.

Female↗
PubMed1989

Purification and properties of a Ca2+-independent sialic acid-binding lectin from human placenta with preferential affinity to O-acetylsialic acids.

A Ca2+-independent sialic acid-specific lectin from two developmental stages of human placenta was similarly purified to apparent homogeneity by DEAE-cellulose chromatography, affinity chromatography on bovine submaxillary mucin, and gel filtration. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis and gel filtration disclosed a molecular mass of 53 kDa. The specificity of the lectin for O-acetylsialic acids was substantiated by the dependence of hemagglutination on the presence of acetylated sialic acids on the surface of mammalian erythrocytes of various sources, by hapten inhibition in hemagglutination assays with protease-treated rabbit erythrocytes and by hapten inhibition of binding of labeled N-acetylneuraminic acid-bovine serum albumin to the lectin in a solid-phase assay. Bovine and equine submaxillary mucins that contain 9(7,8)-O-acetyl and 4-O-acetylsialic acids were potent inhibitors in contrast to the non-acetylated sialic acids of ovine submaxillary mucin. Absence of inhibitory efficiency of other negatively charged substances like phosphorylated sugars, glucuronic acid, heparin, or oligodeoxynucleotides emphasized the importance of structural features instead of simple ionic interaction. In the presence of acetylation, the pattern of inhibition by gangliosides in the solid-phase assay indicated a preference to alpha-2,8- or alpha-2,6-linked sialic acids in comparison to alpha-2,3-linked moieties. Chemical modification of the lectin by group-specific reagents allowed to emphasize the role of primarily lysine residues, but also, although less pronounced, arginine, tryptophan, and carboxyl groups for ligand binding and/or maintenance of the active conformational state. Application of reagents, specific for histidine or tyrosine residues, failed to affect lectin activity.

Acetylation↗
PubMed1989

Neoglycoenzymes: a versatile tool for lectin detection in solid-phase assays and glycohistochemistry.

Carbodiimide-mediated coupling of p-aminophenyl glycosides to a naturally nonglycosylated enzyme yields a neoglycoenzyme. This compound combines inherent enzymatic activity with synthetically conferred ligand properties to lectins. Appropriate choice of the ligand allows custom-made synthesis to reliably detect various types of lectins. To exemplify practical applications of this class of compounds, glycosylated bacterial beta-galactosidase has been employed to quantitate plant lectins, immobilized on plastic surfaces as well as on nitrocellulose. Competitive inhibition by specific sugar ascertained the dependence of binding on protein--carbohydrate interactions. In view of lectins as tools, a sandwich lectin-binding assay for high mannose-type glycoprotein detection has been modified to principally facilitate wide application to other lectin-reactive sugar chains by introducing the neoglycoenzyme. In addition to lectin determination in solid-phase assays, neoglycoenzymes allow one to glycohistochemically localize endogenous lectins in tissue prints and tissue sections with a minimum number of steps. This nonradioactive, rapid, sensitive, and convenient assay concept, based on conjugation of a ligand to an enzyme with maintenance of its receptor-binding activity, may find extended application beyond lectinology in receptor analysis.

Biotin↗
PubMed1989

Site-associated expression of endogenous tumor lectins.

The expression of endogenous lectins was compared in mouse Lewis lung carcinomas growing in the kidney or subcutaneously. A panel of carbohydrates coupled to a biotinylated carrier molecule (bovine serum albumin), biotinylated desialylated glycoproteins and sulfated polysaccharides were used in histochemical assays to detect the presence and distribution of carbohydrate receptors. Heterogeneous staining patterns were observed with most carbohydrates in sections of tumor tissue from both anatomic sites, and staining intensities also varied within each section. At least 2 populations of cells were identified at each site, of which one had receptors for all carbohydrates, while the other had no receptors for melibiose, sialic acid, or alpha-glucosides (maltose and glucose). There were quantitative differences in expression of endogenous lectins by tumors growing s.c. or in the kidney; 3LL cells growing in the kidney bound 6 out of 10 carbohydrates to a greater extent than 3LL cells in s.c. tumors. Conversely, 3LL cells in s.c. tumors bound heparin and asialofetuin to greater extents than cancer cells in kidney tumors. Biochemical analyses of detergent extracts of tissues subjected to affinity chromatography and subsequent SDS-PAGE revealed quantitative and also qualitative differences in lectins between tumors growing in the 2 anatomic sites.

Animals↗
PubMed1989

Modulatory potency of the beta-galactoside-specific lectin from mistletoe extract (Iscador) on the host defense system in vivo in rabbits and patients.

Proprietary extract of mistletoe (Iscador) that has federal approval for clinical application can exhibit immunomodulatory capacity. However, the nature of this responsible substance has still remained elusive. To validate the hypothesis that specific lectin-carbohydrate interactions at least participate in eliciting immunomodulation, the modulatory efficiency of the major beta-galactoside-specific mistletoe lectin (ML I) from the clinically applied extract on selected immunological parameters was monitored "in vivo" in rabbits. Injections of nontoxic doses of the purified lectin or even only of its carbohydrate-binding subunit (0.25-1.0 ng/kg) into rabbits yielded significant increases in natural killer cytotoxicity, frequency of large granular lymphocytes, and phagocytic activity of granulocytes. In the clinically relevant situation, changes in these parameters were also determined in cancer patients after extract (Iscador) injection s.c. as well as i.v., emphasizing the potential relevance of the lectin. Comparative analyses of the changes in the selected parameters following injection of extract with normal lectin content as well as of extract, selectively depleted of lectin, into healthy volunteers corroborated this inference. Lectin depletion by affinity chromatography was highly specific and did not affect any other substance in the extract. Remarkably, contamination by endotoxin has been rigorously excluded in each applied specimen. These results encourage detailed elucidation of lectin action on various parts of the tumor defense system "in vitro" with the long range goal of achieving progress in the treatment of cancer through immunological strategies, exploring selective mediatory lectin-carbohydrate interactions.

Adjuvants, Immunologic↗
PubMed1989

Identification of a cell cycle-dependent gene product as a sialic acid-binding protein.

A Ca2+-dependent sialic acid-binding protein was purified on fetuin-Sepharose from various types of human tissue. The molecular mass was determined to be 10,315 Da by laser desorption mass spectrometry. Partial sequence analysis after cyanogen bromide cleavage that yielded one N-terminus accessible for Edman degradation revealed an identity to an internal stretch following the only methionine residue within a putative amino acid sequence (Mr 10,048), deduced from the cDNA of a cell cycle-specific gene. The reported biochemical identification is a prerequisite to infer the biological role of the so far undetected gene product. Initial glycohistochemical studies with sialic acid-(BSA-biotin) raised evidence for nuclear localization of sialic acid-binding sites that might reflect, at least in part, detection of this protein.

Amino Acid Sequence↗
PubMed1989

Heparin-binding lectin from human placenta: purification and partial molecular characterization and its relationship to basic fibroblast growth factors.

The heparin-binding lectin from human placenta is isolated on the basis of its tendency to form large aggregates by gel filtration and on the basis of its affinity for heparin by affinity chromatography. The purified lectin dissociates into up to four distinct polypeptides with molecular weight values of 14,400, 15,000, 16,200, and 16,700 and a single isoelectric point of 9.0. Molecular heterogeneity is not due to different degrees of glycosylation, as evidenced by gel electrophoretic analysis after extensive treatment with various endoglycosidases. Despite its similarities of affinity to heparin, molecular size, and isoelectric point to the basic fibroblast growth factor (bFGF), the comparatively high yield of the lectin (approximately 1.5 mg/100 g of placenta), the occurrence of proteolytic fragmentation in the presence of heparin, and the lack of homology to the amino-terminal sequence of the lectin argue against any notable relationship to bFGF. Most importantly, the lack of mitogenic activity in a commonly used bioassay with quiescent 3T3 fibroblasts rules out any FGF-like activity on cell proliferation. The heparin-binding lectin is thus clearly distinguishable from heparin-binding growth factors. By employing biotinylated heparin as labeled ligand to visualize and quantify heparin binding, hapten inhibition in a solid-phase assay reveals that except for heparin no other vertebrate glycosaminoglycan but the sulfated fucan fucoidan can effectively reduce the Ca2+-independent ligand binding. Proteolytic fragmentation by chymotrypsin in two independent assays demonstrates that a fragment of Mr 7800 still retains ability to bind heparin. The interaction of this lectin with naturally occurring heparin-like molecules may physiologically be involved in modulatory regulation of heparin-mediated processes.

Amino Acid Sequence↗
PubMed1989

Is part of the molecular basis of the perineurial barrier function the lack of endogenous carbohydrate-binding proteins?

The sugar part of cellular glycoconjugates and specific endogenous sugar receptors, i.e., lectins, can establish a system of biological recognition based on protein-carbohydrate interactions. An assortment of labelled (neo)glycoproteins, carrying different types of sugar moieties, is synthesized to localize respective sugar receptors. With these tools, the histochemical patterns of endogenous carbohydrate-binding receptors of the epi-, peri-, and endoneurium were analyzed in human sural and accessory nerves and in swine sciatic nerve. This approach is complementary to the application of plant lectins, focusing on endogenous carbohydrate-binding proteins (lectins). In contrast to the epi- and endoneurium, which bound certain types of carbohydrates, such endogenous sugar receptors were histochemically not detectable in the perineurial cells. Moreover, no histochemical reaction was present in the "connective tissue septa" localized in the endoneurium in which the endoneurial vessels were embedded. This common property supplies evidence that these septa are composed of perineurial cells. They may represent a barrier in addition to the capillary endothelium. Our observations suggest histogenetical differences between the cell populations of epi- and endoneurium vs. perineurium. This significant difference in the ability to bind carbohydrate residues, conjugated to a carrier protein, is contradictory to the assumption that perineurial cells and fibroblasts are functional variants of the same cell type. The histochemical patterns of endogenous carbohydrate-binding receptors found in human and swine nerves were similar but not identical, with exception of the perineurium, reflecting phylogenetic differences in the expression of sugar-binding proteins. The absence of specific sugar receptors in perineurial cells, however, seems to be a more general phenomenon.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗