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Journal ArticleDOI

Expression of M-N#1, a histo-blood group B–like antigen, is strongly up-regulated in nonapoptosing mammary epithelial cells during rat mammary gland involution

01 Jun 2001-Glycobiology (Oxford University Press)-Vol. 11, Iss: 6, pp 441-449
TL;DR: It is shown that up-regulation of carbohydrate antigens is not a general phenomenon during mammary gland involution, and thus that M-N#1 antigen expression is specifically regulated, and that alpha(1,2)fucosyltransferase A, an enzyme required for M-M-N-1 antigen synthesis, is at least partly responsible for regulated M- N# 1 antigen expression postlactation.
Abstract: Antibodies against the histo-blood group B–like antigen M-N#1 efficiently block the growth in vivo of rat mammary carcinoma cells that bear the antigen (Sleeman et al., 1999, Oncogene 18, 4485–4494). To try to understand the function of the M-N#1 antigen, we investigated when and where the antigen is expressed during the normal function of the rat mammary gland. Expression was virtually only seen during mammary gland involution. Here, strong expression of the antigen was observed in mammary epithelial cells, beginning around 2 days postweaning and lasting throughout the involution process. Dexamethasone treatment of animals postlactation inhibited alveolar collapse and remodeling in the mammary gland but inhibited neither the apoptosis of mammary epithelial cells nor the expression of the M-N#1 antigen. We show that up-regulation of carbohydrate antigens is not a general phenomenon during mammary gland involution, and thus that M-N#1 antigen expression is specifically regulated. Up-regulation of α(1,2)fucosyltransferase A, an enzyme required for M-N#1 antigen synthesis, is at least partly responsible for regulated M-N#1 antigen expression postlactation. Most significantly, we observed that the M-N#1 antigen is virtually exclusively expressed on nonapoptosing epithelial cells in the involuting mammary gland. These data suggest that M-N#1 antigen expression might either provide a survival function and/or be expressed in epithelial cells that are destined to grow and remodel mammary duct structures.

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Citations
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Journal ArticleDOI
TL;DR: A polymeric conjugate is synthesized in such a way that de-N-acetylated A-trisaccharide is attached to a polymer via the nitrogen in position C-2 of the galactosamine residue, in order to test the hypothesis that the supposed AB-epitope should be situated in the part of the molecule that is opposite to the NHAc group of GalNAc residue.
Abstract: Although the nature of the blood groups A and B has been comprehensively studied for a long time, it is still unclear as to what exactly is the epitope that is recognized by antibodies having AB specificity, i.e. monoclonal and polyclonal antibodies which are capable of interacting equally well with the antigens GalNAcalpha 1-3(Fucalpha 1-2)Gal (A trisaccharide) and Galalpha 1-3(Fucalpha 1-2)Gal (B trisaccharide), but do not react with their common fragment Fucalpha 1-2Gal. We have supposed that besides Fucalpha 1-2Gal, A and B antigens have one more shared epitope. The trisaccharides A and B are practically identical from the conformational point of view, the only difference being situated at position 2 of Galalpha residue, i.e. trisaccharide A has a NHAc group, whereas trisaccharide B has a hydroxyl group (see formulas). We have hypothesized that the AB-epitope should be situated in the part of the molecule that is opposite to the NHAc group of GalNAc residue. In order to test this hypothesis we have synthesized a polymeric conjugate in such a way that de-N-acetylated A-trisaccharide is attached to a polymer via the nitrogen in position C-2 of the galactosamine residue. In this conjugate the supposed AB-epitope should be maximally accessible for antibodies from the solution, whereas the discrimination site of antigens A and B by the antibodies should be maximally hidden due to the close proximity of the polymer. Interaction with several anti-AB monoclonal antibodies revealed that a part of them really interacted with the synthetic AB-glycotope, thus confirming our hypothesis. Moreover, similar antibodies were revealed in the blood of healthy blood group 0 donors. Analysis of spatial models was performed in addition to identify the hydroxyl groups of Fuc, Galalpha, and Galbeta residues, which are particularly involved in the composition of the AB-glycotope.

20 citations

Journal ArticleDOI
TL;DR: The data suggest that the up-regulation of galectin-3 in the involuting mammary gland is not only controlled transcriptionally but also regulated posttranscriptionally under the control of systemic glucocorticoid hormones involved in coordinating the involution process.
Abstract: Galectin-3 is an endogenous mammalian lectin that binds to ABH carbohydrate antigens. Here we show that galectin-3 is strongly up-regulated during mammary gland involution and that it is expressed virtually exclusively on nonapoptotic cells. We demonstrate that dexamethasone, an inhibitor of the second phase of mammary gland involution, potently suppresses up-regulation of galectin-3 as judged immunohistochemically and on western blots, suggesting that systemic hormone levels regulate galectin-3 expression during involution. However, at the RNA level galectin-3 expression is rapidly up-regulated on the onset of involution but remains consistantly high during the first and second phase of involution regardless of dexamethasone treatment. These data suggest that the up-regulation of galectin-3 in the involuting mammary gland is not only controlled transcriptionally but also regulated posttranscriptionally under the control of systemic glucocorticoid hormones involved in coordinating the involution process.

20 citations


Cites background from "Expression of M-N#1, a histo-blood ..."

  • ...Note that the lactating mammary gland sample is overloaded to permit a glyceraldehyde-3-phosphate dehydrogenase (GAPDH) signal to be seen because we have previously shown that GAPDH expression is up-regulated during mammary gland involution (Mengwasser and Sleeman, 2001)....

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  • ...The essentially blood group B antigen M-N1 is strongly up-regulated in involuting mammary glands (Mengwasser and Sleeman, 2001)....

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  • ...Additionally, dexamethasone treatment inhibits tissue remodeling but not apoptosis in involuting mammary glands (Lund et al., 1996; Li et al., 1997; Mengwasser and Sleeman, 2001; data presented herein) and also suppresses galectin-3 expression (Figure 3, Figure 5, Table II)....

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  • ...We have previously demonstrated that the M-N1 antigen is not expressed on apoptosing cells in this context (Mengwasser and Sleeman, 2001)....

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  • ...Glucocorticoid hormones have been reported to be able to inhibit the second stage of involution in postlactating animals (Lund et al., 1996; Li et al., 1997; Mengwasser and Sleeman, 2001)....

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Journal ArticleDOI
TL;DR: Novel evidence for altered expression of proliferative antigens and enhanced expression of HBGA B in fibroblasts treated with dental monomers bis-GA and bis-GMA is presented suggesting that these substances affect cell morphology, proliferative activity, and antigenic profile.
Abstract: The present work is focused on examining the effect of the structurally similar dental monomers bis-GA and bis-GMA on the expression of histo-blood group antigens (HBGAs) in comparison with fibroblast vitality and proliferation. The fibroblast cell line McCoy-Plovdiv was cultivated in a serum-free medium and was treated with both monomers. Cell vitality was measured by the crystal violet test. Mitotic index and cell morphology were assessed. An immunocytochemical technique was applied to follow the expression of proliferative antigens PCNA and Ki-67 and of HBGA. The expression level of HBGA was measured by an improved pixel selection algorithm with proprietary software. The lowest concentration of 2.5 μmol/L did not significantly affect morphology, vitality, or proliferation activity. Interestingly, the quantitative analysis revealed augmented expression of HBGA B at 2.5 μmol/L. The higher concentrations of the dental monomers reduced cell vitality and mitotic indices and altered proliferative antigen expression. Bis-GA proved to be more toxic than bis-GMA and caused more prominent alterations including greater enhancement of HBGA B expression. We present novel evidence for altered expression of proliferative antigens and enhanced expression of HBGA B in fibroblasts treated with dental monomers bis-GA and bis-GMA suggesting that these substances affect cell morphology, proliferative activity, and antigenic profile.

3 citations


Cites background from "Expression of M-N#1, a histo-blood ..."

  • ...It was shown that a histo-blood group B-like antigen is strongly upregulated during rat mammary gland involution, implying that this antigen might either provide a survival function and/ or be expressed in epithelial cells destined to remodel mammary ducts (Mengwasser and Sleeman 2001)....

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References
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Journal ArticleDOI
17 May 1990-Nature
TL;DR: A critical single-base deletion was found in the 0 gene, which results in an entirely different, inactive protein incapable of modifying the H antigen, and this work presents a molecular basis for the ABO genotypes.
Abstract: The histo-blood group ABO, the major human alloantigen system, involves three carbohydrate antigens (ABH). A, B and AB individuals express glycosyltransferase activities converting the H antigen into A or B antigens, whereas O(H) individuals lack such activity. Here we present a molecular basis for the ABO genotypes. The A and B genes differ in a few single-base substitutions, changing four amino-acid residues that may cause differences in A and B transferase specificity. A critical single-base deletion was found in the O gene, which results in an entirely different, inactive protein incapable of modifying the H antigen.

1,066 citations

Journal Article
TL;DR: Glycosylation analogues and sphingolipid analogues that inhibit protein kinase C and receptor-associated tyrosine kinase have been applied for inhibition of metastasis and provide the basis for development of "anti-adhesion therapy."
Abstract: Aberrant glycosylation expressed in glycosphingolipids and glycoproteins in tumor cells has been implicated as an essential mechanism in defining stage, direction, and fate of tumor progression This general concept is supported by results from three lines of study: (a) Numerous clinicopathological studies have shown a clear correlation between aberrant glycosylation status of primary tumor and invasive/metastatic potential of human cancer as reflected by 5- or 10-year survival rates of patients (b) Carbohydrates expressed in tumor cells are either adhesion molecules per se or modulate adhesion receptor function Some are directly involved in cell adhesion They are recognized by selectins or other carbohydrate-binding proteins or by complementary carbohydrates (through carbohydrate-carbohydrate interaction) N- or O-glycosylation of functionally important membrane components may alter tumor cell adhesion or motility in a direction that either promotes or inhibits invasion and metastasis Examples of such receptors are E-cadherin, integrins, immunoglobulin family receptors (eg, CD44), and lysosome-associated membrane protein (c) Gangliosides and sphingolipids modulate transmembrane signaling essential for tumor cell growth, invasion, and metastasis The transducer molecules susceptible to gangliosides and sphingolipids include integrin receptors, tyrosine kinase-linked growth factor receptors, protein kinase C, and G-protein-linked receptor affecting protein kinase A Some glycosphingolipids (eg, Gb3Cer, Le(y), ceramide, and sphingosine induce tumor cell differentiation and subsequent apoptosis Shedded gangliosides may block immunogenicity of tumor cells, providing conditions favorable for "escape" from immunological suppression of tumor growth by the host Various reagents that block carbohydrate-mediated tumor cell adhesion or block glycosylation processing have been shown to inhibit tumor cell metastasis This provides the basis for further development of "anti-adhesion therapy" Ganglioside analogues and sphingolipid analogues that inhibit protein kinase C and receptor-associated tyrosine kinase have been applied for inhibition of metastasis A crucial mechanism for inhibition of metastasis by these reagents may involve blocking of transmembrane signaling for expression of P- and E-selectin This provides the basis for development of "ortho-signaling therapy"

986 citations

Journal ArticleDOI
TL;DR: Galectin-3 is a regulator of cell growth and apoptosis and it may function through a cell death inhibition pathway that involves Bcl-2, a well-characterized suppressor of apoptosis.
Abstract: Galectin-3 is a member (if a large family of beta-galactoside-binding animal lectins. It has been shown that the expression of galectin-3 is upregulated in proliferating cells, suggesting a possible role for this lectin in regulation of cell growth. Previously, we have shown that T cells infected with human T-cell leukemia virus type I express high levels of galectin-3, in contrast to uninfected cells, which do not express detectable amounts of this protein. In this study, we examined growth properties of human leukemia T cells transfected with galectin-3 cDNA, and thus constitutively overexpressing this lectin. Transfectants expressing galectin-3 displayed higher growth rates than control transfectants, which do not express this lectin. Furthermore, galectin-3 expression in these cells confers resistance to apoptosis induced by anti-Fas antibody and staurosporine. Galectin-3 was found to have significant sequence similarity with Bcl-2, a well-characterized suppressor of apoptosis. In particular, the lectin contains the NWGR motif that is highly conserved among members of the Bcl-2 family and shown to be critical for the apoptosis-suppressing activity. We further demonstrated that galectin-3 interacts with Bc1-2 in a lactose-inhibitable manner. We conclude that galectin-3 is a regulator of cell growth and apoptosis and it may function through a cell death inhibition pathway that involves Bcl-2.

750 citations

Journal ArticleDOI
R. Strange1, Feng Li1, Susanne Saurer1, A. Burkhardt1, Robert R. Friis1 
TL;DR: Wyllie et al. as discussed by the authors showed that post-lactational mammary gland involution results in a dramatic drop in ODC, a gene involved in synthesis of a component of milk, and nearly simultaneous induction of SGP-2, associated with apoptotic cell death.
Abstract: During post-lactational mammary gland involution, the bulk of mammary epithelium dies and is reabsorbed. This massive cell death and tissue restructuring was found to be accompanied by a specific pattern of gene expression. Northern blot analysis showed that weaning resulted in a dramatic drop in ODC, a gene involved in synthesis of a component of milk, and the nearly simultaneous induction of SGP-2, a gene associated with apoptotic cell death. These changes were followed by decreases in expression of milk protein genes to basal levels and expression of genes associated with regulation of cell proliferation and differentiation, p53, c-myc and TGF-beta 1. Subsequently, additional genes implicated in stress response, tissue remodelling, and apoptotic cell death were transiently expressed, expression peaking at about 6 days post-weaning. A non-random degradation of DNA yielding the oligonucleosomal length fragmentation pattern typical of apoptotic cell death (Wyllie, 1980; Wyllie et al., 1980) was detected in association with morphological changes and gene expression. The correlations between: (a) changes in morphology, (b) pattern of gene expression and (c) changes in DNA integrity suggest that complementary programs for cell death and tissue remodelling direct post-lactational mammary gland involution.

531 citations

Journal ArticleDOI
TL;DR: The data support the hypothesis that there are at least two distinct phases of involution: an initial phase, characterized by induction of the apoptosis-associated genes SGP-2 and ICE and apoptosis of fully differentiated mammary epithelial cells without visible degradation of the extracellular matrix, and a second phase, characterize by extrace cellular matrix remodeling and altered mesenchymal-epithelial interactions, followed by apoptotic cells that are losing differentiated functions.
Abstract: Postlactational involution of the mammary gland is characterized by two distinct physiological events: apoptosis of the secretory, epithelial cells undergoing programmed cell death, and proteolytic degradation of the mammary gland basement membrane. We examined the spatial and temporal patterns of apoptotic cells in relation to those of proteinases during involution of the BALB/c mouse mammary gland. Apoptosis was almost absent during lactation but became evident at day 2 of involution, when beta-casein gene expression was still high. Apoptotic cells were then seen at least up to day 8 of involution, when beta-casein gene expression was being extinguished. Expression of sulfated glycoprotein-2 (SGP-2), interleukin-1 beta converting enzyme (ICE) and tissue inhibitor of metalloproteinases-1 was upregulated at day 2, when apoptotic cells were seen initially. Expression of the matrix metalloproteinases gelatinase A and stromelysin-1 and the serine proteinase urokinase-type plasminogen activator, which was low during lactation, was strongly upregulated in parallel starting at day 4 after weaning, coinciding with start of the collapse of the lobulo-alveolar structures and the intensive tissue remodeling in involution. The major sites of mRNA synthesis for these proteinases were fibroblast-like cells in the periductal stroma and stromal cells surrounding the collapsed alveoli, suggesting that the degradative phase of involution is due to a specialized mesenchymal-epithelial interaction. To elucidate the functional role of these proteinases during involution, at the onset of weaning we treated mice systemically with the glucocorticoid hydrocortisone, which is known to inhibit mammary gland involution. Although the initial wave of apoptotic cells appeared in the lumina of the gland, the dramatic regression and tissue remodeling usually evident by day 5 was substantially inhibited by systemic treatment with hydrocortisone. mRNA and protein for gelatinase A, stromelysin-1 and uPA were weakly induced, if at all, in hydrocortisone-treated mice. Furthermore, mRNA for membrane-type matrix metalloproteinase decreased after hydrocortisone treatment and paralleled the almost complete inhibition of activation of latent gelatinase A. Concomitantly, the gland filled with an overabundance of milk. Our data support the hypothesis that there are at least two distinct phases of involution: an initial phase, characterized by induction of the apoptosis-associated genes SGP-2 and ICE and apoptosis of fully differentiated mammary epithelial cells without visible degradation of the extracellular matrix, and a second phase, characterized by extracellular matrix remodeling and altered mesenchymal-epithelial interactions, followed by apoptosis of cells that are losing differentiated functions.

526 citations


"Expression of M-N#1, a histo-blood ..." refers background in this paper

  • ...On the other hand, subcutaneous injection of hydrocortisone was observed to inhibit the second phase on involution without affecting apoptosis induction (Lund et al., 1996; Li et al., 1996)....

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  • ...Changes in the expression of a number of different genes during mammary gland involution have been described, including decreased expression of milk protein genes and up-regulated expression of bcl-2 family members, interleukin-1β converting enzyme, sulfated glycoprotein-2, and TIMP-1 (Lund et al., 1996; Li et al., 1996)....

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  • ...In the second phase, proteases degrade extracellular matrix and basement membrane components, and the lobular-alveolar structures collapse (Lund et al., 1996)....

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  • ...Mammary involution occurs in two stages: a first, reversible stage controlled by local factors in which alveolar structure remains intact; and a second stage dependent on systemic hormone levels, which is characterized by alveolar collapse and tissue remodeling and which begins around 4 days postweaning (Lund et al., 1996; Li et al., 1997)....

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  • ...During involution the mammary gland is restructured through the coordinated processes of apoptosis of luminal epithelial, myoeptithelial, and endothelial cells and lobular-alveolar remodeling (Pitelka, 1988; Lund et al., 1996; Strange et al., 1992)....

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