Iwona Kątnik-Prastowska

Alterations of branching and differential expression of sialic acid on alpha-1-acid glycoprotein in human seminal plasma

BACKGROUND The degree of branching and types of fucosylation of glycans on alpha(1)-acid glycoprotein (AGP) have been found to be associated with alpha(1)-acid glycoprotein concentrations in human seminal plasma. The glycosylation pattern of alpha(1)-acid glycoprotein in seminal plasma obtained from men living in infertile couples can undergo alterations in relation to sperm analysis and/or alpha(1)-acid glycoprotein concentrations. METHODS The glycosylation of alpha(1)-acid glycoprotein was studied upon the reactivity with specific lectins by crossed affinity immunoelectrophoresis (concanavalin A), and by glycoprotein lectin immunosorbent assay (Maackia amurensis and Sambucus nigra lectins), as well as high pH anion-exchange chromatography with pulsed amperometric detection. RESULTS Nonsignificant differences in alpha(1)-acid glycoprotein glycan branching and degree of its sialylation were observed among the AGP derived from seminal plasmas in relation to spermiogram and sperm morphology. However, significant concentration-dependent differences were found in extent of branching and type of sialylation. CONCLUSIONS The presence in seminal plasma of high concentrations of aberrantly glycosylated AGP molecules might be indicative for a chronic inflammatory condition in the reproductive tract, and can be used as additional tool to subdivide the seminal plasmas of men living in infertile couples.

Alterations in molecular status of plasma fibronectin associated with aging of normal human individuals

OBJECTIVES Senescence, progressive deterioration of many bodily functions might be associated with age-dependent alterations of plasma fibronectin (FN) molecular status (i.e., domain, glycotope, and molecular form expressions). DESIGN AND METHODS FN molecular status was analyzed in 127 plasma samples of healthy individuals in groups of newborns, and subjects aged 3-14, 15-39, 41-59, and 60-82 years by FN-ELISA, lectin-FN-ELISA, and immunoblotting using a set of domain-specific monoclonal antibodies, specific lectins, and monoclonal antibody to FN, respectively. RESULTS During the first four decades of human life the levels of cell-binding-, carboxyl-terminal-, collagen-, heparin-, and fibrin-domains of plasma FN gradually increased. In subjects aged up to 82 years the cell-binding and carboxyl-terminal FN domain concentrations did not change, while the heparin, fibrin, and collagen domains significantly increased. The relative reactivity of plasma FN with Maackia amurensis lectin, specific to α2,3-linked sialic acid, significantly decreased after birth, reaching a stable level in the subsequent life period, whereas with Sambucus nigra lectin, specific to α2,6-linked sialic acid, it significantly decreased in the 60-82 year old group. Moreover, the appearance of 280-kDa and 320-kDa FN bands, absent in young and mature healthy individuals, was found in the groups of 41-59 and 60-82 year olds. CONCLUSIONS The alterations of FN molecular status throughout growth, maturation and senescence might be associated not only with disturbances in the balance of FN production rate and degradation, but concomitantly with conformational rearrangements of FN and its engagement in age-related vascular remodeling processes.

Relative sialylation and fucosylation of synovial and plasma fibronectins in relation to the progression and activity of rheumatoid arthritis

The expressions of terminal sugars in synovial and plasma fibronectins were studied in relation to rheumatoid arthritis (RA) progression defined according to the early, established and late radiological changes in the patients’ hands. The relative amounts of sialic acid and fucose were analyzed by lectin-ELISA using appropriate sialic acid-linked α2-3 (Maackia amurensis) and α2-6 (Sambucus nigra) lectins as well as fucose-linked α1-6 (Aleuria aurantia), α1-2 (Ulex europaeus), and α1-3 (Tetragonolobus purpureus). In the early RA group, the synovial fibronectin reactivities were the lowest with the all lectins used. In the established and late groups, relative sialylation and fucosylation significantly increased. However, sialylation negligibly decreased, whereas fucosylation remained at nearly the same level in the late group. Moreover, the expression of α1-6-linked fucose was found to be related to disease activity. In contrast, plasma fibronectin reactivity with lectins showed different dynamic alterations. In the early RA group, the reactivity of fibronectin with the lectins used was similar to that of healthy individuals, whereas it increased significantly in the established RA group compared with the early and normal plasma groups. In the late RA group it decreased to a level similar to that of the normal group. The lower expressions of terminal sugars in synovial fibronectin were mainly associated with the early degenerative processes of RA. In conclusion, such alterations may be applicable as a stage-specific marker for diagnosis and therapy of RA patients. The higher expression of terminal sugars in fibronectin could be associated with repair and adaptation processes in longstanding disease.

The expression of fucose isoforms of amniotic and plasma alpha-1-acid glycoprotein derived from 2nd and 3rd trimester normal pregnancies

OBJECTIVES To analyse modifications in AGP fucosylation in relation to different stages of human pregnancy. DESIGN AND METHODS The relative amounts of three fucosyl-glycotopes on AGP were analysed by lectin-ELISA using fucose-specific biotinylated lectins in 169 plasma and 178 amniotic fluid samples from normal pregnancies with gestational ages of 14 to 42 weeks. RESULTS The plasma AGPs of all the pregnant women and amniotic AGPs from the 2nd trimester lacked fucoses. In contrast, in the 3rd trimester the amniotic AGPs were highly decorated by the innermost alpha1,6-fucose as well as alpha1,2- and alpha1,3-fucoses of the outer arms, reaching the highest expression around the perinatal period. At delivery the relative amounts of the alpha1,3- and alpha1,2-AGP isoforms, but not the alpha1,6 isoform, significantly decreased. CONCLUSIONS The highly fucosylated amniotic AGP isoforms could be implicated in regulatory processes to ensure homeostasis during pregnancy and to protect the fetus. They have the potential of becoming laboratory markers in obstetrics to monitor pregnancy.

Presence of high-molecular-weight forms and domain alterations of fibronectin in pleural effusion of patients with lung cancer

OBJECTIVES Appearance of fibronectin (FN) molecular forms and alterations of domain expositions can be associated with lung cancer. DESIGN AND METHODS The presence of the FN molecular forms and epitopes in its cell-binding, carboxyl-, and amino-terminal domains was determined in the plasma and pleural effusion of patients suffering from small- and non-small-cell lung cancer, and lung inflammation by immunoblotting and FN-ELISA. RESULTS The 320-kDa and 280-kDa FN forms as well as FN fragments appeared in the pleural effusion and plasma of patients suffering from lung inflammation or cancer in significantly higher relative amounts in both lung cancer groups than in the inflammation. The domain concentrations were higher in the cancer and inflammatory plasma groups than those in the control group. The higher N-terminal epitope expression in pleural effusion than in plasma indicates different epitope accessibility for the monoclonal antibody. CONCLUSIONS The molecular status of FN probably reflects the dynamic changes which occur in cancer and inflammatory tissue.

Expression and potential biological role of α(1,2)fucosylated glycotopes on amniotic and seminal fibronectins.

The present paper describes concisely the expression and role of α(1,2)-linked fucose on some glycoconjugates as well as the detection, distribution and potential role of that glycotope on human soluble plasma and cellular fibronectins in addition to the expression on both normal and pathological amniotic fluid and seminal plasma fibronectins. The determination of α(1,2)fucosylated glycans is considered with respect to its usefulness as a potential clinically applicable biomarker in obstetrics to monitor pregnancy and in andrology to evaluate the ejaculate of infertile men and in vitro fertilization.

Fucose and sialic acid expressions in human seminal fibronectin and α{1} -acid glycoprotein associated with leukocytospermia of infertile men.

Introduction: The aim of this study was to compare fucose and sialic acid residue expression on fibronectin and α1-acid glycoprotein in the seminal plasma of men suspected of infertility and suffering from leukocytospermia. Subjects and methods: Seminal ejaculates were collected from 27 leukocytospermic and 18 healthy, normozoospermic men. The relative degree of fucosylation and sialylation of fibronectin and α1-acid glycoprotein was estimated by ELISA using fucose and sialic acid specific lectins from Aleuria aurantia, Lotus tetragonolobus, and Ulex europaeus as well as Maackia amurensis and Sambucus nigra, respectively. Results: Leukocytospermic seminal fibronectin, in comparison with fibronectin of normal fertile group, showed lower relative reactivity with AAL, LTA and UEA, and higher reactivity with MAA and SNA, while the AGP of the leukocytospermic group was less reactive with AAL, and the relative reactivity with LTA and MAA was significantly higher. Fibronectin and α1-acid glycoprotein reactivity with UEA and MAA showed high positive correlations. Discussion: Leukocytospermia was associated with the alterations of terminal monosaccharide expression in human seminal fibronectin and α1-acid glycoprotein. The increase of sialyl-Lewisx antigen in α1-acid glycoprotein can be used as a marker of genital tract inflammation manifested by leukocytospermia.

Lectin-based analysis of fucose and sialic acid expressions on human amniotic IgA during normal pregnancy

The sugar moiety of IgA is known to provide a link between the innate and adaptive immune systems. Terminally located glycotopes on IgA are potential ligands engaged in the interactions which may modulate the biological activities of IgA. In the present work the expressions of Maackia amurensis (MAA), Sambucus nigra (SNA), Lens culinaris (LCA), Tetragonolobus purpureus (LTA), and Ulex europaeus (UEA) reactive glycotopes on maternal plasma and amniotic IgA were evaluated in relation to the progression of a normal human pregnancy, from the 2nd trimester, throughout the 3rd trimester, perinatal period, post-date pregnancy and delivery, by lectin-IgA-ELISA, using specific biotinylated lectins. The amniotic and maternal plasma IgA concentrations and a degree of SNA and LCA reactivity of maternal plasma IgA were almost unaltered during the normal pregnancy. The amniotic IgA from the 2nd trimester was decorated by MAA-, SNA-reactive and LCA-, LTA-, and UEA-reactive glycotopes. At the turn of the 2nd and 3rd trimesters the expression of MAA-, SNA-, LTA-, and UEA-reactive glycotopes, except for LCA-reactive, increased and remained almost at unaltered levels throughout the perinatal period and delivery. However, in the post-date pregnancy the expression of LCA-, LTA-, and UEA-reactive and SNA-reactive glycotopes were significantly higher. The unique fucosylated and sialylated glycovariants of amniotic IgA associated with the progression of the normal pregnancy may illustrate a general importance of carbohydrate-lectin receptor interactions in the control and modulation of biological events to ensuring homeostasis during pregnancy, protection and well-being of fetus.

Terminal monosaccharide expression on amniotic glycoproteins as biomarkers of fetus maturity.

Glycotypes, particularly those that terminate with sialic acid and fucose are known to play a fundamental role in human development, during implantation, growth and differentiation of fetal tissues. The present review describes changes in the exposition of terminal sialic acid and fucose isoforms in the amniotic fluid glycoconjugates, α1-acid glycoprotein and fibronectin during critical stages of pregnancy, i.e. second and third trimester, perinatal period, delivery and post-date pregnancy. The distinct amniotic glycoforms are suggested to be implicated in regulatory processes to ensure homoeostasis during pregnancy and to protect the fetus. These may have the potential of becoming additional laboratory makers in obstetrics to monitor pregnancy.

Terminal glycotope expression on milk fibronectin differs from plasma fibronectin and changes over lactation

OBJECTIVES Fibronectin (FN) is a multifunctional glycoprotein appearing in various glycovariants with potential biological activities. Using lectins we analyzed the expression of terminal glycotopes on human milk fibronectin over lactation and compared it with that of the mothers plasma. DESIGN AND METHODS FN concentration and relative amounts of its fucosylated and sialylated glycovariants as well as O-glycans were analyzed in early colostrum, colostrum, transitional and mature milk samples of 132 healthy mothers by lectin-FN-ELISA using α2,3- and α2,6-sialic acid, α1,2-, α1,3-, and α1,6-fucose, and sialyl-T, asialyl-T and Tn antigen specific biotinylated Maackia amurensis, Sambucus nigra, Ulex europaeus, Tetragonolobus purpureus, Lens culinaris, Artocarpus integrifolia, Arachis hypogaea, and Vicia villosa lectins, respectively. RESULTS FN concentration was almost unchanged during human milk maturation and was about 150 times lower than in plasma of lactating mothers. Milk FN elicited significantly higher expression of sialylated glycotopes including sialyl-T antigen than plasma FN, and contained fucose-linked glycans, as well as T and Tn antigens absent in plasma FN. With milk maturation the expression of α2,6-sialylated, sialyl-T, α1,6- and α1,2-fucosylated epitopes decreased in transitional milk compared with colostrum, whereas that of asialyl-T antigen increased. The expression levels of α2,3-sialyl- and α1,3-fucosyl-glycotopes and Tn antigen on FN were low and did not change over lactation. CONCLUSION The expression of terminal sugars on milk FN is different from that of plasma FN of the lactating mother and is associated with milk maturation. The analysis of degree of milk sialylation and fucosylation should be considered during control of biochemical quality of milk collected in milk banks.