Expression and secretion of galectin-12 in the context of neutrophilic differentiation of human promyeloblastic HL-60 cells.

Galectin-12 is a tissue-specific galectin that has been largely defined by its role in the regulation of adipocyte differentiation and lipogenesis. This study aimed to evaluate the role of galectin-12 in the differentiation and polarization of neutrophils within a model of acute myeloid leukemia HL-60 cells. All-trans retinoic acid and dimethyl sulfoxide were used to induce differentiation of HL-60 cells which led to the generation of two phenotypes of neutrophil-like cells with opposite changes in galectin-12 gene (LGALS12) expression and different functional responses to N-formyl- l-methionyl- l-leucyl- l-phenylalanine. These phenotypes showed significant differences of differentially expressed genes on a global scale based on bioinformatics analysis of available Gene Expression Omnibus (GEO) data sets. We also demonstrated that HL-60 cells could secrete and accumulate galectin-12 in cell culture medium under normal growth conditions. This secretion was found to be entirely inhibited upon neutrophilic differentiation and was accompanied by an increase in intracellular lipid droplet content and significant enrichment of 22 lipid gene ontology terms related to lipid metabolism in differentiated cells. These findings suggest that galectin-12 could serve as a marker of neutrophilic plasticity or polarization into different phenotypes and that galectin-12 secretion may be influenced by lipid droplet biogenesis.

Yeast sexes: mating types do not determine the sexes in Metschnikowia species.

Although filamentous Ascomycetes may produce structures that are interpreted as male and female gametangia, ascomycetous yeasts are generally not considered to possess male and female sexes. In haplontic yeasts of the genus Metschnikowia, the sexual cycle begins with the fusion of two morphologically identical cells of complementary mating types. Soon after conjugation, a protuberance emerges from one of the conjugants, eventually maturing into an ascus. The originating cell can be regarded as an ascus mother cell, hence as female. We tested the hypothesis that the sexes, female or male, are determined by the mating types. There were good reasons to hypothesize further that mating type α cells are male. In a conceptually simple experiment, we observed the early stages of the mating reaction of mating types differentially labeled with fluorescent concanavalin A conjugates. Three large-spored Metschnikowia species, M. amazonensis, M. continentalis, and M. matae, were examined. In all three, the sexes were found to be independent of mating type, cautioning that the two terms should not be used interchangeably.

Insights into cAMP-dependent molecular mechanisms regulating expression and function of LGALS16 gene in choriocarcinoma JEG-3 cells.

The human choriocarcinoma cell line JEG-3 offers a valuable model to study galectin-16 gene (LGALS16) expression and functions in the context of placental cell differentiation and cancer cell biology. Recent evidence indicates that cAMP-mediated signaling pathways might be responsible for the upregulation of LGALS16; however, the underlying mechanisms are unknown. Here, we employed biochemical inhibitors of the cAMP cascade and CRISPR/Cas9 engineered cells to assess regulatory patterns and associations between cAMP-induced trophoblast differentiation and LGALS16 expression in JEG-3 cells. The expression of LGALS16 was significantly upregulated in parallel with human chorionic gonadotropin beta (CGB), a biomarker of syncytiotrophoblast differentiation, in response to 8-Br-cAMP. Inhibition of p38 MAPK and EPAC significantly altered LGALS16 expression during differentiation, while PKA inhibition failed to change LGALS16 and CGB3/5 expression in our cell model. The CRISPR/Cas9 LGALS16 knockout cell pool expressed a significantly lower amount of CGB3/5, a reduced level of CGB protein, and an unaltered cell growth rate in response to 8-Br-cAMP in comparison with wild-type JEG-3 cells. Collectively, these findings suggest that LGALS16 is required for the trophoblast-like differentiation of JEG-3 cells, and its expression is mediated through p38 MAPK and EPAC signaling pathway branches.

O-GlcNAc-Mediated Regulation of Galectin Expression and Secretion in Human Promyelocytic HL-60 Cells Undergoing Neutrophilic Differentiation.

In this study, we have tested the hypothesis that the expression and secretion of galectins are driven through mechanisms globally impacted by homeostatic regulation involving the post-translational modification of intracellular proteins with O-linked N-acetylglucosamine (O-GlcNAc). We showed that neutrophilic differentiation of HL-60 cells induced by all-trans retinoic acid (ATRA) and 6-diazo-5-oxo-L-norleucine (DON) was associated with a significant drop of cellular O-GlcNAc levels in serum-contained and serum-free cell culture media. Galectin gene and protein expression profiles in HL-60 cells were specifically modified by ATRA and by inhibitors of O-GlcNAc cycle enzymes, however overall trends for each drug were similar between cells growing in the presence or absence of serum except for LGALS9 and LGALS12. The secretion of four galectins (-1, -3, -9, and -10) by HL-60 cells in a serum-free medium was stimulated by O-GlcNAc-reducing ATRA and DON while O-GlcNAc-elevating thiamet G (O-GlcNAcase inhibitor) failed to change the basal levels of extracellular galectins. Taken together, these results demonstrate that O-GlcNAc homeostasis is essential not only for regulation of galectin expression in cells but also for the secretion of multiple members of this protein family, which can be an important novel aspect of unconventional secretion mechanisms.

Cell Biology of Galectins: Novel Aspects and Emerging Challenges.

Galectins are a family of soluble ß-galactoside-binding proteins with diverse glycan-dependent and glycan-independent functions outside and inside the cell [...].

O-GlcNAcylation and Regulation of Galectin-3 in Extraembryonic Endoderm Differentiation.

The regulation of proteins through the addition and removal of O-linked ß-N-acetylglucosamine (O-GlcNAc) plays a role in many signaling events, specifically in stem cell pluripotency and the regulation of differentiation. However, these post-translational modifications have not been explored in extraembryonic endoderm (XEN) differentiation. Of the plethora of proteins regulated through O-GlcNAc, we explored galectin-3 as a candidate protein known to have various intracellular and extracellular functions. Based on other studies, we predicted a reduction in global O-GlcNAcylation levels and a distinct galectin expression profile in XEN cells relative to embryonic stem (ES) cells. By conducting dot blot analysis, XEN cells had decreased levels of global O-GlcNAc than ES cells, which reflected a disbalance in the expression of genes encoding O-GlcNAc cycle enzymes. Immunoassays (Western blot and ELISA) revealed that although XEN cells (low O-GlcNAc) had lower concentrations of both intracellular and extracellular galectin-3 than ES cells (high O-GlcNAc), the relative secretion of galectin-3 was significantly increased by XEN cells. Inducing ES cells toward XEN in the presence of an O-GlcNAcase inhibitor was not sufficient to inhibit XEN differentiation. However, global O-GlcNAcylation was found to decrease in differentiated cells and the extracellular localization of galectin-3 accompanies these changes. Inhibiting global O-GlcNAcylation status does not, however, impact pluripotency and the ability of ES cells to differentiate to the XEN lineage.

Lactoferrin modified by hypohalous acids: Partial loss in activation of human neutrophils.

Previously we have shown that lactoferrin (LTF), a protein of secondary neutrophilic granules, can be efficiently modified by hypohalous acids (HOCl and HOBr), which are produced at high concentrations during inflammation and oxidative/halogenative stress by myeloperoxidase, an enzyme of azurophilic neutrophilic granules. Here we compared the effects of recombinant human lactoferrin (rhLTF) and its halogenated derivatives (rhLTF-Cl and rhLTF-Br) on functional responses of neutrophils. Our results demonstrated that after halogenative modification, rhLTF lost its ability to induce mobilization of intracellular calcium, actin cytoskeleton reorganization, and morphological changes in human neutrophils. Moreover, both forms of the halogenated rhLTF prevented binding of N-acetylglucosamine-specific plant lectin Triticum vulgaris agglutinin (WGA) to neutrophils and, in contrast to native rhLTF, inhibited respiratory burst of neutrophils induced by N-formyl-L-methionyl-L-leucyl-L-phenylalanine and by two plant lectins (WGA and PHA-L). However, we observed no differences between the effects of rhLTF, rhLTF-Cl, and rhLTF-Br on respiratory burst of neutrophils induced by phorbol 12-myristate 13-acetate (PMA), digitonin, and number of plant lectins with different glycan-binding specificity. Furthermore, all rhLTF forms interfered with PMA- and ionomycin-induced formation of neutrophil extracellular traps. Thus, halogenative modification of LTF is one of the mechanisms involved in modulating a variety of signaling pathways in neutrophils to control their pro-inflammatory activity.

Expression, Regulation, and Functions of the Galectin-16 Gene in Human Cells and Tissues.

Galectins comprise a family of soluble ß-galactoside-binding proteins, which regulate a variety of key biological processes including cell growth, differentiation, survival, and death. This paper aims to address the current knowledge on the unique properties, regulation, and expression of the galectin-16 gene (LGALS16) in human cells and tissues. To date, there are limited studies on this galectin, with most focusing on its tissue specificity to the placenta. Here, we report the expression and 8-Br-cAMP-induced upregulation of LGALS16 in two placental cell lines (BeWo and JEG-3) in the context of trophoblastic differentiation. In addition, we provide the results of a bioinformatics search for LGALS16 using datasets available at GEO, Human Protein Atlas, and prediction tools for relevant transcription factors and miRNAs. Our findings indicate that LGALS16 is detected by microarrays in diverse human cells/tissues and alters expression in association with cancer, diabetes, and brain diseases. Molecular mechanisms of the transcriptional and post-transcriptional regulation of LGALS16 are also discussed based on the available bioinformatics resources.

The Emerging Role of Galectins and O-GlcNAc Homeostasis in Processes of Cellular Differentiation.

Galectins are a family of soluble ß-galactoside-binding proteins with diverse glycan-dependent and glycan-independent functions outside and inside the cell. Human cells express twelve out of sixteen recognized mammalian galectin genes and their expression profiles are very different between cell types and tissues. In this review, we summarize the current knowledge on the changes in the expression of individual galectins at mRNA and protein levels in different types of differentiating cells and the effects of recombinant galectins on cellular differentiation. A new model of galectin regulation is proposed considering the change in O-GlcNAc homeostasis between progenitor/stem cells and mature differentiated cells. The recognition of galectins as regulatory factors controlling cell differentiation and self-renewal is essential for developmental and cancer biology to develop innovative strategies for prevention and targeted treatment of proliferative diseases, tissue regeneration, and stem-cell therapy.

The association between increasing levels of O-GlcNAc and galectins in the liver tissue of hibernating thirteen-lined ground squirrels (Ictidomys tridecemlineatus).

Post-translational glycosylation of proteins with O-linked ß-N-acetylglucosamine (O-GlcNAcylation) and changes of galectin expression profiles are essential in many cellular stress responses. We examine this regulation in the liver tissue of hibernating thirteen-lined ground squirrels (Ictidomys tridecemlineatus) representing a biological model of hypometabolism and physiological stress resistance. The tissue levels of O-GlcNAcylated proteins as well as galectin-1 and galectin-3 proteins detected by immunodot blot assay were significantly lower by 4.6-5.4-, 2.2-2.3- and 2.5-2.9-fold, respectively, in the non-hibernating summer squirrels compared with those in winter, whether hibernating or aroused. However, there were no differences in the expression of genes encoding enzymes involved in O-GlcNAc cycle (O-GlcNAc transferase and O-GlcNAcase) and such galectins as LGALS1, LGALS2, LGALS3, LGALS4 and LGALS9. Only the expression of LGALS8 gene in the liver tissue was significantly decreased by 37.6 ± 0.1% in hibernating ground squirrels relative to summer animals. Considering that the expression of a proven genetic biomarker ELOVL6 encoding ELOVL fatty acid elongase 6 was readily upregulated in non-hibernating animals by 11.3-32.9-fold, marginal differential changes in the expression of galectin genes cannot be classified as biomarkers of hibernation. Thus, this study provides evidence that hibernation in Ictidomys tridecemlineatus is associated with increasing O-GlcNAcylation of liver proteins and suggests that the contribution of galectins deserves further studies at the protein level.

Effects of recombinant human lactoferrin on calcium signaling and functional responses of human neutrophils.

Lactoferrin is a non-heme iron-binding glycoprotein with multiple health-beneficial functions including antimicrobial, antioxidant, anticarcinogenic, and immunomodulatory effects. There is emerging evidence that neutrophils may serve as targets of lactoferrin in vivo, and here we show how recombinant human lactoferrin (rhLf) can contribute to this regulation. Indeed, our results demonstrate that rhLf binds efficiently to human neutrophils and induces a variety of early cellular responses such as mobilization of intracellular Ca2+, remodeling of actin cytoskeleton, and degranulation (release of lysozyme and myeloperoxidase). In addition, rhLf facilitates lectin-induced H2O2 production and stabilization of lectin-induced cellular aggregates. The role of calcium signaling seems to be essential for rhLf-induced activation of neutrophils, as Ca2+-chelators inhibit degranulation response while lectin-induced H2O2 production correlates significantly with cytoplasmic Ca2+ elevation. Taken together, our findings justify that rhLf can activate neutrophil functions in a calcium-dependent manner and hence, can potentiate innate immune responses.

Effects of Global O-GlcNAcylation on Galectin Gene-expression Profiles in Human Cancer Cell Lines.

BACKGROUND/AIM: The effects of O-linked ß-N-acetyl-D-glucosamine (O-GlcNAc) transferase (OGT) and O-GlcNAcase (OGA) inhibitors on galectin gene expression profiles were examined in MCF7, HT-29, and HL-60 cancer cell lines. MATERIALS AND METHODS: Cell cultures were treated for 24 h with OGA inhibitor thiamet G or OGT inhibitor 2-acetamido-1,3,4,6-tetra-O-acetyl-2-deoxy-5-thio-α-D-glucopyranose, and global O-GlcNAc levels and expression of galectin genes were determined using an immunodot blot assay and real-time quantitative polymerase chain reaction. RESULTS: Two galectin genes, LGALS3 in MCF7 cells and LGALS12 in HL-60 cells, were up-regulated by O-GlcNAc, whereas other cell-specific galectins were unresponsive to changes in O-GlcNAc level. Of interest, basal levels of O-GlcNAc in resting HL-60 and HT-29 cells were significantly higher than those in cells differentiated into neutrophilic or enterocytic lineages, respectively. CONCLUSION: O-GlcNAc-mediated signaling pathways may be involved in regulating the expression of only a limited number of galectin genes. Additional O-GlcNAc-dependent mechanisms may work at the protein level (galectin secretion and intracellular localization) and warrant further investigation.

Seasonal and flight-related variation of galectin expression in heart, liver and flight muscles of yellow-rumped warblers (Setophaga coronata).

Galectins, a family of multifunctional glycan-binding proteins, are proposed as biomarkers of cellular stress responses. Avian migration is an energetically challenging physical stress, which represents a physiological model of muscular endurance exercises. This study assesses change in galectin gene expression profiles associated with seasonal variation in migratory state and endurance flight in yellow-rumped warblers (Setophaga coronata). Bioinformatics analysis and real-time qPCR were used to analyse the expression of galectins in flight muscle, heart and liver tissues of 15 warblers separated into three groups of winter unflown, and fall migratory flown/unflown birds. Five transcripts similar to chicken and human galectins -1, -2, -3, -4, and -8 were identified in warbler tissues. The expression of these galectins showed no seasonal changes between two experimental groups of birds maintained under unflown winter and fall conditions indicating a minor role of galectins in preparation for migration. However, endurance flight led to a significant elevation of galectin-1 and galectin-3 mRNAs in flight muscles and galectin-3 mRNA in heart tissue while no changes were observed in liver. Different changes were observed for the level of O-GlcNAcylated proteins, which were elevated in flight muscles under winter conditions. These results suggest that secreted galectin-1 and galectin-3 may be active in repair of bird muscles during and following migratory flight and serve as molecular biomarkers of recent arrival from migratory flights in field studies.

The association between oxidative stress-induced galectins and differentiation of human promyelocytic HL-60 cells.

Galectins are multifunctional ß-galactoside-binding proteins that are involved in the regulation of cellular stress responses and differentiation. The relationship between these processes is unclear and we report here that galectins display oxidative-stress specific expression patterns in neutrophil-like differentiated HL-60 cells. Three galectins (-1, -3, and -10) are upregulated in response to either menadione or DMSO exposure whereas galectins -9 and -12 exhibited a stimulus-dependent downregulation. Changes in galectin expression are oxidant dependent based on the observations that 1) oxidative stress biomarkers HMOX1 (heme oxygenase-1) and NCF1 (neutrophil cytosolic factor 1, which is also a biomarker of neutrophil differentiation) are elevated in both cases, and 2) the antioxidant N-acetyl-L-cysteine restores basal expression of galectin-3 following oxidant exposure. In addition, our results suggest that the regulation of oxidative stress-sensitive galectins involves DNA hypomethylation mechanisms. Expression of galectin-3 and galectin-12 exhibits an opposite relationship to the expression of HMOX1/NCF1, suggesting a stimulatory and inhibitory role of these galectins in neutrophil-like differentiation of HL-60 cells. We also show that the inhibition of galectins reduces the growth rate of HL-60 cells, and facilitates their neutrophil-like differentiation. Collectively, our findings indicate that the process of cellular differentiation implicates, in part, oxidative stress-sensitive galectins, which further highlights a biological significance of galectin network remodeling in cells.

Binding of human myeloperoxidase to red blood cells: Molecular targets and biophysical consequences at the plasma membrane level.

Myeloperoxidase (MPO) is an oxidant-producing enzyme that can also bind to cellular surface proteins. We found that band 3 protein and glycophorins A and B were the key MPO-binding targets of human red blood cells (RBCs). The interaction of MPO with RBC proteins was mostly electrostatic in nature because it was inhibited by desialation, exogenic sialic acid, high ionic strength, and extreme pH. In addition, MPO failed to interfere with the lectin-induced agglutination of RBCs, suggesting a minor role of glycan-recognizing mechanisms in MPO binding. Multiple biophysical properties of RBCs were altered in the presence of native (i.e., not hypochlorous acid-damaged) MPO. These changes included transmembrane potential, availability of intracellular Ca(2+), and lipid organization in the plasma membrane. MPO-treated erythrocytes became larger in size, structurally more rigid, and hypersensitive to acidic and osmotic hemolysis. Furthermore, we found a significant correlation between the plasma MPO concentration and RBC rigidity index in type-2 diabetes patients with coronary heart disease. These findings suggest that MPO functions as a mediator of novel regulatory mechanism in microcirculation, indicating the influence of MPO-induced abnormalities on RBC deformability under pathological stress conditions.

Towards molecular mechanisms regulating the expression of galectins in cancer cells under microenvironmental stress conditions.

Galectins, a family of soluble ß-galactoside-binding proteins, serve as mediators of fundamental biological processes, such as cell growth, differentiation, adhesion, migration, survival, and death. The purpose of this review is to summarize the current knowledge regarding the ways in which the expression of individual galectins differs in normal and transformed human cells exposed to various stimuli mimicking physiological and pathological microenvironmental stress conditions. A conceptual point is being made and grounded that the modulation of galectin expression profiles is a key aspect of cellular stress responses. Moreover, this modulation might be precisely regulated at transcriptional and post-transcriptional levels in the context of non-overlapping transcription factors and miRNAs specific to galectins.

Co-expression of α9ß1 integrin and VEGF-D confers lymphatic metastatic ability to a human breast cancer cell line MDA-MB-468LN.

INTRODUCTION AND OBJECTIVES: Lymphatic metastasis is a common occurrence in human breast cancer, mechanisms remaining poorly understood. MDA-MB-468LN (468LN), a variant of the MDA-MB-468GFP (468GFP) human breast cancer cell line, produces extensive lymphatic metastasis in nude mice. 468LN cells differentially express α9ß1 integrin, a receptor for lymphangiogenic factors VEGF-C/-D. We explored whether (1) differential production of VEGF-C/-D by 468LN cells provides an autocrine stimulus for cellular motility by interacting with α9ß1 and a paracrine stimulus for lymphangiogenesis in vitro as measured with capillary-like tube formation by human lymphatic endothelial cells (HMVEC-dLy); (2) differential expression of α9 also promotes cellular motility/invasiveness by interacting with macrophage derived factors; (3) stable knock-down of VEGF-D or α9 in 468LN cells abrogates lymphangiogenesis and lymphatic metastasis in vivo in nude mice. RESULTS: A comparison of expression of cyclo-oxygenase (COX)-2 (a VEGF-C/-D inducer), VEGF-C/-D and their receptors revealed little COX-2 expression by either cells. However, 468LN cells showed differential VEGF-D and α9ß1 expression, VEGF-D secretion, proliferative, migratory/invasive capacities, latter functions being stimulated further with VEGF-D. The requirement of α9ß1 for native and VEGF-D-stimulated proliferation, migration and Erk activation was demonstrated by treating with α9ß1 blocking antibody or knock-down of α9. An autocrine role of VEGF-D in migration was shown by its impairment by silencing VEGF-D and restoration with VEGF-D. 468LN cells and their soluble products stimulated tube formation, migration/invasiveness of HMVEC-dLy cell in a VEGF-D dependent manner as indicated by the loss of stimulation by silencing VEGF-D in 468LN cells. Furthermore, 468LN cells showed α9-dependent stimulation of migration/invasiveness by macrophage products. Finally, capacity for intra-tumoral lymphangiogenesis and lymphatic metastasis in nude mice was completely abrogated by stable knock-down of either VEGF-D or α9 in 468LN cells. CONCLUSION: Differential capacity for VEGF-D production and α9ß1 integrin expression by 468LN cells jointly contributed to their lymphatic metastatic phenotype.

Lectin-induced activation of plasma membrane NADPH oxidase in cholesterol-depleted human neutrophils.

The gp91phox subunit of flavocytochrome b(558) is the catalytic core of the phagocyte plasma membrane NADPH oxidase. Its activation occurs within lipid rafts and requires translocation of four subunits to flavocytochrome b(558). gp91phox is the only glycosylated subunit of NADPH oxidase and no data exist about the structure or function of its glycans. Glycans, however, bind to lectins and this can stimulate NADPH oxidase activity. Given this information, we hypothesized that lectin-gp91phox interactions would facilitate the assembly of a functionally active NADPH oxidase in the absence of lipid rafts. To test this, we used lectins with different carbohydrate-binding specificity to examine the effects on H(2)O(2) generation by human neutrophils treated with the lipid raft disrupting agent methyl-ß-cyclodextrin (MßCD). MßCD treatment removed membrane cholesterol, caused changes in cell morphology, inhibited lectin-induced cell aggregation, and delayed lectin-induced assembly of the NADPH oxidase complex. More importantly, MßCD treatment either stimulated or inhibited H(2)O(2) production in a lectin-dependent manner. Together, these results show selectivity in lectin binding to gp91phox, and provide evidence for the biochemical structures of the gp91phox glycans. Furthermore, the data also indicate that in the absence of lipid rafts, neutrophil NADPH oxidase activity can be altered by these select lectins.

Chitin hydrolysate stimulates VEGF-C synthesis by MDA-MB-231 breast cancer cells.

Up-regulation of VEGF-C (vascular endothelial growth factor C), a most potent lymphangiogenic factor, is associated with inflammation and cancer metastasis. Identification of stimuli contributing to these processes is a challenging task. I demonstrate in this paper that chitin hydrolysate served as a strong inducer of VEGF-C synthesis by human breast cancer MDA-MB-231 cells, increasing the secretion of VEGF-C to the cell culture medium as much as by 10-fold in comparison with the basal production. A moderate increase of VEGF-C secretion was also observed in the presence of hypertonic doses of NaCl, which mimicked the matrix of chitin hydrolysate stock solution, and in the presence of chitin-binding lectin, WGA (wheat germ agglutinin). WGA, but not chitin hydrolysate, significantly affected the morphology of cells, which become smaller and rounded as assessed by viewing the actin cytoskeleton. Moreover, chitin hydrolysate inhibited the lectin effect on the cytoskeleton and sustained the overproduction of VEGF-C indicating that WGA-independent receptors were responsible for chitin-mediated stimulation of VEGF-C synthesis. These results suggest a novel function of chitin-derived oligosaccharides as VEGF-C stimuli.

Redox regulation of morphology, cell stiffness, and lectin-induced aggregation of human platelets.

Redox regulation and carbohydrate recognition are potent molecular mechanisms which can contribute to platelet aggregation in response to various stimuli. The purpose of this study is to investigate the relationship between these mechanisms and to examine whether cell surface glycocalyx and cell stiffness of human platelets are sensitive to the redox potential formed by glutathione. To this end, human platelets were treated with different concentrations (0.05 µM to 6 mM) and ratios of reduced or oxidized glutathione (GSH or GSSG), and platelet morphological, mechanical, and functional properties were determined using conventional light microscopy, atomic force microscopy, and lectin-induced cell aggregation analysis. It was found that lowering the glutathione redox potential changed platelet morphology and increased platelet stiffness as well as modulated nonuniformly platelet aggregation in response to plant lectins with different carbohydrate-binding specificity including wheat germ agglutinin, Sambucus nigra agglutinin, and Canavalia ensiformis agglutinin. Extracellular redox potential and redox buffering capacity of the GSSG/2GSH couple were shown to control the availability of specific lectin-binding glycoligands on the cell surface, while the intracellular glutathione redox state affected the general functional ability of platelets to be aggregated independently of the type of lectins. Our data provide the first experimental evidence that glutathione as a redox molecule can affect the mechanical stiffness of human platelets and induce changes of the cell surface glycocalyx, which may represent a new mechanism of redox regulation of intercellular contacts.