Ricardo Lascurain

Bacterial agglutination by the sialic acid specific serum lectin from Macrobrachium rosenbergii

We have isolated a serum lectin from the freshwater prawn, Macrobrachium rosenbergii that agglutinates Bacillus cereus and Aeromona sp. This lectin also agglutinates other bacteria such as Pasteurella haemolytica biotype A (capsular serotype 12), several serotypes from P. multocida and Staphylococcus aureus and to a lesser extent Escherichia coli and Salmonella arizona.Lectin recognition of well known polysaccharide components seems to involve several bacterial O-keto and O-methyl containing sugars, the N-acetyl-sugar residues and teichcoic from the polysaccharide cell wall.

MORPHOLOGY OF HEMOCYTES FROM THE FRESHWATER PRAWN MACROBRACHIUM ROSENBERGII

Using morphological criteria, we identified three types of blood cells in the freshwater prawn Macrobrachium rosenbergii. Hyaline hemocytes, the most abundant type, have few large cytoplasmic granules, a large nucleocytoplasmic ratio, and lyse spontaneously in the absence of anticoagulant. Granular hemocytes are heterogeneous in size and in density of their granules. They are phagocytic and readily spread on substrates. The third type of hemocytes, identified as undifferentiated hemocytes, are the least abundant. The hemocytes of this economically relevant crustacean are compared with blood cells of other decapods. J. Morphol. 234:147–153, 1997.

Intracellular expression of interleukin-4 and interferon-γ by a Mycobacterium tuberculosis antigen-stimulated CD4+ CD57+ T-cell subpopulation with memory phenotype in tuberculosis patients

In some chronic pathological conditions, antigen persistence activates and expands the CD4+ CD57+ T‐cell subset. The host immune response against tuberculosis infection is maintained through the continuous presence of antigen‐stimulated effector/memory helper T cells. To determine whether CD4+ CD57+ T cells were also expanded in human tuberculosis, we analysed (by flow cytometry) the phenotype of peripheral blood CD4+ T cells from 30 tuberculosis patients and 30 healthy controls. We observed a significant increase in the CD4+ CD57+ T‐cell subset in tuberculosis patients in comparison to healthy controls (P < 0·001). Most CD4+ CD57+ T cells exhibited a CD28− CD45RO+ CD62L− phenotype, which is associated with memory cells. In vitro, a higher number of antigen‐stimulated CD4+ CD57+ T cells produced intracellular interferon‐γ and interleukin‐4 compared with antigen‐stimulated CD4+ CD57− T cells (P < 0·001). These findings suggest that the majority of CD4+ CD57+ T cells correspond to a phenotype of activated memory T cells.

Specificity of Amaranthus leucocarpus lectin

We have demonstrated thatAmaranthus leucocarpus lectin hemagglutinating activity was powerfully inhibited by the T-antigen, containing Gal(β1–3)GalNAc(α1–3)Ser/Thr, and the Tn-antigen, which contains GalNAc(α1–3)Ser/Thr. This suggests that the acetamido group at C-2 and the axial -OH at C-4 of theN-acetyl-D-galactopyranosylamine ring are important for lectin binding. The hemagglutination assays also established that desialylated and Pronase-treated human typeO erythrocytes with an M phenotype were better recognized than erythrocytes from all other blood groups. The recognition was dependent on pH and ionic strength.

Pars planitis is associated with an increased frequency of effector-memory CD57+ T cells

Aim: To evaluate the frequency, phenotype and the potential function of CD57+ T cell subsets in patients with pars planitis. Methods: CD4+CD57+ and CD8+CD57+ T cells were quantitated in peripheral blood from 15 patients with pars planitis and 15 healthy controls. To evaluate the phenotype and potential function of CD57+ T cell subsets CCR7, CD27, CD28, CD45RA, CD45RO, intracellular IFN-&ggr;, IL-4, perforin and granzyme-A expression were assessed by flow cytometry. Results: CD57+ T cells subsets were increased in patients with pars planitis (p = 0.002). The majority of CD4+CD57+ T cells were CCR7−CD27−CD28−CD45RO+, while the most CD8+CD57+ T cells were CCR7−CD27−CD28−CD45RA+. The number of cells positive for intracellular IFN-&ggr; and IL-4 was higher in the CD57+ T cell populations. A greater number of CD8+CD57+ T cells than CD8+CD57− T cells were positive to perforin (p = 0.006) and granzyme-A (p = 0.01). Conclusions: CD57+ T cells had a phenotype associated with peripheral memory (CCR7−CD27−CD28−). Cytokine production by CD57+ T cells suggests that these cells may play a role in helper cell regulation. High expression of intracellular proteins involved in cytotoxicity suggests that CD8+CD57+ T cells may play an effector role. Taken together, this study proposes that CD57+ T cells function as memory-effector T cell subsets during pars planitis pathogenesis.

Sialylation is modulated through maturation in hemocytes from Macrobrachium rosenbergii.

In this work we identified in adult and juvenile freshwater prawn, Macrobrachium rosenbergii, three major type of circulating hemocytes: fusiform; rounded; and large ovoid hemocytes. Rounded and large hemocytes represent the first defense line, since this type of cells exerts phagocytic activity as well as lectin synthesis. Considering that glycosylation plays important roles in cell communication and as a target for pathogenic microorganisms, in this report was also described the main glycosidic modifications that occur in the large and rounded hemocytes from the freshwater prawn during maturation as determined with lectins. Neu5Acalpha2,6Gal, was identified homogeneously distributed in the membrane in 90% of hemocytes from juvenile organisms. Maturation of the freshwater prawn induced a decrease or complete loss of Neu5Acalpha2,6Gal residues that were replaced with Neu5Acalpha2,3 molecules in practically all hemocytes from adult organisms. This change was paralleled by a diminution in 9-O-acetyl-neuraminic acid (Neu5,9Ac(2)) expression. T and Tn antigens (Galbetal,3 GalNAcalpha1-0-Ser/Thr or GalNAcalpha1-0-Ser/Thr, respectively), as well as N-glycosidically linked glycans, seem to be highly conserved throughout maturation. Our results show that sialylation of freshwater prawn hemocytes is modulated throughout the maturation process.

Relevance of sialoglycoconjugates in murine thymocytes during maturation and selection in the thymus

Differentiation of most T lymphocytes is characterized not only by the variable expression of CD4/CD8 coreceptor molecules and increased surface density of the T cell antigen receptor, but also by changes in the glycosylation pattern of cell surface glycolipids or glycoproteins. In this work we evaluated the changes in the sialylation pattern in thymus sections from normal and dexamethasone treated mice. We used sialic acid specific lectins, such as Sambucus nigra agglutinin (SNA, NeuAcalpha2,6-Gal specific) and Maackia amurensis agglutinin (MAA, NeuAcalpha2,3-Gal specific). Our results indicate that the sialylation pattern was modified during the maturation process of thymic cells. The immature CD4-CD8- and CD4+CD8+ cortical thymocytes were recognized by SNA, whereas the mature single positive (CD4+ or CD8+) medullary cells, preferentially bound MAA lectin. However, in the corticomedullary region we found not only SNA+ cells, but also MAA+ cells. In the thymus of dexamethasone treated mice, the clusters of thymocytes undergoing apoptosis in the cortex were characteristically stained by SNA. These results suggest that in the initial stages of the differentiation pathway, a great number of thymocytes express an alpha2,6 linked sialic acid on their surface and as they progress to more mature stages there is a change in the sialylation pattern to alpha2,3 linked sialic acids probably due to a regulated expression of different sialyltransferases, which could be modulated by the thymic microenvironment.

Amaranthus leucocarpus Lectin Recognizes Human Naive T Cell Subpopulations

Amaranthus leucocarpus lectin (ALL) is specific for GalNAc residue found in the inner core of Gal beta 1,3GalNAc alpha 1,O-Ser/Thr disaccharide (T-antigen) or GalNAc alpha 1,O-Ser/Thr (Tn-antigen). Flow cytometric analysis using fluorescein-labeled lectin and monoclonal antibodies against human cell surface markers indicated that 5.7% of mononuclear cells from human healthy donors are recognized by ALL. These cells have the phenotype CD2+CD4+CD19- and most of the lymphocytes recognized are also CD27+, CD45RA+ and CD43+. ALL possesses mitogenic activity on lymphocytes after neuraminidase treatment. Our results indicate that the receptors recognized by ALL could be considered surface markers for naive human T lymphocyte subsets.

The interaction between Histoplasma capsulatum cell wall carbohydrates and host components: relevance in the immunomodulatory role of histoplasmosis

Histoplasma capsulatum is an intracellular fungal pathogen that causes respiratory and systemic disease by proliferating within phagocytic cells. The binding of H. capsulatum to phagocytes may be mediated by the pathogens cell wall carbohydrates, glucans, which consist of glucose homo and hetero-polymers and whose glycosydic linkage types differ between the yeast and mycelial phases. The alpha-1,3-glucan is considered relevant for H. capsulatum virulence, whereas the beta-1,3-glucan is antigenic and participates in the modulation of the host immune response. H. capsulatum cell wall components with lectin-like activity seem to interact with the host cell surface, while host membrane lectin-like receptors can recognize a particular fungal carbohydrate ligand. This review emphasizes the relevance of the main H. capsulatum and host carbohydrate-driven interactions that allow for binding and internalization of the fungal cell into phagocytes and its subsequent avoidance of intracellular elimination.

Differential expression of a 70 kDa O-glycoprotein on T cells: a possible marker for naive and early activated murine T cells.

We purified a 70 kDa O-glycoprotein that binds to the GalNAc specific lectin from Amaranthus leucocarpus (ALLr) and determined its expression pattern on T lymphocytes from different murine lymphoid organs. High level of ALLr expression was demonstrated in 95-98% of both CD4(+)8(+) and CD4(-)8(+) thymocytes, and in 80-95% of CD8(+) T cells from peripheral blood, lymph nodes, and spleen, whereas a minor fraction of CD4(+)8(-) thymocytes (46-67%) and peripheral CD4(+) T cells (9-40%) showed low ALLr expression. Peripheral CD19(+) B cells were ALLr negative and most of the peripheral ALL(+) T cells showed a CD62L(hi)CD45RB(hi)CD44(lo/-) phenotype, indicating features of naive cells. Mitogenic activation of peripheral T cells increased 3-fold the number of ALL(+)CD4(+) T cells 24 h after stimulation, as opposed to a >80% decrease in CD8(+) T cells 72 h after stimulation. Our results suggest that ALL detects a non-described surface O-glycoprotein selectively expressed by naive CD8(+) T cells and by early activated CD4(+) T cells.