Éva Monostori

Biphasic Effect of Recombinant Galectin‐1 on the Growth and Death of Early Hematopoietic Cells

Galectin‐1 is a member of the family of β‐galactoside binding animal lectins, galectins. Its presence in the bone marrow has been detected; however, its role in the regulation of hematopoiesis is unknown. In the present study, we have evaluated the effect of recombinant human galectin‐1 on the proliferation and survival of murine and human hematopoietic stem and progenitor cells. We show that low amount of galectin‐1 (10 ng/ml) increases the formation of granulocyte‐macrophage and erythroid colonies and the frequencies of day‐7 cobblestone area–forming cells on a lactose‐inhibitable fashion. In contrast, high amount of galectin‐1 (10 μg/ml) dramatically reduces the growth of the committed blood‐forming progenitor cells as well as the much younger, lineage‐negative hematopoietic cells (day‐28 to −35 cobblestone area–forming cells). This inhibition is not blocked by lactose and, therefore, is largely independent of the β‐galactoside–binding site of the lectin. Furthermore, assays to detect apoptosis render it likely that the high amount of galectin‐1 acts as a classical proapoptotic factor for the premature hematopoietic cells.

Mechanism of tumor cell-induced T-cell apoptosis mediated by galectin-1.

Galectin-1 (Gal-1) has been implicated in tumor progression partly via the induction of T-cell apoptosis. However the mechanism of Gal-1 induced T-cell death was mostly studied using recombinant, soluble Gal-1 producing controversial results. To explore the true mechanism of Gal-1 and hence tumor cell-induced T-cell death, we applied co-cultures of tumor cells and T-cells thus avoiding artificial circumstances generated using recombinant protein. T-cells died when co-cultured with Gal-1-expressing but survived with Gal-1 non-expressing tumor cells. Removing tumor cell surface Gal-1 or knocking down Gal-1 expression resulted in diminution of T-cell apoptosis. Gal-1 transgenic or soluble Gal-1 treated HeLa cells became cytotoxic. Stimulation of apoptosis required interaction between the tumor and T-cells, presence of p56lck and ZAP70, decrease of mitochondrial membrane potential and caspase activation. Hence tumor cell-derived Gal-1 might efficiently contribute to tumor self-defense. Moreover this system resolves the discrepancies obtained using recombinant Gal-1 in T-cell apoptosis studies.

Activated T-cells and pro-inflammatory cytokines differentially regulate prostaglandin E2 secretion by mesenchymal stem cells

In recent years it has become clear that mesenchymal stem or stromal cells (MSCs) are capable of modulating inflammatory and immune responses through interaction with a wide variety of cells. Whereas several studies indicated that PGE2 is one of the chief soluble mediators involved in these processes, here we investigated prostaglandin E2 (PGE2) production of murine bone marrow- (BM-) and adipose tissue- (Ad-) derived MSCs stimulated with pro-inflammatory cytokines TNF-α and IFN-γ, or co-cultured with ConA-induced T-cell blasts. We found that both MSC populations are able to produce high amounts of PGE2 in MSC/activated T-cell co-cultures. This effect was markedly attenuated when direct cell-cell contact was prevented in transwell system, indicating that the elicitation of the PGE2 secretion of MSCs is contact-dependent in this experimental setting. In contrast, when soluble recombinant pro-inflammatory cytokines were added to the MSC cultures, TNF-α and IFN-γ act synergistically to induce PGE2 production, whereas only high amount of TNF-α but not IFN-γ was able to do so alone. Although the PGE2 secretion by MSCs was completely abrogated by addition of indomethacin under all culture conditions tested, L-NMA, a NOS inhibitor could only partially inhibit it when the cells were elicited in the concomitant presence of TNF-α and IFN-γ. These results, combined with others, suggest that NO acts downstream of IFN-γ but upstream of COX2. Taken together, our findings demonstrate that the induction of PGE2 secretion by BM- and Ad-MSCs is not mediated by a single or unique, nonredundant molecular mechanism under different experimental conditions.

Receptor tyrosine phosphatase, CD45 binds galectin-1 but does not mediate its apoptotic signal in T cell lines

Galectin-1 (Gal-1) is an endogenous mammalian S-type lectin with highly pleiotropic effect on different tissues. The viability of the lymphoid cells is reduced by gal-1 by triggering apoptosis, however, the mechanism of the gal-1 induced apoptosis is still under investigation. The receptor tyrosine phosphatase, CD45, a heavily glycosylated cell surface molecule binds to gal-1 with high affinity, however, its contribution to the gal-1 induced apoptosis is still controversial. In this study we show that galectin-1 binds to cells deficient for CD45, although CD45 is one of the galectin-1-binding cell surface proteins on T cells. Moreover, the CD45 deficient Jurkat variant, J45.01 responds readily with tyrosine phosphorylation and subsequent apoptosis to galectin-1 treatment in a similar degree as its wild type counterpart, Jurkat does. These results strongly indicate that CD45 is not the receptor via gal-1 mediates the apoptotic signal into the cells as it was suggested in previous studies.

Role of p56lck and ZAP70-mediated tyrosine phosphorylation in galectin-1-induced cell death

Role of p56 lck and ZAP70-mediated tyrosine phosphorylation in galectin-1-induced cell death

Digestion rate in relation to starvation in the larva of a carabid predator, Poecilus cupreus

Prey antigen levels were determined by a quantitative enzyme‐linked immunosorbent assay in larvae of a carabid predator, Poecilus cupreus L. under laboratory conditions. We determined prey consumption and rate of prey protein decay in larvae having starved for different periods. Prey consumption increased exponentially over 7 days of starvation. The rate of prey immunoreactive material decay seemed to be logarithmic, similar in larvae which had starved for different periods. Most of the digestion took place in the first 3 days period after feeding, irrespective of the amounts ingested. The rate of decay increased with increasing period of starvation. There were also signs of food retention in the alimentary canal in larvae which starved longer.

Identical, similar or different? Learning about immunomodulatory function of mesenchymal stem cells isolated from various mouse tissues: bone marrow, spleen, thymus and aorta wall

Mesenchymal stem or multipotent stromal cells (MSCs) have been implicated in tissue maintenance and repair and regulating immune effector cells through different mechanisms. These functions in mouse were primarily described for bone marrow (BM)-derived MSCs. To learn more about MSCs of different tissue origin, we compared the immunophenotype, differentiation ability to adipocyte and bone and immunomodulatory activity of MSCs isolated from BM, spleen, thymus and aorta wall of 14-day-old C57Bl/6 mice. The established cell lines fulfilled the requirements described for MSCs in terms of morphology, surface marker expression and differentiation potential although they were distinguishable regarding the expression pattern of the MSC markers and ability generating other cell types. Most importantly, a remarkable diversity was shown in the capacity of inhibition of mitogen- and alloantigen-induced T-cell proliferation, since BM- and spleen-derived MSCs were the most powerful aorta-derived MSCs were less effective, whereas thymus-derived mesenchymal cells were unable to block T-cell growth in vitro. Accordingly, BM, spleen and aorta, but not thymus-derived MSCs, in combination with BM hematopoietic cells were equally efficient to prevent streptozotocin-induced diabetes in vivo. These findings suggested that MSCs residing in different organs might stem from common ancestor; however, once populating into a given tissue microenvironment, they acquire specific properties mainly in the term of the immunoregulatory function.

Co-localization of galectin-1 with GM1 ganglioside in the course of its clathrin- and raft-dependent endocytosis

Abstract.Mammalian galectin-1 (Gal-1), a β-galactoside-binding lectin has a prominent role in regulating cell adhesion, cell growth and immune responses. Downregulation of these biological functions may occur via internalization of Gal-1. In the present study we have investigated the mechanism and possible mediator(s) of Gal-1 endocytosis. We show that internalization occurs at a temperature higher than 22 °C in an energy dependent fashion. After one hour incubation Gal-1 localizes in the Golgi system within the cells, and then disappears without accumulation in degradation compartments, such as lysosomes. Based on their strong intracellular co-localization, two glycoconjugates, GM1 ganglioside and CD7 are implicated in the sorting of internalized Gal-1 into Golgi. Other known Gal-1 binding glycoproteins on T cells (CD2, CD3, CD43 and CD45) do not cointernalize with the lectin. Internalization of Gal-1 depends on its lectin activity and follows dual pathways involving clathrin-coated vesicles and raft-dependent endocytosis.

Foldameric α/β-peptide analogs of the β-sheet-forming antiangiogenic anginex: structure and bioactivity.

The principles of β-sheet folding and design for α-peptidic sequences are well established, while those for sheet mimetics containing homologated amino acid building blocks are still under investigation. To reveal the structure-function relations of β-amino-acid-containing foldamers, we followed a top-down approach to study a series of α/β-peptidic analogs of anginex, a β-sheet-forming antiangiogenic peptide. Eight anginex analogs were developed by systematic α → β(3) substitutions and analyzed by using NMR and CD spectroscopy. The foldamers retained the β-sheet tendency, though with a decreased folding propensity. β-Sheet formation could be induced by a micellar environment, similarly to that of the parent peptide. The destructuring effect was higher when the α → β(3) exchange was located in the β-sheet core. Analysis of the β-sheet stability versus substitution pattern and the local conformational bias of the bulky β(3)V and β(3)I residues revealed that a mismatch between the H-bonding preferences of the α- and β-residues played a minor role in the structure-breaking effect. Temperature-dependent CD and NMR measurements showed that the hydrophobic stabilization was scaled-down for the α/β-peptides. Analysis of the biological activity of the foldamer peptides showed that four anginex derivatives dose-dependently inhibited the proliferation of a mouse endothelial cell line. The α → β(3) substitution strategy applied in this work can be a useful approach to the construction of bioactive β-sheet mimetics with a reduced aggregation tendency and improved pharmacokinetic properties.

Identification of galectin-1 as a critical factor in function of mouse mesenchymal stromal cell-mediated tumor promotion.

Bone marrow derived mesenchymal stromal cells (MSCs) have recently been implicated as one source of the tumor-associated stroma, which plays essential role in regulating tumor progression. In spite of the intensive research, the individual factors in MSCs controlling tumor progression have not been adequately defined. In the present study we have examined the role of galectin-1 (Gal-1), a protein highly expressed in tumors with poor prognosis, in MSCs in the course of tumor development. Co-transplantation of wild type MSCs with 4T1 mouse breast carcinoma cells enhances the incidence of palpable tumors, growth, vascularization and metastasis. It also reduces survival compared to animals treated with tumor cells alone or in combination with Gal-1 knockout MSCs. In vitro studies show that the absence of Gal-1 in MSCs does not affect the number of migrating MSCs toward the tumor cells, which is supported by the in vivo migration of intravenously injected MSCs into the tumor. Moreover, differentiation of endothelial cells into blood vessel-like structures strongly depends on the expression of Gal-1 in MSCs. Vital role of Gal-1 in MSCs has been further verified in Gal-1 knockout mice. By administering B16F10 melanoma cells into Gal-1 deficient animals, tumor growth is highly reduced compared to wild type animals. Nevertheless, co-injection of wild type but not Gal-1 deficient MSCs results in dramatic tumor growth and development. These results confirm that galectin-1 is one of the critical factors in MSCs regulating tumor progression.