Junctional Adhesion Molecules are required for melanoma cell lines transendothelial migration in vitro.

International audienceOne of the main steps of metastasis is extravasation, a phenomenon well described in lymphocytes, but remaining to be fully uncovered for melanoma. Junctional Adhesion Molecules (JAMs) are controlling the transendothelial migration of leukocytes. To date the role of the JAM proteins, notably JAM-A and JAM-C, has not been examined in melanoma. Here, we compared two melanoma tumor cell lines, A375 and SLM8 cells, the A375 cell line being four times more efficient than the SLM8 cells in the crossing of the endothelial monolayer. We evidence the differential expression of JAM-A and JAM-C in these cell lines with JAM-C mainly expressed in the A375 cell line, and JAM-A detected preferentially in the SLM8 cells. To further dissect the respective roles of these proteins, we used both siRNA and blocking antibodies to decrease JAM-A and JAM-C expression

Link between the EZH2 noncanonical pathway and microtubule organization center polarization during early T lymphopoiesis

International audienceAbstract EZH2 plays an essential role at the β-selection checkpoint of T lymphopoiesis by regulating histone H3 lysine 27 trimethylation (H3K27me3) via its canonical mode of action. Increasing data suggest that EZH2 could also regulate other cellular functions, such as cytoskeletal reorganization, via its noncanonical pathway. Consequently, we investigated whether the EZH2 noncanonical pathway could be involved in early T-cell maturation, which requires cell polarization. We observed that EZH2 localization is tightly regulated during the early stages of T-cell development and that EZH2 relocalizes in the nucleus of double-negative thymocytes enduring TCRβ recombination and β-selection processes. Furthermore, we observed that EZH2 and EED, but not Suz12, colocalize with the microtubule organization center (MTOC), which might prevent its inappropriate polarization in double negative cells. In accordance with these results, we evidenced the existence of direct or indirect interaction between EED and α-tubulin. Taken together, these results suggest that the EZH2 noncanonical pathway, in association with EED, is involved in the early stages of T-cell maturation

Nanog and Oct4 overexpression increases motility and transmigration of melanoma cells.

International audiencePURPOSE: Melanoma tumors are highly heterogeneous and can undergo phenotypic modifications depending on their plasticity and the microenvironment, with shifts between proliferative and invasive states. We have shown that melanoma cells, grown as spheroids in a neural crest cell medium, polarize toward an invasive and motile phenotype, in agreement with transcriptomic modulations, including the up-regulation of Nanog and Oct4. Overexpression of these genes was shown to be associated with poor prognosis and metastatic forms of some cancers. We thus investigated implication of Nanog and Oct4, two embryonic transcription factors, in melanoma motility. METHODS: Our team used stable transfection of Nanog or Oct4 in A375 melanoma cell line to investigate motility in a wound healing assay and a transendothelial migration assay. Using semiquantitative RT-PCR, expression of two gene panels involved either in mesenchymal motility or in amoeboid migration was studied. RESULTS: Strongly enhanced capacities of motility and extravasation were observed with cells overexpressing Oct4 and Nanog. The A375 cell line has been described as having a mesenchymal migration type. However, in the Oct4 and Nanog transfectants, several amoeboid migration markers are strongly induced. Accordingly, amoeboid migration inhibitors decrease significantly the transmigration of Oct4- and Nanog-expressing cells through endothelial cells. CONCLUSIONS: We propose here that Nanog and Oct4 pluripotency marker expression in melanoma cells increases the transmigration capacity of these cells through the gain of amoeboid motility, leading to higher invasiveness and aggressiveness

Socioenvironmental stressors encountered during spaceflight partially affect the murine TCR-β repertoire and increase its self-reactivity

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Analysis of femurs from mice embarked on board BION-M1 biosatellite reveals a decrease in immune cell development, including B cells, after 1 wk of recovery on Earth

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Coexpression of major histocompatibility complex class II with chemokines and nuclear NFkappaB p50 in melanoma: a rational for their association with poor prognosis.

International audienceThe constitutive expression of major histocompatibility complex class II (MHC II) molecules in melanoma is highly unusual and has been associated with unfavorable clinical outcome and higher metastatic dissemination. This association remains poorly understood and therefore, in this study we looked to whether it is caused by intracellular events that promote tumor progression. We previously reported that MHC II expression in melanoma cells requires active mitogen-activated protein kinase/extracellular signal-related kinase. However, our comparative and molecular analyses of a panel of melanoma cell lines herein provide clear evidence that mitogen-activated protein kinase/extracellular signal-related kinase is not sufficient for HLA-DR expression. We found that the expression of HLA-DR in these tumors rather coincides with the expression of CXCL-1 and CXCL-8 chemokines, both known to be expressed in tumors that invade early and are related to invasive stages of melanoma. The expression of HLA-DR also nicely paralleled that of the nuclear NFkappaB p50 subunit, regulating the expression of these chemokines in melanoma and previously correlated with poor prognosis of melanoma patients, although we provide evidence that NFkappaB is not directly regulating MHC II expression level. The molecular basis for class II transactivator and HLA-DR expression in melanoma therefore remains unsolved, but our findings linking together the expression of HLA-DR, of chemokines involved in invasiveness, and of nuclear NFkappaB p50 strongly support the content that MHC II may be a marker of invasive primary melanoma, and could explain the long-standing association of MHC II expression with overall poor prognosis and unfavorable clinical outcome

LFA-1 and ICAM-1 expression induced during melanoma-endothelial cell co-culture favors the transendothelial migration of melanoma cell lines in vitro

<p>Abstract</p> <p>Background</p> <p>Patients with metastatic melanoma have a poor median rate of survival. It is therefore necessary to increase our knowledge about melanoma cell dissemination which includes extravasation, where cancer cells cross the endothelial barrier. Extravasation is well understood during travelling of white blood cells, and involves integrins such as LFA-1 (composed of two chains, CD11a and CD18) expressed by T cells, while ICAM-1 is induced during inflammation by endothelial cells. Although melanoma cell lines cross endothelial cell barriers, they do not express LFA-1. We therefore hypothesized that melanoma-endothelial cell co-culture might induce the LFA-1/ICAM ligand/receptor couple during melanoma transmigration.</p> <p>Methods</p> <p>A transwell approach has been used as well as blocking antibodies against CD11a, CD18 and ICAM-1. Data were analyzed with an epifluorescence microscope. Fluorescence intensity was quantified with the ImageJ software.</p> <p>Results</p> <p>We show here that HUVEC-conditioned medium induce cell-surface expression of LFA-1 on melanoma cell lines. Similarly melanoma-conditioned medium activates ICAM-1 expression in endothelial cells. Accordingly blocking antibodies of ICAM-1, CD11a or CD18 strongly decrease melanoma transmigration. We therefore demonstrate that melanoma cells can cross endothelial monolayers in vitro due to the induction of ICAM-1 and LFA-1 occurring during the co-culture of melanoma and endothelial cells. Our data further suggest a role of LFA-1 and ICAM-1 in the formation of melanoma cell clumps enhancing tumor cell transmigration.</p> <p>Conclusion</p> <p>Melanoma-endothelial cell co-culture induces LFA-1 and ICAM-1 expression, thereby favoring in vitro melanoma trans-migration.</p