270 research outputs found

    Immuno-Modulatory Properties of a Quinolin-2-(1H)-on-3-Carboxamide Derivative: Relevance in Multiple Sclerosis

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    Background: We have recently released the structure of a class of quinolin-2-(1H)-on-3-carboxamide derivatives and among them; the drug A2 has the highest CB2 receptor selectivity. Objective: In this work we assessed the immuno-modulatory properties of A2 in lymphocytes isolated from peripheral blood of multiple sclerosis patients and healthy donors. Methods: Cell proliferative response was measured by 3H-thymidine incorporation, cell viability and apoptosis by trypan blue, annexin V staining and western blot. Cell activation was investigated by flow cytometry and molecular pathways by western blot. Results: A2 exerted anti-proliferative effects with down-regulation of TNF-α , IL-10 and Rantes in both cell types. No relevant changes were observed in cell viability between the two cell types. In cells from healthy subjects, A2 did not induce apoptosis, inhibited the cell cycle and similarly down-regulated in CD4+T cells the markers CD69, CD25, CD49d and CD54. Indeed, A2 also inhibited the phosphorylation of Akt, NF-kB, IKKα/β, ERK and blocked the expression of Cox-2 and CB2 receptor. Published patents also describe CB2 receptor agonists like purine derivatives. Differently, in cells from patients, A2 did not affect CD49d, while potently blocked CD54 expression. A2 inhibitory effects of Akt and Cox-2 expression were confirmed, whereas unchanged level of the CB2 receptor was observed in these cells. Conclusion: We reported similar effects of A2 in both cell types; however, a different mechanism of action might be suggested in cells from patients concerning cell activation and CB2 receptor expression. Overall, these data suggest an anti-inflammatory profile of A2 with potential implication in multiple sclerosis

    Thyrotropin modulates low density lipoprotein binding activity in FRTL-5 thyroid cells.

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    Abstract FRTL-5 cells possess high affinity low density lipoprotein (LDL) receptors which bind, internalize, and degrade LDL. When FRTL-5 cells are deprived of thyrotropin (TSH) the binding of LDL increases more than 2-fold. Upon addition of TSH, at a concentration of 1 x 10(-10) M or greater, LDL binding decreases rapidly and within 24 h reaches the level which is typical of FRTL-5 cells chronically stimulated by TSH. The data available suggest that TSH-dependent down-regulation of LDL receptor activity is exerted through a reduction of the number of active LDL receptors, with no change in affinity. It is unlikely that the synthesis of LDL receptors is impaired, since LDL receptor messenger RNA is not decreased by TSH. The effect of the hormone on LDL receptor activity can be mimicked by 8-Br-cAMP and is completely abolished by the protein synthesis inhibitor cycloheximide but not by actinomycin D. TSH regulation of LDL receptor activity is lost in v-ras Ki-transformed FRTL-5 cells (Ki Mol) which also have lost TSH dependence for adenylate cyclase activation and growth. However, 8-Br-cAMP decreases LDL binding in Ki Mol FRTL-5 cells. The reduced availability of LDL receptor in TSH-stimulated FRTL-5 cells may be related to the increased membrane fluidity (Beguinot, F., Beguinot, L., Tramontano, D., Duilio, C., Formisano, S., Bifulco, M., Ambesi-Impiombato, F. S., and Aloj, S. M. (1987) J. Biol. Chem. 262, 1575-1582) or may reflect increased degradation of LDL receptors. We propose that a lower cholesterol uptake is needed in an actively proliferating cell population, to increase the production of isoprenoids whether it be for cholesterol biosynthesis or for the synthesis of other compounds requiring isoprenoid precursors

    Role of the small GTPase Rab7 in the late endocytic pathway.

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    Rab7 is a small GTPase localized to the late endosomal compartment. Its function was investigated by overexpressing dominant negative or constitutively active mutants in BHK-21 cells. The effects of such overexpression on the internalization and/or degradation of different endocytic markers and on the morphology of the late endosomal compartment were analyzed. We observed a marked inhibition of the degradation of 125I-low density lipoproteins in cells transfected with the Rab7 dominant negative mutants while the rate of internalization was not affected. Moreover in these cells there was an accumulation of many small vesicles scattered throughout the cytoplasm. In contrast, overexpression of the activating mutants led to the appearance of atypically large endocytic structures and caused a dramatic change in the distribution of the cation-independent mannose 6-phosphate receptor. Our data indicate that the Rab7 protein in mammalian cells is present on a late endosomal compartment much larger than the compartment labeled by the cation-independent mannose 6-phosphate receptor. Rab7 also appears to play a fundamental role in controlling late endocytic membrane traffic

    The endocannabinoid anandamide inhibits colon cancer cell growth by modulating different survival and proliferating pathways

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    The Endocannabinoid System (ECS) comprising the CB1 and CB2 receptors and their endogenous ligands is a central signalling system regulating food intake and energy balance. It is also present in peripheral tissues where it is involved in cell proliferation and survival. It has been shown that in colon cancer cells, the CB1 receptor antagonist SR171416 reduces colon cancer cell growth by acting as an inverse agonist rather than an antagonist [1]. Starting from this observation and from evidence indicating that some biological responses to cannabinoids depend on estrogen levels and some selective estrogen receptor modulators can bind the CB1 receptor [2], we aimed to study the effects of the CB1 receptor ligand anandamide (AEA) on colon cancer cell proliferation and its ability to modulate some survival and proliferating pathways including Akt, MAPK/ERK and estrogen receptor (ER) b signalling which is the predominant ER pathway in colonic epithelium. We used an AEA-analogue and a selective inhibitor of fatty acid amide hydrolase (FAAH) that enhances intracellular levels of AEA and studied proliferation and cell cycle progression on human adenocarcinoma cells DLD1 and SW620. Our results showed that increased levels of AEA significantly reduced cell proliferation in both cell lines at 24 and 48 h also inducing an S phase cell cycle accumulation. The AEA-induced inhibition of cell growth was mediated by a reduced expression of phoshoAkt and phosphoERK and, at the same time, by an induction of ERβ expression. These data suggest that AEA can reduces colon cancer cell proliferation by interfering with different signalling pathways
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