Morphological and biochemical characterization of macrophages activated by carrageenan and lipopolysaccharide in vivo.
ABSTRACT Macrophages are able to recognize, internalize and destroy a large number of pathogens, thus restricting the infection until adaptive immunity is initiated. In this work our aim was to analyze the surface charge of cells activated by carrageenan (CAR) and lipopolysaccharide (LPS) through light and electron microscopy approaches as well as the release of inflammatory mediators in vitro. The ultrastuctural analysis and the light microscopy data showed that in vivo administration of CAR represents a potent inflammatory stimulation for macrophages leading to a high degree of spreading, an increase in their size, in the number of the intracellular vacuoles and membrane projections as compared to the macrophages collected from untreated animals as well as mice submitted to LPS. Our data demonstrated that CAR stimulated-macrophages displayed a remarkable increase in nitric oxide production and PGE2 release as compared to the cells collected from non-stimulated and stimulated mice with LPS in vivo. On the other hand, non-stimulated macrophages as well as macrophages stimulated by LPS produce almost the same quantities of TNF-alpha, while in vivo stimulation by CAR leads to a 30-40% increase of cytokine release in vitro compared to the other groups. In conclusion, our morphological and biochemical data clearly showed that in vivo stimulation with CAR induces a potent inflammatory response in macrophages representing an interesting model to analyze inflammatory responses.
Article: Cytoplasmic linker protein-170 enhances spreading and phagocytosis in activated macrophages by stabilizing microtubules.[show abstract] [hide abstract]
ABSTRACT: Activation of macrophages causes increased cell spreading, increased secretion of cytokines and matrix metalloproteinases, and enhanced phagocytosis. The intracellular mechanisms driving the up-regulation of these activities have not been completely clarified. We observe that classical activation of murine resident peritoneal or RAW 264.7 macrophages with a combination of IFN-gamma and LPS induces an increase in stabilized cytoplasmic microtubules (MTs), measured with an anti-acetylated alpha-tubulin Ab. We examined the mechanism of this MT stabilization and find that macrophage activation causes redistribution of the MT plus-end tracking protein, cytoplasmic linker protein-170 (CLIP-170). CLIP-170 is localized at the distal plus-ends of MTs in resting macrophages, but accumulates along the length of MTs in IFN-gamma/LPS-activated cells. A direct involvement of CLIP-170 in MT stabilization has not been thoroughly established. In this study, we show that expression of a mutant CLIP-170 chimeric protein (dominant-negative CLIP-170-GFP), lacking the MT-binding domain, prevents MT stabilization in activated RAW 264.7 macrophages. Furthermore, we find enhanced CLIP-170 association with MTs and MT stabilization by treating resting macrophages with okadaic acid, implicating the protein phosphatase 2A in CLIP-170 binding and MT stabilization in RAW 264.7 cells. Finally, we observed enhanced cell spreading and phagocytosis in both IFN-gamma/LPS-activated and okadaic acid-treated resting RAW 264.7 cells, which are markedly reduced in activated cells expressing dominant-negative CLIP-170-GFP. These results identify CLIP-170 as a key regulator of MT stabilization and establish a prominent role for stabilized MTs in cell spreading and phagocytosis in activated macrophages.The Journal of Immunology 10/2007; 179(6):3780-91. · 5.79 Impact Factor
[show abstract] [hide abstract]
ABSTRACT: Effects of amlodipine, lacidipine and nicardipine on acute phase of inflammation in the carregeenan model in intact rats were investigated in this study. In addition, the most effective dose of nicardipine that had the highest anti-inflammatory impact was investigated in carregeenan test in adrenalectomized rats. The effective dose of nicardipine was tested in the chronic phase of inflammation in the model of cotton pellet granuloma, and its efficiency was compared with diclofenac sodium. Amlodipine at 5 and 10 mg/kg doses showed 61%, 80%, lacidipine 73%, 34% and nicardipine 38%, 87% inhibition of carrageenan-induced inflammation, respectively. Nicardipine (10 mg/kg) and diclofenac sodium (25 mg/kg) showed 11.6% and 16.2% inhibition, respectively against carrageenan-induced edema. Diclofenac at 10 mg/kg showed 43% inhibition of the inflammation. In cotton pellet test, antiproliferative effects of nicardipine (10 mg/kg) and diclofenac sodium (10 mg/kg) were evaluated as 60% and 39.5%, respectively. The obtained results showed that calcium channel blockers and diclofenac sodium significantly blocked acute and chronic phases of inflammation in intact rats, but in adrenalectomized rats calcium channel blockers and diclofenac sodium had no significant antiinflammatory effect.Pharmacological reports: PR 58(5):692-9. · 2.44 Impact Factor