Diffuse vesicular distribution of Rab3D in the polarized neuroendocrine cell line AtT-20

AbstractThe neuroendocrine cell line AtT-20 has two types of storage vesicles: dense core granules and synaptic vesicles, both sequestered at the tip of the processes. Here we show that Rab3D protein, which is abundant in fat cells, is also expressed in AtT-20 cells. Differently from Rab3A, which is localized in secretory vesicles accumulated at the tips, Rab3D has a diffuse vesicular distribution in the cytoplasm of the cell body, the processes and the tips. In AtT-20 cells, Rab3D may define a regulated secretory pathway which functions independently from cell polarity

Physiology and pathophysiology of the RANKL/RANK system

The bone resorption activity of osteoclasts is regulated at many levels, including differentiation of their monocytes precursors, fusion into multi-nucleated cells, migration to the resorption site, polarization of the mature osteoclasts and assembly of a podosome-based sealing zone. Another function of osteoclasts is relative to the integrity of the actin cytoskeleton, depending on the substratum upon which the osteoclasts are spread. There are two different structures of actin known as podosomes and sealing zone, actived in specialized matrix contacts and delimiting the membrane domain, where the ruffled border is formed. When a dual coculture of murine osteoblasts and murine mononuclear monocytes, in absolute absence of exogenous cytokines and other growth factors, was cultured on glass, the basic architecture of podosomes units and ruffled border was maintained regularly (1). We studied the osteoclast morphology and its behaviour in adhesion and in vesicle traffic by combination of light microscopy immunohistochemistry and transmission electron microscopy (TEM) immunolabeling. The adhesion and the fusion of preosteoclasts were observed by scanner electron microscopy (SEM). The osteoclasts produced by our physiological dual co-culture (without interaction of specific cytokines) are functionally and biologically active TRAP + and multinucleated cells. In fact the role of RANK, expressed by osteoblasts, controls the modulation of OPG bioavailability in the extracellular compartment. The fusion of monocytes is influenced by the presence of osteoblasts, that is based on RANK-RANKL interaction and communication between osteoblasts and preosteoclasts mediated by several molecules (2). Osteoclasts and osteoblasts can make direct contact, allowing membrane-bound ligands and receptors to interact and initiate intracellular signalling. RANKL-RANK complexes are likely internalized via rafts and then degraded in lysosomes. A recent study has shown that membrane-bound RANKL complexes to OPG is internalized by endocytosis process. A potential interaction of RANKL with clathrin components prior to the OPG binding, as shown by the kinetic results, OPG is intracellularly degraded after being internalized: our observation of osteoclast membrane at TEM has shown that immunogold labelled RANKL colocalizes with immunoglod labelled clathrin via the clathrin-coated-pit-mediated (3) pathway and both proteins are degraded by lysosome and proteasome pathway. 1) Nicolin V. et al.,. 2006 J.Mol.Histol.37 :171-177 2) Narducci P., et al.,. Acta Histochem.113(2):73-81. 3) Narducci P., et al., 2010, Eur J Histochem 54(1):e6

Re‐examination of the mechanisms regulating nuclear inositol lipid metabolism

Although inositol lipids constitute only a very minor proportion of total cellular lipids, they have received immense attention by scientists since it was discovered that they play key roles in a wide range of important cellular processes. In the late 1980s, it was suggested that these lipids are also present within the cell nucleus. Albeit the early reports about the intranuclear localization of phosphoinositides were met by skepticism and disbelief, compelling evidence has subsequently been accumulated convincingly showing that a phosphoinositide cycle is present at the nuclear level and may be activated in response to stimuli that do not activate the inositol lipid metabolism localized at the plasma membrane. Very recently, intriguing new data have highlighted that some of the mechanisms regulating nuclear inositol lipid metabolism differ in a substantial way from those operating at the cell periphery. Here, we provide an overview of recent findings regarding the regulation of both nuclear phosphatidylinositol 3‐kinase and phosphoinositide‐specific phospholipase C‐β1

In vitro evaluation of the biocompatibility of dental alloys: fibronectin expression patterns and relationships to cellular proliferation rates.

This short-term (72- to 96-hour) in vitro study on fibroblasts evaluated the biocompatibility of 3 single-phase dental alloys by determining cellular proliferation rates and the expression of a glycoprotein, fibronectin, which is involved in cellular adhesion processes. METHOD AND MATERIALS: Flow 2002 fibroblasts were cultured together with 3 single-phase dental alloys of different composition. Proliferation rates were determined by 5-bromodeoxyuridine incorporation. Fibronectin expression was determined by indirect immunofluorescence. RESULTS: At 72 hours, cells cultured with the alloy containing the lowest amount of noble elements (gold, platinum, and palladium) and the highest amount of silver exhibited significantly less proliferation than did controls. At 96 hours, only cultures with the alloy containing the greatest amount of noble elements behaved in a way similar to controls. Fibronectin organization in fibrils and in focal adhesions was correlated to higher cellular proliferation rates. CONCLUSION: Fibronectin organization could be a useful tool to determine the biocompatibility of dental alloys. Among the noble elements, palladium by itself exhibits very good biocompatibility. These indications could be useful for practitioners in the choice of the best alloy for specific clinical applications