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L Naldini

Publications and source records attributed to L Naldini.

62 records · Page 4Linked to original sources

Cell-substratum interaction of cultured avian osteoclasts is mediated by specific adhesion structures.

The cell-substratum interaction was studied in cultures of osteoclasts isolated from the medullary bone of laying hens kept on low calcium diet. In fully spread osteoclasts, cell-substratum adhesion mostly occurred within a continuous paramarginal area that corresponded also to the location of a thick network of intermediate filaments of the vimentin type. In this area, regular rows of short protrusions contacting the substratum and often forming a cup-shaped adhesion area were observed in the electron microscope. These short protrusions showed a core of F-actin-containing material presumably organized as a network of microfilaments and surrounded by a rosette-like structure in which vinculin and alpha-actinin were found by immunofluorescence microscopy. Rosettes were superposable to dark circles in interference-reflection microscopy and thus represented circular forms of close cell-substratum contact. The core of ventral protrusions also contained, beside F-actin, fimbrin and alpha-actinin. Villin was absent. This form of cell-substratum contact occurring at the tip of a short ventral protrusion differed from other forms of cell-substratum contact and represented an osteoclast-specific adhesion device that might also be present in in vivo osteoclasts as well as in other normal and transformed cell types.

Actinin↗

Osteoclasts and monocytes have similar cytoskeletal structures and adhesion property in vitro.

The distribution of some cytoskeletal structures (microtubules, microfilaments, intermediate filaments) has been studied by indirect immunofluorescence microscopy and affinity purified antibodies in osteoclasts isolated from medullary bone of laying hens and in hen blood monocytes cultured in vitro. Both cell types show similar patterns of distribution of cytoskeletal structures and this further supports the concept that these cells are closely related. Osteoclasts and monocytes are also similar in their adhesion patterns, because they adhere to fibronectin-free areas and show closely comparable cell-to-substrate interactions when observed with interference reflection microscopy.

Animals↗

Microtubules and microfilaments in fixed and permeabilized cells are selectively decorated by nerve growth factor.

A specific antibody against nerve growth factor (NGF) and indirect immunofluorescence microscopy have been used to follow the in vitro binding of NGF to cells made permeable to large molecules. All cells tested, both target (sensory neurons and PCI2 cells) and nontarget (3T3, BKH 2I, C6 glioma cells), revealed a decoration of cytoskeletal structures which on the basis of their form, reactivity with antibodies, and sensitivity to specific drugs may be identified as microtubules (MTs) and microfilaments (MFs). The decoration of either structure depends on the fixation and permeabilization conditions: MFs, in the form of stress fibers, are stained by NGF when the plasma membrane is permeabilized with methanol/acetone; MTs become intensely stained when the plasma membrane is solubilized with a nonionic detergent in the presence of a MT-stabilizing medium. The two procedures do not affect the staining of these structures with specific antibodies. Binding of 125I-labeled NGF to PCI2 cells was not competitively inhibited by a 100-fold excess of several positively charged proteins but it was markedly decreased in the presence of DNase I. 125I-Labeled NGF interacted with MTs and F-actin (fixed with paraformaldehyde) in a range of concentrations similar to that used for their cellular localization with NGF-anti-NGF. Our studies show that the specificity and affinity of NGF binding to MTs and MFs is in the range of that of antibodies against tubulin and actin. The possible relevance of these findings to the mechanism of action of NGF in target cells is discussed.

Actins↗

Distribution of nerve growth factor in chick embryo sympathetic neurons in vitro.

The distribution of nerve growth factor (NGF) in chick embryo sympathetic neurons has been followed by two distinct procedures, that is, indirect immunofluorescence microscopy employing purified NGF antibodies and autoradiography after exposure of cells to [125I]NGF. This study shows that NGF bound to sympathetic neurons is not uniformly distributed but appears in spots over the surface of perikarya and along processes. If the same cells, after incubation with NGF and fixation, are treated with methanol-acetone to allow permeation of immunoglobulins across the plasma membrane, NGF antibody immunoreactive material is also found within the cytoplasm and notably in the paranuclear area only in sympathetic neurons. Analogous findings are obtained when sympathetic neurons are incubated in the presence of [125I]NGF, fixed, sectioned and processed for autoradiography. Also with this technique NGF appears to be localized in the cytoplasmic compartment and is found around the nucleus. These studies are discussed in connection with the results of similar experiments performed on a clonal line of NGF target cells known as PC12.

Animals↗

Intracellular distribution of nerve growth factor in rat pheochromocytoma PC12 cells: evidence for a perinuclear and intranuclear location.

The distribution of nerve growth factor (NGF) in rat pheochromocytoma cells (clone PC12) has been studied with two different techniques: immunofluorescence and autoradiography. It was found that NGF is progressively internalized in the cytoplasmic compartment and eventually accumulates in the form of discrete dots around the nucleus. A fraction of the internalized NGF appears within the nucleoplasm, often contiguous with the nucleolus. It is suggested that cytoplasmic and perinuclear NGF may be be in contact with a pool of tubulin or actin-like proteins in their soluble or organized form and play a key role in the process of arrest of division and neurite growth.

Actins↗

Lentiviral vectors: excellent tools for experimental gene transfer and promising candidates for gene therapy.

Lentiviral vectors are tools for gene transfer derived from lentiviruses. From their first application to now they have been strongly developed in design, in biosafety and in their ability of transgene expression into target cells. Primate and non-primate derived lentiviral vectors are now available and with both types of systems a lot of studies tuned to improve their performances in a large number of tissues are ongoing. Here we review the state of the art of lentiviral vector systems discussing their potential for gene therapy.

Animals↗

Characterization of the detergent solubilized receptor for gastrin-releasing peptide.

Properties of detergent solubilized gastrin-releasing peptide receptor were investigated. Swiss 3T3 membranes were covalently labeled with [125I]GRP and homobifunctional cross-linkers. A major labeled protein of 75 kDa was resolved using SDS-polyacrylamide gel electrophoresis. When the same preparation was solubilized with zwitterionic detergent and analyzed under nondenaturing conditions the protein bound radioactivity was resolved in two different peaks, a major one of apparent molecular weight 220,000 (peak 1) and a minor one of 80,000 (peak 2) both containing the 75 kDa protein. Specific ligand binding activity also eluted with peak 1. These results indicate that the active form of bombesin/GRP receptor is a large complex containing the 75 kDa ligand binding domain.

Animals↗