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At least 1,045 records · Page 58Linked to original sources
PubMed2003

Inactivation of Src family kinases inhibits angiogenesis in vivo: implications for a mechanism involving organization of the actin cytoskeleton.

Inhibition of angiogenesis could be a treatment strategy for diseases such as cancer, rheumatoid arthritis, and diabetic retinopathy. PP2 is a pharmacological inhibitor of Src family kinases and was found to inhibit FGF-2 induced angiogenesis in vivo. Experiments in vitro showed that PP2 inhibited invasive growth and sprouting of both endothelial and vascular smooth muscle cells into a fibrin matrix. PP2 inhibited the formation of lamellopodia and expression of kinase inactive c-Src reduced phosphorylation of cortactin and paxillin, suggesting a model in which Src kinases are involved in organization of the actin cytoskeleton. Consequently, endothelial cells expressing kinase inactive c-Src failed to spread and form cord-like structures on a collagen matrix. These data suggest that pharmacological inactivation of Src family kinases inhibits FGF-2 stimulated angiogenesis by interference with organization of the actin cytoskeleton in both endothelial and vascular smooth muscle cells, which affects cell migration.

Actin Cytoskeleton↗
PubMed1986

The membrane-associated cytoskeleton in cultured lens cells. Electron microscopical visualization in cleaved whole-mount preparations and platinum replicas.

The membrane-bound cytoskeleton of bovine lens cells was investigated using different electron-microscopic methods. Cells were grown on glass coverslips and fixed in 0.5% glutaraldehyde, post-fixed and prepared for critical-point drying following standardized methods. Cells were broken open before post-fixation or after critical-point drying in order to expose the cortical cytoplasm. Cleaved, critical-point-dried cells were subsequently rotary replicated. Cells were also cleaved on grids before post-fixation. These cells were examined directly in the microscope. All methods showed the presence of a cytoskeletal network. Bundles and starlike foci of microfilaments and microtubules were usually found in close association with the membrane. Occasionally also intermediate filaments were observed, in a few cases running close to the microtubules. The reliability of the methods to visualize the membrane-bound cytoskeleton of lens cells is discussed.

Actin Cytoskeleton↗
PubMed1999

Unique sequence of a high molecular weight myosin light chain kinase is involved in interaction with actin cytoskeleton.

Myosin light chain kinase (MLCK) is the key regulator of cell motility and smooth muscle contraction in higher vertebrates. We searched for the features of the high molecular weight MLCK (MLCK-210) associated with its unique N-terminal sequence not found in a more ubiquitous lower molecular weight MLCK (MLCK-108). MLCK-210 demonstrates stronger association with the Triton-insoluble cytoskeletons than MLCK-108, suggesting the role for this sequence in subcellular targeting. Indeed, the expressed unique domain of MLCK-210 binds and bundles F-actin in vitro and colocalises with the microfilaments in transfected cells reproducing endogenous MLCK-210 distribution. Thus, MLCK-210 features an extensive actin binding interface and, perhaps, acts as an actin cytoskeleton stabiliser.

Actin Cytoskeleton↗
PubMedDate not supplied

The role of cytoskeleton in cell changes under condition of simulated microgravity.

Single cells and cell culture are very good model for estimation of primary effects of gravitational changes. It is suggested that cell cytoskeleton plays a key role in mechanisms of adaptation to mechanical influences including gravitational ones. Our results demonstrated that cultured cells of human vascular endothelium (correction of endotheliun) are highly sensitive to hypogravity (clinorotation) and respond by significant decrease of cell proliferative activity. Simultaneously it was noted that the formation of confluent monolayer appeared early in cultures exposed to simulated microgravity due to accelerated cells spreading. Long-term hypogravity (several hours or days) leads to significant changes of cell cytoskeleton revealed as microfilament thinning and their redistribution within cell. Such changes were observed only in monolayer cells and not in cell suspensions. Gravitational forces as known to be modificators of cell adhesive ability and determine their mobility. Hypogravity environment stimulated endothelial cell migration in culture: 24-48 hrs pre-exposition to hypogravity significantly increased endothelial cell migration resulting in 2-3-fold acceleration of mechanically injured monolayer repair. Obtained results suggest that the effects of hypogravity on cultured human endothelial cells are, possibly, associated with protein kinase C and/or adenylate cyclase activity and are accompanied by noticeable functional cell changes.

Actin Cytoskeleton↗
PubMed1997

Organization of cytoskeleton controls the changes in cytosolic calcium of cold-shocked Nicotiana plumbaginifolia protoplasts.

Using Nicotiana plumbaginifolia constitutively expressing the recombinant bioluminescent calcium indicator, aequorin, it has been previously demonstrated that plant cells react to cold-shock by an immediate rise in cytosolic calcium. Such an opportune system has been exploited to address the regulatory pathway involved in the calcium response. For this purpose, we have used protoplasts derived from N. plumbaginifolia leaves that behave as the whole plant but with a better reproducibility. By both immunodetecting cytoskeletal components on membrane ghosts and measuring the relative change in cytosolic calcium, we demonstrate that the organization of the cytoskeleton has profound influences on the calcium response. The disruption of the microtubule meshwork by various active drugs, such as colchicin, oryzalin and vinblastin, leads to an important increase in the cytosolic calcium (up to 400 nM) in cold-shocked protoplasts over control. beta-Lumicolchicin, an inactive analogue of colchicin, is ineffective either on cytoplasmic calcium increase or on microtubule organization. A microfilament disrupting drug, cytochalasin D, exerts a slight stimulatory effect, whereas the simultaneous disruption of microtubule and microfilament meshworks results in a dramatic increase in the calcium response to cold-shock. The results described in the present paper illustrate the role of the intracellular organization and, more specifically, the role of cytoskeleton in controlling the intensity of calcium response to an extracellular stimulus.

Actin Cytoskeleton↗
PubMed1999

Drug-induced alterations in rat peritubular cell cytoskeleton result in proteoglycan synthesis modifications. Comparison with some intracellular signaling pathways.

The influence of phorbol myristate acetate (PMA), dibutyryl cAMP and insulin-like growth factor (IGF-1) as well as cytoskeletal disrupting drugs on morphological changes has been studied in peritubular cells isolated from immature rat testis. Morphological studies were combined with immunofluorescence investigations of cytoskeletal elements and their rearrangements by various agents. The results were correlated with modulation of proteoglycan synthesis. Peritubular cells exposed to dibutyryl cAMP or cytochalasin D were transformed from flattened, fibroblast-like into neuronal-like morphology. In such cells, destruction of actin filaments was accompanied with a 50% decrease in cell-associated proteoglycan synthesis as well as with oversulfation of total proteoglycans. On the contrary, peritubular cell shape has been slightly altered after addition of PMA, IGF-1, vinblastine or colchicine. After these treatments, destruction or rearrangement of cytoskeletal elements was observed; cell-layer proteoglycan synthesis remained either unchanged or increased while total proteoglycans were always undersulfated. IGF-1, PMA and dibutyryl cAMP modified the peritubular cell morphology, cytoskeletal organization and proteoglycan production; the cytoskeleton disrupting drugs such as vinblastine, colchicine and cytochalasin D mimicked some of these effects. These observations suggest that alterations in proteoglycan biosynthesis, after activation of tyrosine kinase, protein kinase C and protein kinase A pathways might be mediated, at least in part, by the disorganization of the cytoskeleton structure.

Actin Cytoskeleton↗
PubMed2003

Nanomolar concentrations of epothilone D inhibit the proliferation of glioma cells and severely affect their tubulin cytoskeleton.

The purpose of this study was to investigate the potential effects of epothilones (EPOs), a new class of microtubule stabilizing cytotoxic drugs, on glioma cells in vitro. The effects of 1, 10 and 100 nM concentrations of EPO D in four malignant human glioma cell lines were measured using a microtiter-tetrazolium assay. Besides the cell lines U87MG, U138MG and LN405, one cell line was used, which had been derived from a recurrent and therapy-resistant glioblastoma in our laboratory. In addition, changes of the cell morphology were followed by light microscopy and changes in the microtubule and actin cytoskeleton were visualized by a confocal laser microscope. In all four human glioma cell lines, 10 and 100 nM concentrations of the drug, applied for 96 h, lead to a highly significant decrease in the viable cell number (p < 0.001). A mean reduction of the viable cell number between 30% and 40% (60% and 90%) was observed for a drug concentration of 10 nM (100 nM). A round cell morphology occured in most EPO treated cells and the organized network of microtubules was shrunk in these round cells. The tubulin immunostaining now appeared amorphous and was restricted to small perinuclear regions. Large actin filaments also disappeared, but actin staining was present in the whole cytosplasm. These results prove that EPOs have antiproliferative effects in glioma cells and affect their tubulin cytoskeleton, as it was previously observed in several types of carcinoma cells.

Actin Cytoskeleton↗
PubMed2002

The volatile fraction of cigarette smoke induces alterations in the human gingival fibroblast cytoskeleton.

Several in vitro investigations have indicated that the particulate phase of cigarette smoke, such as nicotine, affects many cell types, including gingival fibroblasts. However, few studies have been performed on the effects of the volatile fraction on the cellular structures that are involved in cell functions, such as adhesion and proliferation. Since the survival and reproduction of gingival fibroblasts are fundamental in maintaining the integrity of the oral connective tissue, as well as in wound healing, the effects on the cytoskeleton of acrolein and acetaldehyde, which are the volatile fractions of cigarette smoke, were examined in vitro for human gingival fibroblasts (HGFs). HGF strains that were taken from healthy subjects with non-inflamed-gingiva were utilized in this investigation. The cells were incubated in the presence of different concentrations of acrolein and acetaldehyde. Cell adhesion and viability were evaluated after incubation for 3 h and 5 days, respectively. The influence on cytoskeletal structures (tubulin, actin and vimentin intermediate filaments) was investigated with the indirect immunofluorescence technique. The results show that both substances produced similar effects, which resulted in a dose-dependent inhibition of HGF adhesion and viability. Disturbance of the HGF cytoskeleton consisted of disruption of the microtubules, actin filaments and vimentin microfilaments, which was accompanied by alterations to cell shape. Our experimental findings suggest that the volatile fractions of cigarette smoke, such as acrolein and acetaldehyde, have a cytotoxic effect on HGFs, with the result that they lose their capacity for adhesion and proliferation. The consequences of this could be impairment of the maintenance, integrity and remodelling of the oral connective tissue. According to our morphological evidence, these findings show that cigarette smoke can lead to the development and progression of periodontal disease, and indicate the need for appropriate therapy.

Acetaldehyde↗
PubMed2004

Role of the cytoskeleton during leukocyte responses.

The cytoskeleton is a cellular network of structural, adaptor and signalling molecules that regulates most cellular functions that are related to the immune response, including migration, extravasation, antien recognition, activation and phagocytosis by different subsets of leukocytes. Recently, a large number of regulatory elements and structural constituents of the leukocyte cytoskeleton have been identified. In this review, we discuss the composition and regulation of the different cytoskeletal elements and their role in immune responses.

Actin Cytoskeleton↗
PubMed1998

Role of the actin cytoskeleton in regulating the outer phosphatidylethanolamine levels in yeast plasma membrane.

Transbilayer phosphatidylethanolamine (PtdEtn) movements in the plasma membrane of Saccharomyces cerevisiae are regulated by an ATP-dependent, protein-mediated process(es). To examine whether this process is influenced by the actin cytoskeleton, we have studied the PtdEtn translocation in S. cerevisiae cells after treatment with microfilament disrupting and microtubule-disrupting agents. PtdEtn translocation was studied by measuring the external PtdEtn levels, using fluorescamine as the external membrane probe, in the ATP-depleted, ATP-depleted and repleted, and N-ethylmaleimide-treated cells. The microfilaments and microtubules were disrupted by treatment with various cytochalasins and colchicine (or benomyl) respectively PtdEtn translocation became abnormal in the cytochalasin-treated cells but not in cells that were treated with microtubule-disrupting agents, such as colchicine or benomyl. These results have been interpreted to suggest that the actin cytoskeleton is involved in regulating the PtdEtn translocase activity in the yeast cell plasma membrane.

Actin Cytoskeleton↗
PubMed1999

Coronin promotes the rapid assembly and cross-linking of actin filaments and may link the actin and microtubule cytoskeletons in yeast.

Coronin is a highly conserved actin-associated protein that until now has had unknown biochemical activities. Using microtubule affinity chromatography, we coisolated actin and a homologue of coronin, Crn1p, from Saccharomyces cerevisiae cell extracts. Crn1p is an abundant component of the cortical actin cytoskeleton and binds to F-actin with high affinity (Kd 6 x 10(-9) M). Crn1p promotes the rapid barbed-end assembly of actin filaments and cross-links filaments into bundles and more complex networks, but does not stabilize them. Genetic analyses with a crn1Delta deletion mutation also are consistent with Crn1p regulating filament assembly rather than stability. Filament cross-linking depends on the coiled coil domain of Crn1p, suggesting a requirement for Crn1p dimerization. Assembly-promoting activity is independent of cross-linking and could be due to nucleation and/or accelerated polymerization. Crn1p also binds to microtubules in vitro, and microtubule binding is enhanced by the presence of actin filaments. Microtubule binding is mediated by a region of Crn1p that contains sequences (not found in other coronins) homologous to the microtubule binding region of MAP1B. These activities, considered with microtubule defects observed in crn1Delta cells and in cells overexpressing Crn1p, suggest that Crn1p may provide a functional link between the actin and microtubule cytoskeletons in yeast.

Actin Cytoskeleton↗
PubMed1999

Regulated actin cytoskeleton assembly at filopodium tips controls their extension and retraction.

The extension and retraction of filopodia in response to extracellular cues is thought to be an important initial step that determines the direction of growth cone advance. We sought to understand how the dynamic behavior of the actin cytoskeleton is regulated to produce extension or retraction. By observing the movement of fiduciary marks on actin filaments in growth cones of a neuroblastoma cell line, we found that filopodium extension and retraction are governed by a balance between the rate of actin cytoskeleton assembly at the tip and retrograde flow. Both assembly and flow rate can vary with time in a single filopodium and between filopodia in a single growth cone. Regulation of assembly rate is the dominant factor in controlling filopodia behavior in our system.

Actin Cytoskeleton↗
PubMed2006

Cytoskeleton and morphogenesis in brown algae.

BACKGROUND: Morphogenesis on a cellular level includes processes in which cytoskeleton and cell wall expansion are strongly involved. In brown algal zygotes, microtubules (MTs) and actin filaments (AFs) participate in polarity axis fixation, cell division and tip growth. Brown algal vegetative cells lack a cortical MT cytoskeleton, and are characterized by centriole-bearing centrosomes, which function as microtubule organizing centres. SCOPE: Extensive electron microscope and immunofluorescence studies of MT organization in different types of brown algal cells have shown that MTs constitute a major cytoskeletal component, indispensable for cell morphogenesis. Apart from participating in mitosis and cytokinesis, they are also involved in the expression and maintenance of polarity of particular cell types. Disruption of MTs after Nocodazole treatment inhibits cell growth, causing bulging and/or bending of apical cells, thickening of the tip cell wall, and affecting the nuclear positioning. Staining of F-actin using Rhodamine-Phalloidin, revealed a rich network consisting of perinuclear, endoplasmic and cortical AFs. AFs participate in mitosis by the organization of an F-actin spindle and in cytokinesis by an F-actin disc. They are also involved in the maintenance of polarity of apical cells, as well as in lateral branch initiation. The cortical system of AFs was found related to the orientation of cellulose microfibrils (MFs), and therefore to cell wall morphogenesis. This is expressed by the coincidence in the orientation between cortical AFs and the depositing MFs. Treatment with cytochalasin B inhibits mitosis and cytokinesis, as well as tip growth of apical cells, and causes abnormal deposition of MFs. CONCLUSIONS: Both the cytoskeletal elements studied so far, i.e. MTs and AFs are implicated in brown algal cell morphogenesis, expressed in their relationship with cell wall morphogenesis, polarization, spindle organization and cytokinetic mechanism. The novelty is the role of AFs and their possible co-operation with MTs.

Actin Cytoskeleton↗
PubMed2002

Involvement of calcium signaling and the actin cytoskeleton in the membrane block to polyspermy in mouse eggs.

This study examines the effects of actin microfilament-disrupting drugs on events of fertilization, with emphasis on gamete membrane interactions. Mouse eggs, freed of their zonae pellucidae, were treated with drugs that perturb the actin cytoskeleton by different mechanisms (cytochalasin B, cytochalasin D, jasplakinolide, latrunculin B) and then inseminated. Cytochalasin B, jasplakinolide, and latrunculin B treatments resulted in a decrease in the percentage of eggs fertilized and the average number of sperm fused per egg. However, cytochalasin D treatment resulted in an increase in the average number of sperm fused per egg and the percentage of polyspermic eggs. This increase in polyspermy occurred despite the observation that cytochalasin D treatment caused a decrease in sperm-egg binding and did not affect spontaneous acrosome reactions or sperm motility. This suggested that cytochalasin D-treated eggs had an impaired ability to establish a block to polyspermy at the level of the plasma membrane. The effect of cytochalasin D on the block to polyspermy was not due to a general disruption of egg activation because sperm-induced calcium oscillations and cortical granule exocytosis were similar in cytochalasin D-treated and control eggs. However, buffering of intracellular calcium levels with the calcium chelator BAPTA-AM resulted in an increase in polyspermy. Together, these data suggest that a postfertilization decrease in egg membrane receptivity to sperm requires functions of the egg actin cytoskeleton that are disrupted by cytochalasin D. Furthermore, egg activation-associated increased intracellular calcium levels are necessary but not sufficient to affect postfertilization membrane dynamics that contribute to a membrane block to polyspermy.

Acrosome↗
PubMed2003

Paradoxical cytoskeleton and microparticle formation changes in monocytes and polymorphonuclear leukocytes in severe systemic inflammatory response syndrome patients.

BACKGROUND: Circulating monocytes and polymorphonuclear leukocytes (PMNLs) are considered as central regulators controlling systemic inflammatory response after severe insults. Recently, activated monocytes and PMNLs have been reported to produce microparticles (MPs) in vitro. The objective of this study was to evaluate production of MPs and changes of cytoskeleton in monocytes from severe systemic inflammatory response syndrome (SIRS) patients, and to compare them with those in PMNLs. METHODS: Twenty severe SIRS patients (SIRS criteria and serum C-reactive protein > 10 mg/dL) and 15 healthy volunteers were included. MP formation and F-actin content in monocytes and PMNLs were measured by flow cytometry in the presence or absence of lipopolysaccharide or formylmethionyl-leucyl-phenylalanine (FMLP). The membrane expression of human leukocyte antigen-DR and CD64 in monocytes and O2- production in PMNLs were also measured by flow cytometry. RESULTS: In severe SIRS patients, MP formation with and without lipopolysaccharide in monocytes significantly decreased in comparison with those in normal controls (p < 0.05), whereas those with and without FMLP in PMNLs increased (p < 0.05). F-actin content with and without FMLP in monocytes also significantly decreased in patients (p < 0.05), whereas those in PMNLs increased as compared with normal controls (p < 0.05). The expression of human leukocyte antigen-DR in monocytes significantly decreased in patients (p < 0.05), which indicated monocyte modulation. The O2- production in PMNLs increased in patients (p < 0.05), which showed PMNL activation. CONCLUSION: The changes of MP formation and cytoskeleton in circulating monocytes and PMNLs were paradoxically different in severe SIRS patients.

Actin Cytoskeleton↗
PubMed2003

Involvement of the secretory pathway and the cytoskeleton in intracellular targeting and tubule assembly of Grapevine fanleaf virus movement protein in tobacco BY-2 cells.

Grapevine fanleaf virus (GFLV) is one of a large class of plant viruses whose cell-to-cell transport involves the passage of virions through tubules composed of virus-encoded movement protein (MP). The tubules are embedded within modified plasmodesmata, but the mechanism of targeting of MP to these sites is unknown. To study intracellular GFLV MP trafficking, a green fluorescent protein-MP fusion (GFP:MP) was expressed in transgenic tobacco BY-2 suspension cells under the control of an inducible promoter. We show that GFP:MP is targeted preferentially to calreticulin-labeled foci within the youngest cross walls, where it assembles into tubules. During cell division, GFP:MP colocalizes in the cell plate with KNOLLE, a cytokinesis-specific syntaxin, and both proteins are linked physically, as shown by coimmunoprecipitation of the two proteins from the same microsomal fraction. In addition, treatment with various drugs has revealed that a functional secretory pathway, but not the cytoskeleton, is required for tubule formation. However, correct GFP:MP targeting to calreticulin-labeled foci seems to be cytoskeleton dependent. Finally, biochemical analyses have revealed that at least a fraction of the MP behaves as an intrinsic membrane protein. These findings support a model in which GFP:MP would be transported to specific sites via Golgi-derived vesicles along two different pathways: a microtubule-dependent pathway in normal cells and a microfilament-dependent default pathway when microtubules are depolymerized.

Actin Cytoskeleton↗
PubMed1998

Nuclear localization signal receptor importin alpha associates with the cytoskeleton.

Importin alpha is the nuclear localization signal (NLS) receptor that is involved in the nuclear import of proteins containing basic NLSs. Using importin alpha as a tool, we were interested in determining whether the cytoskeleton could function in the transport of NLS-containing proteins from the cytoplasm to the nucleus. Double-labeling immunofluorescence studies showed that most of the cytoplasmic importin alpha coaligned with microtubules and microfilaments in tobacco protoplasts. Treatment of tobacco protoplasts with microtubule- or microfilament-depolymerizing agents disrupted the strands of importin alpha in the cytoplasm, whereas a microtubule-stabilizing agent had no effect. Biochemical analysis showed that importin alpha associated with microtubules and microfilaments in vitro in an NLS-dependent manner. The interaction of importin alpha with the cytoskeleton could be an essential element of protein transport from the cytoplasm to the nucleus in vivo.

Actin Cytoskeleton↗
PubMed1990

High resolution scanning electron microscopy of the subplasmalemmal cytoskeleton in human odontoblasts.

The subplasmalemmal cytoskeleton in human odontoblasts was studied by high resolution scanning electron microscopy. The odontoblast layer was isolated and exposed to formaldehyde, glutaraldehyde, and OsO4 for some specimens, while the membraneous structures and soluble proteins in the dental tissue were removed by Zenker's solution and 1% OsO4 for other specimens, without further fixation of the remaining components. The cytoskeletal elements comprised a dense network of interlacing filaments of different diameters in the cell body. Most cytoskeletal elements were parallel to the axis of the cell processes situated inside the dentinal tubules. The appearance and orientation of the investigated subplasmalemmal cytoskeletal elements was unaffected by the choice of method. Both methods confirm the presence of a subplasmalemmal cytoskeleton in human odontoblasts.

Actin Cytoskeleton↗