Seven fibrovascular diabetic preretinal membranes were examined with lightmicroscophic immunohistochemical stain and electron-microscopy to evaluate the possibility of pericytes to be involved in membrane contraction. Pericytes were positively stained with anti-actin antibody together with some stromal cells thought to be myofibroblasts presumedly. On transmission electron microscopic study, pericytes were highly active with numerous cytoplasmic processes and contained abundant microfilaments considered as actin in their cytoplasm. Pericyte/endothelial cell ratio of vascular channels were increased in some actively proliferative portion of the membranes. Myofibroblasts that contain abundant cytoplasmic microfilaments were also demonstrated in the extravascular stroma of the membranes and were very similar to the pericytes morphologically. Although the evidence that the pericytes are related to the origin of the myofibroblasts could not be demonstrated, this study suggested that the pericytes may play important roles in development and contraction mechanism of fibrovascular diabetic preretinal membranes.
Actin Cytoskeleton ; Actins ; Cytoplasm ; Membranes* ; Myofibroblasts ; Pericytes* ; Stromal Cells

Mice ; Animals ; Actins/metabolism* ; Actin Depolymerizing Factors/metabolism* ; TRPM Cation Channels ; Actin Cytoskeleton/metabolism* ; Nervous System/metabolism*

UNLABELLEDOBJECTIVE To evaluate the effects of grooved, alkali- and heat-treated, acid-etched and TiO2 blasted surfaces of titanium substrates on F-actin cytoskeleton of osteoblasts in vitro.

METHODSOsteoblasts derived from fetal rat calvarial were cultured on 6 different commercially pure titanium discs-grooved(G), sandblasted (SB), sand-blasted and acid-etching (SLA) surfaces and alkali- and heat-treated (AH1, AH2, AH3) surfaces. For F-actin cytoskeleton measurement, osteoblasts whose filamentous actin was stained with phalloidin-TRITC were cultured for 1, 2, 4, 12 h, evaluated by CLSM observation.

RESULTSOsteoblasts attached to the different types of surfaces after 1 hour culture were similar. The actin cytoskeleton formed a ring of cortical filaments around the nucleus after 1 hour on SB, AH2, AH3, SLA surfaces. Actin filaments condensed along edges of pits. The actin filaments of seeded cells were spread after 2 h. The actin filaments on G formed bundles around the nucleus. The filaments began to parallel to the grooves. On AH1, the fibres formed a ring of cortical filaments around the nucleus with some cytoplasmic fibres radially oriented. On AH2, AH3, SB, the fibres orignised in a cytoplasmic meshwork with fibres which terminate at the ridge of depressions. The cell were suspending itself over the depressed areas. Actin filaments on SB were distinct and well formed that were oriented paralled to one another and the long axis of cells. After 4 h, actin filaments appeared organised in a parallel to one another and the long axis of cells. After 12 h, the actin filaments on all surfaces were well spread and were oriented paralled to another and to the long axis of the cell. The filaments formed bundles which reached to holes or adhered to the ridge of raised points, suspending cells over depressed areas.

CONCLUSIONAfter 12 h, the actin filaments on all surfaces were well spread and were oriented parallel to another and to the long axis of the cell. It was concluded that F-actin cytoskeleton of osteoblasts were spread best on SB surfaces among all surfaces.

Actin Cytoskeleton ; Actins ; Animals ; Cytoskeleton ; Microtubules ; Osteoblasts ; Prostheses and Implants ; Rats ; Surface Properties ; Titanium

It is believed that there exists some relationship between the distribution and morphology of intracellular actin and cell adherence. Cells are likely to be deteched when the quantity of actin filament decreases. Actin filaments locate in the fringe of cancer cells and cells cultured in static state, so that these filaments can stretch out and form pseudopodia to adhere to the matrix. When these cells are stimulated their pseudopodia retract so that they can easily be detached from the matrix. When external forces are exerted on cells to adhere and deadhere from the matrix, the morphology and distribution of skeleton actin will change, so as the cells' morphology. The skeleton actins in cells are changed differently to adapt to different external forces which are imposed on the cells. It is obvious that the relationship between the mechanism of cell adhering to the matrix and the morphology & distribution of actins needs more attention.
Actin Cytoskeleton ; metabolism ; Actins ; metabolism ; Cell Adhesion ; Humans ; Neoplasms ; metabolism ; pathology ; Pseudopodia ; metabolism ; Shear Strength

OBJECTIVETo investigate the relationship between c-fos gene and filamentous actin (F-actin) in MG-63 osteoblasts under cyclic tensile stress.

METHODSMG-63 osteoblasts were subjected to cyclic tensile stress (0.5 Hz, 2 000 microstrain) for 3, 6, and 12 h. The changes of c-fos gene were investigated by fluorescent quantitation polymerase chain reaction. Then the best loading time group was screened as the experimental group compared with 0 h group. The changes of F-actin and c-fos were investigated with or without cytochalasin D treatment.

RESULTSCyclic tensile stress induced high expression of c-fos mRNA, and peaked at 3 h. After loading, F-actin had a structure reorganization, but had no change in expression. After cytochalasin D treatment, the formation of stress fibers and the fluorescence intensity of F-actin cytoskeleton significantly reduced, meanwhile the c-fos mRNA expression was inhibited.

CONCLUSIONAfter loading, there is only structure reorganization for F-actin, and the expression has not any change. That means the remodeling F-actin is the existing one. F-actin reorganization is an important part in c-fos gene expression induced by stress.

Actin Cytoskeleton ; Actins ; Cytochalasin D ; Cytoskeleton ; Genes, fos ; Humans ; Microtubules ; Osteoblasts ; RNA, Messenger ; Stress, Mechanical

PURPOSE: In osteoclast differentiation, actin-rich membrane protrusions play a crucial role in cell adhesion. Odontogenic ameloblast-associated protein (Odam) contributes to cell adhesion by inducing actin rearrangement. Odam-mediated RhoA activity may play a significant role in multinucleation of osteoclasts. However, the precise function of Odam in osteoclast cell adhesion and differentiation remains largely unknown. Here, we identify a critical role for Odam in inducing osteoclast adhesion and differentiation. MATERIALS AND METHODS: The expression of Odam in osteoclasts was evaluated by immunohistochemistry. Primary mouse bone marrow and RAW264.7 cells were used to test the cell adhesion and actin ring formation induced by Odam. RESULT: Odam was expressed in osteoclasts around alveolar bone. Odam transfection induced actin filament rearrangement and cell adhesion compared with the control or collagen groups. Overexpression of Odam promoted actin stress fiber remodeling and cell adhesion, resulting in increased osteoclast fusion. CONCLUSION: These results suggest that Odam expression in primary mouse osteoclasts and RAW264.7 cells promotes their adhesion, resulting in the induction of osteoclast differentiation.
Actin Cytoskeleton ; Actins* ; Animals ; Bone Marrow ; Cell Adhesion ; Collagen ; Immunohistochemistry ; Membranes ; Mice ; Osteoclasts* ; Stress Fibers ; Transfection

T cell activation and function require physical contact with antigen presenting cells at a specialized junctional structure known as the immunological synapse. Once formed, the immunological synapse leads to sustained T cell receptor-mediated signalling and stabilized adhesion. High resolution microscopy indeed had a great impact in understanding the function and dynamic structure of immunological synapse. Trends of recent research are now moving towards understanding the mechanical part of immune system, expanding our knowledge in mechanosensitivity, force generation, and biophysics of cell-cell interaction. Actin cytoskeleton plays inevitable role in adaptive immune system, allowing it to bear dynamic and precise characteristics at the same time. The regulation of mechanical engine seems very complicated and overlapping, but it enables cells to be very sensitive to external signals such as surface rigidity. In this review, we focus on actin regulators and how immune cells regulate dynamic actin rearrangement process to drive the formation of immunological synapse.
Actin Cytoskeleton ; Actins ; Antigen-Presenting Cells ; Biophysics ; Immune System ; Immunological Synapses ; Microscopy ; T-Lymphocytes ; Ursidae

BACKGROUND: Regulation of cell volume is one of the major physiologic functions in living cells. Intracellular calcium ion and cytoskeletal component of cell membrane play pivotal roles in cell volume regulation. The significance of astroglial cell swelling is in its relation to delayed and permanent neuronal death. The aim of the current study is therefore to elucidate the effect of propofol on the cell volume, [Ca2+]i and actin filaments in astroglial cells. METHODS: Astroglial cell line U1242MG astrocytoma cells were used in vitro. To investigate the alterations of cell volume and [Ca2+]i by propofol, flow cytometry system was used. Actin filaments were determined by tetramethylrhodamine isothiocyanate (TRITC)-phalloidin staining. RESULTS: Treatment with propofol 10 microgram/ml for 15 min or 2 h perfusion reduced significantly both cell volume and [Ca2+]i. Cell volume was reduced 3-4% in either duration of perfusion with propofol. Decreases in [Ca2+]i level in the presence of propofol was duration-dependent. Fifteen min perfusion with propofol caused 5% decrease in [Ca2+]i, but 2 h perfusion decreased [Ca2+]i further to 10%. The changes of actin filaments after propofol treatment for 30 min were apparently observed under fluorescent microscope. CONCLUSIONS: Changes of cell volume, [Ca2+]i and actin filaments are occurred by propofol in astroglial cells. Rearrangement of actin filaments may be associated with volume changes by propofol in astroglial cells.
Actin Cytoskeleton* ; Actins* ; Astrocytoma ; Calcium* ; Cell Line ; Cell Membrane ; Cell Size* ; Flow Cytometry ; Neurons ; Perfusion ; Propofol*

OBJECTIVETo explore the response of rat bone marrow mesenchymal stem cells (MSCs and calvarial osteoblasts to mechanical strain and the consequent changes of cytoskeleton F-actin.

METHODSBone marrow MSCs and calvarial osteoblasts were isolated from SD rats and cultured in vitro. Mechanical stretch was performed on passage 3 cells at 2 000 microepsilon for 0, 2, 6 and 12 hours using four-point bending system. The response of cells and the distribution of F-actin were observed using fluorescent staining under laser scanning confocal microscope and the morphological parameters were quantified using image analysis software Laserpix.

RESULTSUnder mechanical stretch, the fluorescent staining decreased obviously at both MSCs and osteoblasts, and F-actin filaments were rearranged and became tenuous, thinner, and abnormally distributed. The outline of nucleus became unclear and apoptotic changes were observed at some of both cells. Cellular size decreased more significantly in MSCs than in osteoblasts. Quantity analysis showed that total area of cells, total fluorescent density and green fluorescent density (F-actin) were all significantly decreased in MSCs (P < 0.05 or P < 0.01), and total fluorescent density, green fluorescent density and red fluorescent density (nuclei) did also in osteoblasts (P < 0.05 or P < 0.01).

CONCLUSIONMechanical stretch caused extensive response on both MSCs and osteoblasts which led to the rearrangement of F-actin filament and apoptosis in some of these cells. MSCs were more sensitive to mechanical strain than osteoblasts.

Actin Cytoskeleton ; metabolism ; Actins ; metabolism ; Animals ; Bone Marrow Cells ; Cells, Cultured ; Cytoskeleton ; Mesenchymal Stromal Cells ; Microtubules ; Osteoblasts ; Rats ; Stress, Mechanical

To investigate the changing patterns of microfilament and microtubule arrangement and influence of myocardial cells and colchicines to microfilament and microtubule formation in cardiac endothelial cells the authors carried out indirect immunofluorescence stain for actin and tubulin with supernatant monoclonal antibodies. Secondary antibodies were IgG FITC conjugate. The results were summarized as follows. Fiberform reactions were stronger in the cells with many processes and spread cytoplasm and they became weaker after the endothelial cells formed monolayer. In the endothelial cells cocultured with myocardial cells the fiberform of the microtubule became less visible compared to control group but fiberform of the microtubule maintained strong intensity as endothelial cells formed monolayer. In the group treated with colchicines, there were no visible differences in microfilaments compared to control group but fiberform of microtubule revealed weaker intensity after colchicines treatment. The intensity of microtubule fiberform returned to control level after 2 days.
Actin Cytoskeleton ; Actins ; Antibodies ; Antibodies, Monoclonal* ; Cytoplasm ; Endothelial Cells* ; Fluorescein-5-isothiocyanate ; Fluorescent Antibody Technique, Indirect ; Immunoglobulin G ; Microtubules ; Tubulin