Number Of Pseudopodia Research Articles

Regulation of Tendon Stem Cell Behavior by Designed Nanoporous Topography of Microfibers.

Nano scale topography scaffold is more bioactive and biomimetic than smooth fiber topographies. Tendon stem cells (TSCs) play important roles in the tendinogenesis of tendon tissue engineering, but the effects and mechanisms of nano topography on TSC behavior are still unclear. This study determined whether the morphology, proliferation, cytoskeleton, and differentiation of TSCs are affected by topography of scaffold in vitro. The porous PA56 scaffolds were prepared with different concentration ratios of glycerol as the molecular template by electrospinning. Its topological characteristics, hydrophilicity, and degradation properties varied with glycerol proportion and movement rate of the receiving plate. Porous fibers promoted the proliferation of TSCs and the number of TSCs varied with topography. Although there was no significant difference due to the small sample size, the number of pseudopodia and cell polarizability still showed differences among different topographies. The morphology of actin cytoskeleton of TSCs showed difference among cultured on porous fibers, smooth fibers, and in culture media with no fiber, suggesting the orientation growth of cells on porous fiber. Moreover, porous fibers promoted teno-lineage differentiation of TSCs by upregulating tendon-specific gene expression. These findings provide evidence that nano porous topography scaffold promotes TSC proliferation, cytoskeleton orientation, and tenogenic differentiation.

Dextran‑coated superparamagnetic iron oxide nanoparticles activate the MAPK pathway in human primary monocyte cells.

With the increase in applications of superparamagnetic iron oxide nanoparticles (SPIONs) in biomedicine, it is essential to investigate the bio‑security of these nanoparticles, especially with respect to the human immune system. In the present study, the biological effects of dextran‑coated superparamagnetic iron oxide nanoparticles (Dex‑SPIONs) on human primary monocyte cells were evaluated. The results of the present study demonstrated that Dex‑SPIONs can be identified in phagosomes or freed in the cytoplasm and did not affect cell viability or induce apoptosis. Notably, there were certain bulky vacuoles and a number of pseudopodia from the cell membrane, suggesting potential activation of human monocyte cells. In addition, the expression levels of pro‑inflammatory cytokines interleukin (IL)‑1β and tumor necrosis factor (TNF)‑α were also increased following treatment with Dex‑SPIONs. Simultaneously, the phosphorylation levels of mitogen‑activated protein kinase (MAPK) p38, c‑Jun N‑terminal kinase 1 and extracellular signal regulated kinase were markedly enhanced following nanoparticle exposure and MAPK inhibitors could abate the production of IL‑1β and TNF‑α. The results of the present study demonstrated that Dex‑SPIONs could activate human monocyte cells and that activation of MAPK pathway may be involved in these effects.

Relationship between TLR4/NF-κB signaling pathway and propofol-induced inhibition of endotoxin-induced release of TNF-α from alveolar macrophages of rats

Objective To evaluate the relationship between Toll-like receptor 4(TLR4)/nuclear factor kappa B(NF-κB)signaling pathway and propofol-induced inhibition of endotoxin-induced release of tumor necrosis factor-alpha(TNF-α)from alveolar macrophages(AMs)of rats. Methods AMs extracted from adult male Sprague-Dawley rats were cultured and inoculated in 6-well plates(1×106 cells/well)and in 96-well plates(1×104 cells/well). The cells were divided into 5 groups(n=18 each)using a random number table: control group(group C), dimethyl sulfoxide group(group D), lipopolysaccharide(LPS)group(group L), propofol group(group P)and LPS plus propofol group(group L+ P). The cells were continuously cultured with phosphate buffer solution in group C. Dimethyl sulfoxide was added at the final concentration of 5 mg/ml in group D. LPS was added at the final concentration of 1 μg/ml in group L. Propofol was added at the final concentration of 25 μmol/L(4.46 μg/ml)in group P. LPS and propofol were added at the final concentration of 1 μg/ml and 25 μmol/L(4.46 μg/ml), respectively, in group L+ P.At 24 h of culture or incubation, the cell viability was detected by CCK-8 assay, the morphological changes of cells were observed using Wright′s staining, the concentration of TNF-α in the supernatant was determined by enzyme-linked immunosorbent assay, and TLR4 expression and NF-κB activities were measured by Western blot. Results Compared with group C, the cell viability and concentration of TNF-α in the supernatant were significantly increased, the expression of TLR4 was up-regulated, and the activity of NF-κB was enhanced in L and L+ P groups(P 0.05). Compared with group L, the cell viability and concentration of TNF-α in the supernatant were significantly decreased, the expression of TLR4 was down-regulated, and the activity of NF-κB was weakened(P<0.05), the morphological changes of cells were significantly attenuated, and the number of pseudopodia was reduced in group L+ P. Conclusion The mechanism by which propofol inhibits endotoxin-induced release of TNF-α from AMs is related to inhibited activation of TLR4/NF-κB signaling pathway in rats. Key words: Toll-like receptor 4; NF-kappa B; Propofol; Endotoxins; Macrophages, alveolar; Tumor necrosis factor-alpha

Study on inflammatory effect of toxic raphides from Pinellia ternate and its correlation with macrophages

To study the toxic mechanism of toxic raphides from Pinellia ternata. Mouse peritoneal macrophage in vitro culture model was adopted to study dose-dependent and time-dependent curves of toxic raphides, with TNF-alpha, IL-1beta and IL-6 in supernatant as indexes. Scanning electron microscopy was used to observe the changes in surface morphology of raphides-treated macrophages. Macrophages-neutrophils co-cultured the transport model to study the effect of toxic raphides' stimulation of macrophages on neutrophils migration. Toxic raphides' stimulation of macrophages could cause the increase in the levels of TNF-alpha, IL-1beta and IL-6 released, and showed dose dependence and time dependence. Scanning electron microscopy showed that toxic raphides were swallowed by macrophages, with notable cell membrane creases, increase in the number of pseudopods and decrease in integrity of cell membranes, and could significantly induce migration of neutrophils. The inflammatory process induced by toxic raphides is mainly mediated by macrophages. The toxic mechanism of toxic raphides from P. ternata is that toxic raphides penetrate into tissues to activate resident macrophages, release phagocytic and inflammatory cytokines, and cause migration of neutrophils, which finally results in acute inflammatory response.

First Record of Hemocytes and Oenocytes in Freshwater Ostracodes

This study provides morphological descriptions of three distinct types of hemocytes and one type of oenocyte newly observed in the body cavity of several freshwater ostracode species (Candona candida, C. neglecta, Herpetocypris reptans, H. incongruens, Eucypris virens, Psychrodromus olivaceus, Notodromas monacha, N. persica and Scottia birigida, all Podocopida, Cypridoidea). These cells were found in both males and females and in both juveniles and adults predominantly in inter-lamellar space, along the dorsal margin of the body cavity and in the anteroventral part of the body cavity. In TEM, large granulocytes with a number of pseudopodia and remarkable rounded granula were observed. Their affinity to muscle syncytia and a role in phagocytosis of exogenous particles is distinct. Oenocytoid-like cells with homogenous cytoplasm, rich in polyribosomes and with few membrane-delimited vesicles, were caught in the peripheral parts of body cavity accumulating in special multicellular formations. The oenocytoid-like cells were observed in two cases also by light microscopy. Two types of plasmatocytes (agranular and granular) with numerous pseudopodia were observed in TEM to be involved in phagocytosis of cell debris. Moreover, the plasmatocytes are located in close vicinity with the multicellular formations of oenocytoid-like cells and envelope them or form a sheath. Oenocytes were shown by light microscopy using histological staining to occur in huge amounts in juveniles. In adults their frequency becomes considerably lower. Possible functions of these cells are discussed in the context of invertebrate haematology, physiology and immunology.

In vitro biocompatibility and degradation of terpolyester 3HB-co-4HB-co-3HHx, consisting of 3-hydroxybutyrate, 4-hydroxybutyrate and 3-hydroxyhexanoate

A terpolyester consisting of 3-hydroxybutyrate (3HB), 4-hydroxybutyrate (4HB) and 3-hydroxyhexanoate (3HHx), abbreviated as P(3HB-co-4HB-co-3HHx), was studied for possible application as an implant biomaterial. L929 mouse fibroblasts, MC3T3-E1 murine osteoblasts and a human cell line of immortalized human keratinocyte (HaCat cells) were used to study the biocompatibility of P(3HB-co-4HB-co-3HHx). Cell morphology and cell activity were studied using scanning electron microscopy (SEM) and the MTT assay, respectively. All three cell types showed higher activities when grown on films of P(3HB-co-4HB-co-3HHx) compared with their growth on poly(lactic acid) (PLA), co-polyester PHBHHx films and on polylysine-coated plates (blank), respectively. The three cell types grown on the terpolyester also demonstrated a well-spread cell shape and large number of pseudopods due to strong cell–cell and cell–material interactions. It was clearly observed that P(3HB-co-4HB-co-3HHx) had a much faster degradation rate than PHBHHx after 15 weeks of incubation in phosphate-buffered saline under dynamic conditions. The results proved that the terpolyester had favorable biocompatibility and biodegradability compared with the well-studied polyesters PLA and PHBHHx.

Cellular Response to Gelatin- and Fibronectin-Coated Multilayer Polyelectrolyte Nanofilms

Surface engineering is a critical effort in defining substrates for cell culture and tissue engineering. In this context, multilayer self-assembly is an attractive method for creating novel composites with specialized chemical and physical properties that is currently drawing attention for potential application in this area. In this work, effects of thickness, surface roughness, and surface material of multilayer polymer nanofilms on the growth of rat aortic smooth muscle cells were studied. Polyelectrolyte multilayers (PEMs) electrostatically constructed from poly(allylamine hydrochloride) and poly(sodium 4-styrenesulfonate) (PSS) with gelatin, fibronectin, and PSS surface coatings were evaluated for interactions with smooth muscle cells (SMCs) in an in vitro environment. The results prove that PEMs terminated with cell-adhesive proteins promote the attachment and further growth of SMCs, and that this property is dependent upon the number of layers in the underlying multilayer film architecture. Cell roundness and number of pseudopodia were also influenced by the number of layers in the nanofilms. These findings are significant in that they demonstrate that both surface coatings and underlying architecture of nanofilms affect the morphology and growth of SMCs, which means additional degrees of freedom are available for design of biomaterials. This work supports the excellent potential of nanoassembled ultrathin films for biosurface engineering, and points to a novel perspective for controlling cell-material interaction that can lead to an elegant system for defining the extracellular in vitro environment.

Disaggregation of aggregated platelets by savignygrin, a alphaIIbeta3 antagonist from Ornithodoros savignyi.

Ticks control their host's hemostatic system by secretion of bioactive components during feeding that inhibit blood coagulation and platelet aggregation. Dissolution of platelets that have already aggregated can enhance control over the hemostatic system. It has been shown that disaggregation of aggregated platelets by the enzyme apyrase was accompanied by a shape change from the aggregated spherical form back to the discoid form associated with un-activated platelets. The present study concerns the disaggregation effect of the alpha IIb/beta3 antagonist, savignygrin. Aggregated platelets that were disaggregated by savignygrin and platelets pre-incubated with savignygrin before activation with ADP, retained a spherical form similar to platelets disaggregated by the fibrinogenolytic enzyme plasmin. The number of pseudopods were however, markedly reduced suggesting a disruption of the focal adhesion points that act as a localization point of alpha IIb/beta3. These results are concurrent with targeting of alpha IIb/beta3 and dissociation of fibrinogen from its receptor, once aggregation has taken place. This is the second mediator of platelet disaggregation found in soft ticks and suggests that disaggregation of aggregated platelets might play an important part in the anti-hemostatic strategy of ticks.

Spreading and motility of human glioblastoma cells on sheets of silicone rubber depend on substratum compliance

Although there is a substantial quantity of experimental data examining the effects of adhesion on the morphology and migration of tissue cells, little attention has been focused on how changes in substratum mechanical properties affect these cellular behaviours. To determine whether the ability of a substratum mechanically to support traction influences cell morphology and motility, measurements are taken of the spreading, the fraction of a population with pseudopodia, the number of pseudopodia and the translocation of human SNB-19 glioblastoma cells cultured on films of poly(methylphenyl)siloxane possessing a range of mechanical compliances. Cells cultured on these films generate deformations (i.e. 'wrinkles') that are used as a basis to estimate effective substratum compliances. The average projected cell area decreases by over 60%, with a two-orders-of-magnitude increase in compliance. Time-lapse videomicroscopy reveals that cell migration also decreases with increasing compliance: the average cell speed decreases from approximately 8 microns h-1 on the most rigid substrata to 1.2 microns h-1 on the most compliant substrata examined. Changes in compliance do not alter mean directional persistence time. These results are interpreted in terms of the predictions of mathematical models for the effects of substratum compliance on motility.

Benzalkonium chloride interferes with energy production, secretion and morphology in human blood platelets

Benzalkonium chloride (BC) is a bactericidal compound used as a topical antiseptic and as a preservative in various products for local treatment, e.g., eye and nose drops. BC is toxic to human cells, including those of the respiratory mucosa. Few studies have, however, focused on what cellular functions BC interferes with. The effects of BC were studied on washed human blood platelets in vitro . Cellular energy production as well as secretion were studied. Incubation of platelets with BC resulted in rapid swelling and toxic morphological changes. After incubation with BC oxidation of [1-14C] palmitate was inhibited, and both lactate dehydrogenase and endogenous serotonin were spontaneously released. Thrombin-induced secretion of serotonin was strongly reduced after BC exposure. Histological changes with increased size, spherical form, decreased numbers of pseudopodia, loss of an intact continuous tubulus system and reduced number of granules were found by scanning and transmission electron microscopy. It is concluded that the toxic effects of BC are because of interference with membrane function and energy production.

Benzalkonium chloride interferes with energy production, secretion and morphology in human blood platelets

Benzalkonium chloride (BC) is a bactericidal compound used as a topical antiseptic and as a preservative in various products for local treatment, e.g., eye and nose drops. BC is toxic to human cells, including those of the respiratory mucosa. Few studies have, however, focused on what cellular functions BC interferes with. The effects of BC were studied on washed human blood platelets in vitro. Cellular energy production as well as secretion were studied. Incubation of platelets with BC resulted in rapid swelling and toxic morphological changes. After incubation with BC oxidation of [1-14C] palmitate was inhibited, and both lactate dehydrogenase and endogenous serotonin were spontaneously released. Thrombin-induced secretion of serotonin was strongly reduced after BC exposure. Histological changes with increased size, spherical form, decreased numbers of pseudopodia, loss of an intact continuous tubulus system and reduced number of granules were found by scanning and transmission electron microscopy. It is concluded that the toxic effects of BC are because of interference with membrane function and energy production.

Calculation of pulsatility index of flow volume independent of vessel diameter and flow profile

An application of a morphometric model is used to study the stroma epithelial interface in human cervical epithelium. This model derived by Okagaki and associates estimates from two dimensional transmission electron photomicrographs the number of basal cell pseudopodia per basal cell. The investigation of the full spectrum of cervical intraepithelial neoplasia (CIN) focusing on the basal cell-stroma interface with the “multiple cell-single section” model is discussed. Significant decreases in histopathologic grades of CIN are noted in the number of pseudopodia per basal cell (N¯) and the ratio (F¯) of the area of the stems of pseudopodia to the area of the base of the basal cell.

Electron microscopic analysis of the normal and the activated pleural macrophage.

Despite an apparent role in pleural pathophysiology, little information is known about pleural macrophage morphology. Intrapleural tetracycline (TCN) results in pleural macrophage influx and pleural fibrosis; intrapleural carrageenan (CAR) induces macrophage influx without ensuing fibrosis. Pleural macrophages collected from normal (NL) and TCN- or CAR-exposed rabbit pleural spaces were examined with electron microscopy. Cellular size; number of microvilli; pseudopods; coated pits (CP) and coated vesicles (CV); and prevalence of golgi, rough endoplasmic reticulum (RER), and intermediate filaments (IF) were determined. The means of each variable in each group were assessed by one-way analysis of variance, with post hoc testing performed by Scheffe F test; p < or = .05 was considered significant. TCN-stimulated pleural macrophages were characterized by their small perimeters. CAR-induced pleural macrophages were marked by their large size and abundant intracellular amorphous material. They had larger perimeters, areas, and diameters than the TCN-induced or normal macrophages and thus smaller numbers of CV + CP per area. The normal pleural macrophages were characterized by more IF, microvilli, and microvilli per perimeter than either the CAR- or TCN-induced pleural macrophages. No differences between groups were found in nuclear cytoplasmic ratios, number of pseudopods, and content of golgi or of RER. The results suggest that normal pleural macrophages and TCN- and CAR-induced pleural macrophages differ morphologically and that these morphologic differences reflect functional differences.

Amphotericin B‐induced injury in stored human platelets

Administration of intravenous amphotericin B is a major factor in the reduction of the recovery of transfused platelet concentrates (PCs). For the study of possible drug-induced platelet lesions, fresh platelets and stored PCs were incubated with therapeutic concentrations of amphotericin B (10-50 micrograms/mL) and examined for membrane changes by scanning electron microscopy. Stored platelets demonstrated the formation of "pits" on the surface membrane, which were maximal immediately after preparation and gradually decreased with storage. The number of pits was significantly higher after exposure to amphotericin B. No increase was detected in fresh platelets following exposure to amphotericin B. The maximal effect of the drug was seen after 5 days of storage as PCs. Amphotericin B did not affect platelet shape or the number of pseudopodia. There was no correlation between pit formation and the pH, pO2, or pCO2 of the concentrates. Amphotericin B did not release 51Cr from prelabeled stored platelets after 2 hours' incubation at 37 degrees C. Thus, amphotericin B appears to exacerbate a membrane lesion induced by the preparation and storage of PCs.

Inhibition by lysosomotropic amines of dog thyroid secretion in vitro.

The lysosomotropic amines are well-known inhibitors of lysosomal protein degradation. These drugs were used in dog thyroid slices to ascertain the role of the lysosome in thyroglobulin hydrolysis and in hormone secretion. NH4Cl (1-20 mM) and chloroquine (5-500 microM) inhibited, in a concentration-dependent manner, the TSH-stimulated secretion. The high concentrations of these compounds also inhibited the basal secretion. The inhibition was not toxic since 1) the nonbutanol extractable iodine, an index of follicle disruption, was not increased, except slightly by 20 mM NH4Cl; 2) the compounds did not inhibit the TSH-induced stimulation of protein iodination; 3) the inhibition of secretion caused by these compounds was reversible; 4) the ultrastructural changes induced by NH4Cl were reversed after its withdrawal. The inhibition of secretion was presumably related to the lysosomal trapping of the drugs because: 1) the time lag for the fall in secretion rate after drug addition was shorter than for the inhibition of secretion at the level of pseudopod formation by carbamylcholine and cytochalasin B; 2) for NH4Cl this delay and the degree of inhibition were modulated by the [H+] gradient between the medium and the lysosome; 3) 20 min after NH4Cl addition, 92% of the lysosomes were vacuolated and swollen (median section area, 126 mu2 vs. 50 mu2 for the controls) and 8% of the lysosomes were swollen and still dense (median, 206 mu2); 20 min after chloroquine addition, 90% of the lysosomes remained dense and had a significantly higher section area (median, 79 mu2) than the controls (P less than 0.001), whereas 10% of the lysosomes were vacuolated and large (median area, 438 mu2). The number of pseudopods measured by scanning electron microscopy significantly decreased only after 1 h (P less than 0.001). This late decrease could not account for the early block of secretion and suggested a lack of membrane recycling. In conclusion, lysosomotropic amines interfere with a post phagocytotic, presumably lysosomal, step in secretion by dog thyroid. These data constitute the first biochemical evidence in the intact cell of the role of lysosomes in TSH-induced thyroglobulin hydrolysis and thyroid hormone secretion.

Enumeration of microprocesses of T-lymphoma cells by two morphometric methods using SEM and TEM

We have demonstrated that the number of pseudopodia is directly correlated with cell differentiation and carcinogenesis in the squamous epithelium. Microprocesses over T-lymphocytes may also reflect cell differentiation or a functional state of the cells as seen in the epithelial cell model, although their functions are not well understood. Visual quantification of spatially distributed bodies such as microprocesses observed with a scanning electron microscope (SEM) is inherently poor. That is, exact quantitation of the number of microprocesses per cell may be necessary to demonstrate a smaller difference in frequency of such structure. One drawback of previous attempts to use such a method was that the quantitation of a structure over a surface which was at a steep angle to the film plane could not be accurate. We developed a modified method of enumeration with SEM and another method with a transmission electron microscope (TEM), using cultured murine T-lymphoma cells (EL-4) as a model, and the results were compared.

Progressive Reduction of Pseudopodia in Advancing Grades of Uterine Cervical Intraepithelial Neoplasia (CIN) - An Application of a New Morphometric Method

It has been suggested that stroma epithelial interaction may play a significant role in epithelial differentiation and carcinogenesis. Squamous epithelium normally possesses pseudopodia (foot processes) which protrude in the underlying stromal tissue. Whether the pseudopodia facilitate the communication between the epithelium and stroma is an open question. Nevertheless, it has been demonstrated by a semi-quantitative observation that the number of pseudopodia on the basal cells of the advanced premalignant condition (carcinoma in situ or grade 4 cervical intraepithelial neoplasia (CIN)) of the uterine cervix was reduced.We have developed a method to measure the number of pseudopodia on the basal cells of the epithelium, their cross sectional widths, and the cross section widths of basal cells appearing in transmission electron micrographs. First on each observed basal cell, the number of pseudopodia is calculated from formula derived by probabilistic (morphometric) calculation:

A preliminary report on a technique to analyze the activity of Entamoeba histolytica trophozoites in vitro

A quantitative method for measuring the activity of individual amoebae in vitro is described. Trophozoites of three strains of Entamoeba histolytica, IP-106, NIH:200, and DKB, were compared and measurable differences in behaviour and activity between the three strains were observed. Although the total number of pseudopodia produced by amoebae was the same for the three strains tested, amoebae of strains IP-106 and NIH:200 moved farther during a 5-min period than did those of strain DKB. This correlates with a more tortuous track of the latter strain.

Basal cell pseudopodia in cervical intraepithelial neoplasia; progressive reduction of number with severity: A morphometric quantification

An application of a morphometric model is used to study the stroma epithelial interface in human cervical epithelium. This model derived by Okagaki and associates estimates from two dimensional transmission electron photomicrographs the number of basal cell pseudopodia per basal cell. The investigation of the full spectrum of cervical intraepithelial neoplasia (CIN) focusing on the basal cell-stroma interface with the “multiple cell-single section” model is discussed. Significant decreases in histopathologic grades of CIN are noted in the number of pseudopodia per basal cell (N¯) and the ratio (F¯) of the area of the stems of pseudopodia to the area of the base of the basal cell.

Measurement of number and cross-sectional area of basal cell pseudopodia: a new morphometric method.

A method of morphometric quantitative of the number of pseudopodia per individual basal cell and the ratio of the total cross-sectional area of the pseudopodia to the base area of the basal cell, using the transmission electron microscope, was developed. The diameters and areas of the bases of basal cells and the pseudopodia were also obtained. The number of pseudopodia per basal cell (N) and the ratio of the areas (F) measured in normal human uterine cervical epithelium were 34.22 and 0.338, respectively. The values observed in reactive atypia were 23.62 and 0.188; and those in mild dysplasia of the cervical epithelium (the earliest premalignant condition of the cervical epithelium), 26.98 and 0.226. There were statistically significant reductions in the number of pseudopodia per cell (N) and the ratio of areas (F) in the latter two pathological conditions compared to the controls. This morphometric method provides higher sensitive means by which one can quantify the characteristics of pseudopodia in various premalignant epithelia.