The ratio of nuclear area to cytoplasm area represents how much stresses are transmitted to nucleus. MDA-MB-231 cells exhibit lower value of the ratios (Nuclear/Cytoplasm) for both substrates in comparison with MCF-7 cells, indicating the prominent cell stretching and more non-circular shape of the cellularmorphology. For both cells, the spreading parameter of Nuclear/Cytoplasm becomes greater for cells cultured on softer collagen gel substrate than stiffer collagen coat substrate. This indicates that the cell stretching is restricted and the more circular morphology of the cells is overwhelming. In addition, both cells were more spread accompanied by lower values of spreading parameter with incubation time from 24 to 72 h, except MCF-7 cells cultured on collagen gel substrate. The response against collagen gel substrate for incubation of MCF-7 cells does not exactly follow the same trend of the feature of MDA-MB-231 cells.
we examined whether the subcellular localization of wildtype MLC1 can affect alterations in both morph- ology and motility during the surface-targeting process. Indeed, some cells transfected with wildtype MLC1 also exhibited mutant-like intracellular localization of MLC1 proteins. From the randomly selected regions of interest (ROIs), 17.5% ( n = 30) of cells showed that the wildtype MLC1 proteins was trapped intracellularly with a small fraction of surface expression at 24 h after transfection. We classified MLC-expressing cells based on the subcellu- lar distribution of MLC1 proteins: PM-MLC1 cells, in which MLC1 is found at the plasma membrane, and ER- MLC1 cells, in which the significant fraction of MLC1 proteins are trapped in the ER. To detect surface- expressed MLC1 accurately, we placed the Myc-tag in the putative extracellular loop between the third and the fourth transmembrane domains. Immunofluorescence staining confirmed the exclusive accessibility of the Myc- tag from the extracellular side (Additional file 3: Figure S3). Interestingly, the PM-MLC1 cells exhibited abnormal spiky membrane protrusions with the significant surface expression of MLC1 (Fig. 4a, marked with triple asterisks), while the ER-MLC1 cells exhibited normal morphology with feeble MLC1 signals at the cell surface (Fig. 4a, an as- terisk), which is somewhat similar to cells expressing patient-derived mutants (Fig. 2a). An intermediate distri- bution of MLC1 with weak surface MLC1 signals and relatively few filopodia was also observed (Fig. 4a, double asterisks). Moreover, the polarized distribution of surface MLC1 resulted in asymmetric filopodia formation: The re- gion with higher surface expression of MLC1 induced well-developed filopodia formation while reduced surface expression could not. These asymmetric filopodia forma- tion indicates that localization of MLC1 at the plasma membrane is critical to the morphological changes of the cell membrane (Fig. 4b). We also compared cellular motil- ity between PM- and ER-MLC1 cells via live-cell imaging over 500 min. Kymographs of the PM- and ER-MLC1 cells indicated that cellular motility is regulated by the subcel- lular localization of MLC1 (Fig. 4c). The mean velocity of ER-MLC1 cells was significantly higher than that of PM- MLC1 cells (Fig. 4d), although no changes in directionality were observed as in the cases of patient-derived MLC1 mutants (Fig. 4e). The live-cell imaging allowed us to gain
Figure 11 Microphotographs of histopathological sections of SeNPs-administered (2.5, 5, 10, and 20 mg/kg) mice spleen at 10x magniﬁcation. (A) Healthy control showing normal splenic architecture with well-de ﬁ ned red and white pulp areas. (B – E) Spleen of SeNPs-administered mice preserved the normal splenic architecture in all groups, respectively. Microphotographs of histopathological sections of SeNPs-administered (2.5, 5, 10, and 20 mg/kg) mice liver at 40x magniﬁcation. (F) Liver section of healthy control mice with intact cellularmorphology, normal hepatocytes, and kupffer cells. (G – J) SeNPs administration led to occasional mononuclear lymphocytic cellular inﬁltration (arrows) along with normal cellularmorphology at all the tested doses. Microphotographs of histopathological sections of SeNPs-administered (2.5, 5, 10, and 20 mg/kg) mice kidney at 40x magni ﬁ cation. (K) kidney section of healthy mice showed normal morphology with intact glomeruli (black arrow) and tubular structures (*). (L–O) Normal morphology of kidney at the doses of 2.5 and 5 mg/kg but mild cellular inﬁltration was observed in the glomerular surroundings (thick arrows) at 10 and 20 mg/kg tested doses.
Our biochemical assays have identified a number of downstream events following Sema6A activation, involving phosphorylation of multiple proteins with functions in cytoskeletal regu- lation. The most direct interaction is with the cytoplasmic tyrosine kinase, Abl, which acts as an intermediary between transmembrane receptors and the cytoskeleton, affecting cellularmorphology, adhesion and migration . Sema6A activation by ligand binding leads to recruitment of Abl kinase to the cytoplasmic domain and to Abl phosphorylation. We show, using a deletion construct, that Abl recruitment is required for PlxnA2-induced cellular col- lapse of NIH3T3 cells, suggesting it is a proximal event in Sema6A ligand-induced reverse sig- naling, though this role remains to be demonstrated directly in neurons. It is not however required for the increase in cellular area and complexity that accompanies full-length Sema6A expression, providing additional evidence for a biochemically distinct, constitutive mode of reverse signaling by Sema6A.
Results from fluorescence microscopy revealed that infection by M. pachy- dermatis induces changes in the actin cytoskeleton. Uninfected cells (groups I and II) displayed a normal structure of the actin cytoskeleton (Figure 1(b)). In- fected cells exhibited loss of cellularmorphology on the yeast attachment sites to the cell, apparently overgrown cellular clusters in circular arrangements were formed, cells were elongated and rounded with loss of stress fibers and thickened cellular cortex (Figure 1(d)). When performing the analysis of the cultures in which ac- tin filaments and nuclei were detected by the direct fluorescence assay, autofluo- rescence was observed in the yeasts.
Novel physiological challenges in different environments can promote the evolution of divergent phenotypes, either through plastic or genetic changes. Environmental salinity serves as a key barrier to the distribution of nearly all aquatic organisms, and species diversification is likely to be enabled by adaptation to alternative osmotic environments. The threespine stickleback (Gasterosteus aculeatus) is a euryhaline species with populations found both in marine and freshwater environments. It has evolved both highly plastic and locally adapted phenotypes due to salinity-derived selection, but the physiological and genetic basis of adaptation to salinity is not fully understood. We integrated comparative cellularmorphology of the kidney, a key organ for osmoregulation, and candidate gene expression to explore the underpinnings of evolved variation in osmotic plasticity within two populations of sticklebacks from distinct salinity zones in the Baltic Sea: the high salinity Kattegat, representative of the ancestral marine habitat; and the low salinity Bay of Bothnia. A common-garden experiment revealed that kidney morphology in the ancestral high-salinity population had a highly plastic response to salinity conditions whereas this plastic response was reduced in the low-salinity population. Candidate gene expression in kidney tissue revealed a similar pattern of population- specific differences, with a higher degree of plasticity in the native high-salinity population. Together these results suggest that renal cellularmorphology has become canalized to low salinity, and that these structural differences may have functional implications for osmoregulation.
Abstract: Objective: This study is to investigate the effects of Guiqi polysaccharide (GQP) on H 2 O 2 -induced premature senescence in normal human fetal lung fibroblast WI-38 cells. Methods: WI-38 cells were subjected to treatments of GQP, Angelica sinensis polysaccharide (ASP), and Astragalus membranaceus polysaccharide (AMP), and then treated with H 2 O 2 to induce premature senescence. Morphological observation, MTT assay, senescence-associated β-galactosidase activity assessment, telomerase activity determination, cell cycle analysis, and Western blot analy- sis were performed to evaluate cellular senescence. Results: H 2 O 2 treatment induced premature senescence in WI-38 cells, as indicated by the decreased fibroblast proliferation activity and changed cellularmorphology. When treated with GQP, ASP, or AMP, the morphological changes in WI-38 cells induced by H 2 O 2 could be restored. SA-β-gal activity was elevated in H 2 O 2 -treated WI-38 cells, which could be decreased by GQP treatment. Moreover, compared with the normal control, H 2 O 2 treatment significantly inhibited the telomerase activity of WI-38 cells. However, GQP effectively elevated the telomerase activity of these senescent cells. Furthermore, flow cytometry and cell cycle analysis showed that GQP treatment could abrogate the cell cycle arrest in H 2 O 2 -treated WI-38 cells, which might contribute to the anti-senescent effects. In addition, GQP significantly affected the p53-p21 and p16-pRb pathways in H 2 O 2 -treated WI-38 cells. The effectiveness of GQP was superior to AMP or ASP treatment alone. Conclusion: GQP has protective effects in oxidative stress-induced senescence. Our findings suggest the promising role of GQP as an attractive and bio-safe agent with the potential to retard senescence and attenuate senescence-related diseases.
Glyco-engineered strains often show a decrease in pro- ductivity over time [18, 21, 25], which might be linked to their altered glycosylation machinery and therefore a stressed metabolism. Based on our recent findings with the ∆OCH1 strain [12, 26], we found that cellu- lar agglomeration and therefore an altered morphology, affected process performance over time. Therefore, we initially hypothesized that an altered glycosylation machinery might be the trigger for morphological devi- ations, due to an overall decrease in glycan length on the cell surface resulting in cellular agglomeration, as it was also shown for glyco-proteins . Based on this hypothesis, we performed a shake-flask screening under inducing conditions to produce HRP C1A as recombi- nant model product in different P. pastoris strains and monitored cellularmorphology by microscopy and flow cytometry. A hypermannosylating wt strain, the ∆OCH1 strain (Man 8–10 GlcNAc 2 glycans) and the SuperMan 5
The cytotoxicity of the Y 2 O 3 NPs in the HEK cells was evaluated by IC 50 determination, MTT assay measurement, measurement of LDH leakage, and an analysis of cellularmorphology. Figure 3A shows the data of the MTT assay to determine the cytotoxicity dose (IC 50 of Y 2 O 3 NPs). The IC 50 value for the Y 2 O 3 NPs was found to be 108 µ g/mL. Based on this IC 50 value, the decision was made to vary the concentration of Y 2 O 3 NPs from 0.25 to 50 µ g/mL for further cytotoxicity studies. Compared to that observed in the control cells, the exposure to Y 2 O 3 NPs was associated with cell clustering, cell rounding, reductions in cell size, and cell detachment from the substratum (Figure 3B). We repeated the MTT assay using similar time points and con- centrations to those used to assess morphological change. The inhibition of cell survival by Y 2 O 3 NPs was time and concentration dependent (Figure 3C). At 10 hours, only higher concentrations $6.125 µ g decreased cell survival significantly (P,0.05); whereas, at 24 hours and at 48 hours of exposure, there were significant differences at all concentrations examined. We also found that cotreatment with NAC appeared to provide a protective effect against Y 2 O 3 NPs at 24 hours and at 48 hours. As shown in Figure 3D, LDH leakage due to Y 2 O 3 NPs was time and concentra- tion dependent. Compared to untreated controls, paraquat showed significant changes in the MTT assay but not LDH leakage at each time point. Similar to our MTT analysis, we also found that a cotreatment with NAC appeared to provide a protective effect against Y 2 O 3 NPs at 48 hours (P,0.05).
Bone marrow aspiration is comparatively easy to perform with less discomfort to the patient, better appreciation of cellularmorphology, being more suitable for cytochemical stains and other ancillary studies like flow cytometry, cytogenetics, culture etc ; however, the disadvantages included are, that cellularity cannot be assessed properly, some times we can get dry taps because of marrow fibrosis or if marrow is packed with blasts as in case of CML- Blast phase. Megakaryocytic morphology and fibrosis and cell topography is best assessed on bone marrow biopsy. Ancillary Techniques like immunohistochemistry can be used on biosy sections . Assessment of marrow fibrosis has been shown to have clinical and prognostic implication in different neoplasms. In chronic myelogenous leukaemia, this parameter has been demonstrated to have predictive value on therapy and outcome. CML is one of the most common leukaemia’s in adults and was the first hematological neoplasm where cytogenetic alterations and the development of leukemia could be associated . In this study, there were 13 cases of chronic myeloid leukemia [CML] [n = 13], which were concordant on aspiration and biopsy similar to Ghodasara and Gonsai. The aspirates are better able to classify the phases of CML as compared to biopsy . Evaluation of megakaryopoiesis, grading of fibrosis and localization of blasts are possible on a trephine biopsy.
We hypothesized that the expression of ERC/mesothelin influences the morphology of cells, as its expression is well correlated to the histological subtypes of mesotheli- oma. Thus, we examined the effect of ERC/mesothelin overexpression or knockdown on cellularmorphology. As shown in Fig. 1a, ERC/mesothelin overexpression in H2452 (spindle-shaped) or knockdown in H226 (polyg- onal) cells did not affect cell morphology. The overex- pression or knockdown of ERC/mesothelin was confirmed in Fig. 1b and c. The manipulation of ERC/ mesothelin expression did not have any effect on epithe- lial–mesenchymal transition (EMT) markers such as E- cadherin, vimentin (Fig. 1b), or ZEB1, or Twist (Add- itional file 1: Figure S1). We then examined the effects of ERC/mesothelin on cellular activities. We found that ERC/mesothelin overexpression enhanced cellular adhe- sion (Fig. 2a) and migration/invasion (Fig. 2b and c) with regard to the extracellular matrix (ECM), but did not in- fluence cellular proliferation (Fig. 2d and e). The expres- sion of MMP-9 was enhanced in ERC/mesothelin- overexpressing cells (Fig. 1b), but that of integrin α5 and integrin β1 remained unchanged (Additional file 1: Fig- ure S1).
targeting ligands and drug molecules), increased hydro- philicity, molecular gate-keeping, stimuli responsiveness, controlled cleavage of surface-bound coating or moieties, and so forth. On a less sophisticated level, it is generally believed that a net positive charge is beneficial in maxi- mizing cellular uptake due to attraction to the negatively charged cell membrane. Thus, functionalization with posi- tively charged surface groups is frequently applied to en- hance cellular uptake. This is also the primary reason why we decided to functionalize our particles with the cationic polyelectrolyte PEI. Aside from enhanced cellular uptake, the branched structure of PEI also offers a higher amount of terminal primary amino groups than possible to attain via conventional amino functionalization, simultaneously providing increased electrostatic suspension stability to the system and possibly introducing pH-dependent mo- lecular gate properties along with enabling endosomal es- cape ability, both of which would be utilized when carrying drug cargo for intracellular release . To inves- tigate the effect on cellular uptake upon derivatization of the PEI layer, we also capped the terminal primary amino groups with either uncharged acetyl groups or acidic (negatively charged under neutral conditions) succinic acid groups. The uptake in terms of positive cell percent- age of the whole series of particles with (or without) differ- ent functionalization is presented in Figure 7.
Computational simulations have become an indispensable tool for solving complex problems in engineering and science. One of the new computational techniques are the meshless methods, covering several application fields in engineering. In this paper the Smoothed Particle Hydrodynamics (SPH) method and its implementation in the explicit finite element code LS- DYNA is discussed. Its application and efficiency is shown with two practical engineering application examples. The first example describes the modeling of fuel sloshing in a reservoir, where different formulations, using mesh-based and meshless methods, are compared and evaluated according to experimental measurements. The second example describes the impact analysis of a cellular structure, where the influence of viscous fluid pore filler flow has been studied. The SPH method proved to become a reliable and efficient tool, especially for solving large scale and advanced engineering problems.
continuously with incubation solution at room temperature. Cells were electropo- rated under visual control of a CCD camera (TILL Photonics, Gräfelfing, Germany) mounted onto an upright micro scope (BX50WI, Olympus, Hamburg, Germany) equipped with gradient contrast illumination (Luigs and Neumann, Ratingen, Ger- many). Visually identified cells were approached with a glass pipette (tip diameter ∼ 1 µm) filled with either Alexa Fluor 488 sodium hydrazide or its 568 analog (1 mM, Molecular Probes, Eugene, OR). Alexa hydrazide derivatives with low molecular weight were used so that small cellular processes such as spines and filopodia were readily labeled, allowing accurate detection (Lang et al., 2006; Nevian and Helm- chen, 2007). The fluorescent dye-loaded pipette was pressed onto a selected cell and a single 18–20 ms long voltage pulse (15–20 V) was applied. The voltage pulse was generated by an EPC10/2 amplifier (HEKA Elektronik, Lambrecht, Germany) and post-amplified 10 times by a linear stimulus isolator (A395; World Precision Instru- ments, Berlin, Germany). Cell loading was visualized with a monochromator sys- tem (TILL Photonics). After 10–30 seconds the dye appeared evenly distributed within the dendrites of a given cell. Slices containing loaded cells were fixed in 4% paraformaldehyde overnight. After a 2× 5-minute wash with phosphate-buffered saline (PBS; pH 7.4), the slices were mounted in Vectrashield medium (H-100, Vec- tor, AXXORA, Lörach, Germany) on glass slides with a small spatial separation of plasticine sealed under a coverslip with nail polish.
Previous morphological studies on the mantle ultra- structure of various abalone species have reported the presence of pigments in tubular structures found in the region of the mantle thought to give rise to the prismatic layer [12,24]. Similar tubular structures have been found in the mantle of juvenile H. asinina . It has also been reported that, at least for land snails, pigmenta- tion patterns in the mantle match those found upon the shell . These findings hint that shell pigmenta- tion may not be controlled by the simple ‘line of cells’ that is assumed in the mathematical models, and that the system controlling pigmentation may be signifi- cantly more complex. In this paper, we investigate the cellular basis for control of pigmentation patterns in H. asinina , and extend these findings to other gastropod species.
Abstract Some experiments evidence that Microgravity (Micro-g) has influence in the morphology growth rate of materials and physiology of biological systems. Some Micro-g systems have been developed for experimentation under these conditions. The 2-D, 3-D clinostats and Random Positioning machines stimulate weightlessness. The aim of this review is to show the state of the art of the research related to Micro-g effect on inorganic and organic materials. Some noticeable examples related to changes in nanomaterial morphology; physiological effect, and cellular response in biological systems of plants cells as maize (Zea mays), rice (Oryza sativa) and bean (Phaseolus vulgaris); and osseous and muscular tissue will be shown. There has been identified the Micro-g as a new condition of significant structural and functional changes in the organization cellular.
We were interested in studying the mechanisms by which mutated TRNT1 might cause disease. Since some of the symptoms associated with SIFD appear to be re- lated to metabolic syndrome, we were interested in de- termining how the patient-derived TRNT1 mutations affect mitochondrial biology, and we focused our studies on mitochondrial processes in patient-derived fibro- blasts. Interestingly, immunofluorescence analysis of TRNT1 subcellular localization and electron microscopy of cells showed no gross morphological differences be- tween patient and control cells, and that the number of mitochondria per cell and their structure are similar. Our initial findings using micro-oximetry analysis dem- onstrated that basal and maximal respiration are de- creased in patient cells as compared to healthy control cells. Furthermore, our western blot analyses further suggest a defect in cellular respiration by demonstrating a decrease in expression of some OXPHOS complex subunits. Notably, at least two of the affected proteins (NDUFB8 (complex I) and SDHB (complex II)) are encoded in the nucleus, an observation which would be consistent with the global role of TRNT1 in tRNA mat- uration. However, we did not observe any defect in cyto- plasmic translation, suggesting that the decrease in abundance of these proteins might be due to complex assembly, rather than protein synthesis. Because the OXPHOS system is assembled from both nuclear- and mitochondrial-encoded components, proper assembly of these complexes is essential and, in fact mutations in assembly factors are associated with human pathologies . We have also observed a specific decrease in the abundance of the mitochondrial–encoded COX II (com- plex IV) protein. We are currently conducting SILAC experiments to identify the full spectrum of changes in the abundance and stoichiometry of all OXPHOS com- plexes as well as changes in the abundance of additional mitochondrial proteins.
As we saw a change in the astroglia morphology in the ACTG1 variant case, we sought to determine if other glial populations were affected. Hence, we stained for oligodendroglia using anti- OLIG2 and microglia with anti-Iba-1. In the CGE of the control case, there was a moderate to frequent expression of the OLIG2 protein (Figures 5A–Aii). Although oligodendrocytes are present in the ACTG1 variant, OLIG2 immunolabeled cells labeled with mouse anti- OLIG2 immunoreactivity were scant (Figures 5B– Bii). Surprisingly, with edema in the ACTG1 variant seen around the ventricular endothelial and white matter regions, there was only a low level of Iba-1 expression (Figures 5D–Dii). The control had ameboidal Iba1-positive microglia seen along the ventricular lining (Figures 5C,Ci) and in the CGE regions that were adjacent to the white matter (Figures 5C,Cii). The absence of oligodendroglia and microglia may indicate that there may be significant deficits in this brain with heterotopia which may interfere with the migrating neurons and the proliferation of supporting glia; how this attributes to the formation of the heterotopia remains unclear.