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PubMed1997

Ras1-mediated modulation of Drosophila homeotic function in cell and segment identity.

Mutations of the Drosophila homeotic proboscipedia gene (pb, the Hox-A2/B2 homologue) provoke dose-sensitive defects. These were used to search for dose-sensitive dominant modifiers of pb function. Two identified interacting genes were the proto-oncogene Ras1 and its functional antagonist Gap1, prominent intermediaries in known signal transduction pathways. Ras1+ is a positive modifier of pb activity both in normal and ectopic cell contexts, while the Ras1-antagonist Gap1 has an opposite effect. A general role for Ras1 in homeotic function is likely, since Ras1+ activity also modulates functions of the homeotic loci Sex combs reduced and Ultrabithorax. Our data suggest that the modulation occurs by a mechanism independent of transcriptional control of the homeotic loci themselves, or of the Ras1/Gap1 genes. Taken together our data support a role for Ras1-mediated cell signaling in the homeotic control of segmental differentiation.

Animals↗
PubMed2005

Segmentation of complex cell clusters in microscopic images: application to bone marrow samples.

BACKGROUND: Morphologic examination of bone marrow and peripheral blood samples continues to be the cornerstone in diagnostic hematology. In recent years, interest in automatic leukocyte classification using image analysis has increased rapidly. Such systems collect a series of images in which each cell must be segmented accurately to be classified correctly. Although segmentation algorithms have been developed for sparse cells in peripheral blood, the problem of segmenting the complex cell clusters characterizing bone marrow images is harder and has not been addressed previously. METHODS: We present a novel algorithm for segmenting clusters of any number of densely packed cells. The algorithm first oversegments the image into cell subparts. These parts are then assembled into complete cells by solving a combinatorial optimization problem in an efficient way. RESULTS: Our experimental results show that the algorithm succeeds in correctly segmenting densely clustered leukocytes in bone marrow images. CONCLUSIONS: The presented algorithm enables image analysis-based analysis of bone marrow samples for the first time and may also be adopted for other digital cytometric applications where separation of complex cell clusters is required.

Algorithms↗
PubMed2025

Measuring Cell Dimensions in Fission Yeast Using Machine Learning.

In fission yeast (Schizosaccharomyces pombe), cell length is a crucial indicator of cell cycle progression. Microscopy screens that examine the effect of agents or genotypes suspected of altering genomic or metabolic stability and thus cell size are crucial for studying disruptions to cell cycle dynamics. This method is based on using an automated cell segmentation algorithm to measure S. pombe cells imaged by brightfield (BF) microscopy methods. PhotoPhenosizer (PP) is a machine learning-based tool designed for automated cell measuring and dimensional analysis of morphology frequency distributions. Integration of this method into large-scale pipelines for tracking cell dimension change streamlines morphological measurements, which facilitates the examination of cellular responses to genomic and metabolic stresses. In this protocol, we use PP to observe the effect of genomic instability on cell size dynamics over a 12-day chronological lifespan assay. Our results show that relative to wild-type cells, a replication stress mutant shows larger cells during chronological aging in excess glucose media. Our results are consistent with activation of checkpoints that regulate cell morphology in response to DNA damage. This method's application highlights the relevance of its incorporation in experimental routines that require large-scale image processing and its adoption by users with routine needs in S. pombe molecular research projects.

Schizosaccharomyces↗
PubMed2002

Cold storage preservation and warm ischaemic injury to isolated arterial segments: endothelial cell injury.

Injury to endothelial cells is thought to be important to the development of the vascular lesion of chronic rejection. It was the aim of this study to develop a semiquantitative method to assess endothelial injury in arterial grafts and to document the injury produced by cold storage preservation and additional warm ischaemia. Twelve- and 24-h cold preservation of rat aortic segments, together with an additional 1 h of warm ischaemia, were assessed. Electron micrographs of representative endothelial cells were scored for cytoplasmic, nuclear and mitochondrial injury. The overall injury score was obtained by addition of the individual scores. Storage for up to 24 h in University of Wisconsin (UW) and Terasaki did not produce any injury. Twenty-four hours of storage in Euro-Collins resulted in endothelial cell death. Injury occurred after 12 h of storage in Ross, Collins and normal saline, and the injury increased following 24 h of storage. One hour of warm ischaemia did not increase the injury. Injury to endothelial cells varies with the preservation solution used and the time of cold storage, so that both the type of solution and the storage time should be taken into account in clinical studies looking at the influence of cold ischaemia time and graft outcome.

Animals↗
PubMed1977

The effect of transposition to jejunum on epithelial cell kinetics in an ileal segment.

Epithelial cell kinetics were studied in an ileal segment after transposition to proximal jejunum. The number of cells per villus column in the transposed ileum increased after 4--7 days to reach values normal for jejunum after 14--30 days. This increase was accompanied by a simultaneous increase in the number of cells per crypt column up to 130% of values in jejunum and ileum in situ. The percentage of labelled crypt cells, after labelling with 3H-thymidine, and the relative size of the proliferative cell compartment in the crypt in the transposed ileum did not differ from values in the ileum in situ at any time interval after surgery. The total proliferative activity per crypt, which was determined by scintillation counting of isolated crypts after 3H-thymidine labelling, increased two-fold from 7 days after surgery. Cell migration studies showed that the increase in the number of villus cells was probably not caused by a change in the life span of the epithelial cells. It seems that the increase in the number of villus cells in ileal epithelium after transposition to proximal jejunum is brought about by an enlargement of the crypt, while the relative size of the proliferative cell compartment in the crypt remains unchanged.

Animals↗
PubMed1989

Localization of alpha 2u-globulin within protein droplets of male rat kidney: immunohistochemistry using perfusion-fixed, GMA-embedded tissue sections.

We investigated the light microscopic subcellular localization and quantitation of alpha 2u-globulin (alpha 2uG) in male rat kidney after 2,2,4-trimethylpentane exposure using a monoclonal antibody to alpha 2uG. Slices of perfusion-fixed kidney were cold-processed in glycolmethacrylate and the antigen localized after an avidin-biotin-horseradish peroxidase procedure with Hanker-Yates reagent as the chromagen. Light microscopic examination revealed resolution comparable to low-magnification electron microscopy, with excellent morphological detail of tissue architecture, including subcellular localization of alpha 2uG within lysosomes of P2 segment cells of the proximal tubule epithelium. The anatomical relationship between alpha 2uG and TMP-induced protein droplets observed after staining with Lee's methylene blue-basic fuchsin was studied using serial sections. Image analysis of selected P2 segments in treated and control rats revealed a high correlation between subcellular localization of alpha 2uG and protein droplet deposition in the cytoplasm of P2 segment cells of the proximal tubule epithelium. Quantitative morphometry of alpha 2uG-stained proximal tubule epithelium 72 hr after treatment with 50 mg/kg 2,2,4-trimethylpentane p.o. demonstrated a 1.5- to 2-fold increase in staining area of tubules from treated rats compared with controls. Similar increases of Lee's methylene blue-basic fuchsin-stained protein droplets were also observed, but quantitative morphometry of the protein droplets was technically more difficult owing to a lower staining contrast between droplets and the surrounding cytoplasm. This immunohistochemical procedure provides a valuable technique for further studies on the pathological role of alpha 2uG in protein droplet nephropathy of male rats induced by many environmental chemicals, and demonstrates the value of cold glycolmethacrylate processing to improve morphological detail.

Alpha-Globulins↗
PubMedDate not supplied

[Change in the activity of calcium ions during illumination of a suspension of visual cell outer segment fragments].

Changes in the activity of calcium ions in the medium containing outer fragments suspension of bovine eye retina rods have been studied by the method of calcium-selective electrodes. Illumination of the suspension increases calcium ion activity in the incubation medium. Photoinduced yield of calcium ions depends on Ca+2 concentration: it equals 0.11+/-0.015 M Ca2+/1m rodopsin in the medium containing 0.1 mM CaCl2 and 0.046+/-0.002Ca2+/1M rodopsin in the medium containing 0.05 mM CaCl2. In the medium containing more than 10(-4) M CaCl2 both an increase and a decrease of Ca2+ ions have been observed.

Animals↗
PubMed2006

A biased median filtering algorithm for segmentation of intestinal cell gland images.

In this paper, we introduce a biased median filtering image segmentation algorithm for intestinal cell glands consisting of goblet cells. While segmentation of individual cells are generally based on the dissimilarities in intensities, textures, and shapes between cell regions and background, the proposed segmentation algorithm of intestine cell glands is based on the differences in cell distributions. The intestine cell glands consist of goblet cells that are distributed in the chain-organized patterns in contrast to the more randomly distributed nongoblet cells scattered in the bright background. Four biased median filters with long rectangular windows of identical dimension, but different orientations, are designed based on the shapes and distributions of cells. Each biased median filter identifies a part of gland segments in a particular direction. The complete gland regions are the combined responses of the four biased median filters. A postprocessing procedure is designed to reduce the defects that may occur when glands are located very close together and to narrow the gapping areas because of the thin distribution of goblet cells. Segmentation results of real intestinal cell gland images are provided to show the effectiveness of the proposed algorithm.

Algorithms↗
PubMed1982

Cell clones and segmentation in leech development.

Cell lineage tracer dyes rhodamine-D-peptide and fluorescein-D-peptide were used to study the development of segmentation in embryos of the leech Helobdella triserialis. The earliest overt manifestation of segmentation is in the mesodermal cell layer of the germinal bands, where a repeating pattern of nearly isomorphic mesoblast clusters is seen at an early stage of development. With joint use of both lineage tracers, it is shown that each mesoblast cluster is a clone derived from a single primary mesoblast, as well as being the precursor to an adult mesodermal hemisegment. Not all of the primary mesoblasts give rise to hemisegments, however. Cell clones derived from the first primary mesoblasts produced contribute to the nonmetameric prostomial region, whereas the last primary mesoblasts produced appear to be supernumerary.

Animals↗
PubMed1989

[Ultrastructural observations of the retinal pigment epithelial cells and the outer segments of the rod visual cells after vortex vein occlusion in rabbit eyes].

The effect of occlusion of the vortex vein to the retinal pigment epithelial cells (RPE) and the outer segments of the rod visual cells (OS) of albino rabbits was observed by scanning electron microscopy. Three vortex veins of the left eye and two vortex veins of the right eye in each rabbit were ligated with 6-0 silk suture. The morphological changes were observed at 24 hours, 1, 2, and 4 weeks after the ligation. The changes of RPE showed no significant difference between ligation of 2 and 3 vortex veins. The RPE showed degenerative changes 1 week after the ligation and the degenerative changes became severe after 2 weeks. The OS, showed no change after 2 vortex veins occlusion, while after occlusion of 3 vortex veins OS revealed degenerative changes after 1 week and the degenerative changes reached a maximum after 2 weeks. The changes of both RPE and OS tended to return to normal at 4 weeks. The changes of both RPE and OS may be caused by reduction of the blood flow in the choroidal circulation.

Animals↗
PubMed1984

Regional differences of the proximal part of mouse epididymis: morphological and histochemical characterization.

Regional differences in the proximal part of mouse epididymis were reported to provide a morphological baseline for studies on functional zonation of this part that is critical in sperm maturation. Macroscopical, histological, ultrastructural, and histochemical observations permitted us to subdivide this part into five segments, characterized by epithelial height, nuclear position, cytological and histochemical features of principal cells. Segment I corresponded to the initial segment previously described in rodents. Segment II differed from segment I by endoplasmic reticulum (ER) and dictyosomes aspect in principal cells, apical alkaline phosphatase and Ca2+-dependent ATPase activities. Segment III was characterized by spermatozoa package, high content of cells in multivesicular bodies, mitochondria shape, complex interdigitating membranes, and strong periodic acid-Schiff (PAS)-positive cell border. Segments IV and V presented the same cytological features but differed by their esterase activity. In the principal cells of each segment, dense spherical concretions were scattered in ER caveolae. Cells with apical nuclei were classified into two groups. The cells of the first group presented the same morphological and histochemical features as the adjacent principal cells and were scattered in the five segments ("apical cells"). The cells of the second group differed from the others by their goblet shape, a dense cytoplasm, and a high mitochondria succinate-D activity. They presented different cytological and histochemical features depending on their localization in segments I ("narrow cells"), II ("prominent cells"), or III, IV, V ("mitochondria goblet-cells"). The possible relationships between epithelium structure and epididymal functions were herein discussed.

Animals↗
PubMed2001

Evaluation of the expression of NADPH oxidase components during maturation of HL-60 clone 15 cells to eosinophilic lineage.

OBJECTIVE: Superoxide-generating NADPH oxidase consists of the membrane-bound cytochrome b558 (gp91phox and p22Phox) and the cytosolic components (p67phox, p47phox, p40phox and rac). In this study, we evaluated the superoxide-generating activity and the expression of NADPH oxidase components during eosinophilic maturation using HL-60 clone 15 cell line. MATERIALS AND METHODS: HL-60 clone 15 cells were matured to eosinophils by incubation with 0.5 mM butyrate for 7 days, and NADPH oxidase components were detected by Northern blot, Western blot analyses and immunocytochemical staining. Moreover, superoxide-generating activity was examined by nitro blue tetrazolium (NBT) assay. RESULTS: Northern blot and Western blot analyses revealed that mRNAs and proteins for gp91phox, p67phox and p47phox were expressed after eosinophilic myelocyte stages, whereas mRNAs and proteins for p40phox and rac-2 were expressed from the promyelocyte stage. Interestingly, p22phox mRNA was expressed from the promyelocyte stage, but its protein was expressed after eosinophilic myelocyte stages. Consistent with the results of Western blotting, immunocytochemical staining of butyrate-induced HL-60 clone 15 cells indicated that gp91phox, p22phox, p67phox and p47phox were detected after eosinophilic myelocyte stages (eosinophilic myelocytes, eosinophilic metamyelocytes, eosinophilic band cells and eosinophilic-segmented cells), whereas p40phox and rac-2 were expressed from the promyelocyte stage. Moreover, almost the same results as those with butyrate-treated HL-60 clone 15 cells were obtained using human bone marrow cells by immunocytochemical staining. Furthermore, nitro blue tetrazolium (NBT) assay indicated that superoxide could be produced after eosinophilic myelocyte stages but not produced before the promyelocyte stage. CONCLUSIONS: Together these observations indicate that all the components for NADPH oxidase are expressed, and the superoxide-producing activity is obtained after myelocyte stages during eosinophilic maturation.

Amino Acid Sequence↗