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| 2. |
- Gustavsson, Johanna, 1956-, et al.
(författare)
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Localization of the insulin receptor in caveolae of adipocyte plasma membrane
- 1999
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Ingår i: The FASEB Journal. - 0892-6638 .- 1530-6860. ; 13:14, s. 1961-1971
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Tidskriftsartikel (refereegranskat)abstract
- The insulin receptor is a transmembrane protein of the plasma membrane, where it recognizes extracellular insulin and transmits signals into the cellular signaling network. We report that insulin receptors are localized and signal in caveolae microdomains of adipocyte plasma membrane. Immunogold electron microscopy and immunofluorescence microscopy show that insulin receptors are restricted to caveolae and are colocalized with caveolin over the plasma membrane. Insulin receptor was enriched in a caveolae-enriched fraction of plasma membrane. By extraction with β-cyclodextrin or destruction with cholesterol oxidase, cholesterol reduction attenuated insulin receptor signaling to protein phosphorylation or glucose transport. Insulin signaling was regained by spontaneous recovery or by exogenous replenishment of cholesterol. β-Cyclodextrin treatment caused a nearly complete annihilation of caveolae invaginations as examined by electron microscopy. This suggests that the receptor is dependent on the caveolae environment for signaling. Insulin stimulation of cells prior to isolation of caveolae or insulin stimulation of the isolated caveolae fraction increased tyrosine phosphorylation of the insulin receptor in caveolae, demonstrating that insulin receptors in caveolae are functional. Our results indicate that insulin receptors are localized to caveolae in the plasma membrane of adipocytes, are signaling in caveolae, and are dependent on caveolae for signaling.
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| 3. |
- Holmgren-Peterson, Kajsa, 1964-
(författare)
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Structure and dynamics of epithelial cells : Studied with confocal microscopy and flourescence recovery after photobleaching
- 1995
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Epithelial cells of the human body form a physical barrier to harmful agents and potential invading microorganisms. In the small intestine they must also produce enzymes for digestion of food and be able to absorb nutrients.The aim of this work was to study properties of epithelial cells in model systems using cell lines and toad bladder cells, and to study the effect of gluten intolerance (celiac disease, CD), viz. a pathological condition, on human small-intestine epithelial cells, enterocytes. Fluores-cence microscopy techniques, and primarily confocallaser scanning microscopy (CLSM), have beeen used as the major methods in the investigation. Part of the work has also been to develop, and apply, tools for measurements in confocal rnicroscopy images to obtain semi-quantitative information on structures.Fluorescence recovery after photobleaching (FRAP) was used to study the effect of maturation of small intestine-like epithelial cells. Lateral diffusion of membrane components was measured to reveal alterations in membrane fluidity induced by differentiation. No general effects on the lateral mobility of membrane components was observed, rather distinct effects were noticed on protein diffusion.Vasopressin induces the fusion of vesicles containing water channels with the apical membrane of toad bladder epithelial cells. This fusion is known to result in depolymerization of fllamentous actin (F-actin) of the cell. CLSM was used to assess where in the cell the depolymerization occurs. It was demonstrated that the depolymerization is not evenly distributed, but confined only to the apical region of the cells.In children suffering from celiac disease the mucosa of the small intestine is severely damaged. The damage at the enterocyte level is, however, less investigated. In the present work, CLSM was applied to compare the distributions ofF-actin and of glycoconjugates in enterocytes from children with CD to enterocytes from children not suffering from the disease. The results show that in children with active CD the distribution of both structures was altered, but also that compliance to a gluten-free diet results in the return to normal-looking enterocytes.
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- Immerstrand, Charlotte, 1974-, et al.
(författare)
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Height changes associated with pigment aggregation in Xenopus laevis melanophores
- 2004
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Ingår i: Bioscience Reports. - : Portland Press Ltd.. - 0144-8463 .- 1573-4935. ; 24:3, s. 203-214
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Tidskriftsartikel (refereegranskat)abstract
- Melanophores are pigment cells found in the skin of lower vertebrates. The brownish-black pigment melanin is stored in organelles called melanosomes. In response to different stimuli, the cells can redistribute the melanosomes, and thereby change colour. During melanosome aggregation, a height increase has been observed in fish and frog melanophores across the cell centre. The mechanism by which the cell increases its height is unknown. Changes in cell shape can alter the electrical properties of the cell, and thereby be detected in impedance measurements. We have in earlier studies of Xenopus laevis melanophores shown that pigment aggregation can be revealed as impedance changes, and therefore we were interested in investigating the height changes associated with pigment aggregation further. Accordingly, we quantified the changes in cell height by performing vertical sectioning with confocal microscopy. In analogy with theories explaining the leading edge of migrating cells, we investigated the possibility that the elevation of plasma membrane is caused by local swelling due to influx of water through HgC12-sensitive aquaporins. We also measured the height of the microtubule structures to assess whether they are involved in the height increase. Our results show that pigment aggregation in X. laevis melanophores resulted in a significant height increase, which was substantially larger when aggregation was induced by latrunculin than with melatonin. Moreover, the elevation of the plasma membrane did not correlate with influx of water through aquaporins or formation of new microtubules, Rather, the accumulation of granules seemed to drive the change in cell height.
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| 5. |
- Immerstrand, Charlotte, 1974-, et al.
(författare)
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Organelle transport in melanophores analyzed by white light image correlation spectroscopy
- 2007
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Ingår i: Journal of Microscopy. - : Wiley. - 0022-2720 .- 1365-2818. ; 225:3, s. 275-282
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Tidskriftsartikel (refereegranskat)abstract
- Intracellular transport of organelles, vesicles and proteins is crucial in all eukaryotic cells, and is accomplished by motor proteins that move along cytoskeletal filaments. A widely used model of intracellular transport is Xenopus laevis melanophores. These cells help the frog to change color by redistributing melanin-containing organelles in the cytoplasm. The high contrast of the pigment organelles permits changes in distribution to be observed by ordinary light microscopy; other intracellular transport systems often require fluorescence labeling. Here we have developed white light Image Correlation Spectroscopy (ICS) to monitor aggregation and dispersion of pigment. Hitherto in ICS, images of fluorescent particles from Confocal Laser Scanning Microscopy (CLSM) have been used to calculate autocorrelation functions from which the density can be obtained. In the present study we show that ICS can be modified to enable analysis of light-microscopy images; it can be used to monitor pigment aggregation and dispersion, and distinguish between different stimuli. This new approach makes ICS applicable not only to fluorescent but also to black-and-white images from light or electron microscopy, and is thus very versatile in different studies of movement of particles on the membrane or in the cytoplasm of cells without potentially harmful fluorescence labeling and activation.
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| 6. |
- Jager, Edwin W.H., et al.
(författare)
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The cell clinic : closable microvials for single cell studies
- 2002
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Ingår i: Biomedical microdevices (Print). - 1387-2176 .- 1572-8781. ; 4:3, s. 177-187
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Tidskriftsartikel (refereegranskat)abstract
- We present the development of a cell clinic. This is a micromachined cavity, or microvial, that can be closed with a lid. The lid is activated by two polypyrrole/Au microactuators. Inside the microvials two Au electrodes have been placed in order to perform impedance studies on single or a small number of cells. We report on impedance measurements on Xenopus leavis melanophores. We could measure a change in the impedance upon cell spreading and identify intracellular events such as the aggregation of pigment granules. The electrical data is correlated to optical microscopy.
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| 7. |
- Sjö, Anita, 1944-, et al.
(författare)
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Distinct Effects of Protein Kinase C on the Barrier Function at Different Developmental Stages
- 2003
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Ingår i: Bioscience Reports. - 0144-8463 .- 1573-4935. ; 23:2-3, s. 87-102
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Tidskriftsartikel (refereegranskat)abstract
- We show here, that activation of protein kinase C by the phorbol ester PMA improves barrier function in colon carcinoma (HT 29) cells. By contrast, in canine kidney (MDCK I) cells it caused increased permeability and opening of tight junctions; the latter has also been noticed in other studies. Thus, with PMA confluent HT 29 cells responded with a reduced passage of 330 kDa sodium fluorescein, increased transepithelial electrical resistance, and a change in the cell shape of the HT 29 cells from an irregular to a regular, hexagonal form. Confocal imaging revealed parallel distinct changes in the staining of occludin and caludin-1, viz. a translocation from cytoplasmic clusters to apical cell–cell contacts. Interestingly, in both cell lines protein kinase A activation caused a decreased in the threonine phosphorylation of occludin that correlated with tight junction assembly in HT 29 cells and tight junction disassembly in MDCK I cells. We conclude that protein kinase C regulation of the epithelial barrier involves specific molecular mechanisms and achieves distinct effects at different developmental stages.
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| 8. |
- Söderholm, Johan D, 1958-, et al.
(författare)
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Epithelial permeability to proteins in the noninflamed ileum of Crohn's disease?
- 1999
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Ingår i: Gastroenterology. - 0016-5085 .- 1528-0012. ; 117:1, s. 65-72
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Tidskriftsartikel (refereegranskat)abstract
- Background & Aims: Crohn's disease (CD) is associated with a disturbed intestinal barrier. Permeability studies have focused on inert molecules, but little is known about transepithelial transport of macromolecules with antigenic potential in humans. The aim of this study was to quantify permeation and to characterize passage routes for macromolecules in ileal mucosa in CD.Methods: Noninflamed and inflamed ileal mucosa specimens from patients with CD (n = 12) and ileal specimens from patients with colon cancer (n = 7) were studied regarding transmucosal permeation of ovalbumin, dextran (mol wt, 40,000), and 51Cr-EDTA for 90 minutes in vitro in Ussing chambers. Transepithelial passage routes for fluorescent ovalbumin and dextran 40,000 were investigated by confocal microscopy.Results: Noninflamed ileum from CD patients showed increased permeation of ovalbumin compared with ileum from colon cancer patients (P < 0.05). Dextran permeation was equal in the three groups, whereas 51Cr-EDTA permeability was increased in inflamed ileum. Ovalbumin passed both transcellularly and paracellularly, but dextran followed a strictly paracellular route. Both markers were subsequently endocytosed by cells of the lamina propria.Conclusions: Noninflamed ileal mucosa from patients with CD shows increased epithelial permeability to ovalbumin, probably by augmented transcytosis. This increase in antigen load to the lamina propria could be an initiating pathogenic event in CD.
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