| 1. |
- Sundqvist, Tommy, et al.
(författare)
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Significantly increased levels of nitric oxide products in urine of children with celiac disease
- 1998
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Ingår i: Journal of Pediatric Gastroenterology and Nutrition - JPGN. - : Ovid Technologies (Wolters Kluwer Health). - 0277-2116 .- 1536-4801. ; 27:2, s. 196-198
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Tidskriftsartikel (refereegranskat)abstract
- Background: Celiac disease is characterized by morphologic and functional aberrations of the small intestinal mucosa, i.e. crypt hyperplasia, villous atrophy, infiltration of intraepithelial lymphocytes, and alteration of permeability. Nitric oxide has been shown to affect mucosal permeability after ischemia-reperfusion, but little is known about the regulatory role of nitric oxide in celiac disease. The purpose of this study was to assess nitric oxide production in children with celiac disease and in control subjects.Methods: The sum of nitrite and nitrate in the urine was measured with a colorimetric method in 137 children with a median age of 3 years, 84 patients and 53 reference children, all of whom underwent a small intestinal biopsy to confirm or overrule suspicion of celiac disease.Results: Median urinary nitrite-nitrate concentration in celiac children was 3323µM (4147 ± 1102; mean ± SEM) at first clinical examination and 2501 µM (2939 ± 386) after gluten challenge, which was significantly higher than concentrations in reference children(1029 µM; 1174 ± 116) and in children with celiac disease on a gluten-free diet (882 µM; 1369 ± 360) (p< 0.0001).Conclusions: A gluten-containing diet is associated with an increased nitrite-nitrate secretion in the urine in children with celiac disease, presumably as a result of nitric oxide synthase activation and nitric oxide production in the diseased small intestinal mucosa.
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| 2. |
- Andersson, Kerstin, et al.
(författare)
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Yersinia pseudotuberculosis-induced calcium signaling in neutrophils is blocked by the virulence effector YopH
- 1999
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Ingår i: Infection and Immunity. - 0019-9567 .- 1098-5522. ; 67:5, s. 2567-2574
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Tidskriftsartikel (refereegranskat)abstract
- Pathogenic species of the genus Yersinia evade the bactericidal functions of phagocytes. This evasion is mediated through their virulence effectors, Yops, which act within target cells. In this study we investigated the effect of Yersinia pseudotuberculosis on Ca 2+ signaling in polymorphonuclear neutrophils. The intracellular free calcium concentration in single adherent human neutrophils was monitored during bacterial infection and, in parallel, the encounter between the bacteria and cells was observed. When a plasmid-cured strain was used for infection, adherence of a single bacterium to the cellular surface induced a β 1 integrin-dependent transient increase in the intracellular concentration of free calcium. This was, however, not seen with Yop-expressing wild-type bacteria, which adhered to the cell surface without generating any Ca 2+ signal. Importantly, the overall Ca 2+ homeostasis was not affected by the wild-type strain; the Ca 2+ signal mediated by the G-protein-coupled formyl-methionyl-leucyl- phenylalanine receptor was still functioning. Hence, the blocking effect was restricted to certain receptors and their signaling pathways. The use of different Yop mutant strains revealed that the protein tyrosine phosphatase YopH was responsible for the inhibition. This virulence determinant has previously been implicated in very rapid Yersinia-mediated effects on target cells as the key effector in the blockage of phagocytic uptake. The present finding, that Y. pseudotuberculosis, via YopH, specifically inhibits a self- induced immediate-early Ca 2+ signal in neutrophils, offers more-detailed information concerning the effectiveness of this virulence effector and implies an effect on Ca 2+-dependent, downstream signals.
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| 3. |
- 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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| 4. |
- Hájková, Lucie, 1967-
(författare)
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The dynamic microfilament system : cellular organization of actin and profilin and their association with cell signalling
- 1999
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Active shape changes and movement of whole cells, and translocations within cells are essential to life. For instance, during embryogenesis cell migration is a central theme. Wound healing depends on increased and directed motile activity of several cell types. Platelets change their shape dramatically during blood clotting and restoration of blood flow. Cells of the immune system extravasate (leave blood vessels) and migrate to sites of infection, where they carry out their defense reactions etc. Furthermore, increased motile activity is one of the hallmarks of malignant tumor cells.All these movements are carried out by a system of muscle proteins. They are present in all our cells, in fact in all eukaryotic cells. The proteins actin and myosin play central roles in this system. They form superstructures inside the cell, converting chemical energy into movements. In non-muscle cells, they are part of a highly dynamic system called the microfilament system, where the basic component are filaments formed from actin. Microfilaments are particularly concentrated beneath the plasma membrane, where they are linked, directly or indirectly, to transmembrane proteins - receptors, adhesion proteins, and ion channels. The dynamic activity of the microfilament system is based on the ability the cell has to change the system, both with respect to organization and activity, and it does so in response to the interactions that it has with the immediate surroundings.The protein profilin is one of the crucial components in the regulation of the microfilament system. It binds to actin and controls its polymerizability. In addition to that, it appears to be an important link between the transmembrane signalling and the mechanisms that polymerize actin into filaments.This thesis deals with the role of profilin in vivo. It reports on the localization of profilin in cells and describes the effects of microinjecting normal and mutant profilins, and a chemically crosslinked profilin:actin complex into cells. The results strengthen the view that profilin plays a role in the activation of the microfilament system, and provides evidence that the profilin:actin complex is the precursor for filament formation and that dissociation of the complex is a crucial step in the polymerization reaction. The thesis also describes effects on the microfilament system by microinjection of antibodies against two isoforms of PI3-kinase, an enzyme that is involved in the metabolism of a particular type of phospholipid present in the plasma membrane and linked to the control of the microfilament system.
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| 5. |
- Holm, Åsa, 1969-, et al.
(författare)
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Mechanical manipulation of polymorphonuclear leukocyte plasma membranes with optical tweezers causes influx of extracellular calcium through membrane channels
- 1999
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Ingår i: Medical and Biological Engineering and Computing. - 0140-0118 .- 1741-0444. ; 37:3, s. 410-412
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Tidskriftsartikel (refereegranskat)abstract
- Optical tweezers are used mechanically to manipulate the plasma membrane of polymophonuclear leukocytes attached to the bottom of a glass manipulation chamber. The laser trapping beam is dragged across the membrane of cells in calcium-containing and calcium-depleted extracellular medium. This treatment causes a significant rise in the intracellular calcium concentration compared with controls, in cells in calcium-containing medium (239.8±49.0% against 75.4±16.4%, respectively), but not in cells in calcium-depeleted medium (69.1±9.6% against 83.4±18.5%, respectively), indicating that the calcium rise is caused by an influx of calcium from the environment. The rise in calcium concentration is blocked (23.5±7.1% against 17.1±4.1%, respectively) by the addition of lansoprazole, indicating that the influx is not due to unspecific membrane damage caused by the mechanical manipulation of the cell. It can therefore be concluded that mechanical manipulation of the neutrophil membrane, in the piconewton force range exerted by the optical tweezer, does not damage the plasma membrane but stimulates a mechanically inducible, membrane channel-mediated influx of extracellular calcium.
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| 6. |
- Holmström, Anna, et al.
(författare)
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YopK of Yersinia pseudotuberculosis controls translocation of Yop effectors across the eukaryotic cell membrane
- 1997
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Ingår i: Molecular Microbiology. - : Wiley. - 0950-382X .- 1365-2958. ; 24:1, s. 73-91
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Tidskriftsartikel (refereegranskat)abstract
- Introduction of anti-host factors into eukaryotic cells by extracellular bacteria is a strategy evolved by several Gram-negative pathogens. In these pathogens, the transport of virulence proteins across the bacterial membranes is governed by closely related type III secretion systems. For pathogenic Yersinia, the protein transport across the eukaryotic cell membrane occurs by a polarized mechanism requiring two secreted proteins, YopB and YopD. YopB was recently shown to induce the formation of a pore in the eukaryotic cell membrane, and through this pore, translocation of Yop effectors is believed to occur (Håkansson et al., 1996b). We have previously shown that YopK of Yersinia pseudotuberculosis is required for the development of a systemic infection in mice. Here, we have analysed the role of YopK in the virulence process in more detail. A yopK-mutant strain was found to induce a more rapid YopE-mediated cytotoxic response in HeLa cells as well as in MDCK-1 cells compared to the wild-type strain. We found that this was the result of a cell-contact-dependent increase in translocation of YopE into HeLa cells. In contrast, overexpression of YopK resulted in impaired translocation. In addition, we found that YopK also influenced the YopB-dependent lytic effect on sheep erythrocytes as well as on HeLa cells. A yopK-mutant strain showed a higher lytic activity and the induced pore was larger compared to the corresponding wild-type strain, whereas a strain overexpressing YopK reduced the lytic activity and the apparent pore size was smaller. The secreted YopK protein was found not to be translocated but, similar to YopB, localized to cell-associated bacteria during infection of HeLa cells. Based on these results, we propose a model where YopK controls the translocation of Yop effectors into eukaryotic cells.
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| 7. |
- Lirvall, Margareta, et al.
(författare)
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UVB radiation affects the mobility of epidermal growth factor receptors in human keratinocytes and fibroblasts
- 1996
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Ingår i: Bioscience Reports. - 0144-8463 .- 1573-4935. ; 16:3, s. 227-238
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Tidskriftsartikel (refereegranskat)abstract
- Growth factor receptors transmit biological signals for the stimulation of cell growth in vitro and in vivo and their autocrine stimulation may be involved in tumorigenesis. It is therefore, of great value to understand receptor reactions in response to ultraviolet (UV) light which certain normal human cells are invaribly exposed to during their growth cycle. UV irradiation has recently been shown to deplete antioxidant enzymes in human skin. The aims of the present study were a) to compare the lateral mobility of epidermal growth factor receptors (EGF-R) in cultured human keratinocytes and human foreskin fibroblasts, b) to investigate effects of ultraviolet B radiation on the mobility of EGF-R in these cells, and c) study the response of EGF-R on addition of antioxidant enzymes. The epidermal growth factor receptors were labeled with rhodaminated EGF, the lateral diffusion was determined and the fraction of mobile EGF-R assessed with the fluorescence recovery after photobleaching (FRAP). We found that human keratinocytes display a higher basal level of EGF-R mobility than human skin fibroblasts, viz, with diffusion coefficients (D ± standard error of the mean, SEM) of 4.2 ± 0.2 x 10-10 cm2/s, and 1.8 ± 0.2 x 10-10 cm2/s, respectively. UVB-irradiated fibroblasts showed an almost four-fold increase in the diffusion coefficient; D was 6.3 ± 0.3 x 10-10 cm2/s. The keratinocytes, however, displayed no significant increase in receptor diffusion after irradiation; D was 5.1 ± 0.8 x 10-10 cm2/s. In both cell types the percentage of EGF-R fluorescence recovery after photobleaching, i.e. the fraction of mobile receptors, was significantly increased after irradiation. In keratinocytes it increased from 69% before irradiation to 78% after irradiation. Analogous figures for fibroblasts were 61% and 73%. The effect of UVB on fibroblast receptors was abolished by prior addition of superoxide dismutase (SOD) and catalase (CAT). It is concluded that UVB radiation of fibroblasts and keratinocytes can affect their biophysical properties of EGF-R. The finding that addition of antioxidant enzymes prevented the UVB effect in fibroblasts may indicate the involvement of reactive oxygen metabolites.
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| 8. |
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| 9. |
- Pettersson, Jonas, et al.
(författare)
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Modulation of Virulence Factor Expression by Pathogen Target Cell Contact
- 1996
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Ingår i: Science. - : American Association for the Advancement of Science. - 0036-8075 .- 1095-9203. ; 273:5279, s. 1231-1233
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Tidskriftsartikel (refereegranskat)abstract
- Upon contact with the eukaryotic cell, Yersinia pseudotuberculosis increased the rate of transcription of virulence genes (yop), as determined by in situ monitoring of light emission from individual bacteria expressing luciferase under the control of the yopE promoter. The microbe-host interaction triggered export of LcrQ, a negative regulator of Yop expression, via the Yop-type III secretion system. The intracellular concentration of LcrQ was thereby lowered, resulting in increased expression of Yops. These results suggest a key role for the type III secretion system of pathogenic bacteria to coordinate secretion with expression of virulence factors after physical contact with the target cell.
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| 10. |
- 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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