| 1. |
- Andersson, Madeleine, et al.
(författare)
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Calcium-dependent proteolysis in rabbit lens epithelium after oxidative stress
- 1998
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Ingår i: Ophthalmic Res. ; 30:3, s. 157-67
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Tidskriftsartikel (refereegranskat)abstract
- The purpose of this study was to examine changes in calcium-dependent proteolytic activity in the lens epithelium from whole rabbit lenses exposed to long-term oxidative stress at near physiological levels. Rabbit lenses, incubated in 50 microM H2O2 for 1 or 24 h, were checked for clarity and morphological changes in the epithelium. Proteolytic activity was measured in the epithelium using a fluorogenic synthetic substrate; N-succinyl-Leu-Tyr-7-amino-4-methylocoumarin, both in the presence and the absence of calcium (1 mM Ca2+ and 5 mM EDTA respectively). The effect on transparency and morphology of the epithelium following a 1-hour incubation in 100 microM H2O2 was also studied. Lenses incubated in 50 microM H2O2 were clear even after 24h. After a 1-hour incubation in 50 microM H2O2 the epithelium of the exposed lens appeared normal. However, after 24 h the epithelium cells appeared swollen and microscopical examination showed extensive intracellular and subepithelial vacuolization. Incubation in 100 microM H2O2 for 1 h caused loss of transparency; vacuole formation, globulization of the superficial lens fibers and death of the epithelial cells. There was a 55% increase in calcium-dependent proteolytic activity after 1 h in 50 microM H2O2, implying a role for the calcium-activated protease calpain in oxidatively induced cataract.
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| 2. |
- Andersson, Madeleine, et al.
(författare)
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Caspase and proteasome activity during staurosporin-induced apoptosis in lens epithelial cells
- 2000
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Ingår i: Investigative Ophthalmology and Visual Science. - 0146-0404. ; 41:9, s. 2623-32
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Tidskriftsartikel (refereegranskat)abstract
- PURPOSE: To determine what caspases are activated during staurosporin-induced apoptosis in cultured bovine lens epithelial cells (BLECs), to study the time course of caspase activation in relation to morphologic changes, and to investigate the effect of caspase and/or proteasome inhibition on apoptosis. METHODS: BLECs were incubated with staurosporin at different concentrations or for different times. Phosphatidylserine (PS) externalization was detected by annexin-V labeling, nuclear morphology was studied by staining with Hoechst 33342 stain (Hoechst, Frankfurt, Germany), and the percentage of apoptotic cells was determined by the TdT-dUTP terminal nick-end labeling (TUNEL) assay. The activity of caspase-1, -2, -3, -4, -8, and -9 as well as the chymotrypsin-like activity of the proteasome was measured by the use of fluorogenic peptide substrates. Inhibition of the proteasome was performed by incubation with 10 microM lactacystin, and caspases were inhibited by 1 microM Z-DEVD-FMK or 20 microM Z-VAD-FMK. RESULTS: Staurosporin treatment caused a dose- and time-dependent increase in the number of apoptotic cells and in caspase-3 activity. Activation of caspase-2, -4, -8, and -9 was also seen. Caspase activity was increased after 3 hours' incubation with 1 microM staurosporin, which is also the time when most cells became annexin-V-positive. Nuclear changes indicative of apoptosis, viewed with both Hoechst and TUNEL staining, appeared after 4 to 6 hours of staurosporin incubation. Incubation of BLECs with lactacystin caused reduction of proteasome activity and increased apoptosis, evidenced in both the TUNEL assay and caspase-3 activation. Preincubation of lens epithelial cells with caspase inhibitors caused complete inhibition of lactacystin- or staurosporin-induced caspase-3 activation (Z-DEVD-FMK/Z-VAD-FMK) and also of caspase-2, -4, -8, and -9 (Z-VAD-FMK), but the reduction in TUNEL-positive cells was only partial. PS translocation and DNA fragmentation after staurosporin treatment occurred despite complete caspase blockade. CONCLUSIONS: Staurosporin-induced apoptosis in BLECs involves activation of several caspases. Inhibition of the proteasome causes caspase-3 activation and apoptosis. Both staurosporin- and lactacystin-induced apoptosis can be executed in a caspase-independent manner. The present data are useful for understanding of proteolytic mechanisms during apoptosis in lens epithelial cells, which may be an important event in normal lens development as well as in some types of cataract.
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| 3. |
- Andersson, Madeleine, et al.
(författare)
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Decreased caspase-3 activity in human lens epithelium from posterior subcapsular cataracts
- 2003
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Ingår i: Experimental Eye Research. - 0014-4835. ; 76:2, s. 175-82
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Tidskriftsartikel (refereegranskat)abstract
- Apoptosis has been implied in normal lens development in the embryo as well as in lens fibre differentiation. It has also been suggested to play a role in non-congenital cataract and in the formation of posterior subcapsular opacification, but data on the presence of apoptosis in human lens epithelium from cataractous lenses are scarce and conflicting. The present study aimed to investigate apoptosis in lens epithelium from patients undergoing cataract surgery. The amount of apoptosis detected was correlated to age, gender, type of cataract, medications and disease. Moreover, the ability of human lens epithelial cells in culture to respond to the apoptosis-inducing agent staurosporin by activation of caspase-3 was investigated. Human lens capsulotomy specimens were collected immediately after surgery, frozen and later analysed with respect to caspase-3 activity, using the fluorogenic substrate Ac-DEVD-AMC. Generally, the activity of caspase-3 detected in this manner was very low and in 23% of the specimens it was non-detectable. However, there were differences in caspase activity between lens epithelial cells from different types of cataract, where samples from lenses with posterior subcapsular cataract exhibited significantly lower caspase-3 activity than lenses with a clear subcapsular zone. Age, gender or medications did not show any correlation with caspase activity but human capsulotomy specimens from diabetic patients exhibited significantly lower caspase-3 activity. Staurosporin caused a concentration-dependent increase in caspase activity in cultured human lens epithelial cells and the amount of apoptotic nuclei was also increased as viewed by staining with Hoechst 33342, showing chromatin condensation and nuclear fragmentation. Similar results were obtained when fresh human lens capsulotomy specimens were exposed to 1000 nM staurosporin for 24 hr. To conclude, the present data indicate that human lens epithelial cells have the ability to respond to apoptosis-inducing agents with caspase-3 dependent apoptosis, and that even though the general level of apoptosis in human lens epithelium in vivo is low, there are differences in caspase-3 activity levels in lenses with or without posterior subcapsular cataract. The latter finding supports previous studies indicating that this type of cataract may result from defective differentiation, in which apoptosis may play an important role.
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| 4. |
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| 5. |
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| 6. |
- Billger, Martin, et al.
(författare)
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Calpain processing of brain microtubules from the Atlantic cod, Gadus morhua.
- 1993
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Ingår i: Molecular and cellular biochemistry. - 0300-8177. ; 121:1, s. 85-92
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Tidskriftsartikel (refereegranskat)abstract
- Microtubules isolated from Atlantic cod (Gadus morhua) brains retained assembly competence and ultraculture, although treatment with rabbit calpain resulted in loss of MAPs. In addition, spirals and aberrant structures formed when calpain I was activated post assembly. No such effect was seen with calpain II. Soluble fractions from cod brain were found to contain proteolytic activity that could be blocked by exogenously added calpastatin. Calpain was also isolated from cod muscle tissue with 10 times less yield, compared to rabbit lung. On the basis of Ca(2+)-requirements for activation in the mM range, electrophoretic mobility, antigenicity and hydrophobicity, we conclude that the proteolytic activity was attributable to calpain II. There was no difference in effects of rabbit and cod calpain II on cod microtubule proteins, indicating that calpain is a conserved protein. Our results suggest that calpains might be involved in the Ca(2+)-dependent irreversible regulation of cod brain microtubules.
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| 7. |
- Celojevic, Dragana, 1985, et al.
(författare)
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Effects of 17beta-estradiol in human lens epithelial cells
- 2010
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Ingår i: Acta Ophthalmologica. 2010;88:S246. Presented at European Vision and Eye Research (EVER) Annual Meeting 2010, 6-9 Oct, Crete, Greece.. - : Wiley.
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Konferensbidrag (refereegranskat)
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| 8. |
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| 9. |
- Celojevic, Dragana, 1985, et al.
(författare)
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Superoxide dismutase gene polymorphisms in patients with age-related cataract
- 2013
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Ingår i: Ophthalmic genetics. - : Informa UK Limited. - 1744-5094 .- 1381-6810. ; 34:3, s. 140-145
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Tidskriftsartikel (refereegranskat)abstract
- BACKGROUND: Functional polymorphisms in genes encoding antioxidant enzymes may result in reduced enzyme activity and increased levels of reactive oxygen species, such as superoxide radicals, which in turn may contribute to increased risk of age-related disorders. Copper-zinc superoxide dismutases, SOD-1 and SOD-3, and manganese superoxide dismutase, SOD-2, are enzymes involved in the protection against oxidative stress and detoxification of superoxide. In this study, we investigated a number of disease-associated single nucleotide polymorphisms (SNPs) of SOD1, SOD2 and SOD3, in patients with age-related cataract. MATERIALS AND METHODS: The study included an Estonian sample of 492 patients with age-related cataract, subgrouped into nuclear, cortical, posterior subcapsular and mixed cataract, and 185 controls. Twelve SNPs in SOD1, SOD2 and SOD3 were genotyped using TaqMan Allelic Discrimination. Haplotype analysis was performed on the SNPs in SOD2. RESULTS: None of the studied SNPs showed an association with risk of cataract. These results were consistent after adding known risk factors (age, sex and smoking) as covariates in the multivariate analyses and after stratification by cataract subtype. Analysis of SOD2 haplotypes did not show any associations with risk of cataract. CONCLUSIONS: If genetic variation in genes encoding SOD-1, SOD-2 and SOD-3 contributes to cataract formation, there is no major contribution of the SNPs analyzed in the present study.
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| 10. |
- Demelash, Abeba, 1962, et al.
(författare)
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Selenium has a protective role in caspase-3-dependent apoptosis induced by H2O2 in primary cultured pig thyrocytes.
- 2004
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Ingår i: European journal of endocrinology / European Federation of Endocrine Societies. - 0804-4643. ; 150:6, s. 841-9
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Tidskriftsartikel (refereegranskat)abstract
- OBJECTIVE: Hydrogen peroxide (H2O2), necessary for thyroid hormonogenesis, is produced at the apical surface of the thyroid follicular epithelium. Excess H2O2 is potentially cytotoxic and may contribute to the development of hypothyroidism, e.g. in severe selenium deficiency. Yet it is unclear how H2O2 contributes to thyroid cell death. DESIGN AND METHODS: H2O2-induced apoptosis and necrosis were studied in primary cultured pig thyroid cells. Glutathione peroxidase (GPx) activity was altered by culture in low serum with or without selenite substitution. Apoptosis was evaluated by spectrofluorometric measurement of caspase-3-specific substrate cleavage, and by analysis of DNA fragmentation by agarose gel electrophoresis. Necrosis was detected by 51Cr release from prelabeled cells. RESULTS: Exogenous H2O2 dose-dependently (100-400 micromol/l) activated caspase-3 within 3-12 h, and DNA degradation was observed after 24 h. The potency of H2O2 to induce apoptosis was low compared with that of staurosporine, a strong proapoptotic agent. H2O2-treated cells with reduced GPx activity showed increased caspase-3 activation. Incubation of serum-starved cells with selenite (10-100 nmol/l) normalized the GPx activity and reduced the activation of caspase-3 by H2O2. High H2O2 concentrations (400-800 micromol/l) were required to obtain necrosis. The H2O2-induced necrosis was exaggerated by both low GPx activity and catalase inhibition. CONCLUSIONS: Cytotoxic effects of H2O2 on thyroid cells include caspase-3-dependent apoptosis that occurs at H2O2 concentrations insufficient to induce necrosis. Selenium deficiency aggravates the apoptotic response, probably due to impaired capacity of GPx to degrade H2O2.
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| 11. |
- Jonhede, Sofia, et al.
(författare)
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Acute effects of the sigma-2 receptor agonist siramesine on lysosomal and extra-lysosomal proteolytic systems in lens epithelial cells
- 2010
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Ingår i: Molecular Vision. - 1090-0535. ; 16, s. 819-827
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Tidskriftsartikel (refereegranskat)abstract
- Purpose: The aim of the present study was to examine the effects of the sigma-2 receptor agonist, siramesine, on morphology, growth, cell death, lysosomal function, and effects on extra-lysosomal proteolytic systems in human lens epithelial cells. Methods: Human lens epithelial cells in culture were exposed to siramesine and examined for morphological changes using Nomarski optics or calcein. Lysosomes were evaluated using acridine orange and Magic Red (RR-cresyl violet). Nuclear morphology was studied using Hoechst 33342 and propidium iodide. Enzymatic activities in living cells or cell lysates were studied using fluorogenic substrates. Results: Siramesine at low concentrations increased the cytoplasmic proteolytic activity of the proteasome and the calpain system. Effects were also observed with respect to lysosomal morphology, acidity and function. In addition, activation of caspase-3 and the appearance of nuclei with an apoptotic morphology was found. Conclusions: Siramesine at low concentrations affects lens epithelial cells with perturbations of the major proteolytic systems and lysosomal morphology, resulting in caspase activation and cell death. Siramesine may be a possible substance for the treatment or prevention of posterior capsular opacification (PCO).
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| 12. |
- Karlsson, Jan-Olof, 1944, et al.
(författare)
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Acute Effects of Indomethacin on Mitochondrial Function and Glutathione Levels in the Mouse Lens in Organ Culture
- 2003
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Ingår i: Invest. Ophthalmol. Vis. Sci.. ; 44:5, s. 3500-
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Konferensbidrag (refereegranskat)abstract
- Purpose, Experiments were carried out to study the effects of indomethacin on markers for oxidative damage and mitochondrial function in the intact mouse lens. Methods, Mouse lenses were incubated with fluorogenic indicator dyes prior to administration of 50 {micro}M indomethacin. The response was monitored, in real time, for several hours. Mitochondrial depolarization was followed by preloading the lens with Rhodamine 123 (10 {micro}g/ml). Peroxide production was studied in lenses loaded with 50{micro}M DCF-DA and superoxide levels with 10 {micro}M hydroethidium. Glutathione levels were assayed with 50 {micro}M monochlorobimane following short-term administration of indomethacin. Results, No significant changes in the production of peroxide or superoxide were found up to 3 hours after administration of indomethacin. The level of reduced glutathione was reduced by approximately 10 percent 3 h after drug administration. There was a significant increase (20%) of mitochondrial depolarization, compared to control lenses, one hour following indomethacin administration. Conclusions, The frequent use of nonsteroidal anti-inflammatory drugs in ophthalmology necessitates a close examination of effects besides the anti-inflammatory ones. This study points to relatively rapid effects on mitochondrial function and glutathione levels in the cultured mouse lens. The possible relation between these changes and the development of cataract remains to be studied.
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| 13. |
- Karlsson, Jan-Olof, 1944, et al.
(författare)
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Acute Effects of the Sigma-2 Receptor Agonist Siramesine on Lens Epithelial Cells
- 2007
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Ingår i: Invest. Ophthalmol. Vis. Sci.. ; 48:5, s. 2432-
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Konferensbidrag (refereegranskat)abstract
- PurposeExperiments were carried out to study the effects of siramesine on markers for apoptosis, oxidative damage and mitochondrial function in primary cultures of human lens epithelial cells (HLEC). MethodsHLEC were incubated with 25 {micro}M siramesine for 1, 2, 3, 4, 6 and 8 hours. Caspase-3 was assayed in cell extracts with DEVD-AMC. Mitochondrial depolarization was assayed with JC-1. Peroxide production was studied with DCF-DA and superoxide levels with hydroethidium. Glutathione levels were assayed with monochlorobimane. ResultsSiramesine induced a significant increase of caspase-3 activity after 6h exposure to 25 {micro}M siramesine. Nuclear morphology examined with Hoechst 33342 showed signs of apoptosis after the same time intervals. A significant increase in the production of peroxide and superoxide were found up to 4 -8 hours after administration of siramesine. ConclusionsSiramesine, a piperidine analogue, was developed for the treatment of psychiatic disorders and is considered to be relatively nontoxic. This study, and others, indicate effects on cell growth, apoptosis and production of ROS. The sigma-2 receptor may be a regulator of HLEC growth and apoptosis.
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| 14. |
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| 15. |
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| 16. |
- Karlsson, Jan-Olof, 1944, et al.
(författare)
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Inhibition Of Glycogen Synthase Kinase (gsk-3) Affects Markers Of Oxidative Stress And Attenuates Apoptosis In Human Lens Epithelial Cells
- 2004
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Ingår i: Invest. Ophthalmol. Vis. Sci.. ; 45:5, s. 3508-
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Konferensbidrag (refereegranskat)abstract
- Purpose: GSK-3, an evolutionary conserved S/T kinase which regulates cell fate determination in diverse organisms have been implicated in the formation of amyloid b-peptides and the phosphorylation of tau and catenin. Inhibition of GSK-3 can be obtained via the structurally unrelated substances lithum or Kenpaullone. The lens and the lens epithelial cells are excellent models to study the role of this enzyme. Methods: Confluent human lens epithelial cells (HLEC) were exposed to the GSK-3 inhibitors lithium (2 mM) or Kenpaullone (2 {micro}M) for times upp to 24h. The cells were, before or after treatment, placed in medium containing fluorogenic indicators of oxidative damage. DCFH-DA was used to assay peroxides, mitochondrial function was evaluated with Rhodamine 123, monochlorobimane was used to assay intracellular glutathione (GSH) levels, Proteolytic activities were assayed, on line, with cell-permeable fluorogenic substrates.Proteasome and calpain activities were assayed with LLVY-AMC, Cathepsin B with RR-AMC or FR-AMC. Metalloproteases were assayed with AAF-AMC. Caspase-3, 8 and 9 were assayed in cell extracts with DEVD-, IETD- or LEHD-AMC, respectively. Results: The mitochondrial membrane potential and the level of GSH increased by 10% after treatment with Li or Kenpaullone for 24h. No change was observed for peroxide production. The basal (low) level of caspase-3 activity was decreased by 20%. No significant effects were found concerning caspase-8 or 9 activities. No effect was observed on the activity of calpain, the proteasome, metalloproteases and cathepsin D/E activity. Conclusions:Inhibition of GSK-3 may protect against oxidative damage and attenuate apoptosis in HLEC. No changes of the other major proteolytic systems in the cell were detected. These data may be important for the interpretation of Wnt signaling and cell growth in HLEC as well as for the formation of amyloid in the lens.
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| 17. |
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| 18. |
- Karlsson, Jan-Olof, 1944, et al.
(författare)
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Inhibition of Glycogen Synthase Kinase (GSK-3) Protects Against Oxidative Stress and Attenuates Apoptosis in Human Lens Epithelial Cells and the Mouse Lens in Organ Culure
- 2005
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Ingår i: Invest. Ophthalmol. Vis. Sci.. ; 46:5, s. 3867-
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Konferensbidrag (refereegranskat)abstract
- Purpose: GSK-3 may regulate Wnt signaling, gene expression, the cell cycle, cell differentiation and apoptosis. Inhibition of GSK-3 can be obtained via the structurally unrelated substances lithum or Kenpaullone. The lens and the lens epithelial cells are excellent models to study the role of this enzyme. Methods: Primary cultures of human lens epithelial cells (HLEC) or the mouse lens in organ culture were exposed to the GSK-3 inhibitors lithium (2 mM) or Kenpaullone (2 {micro}M) for times upp to 24h.The cells were, before or after treatment, placed in medium containing fluorogenic indicators of oxidative damage. DCFH-DA was used to assay peroxides, mitochondrial function was evaluated with Rhodamine 123 and JC-1, monochlorobimane was used to assay intracellular glutathione (GSH) levels, Proteolytic activities were assayed, on line, with cell-permeable fluorogenic substrates.Proteasome and calpain activities were assayed with LLVY-AMC, Cathepsin B with RR-AMC or FR-AMC. Metalloproteases were assayed with AAF-AMC. Caspase-3, 8 and 9 were assayed in cell extracts with DEVD-, IETD- or LEHD-AMC, respectively. Results: The mitochondrial membrane potential and the level of GSH increased by 10% after treatment of HLEC with Li or Kenpaullone for 24h. No change was observed for peroxide production. The basal (low) level of caspase-3 activity was decreased by at least 20%. No significant effects were found concerning caspase-8 or 9 activities. No effect was observed on the activity of calpain, the proteasome, metalloproteases and cathepsin D/E activity.The whole mouse lens in organ culture showed essentially the same elevated mitochondrial potential. The GSH increase was even more evident in the whole lens preparation. Conclusions: Inhibition of GSK-3 may protect against oxidative stress (and cataract) via prevention of MPT induction and attenuate apoptosis in HLEC.
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| 19. |
- Karlsson, Jan-Olof, 1944, et al.
(författare)
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Proteolysis in human lens epithelium as determined by a cell-permeable substrate
- 1998
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Ingår i: Investigative Ophthalmology and Visual Science. ; 40:1, s. 261-264
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Tidskriftsartikel (refereegranskat)abstract
- PURPOSE: To develop a system for continuous evaluation of proteolytic activity in human lens epithelium and to characterize factors of importance for the regulation of proteolytic activity in lens epithelial cells. METHODS: Human lens epithelial cells were obtained during cataract surgery. Capsule epithelium specimens consisted of the central parts of the anterior capsule and the underlying lens epithelium. The sample, with the cell-permeable substrate Suc-Leu-Leu-Val-Tyr-7-amino-4-methylcoumarin, was placed in a chamber, which was placed in a thermostat-controlled aluminum block. Fluorescence changes were continuously measured by the fiber optics of the luminometer, which was placed 5 mm above the buffer surface. RESULTS: After administration of substrate to the medium overlying the cells, the substrate was degraded at a relatively slow rate. Approximately 10 picomoles of amino-4-methylcoumarin were formed per minute. A significant increase of proteolytic activity could be observed after application of 1 microM ionomycin or 2 microM thapsigargin. No leakage of lactate dehydrogenase from the cells was observed during these procedures. Basal proteolytic activity was totally inhibited by the proteasome inhibitor lactacystin. Lactacystin also attenuated the response to ionomycin and thapsigargin. CONCLUSIONS: Human lens epithelium responds to increased Ca levels from external or internal stores with an increased proteolytic activity that may be mediated by calpain, by the proteasome, or by both. This calcium-dependent change in proteolytic activity may be of importance in the development of cataract.
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| 20. |
- Karlsson, Jan-Olof, 1944, et al.
(författare)
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Proteolysis in human lens epithelium determined by a cell-permeable substrate
- 1999
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Ingår i: Invest Ophthalmol Vis Sci. ; 40:1, s. 261-4
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Tidskriftsartikel (refereegranskat)abstract
- PURPOSE: To develop a system for continuous evaluation of proteolytic activity in human lens epithelium and to characterize factors of importance for the regulation of proteolytic activity in lens epithelial cells. METHODS: Human lens epithelial cells were obtained during cataract surgery. Capsule epithelium specimens consisted of the central parts of the anterior capsule and the underlying lens epithelium. The sample, with the cell-permeable substrate Suc-Leu-Leu-Val-Tyr-7-amino-4-methylcoumarin, was placed in a chamber, which was placed in a thermostat-controlled aluminum block. Fluorescence changes were continuously measured by the fiber optics of the luminometer, which was placed 5 mm above the buffer surface. RESULTS: After administration of substrate to the medium overlying the cells, the substrate was degraded at a relatively slow rate. Approximately 10 picomoles of amino-4-methylcoumarin were formed per minute. A significant increase of proteolytic activity could be observed after application of 1 microM ionomycin or 2 microM thapsigargin. No leakage of lactate dehydrogenase from the cells was observed during these procedures. Basal proteolytic activity was totally inhibited by the proteasome inhibitor lactacystin. Lactacystin also attenuated the response to ionomycin and thapsigargin. CONCLUSIONS: Human lens epithelium responds to increased Ca levels from external or internal stores with an increased proteolytic activity that may be mediated by calpain, by the proteasome, or by both. This calcium-dependent change in proteolytic activity may be of importance in the development of cataract.
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| 21. |
- Karlsson, Jan-Olof, 1944, et al.
(författare)
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Sigma-2 receptor agonists kill lens epithelial cells
- 2010
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Ingår i: Acta Ophthalmologica. 2010;88:S246. Presented at European Vision and Eye Research (EVER) Annual Meeting 2010, 6-9 Oct, Crete, Greece.. - : Wiley.
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Konferensbidrag (refereegranskat)
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| 22. |
- Lyckesvärd, Madeleine Nordén, et al.
(författare)
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Linking loss of sodium-iodide symporter expression to DNA damage
- 2016
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Ingår i: Experimental Cell Research. - : Elsevier BV. - 0014-4827. ; 344:1, s. 120-131
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Tidskriftsartikel (refereegranskat)abstract
- Radiotherapy of thyroid cancer with I-131 is abrogated by inherent loss of radioiodine uptake due to loss of sodium iodide symporter (NIS) expression in poorly differentiated tumor cells. It is also known that ionizing radiation per se down-regulates NIS (the stunning effect), but the mechanism is unknown. Here we investigated whether loss of NIS-mediated iodide transport may be elicited by DNA damage. Calicheamicin, a fungal toxin that specifically cleaves double-stranded DNA, induced a full scale DNA damage response mediated by the ataxia-telangiectasia mutated (ATM) kinase in quiescent normal thyrocytes. At sublethal concentrations (< 1 nM) calicheamicin blocked NIS mRNA expression and transepithelial iodide transport as stimulated by thyrotropin; loss of function occurred at a much faster rate than after I-131 irradiation. KU-55933, a selective ATM kinase inhibitor, partly rescued NIS expression and iodide transport in DNA-damaged cells. Prolonged ATM inhibition in healthy cells also repressed NIS-mediated iodide transport. ATM-dependent loss of iodide transport was counteracted by IGF-1. Together, these findings indicate that NIS, the major iodide transporter of the thyroid gland, is susceptible to DNA damage involving ATM-mediated mechanisms. This uncovers novel means of poor radioiodine uptake in thyroid cells subjected to extrinsic or intrinsic genotoxic stress. (C) 2016 Elsevier Inc. All rights reserved.
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| 23. |
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| 24. |
- Petersen, Anne, 1962, et al.
(författare)
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A new model for assessing proteolysis in the intact mouse lens in organ culture
- 2004
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Ingår i: Ophthalmic Res. - 1423-0259. ; 36:1, s. 25-30
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Tidskriftsartikel (refereegranskat)abstract
- PURPOSE: To develop a new method to investigate proteolysis in the intact lens in organ culture. METHODS: Intact mouse lenses were assayed at regular intervals for proteolytic activity using fluorogenic peptide substrates +/- addition of ionomycin. Specific inhibitors were used to determine the activity of calpains, the proteasome and acid lysosomal enzymes. RESULTS: Significant levels of proteolytic activity were present in the intact lens. Proteolysis was stimulated by ionomycin. Preincubation with an inhibitor to the proteasome significantly decreased proteolysis whereas inhibitors of calpain and acid lysosomal enzymes did not. CONCLUSION: This study indicates that in the intact mouse lens in culture, the proteasome is an important protease. Its activity is at least partially regulated by calcium.
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| 25. |
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| 26. |
- Petersen, Anne, 1962, et al.
(författare)
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COX-inhibitors and ASA; influence on oxidative stress in cultured mouse lens
- 2004
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Ingår i: Invest. Ophthalmol. Vis. Sci.. ; 45:5, s. 395-
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Konferensbidrag (refereegranskat)abstract
- Purpose: Several substances such as ASA and COX-inhibitors have been attributed antioxidant properties. The main objectives of this study were to examine possible protection against oxidative damage in cultured mouse lens by ASA, specific COX-2 as well as unspecific COX-1-2 inhibitors. Methods: Intact mouse lenses were exposed to acetylsalicylic acid (0.03 mM), celecoxib (0.5 {micro}M), indomethacin (0.5 {micro}M), diclofenac (0.5 {micro}M), and/or H2O2 (100 {micro}M) for 8 days. H2O2 were administered as a single dose each day. Lens transparency was examined every day by light microscopy. The intact lenses were assayed at regular intervals during 6 h for proteolytic activity using the fluorogenic peptide substrate Suc-Leu-Leu-Val-Tyr-AMC (LLVY-AMC) (50 {micro}M) suitable for the calpain and proteasome proteolytic systems. Prior to the oxidative stress experiment, dose-response curves had been performed in a similar manner using doses ranging between 0.5-50 {micro}M for COX-inhibitors, 0.03-3 mM for ASA and 10-100 {micro}M for H2O2. Results: Dose-response experiments showed no effect by COX-inhibitors at 0.5 {micro}M or ASA at 0.03 mM on proteolysis or lens transparency compared to control. Hence, these doses of drugs were used to look at possible protective effects in oxidative stress experiments. Opacification of lenses exposed to a single dose of 100 {micro}M H2O2 each day appeared at day 5. Proteolytic activity of the calpain - proteasome systems in lenses exposed to H2O2 in the presence of ASA, COX-2 or COX-1-2 inhibitors did not differ significantly from lenses exposed to H2O2. No protection against oxidative stress by these substances could be detected with regard to change in lens transparency. Conclusions: The protective effect of ASA, COX-2 and COX-1-2 inhibitors against oxidative damage in the lens may be limited. The mechanisms through which these substances exert their actions with respect to oxidative damage require further studies.
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| 27. |
- Petersen, Anne, 1962, et al.
(författare)
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Effect of NSAIDs on Proteolytic Activity and Morphology of the Mouse Lens in Organ Culture
- 2003
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Ingår i: Invest. Ophthalmol. Vis. Sci.. ; 44:5, s. 3499-
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Konferensbidrag (refereegranskat)abstract
- Purpose: To study the influence of nonsteroidal anti-inflammatory drugs (NSAIDs) on proteolytic activity in the mouse lens in organ culture and to examine possible morphological changes indicative of cataract induction. Methods: Mouse lenses were incubated in Earle's balanced salt solution for 24 hours prior to addition of indomethacin or diclofenac at 0.5, 5, or 50 {micro}M. The lenses were exposed to the NSAIDs from 1 to 12 days. Clarity of the lenses was studied every day. The proteolytic assay was performed on a microtiter plate using a fluorogenic substrate, Suc-Leu-Leu-Val-Tyr-AMC (LLVY-AMC) (50 {micro}M) suitable for the calpain and proteasome proteolytic systems. The preparation was assayed at regular intervals for degradation of the substrate during 24 hours. Morphology of the lenses was studied after the experiment by light microscopy. Results: Opacification of the lenses appeared at day 8 at 50{micro}M and at day 12 at 5{micro}M NSAID concentration for indomethacin as well as for diclofenac. However, the indomethacin-incubated lenses expressed a more severe turbidity than diclofenac-incubated ones. Following an initial decrease in proteolysis (day 1-3), incubation with indomethacin resulted in increased proteolytic activity (4-12 days) of calpain and/or the proteasome. Lenses exposed to diclofenac exhibited increased degradation of LLVY as compared to control lenses during the whole incubation period. Morphological examination of the lenses exposed to NSAIDs showed swelling of outer cortical fibres at 0.5 {micro}M. Higher concentrations presented major morphological changes, such as disrupted epithelial cells and swollen, partly globulized cortical fibres. Conclusion: The role of NSAIDs in cataract formation is still unclear; previous data has indicated a cataractogenic as well as a potential protective effect of NSAIDs against cataract formation. Our study demonstrates proteolytic disturbances in the calpain and proteasome system after NSAID administration and also morphological changes. Further studies are required to evaluate NSAID as a potential cataractogenic risk factor.
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| 28. |
- Petersen, Anne, 1962, et al.
(författare)
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Effects Of Dexamethasone In The Lens
- 2006
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Ingår i: Invest. Ophthalmol. Vis. Sci.. ; 47:5, s. 4110-
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Konferensbidrag (refereegranskat)abstract
- Purpose: The aim of the study was to investigate effects of glucocorticoids in the lens. Methods: Lens epithelial cells (HLEC) were exposed to dexamethasone for 24 hours. Cells were assayed for changes in superoxide production using dihydroethidium (HET), for alterations in peroxide production using DCFH-DA or for GSH variations using monochlorobimane (MCB). Apoptosis was determined by Caspase-3 assay and by nuclear morphology of Hoechst stained cells. Mitochondria depolarisation was measured using the potential-sensitive colour JC-1. Morphology was examined by transmission electron microscopy (TEM). Results: Apoptosis were increased in HLEC exposed to 1, 10, 100 and 1000 {micro}M dexamethasone as revealed by nuclear morphology studies. Caspase-3 activity was increased at 100 and 1000 {micro}M dexamethasone. No effect on GSH, superoxide or peroxide production by dexamethasone was present. High concentrations of dexamethasone (1000 {micro}M) depolarised the mitochondria. TEM showed multilayering of cells, mitochondrial changes and accumulation of membrane delimited electron dense material. Conclusions: The mechanism underlying dexamethasone induced apoptosis and morphological changes in HLEC are probably not due to oxidative effects.
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| 29. |
- Petersen, Anne, 1962, et al.
(författare)
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Effects of dexamethasone on human lens epithelial cells in culture
- 2008
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Ingår i: Mol Vis. - 1090-0535. ; 14, s. 1344-52
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Tidskriftsartikel (refereegranskat)abstract
- PURPOSE: Treatment with glucocorticoids is a well known risk factor for cataract development, although the pathogenic mechanism has not been elucidated. The aim of the study was to investigate the effects of glucocorticoids in cultured human lens epithelial cells. METHODS: Human lens epithelial cells (HLECs) were exposed to dexamethasone for 24 h. The number of viable cells was determined using the 3-[4, 5-dimethylthiazolyl-2]-2, 5-diphenyltetrazolium bromide (MTT) assay, and proliferation was quantified using Ki-67. Apoptosis was investigated by measuring caspase-3 activity and by evaluating nuclear morphology of cells stained with Hoechst 33342. Mitochondria depolarization was measured using the potential-sensitive color, JC-1. Cells were assayed for changes in superoxide production using dihydroethidium (HET), for alterations in peroxide production using dichlorofluorescein diacetate (DCFH-DA), and for glutathione (GSH) variations using monochlorobimane (MCB). Caspase-3 activity was also measured in HLECs simultaneously exposed to dexamethasone and the glucocorticoid antagonist, RU486. RESULTS: Low doses of dexamethasone (0.1 microM) resulted in increased proliferation of HLECs. Apoptosis was increased in HLECs exposed to 1 microM, 10 microM, and 100 microM of dexamethasone as revealed by nuclear morphology studies. Apoptosis was also confirmed by measuring caspase-3 activation. No effect on superoxide production by dexamethasone was seen. There were no effects on GSH levels or mitochondrial depolarization either. Only the highest concentration of dexamethasone (100 microM) caused an increase in peroxide production. In HLECs incubated with the glucocorticoid antagonist, RU486, apoptosis was induced at a lower concentration of dexamethasone (0.1 microM) than with dexamethasone alone. CONCLUSIONS: Low doses of dexamethasone cause a moderate increase in proliferation of cultured HLECs. Slightly higher but still physiologically relevant concentrations of dexamethasone result in a dose-dependent increase in apoptosis. Dexamethasone-induced apoptosis in HLECs does not seem to involve oxidative mechanisms. The proapoptotic effect of dexamethasone does not appear to act through the glucocorticoid receptor. Effects on proliferation and/or dysregulation of apoptosis in lens epithelial cells may be an important factor in human steroid-induced posterior subcapsular cataract.
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| 30. |
- Petersen, Anne, 1962, et al.
(författare)
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Intracellular effects of NSAIDs/ASA in oxidatively stressed human lens epithelial cells in culture.
- 2008
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Ingår i: Ophthalmic research. - : S. Karger AG. - 1423-0259 .- 0030-3747. ; 40:2, s. 77-85
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Tidskriftsartikel (refereegranskat)abstract
- The aim of the study was to examine the effects of nonsteroidal anti-inflammatory drugs (NSAIDs)/acetylsalicylic acid (ASA) on human lens epithelial cells (HLECs) during oxidative stress. HLECs were exposed to H2O2 in the absence or presence of indomethacin, diclofenac, celecoxib (NSAIDs) or ASA for 24 h. HLECs were assayed for changes in superoxide and peroxide production and for variations in glutathione. Mitochondrial depolarization was measured using the membrane potential-sensitive dye JC-1. The results of the study include reduction in superoxide and peroxide production as well as reduction in glutathione depletion in oxidatively stressed HLECs incubated with low concentrations of NSAIDs/ASA. However, no protection against H2O2-induced mitochondrial depolarization by NSAIDs/ASA could be seen. In conclusion, NSAIDs/ASA display reactive oxygen species-scavenging properties in H2O2-exposed HLECs in culture.
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| 31. |
- Petersen, Anne, 1962, et al.
(författare)
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Potential Protective Effects of NSAIDs/ASA in Oxidatively Stressed Human Lens Epithelial Cells and Intact Mouse Lenses in Culture
- 2005
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Ingår i: Ophthalmic Res. - : S. Karger AG. - 0030-3747. ; 37:6, s. 318-327
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Tidskriftsartikel (refereegranskat)abstract
- Purpose: To study possible toxic effects of indomethacin, diclofenac, and celecoxib (NSAIDs) and acetylsalicylic acid (ASA) as well as potentially protective effects of these substances in oxidatively stressed human lens epithelial cells (HLEC) and in intact mouse lenses in culture. Methods: HLEC and mouse lenses were incubated with NSAIDs or ASA alone or in the presence of H(2)O(2). To study apoptosis the cells were then either stained with Hoechst 33342 or assayed for caspase-3 activity. Mouse lenses were studied with respect to lens transparency. Results: Low concentrations of NSAIDs/ASA caused a significant protection against H(2)O(2)-induced apoptosis in HLEC whereas higher concentrations were toxic. Conclusion: The protective effects of NSAIDs/ASA against oxidative damage are confined to a relatively small therapeutic window. Copyright (c) 2005 S. Karger AG, Basel.
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| 32. |
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| 33. |
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| 34. |
- Petersen, Anne, 1962, et al.
(författare)
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Proteolytic Systems of the Mouse Lens. Effects of Ionophors and Growth Factors
- 2002
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Ingår i: Invest. Ophthalmol. Vis. Sci.. ; 43:12, s. 462-
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Konferensbidrag (refereegranskat)abstract
- Purpose: To characterize the basal proteolytic activity in the intact mouse lens in organ culture. To determine the effect of calcium ionophors, growth factors and insulin on proteolysis. Methods: Lenses from adult mice were placed in a medium containing a fluorogenic protease substrate. The preparation was placed in a fluorometer and continuously assayed for the degradation of the substrate during 4 hours. The following proteolytic systems were studied; calpain, the proteasome, metalloproteases, cathepsin B and cathepsin D/E. Results: The level of basal calpain and proteasome activity, as measured by degradation of LLVY-AMC, was low. This basal activity could not be inhibited by inhibitors of calpain (calpeptin), the proteasome (lactacystin) and the lysosomal enzymes (monensin). An increase of the proteolytic activity could be observed after application of thapsigargin (10{micro}M) and the Ca-ionophore, ionomycin (10{micro}M). This increase could be inhibited by the pretreatment with lactacystin. The growth factors, TGF-{beta} (50 ng/ml), FGF (50 ng/ml) and insulin (20 {micro}g/ml) had no significant effect on the activity of calpain or of the proteasome during 24 hours. The proteolytic activity of metalloproteases was very low using non-cellpermeable substrate (AAA-AMC, DQ gelatine and Suc-Gly-Pro-Leu-Gly-Pro-MCA). Significant activity was obtained with a cell-permeable substrate (AAF-AMC and with MOCAc-Pro-Leu-Gly-Leu-A2Pr(Dnp)-Ala-Arg-NH2). Cathepsin B and D/E activity was relatively low. After the proteolysis assay, the morphology of the lenses was examined using metacarn fixation and plastic resin embedding. Conclusion: The basal proteolytic activity of the lens was relatively low. Increased Ca-levels increased the proteolytic activity, which may derive from the proteasome and possibly from the calpain system. The growth factors did not affect the proteolysis in the lens for up to one day. Some swelling of the outer cortical fibres was seen in ionomycin and thapsigargin treated lenses.
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| 35. |
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| 36. |
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| 37. |
- Petersen, Anne, 1962, et al.
(författare)
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The Mechanism of Antioxidant Actions by NSAIDs/ASA in Cultured Human Lens Epithelial Cells
- 2005
-
Ingår i: Invest. Ophthalmol. Vis. Sci.. ; 46:5, s. 3854-
-
Konferensbidrag (refereegranskat)abstract
- Purpose: To study the possible mechanisms for protection against oxidative stress by indomethacin, diclofenac, celecoxib (NSAIDs) or acetylsalicylic acid (ASA) in cultured human lens epithelial cells (HLEC). Methods: Changes in peroxide levels in HLEC was measured after exposure to low concentrations (0, 0.005, 0.05 or 0,5 {micro}M) of indomethacin, diclofenac, celecoxib or acetylsalicylic acid for 24 hours. The cells were assayed at regular intervals during 24 h using 2',7'-dichlorodihydrofluorescein diacetate (DCFH-DA). Cultured HLEC were incubated with H2O2 (200 {micro}M) alone or in the presence of NSAIDs/ASA. The cells were then assayed for changes in GSH levels using monochlorobimane (MCB). Results: NSAIDs/ASA at concentrations previous described to be effective against oxidative stress in HLEC did not affect peroxide levels in cultured HLEC. The reducing capacity as measured as the GSH levels, were not significantly changed in oxidatively stressed HLEC. No toxic effects on GSH or peroxide levels were present. Conclusions: Indomethacin, diclofenac, celecoxib or acetylsalicylic acid does not exert their protective actions against oxidative stress via changed levels of endogenous peroxide or GSH.
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| 38. |
- Petersen, Anne, 1962, et al.
(författare)
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The proteasome and intracellular redox status: implications for apoptotic regulation in lens epithelial cells.
- 2007
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Ingår i: Current eye research. - : Informa UK Limited. - 0271-3683 .- 1460-2202. ; 32:10, s. 871-82
-
Tidskriftsartikel (refereegranskat)abstract
- Purpose: This study aimed to investigate redox regulation of the proteasome as well as the effect of proteasome inhibition on intracellular oxidative status and apoptosis. Methods: Oxidative stress was induced in cultured human lens epithelial cells (HLECs) and intact mouse lenses by 100 mu M H(2)O(2). HLECs were also exposed to the reduced and the oxidized forms of glutathione (GSH/GSSG) and the reducing agent dithiotreitol (DTT). The chymotrypsin-like, the trypsin-like, and the peptidylglutamyl peptidase activities of the proteasome were measured using synthetic fluorogenic substrates. Superoxide as well as peroxide production, mitochondrial membrane potential, and the level of GSH was measured in HLECs after proteasome inhibition by MG-132 or lactacystin. Apoptosis was determined by measuring caspase-3 activation and by studying apoptotic nuclei after staining with Hoechst 33342. Results: All three peptidase activities of the proteasome were inhibited by 100 mu M H(2)O(2) and by the oxidized form of glutathione (GSSG), whereas the reduced form (GSH) stimulated chymotrypsin-like and peptidylglutamyl peptidase activities in HLECs lysates. Intact mouse lenses exposed to 100 mu M H(2)O(2) exhibited loss of transparency and trends of decreased chymotrypsin-like proteasome activity as well as decreased GSH levels. Inhibition of the proteasome in cultured HLECs caused significant increase in apoptosis and disturbed intracellular redox balance. Simultaneous addition of exogenous GSH completely abolished the increased apoptosis seen after MG-132 treatment. Conclusions: This study supports the hypothesis that intracellular proteolytic and oxidative regulatory systems are tightly coupled. The current data also indicate that apoptosis by proteasome inhibition is mediated through oxidative mechanisms.
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| 39. |
- Pålsson, Ann-Zophi, 1971, et al.
(författare)
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Glucose and oxidative cell damage in intact mouse lenses and human epithelial cells
- 2004
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Ingår i: Invest. Ophthalmol. Vis. Sci.. ; 45:5, s. 1700-
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Konferensbidrag (refereegranskat)abstract
- Purpose: To develop an experimental system for continuous evaluation of oxidative stress-induced damage in cultured human epithelial cells (HLEC) and intact mouse lenses. Especially we want to analyze effects of glucose overexposure in order to elucidate the mechanism behind diabetic cataract. Methods: Confluent HLEC and intact mouse lenses were exposed to 50 mM glucose during 24 hours prior to analysis. The use of cell permeable fluorogenic substrates allowed us to follow, in real time, biochemical intracellular changes. For this propose 5 {micro}g/ml rhodamine 123 (R123) and 25 {micro}M monochlorobimane (MCB) were used for analysis of alterations in mitochondrial membrane potential and reduced glutathione (GSH) levels respectively. The production of superoxide was monitored via 5 {micro}M hydroethidium (HET), peroxides were asseyed with 20 {micro}M 2',7'-dichlorofluorescein-diacetate (DCFH-DA). 50 {micro}M Suc-Leu-Leu-Tyr-Val-7-amido-4-methylcoumarin (LLVY-AMC) was used for detection of proteolytic activity. Histological sections of cultured mouse lenses were examined under a light microscope. Results: Histological sections showed fiber cell swelling, globulization and liquefaction of equatorial cortex. The epithelial layer in the germinative zone shows an increase in cell height and signs of degeneration. However the lenses were still clear before fixation. In the presence of 50 mM glucose proteolytic activities were 1,7 and 3,8 fold higher in intact mouse lens and HLEC, respectively. The level of GSH was significantly increased by 18,1 % during high glucose. There was also a significant decrease by 16,9% in peroxide levels in HLEC but an increase by 31,6% in the intact mouse lenses. There were almost no changes in superoxide levels and mitochondrial membrane potential in comparison with the control. Conclusions: High glucose induced an adaptive antioxidant response in HLEC. In the formation of diabetic cataract it is possible that activation of protease systems are due to oxidative stress that might originate from changes in the intracellular levels of glucose or/and its metabolites.
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| 40. |
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| 41. |
- Skiljic, Dragana, 1985, et al.
(författare)
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Effects of 17β-Estradiol on Activity, Gene and Protein Expression of Superoxide Dismutases in Primary Cultured Human Lens Epithelial Cells
- 2018
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Ingår i: Current Eye Research. - : Informa UK Limited. - 1460-2202 .- 0271-3683. ; 43:5, s. 639-646
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Tidskriftsartikel (refereegranskat)abstract
- Purpose: Protective effects of estradiol against H 2 O 2 -induced oxidative stress have been demonstrated in lens epithelial cells. The purpose of this study was to investigate the effects of 17β-estradiol (E2) on the different superoxide dismutase (SOD) isoenzymes, SOD-1, SOD-2, and SOD-3, as well as estrogen receptors (ERs), ERα and ERβ, in primary cultured human lens epithelial cells (HLECs). Materia ls and methods: HLECs were exposed to 0.1 µM or 1 µM E2 for 1.5 h and 24 h after which the effects were studied. Protein expression and immunolocalization of SOD-1, SOD-2, ERα, and ERβ were studied with Western blot and immunocytochemistry. Total SOD activity was measured, and gene expression analyses were performed for SOD1, SOD2, and SOD3. Results: Increased SOD activity was seen after 1.5 h exposure to both 0.1 µM and 1 µM E2. There were no significant changes in protein or gene expression of the different SODs. Immunolabeling of SOD-1 was evident in the cytosol and nucleus; whereas, SOD-2 was localized in the mitochondria. Both ERα and ERβ were immunolocalized to the nucleus, and mitochondrial localization of ERβ was evident by colocalization with MitoTracker. Both ERα and ERβ showed altered protein expression levels after exposure to E2. Conclusions: The observed increase in SOD activity after exposure to E2 without accompanying increase in gene or protein expression supports a role for E2 in protection against oxidative stress mediated through non-genomic mechanisms.
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| 42. |
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| 43. |
- Skiljic, Dragana, 1985, et al.
(författare)
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Oestradiol levels and superoxide dismutase activity in age-related cataract : a case-control study
- 2016
-
Ingår i: BMC Ophthalmology. - : Springer Science and Business Media LLC. - 1471-2415. ; 16
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Tidskriftsartikel (refereegranskat)abstract
- Background: It has been suggested that the higher prevalence of cataract in women is caused by a withdrawal effect of oestrogen at menopause. In vitro studies have demonstrated protection of serum oestradiol (E2) against oxidative stress through upregulation of antioxidant enzymes, including superoxide dismutase (SOD). The purpose of the present study was to investigate E2 levels and SOD erythrocyte activity in patients with age-related cataract.Methods: The studied subjects consisted of 103 patients with age-related cataract and 22 controls. Cataracts were classified as nuclear, cortical, or posterior subcapsular. Blood samples were collected and data on smoking, hormonal use, diabetes and age at menarche/menopause was obtained for all individuals. Serum oestradiol analyses were performed with radioimmunoassay (RIA) and SOD activity was measured in erythrocyte lysates.Results: A negative correlation between age and E2 concentration was seen in a linear regression analysis. No correlation was seen between SOD activity and age or gender and no correlation between E2 levels and SOD activity was found using multiple linear regression. The mean level of E2 for all male subjects was 50.1 +/- 16.3 pmol/L, significantly higher compared to 13.8 +/- 11.8 pmol/L for postmenopausal women.Conclusion: The present study does not support a role for E2-induced effects on SOD in cataract formation. The findings of higher E2 levels in men than in postmenopausal women may suggest that decreased oestrogen at menopause is partially responsible for the gender-related difference in cataract prevalence. However, the latter can only be verified through prospective randomized trials using hormonal replacement therapy.
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| 44. |
- Wang, Xiaoyang, 1965, et al.
(författare)
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N-acetylcysteine reduces lipopolysaccharide-sensitized hypoxic-ischemic brain injury.
- 2007
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Ingår i: Annals of neurology. - : Wiley. - 0364-5134 .- 1531-8249. ; 61:3, s. 263-71
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Tidskriftsartikel (refereegranskat)abstract
- OBJECTIVE: Maternal inflammation/infection alone or in combination with birth asphyxia increases the risk for perinatal brain injury. Free radicals are implicated as major mediators of inflammation and hypoxia-ischemia (HI)-induced perinatal brain injury. This study evaluated the neuroprotective efficacy of a scavenging agent, N-acetylcysteine (NAC), in a clinically relevant model. METHODS: Lipopolysaccharide (LPS)-sensitized HI brain injury was induced in 8-day-old neonatal rats. NAC was administered in multiple doses, and brain injury was evaluated at 7 days after HI. RESULTS: NAC (200mg/kg) provided marked neuroprotection with up to 78% reduction of brain injury in the pre+post-HI treatment group and 41% in the early (0 hour) post-HI treatment group, which was much more pronounced protection than another free radical scavenger, melatonin. Protection by NAC was associated with the following factors: (1) reduced isoprostane activation and nitrotyrosine formation; (2) increased levels of the antioxidants glutathione, thioredoxin-2, and (3) inhibition of caspase-3, calpain, and caspase-1 activation. INTERPRETATION: NAC provides substantial neuroprotection against brain injury in a model that combines infection/inflammation and HI. Protection by NAC was associated with improvement of the redox state and inhibition of apoptosis, suggesting that these events play critical roles in the development of lipopolysaccharide-sensitized HI brain injury.
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| 45. |
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| 46. |
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| 47. |
- Zetterberg, Madeleine, 1969, et al.
(författare)
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Methylenetetrahydrofolate reductase genetic polymorphisms in patients with cataract.
- 2005
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Ingår i: American journal of ophthalmology. - : Elsevier BV. - 0002-9394. ; 140:5, s. 932-4
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Tidskriftsartikel (refereegranskat)abstract
- PURPOSE: Hyperhomocysteinemia is commonly associated with polymorphisms in the methylenetetrahydrofolate reductase (MTHFR) gene. The level of homocysteine can be lowered by dietary intake of folate. A protective effect of folate supplementation has been reported against cataract. Here we investigate MTHFR polymorphisms in human cataract. DESIGN: Retrospective case-control association study. METHODS: Patients with nuclear (n = 77), cortical (n = 155), posterior subcapsular (n = 119), and mixed (n = 151) cataract, and 187 controls were analyzed for the MTHFR 677C-->T and 1298A-->C polymorphisms using minisequencing technique. RESULTS: The wild-type MTHFR 677CC/1298AA genotype was strongly overrepresented among cataract cases (P = .003). This effect was most pronounced in the mixed cataract group (P < .001). Hyperhomocysteinemia-associated genotypes had similar frequencies in cataract and control groups. CONCLUSIONS: The previously reported protective effect of folate against cataract is not due to overrepresentation of hyperhomocysteinemia-associated MTHFR genotypes. Instead, the strong predominance of wild-type MTHFR in cataract may suggest impaired DNA synthesis as a cataractogenic factor.
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| 48. |
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| 49. |
- Zetterberg, Madeleine, 1969, et al.
(författare)
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Proteasome activity in human lens nuclei and correlation with age, gender and severity of cataract
- 2003
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Ingår i: Curr Eye Res. ; 27:1, s. 45-53
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Tidskriftsartikel (refereegranskat)abstract
- PURPOSE: The aim of this study was to measure proteasome activity in human lens nuclei resulting from cataract surgery and in different regions of donor lenses. METHODS: The chymotrypsin-like, the trypsin-like and the peptidylglutamyl-peptide hydrolysing activities of the proteasome were studied using synthetic flourogenic substrates. RESULTS: Proteasome activity did not show any correlation with age of the patients or with gender. Increased opacification of the lens nucleus, as estimated prior to surgery using a 4-grade scale, was significantly correlated with decreased activity of all peptidase activities in the insoluble fraction. In the donor lenses, all peptidase activities were highest in the epithelium and decreased rapidly towards the nucleus. CONCLUSIONS: The present study demonstrates that proteasome activity is preserved in the nucleus of lenses from elderly individuals, although a decrease can be seen with cataract formation. This finding may be of importance for elucidating the mechanism behind the formation of nuclear cataract.
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| 50. |
- Zhu, Changlian, 1964, et al.
(författare)
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Nuclear translocation and calpain-dependent reduction of Bcl-2 after neonatal cerebral hypoxia-ischemia
- 2010
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Ingår i: Brain, Behavior, and Immunity. - : Elsevier BV. - 0889-1591. ; 24:5, s. 822-30
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Tidskriftsartikel (refereegranskat)abstract
- Apoptosis-related mechanisms are important in the pathophysiology of hypoxic-ischemic injury in the neonatal brain. Caspases are the major executioners of apoptosis, but there are a number of upstream players that influence the cell death pathways. The Bcl-2 family proteins are important modulators of mitochondrial permeability, working either to promote or prevent apoptosis. In this study we focused on the anti-apoptotic Bcl-2 protein after neonatal cerebral hypoxia-ischemia (HI) in 8-day-old rats. Bcl-2 translocated to nuclei and accumulated there over the first 24h of reperfusion after HI, as judged by immunohistochemistry and immuno-electron microscopy. We also found that the total level of Bcl-2 decreased after HI in vivo and after ionophore challenge in cultured human neuroblastoma (IMR-32) cells in vitro. Furthermore, the Bcl-2 reduction was calpain-dependent, because it could be prevented by the calpain inhibitor CX295 both in vivo and in vitro, suggesting cross-talk between excitotoxic and apoptotic mechanisms.
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