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| 2. |
- Bergkvist, Liza, 1985-
(författare)
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Amyloid-β and lysozyme proteotoxicity in Drosophila : Beneficial effects of lysozyme and serum amyloid P component in models of Alzheimer’s disease and lysozyme amyloidosis
- 2017
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- In the work presented this thesis, two different conditions that are classified as protein misfolding diseases: Alzheimer's disease and lysozyme amyloidosis and proteins that could have a beneficial effect in these diseases, have been studied using Drosophila melanogaster, commonly known as the fruit fly. The fruit fly has been used for over 100 years to study and better understand fundamental biological processes. Although the fruit fly, unlike humans, is an invertebrate, many of its central biological mechanisms are very similar to ours. The first transgenic flies were designed in the early 1980s, and since then, the fruit fly has been one of the most widely used model organisms in studies on the effects of over-expressed human proteins in a biological system; one can regard the fly as a living, biological test tube. For most proteins, it is necessary that they fold into a three-dimensional structure to function properly. But sometimes the folding goes wrong; this may be due to mutations that make the protein unstable and subject to misfolding. A misfolded protein molecule can then aggregate with other misfolded proteins. In Alzheimer's disease, which is the most common form of dementia, protein aggregates are present in the brains of patients. These aggregates are composed of the amyloid-β (Aβ) peptide, a small peptide of around 42 amino acids which is cleaved from the larger, membrane-bound, protein AβPP by two different enzymes, BACE1 and γ-secretase. In the first part of this thesis, two different fly models for Alzheimer’s disease were used: the Aβ fly model, which directly expresses the Aβ peptide, and the AβPP-BACE1 fly model, in which all the components necessary to produce the Aβ peptide in the fly are expressed in the fly central nervous system (CNS). The two different fly models were compared and the results show that a significantly smaller amount of the Aβ peptide is needed to achieve the same, or an even greater, toxic effect in the AβPP-BACE1 model compared to the Aβ model. In the second part of the thesis, these two fly models for Alzheimer’s disease were again used, but now to investigate whether lysozyme, a protein involved in our innate immune system, can counteract the toxic effect of Aβ generated in the fly models. And indeed, lysozyme is able to save the flies from Aβ-induced toxicity. Aβ and lysozyme were found to interact with each other in vivo. The second misfolding disease studied in this thesis is lysozyme amyloidosis. It is a rare, dominantly inherited amyloid disease in which mutant variants of lysozyme give rise to aggregates, weighing up to several kilograms, that accumulate around the kidneys and liver, eventually leading to organ failure. In the third part of this thesis, a fly model for lysozyme amyloidosis was used to study the effect of co-expressing the serum amyloid P component (SAP), a protein that is part of all protein aggregates found within this disease class. SAP is able to rescue the toxicity induced by expressing the mutant variant of lysozyme, F57I, in the fly's CNS. To further investigate how SAP was able to do this, double-expressing lysozyme flies, which exhibit stronger disease phenotypes than those of the single-expressing lysozyme flies previously studied, were used in the fourth part of this thesis. SAP was observed to reduce F57I toxicity and promote F57I to form aggregates with more distinct amyloid characteristics. In conclusion, the work included in this thesis demonstrates that: i) Aβ generated from AβPP processing in the fly CNS results in higher proteotoxicity compared with direct expression of Aβ from the transgene, ii) lysozyme can prevent Aβ proteotoxicity in Drosophila and could thus be a potential therapeutic molecule to treat Alzheimer’s disease and iii) in a Drosophila model of lysozyme amyloidosis, SAP can prevent toxicity from the disease-associated lysozyme variant F57I and promote formation of aggregated lysozyme morphotypes with amyloid properties; this is important to take into account when a reduced level of SAP is considered as a treatment strategy for lysozyme amyloidosis.
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| 3. |
- Fahlgren, Anna, 1972-, et al.
(författare)
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Design and Outcome of a CDIO Syllabus Survey for a Biomedicine Program
- 2019
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Ingår i: The 15th International CDIO Conference: Proceedings – Full Papers. - Aarhus : Aarhus University. - 9788775074594 ; , s. 191-200
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Konferensbidrag (refereegranskat)abstract
- The CDIO Syllabus survey has successfully been applied to the Bachelor’s and Master’s programs in Experimental and Medical Biosciences, within the Faculty of Medicine and Health Sciences at Linköping University, Sweden. The programs are and have been, subject to considerable redesign with strong influence from the CDIO framework. One of the main drivers for the redesign is a shift concerning the main job market after graduation, from an academic career to industry and healthcare. One of the steps in the development process has been to carry out a Syllabus survey based on an adapted version of the CDIO Syllabus. The survey was sent out to students and to various categories of professionals, and in total 87 responses were received. The adapted version of the Syllabus and the design, execution, and outcome of the survey is presented.
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| 4. |
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| 5. |
- Kågedal, Katarina, 1970-
(författare)
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Cathepsin D released from lysosomes mediates apoptosis
- 2003
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Last year (2002), the Nobel Prize in Physiology or Medicine was awarded to three scientists who have conducted pioneer research on programmed cell death. In the human body, more than a thousand billion cells are created every day, and an equal number die, thus programmed cell death, or apoptosis, is an important mechanism for maintaining tissue homeostasis and protecting against disease. Malfunctioning apoptosis is associated with many pathological conditions, for example, excess apoptosis is characteristic of AIDS, stroke, neurodegenerative diseases, and myocardinal infarction, and insufficient apoptosis is seen in autoimmune conditions and cancer. Robert Horvitz, one of the mentioned Nobel Prize Laureates, was the first to identify death genes, namely ced-3, -4, and -9 in the nematode Caenorhabditis elegans, which were later discovered to have counterparts in humans.The aim of this thesis is to clarify the participation of lysosomes and lysosomal proteases in the initiation of apoptosis. The lysosomal enzyme cathepsin D regulates the human homologue of ced-3, which encoded the caspase family of proteases. Moreover, the human homologue of ced-9 encodes the Bcl-2 family of proteins such as Bax, which was involved in regulating the release of cathepsin D from lysosomes during apoptosis. In the present studies, apoptosis was induced by various substances, all of which first caused damage to lysosomes with ensuing release of lysosomal proteases. Fibroblasts exposed either to free radicals generated by the redox cycling quinone naphthazarin or to the kinase inhibitor staurosporine exhibited rapid translocation of cathepsin D from lysosomes to the cytosol and subsequent apoptosis. Malignant macrophages (J774 cells) and T lymphocytes (Jurkat cells) exposed to the lysosomotropic detergent sphingosine displayed early lysosomal destabilization and later apoptosis. Sphingosine also destabilized isolated lysosomes. Moreover, mimicking the translocation of cathepsin D by microinjecting cathepsin D into the cytosol induced apoptosis in fibroblasts.In the mentioned systems, lysosomes were destabilized before mitochondrial changes occurred and caspases were activated. Furthermore, apoptosis was prevented by inhibition of cathepsin D in the naphthazarin, staurosporine, and sphingosine systems and by inhibition of cysteine proteases such as cathepsins B and L in the sphingosine system. These results emphasize that cytosolic localization of lysosomal proteases is necessary for the ability of these enzymes to induce apoptosis.The present results also demonstrate that, during apoptosis, lysosomal membranes are destabilized by the following: (i) free-radical-mediated lipid peroxidation; (ii) pore formation through the Bcl-2 family member Bax; (iii) the impact of the lysosomotropic detergent sphingosine. All three of these events have been implicated in numerous other apoptosis systems. Accordingly, the participation of lysosomal enzymes in apoptosis may be more widespread than previously assumed. This new perspective on lysosomes as regulators of apoptosis may lead to novel treatment strategies for diseases associated with malfunctioning apoptosis.
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| 6. |
- Kågedal, Katarina, 1970-, et al.
(författare)
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Sphingosine-induced apoptosis is dependent on lysosomal proteases
- 2001
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Ingår i: Biochemical Journal. - 0264-6021 .- 1470-8728. ; 359:2, s. 335-343
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Tidskriftsartikel (refereegranskat)abstract
- We propose a new mechanism for sphingosine-induced apoptosis, involving relocation of lysosomal hydrolases to the cytosol. Owing to its lysosomotropic properties, sphingosine, which is also a detergent, especially when protonated, accumulates by proton trapping within the acidic vacuolar apparatus, where most of its action as a detergent would be exerted. When sphingosine was added in low-to-moderate concentrations to Jurkat and J774 cells, partial lysosomal rupture occurred dose-dependently, starting within a few minutes. This phenomenon preceded caspase activation, as well as changes of mitochondrial membrane potential. High sphingosine doses rapidly caused extensive lysosomal rupture and ensuing necrosis, without antecedent apoptosis or caspase activation. The sphingosine effect was prevented by pre-treatment with another, non-toxic, lysosomotropic base, ammonium chloride, at 10mM. The lysosomal protease inhibitors, pepstatin A and epoxysuccinyl-L-leucylamido-3-methyl-butane ethyl ester ('E-64d'), inhibited markedly sphingosine-induced caspase activity to almost the same degree as the general caspase inhibitor benzyloxycarbonyl-Val-Ala-DL-Asp-fluoromethylketone ('Z-VAD-FMK'), although they did not by themselves inhibit caspases. We conclude that cathepsin D and one or more cysteine proteases, such as cathepsins B or L, are important mediators of sphingosine-induced apoptosis, working upstream of the caspase cascade and mitochondrial membrane-potential changes.
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| 7. |
- Neuzil, Jiri, 1958-, et al.
(författare)
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Vitamin E analogs : A new class of multiple action agents with anti-neoplastic and anti-atherogenic activity
- 2002
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Ingår i: Apoptosis (London). - 1360-8185 .- 1573-675X. ; 7:2, s. 179-187
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Tidskriftsartikel (refereegranskat)abstract
- The incidence of cancer and atherosclerosis, two most common causes of death in developed countries, has been stagnating or, even, increasing. Drugs effective against such conditions are needed and, in this regard, the potential anti-atherosclerotic activity of vitamin E analogs has been studied extensively. Surprisingly, recent results indicate that these agents may also exert anti-neoplastic effects. Here we review the evidence that particular analogs of vitamin E may act as both anti-atherogenic and anti-cancer agents, and discuss the possible molecular bases for these actions.
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| 8. |
- Quinn, Carmel M., et al.
(författare)
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Induction of fibroblast apolipoprotein E expression during apoptosis, starvation-induced growth arrest and mitosis
- 2004
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Ingår i: Biochemical Journal. - 0264-6021 .- 1470-8728. ; 378:3, s. 753-761
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Tidskriftsartikel (refereegranskat)abstract
- Apolipoprotein E (apoE) mediates the hepatic clearance of plasma lipoproteins, facilitates cholesterol efflux from macrophages and aids neuronal lipid transport. ApoE is expressed at high levels in hepatocytes, macrophages and astrocytes. In the present study, we identify nuclear and cytosolic pools of apoE in human fibroblasts. Fibroblast apoE mRNA and protein levels were up-regulated during staurosporine-induced apoptosis and this was correlated with increased caspase-3 activity and apoptotic morphological alterations. Because the transcription of apoE and specific pro-apoptotic genes is regulated by the nuclear receptor LXR (liver X receptor) α, we analysed LXRα mRNA expression by quantitative real-time PCR and found it to be increased before apoE mRNA induction. The expression of ABCA1 (ATP-binding cassette transporter A1) mRNA, which is also regulated by LXRα, was increased in parallel with apoE mRNA, indicating that LXRα probably promotes apoE and ABCA1 transcription during apoptosis. Fibroblast apoE levels were increased under conditions of serum-starvation-induced growth arrest and hyperoxia-induced senescence. In both cases, an increased nuclear apoE level was observed, particularly in cells that accumulated lipofuscin. Nuclear apoE was translocated to the cytosol when mitotic nuclear disassembly occurred and this was associated with an increase in total cellular apoE levels. ApoE amino acid sequence analysis indicated several potential sites for phosphorylation. In vivo studies, using 32P-labelling and immunoprecipitation, revealed that fibroblast apoE can be phosphorylated. These studies reveal novel associations and potential roles for apoE in fundamental cellular processes.
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| 9. |
- Terman, Alexei, 1957-, et al.
(författare)
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Decreased apoptotic response of inclusion-cell disease fibroblasts : A consequence of lysosomal enzyme missorting?
- 2002
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Ingår i: Experimental Cell Research. - : Elsevier BV. - 0014-4827 .- 1090-2422. ; 274:1
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Tidskriftsartikel (refereegranskat)abstract
- To better understand the role of lysosomes in apoptosis, we compared the responses to apoptotic stimuli of normal fibroblasts with those of inclusion cells (I-cells), i.e., fibroblasts with impaired function of lysosomal enzymes due to their missorting and ensuing nonlysosomal localization. Although both cell types did undergo apoptosis when exposed to the lysosomotropic detergent MSDH, the redox-cycling quinone naphthazarin, or the protein kinase inhibitor staurosporine, I-cells exerted a markedly decreased response to these agonists than did normal fibroblasts. Furthermore, leupeptin and pepstatin A (inhibitors of cysteine and aspartic proteases, respectively) suppressed staurosporine-induced apoptosis of normal fibroblasts, whereas survival of I-cells was unaffected. These findings give further support for the involvement of lysosomal enzymes in apoptosis and suggest I-cells as a suitable model for studying the role of lysosomes in programmed cell death.
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| 10. |
- Öllinger, Karin, 1962-, et al.
(författare)
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Induction of apoptosis by redoxcycling quinones
- 2002
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Ingår i: Phospholipid metabolism in apoptosis. - Linköping : Linköpings universitet. - 0306467828 - 9780306479311 ; , s. 151-170
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Bokkapitel (övrigt vetenskapligt/konstnärligt)abstract
- The last few years have witnessed an explosion of both interest and knowledge about apoptosis, the process by which a cell actively commits suicide. It is now well recognised that apoptosis is essential in many aspects of normal development and is required for maintaining tissue homeostasis. The molecular mechanisms of apoptosis are presently unknown and the subject of focused research effort. It is clear that cell membrane structure and properties play an early part in the induction process. There is increasing evidence that the arrangement of polar lipids in the membrane lipid matrix is an important factor coupled with the homeostatic mechanisms responsible for preserving membrane lipid composition and asymmetry. Changes in membrane permeability are also likely to be involved, possibly as a direct consequence of disturbances in the lipid bilayer matrix. The purpose of this volume is to examine the involvement of membrane lipids in early events of apoptosis. In particular, the role of phospholipids in mitochondrial permeability, membrane lipid asymmetry, and sphingolipid and phospholipid signalling processes in early apoptotic events are reviewed by current researchers in these fields
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