| 51. |
- Sörgjerd, Karin, 1977-, et al.
(författare)
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BiP can function as a molecular shepherd that alleviates oligomer toxicity and amass amyloid
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Annan publikation (övrigt vetenskapligt/konstnärligt)abstract
- A wide range of diseases are linked to protein misfolding and aggregation inside and outside the cell. It is of utmost interest to understand how the molecular chaperone machinery of the endoplasmic reticulum (ER) handles the expression of highly amyloidogenic proteins. We explored the hypothesis that the ER located Hsp70 molecular chaperone BiP plays a crucial role in amyloid diseases and influence the misfolding process and disease progression. We used the transthyretin mutant TTR D18G associated with an unusual central nervous system amyloid disease as the model substrate because it represents the most destabilized and degraded TTR variant known. Over-expression of TTR D18G in concert with BiP showed that BiP selectively recognize the amyloidogenic mutant protein as compared to wild type in human cells and collects the mutant in stable intermediate size oligomers within the ER. Furthermore, whereas TTR D18G was found to be highly cytotoxic to neuroblastoma cells, TTR D18G preincubated with BiP was non-toxic indicating that BiP protects the cell from cytotoxicity. BiP was also found present in cerebellar amyloid deposits and co-localized with TTR in a TTR D18G patient suggesting that the complex can be found in the extracellular space. We promote a fundamental role of BiP in misfolding diseases and describe a molecular shepharding function of BiP in sequestrating amyloidogenic protein molecules in benign oligomeric states.
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| 52. |
- Sörgjerd, Karin, 1977-, et al.
(författare)
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Prefibrillar Amyloid Aggregates and Cold Shocked Tetrameric Wild Type Transthyretin are Cytotoxic
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Annan publikation (övrigt vetenskapligt/konstnärligt)abstract
- Recent studies suggest that soluble, oligomeric species, which are intermediates in the fibril formation process in amyloid disease, might be the key species in amyloid pathogenesis. Soluble oligomers of TTR were produced by kinetic sampling from a TTR fibrillation reaction (A-state TTR, pH 2, 100 mM NaCl). The reaction was terminated at different time points, and different states in the aggregation process were captured and analyzed to elucidate the oligomer properties followed by sampling for cytotoxicity using exposure towards human SH-SYY5 neuroblastoma cells. Employing ThT fluorescence, time-resolved fluorescence anisotropy of pyrenelabeled TTR, chemical cross-linking and electron microscopy we demonstrated that early formed oligomers from A-state TTR were soluble and comprised on the average 20-30 TTR monomers. Early oligomers were highly cytotoxic and induced apoptosis as indicated by the MTT assay and caspase-3 activation, whereas mature fibrils were non-toxic. We also indicate an activated unfolded protein response in cells exposed to oligomers as evidenced by an increased expression of the endoplasmic reticulum located molecular chaperone BiP. Following exposure, BiP appeared relocalized to the cytoplasm. Surprisingly, we also found that native tetrameric TTR purified and stored under cold conditions (4 °C) was highly cytotoxic. The effect could be partially restored by increasing the temperature of the protein. The molecular basis for this pathogenicity is rather unclear but likely stems from previously reported increased sensitivity towards dissociation and denaturation of TTR at low temperatures and opens the possibility that rearranged tetrameric TTR is cytotoxic towards neuroblastoma cells.
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| 53. |
- Sörgjerd, Karin, et al.
(författare)
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Prefibrillar transthyretin oligomers and cold stored native tetrameric transthyretin are cytotoxic in cell culture
- 2008
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Ingår i: Biochemical and Biophysical Research Communications - BBRC. - : Elsevier BV. - 0006-291X .- 1090-2104. ; 377:4, s. 1072-1078
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Tidskriftsartikel (refereegranskat)abstract
- Recent studies Suggest that Soluble, oligomeric species, which are intermediates in the fibril formation process in amyloid disease, might be the key species in amyloid pathogenesis. Soluble oligomers of human wild type transthyretin (TTR) were produced to elucidate oligomer properties. Employing ThT fluorescence, time-resolved fluorescence anisotropy of pyrene-labeled TTR, chemical cross-linking, and electron microscopy we demonstrated that early formed soluble oligomers (within minutes) from A-state TTR comprised on the average 20-30 TTR monomers. When administered to neuroblastoma cells these early oligomers proved highly cytotoxic and induced apoptosis after 48 h of incubation. More Mature fibrils (> 24 h of fibrillation) were non-toxic. Surprisingly, we also found that native tetrameric TTR, when purified and stored under cold conditions (4 degrees C) was highly cytotoxic. The effect Could be partially restored by increasing the temperature of the protein. The cytotoxic effects of native tetrameric TTR likely stems from a hitherto unexplored low temperature induced rearrangement of the tetramer conformation that possibly is related to the conformation of misfolded TTR in amyloigogenic oligomers.
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| 54. |
- 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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| 55. |
- Zheng, Lin, et al.
(författare)
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Macroautophagy-generated increase of lysosomal amyloid β-protein mediates oxidant-induced apoptosis of cultured neuroblastoma cells
- 2011
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Ingår i: Autophagy. - : Landes Bioscience. - 1554-8627 .- 1554-8635. ; 7:12, s. 1528-1545
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Tidskriftsartikel (refereegranskat)abstract
- Increasing evidence suggests the toxicity of intracellular amyloid β-protein (Aβ) to neurons, as well as the involvement of oxidative stress in Alzheimer disease (AD). Here we show that normobaric hyperoxia (exposure of cells to 40% oxygen for five days, and consequent activation of macroautophagy and accumulation of Aβ within lysosomes, induced apoptosis in differentiated SH-SY5Y neuroblastoma cells. Cells under hyperoxia showed: (1) increased numbers of autophagic vacuoles that contained amyloid precursor protein (APP) as well as Aβ monomers and oligomers, (2) increased reactive oxygen species production, and (3) enhanced apoptosis. Oxidant-induced apoptosis positively correlated with cellular Aβ production, being the highest in cells that were stably transfected with APP Swedish KM670/671NL double mutation. Inhibition of γ-secretase, prior and/or in parallel to hyperoxia, suggested that the increase of lysosomal Aβ resulted mainly from its autophagic uptake, but also from APP processing within autophagic vacuoles. The oxidative stress-mediated effects were prevented by macroautophagy inhibition using 3-methyladenine or ATG5 downregulation. Our results suggest that upregulation of macroautophagy and resulting lysosomal Aβ accumulation are essential for oxidant-induced apoptosis in cultured neuroblastoma cells and provide aditional support for the interactive role of oxidative stress and the lysosomal system in AD-related neurodegeneration.
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| 56. |
- Zheng, Lin, et al.
(författare)
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Oxidative stress induces macroautophagy of amyloid beta-protein and ensuing apoptosis
- 2009
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Ingår i: Free Radical Biology & Medicine. - : Elsevier BV. - 0891-5849 .- 1873-4596. ; 46:3, s. 422-429
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Tidskriftsartikel (refereegranskat)abstract
- There is increasing evidence for the toxicity of intracellular amyloid beta-protein (A beta) to neurons and the involvement of lysosomes in this process in Alzheimer disease (AD). We have recently shown that oxidative stress, a recognized determinant of AD. enhances macroautophagy and leads to intralysosomal accumulation of A beta in Cultured neuroblastoma cells. We hypothesized that oxidative stress promotes AD by stimulating macroautophagy of A that further may induce cell death by destabilizing lysosomal membranes. To investigate such possibility, we compared the effects of hyperoxia (40% ambient oxygen) in cultured HEK293 cells that were transfected with an empty vector (Vector), wild-type APP (APPwt), or Swedish mutant APP (APPswe). Exposure to hyperoxia for 5 days increased the number of cells with A beta-containing lysosomes, as well as the number of apoptotic cells, compared to normoxic conditions. The rate of apoptosis in all three cell lines demonstrated dependence on intralysosomal A beta content (Vector
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| 57. |
- 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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