Alzheimer Disease: Ankarcrona M

Cowburn RF, Popescu BO, Ankarcrona M, Dehvari N, Cedazo-Minguez A. · Karolinska Institutet, Department of Neurobiology, Care Sciences and Society, KI-Alzheimer's Disease Research Center, Novum, plan 5, S-141 57 Huddinge, Sweden. · Physiol Behav. · Pubmed #17568632 No free full text.

Abstract: Presenilin proteins, mutated forms of which cause early onset familial Alzheimer's disease, are capable of modulating various cell signal transduction pathways, the most extensively studied of which has been intracellular calcium signalling. Disease causing presenilin mutations can potentiate inositol(1,4,5)trisphosphate (InsP3) mediated endoplasmic reticulum release due to calcium overload in this organelle, as well as attenuate capacitative calcium entry. Our own studies have shown a novel function for presenilins that involves regulation of acetylcholine muscarinic receptor-stimulated phospholipase C upstream of InsP3 regulated calcium release. This article reviews the mechanisms by which presenilins modulate intracellular calcium signalling and the role that deregulated calcium homeostasis could play in the pathogenesis of Alzheimer's disease.

Ankarcrona M, Winblad B. · Karolinska Institutet, Neurotec, Section of Experimental Geriatrics, Novum, Huddinge, Sweden. · Int J Geriatr Psychiatry. · Pubmed #15660410 No free full text.

Abstract: Alzheimer's disease (AD) affects millions of people worldwide and the number of AD cases will increase with increased life expectancy. Today there is no cure for this devastating and always lethal disease and therefore it is of great interest for patients, relatives and societies to find new drugs that can hinder the disease process. During the progression of AD a substantial amount of neurons degenerate in the brain. The mechanisms of cell death involved in AD have not been fully elucidated. However, there are several reports showing that neurons die partly by apoptosis in the AD brain. Drugs blocking apoptosis could therefore be potentially useful for early prevention of neuronal cell death. Biomarkers for apoptosis should be important tools in the evaluation of drug effects and in the diagnostics of AD. Here we review the current knowledge in the field and discuss potential biomarkers for apoptosis in AD.

Popescu BO, Ankarcrona M. · Karolinska Institutet, Neurotec, Section of Experimental Geriatrics, Novum, Huddinge, Sweden. · J Alzheimers Dis. · Pubmed #15096695 No free full text.

Abstract: Presenilins are often mutated in familial forms of Alzheimer's disease (AD). Such mutations sensitize cells in culture to different apoptotic stimuli eg. staurosporine, calcium ionophore, growth factor withdrawal. The altered responses to apoptotic stimuli in cells carrying presenilin mutations include increased intracellular calcium concentrations and enhanced production of reactive oxygen species. Presenilin mutations also result in increased production of amyloid beta (Abeta) indicating that presenilins participate in the cleavage of amyloid beta-protein precursor (AbetaPP). In fact, presenilin is part of the gamma-secretase complex which together with beta-secretase cleaves AbetaPP and produce Abeta, later forming the senile plaques typical for AD pathology. Here we review the current knowledge about the mechanisms of cell death in AD with focus on the role of presenilin and presenilin mutations in apoptosis. It appears that presenilin and its different fragments, generated after proteolytic cleavage, have a regulatory role in apoptosis. In addition, different studies show that the cellular levels of presenilin are controlled by proteasomal degradation both under normal and stress conditions.

Hansson Petersen CA, Alikhani N, Behbahani H, Wiehager B, Pavlov PF, Alafuzoff I, Leinonen V, Ito A, Winblad B, Glaser E, Ankarcrona M. · Karolinska Institutet Dainippon Sumitomo Pharma Alzheimer Center, NVS, Novum, 141 57 Huddinge, Sweden. · Proc Natl Acad Sci U S A. · Pubmed #18757748 links to  free full text

Abstract: The amyloid beta-peptide (Abeta) has been suggested to exert its toxicity intracellularly. Mitochondrial functions can be negatively affected by Abeta and accumulation of Abeta has been detected in mitochondria. Because Abeta is not likely to be produced locally in mitochondria, we decided to investigate the mechanisms for mitochondrial Abeta uptake. Our results from rat mitochondria show that Abeta is transported into mitochondria via the translocase of the outer membrane (TOM) machinery. The import was insensitive to valinomycin, indicating that it is independent of the mitochondrial membrane potential. Subfractionation studies following the import experiments revealed Abeta association with the inner membrane fraction, and immunoelectron microscopy after import showed localization of Abeta to mitochondrial cristae. A similar distribution pattern of Abeta in mitochondria was shown by immunoelectron microscopy in human cortical brain biopsies obtained from living subjects with normal pressure hydrocephalus. Thus, we present a unique import mechanism for Abeta in mitochondria and demonstrate both in vitro and in vivo that Abeta is located to the mitochondrial cristae. Importantly, we also show that extracellulary applied Abeta can be internalized by human neuroblastoma cells and can colocalize with mitochondrial markers. Together, these results provide further insight into the mitochondrial uptake of Abeta, a peptide considered to be of major significance in Alzheimer's disease.

Oprica M, Hjorth E, Spulber S, Popescu BO, Ankarcrona M, Winblad B, Schultzberg M. · Department of Neurobiology, Care Sciences and Society, Division of Neurodegeneration & Neuroinflammation, Karolinska Institutet, Huddinge, SE-14186 Stockholm, Sweden. · J Cell Mol Med. · Pubmed #17760842 No free full text.

Abstract: Inflammation is associated with both acute and chronic neurological disorders, including stroke and Alzheimer's disease (AD). Cytokines such as interleukin (IL)-1 have several activities in the brain both under physiological and pathophysiological conditions. The objective of this study was to evaluate consequences of the central blockade of IL-1 transmission in a previously developed transgenic mouse strain with brain-directed overexpression of human soluble IL-1 receptor antagonist (Tg hsIL-1ra). Effects on brain morphology and brain levels of the AD-related proteins beta-amyloid precursor protein (APP) and presenilin 1(PS1), as well as the levels of IL-1beta, IL-6 and tumour necrosis factor-alpha (TNF-alpha) were analysed in homozygotic and heterozygotic mice and wild type (WT) controls, of both genders and of young (30-40 days) and adult (13-14 months) age. A marked reduction in brain volume was observed in transgenic mice as determined by volumetry. Western blot analysis showed higher levels of APP, but lower levels of PS1, in adult animals than in young ones. In the cerebellum, heterozygotic (Tg hsIL-1ra(+/-)) mice had lower levels of APP and PS1 than WT mice. With one exception, there were no genotypic differences in the levels of IL-1beta, IL-6 and TNF-alpha. The cytokine levels were generally higher in adult than in young mice. In conclusion, the chronic blockade of IL-1 signalling in the brain was associated with an atrophic phenotype of the brain, and with modified levels of APP and PS1. Brain-directed overexpression of hsIL-1ra was not followed by major compensatory changes in the levels of pro-inflammatory cytokines.

Selivanova A, Winblad B, Farmery MR, Dantuma NP, Ankarcrona M. · Department of Neurobiology, Caring Sciences and Society (NVS), KI Alzheimer Disease Research Center, Karolinska Institutet, Novum 5th floor, S-141 57 Stockholm, Sweden. · Biochem Biophys Res Commun. · Pubmed #16999935 No free full text.

Abstract: Sequential cleavage of the amyloid precursor protein (APP) by beta- and gamma-secretases results in the production of beta-amyloid peptide, which is a key determinant in Alzheimer's disease. Since several putative locations for gamma-secretase cleavage have been identified along the secretory pathway, trafficking of APP may be of importance for beta-amyloid peptide production. Here we have studied the role of retrograde transport in APP processing. We found that APP interacts with the beta subunit of the coatomer protein I (COPI) complex, which is involved in retrograde transport. In line with a role of retrograde trafficking in APP transport, inhibition of COPI-dependent transport altered APP trafficking, decreased APP cell surface expression, and coincided with a profound reduction in gamma-secretase cleavage. These results suggest that COPI-dependent retrograde transport is important for APP processing and influences production of beta-amyloid peptide.

Behbahani H, Shabalina IG, Wiehager B, Concha H, Hultenby K, Petrovic N, Nedergaard J, Winblad B, Cowburn RF, Ankarcrona M. · Department of Neurobiology, Karolinska Institutet Dainippon Sumitomo Pharmaceuticals Alzheimer Center, Caring Sciences and Society, Novum, Huddinge, Sweden. · J Neurosci Res. · Pubmed #16883555 No free full text.

Abstract: Increasing evidence indicates that mitochondrial alterations contribute to the neuronal death in Alzheimer's disease (AD). Presenilin 1 (PS1) and Presenilin 2 (PS2) mutations have been shown to sensitize cells to apoptosis by mechanisms suggested to involve impaired mitochondrial function. We have previously detected active gamma-secretase complexes in mitochondria. We investigated the impact of PS/gamma-secretase on mitochondrial function using mouse embryonal fibroblasts derived from wild-type, PS1-/-, PS2-/- and PS double knock-out (PSKO) embryos. Measurements of mitochondrial membrane potential (DeltaPsim) showed a higher percentage of fully functional mitochondria in PS1-/- and PSwt as compared to PS2-/- and PSKO cells. This result was evident both in whole cell preparations and in isolated mitochondria. Interestingly, pre-treatment of isolated mitochondria with the gamma-secretase inhibitor L-685,458 resulted in a decreased population of mitochondria with high DeltaPsim in PSwt and PS1-/- cells, indicating that PS2/gamma-secretase activity can modify DeltaPsim. PS2-/- cells showed a significantly lower basal respiratory rate as compared to other cell lines. However, all cell lines demonstrated competent bioenergetic function. These data point toward a specific role of PS2/gamma-secretase activity for proper mitochondrial function and indicate interplay between PS1 and PS2 in mitochondrial functionality.

Falkevall A, Alikhani N, Bhushan S, Pavlov PF, Busch K, Johnson KA, Eneqvist T, Tjernberg L, Ankarcrona M, Glaser E. · Department of Biochemistry and Biophysics, Stockholm University SE-106 91 Stockholm, Sweden. · J Biol Chem. · Pubmed #16849325 links to  free full text

Abstract: Recently we have identified the novel mitochondrial peptidase responsible for degrading presequences and other short unstructured peptides in mitochondria, the presequence peptidase, which we named PreP peptidasome. In the present study we have identified and characterized the human PreP homologue, hPreP, in brain mitochondria, and we show its capacity to degrade the amyloid beta-protein (Abeta). PreP belongs to the pitrilysin oligopeptidase family M16C containing an inverted zinc-binding motif. We show that hPreP is localized to the mitochondrial matrix. In situ immuno-inactivation studies in human brain mitochondria using anti-hPreP antibodies showed complete inhibition of proteolytic activity against Abeta. We have cloned, overexpressed, and purified recombinant hPreP and its mutant with catalytic base Glu(78) in the inverted zinc-binding motif replaced by Gln. In vitro studies using recombinant hPreP and liquid chromatography nanospray tandem mass spectrometry revealed novel cleavage specificities against Abeta-(1-42), Abeta-(1-40), and Abeta Arctic, a protein that causes increased protofibril formation an early onset familial variant of Alzheimer disease. In contrast to insulin degrading enzyme, which is a functional analogue of hPreP, hPreP does not degrade insulin but does degrade insulin B-chain. Molecular modeling of hPreP based on the crystal structure at 2.1 A resolution of AtPreP allowed us to identify Cys(90) and Cys(527) that form disulfide bridges under oxidized conditions and might be involved in redox regulation of the enzyme. Degradation of the mitochondrial Abeta by hPreP may potentially be of importance in the pathology of Alzheimer disease.

Hansson CA, Frykman S, Farmery MR, Tjernberg LO, Nilsberth C, Pursglove SE, Ito A, Winblad B, Cowburn RF, Thyberg J, Ankarcrona M. · Karolinska Institutet and Sumitomo Pharmaceuticals Alzheimer Center (KASPAC), Neurotec, Novum, SE-141 57 Huddinge, Sweden. · J Biol Chem. · Pubmed #15456764 links to  free full text

Abstract: Mitochondria are central in the regulation of cell death. Apart from providing the cell with ATP, mitochondria also harbor several death factors that are released upon apoptotic stimuli. Alterations in mitochondrial functions, increased oxidative stress, and neurons dying by apoptosis have been detected in Alzheimer's disease patients. These findings suggest that mitochondria may trigger the abnormal onset of neuronal cell death in Alzheimer's disease. We previously reported that presenilin 1 (PS1), which is often mutated in familial forms of Alzheimer's disease, is located in mitochondria and hypothesized that presenilin mutations may sensitize cells to apoptotic stimuli at the mitochondrial level. Presenilin forms an active gamma-secretase complex together with Nicastrin (NCT), APH-1, and PEN-2, which among other substrates cleaves the beta-amyloid precursor protein (beta-APP) generating the amyloid beta-peptide and the beta-APP intracellular domain. Here we have identified dual targeting sequences (for endoplasmic reticulum and mitochondria) in NCT and showed expression of NCT in mitochondria by immunoelectron microscopy. We also showed that NCT together with APH-1, PEN-2, and PS1 form a high molecular weight complex located in mitochondria. gamma-secretase activity in isolated mitochondria was demonstrated using C83 (alpha-secretase-cleaved C-terminal 83-residue beta-APP fragment from BD8 cells lacking presenilin and thus gamma-secretase activity) or recombinant C100-Flag (C-terminal 100-residue beta-APP fragment) as substrates. Both systems generated an APP intracellular domain, and the activity was inhibited by the gamma-secretase inhibitors l-685,458 or Compound E. This novel localization of NCT, PS1, APH-1, and PEN-2 expands the role and importance of gamma-secretase activity to mitochondria.

Popescu BO, Cedazo-Minguez A, Popescu LM, Winblad B, Cowburn RF, Ankarcrona M. · Karolinska Institute, NEUROTEC, Division of Geriatric Medicine, KFC, NOVUM, Huddinge, Sweden. · J Neurosci Res. · Pubmed #11599009 No free full text.

Abstract: Presenilins (PSs) are mutated in a majority of familial Alzheimer disease (FAD) cases. Mutated PSs may cause FAD by a number of pro-apoptotic mechanisms, or by regulating gamma-secretase activity, a protease involved in beta-amyloid precursor protein processing to the neurotoxic beta-amyloid peptide. Besides their normal endoproteolytic processing, PSs are substrates for caspases, being cleaved to alternative N-terminal and C-terminal fragments. So far little is known about the role of PSs cleavage in the apoptotic machinery. Here, we used SH-SY5Y neuroblastoma cells stably transfected with wild-type or exon 9 deleted presenilin 1 (PS1) in a time-course study after the exposure to the calcium ionophore A23187. During and after exposure to A 23187, intracellular calcium levels were higher in exon 9 deleted PS1 cells as compared with non-transfected and wild-type PS1 transfected cells. Cell death and the enrichment of apoptotic cells after A23187 exposure were increased by overexpression of exon 9 deleted PS1 as compared with the control cell lines. Wild-type PS1 cells were compared with exon 9 deleted PS1 cells and the temporal relationship between PS1 and other caspase substrates cleavages was analyzed. Exon 9 deleted PS1 cells exhibited a higher caspase-3 activation and a greater cleavage of PS1 and poly(ADP-ribose) polymerase (PARP) compared with wild-type PS1 cells. Exon 9 deleted PS1 cleavage occurred earlier than other caspase substrate cleavages (i.e., PARP and gelsolin), simultaneous with minimum detectable caspase-3 activation. Therefore, alternative cleavage of PS1 may play an important role for the regulation of the proteolytic cascade activated during apoptosis.

Tanii H, Ankarcrona M, Flood F, Nilsberth C, Mehta ND, Perez-Tur J, Winblad B, Benedikz E, Cowburn RF. · Karolinska Institutet, Department of Geriatric Medicine, Huddinge, Sweden. · Neuroscience. · Pubmed #10658639 No free full text.

Abstract: Mutations in the presenilin-1 (PS1) and presenilin-2 (PS2) genes account for the majority of early-onset familial Alzheimer's disease cases. Recent studies suggest that presenilin gene mutations predispose cells to apoptosis by mechanisms involving altered calcium homeostasis and oxidative damage. In the present study, we determined whether PS1 mutations also sensitize cells to hyperosmotic stress-induced apoptosis. For this, we established SH-SY5Y neuroblastoma cell lines stably transfected with wild-type PS1 or either the PS1 exon 9 deletion (deltaE9) or PS1 L250S mutants. Cultured cells were exposed to an overnight (17 h) serum deprivation, followed by a 30 min treatment with either 20 mM glucose, 10 nM insulin-like growth factor-1 or 20 mM glucose + 10 nM insulin-like growth factor-1. Cells were then cultured for a further 3, 6 or 24 h and stained for apoptotic condensed nuclei using propidium iodide. Confirmation that cells were undergoing an active apoptotic process was achieved by labelling of DNA strand breaks using the terminal dUTP nick end labelling (TUNEL) technique. We also determined cell viability using 3-(4,5-dimethyl-thiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) reduction. Propidium iodide staining revealed that all cell lines and controls showed an increased number of apoptotic cells appearing with condensed nuclei at 24 h compared with 6 h and 3 h. High glucose-induced hyperosmotic stress resulted in significantly more apoptotic cells in the PS1 deltaE9 and PS1 L250S mutation cell lines at 24 h, compared with the wild-type PS1 lines (P < 0.001, ANOVA for both comparisons). Mean values (+/-S.D.) for the percentage number of apoptotic cells at 24 h following high glucose treatment were 16.1 +/- 3.5%, 26.7 +/- 5.5% and 31.0 +/- 5.7% for the wild-type PS1, PS1 deltaE9 and PS1 L250S lines, respectively. The pro-apoptotic effects of high glucose treatment were reversed by 10 nM insulin-like growth factor-1, although to a lesser extent in the mutation cell lines (5.8 +/- 2.4%, 15.2 +/- 7.3% and 13.2 +/- 2.0% for the wild-type PS1, PS1 deltaE9 (P < 0.01 for comparison with wild-type PS1) and PS1 L250S (P < 0.01 for comparison with wild-type PS1) transfected lines, respectively. TUNEL labelling of cells at 24 h following treatment gave essentially the same results pattern as obtained using propidium iodide. The percentage number of apoptotic cells with DNA strand breaks (means +/- S.D.) following high glucose treatment was 15.4 +/- 2.6% for the wild-type PS1, 26.8 +/- 3.2% for the PS1 deltaE9 (P < 0.001 for comparison with wild-type PS1) and 29.7 +/- 6.1% for the PS1 L250S transfected lines (P < 0.001 for comparison with wild-type PS1). The PS1 deltaE9 and PS1 L250S transfected lines also showed a higher number of apoptotic cells with DNA strand breaks at 24 h following high glucose plus insulin-like growth factor-1 treatment (11.4 +/- 2.0% and 14.3 +/- 2.8%, respectively), compared with values for the wild-type PS1 lines (8.5 +/- 2.4%). These differences were significant (P < 0.01) for the comparison of wild-type PS1 and PS1 L250S, but not PS1 deltaE9 lines. The mutation-related increases in number of apoptotic cells at 24 h following high glucose treatment were not accompanied by significant differences in cell viability at this time-point. Our results indicate that PS1 mutations predispose to hyperosmotic stress-induced apoptosis and that the anti-apoptotic effects of insulin-like growth factor-1 are compromised by these mutations. Perturbations of insulin-like growth factor-1 signalling may be involved in PS1 mutation-related apoptotic neuronal cell death in Alzheimer's disease.