Does the cytotoxic effect of transient amyloid oligomers from common equine lysozyme in vitro imply innate amyloid toxicity?

被引:60
作者
Malisauskas, M
Ostman, J
Darinskas, A
Zamotin, V
Liutkevicius, E
Lundgren, E
Morozova-Roche, LA [1 ]
机构
[1] Umea Univ, Dept Med Biochem & Biophys, S-90187 Umea, Sweden
[2] Umea Univ, Dept Cell & Mol Biol, S-90187 Umea, Sweden
[3] Vilnius Univ, Inst Immunol, Pharmacol Lab, LT-08409 Vilnius, Lithuania
关键词
D O I
10.1074/jbc.M407273200
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
In amyloid diseases, it is not evident which protein aggregates induce cell death via specific molecular mechanisms and which cause damage because of their mass accumulation and mechanical properties. We showed that equine lysozyme assembles into soluble amyloid oligomers and protofilaments at pH 2.0 and 4.5, 57 degreesC. They bind thioflavin-T and Congo red similar to common amyloid structures, and their morphology was monitored by atomic force microscopy. Molecular volume evaluation from microscopic measurements allowed us to identify distinct types of oligomers, ranging from tetramer to octamer and 20-mer. Monomeric lysozyme and protofilaments are not cytotoxic, whereas the oligomers induce cell death in primary neuronal cells, primary fibroblasts, and the neuroblastoma IMR-32 cell line. Cytotoxicity was accessed by ethidium bromide staining, MTT reduction, and TUNEL assays. Primary cultures were more susceptible to the toxic effect induced by soluble amyloid oligomers than the neuroblastoma cell line. The cytotoxicity correlates with the size of oligomers; the sample incubated at pH 4.5 and containing larger oligomers, including 20-mer, appears to be more cytotoxic than the lysozyme sample kept at pH 2.0, in which only tetramers and octamers were found. Soluble amyloid oligomers may assemble into rings; however, there was no correlation between the quantity of rings in the sample and its toxicity. The cytotoxicity of transient oligomeric species of the ubiquitous protein lysozyme indicates that this is an intrinsic feature of protein amyloid aggregation, and therefore soluble amyloid oligomers can be used as a primary therapeutic target and marker of amyloid disease.
引用
收藏
页码:6269 / 6275
页数:7
相关论文
共 34 条
[1]  
Alberts B., 1994, MOL BIOL CELL
[2]   Only amyloidogenic intermediates of transthyretin induce apoptosis [J].
Andersson, K ;
Olofsson, A ;
Nielsen, EH ;
Svehag, SE ;
Lundgren, E .
BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS, 2002, 294 (02) :309-314
[3]   Cytotoxic effects of triphenylbismuth on rat thymocytes: Comparisons with bismuth chloride and triphenyltin chloride [J].
Arata, T ;
Oyama, Y ;
Tabaru, K ;
Satoh, M ;
Hayashi, H ;
Ishida, S ;
Okano, Y .
ENVIRONMENTAL TOXICOLOGY, 2002, 17 (05) :472-477
[4]   Inherent toxicity of aggregates implies a common mechanism for protein misfolding diseases [J].
Bucciantini, M ;
Giannoni, E ;
Chiti, F ;
Baroni, F ;
Formigli, L ;
Zurdo, JS ;
Taddei, N ;
Ramponi, G ;
Dobson, CM ;
Stefani, M .
NATURE, 2002, 416 (6880) :507-511
[5]   Self-assembly of Aβ1-42 into globular neurotoxins [J].
Chromy, BA ;
Nowak, RJ ;
Lambert, MP ;
Viola, KL ;
Chang, L ;
Velasco, PT ;
Jones, BW ;
Fernandez, SJ ;
Lacor, PN ;
Horowitz, P ;
Finch, CE ;
Krafft, GA ;
Klein, WL .
BIOCHEMISTRY, 2003, 42 (44) :12749-12760
[6]   Method for measuring neurotoxicity of aggregating polypeptides with the MTT assay on differentiated neuroblastoma cells [J].
Datki, Z ;
Juhász, A ;
Gálfi, M ;
Soós, K ;
Papp, R ;
Zádori, D ;
Penke, B .
BRAIN RESEARCH BULLETIN, 2003, 62 (03) :223-229
[7]   Annular α-synuclein protofibrils are produced when spherical protofibrils are incubated in solution or bound to brain-derived membranes [J].
Ding, TT ;
Lee, SJ ;
Rochet, JC ;
Lansbury, PT .
BIOCHEMISTRY, 2002, 41 (32) :10209-10217
[8]   Protein folding and misfolding [J].
Dobson, CM .
NATURE, 2003, 426 (6968) :884-890
[9]   THE TOXIC EFFECTS OF FORMATE IN DISSOCIATED PRIMARY MOUSE NEURAL CELL-CULTURES [J].
DORMAN, DC ;
BOLON, B ;
MORGAN, KT .
TOXICOLOGY AND APPLIED PHARMACOLOGY, 1993, 122 (02) :265-272
[10]   Syncollin homo-oligomers associate with lipid bilayers in the form of doughnut-shaped structures [J].
Geisse, NA ;
Wäsle, B ;
Saslowsky, DE ;
Henderson, RM ;
Edwardson, JM .
JOURNAL OF MEMBRANE BIOLOGY, 2002, 189 (02) :83-92