Abstract
Zebrafish, a diurnal vertebrate characterized by gradual senescence, is an excellent model for studying age-dependent diseases, such as neurodegenerative diseases. Cerebral amyloid angiopathy (CAA) caused by amyloid β (Aβ) deposition around brain microvessels is a human neurovascular degenerative disease that is characterized by an early onset of recurrent stroke episodes, vascular brain degenerative changes, and moderate to severe clinical presentations. Recently, by using the zebrafish model, we investigated whether Aβ peptides cause endothelial cells to enter senescence at an early stage of vascular development. During early embryonic zebrafish development, the presence of senescence-associated biomarkers, such as β-galactosidase and the cyclin-dependent kinase inhibitor p21, has been shown to be predictive of the premature aging phenotype. By measuring β-galactosidase activity and p21 expression in whole-mount zebrafish embryos exposed to Aβ, we demonstrated that these oxidative peptides promote vascular senescence at an early stage of development, a harbinger of vascular clinical symptoms in adult. This chapter describes the methods for studying cell senescence in zebrafish, detailing protocols for β-gal activity and the in situ p21 hybridization in whole-mount zebrafish embryos.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Revesz T, Holton JL, Lashley T, Plant G, Frangione B, Rostagno A, Ghiso J (2009) Genetics and molecular pathogenesis of sporadic and hereditary cerebral amyloid angiopathies. Acta Neuropathol 118:115–130
Wisniewski T, Ghiso J, Frangione B (1991) Peptides homologous to the amyloid protein of Alzheimer’s disease containing a glutamine for glutamic acid substitution have accelerated amyloid fibril formation. Biochem Biophys Res Commun 179:1247–1254
Solito R, Corti F, Fossati S, Mezhericher E, Donnini S, Ghiso J, Giachetti A, Rostagno A, Ziche M (2009) Dutch and Arctic mutant peptides of beta amyloid(1-40) differentially affect the FGF-2 pathway in brain endothelium. Exp Cell Res 315:385–395
Maat-Schieman M, Roos R, van Duinen S (2005) Hereditary cerebral hemorrhage with amyloidosis-Dutch type. Neuropathology (J Japan Soc Neuropathol) 25:288–297
Donnini S, Solito R, Cetti E, Corti F, Giachetti A, Carra S, Beltrame M, Cotelli F, Ziche M (2010) Abeta peptides accelerate the senescence of endothelial cells in vitro and in vivo, impairing angiogenesis. FASEB J (Fed Am Soc Exp Biol) 24:2385–2395
Kishi S, Uchiyama J, Baughman AM, Goto T, Lin MC, Tsai SB (2003) The zebrafish as a vertebrate model of functional aging and very gradual senescence. Exp Gerontol 38:777–786
Donnini S, Cantara S, Morbidelli L, Giachetti A, Ziche M (2006) FGF-2 overexpression opposes the beta amyloid toxic injuries to the vascular endothelium. Cell Death Differ 13:1088–1096
Poleo G, Brown CW, Laforest L, Akimenko MA (2001) Cell proliferation and movement during early fin regeneration in zebrafish. Dev Dyn (Am Assoc Anat) 221:380–390
Jung T, Bader N, Grune T (2007) Lipofuscin: formation, distribution, and metabolic consequences. Ann N Y Acad Sci 1119:97–111
Kishi S, Bayliss PE, Uchiyama J, Koshimizu E, Qi J, Nanjappa P, Imamura S, Islam A, Neuberg D, Amsterdam A, Roberts TM (2008) The identification of zebrafish mutants showing alterations in senescence-associated biomarkers. PLoS Genet 4:e1000152
Jowett T, Lettice L (1994) Whole-mount in situ hybridizations on zebrafish embryos using a mixture of digoxigenin- and fluorescein-labelled probes. Trends Genetics TIG 10:73–74
Santoriello C, Deflorian G, Pezzimenti F, Kawakami K, Lanfrancone L, d’Adda di Fagagna F, Mione M (2009) Expression of H-RASV12 in a zebrafish model of Costello syndrome causes cellular senescence in adult proliferating cells. Dis Model Mech 2:56–67
Lu WJ, Abrams JM (2006) Lessons from p53 in non-mammalian models. Cell Death Differ 13:909–912
Kimmel CB, Ballard WW, Kimmel SR, Ullmann B, Schilling TF (1995) Stages of embryonic development of the zebrafish. Dev Dyn (Am Assoc Anat) 203:253–310
Furthauer M, Lin W, Ang SL, Thisse B, Thisse C (2002) Sef is a feedback-induced antagonist of Ras/MAPK-mediated FGF signalling. Nat Cell Biol 4:170–174
Furthauer M, Reifers F, Brand M, Thisse B, Thisse C (2001) Sprouty4 acts in vivo as a feedback-induced antagonist of FGF signaling in zebrafish. Development 128:2175–2186
Furthauer M, Thisse C, Thisse B (1997) A role for FGF-8 in the dorsoventral patterning of the zebrafish gastrula. Development 124:4253–4264
Tsang M, Friesel R, Kudoh T, Dawid IB (2002) Identification of Sef, a novel modulator of FGF signalling. Nat Cell Biol 4:165–169
Acknowledgments
This work was supported by Ministero dell’Istruzione, dell’Università e della Ricerca (MIUR), PRIN project 2008.
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2013 Springer Science+Busincess Media, LLC
About this protocol
Cite this protocol
Donnini, S., Giachetti, A., Ziche, M. (2013). Assessing Vascular Senescence in Zebrafish. In: Galluzzi, L., Vitale, I., Kepp, O., Kroemer, G. (eds) Cell Senescence. Methods in Molecular Biology, vol 965. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-62703-239-1_34
Download citation
DOI: https://doi.org/10.1007/978-1-62703-239-1_34
Published:
Publisher Name: Humana Press, Totowa, NJ
Print ISBN: 978-1-62703-238-4
Online ISBN: 978-1-62703-239-1
eBook Packages: Springer Protocols