Using Ambystoma mexicanum (Mexican axolotl) embryos, chemical genetics, and microarray analysis to identify signaling pathways associated with tissue regeneration

Larissa V. Ponomareva, Antony Athippozhy, Jon S. Thorson, S. Randal Voss

Research output: Contribution to journalArticlepeer-review

31 Scopus citations

Abstract

Amphibian vertebrates are important models in regenerative biology because they present exceptional regenerative capabilities throughout life. However, it takes considerable effort to rear amphibians to juvenile and adult stages for regeneration studies, and the relatively large sizes that frogs and salamanders achieve during development make them difficult to use in chemical screens. Here, we introduce a new tail regeneration model using late stage Mexican axolotl embryos. We show that axolotl embryos completely regenerate amputated tails in 7 days before they exhaust their yolk supply and begin to feed. Further, we show that axolotl embryos can be efficiently reared in microtiter plates to achieve moderate throughput screening of soluble chemicals to investigate toxicity and identify molecules that alter regenerative outcome. As proof of principle, we identified integration 1 / wingless (Wnt), transforming growth factor beta (Tgf-β), and fibroblast growth factor (Fgf) pathway antagonists that completely block tail regeneration and additional chemicals that significantly affected tail outgrowth. Furthermore, we used microarray analysis to show that inhibition of Wnt signaling broadly affects transcription of genes associated with Wnt, Fgf, Tgf-β, epidermal growth factor (Egf), Notch, nerve growth factor (Ngf), homeotic gene (Hox), rat sarcoma/mitogen-activated protein kinase (Ras/Mapk), myelocytomatosis viral oncogene (Myc), tumor protein 53 (p53), and retinoic acid (RA) pathways. Punctuated changes in the expression of genes known to regulate vertebrate development were observed; this suggests the tail regeneration transcriptional program is hierarchically structured and temporally ordered. Our study establishes the axolotl as a chemical screening model to investigate signaling pathways associated with tissue regeneration.

Original languageEnglish
Pages (from-to)128-135
Number of pages8
JournalComparative Biochemistry and Physiology Part - C: Toxicology and Pharmacology
Volume178
DOIs
StatePublished - Dec 1 2015

Bibliographical note

Funding Information:
This research was funded by the Army Research Office ( 56157-LS-MUR ), the National Institutes of Health ( R24OD010435 ), the National Center for Advancing Translational Sciences ( UL1TR000117 ) and the National Science Foundation ( DBI-0951484 ) through its support of the Ambystoma Genetic Stock Center at the University of Kentucky. The contents of this paper are solely the responsibility of the authors and do not necessarily represent the official views of the ARO, NIH, or NSF.

Publisher Copyright:
© 2015 Elsevier Inc. All rights reserved.

Keywords

  • Axolotl
  • C59
  • Chemical screening
  • Regeneration
  • Wnt

ASJC Scopus subject areas

  • Biochemistry
  • Physiology
  • Toxicology
  • Cell Biology
  • Health, Toxicology and Mutagenesis

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