Development of a marine fish model for studying in vivo molecular responses in ecotoxicology

R. Y. C. KONG, J. P. GIESY, Shiu Sun Rudolf WU, E. X. H. CHEN, M. W. L. CHIANG, P. L. LIM, B. B. H. YUEN, B. W. P. YIP, H. O. L. MOK, D. W. T. AU

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Abstract

A protocol for fixation and processing of whole adult marine medaka (Oryzias melastigma) was developed in parallel with in situ hybridization (ISH) and immunohistochemistry (IHC) for molecular analysis of in vivo gene and protein responses in fish. Over 200 serial sagittal sections (5 μm) can be produced from a single adult medaka to facilitate simultaneous localization and quantification of gene-specific mRNAs and proteins in different tissues and subcellular compartments of a single fish. Stereological analysis (as measured by volume density, Vᵥ) was used to quantify ISH and IHC signals on tissue sections. Using the telomerase reverse transcriptase (omTERT) gene, omTERT and proliferating cell nuclear antigen (PCNA) proteins as examples, we demonstrated that it is possible to localize, quantify and correlate their tissue expression profiles in a whole fish system. Using chronic hypoxia (1.8 ± 0.2 mg O₂ L⁻¹ for 3 months) as an environmental stressor, we were able to identify significant alterations in levels of omTERT mRNA, omTERT protein, PCNA (cell proliferation marker) and TUNEL (apoptosis) in livers of hypoxic O. melastigma (p < 0.05). Overall, the results suggest that O. melastigma can serve as a model marine fish for assessing multiple in vivo molecular responses to stresses in the marine environment. Copyright © 2007 Elsevier B.V. All rights reserved.

Original languageEnglish
Pages (from-to)131-141
JournalAquatic Toxicology
Volume86
Issue number2
DOIs
Publication statusPublished - 31 Jan 2008

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Ecotoxicology
ecotoxicology
Oryzias
marine fish
Fishes
proliferating cell nuclear antigen
protein
Proliferating Cell Nuclear Antigen
fish
antigen
in situ hybridization
In Situ Hybridization
immunohistochemistry
gene
proteins
Immunohistochemistry
telomerase
Messenger RNA
Proteins
genes

Citation

Kong, R. Y. C., Giesy, J. P., Wu, R. S. S., Chen, E. X. H., Chiang, M. W. L., Lim, P. L., . . . Au, D. W. T. (2008). Development of a marine fish model for studying in vivo molecular responses in ecotoxicology. Aquatic Toxicology, 86(2), 131-141. doi: 10.1016/j.aquatox.2007.10.011

Keywords

  • Hypoxic stress
  • Histology
  • Immunohistochemistry
  • ISH
  • Telomerase