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Using Laser Scanning Cytometry to Measure PPAR-Mediated Peroxisome Proliferation and β OxidationPfizer Global Research and Development, Eastern Point Road, Groton, Connecticut 06340-8014, USA Correspondence: Address correspondence to: Dr. Ingrid M. Pruimboom-Brees, Pfizer Global Research and Development, Groton Safety Sciences, Eastern Point Road MS 8274–1219, Groton, Connecticut 06340–8014, USA; e-mail: ingrid_m_pruimboom-brees{at}groton.pfizer.com Laser scanning cytometry (LSC) is a new technology that combines the properties and advantages of flow cytometry (FC) and immunohistochemistry (IHC), thus providing qualitative and quantitative information on protein expression with the additional perspective provided by cell and tissue localization. Formalin-fixed, paraffin embedded liver sections from rats exposed to a Peroxisome Proliferator Activated Receptor (PPAR) agonist were stained with antibodies against peroxisomal targeting signal-1 (PTS-1) (a highly conserved tripeptide contained within all peroxisomal enzymes), Acyl CoA oxidase (AOX) (the rate limiting enzyme of peroxisomal β oxidation), and catalase (an inducible peroxisomal antioxidant enzyme) to evaluate peroxisomal β oxidation, oxidative stress, and peroxisome proliferation. The LSC showed increased AOX, catalase, and PTS-1 expression in centrilobular hepatocytes that correlated favorably with the microscopic observation of centrilobular hepatocellular hypertrophy and with the palmitoyl CoA biochemical assay for peroxisomal β oxidation, and provided additional morphologic information about peroxisome proliferation and tissue patterns of activation. Therefore, the LSC provides qualitative and quantitative evaluation of peroxisome activity with similar sensitivity but higher throughput than the traditional biochemical methods. The additional benefits of the LSC include the direct correlation between histopathologic observations and peroxisomal alterations and the potential utilization of archived formalin-fixed tissues from a variety of organs and species.
Key Words: Peroxisome • proliferation • β-oxidation • PTS1 • Acyl CoA oxidase Abbreviations: LSC, Laser Scanning Cytometry • FC, Flow cytometry • IHC, Immunohistochemistry • PPAR, Peroxisome Proliferator Activated Receptor • PTS-1, Peroxisome Targeting Signal-1 • AOX, Acyl CoA Oxidase • MPI, Maximum Pixel Intensity •
Toxicologic Pathology, Vol. 33, No. 1,
86-91 (2005) This article has been cited by other articles:
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Abs, Change in absorbance • KLH, keyhole limpet hemocyanine • FITC, Fluorescein Isothyocyanate • H&E, Hematoxylin and eosin • OWB, Optimax Wash Buffer • DAPI, 4' 6-diamino-2-phenylindole
