Protein oxidation in the brain in Alzheimer's disease
Section snippets
Tissue samples
For protein carbonyl immunohistochemical analysis, specimens of the hippocampus and parahippocampal gyrus (HPG), superior and middle temporal gyri (SMT), and cerebellum from six AD patients (mean age 80.8±5.0 years, mean post mortem interval 3.0±0.3 h) and six age-matched control (mean age 78.8±4.8 years, mean post mortem interval 2.8±0.2 h) subjects were fixed in Methacarn (methanol:chloroform:acetic acid, 60:30:10) at 4°C for 24 h. Tissue was dehydrated through ascending ethanol solutions and
Results
Anti-DNP immunostaining of the AD brain sections showed increased protein carbonyl immunoreactivity in the HPG and SMT compared with the cerebellum. Severely affected brain regions in AD exhibited increased staining intensity compared with controls (Fig. 1).
Intense anti-DNP immunoreactivity localized in neuronal cell bodies was consistently found in HPG and SMT sections from AD patients. In control sections from the same brain regions, neurons with excessively oxidized proteins in the cytoplasm
Discussion
This study used immunohistochemical and 2D immunoblotting analyses together for the first time to study protein carbonyl formation in the brain of AD patients. We observed that in the severely affected regions of the AD brain, increased protein carbonyl immunoreactivity occurred in the cell bodies of neurons without visual pathomorphological changes and neurons with neurofibrillary abnormalities. Our results of the immunohistochemical analysis of protein carbonyls in brain regions severely
Conclusions
Immunohistochemical detection of oxidative damage markers demonstrates their preferential localization in neurons versus glia39., 43. in AD. In this study, we did not identify glial-specific GFAP as carbonyl-containing protein in AD or control brain extracts, which is consistent with the suggestion that glia are less sensitive to oxidative stress than neurons. Nevertheless, our results cannot imply that glial cells in the AD brain completely escape protein oxidation. Individual brain proteins,
Acknowledgements
This work was supported by NIH grants 5P01 AG05119, 5P50 AG05144, and grants from the Abercrombie Foundation and the Kleberg Foundation. The authors thank Dianna Tudor and Ela Patel for technical assistance, Jane Meara and Paula Thomason for assistance in manuscript preparation, and Cecil Runyons for subject demographic data. The authors also thank Elan Pharmaceutical for the gift of antibodies.
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