PROCESSING OF OXIDATIVE STRESS IN ALZHEIMER

Aging

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Abstract

Summary of work: Studies have demonstrated that several cellular markers of oxidative stress are higher in cells from Alzheimer disease (AD) patients as compared to normal age-matched controls. These markers include oxidative damage to lipids, proteins, and DNA in various tissues from AD patients. It has been proposed that AD cells may have a defect in the DNA repair processing of oxidative base lesion leading to accumulation of DNA damage in AD cells. We have investigated the repair of oxidative base lesions using whole cell extracts from cultured AD lymphoblasts. DNA substrates containing both pyrimidine and purine lesions were obtained by treatment of plasmid with either gamma irradiation or fluorescent light (FL). Plasmid DNA containing primarily thymine glycol or 8-hydroxyguanine was prepared by damaging DNA with either OsO4 or methylene blue plus light ,respectively. The DNA substrates were purified free of strand breaks and were used in DNA repair synthesis assays. FAD cells were proficient in repair of these substrates containing various oxidative base lesions. Extracts from FAD cells repaired the plasmids damaged by gamma or FL-irradiation with equal efficiency as extracts from unaffected individuals. Furthermore, DNA damaged with methylene blue plus light and OsO4 were repaired with greater efficiency using FAD extracts (approximately 0.5-fold increase) as compared to cells from unaffected individuals. Our data indicate that the DNA damages resulting from the oxidative stresses used here are repaired efficiently in FAD cells. In our experiments, we have compared the repair in FAD cells with cell lines from patients known to be effective in DNA repair capacity. For example, we compared the repair in FAD cells to that in xeroderma pigmentosum group A (XPA) cells known to be defective in nucleotide excision repair. Interestingly, the XPA cells were defective in teh repair of certain oxidative DNA base modifications. These lesions may thus be repaired by nucleotide excision repair rather than base excision repair. It is possible that the repair of specific oxidative base damages, such as that seen for thymine glycol and 8-hydroxyguanine, may be upregulated in FAD due to chronic oxidative stress which has been previously implicated in this disease. There have been reports of unusual accummulation of oxidative DNA damage in mitochondrial DNA from patients with AD, and we are have recently developed novel techniques to study DNA repair in these organelles. We thus plan to investigate the DNA repair efficiency in mitochondria from individuals with AD. In addition studies are underway in knockout mice that are defective in the APP1 gene for their processing of oxidative DNA damage.