Quinone-Induced Mitochondrial Dysfunction and DamageEPA Grant Number: U915002
Title: Quinone-Induced Mitochondrial Dysfunction and Damage
Investigators: Stone, Laura A.
Institution: University of Nevada - Reno
EPA Project Officer: Carleton, James N
Project Period: January 1, 1996 through January 1, 1999
Project Amount: $102,000
RFA: STAR Graduate Fellowships (1996) RFA Text | Recipients Lists
Research Category: Fellowship - Toxicology , Academic Fellowships , Health Effects
The objective of this research project is to determine whether exposure to quinines commonly found in urban air and cigarette smoke result in compromised mitochondrial DNA (mtDNA) function, decreased respiratory capacity, and damage to the mitochondrial DNA. Xenobiotic quinines are ubiquitous and can lead to or contribute to numerous diseases in humans. Previous research has suggested that quinine toxicity is a function of damage to the mitochondrial DNA.
The quinines used in this research project include 1,4-benzoquinone, 2-methyl-1,4-benzoquinone, 1,4-naphthoquinone, and 2-methyl-1,4-naphthoquinone. The models for these studies are Chinese Hamster Ovary (CHO) cells and Balb/c mice. Interactions with mtDNA will be determined by assessing the degress of conformational change from the supercoiled to the relaxed state under both in vitro and in vivo conditions. MtDNA isolated from exposed models will be electrophoresed, transferred to a nylon membrane by Southern techniques, hybridized with an mtDNA-specific probe labeled with digoxigenin, and detected by chemiluminescence. The effects of exposure on adenosine triphosphate (ATP) levels will be determined using the luciferin-luciferase assay and quantified using a luminometer. Heart and liver tissues and CHO cells will be assessed for ATP levels. Oxidative damage to the mtDNA will be analyzed using formamido-pyrimidine-DNA glycosylase (FPG), which predominately recognizes 7,8-dihydro-8-oxo-2'-deoxyguanosine. In these studies, extracted DNA will be restricted with FPG and electrophoresed as described above. The DNA will be transferred to nylon membranes and probed with mtDNA-specific probes as described above.