(verbatim from application) To elucidate the molecular basis of aging and its retardation through caloric restriction (CR), we employed high density oligonucleotide arrays representing 6347 genes to study aging of the gastrocnemius muscle of C57BL/6J mice (Science 285:1390, 1999). Aging resulted in a differential gene expression pattern indicative of a marked stress response and lower expression of metabolic and biosynthetic genes. Most alterations were either partially or completely prevented by CR. Taken together, our finding suggest that aging, at the transcriptional level is associated with a chronic state of oxidative stress (OS). Thus, the central hypothesis to be tested is that OS is a major causative factor of aging in skeletal and cardiac muscle. We propose two specific aims:
Specific aim 1 : Gene expression profiling of greater than 30,000 genes in aging skeletal and cardiac muscle. By using newly developed Affymetrix DNA chips, we will expand our analysis of the GN muscle to include greater than 30,000 genes. Cardiac muscle (left ventricle (LV)) will also be studied. Adult (5 mo) and old (30 month control fed and CR) B6 mice will be studied. Results will be validated by comparing the gene expression profile for control fed mice with those on CR. Clustering algorithms will be applied to define families of genes that are differentially responsive to aging and CR. Also statistical methods we have developed for evaluating differences in microarray expression data will be applied.
Specific aim 2 : A genetic test of the OS hypothesis of aging in mammals. Although aging is associated with increased OS, a causal role for OS has not been shown in mammals. Using newly developed transgenic mice, we will determine whether aging is retarded in either a strain over expressing both catalase and CuZn superoxide dismutase (SOD) or another over expressing mitochondrially-targeted catalase and MnSOD. Aging rates will be evaluated at the 1) organism level by survival and disease patterns, 2) morphological level by electronmicroscopy (EM) immunogold techniques to localize and quantify antioxidant enzyme levels and oxidative damage products in the GN and LV, 3) biochemical level by measure of antioxidant enzyme activities and indicators of oxidative damage in the GN and LV, and 4) molecular level through gene expression profiling of the GN and LV. This investigation would be the first to employ a global set of molecular (transcriptional) markers of aging coupled with biochemical and histological analyses to study the aging process and its possible retardation by genetic interventions designed to lower OS.
Showing the most recent 10 out of 21 publications
