Context: Aging in humans is characterized by a selective decline in circulating levels of adrenal androgens. The results of in vivo studies are suggestive of reduced adrenal 17,20-lyase activity in aging men and women. Objective: We sought to determine whether there are changes in the distribution and/or expression of cytochrome B5 (CytB5), an accessory protein important in the regulation of 17,20-lyase activity, in the adrenals of aging humans. Design: Comparison between younger and older adrenal glands. Setting: The study was conducted in a University Center. Patients or Other Participants: Adrenal glands obtained at autopsy after sudden death as a result of trauma from 46 young (age 20-40 yr) and 26 older (age 50-91 yr) humans were obtained and fixed within 24 h postmortem. Interventions: Paraffin sections were stained with hematoxylin and eosin and also were subjected to immunohistochemical staining for CytB5. All sections were quantitatively evaluated using an image capture and analysis program and qualitatively evaluated with respect to staining intensity. Main Outcome Measures: To determine whether there are any changes in CytB5 distribution in the adult human adrenal cortex during the aging process using qualitative and quantitative analysis with respect to age, gender, race, and postmortem interval. Results: CytB5 immunoreactivity was found in cells that corresponded to those of the zona reticularis. The percentage of the adrenal cortex immunoreactive for CytB5 decreased with aging (38.6 ± 7.6% for young and 30.1 ± 5.9% for older, mean ± SD; P < 0.0001) as did the percentage of adrenocortical tissue comprising the zona reticularis (36.8 ± 10.8% for young and 27.2 ± 5.9% for older; P < 0.001). However, there was no apparent change in the staining intensity of CytB5 among those cells that were immunopositive for this factor with aging. Conclusions: There appears to be a reduction in the proportion of the adrenal cortex that expresses CytB5 with aging, and this likely corresponds to a shrinkage of the zona reticularis. The mechanism and cause for this cortical regression are unknown. Copyright © 2005 by The Endocrine Society.