The maximal capacity of Brown Norway rat Leydig cells to produce testosterone decreases significantly with age. Our major objective is to elucidate the molecular mechanisms that are responsible. A central hypothesis of this application is that reactive oxygen species (ROS), produced by the Leydig cells themselves, play an important role in age-related reductions in Leydig cell testosterone production.
Three specific aims are proposed. The first is to determine whether age-related reductions in Leydig cell testosterone production are reversed by administering LH directly to the testes of old rats, or by encapsulating old cells and implanting them into young rats. These studies will test the hypothesis that factors outside the Leydig cells might be responsible for the reduced ability of old Leydig cells to produce testosterone. In the second specific aim, we will determine whether the increases in Leydig cell reactive oxygen production that occur as these cells age are from the mitochondrial transport chain, the P450 reactions of steroidogenesis, or both; and whether there are age-related changes in mRNA, protein, and/or activity levels of the major enzymatic scavengers of reactive oxygen species in Leydig cells - SOD, glutathione peroxidase and catalase.
The third aim i s to examine the effects of manipulating reactive oxygen load on Leydig cell steroidogenesis during aging, based on the hypothesis that reactive oxygen, whether derived from the electron transport chain, steroidogenesis, or both, plays an important role in the reduced testosterone produced by aging Leydig cells. We will test this hypothesis by examining the consequences of manipulating oxidative stress load in vivo on Leydig cell function, including: the effects of vitamin E supplementation and deprivation; the effects of age on the acute response of Leydig cells to depletion of its major non-enzymatic antioxidant, glutathione; and the effects of caloric restriction. Together, these studies will provide new insights into how Leydig cells cope with stressors that are present or increase during aging, and will shed light on the underlying molecular basis for functional changes in Leydig cells that occur during aging.
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