The hypothesis to be tested in this proposal is that glucocorticoids (GC) exert a biphasic effect on the control of key components of the lipopolysaccharride (LPS)-induced monocyte/ macrophage (MOMac) activation program. A model was proposed by the PI and Co-I to reconcile the stimulatory (""""""""permissive"""""""") actions of moderate cortisol levels with the suppressive actions of high cortisol levels. This model predicts a bell-shaped curve for the effects of cortisol on inflammatory mediators that peaks at or near the upper levels of diurnal cortisol variation. As the permissive effects of GC are less well understood than the inhibitory actions, they provide the focus of our proposed studies on mediators and regulators of MOMac activation.
The Specific Aims are: (1) To test the hypothesis that GC exert a biphasic influence on cytokine production and MOMac phenotype in response to LPS, and (2) To elucidate the molecular mechanisms of permissive and suppressive effects of GC on the MOMac activation program. We propose to address these questions employing both in vitro and human in vivo systems. Initially, we will use cultures of elutriated MO that will be exposed to LPS under conditions in which cortisol is maintained at levels predicted to be sub-permissive, permissive or suppressive. We will use ELISA and flow cytometric analyses to determine the interactive effects of cortisol and LPS on two key components of the MOMac activation program--IL-6 and CD163. Using LPS and cortisol at dosages and times that are first determined to result in maximal enhancement or inhibition of CD 163 and IL-6, we will examine additional cytokines, signaling molecules and receptors that have been shown to be induced or suppressed by LPS. We will also use a well-controlled paradigm of human experimental endotoxemia to elucidate the physiologic and molecular mechanisms that lead GC to either suppress or enhance the production and actions of putative pro- and anti-inflammatory molecules during the response to endotoxin in vivo. We believe that these studies will yield important insights into the mechanisms by which GC can both enhance and suppress immune and inflammatory functions, leading to more effective approaches to their use in clinical settings of inflammation.
