Granulocyte-macrophage colony-stimulating factor (GM-CSF) was established as the constitutive and elicited human umbilical vein endothelial cell-derived eosinophil viability-sustaining factor. Stimulation of endothelium cell monolayers with IL-la (5 U/ml) increased the 48-h elaboration of GM-CSF from a mean of3.2 to a mean of8.2 pM (P < 0.05). Dexamethasone (100 nM) decreased the constitutive GM-CSF elaboration by 49% (P < 0.001) but did not diminish production by IL-lastimulated endothelium. However, eosinophil viability decreased by 21% in dexamethasone-pretreated IL-la-stimulated endothelial cell-conditioned medium (P < 0.05), which suggested viability antagonism by glucocorticoids. After 24 h of culture, eosinophil viability for replicate cells in enriched medium alone or with 1 pM GM-CSF decreased from means of 43 and 75% to means of 21 and 54%, respectively, when dexamethasone was included (P < 0.05). However, 10 pM GM-CSF, IL-3, or IL-5 protected the cells against dexamethasone and against endonuclease-specific DNA fragmentation. In this model system of eosinophil-tissue interactions, dexamethasone prevents the endothelial cells from inducing a pathobiologic phenotypic change in the eosinophil by suppression of GM-CSF elaboration to concentrations that are not cytoprotective. Cytokine priming by GM-CSF, IL-3, or IL-5 may account for the differential responsiveness of select eosinophilic disorders to glucocorticoids. (J. Clin.

Chemokine receptors are members of the G protein-coupled receptor (GPCR) family. CCR5 and CXCR4 act as co-receptors for human immunodeficiency virus (HIV) and several efforts have been made to develop ligands to inhibit HIV infection by blocking those receptors. Removal of chemokine receptors from the cell surface using polymorphisms or other means confers some levels of immunity against HIV infection. Up to now, very limited success has been obtained using ligand therapies so we explored potential avenues to regulate chemokine receptor expression at the plasma membrane. We identified a molecular chaperone, DRiP78, that interacts with both CXCR4 and CCR5, but not the heterodimer formed by these receptors. We further characterized the effects of DRiP78 on CCR5 function. We show that the molecular chaperone inhibits CCR5 localization to the plasma membrane. We identified the interaction region on the receptor, the F(x)6LL motif, and show that upon mutation of this motif the chaperone cannot interact with the receptor. We also show that DRiP78 is involved in the assembly of CCR5 chemokine signaling complex as a homodimer, as well as with the Gαi protein. Finally, modulation of DRiP78 levels will affect receptor functions, such as cell migration in cells that endogenously express CCR5. Our results demonstrate that modulation of the functions of a chaperone can affect signal transduction at the cell surface.

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