Reactivation of latent human cytomegalovirus in CD14⁺ monocytes is differentiation dependent

We have previously demonstrated reactivation of latent human cytomegalovirus (HCMV) in myeloid lineage cells obtained from healthy donors. Virus was obtained from allogenically stimulated monocyte-derived macrophages (Allo-MDM), but not from macrophages differentiated by mitogenic stimulation (ConA-MDM). In the present study, the cellular and cytokine components essential for HCMV replication and reactivation were examined in Allo-MDM. The importance of both CD4⁺ and CD8⁺ T cells in the generation of HCMV-permissive Allo-MDM was demonstrated by negative selection or blocking experiments using antibodies directed against both HLA class I and HLA class II molecules. Interestingly, contact of monocytes with CD4 or CD8 T cells was not essential for reactivation of HCMV, since virus was observed in macrophages derived from CD14⁺ monocytes stimulated by supernatants produced by allogeneic stimulation of peripheral blood mononuclear cells. Examination of the cytokines produced in Allo-MDM and ConA-MDM cultures indicated a significant difference in the kinetics of production and quantity of these factors. Further examination of the cytokines essential for the generation of HCMV-permissive Allo-MDM identified gamma interferon (IFN-γ) but not interleukin-1 or -2, tumor necrosis factor alpha, or granulocyte-macrophage colony-stimulating factor as critical components in the generation of these macrophages. In addition, although IFN-γ was crucial for reactivation of latent HCMV, addition of IFN-γ to unstimulated macrophage cultures was insufficient to reactivate virus. Thus, this study characterizes two distinct monocyte-derived cell types which can be distinguished by their ability to reactivate and support HCMV replication and identifies the critical importance of IFN-γ in the reactivation of HCMV.