Incoming Human Cytomegalovirus pp65 (UL83) Contained in Apoptotic Infected Fibroblasts Is Cross-Presented to CD8+ T Cells by Dendritic Cells

CD8+ T-cell response is a major effector mechanism to contain viral infections. The importance of dendritic cells (DCs) for the initiation of CD8+ T-cell responses against viruses has been recently emphasized by the description of molecular mechanisms involved in naive T-cell stimulation. It has been suggested that CD8+ T-cell expansion could first require CD4+ T-cell help through CD40 ligation on DCs and that this step can be bypassed by direct infection of DCs by a virus (17). Nevertheless, this finding did not fully explain how antiviral CD8+ T cells could be generated in cases where DCs were not susceptible to infection. DCs function as sentinels of the immune system and are characterized by their ability to activate T cells through unusual pathways of antigen capture (2). Since DCs can deliver exogenous antigens in either the soluble or particulate form into the major histocompatibility complex (MHC) class I pathway, it is conceivable that antiviral cytotoxic T-lymphocyte (CTL) priming might be achieved in this way in cases of infection by cytopathic viruses. Moreover, DCs are able to acquire antigens through phagocytosis of infected apoptotic cells and process them into the MHC class I pathway for presentation to CD8+ T cells (1). Although there is evidence that DCs are susceptible to infection by viruses such as influenza virus, human immunodeficiency virus, and measles virus (1, 12), the extent to which each antigen capture mechanism contributes to the activation of virus-specific immune response is not fully known. Nevertheless, one may suggest that the cross-presentation mechanism has direct implications in expansion of CD8+ T cells targeted against noncytopathic viruses which do not infect DCs. We then asked how this could take part in activation of CD8+ T cells against human cytomegalovirus (HCMV). Infection by HCMV, a latent betaherpesvirus, is well controlled by T cells mainly of the CD8+ subset, whose major target is the matrix protein pp65 (for reviews, see references 3 and 15). Indeed, very high frequencies of anti-pp65 CD8+ T-cell precursors have been found in healthy blood donors compared to other viral proteins (24). Whereas it has been shown that HCMV is latent in DC progenitors (11), it seems that susceptibility of either immature or mature DCs to infection is not well established and may vary according to both cell subset and HCMV strain (18, 21). It was recently demonstrated that HCMV infection could either induce or inhibit apoptosis mediated by Fas, tumor necrosis factor alpha (TNF-α) receptor 1 (TNF-R1), and TRAIL (TNF-related apoptosis-inducing ligand) receptor in vitro (4, 9, 20), and that apoptosis of infected cells in Fas and TNF-R1 knockout mice played a pivotal role in the clearance of murine CMV in vivo (7). In this study, we examined whether cross-presentation of apoptotic cells by DCs could be involved in activation of anti-HCMV-specific CD8+ T cells. We showed that TNF-α could induce apoptosis of HCMV AD169-infected MRC5 fibroblasts very early after infection, providing a suitable source of antigen for phagocytosis by immature DCs. We then derived immature DCs from peripheral blood mononuclear cells (PBMC) and demonstrated that they did not synthesize HCMV immediate-early (IE) antigens even after prolonged incubation with the virus. Finally, we showed that HLA-A2- and HLA-B35-restricted anti-pp65 CD8+ CTL were stimulated following exposure of DCs to HCMV-infected apoptotic fibroblasts, suggesting that DCs acquired and processed properly the matrix protein pp65 through phagocytosis of apoptotic bodies. Thus, seeing that pp65 was internalized by fibroblasts immediately after viral input without de novo synthesis, processing of this incoming antigen by DCs may provide a quick and efficient way to prime anti-HCMV CTL before viral replication.