Human Cytomegalovirus pp65- and Immediate Early 1 Antigen-Specific HLA Class I-Restricted Cytotoxic T Cell Responses Induced by Cross-Presentation of Viral Antigens

Abstract

Dendritic cells (DCs) play a pivotal role in the development of anti-viral CD8+ CTL responses. This is straightforward if they are directly infected with virus, but is less clear in response to viruses that cannot productively infect DCs. Human CMV (HCMV) shows strain-specific cell tropism: fibroblast (Fb)-adapted laboratory strains (AD169) and recent clinical isolates do not infect DCs, whereas endothelial cell-adapted strains (TB40/E) result in productive lytic DC infection. However, we show here that uninfected DCs induce CD8+ T cell cytotoxicity and IFN-γ production against HCMV pp65 and immediate early 1 Ags following in vitro coculture with HCMV-AD169-infected Fbs, regardless of the HLA type of these Fbs. CD8+ T cell stimulation was inhibited by pretreatment of DCs with cytochalasin B or brefeldin A, indicating a phagosome/endosome to cytosol pathway. HCMV-infected Fbs were not apoptotic as measured by annexin V binding, and induction of apoptosis of infected Fbs in vitro did not augment CTL induction by DCs, suggesting a mechanism other than apoptosis in the initiation of cross-presentation. Furthermore, HCMV-infected Fbs provided a maturation signal for immature DCs during coculture, as evidenced by increased CD83 and HLA class II expression. Cross-presentation of HCMV Ags by host DCs enables these professional APCs to bypass some of the evasion mechanisms HCMV has developed to avoid T cell recognition. It may also serve to explain the presence of immediate early 1 Ag-specific CTLs in the face of pp65-induced inhibition of Ag presentation at the level of the infected cell.

Efficient Generation of Antigen-Specific Cytotoxic T Cells Using Retrovirally Transduced CD40-Activated B Cells

“…For example, infusion of CMVspecific CD8 ϩ T cell clones to hemopoietic stem cell recipients resulted in the restoration of CMV-specific immunity (12,19). Various combinations of APC and CMV Ags have been developed for the generation of CMV-specific T cell responses including CMV-infected fibroblasts (20); CMV-infected MoDC (21,22); MoDC pulsed with pp65 peptides (23,24); MoDC pulsed with recombinant pp65 protein (25); adenovirus vector-infected MoDC (26); vaccinia virus vector-infected MoDC (27); LCL transfected with pp65 gene (28); and retrovirus vector-infected LCL (29). All of the methods described to date have limitations.…”

Cross-Presentation of the Oncofetal Tumor Antigen 5T4 from Irradiated Prostate Cancer Cells—A Key Role for Heat-Shock Protein 70 and Receptor CD91

“…Furthermore, DCs can acquire antigen from live cells for antigen cross-presentation in both tumor and viral settings (44)(45)(46). In the latter, while apoptosis is inhibited by the virus, Hsp70 expression is significantly upregulated (47).…”

Definition of an HPV18/45 cross‐reactive human T‐cell epitope after DNA immunisation of HLA‐A2/KB transgenic mice

“…Dendritic cells were grown from adherent monocytes as previously described. 37,38 Briefly, dendritic cells were grown from adherent fraction of PBMC and grown in hCM supplemented with 500 u/ml IL-4 (R&D systems, UK) and 800 u/ml GM-CSF (Leucomax, UK). Media and cytokines were replenished on day 3.…”