Heat Shock Fusion Protein gp96-Ig Mediates Strong CD8 CTL Expansion in vivo
Article first published online: 5 SEP 2002
American Journal of Reproductive Immunology
Volume 48, Issue 4, pages 220–225, October 2002
How to Cite
ŠTRBO, N., YAMAZAKI, K., LEE, K., RUKAVINA, D. and PODACK, E. R. (2002), Heat Shock Fusion Protein gp96-Ig Mediates Strong CD8 CTL Expansion in vivo. American Journal of Reproductive Immunology, 48: 220–225. doi: 10.1034/j.1600-0897.2002.01118.x
- Issue published online: 5 SEP 2002
- Article first published online: 5 SEP 2002
- dendritic cells;
- MHC tetramer;
Štrbo N, Yamazaki K, Lee K, Rukavina D, Podack ER. Heat shock fusion protein gp96-Ig mediates strong CD8 CTL expansion in vivo. AJRI 2002; 48:220–225 © Blackwell Munksgaard, 2002
PROBLEM: As shown previously, gp96-Ig peptide complexes secreted by an ovalbumin transfected tumor (EG7) mediate strong, specific tumor immunity through a CD4 T cell independent CD8+ CTL response. In this study, we set out to develop a system to quantitatively determine the CD8 CTL response to gp96-Ig and to evaluate the influence of an established wild type tumor.
METHODS: Secreted heat shock protein gp96-Ig was constructed by replacement of the endoplasmic reticulum retention signal with the Fc portion of IgG1, transfected into EG7 (EG7-gp96-Ig) and used to induce CD8+ CTL expansion in vivo. Adoptively transferred, ovalbumin specific T-cell receptor (TCR) transgenic CD8+ cells (OT-1) responded with clonal expansion to the immunization with EG7-gp96-Ig. OT-1 expansion was quantitated with Kb-peptide-tetramers by flow cytometry.
RESULTS: In response to primary immunization with EG7-gp96-Ig, OT-1 expand from an initial frequency of 0.5 to 25% of all CD8 cells, and to 50% of all CD8 cells after a booster immunization. Endogenous ovalbumin specific CD8 cells also expand strongly. Antigen specific effector function was measured by enzyme-linked immunosorbent spot-forming cell assay (ELISPOT) for interferon-γ (IFN-γ). While effector function was strongly induced by secreted gp96-Ig, not all expanded OT-1 produce IFN-γ. EG7 does not cause OT-1 expansion, but rather induces anergy. If OT-1 are transferred into wild type EG7 tumor bearing mice to induce anergy of OT-1, immunization with EG7-gp96-Ig can partly overcome unresponsiveness.
CONCLUSIONS: We conclude that secreted gp96-Ig is a powerful mediator of specific CD8+ CTL responses in vivo. Secretory gp96 mimics release of gp96 by damaged or necrotic cells that is able to activate dendritic cells without CD4 help. Gp96-Ig associated peptides have not been selected by binding to major histocompatibility complex (MHC). Specific immunization by secreted gp96-Ig therefore is expected to occur also in allogeneic settings.