Injection of Soluble but Not Tumor-Cell-Associated Antigens Into the Anterior Chamber of the Eye Inhibits Antigen-Specific CTL Responses

Abstract

Abstract: : Purpose:A previous report has shown that the injection of P815 mastocytoma cells, into the anterior chamber (a.c.) of Balb/c mice induces a unique form of systemic tolerance termed anterior chamber associated immune deviation or ACAID, characterized by inhibited DTH responses and enhanced CTL responses. The injection of soluble antigen into the a.c. also results in an inhibition of DTH responses. The goal of the studies reported here, was to determine whether soluble antigens injected into the a.c. inhibit CTL responses. Methods:A standard adoptive transfer technique was employed to track antigen specific CTL precursors in vivo. Splenocytes (1.0 X 10₇) from OT-I mice, transgenic for a TCR specific to OVA peptide 257-264 complexed with H-2 K_b, were used as a source of OVA-specific CD8₊ T cells, and were injected into naïve syngeneic C57Bl/6 mice by tail vein injection. Two days later, soluble OVA, E.G7-OVA or EL-4 (1.0 X 10₅) cells were injected into the a.c. of the right eye of recipient mice. E.G7-OVA cells are derived from the EL-4 thymoma that has been transfected with the gene for OVA. As a result, OVA is expressed by the tumor cells as a tumor-cell-associated antigen. As a positive control for the in vivo expansion of OT-I T cells, mice were given OVA in the subconjunctiva (scon.). Three days after antigen administration, OT-I T cells were enumerated in the submandibular lymph nodes. In addition, submandibular lymph node cells were cultured with 150 µg / ml OVA for three days to activate OVA-specific CTL responses that were measured by a standard Cr₅₁ release assay with E.G7-OVA cells as targets. Results:OT-I T cells in the right or left submandibular lymph nodes did not expand in mice given EL-4 a.c. The number of OT-I T cells increased 40-fold in the right submandibular lymph node of mice given OVA scon. when compared to negative control mice given EL-4 a.c. OT-I T cells in the right submandibular lymph nodes of mice given OVA or EG7-OVA a.c. increased 3.4-fold and 4.6-fold respectively. However, CTL responses were markedly higher in right submandibular lymph node cultures from mice given E.G7-OVA (56 % lysis) compared to mice given OVA a.c. (3 % lysis). The lytic activity in cultures from mice given EG7-OVA was comparable to mice given OVA scon. (48 % lysis) whereas the lytic activity in cultures from mice given OVA a.c. was comparable to negative control mice (4% lysis). Conclusion:These data suggest that CTL responses are induced if tumor-cell-associated antigens are injected into the a.c. In constrast, soluble antigens injected into the a.c. inhibit CTL responses.

Keywords: 301 ACAID • 433 immune tolerance/privilege • 317 anterior chamber