Original H‐2^d and alien H‐2^k‐like antigens of a BALB/c fibrosarcoma as defined by in vitro and in vivo studies of cell‐mediated immunity

Abstract

The present study examines the immunosensitivity and the immunogenicity of both original H‐2d and alien H‐2k‐like antigens of the BALB/c (H‐2d) fibrosarcoma C‐1 as detected by in vitro and in vivo cell‐mediated cytotoxicity (CMC) assays. It was found that 51Cr‐labeled C‐1 cells were lysed in vitro by C 57 BL/6 anti‐H‐2d lymphocytes. The specificity of this reaction was shown by cold inhibition experiments in which the anti‐H‐2d cytotoxic activity on YC8 (H‐2d) targets was inhibited by unlabeled YC8 or C‐1 but not by C3UR11 (H‐2k) tumor cells. Both Dd‐ and Kd‐encoded antigens were recognized by appropriate cytotoxic effectors. The immunogenicity of H‐2d antigens of C‐1 was revealed by the ability of C 57 BL/6 anti‐C‐1 lymphocytes to lyse YC8 targets. The expression of H‐2k‐like alien alloantigens on C‐1 was indicated by the finding that anti‐H‐2k cytotoxic T lymphocytes (CTL), generated by culturing BALB/c spleen cells immune to BALB.K (H‐2k), C3Hf (H‐2k) or A (H‐2a = H‐2k/d) tissues with the cells of the same strain used for immunization, lysed C‐1 targets. The cytotoxicity of these anti‐H‐2k CTL against C 3 UR11 (H‐2k) targets could be specifically inhibited by cold C 3 UR 11 or C‐1 cells but not by two other BALB/c tumors. Using recombinant H‐2‐congenic mice, it was shown that both Dk and Kk antigens were recognized by CTL on C‐1 cells. The immunogenicity of the H‐2k‐like antigens, however, could not be detected in vitro. In fact, effector spleen cells from BALB/c mice immune to C‐1 did not develop any detectable cytotoxicity against C 3 UR 11 targets as assayed either by a direct in vitro test or after in vitro restimulation with C‐1 sarcoma cells. A similar experimental design was adopted in Winn assays carried out by mixing spleen cells of BALB/c immune mice with either C‐1 or C3 UR 11 targets and injecting the mixtures in BALB/c or hybrid recipients. These in vivo tests revealed the presence of both H‐2d (Kd and Dd) and H‐2k‐like (Kk and Dk) antigens on C‐1. At variance with the in vitro CMC assays, however, the Winn assay also detected the immunogenicity of the H‐2k alien antigens, since BALB/c anti‐C‐1 spleen cells were able to significantly reduce the growth of C 3 UR 11 lymphoma cells in (BALB/c × C 3 Hf)F1 hosts.

Lysis of autologous human melanoma cells by in vitro allosensitized peripheral blood lymphocytes

“…The cytotoxicity of in vitro stimulated lymphocytes was tested against tumor target cells in 4-or 18 h 51Cr-release assay, as previously described [13]. The cytotoxicity of in vitro stimulated lymphocytes was tested against tumor target cells in 4-or 18 h 51Cr-release assay, as previously described [13].…”

Mouse tumors are heterogeneous in their susceptibility to syngeneic lymphokine-activated killer cells and delineate functional subsets in such effectors

“…The assay was carried out as previously described [24]. Briefly, effectors and »lCrlabelled targets were added to triplicate U-bottom wells of 96-well microplates (Costar 3799) in a final volume of 200 l-tl and incubated at 37 ° C for 4 h. In cold-target inhibition studies, unlabelled competitor cells were added first, followed by »lCr-labelled targets and an appropriate number of effectors.…”

Cross‐reactions between tumor cells and allogeneic normal tissues. inhibition of a syngeneic lymphoma outgrowth in h‐2 and non‐h‐2 alloimmune balb/c mice

“…The outgrowth of otherwise lethal doses of the Moloney virusinduced lymphoma YC8 and of its clones was inhibited in BALBlc mice immune to DBAl2 (H-2'9, C3Hf (H-ZC), C3H.SW (H-Zb), C3H It has been reported that murine tumors may express alien histocompatibility antigens (HA) encoded both by the MHC or by non-H-2 genes (Parmiani et al, 1979). One approach used to detect cross-reactions between tumors and allogeneic normal cells was to see whether alloimmunization of recipient mice would generate in vivo resistance to subsequent challenges of syngeneic tumors or in vitro cytotoxic activity to tumor targets (Gipson et al, 1978;Fujiwara et al, 1978;Parmiani et al, 1980;Russell et al, 1979;Schinmacher et al, 1980;Sensi et al, 1980). Without identifying the cross-reaction antigens, Zarling et al (1978a) were able to elicit significant cell-mediated killing on human autologous neoplastic targets by peripheral blood lymphocytes of certain patients sensitized in vitro to a pool of 20 different allogeneic normal lymphocytes, a panel representative of the whole spectrum of HLA antigens or of their lymphocyte-defined determinants.…”