Giusi Carbone

MULTIPLE FOREIGN NON-H-2 DETERMINANTS ON THE SURFACE OF A CHEMICALLY-INDUCED MURINE SARCOMA*

By immunizing BALB/c (H‐2d mice against normal tissues from C57BL/6J (H‐2b), C3Hf (H‐2k) and DBA/2 (H‐2d), but not from AKR (H‐2k) strains, resistance was induced to the subsequent challenge of the ‘syngeneic’ methyl‐cholanthrene‐induced BALB/c sarcoma ST5; lymph node cells from allo‐immune BALB/c mice were also able to exert a parallel cytotoxic effect against in vitro cultured ST5 cells. The involvement of foreign H‐2 specificities in the observed cross‐reactions was ruled out by absorptions of H‐2 monospecific sera and by the interallelic combinations used, thus suggesting that non‐H‐2 histocompatibility antigens were responsible for the above findings.

Lack of anti-TSTA antibodies in mice immunized with a methylcholanthrene-induced fibrosarcoma

SummaryThe 3-methylcholanthrene (MCA)-induced BALB/c (H-2d) fibrosarcoma C-1 bears a strong tumor-specific transplantation antigen (TSTA) which, in previous studies, appeared to be distinct from H-2k alien antigens expressed by this tumor. To see whether a syngeneic anti-C-1 serum obtained by multiple immunizations with C-1 tumor cells contained anti-TSTA-specific antibodies, in vitro cytotoxicity tests were performed. The syngeneic anti-C-1 serum had a high cytotoxic activity on C-1 cells, which allowed an absorption analysis to be carried out. Absorption of the serum with C3Hf, AKR, or B10.BR normal lymphoid cells (all sharing H-2k antigens) reduced the cytotoxic activity on C-1 cells to 30%–50%. This residual activity could not be absorbed by FMR+ or G+ murine leukemias, by ecotropic endogenous virus obtained from SC-1 cells infected with the C-1 virus, by embryonic cells, or by normal BALB/c or C57BL/6 lymphoid cells. Conversely, the serum activity was abrogated by absorbing with the MCA-induced BALB/c fibrosarcomas C-1, ST2, C-3, GI-17, or CMS-1, and significantly lowered by the MCA-induced C3Hf fibrosarcoma C3H-7. A significant reduction of the anti-C-1 cytotoxicity was also obtained by absorbing with the two BALB/c fibrosarcomas teflon-9 and SCS (both lacking TSTA), by means of fresh newborn BALB/c or C3Hf muscle cells or of in vitro-cultured newborn BALB/c fibroblasts. These results suggest that, in spite of the strong transplantation immunity elicited by C-1 cells, antibodies to the individual TSTA of C-1 were undetectable in the syngeneic anti-C-1 serum obtained from animals highly resistant to the challenge of C-1 cells.

Increased oncogenic effect of a low dose of methylcholanthrene in immunodepressed mice.

Two different doses of MCA were used to elicit subcutaneous sarcomas in intact vs. immunodepressed adult (C57BL/6xBALB/c)F1 female mice. Immunodepression was achieved by adult thymectomy followed by a short treatment with antilymphocyte serum (4 i.p. injections of 0.25 ml at days −1, +1, +3, +5). The lower dose of MCA (30 μg) gave rise to sarcomas in 11 of 18 intact mice, and in 17 out of 17 immunodepressed animals. No such difference was found with 150 μg of the carcinogen which induced tumors in 16 of 17 immunodepressed mice and in 17 of 17 intact counterparts. These results clearly show the importance of the dosage of MCA in visualizing the funcion of immunologic surveillance in this system of chemical carcinogenesis.

In vivo and in vitro methylcholanthrene carcinogenesis in diffusion chambers.

Two-compartment diffusion chambers containing murine fibroblasts and a 5% (150 μg) or 1% (30 μg) MCA-discs were placed either in the peritoneal cavity of syngeneic adult mice or incubated in vitro. After 4–28 weeks of culture, groups of 5–6 DCs were recovered at 4 weeks intervals, the MCA discarded, and the contents individually implanted into immunodepressed syngeneic mice. The carcinogen displayed a strong cytotoxicity in vitro but not in vivo with both doses. On the whole, when 5% MCA was applied sarcomas appeared in 22/23 mice implanted with cells of the in vivo DC cultures and in 11/16 mice which received the in vitro cultured cells; with the lower dose, the in vivo induced tumors were 19/22 and the in vitro ones 6/12. In all groups the tumor outgrowth into the assay animals was inversely correlated with the time of exposure to MCA during the culture period.

[Role of the immune system in the growth of an isotransplanted urethan-induced lymphosarcoma].

Two groups of (SWR×C57BL)F1 mice were injected subcutaneously with two quantitatively different cell suspensions from a uretan-induced thymic lymphoma of the same hybrid combination. Each of these groups was divided before the neoplastic isograft into the following sub-groups: controls, cyclophosphamide injected, X-irradiated and BCG injected mice. Percentage of takes, tumor diameters and mortality provided the parameters for the evaluation of the tumor growth. With the higher inoculum of neoplastic cells (5×105) only cyclophosphamide was effective in enhancing the take percentage 10 days after the isograft. With the smaller inoculum (5 × 103) all treatments were effective on tumor growth although the inhibitory action of BCG was not as strong as the enhancing action of the immune depressive treatments. The results are interpreted as evidence of a specific antigenicity of urethan-induced lymphoma.

Immunogenic neoplastic clones derived in vitro from an originally non-immunogenic BALB/c fibrosarcoma.

A non-immunogenic fibrosarcoma (SDC2), obtained by spontaneous neoplastic transformation of BALB/c fibroblasts cultured within a diffusion chamber kept in the peritoneal cavity of (BALB/c × C3Hf)F1 mice, was maintained in tissue culture for 10 passages. Three different clones were then derived from the in vitro neoplastic population. Each of the clones, the original in vivo tumor, and its in vitro line taken at the 10th passage (before cloning) were tested for the presence of individual tumor-associated transplantation antigens (TATA) by in vivo growth and excision assay. Contrary to the original SDC2 neoplasm, its in vitro line and 2 out of 3 clones (CL1-SDC2 and CL3-SDC2) were immunogenic, whereas the other clone (CL6-SDC2) displayed no immunogenicity. In addition, CL1-SDC2 and CL3-SDC2 were able to induce a reciprocal cross-protection in in vivo transplantation tests, thus showing common TATA; no cross-reactions were found between these clones and 2 other chemically-induced immunogenic sarcomas. The results suggest an antigenic heterogeneity in the original population of the nonimmunogenic SDC2 sarcoma, with the presence of antigenic but sub-immunogenic neoplastic cells.