We have previously reported the following: 1) induction of a Shwartzman-type biological reaction in a mass of Ehrlich solid carcinoma in mice using bacterial components resulted in necrosis of the carcinoma tissue 1); 2) mice that achieved reduction and subsequent resolution of a mass became resistant to the second subcutaneous challenge with Ehrlich carcinoma cells and transplantation of skin isografts 2); 3) administration of non-bacterial components, such as gelatin and its derivatives, induced the same type of immunity 3); and 4) Ehrlich carcinoma cells transplanted subcutaneously into mice with acquired anti-transplant immunity to Ehrlich carcinoma transplants underwent coagulative necrosis 4).
Although it is interesting to see the type of immune response observed when Ehrlich carcinoma cells are intraperitoneally transplanted into mice which have become resistant to transplantation of these cells, such immunity is only effective against solid carcinoma and not against ascites carcinoma. More specifically, mice that tolerated subcutaneous transplantation of 5 x 106 carcinoma cells could not escape carcinoma-related death after intraperitoneal transplantation of the same number of carcinoma cells. We then attempted to enhance the immunity of these mice to create animals also resistant to ascites carcinoma transplants. Described below are the results obtained by intraperitoneally transplanting ascites carcinoma cells into these mice.

Methods and results

Porcine skin gelatin with amino groups methylene polymerized by formaldehyde was dissolved in water to make a 1% solution, and 0.3 cc of this solution was repeatedly injected subcutaneously into the inguinal region of mice every 7 days for a total of 5 doses. Seven days after the last injection, 5 x 106 Ehrlich carcinoma cells were subcutaneously transplanted into the gluteal region.
After 7 days, masses were palpable in all mice. In 2 of 3 mice, the masses gradually reduced in size and eventually disappeared; thus, transplantation failed. In the remaining mouse, carcinoma cells engrafted and the mouse died due to carcinoma. When the mice which had survived for 1 month were subcutaneously rechallenged with 5 x 106 Ehrlich carcinoma cells, the cells did not engraft and all mice survived. After 1 month, the mice were subcutaneously rechallenged with the same number of cells and again all mice survived. When these mice, which had tolerated subcutaneous challenge with a total of 15 x 106 cells, were intraperitoneally transplanted with 5 x 106 cells, half developed ascites and died due to carcinoma while the remaining mouse survived without ascites, with non-engraftment of carcinoma cells. The mouse that had tolerated intraperitoneal transplantation of 5 x 106 cells also tolerated and survived subsequent intraperitoneal rechallenge with 5 x 107 carcinoma cells.
To the mice that had tolerated subcutaneous and intraperitoneal transplantation of Ehrlich carcinoma cells, 1.2 x 107 Ehrlich carcinoma cells were intraperitoneally transplanted and small amounts of peritoneal content were collected over time by abdominal puncture. The samples were examined with a phase-contrast microscope for observation of cells.
By 2 h after transplantation, round cells were observed attached to carcinoma cells. By 3 h and 30 min after transplantation, 60% of carcinoma cells were found to be surrounded by 1-15 peritoneal round cells in the form of rosettes (Fig. 1). By 5 h after transplantation, carcinoma cells were lyzed and became uncollectible.

Methods and results

The fact that immunity acquired by a tumor-bearing animal is not sufficient to cause necrosis of carcinoma explains why carcinoma can be intractable. In experimental settings, coagulative necrosis of carcinoma cells was observed in subcutaneous carcinoma transplantation and cellular immune response medicated by round cells was observed in intraperitoneal carcinoma transplantation. The fact that such immunity can be transferred via serum suggests the presence of a humoral mechanism that functions prior to cellular immune response. If cancer immunity is considered as a form of tissue transplantation immunity, biological reactions mediating the rejection of transplanted carcinoma tissue that would otherwise be accepted may eventually induce degenerative necrosis of autologous tissue. Until now, none of the studied mice have developed autoimmune disease.