We have previously reported that pre-treatment with gelatin or its derivatives induced resistance to Ehrlich carcinoma transplants in mice^1-4). The effect varied according to the type of gelatin, was not related to the pharmacological- or macrophage-inducing activity of gelatin, and was enhanced by methylene-polymerization of amino groups with formaldehyde. In previous studies, all gelatin products were injected as respective aqueous solutions. In the present study, we demonstrated induction of anti-tumor immunity following the subcutaneous injection of insoluble gelatin power, which was subsequently and spontaneously eliminated from the mouse body.

Materials and methods

A commercially available 20% aqueous gelatin solution was sterilized at 140 °C for 5 min, cooled to 10 °C, cut into blocks, and allowed to gelate in 10% formalin solution at room temperature for 48 h. The blocks were then cut into small pieces, washed with running water for 3 days to remove excess formalin, dried with hot air, and powderized with a ball mill. The power was filtered through a 200-μm mesh and used in the following experiment. The residual formalin content in the dried power was 3.36 mg/1000 mg. This gelatin powder swelled when exposed to water, but was insoluble in water even after heating at 100 °C for 30 min. Subcutaneous injection of 30 mg of this power into the gluteal region of a mouse resulted in the formation of 2-4 mm-thick granulation tissue around the injection site within 2 weeks. After 1 month, the granulation tissue disintegrated where it came into contact with the skin and was gradually eliminated as unchanged dry powder from the skin surface. Injection of non-polymerized gelatin powder or formalin solution alone resulted in no formation of granulation tissue. Next, 30 mg of the same insoluble gelatin power filtered through a 200-μm mesh was mixed in sterilized water for convenience and injected subcutaneously into the gluteal region of mice (ddY) through a thick needle connected to a syringe. After 1 month, the granulation tissue which had formed around the injection site disintegrated where it came into contact with the skin and was gradually eliminated as unchanged powder from the skin surface over a few days without being dissolved or absorbed. Then, 5 x 106 Ehrlich carcinoma cells were subcutaneously transplanted in the contralateral gluteal region.

Results and discussion

Carcinoma cells did not engraft in 6 of 7 transplanted mice in the experimental group and in 1 of 7 mice in the control group. The gelatin power swelled when mixed with water for injection but was re-dehydrated in the mouse body and eliminated as dried and unchanged powder without being dissolved or absorbed. The powder was not biocompatible and was eliminated from the mouse body, leaving an anti-transplant immunity against subsequently transplanted Ehrlich carcinoma cells.

This study was conducted at the Department of Microbiology, Osaka Medical College.