Effects of Bacillus Calmette-Guerin (BCG) on the Spleen of Mouse Inoculated with Ehrlich Carcinoma Cells: A Morphological Study

Ko, Jeong Sik; Park, Sook Hyun; Ahn, E Tay; Park, Kyung Ho; Kim, Jin Gook

Abstract

This experiment was performed to study the morphological responses of the spleen of mouse inoculated with Ehrlich carcinoma cells, following administration of Bacillus Calmette-Guerin (BCG). Healthy adult ICR mice weighing 25 g each were divided into normal and experimental groups (experimental control group and BCG treated group). In the experimental groups, each mouse was inoculated with 1X10(7) Ehrlich carcinoma cells subcutaneously in the inguinal area. From next day, 0.2 ml of saline or BCG (0.03X10(8)-0.32X10(8) CFU) were injected subcutaneously to the animals every other day. The day following the 7th injection, each mouse was injected with a single dose of 0.7 microCi/gm of methyl-3H-thymidine (25 Ci/mmol, Amersham Lab., England) through tail vein. Seventy minutes after the 3H-thymidine injection, animals were sacrificed. Pieces of the splenic tissue, fixed in 10% neutral formalin for light microscopy. The sections were coated with autoradiographic emulsion EM-1 (Amersham Lab., England) and the coated sections were exposured for 5 weeks in the dark room. For electron microscopy, tissues were prefixed with phosphate buffered 2,5% glutaraldehyde-1.5% paraformaldehyde (pH 7.3), and post-fixed with phosphate buffered 1% osmium tetroxide solution (pH 7.3). Ultrathin sections of the white pulp area stained with uranyl acetate and lead citrate were observed with a JEM 100CX-II electron microscope. On histological study in the splenic white pulp, BCG treated mice showed more macrophages containing pyknotic nuclei than normal or experimental control mice showed. On autoradiographic study, a large number of the 3Hthymidine labeled cells were seen near the marginal zone, whereas only a small number of labeled cells were seen in the red pulp or the white pulp of the spleen. The number of the labeled cells in experimental control group was similar to that in the normal control mice, whereas that in BCG-treated mice was significantly increased as compared with that of normal control one. On electron microscopic study, in the white pulp of BCG treated mouse, mitotic cells were observed more frequently than in those of the normal or experimental control mice. In the BCG treated mice, macrophages and plasma cells in the white pulp were observed more frequently than in those of the normal or experimental control mice, whereas a few eosinopile leucocytes were observed, and perichromatin granules within the nuclei of the lymphocytes and the reticular cells were observed frequently. From the above results, it was concluded that DNA syntheses were more active in the cells of the marginal zone than in the cells of the white pulp or the red pulp. And repeated treatment with BCG could activate the DNA syntheses of splenic cells and increase the number of the macrophages and the plasma cells in the white pulp.