ラットのDMBA誘発乳癌の発癌に及ぼす蛋自分解酵素阻害剤(Leupeptin)の影響に関する実験的研究

Abstract

In recent years, there have been reports that various protease inhibitors depress the carcinogenesis in mouse skin induced by DMBA and croton oil, and that protease enhances the transformation and proliferation of cultured cells. Thus, it has been shown that protease plays an important role in carcinogenesis.This study was undertaken to elucidate the effect of Leupeptin, a protease inhibitor isolated from Actinomycetes, on rat mammary carcinogenesis induced by DMBA. Groups of Sprague-Dawley female rats were given the usual laboratory diet (control diet) and others the same diet containing 0.1% Leupeptin (Leupeptin diet).In the first experiment, all rats were injected intravenously with 3 mg of DMBA at 55 and 58 days of age. Administration of the Leupeptin diet was begun 7 days before DMBA treatment and continued for 120 days after DMBA treatment. Systemic toxicity combined with adrenal damage caused by DMBA treatment led to 41% mortality in the control diet group and 17% mortality in the Leupeptin diet group. The difference was significant (P<0.05). At the end of the experiment, a comparison was made of tumor development in the two groups. Significantly fewer tumors and lower tumor weight per rat were observed and the incubation time was longer in the Leupeptin diet group than in the control diet group.In the second experiment, rats were injected with 2 mg of DMBA at 55 and 58 days of age. The periods of feeding with the Leupeptin diet were as follows;L-1 group; the same period as in the first experimentL-2 group; 40 days after DMBA treatmentL-3 group; 7 days before DMBA treatment The L-2 group showed significant decrease of tumor number and prolongation of incubation time as compared with the control group. Tumor development in the L-3 group was similar to that in the control diet group.In the third experiment, protease activities of rat mammary fat pads were assayed and compared 3,7,14,21 and 28 days after DMBA treatment. A 2500×g pellet was active for neutral protease digesting Calf thymus histon and the 20000×g supernatant for Cathepsin D and caseinolytic activities. Caseinolytic activity at 3 days and neutral protease activities at 3,7 and 21 days were significantly decreased in the Leupeptin diet group. However, caseinolytic activity was significantly increased only at 21 days in the Leupeptin diet group. No differencein Cathepsin D activities was observed between the control and Leupeptin diet groups.Microscopic examination of established mammary tumors showed that majority were papillary or tubular adenocarcinoma, and no differences in histological features were observed between the two groups.The results indicate that the administration of Leupeptin inhibits rat mammary carcinogenesis, especially when Leupeptin is given during DMBA induction of mammary tumor, and further the effect of Leupeptin is considered to be one of promotion rather than initiation. In addition, it seems that proteases which digest casein and Calf thymus histon play an important role in rat mammary carcinogenesis.