Derivatives of dehydrocrotonin, a diterpene lactone isolated from Croton cajucara: cytotoxicity in rat cultured hepatocytes and in V79 cells.
Derivatives of dehydrocrotonin (DHC; Compound I) with different anti-ulcerogenic properties but less toxicity were produced by reducing the cyclohexenone moiety of DHC with NaBH4 (Compound II), reducing the cyclohexenone and lactone moieties with LiAlH4 (Compound III) and transforming the lactone moiety into an amide (Compound IV) using dimethylamine. Derivatives of DHC were assayed in cultured hepatocytes and V79 fibroblasts. Three independent endpoints assays for cytotoxicity were used, namely, the DNA content, tetrazolium reduction (MTT) and neutral red uptake (NRU). Compound III was less toxic than the other DHC derivatives in both cell cultures. ICso values ranging from 250 to 600 microM were obtained for Compounds II and IV in the NRU and DNA content tests evaluated in 4-hour hepatocyte cultures. Although Compound II showed relatively low cytotoxicity in rat hepatocytes based on the NRU and DNA content assays, a very high toxicity (IC50=10 microM) was observed in the MTT test. Metabolites of Compound II in conditioned medium from 4-hour old hepatocyte cultures enhanced the MTT-reducing ability of V79 fibroblasts. The cytotoxicity of the derivatives was greater in recently isolated hepatocytes (only a 4-hour incubation for cell attachment prior to treating with the derivatives) than in hepatocytes previously cultured (24-hour incubation) before the treatment. Thus, aging reduced the cytotoxic effects of DHC derivatives in isolated hepatocytes, suggesting that P450-mediated biotransformation of such derivatives may lead to the formation of more toxic metabolites.
Departamento de Bioquímica, Instituto de Biologia, Universidade Estadual de Campinas, SP, Brazil. pmelo@unicamp.br
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D000818:Animals; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects; D016922:Cellular Senescence / Q000502:physiology; D002927:Cimetidine / Q000494:pharmacology; D006224:Cricetinae; D003412:Cricetulus; D029802:Croton; D017077:Culture Media, Conditioned / Q000494:pharmacology; D004247:DNA / Q000032:analysis; D004224:Diterpenes / Q000008:administration & dosage / Q000633:toxicity*; D045785:Diterpenes, Clerodane; D004305:Dose-Response Relationship, Drug; D005347:Fibroblasts / Q000187:drug effects* / Q000378:metabolism / Q000473:pathology; D022781:Hepatocytes / Q000187:drug effects* / Q000378:metabolism / Q000473:pathology; D008297:Male; D009499:Neutral Red / Q000378:metabolism; D010946:Plants, Medicinal; D051381:Rats; D017208:Rats, Wistar; D013385:Succinate Dehydrogenase / Q000378:metabolism; D013778:Tetrazolium Salts / Q000378:metabolism; D013844:Thiazoles / Q000378:metabolism
D017077:Culture Media, Conditioned; D004224:Diterpenes; D045785:Diterpenes, Clerodane; D013778:Tetrazolium Salts; D013844:Thiazoles; C103060:dehydrocrotonin; D009499:Neutral Red; D002927:Cimetidine; D004247:DNA; D013385:Succinate Dehydrogenase; C022616:thiazolyl blue