RESUMO
For estrogen-responsive B-1F cells, established from estrogen-responsive mouse Leydig cell tumor, it has been reported that the 5-lipoxygenase (5-LOX) metabolic pathway appears to be associated with cell growth. The addition of 5-LOX inhibitor 2-(12-hydroxydodeca-5,10-diyl)-3,5,6-trimethyl-1,4-benzoquinone (AA861) to the medium resulted in a dose-dependent increase in cell yield as described previously. When the growth of the palpable tumors was measured, AA861 had stimulated in vivo tumor growth in adult male mouse inoculated B-1F cells. The effects of AA861 and 17beta-estradiol (E2) on the contents of various arachidonic acid metabolites in B-1F cells and their conditioned medium were examined. Although AA861 and E2 decreased the contents of leukotrienes (LTs), the two did not significantly change those of prostaglandins, thromboxan, prostacyclin, 12-hydroxyeicosatetraenoic acid (HETE) and 15-HETE. In immunohistochemical study B-1F cells show positive staining for 5-LOX in the E2-depleted condition, while E2 decreased the expression of 5-LOX. The decrease of the intensities of 79-kDa 5-LOX protein and 403-bp RT-PCR product bands was observed. The growth of Morpholino-anti oligo delivered B-1F cells was higher than that of Standard control oligo delivered cells. The delivery of Morpholino-anti oligo into B-1F cells caused the decrease of contents of LTs and 5-HETE in the cells and medium, and the reduction of 5-LOX activity. When LTD4 was added in the culture medium, the increasing concentrations of LTD4 resulted in a significant inhibition of cell yields of E2-treated B-1F cells. Morphological changes such as nuclear condensation and fragmentation, and DNA ladder pattern were demonstrated in E2-stimulated B-1F cells treated with LTD4 as well as in control cells cultured in the basal medium. These results implicate that 5-LOX at least plays an important role in the growth of B-1F cells and LD4 induces the apoptosis of B-1F cells.
Assuntos
Apoptose/efeitos dos fármacos , Estradiol/farmacologia , Leucotrieno D4/farmacologia , Tumor de Células de Leydig/patologia , Animais , Araquidonato 5-Lipoxigenase/biossíntese , Araquidonato 5-Lipoxigenase/genética , Araquidonato 5-Lipoxigenase/metabolismo , Ácido Araquidônico/antagonistas & inibidores , Ácido Araquidônico/metabolismo , Benzoquinonas/farmacologia , Processos de Crescimento Celular/efeitos dos fármacos , Tumor de Células de Leydig/tratamento farmacológico , Tumor de Células de Leydig/metabolismo , Inibidores de Lipoxigenase , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Oligonucleotídeos Antissenso/genética , Oligonucleotídeos Antissenso/farmacologia , RNA Mensageiro/biossíntese , RNA Mensageiro/genética , Células Tumorais CultivadasRESUMO
SC-3 is a cloned cell line derived from an androgen-dependent mouse mammary tumor (Shionogi Carcinoma 115). A physiological level of androgen stimulates the growth of SC-3 cells through the production of androgen-induced growth factor. Methylcobalamin (MeCbl), one of the active cobalamins, inhibits the growth of SC-3 cells stimulated by androgen. It is known that apoptosis has an important role in tumor growth. The specific aim of this study is to examine the effects of MeCbl, in the presence of androgen, on apoptosis in SC-3 cells. Morphological analysis revealed budding nuclei and chromatin condensation in cells cultured with MeCbl, but few in cells cultured without MeCbl. Low molecular weight DNA extracted from cells cultured with or without MeCbl was analysed by gel electrophoresis. A characteristic nucleosomal size ladder was detected in the culture with MeCbl. The terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate-biotin nick end-labelling method was also used to evaluate apoptotic cell death in SC-3 cells. Apoptosis was observed more frequently in SC-3 cells treated with MeCbl than in those without MeCbl. These results demonstrate that androgen-dependent SC-3 cells undergo apoptosis by MeCbl even if in the presence of androgen.
