Inhibitory effects of epidermal growth factor on progesterone production of ovarian granulosa cells in Tsaiya duck (Anas platyrhynchos var. domestica).

1. The purpose of this study was to investigate the effects of epidermal growth factor (EGF) on progesterone (P(4)) secretion from ovarian granulosa cells in Tsaiya ducks (Anas platyrhynchos var. domestica). 2. We obtained the largest (F1) follicle from Tsaiya duck, the granulosa layer was separated and the cells were isolated according to their proximity to the germinal disc. 3. The granulosa cells were cultured in vitro, the culture media and the cells were used to determine P(4) and steroidogenic enzyme concentrations, respectively. 4. P(4) concentrations were decreased in cultured granulosa cells taken proximal to the germinal disc (GD) compared to those distal to the germinal disc (NGD). 5. EGF inhibited both basal and ovine luteinising hormone (oLH)-induced P(4) concentrations. It also inhibited the P(4) secretion via protein kinase A (PKA) pathway when cultured with GD and NGD granulose cells (mixed together) in vitro. 6. Western blot results showed decreased concentrations of cytochrome P450 side-chain cleavage (P450scc) enzyme, 3ß-hydroxysteroid dehydrogenase (3ß-HSD) enzyme and steroid acute regulatory (StAR) protein expression, when the cells were co-treated with EGF and oLH. 7. The inhibitory effect of oLH-induced P(4) production was attenuated by EGF by the addition of MAP-erk kinase (MEK) inhibitor (PD98059) suggests EGF may inhibit P(4) production by affecting via the mitogen-activated protein kinase (MAPK) pathway.

Taxol induces concomitant hyperphosphorylation and reorganization of vimentin intermediate filaments in 9L rat brain tumor cells.

Taxol, a microtubule stabilizing agent, has been extensively investigated for its antitumor activity. The cytotoxic effect of taxol is generally attributed to its antimicrotubule activity and is believed to be cell cycle dependent. Herein, we report that taxol induces hyperphosphorylation and reorganization of the vimentin intermediate filament in 9L rat brain tumor cells, in concentration- and time-dependent manner. Phosphorylation of vimentin was maximum at 10(-6) M of taxol treatment for 8 h and diminished at higher (10(-5) M) concentration. Enhanced phosphorylation of vimentin was detectable at 2 h treatment with 10(-6) M taxol and was maximum after 12 h of treatment. Taxol-induced phosphorylation of vimentin was largely abolished in cells pretreated with staurosporine and bisindolymaleimide but was unaffected by H-89, KT-5926, SB203580, genistein, and olomoucine. Thus, protein kinase C may be involved in this process. Hyperphosphorylation of vimentin was accompanied by rounding up of cells as revealed by scanning electron microscopy. Moreover, there was a concomitant reorganization of the vimentin intermediate filament in the taxol-treated cells, whereas the microtubules and the actin microfilaments were less affected. Taken together, our data demonstrate that taxol induces hyperphosphorylation of vimentin with concomitant reorganization of the vimentin intermediate filament and that this process may be mediated via a protein kinase C signaling pathway.