| Fenvalerate, a synthetic pyrethroid, is widely used in agricultural and other domestic applications in China. Fenvalerate has been found to cause severe neurotoxic effects. However, Fenvalerate has been suspected as one kind of endocrine disrupting chemicals(EDCs) in recent years. EDCs are known to act at multiple sites through multiple modes of action. It is necessary to explore the mechanisms of action from multiple viewpoints.Intracellular Ca2+ homeostasis and the messenger role of the Ca2+ ion in the regulation and control of cell functions have been a very active and fruitful areas of basic biological research. The chemical-physical events mediated by calcium are various such as the release of neurotransmitter, steroidogenesis, fertilization, and synthesis of DNA, etc. Based on thecentral role of the Ca2+ messenger system in these aspects of cell functions, it is logical to examine possible disturbances in Ca2+ homeostasis and Ca2+-mediated functions as underlying mechanisms of toxicant action.To our knowledge, no studies have been carried out to address the effects of pyrethroid on ovarian calcium homeostasis. If fenvalerate directly or indirectly, or secondarily alter Ca2+ homeostasis in ovarian cells, it may impair the reproductive endocrine functions. Furthermore, accumulating evidences indicate calcium ions modulate gonadotropin-stimulated steroidogenesis by granulosa cells.In the present work, we first investigatd the effects of fenvalerate on calcium homeostasis in ovary both in vivo and in vitro. And then, we studied the effects of fenvalerate on serum steroid levels in female SD rats exposed for 30 days. To further study the relationship between fenvalerate and FSH-stimulated progesterone biosynthesis, we used primarily cultured human ovarian granulosa-luteinizing cell (hGLCs) model, which is undertaking multiple and important functions in ovary involving steroidogenesis, oocyte development and so on.1. The effect of fenvalerate on calcium homeostasis in ovaryThe effects of fenvalerate on calcium homeostasis in ovary were studied. In vivo study, female Sprague-Dawley rats were treated withfenvalerate at doses of 0.00, 1.91, 9.55 and 31.80mg/kg for 30 days. Compared with the control, an significant increase in the serum free calcium concentrations in rats treated with 31.8mg/kg fenvalerate was observed (P<0.05). The activities of phosphorylase a, a valid indicator of fluctuations in cytosolic Ca2+ level, were enhanced in rat ovary in all treated groups. Another calcium related enzyme, Ca2+-ATPase, was inhibited by fenvalerate. Calmodulin, the primary calcium receptor, was measured in rat ovary by ELISA at each stage of estrous cycle. The results showed that fenvalerate increased the total concentration of calmodulin in estrus period. In vitro study, the effects of fenvalerate on calcium homeostasis in hGLCs were studied. Fenvalerate (20.0u mol/L, 2.0u mol/L) induced the increases of [Ca2+]i in hGLCs by using a fluorescent Ca2+ indicator fluo-3/AM. This [Ca2+]i increase mostly resulted from Ca2+ influx in the studied concentration. The total concentration of CaM (free and bound fraction) in hGLCs induced by fenvalerate at doses of 5, 25 and 125 u M were significantly increased by a method of immunofluorescence. The relative activitities of CaM (bound fraction) in hGLCs incubated with fenvalerate for 24h with trifluoperazine as fluoresence probe were enhanced by flow cytometry analysis. In conclusion, fenvalerate interferes with calcium homeostasis in rat ovary and hGLCs. 2. The effect of fenvalerate on FSH-stimulated steroidogenesisIn this study, we first examined serum estradiol and progesterone level in female SD rats after fenvalerate exposure (0.00, 1.91, 9.55, 31.80mg/kg) for 30 days. During the study period, daily vaginal smears were taken to monitor estrous cycle. A decrease in serum progesterone (P4) concentration and a increase in serum estradiol (E2) concentration in rats treated with fenvalerate which were in diestrus were observed. To access whether it is relevant to steroi... |
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