为揭示LH/hCG受体和cAMP在玉米赤霉烯酮(ZEN)抑制小鼠睾丸间质细胞(leydig cells)分泌睾酮中的作用,本研究以原代培养的小鼠Leydig细胞为材料,用ELISA法分别测定ZEN暴露浓度为0、1、5、10、20 μg·mL-1及暴露时间为1、6、12、24h时睾酮的分泌量(hCG环境及cAMP环境)和胞质内cAMP水平(hCG环境),用化学发光标记免疫分析法(CLIA)测定其细胞表面的LH/hCG受体结合力,用Western Blot法测定LH/hCG受体蛋白的表达量.结果表明,hCG环境中高浓度及长时间ZEN暴露对睾酮分泌的抑制效应极显著(P＜0.01),而cAMP环境中,不同浓度及时间的ZEN暴露均可显著抑制Leydig细胞分泌睾酮(P＜0.05),且浓度及时间-效应关系明显.不同浓度及时间的ZEN暴露对Leydig细胞LH/hCG受体的结合力及表达量的影响不显著(P＞0.05),而低浓度及长时间ZEN暴露使胞质内cAMP水平显著升高(P＜0.05).结果显示,玉米赤霉烯酮可通过cAMP途径抑制小鼠Leydig细胞分泌睾酮,与LH/hCG受体途径无关.%To reveal the reproductive toxicity mechanism of zearalenone(ZEN) to male animals,the role of LH/hCG receptor and cAMP in zearalenone inhibiting testosterone secretion in mouse Leydig cells has been researched.Primary Leydig cells in vivo were taken to determine the testosterone level (hCG or cAMP stimulated) and cAMP level (hCG stimulated) after exposed to 0,1,5,10,20 μg · mL-1 zearalenone for 12 h or to 2.5 μg · mL-1 for 1,6,12,24 h by ELISA,determined binding activity of LH/hCG receptor by Chemiluminescence Immunoassay (CLIA),and determined LH/hCG protein expression level by Western Blot.The results showed that in hCG environment long time and high dose zearalenone exposure inhibited testosterone secretion extremely significant (P ＜ 0.01),in cAMP environment every dosage and time exposure for zearalenone reduced testosterone secretion significant (P＜0.05),and the time-or concentrationeffect relationship was obvious.Every exposure condition effected neither LH/hCG receptor binding activity nor protein expression obviously (P＞0.05),and low concentration and long time zearalenone exposure could increase the intercellular cAMP level significantly (P＜0.05).These results indicate that zearalenone can inhibit testosterone secretion through cAMP pathway but not LH/hCG pathway.
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