薏苡麩皮及薏苡殼甲醇萃取物對大鼠卵巢顆粒細胞分泌孕酮之效應

Effects of Methanol Extract of Adlay Bran and Adlay Hull on Progesterone Release in Rat Ovarian Granulosa Cells

夏詩閔(Shih-Min Hsia)

指導教授 ： 江文章

共同指導教授 ： 王錫崗

國立臺灣大學/生物資源暨農學院/食品科技研究所/博士(2006年)

https://doi.org/10.6342/NTU.2006.01185

摘要

薏苡為禾本科一年生草本植物，在中國傳統藥書中對於其功效有很多的記載。在日本有研究顯示，薏苡中具有一種可以誘發女性排卵的化學物質，其他研究也顯示薏苡對於女性排卵障礙有其治療的效果，故推測可能與生殖內分泌有重要的關係。但是目前薏苡萃取物對於性腺類固醇激素的影響及作用機制，瞭解非常有限。本實驗計畫希望深入探討薏苡萃取物對性腺細胞類固醇激素合成與分泌之影響，並探討薏苡萃取物對性腺細胞類固醇激素之作用機轉。實驗將依以下步驟執行，包括:[1] 雌鼠顆粒細胞 (Granulosa cell)與薏仁萃取物，人絨性促素 (hCG)，或forskolin、SQ22536、8-Br-cAMP 等共同培養，培養液中助孕酮 (progesterone) 或pregnenolone 的濃度均以放射免疫測定法(radioimmunoassay，RIA) 檢測；[2] 在機轉的研究方面，給予類固醇激素之前驅物：25-OH-cholesterol、pregnenolone，探討類固醇激素合成酵素，StAR protein及P450scc的活性是否會受到薏仁萃取物之影響；[3] 利用西方點墨法 (Western blotting) 及RT-PCR之方式偵測類固醇激素合成相關蛋白P450scc、StAR之蛋白質表現及基因表現。由實驗結果發現薏苡麩皮甲醇萃取物正丁醇區分層(ABM-Bu)具有抑制助孕酮分泌之效果，其可能的抑制機轉: (一) 抑制cAMP-PKA路徑；(二) 抑制P450scc及3β-HSD之活性；(三) StAR蛋白之蛋白質表現及mRNA表現。本計劃所獲成果可以更進一步證實薏苡萃取物確實對於女性內分泌具有影響的效果。人體一些疾病可能由於內分泌失調的結果，如女性常見的乳癌可能是因為助孕酮分泌過多所導致，而本研究結果發現薏苡萃取物可以抑制助孕酮之分泌，因此可能可以將薏苡開發為一特殊機能性之保健食品，亦可以利用此實驗模式來進行活性測試，進而分離純化出一些具有抑制或促進助孕酮分泌之純化合物，進行新藥的開發工作。

關鍵字

薏苡 ； 顆粒細胞 ； 孕酮 ； 雌二醇

並列摘要

Adlay (Coix lachryma-jobi L. var. ma-yuen Stapf.) has long been used as traditional Chinese medicine for the dysfunction of endocrine system and inflammation condition. However, the effect of adlay seed on the endocrine system has not yet been reported. In the present study, the effects and the mechanisms of methanolic extract of adlay bran (ABM) on progesterone synthesis in rat granulosa cell were studied. ABM was further partitioned with different solvent including water, 1-butanol, ethyl acetate and n-hexane. Four subfractions named ABM-Wa (water fraction), ABM-Bu (1-butanol fraction), ABM-EA (ethyl acetate fraction) and ABM-Hex (n-hexane fraction) were obtained. ABM-Bu was further fractionated using Diaion HP-20 resin column chromatography with gradient elution. Granulosa cells were prepared from pregnant mare serum gonadotropin-primed immature female rats and challenged with different reagents including human chorionic gonadotropin (hCG, 0.5 IU/ml), forskolin (10 μM), 8-bromo-adenosine-3’,5’-cyclic monophosphate (8-Br-cAMP, 1 mM), A23187 (10 μM), phorbol 12-myristate 13-acetate (PMA, 0.01 μM), 25-OH-cholesterol (0.1-10 μM) and pregnenolone (0.1-10 μM) in the presence or absence of ABM-Bu (100 μg/ml). The functions of steroidogenic enzyme including protein expression of the steroidogenic acute regulatory protein (StAR) and cytochrome P450 side chain cleavage enzyme (P450scc) protein were investigated. Both P450scc and StAR mRNA expressions have also been explored. We found that ABM decreased progesterone production via an inhibition on (1) the cAMP-PKA Uand PKCU signal transduction pathway, (2) P450scc and 3β-hydroxysteroid dehydrogenase (3β-HSD) enzyme activity, (3) P450scc and StAR protein and mRNA expressions, and (4) the phosphorylation of ERK1/2 in rat granulosa cells.

並列關鍵字

adlay ； granulosa cell ； progesterobe ； estradiol

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