Expression and activity of microsomal epoxide hydrolase in follicles isolated from mouse ovaries
Expression and activity of microsomal epoxide hydrolase in follicles isolated from mouse ovaries
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DOI：
10.1093/toxsci/68.1.24
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发表时间：
2002-07-01
影响因子：
3.8
通讯作者：
Hoyer, PB
中科院分区：
文献类型：
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作者：
Cannady, EA;Dyer, CA;Hoyer, PB
微粒体环氧化物水解酶(Meh)参与外源物质的解毒作用，这些外源物质是或可以形成环氧化物代谢物，包括卵毒物质4-乙烯基环己烯(VCH)。这种工业化学品被肝脏的细胞色素P450激活为二环氧化物的代谢物VCD，它破坏小鼠腔前卵泡(F1)。由于卵巢mEH可能在VCD的解毒中起作用，这些研究调查了mEH在分离的卵巢部分的表达和活性。小鼠每天ip VCH(7.4 mmol/kg/d)或VCD(0.57 mmol/kg/d)，4h后摘除卵巢，分离不同数量的完整卵泡(F1，25-100um；F2,100-250um；F3，&Gt；250um)和间质细胞(Int)，提取总RNA和蛋白质。用实时定量聚合酶链式反应和底物二苯乙烯氧化物(CSO；12.5um)分别检测mEH的表达和比活性。共聚焦显微镜观察mEH蛋白在卵巢中的分布。重复给予VCH或VCD后，F1(410+/-5%VCH；292+/-5%VCD)和F2(1379+/-4%VCH；381+/-11%VCD)卵泡的meH基因表达增加。重复给予VCH/VCD(381+/-11%VCH；384+/-27%VCD)后，F1卵泡MeH的催化活性增强。共聚焦显微镜下可见meh蛋白分布于卵巢各处，以间质细胞和膜细胞染色强度最强，且随着剂量的增加(56+/-0.8%VCH和29+/-0.9%VCD)染色增强。我们的结论是，MeH在小鼠卵巢卵泡中表达并具有功能。此外，VCH和VCD的体内剂量会影响这些参数。
Microsomal epoxide hydrolase (mEH) is involved in the detoxification of xenobiotics that are or can form epoxide metabolites, including the ovotoxicant, 4-vinylcyclohexene (VCH). This industrial chemical is bioactivated by hepatic CYP450 to the diepoxide metabolite, VCD, which destroys mouse small preantral follicles (F1). Since ovarian mEH may play a role in VCD detoxification, these studies investigated the expression and activity of mEH in isolated ovarian fractions. Mice were given 1 or 15 daily doses (ip) of VCH (7.4 mmol/kg/day) or VCD (0.57 mmol/kg/day); 4 h following the final dose, ovaries were removed, distinct populations of intact follicles (F1, 25-100 mum; F2, 100-250 mum; F3, > 250 mum) and interstitial cells (Int) were isolated, and total RNA and protein were extracted. Real-time polymerase chain reaction and the substrate cis-stilbene oxide (CSO; 12.5 muM) were used to evaluate expression and specific activity of mEH, respectively. Confocal microscopy evaluated ovarian distribution of mEH protein. Expression of mRNA encoding mEH was increased in F1 (410 +/- 5% VCH; 292 +/- 5% VCD) and F2 (1379 +/- 4% VCH; 381 +/- 11% VCD) follicles following repeated dosing with VCH or VCD. Catalytic activity of mEH increased in F1 follicles following repeated dosing with VCH/VCD (381 +/- 11% VCH; 384 +/- 27% VCD). Visualized by confocal microscopy, mEH protein was distributed throughout the ovary with the greatest staining intensity in the interstitial cells and staining in the theca cells that was increased by dosing (56 +/- 0.8% VCH; 29 +/- 0.9% VCD). We conclude that mEH is expressed and is functional in mouse ovarian follicles. Additionally, in vivo dosing with VCH and VCD affects these parameters.