Ptak Anna, Ludewig Gabriele, Kapiszewska Maria, Magnowska Zofia, Lehmler Hans-Joachim, Robertson Larry W, Gregoraszczuk Ewa L
Laboratory of Physiology and Toxicology of Reproduction, Department of Animal Physiology, Institute of Zoology, Jagiellonian University, Ingardena 6, Krakow, Poland.
Toxicol Lett. 2006 Oct 25;166(3):200-11. doi: 10.1016/j.toxlet.2006.07.304. Epub 2006 Jul 14.
Polychlorinated biphenyl (PCBs) levels of tens and hundreds of pg/ml for individual congeners are measured in human follicular fluid. PCB3 (4-chlorobiphenyl), caused a significant increase in estradiol secretion in porcine granulose-theca cell co-cultures and its two metabolites, 4-OH-PCB3 and 3,4-diOH-PCB3, were even more potent than PCB3 itself [Ptak, A., Ludewig, G., Lehmler, H.J., Wojtowicz, A.K., Robertson, L.W., Gregoraszczuk, E.L. 2005. Comparison of the actions of 4-chlorobiphenyl and its hydroxylated metabolites on estradiol secretion by ovarian follicles in primary cells in culture. Reprod. Toxicol. 20, 57-64]. The question is whether these follicle cells are potentially able to metabolize PCB3 to hydroxylated and genotoxic or cytotoxic intermediates. We report here that granulose-theca co-cultures express xenobiotic-metabolizing cytochrome P450 activities, with CYP1A1>CYP2B>>CYP1A2. A significant increase in CYP1A1 and 2B, but not CYP1A2, activity was seen in cells that were exposed to 6 ng/ml PCB3 or 20 nM 17-beta-estradiol. An increase in caspase-3 activity, indicative for apoptosis, was only observed in PCB3-exposed cells after 24 h exposure. Genotoxicity, determined with the Comet assay, was initially reduced after 24 h exposure to PCB3 and both metabolites compared to untreated controls, followed by a significant transient increase in Comets at the 4 and 24 h time point with PCB3 and 4-OH-PCB3. 3,4-diOH-PCB3 induced a significant increase only after 72 h of recovery. We hypothesize that these biphasic damage kinetics may be due to cross-links caused by adduct formation. These results show for the first time that granulose-theca cells in co-culture express CYP1A1, 2B and 1A2 activities and that PCBs at concentrations that are reached in the environment induce genotoxicity in granulosa cells.
在人类卵泡液中检测到单个多氯联苯（PCB）同系物的含量为数十至数百皮克/毫升。PCB3（4-氯联苯）可使猪颗粒-卵泡膜细胞共培养物中的雌二醇分泌显著增加，其两种代谢产物4-羟基-PCB3和3,4-二羟基-PCB3的作用甚至比PCB3本身更强[Ptak, A., Ludewig, G., Lehmler, H.J., Wojtowicz, A.K., Robertson, L.W., Gregoraszczuk, E.L. 2005. 4-氯联苯及其羟基化代谢产物对培养的原代细胞中卵巢卵泡雌二醇分泌作用的比较。生殖毒理学。20, 57 - 64]。问题在于这些卵泡细胞是否有可能将PCB3代谢为羟基化且具有遗传毒性或细胞毒性的中间体。我们在此报告，颗粒-卵泡膜细胞共培养物表达外源性物质代谢细胞色素P450活性，其中CYP1A1 > CYP2B >> CYP1A2。在暴露于6纳克/毫升PCB3或20纳摩尔17-β-雌二醇的细胞中，CYP1A1和2B的活性显著增加，但CYP1A2的活性未增加。仅在暴露于PCB3 24小时后的细胞中观察到caspase-3活性增加，这表明细胞发生凋亡。与未处理的对照相比，在暴露于PCB3及其两种代谢产物2小时后，用彗星试验测定的遗传毒性最初降低，随后在4小时和24小时时间点，PCB3和4-羟基-PCB3导致彗星数量显著短暂增加。3,4-二羟基-PCB3仅在恢复72小时后才导致显著增加。我们推测这些双相损伤动力学可能是由于加合物形成导致的交联所致。这些结果首次表明，共培养的颗粒-卵泡膜细胞表达CYP1A1、2B和1A2活性，并且环境中达到的浓度的多氯联苯会在颗粒细胞中诱导遗传毒性。
