Pisani Cédric, Voisin Sébastien, Arafah Karim, Durand Philippe, Perrard Marie-Hélène, Guichaoua Marie-Roberte, Bulet Philippe, Prat Odette
CEA, Direction de la Recherche Fondamentale, BIAM-Marcoule, Bagnols-sur-Cèze, France.
Plateforme BioPark d'Archamps, Archamps Technopole, Saint Julien en Genevois, France.
ALTEX. 2016;33(4):393-413. doi: 10.14573/altex.1601253. Epub 2016 May 8.
To measure the testicular toxicity of two fungicides (carbendazim and iprodione), alone or in a mixture, we used a rat ex vivo model of seminiferous tubules, greatly reducing the number of rodents used, in accordance with the 3R rule (Replacement, Reduction, and Refinement). This model allows the representation of puberty, a critical life period with regard to endocrine disruptors. The cellular modifications were followed for three weeks through transcriptomic and proteomic profiling analysis. A quantitative and comparative method was developed to estimate how known pathways were disturbed by each substance. This pathway-driven analysis revealed a strong alteration of steroidogenesis and an impairment of meiosis in all cases, albeit the initial molecular events were different for both substances. The ex vivo cytogenetic analysis confirmed that both fungicides alter the course of the first meiotic prophase. In addition, the mixture of both substances triggered effects greater than the sum of their cumulative effects and compromised future sperm motility after a shorter time of exposure compared with the fungicides tested separately. The alliance of an ex vivo culture with "omics" strategies complemented with a physiological examination is a powerful combination of tools for testing substances, separately or in a mixture, for their testicular toxicity. In particular, proteomics allowed the identification of systematically differentially expressed proteins in the secretomes of exposed cultures, such as FUCO and PEBP1, two proteins linked with the motility and fertilizing ability of spermatozoa, respectively. These proteins may be potential biomarkers of testicular dysfunction and infertility.
为了测量两种杀菌剂（多菌灵和异菌脲）单独或混合使用时的睾丸毒性，我们使用了大鼠生精小管的离体模型，根据3R原则（替代、减少和优化）大幅减少了所用啮齿动物的数量。该模型能够模拟青春期，这是一个对于内分泌干扰物而言至关重要的生命阶段。通过转录组学和蛋白质组学分析对细胞变化进行了为期三周的跟踪。开发了一种定量和比较方法来估计每种物质对已知途径的干扰程度。这种基于途径的分析揭示了在所有情况下类固醇生成的强烈改变和减数分裂的受损，尽管两种物质的初始分子事件有所不同。离体细胞遗传学分析证实，这两种杀菌剂都会改变第一次减数分裂前期的进程。此外，与单独测试的杀菌剂相比，两种物质的混合物引发的效应大于其累积效应之和，并且在较短的暴露时间后就损害了未来精子的活力。将离体培养与“组学”策略相结合并辅以生理检查，是一种用于单独或混合测试物质睾丸毒性的强大工具组合。特别是，蛋白质组学能够识别暴露培养物分泌组中系统差异表达的蛋白质，例如FUCO和PEBP1，这两种蛋白质分别与精子的活力和受精能力相关。这些蛋白质可能是睾丸功能障碍和不育的潜在生物标志物。
