Witek Patrycja, Grzesiak Malgorzata, Kotula-Balak Malgorzata, Koziorowski Marek, Slomczynska Maria, Knapczyk-Stwora Katarzyna
Department of Endocrinology, Institute of Zoology and Biomedical Research, Jagiellonian University, Gronostajowa 9, 30-387 Krakow, Poland.
University Centre of Veterinary Medicine, University of Agriculture in Krakow, 30-059, Krakow, Poland.
Theriogenology. 2021 Jan 1;159:45-52. doi: 10.1016/j.theriogenology.2020.10.010. Epub 2020 Oct 8.
Recently, we have demonstrated that neonatal exposure to environmental endocrine-active compounds (EACs) with androgenic/antiandrogenic and estrogenic/antiestrogenic activities led to morphological and functional changes in the porcine corpus luteum (CL). To gain insight into the regulatory mechanisms of the long-term effects of EACs, we analyzed the impact of neonatal exposure of such compounds on global DNA methylation and the expression of miRNA biogenesis components in the porcine CL. Piglets were injected subcutaneously with testosterone propionate (TP, an androgen), flutamide (FLU, an antiandrogen), 4-tert-octylphenol (OP, an estrogenic compound), ICI 182,780 (ICI, an antiestrogen), methoxychlor (MXC, a compound with mixed activities) or corn oil (control) between postnatal days 1 and 10 (n = 5/group). The CLs from sexually mature gilts were examined for global DNA methylation and for the abundance of proteins related to DNA methylation (DNMT1, DNMT3A and DNMT3B) and miRNA biogenesis (DROSHA, XPO5, DICER1, AGO2) using an enzyme-linked immunosorbent assay, Western blotting and immunohistochemical staining. ICI and MXC increased the global DNA methylation levels and DNMT1 protein abundance in the luteal tissue. OP treatment led to a lower DROSHA protein abundance, while ICI treatment resulted in a greater DROSHA protein abundance. Both FLU and ICI increased DICER1 protein abundance in the luteal tissue. In addition, XPO5 showed immunolocalization exclusively in small luteal cells in the OP-treated pigs, in contrast to localization in both small and large luteal cells in the controls. In conclusion, the changes in DNA methylation, as well as the altered miRNA biogenesis components, seem to be a part of the regulatory network that mediates the long-term effects of EACs on CL function in pigs.
最近，我们已经证明，新生仔猪暴露于具有雄激素/抗雄激素和雌激素/抗雌激素活性的环境内分泌活性化合物（EACs）会导致猪黄体（CL）的形态和功能发生变化。为了深入了解EACs长期影响的调控机制，我们分析了新生仔猪暴露于此类化合物对猪CL中整体DNA甲基化和miRNA生物合成成分表达的影响。在出生后第1天至第10天之间，给仔猪皮下注射丙酸睾酮（TP，一种雄激素）、氟他胺（FLU，一种抗雄激素）、4-叔辛基苯酚（OP，一种雌激素化合物）、ICI 182,780（ICI，一种抗雌激素）、甲氧滴滴涕（MXC，一种具有混合活性的化合物）或玉米油（对照）（每组n = 5）。使用酶联免疫吸附测定、蛋白质印迹和免疫组织化学染色，对性成熟后备母猪的CL进行整体DNA甲基化以及与DNA甲基化（DNMT1、DNMT3A和DNMT3B）和miRNA生物合成（DROSHA、XPO5、DICER1、AGO2）相关的蛋白质丰度检测。ICI和MXC提高了黄体组织中的整体DNA甲基化水平和DNMT1蛋白丰度。OP处理导致DROSHA蛋白丰度降低，而ICI处理导致DROSHA蛋白丰度增加。FLU和ICI均增加了黄体组织中DICER1蛋白丰度。此外，与对照组中小黄体细胞和大黄体细胞均有定位不同，XPO5在OP处理的猪中仅在小黄体细胞中显示免疫定位。总之，DNA甲基化的变化以及miRNA生物合成成分的改变似乎是介导EACs对猪CL功能长期影响的调控网络的一部分。
