Atli Mehmet O, Kose Mehmet, Hitit Mustafa, Kaya M Salih, Bozkaya Faruk
Department of Obstetrics and Gynaecology, Faculty of Veterinary Medicine, Dicle University, Diyarbakır, Turkey.
Department of Obstetrics and Gynaecology, Faculty of Veterinary Medicine, Dicle University, Diyarbakır, Turkey.
Theriogenology. 2018 Apr 15;111:25-33. doi: 10.1016/j.theriogenology.2018.01.010. Epub 2018 Jan 11.
The aim of this study was to elucidate the expression profiles of Toll-like receptors (TLRs) in the ovine corpus luteum (CL) during early pregnancy and prostaglandin F2α (PGF2α)-induced luteolysis. For this purpose, multiparous Anatolian Merino ewes were selected and randomly allotted into cyclic (including those in the induced luteolysis group, n = 20) and pregnant (n = 12) groups. All of the ewes were scheduled to be slaughtered for predetermined days/hour during the estrous cycle, early pregnancy, and PGF2α induced luteolysis. The CLs were collected from both cyclic and pregnant ewes on days 12 (C12 and P12; n = 8) and 16 (C16 and P16; n = 8) and pregnant ewes on day 22 (P22; n = 4). For the induced luteolysis model, ewes were injected with PGF2α on day 12 of the estrous cycle and CLs were collected at 1 h (PG1h; n = 4), 4 h (PG4h; n = 4), and 16 h (PG16h, n = 4) after injection. Quantitative polymerase chain reaction (qPCR) was used to evaluate the expression profiles of TLR2, TLR4, TLR6, TLR8, and TLR10, while free-floating in situ hybridization and immunohistochemistry were used to define the spatial localization of TLR2, TLR4, and TLR7 in the CL. Data were then analyzed by one-way ANOVA and were considered statistically significant when P values were lower than 0.05. Expression of TLR2 was upregulated in both early and late stages of luteolysis (P < .05). An upregulation of TLR4 was detected at PG16h, while TLR6 was decreased at PG4h (P < .05). Expression of TLR7 and TLR8 was significantly increased during early pregnancy, at both PG16h and regressed groups (C16, P < .05). In contrast, TLR10 was downregulated during PGF2α-induced luteolysis and on P16 (P < .05). TLR4 and TLR7 proteins were particularly localized in endothelial cells on C12/PG0h, but prominent signals corresponding to TLR4 and TLR7 were detected in luteal cells at PG16h. The results suggest an involvement of TLRs in the luteolytic mechanism in ovine CL, as indicated by differential expression levels of TLRs during PGF2α-induced luteolysis. Moreover, the present study indicates that early pregnancy-mediated changes in TLR expression in the CL may contribute to the establishment and maintenance of ovine pregnancy.
本研究的目的是阐明妊娠早期和前列腺素F2α（PGF2α）诱导的黄体溶解过程中，绵羊黄体（CL）中Toll样受体（TLR）的表达谱。为此，选择经产安纳托利亚美利奴母羊，并随机分为发情周期组（包括诱导黄体溶解组，n = 20）和妊娠组（n = 12）。所有母羊计划在发情周期、妊娠早期和PGF2α诱导的黄体溶解期间的预定天数/小时进行屠宰。在第12天（C12和P12；n = 8）和第16天（C16和P16；n = 8）从发情周期和妊娠母羊中收集CL，在第22天（P22；n = 4）从妊娠母羊中收集CL。对于诱导黄体溶解模型，在发情周期的第12天给母羊注射PGF2α，并在注射后1小时（PG1h；n = 4）、4小时（PG4h；n = 4）和16小时（PG16h，n = 4）收集CL。采用定量聚合酶链反应（qPCR）评估TLR2、TLR4、TLR6、TLR8和TLR10的表达谱，同时采用游离原位杂交和免疫组织化学法确定CL中TLR2、TLR4和TLR7的空间定位。然后通过单因素方差分析对数据进行分析，当P值低于0.05时认为具有统计学意义。TLR2的表达在黄体溶解的早期和晚期均上调（P < 0.05）。在PG16h检测到TLR4上调，而在PG4h TLR6下调（P < 0.05）。在妊娠早期、PG16h和退化组（C16）中，TLR7和TLR8 的表达均显著增加（P < 0.05）。相反，在PGF2α诱导的黄体溶解期间和P16时，TLR10下调（P < 0.05）。TLR4和TLR7蛋白在C12/PG0h时特别定位于内皮细胞，但在PG16h时在黄体细胞中检测到与TLR4和TLR7对应的明显信号。结果表明，PGF2α诱导的黄体溶解过程中TLR的差异表达水平表明TLR参与了绵羊CL的黄体溶解机制。此外，本研究表明，妊娠早期介导的CL中TLR表达变化可能有助于绵羊妊娠的建立和维持。
