Swedish University of Agricultural Sciences, Department of Clinical Sciences, Division of Reproduction, Uppsala, Sweden.
Swedish University of Agricultural Sciences, Department of Animal Breeding and Genetics, Section of Molecular Genetics, Uppsala, Sweden.
PLoS One. 2019 Sep 5;14(9):e0222081. doi: 10.1371/journal.pone.0222081. eCollection 2019.
Lipopolysaccharide (LPS) expressed on the surface of Gram-negative bacteria activates pro-inflammatory pathways, dys-regulates the function of endometrial cells and is a key player in the mechanisms involved in endometritis. This study aimed to investigate the effects of LPS on bovine endometrial epithelial cells (bEEC) from whole transcriptome with a special focus on genes involved in embryo-maternal interactions. Following in vitro culture, bEEC from three cows were exposed to 0, 2, and 8 μg/mL LPS for 24h. RNA samples extracted at 0 and 24 hours were analyzed by RNA sequencing (RNA-seq). At 24h, 2035 differentially expressed genes (DEGs) were identified between controls and samples treated with 2 μg/mL LPS. Gene ontology analysis showed that over-expressed DEGs were associated to immune response, response to stress and external stimuli, catalytic activity, and cell cycle. Genes associated with cell membrane and cell adhesion pathways were under-expressed. LPS induced changes in expression of specific genes related to embryo-maternal interactions including under-expression of eight members of the cadherin superfamily, over-expression of six members of the mucin family, and differential expression of a large set of genes binding the above molecules and of more than 20 transcripts coding for cytokines and their receptors. Type I interferon-τ dependent genes were also over-expressed. From a sub-set of 19 genes, (biological replicates of bEEC from cows taken at time 6 (n = 3), 24 (n = 6) and 48 hours (n = 3), and 2 technical replicates per sample) differential gene expression was confirmed by RT2-qPCR (r2 between fold changes at 24 hours by RT2-qPCR and RNA-seq = 0.97). These results indicate that LPS affects the function of bEEC in many ways by differential transcription, glycolytic metabolism and oxidative stress. Many transcriptomic signatures related to implantation and embryo maternal interactions were strongly affected by LPS. These results pave the way for further studies to investigate the duration of these changes and their possible impact on endometrial function and fertility.
脂多糖(LPS)表达在革兰氏阴性菌的表面,激活促炎途径,失调子宫内膜细胞的功能,是子宫内膜炎相关机制中的关键因素。本研究旨在从全转录组水平研究 LPS 对牛子宫内膜上皮细胞(bEEC)的影响,特别关注与胚胎-母体相互作用相关的基因。体外培养后,将来自 3 头奶牛的 bEEC 分别用 0、2 和 8μg/mL LPS 孵育 24 小时。分别在 0 和 24 小时提取 RNA 样本,并进行 RNA 测序(RNA-seq)分析。在 24 小时时,在对照组和用 2μg/mL LPS 处理的样本之间鉴定出 2035 个差异表达基因(DEGs)。基因本体分析表明,过表达的 DEGs 与免疫反应、应激和外部刺激反应、催化活性和细胞周期有关。与细胞膜和细胞黏附途径相关的基因表达下调。LPS 诱导与胚胎-母体相互作用相关的特定基因表达变化,包括钙黏蛋白超家族的 8 个成员表达下调、粘蛋白家族的 6 个成员表达上调,以及与上述分子结合的一大组基因和 20 多个编码细胞因子及其受体的转录本的差异表达。I 型干扰素-τ 依赖性基因也过表达。从一组 19 个基因(来自于第 6 小时(n=3)、24 小时(n=6)和 48 小时(n=3)的奶牛 bEEC 的生物学重复,每个样本 2 个技术重复)中,通过 RT2-qPCR (24 小时时 RT2-qPCR 和 RNA-seq 的折叠变化之间的 r2=0.97)验证了基因表达的差异。这些结果表明,LPS 通过差异转录、糖酵解代谢和氧化应激等多种方式影响 bEEC 的功能。许多与着床和胚胎-母体相互作用相关的转录组特征受到 LPS 的强烈影响。这些结果为进一步研究这些变化的持续时间及其对子宫内膜功能和生育能力的可能影响铺平了道路。