Bovine endometrial stromal cells (bESCs) are exposed to a complex environment of bacteria and viruses due to the rupture of epithelial cells after delivery. Inflammatory responses are elicited by the activation of host pattern recognition receptors through pathogen-related molecules such as lipopolysaccharides (LPS) on the cell membrane. Forsythoside A (FTA) is a major active constituent of Forsythia suspensa (Thunb.) Vahl. is a flowering plant widely employed as a traditional Chinese herbal medicine to treat various inflammatory diseases such as nephritis, eye swelling, scabies, ulcers, and mastitis; however, the molecular mechanisms underlying its therapeutic effects on bovine endometritis are still unclear. The aim of this study was to explore the role of miRNA and the mechanisms underlying the protective activity of FTA on the inflammation of bovine endometrial stromal cells induced by LPS. Based on previous research, we isolated and cultured bESCs in vitro and categorized them into LPS and LPS+FTA groups with three replicates. Upon reaching 80% confluence, the bESCs were treated with 0.5 μg/mL of LPS or 0.5 μg/mL of LPS + 100 μg/mL of FTA. We, then, performed high-throughput sequencing (RNA-Seq) to investigate the effects of FTA on LPS-stimulated primary bESCs and their underlying mechanisms. We identified 167 miRNAs differentially expressed in the LPS groups; 72 miRNAs were up-regulated, and 95 were down-regulated. Gene ontology enrichment analysis revealed that differentially expressed microRNA (DEGs) were most enriched during the cellular metabolic process; they were mostly located intracellularly and participated in protein, enzyme, and ion binding. Kyoto Encyclopedia of Genes and Genomes pathway analysis revealed that the DEGs were most enriched in the mitogen-activated protein kinase, tumor necrosis factor, and Interleukin-17 signaling pathways. These results reveal the complex molecular mechanism involved in the FTA and provide a basis for future studies of bovine endometritis treatment with traditional Chinese medicine monomer.

由于分娩后上皮细胞的破裂，牛子宫内膜基质细胞（bESC）暴露于细菌和病毒的复杂环境中。通过病原体相关分子（例如细胞膜上的脂多糖（LPS））激活宿主模式识别受体，引起炎症反应。连翘苷A（FTA）是其主要活性成分连翘（大号）瓦尔。是一种开花植物，被广泛用作中草药，用于治疗各种炎性疾病，例如肾炎，眼睛肿胀，sc疮，溃疡和乳腺炎；然而，其对牛子宫内膜炎治疗作用的分子机制仍不清楚。这项研究的目的是探索miRNA的作用和FTA保护活性对LPS诱导的牛子宫内膜基质细胞发炎的保护作用的机制。根据先前的研究，我们分离并培养了bESC体外并将它们分为三个重复的LPS和LPS + FTA组。达到80％融合后，用0.5μg/ mL的LPS或0.5μg/ mL的LPS + 100μg/ mL的FTA处理bESC。然后，我们进行了高通量测序（RNA-Seq），以研究FTA对LPS刺激的初级bESC及其基础机制的影响。我们确定了LPS组中差异表达的167个miRNA。72个miRNA上调，而95个下调。基因本体富集分析表明，差异表达的microRNA（DEG）在细胞代谢过程中最富集。它们大多位于细胞内，并参与蛋白质，酶和离子结合。京都基因与基因组百科全书通路分析表明，DEG富含丝裂原激活的蛋白激酶，肿瘤坏死因子和白介素17信号通路。这些结果揭示了FTA中涉及的复杂分子机制，并为将来用中药单体治疗牛子宫内膜炎提供了基础。