Table5_Identification of microRNAs implicated in modulating resveratrol-induced apoptosis in porcine granulosa cells.XLSX

MicroRNAs (miRNAs) are small, noncoding RNAs that play a crucial role in the complex and dynamic network that regulates the apoptosis of porcine ovarian granulosa cells (POGCs). Resveratrol (RSV) is a nonflavonoid polyphenol compound that is involved in follicular development and ovulation. In previous study, we established a model of RSV treatment of POGCs, confirming the regulatory effect of RSV in POGCs. To investigate the miRNA-level effects of RSV on POGCs to reveal differentially expressed miRNAs, a control group (n = 3, 0 μM RSV group), a low RSV group (n = 3, 50 μM RSV group), and a high RSV group (n = 3, 100 μM RSV group) were created for small RNA-seq. In total, 113 differentially expressed miRNAs (DE-miRNAs) were identified, and a RT-qPCR analysis showed a correlation with the sequencing data. Functional annotation analysis revealed that DE-miRNAs in the LOW vs. CON group may be involved in cell development, proliferation, and apoptosis. In the HIGH vs. CON group, RSV functions were associated with metabolic processes and responses to stimuli, while the pathways were related to PI3K24, Akt, Wnt, and apoptosis. In addition, we constructed miRNA-mRNA networks related to Apoptosis and Metabolism. Then, ssc-miR-34a and ssc-miR-143-5p were selected as key miRNAs. In conclusion, this study provided an improved understanding of effects of RSV on POGCs apoptosis through the miRNA modulations. The results suggest that RSV may promote POGCs apoptosis by stimulating the miRNA expressions and provided a better understanding of the role of miRNAs combined with RSV in ovarian granulosa cell development in pigs.

微小RNA（MicroRNAs，miRNAs）是一类小型非编码RNA，在调控猪卵巢颗粒细胞（porcine ovarian granulosa cells, POGCs）凋亡的复杂动态网络中发挥关键作用。白藜芦醇（Resveratrol, RSV）是一种非黄酮类多酚化合物，可参与卵泡发育与排卵过程。本课题组前期研究构建了RSV处理猪卵巢颗粒细胞的模型，证实了RSV对猪卵巢颗粒细胞的调控效应。为探究RSV对猪卵巢颗粒细胞的miRNA层面调控效应、筛选差异表达miRNAs，本研究设置空白对照组（n=3，0 μM RSV组）、低剂量RSV组（n=3，50 μM RSV组）与高剂量RSV组（n=3，100 μM RSV组），开展小RNA测序（small RNA-seq）。共计鉴定出113个差异表达miRNAs（differentially expressed miRNAs, DE-miRNAs），实时定量荧光PCR（RT-qPCR）验证结果与测序数据具有良好一致性。功能注释分析显示，低剂量RSV组与对照组比较（LOW vs. CON）的差异表达miRNAs可能参与细胞发育、增殖及凋亡过程；高剂量RSV组与对照组比较（HIGH vs. CON）的差异表达miRNAs则与代谢过程、应激应答相关，涉及的信号通路包括PI3K24、Akt、Wnt及凋亡通路。此外，本研究构建了与凋亡及代谢相关的miRNA-mRNA调控网络，并筛选出ssc-miR-34a与ssc-miR-143-5p作为核心miRNAs。综上，本研究深化了对RSV通过调控miRNA影响猪卵巢颗粒细胞凋亡的认知；研究结果表明，RSV可能通过上调miRNA表达促进猪卵巢颗粒细胞凋亡，同时为阐明miRNA联合RSV在猪卵巢颗粒细胞发育中的调控作用提供了新的理论依据。