Laboratory of Theriogenology, College of Veterinary Medicine, Chungnam National University, 99, Daehak-Ro, Daejeon, 34134, Republic of Korea.
Research Institute of Veterinary Medicine, Chungnam National University, Daejeon, 34134, Republic of Korea.
Funct Integr Genomics. 2023 Jun 7;23(3):200. doi: 10.1007/s10142-023-01126-9.
MiRNAs are small non-coding RNA molecules that play important regulatory roles in diverse biological processes. Royal jelly, a milky-white substance produced by nurse honeybees (Apis mellifera), is the primary food of queen bees and plays a crucial role in their development. However, little is known about the microRNA (miRNAs) content of royal jelly and their potential functions. In this study, we isolated extracellular vesicles from the royal jelly of 36 samples through sequential centrifugation and targeted nanofiltration and performed high-throughput sequencing to identify and quantify the miRNA content of honeybee royal jelly extracellular vesicles (RJEVs). We found a total of 29 known mature miRNAs and 17 novel miRNAs. Through bioinformatic analysis, we identified several potential target genes of the miRNAs present in royal jelly, including those involved in developmental processes and cell differentiation. To investigate the potential roles of RJEVs in cell viability, RJEVs were supplemented to apoptotic porcine kidney fibroblasts induced by ethanol 6% exposure for 30 min. TUNEL assay showed a significant reduction in the apoptosis percentage after RJEV supplementation when compared with the non-supplemented control group. Moreover, the wound healing assay performed on the apoptotic cells showed a rapid healing capacity of RJEV-supplemented cells compared to the control group. We observed a significant reduction in the expression of the miRNA target genes such as FAM131B, ZEB1, COL5A1, TRIB2, YBX3, MAP2, CTNNA1, and ADAMTS9 suggesting that RJEVs may regulate the target gene expression associated with cellular motility and cell viability. Moreover, RJEVs reduced the expression of apoptotic genes (CASP3, TP53, BAX, and BAK), while significantly increasing the expression of anti-apoptotic genes (BCL2 and BCL-XL). Our findings provide the first comprehensive analysis of the miRNA content of RJEVs and suggest a potential role for these vesicles in the regulation of gene expression and cell survival as well as augmenting cell resurrection or anastasis.
miRNAs 是一类小的非编码 RNA 分子，在多种生物过程中发挥着重要的调节作用。蜂王浆是由工蜂（Apis mellifera）分泌的一种乳白色物质，是蜂王的主要食物，对其发育起着至关重要的作用。然而，蜂王浆中 microRNA（miRNAs）的含量及其潜在功能知之甚少。在这项研究中，我们通过连续离心和靶向纳米过滤从 36 个样本的蜂王浆中分离出细胞外囊泡，并进行高通量测序，以鉴定和定量蜜蜂蜂王浆细胞外囊泡（RJEVs）中的 miRNA 含量。我们共发现了 29 个已知成熟的 miRNA 和 17 个新的 miRNA。通过生物信息学分析，我们鉴定出了蜂王浆中存在的 miRNA 的几个潜在靶基因，包括参与发育过程和细胞分化的靶基因。为了研究 RJEVs 在细胞活力中的潜在作用，我们将 RJEVs 添加到经 6%乙醇暴露 30 分钟诱导凋亡的猪肾成纤维细胞中。TUNEL 实验表明，与未添加对照组相比，添加 RJEV 后凋亡细胞的凋亡比例显著降低。此外，在凋亡细胞上进行的划痕愈合实验表明，添加 RJEV 的细胞具有比对照组更快的愈合能力。我们观察到 miRNA 靶基因如 FAM131B、ZEB1、COL5A1、TRIB2、YBX3、MAP2、CTNNA1 和 ADAMTS9 的表达显著降低，这表明 RJEVs 可能调节与细胞运动和细胞活力相关的靶基因表达。此外，RJEVs 降低了凋亡基因（CASP3、TP53、BAX 和 BAK）的表达，同时显著增加了抗凋亡基因（BCL2 和 BCL-XL）的表达。我们的研究结果提供了 RJEVs 中 miRNA 含量的首次全面分析，并表明这些囊泡在调节基因表达和细胞存活以及增强细胞复活或复苏方面可能发挥作用。
