背景:越来越多的证据表明输卵管细胞外囊泡(oEVs)在早期胚胎-母体对话中具有积极作用。然而,目前尚不清楚oEV是否有助于识别输卵管中胚胎的存在及其质量.因此,我们检查了牛输卵管上皮细胞(BOEC)分泌的oEV的分子货物是否根据良好的存在而有所不同(≥8个细胞,G)或较差(<8个细胞,P)优质胚胎。此外,分析了G和P胚胎之间RNA谱的差异,试图区分oEV和胚胎EV货物。
方法:为此,在受精后53小时将原代BOEC与体外产生的胚胎(IVP)共培养,如下所示:BOEC与G胚胎(BGE);BOEC与P胚胎(BPE);单独的G胚胎(GE);单独的P胚胎(PE);单独的BOEC(B)和培养基对照(M)。共培养24小时后,收集所有组的条件培养基,并分离和表征EV.通过小RNA测序进行EV和胚胎的MicroRNA分析。
结果:在电动汽车中,鉴定了84个miRNA,具有8个差异丰富的(DA)miRNA的BGE与B和4用于BPE与B(P值<0.01)。在胚胎中,鉴定了187个miRNA,有12个DAmiRNA用于BGE与BPE,3为Gvs.P,8为BGEvs.GE,和11为BPEvs.PE(P值<0.01)。
结论:这些结果表明,oEV参与输卵管-胚胎识别,并指出在早期胚胎发育过程中具有信号传导和支持作用的特定miRNA。
BACKGROUND: Increasing evidence points to an active role of oviductal extracellular vesicles (oEVs) in the early embryo-maternal dialogue. However, it remains unclear whether oEVs contribute to the recognition of the presence of embryos and their quality in the oviduct. Hence, we examined whether the molecular cargo of oEVs secreted by bovine oviduct epithelial cells (BOEC) differs depending on the presence of good (≥ 8 cells, G) or poor (< 8 cells, P) quality embryos. In addition, differences in RNA profiles between G and P embryos were analyzed in attempt to distinguish oEVs and embryonic EVs cargos.
METHODS: For this purpose, primary BOEC were co-cultured with in vitro produced embryos (IVP) 53 h post fertilization as follows: BOEC with G embryos (BGE); BOEC with P embryos (BPE); G embryos alone (GE); P embryos alone (PE); BOEC alone (B) and medium control (M). After 24 h of co-culture, conditioned media were collected from all groups and EVs were isolated and characterized. MicroRNA profiling of EVs and embryos was performed by small RNA-sequencing.
RESULTS: In EVs, 84 miRNAs were identified, with 8 differentially abundant (DA) miRNAs for BGE vs. B and 4 for BPE vs. B (P-value < 0.01). In embryos, 187 miRNAs were identified, with 12 DA miRNAs for BGE vs. BPE, 3 for G vs. P, 8 for BGE vs. GE, and 11 for BPE vs. PE (P-value < 0.01).
CONCLUSIONS: These results indicated that oEVs are involved in the oviductal-embryo recognition and pointed to specific miRNAs with signaling and supporting roles during early embryo development.