Shuttle Transfer of mRNA Transcripts via Extracellular Vesicles From Male Reproductive Tract Cells to the Cumulus-Oocyte Complex in Rabbits (Oryctolagus cuniculus).
Shuttle Transfer of mRNA Transcripts via Extracellular Vesicles From Male Reproductive Tract Cells to the Cumulus-Oocyte Complex in Rabbits (Oryctolagus cuniculus).
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DOI：
10.3389/fvets.2022.816080
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发表时间：
2022
影响因子：
3.2
通讯作者：
Saadeldin IM
中科院分区：
文献类型：
--
作者：
Abumaghaid MM;Abdelazim AM;Belali TM;Alhujaily M;Saadeldin IM
已知精液中含有一种排卵诱导因子（鉴定为神经生长因子，NGF），在诱导排卵物种中，精液沉积后排卵显着增加。然而，男性生殖道细胞和卵母细胞成熟之间的相互作用，通过信使RNA（mRNA）货物还有待研究。分离兔前列腺（pEVs）、附睾（eEVs）和睾丸（tEVs）原代培养物的细胞外囊泡（EVs），通过相对定量PCR（RT-qPCR）检测其几种mRNA转录本的含量。检测神经生长因子（NGF）、神经营养因子3（NTF 3）、血管内皮生长因子A（VEGFA）、去整合素和金属蛋白酶17（ADAM 17）、中期因子（MDK）、kisspeptin（KISS 1）和促性腺激素释放激素（GNRH 1）的表达。通过透射电子显微镜表征EV。通过PKH 26染色的EV的显微镜检测证实卵丘细胞培养物的EV摄取。此外，比较了pEV、eEV和tEV与补充NGF（10、20和30 ng/ml）对卵母细胞体外成熟（IVM）和转录物表达的影响。KISS 1、NTF 3、MDK、ADAM 17、GAPDH和ACTB在所有EV类型中都有表达。在tEV中检测到GNRH 1。在pEV中检测到NGF，而在eEV中检测到VEGFA。pEV、eEV和20 ng/ml NGF显示最高级别的卵丘扩张，其次是tEV和10 ng/ml NGF。对照组和30 ng/ml NGF组的卵丘扩张程度最低。类似地，在用eEV、pEV、tEV、NGF 20（20 ng/ml NGF）、NGF 10（10 ng/ml NGF）、对照和NGF 30（30 ng/ml NGF）成熟的卵母细胞中，第一极体（PB）排出显著增加。此外，与对照组相比，补充eEV的卵丘细胞中NGFR的表达增加了1.5倍，而补充NGF 20的卵丘-卵母细胞复合物（COCs）中PTGS 2（COX 2）和NTRK的表达分别增加了3倍和5倍。卵母细胞PMP 15表达在补充有eEV的IVM培养基中显示1.8倍的增加。此外，卵母细胞NGFR和NTRK表达在补充有pEVS（分别为3.2倍和1.6倍）和tEV（分别为4倍和1.7倍）的IVM培养基中显著增加。这是第一份报告，以检查mRNA货物的存在下，在电动汽车的雄性兔生殖道细胞，提供了一个模型，刺激雌性兔精液沉积后。
Semen is known to contain an ovulation-inducing factor (identified as a nerve growth factor, NGF) that shows a significant increase in ovulation after semen deposition in induced ovulatory species. However, the interplay between the male reproductive tract cells and oocyte maturation through messenger RNA (mRNA) cargo is yet to be investigated. Extracellular vesicles (EVs) from the primary culture of rabbit prostate (pEVs), epididymis (eEVs), and testis (tEVs) were isolated to examine their contents for several mRNA transcripts through relative quantitative PCR (RT-qPCR). The expressions of NGF, neurotrophin (NTF3), vascular endothelial growth factor A (VEGFA), A disintegrin and metalloprotease 17 (ADAM17), midkine (MDK), kisspeptin (KISS1), and gonadotrophin-releasing hormone (GNRH1) were examined in isolated EVs. EVs were characterized through transmission electron microscopy. EV uptake by cumulus cell culture was confirmed through microscopic detection of PKH26-stained EVs. Furthermore, the effects of pEVs, eEVs, and tEVs were compared with NGF (10, 20, and 30 ng/ml) supplementation on oocyte in vitro maturation (IVM) and transcript expression. KISS1, NTF3, MDK, ADAM17, GAPDH, and ACTB were detected in all EV types. GNRH1 was detected in tEVs. NGF was detected in pEVs, whereas VEGFA was detected in eEVs. pEVs, eEVs, and 20 ng/ml NGF showed the highest grade of cumulus expansion, followed by tEVs and 10 ng/ml NGF. Control groups and 30 ng/ml NGF showed the least grade of cumulus expansion. Similarly, first polar body (PB) extrusion was significantly increased in oocytes matured with eEVs, pEVs, tEVs, NGF20 (20 ng/ml NGF), NGF10 (10 ng/ml NGF), control, and NGF30 (30 ng/ml NGF). Additionally, the expression of NGFR showed a 1.5-fold increase in cumulus cells supplemented with eEVs compared with the control group, while the expression of PTGS2 (COX2) and NTRK showed 3-fold and 5-fold increase in NGF20-supplemented cumulus-oocyte complexes (COCs), respectively. Oocyte PMP15 expression showed a 1.8-fold increase in IVM medium supplemented with eEVs. Additionally, oocyte NGFR and NTRK expressions were drastically increased in IVM medium supplemented with pEVS (3.2- and 1.6-fold, respectively) and tEVs (4- and 1.7-fold, respectively). This is the first report to examine the presence of mRNA cargo in the EVs of male rabbit reproductive tract cells that provides a model for the stimulation of female rabbits after semen deposition.
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影响因子：
3.7
作者：
Al-Dossary AA;Strehler EE;Martin-Deleon PA
通讯作者：
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影响因子：
1.2
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