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上皮钙黏蛋白存在于牛输卵管上皮细胞和配子中,并参与受精相关事件。

Epithelial cadherin is present in bovine oviduct epithelial cells and gametes, and is involved in fertilization-related events.

作者信息

Caballero Julieta N, Gervasi María G, Veiga María F, Dalvit Gabriel C, Perez-Martínez Silvina, Cetica Pablo D, Vazquez-Levin Mónica H

机构信息

Instituto de Biología y Medicina Experimental (IBYME), National Research Council of Argentina (CONICET), University of Buenos Aires, Buenos Aires, Argentina.

Centro de Estudios Farmacológicos y Botánicos (CEFyBO), National Research Council of Argentina (CONICET), University of Buenos Aires, Buenos Aires, Argentina.

出版信息

Theriogenology. 2014 Jun;81(9):1189-206. doi: 10.1016/j.theriogenology.2014.01.028. Epub 2014 Jan 31.

Abstract

Fertilization is a calcium-dependent process that involves sequential cell-cell adhesion events of spermatozoa with oviduct epithelial cells (OECs) and with cumulus-oocyte complexes (COCs). Epithelial cadherin (E-cadherin) participates in calcium-dependent somatic cell adhesion; the adaptor protein β-catenin binds to the E-cadherin cytoplasmic domain and links the adhesion protein to the cytoskeleton. The study was conducted to immunodetect E-cadherin and β-catenin in bovine gametes and oviduct (tissue sections and OEC monolayers), and to assess E-cadherin participation in fertilization-related events. Epithelial cadherin was found in spermatozoa, oocytes, cumulus cells, and OEC. In acrosome-intact noncapacitated spermatozoa, E-cadherin was mainly localized in the apical ridge and acrosomal cap (E1-pattern; 84 ± 9%; mean ± standard deviation of the mean). After sperm treatment with heparin to promote capacitation, the percentage of cells with E1-pattern (56 ± 12%) significantly decreased; concomitantly, the percentage of spermatozoa depicting an E-cadherin staining pattern similar to E1-pattern but showing a signal loss in the acrosomal cap (E2-pattern: 40 ± 11%) increased. After l-α-lysophosphatidylcholine-induced acrosome reaction, E-cadherin signal was mainly localized in the inner acrosomal membrane (E3-pattern: 67 ± 22%). In IVM COC, E-cadherin was immunodetected in the plasma membrane of cumulus cells and oocytes, but was absent in the polar body. The 120 KDa mature protein form was found in protein extracts from spermatozoa, oocytes, cumulus cells, and OEC. β-Catenin distribution followed E-cadherin's in all cells evaluated. Epithelial cadherin participation in cell-cell interaction was evaluated using specific blocking monoclonal antibody DECMA-1. Sperm incubation with DECMA-1 impaired sperm-OEC binding (the number of sperm bound to OEC: DECMA-1 = 6.7 ± 6.1 vs. control = 29.6 ± 20.1; P < 0.001), fertilization with COC (% fertilized COC: DECMA-1 = 68.8 ± 10.4 vs. control = 90.7 ± 3.1; P < 0.05) or denuded oocytes (% fertilized oocytes: DECMA-1 = 57.0 ± 15.2 vs. control = 89.2 ± 9.8; P < 0.05) and binding to the oolemma (the number of sperm bound to oolemma: DECMA-1 = 2.2 ± 1.1 vs. control = 11.1 ± 4.8; P < 0.05). This study describes, for the first time, the presence of E-cadherin in bovine spermatozoa, COC, and OEC, and shows evidence of its participation in sperm interaction with the oviduct and the oocyte during fertilization.

摘要

受精是一个依赖钙的过程,涉及精子与输卵管上皮细胞(OECs)以及与卵丘 - 卵母细胞复合体(COCs)之间连续的细胞 - 细胞黏附事件。上皮钙黏蛋白(E - cadherin)参与依赖钙的体细胞黏附;衔接蛋白β - 连环蛋白与E - cadherin的细胞质结构域结合,并将黏附蛋白与细胞骨架相连。本研究旨在免疫检测牛配子和输卵管(组织切片及OEC单层细胞)中的E - cadherin和β - 连环蛋白,并评估E - cadherin在受精相关事件中的作用。在精子、卵母细胞、卵丘细胞和OEC中均发现了上皮钙黏蛋白。在顶体完整的未获能精子中,E - cadherin主要定位于顶脊和顶体帽(E1模式;84±9%;平均值±平均值的标准差)。用肝素处理精子以促进获能后,具有E1模式的细胞百分比(56±12%)显著降低;同时,呈现与E1模式相似但顶体帽信号缺失的E - cadherin染色模式的精子百分比(E2模式:40±11%)增加。在l-α-溶血磷脂酰胆碱诱导顶体反应后,E - cadherin信号主要定位于顶体内膜(E3模式:67±22%)。在体外成熟(IVM)的COC中,在卵丘细胞和卵母细胞的质膜中免疫检测到E - cadherin,但在极体中未检测到。在精子、卵母细胞、卵丘细胞和OEC的蛋白质提取物中发现了120 KDa的成熟蛋白形式。在所有评估的细胞中,β - 连环蛋白的分布与E - cadherin一致。使用特异性阻断单克隆抗体DECMA - 1评估E - cadherin在细胞 - 细胞相互作用中的作用。用DECMA - 1孵育精子会损害精子与OEC的结合(与OEC结合的精子数量:DECMA - 1 = 6.7±6.1,对照组 = 29.6±20.1;P < 0.001)、与COC的受精(受精的COC百分比:DECMA - 1 = 68.8±10.4,对照组 = 90.7±3.1;P < 0.05)或与裸卵的受精(受精的卵母细胞百分比:DECMA - 1 = 57.0±15.2,对照组 = 89.2±9.8;P < 0.05)以及与卵膜结合(与卵膜结合的精子数量:DECMA - 1 = 2.2±1.1,对照组 = 11.1±4.8;P < 0.05)。本研究首次描述了牛精子、COC和OEC中存在E - cadherin,并显示了其在受精过程中参与精子与输卵管及卵母细胞相互作用的证据。

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