Pitangui-Molina Caroline P, Vireque Alessandra A, Tata Alessandra, Belaz Katia Roberta A, Santos Vanessa G, Ferreira Christina R, Eberlin Marcos N, Silva-de-Sá Marcos Felipe, Ferriani Rui A, Rosa-E-Silva Ana Carolina J S
Department of Obstetrics and Gynecology, School of Medicine of Ribeirão Preto, University of São Paulo, 14049-900 Ribeirão Preto, SP, Brazil.
Thomson Mass Spectrometry Laboratory, Institute of Chemistry, University of Campinas, 13083-970 Campinas, SP, Brazil.
Chem Phys Lipids. 2017 Apr;204:76-84. doi: 10.1016/j.chemphyslip.2017.03.003. Epub 2017 Mar 21.
The phospholipid (PL) composition of embryo and oocyte membranes affects thermal phase behavior and several physicochemical properties such as fluidity and permeability. The characterization of PL profiles and the development of suitable in vitro maturation (IVM) protocols, that are able to modify membrane's composition, may result in significant improvements in oocyte developmental potential and cryotolerance. Using soybean phosphatidylcholine (PC) as a model supplement, we evaluated the effect of PL supplementation during IVM on bovine cumulus-oocyte-complex (COC). Substantial changes in the lipid profiles of oocyte membrane were observed and associated with pre-implantation data. The propensity of the PC supplement to become soluble in the maturation medium and/or diffuse into mineral oil was also assessed. Oocytes were matured in TCM without supplementation, i.e. control, (n=922) or supplemented with 50 or 100μM PC (n=994). The maturation media and mineral oil pre- and post- IVM, along with control and PC-treated oocytes were then analyzed using matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS), and the lipid profiles were compared via principal component analysis (PCA). Soybean PCs are bioavailable and stable in IVM medium; further, PCs did not diffuse to the mineral oil, which also remained unaltered by the metabolism of treated oocytes. PC supplementation at 100μM resulted in substantially greater relative abundances of polyunsatured PL, namely PC (32:1), PC (34:2), PC (36:6), PC (36:4), and PC (38:6), in oocyte membrane. These differences indicated that short-term exposure to the PC supplement could indeed modify the lipid composition of IVM-oocytes in a dose-dependent manner. Membrane incorporation of polyunsaturated molecular species of PC was favored, and does so without compromising the viability of the subsequent embryo in regards to cleavage, blastocyst development and hatching rate. The reported approach will allow for the development of novel strategies to modulate oocyte membrane dynamics and structure.
胚胎和卵母细胞膜的磷脂（PL）组成会影响热相行为以及一些物理化学性质，如流动性和通透性。对PL谱进行表征并开发能够改变膜组成的合适体外成熟（IVM）方案，可能会显著提高卵母细胞的发育潜力和耐冻性。我们以大豆磷脂酰胆碱（PC）作为模型补充剂，评估了IVM期间PL补充对牛卵丘-卵母细胞复合体（COC）的影响。观察到卵母细胞膜脂质谱有显著变化，并与着床前数据相关。还评估了PC补充剂在成熟培养基中溶解和/或扩散到矿物油中的倾向。卵母细胞在未补充的TCM中成熟，即作为对照（n = 922），或补充50或100μM PC（n = 994）。然后使用基质辅助激光解吸/电离质谱（MALDI-MS）分析IVM前后的成熟培养基和矿物油，以及对照和PC处理的卵母细胞，并通过主成分分析（PCA）比较脂质谱。大豆PC在IVM培养基中具有生物可利用性且稳定；此外，PC没有扩散到矿物油中，矿物油也未因处理过的卵母细胞的代谢而改变。100μM的PC补充导致卵母细胞膜中多不饱和PL，即PC（32:1）、PC（34:2）、PC（36:6）、PC（36:4）和PC（38:6）的相对丰度显著更高。这些差异表明，短期暴露于PC补充剂确实可以以剂量依赖的方式改变IVM卵母细胞的脂质组成。PC的多不饱和分子种类有利于整合到膜中，并且在不影响后续胚胎在分裂、囊胚发育和孵化率方面的活力的情况下做到了这一点。所报道的方法将有助于开发调节卵母细胞膜动力学和结构的新策略。
