Intracellular oxygen metabolism during bovine oocyte and preimplantation embryo development.
Intracellular oxygen metabolism during bovine oocyte and preimplantation embryo development.
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DOI：
10.1038/s41598-021-99512-5
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发表时间：
2021-10-28
影响因子：
4.6
通讯作者：
Picton HM
中科院分区：
文献类型：
--
作者：
McKeegan PJ;Boardman SF;Wanless AA;Boyd G;Warwick LJ;Lu J;Gnanaprabha K;Picton HM
我们报告了一种新的方法，在体外哺乳动物卵母细胞和植入前胚胎发育过程中，使用市售的多模式磷光纳米传感器（MM 2）的轮廓胞内氧浓度（icO 2）。将屠宰场来源的牛卵母细胞和胚胎与MM 2进行体外孵育。在活细胞多光子成像过程中应用一系列抑制剂来记录与线粒体过程相关的icO 2变化。解偶联剂羰基氰化物-对三氟甲氧基苯腙（FCCP）解偶联线粒体耗氧量的最大值，而抗霉素抑制复合物III消融线粒体耗氧量。预计增加耗氧量可降低icO 2，减少耗氧量可增加icO 2。这些抑制剂的使用量化了与线粒体功能的上限和下限相比在基础时消耗了多少氧。将icO 2测量值与通过qPCR分析的线粒体DNA拷贝数进行比较。抗霉素治疗增加icO 2的所有阶段的测试，表明显着的线粒体耗氧量在基础。卵母细胞和植入前胚胎的icO 2不受FCCP处理的影响。内细胞团icO 2低于滋养外胚层，可能反映了扩散的局限性。线粒体DNA拷贝数在0.9-4 × 106拷贝范围内在各阶段之间相似，与icO 2无关。这些结果验证了MM 2探针作为哺乳动物卵母细胞和植入前胚胎中细胞内氧浓度的灵敏、无毒探针。
We report a novel method to profile intrcellular oxygen concentration (icO2) during in vitro mammalian oocyte and preimplantation embryo development using a commercially available multimodal phosphorescent nanosensor (MM2). Abattoir-derived bovine oocytes and embryos were incubated with MM2 in vitro. A series of inhibitors were applied during live-cell multiphoton imaging to record changes in icO2 associated with mitochondrial processes. The uncoupler carbonyl cyanide-p-trifluoromethoxyphenylhydrazone (FCCP) uncouples mitochondrial oxygen consumption to its maximum, while antimycin inhibits complex III to ablate mitochondrial oxygen consumption. Increasing oxygen consumption was expected to reduce icO2 and decreasing oxygen consumption to increase icO2. Use of these inhibitors quantifies how much oxygen is consumed at basal in comparison to the upper and lower limits of mitochondrial function. icO2 measurements were compared to mitochondrial DNA copy number analysed by qPCR. Antimycin treatment increased icO2 for all stages tested, suggesting significant mitochondrial oxygen consumption at basal. icO2 of oocytes and preimplantation embryos were unaffected by FCCP treatment. Inner cell mass icO2 was lower than trophectoderm, perhaps reflecting limitations of diffusion. Mitochondrial DNA copy numbers were similar between stages in the range 0.9–4 × 106 copies and did not correlate with icO2. These results validate the MM2 probe as a sensitive, non-toxic probe of intracellular oxygen concentration in mammalian oocytes and preimplantation embryos.
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影响因子：
4.5
作者：
Fragouli E;Spath K;Alfarawati S;Kaper F;Craig A;Michel CE;Kokocinski F;Cohen J;Munne S;Wells D
通讯作者：
Wells D
影响因子：
8
作者：
Dmitriev, Ruslan I.;Papkovsky, Dmitri B.
通讯作者：
Papkovsky, Dmitri B.
影响因子：
4
作者：
Cotterill M;Harris SE;Collado Fernandez E;Lu J;Huntriss JD;Campbell BK;Picton HM
通讯作者：
Picton HM
影响因子：
14.8
作者：
Ferreira F;Luxardi G;Reid B;Ma L;Raghunathan V;Zhao M
通讯作者：
Zhao M
影响因子：
3
作者：
Coskun, Pinar;Wyrembak, Joanne;Schriner, Samual E.;Chen, Hsiao-Wen;Marciniack, Christine;LaFerla, Frank;Wallace, Douglas C.
通讯作者：
Wallace, Douglas C.