Metabolic Comparison of Dorsal versus Ventral Cells Directly in the Live 8-cell Frog Embryo by Microprobe Single-cell CE-ESI-MS.
Metabolic Comparison of Dorsal versus Ventral Cells Directly in the Live 8-cell Frog Embryo by Microprobe Single-cell CE-ESI-MS.
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DOI：
10.1039/c7ay00834a
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发表时间：
2017-09-14
期刊：
影响因子：
--
通讯作者：
Nemes P
中科院分区：
文献类型：
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作者：
Onjiko RM;Plotnick DO;Moody SA;Nemes P
单细胞质谱 (MS) 能够表征早期胚胎发生过程中细胞分化为不同组织时的代谢组变化。利用全细胞解剖和毛细管电泳电喷雾电离 (CE-ESI) MS，我们最近发现了南非爪蛙 (Xenopus laevis) 8 细胞和 16 细胞胚胎中代谢细胞间的差异，提出了一个问题：代谢细胞异质性是否也可以在 8 细胞胚胎的背腹轴上检测到。在这里，我们通过使用微探针单细胞 CE-ESI-MS 在正离子模式下量化同一胚胎中左背侧动物 (D1L) 和左腹侧动物 (V1L) 细胞对之间的单细胞代谢，直接在活胚胎中测试了这一假设。在对 n = 5 个不同胚胎的两个细胞中可重复检测到的约 70 种分子特征（包括 52 种已鉴定的代谢物）进行量化后，我们采用基于偏最小二乘判别分析 (PLSDA) 的监督多变量数据分析来比较细胞类型之间的代谢。统计分析显示，与 V1L 相比，天冬酰胺、甘氨酸甜菜碱和尚未鉴定的分子在 D1L 细胞中显着富集（p < 0.05 且倍数变化 ≥ 1.5）。这些结果表明，源自同一半球（动物极）的细胞早在 8 细胞阶段就沿着背腹轴具有不同的代谢活动。除了提供早期胚胎发生过程中代谢细胞异质性的新证据外，这项研究还表明，微探针单细胞 CE-ESI-MS 能够分析同一活脊椎动物胚胎中的多个单细胞。
Single-cell mass spectrometry (MS) empowers the characterization of metabolomic changes as cells differentiate to different tissues during early embryogenesis. Using whole-cell dissection and capillary electrophoresis electrospray ionization (CE-ESI) MS, we recently uncovered metabolic cell-to-cell differences in the 8- and 16-cell embryo of the South African clawed frog (Xenopus laevis), raising the question whether metabolic cell heterogeneity is also detectable across the dorsal–ventral axis of the 8-cell embryo. Here, we tested this hypothesis directly in the live embryo by quantifying single-cell metabolism between the left dorsal-animal (D1L) and left ventral-animal (V1L) cell pairs in the same embryo using microprobe single-cell CE-ESI-MS in the positive ion mode. After quantifying ~70 molecular features, including 52 identified metabolites, that were reproducibly detected in both cells among n = 5 different embryos, we employed supervised multivariate data analysis based on partial least squares discriminant analysis (PLSDA) to compare metabolism between the cell types. Statistical analysis revealed that asparagine, glycine betaine, and a yet-unidentified molecule were statistically significantly enriched in the D1L cell compared to V1L (p < 0.05 and fold change ≥ 1.5). These results demonstrate that cells derived from the same hemisphere (animal pole) harbor different metabolic activity along the dorsal–ventral axis as early as the 8-cell stage. Apart from providing new evidence of metabolic cell heterogeneity during early embryogenesis, this study demonstrates that microprobe single-cell CE-ESI-MS enables the analysis of multiple single cells in the same live vertebrate embryo.