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マウス着床後発生における胚ー子宮相互作用の研究

小鼠着床后发育过程中胚胎与子宫相互作用的研究

基本信息

批准号：
16F16089
负责人：
藤森俊彦
金额：
$ 1.54万
依托单位：
National Institute for Basic Biology
依托单位国家：
日本
项目类别：
Grant-in-Aid for JSPS Fellows
财政年份：
2016
资助国家：
日本
起止时间：
2016-04-22 至 2018-03-31
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-16F16089/
关键词：
鉄イオン

项目摘要

I investigated the morphological changes of both the uterus and the extraembryonic tissues through analysis of 3D rendered images of post-implantation embryos. Proof of principle was determined by making 3D reconstructions of heart tissues from serially sectioned 9.5 dpc embryos, scanned at high resolution, aligned by unique and precise automated software, and then delineated from surrounding tissues by manual tracing throughout 241 serial sections using the free open-source software FIJI/TrakEM2. Gene expression analysis with reverse-transcription quantitative PCR of iron regulatory genes expression was performed, and analysis through immunofluorescence of non-pregnant and pregnant stages of 4.5-9.5 days. SLC40a1 protein was upregulated in non-pregnant females is particularly high in the LE and GE of the uterus. Female SLC40a1 transgenic uteri were collected at several stages of the uterine cycle and stained for LacZ. This showed that SLC40a1 is most highly expressed during the estrus cycle. High expression of Lacz staining is exhibited in the luminal epithelium and glandular epithelium with little or no expression in the underlying stromal tissue. Concurrently, analysis of the ovary and oviduct revealed consistent expression of SLC40a1-driven LacZ independent of estrus cycle, suggesting a differential expression mechanism of ovarian SlC40a1 vs uterine SLC40a1. DAB staining of both non-pregnant and pregnant female WT mice exhibited differential expression patterns consistent with promoter driven Lacz staining and immunofluorescence.

我通过分析植入后胚胎的3D渲染图像，研究了子宫和胚外组织的形态学变化。原理的证明是通过从连续切片的9.5 dpc胚胎中进行心脏组织的3D重建来确定的，以高分辨率扫描，通过独特而精确的自动化软件进行对齐，然后使用免费的开源软件FIJI/TrakEM 2通过在241个连续切片中手动追踪从周围组织中划定。采用逆转录定量PCR技术对铁调控基因的表达进行基因表达分析，并通过免疫荧光技术对非妊娠期和妊娠4.5-9.5天的妊娠期进行分析。 SLC 40 a1蛋白在非妊娠女性中上调，在子宫的LE和GE中特别高。在子宫周期的几个阶段收集雌性SLC 40 a1转基因子宫，并对LacZ染色。这表明SLC 40 a1在发情周期中表达最高。 Lacz染色在腔上皮和腺上皮中表现出高表达，而在下面的基质组织中很少或没有表达。同时，对卵巢和输卵管的分析显示，SLC 40 a1驱动的LacZ的一致表达与动情周期无关，这表明卵巢SLC 40 a1与子宫SLC 40 a1的表达机制存在差异。非妊娠和妊娠雌性WT小鼠的DAB染色显示出与启动子驱动的Lacz染色和免疫荧光一致的差异表达模式。