LAG3 regulates the homeostasis of brain white matter microglia and affects astrocyte morphogenesis in the early postnatal period

In order to further understand how microglia-derived LAG3 regulates microglia homeostasis and affects the morphogenesis of astrocytes, RNA-seq was used to analyze the transcriptome changes of microglia isolated from Lag3 conditional knockout mice and littermate-born wild-type mice in P7 period. About 7000 transcripts showed the differential expression between the microglia of the corpus callosum of Lag3fl/fl and Lag3cKO-Cx3 mice, with a fold change 2, and a p value < 0.05. Gene ontology analysis of downregulated genes were enriched in terms related to actin recombination, ciliary recombination and others, while upregulated genes are enriched in inflammatory response and other terms. These results reflected the role of microglia Lag3 in microglia development. mRNA was extracted from microglia isolated from the corpus callosum of Lag3fl/fl and Lag3cKO-Cx3 mice at P7. Specifically, Agilent 2100 Bioanalyze was used to quality controlled and quantified. Then, the mRNA is converted into cDNA and bound the library. RNA-sequencing analysis was carried out using Illumina HiSeq 2500 platform in anno.txtroad Genomics.为深入探究小胶质细胞（microglia）来源的淋巴细胞活化基因3（LAG3）对小胶质细胞稳态的调控作用，及其对星形胶质细胞（astrocytes）形态发生的影响，本研究采用RNA测序（RNA-seq）技术，分析了出生后第7天（P7）的Lag3条件性敲除小鼠与同窝野生型小鼠分离获得的小胶质细胞的转录组变化。 在Lag3fl/fl与Lag3cKO-Cx3小鼠胼胝体（corpus callosum）的小胶质细胞中，共鉴定出约7000个差异表达转录本，其差异倍数为2，p值<0.05。对下调基因进行基因本体论（Gene Ontology, GO）富集分析发现，其显著富集于肌动蛋白重组（actin recombination）、纤毛重组（ciliary recombination）等相关生物学过程；而上调基因则显著富集于炎症应答（inflammatory response）等相关条目。上述结果揭示了小胶质细胞LAG3在小胶质细胞发育中的关键作用。 本研究提取了P7时期Lag3fl/fl与Lag3cKO-Cx3小鼠胼胝体分离得到的小胶质细胞的总mRNA。具体而言，采用安捷伦2100生物分析仪（Agilent 2100 Bioanalyzer）对mRNA进行质量控制与定量分析；随后将mRNA反转录为互补DNA（cDNA）并构建测序文库；最终依托Illumina HiSeq 2500测序平台，在anno.txtroad Genomics完成RNA测序分析。