TREM2 Gene Dosage Increase Reprograms Microglia Responsivity and Ameliorates Pathological Phenotypes in Alzheimer's Disease Models

TREM2 BAC transgenic mice with elevated expression of human TREM2 in microglia under its endogenous regulation without overexpression of other TREM-like genes on the BAC were generated and crossed with 5xFAD mice, an mouse model of AD. Transcriptome and gene coexpression analyses were performed to obtain chronical view of TREM gene dosage dependent changes in the context of amyloid pathology. The results confer strong evidence that increased TREM2 alters brain transcriptome network response only in the context of a disease state and with an overall rescuing effect in 5xFAD mice. Overall design: Brain cortical tissues were dissected from WT, BAC-TREM2, 5xFAD and 5xFAD/BAC-TREM2 at 2, 4 and 7 months of age. RNA was extracted using Qiagen RNeasy kit. Libraries prepared using the Illumina TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq4000 sequencer using strand-specific, paired-end, 69-mer sequencing protocols to a minimum read depth of 30 million reads per sample. Reads were aligned to mouse genome mm10 using the STAR aligner with default settings. Read counts for individual genes were obtained using HTSeq.本研究构建了在小胶质细胞中受内源调控且高表达人类髓系细胞触发受体2（TREM2）的细菌人工染色体（BAC）转基因小鼠，且该BAC载体未过度表达其他TREM样基因；随后将该小鼠与阿尔茨海默病（AD）模型小鼠5xFAD进行杂交。通过转录组与基因共表达分析，以获取淀粉样蛋白病理背景下TREM基因剂量依赖性变化的时序特征。研究结果有力证实：TREM2表达升高仅在疾病状态下会改变大脑转录组网络应答，并在5xFAD小鼠中整体发挥挽救作用。 实验设计： 分别于2、4、7月龄时，从野生型（WT）、BAC-TREM2、5xFAD及5xFAD/BAC-TREM2小鼠中取材大脑皮层组织。采用Qiagen RNeasy试剂盒提取总RNA，利用Illumina TruSeq RNA Library Prep Kit v2构建测序文库，随后采用链特异性双端69-mer测序策略，在Illumina HiSeq4000测序仪上完成测序，每个样本的测序深度不低于3000万条reads。使用STAR比对软件以默认参数将测序reads比对至小鼠基因组mm10版本，并通过HTSeq工具获取各基因的读段计数。