Transcriptomic analysis of purified human cortical microglia reveals age-associated changes

Purpose: Microglia are essential for central nervous system (CNS) homeostasis and innate neuroimmune function, and play important roles in neurodegeneration and brain aging. Here, we present gene expression profiles of purified microglia isolated at autopsy from the parietal cortex of 39 human subjects with intact cognition. We identified an age-associated gene signature in human microglia that was enriched for genes involved in cell adhesion, axonal guidance, cell surface receptor expression, and actin disassembly. Methods: mRNA profiles of 39 human microglia samples isolated from subjects with intact cognition, 16 corresponding superior parietal cortex tissue and 10 epilepsy surgical samples were generated in a Illumina HiSEQ 2500 sequencer. mRNA profiles of 6 parietal cortex from mice were prepared with a Quantseq 3' mRNA-Seq kit (Lexogen, USA). Reads were aligned to the hg38 assembly of the human genome using STAR and quantified at the gene level by featureCounts. Differential expression between whole brain tissue, surgical samples and isolated microglia was assessed with limma. Results: Overall, genes expressed by human microglia are similar to those in mouse, including established microglia genes CX3CR1, P2YR12, and ITGAM/CD11B. However, a number of immune genes, not identified as part of the mouse microglial signature, were abundantly expressed in human microglia, including TLR, Fc-gamma and SIGLEC receptors, as well as TAL1 and IFI16, regulators of proliferation and cell cycle. Age-associated changes in human microglia were enriched for genes involved in cell adhesion, axonal guidance, cell surface receptor expression, and actin (dis)assembly. Limited overlap was observed in microglial genes regulated during aging between mice and humans, indicating that human and mouse microglia age differently. Conclusions: Here we present the an extensive human microglia gene expression profile. Critical differences with mouse microglia, especially in the context of aging, were observed which highlight the necessity to independently study human microglia. These data and analyses serve as a starting point to address human-specific microglia genes and functions under physiological and neuropathological conditions. mRNA profiles of 39 human microglia samples isolated from subjects with intact cognition, 16 corresponding superior parietal cortex tissue and 10 epilepsy surgical samples were generated in a Illumina HiSEQ 2500 sequencer. mRNA profiles of 6 parietal cortex from mice were prepared with a Quantseq 3' mRNA-Seq kit (Lexogen, USA).研究目的：小胶质细胞（Microglia）是中枢神经系统（CNS）稳态与先天神经免疫功能的核心调控因子，在神经退行性病变及脑衰老过程中发挥关键作用。本研究针对39例认知完整的人类受试者尸检顶叶皮层中分离纯化的小胶质细胞构建基因表达谱，并鉴定出与人小胶质细胞衰老相关的基因特征，该特征显著富集于细胞黏附、轴突导向、细胞表面受体表达及肌动蛋白解聚相关基因。 研究方法：本研究采用Illumina HiSEQ 2500测序仪，对39例来自认知完整受试者的分离人小胶质细胞样本、16例对应顶上叶皮层组织样本及10例癫痫手术样本进行mRNA表达谱分析；同时采用Quantseq 3'端mRNA测序试剂盒（Lexogen，美国）构建6例小鼠顶叶皮层的mRNA表达谱。测序reads通过STAR软件比对至人类基因组hg38组装版本，并通过featureCounts工具完成基因水平定量。采用limma包分析全脑组织、手术样本与分离小胶质细胞之间的差异表达基因。 研究结果：整体而言，人类小胶质细胞的表达基因谱与小鼠小胶质细胞具有较高相似性，包括已被证实的小胶质细胞标志基因CX3CR1、P2YR12及ITGAM/CD11B。然而，大量未被纳入小鼠小胶质细胞特征集的免疫相关基因在人类小胶质细胞中高表达，包括TLR（Toll样受体）、Fcγ受体及SIGLEC（唾液酸结合免疫球蛋白样凝集素）家族受体，以及调控细胞增殖与细胞周期的TAL1与IFI16。人类小胶质细胞的衰老相关表达变化显著富集于细胞黏附、轴突导向、细胞表面受体表达及肌动蛋白（解）聚相关基因。小鼠与人类小胶质细胞在衰老过程中调控的基因重叠度极低，提示人类与小鼠小胶质细胞的衰老机制存在显著差异。 研究结论：本研究构建了一套全面的人类小胶质细胞基因表达谱。研究发现人类与小鼠小胶质细胞存在诸多关键差异，尤其在衰老相关生物学过程中，这凸显了独立开展人类小胶质细胞研究的必要性。本数据集与分析结果可为探究生理及神经病理条件下人类特异性小胶质细胞基因与功能提供基础参考。 本研究采用Illumina HiSEQ 2500测序仪，对39例来自认知完整受试者的分离人小胶质细胞样本、16例对应顶上叶皮层组织样本及10例癫痫手术样本进行mRNA表达谱分析。采用Quantseq 3'端mRNA测序试剂盒（Lexogen，美国）构建6例小鼠顶叶皮层的mRNA表达谱。