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Characterization and Isolation of Mouse Primary Microglia by Density Gradient Centrifugation
Published on: February 16, 2018
Exploring the feasibility of using mice as a substitute model for investigating microglia in aging and Alzheimer's
Rong He1, Qiang Zhang1, Limei Wang1
1Laboratory Animal Department, Kunming Medical University, Kunming, Yunnan, China.
Objective:
To guide animal experiments, we investigated the similarities and differences between humans and mice in aging and Alzheimer's disease (AD) at the single-nucleus RNA sequencing (snRNA-seq) or single-cell RNA sequencing (scRNA-seq) level.
Methods:
Microglia cells were extracted from dataset GSE198323 of human post-mortem hippocampus. The distributions and proportions of microglia subpopulation cell numbers related to AD or age were compared. This comparison was done between GSE198323 for humans and GSE127892 for mice, respectively. The Seurat R package and harmony R package were used for data analysis and batch effect correction. Differentially expressed genes (DEGs) were identified by FindMarkers function with MAST test. Comparative analyses were conducted on shared genes in DEGs associated with age and AD. The analyses were done between human and mouse using various bioinformatics techniques. The analysis of genes in DEGs related to age was conducted. Similarly, the analysis of genes in DEGs related to AD was performed. Cross-species analyses were conducted using orthologous genes. Comparative analyses of pseudotime between humans and mice were performed using Monocle2.
Results:
(1) Similarities: The proportion of microglial subpopulation Cell_APOE/Apoe shows consistent trends, whether in AD or normal control (NC) groups in both humans and mice. The proportion of Cell_CX3CR1/Cx3cr1, representing homeostatic microglia, remains stable with age in NC groups across species. Tuberculosis and Fc gamma R-mediated phagocytosis pathways are shared in microglia responses to age and AD across species, respectively. (2) Differences: IL1RAPL1 and SPP1 as marker genes are more identifiable in human microglia compared to their mouse counterparts. Most genes of DEGs associated with age or AD exhibit different trends between humans and mice. Pseudotime analyses demonstrate varying cell density trends in microglial subpopulations, depending on age or AD across species.
Conclusions:
Mouse Apoe and Cell_Apoe maybe serve as proxies for studying human AD, while Cx3cr1 and Cell_Cx3cr1 are suitable for human aging studies. However, AD mouse models (App_NL_G_F) have limitations in studying human genes like IL1RAPL1 and SPP1 related to AD. Thus, mouse models cannot fully replace human samples for AD and aging research.
Insights
Mouse Apoe and Cx3cr1 show promise for studying Alzheimer's disease (AD) and aging in humans, respectively. However, current AD mouse models have limitations in fully replicating human AD and aging gene expressions, highlighting the need for human samples in research.
Area of Science:
- Neuroscience
- Genomics
- Immunology
Background:
- Aging and Alzheimer's disease (AD) are complex neurological conditions.
- Understanding cross-species similarities and differences in these conditions is crucial for developing effective research models.
- Microglia play a key role in brain aging and AD pathogenesis.
Purpose of the Study:
- To compare human and mouse microglia at the single-cell level.
- To identify similarities and differences in gene expression related to aging and AD.
- To guide the development and application of animal models for human neurological research.
Main Methods:
- Single-nucleus RNA sequencing (snRNA-seq) and single-cell RNA sequencing (scRNA-seq) data from human and mouse hippocampus.
- Bioinformatic analysis using Seurat and harmony R packages for differential gene expression and batch correction.
- Cross-species comparative analysis of orthologous genes and pseudotime analysis using Monocle2.
Main Results:
- Microglial subpopulations (Cell_APOE/Apoe, Cell_CX3CR1/Cx3cr1) show conserved proportions related to AD and aging across species.
- Shared pathways like Tuberculosis and Fc gamma R-mediated phagocytosis are involved in microglia responses to aging and AD.
- Significant differences exist in age- and AD-associated gene expression trends and specific marker genes (e.g., IL1RAPL1, SPP1) between humans and mice.
Conclusions:
- Mouse Apoe/Cell_Apoe and Cx3cr1/Cell_Cx3cr1 may serve as valuable proxies for human AD and aging research, respectively.
- Current AD mouse models (App_NL_G_F) have limitations in recapitulating specific human gene expressions (IL1RAPL1, SPP1).
- Mouse models cannot entirely substitute for human samples in comprehensive AD and aging research.
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