Project description:We used primary mouse microglia to determine whether Bruton's tyrosine kinase (BTK) inhibition could attenuate differential gene expression induced by Fc receptor stimulation.
Project description:To test the functional role of ApoA-I on microglia in the context of stress, we treated primary microglia with LPS stress with or without ApoA-I protein, and used bulk RNA-seq to evaluate if ApoA-I exhibits an anti-inflammatory role.
Project description:Preparation of primary microglial cultures from postnatal mice is tedious with a low yield, high variability and risk of astrocytic contamination. Microglia derived from embryonic stem cells (ESdM) have been suggested as alternative source, but it is unclear how closely ESdM resemble the molecular phenotype of primary microglia. Here, we performed a whole transcriptome analysis of ESdM in comparison to primary cultured and flow cytometry-sorted microglia and compared the microglial transcriptome to other cell types. Cultured microglia and ESdM were related to sorted microglia, but clearly distinct from other myeloid cell types, T cells, astrocytes and neurons. ESdM and primary cultured microglia showed strong overlap in their transcriptome. Only 144 gene transcripts were differentially expressed between both cell types, mainly derived from immune-related genes with a higher activation status of pro-inflammatory and immune defense genes in primary microglia compared to ESdM. Flow cytometry analysis of cell surface markers CD54, CD74 and CD274 selected from the microarray confirmed the close phenotypic relation between ESdM and primary cultured microglia. Thus, assessment of genome-wide transcriptional regulation demonstrates that microglia are distinct from other macrophage cell types and that mouse pluripotent stem cell-derived microglia are closely related to cultured postnatal microglia. Comparison of different primary neuronal cells with ES-cell derived microglial cells
Project description:We performed brain bulk RNA-seq aiming to study how microglia TREM2-WT or TREM2-R47H expression affect the overall brain transcriptomic profiles. Using WGCNA gene network analysis, we found the presynaptic transmission pathway was dysregulated after TREM2-R47H expression, not TREM2-WT. This finding was consistent with the impaired presynaptic transmission detected by electrophysiological recording in TREM2-R47H expressing mice. In addition, dysregulated circadian pathways were also identified in TREM2-R47H mice, not in TREM2-WT mice. Altogether, our bulk RNA-seq results provide additional evidence at transcriptomics level that TREM2-R47H expression in microglia affects neuronal functions and brain networks.
Project description:Microglia, the resident immune cells of the brain, can exhibit a broad range of activation phenotypes, many of which have been implicated in several diseases and disorders of the central nervous system including alcohol use disorders and disorders. By utilizing a method optimized for sensitive and rapid quantitative proteomic analysis of microglia involving suspension trapping (S-Trap), we were able to produce efficient and reproducible protein extraction from low cell yielding primary mouse brains. Using a ~2 h gradient on a 75 cm UPLC column with a modified data dependent acquisition method on a hybrid quadrupole-Orbitrap mass spectrometer (QE Plus), 5,062 total proteins were identified where 4,928 of those proteins were quantifiable by label-free quantitation (with 5 biological replicates). This analysis resulted in the most comprehensive proteomic dataset for ethanol- and LPS-treated primary mouse microglia to date and even expanded upon the well-characterized macrophage/microglia response to LPS treatment. This study also highlights the subtle, yet significant changes ethanol exposure can induce when compared to control. Interestingly, these changes are not consistent with the robust classical activation induced by LPS treatment, but instead align with the emerging theory that ethanol-treated microglia yield an alternative activation response. The contrast to LPS-treated microglia leads us to conclude that ethanol does not elicit a strong inflammatory response but rather might have a general inhibitory effect on multiple pathways such as phagocytosis and cell migration.
Project description:Human iPSC-derived microglia assume a primary microglia-like state after transplantation into the neonatal mouse brain [Bulk RNAseq]
Project description:We assessed the roles of repopulating microglia in brain repair using mouse models. In this project, we show that removal of microglia from the mouse brain has little impact on the outcome of TBI but inducing the turnover of these cells through either pharmacologic or genetic approaches can yield a neuroprotective microglial phenotype that profoundly aids recovery. As a part of the experimental approaches, we perform bulk RNA sequencing experiments to unbiasedly profile the transcriptome of repopulating microglia. We identified unique gene signatures from repopulating microglia cells and infer how these cells modulate the microenvironment after TBI.