Project description:To test the effect of saracatinib on gene expression patterns in microglia, we treated mouse (C57BL/6) primary microglial cultures with saracatinib (1microM, 24h) versus DMSO vehicle control and quantified gene expression. Sequencing libraries were prepared using the Lexogen QuantSeq 3'Forward kit with sequencing performed on an Illumina HiSeq 4000 (single end reads 1 x 65; 5M reads per sample). For processed data, reads were aligned against GRCm38 and quantified using STAR. Samples were normalized with CQN. Batch correction was completed using ComBat from sva version 3.6.0.
Project description:To test the effect of fatostatin on gene expression patterns in microglia, we treated mouse (C57BL/6) primary microglial cultures with fatostatin (5microM, 72h) versus DMSO-only control and quantified gene expression using 3' end labeling. Sequencing libraries were prepared using the Lexogen QuantSeq 3'Forward kit and sequencing of single end reads (1 x 65) was performed on an Illumina HiSeq 4000 at 5M reads per sample. For processed data, reads were aligned against GRCm38 using STAR. Transcripts were quantified and annotated against GencodeM11 using Rsubread. Samples were normalized with CQN.
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:Primary microglia were derived from neonatal mice and were activated via LPS+ATP treatment. test mice were treated with the drug Ladostigil and transcriptome was compared to untreated controls
Project description:The main aim of this study is to evaluate how comparable the established colorectal primary tumor cultures are to the original tumor tissues obtained directly from patients in terms of their genomic profile for which they can be eventually utilized to improve in vitro drug assessment and facilitate personalized treatment. Here, we used colorectal cancer patient samples collected after surgery to perform a gene expression comparison study between original tissues and self-established primary cultures. Subsequently, we also investigated the possibility of using primary cells from patients’ tumors as a model for assessing drug response by characterizing the modifications in gene-drug associations of the primary cells which occurred during the establishment of cell cultures. On an auxiliary note, cytogenetic studies are only possible in cultured cells and not in tissue samples, hence, in this study we also evaluated the potential of genome stability analysis on primary cells as an alternative method to improve our understanding of the genomic changes which occur in cancer initiation and progression.