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High-throughput sequencing Analysis of Transcriptomes of depolarized mouse BMDM cell

ABSTRACT: Purpose:Mouse BMDM is the universal cell type of studying innate immunity.This study was to analyze LPS induced innate immune respone and the relationship between the membrane potential and LPS induced innate immune respone Methods: WT BMDM cells were treated with or without LPS for 6 hours in the pressence of membranepotential depolarization stimuli. Then mRNA profiles of these samles were generated by High-throughput sequencing analysis, in triplicate, using Illumina HiSeq /Novaseq or MGI2000. And the differencial mRNA profiles were analyzed. Results: mRNA profiles of more than 20000 genes were analyzed, and differencial expression profiles were compared . Conclusions: Our study represents the first detailed analysis of transcriptomes of WT and membrane potential depolarized macrophages, with biologic replicates, generated by RNA-seq technology. This data was useful to analyze the relationship between membrane potential and LPS induced innate immune respone

ORGANISM(S): Mus musculus

PROVIDER: GSE183067 | GEO | 2022/04/05

REPOSITORIES: GEO

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Project description:BMDMs or iMACs were generated via differentiation of bone marrow cells or immortalized precursors, respectively, in culture media containing m-CSF for 7 day. Upon differentiation, BMDMs or iMacs were left untreated or stimulated as described below. 1) BMDM WT were stimulated with IFNa, LPS, PGE2, IL-4, IL-10, LPS+PGE2, LPS+IL-4, LPS+IL-10, IFNa+PGE2, IFNa+IL-10: each condition in duplicate. This experiment was performed to investigate transcriptional cross-antagonism in the concomitant presence of pro-inflammatory (LPS or IFNa) and anti-inflammatory stimuli (IL-4, IL-10, or PGE2). 2) BMDM WT were stimulated with LPS, in the presence or absence of PFI-1 (Brd2/4 inhibitor) or SGC-CBP30 (CBP/p300 inhibitor): each condition in triplicate. This experiment was performed to define pro-inflammatory genes dependent or not on chromatin remodeling for their induction. 3) BMDM Mef2C/D proficient (Vav-Cre) or BMDM Mef2C/D KO (obtained by crossing Mef2d-/- Mef2cfl/fl with Vav-Cre mice) were stimulated with LPS, PGE2, or LPS+PGE2: each condition in triplicate. This experiment was performed to investigate the role of MEF2C-D on LPS transcritptional response. 4) BMDM WT were pretreated for 40 minutes with an anti-IL10R antibody or the corresponding isotope control, and then left untreated or stimulated with LPS, PGE2 or LPS+PGE2: each condition in duplicate. This experiment was performed to investigate the crosstalk between IL-10 and PGE2. 5) BMDM WT were left untreated or stimulated with PGE2, LPS, LPS+PGE2, or LPS+PGE2+IFNb. In the latter condition, IFNb was added 1 hour after the beginning of the costimulation: each condition in triplicate. This experiment was performed to asses the impact of PGE2 into the LPS-mediated IFNb induction. 5) MEF2A-deficient iMac clones (D7, A7, A8, C7) and MEF2A-proficient (referred to as wild-type) iMac clones (NE, B3 and D10) were generated via CRISPR/Cas9 and stimulated or not with LPS: each condition in single. This experiment was performed to investigate the role of MEF2A on LPS transcritptional response.

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High-throughput sequencing Analysis of Transcriptomes of depolarized mouse BMDM cell

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