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High-throughput sequencing Analysis of mouse BMDM cell Transcriptomes after LPS Challenge

ABSTRACT: Purpose: Mouse BMDM is the universal cell type of studying innate immunity.This study was designed to analyze LPS induced innate immune respone and the gene expression in WT and HDAC3 KO BMDMs. Methods: BMDM cells from Hdac3f/f and Lyz2-Cre-Hdac3f/f were treated with or without LPS for 4 hours. Then mRNA profiles of these samles were generated by High-throughput sequencing analysis, in triplicate, using illumina HiSeqTM2000/MiSeq. And the differencial mRNA profiles were analyzed. Results: mRNA profiles were analyzed, and differencial expression profiles were compared . Conclusions: Our study demostrated the transcriptional profiles of BMDMs from Hdac3f/f and Lyz2-Cre-Hdac3f/f mice upon LPS challenge, with biologic replicates, generated by RNA-seq technology.

ORGANISM(S): Mus musculus

PROVIDER: GSE155801 | GEO | 2021/08/06

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 mouse BMDM cell Transcriptomes after LPS Challenge

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