Session: 201. Granulocytes, Monocytes, and Macrophages: Poster III
Hematology Disease Topics & Pathways:
cell division, Biological Processes, white blood cells, Cell Lineage, inflammation
We utilised the Klf3-null mouse model  to more closely examine the role of Klf3 in innate inflammatory cells. These mice exhibit elevated white cell counts, including monocytes (Figure 1A), and inflammation of the skin. Conditional knockout of Klf4 in myeloid cells leads to a deficiency of inflammatory macrophages . To test our hypothesis KLF3 normally represses inflammation, perhaps by antagonising the action of KLF4, bone-marrow derived macrophages (BMDM) were generated from wild-type or Klf3-null mice and stimulated with the bacterial toxin lipopolysaccharide (LPS). In wild type BMDM, LPS induces Klf3 gene expression and activation then delayed repression of target genes such as Lgals3 (galectin-3) over a 21 hour time course (Figure 1B). Quantitative real-time PCR and mRNA-seq of WT v Klf3-null macrophages identified ~100 differentially expressed genes involved in proliferation, macrophage activation and inflammation.
We transduced the monocyte cell line, RAW264.7 (that expresses Klf4, Klf3 and Klf2), with a retroviral vector expressing a tamoxifen-inducible KLF3-ER fusion construct. KLF3 induced cell cycle arrest and macrophage differentiation. We will report on KLF3-induced gene expression changes (repression and activation), and ChIP-seq for KLF3, in RAW cells. The results shed light on the mechanism by which KLF3 normally represses monocyte/macrophage responses to infection.
This study highlights the importance of key transcriptional regulators that tightly control gene expression during inflammation. Loss of Klf3 leads to alterations in this process, resulting in hyper-activation of inflammatory macrophages, increased white cell counts and inflammation of the skin. A greater knowledge of the inflammatory process and how it is regulated is important for our understanding of acute infection and inflammatory disease. Further studies are planned to investigate the role of the KLF3 transcription factor in response to inflammation in vivo.
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Figure 1: Elevated WCC (A) and inflammatory markers (B) in BMDM after LPS stimulation.
- Total WCC in adult mice (3-6 months old) of the indicated genotypes. There is a statistically significant increase in the WCC in Klf3-/- v wild type mice (P<0.001 by student’s t test). B. Time course (hours) after LPS stimulation of confluent BMDM. Klf3 is induced 3-fold by LPS and KLF3-target genes such as Lgals3 are not fully repressed by 21 hours in knockout mice.
Disclosures: Perkins: Novartis Oncology: Honoraria, Membership on an entity's Board of Directors or advisory committees.
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