Cell cycle kinase CHEK2 in macrophages alleviates the inflammatory response to Staphylococcus aureus-induced
Fei Xie1, Ruidong Chen1, Jie Zhao1
1Department of Pediatrics, Cangzhou Central Hospital, Cangzhou, Hebei, China.
Background:
Excessive macrophage-mediated inflammation participates in the development of Staphylococcus aureus (S. aureus)-induced pneumonia. Checkpoint kinase 2 (Chek2) was screened out as macrophage-related infantile pneumonia gene after the differentially expressed analysis of RNAseq data derived from pam3CSK4 stimulated bone marrow-derived macrophages (BMDMs).
Methods:
RAW264.7 macrophage cells were transfected with Chek2-specific gRNA, which were further overexpressed with wide-type Chek2 or Chek2 kinase activity mutant (Chek2 KD, D368N). At the same time, the relative protein and mRNA expression of inflammatory cytokines were determined. C57BL/6J WT mice were intranasally infected with S. aureus to induce S. aureus-induced pneumonia, which was treated with BML-277, an inhibitor of Chek2. The symptoms of pneumonia mice and inflammatory cytokines associated with the nuclear factor kappa B (NF-κB) signaling pathways were further examined.
Results:
In vivo, BML-277 significantly promoted pneumonia symptoms, including mortality, lung infiltration of immune cells, and the abundance of lung pro-inflammatory cytokines. Mechanically, BML-277 did not affect BMDMs survival but up-regulated the mRNA expression of tumor necrosis factor (Tnf), nitric oxide synthase 2 (Nos2), interleukin (Il)23a, and the secretion of Tnf-α and Il-23a. At the same time, genetic complementation experiment testified that Chek2 KD did not inhibit NF-κB and relevant inflammatory cytokines expression.
Conclusion:
Chek2 functions through the kinase mechanism to down-regulate the NF-κB pathway in macrophages to alleviate S. aureus-induced pneumonia in mice.
Insights
Checkpoint kinase 2 (Chek2) down-regulates the NF-κB pathway in macrophages, alleviating Staphylococcus aureus-induced pneumonia. Inhibiting Chek2 worsened pneumonia symptoms in mice.
Area of Science:
- Immunology
- Molecular Biology
- Pathology
Background:
- Macrophage-driven inflammation is central to Staphylococcus aureus pneumonia.
- Checkpoint kinase 2 (Chek2) was identified as a key gene in macrophage-related pneumonia via RNA sequencing.
- Bone marrow-derived macrophages (BMDMs) were used to screen for pneumonia-associated genes.
Purpose of the Study:
- To investigate the role of Checkpoint kinase 2 (Chek2) in Staphylococcus aureus-induced pneumonia.
- To elucidate the mechanism by which Chek2 influences macrophage inflammatory responses.
- To evaluate the therapeutic potential of Chek2 inhibition in pneumonia models.
Main Methods:
- RAW264.7 macrophages were manipulated to overexpress wild-type or mutant Chek2.
- Inflammatory cytokine expression was measured at mRNA and protein levels.
- A mouse model of S. aureus pneumonia was established and treated with the Chek2 inhibitor BML-277.
Main Results:
- Inhibition of Chek2 by BML-277 exacerbated pneumonia symptoms, increasing mortality and lung inflammation.
- BML-277 upregulated pro-inflammatory mediators including TNF, iNOS, and IL-23 in macrophages.
- Genetic experiments confirmed that a kinase-dead Chek2 mutant did not suppress NF-κB signaling.
Conclusions:
- Chek2 acts as a negative regulator of the NF-κB pathway in macrophages.
- Chek2's kinase activity is crucial for its anti-inflammatory function in S. aureus pneumonia.
- Targeting Chek2 may offer a novel therapeutic strategy for bacterial pneumonia.
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