ABSTRACT
BACKGROUND: Macrophage pyroptosis is a pivotal inflammatory mechanism in sepsis-induced lung injury, however, the underlying mechanisms remain inadequately elucidated. METHODS: Lipopolysaccharides (LPS)/adenosine triphosphate (ATP)-stimulated macrophages and cecal ligation and puncture (CLP)-induced mouse model for sepsis were established. The levels of key molecules were examined by qRT-PCR, Western blotting, immunohistochemistry (IHC) and ELISA assay. The subcellular localization of circMAPK1 was detected by RNA fluorescence in situ hybridization (FISH). Cell viability, LDH release and caspase-1 activity were monitored by CCK-8, LDH assays, and flow cytometry. The bindings between KDM2B/H3K36me2 and WNK1 promoter was detected by chromatin immunoprecipitation (ChIP) assay and luciferase assay, and associations among circMAPK1, UPF1 and KDM2B mRNA were assessed by RNA pull-down or RNA immunoprecipitation (RIP) assays. The pathological injury of lung tissues was evaluated by lung wet/dry weight ratio and hematoxylin and eosin (H&E) staining. RESULTS: CircMAPK1 was elevated in patients with septic lung injury. Knockdown of circMAPK1 protected against LPS/ATP-impaired cell viability and macrophage pyroptosis via WNK1/NLRP3 axis. Mechanistically, loss of circMAPK1 enhanced the association between KDM2B and WNK1 promoter to promote the demethylation of WNK1 and increase its expression. CircMAPK1 facilitated KDM2B mRNA decay by recruiting UPF1. Functional experiments showed that silencing of KDM2B or WNK1 counteracted circMAPK1 knockdown-suppressed macrophage pyroptosis. In addition, silencing of circMAPK1 alleviated CLP-induced lung injury in mice via KDM2B/WNK1/NLRP3 axis. CONCLUSION: CircMAPK1 exacerbates sepsis-induced lung injury by destabilizing KDM2B mRNA to suppress WNK1 expression, thus facilitating NLRP3-driven macrophage pyroptosis.
Subject(s)
Epigenesis, Genetic , Jumonji Domain-Containing Histone Demethylases , Pyroptosis , Sepsis , WNK Lysine-Deficient Protein Kinase 1 , Animals , Pyroptosis/genetics , Sepsis/complications , Sepsis/genetics , Sepsis/metabolism , Mice , Jumonji Domain-Containing Histone Demethylases/metabolism , Jumonji Domain-Containing Histone Demethylases/genetics , Male , WNK Lysine-Deficient Protein Kinase 1/metabolism , WNK Lysine-Deficient Protein Kinase 1/genetics , Humans , RNA Stability , Lung Injury/etiology , Lung Injury/metabolism , Lung Injury/genetics , Disease Models, Animal , Female , Macrophages/metabolism , Mice, Inbred C57BL , F-Box ProteinsABSTRACT
Lung adenocarcinoma (LUAD) is the most common pathological subtype of lung cancer and has a poor prognosis. Immune Checkpoint Blockage (ICB) have been shown to improve the survival of LUAD in the last decade. CD8 + T cell infiltration is significantly related to LUAD prognosis and plays a critical role in ICB response efficiency. Chemokines expressed and secreted by tumor and microenvironment cells regulate the recruitment of CD8 + T cells. A cytoplasm-dominant circRNA, termed circMAPK1, was found to be down-regulated in LUAD and dramatically suppressed the growth of LUAD upon circMAPK1 overexpression in immunocompetent mice. Meanwhile, it was found that circMAPK1 significantly promoted the CD8 + T cell intratumoral infiltration in vitro and in vivo. CircMAPK1 was identified as binding IGF2BP1 in the cytoplasm and inducing IGF2BP1 to occupy the 3'UTR of CCL5 mRNA, resulting in retained stability of CCL5 mRNA. In general, circMAPK1 is a microenvironment-associated circRNA that recruits CD8 + T cells in LUAD. CircMAPK1 is an effective microenvironment regulator and a potential nucleic acid drug that can be combined with ICB to improve immunotherapy response efficiency.