TET1 deficiency amplifies macrophage inflammatory signaling associated with Crohn’s disease

Abstract

Objective and design To define the role of Ten-Eleven Translocation (TET) proteins in Crohn’s disease (CD)-associated inflammation through integrative human and mechanistic studies. Material Publicly available CD transcriptomic and DNA methylation datasets, and primary mononuclear cells and ileal biopsies were analyzed for TET gene expression and signatures. TET1 and TET2 CRISPR/Cas9 knockout macrophage cell lines were generated. Treatment Macrophages were stimulated with LPS in the presence or absence of kinase inhibitors. Conditioned media from macrophages were applied to primary human neutrophils. PBMCs from CD patients and healthy donors were stimulated with LPS for validation. Methods Macrophages or primary patients samples were analyzed by high-throughput surface marker profiling, RNA sequencing, 5hmC sequencing, assays of effector function, qRT-PCR, phosphoflow, and cytokine/chemokine release by ELISA. Results TET1 was the most downregulated TET enzyme in CD blood and ileal tissues, correlating with reduced TET-associated gene signatures and elevated inflammatory mediators. TET1 -deficient macrophages exhibited distinct surface phenotypes, reduced PTEN expression, altered 5hmC distribution, and heightened IFN gene expression, ERK activation, and chemokine release associated with enhanced neutrophil migration. PBMCs from CD patients mirrored reduced TET1 expression and exaggerated inflammatory responses. Conclusions TET1 functions as a non-redundant regulator of inflammatory macrophages and aberrant chemokine signaling linked to immune cell recruitment in Crohn’s disease.

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