EGCG Prevents the Transcriptional Reprogramming of an Inflammatory and Immune-Suppressive Molecular Signature in Macrophage-like Differentiated Human HL60 Promyelocytic Leukemia Cells

Abstract

Background: The promyelocytic leukemia cell differentiation process enables recapitulation of the polarized M1 or M2 macrophage-like phenotype with inflammatory and immune-suppressive properties. While evidence supports the anti-inflammatory effect of dietary-derived epigallocatechin-3-gallate (EGCG), its impact on the onset of immune phenotype molecular signature remains unclear. Methods: Human HL60 promyelocytic cells grown in suspension were differentiated into CD11bHigh/CD14Low adherent macrophages with phorbol 12-myristate 13-acetate (PMA). Gelatin zymography was used to assess the levels of matrix metalloproteinase (MMP)-9, and total RNA was isolated for RNAseq and RT-qPCR assessment of differentially expressed gene levels involved in inflammation and immunity. Protein lysates were used to assess the phosphorylation status of signaling intermediates involved in macrophage-like cell differentiation. Results: Cell adhesion and induction of MMP-9 were indicative of HL60 cell differentiation into a macrophage-like phenotype. The extracellular signal-regulated kinase (ERK), glycogen synthase kinase (GSK)-3, p90 ribosomal S6 kinases (RSK), and cAMP-response-element-binding protein (CREB) were all phosphorylated, and EGCG reduced such phosphorylation status. Increases in inflammation and immunity genes included, among others, CCL22, CSF1, CSF2, IL1B, and TNF, which inductions were prevented by EGCG. This was corroborated by unbiased transcriptomic analysis which further highlighted the capacity of EGCG to downregulate the hematopoietic stem cell regulator CBFA2T3. Conclusion: EGCG inhibits inflammatory signaling crosstalk and prevents the onset of an immune phenotype in macrophage-like differentiated cells.