Genistein promotes M1 macrophage apoptosis and reduces inflammatory response by disrupting miR-21/TIPE2 pathway

Abstract

Cardiovascular diseases are a major cause of mortality, and vascular injury, a common pathological basis of cardiovascular disease, is deeply correlated with macrophage apoptosis and inflammatory response. Genistein, a type of phytoestrogen, exerts cardiovascular protective activities, but the underlying mechanism has not been fully elucidated. In this study, RAW264.7 cells were treated with genistein, lipopolysaccharide (LPS), nuclear factor-kappa B (NF-κB) inhibitor, and/or protein kinase B (AKT) agonist to determine the role of genistein in apoptosis and inflammation in LPS-stimulated cells. Simultaneously, high fat diet-fed C57BL/6 mice were administered genistein to evaluate the function of genistein on LPS-induced cardiovascular injury mouse model. Here, we demonstrated that LPS obviously increased apoptosis resistance and inflammatory response of macrophages by promoting miR-21 expression, and miR-21 downregulated tumor necrosis factor-α-induced protein 8-like 2 (TIPE2) expression by targeting the coding region. Genistein reduced miR-21 expression by inhibiting NF-κB, then blocked toll-like receptor 4 (TLR4) pathway and AKT phosphorylation dependent on TIPE2, resulting in inhibition of LPS. Our research suggests that miR-21/TIPE2 pathway is involved in M1 macrophage apoptosis and inflammatory response, and genistein inhibits the progression of LPS-induced cardiovascular injury at the epigenetic level via regulating the promoter region of Vmp1 by NF-κB.

Keywords: 3′ UTR, 3′ untranslated region; ABCG1, ATP‐binding cassette transporter G1; AKT, protein kinase B; AP-1, activating protein-1; BAX, BCL-2 associated X; BCL-2, B cell lymphoma-2; CCK-8, cell counting kit-8; CHOP, C/EBP homologous protein; COX-2, cyclooxygenase-2; CVDs, cardiovascular diseases; ChIP, chromatin immunoprecipitation; DMEM, Dulbecco's modified eagle's medium; ELISA, enzyme-linked immunosorbent assay; FBS, fetal bovine serum; GEN, genistein; Genistein; HSP90, heat shock protein 90; IF, immunofluorescence; IGF-1, insulin-like growth factor-1; IL-6, interleukin 6; Inflammatory response; JNK, c-Jun N-terminal kinase; LDH, lactate dehydrogenase; LPS, lipopolysaccharide; Lipopolysaccharide; MUT, mutant; Macrophage apoptosis; MyD88, myeloid differentiation primary response 88; NF-κB, nuclear factor-kappa B; PBMCs, peripheral blood mononuclear cells; PBS, phosphate buffered saline; PDCD4, programmed cell death 4; PDTC, pyrrolidine dithiocarbamate; PMφs, peritoneal macrophages; STAT3, signal transducer and activator of transcription 3; TIPE2, tumor necrosis factor-α-induced protein 8-like 2; TLR4, toll-like receptor 4; TNF-α, tumor necrosis factor-α; TRIF, Toll/IL-1R domain-containing adaptor-inducing IFN-β; TiO2 NPs, titanium dioxide nanoparticles; Tumor necrosis factor-α-induced protein 8-like 2; WT, wild type; iNOS, inducible nitric oxide synthase; miR-21; qRT-PCR, quantitative real-time polymerase chain reaction.