hy  (MedChemExpress)

Journal: Redox Biology

Article Title: Epithelial redox stress programs macrophage immunometabolism through a ZNF24-MIF–NF–κB pathway in chronic nonbacterial prostatitis

doi: 10.1016/j.redox.2026.104042

Figure Lengend Snippet: Pharmacological inhibition of MIF by ISO-1 attenuates prostatitis severity and suppresses M1 macrophage polarization in vivo . (A) Mechanical allodynia assessed by von Frey testing, showing reduced response frequency in ISO-1-treated mice. (B) Representative H&E staining of prostate sections showing decreased inflammatory infiltration after ISO-1 treatment. (C) Quantification of histopathological inflammation scores. (D) ELISA quantification of TNF-α, IL-6, and IL-1β in serum confirming cytokine downregulation. (E) Immunofluorescence staining for CD45 (red) indicating reduced immune cell infiltration. (F) Quantification of CD45 + immune cell infiltration in prostate tissues. (G–H) Flow cytometry of F4/80+CD11b + CD86 + macrophages showing reduced M1 subset after ISO-1. (I) Co-immunofluorescence for CD68 (red) and iNOS (green) showing reduced M1 macrophages in ISO-1-treated mice. (J) Western blot confirming downregulation of CD86 and iNOS in prostate tissues. Data are presented as the mean ± SD. ns, not significant; ∗p < 0.05; ∗∗p < 0.01; ∗∗∗p < 0.001. Abbreviations: CTR, control; EAP, experimental autoimmune prostatitis; qRT‒PCR, quantitative real-time PCR.

Article Snippet: Following the second immunization, the mice were treated daily for 14 days with ISO-1 (3.5 mg/kg in 10 % dimethyl sulfoxide (DMSO)/90 % corn oil; Cat. No. HY-16692; MCE) [ ] via intraperitoneal injection, whereas the controls received vehicle alone.

Techniques: Inhibition, In Vivo, Staining, Enzyme-linked Immunosorbent Assay, Immunofluorescence, Flow Cytometry, Western Blot, Control, Real-time Polymerase Chain Reaction

Journal: Redox Biology

Article Title: Epithelial redox stress programs macrophage immunometabolism through a ZNF24-MIF–NF–κB pathway in chronic nonbacterial prostatitis

doi: 10.1016/j.redox.2026.104042

Figure Lengend Snippet: Epithelial ROS-ZNF24 axis drives MIF transcription and promotes CD74-dependent M1 macrophage polarization. (A–E) CD74 knockdown attenuates MIF-induced M1 macrophage polarization, as assessed by CD86 and iNOS expression (A–B), proinflammatory cytokine secretion (C), and flow cytometry analysis of F4/80 + CD86 + macrophages (D–E). (F–H) LPS stimulation induces MIF mRNA expression (F), protein expression (G), and secretion (H) in RWPE-1 prostate epithelial cells. (I–K) Transwell co-culture system showing that LPS-stimulated prostate epithelial cells promote M1 polarization and cytokine secretion in iBMDMs, which is suppressed by the MIF inhibitor ISO-1. (L–M) Increased epithelial oxidative stress in EAP mice and LPS-stimulated RWPE-1 cells, indicated by 8-OHdG staining (L) and intracellular ROS levels (M), respectively; NAC effectively reduces ROS accumulation. (N–O) ROS scavenging with NAC suppresses epithelial MIF expression and attenuates M1 marker expression in co-cultured macrophages. (P–T) ZNF24 is induced by epithelial ROS and directly regulates MIF transcription, as shown by ZNF24 expression (P), ZNF24 knockdown (Q), predicted ZNF24 binding motifs in the MIF promoter (R), and ChIP assays demonstrating enhanced ZNF24 binding upon LPS stimulation (S–T). Data are presented as mean ± SD. ns, not significant; ∗p < 0.05; ∗∗p < 0.01; ∗∗∗p < 0.001. Abbreviations: iBMDMs, immortalized bone marrow-derived macrophages; RWPE-1, human prostate epithelial cell line; siCD74, small interfering RNA targeting CD74; LPS, lipopolysaccharide; NAC, N-acetylcysteine; ROS, reactive oxygen species.

Article Snippet: Following the second immunization, the mice were treated daily for 14 days with ISO-1 (3.5 mg/kg in 10 % dimethyl sulfoxide (DMSO)/90 % corn oil; Cat. No. HY-16692; MCE) [ ] via intraperitoneal injection, whereas the controls received vehicle alone.

Techniques: Knockdown, Expressing, Flow Cytometry, Co-Culture Assay, Staining, Marker, Cell Culture, Binding Assay, Derivative Assay, Small Interfering RNA

Journal: Redox Biology

Article Title: Epithelial redox stress programs macrophage immunometabolism through a ZNF24-MIF–NF–κB pathway in chronic nonbacterial prostatitis

doi: 10.1016/j.redox.2026.104042

Figure Lengend Snippet: Pharmacological inhibition of NF-κB mitigates MIF-CD74-driven inflammation and M1 polarization in chronic prostatitis. (A) Heatmap of GSEA HALLMARK pathways showing increased inflammatory and NF-κB-related signaling in EAP, reversed by ISO-1. (B) Quantification of TNFA signaling via NF-κB scores from HALLMARK enrichment based on RNA-seq results derived from mouse model. (C) Heatmap of key NF-κB pathway genes, upregulated in EAP and suppressed by ISO-1 based on RNA-seq results derived from mouse model. (D–E) Single-cell transcriptomic analysis showing increased NF-κB pathway activity in EAP prostates using AddModuleScore. (F) IHC of p-p65 confirming NF-κB activation in EAP and inhibition by ISO-1 or CD74 blockade. (G) In vitro validation in iBMDMs: MIF-induced p65 phosphorylation reversed by si CD74 . (H) Confocal immunofluorescence showing MIF-induced nuclear translocation of p65 in iBMDMs, abolished by si CD74 . (I) Western blot showing suppression of MIF-induced CD86 and iNOS upregulation by the NF-κB inhibitor JSH-23. (J) Confocal staining showing decreased iNOS after JSH-23 treatment in MIF-stimulated iBMDMs. (K–L) Flow cytometry and quantification of F4/80+CD86 + macrophages showing reduced M1 polarization with JSH-23. (M) ELISA showing suppression of IL-6, IL-1β, and TNF-α secretion by JSH-23. Data are presented as the mean ± SD. ns, not significant; ∗p < 0.05; ∗∗p < 0.01; ∗∗∗p < 0.001. Abbreviations: GSEA, Gene Set Enrichment Analysis; HALLMARK, curated gene set collection; AddModuleScore, single-cell pathway activity scoring method; JSH-23, NF-κB inhibitor.

Article Snippet: Following the second immunization, the mice were treated daily for 14 days with ISO-1 (3.5 mg/kg in 10 % dimethyl sulfoxide (DMSO)/90 % corn oil; Cat. No. HY-16692; MCE) [ ] via intraperitoneal injection, whereas the controls received vehicle alone.

Techniques: Inhibition, RNA Sequencing, Derivative Assay, Activity Assay, Paraffin-embedded Immunohistochemistry, Activation Assay, In Vitro, Biomarker Discovery, Phospho-proteomics, Immunofluorescence, Translocation Assay, Western Blot, Staining, Flow Cytometry, Enzyme-linked Immunosorbent Assay

Journal: Redox Biology

Article Title: Epithelial redox stress programs macrophage immunometabolism through a ZNF24-MIF–NF–κB pathway in chronic nonbacterial prostatitis

doi: 10.1016/j.redox.2026.104042

Figure Lengend Snippet: Schematic illustration of the epithelial ROS-ZNF24-MIF-CD74-PKM2-NF-κB signaling axis in chronic prostatitis. In response to epithelial injury or inflammatory stimuli, excessive ROS accumulate in prostatic epithelial cells, leading to the activation of the redox-responsive transcription factor ZNF24. Activated ZNF24 directly binds to the MIF promoter and drives MIF transcription, resulting in enhanced MIF production and release. Epithelial-derived MIF subsequently acts in a paracrine manner by binding to CD74 on macrophages. This interaction promotes the stabilization and nuclear translocation of PKM2, which acts as a co-activator to increase NF-κB signaling via p65 nuclear translocation. PKM2 activation also enhances glycolytic flux and contributes to mitochondrial dysfunction, further promoting M1 macrophage polarization and inflammatory cytokine production. Disruption of this axis via ISO-1, CD74 blockade, or PKM2 modulation (DASA-58) effectively mitigates chronic inflammation in the prostate.

Article Snippet: Following the second immunization, the mice were treated daily for 14 days with ISO-1 (3.5 mg/kg in 10 % dimethyl sulfoxide (DMSO)/90 % corn oil; Cat. No. HY-16692; MCE) [ ] via intraperitoneal injection, whereas the controls received vehicle alone.

Techniques: Activation Assay, Derivative Assay, Binding Assay, Translocation Assay, Disruption