Journal: JCI Insight

Article Title: ANGPTL8 links refeeding to monocyte dynamics and metabolic inflammation via the CCL5-CCR5 axis

doi: 10.1172/jci.insight.196605

Figure Lengend Snippet: ( A ) Representative images and quantification of monocyte migration induced by conditioned medium (CM) from BMDMs pretreated with or without rA8 over time ( n = 4). ( B ) Heatmap of chemokine-related DEGs (adjusted P < 0.05) in BMDMs cultured with or without rA8 (40 nM) stimulation for 24 hours ( n = 3). ( C ) mRNA expression of chemokines and their receptors after BMDMs were treated with rA8 or scrambled protein ( n = 6 cells examined over 3 independent experiments). ( D ) CCL5 levels in CM from BMDMs with rA8 stimulation over time ( n = 4). ( E ) Migration of BMDMs with or without CCR5 antagonist (TAK-220) before rA8 stimulation ( n = 5 cells examined over 3 independent experiments). ( F ) Effects of chemokine antagonists on migration of BMDMs with rA8 treatment ( n = 4). ( G ) Experimental scheme for CCR5 antagonist (TAK-220) gavage and rA8 injection in Loxp and Angptl8 HepOE mice. ( H ) Plasma proinflammation cytokine levels and ( I ) circulating monocyte populations of indicated groups ( n = 8 mice/group). ( J ) CCL5 levels in CM from human MDMs with or without rA8 stimulation ( n = 4). ( K ) Migration of U937 and THP-1 cells pretreated with or without CCR5 antagonist (TAK-220) before rA8 stimulation ( n = 5 cells examined over 3 independent experiments). Migrated cells were fixed and stained with 0.1% crystal violet ( A and F ). The data are shown as the mean ± SEM and were statistically analyzed by 2-tailed Student’s t test ( A , C , D , F , and J ) or 1-way ANOVA with Tukey’s multiple-comparison test ( E , H , I , and K ). All samples are biologically independent replicates, and n indicates the number of biologically independent samples examined. All the P values were 2 sided, and adjustments were made for multiple comparisons. * P < 0.05, ** P < 0.01, *** P < 0.001, **** P < 0.0001. DEGs, differentially expressed genes; MDMs, monocyte-derived macrophages; rA8, recombinant ANGPTL8; MCP1, monocyte chemoattractant protein-1 (CCL2); MIPs, macrophage inflammatory proteins, RANTES, regulated on activation normal T cell expressed and secreted (CCL5); CM, conditioned medium; TAK-220, CCR5 antagonist; R243, CCR8 antagonist; AMR69, CCL12 inhibitor; SB225002, CXCR2 antagonist; AMG487, CXCR3 antagonist; MIG-2F5.5, anti-CXCL9 antibody; SLW131, CCR7 antagonist.

Article Snippet: The CCR5 antagonist TAK-220 (MedChem Express, catalog HY-19974) was dissolved in DMSO and diluted in saline to achieve a maximal DMSO concentration of 1% and was applied daily by oral gavage.

Techniques: Migration, Cell Culture, Expressing, Injection, Clinical Proteomics, Staining, Comparison, Derivative Assay, Recombinant, Activation Assay

Journal: JCI Insight

Article Title: ANGPTL8 links refeeding to monocyte dynamics and metabolic inflammation via the CCL5-CCR5 axis

doi: 10.1172/jci.insight.196605

Figure Lengend Snippet: Effect of fasting-refeeding in the ( A ) plasma ANGPTL8 and ( B ) CCL5 levels of HFD-induced obese mice ( n = 8 mice/group). ( C ) Circulating monocyte populations and ( D ) proinflammation cytokines of fasting-refeeding mice fed with HFD. ( E ) Experimental scheme for CCR5 antagonist (TAK-220) gavage in Loxp and Angptl8 HepOE mice with fed with HFD. ( F ) Circulating monocyte populations and ( G ) proinflammation cytokine levels of the indicated groups ( n = 8 mice/group). ( H ) IPGTTs and ( I ) ITTs of the indicated groups ( n = 4 mice/group). All fasting/refeeding experiments were performed using a 12-hour fast during the dark phase (ZT2–14) followed by a 2-hour refeed. The data are shown as the mean ± SEM and were statistically analyzed by 1-way ANOVA with Tukey’s multiple-comparison test. All samples are biologically independent replicates, and n indicates the number of biologically independent samples examined. All the P values were 2 sided, and adjustments were made for multiple comparisons. ** P < 0.01, *** P < 0.001, **** P < 0.0001. HFD, high-fat diet; TAK-220, CCR5 antagonist; IPGTTs, i.p. glucose tolerance tests; ITTs, insulin tolerance tests; AUC, area under curve.

Article Snippet: The CCR5 antagonist TAK-220 (MedChem Express, catalog HY-19974) was dissolved in DMSO and diluted in saline to achieve a maximal DMSO concentration of 1% and was applied daily by oral gavage.

Techniques: Clinical Proteomics, Comparison

Journal: Advanced Science

Article Title: USP11‐PGAM5 Axis Promotes Neurotoxic Astrocyte Reactivity by Aggravating the mtDNA‐cGAS‐STING Pathway After Intracerebral Hemorrhage

doi: 10.1002/advs.202514283

Figure Lengend Snippet: Astrocytic PGAM5 mediates neutrophil TEM and NET release via the CCL5‐CCR5 chemotactic axis. A,B) Immunofluorescence staining of IBA1 and CD86, and statistical analysis of CD86+ cells in IBA1+ cells after ICH (n = 12 slices from 6 mice per group, Student's t test). Scale bar: 100 µm. Hemorrhage borders are demarcated by distinct dashed lines. C) Example gating strategy for flow cytometric analysis of brain immune cells. D–F) Fluorescent‐conjugated surface antibodies were employed to distinguish distinct cell populations: D) Ly6G⁺ neutrophils, E) Ly6G − Ly6C⁺ macrophages/monocytes, and F) CD3e⁺ T cells ( n = 6 per group, Student's t test). G) The heatmap demonstrated alterations in chemokine expression across experimental groups. H) Levels of chemokines CXCL16, CXCL10, CCL7, CXCL1, CXCL9, CCL2, CCL5, and CCL25 in the post‐ICH brain at 72 h were quantified via ELISA ( n = 6 per group, Student's t test). I,J) Flow cytometry assessed the impact of anti‐CCL5 antibody on Ly6G⁺ neutrophil infiltration levels in mouse brains at 24 h after ICH ( n = 6 per group, one‐way ANOVA). (K‐L) Quantification analysis of the CitH3 + cell number per mm 2 ( n = 6 per group, Student's t ‐test). Scale bar: 50 µm. M,N) Representative micrographs of IBA1 and CD86 expression, along with quantitative analysis of CD86+ cells in IBA1+ cells following anti‐CCL5 treatment ( n = 12 slices from 6 mice per group, Student's t ‐test). Scale bar: 100 µm. Data are presented as means ± SD. * p < 0.05, ** p < 0.01, *** p < 0.001.

Article Snippet: Moreover, the therapeutic potential of the CCL5‐CCR5 axis was evaluated by intraperitoneal administration of a CCL5‐neutralizing antibody (50 μg/mouse, R&D Systems, MAB478‐500) or a CCR5 antagonist (100 mg kg −1 , Selleckchem, S2003).

Techniques: Immunofluorescence, Staining, Expressing, Enzyme-linked Immunosorbent Assay, Flow Cytometry