Journal: Nature Communications
Article Title: Tissue-resident macrophage survival depends on mitochondrial function regulated by SerpinB2 in chronic inflammation
doi: 10.1038/s41467-026-69196-4
Figure Lengend Snippet: A , B SerpinB2 was quantified in THP-1 macrophages after stimulation with IFN-γ in the presence or absence of Bay-118072, an NF-kB inhibitor, by qPCR ( A ) (n = 8/group) and confocal imaging ( B ) (n = 10/group). A , B each dot represents cells cultured in one well. C VAT from lean and obese mice were stained for the markers of fibroblasts, macrophages, and adipocytes along with IFN-γ. The frequencies of IFN-γ-expressing cells were assessed by confocal microscopy (n = 5/group) Scale bar=20 µm. D – J LysM +/+ Ifngr1 fl/fl and LysM cre/+ Ifngr1 fl/fl mice were fed with an HFD for four months. D VAT resident macrophages, SerpinB2-expressing macrophages, and caspase 3 + resident macrophages were measured by confocal microscopy (n = 12/group). E The frequencies of apoptotic (annexin V + PI − ) macrophages were measured by flow cytometry (n = 5 for WT and 4 for KO). F GTT and ITT were performed, and fasting blood glucose, serum insulin, lipid concentrations, and body weights were evaluated (n = 10/group, combined data of at least 2 independent experiments). G , H Immunoblot showing pAkt, total Akt, Glut4, adiponectin, and Ppar y expressions in muscle, visceral adipose. and liver (n = 6/group). I , J qPCR was carried out to measure the expression of the metabolic and inflammatory genes in VAT (n = 7/group). K , L The top four transcription factors predicted to bind to the SerpinB2 promoter ( K ), and ChIP validation of Ikaros binding to the SerpinB2 promoter ( L ) (n = 4/group, with two sets of primers) are shown. M Ikaros quantification by qPCR in BMDM cultured in the presence or absence of 250 milli units IFN-γ (n = 10 and 8 for with and without IFN-γ, respectively). L , M each dot represents cells cultured in one well. N SerpinB2 expression in BMDM treated with or without IFN-γ and lenalidomide (representative image of the 3 independent experiments). O Ifngr1 and Ifngr2 were quantified by RNA seq in the adipose macrophage subsets (n = 3/group, each dot represents one mouse.). P – R Evaluation of apoptosis in BMDM lacking SerpinB2 (P) (n = 6/group), THP-1 macrophages overexpressing SerpinB2 ( Q ) (n = 4/group), and VAT resident macrophages in obese SerpinB2 +/+ and SerpinB2 − / − mice ( R ) (n = 4 for WT and 6 for KO) by flow cytometry. * P < 0.05, ** P < 0.01, *** P < 0.001. The Mann Whitney test (two-tailed) was used to determine the significance between two groups. One-way ANOVA with Bonferoni’s post hoc correction test was performed to determine differences among data obtained from more than two groups (Fig. 4B).
Article Snippet: In a separate experiment, BMDMs were treated with or without Ikaros inhibitor lenalidomide (5 μM) (MedChemExpress, #HYA0003) for 2 hours followed by stimulation with 250 milliunits of IFN- γ for 24 hours.
Techniques: Imaging, Cell Culture, Staining, Expressing, Confocal Microscopy, Flow Cytometry, Western Blot, Biomarker Discovery, Binding Assay, RNA Sequencing, MANN-WHITNEY, Two Tailed Test
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