anti-mac3  (Bio-Rad)

Journal: Biochimica et biophysica acta. Molecular basis of disease

Article Title: Dietary apigenin ameliorates obesity-related hypertension through TRPV4-dependent vasorelaxation and TRPV4-independent adiponectin secretion.

doi: 10.1016/j.bbadis.2024.167488

Figure Lengend Snippet: Fig. 6. Apigenin reversed inflammatory responses and restored NAD+ levels in BAT. A Immunochemical staining for Mac3, CD45, and CD38 in BAT of mice fed a ND, HFD, or HFD + Api for 16 weeks. Arrows indicate positively enhanced signals of MAC, CD45 and CD38 in HFD mice, respectively. The scale bar indicates 50 μm. B-C Immunoblots of Mac3, CD45, and CD38 in the BAT of mice. D qPCR analyses of the mRNA levels of Mac3, CD45, and CD38 in the BAT of mice. E-F Immunoblots of CD38 in cultured brown adipocytes treated with BSA (300 μM) or PA (300 μM) with or without apigenin (5 μM) for 12 h. G qPCR analyses of mRNA levels of CD38 in cultured brown adipocytes treated with BSA or PA with or without apigenin. H Total intracellular NAD+ levels were measured in cell lysates (E). I Total intracellular NAD+ levels were measured in the BAT of mice (B). The data represent the mean ± SEM (n = 5), and the densitometric values for protein expression were normalized to β-tubulin. **P < 0.01, ***P < 0.001 compared with the ND (C-D, I) group or BSA (F-H) group,#P < 0.05, ##P < 0.01, ###P < 0.001 compared with the HFD (C-D, I) group or PA (F-H) group.

Article Snippet: Subsequently, the sections underwent microwave treatment in boiling antigen retrieval buffer (#C1034, Solarbio) for 20 min and were then blocked with QuickBlockTM Blocking Buffer (#P0260, Beyotime) for 1 h. Primary antibodies against Mac3 (Santa Cruz Biotechnology, sc-18,822, 1:600), CD45 (BD Biosciences, 550,539, 1:200), and CD38 (Abcam, ab108403, 1:200) were incubated with the sections overnight at 4 ◦C.

Techniques: Staining, Western Blot, Cell Culture, Expressing

Journal: BMC Cardiovascular Disorders

Article Title: Coenzyme Q10 mitigates macrophage mediated inflammation in heart following myocardial infarction via the NLRP3/IL1β pathway

doi: 10.1186/s12872-024-03729-x

Figure Lengend Snippet: IL1β production-related post-infarction cardiac inflammation was suppressed by CoQ10 treatment. (A) Analysis of gene expression in the infarct myocardium from each group of mice at 3 days after sham/LAD-ligation surgeries (Statistical analysis was performed using 2-way ANOVA with Sidak’s multiple comparison tests). Mice were treated with either vehicle or CoQ10. (B)(C) Representative images and analysis of the IL1β immunofluorescence staining in the Mac3-positive macrophages from mice treated with vehicle or CoQ10 at 28 days after LAD ligation (Statistical analysis was performed using a two-tailed unpaired Student’s t-test). 630×. Scale bars indicate 20 μm. (D) Representative flow cytometry analysis of left ventricles after sham/LAD ligation surgeries. Cells were defined as neutrophils (Neut, CD45.2 + CD64 − Ly6G + ), monocytes (Mono, CD45.2 + Ly6G − CD64 int Ly6C hi ), macrophages (Mac, CD45.2 + Ly6G − CD64 + Ly6C lo ), or CCR2 + macrophages (CCR2 + Mac, CD45.2 + Ly6G − CD64 + Ly6C lo CCR2 + ). (E) Quantification of the mean fluorescence intensity (MFI) of IL-1β in the CCR2 + macrophages of mice 3 days after LAD ligation, either treated with vehicle or CoQ10 (Statistical analysis was performed using a two-tailed unpaired Student’s t-test). MI: myocardial infarction; CoQ10: Coenzyme Q10; MFI: mean fluorescent intensity. * p < 0.05, ** p < 0.01, *** p < 0.001

Article Snippet: For Mac3 and IL-1β co-immunofluorescence staining, antigen retrieval with citric acid was performed, followed by blocking with 2.5% goat serum at room temperature for 1 h. Sections were incubated with rabbit anti-IL-1β polyclonal antibody (Bioss) at 1:100 dilution and anti-rat Mac3 antibody (Santa Cruz) at 1:100 dilution at 4 °C overnight.

Techniques: Expressing, Ligation, Comparison, Immunofluorescence, Staining, Two Tailed Test, Flow Cytometry, Fluorescence

Journal: Journal of Neuroinflammation

Article Title: IKKβ deletion from CNS macrophages increases neuronal excitability and accelerates the onset of EAE, while from peripheral macrophages reduces disease severity

doi: 10.1186/s12974-024-03023-9

Figure Lengend Snippet: Global depletion of myeloid IKKβ results in reduced axonal damage and oxidative stress in EAE. A Specimen images of paraffin-embedded coronal spinal cord sections from IKKβF/F (upper panel) and ΜφΙΚΚβKO (lower panel) mice with MOG 35-55 -induced EAE at 23 dpi (chronic phase). Serial sections were stained for markers of inflammation and demyelination, such as hematoxylin and eosin (H&E) and Luxol fast blue (LFB), respectively. Representative images from immunohistochemistry for markers of axonal damage (APP and Biel. Spheroids), immune cell infiltration (CD3 and Mac3), myeloid cell activation (iNOS) and macrophagic oxidative stress (p22 phox) are also shown. B Semi-quantitative scoring of immune cell infiltration (increased intensity of H/E staining) and demyelination (loss of LFB staining) of the same groups of mice shown in A . C Quantification index of axonal damage measured as levels of axonal APP and number of Biel. Spheroids of the same groups of mice shown in A . D Quantification index of myeloid cell activation measured as levels of iNOS and number of macrophagic p22 phox of the same groups of mice shown in A. Numbers of mice are annotated as scatter dots on the bars. All mice were adult females 2–4 months old

Article Snippet: The slices were then washed in PBS, blocked in 10% FBS in PBS for 1 h at room temperature and incubated in the following primary antibodies, rabbit anti-Iba1 (1/500; Wako chemicals; catalogue 019–19741), rabbit anti-GFAP (1/300; Dako; catalogue Z0334), rabbit anti-CD3 (1/2000; Neomarkers; RM-9107), anti-Mac3 (1/200; clone M3/84; Serotec), rabbit anti-rat iNOS (1/375; Chemicon; AB1631) or polyclonal rabbit anti-p22phox (1/100; Santa Cruz Biotexh; sc-20781), overnight at 4 °C.

Techniques: Staining, Immunohistochemistry, Activation Assay