Journal: Biomolecules

Article Title: Palmitic Acid Esterification Boosts Epigallocatechin Gallate’s Immunomodulatory Effects in Intestinal Inflammation

doi: 10.3390/biom15081208

Figure Lengend Snippet: Dot plots of representative pro-inflammatory (CD86-Coralite ® Plus 647, M1) and propidium iodide double stains of M0 macrophages (differentiated from THP-1 monocytic cells treated with Phorbol-Myristate Acetate) exposed to pro-inflammatory growth media of Caco-2 cells in the presence/absence of epigallocatechin gallate and palmitoyl epigallocatechin gallate. Distinct lowercase letters indicate significantly different values at p < 0.05 according to one-way analyses of variance (ANOVA) and Duncan’s multiple range test ( n = 3).

Article Snippet: CoraLite ® Plus 647 Anti-Human CD86 and CD206 antibodies were obtained from Proteintech ® (ThermoFisher Scientific, Madrid, Spain; Catalog #CL647-65165 and #CL647-65155, respectively).

Techniques:

Journal: Journal of photochemistry and photobiology. B, Biology

Article Title: ATG5-mediated keratinocyte ferroptosis promotes M1 polarization of macrophages to aggravate UVB-induced skin inflammation.

doi: 10.1016/j.jphotobiol.2024.112948

Figure Lengend Snippet: Fig. 6. ATG5-related keratinocyte ferroptosis contributes to M1 polarization of macrophages after UVB irradiation. (A) Macrophage chemotaxis was detected using Calcein AM fluorescence assay. Representative fluorescence images of adherent THP-1 cells (left) and quantification of adherent THP-1 cells (right). n = 3. (B) The recruitment of M1 macrophages in mice skin at 48 h after UVB exposure was analyzed by flow cytometry. The frequencies of CD86+ cells in CD45+CD11b+F4/80+ gated live cells were used for quantitative analysis. n = 3/group. (C) Representative images of immunofluo rescence staining of CD86 (green), F4/80 (red) and nuclear staining with Hoechst (blue). n = 3/group. (D) mRNA levels of indicators of M1 macrophages were assessed by qRT-PCR (n = 3). (E) Representative immunoblots of M1 macrophage marker CD86 from membrane protein were shown. ATP1A1 served as loading control, and GAPDH was used as a quality control for purity of the extracted membrane protein. Quantification of CD86 protein was shown (n = 3). *: p < 0.05; **: p < 0.01; ***: p < 0.001. (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article.)

Article Snippet: Primary antibodies against CD86 (#NBP2–25208) and F4/80 (#NB600–404) were purchased from Novus Biologicals (USA).

Techniques: Irradiation, Chemotaxis Assay, Fluorescence, Flow Cytometry, Staining, Quantitative RT-PCR, Western Blot, Marker, Membrane, Control

Journal: Journal of photochemistry and photobiology. B, Biology

Article Title: ATG5-mediated keratinocyte ferroptosis promotes M1 polarization of macrophages to aggravate UVB-induced skin inflammation.

doi: 10.1016/j.jphotobiol.2024.112948

Figure Lengend Snippet: Fig. 7. ATG5-related ferroptosis contributes to UVB-induced IFN-γ secretion, which exacerbates M1 polarization and skin inflammation. (A) Representative immunoblots of IFN-γ in mice dorsal skin (left) and quantification of it (right). n = 3/group. (B) The secretion of IFN-γ in supernatants of HaCaT cells was determined by ELISA at 24 h after 25 mJ/cm2 UVB exposure. n = 4/group. (C-F) WT C57Bl/6 J mice were pre-treated with either vehicle (UVB irradiated/ unirradiated groups) or Fer-1 (10 μm, UVB irradiated group) before 430 mJ/cm2 UVB radiation on dorsal skin. Every 12 h after irradiation, mice were treated with either vehicle or Fer-1 (10 μm). (C) Images showed the skin appearance at 24 and 48 h after exposure. Skin damage was evaluated by scores. n = 5/group. (D) Representative tissue morphology of dorsal skin from each group (H&E staining). Epidermal thickness (μm) was determined in skin sections. n = 5/group. (E) Representative immunoblots of GXP4, xCT, and IFN-γ. Quantification of protein bands of interest was shown (n = 3). (F) Recruitment of M1 macrophages in the dorsal skin at 48 h after UVB exposure was analyzed by flow cytometry. The frequencies of CD86+ cells in CD45+CD11b+F4/80+ gated live cells were used for quantitative analysis. n = 4/group. (G) The flowchart shows the process of the experiment of UVB-challenged control/Atg5 cKO mice with intradermal injection of 3000 IU recombinant mouse IFN-γ. (H) Representative images of the dorsal back from mice at 48 h after IFN-γ injection and 430 mJ/cm2 UVB radiation. Skin damage was evaluated by scores. n = 4/group. (I) Representative tissue morphology of dorsal skin from each group (H&E staining) and quantification of epidermal thickness (μm) of dorsal epidermis was shown. n = 4/group. (J) Recruitment of M1 macrophages in the dorsal skin at 48 h after UVB exposure was analyzed by flow cytometry. The frequencies of CD86+ cells in CD45+CD11b+F4/80+ gated live cells were used for quantitative analysis. n = 3/group. *: p < 0.05; **: p < 0.01; ***: p < 0.001. ns: nonsense.

Article Snippet: Primary antibodies against CD86 (#NBP2–25208) and F4/80 (#NB600–404) were purchased from Novus Biologicals (USA).

Techniques: Western Blot, Enzyme-linked Immunosorbent Assay, Irradiation, Staining, Flow Cytometry, Control, Injection, Recombinant

Journal: iScience

Article Title: Reduction of IFN-I responses by plasmacytoid dendritic cells in a longitudinal trans men cohort

doi: 10.1016/j.isci.2023.108209

Figure Lengend Snippet:

Article Snippet: CD86 / FITC / BU63 , BD Biosciences , Cat#: 374204 / RRID: AB_2721574.

Techniques: Recombinant, Enzyme-linked Immunosorbent Assay, Software

Journal: Cancers

Article Title: Hypoxia and Macrophages Act in Concert Towards a Beneficial Outcome in Colon Cancer

doi: 10.3390/cancers12040818

Figure Lengend Snippet: Hypoxia impacts macrophage antigen-presentation associated molecules. Macrophages were indirectly co-cultured with colon cancer cells RKO (MAC CC), at 20% or 1% O 2 for 72 h. Expression of the monocyte/macrophage lineage marker CD14, HLA-ABC (MHC-I), HLA-DR (MHC-II), and CD86 was determined by flow cytometry. ( A ) Pseudo-color plots display the gating strategy for flow cytometry created with FlowJo. ( B – D ) Graphs represent the percentage and median fluorescence intensity, which is presented as fold-change relatively to 20% O 2 condition of ( B ) CD14+HLA-ABC+, ( C ) CD14+HLA-DR+, or ( D ) CD14+CD86+ cells. Graphs represent the mean values with standard deviations, and are representative of n = 7 (HLA-ABC); n = 10 (HLA-DR); n = 6 (CD86) independent experiments. The statistical tests Wilcox or paired t -test were used; * p < 0.05. The p -values between 0.05 and 0.1 were presented and considered a tendency.

Article Snippet: Macrophages were immunostained with the following antibodies: anti-human CD14-APC (clone MEM-12), CD14-PE/FITC (clone 18D11), CD14-PerCP/Cy5.5 (clone OFC14D), CD86-FITC (clone BU63), HLA-ABC-PE (clone W6/32), HLA-DR-PE/FITC (clone MEM-12) (Immunotools), CCR7-PerCP/Cy5.5 (G043H7), SIRPα-APC (clone SE5A5) (Biolegend, San Diego, CA, USA), CD163-PE (clone GHI/61) (BD Bioscience, Franklin Lakes, NJ, USA).

Techniques: Immunopeptidomics, Cell Culture, Expressing, Marker, Flow Cytometry, Fluorescence

Journal: Journal of translational medicine

Article Title: SIRPB1 regulates inflammatory factor expression in the glioma microenvironment via SYK: functional and bioinformatics insights.

doi: 10.1186/s12967-024-05149-z

Figure Lengend Snippet: Fig. 2 A, B SIRPB1 expression in astrocytoma and glioblastoma cell clusters. C Correlation of SIRPB1 with immune cell infiltration. D Association between macrophage infiltration and SIRPB1; Wilcoxon and Spearman tests. E SIRPB1 and macrophage correlation in TCGA-GBM via TIMER2.0. F Kaplan–Meier curves for OS of TCGA-GBMLGG with ICR-high. G Co-localization of SIRPB1 with TMEM119, CD86, and CD163 in glioma. H SIRPB1 levels in glioma and monocyte lines from CCLE. I SIRPB1 expression in human and mouse glioma, microglia, and monocyte lines

Article Snippet: The induced THP-1 macrophages were collected, Fc receptors were blocked by human Fc Receptor Blocking Solution (Maokangbio), stained with FITC Anti-Mouse/ Human CD11b Antibody (Elabscience, clone:M1/70), 7-AAD (Elabscience), APC Anti-Human CD206 Antibody (Elabscience, clone:15–2) and PE Anti-Human CD86 Antibody (Elabscience, clone:BU63), and detected and analyzed by Beckman cytoflex flow cytometry.

Techniques: Expressing

Journal: Journal of translational medicine

Article Title: SIRPB1 regulates inflammatory factor expression in the glioma microenvironment via SYK: functional and bioinformatics insights.

doi: 10.1186/s12967-024-05149-z

Figure Lengend Snippet: Fig. 4 SIRPB1 Knockout and Macrophage Polarization. A T7E1 assay results. WT wild-type, NC negative control, PC positive control. B SIRPB1 and FLAG-cas9 expression in THP-1 lines. C Sanger sequencing of SIRPB1WT and SIRPB1KO. D Protein expression post-M1/M2 treatments. E mRNA levels of M1/M2 markers (*P < 0.05, **P < 0.01, ***P < 0.001, Dunnett’s test). F Flow cytometry of CD11b, CD86, CD206 in THP-1 lines

Article Snippet: The induced THP-1 macrophages were collected, Fc receptors were blocked by human Fc Receptor Blocking Solution (Maokangbio), stained with FITC Anti-Mouse/ Human CD11b Antibody (Elabscience, clone:M1/70), 7-AAD (Elabscience), APC Anti-Human CD206 Antibody (Elabscience, clone:15–2) and PE Anti-Human CD86 Antibody (Elabscience, clone:BU63), and detected and analyzed by Beckman cytoflex flow cytometry.

Techniques: Knock-Out, Negative Control, Positive Control, Expressing, Sequencing, Flow Cytometry