Journal: Cells

Article Title: Statins Modulate Microenvironmental Cues Driving Macrophage Polarization in Simulated Periodontal Inflammation

doi: 10.3390/cells12151961

Figure Lengend Snippet: MyD88−dependent modulation of M2 polarization by simvastatin. ( A ) A schematic representation of simvastatin, MyD88 inhibitor treatment, and THP-1-macrophages (M0–M1–M2). ( B ) Cell surface antigen analysis of M2 macrophage markers CD68, CD163, and CD206 was conducted using flow cytometry. Expression levels were normalized using IgG isotype control, and values were expressed in percentage of positive cells (mean ± SD) from three independent preparations. ( C ) RT-PCR analysis and heat maps summarizing TLR and NFkB signaling from M0 (CCL2), M1 (LPS, IFNγ, TNFα), and M2 (IL4, IL10, TGFβ) macrophages. (n = 3) preparations per treatment group, and data found to be significantly different were plotted as heat maps. ( D ) Graphical summary of statin-mediated suppression of TLR and NFkB signaling.

Article Snippet: Phenotyping of macrophages was performed in 1% BSA and 3% human serum PBS according to standard methods using a panel of antibodies targeting CD68 (R and D Systems Cat# IC20401P, Minneapolis, MN, USA, RRID: http://scicrunch.org/resolver/AB_2074835 , accessed on 28 April 2023), CD163 (R and D Systems Cat# FAB1607P, RRID: http://scicrunch.org/resolver/AB_2074536 , accessed on 28 April 2023), and CD206 (R and D Systems Cat# FAB25342P, RRID: http://scicrunch.org/resolver/AB_10889015 , accessed on 28 April 2023); antibodies all from R and D Systems.

Techniques: Flow Cytometry, Expressing, Control, Reverse Transcription Polymerase Chain Reaction

Journal: Antioxidants

Article Title: Caffeic Acid Phenethyl Ester Suppresses Oxidative Stress and Regulates M1/M2 Microglia Polarization via Sirt6/Nrf2 Pathway to Mitigate Cognitive Impairment in Aged Mice following Anesthesia and Surgery

doi: 10.3390/antiox12030714

Figure Lengend Snippet: All primary and secondary antibodies used in this study.

Article Snippet: Mouse MMR/CD206 APC-conjugated Antibody , Goat , , 1:20 , R & D Systems , FAB2535A.

Techniques:

Journal: Marine Drugs

Article Title: Karenia brevis Extract Induces Cellular Entry through Distinct Mechanisms in Phagocytic RAW 264.7 Macrophages versus Non-Phagocytic Vero Cells

doi: 10.3390/md22010004

Figure Lengend Snippet: Assessment of macrophage alternative activation states by cytokine secretion (( A ) n-3) IL-10, TNF α and IL-6 and cell surface expression ( B ) of CD206 (n-10), CD80 (n-3) and IL4Rα (n-3). RAW 264.7 macrophages were treated with algal extracts for 24 h and stained with fluorescent antibodies prior to analysis on a flow cytometer. * indicates a statistically significant difference from vehicle control (VC) with p < 0.05.

Article Snippet: Antibodies included: (rat) anti-mannose receptor CD206 conjugated to Alexa Fluor 488 (R&D Systems, Minneapolis, MN, USA), (hampster) anti-mouse CD80 conjugated to PE-CF594 (BD Biosciences, San Jose, CA, USA), and (rat) anti-mouse IL4Rα conjugated to BV421 (BD Biosciences, San Jose, CA, USA).

Techniques: Activation Assay, Expressing, Staining, Flow Cytometry

Journal: Cell Death & Disease

Article Title: The ASIC3-M-CSF-M2 macrophage-positive feedback loop modulates fibroblast-to-myofibroblast differentiation in skin fibrosis pathogenesis

doi: 10.1038/s41419-022-04981-9

Figure Lengend Snippet: A Experimental design of fibroblast and macrophage indirect co-culture assay. B Representative images of THP-1 cells and 100 ng/ml PMA-derived M0 macrophages. C Flow cytometric analysis of CD68 in THP-1 cells and 100 ng/ml PMA-derived M0 macrophages. D Immunofluorescence staining of CD206 (green) in experimental and control macrophages after 48 h of co-cultivation. Scale bars, 100 μm. E , F Western blot and RT-qPCR analysis of CD206 protein levels and relative mRNA levels expression in the indicated groups ( n = 3). GAPDH served as a loading control. G Expressions of CD68 and CD206 were determined by flow cytometry. H Quantification of the flow cytometry assay results (n = 3). Data are expressed as the means ± SD. * P < 0.05, ** P < 0.01, and *** P ≤ 0.001. THP-1, human acute monocytic leukemia cell line; PMA, phorbol 12-myrisate 13-acetate; APC, allophycocyanin; FITC, fluorescein-isothiocyanate; GMQ, 2-guanidine-4-methylquinazoline.

Article Snippet: After blocking in goat serum at 37 °C for 30 min, the samples were incubated with antibodies against CD68 (Novus Biologicals, USA, NBP2-32831), CD206 (NovusBiologicals, USA, MAB2534), α-SMA (Novus Biologicals, USA, NBP2-33006), or Collagen I (Novus Biologicals, USA, NB600-408) in Diluent at 4 °C overnight.

Techniques: Co-culture Assay, Derivative Assay, Immunofluorescence, Staining, Control, Western Blot, Quantitative RT-PCR, Expressing, Flow Cytometry

Journal: Cell Death & Disease

Article Title: The ASIC3-M-CSF-M2 macrophage-positive feedback loop modulates fibroblast-to-myofibroblast differentiation in skin fibrosis pathogenesis

doi: 10.1038/s41419-022-04981-9

Figure Lengend Snippet: A Schematic diagram of establishing rabbit ear hypertrophic scar model and experimental plan. B Immunohistochemistry analysis of ASIC3 expression in rabbit ear tissue 21 days after wounding. Scale bars, 100 μm. C Immunofluorescence staining of rabbit ear tissue 21 days after wounding showing the total (CD68+, green) and M2 subtype (CD206+, red) macrophages after treatment by PBS, DMSO, or GMQ. Scale bars, 100 μm. D , E MFI quantification of CD68 and CD206 in (C) ( n = 3). F , G Western blot and RT-qPCR analysis of rabbit ear tissue 21 days after wounding. Total CD206 protein levels and relative mRNA levels expression in control, vehicle, and GMQ groups ( n = 3). H , I Relative mRNA expressions of M1 macrophage markers (iNOS, TNF-α) as evaluated by RT-qPCR ( n = 3). Data are expressed as the means ± SD. * P < 0.05, *** P ≤ 0.001. NS, not significant. iNOS, inducible nitric oxide synthase; TNF-α, tumor necrosis factor alpha.

Article Snippet: After blocking in goat serum at 37 °C for 30 min, the samples were incubated with antibodies against CD68 (Novus Biologicals, USA, NBP2-32831), CD206 (NovusBiologicals, USA, MAB2534), α-SMA (Novus Biologicals, USA, NBP2-33006), or Collagen I (Novus Biologicals, USA, NB600-408) in Diluent at 4 °C overnight.

Techniques: Immunohistochemistry, Expressing, Immunofluorescence, Staining, Western Blot, Quantitative RT-PCR, Control