Journal: Journal of experimental & clinical cancer research : CR

Article Title: Gastric cancer-derived exosomal let-7 g-5p mediated by SERPINE1 promotes macrophage M2 polarization and gastric cancer progression.

doi: 10.1186/s13046-024-03269-4

Figure Lengend Snippet: Fig. 2 High SERPINE1 expression in GC cells promotes macrophage M2 polarization. tSNE visualization of nine single-cell clusters partitioned by unsu pervised cluster analysis, SERPINE1 expression of each single-cell, and SERPINE1 expression abundance of different single-cell clusters in the GSE134520 (A–C) and GSE167297 (D–F) datasets. (G) Flow cytometry analysis of the proportion of CD68+CD206+ macrophages in a Transwell co-culture system, with MKN45 and AGS cells overexpressing (oe_SERPINE1) or silencing SERPINE1 (shRNA#3 or sh_SERPINE1#3) in the upper chamber, and THP1 cells treated with PMA in the lower chamber. (H) Immunofluorescence staining of xenograft tumor tissues. Comparison of the proportion of M1 or M2 macrophage infiltra tion. Green indicates F4/80. Red indicates iNOS or Arg1 expression

Article Snippet: THP-1 cells were differentiated into macrophages using 150 ng/mL phorbol 12-myristate 13-acetate (PMA, Sigma) for 24 h and subsequently co-cultured with cancer-derived exosomes or GC cells in 6-well plates with 0.4-μm membranes for 72 h. Harvested macrophages were converted into single-cell suspensions, stained with Elab Fluor 488 anti-human CD68 (Mouse, 1:20, ElabScience) and APC anti-human CD206 (Mouse, 1:20, ElabScience) antibodies, and analyzed for CD68+CD206+ populations by flow cytometry (Accuri C6, BD).

Techniques: Expressing, Flow Cytometry, Co-Culture Assay, shRNA, Immunofluorescence, Staining, Comparison

Journal: Journal of experimental & clinical cancer research : CR

Article Title: Gastric cancer-derived exosomal let-7 g-5p mediated by SERPINE1 promotes macrophage M2 polarization and gastric cancer progression.

doi: 10.1186/s13046-024-03269-4

Figure Lengend Snippet: Fig. 5 SERPINE1-mediated gastric cancer-derived exosomes facilitate the polarization of THP1 cells into M2 macrophages. (A) Schematic representation of the extraction and identification of exosomes and the induction of macrophage polarization. Transmission electron microscopy (B), nanoparticle tracking analysis (C), and western blotting (D) were used to identify the morphology, particle size, and markers of exosomes. (E) Confocal laser scanning microscopy detected Dil-labeled exosomes (red) internalized by DAPI-labeled macrophages (blue). (F–G) Immunofluorescence analysis of the proportion of CD206+ cells in THP1 cells treated with exosomes. (H–I) Flow cytometry analysis of the proportion of CD68+CD206+ cells in THP1 cells treated with exosomes. (J–K) qRT-PCR analysis of M1 markers (iNOS and TNF-α) and M2 markers (TGF-β, IL-10, and Arg-1) in THP1 cells treated with exosomes. (L–N) Transwell migration and invasion assays of GC cells (upper chamber) co-cultured with macrophages (lower chamber) ingesting exosomes

Article Snippet: THP-1 cells were differentiated into macrophages using 150 ng/mL phorbol 12-myristate 13-acetate (PMA, Sigma) for 24 h and subsequently co-cultured with cancer-derived exosomes or GC cells in 6-well plates with 0.4-μm membranes for 72 h. Harvested macrophages were converted into single-cell suspensions, stained with Elab Fluor 488 anti-human CD68 (Mouse, 1:20, ElabScience) and APC anti-human CD206 (Mouse, 1:20, ElabScience) antibodies, and analyzed for CD68+CD206+ populations by flow cytometry (Accuri C6, BD).

Techniques: Derivative Assay, Extraction, Transmission Assay, Electron Microscopy, Western Blot, Confocal Laser Scanning Microscopy, Labeling, Immunofluorescence, Flow Cytometry, Quantitative RT-PCR, Migration, Cell Culture

Journal: Cell metabolism

Article Title: Multiplexed in Situ Imaging Mass Cytometry Analysis of the Human Endocrine Pancreas and Immune System in Type 1 Diabetes

doi: 10.1016/j.cmet.2019.01.003

Figure Lengend Snippet: KEY RESOURCES TABLE

Article Snippet: 159Tb-CD68 (Clone KP1) , Fluidigm , Cat#3159035D.

Techniques: Software

Journal: Mucosal immunology

Article Title: A CCL24-dependent Pathway Augments Eosinophilic Airway Inflammation in House Dust Mite-challenged Cd163 −/− Mice

doi: 10.1038/mi.2015.94

Figure Lengend Snippet: (A) Confocal immunofluorescence microscopy images of BALF cells from an asthmatic subject showing CD163 expression by CD68 + AMΦs. The scale bar indicates 5 μm. (B) Gating strategy for identification of human CD163 + alveolar macrophages in BALF. Cellular debris was excluded using a forward light scatter/side scatter plot and doublets were excluded using width parameter on FSC and SSC properties. CD45 + cells, that co-expressed CD14 and CD68, were identified as alveolar macrophages using side scatter and CD45 bivariate plots from which lymphocytes had been excluded. A microscopic image of sorted CD45 + /CD14 + /CD68 + /CD163 + cells shows a cellular population possessing typical cellular characteristics of alveolar macrophages. (C) MFI of cell surface CD163 expression by CD45 + /CD14 + /CD68 + AMΦs in BALF from normal individuals and asthmatic subjects (n = 7, P < 0.008, paired t test). (D) A representative histogram overlay comparing cell surface CD163 expression by AMΦs from a normal individual and an asthmatic subject.

Article Snippet: Human BALF cell cytospin slides were fixed in 4% paraformaldehyde, incubated in blocking buffer (Aurion Blocking Solution, Electron Microscopy Sciences, Hatfield, PA) and reacted overnight with mouse anti-human CD68 (1:50 dilution, R & D Systems, Minneapolis, MN) and rabbit anti-human CD163 antibodies (1:50 dilution, Enzo Life Sciences, Inc., NY) diluted in 0.1% Aurion BSA-c (Electron Microscopy Sciences).

Techniques: Immunofluorescence, Microscopy, Expressing

Journal: Cancers

Article Title: Crosstalk between Macrophages and Myxoid Liposarcoma Cells Increases Spreading and Invasiveness of Tumor Cells

doi: 10.3390/cancers13133298

Figure Lengend Snippet: Immunohistochemical characterization of immune infiltrate in MLPS tissues. Representative images of H&E and IHC staining of CD3+, CD4+, CD8+, CD68+, and CD163+ positive cells in FFPE sections from a low grade MLPS ( a ) from patient #21 and high grade. MLPS tissues ( b ) from patient #4, acquired at 200× magnification.

Article Snippet: Monocytes recovered from co-cultures with/without MLPS cells were analyzed by flow cytometry as described [ ], using PE-conjugated anti-CD68 REAfinity™ (Miltenyi Biotec #130-114-460) and APC-conjugated anti-CD163 REAfinity™ (Miltenyi Biotec, #130-112–129, Bergisch Gladbach, Germany) antibodies.

Techniques: Immunohistochemical staining, Immunohistochemistry

Journal: Cancers

Article Title: Crosstalk between Macrophages and Myxoid Liposarcoma Cells Increases Spreading and Invasiveness of Tumor Cells

doi: 10.3390/cancers13133298

Figure Lengend Snippet: Correlation between immune cell infiltration and microvessel density in 26 low grade versus 24 high grade MLPS tissues. Box plots, showing variation in the average count of CD3+, CD4+, CD8+, FOXP3+, CD68+, and CD163+ cells as well as CD31 positive microvessels according to low or high histologic grade. Dark horizontal lines indicate the medians. Circles indicate outliers.

Article Snippet: Monocytes recovered from co-cultures with/without MLPS cells were analyzed by flow cytometry as described [ ], using PE-conjugated anti-CD68 REAfinity™ (Miltenyi Biotec #130-114-460) and APC-conjugated anti-CD163 REAfinity™ (Miltenyi Biotec, #130-112–129, Bergisch Gladbach, Germany) antibodies.

Techniques:

Journal: Cancers

Article Title: Crosstalk between Macrophages and Myxoid Liposarcoma Cells Increases Spreading and Invasiveness of Tumor Cells

doi: 10.3390/cancers13133298

Figure Lengend Snippet: Pearson correlation between the average count of CD163+ macrophages and ( a ) CD3+ T lymphocytes, ( b ) CD4+ T helper lymphocytes, ( c ) CD8+ cytotoxic lymphocytes, ( d ) CD68+ macrophages, and ( e ) CD31+ microvessels in 50 MLPS tissue sections. Pearson correlation coefficients (r) are indicated.

Article Snippet: Monocytes recovered from co-cultures with/without MLPS cells were analyzed by flow cytometry as described [ ], using PE-conjugated anti-CD68 REAfinity™ (Miltenyi Biotec #130-114-460) and APC-conjugated anti-CD163 REAfinity™ (Miltenyi Biotec, #130-112–129, Bergisch Gladbach, Germany) antibodies.

Techniques:

Journal: Cancers

Article Title: Crosstalk between Macrophages and Myxoid Liposarcoma Cells Increases Spreading and Invasiveness of Tumor Cells

doi: 10.3390/cancers13133298

Figure Lengend Snippet: MLPS cells trigger M2-like polarization of monocytes in non-contact co-cultures. ( a , b ) Representative images of H&E staining of FFPE sections from #37 ( a ) and #47 ( b ) MLPS tissues acquired at 200× magnification. ( c , d ) Primary MLPS cells obtained by enzymatic digestion of 37 ( c ) and #47 ( d ) tumor tissues, visualized by phase contrast microscopy (left) and fluorescent microscopy after immunostaining with anti-vimentin and anti-cytokeratin Abs (right). Nuclei were stained blue with DAPI. Original magnifications: 200× (left) and 400× (right). ( e , f ) Human monocytes were co-cultured with #37 ( e ) and #47 ( f ) primary MLPS cells in an in vitro non-contact co-culture for 72 h and then analyzed for CD68 and CD163 expression by flow cytometry. ( e , f ) Percent variation of CD68 and CD163 on monocytes collected after non-contact co-culture, compared to control monocytes. ( g , h ) After co-cultures with #37 ( g ) and #47 ( h ) primary MLPS cells, CMs from monocytes were analyzed for the content of CC2, IL-10, and IL-12 by a dot plot assay. The pixel density of each spot was measured using NIH Image J 2.0 software developed by the US NIH, USA and positive control spots were used to normalize results between the membranes. The intensity of each spot was averaged over the duplicate spots and expressed as percentage of each cytokine or chemokine spontaneously secreted by control monocytes (monocyte CM), considered as 100% (dashed line). Data represent mean ± SD from three experiments performed in duplicate with * p < 0.05, ** p < 0.005.

Article Snippet: Monocytes recovered from co-cultures with/without MLPS cells were analyzed by flow cytometry as described [ ], using PE-conjugated anti-CD68 REAfinity™ (Miltenyi Biotec #130-114-460) and APC-conjugated anti-CD163 REAfinity™ (Miltenyi Biotec, #130-112–129, Bergisch Gladbach, Germany) antibodies.

Techniques: Staining, Microscopy, Immunostaining, Cell Culture, In Vitro, Co-Culture Assay, Expressing, Flow Cytometry, Software, Positive Control

Journal: JTCVS Open

Article Title: Evaluation of tissue-engineered human acellular vessels as a Blalock–Taussig–Thomas shunt in a juvenile primate model

doi: 10.1016/j.xjon.2023.05.018

Figure Lengend Snippet: Histologic evaluation of host immune response. Midgraft sections from 3.5 mm HAV mBTT shunt explants stained by IHC for CD20 (B-cell), CD3 (T-cell), CD68 (macrophage), and CD11b (monocyte) markers. NHPs with midgraft dilatations (NHP1 and NHP3 [J-O]) exhibited markedly greater levels of host immune infiltration within the mBTT shunt wall than NHPs with no dilatations (NHP2, NHP4, NHP5 [A-I]). Host immune cells in explants from NHP1 and NHP3 were predominantly localized around the regions of dilatation. H&E , Hematoxylin and Eosin; NHP , nonhuman primate; HAV , Human Acellular Vessel; mBTT , modified Blalock–Taussig–Thomas shunt; IHC , immunohistochemistry; DAPI , 4′,6-diamidino-2-phenylindole.

Article Snippet: Explanted tissue sections and tissue slides (American MasterTech) were immunostained for alpha-smooth muscle actin (α-SMA; Dako M0851), von Willebrand Factor (vWF; Abcam 179451), CD3 (Dako A0451), CD20 (Abcam ab9475), CD11b (Abcam 52478), and CD68 (R&D Systems MAB2040).

Techniques: Staining, Modification, Immunohistochemistry

Journal: Cancer cell

Article Title: Dendritic cell paucity leads to dysfunctional immune surveillance in pancreatic cancer

doi: 10.1016/j.ccell.2020.02.008

Figure Lengend Snippet: KEY RESOURCES TABLE

Article Snippet: anti-human CD68 (Y1/82A) , Fluidigm , #3171011B.

Techniques: Recombinant, Cell Stimulation, Enzyme-linked Immunosorbent Assay, CyQUANT Assay, LDH Cytotoxicity Assay, Expressing, Staining, Derivative Assay, Software

Journal: Advanced Materials (Deerfield Beach, Fla.)

Article Title: A Pseudo‐ Mytilus Edulis Foot Protein‐Based Hydrogel Adhesive with Osteo‐Vascular‐Immune Coupling Effects for Osteoporotic Bone‐Implant Integration

doi: 10.1002/adma.202511840

Figure Lengend Snippet: Bioglue effectively modulates the immune microenvironment at the screw‒bone interface. A) Schematic diagram of the animal experimental design (created with BioRender.com). B) H&E staining of the screw‒bone interface tissue. C) Immunofluorescence staining of macrophage infiltration at the screw‐bone interface. D) IHC analysis of the screw‒bone interface. E) Senescence‐associated immunofluorescence staining of the screw‒bone interface tissue. F,G) Proportion of iNOS + and Arg‐1 + macrophages (iNOS + & CD68 + /Arg‐1 + & CD68 + ) in tissue fluorescent‐stained sections ( n = 5). The data are presented as the means ± SDs in the analysis figures, with * p < 0.05 and ** p < 0.01 indicating statistical significance.

Article Snippet: Sections were incubated with primary antibodies, iNOS (ABclonal, A3774), Arg‐1 (ABclonal, A25808), CD68 (Proteintech, 3A9A7), P21 (ABclonal, A19094), VEGF (ABclonal, A19094), IL‐10 (ABclonal, A2171), TNF‐α (ABclonal, A11534), at 4 °C overnight.

Techniques: Staining, Immunofluorescence