Journal: American Journal of Respiratory Cell and Molecular Biology

Article Title: IL-13–regulated Macrophage Polarization during Granuloma Formation in an In Vitro Human Sarcoidosis Model

doi: 10.1165/rcmb.2018-0053oc

Figure Lengend Snippet: Figure 5: Immunofluorescence imaging of granuloma-like structures demonstrates persistent macrophage CD163 expression on sarcoidosis PBMCs following PPD stimulation. Representative photomicrographs of immunostained PBMCs in granuloma-like structures 7 days after bead treatment using

Article Snippet: Copyright © 2018 by the American Thoracic Society Alternatively, following washing and blocking, some coverslips were incubated with 0.5 μg/ml CD163 [unlabeled, rabbit, anti-human CD163 mAb (K20-T, IgG) from Novus Biologicals, LLC (Littleton, CO)] or the matching isotype control antibody [Abcam, (Cambridge, MA)] at room temperature for 1 hour.

Techniques: Immunofluorescence, Imaging, Expressing

Journal: Cell Death & Disease

Article Title: M2 macrophage-mediated interleukin-4 signalling induces myofibroblast phenotype during the progression of benign prostatic hyperplasia

doi: 10.1038/s41419-018-0744-1

Figure Lengend Snippet: a Representative CD68 immunohistochemical staining images in the early-progressed BPH, age-matched prostate and elderly BPH tissues. Scale bar, 200 μm. b Scatter plots showing macrophage clusters in the three analysed groups. The number of macrophage clusters in prostate tissues correlated positively with the stroma-to-epithelium ratio, and percent area densities of α-SMA, collagen I and fibres. c Serial histological sections showing the early-progressed BPH tissues stained for CD68, a macrophage-specific marker, and CD163, an M2 macrophage marker. The distribution of CD68 + and CD163 + cells was almost identical. Scale bar, 200 μm. d Representative immunofluorescence (IF) staining images, showing CD68 (green) and CD163 (red) expression in the infiltrating macrophages in early-progressed BPH tissues. Scale bar, 100 μm. e Quantitative RT-PCR results showing α-SMA, COL1A1 and COL3A1 expression in primary prostate fibroblast (PrPF)-early, PrPF-control and PrPF-old cells co-cultured with THP-1-derived M2 macrophages. Data are shown as relative gene expression compared to that in the respective untreated fibroblasts. f Western blots showing α-SMA and collagen I protein expression in PrPF-early, PrPF-control and PrPF-old cells co-cultured with THP-1-derived M2 macrophages. g Representative IF staining results, showing the co-expression of α-SMA (red) and collagen I (green) following a 48-h co-culture of PrPF-early, PrPF-control and PrPF-old cells with THP-1-derived M2 macrophages. The respective untreated fibroblasts were used as controls. Scale bar, 100 μm. h Solidified collagen-gel shrinkage after the seeding of PrPF-early, PrPF-control and PrPF-old cells, untreated or treated with the conditioned medium. The respective untreated fibroblast samples were used as controls, and all assays were performed in triplicate. * P < 0.05, ** P < 0.01, *** P < 0.001

Article Snippet: The primary antibodies included mouse monoclonal α-SMA (Abcam, ab7817, 1:100 dilution), CD163 (OriGene, TA506380, 1:200 dilution) and EpCAM (Cell Signaling Technology, #2929, 1:800 dilution) antibodies, and the rabbit primary antibodies included collagen I (Abcam, ab138492, 1:800 dilution), CD68 (Proteintech, 25747-1-AP, 1:400 dilution), IL4R (ImmunoWay, YT2337, 1:400 dilution), TGFβ RI (ImmunoWay, YT4627, 1:400 dilution) and vimentin (Cell Signaling Technology, #5741, 1:100 dilution) antibodies.

Techniques: Immunohistochemical staining, Staining, Marker, Immunofluorescence, Expressing, Quantitative RT-PCR, Control, Cell Culture, Derivative Assay, Gene Expression, Western Blot, Co-Culture Assay

Journal: Human reproduction (Oxford, England)

Article Title: Distribution of decidual natural killer cells and macrophages in the neighbourhood of the trophoblast invasion front: a quantitative evaluation.

doi: 10.1093/humrep/det353

Figure Lengend Snippet: Figure 5 Distribution of dMph. (a) Cryostat section from a cell column at a first trimester placentation site double-stained with an anti-CD163 antibody (red) and an anti-HLA-G antibody (dark blue) as a sample image. Scale bar ¼ 100 mm. (b) Box plots of normalized areal cell densities s of dMph at first trimester placentation sites according to Equation (15), in five trophoblast neighbourhood rings according to Equation (17), are depicted. The dashed line s ¼ 1 represents the average areal cell density. #P ¼ 0.05, ##P ¼ 0.01 compared with 1 by Student’s t-tests (H0:m ¼ 1). ANOVA analysis showed a high significance,P ≤0.01.*P ¼ 0.05,**P ¼ 0.01ofpair-wiseTukeyhonestsignificantdifferenceposthoctests(s ¼ 1).(c)DistributionofdMphinco-culturesof first trimester decidua parietalis with first trimester villous explants using cryostat sections from co-cultures double-stained with an anti-CD163 antibody and an anti-HLA-G antibody. Box plots of normalized areal cell densities s. #P ¼ 0.05, ##P ¼ 0.01 compared with 1 by Student’s t-tests (H0:m ¼ 1). ANOVA analysis showed a high significance, P ≤0.01. **P ¼ 0.01 of pair-wise Tukey honest significant difference post hoc tests (s ¼ 1). Open circle is an outlier. (d) Distribution of dMph in first trimester decidua parietalis determined by using cryostat sections from tissue fragments stained with an anti-CD163 antibody. Box plots of normalized areal cell densities s. P . 0.05 by ANOVA test. Filled circle is an extreme value.

Article Snippet: The following antibodies were used: † monoclonal mouse anti-human CD56 (clone 123C3; Santa Cruz Biotechnology, Inc., Santa Cruz, CA, USA), dilution: 1:400, final concentration: 0.5 mg/ml; † monoclonal mouse anti-human CD163 (clone 5C6-FAT; Acris Antibodies GmbH, Hiddenhausen, Germany), dilution: 1:500, final concentration: 0.4 mg/ml; † monoclonal mouse anti-human HLA-G (clone 4H84; Exbio, Praha, Czech Republic; supplied by Eubio, Vienna, Austria), dilution: 1:500, final concentration: 2 mg/ml.

Techniques: Staining

Journal: Toxicologic Pathology

Article Title: Biocompatible Solutions: Evaluating the Safety of Repeated Intra-Articular Injections of pMPCylated Liposomes for Knee Osteoarthritis Therapy in Rat Models

doi: 10.1177/01926233241271400

Figure Lengend Snippet: Antibodies used for the immunofluorescence study.

Article Snippet: Novus (CD163) , NBP3-07981 (CD163).

Techniques: Immunofluorescence, Marker

Journal: Nature Communications

Article Title: The neonatal Fc receptor (FcRn) is a pan-arterivirus receptor

doi: 10.1038/s41467-024-51142-x

Figure Lengend Snippet: A Growth of SHFV and PRRSV-2 in wild-type MA-104 cells, inoculated at MOI = 0.01 ( n = 2 biological replicates). Data shows representative results from one of two independent experiments. B Cytopathic effect (CPE) of arterivirus infection of MA-104 cells at 4 days post-inoculation, taken at ×100 magnification. Data shows representative results from one of two independent experiments. C Frequency of sgRNAs, as determined via deep sequencing of genomic DNA from MA-104 cells, prior to and after infection with arteriviruses. D Robust rank aggregation scores (RRA) scores for gene knockouts enriched in surviving MA-104 cells compared to pre-infection; top 10 hits for each screen are shown as colored datapoints. E RRA scores of SHFV vs. PRRSV-2, with CD163 , FCGRT (FcRn), and B2M circled and labeled. F Infection of cells transduced with single-guide RNA targeting grivet (for MA-104) or human (for ACHN) FCGRT (ΔFcRn), or empty vector control. Viruses tested on these cells are arranged in terms of relatedness to arteriviruses from left to right, with taxonomic relationships shown along the top. Data are presented as mean values ± SEM. Infections were performed with n = 3 biological replicates at an MOI of 0.01. Supernatant was titrated via plaque assay or focus-forming assay at the indicated time points; asterisks represent p-values (**p ≤ 0.01; *** p ≤ 0.001) derived from a two-tailed unpaired t-test with Welch correction and multiple comparisons testing. G Brightfield photographs of arterivirus infections from F taken at ×100 magnification at 3 days post-inoculation. Data shows representative results from one of two independent experiments. Source data are provided as a Source Data file.

Article Snippet: Primary antibodies, including CD163 (goat polyclonal anti-CD163 antibody [R&D Systems, #AF1607]), FcRn (rabbit polyclonal anti-FcRn [Abcam, #ab193148]), and beta-actin (mouse monoclonal 8H10D10 [Cell Signaling Technology, #3700 S]) were diluted in TBST + 5% nonfat dried milk and incubated either overnight at 4 °C (CD163 and FcRn) or for 1 h at ambient temperature (beta-actin).

Techniques: Infection, Sequencing, Labeling, Transduction, Plasmid Preparation, Control, Plaque Assay, Focus Forming Assay, Derivative Assay, Two Tailed Test

Journal: Nature Communications

Article Title: The neonatal Fc receptor (FcRn) is a pan-arterivirus receptor

doi: 10.1038/s41467-024-51142-x

Figure Lengend Snippet: A Western blot showing gCD163 and gFcRn expression in wild-type (WT) MA-104 cells and MA-104 cells with ectopic expression of gCD163, gFcRn, or both. B Cells from A were infected with SHFV (MOI = 30) and assayed for intracellular SHFV RNA 4 h later using single molecule RNA fluorescence in situ hybridization (smRNA FISH). Confocal microscopy (×600 magnification) was used to capture 16–18 fields of view taken at random (one representative shown). Blue = nuclei; red = smFISH detecting viral (v)RNA, Scale bar = 25 μm. C Quantification of microscopy performed in B . From each field of view, the percent vRNA positive cells, average number of puncta per infected cell, and average puncta FISH intensity per infected cell were determined and graphed as independent points. Error bars represent the mean ± SEM. Data shows results from one experiment. D Time course of SHFV production from wild-type (WT) MA-104 cells (red) compared to dual-overexpressing gCD163/gFcRn cells (black) at a low (0.03, left) and high (3, right) MOI, n = 3 biological replicates. E Cytopathic effect of SHFV infection of wild-type (WT) MA-104 cells (red) compared to dual-overexpressing gCD163/gFcRn cells (black) at a low MOI (0.03) 24 hrs after inoculation. Data shows results from one representative experiment in D . F Photographs of plaque assay wells (6-well plate) fixed and stained with crystal violet 2 days after inoculation. Note the larger and more well-circumscribed plaques in the CD163/FcRn-dual-expressing cells. G Quantification of SHFV via plaque assay using MA-104 cells (WT, +gCD163, +gFcRn, or +gCD163 & +gFcRn) as substrate for infection with data presented as mean values ± SEM, n = 3 biological replicates, normalized to WT cells and analyzed using one-way ANOVA with multiple comparisons relative to WT cells: ns, not significant; ***p ≤ 0.001; ****p ≤ 0.0001. Source data are provided as a Source Data file.

Article Snippet: Primary antibodies, including CD163 (goat polyclonal anti-CD163 antibody [R&D Systems, #AF1607]), FcRn (rabbit polyclonal anti-FcRn [Abcam, #ab193148]), and beta-actin (mouse monoclonal 8H10D10 [Cell Signaling Technology, #3700 S]) were diluted in TBST + 5% nonfat dried milk and incubated either overnight at 4 °C (CD163 and FcRn) or for 1 h at ambient temperature (beta-actin).

Techniques: Western Blot, Expressing, Infection, Fluorescence, In Situ Hybridization, Confocal Microscopy, Microscopy, Plaque Assay, Staining

Journal: Nature Communications

Article Title: The neonatal Fc receptor (FcRn) is a pan-arterivirus receptor

doi: 10.1038/s41467-024-51142-x

Figure Lengend Snippet: A FcRn orthologs from natural and potential arterivirus hosts—grivet (g), human (hu), mouse (m), pig (p), and horse (ho)–were introduced into MA-104ΔFcRn cells and inoculated with SHFV (red), PRRSV-2 (green), or EAV (blue) at an MOI of 0.01, n = 2 biological replicates. FcRn orthologs derived from the natural host of an arterivirus (i.e., positive control) are denoted by a black outline. Productive infection was then assessed via plaque assay of supernatant at 2 days post inoculation. B Similar experimental setup to A , except with CD163 orthologs in MA-104ΔCD163 cells. Source data are provided as a Source Data file.

Article Snippet: Primary antibodies, including CD163 (goat polyclonal anti-CD163 antibody [R&D Systems, #AF1607]), FcRn (rabbit polyclonal anti-FcRn [Abcam, #ab193148]), and beta-actin (mouse monoclonal 8H10D10 [Cell Signaling Technology, #3700 S]) were diluted in TBST + 5% nonfat dried milk and incubated either overnight at 4 °C (CD163 and FcRn) or for 1 h at ambient temperature (beta-actin).

Techniques: Derivative Assay, Positive Control, Infection, Plaque Assay