Journal: Frontiers in Bioengineering and Biotechnology

Article Title: An Inflamed Human Alveolar Model for Testing the Efficiency of Anti-inflammatory Drugs in vitro

doi: 10.3389/fbioe.2020.00987

Figure Lengend Snippet: Phenotype shift in macrophage monocultures upon exposure to pro- and anti-inflammatory stimuli. (A) Secretion of pro-inflammatory mediators (IL-8 and TNF-α; in pg/mL) in cell-culture medium in the basal compartment, assessed via ELISA. (B) Expression of CD206 surface marker (median fluorescence intensity), denoting M2 phenotype, assessed via flow cytometry analysis. The CD206 intensity histogram is shown in . (C) Cell viability of MDMs after all of the treatments, assessed via membrane rupture (LDH assay), presented as fold increase over untreated cells (the dotted line). The data is presented as the mean of the four biological repetitions ± standard deviation, whereas individual values from biological repetitions are presented as circles, color-coded for the data from the same biological repetitions (donors). Statistically significant differences among the groups (One-way ANOVA, Tukey’s post hoc ; α = 0.05): * p ≤ 0.05; ** p ≤ 0.01; *** p ≤ 0.001, **** p ≤ 0.0001. Abbreviations: IL-8, interleukin 8; TNF-α, tumor necrosis factor α; MFI, median fluorescence intensity; CD206, cluster of differentiation 206, mannose receptor; IL-4+IL-13, interleukins 4 and 13 (both applied at 20 ng/mL for 48 h); LDH, lactate dehydrogenase; Pos. ctrl Triton X, positive control for LDH assay, i.e., cells exposed to 0.2% Triton X-100 (v/v; 24 h).

Article Snippet: Cells from two identically treated wells in a 6-well plate were pooled in flow cytometry tubes (FALCON ® 5 mL Polystyrene Round-Bottom Tube, Corning, Switzerland), counted, centrifuged (5702R, Eppendorf; 500 RCF, 5 min), and resuspended in cold flow cytometry buffer (PBS with 1% BSA, 0.1% NaN3, 1 mM ethylenediaminetetraacetic acid [EDTA] at pH 7.4).

Techniques: Cell Culture, Enzyme-linked Immunosorbent Assay, Expressing, Marker, Fluorescence, Flow Cytometry, Membrane, Lactate Dehydrogenase Assay, Standard Deviation, Positive Control

Journal: Stem Cell Reviews and Reports

Article Title: p53 Inhibition in Pancreatic Progenitors Enhances the Differentiation of Human Pluripotent Stem Cells into Pancreatic β-Cells

doi: 10.1007/s12015-023-10509-1

Figure Lengend Snippet: Inhibition of p53 enhances generation of hPSC-pancreatic progenitors. ( A - B ) Flow cytometry analysis and quantification showing the co-expression of PDX1 and NKX6.1 after treating the cells during stage 4 with DMSO, pifithrin-α, and pifithrin-μ. ( C ) Flow cytometry quantification of the co-expression of PDX1 and NKX6.1 after treating the cells during stage 4 with DMSO, and pifithrin-μ (10 μM and 20 μM). Flow cytometry analysis ( D ) and quantification ( E ) of PDX1 and NKX6.1 expression in DMSO or pifithrin-μ - treated pancreatic progenitors ( n = 3). Flow cytometry analysis ( F ) and quantification ( G ) of CHGA and NKX6.1 expression in DMSO or Pifithrin-μ - treated cells ( n = 3). Data are represented as mean ± SD; * p < 0.05, ** p < 0.01, *** p < 0.001

Article Snippet: Stained cells were filtered through a 40-μm nylon mesh into flow cytometry tubes (BD Falcon) and were analyzed with BD Accuri™ C6 flow cytometer (BD Biosciences, USA) and FlowJo for data analysis.

Techniques: Inhibition, Flow Cytometry, Expressing

Journal: Stem Cell Reviews and Reports

Article Title: p53 Inhibition in Pancreatic Progenitors Enhances the Differentiation of Human Pluripotent Stem Cells into Pancreatic β-Cells

doi: 10.1007/s12015-023-10509-1

Figure Lengend Snippet: p53 activation is important for endocrine lineage commitment. ( A ) The cells were treated with DMSO and pifithrin-μ during stage 5 of differentiation. ( B ) Flow cytometry of CHGA and NKX6.1 expression in DMSO or pifithrin 𝜇 -treated. ( C ) Quantification of the proportion of CHGA+/NKX6.1+ expressing cells ( n = 3). Data are represented as mean ± SD; * p < 0.05, ** p < 0.01

Article Snippet: Stained cells were filtered through a 40-μm nylon mesh into flow cytometry tubes (BD Falcon) and were analyzed with BD Accuri™ C6 flow cytometer (BD Biosciences, USA) and FlowJo for data analysis.

Techniques: Activation Assay, Flow Cytometry, Expressing

Journal: Stem Cell Reviews and Reports

Article Title: p53 Inhibition in Pancreatic Progenitors Enhances the Differentiation of Human Pluripotent Stem Cells into Pancreatic β-Cells

doi: 10.1007/s12015-023-10509-1

Figure Lengend Snippet: p53 inhibition enhances pancreatic progenitor differentiation into monohormonal β-cells. ( A ) Experimental design for B and C. Flow cytometry of C-peptide and Glucagon expression in DMSO or pifithrin 𝜇 -treated ( B ). Quantification of proportion of C- Peptide (C-PEP)/Glucagon (GCG) co-positive β-cells ( C )

Article Snippet: Stained cells were filtered through a 40-μm nylon mesh into flow cytometry tubes (BD Falcon) and were analyzed with BD Accuri™ C6 flow cytometer (BD Biosciences, USA) and FlowJo for data analysis.

Techniques: Inhibition, Flow Cytometry, Expressing

Journal: Stem Cell Reviews and Reports

Article Title: p53 Inhibition in Pancreatic Progenitors Enhances the Differentiation of Human Pluripotent Stem Cells into Pancreatic β-Cells

doi: 10.1007/s12015-023-10509-1

Figure Lengend Snippet: NKX6.1 overexpression in hPSC-derived pancreatic progenitors. ( A ) RT-PCR analysis showing the expression of NKX6.1 in hESC-derived pancreatic progenitors, 48 hours after transfection with NKX6.1 and LacZ control . ( B ) RT-qPCR analysis of NKX6.1 expression in hESC-derived pancreatic progenitors, 48 hours after transfection with NKX6.1 and LacZ control. ( C ) Western blot analysis showing the expression of NKX6.1, 48 hours after transfection with NKX6.1 and LacZ control. ( D ) Flow cytometry analysis of key pancreatic markers, NKX6.1, PDX1, SOX9, and CHGA, 48 hours after transduction of hESC-derived pancreatic progenitors with lentiviruses carrying lacZ control or NKX6.1. ( E ) Heatmap of Nanostring gene expression analysis comparing lacZ-transduced pancreatic progenitors compared to NKX6.1-transduced progenitors

Article Snippet: Stained cells were filtered through a 40-μm nylon mesh into flow cytometry tubes (BD Falcon) and were analyzed with BD Accuri™ C6 flow cytometer (BD Biosciences, USA) and FlowJo for data analysis.

Techniques: Over Expression, Derivative Assay, Reverse Transcription Polymerase Chain Reaction, Expressing, Transfection, Quantitative RT-PCR, Western Blot, Flow Cytometry, Transduction

Journal: Journal of Virology

Article Title: A Species-Specific Amino Acid Difference in the Macaque CD4 Receptor Restricts Replication by Global Circulating HIV-1 Variants Representing Viruses from Recent Infection

doi: 10.1128/JVI.02176-12

Figure Lengend Snippet: Effect of single residue changes in CD4 on HIV-1 infection. (a) Infection of 293T cells transiently expressing huCCR5 and huCD4, ptCD4, or huCD4 and ptCD4 point mutants with GFP reporter pseudoviruses bearing the Q23-17 Env. The y axis represents viral infection relative to huCD4. Error bars represent the standard deviation of the mean from duplicate independent experiments. n.d., not determined due to insufficient CD4 expression. (b) Flow cytometric analysis of 293T cells transiently expressing huCCR5 and huCD4 or ptCD4 receptors. The y axis represents CD4 expression, and the x axis represents CCR5 expression as determined by flow cytometry. The CD4 receptor that is expressed in the cells is labeled at the top of each plot. These plots are representative of at least three independent experiments. (c) Infection of Cf2Th/syn CCR5 cells stably expressing ptCD4 I39N relative to huCD4-expressing cells with GFP pseudoviruses expressing the YU-2 Env and acute/early Envs from subtypes A to D (as denoted on the x axis). Error bars represent the standard deviation of the mean from two independent experiments.

Article Snippet: The fixed cells were transferred to flow cytometry tubes (BD Falcon) that were centrifuged at 800 × g for 5 min.

Techniques: Residue, Infection, Expressing, Standard Deviation, Flow Cytometry, Labeling, Stable Transfection