af1002  (R&D Systems)

Journal: bioRxiv

Article Title: A chimeric human-mouse lung vascular model using induced pluripotent stem cells reveals insights into the pathogenesis of BMPR2 -related pulmonary hypertension

doi: 10.64898/2026.04.29.721664

Figure Lengend Snippet: (A) Schematic of directed differentiation protocol to generate hiEndos. (B) Phase contrast images during the indicated day of differentiation. White dashed square represents region magnified in insets. Scale bar: 50um. (C-E) RT-qPCR for the indicated primers using RNA isolated from cells on each day of differentiation, purified hiEndos at P0 and P5, and HUVECs. N=4 experimental replicates from independent wells of the same differentiation, representative results from two independent differentiations. (F) Flow cytometry of hiEndos using antibodies against the human endothelial lineage markers CD31 and CD144 (right) and isotype controls (left), N=4 biological replicates from distinct differentiations. (G) Immunostaining of hiEndos for human CD31 (green, left) and CD144 (red, right). Hoechst stains nuclei (blue). Scale bar 10um. N=3 biological replicates. (H) Capillary tube formation assay on 3D Matrigel. Scale bar 50um. N=4 biological replicates. (I-J) Human CD31 and acetylated LDL uptake in hiEndos measured by flow cytomtetry (I) or fluorescence microscopy (J). Scale bar 50um. N=8 biological replicates. (K) Schematic of experiment to test the impact of Notch (DAPT) and TGF-beta (SB431542) inhibition on hiEndo expansion (top). Quantification of CD31 and CD144 double-positive cells by flow cytometry (left) and cumulative hiEndo yield (right) at each passage grown in the indicated media conditions. N=2 experimental replicates from independent wells of the same differentiation, representative data shown from 3 independent differentiations. (L) Quantification of total hiEndo yield per input hiEndo in each media. ** p<0.01 using one-way Anova and Tukey’s multiple comparison test. (M) RT-qPCR of arterial and venous markers after serial passaging in SD medium. N=4 experimental replicates from independent wells of the same differentiation, representative results from two independent differentiations. ** p<0.01, *** p<0.001, **** p<0.0001, one-way Anova and Tukey’s multiple comparison test. Abbreviations: hiPSC (human induced pluripotent stem cell), CHIR (CHIR99021), P (passage), HUVEC (human umbilical vein endothelial cells), AcLDL (acetylated low-density lipoprotein), SB (SB431542).

Article Snippet: The cell pellet was then incubated with anti-human CD31 (1:10, Miltenyi, #130-091-935) and anti-human CD144 microbeads (1:10, Miltenyi, #130-097-857) diluted in EGM2 medium (50uL total volume per well of a 6-well plate, Lonza, #CC-3162) for 20 minutes.

Techniques: Quantitative RT-PCR, Isolation, Purification, Flow Cytometry, Immunostaining, Capillary Tube Formation Assay, Fluorescence, Microscopy, Inhibition, Comparison, Passaging

Journal: bioRxiv

Article Title: A chimeric human-mouse lung vascular model using induced pluripotent stem cells reveals insights into the pathogenesis of BMPR2 -related pulmonary hypertension

doi: 10.64898/2026.04.29.721664

Figure Lengend Snippet: (A-C) Fold change in the expression of indicated transcripts in hiPSC-derived cells (BU3 clone) on each day of hiEndo differentiation, in purified hiEndos at P0 and P5, and in HUVECs (RT-qPCR). N=4 experimental replicates from independent wells of the same differentiation. (D) Quantitation of hiEndo differentiation efficiency on day 5 of differentiation across 12 distinct differentiations. (E) Flow cytometry of human CD31, human CD144, and isotype controls pre- and post-MACs purification of hiEndos on day 5 of the differentiation. N=1. (F) Phase contrast microscopy of BU3 hiEndos showing cobblestone morphology. Scale bar 50um. (G) Flow cytometry of human CD31, human CD44, and isotype controls in BU3 hiEndos, the plot is representative of 4 experimental replicates of independent wells from the same differentiation. (H) Quantification of the percent of CD31/CD144 double positive hiEndos by flow cytometry at each passage. N=1 differentiation (BU1). (I) Phase contrast microscopy of BU1 hiEndos at sequential passages. Scale bar 50um. Results are representative of 2 experimental replicates of independent wells from the same differentiation. (J) RT-qPCR analysis of fold change in transcript expression of markers of fibroblast and smooth muscle identity in hiEndos at sequential passages. N=2 experimental replicates of independent wells from the same differentiation. **** p<0.0001 by one-way Anova with Tukey’s multiple comparisons.

Article Snippet: The cell pellet was then incubated with anti-human CD31 (1:10, Miltenyi, #130-091-935) and anti-human CD144 microbeads (1:10, Miltenyi, #130-097-857) diluted in EGM2 medium (50uL total volume per well of a 6-well plate, Lonza, #CC-3162) for 20 minutes.

Techniques: Expressing, Derivative Assay, Purification, Quantitative RT-PCR, Quantitation Assay, Flow Cytometry, Microscopy

Journal: Frontiers in Immunology

Article Title: Endothelial cell, but not neutrophil, programmed cell death receptor-ligand 1 loss has a morbid impact on experimental murine shock/sepsis-induced lung injury

doi: 10.3389/fimmu.2026.1816915

Figure Lengend Snippet: Description (A) and characterization of endothelial cell ( ec PD-L1 −/− ) (B, D) or neutrophil ( pmn PD-L1 −/− ) (C, E) restricted constitutive PD-L1 gene-deficient mice. Taconic Knockout Repository [mouse TF0103 (MGI:1926446); Taconic Biosciences Inc., Germantown, NY], CD274/PD-L1 conditional knockout mouse. PD-L1 gene exons 1 to 4 are shown as green boxes and loxP sites flanking exon 2 are depicted as red triangles (A) . Homozygous PD-L1 flox/flox were subsequently crossed to either VE-Cadherin‐Cre (B6.FVB-Tg[Cdh5-cre]7Mlia/J; Strain #:006137; RRID: IMSR_JAX:006137) or the S100a8-Cre (B6.Cg-Tg[S100a8-cre,-EGFP]1IIw/J; Strain #:021614; RRID: IMSR_JAX:021614) mice, obtained from Jackson Labs, to produce either mice homozygous for both the flox PD-L1 and specific cell-lineage Cre promoter genes or mice that were heterozygous for one or both genes that served as “Controls”. Gating strategy and typical flow cytogram/dot plot of select cell-lineage restricted PD-L1 expression [either CD31 and PD-L1 (B) or Ly6G and PD-L1 (C) ] for various breeding outcomes from respective Cre-Lox matings for PD-L1 flox/flox animals with VE-Cadherin‐Cre (B) or PD-L1 flox/flox animals with S100a8-Cre (C) . Summary data for the change in percentage PD-L1 + endothelial cells (CD31 + ) (D) and percentage PD-L1 + neutrophils (Ly6G + ) (E) . A dotted line is provided to indicate median frequency of non-specific antibody binding typically detected using gating strategy during analysis, which was 2.75% ± 1.77% CD31 + PD-L1 + (D) and 2.29 ± 0.50% Ly6G + PD-L1 + (E) , respectively. The presence of a significant difference between groups was established at * p < 0.05 with a Mann–Whittney U test.

Article Snippet: The VE-Cadherin‐Cre (Strain #:006137; RRID: IMSR_JAX:006137) and the S100a8-Cre (Strain #:021614; RRID: IMSR_JAX:021614) breeder mouse strains were also obtained from The Jackson Laboratory (see ).

Techniques: Knock-Out, Expressing, Binding Assay

Journal: iScience

Article Title: Heart failure pleural fluid impairs endothelial barrier through miR 501-3p mediated ZO 1 remodeling

doi: 10.1016/j.isci.2026.115549

Figure Lengend Snippet: HFPF disrupts VE-cadherin-mediated adherens junctions (A) Immunostaining of VE-cadherin (green) and phalloidin (red) after 24 h HFPF treatment. Scale bars, 100 μm. (B) Western blot analysis of VE-cadherin expression. (C) Fractionation analysis shows membranous and cytosolic VE-cadherin levels. Na + /K + -ATPase and GAPDH served as loading controls. Data means SEM. ∗ p < 0.05 and ∗∗ p < 0.01.

Article Snippet: Adheren junctions modulate cell adhesion via cadherins, and VE-cadherin is one of the major cadherins in endothelial cells.

Techniques: Immunostaining, Western Blot, Expressing, Fractionation

Journal: Bioactive Materials

Article Title: Ribbon-shaped microgels as bioinks for 3D bioprinting of anisotropic tissue structures

doi: 10.1016/j.bioactmat.2025.12.040

Figure Lengend Snippet: Effect of extrusion process on μRB bioink and cell alignment. (A) Schematic of MSCs encapsulated in μRB bioink with HUVECs seeded on top of the printed scaffolds. (B) Live cell staining of MSC alignment on individual μRBs (Scale bar = 100 μm). (C) Distribution of MSC cell length (n = 250 per group); p-values were determined by one-way analysis of variance (ANOVA) with Tukey's multiple comparisons test; ∗∗∗∗p < 0.0001. (D) Cell orientation relative to μRB orientation, where 0° is parallel to the axis of the μRB (n = 250 per group). (E) Confocal images of F-Actin staining for cell morphology and VE-cadherin staining for endothelial cell junctions. Color survey visualization of directional analysis conducted using OrientationJ (Scale bar = 200 μm). (F, G) Quantification of F-actin and VE-Cadherin alignment. For alignment quantification (n = 10 per group) data reported as mean ± S.D., statistical analysis by Watson–Wheeler test, ∗p ≤ 0.05.

Article Snippet: For analyzing cell alignment of HUVEC and MSC, immunostaining of actin (ActinRedTM 555 ReadyProbes, ThermoFisher) and VE-cadherin (1:400, 2500 T, Cell Signaling) were performed on the scaffolds.

Techniques: Staining