ve cadherin  (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: Nature Communications

Article Title: Single-cell multiomics uncovers an endothelial mechanosensitive PIEZO1-IL-33 axis driving pulmonary fibrosis

doi: 10.1038/s41467-026-70193-w

Figure Lengend Snippet: A BLM intratracheal model scheme (n = 3 per group). B H&E and Masson validation of inflammation/fibrosis (n = 3 per group). C Lung scRNA-seq UMAP. D scATAC-seq UMAP subclusters. E Venn showing Piezo1 and Piezo2 as co-regulated genes across silica and BLM bulk, scRNA-seq and scATAC-seq data. F scATAC-seq coverage peaks of Piezo1 and Piezo2 . G CD31 and CD45-sorted EC bulk-RNA-seq heatmap and bar chart (n = 3 per group). Gene expression levels were Z-score normalized. P values were calculated using the two-tailed unpaired t-test. H PIEZO1 (green) and Ve-cad (red) co-staining and quantification in fibrotic mouse lungs (n = 4). UMAP and bar charts (mean ± SEM) of PIEZO1 + ( I ) and PIEZO2 + ( J ) EC counts in normal controls (n = 5) versus IPF (n = 4) patients. P values were calculated using the two-tailed unpaired t-test. K PIEZO1 (green) and CD31 (red) co-staining and quantification in human IPF vs normal lungs (n = 10). L Meta-analysis of five GEO datasets showing elevated PIEZO1 in IPF ECs. Standardized mean difference (SMD) were calculated as Hedges’ g with corresponding 95% confidence intervals. Data were shown as mean ± SEM, H and K: two-tailed unpaired t-tests, Source data are provided as a Source Data file.

Article Snippet: The primary antibodies used were CD31 (Abcam, #ab9498; Servicebio, #GB13063), VE-cadherin (R&D Systems, #AF938), IL-33 (R&D Systems, #AF3626), PIEZO1(proteintech, #15939-1-AP) and αSMA (Abcam, #ab7817) antibodies.

Techniques: Biomarker Discovery, RNA Sequencing, Gene Expression, Two Tailed Test, Staining

Journal: Nature Communications

Article Title: Single-cell multiomics uncovers an endothelial mechanosensitive PIEZO1-IL-33 axis driving pulmonary fibrosis

doi: 10.1038/s41467-026-70193-w

Figure Lengend Snippet: A CellChat analysis of human lung snRNA-seq data. B Venn of 47 shared DEGs from four datasets ( GSE159354 , GSE132771 , GSE128033 , GSE122960 ). C Heatmap of 47-gene expression across cell types. D Correlation analysis (Pearson Correlation) of expression of IGFBP5, IL33, MGP, SPRY1, ACKR1 and mechanical stress scores in ECs. E Correlation analysis (Pearson Correlation) of expression of PIEZO1 and IGFBP5, IL33, MGP, SPRY1, ACKR1 in ECs. F Human snRNA-seq UMAP and violin of IL33 distribution and subpopulations. G Bar chart showed expression of IL33 (mean ± SEM) in IPF (n = 7966 cells) vs control (n = 4174 cells). P value was calculated using the two-tailed unpaired t-test. H Violin of IL33 in PIEZO1 - vs PIEZO1 + ECs. I Bulk-RNA-seq of saline and BLM (7 days and 14 days) induced mice (n = 3 per group). Gene expression levels were Z-score normalized. P value was calculated using the two-tailed unpaired t-test. J qPCR of Il33 in BLM mouse lung ECs. K IL-33 (green) and CD31 (red) co-staining in human IPF vs normal, quantification is shown at right (n = 10). L IL-33 (green) and CD31 (red) co-staining in Yoda1- or GsMTx4-treated fibrotic mouse lungs, quantification is shown at right (n = 3). M IL-33 (green) and CD31 (red)co-staining in Piezo1 WT and Piezo1 ΔEC BLM lungs, quantification is shown at right (n = 3). Error bars represent mean ± SEM, H, K, J, M: two-tailed unpaired t-test; L: one-way ANOVA followed Tukey’s multiple comparwasons test, All n values indicate biologically independent samples. Source data are provided as a Source Data file.

Article Snippet: The primary antibodies used were CD31 (Abcam, #ab9498; Servicebio, #GB13063), VE-cadherin (R&D Systems, #AF938), IL-33 (R&D Systems, #AF3626), PIEZO1(proteintech, #15939-1-AP) and αSMA (Abcam, #ab7817) antibodies.

Techniques: Gene Expression, Expressing, Control, Two Tailed Test, RNA Sequencing, Saline, Staining

Journal: Nature Communications

Article Title: Single-cell multiomics uncovers an endothelial mechanosensitive PIEZO1-IL-33 axis driving pulmonary fibrosis

doi: 10.1038/s41467-026-70193-w

Figure Lengend Snippet: A Schematic of BLM-induced fibrosis in Il33 WT vs Il33 ΔEC mice. B Il33 mRNA (n = 4) and IL-33 protein in lung ECs (n = 3). C Representative Masson/PSR images and quantification (n = 5). D αSMA immunofluorescence and quantification (n = 4). E Lung hydroxyproline content (n = 5). F Schematic: EC-specific Il33 overexpression via AAV-Tie1 in Piezo1 WT and Piezo1 ΔEC mice; 4-week pretreatment, BLM at week 4, sacrifice at week 7. G IL-33 expression in ECs, immune and epithelial cells after AAV-OE- Il33 (n = 3). H Masson/PSR images and quantification (n = 5). I αSMA immunofluorescence and quantification (n = 4). J Hydroxyproline content (n = 5). Error bars: mean ± SEM; B,G: two-tailed unpaired t-test; C,D-E,I: two-way ANOVA followed Tukey’s multiple comparisons test; H,J: three-way ANOVA followed Tukey’s multiple comparisons test. All n values indicate biologically independent samples. Source data are provided as a Source Data file.

Article Snippet: The primary antibodies used were CD31 (Abcam, #ab9498; Servicebio, #GB13063), VE-cadherin (R&D Systems, #AF938), IL-33 (R&D Systems, #AF3626), PIEZO1(proteintech, #15939-1-AP) and αSMA (Abcam, #ab7817) antibodies.

Techniques: Immunofluorescence, Over Expression, Expressing, Two Tailed Test

Journal: Nature Communications

Article Title: Single-cell multiomics uncovers an endothelial mechanosensitive PIEZO1-IL-33 axis driving pulmonary fibrosis

doi: 10.1038/s41467-026-70193-w

Figure Lengend Snippet: HUVECs under 20% stretch for 0,6,24 h or 2,12,25 kPa substrates; IL-33 secretion ( n = 3) A and mRNA (n = 3) B . C PIEZO1 knock-down with sh PIEZO1 ( n = 3). HUVECs were transduced with sh PIEZO1 for 48 h, cultured on 25 kPa substrates for 24 h or subjected to 6 h mechanical stretch before measuring IL-33 secretion ( n = 3) D and mRNA ( n = 3) E . HUVECs were cultured on 2, 12, 25 kPa substrates for 24 h or stretched (20%) for 0, 6, 24 h, followed by measurement of calpain activity F and CAPN2 protein G ( n = 3). HUVECs (sh PIEZO1 ) were cultured on 25 kPa substrates for 24 h or subjected to 6 h mechanical stretch, followed by measurement of calpain activity ( n = 3) H and CAPN2 protein levels ( n = 3) I . J CAPN2 knock-down with shCAPN2 ( n = 3). K HUVECs (sh CAPN2 ) were cultured on 25 kPa substrates for 24 h or subjected to 6 h mechanical stretch, IL-33 secretion(left) and mRNA(right) ( n = 3). L Model for paracrine IL-33 regulation. M Cistrome: STAT3 predicted to drive IL-33 regulation. N intersection of top 100 motifs in BLM-treated mouse ECs with top 10 TFs. O The protein levels of P-STAT3 and STAT3 in HUVECs treated with 20% stretch for 0 and 6 hours ( n = 3). P The protein levels of P-STAT3 and STAT3 in HUVECs (sh PIEZO1 or sh CAPN2 ) treated with 20% stretch for 6 h ( n = 3). Q STAT3 knock-down with sh STAT3 ( n = 3). R HUVECs (sh STAT3 ) were cultured on 25 kPa substrates for 24 h or subjected to 6 h mechanical stretch; IL-33 secretion(left) and mRNA(right) ( n = 3). Error bars: mean ± SEM; C , G , J , O – Q : two-tailed unpaired t-test; A , B , F : one-way ANOVA followed Šídák’s multiple comparisons test, D – E , H – I , K , R : two-way ANOVA Tukey’s multiple comparisons test. All n denote biologically independent samples. Source data are provided as a Source Data file.

Article Snippet: The primary antibodies used were CD31 (Abcam, #ab9498; Servicebio, #GB13063), VE-cadherin (R&D Systems, #AF938), IL-33 (R&D Systems, #AF3626), PIEZO1(proteintech, #15939-1-AP) and αSMA (Abcam, #ab7817) antibodies.

Techniques: Knockdown, Transduction, Cell Culture, Activity Assay, Two Tailed Test