Journal: iScience

Article Title: Epithelial–stromal cell interactions and extracellular matrix mechanics drive the formation of airway-mimetic tubular morphology in lung organoids

doi: 10.1016/j.isci.2021.103061

Figure Lengend Snippet: Normal healthy human bronchial epithelial (NHBE) cell and normal healthy human lung fibroblast (NHLF) coculture NHLF seeding density was increased from 6,000 to 1,350,000 cells/ml and NHLF cells allowed to adhere to the bottom of wells of a 96 well plate before being layered with 25% Matrigel and then seeding NHBE cells in 5% Matrigel. Increases in NHLF resulted in increased numbers of tubular structures formed by epithelial cells. Images are representative of those from 2 wells from each experiment and n = 3 biological repeats. Scale bar = 100μm.

Article Snippet: NHLF cells were purchased from PromoCell.

Techniques:

Journal: Scientific Reports

Article Title: Unphosphorylated STAT1 binds to the BST2 transcription promoter, promoting increased AKBA anchoring on HPMECs to alleviate ARDS

doi: 10.1038/s41598-025-00028-z

Figure Lengend Snippet: Protective effect of AKBA on HPMECs induced by LPS. ( A , B ) Tubule formation assay was performed to evaluate the tubule-forming ability of HPMECs (100×magnification), n = 3. ( C-F ) Colony formation assay and EDU assay were used to detect the relief of AKBA on LPS-induced proliferation inhibition of HPMECs (100×magnification), n = 3. ( G ) Cell viability was evaluated by CCK8 assay, n = 3. ( H-K ) The migration protection of AKBA on HPMECs was confirmed by scratch assay and transwell assay (40×and 100×magnifications), n = 3. ( L , M ) The LC3 expression of HPMECs was determined by immunofluorescence assay, using CD31 as an internal reference. (200×magnification), n = 3. ( N ) Effective inhibition of autophagy by AKBA in HPMECs was elucidated by TEM (15.0k×magnification). Values are presented as mean ± standard deviation (SD), n = 3. Statistical significance levels are denoted as * p < 0.05, ** p < 0.01 and *** p < 0.001.

Article Snippet: Passaged human pulmonary microvascular endothelial cells (HPMECs) from ScienCell (San Diego, CA, USA) and incubated at 37 °C in 5% CO 2 with Dulbecco’s Modified Eagle’s Medium (DMEM; Gibco, Carlsbad, CA, USA) containing with 10% fetal bovine serum (FBS, Gibco, Paisley, UK) and 1% penicillin/streptomycin (Beyotime Shanghai, China).

Techniques: Tube Formation Assay, Colony Assay, EdU Assay, Inhibition, CCK-8 Assay, Migration, Wound Healing Assay, Transwell Assay, Expressing, Immunofluorescence, Standard Deviation

Journal: Scientific Reports

Article Title: Unphosphorylated STAT1 binds to the BST2 transcription promoter, promoting increased AKBA anchoring on HPMECs to alleviate ARDS

doi: 10.1038/s41598-025-00028-z

Figure Lengend Snippet: AKBA may activate U-STAT1 expression and promote BST2 transcription by anchoring BST2. ( A ) Expression of BST2 in peripheral blood of patients with septic shock. ( B ) Expression of ISG15, BST2, OAS2, IFIT1, IFIT3 and STAT1 in LPS + AKBA group, n = 3. ( C ) Molecular docking results and site prediction of AKBA and BST2. ( D ) Top 20 items for differential protein enrichment analysis and a special concentration of STAT1 signaling pathway. ( E , F ) The interaction between AKBA and BST2 was evaluated by CETSA technique and CETSA dissolution curve, n = 3. ( G ) mRNA expression levels of STAT1 and BST2 in HPMECs were evaluated using reverse transcription-quantitative PCR after regulation of STAT1 or BST2, n = 3. ( H ) Relative luciferase activity was assessed using the dual-luciferase reporter assay in HPMECs co-transfected with OE-STAT1 or OE-NC and BST2-WT or BST2-MUT. ( I , J ) The molecular dynamics simulation of AKBA and BST2 complex system was conducted to investigate the binding stability and kinetic behavior. ( K , L ) DARTS-WB assesses the stability of AKBA and BST2 complex systems. ( M , N ) CHlP-gPCR was performed in HPMECs transfected with pcDNA3.1-HA-AP to identify the enrichment of HA-STAT1 onto BST2 promoter region, IgG served as an antibody control. n = 3. * p < 0.05, ** p < 0.01 and *** p < 0.001.

Article Snippet: Passaged human pulmonary microvascular endothelial cells (HPMECs) from ScienCell (San Diego, CA, USA) and incubated at 37 °C in 5% CO 2 with Dulbecco’s Modified Eagle’s Medium (DMEM; Gibco, Carlsbad, CA, USA) containing with 10% fetal bovine serum (FBS, Gibco, Paisley, UK) and 1% penicillin/streptomycin (Beyotime Shanghai, China).

Techniques: Expressing, Protein Enrichment, Concentration Assay, Dissolution, Reverse Transcription, Real-time Polymerase Chain Reaction, Luciferase, Activity Assay, Reporter Assay, Transfection, Binding Assay, Control

Journal: Scientific Reports

Article Title: Unphosphorylated STAT1 binds to the BST2 transcription promoter, promoting increased AKBA anchoring on HPMECs to alleviate ARDS

doi: 10.1038/s41598-025-00028-z

Figure Lengend Snippet: AKBA mediates BST2 to improve HPMECs dysfunction. ( A , B ) Fluorescence intensity and cellular localization of BST2, (630×magnification). ( C ) Fluorescence intensity and cellular localization of U-STAT1, (630× magnification). ( D ) Cell viability was evaluated by CCK8 assay. ( E ) The permeability of HPMECs was evaluated by the vascular permeability marker Evans Blue. ( F , G ) The apoptosis level of HPMECs was evaluated by flow cytometry. n = 3. * p < 0.05, ** p < 0.01 and *** p < 0.001.

Article Snippet: Passaged human pulmonary microvascular endothelial cells (HPMECs) from ScienCell (San Diego, CA, USA) and incubated at 37 °C in 5% CO 2 with Dulbecco’s Modified Eagle’s Medium (DMEM; Gibco, Carlsbad, CA, USA) containing with 10% fetal bovine serum (FBS, Gibco, Paisley, UK) and 1% penicillin/streptomycin (Beyotime Shanghai, China).

Techniques: Fluorescence, CCK-8 Assay, Permeability, Marker, Flow Cytometry

Journal: Scientific Reports

Article Title: Unphosphorylated STAT1 binds to the BST2 transcription promoter, promoting increased AKBA anchoring on HPMECs to alleviate ARDS

doi: 10.1038/s41598-025-00028-z

Figure Lengend Snippet: AKBA may inhibit ARDS progression through U-STAT1/BST2. ( A ) BST2, U-STAT1, YP-STAT1 protein levels in HPMECs were quantified by Western blot analysis, n = 3. ( B ) Statistical analysis of Western blot of BST2 in HPMECs, n = 3. ( C ) Statistical analysis of Western blot of U-STAT1 in HPMECs, n = 3. ( D ) Statistical analysis of Western blot of YP-STAT1 in HPMECs, n = 3. ( E ) Statistical analysis of Western blot of BST2 in mouse lung tissues, n = 5. ( F ) Statistical analysis of Western blot of U-STAT1 in mouse lung tissues, n = 5. ( G ) Statistical analysis of Western blot of YP-STAT in mouse lung tissues, n = 5. ( H ) BST2, U-STAT1, YP-STAT1 protein levels in mice lung tissues were quantified by Western blot, n = 5. ( I-J ) The LC3 expression of HPMECs was determined by immunofluorescence assay, using CD31 as an internal reference. (200×magnification), n = 3. ( K ) The autophagy level of HPMECs was evaluated by transmission electron microscopy. (15.0k×magnification), n = 3. * p < 0.05, ** p < 0.01 and *** p < 0.001.

Article Snippet: Passaged human pulmonary microvascular endothelial cells (HPMECs) from ScienCell (San Diego, CA, USA) and incubated at 37 °C in 5% CO 2 with Dulbecco’s Modified Eagle’s Medium (DMEM; Gibco, Carlsbad, CA, USA) containing with 10% fetal bovine serum (FBS, Gibco, Paisley, UK) and 1% penicillin/streptomycin (Beyotime Shanghai, China).

Techniques: Western Blot, Expressing, Immunofluorescence, Transmission Assay, Electron Microscopy

Journal: Scientific Reports

Article Title: Unphosphorylated STAT1 binds to the BST2 transcription promoter, promoting increased AKBA anchoring on HPMECs to alleviate ARDS

doi: 10.1038/s41598-025-00028-z

Figure Lengend Snippet: AKBA protects LPS-induced HPMECs via U-STAT1/BST2. ( A , B ) The proliferation of HPMECs was verified by EDU assay, (100×magnification), n = 3. ( C , D ) The proliferation of HPMECs was verified by colony formation assay, (100×magnification), n = 3. ( E , F ) The tube forming ability of HPMECs was evaluated by quantifying the number of nodes and the vascular coverage area, (100×magnification), n = 3. ( G , H ) The migration function of HPMECs was verified by wound healing assay. (100×magnification), n = 3. ( I , J ) The migration function of HPMECs was verified by transwell assay. (100×magnification), n = 3. ( K ) Western blot analysis of BST2, U-STAT1, YP-STAT1, Bad, using β-actin expression as the internal reference in HPMECs, n = 3. ( L ) Statistical analysis of Western blot of BST2 in HPMECs, n = 3. ( M ) Statistical analysis of Western blot of U-STAT1 in HPMECs, n = 3. ( N ) Statistical analysis of Western blot of Bad in HPMECs, n = 3. * p < 0.05, ** p < 0.01 and *** p < 0.001.

Article Snippet: Passaged human pulmonary microvascular endothelial cells (HPMECs) from ScienCell (San Diego, CA, USA) and incubated at 37 °C in 5% CO 2 with Dulbecco’s Modified Eagle’s Medium (DMEM; Gibco, Carlsbad, CA, USA) containing with 10% fetal bovine serum (FBS, Gibco, Paisley, UK) and 1% penicillin/streptomycin (Beyotime Shanghai, China).

Techniques: EdU Assay, Colony Assay, Migration, Wound Healing Assay, Transwell Assay, Western Blot, Expressing

Journal: Communications Biology

Article Title: Nipah virus W protein harnesses nuclear 14-3-3 to inhibit NF-κB-induced proinflammatory response

doi: 10.1038/s42003-021-02797-5

Figure Lengend Snippet: a Representative images of HPMEC cells transfected or not (Ø) with plasmids encoding HA-R18 and indicated FLAG-tagged W proteins, and stimulated with 10 ng/mL of IL-1β for 20 min (scale bar = 10 µm). Cells were fixed and stained for NF-κB p65, anti-HA, and anti-FLAG, and analyzed by confocal microscopy. b The mean fluorescence intensity for p65 protein was measured in the cell nucleus. Signal obtained from W expressing cells (as labeled by anti-FLAG) was normalized to the signal from cells not labeled by anti-FLAG and expressed in fold change. Error bars represent the mean’s confidence interval (CI 95%) for nine cells, combined from three independent experiments. Each combination has been done in three different wells performed in two independent experiments. Statistical significance was assessed by a one-way unpaired ANOVA, multiple comparisons Tukey’s test; ** p < 0.01 and **** p < 0.0001.

Article Snippet: HPMEC cells were cultured in Endothelial Cell Growth Medium (PromoCell ® , Cat# C-22121) in flasks coated with 0.1% bovine gelatin in phosphate-buffered saline (PBS) (Gibco, Cat# 14190-094).

Techniques: Transfection, Staining, Confocal Microscopy, Fluorescence, Expressing, Labeling