Journal: Brain : a journal of neurology

Article Title: Connexin43 mimetic peptide reduces vascular leak and retinal ganglion cell death following retinal ischaemia.

doi: 10.1093/brain/awr338

Figure Lengend Snippet: Figure 3 Three hours of hypoxia and 6 h of reperfusion lead to significant endothelial cell death in vitro in rat brain microvascular endothelial cells (RBMVEC; A). Non-specific gap junction blocker carbenoxolone, non-specific hemichannel blocker LaCl3, and connexin43 (Cx43) mimetic peptide protected endothelial cells against hypoxic injury, with the number of viable cells significantly higher than no treatment, while scrambled peptide did not have any protective effects. The number of viable cells was expressed as percentage of the control without hypoxia. Hypoxia and reperfusion also lead to significant propidium iodide dye uptake into primary rat brain microvascular endothelial cells indicating open hemichannels (B). Carbenoxolone, LaCl3 and connexin43 mimetic peptide significantly prevented dye uptake compared to no treatment, indicating hemichannel closure, while scrambled peptide did not have any effect. Stars denote statistical significance when compared to the control group or compared between groups in brackets; P 5 0.05.

Article Snippet: Rat brain microvascular endothelial cells (R840K-05a, Cell Applications) were plated into 24-well plates (1 105 cells/well) in rat brain microvascular endothelial cell growth media (R819K-500, Cell Applications) and allowed to settle for 16 h. Medium was then removed and replaced with Dulbecco’s Modified Eagle’s Medium/F12 containing 0.5% foetal bovine serum and 1% glutamine.

Techniques: In Vitro, Control

Journal: Bioengineering & Translational Medicine

Article Title: Mesenchymal stem cell extracellular vesicle vascularization bioactivity and production yield are responsive to cell culture substrate stiffness

doi: 10.1002/btm2.10743

Figure Lengend Snippet: Substrate stiffness influences BM‐MSC EV production and bioactivity. (a) EV production as quantified by EVs per cell from BM‐MSCs seeded on Sylgard 184 PDMS substrates with different base‐to‐crosslinker ratios. EVs used for this data were from 1 day of collection and isolated and counted separately from the conditioned media from the other 2 days. After media collection, cells were trypsinized and counted ( n = 3). (b) After a scratch was induced, HUVECs were treated with BM‐MSC EVs from the different substrates or growth or basal media, and percent gap closure after 20 h was evaluated via microscopy ( n = 3). (c) HUVECs were resuspended in EV treatments or growth or basal endothelial media, seeded in Matrigel‐coated wells, and tube formation after 3–6 h was quantified by the number of loops that had formed ( n = 3). All values expressed as mean ± SD. All data are representative of at least three independent experiments ( n = 3). Statistical significance was determined by ANOVA; *p < 0.05, **p < 0.01, ***p < 0.001, and ****p < 0.0001.

Article Snippet: To measure in vitro angiogenesis, 48‐well plates were coated with 60 μL of growth factor reduced Matrigel (Corning; 356230) and incubated at 37°C for 30 min. P4 HUVECs were then seeded at 35,000 cells/well with either endothelial growth media (PromoCell; C‐22121) with 1% penicillin–streptomycin (positive control), endothelial basal media (negative control), or endothelial basal media (PromoCell; C‐22221) with 0.1% FBS and 1% penicillin–streptomycin with 5E9 EVs/mL.

Techniques: Isolation, Microscopy

Journal: Bioengineering & Translational Medicine

Article Title: Mesenchymal stem cell extracellular vesicle vascularization bioactivity and production yield are responsive to cell culture substrate stiffness

doi: 10.1002/btm2.10743

Figure Lengend Snippet: Softer 184:527 PDMS substrates improve the angiogenic bioactivity of BM‐MSC EVs. (a) EV production quantified as EV per cell from BM‐MSCs seeded on each substrate made with different ratios of Sylgard 184 and Sylgard 527 ( n = 2). EVs used for this data were from 1 day of collection and isolated and counted separately from the conditioned media from the other 2 days. After media collection, cells were trypsinized and counted. (b) After a scratch was induced, HUVECs were treated with BM‐MSC EVs from the different substrates or growth or basal media, and percent gap closure after 20 h was evaluated via microscopy ( n = 3). (c) HUVECs were resuspended in the different EV treatments or growth or basal endothelial basal media, and tube formation after 3–6 h was quantified by the number of loops that had formed ( n = 3). All values expressed as mean ± SD. Statistical significance was determined by ANOVA; * p < 0.05, ** p < 0.01, and **** p < 0.0001.

Article Snippet: To measure in vitro angiogenesis, 48‐well plates were coated with 60 μL of growth factor reduced Matrigel (Corning; 356230) and incubated at 37°C for 30 min. P4 HUVECs were then seeded at 35,000 cells/well with either endothelial growth media (PromoCell; C‐22121) with 1% penicillin–streptomycin (positive control), endothelial basal media (negative control), or endothelial basal media (PromoCell; C‐22221) with 0.1% FBS and 1% penicillin–streptomycin with 5E9 EVs/mL.

Techniques: Isolation, Microscopy

Journal: Nature Communications

Article Title: Structure and function relationships of mucociliary clearance in human and rat airways

doi: 10.1038/s41467-025-57667-z

Figure Lengend Snippet: a Airway branching generations in human and rat investigated in this study included BG0-6 in humans and BG0-5 in rats. b Workflow for imaging luminal epithelial cell type composition and ciliary beat and clearance function in airway samples. Schematic: Created in BioRender. Nawroth, J. (2025) https://BioRender.com/a01i578 ). c Example IF staining of cilia (ATUB, magenta) and secretory cells (SCGB1A1, green; MUC5AC, gray) in human and rat airway epithelium in BG0 (trachea) and BG5/6. Scale bar: 20 µm. d Quantification of luminal cell proportions labeled with ATUB (ciliated cells) or with MUC5AC and/or SCGB1A1 (secretory cells) as a function of airway branching generation in human and rat airway epithelium. Inset: Percentage of human secretory cell population positive for only MUC5AC (gray), only for SCGB1A1 (green), or for both (white) as a function of branching generation. Solid line: mean, shaded region: SEM. Numbers of human donors (2–3 FOVs each): BG0, n = 3; BG1, n = 3; BG2, n = 5; BG3, n = 3; BG4, n = 7; BG5, n = 2; BG6, n = 2; Numbers of rat donors (2–3 FOVs each): BG0, n = 5; BG1, n = 3; BG2, n = 2; BG4, n = 1; BG5, n = 1. For full donor information see Supplementary Tables and . Source data for ( d ) are provided as a file.

Article Snippet: SAGM: Small Airway Epithelial Growth Medium (Promocell, C-21170) containing 50 nM EC23; and 5. mAir: a 1:1 mix of Dulbecco’s modified eagle medium and Airway epithelial cell growth medium (AECGM, PromoCell, C-21160) with AECGM supplements and 50 nM EC23 (previously described in ref. ).

Techniques: Imaging, Staining, Labeling

Journal: Nature Communications

Article Title: Structure and function relationships of mucociliary clearance in human and rat airways

doi: 10.1038/s41467-025-57667-z

Figure Lengend Snippet: a Representative measurement of ciliary beat frequency (CBF) and associated particle clearance trajectories and speed in a human airway epithelial sample (BG2). b Same measurements in rat airway epithelial sample (BG1). Scalebar in ( a , b ): 100 µm. c Quantification of average CBF, particle clearance speed, and clearance per beat (CPB) in human airways BG0-6 and rat airways BG0-1. Number of human donors: n = 4. Number of rat donors: n = 6. d left: Clearance directionality as a function of distance in human (red) and rat (blue) airways. Thick lines are average curves. Right: Mean directionality over a flow distance of 80 µm. Number of human donors n = 4; number of rat donors: n = 4. Boxplots: Each solid dot is the mean value of one donor (1–3 BGs, 2–4 FOVs each); red line indicates median, bottom and top edges of the box indicate 25th and 75th percentiles, whiskers indicate minimum and maximum; significance was assessed with two-sided unpaired t-test. Source data for ( c , d ) are provided as a file.

Article Snippet: SAGM: Small Airway Epithelial Growth Medium (Promocell, C-21170) containing 50 nM EC23; and 5. mAir: a 1:1 mix of Dulbecco’s modified eagle medium and Airway epithelial cell growth medium (AECGM, PromoCell, C-21160) with AECGM supplements and 50 nM EC23 (previously described in ref. ).

Techniques:

Journal: Nature Communications

Article Title: Structure and function relationships of mucociliary clearance in human and rat airways

doi: 10.1038/s41467-025-57667-z

Figure Lengend Snippet: a Representative IF-images of primary human airway epithelial cultures from 1 donor grown in different differentiation media for 28 days at ALI and stained for cilia (ATUB, magenta) and secretory cell markers (SCGB1A1, green; MUC5AC, white). Scalebar, 40 µm. b Average luminal cell type composition based on IF-staining (in vitro: N = 4–7 donors, 2 inserts each with 3–8 FOVs each; ex vivo: 9 donors, 1–3 BGs, 2–3 FOVs each). c Mapping of IF-staining data of in vitro and ex vivo samples onto three dimensions. y- axis: percentage of MUC5AC+ cells; x-axis: cilia coverage, i.e., percentage of ciliated (ATUB + ) cells; circle diameter: ratio of SCGB1A1+ to MUC5AC+ cell percentages. d Average percentage of MUC5B expressing cells and e relative percentage of MUC5B expressing cells that also express either SCGB1A1 or MUC5AC from IF-stainings (in vitro: n = 2 (SAGM, BD) to n = 4 (PC, PC-S, mAir) donors, 1–2 inserts each with 1–6 FOVs each; ex vivo: n = 3 donors, BG0, 2–4 FOVs each). Boxplot: Each dot represents mean of one donor; red line indicates median, bottom and top edges of the box indicate 25th and 75th percentiles, whiskers indicate minimum and maximum. Source data for ( b – e ) are provided as a file.

Article Snippet: SAGM: Small Airway Epithelial Growth Medium (Promocell, C-21170) containing 50 nM EC23; and 5. mAir: a 1:1 mix of Dulbecco’s modified eagle medium and Airway epithelial cell growth medium (AECGM, PromoCell, C-21160) with AECGM supplements and 50 nM EC23 (previously described in ref. ).

Techniques: Staining, In Vitro, Ex Vivo, Expressing

Journal: Nature Communications

Article Title: Structure and function relationships of mucociliary clearance in human and rat airways

doi: 10.1038/s41467-025-57667-z

Figure Lengend Snippet: a Quantitative analysis of particle CPB and directionality in primary human airway epithelial cultures grown in different differentiation media for 28 days at ALI compared to human and rat benchmark data. Number of donors (2–3 inserts each with 6 FOVs each) for BD, mAir, SAGM, PC, PC-S, respectively: n = 5, 4, 4, 5, 4. b Quantitative analysis of ciliary beat metrics in airway cells cultured and visualized as in ( a ). Number of donors (2–3 inserts each with 6 FOVs each) for BD, mAir, SAGM, PC, PC-S, respectively: Cilia Coverage, n = 7, 4, 4, 7, 4; Ciliary Beat OP, n = 4, 4, 2 (1 not measurable), 4, 4; Ciliary Beat Amplitude: n = 4, 4, 3, 4, 4; Cilia Length, n = 4, 4, 4, 4, 4; Ciliation Gap, n = 4, 4, 3, 4, 4; Crystalline OP, n = 4, 4, 3, 4, 4. Boxplots in ( a , b ): Each solid dot is the mean value of one donor (1–3 BGs, 2–4 FOVs each); red line indicates median, bottom and top edges of the box indicate 25th and 75th percentiles, whiskers indicate minimum and maximum. Dotted lines indicate average human and rat benchmark values. c Top: Predicted CPB and clearance directionality in in vitro cultures compared to predicted human airway performance (red). Shaded regions indicate uncertainty based on spread of input metrics. Measured mean values per donor are overlaid (dots). Bottom: Predicted change in CPB and directionality of in vitro cultures in different media if an individual cilia input parameter is set to match ex vivo values. Source data for ( a – c ) are provided as a file.

Article Snippet: SAGM: Small Airway Epithelial Growth Medium (Promocell, C-21170) containing 50 nM EC23; and 5. mAir: a 1:1 mix of Dulbecco’s modified eagle medium and Airway epithelial cell growth medium (AECGM, PromoCell, C-21160) with AECGM supplements and 50 nM EC23 (previously described in ref. ).

Techniques: Cell Culture, In Vitro, Ex Vivo

Journal: bioRxiv

Article Title: Development of high-affinity, single-domain protein binders for neutralizing household allergens

doi: 10.1101/2025.08.03.668213

Figure Lengend Snippet: (a) Thermostability profile of AVA-1-1-C3 as a representative protein, as determined by differential scanning fluorimetry. (b-g) Cytotoxicity of primary human cells following 48-hour exposure to AVA-1-1-C3 or an equivalent concentration of bovine serum albumin (BSA), evaluated using the CellTiter-Glo® (CTG) luminescent cell viability assay. Cell types included human epithelial keratinocytes (HEK, b), human dermal fibroblasts (HDF, c), human follicle dermal papilla cells (HFDPC, d), human renal cortical epithelial cells (HRCEPC, e), human skeletal muscle cells (HSKMC, f), and human dermal microvascular endothelial cells (HDMEC, g). No significant differences in cell viability were observed. Data in (b-g) represents mean ± s.d. and are representative of two experimental replicates.

Article Snippet: Cells were cultured in their corresponding complete media including HDF growth medium (Cell Applications, Cat# 116-500), human EpiVita serum-free growth medium (Cell Applications, Cat# 141-500a), microvascular endothelial cell growth medium (PromoCells, Cat# C-22120), HSkMC growth medium (Cell Applications, Cat# 151-500), and renal epithelial cell growth media (PromoCell, Cat# C-26130).

Techniques: Concentration Assay, Cell Viability Assay

Journal: Nature Communications

Article Title: Structure and function relationships of mucociliary clearance in human and rat airways

doi: 10.1038/s41467-025-57667-z

Figure Lengend Snippet: a Airway branching generations in human and rat investigated in this study included BG0-6 in humans and BG0-5 in rats. b Workflow for imaging luminal epithelial cell type composition and ciliary beat and clearance function in airway samples. Schematic: Created in BioRender. Nawroth, J. (2025) https://BioRender.com/a01i578 ). c Example IF staining of cilia (ATUB, magenta) and secretory cells (SCGB1A1, green; MUC5AC, gray) in human and rat airway epithelium in BG0 (trachea) and BG5/6. Scale bar: 20 µm. d Quantification of luminal cell proportions labeled with ATUB (ciliated cells) or with MUC5AC and/or SCGB1A1 (secretory cells) as a function of airway branching generation in human and rat airway epithelium. Inset: Percentage of human secretory cell population positive for only MUC5AC (gray), only for SCGB1A1 (green), or for both (white) as a function of branching generation. Solid line: mean, shaded region: SEM. Numbers of human donors (2–3 FOVs each): BG0, n = 3; BG1, n = 3; BG2, n = 5; BG3, n = 3; BG4, n = 7; BG5, n = 2; BG6, n = 2; Numbers of rat donors (2–3 FOVs each): BG0, n = 5; BG1, n = 3; BG2, n = 2; BG4, n = 1; BG5, n = 1. For full donor information see Supplementary Tables and . Source data for ( d ) are provided as a file.

Article Snippet: Cells were then seeded at a density of 30 K cells cm -2 on PureCol (Advanced Biomatrix #5005) coated tissue culture dishes in airway epithelial growth media (Promocell #C-21160) and passaged at 80% confluence.

Techniques: Imaging, Staining, Labeling

Journal: Nature Communications

Article Title: Structure and function relationships of mucociliary clearance in human and rat airways

doi: 10.1038/s41467-025-57667-z

Figure Lengend Snippet: a Representative measurement of ciliary beat frequency (CBF) and associated particle clearance trajectories and speed in a human airway epithelial sample (BG2). b Same measurements in rat airway epithelial sample (BG1). Scalebar in ( a , b ): 100 µm. c Quantification of average CBF, particle clearance speed, and clearance per beat (CPB) in human airways BG0-6 and rat airways BG0-1. Number of human donors: n = 4. Number of rat donors: n = 6. d left: Clearance directionality as a function of distance in human (red) and rat (blue) airways. Thick lines are average curves. Right: Mean directionality over a flow distance of 80 µm. Number of human donors n = 4; number of rat donors: n = 4. Boxplots: Each solid dot is the mean value of one donor (1–3 BGs, 2–4 FOVs each); red line indicates median, bottom and top edges of the box indicate 25th and 75th percentiles, whiskers indicate minimum and maximum; significance was assessed with two-sided unpaired t-test. Source data for ( c , d ) are provided as a file.

Article Snippet: Cells were then seeded at a density of 30 K cells cm -2 on PureCol (Advanced Biomatrix #5005) coated tissue culture dishes in airway epithelial growth media (Promocell #C-21160) and passaged at 80% confluence.

Techniques:

Journal: Nature Communications

Article Title: Structure and function relationships of mucociliary clearance in human and rat airways

doi: 10.1038/s41467-025-57667-z

Figure Lengend Snippet: a Representative IF-images of primary human airway epithelial cultures from 1 donor grown in different differentiation media for 28 days at ALI and stained for cilia (ATUB, magenta) and secretory cell markers (SCGB1A1, green; MUC5AC, white). Scalebar, 40 µm. b Average luminal cell type composition based on IF-staining (in vitro: N = 4–7 donors, 2 inserts each with 3–8 FOVs each; ex vivo: 9 donors, 1–3 BGs, 2–3 FOVs each). c Mapping of IF-staining data of in vitro and ex vivo samples onto three dimensions. y- axis: percentage of MUC5AC+ cells; x-axis: cilia coverage, i.e., percentage of ciliated (ATUB + ) cells; circle diameter: ratio of SCGB1A1+ to MUC5AC+ cell percentages. d Average percentage of MUC5B expressing cells and e relative percentage of MUC5B expressing cells that also express either SCGB1A1 or MUC5AC from IF-stainings (in vitro: n = 2 (SAGM, BD) to n = 4 (PC, PC-S, mAir) donors, 1–2 inserts each with 1–6 FOVs each; ex vivo: n = 3 donors, BG0, 2–4 FOVs each). Boxplot: Each dot represents mean of one donor; red line indicates median, bottom and top edges of the box indicate 25th and 75th percentiles, whiskers indicate minimum and maximum. Source data for ( b – e ) are provided as a file.

Article Snippet: Cells were then seeded at a density of 30 K cells cm -2 on PureCol (Advanced Biomatrix #5005) coated tissue culture dishes in airway epithelial growth media (Promocell #C-21160) and passaged at 80% confluence.

Techniques: Staining, In Vitro, Ex Vivo, Expressing

Journal: Nature Communications

Article Title: Structure and function relationships of mucociliary clearance in human and rat airways

doi: 10.1038/s41467-025-57667-z

Figure Lengend Snippet: a Quantitative analysis of particle CPB and directionality in primary human airway epithelial cultures grown in different differentiation media for 28 days at ALI compared to human and rat benchmark data. Number of donors (2–3 inserts each with 6 FOVs each) for BD, mAir, SAGM, PC, PC-S, respectively: n = 5, 4, 4, 5, 4. b Quantitative analysis of ciliary beat metrics in airway cells cultured and visualized as in ( a ). Number of donors (2–3 inserts each with 6 FOVs each) for BD, mAir, SAGM, PC, PC-S, respectively: Cilia Coverage, n = 7, 4, 4, 7, 4; Ciliary Beat OP, n = 4, 4, 2 (1 not measurable), 4, 4; Ciliary Beat Amplitude: n = 4, 4, 3, 4, 4; Cilia Length, n = 4, 4, 4, 4, 4; Ciliation Gap, n = 4, 4, 3, 4, 4; Crystalline OP, n = 4, 4, 3, 4, 4. Boxplots in ( a , b ): Each solid dot is the mean value of one donor (1–3 BGs, 2–4 FOVs each); red line indicates median, bottom and top edges of the box indicate 25th and 75th percentiles, whiskers indicate minimum and maximum. Dotted lines indicate average human and rat benchmark values. c Top: Predicted CPB and clearance directionality in in vitro cultures compared to predicted human airway performance (red). Shaded regions indicate uncertainty based on spread of input metrics. Measured mean values per donor are overlaid (dots). Bottom: Predicted change in CPB and directionality of in vitro cultures in different media if an individual cilia input parameter is set to match ex vivo values. Source data for ( a – c ) are provided as a file.

Article Snippet: Cells were then seeded at a density of 30 K cells cm -2 on PureCol (Advanced Biomatrix #5005) coated tissue culture dishes in airway epithelial growth media (Promocell #C-21160) and passaged at 80% confluence.

Techniques: Cell Culture, In Vitro, Ex Vivo