Data from: Mouse vocal fold permeability in vivo: Effects of novel low-tech injury and instillation methods
Data files
Dec 11, 2025 version files 1.77 GB
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Figure_1.zip
681.73 MB
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Figure_2-3.zip
106.34 MB
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Figure_4.zip
164.29 MB
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Figure_5.zip
116.36 MB
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Figure_6.zip
375.52 MB
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Figure_7.zip
326.47 MB
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README.md
9.73 KB
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Supporting_Information.zip
17.34 KB
Abstract
Objectives: Vocal fold (VF) injury and instillation are emerging techniques to study laryngeal pathologies in mouse models. Most approaches require high-tech visualization and lengthy anesthesia. Intraperitoneal (IP) naphthalene (NAPH) and topical polidocanol (PDOC) are two chemical methods of injuring murine lower airways. Oropharyngeal aspiration (OA) is used for murine lung instillation. We assessed whether these simple low-tech methods could injure and instill mouse VFs, and whether injury increased in vivo VF epithelial permeability.
Methods: Mouse VFs were injured using IP NAPH at 200, 250, 300, or 350 mg/kg, or OA of PDOC at 0.5% or 2% w/v. Twenty-four hours later, mice received Evans blue (EB) dye OA instillation, then were sacrificed after 30 minutes. Coronal larynx sections were assessed for VF injury. Permeability was measured by imaging EB autofluorescence and quantifying percent positive area and signal intensity. VFs were immunostained for basal cells (p63), tight junctions (ZO-1), and basement membrane (laminin).
Results: VF injury was 100% with ≥350 mg/kg NAPH or high-volume 2% PDOC and 0-40% with other treatments. EB bound VF lamina propria in up to 100% of mice in a volume-dependent manner. Permeability did not differ by injury. Basal cells and tight junctions were decreased in injured VFs. Basement membrane was largely intact.
Conclusions: High-dose NAPH consistently injures mouse VFs. OA reliably instills mouse VFs. Uninjured murine VF epithelium is highly permeable to low molecular weight dye in vivo. Inherent permeability of mouse VFs may facilitate targeted genetic engineering approaches and studies of environmental hazards and drug treatments.
Dataset DOI: 10.5061/dryad.9w0vt4bsc
Description of the data and file structure
The dataset contains microscopy images and data used to generate the graphs and statistical analyses in figures in the manuscript. Images (.TIFF) and data files (.xls) can be viewed and/or analyzed with a number of free or proprietary software programs.
Image naming structure: sample#_slide#_tissue_magnification_filter/stain.
Files and variables
File: Figure_1.zip
Description:
- HE folder: Images used to create Figure 1A-C and analyzed to generate data for Figure 1D. Brightfield images of hematoxylin and eosin (H&E)-stained vocal folds, arranged in folders by injury.
- Figure 1 data.xls: Data underlying Figure 1D. Columns: injury group (categorical), VF sloughing (number of animals per group), no VF sloughing (number of animals per group), % with VF sloughing (percent of animals per group).
File: Figure_2-3.zip
Description:
- EB IF folder: Images used to create Figure 2 and analyzed to generate data for Figure 3. Raw immunofluorescent images of Evans blue red autofluorescence in vocal folds, arranged in folders by administration of dye or mock dye, then by injury.
- Figure 3 data.xls: Data underlying Figure 3.
- Tab 1: Data underlying Figure 3A. Columns: injury group (categorical), EB fluorescence (number of animals per group), no EB fluorescence (number of animals per group), % with EB fluorescence (percent of animals per group).
- Tab 2: Experiment 1 background fluorescence in mock-dyed vocal folds. Columns: injury (categorical), dye (categorical), animal ID (text), %area 1, 2, and 3 (percent area of EB fluorescence in vocal folds in 3 technical replicates), mean %area (mean of %area 1, 2, and 3), MFI 1, 2, and 3 (mean intensity of EB fluorescence (AU) in vocal folds in 3 technical replicates), mean MFI (mean of MFI 1, 2, and 3). Last row: grand mean of background fluorescent %area and MFI for experiment 1.
- Tab 3: Data underlying Figure 3B-C. Columns: injury (categorical), dye (categorical), animal ID (text), %area 1, 2, and 3 (percent area of EB fluorescence in vocal folds in 3 technical replicates), mean %area (mean of %area 1, 2, and 3), background %area (grand mean of background fluorescent %area from tab 2), corrected %area (mean %area - background %area), MFI 1, 2, and 3 (mean intensity of EB fluorescence (AU) in vocal folds in 3 technical replicates), mean MFI (mean of MFI 1, 2, and 3), background MFI (grand mean of background fluorescent MFI from tab 2), corrected MFI (mean MFI - minus background MFI).
File: Figure_4.zip
Description:
- EB-dyed folder: Images used to create Figure 4A-F. Raw immunofluorescent images of Evans blue red autofluorescence in Evans blue-dyed esophagus, lungs, and trachea, arranged in folders by tissue.
- Mock-dyed folder: Images used to create Figure 4G-L. Raw immunofluorescent images of Evans blue red autofluorescence in mock-dyed esophagus, lungs, and trachea, arranged in folders by tissue.
File: Figure_5.zip
Description:
- EB IF folder: Images analyzed to generate data for Figure 5. Raw immunofluorescent images of Evans blue red autofluorescence in vocal folds, arranged in folders by administration of dye or mock dye and by volume.
- Figure 5 data.xls: Data underlying Figure 5.
- Tab 1: Data underlying Figure 5A. Columns: dye volume group (categorical), autofluorescence (number of animals per group), no autofluroescence (number of animals per group), % with autofluorescence (percent of animals per group).
- Tab 2: Experiment 1 background fluorescence in mock-dyed vocal folds. Columns: injury (categorical), dye (categorical), animal ID (text), %area 1, 2, and 3 (percent area of EB fluorescence in vocal folds in 3 technical replicates), mean %area (mean of %area 1, 2, and 3), MFI 1, 2, and 3 (mean intensity of EB fluorescence (AU) in vocal folds in 3 technical replicates), mean MFI (mean of MFI 1, 2, and 3). Last row: grand mean of background fluorescent %area and MFI for experiment 1.
- Tab 3: Experiment 2 background fluorescence in mock-dyed vocal folds. Columns: injury (categorical), dye (categorical), animal ID (text), %area 1, 2, and 3 (percent area of EB fluorescence in vocal folds in 3 technical replicates), mean %area (mean of %area 1, 2, and 3), MFI 1, 2, and 3 (mean intensity of EB fluorescence (AU) in vocal folds in 3 technical replicates), mean MFI (mean of MFI 1, 2, and 3). Last row: grand mean of background fluorescent %area and MFI for experiment 2.
- Tab 4: Data underlying Figure 5B-C. Columns: experiment (categorical), dye (categorical), dye volume (categorical), animal ID (text), %area 1, 2, and 3 (percent area of EB fluorescence in vocal folds in 3 technical replicates), mean %area (mean of %area 1, 2, and 3), background %area (grand mean of background fluorescent %area from tab 2 or 3), corrected %area (mean %area - background %area), MFI 1, 2, and 3 (mean intensity of EB fluorescence (AU) in vocal folds in 3 technical replicates), mean MFI (mean of MFI 1, 2, and 3), background MFI (grand mean of background fluorescent MFI from tab 2 or 3), corrected MFI (mean MFI - minus background MFI).
File: Figure_6.zip
Description:
- HE folder: Images used to create Figure 6A and analyzed to generate data for Figure 6B. Brightfield images of H&E-stained vocal folds, arranged in folders by injury.
- IF folder: Images analyzed to generate data for Figure 6C-D. Raw immunofluorescent images of Evans blue red autofluorescence in vocal folds, arranged in folders by administration of dye or mock dye, then by injury.
- Figure 6 data.xls: Data underlying Figure 6B-D.
- Tab 1: Data underlying Figure 6B. Columns: injury group (categorical), VF sloughing (number of animals per group), no VF sloughing (number of animals per group), % with VF sloughing (percent of animals per group).
- Tab 2: Experiment 3 background fluorescence in mock-dyed vocal folds. Columns: injury (categorical), dye (categorical), animal ID (text), %area 1, 2, and 3 (percent area of EB fluorescence in vocal folds in 3 technical replicates), mean %area (mean of %area 1, 2, and 3), MFI 1, 2, and 3 (mean intensity of EB fluorescence (AU) in vocal folds in 3 technical replicates), mean MFI (mean of MFI 1, 2, and 3). Last row: grand mean of background fluorescent %area and MFI for experiment 3.
- Tab 3: Data underlying Figure 6C-D. Columns: injury (categorical), dye (categorical), animal ID (text), %area 1, 2, and 3 (percent area of EB fluorescence in vocal folds in 3 technical replicates), mean %area (mean of %area 1, 2, and 3), background %area (grand mean of background fluorescent %area from tab 2), corrected %area (mean %area - background %area), MFI 1, 2, and 3 (mean intensity of EB fluorescence (AU) in vocal folds in 3 technical replicates), mean MFI (mean of MFI 1, 2, and 3), background MFI (grand mean of background fluorescent MFI from tab 2), corrected MFI (mean MFI - minus background MFI).
File: Figure_7.zip
Description:
- Folder p63 K14 IF: Images used to create Figure 7A-C and analyzed to generate data for Figure 7D and H. Raw immunofluorescent images of vocal folds stained for p63 (green), cytokeratin 14 (K14; red), and DAPI (blue), arranged in folders by experiment, then by injury. Select images thresholded to remove background signal are also included.
- Figure 7 data.xls: Data underlying Figure 7D and H. Columns: experiment (categorical), injury (categorical), animal ID (text), DAPI+ epithelial cells (number of cells in vocal fold epithelium with blue fluorescent nuclear signal), p63+ epithelial cells (number of cells in vocal fold epithelium with green fluorescent nuclear signal), %p63+ epithelial cells (percent of DAPI+ cells in vocal fold epithelium that are p63+).
- Folder ZO-1 Laminin IF: Images used to create Figure 7E-G. Raw immunofluorescent images of vocal folds stained for zonula occludens 1 (ZO-1, green), laminin (red), and DAPI (blue), arranged in folders by experiment, then by injury. Select images thresholded to remove background signal are also included.
File: Supporting_Information.zip
Description:
- Supporting Information data.xls: Data underlying Table S1 and Figures S3 and S4.
- Tab 1: Data underlying Table S2. Repeated measurements for 20% of quantitative data are presented. Columns: animal ID (text), measurement 1 (number), and measurement 2 (number), presented for raw % area (mean percent area of EB fluorescence in vocal folds averaged from 3 technical replicates), raw MFI (mean fluorescence intensity (AU) of EB fluorescence in vocal folds averaged from 3 technical replicates), p63+ epithelial cells (number of cells in vocal fold epithelium with green fluorescent nuclear signal), and total epithelial cells (number of cells in vocal fold epithelium with blue fluorescent nuclear signal).
- Tab 2: Data underlying Figures S3A and S4A. Columns: experiment (categorical), injury (categorical), animal ID (text), sex (categorical), age in weeks (numerical), VF sloughing (categorical).
- Tab 3: Data underlying Figures S3B-C and S4B-D. Columns: experiment (categorical), dye volume in ul (numerical), animal ID (text), sex (categorical), age in weeks (numerical), corrected %area (mean %area - background %area), corrected MFI (mean MFI - background MFI), total VF area (area of the analyzed region of interest comprising the medial thyroarytenoid muscle and overlying vocal fold lamina propria and epithelium).