README
Whole-slide histology and immunofluorescence images of human ascending thoracic aortic tissue after ex vivo mechanical loading and stent indentation in a biosafe bioreactor platform

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1. GENERAL INFORMATION
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This dataset contains whole-slide images (WSI) of human ascending thoracic aortic tissue stained with hematoxylin and eosin (H&E), Elastica van Gieson (EvG), and multichannel immunofluorescence (DAPI, CD31, VE-cadherin). The tissue was tested in a custom biosafe ex vivo platform that combines multiaxial mechanical loading with a commercial bioreactor, which allows mechanical testing and viability preservation of native tissue at the same time.

The images document tissue structure (elastin and collagen architecture, cellularity) and endothelial coverage and junctional organization in stented, stretched, and control groups at 0 h, 24 h, and 48 h.

Related publication: [DOI to be added]

Ethics: The use of donor specimens was approved by the Ethics Committee of the Medical University of Graz (approval number 32-451 ex19/20).

Location of image acquisition: Graz, Austria.

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2. FILES INCLUDED
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1) Raw whole-slide images
   - 30 brightfield slides (H&E and EvG), SVS format, file names: MY<n>_26_HE.svs and MY<n>_26_EvG.svs
   - 12 immunofluorescence images, VSI format, file names: CD31_VE_MY<n>_26_<section>_20260713.vsi
     Note: VSI files are header files. The pixel data are stored in the accompanying data folder of each file (.ets files). Keep each VSI file together with its folder.
2) WSI_DescriptionTable.csv: description of every file (sample, stain or panel, format, size, MD5 checksum, protocol, group, timepoint, notes)
3) ReadMe.txt: this file

File naming:
   MY<n>   sample number (MY1 to MY15)
   26      year
   HE/EvG  stain
   CD31_VE immunofluorescence (CD31 and VE-cadherin, with DAPI)
   20260713 acquisition date of the fluorescence image (13 July 2026)

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3. HISTOLOGICAL ANALYSIS
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Following biopsy collection, tissue samples were washed three times with pre-warmed PBS (5 min per wash) and then fixed in 4 % formaldehyde solution (ROTI Histofix, Carl Roth, Karlsruhe, Germany) for 2 h at 37 degrees C. After fixation, the samples were washed three times with PBS (5 min per wash) and stored in PBS at 4 degrees C until embedding. The specimens were dehydrated in a graded series of ethanol (70 % to 100 %) and embedded in paraffin. Sections were stained with H&E to identify endothelial and smooth muscle cell nuclei, and with EvG to visualize elastic fibers and collagen content.

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4. IMMUNOFLUORESCENCE STAINING
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Endothelial coverage and junctional organization were evaluated by immunofluorescence on paraffin-embedded tissue sections using markers for CD31 and VE-cadherin.

- Sections were deparaffinized, rehydrated, and subjected to heat-induced epitope retrieval in citrate buffer (pH 6.0) using two 20 min microwave-heating cycles.
- All subsequent incubation steps were performed in a humidified chamber, using approximately 100 to 200 microliters of reagent per section.
- Sections were outlined with a hydrophobic barrier pen and blocked for 10 min with 5 % bovine serum albumin (BSA).
- Primary antibodies (applied simultaneously, 45 min, diluted in antibody diluent):
    mouse monoclonal anti-CD31 (ab9498, Abcam), working concentration 5 micrograms/mL
    rabbit polyclonal anti-VE-cadherin (ab33168, Abcam), working concentration 5 micrograms/mL
- Four washes with PBS containing Tween 20 (PBS-T).
- Secondary antibodies (30 min, protected from light, diluted 1:1000 in PBS):
    Alexa Fluor 488 goat anti-rabbit IgG (ab150113, Abcam), detects VE-cadherin
    Alexa Fluor 594 goat anti-mouse IgG (ab150080, Abcam), detects CD31
- Four washes with PBS-T.
- Nuclear counterstain: 4',6-diamidino-2-phenylindole (DAPI), diluted 1:2000 in PBS, 5 min.
- Four washes with distilled water, air-drying in the dark, and mounting with ProLong Gold antifade mounting medium (P36930, Invitrogen).

CD31 staining was used to evaluate endothelial coverage and continuity along the luminal surface. VE-cadherin staining was used to evaluate the localization and continuity of endothelial cell-cell junctions.

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6. IMAGE ACQUISITION
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Brightfield (H&E, EvG):
   Format: SVS

Fluorescence (DAPI, CD31, VE-cadherin):
   Format: VSI (with companion data folders)
   Channels: DAPI (nuclei), Alexa Fluor 488 (VE-cadherin), Alexa Fluor 594 (CD31)


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7. IMAGE ANALYSIS AND SOFTWARE
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Fluorescence intensity per cell was quantified using QuPath, version 0.7.0 (Bankhead et al., 2017, Scientific Reports, "QuPath: Open source software for digital pathology image analysis").

Recommended software for opening the files:
   - QuPath (SVS and VSI through Bio-Formats)
File sizes range from about 1 MB (VSI header files) to about 900 MB (SVS).
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8. CONTACT AND CITATION
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Contact: Gerhard A. Holzapfel, Institute of Biomechanics, Graz University of Technology (TU Graz), Graz, Austria. E-mail: holzapfel@tugraz.at
Please cite the related publication [DOI to be added] when using this dataset.