Ndp scan

Formalin-fixed, paraffin-embedded tumor sections were labelled using the ultraVIEW DAB system (Ventana, Basel, Switzerland). Briefly, antigen retrieval was carried out by heating slides for 90 minutes at 95°C in 1 mmol/L EDTA at pH 7.8. Slides were then incubated with LXRβ mAb (ab24361 (Abcam) at a 1/20 dilution for 32 min at 37°C. The stained arrays were counterstained with hematoxylin and mounted in Aquamount (Dako, Les Ulis, France). Labeling was detected using a Ventana Benchmark XT automat (Ventana).
Slides were then digitized using Hamamatsu NanoZoomer 2.0HT and NDP Scan software (Hamamatsu Photonics, Massy, France). Nucleus detection and quantification were performed using TissueStudio 3.6.1 software (Definiens, Munich, Germany).

Paraffin-embedded kidney sections (5μm) were stained with standard Sirius Red staining to identify collagen content using reagents from Sigma-Aldrich (St. Louis, MO). Sections were incubated in 0.1% Picro-Sirius Red solution followed by 0.5% Glacial Acetic acid. Slides were scanned with a Hamamatsu Nanozoomer 2.0 HT utilizing NDP.scan and NDP.view as the imaging software at 20X magnification (Hamamatsu). The total area of collagen deposition (red staining) was identified in 12 sections of the digitized images from each kidney (n = 6–7) and measured as the collagen-to-field ratio using Adobe Photoshop 7.0. Analyses were performed by an investigator blinded to the experimental groups.

Images of haematoxylin and eosin-stained slides of gonadal AT were acquired in a Nanozoomer-SQ (Hamamatsu Photonics) using NDP.scan (version 1.0) with a magnification of 20×. Afterwards, at least five random fields per slide were analysed with ImageJ (version 1.51 h) using the Adiposoft (version 1.16) plugin^{45 (link)} using an adipocyte diameter range from 10 to 100 µm.