Microm hm500m cryostar nx70

Paraformaldehyde-fixed kidneys were sectioned into 20 μm thick longitudinal sections with a cryostat (Microm HM500M/Cryostar NX70; Thermo Scientific, Stockholm, Sweden) and collected onto SuperFrost Plus microscope slides (VWR International, Stockholm, Sweden). After 2 h equilibrating at room temperature, tissue sections were rinsed in hematoxylin (Sigma-Aldrich, Stockholm, Sweden; 50% vol/vol diluted in distilled water) for 6 min, washed with distilled water, and stained with eosin (Histolab; Gothenburg, Sweden) for 30 s. After washing, sections were dried and mounted with VectaMount permanent mounting medium (Vector Laboratories, Burlingame, CA, USA). Once the mounting medium was solidified, the preparations were imaged under a 10× magnification objective using an optical microscope (Leica, Stockholm, Sweden). The morphological evaluations were performed in three non-consecutive sections separated by 100 µm. For each section, 3 fields of view were collected randomly in 3 to 4 individual animals per experimental group.

Paraformaldehyde-fixed SAT and VAT were sectioned into 20 μm thick sections with a cryostat (Microm HM500M/Cryostar NX70; Thermo Scientific, Uppsala, Sweden) and collected onto SuperFrost Plus microscope slides (VWR International AB, Stockholm, Sweden). The sections were rinsed in hematoxylin (Sigma-Aldrich, Stockholm, Sweden; 50% vol/vol diluted in distilled water) for 6 min, washed with distilled water and stained with eosin (Histolab, Askim, Sweden) for 30 s. After washing, sections were dried and mounted with VectaMount permanent mounting medium (Vector Laboratories, CA, USA). All procedures were performed in a cold room at 4 °C. Once the mounting medium was solidified, the preparations were brought up to room temperature and imaged under a 20× magnification objective using an optical microscope (Leica, Wetzlar, Germany). The area of the adipocytes was analyzed using the ImageJ program (National Institutes of Health, Bethesda, MD, USA).