Histological and Ultrastructural Liver Analysis

The histological study was performed as described previously [19 (link)]. Briefly, embryonic liver samples were fixed in 10% formalin, dehydrated, embedded in easin, sectioned at 5 μm and stained with hematoxylin and eosin (HE staining). For the ultrastructural study, small tissue fragments were fixed in 2.5% glutaraldehyde in 0.1 mol/L phosphate buffer at 48 ℃ for 1 h, washed in 0.1 mol/L phosphate buffer, and embedded in agar chips, which were further postfixed in 1% osmium tetroxide in 0.1 mol/L phosphate buffer, pH 7.2. they were dehydrated and then embedded in Araldite/Epon (Electron Microscopy Sciences, Washington, USA) and sectioned into semithin slices, that were stained with toluidine blue. The ultrathin sections were contrasted with uranyl acetate and lead citrate and then observed via transmission electron microscopy at 200 kV.