Fig. 2

Hepatocyte proliferation and cholangiocyte apoptosis during liver metamorphosis. a–f: Mitosis marker phospho-histone 3 immunostaining (brown) in liver section at various metamorphic stages. Paraffin sections (4 μm) were stained with DAB and counterstained with hematoxylin (blue/purple) nuclear stain. Note that staining is localized only in hepatocytes. g–l: Apoptosis marker alkaline phosphatase staining (dark purple) in liver section at various metamorphic stages. Paraffin sections (4 μm) were stained with alkaline phosphatase (AP) substrate NBT/BCIP and counterstained with nuclear Fast Red (pink/red stain). In larval (L) and metamorphic stages 1 (M1) and 2 (M2), AP activity was located in canaliculi between hepatocytes and the lumen lining of the cholangiocytes. In metamorphic stage 3 (M3), AP activity was located in canaliculi between hepatocytes, blood cells and whole cholangiocytes. In metamorphic stage 4 (M4), only a few blood cells contain AP activity. In newly transformed juvenile (JV), no AP activity was detected. Scale bar: 50 μm. BD: bile duct; H: hepatocyte