Histopathology and fluorescent immunostaining of larsb-knockout larvae. (A) Lower magnification sagittal views (top row) and higher magnification views (bottom row) of larsb+/+ and larsb−/− larvae. Huge vacuolations, which seemed to disappear in the cytoplasm, were seen in the livers of larsb−/− larvae (black arrows), and some large vacuolations included a bare nucleus (black arrowheads). Scale bar: 25 µm. (B) Western blot analysis of p62 and Lc3b protein expression in larsb+/+ and larsb−/− larvae. β-actin levels served as the loading control. (C) Lower magnification sagittal views of larsb+/+ and larsb−/− larvae (top row). Fluorescent immunostaining against Lc3b (green) and DAPI (blue) of the livers, skeletal muscles, and spinal cords of larsb+/+ and larsb−/− larvae. Livers of larsb−/− larvae had large vacuoles, including floating nuclei and various sized dots with Lc3b immunoreactivity (white arrows). Skeletal muscles and spinal cords of larsb−/− larvae had many dots with Lc3b immunoreactivity (white arrowheads). Scale bar: 10 µm. Lars: leucyl-tRNA synthetase; Lc3b: microtubule-associated protein 1A/1B-light chain 3; DAPI: 4′,6-diamidino-2-phenylindole.
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