Iron accumulation in periportal hepatocytes of neogenin mutant mice. (A) Diagram of the retrotranspon insertion in neogenin gene. Exons 7 and 8, surrounding intron, and location of genotyping primers are shown diagrammatically. Primer sequences are also indicated. (B) Genotyping and decreased neogenin expression in neogenin mutant mice. Livers of indicated genotype were lysed and resulting lysates were subjected to immunoblotting with anti-neogenin antibody. The top panel shows PCR genotyping. The 327-bp fragment was detected in homozygous mice where the 169-bp fragments in wild-type mice. (C-D) Histologic detection of iron content on cryostat sections of liver (C) and spleen (D) of wild-type and neogenin mutant (neogenin^m/m) mice. Five different wild-type and mutant mice were examined, and representative images imaged by Axiophot microscope (Zeiss) and processed using Adobe Photoshop CS2 software are shown. Note iron accumulation was observed in the periportal (portal tract [PT]), but not central vein (CV), regions of P25-old neogenin^m/m mice and absence thereof in the red pulp of the spleen. (E) Western blot analysis of liver and spleen lysates from indicated genotype with anti-ferritin-H antibodies. Molecular weights are indicated. (F) Quantitative determination of iron content (μmol/g dry weight; see “Tissue iron staining and quantification”) in various organs of P25-old wild-type (white), neogenin^+/m (purple), and neogenin^m/m (blue) mice. Means ± SEM of 5 samples for each group were presented. **,#Significant changes (P < .01) in neogenin^m/m mice compared with the wild-type control.