Figure 6.

Ferroportin (Fpn) protein levels and localization, iron deposition, and H-ferritin (HFt) gene expression in kidneys of wild-type (WT) and heme oxygenase 1 (HO1) knockout (KO) mouse neonates. (a) Immunofluorescence (IF) staining of Fpn (red channel) in the kidneys of 3 and 11 day old (WT) and KO neonatal mice, analyzed by confocal microscopy. Cell nuclei were counterstained with DAPI (blue). Scale bars correspond to 20 μm. (b) Fpn protein levels in the kidneys of WT and KO mice from all age groups analyzed by Western blotting. WB PVDF membrane stained with Ponceau S was used as a protein loading control. (c) Histochemical analysis of iron loading in the kidneys of 5 and 11 day old WT and KO mouse neonates by Perl’s Prussian Blue staining. Iron deposits were detected in renal cortex of 11 day old KO mice (visible as blue deposits), whereas, in kidneys from WT mice of the same age and 5 day old mice of both genotypes, iron deposits were absent. Renal tissue was counterstained with pararosaniline (pink) and the scale bar corresponds to 10 µm. (d) RT-qPCR analysis of H-Ft mRNA levels in the kidneys of WT and KO mouse neonates ranging from 3–11 days old. The graph presents relative H-Ft transcript levels in arbitrary units (means ± SD, n = 5 per group). We used Student t-tests for all datasets; * p < 0.05, ** p < 0.01, *** p < 0.001. dpp—days postpartum, MWM—molecular weight marker.