Effects of rhein on intestinal epithelial tight junction in IgA nephropathy

Figure 1 Electron micrograph of intestinal epithelial cells showing tight junction. A: In the control group, the tight junction appeared as an electron-dense belt at the apex of the intestinal epithelial cells (arrow), indicating an intact intestinal mucosal barrie; B: In the IgA nephropathy (IgAN) group group, the intercellular space was widened, the tight junction was indistinct, and the density was reduced (arrow); C and D: In the Rhein-treated and Rhein-prevented groups, the density of the tight junctions was increased compared with that in the IgAN group (arrows). TEM: Transmission electron microscopy.

Figure 2 Location of tight junction protein occludin in rat ileum. Laser confocal microscope immunofluorescence staining of ileum from all four groups of rats. A: Cross-section of a normal intestinal villus. Immunoreactive occludin was localized at the apex of intestinal epithelial cells, consistent with the site of the intestinal mucosal barrier (arrow); B: Cross-section of an intestinal villus in the IgA nephropathy (IgAN) group. Occludin immunofluorescence staining became weak and discontinuous (arrow); C and D: Cross-section of an intestinal villus in the Rhein-treated group and cross-section of an intestinal villus in the Rhein-prevented group. Compared with the IgAN group, occludin immunofluorescence staining became stronger and continuous (arrows). Scale bars = 10 μm. IFC: Integrated fluidic circuit.

Figure 3 Location of tight junction protein zona occludens protein-1 in rat ileum. Laser confocal microscope immunofluorescence staining of ileum from all four groups of rats. A: Cross-section of a normal intestinal villus. Immunoreactive zona occludens protein (ZO)-1 was localized at the apex of intestinal epithelial cells, consistent with the site of the intestinal mucosal barrier (arrow); B: Cross-section of an intestinal villus in the IgA nephropathy (IgAN) group. ZO-1 immunofluorescence staining became weak and discontinuous (arrow); C and D: Cross-section of an intestinal villus in the Rhein-treated group and cross-section of an intestinal villus in the Rhein-prevented group. Compared with the IgAN group, ZO-1 immunofluorescence staining became stronger and continuous (arrows). Scale bars = 10 μm. IFC: Integrated fluidic circuit.

Figure 4 Reverse transcriptase polymerase chain reaction analysis for zona occludens protein-1 and occludin mRNA in rat ileum. By reverse transcriptase polymerase chain reaction, amplification products of expected size [150 bp for zona occludens protein (ZO)-1 and 170 bp for occludin] were obtained in the ileum in all four groups of rats. β-actin was the housekeeping protein. The levels of occludin and ZO-1 expression in the IgA nephropathy (IgAN) group were lower than in the control group. In the Rhein-treated and Rhein-prevented groups, the levels were higher than in the IgAN group. 1: Control; 2: IgAN; 3: Rhein-treated; 4: Rhein-prevented.

Figure 5 Western blotting analysis for zona occludens protein-1 and occludin of rat ileum. Western blotting analysis revealed zona occludens protein (ZO)-1 immunoreactivity by a band of 220 kDa in the control group and by a weaker band at the same level in the IgA nephropathy (IgAN) group. Compared with the IgAN group, there were stronger bands at the same level in the Rhein-treated and Rhein-prevented groups. Occludin immunoreactivity was revealed by a band of 65 kDa in the control group and by a weaker band at the same level in the IgAN group. Compared with the IgAN group, there were stronger bands at the same level in the Rhein-treated and Rhein-prevented groups. β-Actin was the housekeeping protein. 1: Control; 2: IgAN; 3: Rhein-treated; 4: Rhein-prevented.