Figure 5. GRK6-deficient mice spontaneously develop autoimmune disease.

(a) Serum from 40-week-old control WT and GRK6-deficient female (WT: n=14, GRK6-deficient: n=9) and male mice (WT: n=12, GRK6-deficient: n=8) were collected and tested for presence of anti-dsDNA antibodies. Serum from 40-week-old MFG-E8-deficient female mice (n=4) was used as a positive control. (b) Increased deposition of immune complexes in glomeruli of GRK6-deficient mice at 40 weeks of age was assessed by anti-immunoglobulin staining. Scale bar, 100 μm. (c) Paraffin-embedded kidney sections of WT and GRK6-deficient mice at 40 weeks of age were stained using periodic acid-Schiff. Arrowheads indicate a mesangial expansion with increase in matrix and cellularity owing to increased mononuclear cells. Scale bar, 20 μm. (d) Spleens isolated from WT and GRK6-deficient mice of 4-week-old (n=8), 16-week-old (n=4) and 40-week-old (n=9) were weighed, and spleen weight relative to body weight were measured. And the average numbers are shown with s.d. (e) Splenocytes from WT (n=7) and GRK6-deficient mice (n=6) of 40-week-old mice were counted, and the numbers were shown in the graph. (f) Spleen sections from WT mice were stained with anti-CD68 (green), anti-GRK6 (red) and PNA (blue). The fourth column represents the two-colour (anti-CD68 and anti-GRK6) overlay image. Scale bar, 100 μm. (g) Spleen sections from 40-week-old WT and GRK6-deficient mice were double-stained with TUNEL (green) and anti-CD68 antibody (red). Staining profiles were merged in the third column. Scale bar, 50 μm. Enlarged images are shown in the fourth column. Scale bar, 10 μm. (h) The numbers of unengulfed cells associated with CD68-positive macrophages were counted for 40–80 macrophages and are presented as the number per macrophages. The average numbers are shown with s.e.m. All immunohistochemical data are representative of four independent experiments. *P<0.05, ^**P<0.01 and ***P<0.001. KO, knockout.