Figure 4

Rab7l1 and Lrrk2 deficient mice exhibit common lysosomal pathology in renal proximal tubule cells.

(A) Gross appearance of representative kidneys from Rab7l1 KO, Lrrk2 KO, double KO or littermate control mice at 11–14 months of age. Scale bar = 10 mm. (B) Kidney weight and length were significantly greater in Rab7l1 KO, Lrrk2 KO or double KO mice compared with control mice as labeled. The color of kidney was significantly much darker in the mutants. Data represent mean ± SEM. n = 6 (control), 5 (Rab7l1 KO), 5 (Lrrk2 KO), 4 (double KO). *p < 0.05, **p < 0.01. (C) Auto-fluorescent granules were readily apparent in kidney proximal tubule cells of Rab7l1 KO, Lrrk2 KO or double KO mice at 11–14 months of age, but not in kidney tissue from control mice. Scale bar = 50 μm. (D) Histological analyses of kidney from Rab7l1 KO mice. Oil red O staining (top row) revealed the accumulation of lipids in these vacuoles. Jones methenamine silver with eosin counterstaining (middle row) or PAS staining (bottom row) revealed the formation of extensive clear intracytoplasmic vacuoles in kidney proximal tubule cells of 10-month-old Rab7l1 KO mice. Scale bar = 100 μm (Oil red O). Original magnifications: ×400 (HE-silver) and ×600 (PAS). (E) Quantification of the number of the structures with the intracytoplasmic vacuoles per 20× microscopic field. The counts in each indicated kidney tissue were made in at least 20 different 20× microscopic field per kidney. White bar: Control (Rab7l1 (+/−)) mice; black bar: Rab7l1 KO mice. Data represent mean ± SEM. n = 3 per genotype. **p < 0.01. (F) Electron microscopic analysis revealed the accumulation of electron-dense material (arrows) within enlarged secondary lysosomes in proximal tubule cells of 10-month-old Rab7l1 KO mice. These inclusions are rounded, electron dense, of varying size and often have a whorled or lamellated texture (see Figure S3G). P, proximal tubule cells; D, distal tubule cells; BB, brush border. Scale bar = 5 μm. (G) Immunostaining of kidney sections revealed the accumulation of lysosomal proteins Cathepsin B and LAMP2 in 11- to 14-month-old Rab7l1 KO, Lrrk2 KO and double KO mice. Red arrows indicate the immunopositive signals. Scale bars = 20 μm.