A susceptibility gene for kidney disease in an obese mouse model of type II diabetes maps to chromosome 8

Abstract

Most mouse models of diabetes do not fully reproduce features of human diabetic nephropathy, limiting their utility in inferring mechanisms of human disease. Here we performed detailed phenotypic and genetic characterization of leptin-receptor (Lepr) deficient mice on the FVB/NJ background (FVB(db/db)), an obese model of type II diabetes, to determine their suitability to model human diabetic nephropathy. These mice have sustained hyperglycemia, significant albuminuria and characteristic diabetic renal findings including mesangial sclerosis and nodular glomerulosclerosis after 6 months of age. In contrast, equally obese, hyperglycemic Lepr/Sur1 deficient C57BL/6J (Sur1 has defective insulin secretion) mice have minimal evidence of nephropathy. A genome-wide scan in 165 Lepr deficient backcross progeny derived from FVB/NJ and C57BL/6J identified a major locus influencing nephropathy and albuminuria on chromosome 8B1-C5 (Dbnph1 locus, peak lod score 5.0). This locus was distinct from those contrasting susceptibility to beta cell hypertrophy and HIV-nephropathy between the same parental strains, indicating specificity to diabetic kidney disease. Genome-wide expression profiling showed that high and low risk Dbnph1 genotypes were associated with significant enrichment for oxidative phosphorylation and lipid clearance, respectively; molecular pathways shared with human diabetic nephropathy. Hence, we found that the FVB(db/db) mouse recapitulates many clinical, histopathological and molecular features of human diabetic nephropathy. Identifying underlying susceptibility gene(s) and downstream dysregulated pathways in these mice may provide insight into the disease pathogenesis in humans.

Figure 1

A. Tail Cuff measurements show no differences in systolic blood pressure and heart rate between FVB ^db/db and lean FVB^db/+ mice B. Serial SDS-PAGE of spot urine samples from one FVB ^db/db mouse at three different ages. BSA standards are shown on the right panel. C. Progressive increase in the proportion of mice showing albuminuria >1mg/ml, as measured by SDS-PAGE of spot urine samples compared to a dilution of BSA standard (N=12–18 male and famel mice per age group). D. Urine albumin/creatinine ratio at 28 and 52 weeks of age (*p<0.05 FVB ^db/db vs. FVB^db/+)

Figure 2. Histopathology of FVB^db/db Kidneys

A. Diffuse and global mesangial sclerosis affecting all glomeruli (20X). B. Glomerulus with nodular mesangial sclerosis (60X). C. Hyaline accumulation in Bowman's capsule forming a "capsular drop" lesion (60X). D. Severe mesangial sclerosis occluding glomerular capillary lumina in continuity with arteriosclerosis of the preglomerular arteriole (60X).

Figure 3. Comparison of histopathology between FVB^db/db (A–C) and B6^{db/db Sur1−/−} (D–F)

There is diffuse and global mesangial sclerosis in FVB^db/db mice (A, 40X) but minimal mesangial sclerosis in B6^{db/db Sur1−/−} glomeruli (C, 40X). Proteinaceous casts dilate the outer medullary tubules in FVB ^db/db mice (B, 60X), but not B6^{db/db Sur1−/−} mice (D, 60X). Electron micrograph of FVB^db/db glomerulus shows thickened glomerular basement membranes and effaced foot processes (C). A representative capillary of B6^{db/db Sur1−/−} glomerular capillary shows normally thin glomerular basement membrane with intact foot processes (F).

Figure 4. Linkage of diabetic nephropathy to chromosome 8 in backcross mice

Distribution of histology score as quantitative trait (A) and binary trait (B, 1= no disease, 2 = any disease), and Ln-transformed albumin/creatinine ratio (C). Below each histogram, the genome-wide lod plots for these traits for nonparametric analysis of linkage (after fine-mapping with 7 microsatellite markers on chromosome 8) are shown (panels D–F). The X axis denotes genetic distance in centimorgans (cM), chromosomes are listed on the bottom. The Y axis shows the lod score. The dashed lines represent the genome-wide significance thresholds based on 10,000 permutations.

Figure 5. Linkage of nephropathy subphenotypes to chromosome 8

A. Lod plot demonstrating linkage of all nephropathy subphenotypes to chromosome 8 after fine-mapping with 7 microsatellite markers. The X axis denotes genetic distance in centimorgans (cM). The Y axis shows the lod score. The location of genotyped markers is indicated above the plots. The horizontal line above the lod plots shows the lod −2 interval for the Dbnph1 locus B. Phenotypic values associated with FVB/FVB vs FVB/B6 genotype at the peak lod score (D8Mit248).

Figure 6

Genotypic differences in expression of selected transcripts among mice with no evidence of glomerulosclerosis. Jund, Ocel1 Ndufa13. Y axis expression level normalized to a wild type B6 mouse. FF= FVB/FVB genotype (n=7) and FB= FVB/B6 genotype (N=13) at the Dbnph1 locus on chromosome 8. * p<0.05 by T-test.