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A mutation in Ampd2 is associated with nephrotic syndrome and hypercholesterolemia in mice.

Helmering J, Juan T, Li CM, Chhoa M, Baron W, Gyuris T, Richards WG, Turk JR, Lawrence J, Cosgrove PA, Busby J, Kim KW, Kaufman SA, Cummings C, Carlson G, Véniant MM, Lloyd DJ - Lipids Health Dis (2014)

Bottom Line: We identify the third gene as Ampd2, a liver isoform of AMP Deaminase (Ampd), a central component of energy and purine metabolism pathways.The causative mutation was a guanine-to-thymine transversion resulting in an A341S conversion in Ampd2.Metabolomic analysis confirmed an increase in hepatic AMP levels and a decrease in allantoin levels consistent with Ampd2 deficiency, and increases in campesterol and β-sitosterol.

View Article: PubMed Central - PubMed

Affiliation: Department of Metabolic Disorders, Amgen Inc, One Amgen Center Dr, Thousand Oaks, CA 91320, USA. dlloyd@amgen.com.

ABSTRACT

Background: Previously, we identified three loci affecting HDL-cholesterol levels in a screen for ENU-induced mutations in mice and discovered two mutated genes. We sought to identify the third mutated gene and further characterize the mouse phenotype.

Methods: We engaged, DNA sequencing, gene expression profiling, western blotting, lipoprotein characterization, metabolomics assessment, histology and electron microscopy in mouse tissues.

Results: We identify the third gene as Ampd2, a liver isoform of AMP Deaminase (Ampd), a central component of energy and purine metabolism pathways. The causative mutation was a guanine-to-thymine transversion resulting in an A341S conversion in Ampd2. Ampd2 homozygous mutant mice exhibit a labile hypercholesterolemia phenotype, peaking around 9 weeks of age (251 mg/dL vs. wildtype control at 138 mg/dL), and was evidenced by marked increases in HDL, VLDL and LDL. In an attempt to determine the molecular connection between Ampd2 dysfunction and hypercholesterolemia, we analyzed hepatic gene expression and found the downregulation of Ldlr, Hmgcs and Insig1 and upregulation of Cyp7A1 genes. Metabolomic analysis confirmed an increase in hepatic AMP levels and a decrease in allantoin levels consistent with Ampd2 deficiency, and increases in campesterol and β-sitosterol. Additionally, nephrotic syndrome was observed in the mutant mice, through proteinuria, kidney histology and effacement and blebbing of podocyte foot processes by electron microscopy.

Conclusion: In summary we describe the discovery of a novel genetic mouse model of combined transient nephrotic syndrome and hypercholesterolemia, resembling the human disorder.

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Related in: MedlinePlus

Electron micropscopic evaluation of the glomeruli fromAmpd2m/mmice. Glomeruli of Ampd2m/m mice showed diffuse effacement, sloughing, and fusing of podocyte foot processes (FP). Extensive blebbing, fusing, and surface pseudomembrane formation (PSM) was evident along the glomerular tuft endothelium of Ampd2m/m mice (white arrowheads). In addition to a minimal to moderate thickening of the basal lamina (GBM), there was a subtle increase in cellularity of the mesangial matrix (MM) and an increase in inflammatory cells, predominantly lymphocytes (LC) in Ampd2m/m mice. RBC = Red Blood Cell. Upper photomicrographs 4200×; lower photomicrographs 6000×; Bar = 2 μm.
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Fig7: Electron micropscopic evaluation of the glomeruli fromAmpd2m/mmice. Glomeruli of Ampd2m/m mice showed diffuse effacement, sloughing, and fusing of podocyte foot processes (FP). Extensive blebbing, fusing, and surface pseudomembrane formation (PSM) was evident along the glomerular tuft endothelium of Ampd2m/m mice (white arrowheads). In addition to a minimal to moderate thickening of the basal lamina (GBM), there was a subtle increase in cellularity of the mesangial matrix (MM) and an increase in inflammatory cells, predominantly lymphocytes (LC) in Ampd2m/m mice. RBC = Red Blood Cell. Upper photomicrographs 4200×; lower photomicrographs 6000×; Bar = 2 μm.

Mentions: To confirm the effect of the A341S mutation in the kidneys of Ampd2m/m mice we investigated Ampd2 protein abundance in both strains and verified that Ampd2 protein was absent in the liver and kidney of pure B6 Ampd2m/m mice (Figure 6A). NS was verified in Ampd2m/m mice by kidney histology (Figure 6B). Proteinaceous tubular casts and interstitial nephritis were observed in Ampd2m/m mice and not Ampd2+/+ mice (Figure 6B-D). By TEM, glomeruli of selected Ampd2m/m mice and Ampd2+/+ mice were examined to ascertain any ultrastructural differences (Figure 7). Glomeruli from Ampd2m/m mice showed effacement, fusion, and blebbing of podocyte foot processes, minimal to moderate thickening of the glomerular basement membrane, as well as denuding with subsequent pseudomembrane formation of the glomerular endothelium. While damage to foot processes, basal lamina and glomerular endothelium represents impairment of the glomerular filtration membrane, the subtle increase in cellularity of the mesangium, along with a predominant lymphocytic inflammatory infiltrate seen in Ampd2m/m mice, further supports changes secondary to glomerular dysfunction with ensuing proteinuria.Figure 6


A mutation in Ampd2 is associated with nephrotic syndrome and hypercholesterolemia in mice.

Helmering J, Juan T, Li CM, Chhoa M, Baron W, Gyuris T, Richards WG, Turk JR, Lawrence J, Cosgrove PA, Busby J, Kim KW, Kaufman SA, Cummings C, Carlson G, Véniant MM, Lloyd DJ - Lipids Health Dis (2014)

Electron micropscopic evaluation of the glomeruli fromAmpd2m/mmice. Glomeruli of Ampd2m/m mice showed diffuse effacement, sloughing, and fusing of podocyte foot processes (FP). Extensive blebbing, fusing, and surface pseudomembrane formation (PSM) was evident along the glomerular tuft endothelium of Ampd2m/m mice (white arrowheads). In addition to a minimal to moderate thickening of the basal lamina (GBM), there was a subtle increase in cellularity of the mesangial matrix (MM) and an increase in inflammatory cells, predominantly lymphocytes (LC) in Ampd2m/m mice. RBC = Red Blood Cell. Upper photomicrographs 4200×; lower photomicrographs 6000×; Bar = 2 μm.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4232700&req=5

Fig7: Electron micropscopic evaluation of the glomeruli fromAmpd2m/mmice. Glomeruli of Ampd2m/m mice showed diffuse effacement, sloughing, and fusing of podocyte foot processes (FP). Extensive blebbing, fusing, and surface pseudomembrane formation (PSM) was evident along the glomerular tuft endothelium of Ampd2m/m mice (white arrowheads). In addition to a minimal to moderate thickening of the basal lamina (GBM), there was a subtle increase in cellularity of the mesangial matrix (MM) and an increase in inflammatory cells, predominantly lymphocytes (LC) in Ampd2m/m mice. RBC = Red Blood Cell. Upper photomicrographs 4200×; lower photomicrographs 6000×; Bar = 2 μm.
Mentions: To confirm the effect of the A341S mutation in the kidneys of Ampd2m/m mice we investigated Ampd2 protein abundance in both strains and verified that Ampd2 protein was absent in the liver and kidney of pure B6 Ampd2m/m mice (Figure 6A). NS was verified in Ampd2m/m mice by kidney histology (Figure 6B). Proteinaceous tubular casts and interstitial nephritis were observed in Ampd2m/m mice and not Ampd2+/+ mice (Figure 6B-D). By TEM, glomeruli of selected Ampd2m/m mice and Ampd2+/+ mice were examined to ascertain any ultrastructural differences (Figure 7). Glomeruli from Ampd2m/m mice showed effacement, fusion, and blebbing of podocyte foot processes, minimal to moderate thickening of the glomerular basement membrane, as well as denuding with subsequent pseudomembrane formation of the glomerular endothelium. While damage to foot processes, basal lamina and glomerular endothelium represents impairment of the glomerular filtration membrane, the subtle increase in cellularity of the mesangium, along with a predominant lymphocytic inflammatory infiltrate seen in Ampd2m/m mice, further supports changes secondary to glomerular dysfunction with ensuing proteinuria.Figure 6

Bottom Line: We identify the third gene as Ampd2, a liver isoform of AMP Deaminase (Ampd), a central component of energy and purine metabolism pathways.The causative mutation was a guanine-to-thymine transversion resulting in an A341S conversion in Ampd2.Metabolomic analysis confirmed an increase in hepatic AMP levels and a decrease in allantoin levels consistent with Ampd2 deficiency, and increases in campesterol and β-sitosterol.

View Article: PubMed Central - PubMed

Affiliation: Department of Metabolic Disorders, Amgen Inc, One Amgen Center Dr, Thousand Oaks, CA 91320, USA. dlloyd@amgen.com.

ABSTRACT

Background: Previously, we identified three loci affecting HDL-cholesterol levels in a screen for ENU-induced mutations in mice and discovered two mutated genes. We sought to identify the third mutated gene and further characterize the mouse phenotype.

Methods: We engaged, DNA sequencing, gene expression profiling, western blotting, lipoprotein characterization, metabolomics assessment, histology and electron microscopy in mouse tissues.

Results: We identify the third gene as Ampd2, a liver isoform of AMP Deaminase (Ampd), a central component of energy and purine metabolism pathways. The causative mutation was a guanine-to-thymine transversion resulting in an A341S conversion in Ampd2. Ampd2 homozygous mutant mice exhibit a labile hypercholesterolemia phenotype, peaking around 9 weeks of age (251 mg/dL vs. wildtype control at 138 mg/dL), and was evidenced by marked increases in HDL, VLDL and LDL. In an attempt to determine the molecular connection between Ampd2 dysfunction and hypercholesterolemia, we analyzed hepatic gene expression and found the downregulation of Ldlr, Hmgcs and Insig1 and upregulation of Cyp7A1 genes. Metabolomic analysis confirmed an increase in hepatic AMP levels and a decrease in allantoin levels consistent with Ampd2 deficiency, and increases in campesterol and β-sitosterol. Additionally, nephrotic syndrome was observed in the mutant mice, through proteinuria, kidney histology and effacement and blebbing of podocyte foot processes by electron microscopy.

Conclusion: In summary we describe the discovery of a novel genetic mouse model of combined transient nephrotic syndrome and hypercholesterolemia, resembling the human disorder.

Show MeSH
Related in: MedlinePlus