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BACKGROUND: Several studies have indicated the central role of the megalin/cubilin multiligand endocytic receptor complex in protein reabsorption in the kidney proximal tubule. However, the poor viability of the existing megalin-deficient mice precludes further studies and comparison of homogeneous groups of mice. METHODS: Megalin- and/or cubilin-deficient mice were generated using a conditional Cre-loxP system, where the Cre gene is driven by the Wnt4 promoter. Kidney tissues from the mice were analysed for megalin and cubilin expression by quantitative reverse transcription-polymerase chain reaction, western blotting and immunohistochemistry. Renal albumin uptake was visualized by immunohistochemistry. Twenty-four-hour urine samples were collected in metabolic cages and analysed by sodium dodecyl sulphate-polyacrylamide gel electrophoresis and western blotting. Urinary albumin/creatinine ratios were measured by ELISA and the alkaline picrate method. RESULTS: The Meg(lox/lox);Cre(+), Cubn(lox/lox);Cre(+) and Meg(lox/lox), Cubn(lox/lox);Cre(+) mice were all viable, fertile and developed normal kidneys. Megalin and/or cubilin expression, assessed by immunohistology and western blotting, was reduced by >89%. Consistent with this observation, the mice excreted megalin and cubilin ligands such as transferrin and albumin in addition to low-molecular weight proteins. We further show that megalin/cubilin double-deficient mice excrete albumin with an average of 1.45 ± 0.54 mg/day, suggesting a very low albumin concentration in the glomerular ultrafiltrate. CONCLUSIONS: We report here the efficient genetic ablation of megalin, cubilin or both, using a Cre transgene driven by the Wnt4 promoter. The viable megalin/cubilin double-deficient mice now allow for detailed large-scale group analysis, and we anticipate that the mice will be of great value as an animal model for proximal tubulopathies with disrupted endocytosis.

Original publication

DOI

10.1093/ndt/gfr525

Type

Journal article

Journal

Nephrol Dial Transplant

Publication Date

11/2011

Volume

26

Pages

3446 - 3451

Keywords

Albumins, Animals, Blotting, Western, Creatinine, Disease Models, Animal, Endocytosis, Female, Green Fluorescent Proteins, Immunoenzyme Techniques, Integrases, Kidney Tubules, Proximal, Low Density Lipoprotein Receptor-Related Protein-2, Male, Mice, Mice, Inbred C57BL, Mice, Knockout, RNA, Messenger, Real-Time Polymerase Chain Reaction, Receptors, Cell Surface, Wnt4 Protein