Quantitative phenotyping of Nphs1 knockout mice as a prerequisite for gene replacement studies.

Steroid-resistant nephrotic (SRNS) syndrome is the second most frequent cause of chronic kidney disease before the age of 25 years. Nephrin, encoded by NPHS1, localizes to the slit diaphragm of glomerular podocytes and is the predominant structural component of the glomerular filtration barrier. Biallelic variants in NPHS1 can cause congenital nephrotic syndrome of the Finnish type (CNS-1) for which, to date, no causative therapy is available. Recently, adeno-associated virus (AAV) vectors targeting the glomerular podocyte have been assessed as a means for gene replacement therapy. We here established quantitative and reproducible phenotyping of a published, conditional Nphs1 knockout mouse model ( Nphs1tm1.1Pgarg /J and Nphs2-Cre+ ) in preparation of a gene replacement study employing AAV vectors. Nphs1 knockout mice ( Nphs1fl/fl Nphs2-Cre+ ) exhibited: i) A median survival rate of 18 days (range from 9-43 days; males 16.5 days, females 20 days); ii) Average foot process (FP) density of 1.0 FP/µm compared to 2.0 FP/µm in controls, and mean filtration slit density was 2.64 µm/µm2 compared to 4.36 µm/µm2 in controls; iii) A high number of proximal tubular microcysts; iv) Development of proteinuria within the first week of life as evidenced by urine albumin/creatinine ratios; v) Significantly reduced levels of serum albumin, and elevated BUN and creatinine levels. For none of these phenotypes in Nphs1 knockout mice, significant differences between sexes were observed. We quantitatively characterized 5 different phenotypic features of CNS in Nphs1fl/fl Nphs2-Cre+ mice. Our results will facilitate future gene replacement therapy projects by allowing for sensitive detection of, even subtle molecular, effects.