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Background: High blood pressure is the primary risk factor for cardiovascular death worldwide. Autosomal dominant hypertension with brachydactyly clinically resembles salt-resistant essential hypertension and causes death by stroke before 50 years of age. We recently implicated the gene encoding phosphodiesterase 3A ( PDE3A ); however, in vivo modeling of the genetic defect and thus showing an involvement of mutant PDE3A is lacking. Methods: We used genetic mapping, sequencing, transgenic technology, CRISPR-Cas9 gene editing, immunoblotting, and fluorescence resonance energy transfer. We identified new patients, performed extensive animal phenotyping, and explored new signaling pathways. Results: We describe a novel mutation within a 15 base pair (bp) region of the PDE3A gene and define this segment as a mutational hotspot in hypertension with brachydactyly. The mutations cause an increase in enzyme activity. A CRISPR/Cas9-generated rat model, with a 9-bp deletion within the hotspot analogous to a human deletion, recapitulates hypertension with brachydactyly. In mice, mutant transgenic PDE3A overexpression in smooth muscle cells confirmed that mutant PDE3A causes hypertension. The mutant PDE3A enzymes display consistent changes in their phosphorylation and an increased interaction with the 14-3-3θ adaptor protein. This aberrant signaling is associated with an increase in vascular smooth muscle cell proliferation and changes in vessel morphology and function. Conclusions: The mutated PDE3A gene drives mechanisms that increase peripheral vascular resistance causing hypertension. We present 2 new animal models that will serve to elucidate the underlying mechanisms further. Our findings could facilitate the search for new antihypertensive treatments.

Microscopic hematuria is a common feature of patients with Alport syndrome, a familial nephropathy due to mutations in COL4A3, COL4A4 or COL4A5. These genes encode for α3, α4, and α5 type IV collagen polypeptide chains (collagen IV α345), crucial for the structural component of the glomerular basement membrane. Even patients with mild phenotype, namely isolated microhematuria (X-linked females with thin basement membrane on electron microscopy or heterozygous carriers of COL4A3 or COL4A4 mutations), can potentially progress to proteinuria and to end-stage renal disease. Recent pedigree analyses provided evidence for digenic inheritance of Alport syndrome by concomitant mutations in COL4A3/COL4A4 or COL4A4/COL4A5. We describe a Caucasian family with concomitant COL4A3 and COL4A5 mutations, consisting of a novel c.4484A>G COL4A3 (p.Gln1495Arg) mutation and a previously reported c.1871G>A COL4A5 (p.Gly624Asp) mutation. Our segregation analysis raises the possibility that Alport syndrome resembles also digenic inheritance by COL4A3/COL4A5.

Mutations in the ROMK1 potassium channel gene (KCNJ1) cause antenatal/neonatal Bartter syndrome type II (aBS II), a renal disorder that begins in utero, accounting for the polyhydramnios and premature delivery that is typical in affected infants, who develop massive renal salt wasting, hypokalaemic metabolic alkalosis, secondary hyperreninaemic hyperaldosteronism, hypercalciuria and nephrocalcinosis. This BS type is believed to represent a disorder of the infancy, but not in adulthood. We herein describe a female patient with a remarkably late-onset and mild clinical manifestation of BS II with compound heterozygous KCNJ1 missense mutations, consisting of a novel c.197T > A (p.I66N) and a previously reported c.875G > A (p.R292Q) KCNJ1 mutation. We implemented and evaluated the performance of two different bioinformatics-based approaches of targeted massively parallel sequencing [next generation sequencing (NGS)] in defining the molecular diagnosis. Our results demonstrate that aBS II may be suspected in patients with a late-onset phenotype. Our experimental approach of NGS-based mutation screening combined with Sanger sequencing proved to be a reliable molecular approach for defining the clinical diagnosis in our patient, and results in important differential diagnostic and therapeutic implications for patients with BS. Our results could have a significant impact on the diagnosis and methodological approaches of genetic testing in other patients with clinical unclassified phenotypes of nephrocalcinosis and congenital renal electrolyte abnormalities.

Hypertension is the greatest driver of cardiovascular disease, the most common cause of death. Finding novel model mechanisms for blood-pressure (BP) regulation is an important goal. Autosomal-dominant hypertension with brachydactyly (HTNB) clinically resembles salt-resistant essential hypertension and causes death by stroke, commonly before age 50 years. We identified mutated phosphodiesterase 3A as responsible earlier; however, definitive mechanistic proof and animal models were lacking. We now describe new kindreds, one substituted within the earlier-described regulatory region (G449S), and another with a mutation causing an amino acid substitution within the PDE3A enzymatic pocket (R864C). Both exhibit the HTNB phenotype. Mutant PDE3A (T445N) overexpression in mouse vascular smooth muscle cells causes hypertension (mean telemetry BP 130/88 mm Hg vs littermate control 115/84 mm Hg, p

We present a 42-year-old man with a BMI of 32, who was referred because of proteinuria and decreased renal function. We were impressed by his markedly muscular physique. A renal biopsy was performed, which showed focal segmental glomerular sclerosis (FSGS). Is this patient merely an obese person with FSGS or is something else going on here? We performed extensive clinical and laboratory examinations, genetic testing, and anthropometric data monitoring over time. We transferred our methodology for routine FSGS mutation screening (Sanger sequencing) to the Ion Torrent PGM platform with a new custom-targeted NGS gene panel (Ion Ampliseq FSGS panel) and tested the performance of the system in two cohorts of patients with FSGS. We discuss FSGS in bodybuilders, including possible mechanisms, and review the literature.

Mutations in CD2-associated protein (CD2AP) have been identified in patients with focal segmental glomerulosclerosis (FSGS); however, reports of CD2AP mutations remain scarce. We performed Sanger sequencing in a patient with steroid-resistant FSGS and identified a heterozygous CD2AP mutation (p.T374A, c.1120 A > G). Our patient displayed mild cognitive decline, a phenotypic characteristic not previously associated with CD2AP-associated FSGS. His proteinuria was remarkably reduced by treatment with cyclosporine A. Our findings expand the genetic spectrum of CD2AP-associated disorders and broaden the associated phenotype with the co-occurrence of cognitive decline. Our case shows that cyclosporin A is a treatment option for CD2AP-associated nephropathy.