Renin-aldosterone system evaluation over four decades in an extended family with autosomal dominant pseudohypoaldosteronism due to a deletion in the NR3C2 gene.

Renal pseudohypoaldosteronism (PHA1) is a mild form of an aldosterone-resistance syndrome caused by mutations in the NR3C2 gene that codes for the mineralocorticoid receptor (MR). The disease is inherited as an autosomal dominant trait characterized by signs and symptoms of salt-losing in infancy. Disease manifestations could be severe in infancy but improve after the age of 1-3 years. Some affected members are asymptomatic and remain so life-long. In this study, we report the identification of a large deletion in the NR3C2 gene (c.1897+1_1898-1)_(c.*2955+?)del in renal PHA1 patients from an extended family spanning four generations. We prospectively evaluated the plasma renin activity and serum aldosterone profiles over four decades in symptomatic and asymptomatic affected family members. The benefits of early diagnosis on the clinical outcome were assessed as well. The long-term follow-up showed an age-dependent decrease in both plasma renin activity and serum aldosterone levels over the years. However, aldosterone levels remain high life-long. Thus, levels of aldosterone are a reliable marker to detect asymptomatic family members. The diagnosis of the proposita led to early diagnosis and therapy in other affected family members, significantly mitigating the clinical course. Despite the extremely elevated serum aldosterone levels during pregnancy, affected pregnant women did not experience any ill effects. However, this should be verified by observations in other adult patients.

Aldosterone deficiency with a hormone profile mimicking pseudohypoaldosteronism.

Background Aldosterone deficiency (hypoaldosteronism) or aldosterone resistance (pseudohypoaldosteronism) both result in defective aldosterone activity. Case presentation A 42-day-old man presented with failure to thrive, hyponatremia, high urine sodium output, severe hyperkalemia and high plasma renin activity and aldosterone levels. NR3C2, SCNN1A, B and G sequencing showed no variants. Exclusive sodium supplementation resulted in clinical stabilization and growth normalization. His younger sibling had similar clinical and laboratory features, except for low-normal aldosterone. Both patients showed compound heterozygous mutations in CYP11B2 (c.C554T/2802pbE1-E2del). The younger patient needed transient fludrocortisone treatment and higher sodium supplementation, recuperating his weight and a normal growth velocity, although below his brother's and target height (c.10th vs. c.50th). Conclusions On a suggestive clinical picture, high aldosterone plasma levels in early infancy do not rule out aldosterone insufficiency and might mislead differential diagnosis with pseudohypoaldosteronism. Therapeutic requests and growth impairment in hypoaldosteronism vary even with a common genetic background.

A novel SCNN1G mutation in a PHA I infant patient correlates with nephropathy.

Systematic form of pseudohypoaldosteronism Type I (PHA I) is a rare recessive homozygous inherited syndrome characterized by severe salt loss, hyperkalemia, hyponatremia, metabolic acidosis, hyperaldosteronism and hyperreninemia. It is caused by mutations in one of the genes encoding the α, ß and γ subunits of epithelial sodium channels (ENaC). In this study, we performed whole exome sequencing on an infant patient with PHA I as well as nephropathy. The result presented a novel homozygous six-base deletion in the γ subunit encoding gene SCNN1G. Then we correlated the mutant to kidney damage, along with transcriptional alterations of genes involved in inflammation, oxidative stress and apoptosis, via in vitro and in vivo tests. In addition, it was demonstrated that the SCNN1G defects triggered programmed cell death via inhibiting miR-21 and upregulating PTEN, which then orchestrated the key downstream regulators, including Bcl2, Bax2, and cleaved Caspse-3 in a way that favors cell apoptosis. The study enhances our understanding of the pathophysiology of the disorder of PHA I and the mechanisms of renal damage induced by the novel defect.

Plasma Potassium Determines NCC Abundance in Adult Kidney-Specific γENaC Knockout.

The amiloride-sensitive epithelial sodium channel (ENaC) and the thiazide-sensitive sodium chloride cotransporter (NCC) are key regulators of sodium and potassium and colocalize in the late distal convoluted tubule of the kidney. Loss of the αENaC subunit leads to a perinatal lethal phenotype characterized by sodium loss and hyperkalemia resembling the human syndrome pseudohypoaldosteronism type 1 (PHA-I). In adulthood, inducible nephron-specific deletion of αENaC in mice mimics the lethal phenotype observed in neonates, and as in humans, this phenotype is prevented by a high sodium (HNa+)/low potassium (LK+) rescue diet. Rescue reflects activation of NCC, which is suppressed at baseline by elevated plasma potassium concentration. In this study, we investigated the role of the γENaC subunit in the PHA-I phenotype. Nephron-specific γENaC knockout mice also presented with salt-wasting syndrome and severe hyperkalemia. Unlike mice lacking αENaC or ßΕΝaC, an HNa+/LK+ diet did not normalize plasma potassium (K+) concentration or increase NCC activation. However, when K+ was eliminated from the diet at the time that γENaC was deleted, plasma K+ concentration and NCC activity remained normal, and progressive weight loss was prevented. Loss of the late distal convoluted tubule, as well as overall reduced ßENaC subunit expression, may be responsible for the more severe hyperkalemia. We conclude that plasma K+ concentration becomes the determining and limiting factor in regulating NCC activity, regardless of Na+ balance in γENaC-deficient mice.

High aldosterone and cortisol levels in salt wasting congenital adrenal hyperplasia: a clinical conundrum.

BACKGROUND: Salt wasting syndrome (hyponatremia, hyperkalemia, dehydration, metabolic acidosis) in early infancy could be caused by either mineralocorticoid deficiency as in congenital adrenal hyperplasia (CAH) and adrenal insufficiency or mineralocorticoid resistance as in pseudohypoaldosteronism (PHA). In salt wasting CAH, serum aldosterone and cortisol levels are expected to be low. Cross reactivity between high levels of adrenal steroid precursors and aldosterone has recently been reported resulting in elevated aldosterone levels in CAH, leading to difficulty in differentiating between CAH and PHA. CASE PRESENTATION: We report four such cases of salt wasting CAH, where high aldosterone levels and high normal cortisol levels led to initial diagnostic confusion with PHA. Diagnosis of CAH was later established on the basis of significantly elevated adrenocorticotropic hormone (ACTH) stimulated 17-hydroxyprogesterone (17-OHP) values. CONCLUSIONS: By reporting these cases we draw attention to the possibility that high levels of adrenal steroid precursors can cross react with aldosterone and cortisol, and underscore the significance of ACTH stimulated 17-OHP values in differentiating CAH and PHA.

Transient pseudohypoaldosteronism in infancy secondary to urinary tract infection.

AIM: Hyponatraemia with hyperkalaemia in infancy is a typical presentation of congenital adrenal hyperplasia. In the presence of pyelonephritis, the same biochemical picture can occur with transient type 1 pseudohypoaldosteronism (PHA-1) also termed type 4 renal tubular acidosis. Recognition of PHA-1 enables appropriate management thus avoiding unnecessary investigations and treatment. To improve awareness of this condition, we present a case series to highlight the clinical and biochemical features of PHA-1. METHODS: A retrospective chart review of patients diagnosed with transient PHA-1 at a tertiary children's hospital in Western Australia was conducted. RESULTS: Five male infants (32 days to 6 months) with transient PHA-1 were identified. Failure to thrive was the most common symptom with hyponatraemia on presentation. Two infants had antenatally diagnosed bilateral hydronephrosis and urinary tract infection (UTI) on admission. Two infants were treated for congenital adrenal hyperplasia and received hydrocortisone. All infants had UTI and required parenteral antibiotics. The condition was transient and hyponatraemia corrected by day 4 in all infants. There was no correlation between plasma sodium and aldosterone levels. The severity of PHA-1 was independent of the underlying renal anomaly. Four infants had hydronephrosis and vesicoureteric reflux. Surgical intervention was required in two infants. CONCLUSIONS: PHA-1 may be precipitated by UTI or urinary tract anomalies in early infancy. Urine analysis should be performed in infants with hyponatraemia. Diagnosis of PHA-1 facilitates appropriate renal investigations to reduce long-term morbidity.

KLHL3 Knockout Mice Reveal the Physiological Role of KLHL3 and the Pathophysiology of Pseudohypoaldosteronism Type II Caused by Mutant KLHL3.

Mutations in the with-no-lysine kinase 1 (WNK1), WNK4, kelch-like 3 (KLHL3), and cullin3 (CUL3) genes are known to cause the hereditary disease pseudohypoaldosteronism type II (PHAII). It was recently demonstrated that this results from the defective degradation of WNK1 and WNK4 by the KLHL3/CUL3 ubiquitin ligase complex. However, the other physiological in vivo roles of KLHL3 remain unclear. Therefore, here we generated KLHL3-/- mice that expressed ß-galactosidase (ß-Gal) under the control of the endogenous KLHL3 promoter. Immunoblots of ß-Gal and LacZ staining revealed that KLHL3 was expressed in some organs, such as brain. However, the expression levels of WNK kinases were not increased in any of these organs other than the kidney, where WNK1 and WNK4 increased in KLHL3-/- mice but not in KLHL3+/- mice. KLHL3-/- mice also showed PHAII-like phenotypes, whereas KLHL3+/- mice did not. This clearly demonstrates that the heterozygous deletion of KLHL3 was not sufficient to cause PHAII, indicating that autosomal dominant type PHAII is caused by the dominant negative effect of mutant KLHL3. We further demonstrated that the dimerization of KLHL3 can explain this dominant negative effect. These findings could help us to further understand the physiological roles of KLHL3 and the pathophysiology of PHAII caused by mutant KLHL3.

ROMK expression remains unaltered in a mouse model of familial hyperkalemic hypertension caused by the CUL3Δ403-459 mutation.

Familial hyperkalemic hypertension (FHHt) is a rare inherited form of salt-dependent hypertension caused by mutations in proteins that regulate the renal Na(+)-Cl(-) cotransporter NCC Mutations in four genes have been reported to cause FHHt including CUL3 (Cullin3) that encodes a component of a RING E3 ligase. Cullin-3 binds to WNK kinase-bound KLHL3 (the substrate recognition subunit of the ubiquitin ligase complex) to promote ubiquitination and proteasomal degradation of WNK kinases. Deletion of exon 9 from CUL3 (affecting residues 403-459, CUL3(Δ403-459)) causes a severe form of FHHt (PHA2E) that is recapitulated closely in a knock-in mouse model. The loss of functionality of CUL3(Δ403-459) and secondary accumulation of WNK kinases causes substantial NCC activation. This accounts for the hypertension in FHHt but the origin of the hyperkalemia is less clear. Hence, we explored the impact of CUL3(Δ403-459) on expression of the distal secretory K channel, ROMK, both in vitro and in vivo. We found that expressing wild-type but not the CUL3(Δ403-459) mutant form of CUL3 prevented the suppression of ROMK currents by WNK4 expressed in Xenopus oocytes. The mutant CUL3 protein was also unable to affect ROMK-EGFP protein expression at the surface of mouse M-1 cortical collecting duct (CCD) cells. The effects of CUL3 on ROMK expression in both oocytes and M-1 CCD cells was reduced by addition of the neddylation inhibitor, MLN4924. This confirms that neddylation is important for CUL3 activity. Nevertheless, in our knock-in mouse model expressing CUL3(Δ403-459) we could not show any alteration in ROMK expression by either western blotting whole kidney lysates or confocal microscopy of kidney sections. This suggests that the hyperkalemia in our knock-in mouse and human PHA2E subjects with the CUL3(Δ403-459) mutation is not caused by reduced ROMK expression in the distal nephron.

Ocular and skin manifestations in systemic pseudohypoaldosteronism.

Pseudohypoaldosteronism type-1 is a rare disorder characterised by end-organ resistance to aldosterone resulting in salt-losing crisis with hyponatraemic dehydration, hyperkalaemia and metabolic acidosis. We report two siblings with pseudohypoaldosteronism type-1 who presented early in neonatal period with hyponatraemia, severe hyperkalaemia and metabolic acidosis. Both babies had miliaria like skin rash which flared up during episodes of hyperkalaemia and hyponatraemia. They had visible dilated meibomian glands from which a white material was protruding. The clinical presentation of pseudohypoaldosteronism type-1 mimics congenital adrenal hyperplasia. As there is often a delay in obtaining hormonal assay results, the eye and skin manifestations may give an important diagnostic clue which in turn will influence management.

Pseudohypoaldosteronism type 1: management issues.

We report a newborn girl with life-threatening hyperkalemia and salt wasting crisis due to severe autosomal recessive multiple target organ dysfunction pseudohypoaldosteronism type 1 (MTOD PHA1). She was aggressively managed with intravenous fluids, potassium-lowering agents, high-dose sodium chloride supplementation and peritoneal dialysis. Genetic analysis revealed a homozygous mutation of the α- ENaC (epithelial Na(+) channel) gene. She had a stormy clinical course with refractory hyperkalemia and prolonged hospitalization. Eventually, she succumbed to pneumonia and septicemia at 4 months of age. This is probably the first case of PHA1 confirmed by genetic analysis from India.

An inducible transgenic mouse model for familial hypertension with hyperkalaemia (Gordon's syndrome or pseudohypoaldosteronism type II).

Mutations in the novel serine/threonine WNK [With No lysine (=K)] kinases WNK1 and WNK4 cause PHAII (pseudohypoaldosteronism type II or Gordon's syndrome), a rare monogenic syndrome which causes hypertension and hyperkalaemia on a background of a normal glomerular filtration rate. Current animal models for PHAII recapitulate some aspects of the disease phenotype, but give no clues to how rapidly the phenotype emerges or whether it is reversible. To this end we have created an inducible PHAII transgenic animal model that expresses a human disease-causing WNK4 mutation, WNK4 Q565E, under the control of the Tet-On system. Several PHAII inducible transgenic mouse lines were created, each with differing TG (transgene) copy numbers and displaying varying degrees of TG expression (low, medium and high). Each of these transgenic lines demonstrated similar elevations of BP (blood pressure) and plasma potassium after 4 weeks of TG induction. Withdrawal of doxycycline switched off mutant TG expression and the disappearance of the PHAII phenotype. Western blotting of microdissected kidney nephron segments confirmed that expression of the thiazide-sensitive NCC (Na⁺-Cl⁻ co-transporter) was increased, as expected, in the distal convoluted tubule when transgenic mice were induced with doxycycline. The kidneys of these mice also do not show the morphological changes seen in the previous transgenic model expressing the same mutant form of WNK4. This inducible model shows, for the first time, that in vivo expression of a mutant WNK4 protein is sufficient to cause the rapid and reversible appearance of a PHAII disease phenotype in mice.

Neonatal renal venous thrombosis followed by secondary pseudohypoaldosteronism.

A male infant was diagnosed as having renal venous thrombosis (RVT) in association with bilateral flank masses, macroscopic hematuria, and thrombocytopenia. In the course of supportive treatment, hyponatremia, hyperkalemia, and metabolic acidosis became prominent. Plasma renin activity (PRA) and aldosterone increased markedly. Treatment with sufficient sodium chloride and sodium bicarbonate intake was effective. It is important to note that tubular damage by RVT causes secondary pseudohypoaldosteronism.

Structural chromosome disruption of the NR3C2 gene causing pseudohypoaldosteronism type 1 presenting in infancy.

Type I pseudohypoaldosteronism (PHA1) is a rare form of mineralocorticoid resistance presenting in infancy with renal salt wasting and failure to thrive. Here, we present the case of a 6-week-old baby girl who presented with mild hyponatraemia and dehydration with a background of severe failure to thrive. At presentation, urinary sodium was not measurably increased, but plasma aldosterone and renin were increased, and continued to rise during the subsequent week. Despite high calorie feeds the infant weight gain and hyponatraemia did not improve until salt supplements were commenced. Subsequently, the karyotype was reported as 46,XX,inv (4)(q31.2q35). A search of the OMIM database for related genes at or near the inversion breakpoints, showed that the mineralocorticoid receptor gene (NR3C2) at 4q31.23 was a likely candidate. Further FISH analysis showed findings consistent with disruption of the NR3C2 gene by the proximal breakpoint (4q31.23) of the inversion. There was no evidence of deletion or duplication at or near the breakpoint. This is the first report of a structural chromosome disruption of the NR3C2 gene giving rise to the classical clinical manifestations of pseudohypoaldosteronism type 1 in an infant.

Hook effect: a pitfall leading to misdiagnosis of hypoaldosteronism in an infant with pseudohypoaldosteronism.

We report herein the case of a premature infant who presented with failure to thrive, hyponatremia, hyperkalemia and metabolic acidosis. Initial serum hormone profiling suggested isolated hypoaldosteronism (aldosterone: 0.01 pg/ml, normal range: 50-900 pg/ml). A gas chromatography-mass spectrometry spot urinary steroid profile showed grossly elevated levels of 18-hydroxy-tetrahydro-11-dehydrocorticosterone (18-hydroxy-THA: 5,893 microg/l; normal upper limit 36 microg/l) and tetrahydroaldosterone (TH-Aldo: 5,749 microg/l; normal upper limit 36 microg/l) which are aldosterone precursor metabolite and aldosterone metabolite, respectively. Thus, aldosterone synthase deficiency was excluded and pseudohypoaldosteronism (PHA) was suggested. A repeated test after dilution of the serum revealed a very high level of aldosterone (6,490 pg/ml), confirming the diagnosis of PHA in this case.

Aerococcus viridans urinary tract infection in a pediatric patient with secondary pseudohypoaldosteronism.

Aerococcus viridans is a catalase-negative gram-positive bacterium rarely found as human pathogen. Some cases of urinary tract infection (UTI) have been described in immunocompromised adults. In this article we describe a UTI case caused by this agent in a child with severe obstructive uropathy, clinically presented with secondary pseudohypoaldosteronism (SPHA). Although A. viridans is rarely associated with child infection, it can be responsible for life threatening conditions/ situations. To our knowledge, A. viridans UTI has never been reported in pediatric patients.