Stephan C. Reinalter

Clinical presentation of genetically defined patients with hypokalemic salt-losing tubulopathies

PURPOSE Hypokalemic salt-losing tubulopathies (Bartter-like syndromes) comprise a set of clinically and genetically distinct inherited renal disorders. Mutations in four renal membrane proteins involved in electrolyte reabsorption have been identified in these disorders: the furosemide-sensitive sodium-potassium-chloride cotransporter NKCC2, the potassium channel ROMK, the chloride channel ClC-Kb, and the thiazide-sensitive sodium-chloride cotransporter NCCT. The aim of this study was to characterize the clinical features associated with each mutation in a large cohort of genetically defined patients. PATIENTS AND METHODS The phenotypic characteristics of 65 patients with molecular defects in NKCC2, ROMK, ClC-Kb, or NCCT were collected retrospectively. RESULTS ROMK and NKCC2 patients presented with polyhydramnios, nephrocalcinosis, and hypo- or isosthenuria. Hypokalemia was less severe in the ROMK patients compared with the NKCC2 patients. In contrast, NCCT patients had hypocalciuria, hypomagnesemia, and marked hypokalemia. While this dissociation of renal calcium and magnesium handling was also observed in some ClC-Kb patients, a few ClC-Kb patients presented with hypercalciuria and hypo- or isosthenuria. CONCLUSIONS ROMK, NKCC2, and NCCT mutations usually have uniform clinical presentations, whereas mutations in ClC-Kb occasionally lead to phenotypic overlaps with the NCCT or, less commonly, with the ROMK/NKCC2 cohort. Based on these results, we propose an algorithm for the molecular diagnosis of hypokalemic salt-losing tubulopathies.

Impaired response to furosemide in hyperprostaglandin E syndrome: Evidence for a tubular defect in the loop of Henle

In hyperprostaglandin E syndrome (HPS) renal wasting of electrolytes and water is consistently associated with enhanced synthesis of prostaglandin E2. In contrast to Bartter or Gitelman syndrome (BS/GS), HPS is characterized by its severe prenatal manifestation, leading to fetal polyuria, development of polyhydramnios, and premature birth. This disorder mimics furosemide treatment with hypokalemic alkalosis, hypochloremia, isosthenuria, and impaired renal conservation of both calcium and magnesium. Therefore the thick ascending limb of the loop of Henle seems to be involved in HPS. To characterize the tubular defect we investigated the response to furosemide (2 mg/kg) in HPS (n = 8) and BS/GS (n = 3) 1 week after discontinuation of long-term indomethacin treatment. Sensitivity to furosemide was completely maintained in patients with BS/GS. The diuretic, saluretic, and hormonal responses were similar to those of a control group of healthy children (n = 13), indicating an intact function of the thick ascending limb of the loop of Henle in BS/GS. In contrast, patients with HPS had a marked resistance to this loop diuretic. Furosemide treatment increased urine output by 7.5 +/- 0.7 ml/kg per hour in healthy control subjects but only by 4.4 +/- 1.2 ml/kg per hour (p < 0.5) in children with HPS. In parallel, the latter also had a markedly impaired saluretic response (delta Cl(urine) 0.14 +/- 0.04 mmol/kg per hour vs 0.85 +/- 0.09 mmol/kg per hour, p < 0.001; delta Na(urine) 0.23 +/- 0.06 mmol/kg per hour vs 0.77 +/- 0.09 mmol/kg per hour, p < 0.001). Furosemide therapy further enhanced prostaglandin E2 excretion in patients with HPS (54 +/- 17 to 107 +/- 28 ng/hr per 1.73 m2, p < 0.05), whereas no significant effect was observed in healthy children (20 +/- 3 to 12 +/- 3 ng/hr per 1.73 m2). We conclude that a defect of electrolyte reabsorption in the thick ascending limb of the loop of Henle plays a major role in HPS.

Pathogenetic role of cyclooxygenase‐2 in hyperprostaglandin E syndrome/antenatal bartter syndrome: Therapeutic use of the cyclooxygenase‐2 inhibitor nimesulide

Patients with hyperprostaglandin E syndrome/antenatal Bartter syndrome typically have renal salt wasting, hypercalciuria with nephrocalcinosis, and secondary hyperaldosteronism. Antenatally, these patients have fetal polyuria, leading to polyhydramnios and premature birth. Hyperprostaglandin E syndrome/antenatal Bartter syndrome is accompanied by a pathologically elevated synthesis of prostaglandin E2, thought to be responsible for aggravation of clinical symptoms such as salt and water loss, vomiting, diarrhea, and failure to thrive. In this study administration of the cyclooxygenase‐2 (COX‐2) specific inhibitor nimesulide to patients with hyperprostaglandin E syndrome/antenatal Bartter syndrome blocked renal prostaglandin E2 formation and relieved the key parameters hyperprostaglandinuria, secondary hyperaldosteronism, and hypercalciuria. Partial suppression of serum thromboxane B2 synthesis resulting from platelet COX‐1 activity and complete inhibition of urinary 6‐keto‐prostaglandin F1α, reflecting endothelial COX‐2 activity, indicate preferential inhibition of COX‐2 by nimesulide. Amelioration of the clinical symptoms by use of nimesulide indicates that COX‐2 may play an important pathogenetic role in hyperprostaglandin E syndrome/antenatal Bartter syndrome. Moreover, on the basis of our data we postulate that COX‐2‐derived prostaglandin E2 is an important mediator for stimulation of the renin‐angiotensin‐aldosterone system in the kidney.

Late-Onset Manifestation of Antenatal Bartter Syndrome as a Result of Residual Function of the Mutated Renal Na+-K+-2Cl− Co-Transporter

Genetic defects of the Na+-K+-2Cl- (NKCC2) sodium potassium chloride co-transporter result in severe, prenatal-onset renal salt wasting accompanied by polyhydramnios, prematurity, and life-threatening hypovolemia of the neonate (antenatal Bartter syndrome or hyperprostaglandin E syndrome). Herein are described two brothers who presented with hyperuricemia, mild metabolic alkalosis, low serum potassium levels, and bilateral medullary nephrocalcinosis at the ages of 13 and 15 yr. Impaired function of sodium chloride reabsorption along the thick ascending limb of Henles loop was deduced from a reduced increase in diuresis and urinary chloride excretion upon application of furosemide. Molecular genetic analysis revealed that the brothers were compound heterozygotes for mutations in the SLC12A1 gene coding for the NKCC2 co-transporter. Functional analysis of the mutated rat NKCC2 protein by tracer-flux assays after heterologous expression in Xenopus oocytes revealed significant residual transport activity of the NKCC2 p.F177Y mutant construct in contrast to no activity of the NKCC2-D918fs frameshift mutant construct. However, coexpression of the two mutants was not significantly different from that of NKCC2-F177Y alone or wild type. Membrane expression of NKCC2-F177Y as determined by luminometric surface quantification was not significantly different from wild-type protein, pointing to an intrinsic partial transport defect caused by the p.F177Y mutation. The partial function of NKCC2-F177Y, which is not negatively affected by NKCC2-D918fs, therefore explains a mild and late-onset phenotype and for the first time establishes a mild phenotype-associated SLC12A1 gene mutation.

Induction of microsomal prostaglandin E2 synthase in the macula densa in children with hypokalemic salt-losing tubulopathies

In hyperprostaglandin E syndrome (HPGES) and classic Bartter syndrome (cBS), tubular salt and water losses stimulate renin secretion, which is dependent on enhanced cyclooxygenase-2 (COX-2) enzymatic activity. In contrast to other renal COX metabolites, only prostaglandin E2 (PGE2) is selectively up-regulated in these patients. To determine the intrarenal source of PGE2 synthesis, we analyzed the expression of microsomal PGE2 synthase (mPGES; EC: 5.3.99.3), whose product PGE2 has been shown to stimulate renin secretion in vitro. Expression of mPGES was analyzed by immunohistochemistry in eight patients with HPGES, in two patients with cBS, and in six control subjects. Expression of mPGES immunoreactive protein was observed in cells of the macula densa in five of eight HPGES patients and in one of two cBS patients. Expression of mPGES immunoreactive protein was not observed in cells associated with the macula densa in kidneys from control subjects without a history consistent with activation of the renin angiotensin system. Co-induction of COX-2 and mPGES in cells of the macula densa suggests that PGE2 activates renin secretion in humans.

RESISTANCE TO FUROSEMIDE IN HYPERPROSTAGLANDIN E SYNDROME: EVIDENCE FOR A PROSTAGLANDIN-INDEPENDENT DEFECT IN THE LOOP OF HENLE. |[dagger]| 2162

Hyperprostaglandin E syndrome (HPS), a prenatal variant of Bartters syndrome, is characterized by impaired renal salt conservation and fetal polyuria leading to poly-hydramnios and premature birth. Moreover HPS mimics chronic furosemide treatment with isosthenuria, hypochloremia, hypercalciuria, hypokalemic alkalosis, and hyperaldosteronism. To date inadequate formation of PGE2 is considered to be a primary event in HPS. In a group of 9 children with HPS (median age 10.3, range 7.7 to 11.7 years) long-term indomethacin treatment sufficiently suppressed urinary excretion of PGE2 (54±18 to 5±2 ng/h/1.73m2) and corrected hyperreninemia, hyperaldosteronism and hyperkaliuria. However, renal loss of sodium chloride persisted (2.8±0.4 to 3.3±0.3 and 3.2±0.2 to 3.0±0.4 mmol/kg/d, resp.) and the children still remained isosthenuric (227±18 to 270±18 mosmol/kg). In addition polyuria, hypercalciuria and hypermagnesiuria were improved only partially. These observations indicate a tubular defect independent of renal PGE2 release. To study the capacity of electrolyte reabsorption in the thick ascending limb of Henles loop (TALH), furosemide was administered to the patients 7 days after interruption of indomethacin treatment. Compared to a group of 13 healthy children (median age 10.2, range 6.4 to 15.1 years) patients with HPS presented a marked resistance to the loop diuretic. Data were obtained from 3-hour urine collections subsequent to a single oral dose of 2 mg/kg furosemide (means ± SEM; *p<0.05) We conclude that a prostaglandin-independent defect of furosemide-sensitive salt reabsorption in the TALH plays a major pathophysiological role in renal dysfunction of HPS.Table