Nephron Physiology最新文献

Background: The cilium and cilial proteins have emerged as principal mechanosensors of renal epithelial cells responsible for translating mechanical forces into intracellular signals. Polycystin-2 (PC-2), a cilial protein, regulates flow/shear-induced changes in intracellular Ca(2+) ([Ca(2+)](i)) and recently has been implicated in the regulation of mitogen-activated protein (MAP) kinases. We hypothesize that fluid shear stress (FSS) activates PC-2 which regulates MAP kinase and, in turn, induces MAP kinase-dependent gene expression, specifically, monocyte chemoattractant protein-1 (MCP-1).

Methods: To test this, PC-2 expression was constitutively reduced in a murine inner medullary collecting duct (IMCD3) cell line, and the expression of FSS-induced MCP-1 expression and MAP kinase signaling compared between the parental (PC-2-expressing) and PC-2-deficient IMCD3 cells.

Results: FSS induces MAP kinase signaling and downstream MCP-1 mRNA expression in wild-type IMCD3 cells, while inhibitors of MAP kinase prevented the FSS-induced MCP-1 mRNA response. In contradistinction, FSS did not induce MCP-1 mRNA expression in PC-2-deficient cells, but did increase activation of the upstream MAP kinases. Wild-type cells exposed to FSS augmented the nuclear abundance of activated MAP kinase while PC-2-deficient cells did not.

Conclusions: PC-2 regulates FSS-induced MAP kinase trafficking into the nucleus of CD cells.

Background: It has been reported that mutations in WNK1 and WNK4 cause pseudohypoaldosteronism type II (PHA2), an autosomal dominant renal disease. WNK kinase proteins are expressed in the kidney and regulate ion transport including the thiazide-sensitive sodium chloride cotransporter (NCC). In this report, we screened 4 Chinese PHA2 pedigrees for WNK4 mutations, identified a novel mutation, and studied its effects on NCC protein trafficking in vitro.

Methods: The patients' genomic DNA was extracted from peripheral leukocytes. Sequence analysis was performed by PCR amplification of the 19 exons of WNK4. The wild-type or mutant WNK4 was coexpressed with NCC in HEK293 cells. We measured the effect of wild-type WNK4 compared to the mutant WNK4 on NCC protein surface expression.

Results: A novel missense mutation in WNK4, K1169E, was identified in 1 of the 4 pedigrees. Analysis of confocal images showed that K1169E lost its inhibitory effect on NCC surface expression compared to wild-type WNK4 when expressed in HEK293 cells, while it did not change NCC total protein expression.

Conclusions: We identified an unreported disease-causing WNK4 missense mutation, K1169E, in 1 Chinese PHA2 pedigree. This mutation appears to be a 'loss of function' of NCC inhibition and suggests that some important functional roles lie in the 2nd coiled-coil domain of WNK4.

Background/aims: Mutations in the inwardly-rectifying K+ channel KCNJ10/Kir4.1 cause an autosomal recessive disorder characterized by epilepsy, ataxia, sensorineural deafness and tubulopathy (EAST syndrome). KCNJ10 is expressed in the kidney distal convoluted tubule, cochlear stria vascularis and brain glial cells. Patients clinically diagnosed with EAST syndrome were genotyped to identify and study mutations in KCNJ10.

Methods: Patient DNA was sequenced and new mutations identified. Mutant and wild-type KCNJ10 constructs were cloned and heterologously expressed in Xenopus oocytes. Whole-cell K+ currents were measured by two-electrode voltage clamping.

Results: Three new mutations in KCNJ10 (p.R65C, p.F75L and p.V259fs259X) were identified, and mutation p.R297C, previously only seen in a compound heterozygous patient, was found in a homozygous state. Wild-type human KCNJ10-expressing oocytes showed strongly inwardly-rectified currents, which by comparison were significantly reduced in all the mutants (p < 0.001). Specific inhibition of KCNJ10 currents by Ba2+ demonstrated residual function in all mutant channels (p < 0.05) but V259X.

Conclusion: This study confirms that EAST syndrome can be caused by many different mutations in KCNJ10 that significantly reduce K+ conductance. EAST syndrome should be considered in any patient with a renal Gitelman-like phenotype with additional neurological signs and symptoms like ataxia, epilepsy or sensorineural deafness.

Background and aims: The presence of altered plasma Na concentration (PNa) allows calculations of changes in water and electrolyte contents, which are not feasible during normonatremic derangements. We have developed a computational algorithm whereby the changes in solute (ΔNa and ΔCl) and solvent (ΔV) contents can be computed exactly when Na is lost entirely as NaCl (or NaHCO(3)) and nearly exactly in all other circumstances, except when the losses of Na and Cl occur in the same proportions as those of the normal plasma concentration of these ions.

Methods: In computer experiments, we simulated different fluid depletions containing 140 mEq/l of Na (which is to say, ΔNa/ΔV ≈ 140), coupled with variable ratios in Na to Cl losses (variable ΔNa/ΔCl). The data were back-calculated with our algorithms from the ensuing plasma ion concentrations (PNa(1), PCl(1) and POAN(1), where subscript (0) and (1) indicate normal and deranged plasma concentration values, respectively, and OAN indicates anions other than Cl), as if they had been measured on patients, and from known normal values (TBW(0), ECV(0), Na(0)). These were compared to the true values used to build the simulations. This procedure was reproduced in 17 patients suffering from iso-osmolar dehydration, where true data were obtained by balance studies.

Results: True and calculated data were compared with linear regression analysis. We obtained significant correlations both in computer-simulated and real patients (R(2) = 0.83, p < 0.005 and R(2) = 0.63, p < 0.05, respectively).

Conclusion: This new math model and its related computational method are useful in the correct evaluation and treatment of iso-osmolar dehydration.

Background/aims: Renal stone disease may be seen as a clinical symptom of an underlying pathological process predisposing to crystallization within the renal tract. Renal stones may be comprised of calcium salts, uric acid, cystine and various other insoluble complexes. Nephrolithiasis may be the manifestation of rare single gene disorders or part of more common idiopathic renal stone-forming diseases.

Methods and results: Molecular genetics has allowed significant progress to be made in our understanding of certain stone-forming conditions. The molecular defect underlying single gene disorders often contributes to a significant metabolic risk factor for stone formation. In contrast, idiopathic renal stone formation relates to the interplay of environmental, dietary and genetic factors, with hypercalciuria being the most commonly found metabolic risk factor. Candidate genes for idiopathic stone formers have been identified using numerous approaches, some of which are outlined here. Despite this, the genetic basis underlying familial hypercalciuria and calcium stone formation remains elusive. The molecular basis of other metabolic risk factors such as hyperuricosuria, hyperoxaluria and hypocitraturia is being unraveled and is allowing new insights into renal stone pathogenesis.

Conclusion: The discovery of both rare and common molecular defects leading to renal stones will hopefully increase our understanding of the disease pathogenesis. Such knowledge will allow screening for genetic defects and the use of specific drug therapies in order to prevent renal stone formation.