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Nine V A M Knoers

Publications and source records attributed to Nine V A M Knoers.

At least 19 recordsLinked to original sources
PubMed2006

Gitelman syndrome.

In the past decade our understanding of the etiology and pathophysiology of Gitelman syndrome, an autosomal recessive salt-losing tubular disorder with secondary hypokalemia, has increased considerably through the achievements of molecular genetics and cell physiology. In this short review, I will summarize the most recent data on the clinical and biochemical phenotype, the molecular causes, and the pathogenesis of Gitelman syndrome. I will especially focus on the recent elucidation of the mechanisms involved in the pathogenesis of the hypomagnesemia and hypocalciuria that accompanies Gitelman syndrome.

Animals↗
PubMed2006

Cell biological aspects of the vasopressin type-2 receptor and aquaporin 2 water channel in nephrogenic diabetes insipidus.

In the renal collecting duct, water reabsorption is regulated by the antidiuretic hormone vasopressin (AVP). Binding of this hormone to the vasopressin V2 receptor (V2R) leads to insertion of aquaporin-2 (AQP2) water channels in the apical membrane, thereby allowing water reabsorption from the pro-urine to the interstitium. The disorder nephrogenic diabetes insipidus (NDI) is characterized by the kidney's inability to concentrate pro-urine in response to AVP, which is mostly acquired due to electrolyte disturbances or lithium therapy. Alternatively, NDI is inherited in an X-linked or autosomal fashion due to mutations in the genes encoding V2R or AQP2, respectively. This review describes the current knowledge of the cell biological causes of NDI and how these defects may explain the patients' phenotypes. Also, the increased understanding of these cellular defects in NDI has opened exciting initiatives in the development of novel therapies for NDI, which are extensively discussed in this review.

Amino Acid Sequence↗
PubMed2005

Teaching molecular genetics: Chapter 1--Background principles and methods of molecular biology.

In this first chapter of the series "Teaching molecular genetics," an introduction to molecular genetics is presented. We describe the structure of DNA and genes and explain in detail the central dogma of molecular biology, that is, the flow of genetic information from DNA via RNA to polypeptide (protein). In addition, several basic and frequently used general molecular tools, such as restriction enzymes, Southern blotting, DNA amplification and sequencing are discussed, in order to lay the foundations for the forthcoming chapters.

Genetic Phenomena↗
PubMed2005

Genotype-phenotype studies in nail-patella syndrome show that LMX1B mutation location is involved in the risk of developing nephropathy.

Nail-patella syndrome (NPS) is characterized by developmental defects of dorsal limb structures, nephropathy, and glaucoma and is caused by heterozygous mutations in the LIM homeodomain transcription factor LMX1B. In order to identify possible genotype-phenotype correlations, we performed LMX1B mutation analysis and comprehensive investigations of limb, renal, ocular, and audiological characteristics in 106 subjects from 32 NPS families. Remarkable phenotypic variability at the individual, intrafamilial, and interfamilial level was observed for different NPS manifestations. Quantitative urinanalysis revealed proteinuria in 21.3% of individuals. Microalbuminuria was detected in 21.7% of subjects without overt proteinuria. Interestingly, nephropathy appeared significantly more frequent in females. A significant association was established between the presence of clinically relevant renal involvement in an NPS patient and a positive family history of nephropathy. We identified normal-tension glaucoma (NTG) and sensorineural hearing impairment as new symptoms associated with NPS. Sequencing of LMX1B revealed 18 different mutations, including six novel variants, in 28 families. Individuals with an LMX1B mutation located in the homeodomain showed significantly more frequent and higher values of proteinuria compared to subjects carrying mutations in the LIM domains. No clear genotype-phenotype association was apparent for extrarenal manifestations. This is the first study indicating that family history of nephropathy and mutation location might be important in precipitating individual risks for developing NPS renal disease. We suggest that the NPS phenotype is broader than previously described and that NTG and hearing impairment are part of NPS. Further studies on modifier factors are needed to understand the mechanisms underlying phenotypic heterogeneity.

Adolescent↗
PubMed2005

A unique case of PHACES syndrome confirming the assumption that PHACES syndrome and the sternal malformation-vascular dysplasia association are part of the same spectrum of malformations.

PHACES is an acronym for a rare neurocutaneous disorder including the following features: posterior fossa brain malformations; facial hemangiomas; arterial, cardiac, and eye anomalies; and sternal clefts and/or supraumbilical abdominal raphe. Facial hemangiomas are the hallmark of this syndrome. We report a patient with facial hemangiomas, sternal clefting and carotid abnormalities. She is another example of the combined existence of sternal clefting and carotid abnormalities in PHACES syndrome and even more unique since she does not have an aortic abnormality. This report emphasizes that many different combinations of features may be seen in PHACES syndrome.

Abnormalities, Multiple↗
PubMed2004

Mutations in the human TBX4 gene cause small patella syndrome.

Small patella syndrome (SPS) is an autosomal-dominant skeletal dysplasia characterized by patellar aplasia or hypoplasia and by anomalies of the pelvis and feet, including disrupted ossification of the ischia and inferior pubic rami. We identified an SPS critical region of 5.6 cM on chromosome 17q22 by haplotype analysis. Putative loss-of-function mutations were found in a positional gene encoding T-box protein 4 (TBX4) in six families with SPS. TBX4 encodes a transcription factor with a strongly conserved DNA-binding T-box domain that is known to play a crucial role in lower limb development in chickens and mice. The present identification of heterozygous TBX4 mutations in SPS patients, together with the similar skeletal phenotype of animals lacking Tbx4, establish the importance of TBX4 in the developmental pathways of the lower limbs and the pelvis in humans.

Adolescent↗
PubMed2004

Peripheral nerve involvement in spinocerebellar ataxias.

BACKGROUND: In autosomal dominant cerebellar ataxias (ADCAs), it is unclear whether the associated peripheral nerve involvement is always a typical length-dependent axonopathy rather than primary neuronopathy due to neuronal degeneration in the spinal anterior horns and/or dorsal root ganglia. OBJECTIVE: To study the nature and extent of peripheral nerve involvement in patients with ADCA. PATIENTS AND METHODS: Standardized clinical and electrophysiologic studies of 27 genotyped patients with ADCA were conducted prospectively, with special emphasis on the distinction between primary neuronopathy and dying-back axonopathy. RESULTS: Electrophysiologic evidence of involvement of the peripheral nervous system was present in 70% of patients. Findings were compatible with dying-back axonopathy in 30%, while in 40% of patients, neuronopathy was diagnosed. Patients with spinocerebellar ataxia (SCA) 1 and SCA2 mostly displayed features of neuronopathy, while patients with SCA3 and SCA7 displayed both neuronopathy and axonopathy. In SCA6, no significant peripheral nerve involvement was demonstrated. We did not observe an influence of age, disease duration, or ataxia severity on the presence or type of peripheral nerve involvement. CONCLUSIONS: Peripheral nerve involvement in ADCA manifests not only as distal axonal neuropathy, but also as primary neuronopathy. Electrodiagnostic studies in this group of patients should be conducted in such a way that primary neuronopathy is detected.

Adult↗
PubMed2004

Effects of chemical chaperones on partially retarded NaCl cotransporter mutants associated with Gitelman's syndrome in a mouse cortical collecting duct cell line.

BACKGROUND: Epithelial cells lining the distal convoluted tubule express the thiazide-sensitive Na-Cl cotransporter (NCC) that is responsible for the reabsorption of 5-10% of the filtered load of Na(+) and Cl(-). Mutations in NCC cause the autosomal recessive renal disorder Gitelman's syndrome (GS). GS mutations give rise to mutant transporters that are either fully (class I) or partially (class II) retarded. Recent evidence indicates that class II mutations do not alter the intrinsic transport activity of NCC. These findings suggest that in GS caused by class II NCC mutations, pharmacological chaperones may be useful in treatment. METHODS: Initial attempts using 4-phenylbutyrate and glycerol to increase Na(+) uptake in Xenopus laevis oocytes expressing the class II mutant L215P were unsuccessful. To study the effect of the chaperones in a more physiological setting, we next expressed hNCC in the polarized epithelial cell line of distal tubular origin, mpkCCD. RESULTS: mpkCCD cells readily expressed the class II mutant R955Q, but not the class I mutant G741R. Wild-type hNCC was predominantly present in the approximately 120-1403 kD complex glycosylated form. In contrast, the R955Q mutant was predominantly present in a lower molecular weight form of approximately 100 kD. Pretreatment of R955Q expressing cells with 4-phenylbutyrate (5 mM, 16 h), but not thapsigargin (1 microM, 90 min), dimethyl sulfoxide (1%, 16 h) or glycerol (4%, 16 h), increased the expression of the complex glycosylated form and in parallel the number of hNCC positive cells. CONCLUSIONS: Taken together, the data indicate that 4-phenylbutyrate is a promising candidate for rescuing partially retarded but otherwise functional class II GS mutants.

Animals↗
PubMed2003

The structural unit of the thiazide-sensitive NaCl cotransporter is a homodimer.

The thiazide-sensitive NaCl cotransporter (NCC) is responsible for the reabsorption of 5% of the filtered load of NaCl in the kidney. Mutations in NCC cause Gitelman syndrome. To gain insight into its regulation, detailed information on the structural composition of its functional unit is essential. Western blot analysis of total membranes of Xenopus laevis oocytes heterologously expressing FLAG-tagged NCC revealed the presence of both complex-(140-kDa) and core (100-kDa)-glycosylated monomers and a broad band of high molecular mass (250-350-kDa) complexes. Chemical cross-linking with dithiobispropionimidate eliminated the low molecular weight bands and increased the intensity of the high molecular weight bands, indicating that NCC is present in multimeric complexes. Co-expression of HA- and FLAG-tagged NCC followed by co-immunoprecipitation demonstrated that these multimers contained at least two complex-glycosylated NCC proteins. The dimeric nature of the multimers was further substantiated by sucrose gradient centrifugation yielding a peak of approximately 310 kDa. A concatameric construct of two NCC polyproteins exhibited significant 22Na+ uptake, indicating that the transporter is functional as a homodimer. A concatamer of partially retarded G980R- and wild type (wt)-NCC displayed normal Na+ transport. This demonstrates that G980R-NCC, provided that it reaches the surface, is fully active and that wt-NCC is dominant in its association with this mutant. Conversely a concatamer of fully retarded G741R- and wt-NCC did not reach the cell surface, showing that wt-NCC is recessive in its association with this mutant. Oocytes co-expressing G741R- and wt-NCC did not show G741R staining at the plasma membrane, whereas Na+ transport was normal, indicating that wt-NCC dimerizes preferentially with itself. The results are discussed in relation to the recessive nature of NCC mutants in Gitelman syndrome.

Animals↗
PubMed2003

In vivo expression of putative LMX1B targets in nail-patella syndrome kidneys.

The nail-patella syndrome (NPS) is characterized by nail and bone abnormalities, associated with glomerular involvement in approximately 40% of patients. Typical glomerular changes consist of fibrillar material in the irregularly thickened glomerular basement membrane. NPS is inherited as an autosomal dominant trait and caused by heterozygous loss of function mutations in LMX1B, a member of the LIM homeodomain protein family. Mice with homozygous inactivation of the gene exhibit nail and skeletal defects, similar to those observed in patients, associated with glomerular abnormalities. Strong reduction in the glomerular expression of the alpha3 and alpha4 chains of type IV collagen, and of podocin and CD2AP, two podocyte proteins critical for glomerular function, has been observed in Lmx1b null mice. The expression of these proteins appeared to be regulated by Lmx1b. To determine whether these changes in podocyte gene expression are involved in the development of NPS nephropathy, using immunohistological techniques, we analyzed the podocyte phenotype and the renal distribution of type IV collagen chains in the kidneys of seven NPS patients with severe glomerular disease. We also examined the nature of the fibrillar material present within the glomerular extracellular matrix. The glomerular basement membrane fibrillar material was specifically labeled with anti-type III collagen antibodies, suggesting a possible regulation of type III collagen expression by LMX1B. The expression of the alpha3 and alpha4 chains of type IV collagen, and of podocin and CD2AP, was found to be normal in the seven patients. These findings indicate that heterozygous mutations of LMX1B do not appear to dramatically affect the expression of type IV collagen chains, podocin, or CD2AP in NPS patients.

Adaptor Proteins, Signal Transducing↗
PubMed2003

Exclusion of mutations in FXYD2, CLDN16 and SLC12A3 in two families with primary renal Mg2+ loss.

BACKGROUND: Based on genetic studies in families with hereditary renal Mg(2+) reabsorption disorders, several genes were shown to be involved in renal Mg(2+) transport. Mutations in the CLDN16 gene were found to underlie autosomal recessive hypomagnesaemia associated with hypercalciuria and nephrocalcinosis. The FXYD2 gene was implicated in autosomal dominant renal Mg(2+) wasting associated with hypocalciuria. Mutations in the SLC12A3 gene, also known as NCC, cause Gitelman's syndrome. In addition to hypokalaemic metabolic alkalosis, hypomagnesaemia associated with hypocalciuria is considered to be a hallmark feature of this latter disorder. METHODS: We have characterized a new family with presumed dominant renal hypomagnesaemia by detailed clinical examination and mutation analysis of CLDN16, FXYD2 and SLC12A3. In addition, we have performed mutation analysis of these three genes in a previously described family with autosomal recessive renal Mg(2+) wasting. In this family, linkage analysis was performed with polymorphic markers in the vicinity of the FXYD2 gene. RESULTS: The phenotype of the new family closely resembles that of the known dominant families with a mutation in FXYD2, but mutations in this gene were not identified in the new family. No mutations were found in CLDN16 and SLC12A3 either. Sequencing of the three genes in the patients of the recessive family revealed no mutations. In addition, haplotype analysis excluded linkage to the FXYD2 region on chromosome 11q23. CONCLUSION: Our results indicate that, in addition to the currently known loci involved in renal Mg(2+) handling, at least one other gene must be involved.

Adolescent↗
PubMed2003

Mutations in the human Na-K-2Cl cotransporter (NKCC2) identified in Bartter syndrome type I consistently result in nonfunctional transporters.

Bartter syndrome (BS) is a heterogeneous renal tubular disorder affecting Na-K-Cl reabsorption in the thick ascending limb of Henle's loop. BS type I patients typically present with profound hypokalemia and metabolic alkalosis. The main goal of the present study was to elucidate the functional implications of six homozygous mutations (G193R, A267S, G319R, A508T, del526N, and Y998X) in the bumetanide-sensitive Na-K-2Cl cotransporter (hNKCC2) identified in patients diagnosed with BS type I. To this end, capped RNA (cRNA) of FLAG-tagged hNKCC2 and the corresponding mutants was injected in Xenopus laevis oocytes and transporter activity was measured after 72 h by means of a bumetanide-sensitive (22)Na(+) uptake assay at 30 degrees C. Injection of 25 ng of hNKCC2 cRNA resulted in bumetanide-sensitive (22)Na(+) uptake of 2.5 +/- 0.5 nmol/oocyte per 30 min. Injection of 25 ng of mutant cRNA yielded no significant bumetanide-sensitive (22)Na(+) uptake. Expression of wild-type and mutant transporters was confirmed by immunoblotting, showing significantly less mutant protein compared with wild-type at the same cRNA injection levels. However, when the wild-type cRNA injection level was reduced to obtain a protein expression level equal to that of the mutants, the wild-type still exhibited a significant bumetanide-sensitive (22)Na(+) uptake. Immunocytochemical analysis showed immunopositive staining of hNKCC2 at the plasma membrane for wild-type and all studied mutants. In conclusion, mutations in hNKCC2 identified in type I BS patients, when expressed in Xenopus oocytes, result in a low expression of normally routed but functionally impaired transporters. These results are in line with the hypothesis that the mutations in hNKCC2 are the underlying cause of the clinical abnormalities seen in patients with type I BS.

Animals↗
PubMed2003

Functional expression of the human thiazide-sensitive NaCl cotransporter in Madin-Darby canine kidney cells.

The thiazide-sensitive Na(+)-Cl(-) cotransporter (NCC), which is expressed on the apical membrane of epithelial cells lining the distal convoluted tubule, is responsible for the reabsorption of 5% to 10% of filtered Na(+) and Cl(-). To date, functional studies on the structural and regulatory requirements for localized trafficking and ion-transporting activity of NCC have been hampered by lack of a suitable cell system expressing this cotransporter. Reported here is the functional expression of human NCC (hNCC) in a polarized mammalian cell of renal origin-that is, the high-resistance Madin-Darby canine kidney (MDCK) cell. Western blot testing revealed that the cells predominantly expressed the complex glycosylated (approximately 140 kD) form of hNCC. hNCC was present primarily in the apical part of the cell. The functionality of hNCC was demonstrated by the gain of thiazide-sensitive Na(+) uptake and transepithelial transport activity. Na(+) uptake was significantly increased after short-term (15 min) treatment with forskolin, whereas cyclic guanosine monophosphate, wortmannin, phorbol 12-myriatate 13-acetate, and staurosporine were without effect. This indicates that hNCC activity is regulated through cyclic adenosine monophosphate, rather than via cyclic guanosine monophosphate, phospho-inositide 3-kinases or protein kinase C. Aldosterone did not alter Na(+) uptake in the short term (15 min) but significantly increased the transport activity in the long term (16 h). The latter effect of aldosterone was due to an effect on the cytomegalovirus promoter/enhancer driving the expression of hNCC. hNCC-MDCK cells are a good model for the study of the regulation of apical trafficking and ion-transporting activity of hNCC.

Aldosterone↗
PubMed2003

Dimeric architecture of the human bumetanide-sensitive Na-K-Cl Co-transporter.

The primary mediator of NaCl reabsorption in the renal distal tubule is the human bumetanide-sensitive Na(+)-K(+)-2Cl(-) co-transporter (hNKCC2), located at the apical membrane of the thick ascending limb of Henle's loop. The physiologic importance of this transporter is emphasized by the tubular disorder Bartter syndrome type I, which arises from the functional impairment of hNKCC2 as a result of mutations in the SLC12A1 gene. The aim of the present study was to investigate the oligomeric state of hNKCC2 to understand further its operational mechanism. To this end, hNKCC2 was heterologously expressed in Xenopus laevis oocytes. Chemical cross-linking with dimethyl-3,3-dithio-bis-propionamidate indicated that hNKCC2 subunits can reversibly form high molecular weight complexes. Co-immunoprecipitation of tagged hNKCC2 subunits further substantiated a physical interaction between individual hNKCC2 subunits. The size of the hNKCC2 multimers was determined by sucrose gradient centrifugation, and a preference for dimeric complexes (approximately 320 kD) was demonstrated. Finally, concatemeric constructs consisting of two wild-type subunits or a wild-type and a functionally impaired hNKCC2 subunit (G319R) were expressed in oocytes. Subsequently, the concatemers were functionally characterized, resulting in a significant bumetanide-sensitive (22)Na(+) uptake of 2.5 +/- 0.2 nmol/oocyte per 30 min for the wild-type-wild-type concatemer, which was reduced to 1.3 +/- 0.1 nmol/oocyte per 30 min for the wild-type-G319R concatemer. In conclusion, this study suggests that hNKCC2 forms at least functional dimers when expressed in Xenopus laevis oocytes of which the individual subunits transport Na(+) independently.

Acidosis, Renal Tubular↗
PubMed2003

Dominant isolated renal magnesium loss is caused by misrouting of the Na+,K+-ATPase gamma-subunit.

Hereditary primary hypomagnesemia comprises a clinically and genetically heterogeneous group of disorders in which hypomagnesemia is due to either renal or intestinal Mg(2+) wasting. These disorders share the general symptoms of hypomagnesemia, tetany and epileptiformic convulsions, and often include secondary or associated disturbances in calcium excretion. In a large Dutch family with autosomal dominant renal hypomagnesemia, associated with hypocalciuria, we mapped the disease locus to a 5.6-cM region on chromosome 11q23. After candidate screening, we identified a heterozygous mutation in the FXYD2 gene, encoding the Na(+),K(+)-ATPase gamma-subunit, cosegregating with the patients of this family, which was not found in 132 control chromosomes. The mutation leads to a G41R substitution, introducing a charged amino acid residue in the predicted transmembrane region of the gamma-subunit protein. Expression studies in insect Sf9 and COS-1 cells showed that the mutant gamma-subunit protein was incorrectly routed and accumulated in perinuclear structures. In addition to disturbed routing of the G41R mutant, Western blot analysis of Xenopus oocytes expressing wild-type or mutant gamma-subunit showed mutant gamma-subunit lacking a posttranslational modification. Finally, we investigated two individuals lacking one copy of the FXYD2 gene and found their serum Mg(2+) levels to be within the normal range. We conclude that the arrest of mutant gamma-subunit in distinct intracellular structures is associated with aberrant posttranslational processing and that the G41R mutation causes dominant renal hypomagnesemia associated with hypocalciuria through a dominant negative mechanism.

Animals↗
PubMed2003

Confirmation of CLIM2/LMX1B interaction by yeast two-hybrid screening and analysis of its involvement in nail-patella syndrome.

Nail-patella syndrome (NPS), an autosomal dominant disorder characterized by nail dysplasia, absent or hypoplastic patellae, iliac horns, and often associated with nephropathy and, less frequently, with open angle glaucoma, is caused by mutations in the LMX1B gene. Inter-familial and intra-familial phenotypic variability raises the question whether modifier genes can be identified to explain differences in the expression and severity of clinical features of NPS. Genes encoding proteins that interact with the LMX1B protein are good candidates and, therefore, methods to search for interactions can be used to this purpose. By the yeast two-hybrid screening we detected the CLIM2 gene as a LMX1B interactor, confirming previous reports which described the same interaction by biochemical methods. Sequencing of the CLIM2 coding region in seven NPS cases in which no LMX1B mutation had been found, did not detect any molecular variant in these patients. Moreover, by genotyping a polymorphic dinucleotide repeat close to the CLIM2 gene in affected members of a large Dutch NPS family with high incidence of nephropathy, we were unable to find a correlation between the presence of a specific allele and the expression of nephropathy. In conclusion, although the results of this study could not provide any proof of CLIM2 involvement in the pathogenesis of NPS or in determination of the clinical phenotype, we suggest that the CLIM2 gene can be considered as a good candidate for further studies on normal and disturbed kidney development associated with NPS or other hereditary glomerulopathies.

DNA Mutational Analysis↗