Genomic structure of the gene for the human P1 protein (MCM3) and its exclusion as a candidate for autosomal recessive polycystic kidney disease.

The locus PKHD1 (polycystic kidney and hepatic disease 1) has been linked to all typical forms of the autosomal recessive polycystic kidney disease (ARPKD) and maps to chromosome 6p21.1-p12. We previously defined its genetic interval by the flanking markers D6S1714 and D6S1024. In our current work, we have fine-mapped the gene for the human P1 protein (MCM3), thought to be involved in the DNA replication process, to this critical region. We have also established its genomic structure. Mutation analyses using SSCP were performed in ARPKD patients' cDNA samples, leading to the exclusion of this gene as a candidate for this disorder. We also identified two intragenic polymorphisms that allowed families with critical recombination events to be evaluated. Although neither marker was informative in these individuals, they are the closest yet described for PKHD1 and may help to refine the candidate region.

Bartter syndrome: unraveling the pathophysiologic enigma.

Familial hypokalemic, hypochloremic metabolic alkalosis, or Bartter syndrome, is not a single disorder but rather a set of closely related disorders. These Bartter-like syndromes share many of the same physiologic derangements, but differ with regard to the age of onset, the presenting symptoms, the magnitude of urinary potassium (K) and prostaglandin excretion, and the extent of urinary calcium excretion. At least three clinical phenotypes have been distinguished: (1) classic Bartter syndrome; (2) the hypocalciuric-hypomagnesemic Gitelman variant; and (3) the antenatal hypercalciuric variant (also termed hyperprostaglandin E syndrome). The fundamental pathogenesis of this complex set of disorders has long fascinated and stymied investigators. Physiologic investigations have suggested numerous pathogenic models. The cloning of genes encoding renal transport proteins has provided molecular tools to begin testing these hypotheses. To date, molecular genetic analyses have determined that mutations in the gene encoding the thiazide-sensitive sodium-chloride (Na-Cl) cotransporter underlie the pathogenesis of the Gitelman variant. In comparison, the antenatal variant is genetically heterogeneous with mutations in the genes encoding either the bumetanide-sensitive sodium-potassium-chloride (Na-K-2Cl) cotransporter or the luminal, ATP-regulated, K channel. With these data, investigators have begun to unravel the pathophysiologic enigma of the Bartter-like syndromes. Further studies will help refine pathogenic models for this set of disorders as well as provide new insights into the normal mechanisms of renal electrolyte transport.

Prenatal diagnosis of autosomal recessive polycystic kidney disease (ARPKD): molecular genetics, clinical experience, and fetal morphology.

Autosomal recessive polycystic kidney disease (ARPKD) is one of the most common hereditary renal cystic diseases and has a high infant mortality. Prenatal diagnosis using fetal sonography can be unreliable, especially in early pregnancy. The ARPKD locus has been mapped to proximal chromosome 6p allowing haplotype-based prenatal diagnosis in "at-risk" families. From December 1994 to March 1997, we received 258 inquiries regarding prenatal evaluation and we have completed analyses in 212 families. To date, 65 prenatal analyses have been performed in 57 families. In the majority of the requesting families (45/57), the index children are deceased and their DNA was extracted from paraffin-embedded tissue. Eighteen fetuses were homozygous for the disease-associated haplotypes. In 12 of these fetuses, pathoanatomical examination demonstrated typical ARPKD changes consisting of dilated collecting ducts and the characteristic hepatic ductal plate malformation. These changes were detected in two fetuses as early as 13 weeks gestational age. These cases represent the earliest demonstration of ARPKD-associated histopathology reported to date. One high risk fetus was carried to term and turned out to be unaffected. However, the diagnosis of ARPKD remained doubtful in the index patient. Forty-three fetuses were either heterozygous or homozygous for a nondisease-associated haplotype and all infants born were phenotypically unaffected at birth. In four cases, a recombination event occurred between the flanking markers and no genotypic prediction was possible. Three of these pregnancies were terminated and necropsy of the fetuses confirmed ARPKD, while one fetus was carried to term and showed no abnormalities at birth. These results show that haplotype-based prenatal testing is feasible and reliable in pregnancies "at risk" for ARPKD. An absolute prerequisite for these studies is an accurate diagnosis of ARPKD in previously affected sib(s).

Overview: the genetics of renal disease.

Disease susceptibility and, to a certain extent, disease progression, in many renal disorders are determined by specific genetic factors. The Human Genome Project has generated an explosion of gene discovery tools and strategies. These new technologies are being applied to a wide range of renal disorders. The most significant impact to date has been the identification of disease-susceptibility genes for more than 20 monogenic renal disorders. This review summarizes the role of genetics in renal disease, the different modes of inheritance of disease-susceptibility genes, the strategies for gene discovery, and the clinical impact of disease gene identification.

Inherited tubular transport disorders.

Inherited abnormalities of renal tubular transport processes encompass a heterogeneous set of disorders due to single gene defects. Elucidating the molecular basis for these generally rare disorders has provided important insights into disease pathogenesis as well as the complexities of normal renal transport physiology. This review focuses on the clinical features and recent molecular advances in cystinuria, X-linked hypophosphatemic rickets, and the Bartter-Gittelman spectrum of disorders. Also addressed are disorders of calcium homeostasis resulting from loss-of-function and gain-of-function mutations of the extracellular calcium-sensing receptor, as well as the single gene defects that cause distal renal tubular acidosis.

Overexpression of innate immune response genes in a model of recessive polycystic kidney disease.

Defects in the primary cilium/basal body complex of renal tubular cells cause polycystic kidney disease (PKD). To uncover pathways associated with disease progression, we determined the kidney transcriptome of 10-day-old severely and mildly affected cpk mice, a model of recessive PKD. In the severe phenotype, the most highly expressed genes were those associated with the innate immune response including many macrophage markers, particularly those associated with a profibrotic alternative activation pathway. Additionally, gene expression of macrophage activators was dominated by the complement system factors including the central complement component 3. Additional studies confirmed increased complement component 3 protein levels in both cystic and non-cystic epithelia in the kidneys of cpk compared to wild-type mice. We also found elevated complement component 3 activation in two other mouse-recessive models and human-recessive PKD. Our results suggest that abnormal complement component 3 activation is a key element of progression in PKD.

Determinants of renal volume in autosomal-dominant polycystic kidney disease.

The Consortium of Radiologic Imaging Studies of Polycystic Kidney Disease (CRISP) recently showed that renal enlargement in autosomal-dominant polycystic kidney disease mimicked exponential growth. We determined the effects of cyst initiation rate, total number, and growth rate on the time-dependent change of total cyst volume (TCV). Mathematical models with equations integrating cyst surface area, volume, and an invariant growth rate constant were used to compute the time-dependent change in volume of solitary and multiple cysts. Multiple expanding cysts increased TCV in an exponential-like pattern even when individual cysts formed at different rates or exhibited different but constant growth rates. TCV depended on the rate of cyst initiation and on the total number of cysts; however, the compounding effect of exponential-like growth was the most powerful determinant of long-term cyst expansion. Extrapolation of TCV data plots for individual subjects back to an age of 18 predicted TCV values within an established range. We conclude that cysts started early in life were the main contributor to eventual TCV while their growth rate primarily determined renal size; although the rate of formation and the ultimate number of cysts also contributed. The good fit between the exponential models and the extrapolated CRISP data indicates that the TCV growth rate is a defining trait for individual patients and may be used as a prognostic marker.

Molecular insights into the pathogenesis of inherited renal tubular disorders.

Physiologic and biochemical studies have suggested that the inherited disorders cystinuria, Liddle's syndrome, and perhaps Bartter syndrome all result from defects in renal tubular transport processes. With the recent isolation of several candidate transporter genes, these clinically based hypotheses have begun to be confirmed at a molecular level. In addition, the cloning of the water-channel family of proteins has facilitated the characterization of a second gene defect in congenital nephrogenic diabetes insipidus. This review integrates the pathophysiology of these inherited renal tubular disorders with recent molecular genetic discoveries, and provides a starting point for unraveling their pathogenesis at the molecular level.

Toad urinary bladder epithelial cells contain an analogue of cytoskeletal protein 4.1.

Epithelial cell polarity and vectorial transport require cytoskeletal proteins that maintain local cell membrane structure and mediate cytoplasmic vesicle movement. The cytoskeleton of leaky epithelia, such as the intestinal mucosa and renal proximal tubule cells, has been extensively studied. However, cytoskeletal studies in tight epithelia such as the mammalian collecting duct and toad urinary bladder generally have been confined to ultrastructural investigation. Recent research in nonepithelial cell types has identified an interesting family of cytoskeletal proteins. Present in multiple cell types, these protein 4.1 analogues share a number of similar functional characteristics, yet are structurally diverse. They are multiply phosphorylated by several different kinases, and phosphorylation regulates their associations with other cytoskeletal constituents, integral membrane components, and cytoplasmic vesicles. Using a combination of immunochemical and immunofluorescent techniques, we have demonstrated that toad bladder epithelial cells contain a 65-kDa analogue of human erythrocyte protein 4.1. Toad bladder epithelial cell protein 4.1 is structurally similar to its erythrocyte counterpart and is phosphorylated. This protein 4.1 species is present throughout the toad bladder granular cell cytoplasm, suggesting that it participates in multiple granular cell functions.

Reversible Fanconi syndrome associated with valproate therapy.

Two children with developmental delay and seizure disorders had Fanconi syndrome associated with valproate therapy. Both recovered normal proximal tubular function within 4 months of discontinuing valproate therapy.

The mouse polycystic kidney disease mutation (cpk) is located on proximal chromosome 12.

The mouse congenital polycystic kidney (cpk) mutation produces a condition that resembles human autosomal recessive polycystic kidney disease (ARPKD) in its pattern of inheritance, clinical progression, and histopathology. Inheritance of this mouse mutation in crosses segregating the Rb(12.14)8Rma translocation chromosome and various DNA markers of Chromosome 12 have localized cpk to a site near D12Nyu2, approximately 7 cM from the centromere of Chromosome 12. This result suggests that the homologous PKD2 gene should be localized to either human chromosome 2p23-p25 or chromosome 7q22-q31.

Bartter syndrome in Costa Rica: a description of 20 cases.

Bartter syndrome involves an overlapping set of closely related renal tubular disorders which can be subdivided into at least three clinical phenotypes: (1) classic Bartter syndrome (2) Gitelman syndrome, and (3) a neonatal variant of Bartter syndrome. In contrast to classic Bartter syndrome and Gitelman syndrome, the neonatal variant of Bartter syndrome has both the features of renal tubular hypokalemic alkalosis as well as profound systemic manifestations. Specifically, neonatal Bartter syndrome is characterized by intrauterine polyhydramnios, premature delivery, and life-threatening episodes of fever and dehydration. Most of these infants also have severe hypercalciuria with associated nephrocalcinosis and osteopenia. Over a 22-year period, 20 Costa Rican patients with a congenital syndrome that resembles neonatal Bartter syndrome have been identified and characterized. While these patients exhibit some of the clinical characteristics previously described for neonatal Bartter syndrome, this cohort also has a set of distinct features. They are predominantly female, have a later age of diagnosis, manifest a relatively unique set of physical traits, and appear to have milder clinical disease. Given these differences, it will be important to apply the emerging molecular tools to determine whether the phenotypic variability indicates genetic heterogeneity in neonatal-onset Bartter syndrome.

Low peripheral plasma renin activity as a critical marker in pediatric hypertension.

In evaluating hypertensive children and adolescents, the etiological considerations should include a set of inherited disorders that share very low plasma renin activity (PRA) as a common feature. In particular among these disorders, glucocorticoid remediable aldosteronism (GRA) appears to be emerging as an important etiology of hypertension in the pediatric population. We report the evaluation of a 9-year-old Caucasian girl who presented with severe hypertension and a strong family history of early-onset hypertension. Her suppressed PRA, her family history, and her failure to respond to conventional anti-hypertensive therapy raised GRA as a potential etiology. The diagnosis was confirmed by an elevated ratio of urinary 18-oxotetrahydrocortisol to urinary tetrahydroaldosterone and genetic testing, which demonstrated the chimeric gene duplication. The molecular pathogenesis of GRA and the clinical implications are reviewed.

Germline and somatic loss of function of the mouse cpk gene causes biliary ductal pathology that is genetically modulated.

The mouse cpk mutation is the most extensively characterized murine model of polycystic kidney disease (PKD) and closely resembles human autosomal recessive PKD (ARPKD), with the exception that B6-cpk/cpk homozygotes do not express the biliary ductal plate malformation (DPM) lesion. However, homozygous mutants from outcrosses to other strains, e.g. DBA/2J (D2), CD-1, BALB/c and Mus mus castaneus (CAST), express the DPM. The current study was designed: (i) to characterize the cpk-associated biliary disease in affected F(2) homozygotes from intercrosses with either CAST or D2; and (ii) to evaluate focal biliary cysts identified in heterozygotes from a D2-cpk congenic strain. We found that all F(2) cpk/cpk pups expressed both the typical renal cystic disease and the DPM. The DPM severity, assessed using semi-quantitative histopathological analysis, was markedly variable in these F(2) progeny. We found no correlation between the severity of the DPM and the renal cystic disease in either F(2) cohort. In addition, we identified focal cysts, apparently of biliary origin, in the livers of both aged D2-+/cpk and F(1) heterozygotes. Genetic analysis demonstrated loss of heterozygosity at the cpk interval and supports a loss-of-function model for biliary cysts. We conclude that the cpk allele contains an inactivating mutation which disrupts tubulo-epithelial differentiation in the kidney and biliary tract. Expression of the biliary lesion is modulated by genetic background, and the specific biliary phenotype is determined by whether loss of function of the cpk gene occurs as a germline or a somatic event.

The mouse congenital polycystic kidney (cpk) locus maps within 1.3 cM of the chromosome 12 marker D12Nyu2.

The mouse congenital polycystic kidney (cpk) mutation causes bilateral cystic dilatation of the renal collecting tubules and leads to rapidly progressive renal insufficiency in affected homozygotes. The phenotype of the cpk/cpk mutants closely resembles that of human autosomal recessive polycystic kidney disease (ARPKD). Previously, we have reported that the cpk locus maps close to D12Nyu2 on Chromosome (Chr) 12. To determine the cpk map location more precisely, we have extended our previous studies using additional progeny and additional markers of proximal Chr 12. These recent studies position cpk within 1.3 cM of D12Nyu2, closely flanked by (Odc, D12Mit10) and (Tpo, D12Mit12). Our data support an ordered array of seven DNA markers that will provide reference points for building a physical map of the Chr 12 region centered on cpk. Moreover, these data establish that cpk lies within a linkage group that is conserved between mouse Chr 12 and human chr 2p24-2p25. This assignment to a region of homology will facilitate human linkage analyses to determine whether mouse cpk and human ARPKD are mutations of homologous genes.

Dominantly transmitted glomerulocystic kidney disease: a distinct genetic entity.

Glomerulocystic kidney disease (GCKD) is a relatively rare condition with both a sporadic and familial occurrence. Pathologically, GCKD is characterized by cystic dilatation of Bowman's space and the initial proximal convoluted tubule. As a heritable disorder, GCKD has primarily been recognized in infants with a family history of classic, autosomal dominant polycystic kidney disease (ADPKD). Dominantly transmitted GCKD associated with either hypoplastic or normal-sized kidneys has also been reported in older children and adults. A large, three-generation African-American family with familial GCKD is characterized. Of the 20 individuals available for study, seven affected individuals were identified by renal sonogram or renal histopathology. GCKD in this family segregates as an autosomal dominant trait as evidenced by its apparent transmission from a father to his sons. A set of directed linkage strategies indicates that the distinctive GCKD phenotype in this family results from a dominantly acting mutation that disrupts a genetic locus distinct from the ADPKD loci, PKD1 and PKD2, as well the human homologue of mouse jcpk mutation, a newly described murine GCKD. These analyses are the first known genetic studies conducted in a family with heritable GCKD and post-infantile age of onset.

Diffuse renal cystic disease in children: morphologic and genetic correlations.

During a 5-year period, we evaluated seven infants and two fetuses who presented with enlarged, hyperechoic kidneys. In each, the initial clinical diagnosis was autosomal recessive polycystic kidney disease (ARPKD). Among the seven unrelated infants were three Caucasian and four African-American infants. No syndromic stigmata were evident in any of these infants. At the time of the initial evaluation, the family data were incomplete for four infants. The two fetuses were presumed to be at-risk for ARPKD based on the diagnosis in previous siblings. Renal histopathology was evaluated in all nine cases and revealed a spectrum of cystic disease ranging from ARPKD to glomerulocystic kidney disease to autosomal dominant polycystic kidney disease to diffuse cystic dysplasia. In the eight cases for whom liver histopathology was available, varying degrees of biliary dysgenesis were evident. We present a detailed analysis of the key histopathological features in each case and discuss the histopathological findings in an embryological context. In addition, we address the current role of molecular genetics in the diagnostic evaluation.

Characterization of the region containing the jcpk PKD gene on mouse Chromosome 10.

The jcpk gene on mouse Chromosome 10 causes a severe, early onset form of polycystic kidney disease (PKD) when inherited in an autosomal recessive manner. In order to positionally clone this gene, high resolution genetic and radiation hybrid maps were generated along with a detailed physical map of the approximately 500-kb region containing the jcpk gene. Additionally, sixty-nine kidney-specific ESTs were evaluated as candidates for jcpk and subsequently localized throughout the mouse genome by radiation hybrid mapping analysis. Previous studies indicating non-complementation of the jcpk mutation and 67Gso, a new PKD translocation mutant had suggested that 67Gso represents a new allele of jcpk. Fluorescence in situ hybridization (FISH) analysis using key bacterial artificial chromosome clones from the jcpk critical region, refined the 67Gso breakpoint and provided support for the allelism of jcpk and 67Gso.