Charles and Jane Pak Center of Mineral Metabolism and Clinical Research, and the Department of Internal Medicine, University of Texas Southwestern Medical Center, Dallas, Texas
Address correspondence to: Dr. Orson W. Moe, Charles and Jane Pak Center of Mineral Metabolism and Clinical Research, University of Texas Southwestern Medical Center, 5323 Harry Hines Boulevard, Dallas, TX 75390-8855. Phone: 214-648-7993; Fax: 214-645-9993; E-mail: orson.moe{at}utsouthwestern.edu
Hypercalciuria is an important, identifiable, and reversiblerisk factor in stone formation. The foremost and most fundamentalstep in dissecting the genetics of hypercalciuria is understandingits pathophysiology. Hypercalciuria is a complex trait. Thisarticle outlines the various factors that compromise the attemptto dissect the genetics of hypercalciuria, summarizes the clinicaland experimental monogenic causes of hypercalciuria, and outlinesthe initial results from attempts in studying polygenic hypercalciuria.Finally, the problem is set in perspective of the current database,technologic advances and limitations are highlighted, and prospectsof further advances in the field are speculated upon.
The incidence and composition of kidney stones vary considerablyin different parts of the world, but nephrolithiasis remainsa formidable health problem worldwide (1–4). It is importantto emphasize that although nephrolithiasis is accepted as a"diagnosis" in common clinical parlance, its presence conferslittle information about the underlying defect. The occurrenceof a kidney stone can reflect a diverse list of underlying diagnoses.From this standpoint, nephrolithiasis per se is not much moreof a "diagnosis" than for example high BP, ascites, or fever.Regional and etiologic variations not withstanding, calcium-containingstones consistently constitute the majority of nephrolithiasisin all parts of the world. Excessively high concentration ofcalcium in the urine is one distinctly identifiable and correctablefactor in stone formation, although the mechanisms that predisposeto and/or result in precipitation and growth of calcium crystalsin the urinary tract are still incompletely understood.
This monograph aims to achieve several objectives. In the firstpart of the article, we highlight the importance of understandingpathophysiology, accentuating specifically the complexity andmultiorgan nature of hypercalciuria. The intent is to underscorethe critical importance of eliciting pathophysiology when oneattempts to dissect the genetics of a complex trait such ashypercalciuria. We then outline the various factors that compromisethe attempt to dissect the genetics of hypercalciuria. In thesecond part, we summarize the clinical and experimental monogeniccauses of hypercalciuria, not so much as a mere reiterationof the numerous existing excellent reviews on genetic causesof hypercalciuria (5–8) but rather to provide a frameworkto ponder about the formidable challenge that looms ahead—hypercalciuriaas a complex trait. This final topic is the focus of the thirdpart of this article. We poise the problem in perspective ofour current database, highlight technologic advances and limitations,and speculate on prospects of further advances in the field.
Biomineralization: Crystallization of Calcium Salts as a Physiologic Event
"Calcium stone formation" is mineral crystallization in bodytissue or fluid. The deposition of inorganic minerals, crystallineor noncrystalline, around biomolecules is universal in biology,where inorganic crystals are harnessed to become an integralpart of organic tissue to provide hardness and strength. Theseinorganic substances are capable of reversible interaction withbiomolecules so that the crystalline structures can be remodeledfor physiologic needs. Williams (9) discussed the unique abilityof calcium to interact with biomolecules from the point of viewof its concentration, binding strength, rate constants, andmolecular structure. Calcium salts have highly adaptable coordinationgeometry that greatly facilitates binding of calcium, in itssolid state or solution, to the irregular geometry of proteins.
The remarkable material properties of bones and teeth resultfrom the activities of proteins that function at the organic-inorganicinterface (10). Proteins have evolved domains that specificallyinteract with calcium crystals. The aspartate-phosphoserine-phosphoserine-glutamate-glutamate(DpSpSEE) motif described in the saliva protein statherin isalso found in other calcium crystal–interacting proteins,such as osteopontin (11,12). Unlike the EF hand that binds ioniccalcium, this DpSpSEE motif binds solid-phase calcium phosphatecrystals and is conserved down to invertebrates such as crustaceans.This motif is expressed in a protein at the foot pad that allowsthe mussel to attach to calcareous substratum (13).
More than 200 yr ago, Hunter (14) articulated about the similaritybetween stone formation and calcification. He pointed out theequivalence of enamel, eggshell, gallstones, and kidney stones.Thus, mineralization can be arbitrarily divided into physiologicand pathologic. The partition is not always distinct but canbe distinguished on the basis of favorable versus undesiredconsequences to the organism. Physiologic crystallization includesformation of exoskeleton, pearl, endoskeleton, and dentition,whereas pathophysiologic crystallization includes pyrophosphatearthropathy, pigmented gallstones, vascular calcification, andurolithiasis. Pathologic calcium crystallization is a physiologicprocess in the wrong place and the wrong time.
Multiorgan Approach to Hypercalciuria
Because isolated hypercalciuria per se is not detrimental, aclinician’s interest in hypercalciuria concerns the complicationsthat include mainly nephrolithiasis and nephrocalcinosis andperhaps also less morbid conditions such as hematuria (15) andpossibly polyuric enuresis in children (16). The pathophysiologyof calcium nephrolithiasis versus nephrocalcinosis is incompletelyunderstood, but the reader is referred to a recent review bySayer et al. (17). It is not the purpose of this review to tacklethe debate about the relative importance of hypercalciuria versushyperoxaluria in calcium oxalate stones, but a recent studyclearly demonstrates that urinary calcium and oxalate are equallyimportant for stone formation (18).
A majority of patients with this condition have been categorizedas "idiopathic hypercalciuria." The term is decades old andseemed to have secured an indelible place in medical idiom.Although linguistically correct without doubt, it carries animplicit presumption that this is a single condition that isinevitably incorrect and furthermore takes on certain overtonesof futility in unraveling the underlying cause. Although theuse of this term is convenient in common clinical lingo, onemust caution the absolute and unquestioned acceptance of thisterm as a "diagnosis." Calcium homeostasis involves multipleorgans by predominantly endocrine mechanisms and fluxes throughthree target organs in concert effect calcium homeostasis. Anyanalysis of hypercalciuria should at least take the three organsinto consideration. Pak et al. pioneered this attempt back in1975 by introducing a tripartite classification of absorptive,resorptive, and renal hypercalciuria (19). From a pathophysiologicand genetic point of view, this is a very important classification.
Figure 1 depicts three scenarios in which in each, a primarydisturbance occurs in one of the three calcium homeostatic organs.Figure 1A shows an example of pure renal leak. Although a lotof causes of hypercalciuria such as acid load–inducedhypercalciuria has a component of renal leak, multiple mechanismsoften come into play. Pure renal hypercalciuria can be seenin mice with deletion of the TRPV5 calcium channel gene (20).Compensatory intestinal hyperabsorption and bone resorptionmaintains serum calcium at normal levels (20). It is understandablehow the usual battery of clinical tests at the outpatient settingmay not recognize the hypercalciuria as a renal leak. One hasto possess some or all of the following data: (1) Serum parathyroidhormone and 1,25-(OH)2 vitamin D are higher than expected; (2)hypercalciuria is inappropriate for the slightly elevated parathyroidhormone, normal serum calcium, and normal filtered calcium;(3) persistent hypercalciuria is present even during fasting;and (4) increased bone resorption markers and/or reduced bonemineral density. This battery of information may not be feasiblein clinical practice with the time, infrastructure, and financialconstraints. However, it is absolutely imperative that one notabandon the quest for a complete definition of pathophysiologyand strive to uncover a proximal (closer to the underlying lesion)phenotype.
Figure 1. Pathophysiology of hypercalciuria. Hypercalciuric syndromes all are multisystemic diseases and are arbitrarily divided into three overlapping categories according to the primary or predominant disturbance. In each instance, secondary compensatory changes involve multiple hormones such as parathyroid hormone (PTH) and 1,25-dihydroxy-vitamin D3 [1,25-(OH)2vit D]. (A) Renal leak with secondary gut hyperabsorption and increased bone release. (B) Imbalance between bone formation and resorption. (C) Intestinal hyperabsorption. The three primary disturbances can occur simultaneously.
Figure 1, B and C, illustrates two additional scenarios withincreased bone resorption and increased gut absorption as primarydisturbances that cause hypercalciuria. Although the underlyingcauses are diverse and the physiologic profiles are completelydifferent, all three situations culminate in hypercalciuria.Some have argued that because it is so difficult to separateand characterize the defects in these organs, one should foregothat and simply aggregate everything into a pot of heterogeneousconditions. We submit the counter argument that unless one continuouslystrives to dissect and classify on the basis of pathophysiologiccharacteristics, it will be hopeless to advance our understanding.
Dissecting the Genetics of Hypercalciuria
Approximately half of the patients who are labeled as havingidiopathic hypercalciuria have a positive family history ofkidney stones (21). Familial idiopathic hypercalciuria has beendescribed as an autosomal dominant trait in the earlier literature.This was a de facto conclusion based on the observation thatit did not match a recessive pattern, and the male-to-male inheritanceruled out gender-linked transmission (22–26). This isclearly an oversimplification of the genetics of familial hypercalciuria.Not all genetic diseases can be dealt with with a Mendelianapproach. Some unavoidable ambiguity in both genotyping andphenotyping notwithstanding, it is usually possible to discernwhether one has clinical autosomal dominant polycystic kidneydisease and whether one has mutations in polycystin-1 or -2.Such definitive assignments are not possible for hypercalciuriabecause of a number of immanent impediments. Hypercalciuriais a complex trait that cannot be slotted into classical Mendeliancategories. The inherent difficulties of dissecting the geneticsof hypercalciuria are schematically summarized in Figure 2.As the central tenet to all genetic approaches involves someform of genotype-phenotype correlation, phenotypic ambivalencecan deliver a cataclysmic blow to the effort. Figure 2A depictsthe breakdown of the classical Mendelian perfectly co-segregating"one gene locus-one phenotype" paradigm. Each of the followingcharacteristics of hypercalciuria imparts equivocation in thegenotype-phenotype correlation (Figure 2B).
Figure 2. Hypercalciuria as a complex trait. Classical one gene–one protein hence one mutation–one disease model. (A) The underlying mechanisms that account for this non-Mendelian inheritance pattern. (B) Complex traits create exceptions to this linear paradigm such as multiple mutations converging into one disease (middle) or one mutation diverging into several diseases.
Continuous Variable
Whereas the presence or absence of a calcareous calculus inthe urinary tract can theoretically and practically be classifiedas a "yes-no" event, the more proximal and more important phenotype,hypercalciuria, cannot assume such binary status. Akin to parameterssuch as BP and body mass, urinary calcium excretion is a continuousvariable. Although there is little difficulty in assigning diseasestatus at the extreme ends of the spectrum, there is reallyno sharp distinction of margins. "Disease" in this case refersto the bearing of increased statistical risk of nephrolithiasisand/or nephrocalcinosis. Hypercalciuria is an example par excellenceof a quantitative trait (Figure 2). Further compounding thelack of discrete boundary is that the same amount of calciumin a 24-h urine sample can have completely different implicationsdepending on the urine volume; pH; concentration of all of thecalcium-interacting anions such as citrate, phosphate, and oxalate;and concentration of promoters and inhibitors of crystallization.Not to mention that each one of these other factors is alsoa continuous variable. Although there is no good way of creatinga sharp distinction when there is none, one can partially circumventthe problem by analyzing relative supersaturation of calciumoxalate and phosphate in addition to the amount of calcium inclinical samples.
Secondary and Compensatory Effects
As discussed above, hypercalciuria can be accompanied by anextensive panoply of effects that are downstream from or ascompensatory responses to the primary disturbance. This rendersthe use of hypercalciuria singularly as a phenotypic variablevery risky. As shown in Figure 1, three completely differentsyndromes all share the feature of hypercalciuria. Unless onecommits extra effort to dissect out and identify the primaryversus secondary and compensatory changes, using one parameteras "phenotype" will severely undermine the genotype-phenotypecorrelation.
Phenocopy
There are a host of nongenetic factors that can affect gut calciumabsorption, bone calcium release, and renal calcium handling.The cartoon in Figure 2A displays some dietary (representingcompletely nongenetic) factors that can influence urinary calciumexcretion (27). Other than a dietary history, which is semiquantitativeat best, it is very difficult to rule out increased dietarycalcium intake as a cause for hypercalciuria. The cartoon alsoshows that high dietary protein (source of acid) and sodiumcan also cause "physiologic hypercalciuria" (27,28). In theseinstances, one can at least estimate from the 24-h urine samplethe impact of salt (urinary Na+) and acid (urinary NH4+ andSO42–) on urinary calcium. As it is difficult to collectand analyze 24-h urine under the luxury of dietary control inthe setting of a clinical practice, metabolic studies in thesetting of a clinical research center are indispensable. Inaddition to dietary factors, there are multiple other nongeneticfactors that affect intestinal calcium absorption and bone formation/resorption(not covered in Figure 2A). The only safeguard against phenocopicartifacts is sound knowledge and data in pathophysiology.
Polygenic Influence
Adopting an exaggerated view for the sake of emphasizing a point,one can make the extreme statement that there is no such conditionas a pure monogenic disease. Consider some quintessential monogenichuman diseases. Two different individuals may have the identicalmutation in cystic fibrosis transmembrane regulator (CFTR) andyet the clinical severity in cystic fibrosis can differ vastly(29). Similar can be said for the same mutation in -globin causingvery mild or very severe sickle cell anemia (30). There is nodoubt that modifier genes are at work. Consider the idealizedsituation in the laboratory, where one specifically deletesone and only one gene in a murine model with a completely homogeneous(as far as one can tell) genetic background. This is probablythe closest example that one can fathom of a pure monogenicdisorder. It is already well known that the same gene knockoutin the background of different strains can have completely differenteffects. As the investigator delights with the emergence ofthe good news that 80% of the null mice survives and hence canbe studied, one should wonder about the effect of the gene deletionon the 20% of the mortal ones. Penetrance of a singular genemay be small or even negligible, but multiple genes in concertcan have a significant impact on the phenotype. Envision fivehypothetical genes that have an impact on urinary calcium excretion.Each of these genes is polymorphic in Homo sapiens with multiplefully functional alleles that have small quantitative differencesin function. For instance, gene product 1 influences intestinalcalcium absorption, gene product 2 modifies the ability of dietaryacid to release calcium from bone, gene product 3 controls paracellularcalcium permeability in the thick ascending limb, gene product4 influences calcium absorption in the distal convoluted tubule,and gene product 5 alters the sensitivity of the renal calciumsensing receptor to plasma calcium concentration. If each ofthese gene products contributes to a trivial 0.5 mmol/d (20mg) increase in urinary calcium excretion, then inheriting allfive predisposing alleles can theoretically raise urinary calciumby 2.5 mmol/d (100 mg).
Loci Heterogeneity
Similar reasoning along the lines of polygenic influence isloci heterogeneity. Different underlying genetic lesions canstart the pathophysiologic cascade in vastly different manners,but by the time the lesion reaches the whole organism (clinical)level, they may have converged on an end organ in a very restrictedmanner. Renal cysts in humans can result from mutations in ADPKD1,ADPKD2, or ARPKD (31). In animals, there is an enormous collectionof monogenic mutations that converge into a final common phenotypeof renal cyst formation (32). As depicted in Figure 2B, twoindividuals with identical phenotypes may actually have no overlapwhatsoever in their underlying disease-causing genetic loci.Thus, launching a genetic search from hypercalciuria as a startingpoint may lead an investigator to multiple and confusing destinations.However, these different loci will likely have a different pathogenicpathway en route to hypercalciuria and will manifest with differentproximal phenotypes.
The complex nature of hypercalciuria is addressed in more detailbelow. Before one embarks on this formidable exercise, someconsideration will be given to monogenic causes of hypercalciuriaboth in human diseases and in animal models.
Hypercalciuria as a Monogenic Trait
Monogenic diseases often conjure up a sense of disinterest (disgustto some) in many practitioners because of their exotic natureand commonly perceived detachment from the everyday realityof medical practice. This is a misconception but a perfectlyunderstandable one. As one peruses a typical catalogue of thisnature (e.g., tables in this article), one notices that halfof the names of diseases are not recognizable and the otherhalf are not pronounceable, all of which will probably nevershow up in our clinic. With such pretext, how does one justifydevoting the majority of his or her time and effort on sucha minority of medical illnesses? The answer lies in the relatively"clean" nature of the primary lesion. These diseases allow oneto build upon two irrefutable facts: The mutation of a singlegene and an unmistakable phenotype well known and characterizedby clinicians as a delineated syndrome. These two facts allowinvestigators to plant two pillars on solid grounds and workout the elusive black boxes in between.
A number of monogenic causes of hypercalciuria have been identified,and there is no simple classification. Tables 1 through 5 presenta summary of these diseases in terms of their effects on thethree principle organs of calcium homeostasis. We took the libertyto include experimental monogenic diseases (Table 5) becausethese man-made monogenic disease models are rapidly becomingmore common than naturally occurring monogenic diseases. Tables 1 through 4 classify these disorders into whether the primarylesion primarily ails the gastrointestinal tract, kidneys, bone,or other organs. Regardless of the site of the primary lesion,multiple organs are always involved.
Table 5. Animal models of monogenic hypercalciuria
Detailed discussion of each of these conditions is beyond theobjective of this review. Some highlights are presented. Gastrointestinalmonogenic diseases associated with hypercalciuria are uncommon(Table 1) (33–58). There is a group of monogenic diseasesin which intestinal calcium hyperabsorption has been described(Table 1) but the mechanism is undetermined (33–40). Asecond group of monogenic diseases have vitamin D–mediatedintestinal hyperabsorption including primary renal diseasessuch as renal phosphate wasting (41–44). Finally, thereis a group of chromosomal deletion syndromes (not monogenic)in which intestinal hyperabsorption has been described but themechanisms are unknown (45–58). Monogenic diseases thataffect primarily the bone leads to hypercalciuria largely viaincreased calcium release into the circulating pool (Table 2)(59–69). These can be due to conditions related to structuraldefects in bone such as osteogenesis imperfecta (59–61),part of a hyperparathyroid state (62–65), or ricketic-likebone lesion (66–69).
Primary renal monogenic lesions are summarized in Table 3. Severaltreatises of a similar nature have been published (5–8),and we do not delve into this topic any further except to highlighta few notions. There is a large group of monogenic proximaltubulopathies with variable features of the Fanconi syndromethat, when full-blown, represent general proximal tubule dysfunction(70–86). The mechanism of hypercalciuria is not understoodin these diseases, but this is an excellent example of lociheterogeneity. We elected to classify primary renal phosphatewasting from mutations of the Na-phosphate co-transporter NaPi-IIain Table 1 because of the compensatory increase in vitamin D.Thick ascending limb tubulopathies involve reduced paracellularcalcium permeability (87–91), reduction of driving force(92–98), and the erroneous resetting of physiologic controlfor calcium absorption (99–105). The distal tubulopathiesalso represent a heterogeneous group of primary hypercalcurias(106–113). Despite the near identical effects on Na+,K+, and H+ homeostasis by WNK1 and WNK4 mutations, patientswith WNK4Q596E mutation have hypercalciuria and low bone mineraldensity (107), while patients who carry the WNK1 intronic deletionare not reported to have hypercalciuria. The way by which WNKkinase mutations lead to hypercalciuria is not yet clear. Therenal tubular acidoses (108–112) present a complex pictureof the multiorgan origin of hypercalciuria. In addition to gutand bone compensation, acidemia per se can increase bone resorption.Finally, a group of hypercalciuric diseases (116–121)with a wealth of mechanisms are awaiting to be explored (Table 4).
Animal models of monogenic diseases are valuable. The geneticallyaltered animals listed in Table 5 have hypercalciuria from targeteddeletion of known disease-causing genes (122–127) andgenes coding for important calcium homeostatic proteins (20,128–134),and some have hypercalciuria of serendipity (135,136). Animalmodels using ClC5 knockdown (122) or knockout (123,124) havecaptured some but not all of the salient features of Dent’sdisease. This most likely reflects polygenic modifiers in action.Even in a classical monogenic disorder, there may be some lociheterogeneity as evident by the fact that some patients withclassic features of Dent’s do not harbor mutations inClC5 (137). The variable phenotype of calbindin-28K deletionis another testimony of polygenic modifiers (130–133).An intact animal with a known deliberate gene deletion and aphenotype that bears resemblance to human disease is highlyuseful for working out the intermediate steps in the pathogenesis.
Hypercalciuria as a Complex Trait
Calcium nephrolithiasis is a very distal (many steps downstreamfrom the primary lesion) phenotype that can result from a varietyof causes. Even if one takes a step more proximal from calciumstones to hypercalciuria, one is still confronted with a phenotypegoverned by multiple organs with numerous genetic and environmentaldeterminants. There is no doubt that hypercalciuria is a complextrait. After the display of the difficulties in studying a traitas complex as hypercalciuria, one wonders whether there is anyhope in ever unraveling the genetics in this conditions. Theapproaches outlined by the classic review by Lander and Schork(138) a decade ago are still in active use today, and the readeris referred there for an excellent detailed discussion. A morerecent update can be found in the review by Risch and Merikangas(139).
One can fathom ways to bypass or counteract the complexity andenable some degree of informative genotype-phenotype correlation.One can confine the phenotype to the most severe end of thecalciuria continuum, restrict it to a subgroup of fasting hypercalciuriaor hypercalciuria accompanied by heightened intestinal absorptionand reduced bone density, or include only hypercalciuric patientswith a strong family history. All of these efforts are directedto capture a more proximal phenotype and hopefully more homogeneousgenetics. This underscores once again the critical importanceof good metabolic evaluation of patients and that hypercalciuriaper se is not an adequate phenotype for genetic studies. Anotherapproach is to use single large kindreds. The reason is thatthe degree of polygenic influence and loci heterogeneity inthe general population is much reduced in a single pedigreeas long as the number of individuals offers adequate power forthe analysis. Allele-sharing method between sibling pairs isfounded on the rationale that even though a trait is polygenic,the inheritance of a given gene (albeit amid numerous modifiers)is Mendelian (138–140). In this section, we highlighta few issues and review some existing data that are germaneto hypercalciuria.
Association
The easiest method that has been widely used in a lot of complextraits is a population-based association study (Figure 3). Thisis a case-control design that examines the frequency of a particularallele in affected versus unaffected individuals. Unfortunately,this is also the weakest design in terms of generating definitiveinformation. The ambiguity of affected versus unaffected hasalready been discussed. The polygenic nature and loci heterogeneityfurther worsens the problem. In addition, the phenomenon ofpopulation admixture renders this method ridden with artifacts(138). It is highly likely that two populations differ in aspectsother than the disease of interest; hence, any given allelewill have a good chance of having different frequency in twodifferent populations (Figure 3). Positive studies are commonwith the association approach, and the caveats are summarizedin Figure 3. If the different alleles of the gene of interesthave defined differences in function and that difference canaccount for some phenotypic differences, then an associationwill carry more meaning. If the alleles are simply polymorphisms,the one has to be extra cautious in interpreting the outcome.After more than a decade of copious positive case-control studiesof angiotensinogen and angiotensin-converting enzyme gene polymorphismsin more than a dozen clinical conditions, we are hardly closerto the truth. Having stated the caveats and limitations withthis technique, there is no doubt that association studies arereadily and easily performed.
Figure 3. The commonly encountered association study. Cases and controls are defined by phenotypic criteria selected by the investigator. The frequency of a polymorphic candidate gene with two or more alleles is determined in the two populations. A typical positive result is shown with allele A1 more common in affected cases (70%) versus controls (30%). This statistical finding can be due to several possible reasons. (1) Gene A is a disease locus and allele A1 is a mutation that causes hypercalciuria. (2) Gene A is closely situated to gene B and therefore recombines rarely (linkage disequilibrium). In the two study populations, allele A1 is co-segregating with the disease gene B, so A1 seems to occur with higher frequency in the cases. (3) Neither gene A nor any other gene in the region is contributing to hypercalciuria. Allele A1 is occurring with higher frequency in the affect versus unaffected individuals because the two populations are not comparable.
The wealth of information derived from monogenic human diseasesand rodents with single gene deletions provides a valuable databasefor candidate genes to be screened in humans. Genes along thevitamin D axis were examined. Small studies have shown associationof restriction site polymorphisms with one form of phenotypicparameter or another (141–143). However, several otherstudies have found no difference in the phenotype of vitaminD receptor expression, induction, allelic frequencies, codingregion sequence between controls, and well characterized hyperabsorptivehypercalciuric stone formers (144,145). The closest positiveoutcome has been the identification of potential susceptibilitylocus in the vicinity of the vitamin D receptor gene in 47 French-Canadiankindreds using sib-pair analysis (146). One must exercise utmostcaution and rigor before accepting such data as proof of thehypothesis.
Other candidate genes were also pursued. Apart from its knownimplication in Bartter syndrome and autosomal dominant hypocalcemia,the calcium sensing receptor (CaSR) is a candidate gene formultigenic hypercalciuria and bone resorption. One study showedassociative correlation between a single amino acid change (unknownwhether it is a polymorphism or a mutation) and the phenotype(147); there were no point mutations in seven families withidiopathic hypercalciuria (148). In 55 French-Canadian pedigrees,no association was found between CaSR locus and idiopathic hypercalciuriaand calcium nephrolithiasis (149). Direct sequencing of theClC-5 chloride channel was negative in a case-control study(150). Thus far, association studies with candidate genes havenot been particularly informative in genetic hypercalciuria.
Linkage
Linkage is a powerful alternative and complement to associationstudies. As opposed to association, linkage detects genotype-phenotypeco-segregation within families and has been the workhorse fordisease-locus discovery in monogenic diseases (151,152). TheHôpital Maisonneuve-Rosemont group from Montreal has usednonparametric (i.e., a model-free test to avoid a priori assumptionof a particular inheritance pattern) linkage analysis in largenumbers of French-Canadian concordant affected sib-pairs ona large number of candidate genes including the vitamin D receptor(146); 1--hydroxylase (153); the CaSR (148); and crystallizationmodifiers such as osteopontin, Tamm-Horsefall protein, and osteocalcin-relatedgene (154).
Instead of candidate genes, Reed et al. (155) performed a linkageanalysis of three large kindreds with absorptive hypercalciuriausing nonparametric testing. This was the first successful genome-widesearch in this complex trait. One large kindred contributedto the particularly high logarithm of odds score of >12 forthe region in 1q23.4 to 24, which contains several known genesand a number of putative genes. After several known genes weresequenced and no sequence differences were found in affectedversus unaffected individuals, a hypothetical protein with adenylylcyclase motifs that spans 104.5 kB with 33 exons (Gen Bank AL035122)was identified (156). During the course of the work, a homologousrat gene coding for a soluble adenylyl cyclase (sAC) was clonedby Buck et al. (157). The human ortholog of sAC is polymorphicwith 18 base substitutions (156). When this gene was examinedoutside the original kindreds, five sequence variations occurredwith increased frequency in the hypercalciuria population comparedwith normal volunteers, and the number of base substitutionsseems to correlate with lower spinal bone density and intestinalcalcium absorption (156,158), two cardinal features of the absorptivecalciuria syndrome. The pathophysiologic relationship betweenthe human ortholog of sAC and intestinal calcium absorptionremains to be determined. The efforts on human familial hypercalciuricnephrolithiasis has yielded some encouraging but far-from-groundbreakinginformation.
Animal Models
A large number of rodent models of complex traits have beengenerated in the past several decades (159,160), and they arecatalogued at the Rat Genome Database (http://www.rgd.mcw.edu).These models are instrumental for both geneticists and physiologists.The best animal model of polygenic hypercalciuria to date isthe genetic hypercalciuria stone-forming rats (GHS rats) createdby Bushinsky et al. (161) more than one and a half decades ago.That one can breed the physiologic extreme end of calciuriain "normal" rats to a hypercalciuric state with complex pathophysiologylends credence to the polygenic determinants of calcium excretion.Although the physiology differs somewhat from those describedin humans, the GHS rat definitely shows multiorgan pathophysiologywith intestinal hyperabsorption, increased bone calcium mobilization,and primary renal leak, a "full house" coverage of Figure 1.The well-characterized pathophysiology has been summarized inseveral recent reviews (6,162,163).
There are two reasons that the study of these animals is moreutilitarian than humans in studying complex traits. In additionto a valuable source of pathophysiology, a major reason forhaving such strains is identification of disease susceptibilityloci using breeding experiments. A region of the chromosome(perhaps containing one gene) that determines the magnitudeof a continuous trait is called a quantitative trait locus (QTL).With the a priori assumption that the trait is genetic, onecrosses two inbred strains (hypercalciuric and normocalciuric)to produce recombinant strains followed by backcrosses withone of the original strains until one can link the genetic loci(QTL) being delivered, to the phenotypic trait (hypercalciuria).One works on the implicit hypothesis that the QTL confers thehypercalciuria. The loci identification will be followed bymapping of the genes. Hoopes et al. (164) reported on severalQTL that are linked to the hypercalciuria in the GHS rats. Althoughnone of the QTL are in regions that harbor the usual set ofsuspect candidate genes, these are indeed encouraging results,and the hurdle to surmount now is to proceed from locus to gene.
Although traditional QTL approaches are powerful, a recent advancein this field that may revolutionize the dissection of rodentpolygenic traits is the development of chromosome substitutionstrains (165,166). In classical QTL studies, one crosses twoinbred strains with differing phenotypes to create hybrids thatthen are intercrossed and/or backcrossed eventually to generaterecombinant inbred or cogenic strains that carry portions ofthe genome of the original parental disease-carrying strainin the genetic background of the nonaffected parental strain(Figure 4). Because these strains are hypercalciuric, linkagecan be performed to identify loci that segregate with the QTLof interest. With knowledge of the loci, finer structural mappingcan be performed to eventually hone in on the genes. In thechromosomal substitution approach, instead of pieces of DNArandomly scattered through the genome, a panel in which entirechromosomes from one strain are replaced with the correspondingchromosome from another is created (165,166) (Figure 4) in definedand uniform genetic background. The latter approach is morechallenging because the panel of strains has to be generatedand is limited by large DNA regions (entire chromosome versusthe typical 20 cM of QTL). However, no crosses (usually a colossalnumber for QTL) need to be performed and genotyping is not requiredbecause the segregation of the genome is known from the waythe panel is generated.
Figure 4. Comparison of classical quantitative trait loci (QTL) identification to chromosome-specific substitution approach. In classical QTL approach, two inbred strains with differing phenotypes are intercrossed and/or backcrossed to generate recombinant inbred or cogenic strains that carry portions of the genome of the disease-carrying strain in the genetic background of the nonaffected parental strain. Linkage is performed to identify loci that segregate with the QTL of interest. In the chromosomal substitution approach, a panel in which entire chromosomes from one strain are replaced with the corresponding chromosome from another is created (165,166).
Genetic manipulations in rodents are usually performed to examinethe whole organism phenotype of the disruption of a gene productor to create a model of known monogenic diseases for detailedinvasive phenotypic characterization that is not possible inhumans. With the ever-expanding list of artificial rodent strainswith single gene deletions with hypercalciuria (Table 5), anotherapproach will be to take the phenotype of a knockout mouse andsearch large databases such as stone registry to search forhumans who "resemble" the mouse with deletion of gene X andsequence gene X in those individuals.
In summary, hypercalciuria is an important risk factor for nephrolithiasis,and its genetic component is a complex trait that defies simplemethods of human genetics. A combination of approaches are requiredto advance this field, including detailed proximal phenotypedefinition and database management and sharing on a large scale,continued efforts in studying monogenic causes of hypercalciuriain human and genetically altered rodents, and application ofstandard and new polygenic approaches in both human and rodentmodels of hypercalciuria.
Acknowledgments
The authors are supported by the National Institutes of Health(R01-DK48482 and P01-DK to O.W.M.), the Swiss National ScienceFoundation (O.B.), and the National Kidney Foundation (O.B.).
We are grateful to the helpful discussion and advice by Dr.Charles Pak in the preparation of this manuscript.
Footnotes
Published online ahead of print. Publication date availableat www.jasn.org.
Zerwekh JE, Reed-Gitomer BY, Pak CY: Pathogenesis of hypercalciuric nephrolithiasis.
Endocrinol Metab Clin North Am 31
: 869
–884, 2002[CrossRef][Medline]
Bushinsky DA: Kidney stones.
Adv Intern Med 47
: 219
–238, 2001[Medline]
Williams RJP: Calcium chemistry and its relation to biologic function.
Symp Soc Exp Biol 30
: 1
–17, 1976
Fincham AG, Luo W, Moradian-Oldak J, Paine ML, Snead ML, Zeucher-David M: Enamel biomineralization: The assembly and disassembly of the protein extracellular organic matrix. In:
Development, Function, and Evolution of Teeth, edited by Teaford M, Smith MM, and Ferguson M, Cambridge University Press, Cambridge, England, 2000
Stayton PS, Drobny GP, Shaw WJ, Long JR, Gilbert M: Molecular recognition at the protein-hydroxyapatite interface.
Crit Rev Oral Biol Med 14
: 370
–376, 2003[Abstract/Free Full Text]
Drobny GP, Long JR, Karlsson T, Shaw W, Popham J, Oyler N, Bower P, Stringer J, Gregory D, Mehta M, Stayton PS: Structural studies of biomaterials using double-quantum solid-state NMR spectroscopy.
Annu Rev Phys Chem 54
: 531
–571, 2003[CrossRef][Medline]
Waite JH, Qin X: Polyphosphoprotein from the adhesive pads of Mytilus edulis.
Biochemistry 40
: 2887
–2893, 2001[CrossRef][Medline]
Hunter J:
The Natural History of the Human Teeth: Explaining Their Structure, Use, Formation, and Growth, London, Johnson, 1771
Stapleton FB: Hematuria associated with hypercalciuria and hyperuricosuria: A practical approach.
Pediatr Nephrol 8
: 756
–761, 1994[CrossRef][Medline]
Aceto G, Penza R, Coccioli MS, Palumbo F, Cresta L, Cimador M, Chiozza ML, Caione P: Enuresis subtypes based on nocturnal hypercalciuria: A multicenter study.
J Urol 170
: 1670
–1673, 2003[CrossRef][Medline]
Sayer JA, Carr G, Simmons NL: Nephrocalcinosis: Molecular insights into calcium precipitation within the kidney.
Clin Sci (Lond) 106
: 549
–561, 2004[Medline]
Pak CYC, Adams-Huet B, Poindexter JR, Pearle MS, Peterson RD, Moe OW: Relative effect of urinary calcium and oxalate on saturation of calcium oxalate.
Kidney Int 66
: 2032
–2037, 2004[CrossRef][Medline]
Pak CY, Kaplan R, Bone H, Townsend J, Waters O: A simple test for the diagnosis of absorptive, resorptive and renal hypercalciurias.
N Engl J Med 292
: 497
–500, 1975[Abstract]
Hoenderop JG, van Leeuwen JP, van der Eerden BC, Kersten FF, van der Kemp AW, Merillat AM, Waarsing JH, Rossier BC, Vallon V, Hummler E, Bindels RJ: Renal Ca2+ wasting, hyperabsorption, and reduced bone thickness in mice lacking TRPV5.
J Clin Invest 112
: 1906
–1914, 2003[CrossRef][Medline]
Coe FL, Parks JH, Moore ES: Familial idiopathic hypercalciuria.
N Engl J Med 300
: 337
–340, 1979[Abstract]
Favus MJ: Familial forms of hypercalciuria.
J Urol 141
: 719
–722, 1989[Medline]
Pak CY, McGuire J, Peterson R, Britton F, Harrod MJ: Familial absorptive hypercalciuria in a large kindred.
J Urol 126
: 717
–719, 1981[Medline]
Curhan GC, Willett WC, Rimm EB, Stampfer MJ: Family history and risk of kidney stones.
J Am Soc Nephrol 8
: 1568
–1573, 1997[Abstract]
Harangi F, Mehes K: Family investigations in idiopathic hypercalciuria.
Eur J Pediatr 152
: 64
–68, 1993[CrossRef][Medline]
Breslau NA, McGuire JL, Zerwekh JE, Pak CY: The role of dietary sodium on renal excretion and intestinal absorption of calcium and on vitamin D metabolism.
J Clin Endocrinol Metab 55
: 369
–373, 1982[Abstract]
Lemann J Jr, Lennon EJ: Role of diet, gastrointestinal tract and bone in acid-base homeostasis.
Kidney Int 1
: 275
–279, 1972[Medline]
Sontag MK, Accurso FJ: Gene modifiers in pediatrics: Application to cystic fibrosis.
Adv Pediatr 51
: 5
–36, 2004[Medline]
Nagel RL, Steinberg MH: Role of epistatic (modifier) genes in the modulation of the phenotypic diversity of sickle cell anemia.
Pediatr Pathol Mol Med 20
: 123
–136, 2001[CrossRef][Medline]
Wilson PD: Polycystic kidney disease.
N Engl J Med 350
: 151
–164, 2004[Free Full Text]
Guay-Woodford LM: Murine models of polycystic kidney disease: Molecular and therapeutic insights.
Am J Physiol 285
: F1034
–F1049, 2003
Saarela T, Simila S, Koivisto M: Hypercalcemia and nephrocalcinosis in patients with congenital lactase deficiency.
J Pediatr 127
: 920
–923, 1995[CrossRef][Medline]
Belmont JW, Reid B, Taylor W, Baker SS, Moore WH, Morriss MC, Podrebarac SM, Glass N, Schwartz ID: Congenital sucrase-isomaltase deficiency presenting with failure to thrive, hypercalcemia, and nephrocalcinosis.
BMC Pediatr 2
: 4
, 2002[CrossRef][Medline]
Starnes CW, Welsh JD: Intestinal sucrase-isomaltase deficiency and renal calculi.
N Engl J Med 282
: 1023
–1024, 1970
Tasic V, Slaveska N, Blau N, Santer R: Nephrolithiasis in a child with glucose-galactose malabsorption.
Pediatr Nephrol 19
: 244
–246, 2004[CrossRef][Medline]
Abdullah AM, el-Mouzan MI, el Shiekh OK, al Mazyad A: Congenital glucose-galactose malabsorption in Arab children.
J Pediatr Gastroenterol Nutr 23
: 561
–564, 1996[CrossRef][Medline]
Abdullah AM, Abdullah MA, Abdurrahman MB, al Husain MA: Glucose-galactose malabsorption with renal stones in a Saudi child.
Ann Trop Paediatr 12
: 327
–329, 1992[Medline]
Kim do K, Kanai Y, Matsuo H, Kim JY, Chairoungdua A, Kobayashi Y, Enomoto A, Cha SH, Goya T, Endou H: The human T-type amino acid transporter-1: Characterization, gene organization, and chromosomal location.
Genomics 79
: 95
–103, 2002[CrossRef][Medline]
Drummond KN, Michael AF, Ulstrom RA, Good RA: The blue diaper syndrome: Familial hypercalcemia with nephrocalcinosis and indicanuria; a new familial disease, with definition of the metabolic abnormality.
Am J Med 37
: 28
–48, 1964
Prie D, Huart V, Bakouh N, Planelles G, Dellis O, Gerard B, Hulin P, Benque-Blanchet F, Silve C, Grandchamp B, Friedlander G: Nephrolithiasis and osteoporosis associated with hypophosphatemia caused by mutations in the type 2a sodium-phosphate cotransporter.
N Engl J Med 347
: 983
–991, 2002[Abstract/Free Full Text]
van den Heuvel L, Op de Koul K, Knots E, Knoers N, Monnens L: Autosomal recessive hypophosphataemic rickets with hypercalciuria is not caused by mutations in the type II renal sodium/phosphate cotransporter gene.
Nephrol Dial Transplant 16
: 48
–51, 2001[Free Full Text]
Jones A, Tzenova J, Frappier D, Crumley M, Roslin N, Kos C, Tieder M, Langman C, Proesmans W, Carpenter T, Rice A, Anderson D, Morgan K, Fujiwara T, Tenenhouse H: Hereditary hypophosphatemic rickets with hypercalciuria is not caused by mutations in the Na/Pi cotransporter NPT2 gene.
J Am Soc Nephrol 12
: 507
–514, 2001[Abstract/Free Full Text]
Tieder M, Modai D, Samuel R, Arie R, Halabe A, Bab I, Gabizon D, Liberman UA: Hereditary hypophosphatemic rickets with hypercalciuria.
N Engl J Med 312
: 611
–617, 1985[Abstract]
Mathias RS: Rickets in an infant with Williams syndrome.
Pediatr Nephrol 14
: 489
–492, 2000[CrossRef][Medline]
Shultz PK, Strife JL, Strife CF, McDaniel JD: Hyperechoic renal medullary pyramids in infants and children.
Radiology 181
: 163
–167, 1991[Abstract/Free Full Text]
Sforzini C, Milani D, Fossali E, Barbato A, Grumieri G, Bianchetti MG, Selicorni A: Renal tract ultrasonography and calcium homeostasis in Williams-Beuren syndrome.
Pediatr Nephrol 17
: 899
–902, 2002[CrossRef][Medline]
Imamura K, Tonoki H, Wakui K, Fukushima Y, Sasaki S, Yausda K, Takekoshi Y, Tochimaru H: 4q33-qter deletion and absorptive hypercalciuria: Report of two unrelated girls.
Am J Med Genet 78
: 52
–54, 1998[CrossRef][Medline]
Giuffre M, La Placa S, Carta M, Cataliotti A, Marino M, Piccione M, Pusateri F, Meli F, Corsello G: Hypercalciuria and kidney calcifications in terminal 4q deletion syndrome: Further evidence for a putative gene on 4q.
Am J Med Genet 126A
: 186
–190, 2004[CrossRef]
Proesmans W, De Cock P, Eyskens B: A toddler with Down syndrome, hypercalcaemia, hypercalciuria, medullary nephrocalcinosis and renal failure.
Pediatr Nephrol 9
: 112
–114, 1995[CrossRef][Medline]
Andreoli SP, Revkees S, Bull M: Hypercalcemia, hypercalciuria, medullar nephrocalcinosis, and renal insufficiency in a toddler with Down syndrome.
Pediatr Nephrol 9
: 673
, 1995[CrossRef][Medline]
Manz F: A toddler with Down syndrome, hypercalcaemia, hypercalciuria, medullary nephrocalcinosis and renal failure.
Pediatr Nephrol 10
: 251
, 1996[CrossRef][Medline]
Proesmans W: Hypercalcaemia, hypercalciuria and nephrocalcinosis in Down syndrome.
Pediatr Nephrol 10
: 543
–544, 1996[Medline]
Cobenas C, Spizzirri F, Zanetta D: Another toddler with Down syndrome, nephrocalcinosis, hypercalcemia, and hypercalciuria.
Pediatr Nephrol 12
: 432
, 1998[Medline]
Ramage IJ, Durkan A, Walker K, Beattie TJ: Hypercalcaemia in association with trisomy 21 (Down’s syndrome).
J Clin Pathol 55
: 543
–544, 2002[Abstract/Free Full Text]
Baptista F, Varela A, Sardinha LB: Bone mineral mass in males and females with and without Down syndrome.
Osteoporos Int 2004
(Epub ahead of print)
Angelopoulou N, Souftas V, Sakadamis A, Mandroukas K: Bone mineral density in adults with Down’s syndrome.
Eur Radiol 9
: 648
–651, 1999[CrossRef][Medline]
Chines A, Petersen DJ, Schranck FW, Whyte MP: Hypercalciuria in children severely affected with osteogenesis imperfecta.
J Pediatr 119
: 51
–57, 1991[CrossRef][Medline]
Chines A, Boniface A, McAlister W, Whyte M: Hypercalciuria in osteogenesis imperfecta: A follow-up study to assess renal effects.
Bone 16
: 333
–339, 1995[Medline]
Ammenti A, Nitsch M: Hypercalciuria in osteogenesis imperfecta type I.
Klin Padiatr 215
: 283
–285, 2003[CrossRef][Medline]
Thakker RV: Genetics of endocrine and metabolic disorders: Parathyroid.
Rev Endocr Metab Disord 5
: 37
–51, 2004[CrossRef][Medline]
Nishiyama S, Tomoeda S, Inoue F, Ohta T, Matsuda I: Self-limited neonatal familial hyperparathyroidism associated with hypercalciuria and renal tubular acidosis in three siblings.
Pediatrics 86
: 421
–427, 1990[Abstract/Free Full Text]
Kessel D, Hall CM, Shaw DG: Two unusual cases of nephrocalcinosis in infancy.
Pediatr Radiol 22
: 470
–471, 1992[CrossRef][Medline]
Kruse K, Schutz C: Calcium metabolism in the Jansen type of metaphyseal dysplasia.
Eur J Pediatr 152
: 912
–915, 1993[CrossRef][Medline]
Barcia JP, Strife CF, Langman CB: Infantile hypophosphatasia: Treatment options to control hypercalcemia, hypercalciuria, and chronic bone demineralization.
J Pediatr 130
: 825
–828, 1997[Medline]
Lala R, Matarazzo P, Andreo M, Defilippi C, de Sanctis C: Impact of endocrine hyperfunction and phosphate wasting on bone in McCune-Albright syndrome.
J Pediatr Endocrinol Metab 15[Suppl 3]
: 913
–920, 2002
Kirk JM, Brain CE, Carson DJ, Hyde JC, Grant DB: Cushing’s syndrome caused by nodular adrenal hyperplasia in children with McCune-Albright syndrome.
J Pediatr 134
: 789
–792, 1999[CrossRef][Medline]
Fisher SE, Black GC, Lloyd SE, Hatchwell E, Wrong O, Thakker RV, Craig IW: Isolation and partial characterization of a chloride channel gene which is expressed in kidney and is a candidate for Dent’s disease (an X-linked hereditary nephrolithiasis).
Hum Mol Genet 3
: 2053
–2059, 1994
Thakker RV: Molecular pathology of renal chloride channels in Dent’s disease and Bartter’s syndrome.
Exp Nephrol 8
: 351
–360, 2000[CrossRef][Medline]
Sliman GA, Winters WD, Shaw DW, Avner ED: Hypercalciuria and nephrocalcinosis in the oculocerebrorenal syndrome.
J Urol 153
: 1244
–1246, 1995[CrossRef][Medline]
Charnas LR, Bernardini I, Rader D, Hoeg JM, Gahl WA: Clinical and laboratory findings in the oculocerebrorenal syndrome of Lowe, with special reference to growth and renal function.
N Engl J Med 324
: 1318
–1325, 1991[Abstract]
Meier W, Blumberg A, Imahorn W, De Luca F, Wildberger H, Oetliker O: Idiopathic hypercalciuria with bilateral macular colobomata: A new variant of oculo-renal syndrome.
Helv Paediatr Acta 34
: 257
–269, 1979[Medline]
Litin RB, Randall RV, Goldstein NP, Power MH, Diessner GR: Hypercalciuria in hepatolenticular degeneration (Wilson’s disease).
Am J Med Sci 238
: 614
–620, 1959[Medline]
Randall RV, Goldstein NP, Gross JB, Rosevear JW: Hypercalciuria in hepatolenticular degeneration (Wilson’s disease).
Am J Med Sci 252
: 715
–720, 1966[Medline]
Carpenter TO, Carnes DL Jr, Anast CS: Hypoparathyroidism in Wilson’s disease.
N Engl J Med 309
: 873
–877, 1983[Abstract]
Azizi E, Eshel G, Aladjem M: Hypercalciuria and nephrolithiasis as a presenting sign in Wilson disease.
Eur J Pediatr 148
: 548
–549, 1989[CrossRef][Medline]
Itami N, Akutsu Y, Tochimaru H, Takekoshi Y: Hypercalciuria and nephrolithiasis in Wilson disease.
Eur J Pediatr 149
: 145
, 1989[CrossRef][Medline]
Hoppe B, Neuhaus T, Superti-Furga A, Forster I, Leumann E: Hypercalciuria and nephrocalcinosis, a feature of Wilson’s disease.
Nephron 65
: 460
–462, 1993[Medline]
Kalra V, Mahajan S, Kesarwani PK: Rare presentation of Wilson’s disease: A case report.
Int Urol Nephrol 36
: 289
–291, 2004[CrossRef][Medline]
Hegedus D, Ferencz V, Lakatos PL, Meszaros S, Lakatos P, Horvath C, Szalay F: Decreased bone density, elevated serum osteoprotegerin, and beta-cross-laps in Wilson disease.
J Bone Miner Res 17
: 1961
–1967, 2002[CrossRef][Medline]
Laine J, Salo MK, Krogerus L, Karkkainen J, Wahlroos O, Holmberg C: The nephropathy of type I tyrosinemia after liver transplantation.
Pediatr Res 37
: 640
–645, 1995[Medline]
Restaino I, Kaplan BS, Stanley C, Baker L: Nephrolithiasis, hypocitraturia, and a distal renal tubular acidification defect in type 1 glycogen storage disease.
J Pediatr 122
: 392
–396, 1993[Medline]
Weinstein DA, Somers MJ, Wolfsdorf JI: Decreased urinary citrate excretion in type 1a glycogen storage disease.
J Pediatr 138
: 378
–382, 2001[CrossRef][Medline]
Talente GM, Coleman RA, Alter C, Baker L, Brown BI, Cannon RA, Chen YT, Crigler JF Jr, Ferreira P, Haworth JC: Glycogen storage disease in adults.
Ann Intern Med 120
: 218
–226, 1994[Abstract/Free Full Text]
Simon DB, Lu Y, Choate KA, Velazquez H, Al-Sabban E, Praga M, Casari G, Bettinelli A, Colussi G, Rodriguez-Soriano J, McCredie D, Milford D, Sanjad S, Lifton RP: Paracellin-1, a renal tight junction protein required for paracellular Mg2+ resorption.
Science 285
: 103
–106, 1999[Abstract/Free Full Text]
Praga M, Vara J, Gonzalez-Parra E, Andres A, Alamo C, Araque A, Ortiz A, Rodicio JL: Familial hypomagnesemia with hypercalciuria and nephrocalcinosis.
Kidney Int 47
: 1419
–1425, 1995[Medline]
Ulmann A, Hadj S, Lacour B, Bourdeau A, Bader C: Renal magnesium and phosphate wastage in a patient with hypercalciuria and nephrocalcinosis: Effect of oral phosphorus and magnesium supplements.
Nephron 40
: 83
–87, 1985[Medline]
Weber S, Schneider L, Peters M, Misselwitz J, Ronnefarth G, Boswald M, Bonzel KE, Seeman T, Sulakova T, Kuwertz-Broking E, Gregoric A, Palcoux JB, Tasic V, Manz F, Scharer K, Seyberth HW, Konrad M: Novel paracellin-1 mutations in 25 families with familial hypomagnesemia with hypercalciuria and nephrocalcinosis.
J Am Soc Nephrol 12
: 1872
–1881, 2001[Abstract/Free Full Text]
Simon DB, Karet FE, Hamdan JM, DiPietro A, Sanjad SA, Lifton RP: Bartter’s syndrome, hypokalaemic alkalosis with hypercalciuria, is caused by mutations in the Na-K-2Cl cotransporter NKCC2.
Nat Genet 13
: 183
–188, 1996[CrossRef][Medline]
Simon DB, Karet FE, Rodriguez-Soriano J, Hamdan JH, DiPietro A, Trachtman H, Sanjad SA, Lifton RP: Genetic heterogeneity of Bartter’s syndrome revealed by mutations in the K+ channel, ROMK.
Nat Genet 14
: 152
–156, 1996[CrossRef][Medline]
Simon DB, Bindra RS, Mansfield TA, Nelson-Williams C, Mendonca E, Stone R, Schurman S, Nayir A, Alpay H, Bakkaloglu A, Rodriguez-Soriano J, Morales JM, Sanjad SA, Taylor CM, Pilz D, Brem A, Trachtman H, Griswold W, Richard GA, John E, Lifton RP: Mutations in the chloride channel gene, CLCNKB, cause Bartter’s syndrome type III.
Nat Genet 17
: 171
–178, 1997[CrossRef][Medline]
Birkenhager R, Otto E, Schurmann MJ, Vollmer M, Ruf EM, Maier-Lutz I, Beekmann F, Fekete A, Omran H, Feldmann D, Milford DV, Jeck N, Konrad M, Landau D, Knoers NV, Antignac C, Sudbrak R, Kispert A, Hildebrandt F: Mutation of BSND causes Bartter syndrome with sensorineural deafness and kidney failure.
Nat Genet 29
: 310
–314, 2001[CrossRef][Medline]
Watanabe S, Fukumoto S, Chang H, Takeuchi Y, Hasegawa Y, Okazaki R, Chikatsu N, Fujita T: Association between activating mutations of calcium-sensing receptor and Bartter’s syndrome.
Lancet 360
: 692
–694, 2002[CrossRef][Medline]
Vargas-Poussou R, Huang C, Hulin P, Houillier P, Jeunemaitre X, Paillard M, Planelles G, Dechaux M, Miller RT, Antignac C: Functional characterization of a calcium-sensing receptor mutation in severe autosomal dominant hypocalcemia with a Bartter-like syndrome.
J Am Soc Nephrol 13
: 2259
–2266, 2002[Abstract/Free Full Text]
Madrigal G, Saborio P, Mora F, Rincon G, Guay-Woodford LM: Bartter syndrome in Costa Rica: A description of 20 cases.
Pediatr Nephrol 11
: 296
–301, 1997[CrossRef][Medline]
Pollak MR, Brown EM, Estep HL, McLaine PN, Kifor O, Park J, Hebert SC, Seidman CE, Seidman JG: Autosomal dominant hypocalcaemia caused by a Ca(2+)-sensing receptor gene mutation.
Nat Genet 8
: 303
–307, 1994[CrossRef][Medline]
Pearce SH, Williamson C, Kifor O, Bai M, Coulthard MG, Davies M, Lewis-Barned N, McCredie D, Powell H, Kendall-Taylor P, Brown EM, Thakker RV: A familial syndrome of hypocalcemia with hypercalciuria due to mutations in the calcium-sensing receptor.
N Engl J Med 335
: 1115
–1122, 1996[Abstract/Free Full Text]
Hendy GN, Minutti C, Canaff L, Pidasheva S, Yang B, Nouhi Z, Zimmerman D, Wei C, Cole DE: Recurrent familial hypocalcemia due to germline mosaicism for an activating mutation of the calcium-sensing receptor gene.
J Clin Endocrinol Metab 88
: 3674
–3681, 2003[Abstract/Free Full Text]
Arnold A, Horst SA, Gardella TJ, Baba H, Levine MA, Kronenberg HM: Mutation of the signal peptide-encoding region of the preproparathyroid hormone gene in familial isolated hypoparathyroidism.
J Clin Invest 86
: 1084
–1087, 1990
Sunthornthepvarakul T, Churesigaew S, Ngowngarmratana S: A novel mutation of the signal peptide of the preproparathyroid hormone gene associated with autosomal recessive familial isolated hypoparathyroidism.
J Clin Endocrinol Metab 84
: 3792
–3796, 1971
Parkinson DB, Thakker RV: A donor splice site mutation in the parathyroid hormone gene is associated with autosomal recessive hypoparathyroidism.
Nat Genet 1
: 149
–152, 1992[CrossRef][Medline]
Ding C, Buckingham B, Levine MA: Familial isolated hypoparathyroidism caused by a mutation in the gene for the transcription factor GCMB.
J Clin Invest 108
: 1215
–1220, 2001[CrossRef][Medline]
Wilson FH, Disse-Nicodeme S, Choate KA, Ishikawa K, Nelson-Williams C, Desitter I, Gunel M, Milford DV, Lipkin GW, Achard JM, Feely MP, Dussol B, Berland Y, Unwin RJ, Mayan H, Simon DB, Farfel Z, Jeunemaitre X, Lifton RP: Human hypertension caused by mutations in WNK kinases.
Science 293
: 1107
–1112, 2001[Abstract/Free Full Text]
Achard JM, Warnock DG, Disse-Nicodeme S, Fiquet-Kempf B, Corvol P, Fournier A, Jeunemaitre X: Familial hyperkalemic hypertension: Phenotypic analysis in a large family with the WNK1 deletion mutation.
Am J Med 114
: 495
–498, 2003[CrossRef][Medline]
Karet FE, Finberg KE, Nelson RD, Nayir A, Mocan H, Sanjad SA, Rodriguez-Soriano J, Santos F, Cremers CW, Di Pietro A, Hoffbrand BI, Winiarski J, Bakkaloglu A, Ozen S, Dusunsel R, Goodyer P, Hulton SA, Wu DK, Skvorak AB, Morton CC, Cunningham MJ, Jha V, Lifton RP: Mutations in the gene encoding B1 subunit of H+-ATPase cause renal tubular acidosis with sensorineural deafness.
Nat Genet 21
: 84
–90, 1999[CrossRef][Medline]
Smith AN, Skaug J, Choate KA, Nayir A, Bakkaloglu A, Ozen S, Hulton SA, Sanjad SA, Al-Sabban EA, Lifton RP, Scherer SW, Karet FE: Mutations in ATP6N1B, encoding a new kidney vacuolar proton pump 116-kD subunit, cause recessive distal renal tubular acidosis with preserved hearing.
Nat Genet 26
: 71
–75, 2000[CrossRef][Medline]
Karet FE, Gainza FJ, Gyory AZ, Unwin RJ, Wrong O, Tanner MJ, Nayir A, Alpay H, Santos F, Hulton SA, Bakkaloglu A, Ozen S, Cunningham MJ, di Pietro A, Walker WG, Lifton RP: Mutations in the chloride-bicarbonate exchanger gene AE1 cause autosomal dominant but not autosomal recessive distal renal tubular acidosis.
Proc Natl Acad Sci U S A 95
: 6337
–6342, 1998[Abstract/Free Full Text]
Bruce LJ, Cope DL, Jones GK, Schofield AE, Burley M, Povey S, Unwin RJ, Wrong O, Tanner MJ: Familial distal renal tubular acidosis is associated with mutations in the red cell anion exchanger (Band 3, AE1) gene.
J Clin Invest 100
: 1693
–1707, 1997[Medline]
Sly WS, Hewett-Emmett D, Whyte MP, Yu YS, Tashian RE: Carbonic anhydrase II deficiency identified as the primary defect in the autosomal recessive syndrome of osteopetrosis with renal tubular acidosis and cerebral calcification.
Proc Natl Acad Sci U S A 80
: 2752
–2756, 1983[Abstract/Free Full Text]
Ismail EA, Abul Saad S, Sabry MA: Nephrocalcinosis and urolithiasis in carbonic anhydrase II deficiency syndrome.
Eur J Pediatr 156
: 957
–962, 1997[CrossRef][Medline]
Noblins M, Kleinknecht D, Dommergues JP, Nazaret C, Garay RP, Jullien M, Guillot M, Fries D, Charpentier B: Liddle syndrome (or pseudo-hyperaldosteronism). Long-term development and erythrocyte potassium flow study in 4 cases.
Arch Fr Pediatr 49
: 685
–691, 1992[Medline]
Katz SM, Krueger LJ, Falkner B: Microscopic nephrocalcinosis in cystic fibrosis.
N Engl J Med 319
: 263
–266, 1988[Abstract]
Ozcelik U, Besbas N, Gocmen A, Akata D, Akhan O, Ozguc M, Kiper N: Hypercalciuria and nephrocalcinosis in cystic fibrosis patients.
Turk J Pediatr 46
: 22
–27, 2004[Medline]
Cetin T, Oktenli C, Ozgurtas T, Yenicesu M, Sanisoglu SY, Oguz Y, Yildiz O, Kurt I, Musabak U, Bulucu F, Kocar IH: Renal tubular dysfunction in beta-thalassemia minor.
Am J Kidney Dis 42
: 1164
–1168, 2003[CrossRef][Medline]
Mahachoklertwattana P, Chuansumrit A, Sirisriro R, Choubtum L, Sriphrapradang A, Rajatanavin R: Bone mineral density, biochemical and hormonal profiles in suboptimally treated children and adolescents with beta-thalassaemia disease.
Clin Endocrinol 58
: 273
–279, 2003[CrossRef][Medline]
Goldman M, Shuman C, Weksberg R, Rosenblum ND: Hypercalciuria in Beckwith-Wiedemann syndrome.
J Pediatr 142
: 206
–208, 2003[Cross
Top
Abstract
Introduction
-globin causing very mild or very severe sickle cell anemia (30). There is no doubt that modifier genes are at work. Consider the idealized situation in the laboratory, where one specifically deletes one and only one gene in a murine model with a completely homogeneous (as far as one can tell) genetic background. This is probably the closest example that one can fathom of a pure monogenic disorder. It is already well known that the same gene knockout in the background of different strains can have completely different effects. As the investigator delights with the emergence of the good news that 80% of the null mice survives and hence can be studied, one should wonder about the effect of the gene deletion on the 20% of the mortal ones. Penetrance of a singular gene may be small or even negligible, but multiple genes in concert can have a significant impact on the phenotype. Envision five hypothetical genes that have an impact on urinary calcium excretion. Each of these genes is polymorphic in Homo sapiens with multiple fully functional alleles that have small quantitative differences in function. For instance, gene product 1 influences intestinal calcium absorption, gene product 2 modifies the ability of dietary acid to release calcium from bone, gene product 3 controls paracellular calcium permeability in the thick ascending limb, gene product 4 influences calcium absorption in the distal convoluted tubule, and gene product 5 alters the sensitivity of the renal calcium sensing receptor to plasma calcium concentration. If each of these gene products contributes to a trivial 0.5 mmol/d (20 mg) increase in urinary calcium excretion, then inheriting all five predisposing alleles can theoretically raise urinary calcium by 2.5 mmol/d (100 mg). 
-hydroxylase (153); the CaSR (148); and crystallization modifiers such as osteopontin, Tamm-Horsefall protein, and osteocalcin-related gene (154). 
