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J Am Soc Nephrol 12: 2348–2357, 2001

Familial Juvenile Hyperuricemic Nephropathy and Autosomal Dominant Medullary Cystic Kidney Disease Type 2: Two Facets of the Same Disease?

*Center for Human Genetics, §Department of Pathology, and ࿣Division of Nephrology, Catholic University of Louvain, Brussels, Belgium; †University Children’s Hospital, Freiburg, Germany; and ‡Department of Nephrology, Princesse Paola Hospital, Aye, Belgium.

Abstract. Familial juvenile hyperuricemic nephropathy (FJHN) is an autosomal dominant disorder heralded by hyperuricemia during childhood; it is characterized by chronic interstitial nephritis, with marked thickening of tubular basement membranes, and leads to progressive renal failure during adulthood. A gene for FJHN in two Czech families was recently mapped to chromosome 16p11.2, close to the MCKD2 locus, which is responsible for a variant of autosomal dominant medullary cystic kidney disease observed in an Italian family. In a large Belgian family with FJHN, a tight linkage between the disorder

and the marker D16S3060, located within the MCKD2 locus on chromosome 16p12 (maximal two-point logarithmic odds score of 3.74 at a recombination fraction of ␪ ϭ 0), was observed in this study. The candidate region was further narrowed to a 1.3-Mb interval between D16S501 and D16S3036. Together with the striking clinical and pathologic resemblance between previously reported medullary cystic kidney disease type 2 and FJHN occurring in the Belgian family (including the presence of medullary cysts), this study suggests that these two disorders are facets of the same disease.

Familial juvenile hyperuricemic nephropathy (FJHN) [Mendelian Inheritance in Man (MIM) 162000] is an autosomal dominant condition characterized by abnormal tubular handling of urate associated with progressive renal failure. Originally described by Duncan and Dixon (1) in 1960, the disease has been recognized in the Netherlands (2), the United States (3), Australia (4), Switzerland (5), Japan (6), and Spain (7). The Guy’s Hospital team has gathered and characterized the largest number of families (8). The usual presenting feature is gout or hyperuricemia occurring in a young normotensive subject of either gender, in the absence of a purine synthesis disorder. There is grossly reduced fractional uric acid clearance, explaining the hyperuricemia (9). Renal impairment usually appears between 15 and 40 yr of age and leads to end-stage renal failure (ESRF) within 10 to 20 yr (4,7,8,10). Histologic examinations essentially demonstrate chronic interstitial nephritis. The most remarkable feature is thickening and splitting of tubular basement membranes (2–5,7,10). The nature of the molecular defect accounting for FJHN is

Received May 22, 2000. Accepted May 15, 2001. Correspondence to Dr. Karin Dahan, Center for Human Genetics, Universite ´ Catholique de Louvain, Avenue E, Mounier 52, Tour Ve ´ sale 5220, B-1200 Brussels, Belgium. Phone: 0032-2-764-52-20; Fax: 0032-2-764-52-22; E-mail: 1046-6673/1211-2348 Journal of the American Society of Nephrology Copyright © 2001 by the American Society of Nephrology

unknown. The search for candidate genes may be guided by either the biochemical or histologic characteristics of the condition. The hypoexcretion of uric acid indicates a defect involving the tubular transport of uric acid in the proximal tubular epithelium. A candidate gene could be UAT, a gene for a specific urate transporter that was isolated from a rat renal cDNA library (11,12). UAT protein functions as a selective, voltage-sensitive, urate transporter channel; it has been observed to be localized within the brush border membrane of proximal tubules (11). The deduced amino acid sequence of the cDNA predicts a transmembrane protein with a carboxy-terminal domain highly homologous (Ն98%) to the ␤-galactoside-binding lectin domain of galectin 5 (LGALS5), which was isolated from rat erythrocytes (13). The LGALS5 gene has been mapped to mouse chromosome 11, approximately 50 cM from the centromere, in a region syntenic with human chromosome 17q11 (13). Alternatively, hyperuricemia could be the first manifestation of a tubular defect that concurrently leads to interstitial fibrosis and progressive renal failure. The marked thickening of tubular basement membranes observed in FJHN is similar to that observed in the nephronophthisis (NPH)-autosomal medullary cystic kidney disease (MCKD) group of diseases. Juvenile NPH (NPH1, MIM 256100) and adolescent NPH (NPH2, MIM 602088) are autosomal recessive diseases that lead to ESRF in adolescence, whereas autosomal dominant MCKD (MCKD1, MIM 174000; MCKD2, MIM 603860) leads to ESRF during adulthood. Remarkably, a history of gout and/or hyperuricemia

J Am Soc Nephrol 12: 2348–2357, 2001



was also noted for several members from four of 12 kindreds with MCKD reported in the literature (14 –17). Histologic and clinical similarities between FJHN and MCKD thus suggest the possibility of a connection between the two entities. Recent advances in the molecular genetic understanding of MCKD could shed light on its relationship with FJHN. Two different loci for MCKD have been mapped, MCKD1 on chromosome 1q21 in two Cypriot families (18) and MCKD2 on chromosome 16p12 in an Italian family (19). Very recently, FJHN was mapped, in a large Japanese family (20) and in two Czech families (21), to chromosome 16p11 at a location very close to MCKD2, raising the question of whether distinct genes for MCKD2 and FJHN are colocalized within the approximately 10-cM region delimited by the authors or whether these two disorders represent two phenotypic forms of a defect in the same gene (20,21). We report on a large Belgian family with FJHN, including 14 affected members. All affected members met strict criteria for FJHN, and medullary cysts were present in all three available nephrectomy specimens. We performed positional linkage analysis of the specific urate transporter UAT/LGALS5 and the MCKD1 and MCKD2 loci. We observed a significant linkage to the marker D16S3060, located at the MCKD2/FJHN loci. Furthermore, we narrowed the candidate region for the FJHN locus to a 1.3-Mb interval between D16S501 and D16S3036. Together with the striking clinical and pathologic resemblance between previously reported MCKD2 and FJHN in our family (including the presence of medullary cysts), this study suggests that these two disorders are two facets of the same disease.

other at-risk subjects belonging to generations IV and V, none had a history of gout and all exhibited serum creatinine concentrations of Յ1.4 mg/dl. On the basis of serum uric acid concentrations, six of those subjects (subjects IV-1, IV-8, IV-10, V-2, V-3, and V-9) were considered to be affected and five (subjects IV-11, V-4, V-6, V-7, and V-8) were considered to be normal. The status of two subjects (subjects V-1 and V-5) who were not investigated was considered to be undetermined.

Genomic DNA was available for 12 affected individuals, nine unaffected individuals, and two individuals of undetermined status. DNA was extracted from leukocytes by using standard techniques. We performed linkage analysis with eight microsatellites spanning approximately10 to 20 cM (D17S122, D17S805, D17S783, D17S1800, D17S1880, D17S798, D17S933, and D17S800) of the chromosomal region 17q11 (a candidate for LGALS5). The sequence information and the genomic order of microsatellite markers used for linkage analysis of chromosome 17q11 were obtained from the Ge ´ ne ´thon gender-averaged genetic map (23) and the Genome Database. Three polymorphic microsatellite markers from 1q21 (D1S1153, D1S1593, and D1S2125) and three markers from 16p12 (D16S3017, D16S3036, and D16S3041) were selected, on the basis of their demonstrated high logarithmic odds (LOD) scores in the identification of MCKD1 and MCKD2 loci, respectively. Their order was initially based on the works of Christodoulou et al. (18) and Scolari et al. (19). Subsequently, when evidence of linkage was obtained for the 16p12 locus, additional polymorphic markers selected from the available databases (D16S3114, D16S3069, D16S764, D16S3060, D16S287, D16S3103, and D16S499) were analyzed, to determine whether analysis of recombinant individuals within this family would permit refinement of the published FJHN interval (20,21). Fluorescence markers were custom-synthesized by Genset (Paris, France). Expected size information and allele frequencies were obtained from either the Genome Database or the Whitehead Institute for Biomedical Research/MIT Center for Genome Research. In brief, markers were amplified by using a fluorescently labeled primer and AmpliTaq polymerase (Perkin-Elmer, Norwalk, CT), in a 20-␮l reaction volume containing 50 ng of genomic DNA, 10 pmol of each primer, 1 mM dNTP, and standard reaction buffer. The resultant PCR products were electrophoresed through a 4% denaturing polyacrylamide gel in an ABI 373 automated DNA sequencer (Applied Biosystems, Foster City, CA). Gel data were extracted by using ABI Genescan software, and the microsatellite peaks were sized with the same program. For seven of the 10 chromosome 16p markers, the PCR products were radiolabeled by incorporation of [␣-32P]dCTP (Amersham, Arlington Heights, IL). PCR-derived products were loaded, in groups of two or three (according to the expected sizes of the DNA fragments), on a sequencing polyacrylamide gel containing 6 M urea, 37.5% formamide, and Tris-borate-ethylenediaminetetraacetate buffer. Gels were developed at 100 W for 3 h (4°C), dried, and exposed to Kodak x-ray film (Eastman Kodak, Rochester, NY) for 1 to 12 h at room temperature.

Materials and Methods
Clinical Evaluations
This family is a five-generation kindred originating from the southern part of Belgium (Figure 1). The diagnosis of FJHN in this family was established on the basis of the coexistence of (1) autosomal dominant chronic renal failure, (2) similar appearances of chronic interstitial nephritis, with marked thickening of tubular membranes, for all three subjects for whom kidney tissue was available, and (3) a history of gout preceding renal failure for all six investigated subjects with ESRF (Table 1). We defined as affected either patients with ESRF and a history of gout (subjects II-1, III-1, III-5, IV-3, IV-5, IV-13, and IV-14) or patients with serum creatinine concentrations of Յ1. 4 mg/dl and serum uric acid concentrations Ͼ1 SD greater than the normal values for age and gender (Ͻ5 yr, 3.6 Ϯ 0.9 mg/dl; 5 to 10 yr, 4.1 Ϯ 1 mg/dl; male, 12 yr, 4.4 Ϯ 1.1 mg/dl; male, 15 yr, 5.6 Ϯ 1.1 mg/dl; male, Ն18 yr, 6.2 Ϯ 0.8 mg/dl; female, 12 yr, 4.5 Ϯ 0.9 mg/dl; female, 15 yr, 4.5 Ϯ 0.9 mg/dl; female, Ն18 yr, 4 Ϯ 0.7 mg/dl) (22) (subjects IV-1, IV-8, IV-10, V-2, V-3, and V-9). Fourteen subjects were found to be affected. The clinical characteristics of the 12 investigated subjects are presented in Table 1. Subject II-1, who died at 75 yr of age, had a history of gout and renal failure. No information was obtained for subject II-3, who is an obligate carrier. Among subjects belonging to generations III and IV, six (subjects III-1, III-5, IV-3, IV-5, IV-13, and IV-14) experienced ESRF at ages ranging from 28 to 63 yr, with a history of gout preceding ESRF by 4 to 35 yr. Kidney ultrasonography (performed at the stage of renal failure) revealed no cyst for four individuals, one cyst for two individuals, and two cysts for one individual. Among

Linkage Analysis
The alleles were assigned to individuals, which allowed calculation of two-point LOD scores with Cyrillic version 2.0 (Cherwell Scientific) and MLINK (24) software programs. For the two-point LOD scores and the multipoint linkage analyses, FJHN was modeled as an autosomal dominant trait with a disease allele frequency of 0.0001. Full penetrance was assumed for affected individuals. The marker


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J Am Soc Nephrol 12: 2348–2357, 2001

Figure 1. Pedigree of the Belgian family with familial juvenile hyperuricemic nephropathy (FJHN). Haplotypes and recombination for chromosome 16p12 microsatellite markers (ordered from telomere to centromere, top to bottom) are indicated on the left. Individuals with end-stage renal disease are depicted by black symbols, individuals with decreased fractional excretion of uric acid by symbols with a black upper left quadrant, and individuals with reduced GFR by symbols with a black upper right quadrant. Circles denote female subjects, and squares indicate male subjects. Inferred haplotypes are shown in parentheses. The chromosomal region cosegregating with the disease locus is represented by a black box. Vertical lines indicate uninformative regions on the disease chromosome.

allele frequencies were uniformly distributed. Calculations using marker allele frequencies reported in available databases were also performed, but these changes had minimal effects on the LOD scores generated. The number of alleles was estimated on the basis of either information from the Genome Database, if available, or the number of alleles observed in the study samples. Alleles were scored, and genotype data were entered into the pedigree file of the linkage package.

Therefore, at any given locus, results for the 14 affected individuals were used to generate a final LOD score for each marker tested. In addition, multipoint analyses were performed by using a parametric multipoint linkage program included in the Gene Hunter program (25). Mapping information was obtained from the Unified Database (UDB) (, which is based on combined genetic, radiation hybrid, and physical mapping findings (contig

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Table 1. Clinical, biochemical, and histologic characteristics of investigated subjectsa
At Screening Patient No. Gender Phenotype Age (yr) FEur (%) GFR (ml/min) Serum Creatinine Concentration (mg/dl) Serum Uric Acid Concentration (mg/dl) Gout/Age (yr) at First Attack ESRF/ Age (yr) Renal Histologic Features

III-1 III-5 IV-1 IV-3 IV-5 IV-8 IV-10 IV-11 IV-13 IV-14 V-1 V-2 V-3 V-4 V-5 V-6 V-7 V-8 V-9



34 32 19 30 22 36 31 38 10 7 6 14 2 13 16

ND ND 4.6 ND ND 5.5 6.6 ND ND 4.3 ND 5.1 6.4 14.3 ND ND ND 9.8 5.7


88 124

79 43

84 103

1.4 4.4 1.3 0.9 1.1 0.85 1.4 3.0 ND 0.9 0.7 0.6 ND 0.8 0.4 0.8 1

9.5 13.4 High 5.7 6.1 3.7 10.7 3.3 ND 7.2 5.4 3.7 ND 5.1 2.8 4.6 7.4

ϩ/? ϩ/19 Ϫ ϩ/32 ϩ/8 Ϫ Ϫ Ϫ ϩ/18 ϩ/26 Ϫ Ϫ Ϫ Ϫ Ϫ Ϫ Ϫ Ϫ Ϫ

ϩ/63 ϩ/54 Ϫ ϩ/36 ϩ/28 Ϫ Ϫ Ϫ ϩ/40 ϩ/42 Ϫ Ϫ Ϫ Ϫ Ϫ Ϫ Ϫ Ϫ Ϫ


a NL, normal phenotype; ND, not done; UD, undetermined; TIN, tubulointerstitial nephritis; FEur, fractional excretion of uric acid (uric acid clearance/creatinine clearance ϫ 100) (normal values are Ͼ8 Ϯ 1.2% for men, Ͼ12 Ϯ 1.9% for premenopausal women, and Ͼ14 Ϯ 5.3% for children) (9,26); ESRF, end-stage renal failure; FJHN, familial juvenile hyperuricemic nephropathy.

content) and contains very recent data. The marker order, according to the UDB, is as follows (with relative map positions in parentheses): ptel-D16S3114 (25.756 Mb)-D16S3069 (29.196 Mb)-D16S764 (33.204 Mb)-D16S3060 (33.560 Mb)-D16S287 (34.408 Mb)D16S3017 (35.248 Mb)-D16S3103 (35.248 Mb)-D16S499 (35.521 Mb)-D16S3036 (36.472 Mb)-D16S3041 (36.993 Mb)-cen.

Linkage Analysis
We initially genotyped all individuals with eight polymorphic microsatellite repeats (D17S122, D17S805, D17S783, D17S1800, D17S1880, D17S798, D17S933, and D17S800) spanning approximately10 to 20 cM of the chromosomal region 17q11, which is syntenic with the LGALS5 mouse locus. Two-point analysis was performed, and results were consistent with an exclusion. Indeed, this region was excluded on the basis of recombination and LOD scores well below Ϫ2 (data not shown). Linkage analysis of the three markers of MCKD1 located within the approximately 8-cM critical interval between D1S498 and D1S2125 demonstrated no cosegregation between the 12 affected individuals and the haplotype associated with the disease, demonstrating that FJHN is not allelic with MCKD1 in this family (data not shown). Linkage analysis of the three markers of MCKD2 demonstrated significant linkage for marker D16S3017, with a maximal LOD score of 3.2 at a recombination fraction (␪max) of 0, whereas linkage to the markers D16S3041 and D16S3036 was excluded. A two-point analysis was then performed between the putative FJHN locus and four additional polymorphic markers (D16S3060, D16S287, D16S3103, and D16S499) located within the approximately 10.5-cM critical region previously described by Scolari et al. (19) as the MCKD2 locus, as well as three

Renal Pathologic Features
Macroscopic examination of three nephrectomy specimens, obtained at the time of transplantation from subjects IV-3, IV-13, and IV-14, consistently demonstrated small kidneys containing several cysts that were clearly localized in the medulla (Figure 2A). Light-microscopic examination of the kidney biopsy specimen from subject IV-3 demonstrated areas of tubular atrophy and interstitial fibrosis, with marked thickening of tubular basement membranes. More extensive, essentially unchanged lesions were present in the nephrectomy specimens. The same lesions were also observed in the nephrectomy specimens from subjects IV-13 and IV-14; the most remarkable features were the thickening, wrinkling, and splitting of tubular basement membranes (Figure 2B). Ig, complement, or fibrin was not detected with immunofluorescence microscopy.


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telomeric markers (D16S3114, D16S3069, and D16S764). Two markers flanking the D16S3017 marker (D16S3060 and D16S3103) demonstrated significant LOD scores of Ͼ3, with no recombination. The maximal two-point LOD score of 3.74 was obtained with the marker D16S3060, at a maximal recombination fraction (␪max) of 0 (Table 2). We then performed a parametric multipoint linkage analysis with the Gene Hunter version 2.0 beta (r2) program under the autosomal dominant model, as described in the Materials and Methods section. The order of loci obtained from the integrated UDB map (entry, March 19, 2001) (Figure 3A4) was as follows: ptel-D16S3114 (25.7 Mb)-D16S3069 (29.2 Mb)D16S764 (33.2 Mb)-D16S3060 (33.6 Mb)-D16S287 (34.4 Mb)-D16S3017 (35.2 Mb)-D16S3103 (35.2 Mb)-D16S499 (35.5 Mb)-D16S3036 (36.5 Mb) and D16S3041 (37 Mb)-cen. For the program, distances between two markers are assumed on the basis of 1 cM representing 106 bases. The multipoint LOD scores were not different from the two-point LOD scores, with a maximal multipoint LOD score of 3.7 at markers D16S3017 and D16S3103. Multipoint LOD score results are presented in Figure 4.

Haplotype Studies
To refine the FJHN locus, we constructed detailed haplotypes using the same 16p markers that were used for the linkage analyses. The pedigree with haplotypes was drawn using the Cyrillic program and is presented in Figure 1. A common haplotype, i.e., 2-1-1-4-2 for markers D16S764, D16S3060, D16S287, D16S3017, and D16S3103, is shared by all affected individuals, including subjects IV-1, IV-8, IV-10, V-2, and V-9, who exhibit only hyperuricemia resulting from low fractional excretion of uric acid (Table 1). This is in accord with the data in Table 2 that indicated that the LOD scores for the markers D16S3060, D16S3017, and D16S3103 were Ͼ3 at ␪ ϭ 0 when the penetrance was set at 1. In contrast, subject V-4, who is clearly unaffected (Table 1), has received the grandpaternal normal haplotype. Haplotype analysis revealed two recombinant events, nar-

Figure 2. Renal histologic features for subject IV-14. (A) Cut section of the nephrectomy specimen, showing several cysts in the medullary area. Hematoxylin and eosin. (B) Extensive tubular atrophy and interstitial fibrosis, with thickening and splitting of tubular basement membranes. Periodic acid-Schiff stain. Magnifications: ϫ7 in A; ϫ450 in B.

Table 2. Two-point LOD scores obtained between FJHN and eight microsatellite markers spanning the MCKD2 region on chromosome 16p12a
Locus 0.00 0.01 0.05 LOD Scores at ␪ ϭ 0.10 0.20 0.30 0.40

D16S3114 D16S3069 D16S764 D16S3060 D16S3017 D16S3103 D16S287 D16S499 D16S3036 D16S3041

Ϫinf Ϫinf 1.87 3.74 3.2 3.61 1.57 1.15 Ϫinf Ϫinf

0.41 0.60 1.83 3.68 3.1 3.54 1.54 1.13 0.67 0.03

0.98 0.64 1.67 3.43 2.84 3.28 1.42 1.03 1.20 0.53

1.11 1.02 1.48 3.1 2.52 2.93 1.23 0.92 1.29 0.66

1.04 1.15 1.09 2.4 1.85 2.19 0.89 0.69 1.13 0.62

0.80 0.96 0.70 1.65 1.16 1.4 0.56 0.47 0.82 0.44

0.46 0.56 0.34 0.85 0.47 0.59 0.26 0.24 0.44 0.21

LOD, logarithmic odds; inf, infinity.

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Figure 3. (A) Alignment of genetic and physical maps of the FJHN/MCKD2 critical regions used in different studies. Markers studied in the respective publications are underlined. Markers identified as flanking are shown in boxes and connected by dotted lines to their positions on the reference map (4). The telomeric (tel) to centromeric (cen) orientation is indicated. 1, GeneMap 1998 radiation hybrid map (http://, used in MCKD2 mapping by Scolari et al. (19); 2, Marshfield genetic map (http://research.mars), used in FJHN mapping by Stiburkova et al. (21); 3, GeneMap 1999 radiation hybrid map (, used in FJHN mapping by Kamatani et al. (20); those authors also used the Marshfield map (2); 4, Integrated Unified Database (UDB) map (http://, based on data integrated from various radiation hybrid, linkage, and physical mapping resources and used in FJHN mapping in this study. (B) Regions compatible with linkage to the FJHN/MCKD2 loci, as defined in different publications. Horizontal bars end at flanking markers, thus indicating map ranges compatible with linkage. 1 through 4, publications in the same order as in A. It should be noted that there is no critical region flanked by two crossover events with the disease locus, which should represent the outer boundaries of the linked region on chromosome 16p12, for the Japanese family. Bars are drawn in relation to the integrated UDB map (see A4). Flanking markers, as defined in references 19 (A1), 20 (A3), and 21 (A2), are connected to the reference map (A4) by thin dotted lines.

rowing the critical interval for FJHN. As shown in Figure 1, the recombination observed with marker 16S3036 in an affected member (subject IV-8) helps to define the centromeric boundary to marker D16S3036, whereas the recombination event observed for marker D16S3069 in five affected members (subjects III-5, IV-5, IV-13, IV-14, and V-9) defines the telomeric boundary to marker D16S3069. In subjects IV-8 and V-8, the distal marker (D16S499) is homozygous and uninformative; therefore, the possibility of recombination events between markers D16S3103, D16S499, and D16S3036 remains undetermined. The recombination breakpoints were therefore located distal to marker D16S3069 at 29.2 Mb and proximal to marker D16S3036 at 36.5 Mb from the telomere (Figure 3, A4 and B4).

Refinement of the FJHN Locus
The mapping data, with critical recombinations observed in the Japanese and Czech families (20,21) and in this family, are

depicted in Figure 3. This diagram, which allows cross-reference with the available mapping databases, was determined by alignment of genetic and physical maps of the FJHN critical regions for the Japanese (20), Czech (21), and Belgian families. Linkage analysis results reported for our family confirm the localization of the FJHN locus in the p12 region of chromosome 16 (20,21). The relative location of the critical region observed in the single large Japanese family (20) is controversial. Indeed, using a genetic map such as the Marshfield map, the critical region lies between markers D16S403 (at 42.7 cM) and D16S3116 (at 50.6 cM) (Figure 3A3), with a dominant haplotype, shared by all affected individuals, for the five following markers: ptel-D16S417-D16S420-D16S3113D16S401-D16S3133-cen. However, using an integrated database such as the UDB map, which is derived from various mapping and recent sequencing resources for the human genome (26,27), the order of the nine markers used in this


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Figure 4. Results of multipoint logarithmic odds (LOD) score analysis for the Belgian FJHN family reported in this study, using the Gene Hunter program. The x-axis represents the genetic distance, and the y-axis represents the LOD score. The positions of the genetic markers used in the multipoint analysis are drawn to scale on the x-axis. The centromere is located to the right of the graph and is not shown. A maximal parametric LOD score of 3.7 was calculated for D16S3060. Markers identified as flanking in our family are underlined.

study (Figure 3A4) is now (entry of March 19, 2001) as follows: ptel-D16S3133 (37.3 Mb)-D16S3113 (37.3 Mb)D16S403 (39.4 Mb)-D16S3116 (39.6 Mb)-D16S412 (40.1 Mb)-D16S417 (41.6 Mb)-D16S420 (42.1Mb)-D16S401 (42.5Mb) and D16S3093 (42.6 Mb). The recombination events observed by Kamatani et al. (20) for markers D16S403 and D16S3116 (at 39.4 and 39.6 Mb, respectively) probably help define a centromeric boundary, whereas the absence of recombination events for the telomeric markers D16S3113 and D16S3133 (at 37.3 Mb) is consistent with a linkage to the disease at this locus, with a maximal LOD score of 5.13 for D16S3113 at a recombination fraction (␪max) of 0 (Figure 3B3). In contrast, the FJHN critical region defined in this study (Figure 3B4) shows overlap with that reported for two Czech families (Figure 3B2). The FJHN critical interval is flanked by two recombination events, at markers D16S501 (35.2 Mb) and D16S3036 (36.5 Mb) in the Czech and Belgian families, respectively. These two informative recombinations can be used to exclude either centromeric or telomeric portions of 16p12 from carrying the mutated gene. Because the telomeric border for the FJHN locus was previously defined as D16S501 (20) and because, in this study, the centromeric border was delimited to D16S3036, the FJHN locus is now refined to the interval tel-D16S501-FJHN-D16S3036-cen, which measures approximately 4.8 cM on the Marshfield genetic map (Figure 3A2) or 1.3 Mb on the UDB map (Figure 3A4). This

4.8-cM interval is located within the MCKD2 critical interval, which was identified by Scolari et al. (19) as being between D16S500 and SCNN1B1–2 (Figure 3A1).

Using a candidate gene approach, we mapped the gene for FJHN to 16p12, with a maximal two-point LOD score of 3.74 at a recombination fraction (␪max) of 0 for one marker (D16S3060) located within the approximately 10.5-cM critical region previously described by Scolari et al. (19) as the MCKD2 locus. Strong linkage of FJHN to the MCKD2 locus is supported by LOD scores of Ͼ3, with no recombination, for two other markers (D16S3017 and D16S3103) flanking the D16S3060 marker. This location on chromosome 16p confirms the findings from the recent studies by Stiburkova et al. (21) and Kamatani et al. (20), who demonstrated, in two Czech families and a single large Japanese family with FJHN, respectively, strong linkage to the same chromosomal interval (Figure 3). However, with the use of a genetic map such as the Marshfield map, the critical region between markers D16S403 and D16S3116 reported by Kamatani et al. (20) demonstrates no overlap with the critical region between markers D16S3069 and D16S3036 defined for the Belgian family. This finding could be accounted for by the existence of two homologous genes lying in close proximity. Alternatively, genotyping errors or inconsistencies in the genetic maps could lead to misinterpretations in the definition of critical intervals. Interest-

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ingly, by using markers located on chromosome 16p12 between D16S403 and D16S3116, nonoverlapping critical regions have been identified in different families affected by benign familial infantile convulsions (28). This finding raises questions regarding the positions of the markers defining the critical interval in the Japanese family (20). The order of these markers has indeed been recently modified by using more accurate sequencing data resources (Figure 3A4) (26,27), leading to a novel interpretation of the critical region. Using genotype data and two-point LOD scores provided by Kamatani et al. (20), it is impossible to redefine a critical interval flanked by two crossover events with the disease locus, which should represent the outer boundaries of the linked region on chromosome 16p12. Therefore, only haplotype analyses performed for the Czech and Belgian families allow restriction of the candidate region for FJHN within a 4.8-cM region between loci D16S501 and D16S3036, in agreement with the intermarker distances according to the Marshfield genetic map (Figure 3A2) or the UDB map (Figure 3A4). Most interestingly, this interval is located within the MCKD2 critical interval, which was identified by Scolari et al. (19) as being between D16S500 and SCNN1B1–2 (Figure 3A1). The next question is whether the genes for FJHN and MCKD2 are close to each other or the two disorders are actually a single entity (21). A detailed review of the data for families reportedly affected by MCKD, as well as our own findings, strongly argue for the existence of very close, perhaps similar, entities. Among 12 families with MCKD described in the literature (14 –17,29,30), a history of gout and hyperuricemia was mentioned for several subjects from four families (Table 3). Furthermore, low fractional excretion of uric acid was observed for two at-risk teenage subjects affected by MCKD (14), as well as young at-risk subjects from families with FJHN (31). The clinical presentation of the family reported by Scolari et al. (17), in which the MCKD2 locus was mapped (19), shares many similarities with the presentation of our family, i.e., hyperuricemia was observed for five of seven patients, the age of ESRF ranged from 33 to 63 yr, and chronic interstitial nephritis, with marked thickening of tubular basement membranes, was observed for four of four patients. Of note, cysts were detected during imaging for only a minority of patients with MCKD, including only one of the seven patients reported by Scolari et al. (17,19). Only at autopsy, which was performed for a few patients, was the presence of medullary cysts indisputable (14,15).

In our family (which meets all criteria for FJHN), we demonstrate, for the first time for this disease, the existence of medullary cysts characteristic of so-called MCKD. Interestingly, these cysts seem to be a late phenomenon and can easily escape imaging detection. In this family, ultrasonography indeed failed to detect any cysts for four adults with chronic renal failure (age range, 34 to 54 yr), as well as two affected children (ages, 5 and 14 yr). For one adult, ultrasonographic results were negative at 34 yr of age but demonstrated one cyst in each kidney 5 yr later. For another adult, ultrasonography demonstrated two cysts in each kidney but computed tomography demonstrated multiple bilateral cysts 3 yr later. The ultimate proof of the existence of medullary cysts in this family was provided only by the nephrectomy specimens; in each of three cases, several medullary cysts were clearly present. The poor results of conventional imaging in the detection of medullary cysts could explain why this finding was hitherto unreported for FJHN. Our finding of the hallmarks of both FJHN and MCKD in our family supports the hypothesis of allelism, which is consistent with DNA linkage analysis results. It can be imagined that the same disease was termed FJHN by some authors when the first manifestation was early gout, especially when it occurred among young nonobese subjects, and was termed MCKD by other investigators when cysts were detected in shrunken kidneys, especially when hyperuricemia was lacking. FJHN and MCKD could thus be allelic variants. In other families with FJHN, it would be of interest to carefully search for the existence of medullary cysts and to test for linkage to both MCKD1 and MCKD2 loci. Conversely, fractional excretion of uric acid should be measured for affected and at-risk subjects from families with MCKD. A search for candidate genes localized within the approximately 4.8-cM region on the Marshfield genetic map (Figure 3A2) or the 1.3-Mb region on the UDB map identified a large number of expressed-sequence tags mapped by radiation hybrid analysis to the D16S501 to D16S3036 interval, together with four complete cDNA (GeneMap 1999 and UDB). Among the genes noted in the critical region, we observed two complete sequences for cDNA of unknown human genes, termed KIAA0421 and KIAA0419, which were isolated from different sources, including human kidney cDNA libraries. Because these two predicted amino acid sequences have not yet been classified in a functional category, their potential effects remain unknown. A third transcript is a potential central meta-

Table 3. Gout and/or hyperuricemia among members of four families with medullary cystic kidney disease
No. of Patients Affected/Examined Authors No. of Affected Individuals Gout (age at first crisis, yr) Hyperuricemia

Thompson et al., 1978 (14) Burke et al., 1982 (15) Stavrou et al., 1998 (16) Scolari et al., 1998 (17)

5 6 23 10

3/5 (14, 17, 20) 2/6 (18, 38) 4/15 (?) 2/10 (19, 48)

5/5 5/6 7/13 5/7


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bolic regulator belonging to the coenzyme Q7 family, members of which are known to regulate ubiquinone biosynthesis and function as an electron transport component and a lipid-soluble antioxidant (32). The COQ7 gene is well conserved among mammal, bird, and reptile genomes and is dominantly transcribed in heart and skeletal muscle, likely localized in the mitochondrial inner membrane (33). Therefore, the COQ7 gene does not seem to be a candidate gene for FJHN. The fourth cDNA located within the defined critical region is involved in G protein-coupled signaling. Named GPRC5B (G protein-coupled receptor family C, group 5, member B), it contains a signal peptide and seven transmembrane ␣-helices, which are hallmarks of G protein-coupled receptors (34). G protein-coupled receptors constitute a large class of proteins that are divided into three families (family A, rhodopsin receptor-like; family B, secretin receptor-like; family C, metabotropic glutamate receptor-like) and are thought to transmit a variety of signals from the extracellular milieu to the intracellular milieu (35). GPRC5B displays homology to retinoic acid-inducible gene 1 (RAIG1) protein. Both RAIG1 and GPRC5B have short extracellular amino-terminal domains, which is strikingly different from the majority of family C receptors characterized by large amino-terminal domains. The two related transcripts, GPRC5B and RAIG1, are distinctly expressed, with the highest mRNA levels for RAIG1 in lung tissue and the highest levels for GPRC5B in kidney (34). On the basis of their transcriptional induction by retinoic acid, it has been speculated that these transcripts could be involved in modulation of differentiation and maintenance of homeostasis of epithelials cells or in embryonic development and maturation of fetal lung and kidney (36). Interestingly, FJHN and MCKD are diseases characterized by severe changes in the renal interstitium, with accumulation of collagen fibers and disruption of tubular basement membranes. Because these changes could be mediated by G protein-coupled receptors (37), the GPRC5B gene is a potential candidate for mutation analysis. However, the 4.8-cM interval containing the FJHN gene, which spans 1.3 Mb on the integrated map, is still large for sequencing and mutation screening of candidate genes. Analyses of additional families and other genetic markers should help to further restrict the critical interval and establish the possible allelism between MCKD2 and FJHN. The ultimate proof of the latter hypothesis can be provided only by identification of the responsible gene. Elucidation of the molecular bases of MCKD and FJHN should help clarify the classification of hereditary interstitial nephritides and cystic diseases. It should also cast some light on the mechanism of urate hypoexcretion and, it is hoped, expand therapeutic possibilities.

de Belgique et du Fonds National de Recherche Scientifique (Grant 96/PHD/A6/CV/0081LT). Dr. Fuchshuber was supported by the German Research Foundation (Grant FU202/3-1). URL addresses for data described in this article are as follows: Online Mendelian Inheritance in Man,; Genome Database,; UDB, http://bioinformatics.; Genetic Location Database, http://cedar.genetics. (for inferred physical locations of locus markers); GeneMap 1999, (for localization of genes and expressed-sequence tags).

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We express our gratitude to the members of the Belgian family, to Alexandre Irtum for multipoint analyses, to Lucienne Michaux and Genevieve Quenum (Center of Human Genetics, Brussels, Belgium) for fruitful discussions, and to Isabelle Abinet and Chantal Piron for continuous support in the preparation of the manuscript and figures. This work was supported by the Fonds de la Communaute ´ Franc ¸ aise

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