Refinement of the chromosome 5p locus for familial calcium pyrophosphate dihydrate deposition disease.

Familial calcium pyrophosphate dihydrate deposition disease (CPPDD) is a disease of articular cartilage that is radiographically characterized by chondrocalcinosis due to the deposition of calcium-containing crystals in affected joints. We have documented the disease in an Argentinean kindred of northern Italian ancestry and in a French kindred from the Alsace region. Both families presented with a common phenotype including early age at onset and deposition of crystals of calcium pyrophosphate dihydrate in a similar pattern of affected joints. Affected family members were karyotypically normal. Linkage to the short arm of chromosome 5 was observed, consistent with a previous report of linkage of the CPPDD phenotype in a large British kindred to the 5p15 region. However, recombinants in the Argentinean kindred have enabled us to designate a region<1 cM in length between the markers D5S416 and D5S2114 as the CPPDD locus.

Five families with arginine 519-cysteine mutation in COL2A1: evidence for three distinct founders.

Arginine519-cysteine mutation in the type II procollagen gene (COL2A1) is known to be associated with mild spondyloepiphyseal dysplasia (SED) and precocious generalized osteoarthritis (OA). Five families have now been identified with this mutation. To determine whether a common founder was responsible for the mutation in these five families, we defined the haplotype of the mutation-bearing chromosome using four restriction fragment length polymorphisms (RFLPs) and the 3'-untranslated region VNTR. Haplotype frequencies were estimated for 69 control samples. Three distinct mutation-bearing haplotypes were identified, with three families sharing a common haplotype. For three distinct haplotypes to have derived from a single founder, three independent recombination events would have had to occur. Thus the arg519 codon appears to represent a possible site of recurrent mutations in COL2A1, an uncommon phenomenon in collagen genes.

[Antiochian genealogies in which idiopathic epilepsy presents familial conglomeration. Simulations of power for the detection of genetic linkage]. / Genealogías antioqueñas en las que la epilepsia idiopática presenta conglomeración familiar. Simulaciones de poder para detectar ligamiento genético.

OBJECTIVE AND METHODS: We have analyzed a set of multigenerational extended pedigrees ascertained from affected cases of idiopathic epilepsy in the Antioquian Neurologic Institute. All pedigrees show familial aggregation of several forms of non myoclonic idiopathic epilepsy. In a recent paper, we have demonstrated that generalized idiopathic epilepsy of the awakening type is better explained by the existence of a major gene. In this paper, we have explored by simulation techniques the usefulness of the bigger pedigrees for linkage analysis. By using simlink and taking into account the parameters of the major gene, we have estimated that total power of three families is approximately 100 million times favoring the linkage detection. RESULTS AND CONCLUSIONS: These analyses suggest that the major gene accounting by the susceptibility to develop generalized idiopathic epilepsy of the awakening type could be localized by typifying affected families belonging to the Paisa community from Antioquia, Colombia (Acta Neurol Colomb 1997; 13: 69-75).

Genetic heterogeneity in autosomal recessive hereditary motor and sensory neuropathy with focally folded myelin sheaths (CMT4B).

Hereditary motor and sensory neuropathy with focally folded myelin sheaths, or Charcot-Marie-Tooth disease neuropathy type 4B (CMT4B), is a distinct clinical and genetic entity belonging to the heterogeneous group of autosomal recessive demyelinating neuropathies. We previously described a large pedigree with CMT4B and found evidence of linkage to chromosome 11q23. We now describe a second, unrelated family in which two individuals were affected with CMT4B. We exclude the disease locus segregating in this smaller pedigree from the 11q23 region as well as from most of the regions where other CMT loci have been mapped. We thus provide evidence for a second locus causing the CMT4B phenotype.

Alopecia universalis associated with a mutation in the human hairless gene.

There are several forms of hereditary human hair loss, known collectively as alopecias, the molecular bases of which are entirely unknown. A kindred with a rare, recessively inherited type of alopecia universalis was used to search for a locus by homozygosity mapping, and linkage was established in a 6-centimorgan interval on chromosome 8p12 (the logarithm of the odds favoring linkage score was 6.19). The human homolog of a murine gene, hairless, was localized in this interval by radiation hybrid mapping, and a missense mutation was found in affected individuals. Human hairless encodes a putative single zinc finger transcription factor protein with restricted expression in the brain and skin.

The role of MMAC1 mutations in early-onset breast cancer: causative in association with Cowden syndrome and excluded in BRCA1-negative cases.

Cowden syndrome (CS) is an autosomal dominant disorder associated with the development of hamartomas and benign tumors in a variety of tissues, including the skin, thyroid, breast, endometrium, and brain. It has been suggested that women with CS are at increased risk for breast cancer. A locus for CS was recently defined on chromosome 10 in 12 families, resulting in the identification of the CS critical interval, between the markers D10S215 and D10S541. More recently, affected individuals in four families with CS have been shown to have germ-line mutations in a gene known as "PTEN," or "MMAC1," which is located in the CS critical interval on chromosome 10. In this study, we report three novel MMAC1 mutations in CS and demonstrate that MMAC1 mutations are associated with CS and breast cancer. Furthermore, we also show that certain families and individuals with CS do not have mutations in the coding sequence of MMAC1. Finally, we did not detect MMAC1 mutations in a subpopulation of individuals with early-onset breast cancer, suggesting that germ-line mutations in this gene do not appear to be common in this group.

Unusual clinical features and early brain MRI lesions in a family with cerebral autosomal dominant arteriopathy.

Cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL) is a recently described inherited disorder. The pathologic gene maps on chromosome 19. The clinical spectrum of the disease consists of recurrent strokes, migraine, transient ischemic attacks, mood changes, and dementia. We report a genetically assessed CADASIL family with atypical clinical presentations of epileptic seizures. In two asymptomatic family members there were early brain abnormalities on MRI. Our report expands the clinical spectrum of CADASIL and suggests that it is possibly an undiagnosed disorder.

Clinicopathological and genetic studies of two further Italian families with cerebral autosomal dominant arteriopathy.

We report on two Italian families with an early-adult onset autosomal dominant disorder, characterized by leukoencephalopathy, migraine, psychiatric disturbances, stroke and dementia. These findings fulfill the diagnostic criteria for cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL) syndrome. Moreover, to confirm the CADASIL gene location to 19p12, we performed a linkage analysis with four microsatellite markers. The results of the genetic study gave positive but not significant lod scores, indicating only weak evidence of a linkage with 19p12. In one autopsy case, we found extensive ischemic changes due to the selective involvement of the small muscular arteries of the cerebral white matter. The lesions consisted of a thickening of the media with deposition of granular eosinophilic material. Ultrastructural examination of the arterial walls showed graded damage to smooth muscle cells, mostly of the longitudinal layer, and an abnormal proliferation of basal lamina components. Immunocytochemical analysis showed strong reactivity using antibodies to collagen IV and smooth myosin proteins. The results suggest a primary involvement of the smooth muscle cells of small cerebral arteries, with a secondary alteration of basal lamina components and elastic tissue.

Localization of a gene responsible for autosomal recessive demyelinating neuropathy with focally folded myelin sheaths to chromosome 11q23 by homozygosity mapping and haplotype sharing.

Hereditary motor and sensory neuropathy (HMSN) with focally folded myelin sheaths, or Charcot-Marie-Tooth type 4B (CMT4B), is a distinct clinical entity belonging to the heterogeneous group of autosomal recessive demyelinating neuropathies. We first described a large pedigree with CMT4B, which showed a high consanguinity level and an autosomal recessive pattern of inheritance. Through conventional linkage analysis, we excluded linkage of the locus segregating in this pedigree to any of the known genes responsible for other HMSNs. Using homozygosity mapping and haplotype sharing analysis, we were able to localize the disease gene in a 4 cM interval on chromosome 11q23, between the D11S1332 and D11S917 loci. On the basis of the clinical characteristics of the disease, we propose that this locus corresponds to the CMT4B gene.

The locus for a novel syndromic form of neuronal intestinal pseudoobstruction maps to Xq28.

The neuronal type of primary chronic idiopathic intestinal pseudoobstruction (CIIP) results from the developmental failure of enteric neurons to migrate or differentiate correctly. This leads to intestinal motility disorders, which are characterized by symptoms and signs of bowel obstruction in the absence of a mechanical obstacle. Most of these conditions are congenital, and among them some are inherited. One syndromic condition characterized by intestinal pseudoobstruction with morphological abnormalities of the argyrophil neurons in the myenteric plexus, associated with short small bowel, malrotation, and pyloric hypertrophy, has been previously described. We have studied a family affected by this disorder, in which the disease appeared to segregate as an X-linked recessive trait. In order to map the CIIP locus in this family, we performed linkage analysis in 26 family members by use of highly polymorphic microsatellite markers from the X chromosome. One of these markers, DXYS154, located in the distal part of Xq28, shows no recombination with a maximum lod score of 2.32. Multipoint analysis excluded linkage with markers spanning other regions of the X chromosome. Our results, integrated with the current genetic and physical map of Xq28, determine the order of loci as cen-DXS15-(CIIPX)-DXS1108/DXYS154-tel. This study establishes, for the first time, the mapping assignment of a neuropathic form of CIIP other than Hirschsprung disease.

Exclusion of linkage between RET and neuronal intestinal dysplasia type B.

Neuronal Intestinal Dysplasia type B (NID B) is a complex alteration of the enteric nervous system belonging to the group of intestinal dysganglionoses which may involve rectum, colon, and small intestine. Second only to Hirschsprung disease (HSCR), NID B is one of the most frequent causes of chronic constipation and pseudo-obstructive intestinal dysmotility. Since NID B is often associated with HSCR and point mutations in the RET proto-oncogene have been identified in HSCR patients, we analyzed two NID B pedigrees to investigate if RET mutations might cause also the NID B phenotype. Linkage analysis demonstrated that the NID B locus is not linked to RET in the pedigrees analyzed. Further genetic analyses will possibly improve the understanding of the cause and facilitate diagnostic procedures in NID B.

Genetic linkage studies for the identification of cancer-related genes.

The most commonly used method for genetic mapping of disease causing loci is the lod-score method, based on maximum likelihood estimation of the recombination fraction. This method is the most powerful one when the mode of inheritance of the disease is known, but is often used also for genetic mapping of complex traits, although in these cases alternative non parametric procedures are often preferable. In those families where cancer segregates as a Mendelian disease, mapping of the corresponding gene can be carried out as for any other genetic disorder. Modelling of the analysis should also consider the possible occurrence of reduced or age dependent penetrance, phenocopies, and genetic heterogeneity. The application of this method has already led to the identification of the genetic location and eventually to the cloning of several cancer-related genes, including FAP, BRCA1, NF1, and more recently AT. Understanding the molecular mechanisms leading to the disease in the Mendelian forms of cancer may prove helpful in unraveling the basis of the other more common forms of cancer.

Search for mutations in pancreatic sufficient cystic fibrosis Italian patients: detection of 90% of molecular defects and identification of three novel mutations.

A cohort of 31 cystic fibrosis patients showing pancreatic sufficiency and bearing an unidentified mutation on at least one chromosome was analyzed through denaturing gradient gel electrophoresis of the whole coding region of the cystic fibrosis transmembrane conductance regulator gene, including intron-exon boundaries. Three new and 19 previously described mutations were detected. The combination of these with known mutations detected by other methods, allowed the characterization of mutations on 56/62 (90.3%) chromosomes. Among those identified, 17 can be considered responsible for pancreatic sufficiency, since they were found in patients carrying a severe mutation on the other chromosome. Among these presumed mild mutations, eight were detected more than once, R352Q being the most frequent in this sample (4.83%). Intragenic microsatellite analysis revealed that the six chromosomes still bearing unidentified mutations are associated with five different haplotypes. This may indicate that these chromosomes bear different mutations, rarely occurring among cystic fibrosis patients, further underlying the molecular heterogeneity of the genetic defects present in patients having pancreatic sufficiency.

Capillary zone electrophoresis of polymerase chain reaction-amplified DNA fragments in polymer networks: the case of GATT microsatellites in cystic fibrosis.

In cystic fibrosis (CF), the most common mutation, delta F508 (a three-base-pair deletion) accounts for ca. 70% of mutations in the worldwide population. The majority of other mutations (more than 350 reported so far to the Genetic Analysis Consortium) have been detected in single cases, thus rendering quite cumbersome a molecular diagnostic approach for the identification of CF chromosomes. As an alternative, linkage analysis based on intragenic polymorphism can be useful for prenatal diagnosis and CF-carrier detection, provided that the heterozygosity of the allelic forms is very high. For this purpose, DNA microsatellites, consisting of two to epta nucleotide repeat clusters, displaying a high degree of polymorphism, are being increasingly used as markers in linkage studies. Two main allelic forms, one hexameric (111 bp) and one heptameric (115 bp), of a tetranucleotide (GATT) repeat polymorphism, at the junction of intron IVS6a and exon 6b, have been amplified by PCR technology. These two alleles can be separated in a 10-20% T polyacrylamide gradient gel and detected by ethidium bromide staining. As an alternate procedure, these two fragments are efficiently separated by capillary zone electrophoresis in a viscous solution of 6%T linear polyacrylamide and detected by their intrinsic absorbance at 254 nm.