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Background: Haploinsufficiency of the FOXL2 transcription factor in humans causes Blepharophimosis/Ptosis/Epicanthus Inversus syndrome (BPES), characterized by eyelid anomalies and premature ovarian failure. Mice lacking Foxl2 recapitulate human eyelid/forehead defects and undergo female gonadal dysgenesis. We report here that mice lacking Foxl2 also show defects in postnatal growth and embryonic bone and cartilage formation., Methods: Foxl2 (-/-) male mice at different stages of development have been characterized and compared to wild type. Body length and weight were measured and growth curves were created. Skeletons were stained with alcian blue and/or alizarin red. Bone and cartilage formation was analyzed by Von Kossa staining and immunofluorescence using anti-FOXL2 and anti-SOX9 antibodies followed by confocal microscopy. Genes differentially expressed in skull vaults were evaluated by microarray analysis. Analysis of the GH/IGF1 pathway was done evaluating the expression of several hypothalamic-pituitary-bone axis markers by RT-qPCR., Results: Compared to wild-type, Foxl2 null mice are smaller and show skeletal abnormalities and defects in cartilage and bone mineralization, with down-regulation of the GH/IGF1 axis. Consistent with these effects, we find FOXL2 expressed in embryos at 9.5 dpc in neural tube epithelium, in head mesenchyme near the neural tube, and within the first branchial arch; then, starting at 12.5 dpc, expressed in cartilaginous tissue; and at PO and P7, in hypothalamus., Conclusions: Our results support FOXL2 as a master transcription factor in a spectrum of developmental processes, including growth, cartilage and bone formation. Its action overlaps that of SOX9, though they are antagonistic in female vs male gonadal sex determination but conjoint in cartilage and skeletal development.

Background: Despite progress in identifying genes associated with breast cancer, many more risk loci exist. Genome-wide association analyses in genetically-homogeneous populations, such as that of Sardinia (Italy), could represent an additional approach to detect low penetrance alleles., Methods: We performed a genome-wide association study comparing 1431 Sardinian patients with non-familial, BRCA1/2-mutation-negative breast cancer to 2171 healthy Sardinian blood donors. DNA was genotyped using GeneChip Human Mapping 500 K Arrays or Genome-Wide Human SNP Arrays 6.0. To increase genomic coverage, genotypes of additional SNPs were imputed using data from HapMap Phase II. After quality control filtering of genotype data, 1367 cases (9 men) and 1658 controls (1156 men) were analyzed on a total of 2,067,645 SNPs., Results: Overall, 33 genomic regions (67 candidate SNPs) were associated with breast cancer risk at the p <  0(-6) level. Twenty of these regions contained defined genes, including one already associated with breast cancer risk: TOX3. With a lower threshold for preliminary significance to p < 10(-5), we identified 11 additional SNPs in FGFR2, a well-established breast cancer-associated gene. Ten candidate SNPs were selected, excluding those already associated with breast cancer, for technical validation as well as replication in 1668 samples from the same population. Only SNP rs345299, located in intron 1 of VAV3, remained suggestively associated (p-value, 1.16 x 10(-5)), but it did not associate with breast cancer risk in pooled data from two large, mixed-population cohorts., Conclusions: This study indicated the role of TOX3 and FGFR2 as breast cancer susceptibility genes in BRCA1/2-wild-type breast cancer patients from Sardinian population.

Background: In recent years, numerous studies have assessed the prevalence of germline mutations in BRCA1 and BRCA2 genes in various cohorts. We here extensively investigated the prevalence and geographical distribution of BRCA1-2 mutations in the entire genetically-homogeneous Sardinian population. The occurrence of phenotypic characteristics which may be predictive for the presence of BRCA1-2 germline mutations was also evaluated., Methods: Three hundred and forty-eight breast cancer patients presenting a familial recurrence of invasive breast or ovarian carcinoma with at least two affected family members were screened for BRCA1-2 mutations by DHPLC analysis and DNA sequencing. Association of BRCA1 and BRCA2 mutational status with clinical and pathological parameters was evaluated by Pearson's Chi-Squared test., Results and Conclusion: Overall, 8 BRCA1 and 5 BRCA2 deleterious mutations were detected in 35/348 (10%) families; majority (23/35;66%) of mutations was found in BRCA2 gene. The geographical distribution of BRCA1-2 mutations was related to three specific large areas of Sardinia, reflecting its ancient history: a) the Northern area, linguistically different from the rest of the island (where a BRCA2 c.8764&#95;8765delAG mutation with founder effect was predominant); b) the Middle area, land of the ancient Sardinian population (where BRCA2 mutations are still more common than BRCA1 mutations); and c) the South-Western area, with many Phoenician and Carthaginian locations (where BRCA1 mutations are prevalent). We also found that phenotypic features such as high tumor grading and lack of expression of estrogen/progesterone receptors together with age at diagnosis and presence of ovarian cancer in the family may be predictive for the presence of BRCA1-2 germline mutations.

In this study, we report further results of mutation analysis of the ATP7B gene in Wilson disease (WD) patients of Mediterranean origin. A total of 136 WD chromosomes, 73 of which were of Italian, 43 of Turkish, 18 of Sardinian, and two of Spanish origin, were analysed and the mutation characterised in 84.5% of them. We found 50 different mutations of which 19 are novel, including three nonsense, one frameshift, and 15 missense mutations. The mutations detected were rare and mostly found in the compound heterozygous state together with other mutations and only rarely in homozygosity. Most of these mutations lie in the transmembrane and ATP binding loop regions. These data expand our knowledge of both the structure-function relationships of the WD protein and the molecular pathology of WD, thus improving our capability of prevention and genetic counselling.

In this study, we report the results of haplotype and mutation analysis of the ATP7B gene in Wilson disease (WD) patients of Greek origin. We have analysed 25 WD families and two single patients and characterised 94% of the WD chromosomes investigated. We have found 12 different molecular defects (three frameshifts, two splice site, two nonsense, five missense mutations), four of which are novel. Five of the mutations are widely prevalent accounting for 74% of the WD chromosomes analysed. These results may enable preclinical diagnosis in the large majority of WD patients of Greek descent, thereby improving genetic counselling and disease management.

This study reports 12 novel mutations of the Wilson disease (WD) gene which have been detected by the molecular analysis of 29 patients of Mediterranean descent carrying uncommon chromosomal haplotypes at the WD locus. These mutations include two nonsense, one splice site and nine missense. The missense mutations lie in regions of the WD gene critical for its function, such as the transmembrane region, the transduction domain and the ATP loop and ATP-binding domain, indicating that they are disease-causing mutations. These new findings improve our knowledge for the role played by functional domains on the ATP7B function.

Background: Glucose-6-phosphate dehydrogenase (G6PD; E.C.1.1.1.49) deficiency is the most common human enzymopathy; nearly 400 different biochemical variants of the enzyme have been described. Sardinia is the Italian region with the highest frequency of this defect., Methods: We examined genomic DNA of 16 subjects with G6PD Mediterranean, 2 with G6PD Athens-like, 1 with G6PD Ferrara 2 (all as biochemically defined)., Results: All G6PD Mediterranean subjects had a C-->T mutation at nucleotide 563 and a C-->T transition at nucleotide 1311; G6PD Athens-like and Ferrara 2 subjects had a G-->C mutation at nucleotide 844 (the same mutation has been found in G6PD Seattle-like)., Conclusions: This study suggests that in Southern Sardinia G6PD mutations are relatively homogeneous and that the results of biochemical characterization studies must be carefully evaluated, because the same mutations might be responsible for different biochemical behavior.