Single nucleotide polymorphism and FMR1 CGG repeat instability in two Basque valleys.

Fragile X Syndrome (FXS, MIM 309550) is mainly due to the expansion of a CGG trinucleotide repeat sequence, found in the 5' untranslated region of the FMR1 gene. Some studies suggest that stable markers, such as single nucleotide polymorphisms (SNPs) and the study of populations with genetic identity, could provide a distinct advance to investigate the origin of CGG repeat instability. In this study, seven SNPs (WEX28 rs17312728:G>T, WEX70 rs45631657:C>T, WEX1 rs10521868:A>C, ATL1 rs4949:A>G, FMRb rs25707:A>G, WEX17 rs12010481:C>T and WEX10 ss71651741:C>T) have been analyzed in two Basque valleys (Markina and Arratia). We examined the association between these SNPs and the CGG repeat size, the AGG interruption pattern and two microsatellite markers (FRAXAC1 and DXS548). The results suggest that in both valleys WEX28-T, WEX70-C, WEX1-C, ATL1-G, and WEX10-C are preferably associated with cis-acting sequences directly influencing instability. But comparison of the two valleys reveals also important differences with respect to: (1) frequency and structure of "susceptible" alleles and (2) association between "susceptible" alleles and STR and SNP haplotypes. These results may indicate that, in Arratia, SNP status does not identify a pool of susceptible alleles, as it does in Markina. In Arratia valley, the SNP haplotype association reveals also a potential new "protective" factor.

Assessment of the genotoxicity of atenolol in human peripheral blood lymphocytes: correlation between chromosomal fragility and content of micronuclei.

The antihypertensive drug atenolol was found to induce chromosome loss, detected as micronuclei in the peripheral lymphocytes of treated patients. The fundamental question which chromosomes the micronuclei were derived from remains to be answered. Analysis of structural chromosomal aberrations (CAs) and expression of fragile sites (FS) were pursued in this study. They revealed a significantly higher incidence of chromosomal aberrations (chromatid and chromosome breaks) in patients compared with controls, where 10 FS emerged as specific. Also, the band 17q12-21, where known fragile sites have not been reported, was only expressed in atenolol-treated patients. Fluorescence in situ hybridization using chromosome-specific probes revealed the preferential involvement of chromosomes 7, 11, 17 and X in the micronuclei (MN) of patients. The results also suggest a correlation between chromosomal fragility and content of MN, and support the findings for a linkage between hypertension and a locus on chromosome 17.

Study of the Genetic Etiology of Primary Ovarian Insufficiency: FMR1 Gene.

Menopause is a period of women's life characterized by the cessation of menses in a definitive way. The mean age for menopause is approximately 51 years. Primary ovarian insufficiency (POI) refers to ovarian dysfunction defined as irregular menses and elevated gonadotrophin levels before or at the age of 40 years. The etiology of POI is unknown but several genes have been reported as being of significance. The fragile X mental retardation 1 gene (FMR1) is one of the most important genes associated with POI. The FMR1 gene contains a highly polymorphic CGG repeat in the 5' untranslated region of exon 1. Four allelic forms have been defined with respect to CGG repeat length and instability during transmission. Normal (5-44 CGG) alleles are usually transmitted from parent to offspring in a stable manner. The full mutation form consists of over 200 repeats, which induces hypermethylation of the FMR1 gene promoter and the subsequent silencing of the gene, associated with Fragile X Syndrome (FXS). Finally, FMR1 intermediate (45-54 CGG) and premutation (55-200 CGG) alleles have been principally associated with two phenotypes, fragile X tremor ataxia syndrome (FXTAS) and fragile X primary ovarian insufficiency (FXPOI).

Genotoxic evaluation of five Angiotesin II receptor blockers: in vivo and in vitro micronucleus assay.

Angiotensin II receptor blockers (ARBs) are a new class of drugs for the treatment of hypertension. In this study, we studied the potential genotoxic effects of five ARBs in vivo and in vitro in human peripheral blood lymphocytes (PBLs) by means of the cytokinesis-block micronucleous (CBMN) assay in combination with fluorescence in situ hybridization (FISH) with a centromeric probe. The nuclear division index (NDI) was used as a measure of cytotoxicity. We also analyzed the association between sex, age, duration of treatment and MN formation. The in vivo study was carried out in 55 hypertensive patients. The in vitro study was performed in 10 control individuals by adding the drugs to the culture medium at a final concentration similar to the levels found in plasma in patients. Our results showed a significant increase in the frequencies of MN and binucleated cells with MN (BNMN) in vivo and especially in vitro. We observed variability in the mean frequency of MN and BNMN among the five drugs analyzed. In vivo, patients treated with Candesartan, Telmisartan and Valsartan showed a statistical significant increase in these parameters, while Olmesartan showed the highest effect in vitro. We also found that the drugs inhibit the NDI in vitro and that Eprosartan, Olmesartan and Telmisartan are the ARBs studied with the highest effect in decreasing the proliferation of the cells. FISH analysis revealed no significant difference between patients and controls in the frequency of centromeric signals. A slight variability, without statistical significance, in the frequency of micronuclei with a centromere signal (CN(+)MN) was found among the different ARBs analyzed, ruling out an aneugenic potential. When accounting for risk factors, we found that in patients there is a positive correlation between MN, BNMN and sex and a negative correlation with duration of treatment.

Study of FMR1 gene association with ovarian dysfunction in a sample from the Basque Country.

Premature ovarian failure (POF) is defined as cessation of menses before the age of 40. The most significant single gene associated with POF is the Fragile X Mental Retardation 1 gene (FMR1). In the present work we screened women with fertility problems from the Basque Country in order to determine, whether in these women, FMR1 CGG repeat size in the intermediate and premutation range was associated with their pathology, and whether intermediate and premutation carriers had endocrine signs of diminished ovarian function, using the most established measure of ovarian reserve, the gonadotropin FSH. A patient sample of 41 women with ovarian insufficiency and a control sample of 32 women with no fertility problems from the Basque Country were examined. The patient sample was classified into three categories according to the results of the retrospective assessment of their ovarian function. In group 2 of patients, women with irregular cycles, reduced fecundity and FSH levels ≥ 10IU/l, there is a significant increase in the number of intermediate and premutation FMR1 alleles (35-54 CGG repeats). In group 3 of patients, women with amenorrhea for at least four consecutive months and FSH levels ≥ 10IU/l, a significant increase in the number of intermediate FMR1 alleles (35-54 CGG repeats) was found in patients compared with controls. In this group all the patients had a serum concentration > 40 IU/l. The results suggest that in the analysed Basque sample the FMR1 gene has a role in the aetiology of POF. However, elevated FSH levels are more related to the menstrual cycle pattern than to the CGG repeat size.

Dermatoglyphic variation in Spanish Basque populations.

The present study involves the evaluation of digital dermatoglyphic traits of 2185 unrelated individuals (1152 females and 1033 males) from 17 natural valleys of the four Basque provinces (Vizcaya, Guipúzcoa, Navarra, and Alava) in the Spanish Basque Country. Univariate intervalley and between-sex comparisons were carried out by means of chi-square contingency analysis for pattern types and by means of one-way analysis of variance for ridge counts. Multivariate intervalley comparison was carried out by means of correspondence analysis for pattern types and by principal component analysis for ridge counts. The results of this study are notable for the following findings: (1) in general, all variables are significantly heterogeneous among valley populations; (2) there was a greater differentiation among the valley populations than between sexes in one valley population; (3) affinities among the intervalley populations depend on the variables considered; (4) the valley populations from Vizcaya resemble those from the Pyrenees; (5) based on interprovince comparisons, the Vizcaya and Navarra samples are the closest: (6) in general, the valley samples from Alava are the worst clustered; (7) the universality of dermatoglyphic component structure fits better in males.

A new insight into fragile X syndrome among Basque population.

The expansion of a trinucleotide repeat [CGG]n located in the FMR1 X-linked gene is the main cause of fragile X syndrome, the most common form of inherited mental retardation. We have analyzed the factors known, to date, to influence the instability of the repeat in 158 normal X chromosomes from the Spanish Basque population. These factors included length of the repeat, AGG interspersion pattern, length of uninterrupted CGG and DXS548-FRAXAC1 markers associated haplotype. Previous investigations on Basques showed an absence of this disorder among mentally retarded individuals that was likely due to a low prevalence of large CGG alleles and the presence of AGG interruptions on them. The present report suggests that, although the frequency of large alleles is low and they do maintain AGG interruptions, different mutational pathways that might lead to fragile X syndrome could be occurring among Basques. These pathways mainly include alleles with internal sequences 9 + 9 + n and 9 + 12 + 9 that show fragile X associated haplotypes. Besides, the lack of the most proximal AGG interruption, proposed recently as a novel factor involved in CGG repeat instability, was highly identified among alleles with long pure CGG tracts, which showed an internal sequence n + 9. The data suggest that, despite the lower incidence of large alleles, the prevalence of potentially unstable alleles among Basques is similar to that of other Caucasian populations and that these alleles could become fragile X chromosomes.

Chromosomal fragility in a behavioral disorder.

Numerous studies have shown there is consistent evidence implicating genetic factors in the etiology of autism. In some cases chromosomal abnormalities have been identified. One type of these abnormalities is gaps and breaks nonrandomly located in chromosomes, denominated fragile sites (FS). We cytogenetically analyzed a group of autistic individuals and a normal population, and we examined the FS found in both samples with the aim of (1) comparing their FS expression, (2) ascertaining whether any FS could be associated with our autistic sample, and (3) examining if there are differences between individual and pooled-data analyses. Different statistical methods were used to analyse the FS of pooled and individual data. Our results show that there are statistically significant differences in the spontaneous expression of breakages between patients and controls, with a minimal sex difference. Using the method for pooled data, eight autosomal FS have preferential expression in patients and five patients were found to be positive at FS Xq27.3. With the method per-individual analysis, four FS emerged as specific in our autistic sample. Inferences of FS from pooled data were different from those of individual data. The findings suggest that although analysis of pooled data is necessitated by the problem of sparse data, analysis of single individuals is essential to know the significance of FS in autism.