FNDC5/Irisin polymorphisms are associated with osteopenia in postmenopausal Mayan-Mestizo women.

OBJECTIVE: This study aimed to analyze the association between rs3480 and rs16835198 of FNDC5/Irisin and their haplotypes with variations in bone mineral density (BMD) and osteopenia/osteoporosis in postmenopausal Mayan-Mestizo women. METHODS: We studied 547 postmenopausal women of Maya-Mestizo origin. BMD was measured in the lumbar spine and total hip by dual-energy X-ray absorptiometry. DNA was obtained from blood leukocytes. rs3480 and rs16835198 of FNDC5/Irisin were studied using real-time PCR allelic discrimination. Differences between the means of BMD according to genotype were analyzed with covariance. Allele frequency differences were assessed by χ2 and logistic regression was used to test for associations. Pairwise linkage disequilibrium between polymorphisms was calculated by direct correlation r2, and haplotype analysis was conducted. RESULTS: Under a recessive model, we observed a significant association of rs3480 with the presence of osteopenia at the total hip and femoral neck (p = 0.008 and p = 0.003, respectively). For rs16835198, we found an association with osteopenia at the total hip and femoral neck in a dominant model (p = 0.043 and p = 0.009, respectively). CONCLUSIONS: We found an association of rs3480 with risk to present osteopenia at the total hip and femoral neck, while rs16835198 was associated as a protector for presence of osteopenia only at the femoral neck.

The TGFß pathway is a key player for the endothelial-to-hematopoietic transition in the embryonic aorta.

The embryonic aorta produces hematopoietic stem and progenitor cells from a hemogenic endothelium localized in the aortic floor through an endothelial to hematopoietic transition. It has been long proposed that the Bone Morphogenetic Protein (BMP)/Transforming Growth Factor ß (TGFß) signaling pathway was implicated in aortic hematopoiesis but the very nature of the signal was unknown. Here, using thorough expression analysis of the BMP/TGFß signaling pathway members in the endothelial and hematopoietic compartments of the aorta at pre-hematopoietic and hematopoietic stages, we show that the TGFß pathway is preferentially balanced with a prominent role of Alk1/TgfßR2/Smad1 and 5 on both chicken and mouse species. Functional analysis using embryonic stem cells mutated for Acvrl1 revealed an enhanced propensity to produce hematopoietic cells. Collectively, we reveal that TGFß through the Alk1/TgfßR2 receptor axis is acting on endothelial cells to produce hematopoiesis.

(-)-Epicatechin inhibits development of dilated cardiomyopathy in δ sarcoglycan null mouse.

BACKGROUND AND AIMS: Several studies propose that (-)-epicatechin, a flavonol present in high concentration in the cocoa, has cardioprotective effects. This study aimed to evaluate the impact of (-)-epicatechin on the development of dilated cardiomyopathy in a δ sarcoglycan null mouse model. METHODS AND RESULTS: δ Sarcoglycan null mice were treated for 15 days with (-)-epicatechin. Histological and morphometric analysis of the hearts treated mutant mice showed significant reduction of the vasoconstrictions in the coronary arteries as well as fewer areas with fibrosis and a reduction in the loss of the ventricular wall. On the contrary, it was observed a thickening of this region. By Western blot analysis, it was shown, and increment in the phosphorylation level of eNOS and PI3K/AKT/mTOR/p70S6K proteins in the heart of the (-)-epicatechin treated animals. On the other hand, we observed a significantly decreased level of the atrial natriuretic peptide (ANP) and brain natriuretic peptide (BNP) heart failure markers. CONCLUSION: All the results indicate that (-)-epicatechin has the potential to prevent the development of dilated cardiomyopathy of genetic origin and encourages the use of this flavonol as a pharmacological therapy for dilated cardiomyopathy and heart failure diseases.

Polymer waveguide couplers based on metal nanoparticle-polymer nanocomposites.

In this work Au nanoparticles (AuNPs) are incorporated into poly(methyl methacrylate) (PMMA) waveguides to develop optical couplers that are compatible with planar organic polymer photonics. A method for growing AuNPs (of 10 to 100 nm in size) inside the commercially available Novolak resist is proposed with the intention of tuning the plasmon resonance and the absorption/scattering efficiencies inside the patterned structures. The refractive index of the MNP-Novolak nanocomposite (MNPs: noble metal nanoparticles) is carefully analysed both experimentally and numerically in order to find the appropriate fabrication conditions (filling factor and growth time) to optimize the scattering cross section at a desired wavelength. Then the nanocomposite is patterned inside a PMMA waveguide to exploit its scattering properties to couple and guide a normal incident laser light beam along the polymer. In this way, light coupling is experimentally demonstrated in a broad wavelength range (404-780 nm). Due to the elliptical shape of the MNPs the nanocomposite demonstrates a birefringence, which enhances the coupling to the TE mode up to efficiencies of around 1%.

Efficient excitation of photoluminescence in a two-dimensional waveguide consisting of a quantum dot-polymer sandwich-type structure.

In this Letter, we study a new kind of organic polymer waveguide numerically and experimentally by combining an ultrathin (10-50 nm) layer of compactly packed CdSe/ZnS core/shell colloidal quantum dots (QDs) sandwiched between two cladding poly(methyl methacrylate) (PMMA) layers. When a pumping laser beam is coupled into the waveguide edge, light is mostly confined around the QD layer, improving the efficiency of excitation. Moreover, the absence of losses in the claddings allows the propagation of the pumping laser beam along the entire waveguide length; hence, a high-intensity photoluminescence (PL) is produced. Furthermore, a novel fabrication technology is developed to pattern the PMMA into ridge structures by UV lithography in order to provide additional light confinement. The sandwich-type waveguide is analyzed in comparison to a similar one formed by a PMMA film homogeneously doped by the same QDs. A 100-fold enhancement in the waveguided PL is found for the sandwich-type case due to the higher concentration of QDs inside the waveguide.

Duchenne muscular dystrophy in a developing country: challenges in management and genetic counseling.

BACKGROUND AND OBJECTIVE: Multidisciplinary management of Duchenne Muscular Dystrophy (DMD) has achieved outstanding results in developed nations. We aimed to describe the status of diagnosis and management of DMD in a developing country through the experience of non-profit organizations. METHODS: A Multistate, multiple-source, population-based survey was performed from medical records of 432 patients. Data were retrospectively collected, reviewed and curated by health specialists; including clinical features, age at first symptoms, age at diagnosis, disease progression and management, family history, education, age and cause of death. RESULTS: There is a delay in noticing first symptoms and it did not diminish over the past 20 years. Less than 30% of patients obtained definite diagnosis and most of them are in physiotherapy programs but not under steroid treatment. In our study, family history does not anticipate recognition of symptoms compared to sporadic cases (p = 0.05). Approximately 93.33% of our patients attended to education programs. Mean age at death was 18.94 +/- 6.73 years and the most frequent cause was pneumonia. CONCLUSION: Delayed diagnosis of DMD in Mexico is mainly caused by the late detection of first symptoms. There is no difference in early detection of symptoms between familiar and sporadic cases. Lifespan of patients in our cohort is reduced compared to developed countries. The late diagnosis and low percentage of definite cases may affect patient management and genetic counseling and could also preclude participation of patients into novel clinical trials.

Demonstration of near infrared gas sensing using gold nanodisks on functionalized silicon.

In this work, we demonstrate experimentally the use of an array of gold nanodisks on functionalized silicon for chemosensing purposes. The metallic nanostructures are designed to display a very strong plasmonic resonance in the infrared regime, which results in highly sensitive sensing. Unlike usual experiments which are based on the functionalization of the metal surface, we functionalized here the silicon substrate. This silicon surface was modified chemically by buildup of an organosilane self-assembled monolayer (SAM) containing isocyanate as functional group. These groups allow for an easy surface regeneration by simple heating, thanks to the thermally reversible interaction isocyanate-analyte, which allows the cyclic use of the sensor. The technique showed a high sensitivity to surface binding events in gas and allowed the surface regeneration by heating of the sensor at 150 °C. A relative wavelength shift ∆λ(max)λ(0)=0.027 was obtained when the saturation level was reached.

Full-length SRY protein is essential for DNA binding.

SRY directs testicular development. It has been suggested that the only high-mobility group (HMG) box of the SRY is important for the function of this protein; however, other studies have suggested that the N- and C-terminal regions are also involved in this process. Herein, we analysed and compared in vitro the DNA-binding activity of the full-length SRY and three mutants (HMG box alone, N-terminal less and C-terminal less SRY proteins). DNA-binding capability was analysed by mobility shift assays, optical density and dissociation constant by using pure non-fusion SRY proteins. The structure of the full-length SRY was carried out using a protein molecular model. The HMG box SRY alone and C-terminal less SRY proteins had a statistically diminished DNA binding in comparison with the full-length SRY. In contrast, the affinity for DNA of the N-terminal less SRY was relatively similar to the full-length SRY. Likewise, three-dimensional structure of the full-length SRY suggested that some residues of the C-terminal region of the SRY interact with DNA. We demonstrate the importance that full-length SRY has, particularly the C-terminal region of the protein, in DNA binding in vitro. Likewise, the affinity of the HMG box alone is clearly reduced when compared with the full-length SRY.

Association of the rs1801133 variant in the MTHFR gene and sporadic Parkinson's disease.

The MTHFR gene has been reported as a susceptibility locus for sporadic Parkinson's disease (sPD). The functional variant rs1801133 has been linked to hyperhomocysteinemia and dopaminergic cell death. Among different populations, Mexican-Mestizos (most present-day Mexicans) have the highest frequency of this variant. Therefore, we sought to determine a possible association of rs1801133 with SPD. In total, 356 individuals were included: 140 patients with PD, diagnosed according to the Queen Square Brain Bank criteria, and 216 neurologically healthy controls. Genotyping was performed using TaqMan probes for rs1801133 and real-time PCR. Logistic regression analysis with adjustment for smoking and gender was used to test for an association between genotype and SPD. The CC genotype was associated with SPD; exp() = 2.06; 95% CI: 1.101-3.873, p = 0.024. No association with age at onset, cognitive impairment or gender was found in our study group. Our data suggest an important role of MTHFR gene variants in SPD.

Identification of three novel mutations in the KAL1 gene in patients with Kallmann syndrome.

Kallmann's syndrome (KS) is characterized by the association of hypogonadotropic hypogonadism and anosmia or hyposmia. Genetic defects have been observed throughout the KAL1 gene, located on the Xp22.3 region, in less than 50% of the patients. We report the molecular study of the KAL1 gene in 12 males with KS. PCR of the 14 exons of the KAL1 gene was performed on genomic DNA. PCR products of all exons were purified and sequenced. Three novel genetic defects were found. One patient exhibited a complete deletion of exon 5. The second presented a duplication of nucleotides 158-168; this insertion causes a termination codon (TGA) within the same exon. The third presented a mutation in exon 6, in which codon 262 changes from arginine to a stop codon. In the remaining nine individuals, no mutations were found. Three previously reported polymorphic changes were also documented. The deletion of exon 5 occurs within the region encoding the first fibronectin type III-like repeat of the KAL1 protein, this being the first KS patient who exhibits a complete deletion of a single exon of the KAL1 gene. The duplication of nucleotides in exon 1 is located in the conserved cysteine-rich N-terminal region that corresponds to the whey acidic protein motif, affecting the KAL1 protein either by interrupting the normal transcription or stopping the translation at the stop codon. The last novel mutation, a stop codon in exon 6, is located within the region encoding the first fibronectin type III-like repeat of the KAL1 protein. The absence of mutations in the majority of patients suggests the possibility of the existence of other genes involved or that in certain individuals the presence of various polymorphisms within the KAL1 gene could predispose to disease, as has been demonstrated in other pathological entities.

Mutations in the desert hedgehog (DHH) gene in patients with 46,XY complete pure gonadal dysgenesis.

Mutations of SRY are the cause of complete pure gonadal dysgenesis (PGD) in 10-15% of patients. In the remaining individuals, it has been suggested that mutations in other genes involved in the testis-determining pathway could be causative. We describe the first report in which three cases of 46,XY complete PGD are attributed to mutations of the Desert hedgehog (DHH) gene. DHH was sequenced using genomic DNA from paraffin-embedded gonadal tissue from six patients with complete 46,XY PGD. Mutations were found in three patients: a homozygous mutation in exon 2, responsible for a L162P, and a homozygous 1086delG in exon 3. Mutated individuals displayed 46,XY complete PGD, differentiating from the only previously described patient with a homozygous DHH mutation, who exhibited a partial form of PGD with polyneuropathy, suggesting that localization of mutations influence phenotypic expression. This constitutes the first report where mutations of DHH are associated with the presence of 46,XY complete PGD, demonstrating that the genetic origin of this entity is heterogeneous and that disorders in other genes, different from SRY, involved in the testis-determining pathway are implicated in abnormal testicular differentiation in humans. These data extend previous reports demonstrating DHH is a key gene in gonadal differentiation.

Evidence for genetic heterogeneity in male pseudohermaphroditism due to Leydig cell hypoplasia.

Leydig cell aplasia or hypoplasia is a rare form of male pseudohermaphroditism resulting from inadequate fetal testicular Leydig cell differentiation. Affected individuals presented a wide phenotypic spectrum, ranging from complete female external genitalia to males with micropenis. Recessive mutations in the LH receptor gene have been identified as responsible for the condition. The majority of these mutations are point mutations and have been located in exon 11 of the gene. In this study, we report the molecular characterization of the LH receptor gene in three siblings with Leydig cell hypoplasia. After sequencing the 11 exons of the gene, no deleterious mutations were detected in any patient. However, we identified a previously described polymorphism in exon 11. In patients 1 and 3 DNA sequencing revealed a C to T substitution at nucleotide 1065; both patients were homozygous GAT/GAT at codon 355. In contrast, patient 2 was homozygous GAC/GAC, whereas the father and one unaffected sister were heterozygous GAC/GAT at this polymorphic site. These results exclude that Leydig cell hypoplasia in this family is due to a mutation in the LH receptor gene and provide evidence that defects in other loci may also result in failure of Leydig cell differentiation, demonstrating, for the first time, that Leydig cell hypoplasia is a genetically heterogeneous condition.

A mutation in the 5' non-high mobility group box region of the SRY gene in patients with Turner syndrome and Y mosaicism.

In Ullrich-Turner syndrome (UTS) patients, the presence of a Y-chromosome or Y-derived material has been documented in frequencies ranging from 4-61%. Mutations of SRY (testis-determining gene) constitute the cause of XY sex reversal in approximately 10-15% of females with pure gonadal dysgenesis. Most of these mutations have been described in the HMG (high mobility group) box of the gene, which is the region responsible for DNA binding and bending; however, various mutations outside the HMG box have been reported. We carried out molecular studies of the SRY gene in three patients with a UTS phenotype and bilateral streaks; two presented a 45,X/46,XY mosaic, and the third a Y marker chromosome. In two patients a missense mutation, S18N, was identified in the 5' non-HMG box region in DNA from blood and both streaks; this mutation was not identified in 75 normal males. Sequencing of the DNA region of interest was normal in the father and older brother of patient 1, demonstrating that in this patient the mutation was de novo. A previous report of a 46,XY patient with partial gonadal dysgenesis who presented the same mutation as our patients indicates the probable existence of a hot spot in this region of the SRY gene and strengthens the possibility that all gonadal dysgeneses constitute part of a spectrum of the same disorder. It also demonstrates that a single genetic abnormality can result in a wide range of phenotypic expression.

A reliable endocrine test with human menopausal gonadotropins for diagnosis of true hermaphroditism in early infancy.

In true hermaphroditism diverse phenotypes and karyotypes are found; there are no distinctive laboratory features that can distinguish it from other intersex disorders, thus the diagnosis is made by the histological findings. Existence of Leydig cells is demonstrated by testosterone levels above the female range; however, presence of ovarian tissue cannot be ascertained because of the absence of a reliable functional test. Unless appropriate biopsies are performed or the whole gonad is removed, there is a risk of not diagnosing true hermaphroditism. To find a reliable test that can differentiate patients with true hermaphroditism from those with other intersex disorders, we investigated the estradiol (E2) response to human menopausal gonadotropins (hMG) in infants with genital ambiguity. These results were correlated with the histological findings. Eleven infants with genital ambiguity and four with a high scrotal testis were stimulated every 12 h with 2 IU/kg hMG. If E2 rose above 80 pg/mL (cut-off point), the test was discontinued; if after 7 days E2 remained below 80 pg/mL, the hMG dose was doubled and stimulation extended for 7 additional days. In five patients in whom true hermaphroditism was later histologically demonstrated, E2 rose above 80 pg/mL. In two of them, ovarian tissue was removed and hMG stimulation repeated; no response above our cut-off point was observed during the second test. The maximal E2 response to hMG in the remaining 10 individuals was 43 pg/mL; after laparotomy or gonadal biopsies no ovarian tissue was found. The hMG stimulation test can be considered a reliable and safe dynamic procedure for demonstrating the presence or absence of ovarian tissue in infants with genital ambiguity.

No evidence of mutations in the follicle-stimulating hormone receptor gene in Mexican women with 46,XX pure gonadal dysgenesis.

In the ovary FSH is necessary for normal follicular development, binding to its receptor (FSHR) that pertains to the superfamily of G-protein coupled receptors. In the FSHR gene, which consists of 10 exons, an homozygous mutation was reported in six Finnish families with gonadal dysgenesis; whereas two isolated French patients exhibited compound heterozygous mutations. Several groups, however, have searched for FSHR mutations, although in most cases the gene has been studied partially, not finding any genetic abnormalities in German, English, North American or Brazilian women. We performed direct sequencing of all 10 exons of the FSHR gene in seven sporadic patients and two sisters with 46,XX pure gonadal dysgenesis, to investigate the cause of their disorder. No heterozygous or homozygous mutant alleles were present in any of the patients. Although the number of patients evaluated was small, considering all the other previous reports, it seems that except in the Finnish population, the proportion of women with mutations in the encoding region of this gene is very low. Other possibilities for the presence of 46,XX gonadal dysgenesis, such as defects in the regulatory regions of the FSHR gene promoter, in the untranslated regions of exons 1 and 10, and within introns, or the existence of other genes likely to be important for normal ovarian function on the X chromosome or on autosomes, should be considered. In contrast with other studies, we did not find polymorphisms of the FSHR gene, indicating that apparently in Mexicans this gene is not highly polymorphic.

Clinical expression and SRY gene analysis in XY subjects lacking gonadal tissue.

In several syndromes genetic males lack gonadal tissue. A range of phenotypes are seen, which varies from complete female external genitalia to anorchic subjects with sexual infantilism. Differences in phenotypic expression depend on the stage at which testes degenerated during intrauterine development. Although most cases of these syndromes are sporadic, several instances of familial recurrence suggest a genetic origin. To help elucidate the source, we performed molecular analysis of the complete SRY gene open reading frame in two subjects with true agonadism and in two with anorchia. Our results add to previous findings indicating that molecular defects in SRY are not readily identified as a cause of these syndromes.

Molecular analysis of the 5 alpha-steroid reductase type 2 gene in a family with deficiency of the enzyme.

This report describes the identification of a point mutation in the 5 alpha-reductase type 2 (5 alpha-SR2) gene from a family in which both sibs (6 and 3 years old) have steroid 5 alpha-reductase 2 deficiency. The five exons of the gene were individually amplified by the polymerase chain reaction (PCR) and analysed for single-strand conformation polymorphisms (SSCP) to detect mutations. Direct sequencing of the mutant PCR products demonstrated a single C-->T mutation, within exon 4, changing codon 227 from CGA (Arg) to TGA (premature termination signal). Both patients were homozygous for the mutation, but their parents were heterozygous. These results suggest that the mutation at codon 227 impairs normal 5 alpha-SR2 function, thus leading to the phenotypical expression of this rare enzymatic defect.

Partially deleted SRY gene confined to testicular tissue in a 46,XX true hermaphrodite without SRY in leukocytic DNA.

True hermaphroditism is an uncommon form of intersexuality in which testicular and ovarian tissue develop in the same individual. Most true hermaphrodites are 46,XX and lack SRY, the testis-determining gene. We describe results of molecular studies performed in a 46,XX true hermaphrodite SRY-negative in DNA from blood leukocytes but SRY-positive in DNA obtained from the testicular portion of the ovotestis. Surprisingly, the SRY identified in gonadal DNA carries a partial deletion at the 5' end of the gene. Our patient is the first case of a naturally occurring deletion within the SRY ORF (with a normal HMG box) and provides a new explanation for the abnormal gonadal development observed in 46,XX true hermaphrodites.

Molecular analysis in true hermaphrodites with different karyotypes and similar phenotypes.

True hermaphroditism is characterized by the development of ovarian and testicular tissue in the same individual. Müllerian and Wolffian structures are usually present, and external genitalia are often ambiguous. The most frequent karyotype in these patients is 46,XX or various forms of mosaicism, whereas 46,XX is very rarely found. The phenotype in all these subjects is similar. We studied 10 true hermaphrodites. Six of them had a 46,XX chromosomal complement: 3 had been reared as males and 3 as females. The other 4 patients were mosaics: 3 were 46,XX/46,XY and one had a 46,XX/47,XXY karyotype. One of the 46,XX/46,XY mosaics was reared as a female, whereas the other 3 mosaics were reared as males. The sex of assignment in the 10 patients depended only on labio-scrotal differentiation. Molecular studies in 46,XX subjects documented the absence of Y centromeric sequences in all cases, arguing against hidden mosaicism. One patient presented Yp sequences (ZFY+, SRY+), which contrast with South African black 46,XX true hermaphrodites in whom no Y sequences were found. Molecular analysis in the subjects with mosaicism demonstrated the presence of Y centromeric and Yp sequences confirming the presence of a Y chromosome. Gonadal development, endocrine function, and phenotype in the 10 patients did not correlate with the presence of a Y chromosome or Y-derived sequences in the genome, confirming that true hermaphroditism is a heterogeneous condition.

Frequency of Y chromosomal material in Mexican patients with Ullrich-Turner syndrome.

Cytogenetic studies have shown that 40-60% of patients with Ullrich-Turner syndrome (UTS) are 45,X, whereas the rest have structural aberrations of the X chromosome or mosaicism with a second cell line containing a structurally normal or abnormal X or Y chromosome. However, molecular analysis has demonstrated a higher proportion of mosaicism, and studies in different populations have shown an extremely variable frequency of Y mosaicism of 0-61%. We used Southern blot analysis and polymerase chain reaction (PCR) to detect the presence of Ycen, ZFY, SRY, and Yqh in 50 Mexican patients with UTS and different karyotypes to determine the origin of marker chromosomes and the presence of Y sequences. Our results indicated the origin of the marker chromosome in 1 patient and detected the presence of Y sequences in 4 45,X patients. Taken together, we found a 12% incidence of Y sequences in individuals with UTS. The amount of Y-derived material was variable, making the correlation between phenotype and molecular data difficult. Only 1 patient had a gonadoblastoma. We discuss the presence of Y chromosomes or Y sequences in patients with UTS and compare our frequency with that previously reported.