Identification of a rare copy number polymorphic gain at 3q12.2 with candidate genes for familial endometriosis.

Endometriosis is a complex disease that affects 10-15% of women of reproductive age. Familial studies show that relatives of affected patients have a higher risk of developing the disease, implicating a genetic role for this disorder. Little is known about the impact of germline genomic copy number variant (CNV) polymorphisms on the heredity of the disease. In this study, we describe a rare CNV identified in two sisters with familial endometriosis, which contain genes that may increase the susceptibility and progression of this disease. We investigated the presence of CNVs from the endometrium and blood of the sisters with endometriosis and normal endometrium of five women as controls without the disease using array-CGH through the Agilent 2x400K platform. We excluded common CNVs that were present in the database of genomic variation. We identified, in both sisters, a rare CNV gain affecting 113kb at band 3q12.2 involving two candidate genes: ADGRG7 and TFG. The CNV gain was validated by qPCR. ADGRG7 is located at 3q12.2 and encodes a G protein-coupled receptor influencing the NF-kappaß pathway. TFG participates in chromosomal translocations associated with hematologic tumor and soft tissue sarcomas, and is also involved in the NF-kappa B pathway. The CNV gain in this family provides a new candidate genetic marker for future familial endometriosis studies. Additional longitudinal studies of affected families must confirm any associations between this rare CNV gain and genes involved in the NF-kappaß pathway in predisposition to endometriosis.

Identification of a rare copy number polymorphic gain at 3q12.2 with candidate genes for familial endometriosis

Abstract Endometriosis is a complex disease that affects 10-15% of women of reproductive age. Familial studies show that relatives of affected patients have a higher risk of developing the disease, implicating a genetic role for this disorder. Little is known about the impact of germline genomic copy number variant (CNV) polymorphisms on the heredity of the disease. In this study, we describe a rare CNV identified in two sisters with familial endometriosis, which contain genes that may increase the susceptibility and progression of this disease. We investigated the presence of CNVs from the endometrium and blood of the sisters with endometriosis and normal endometrium of five women as controls without the disease using array-CGH through the Agilent 2x400K platform. We excluded common CNVs that were present in the database of genomic variation. We identified, in both sisters, a rare CNV gain affecting 113kb at band 3q12.2 involving two candidate genes: ADGRG7 and TFG. The CNV gain was validated by qPCR. ADGRG7 is located at 3q12.2 and encodes a G protein-coupled receptor influencing the NF-kappaβ pathway. TFG participates in chromosomal translocations associated with hematologic tumor and soft tissue sarcomas, and is also involved in the NF-kappa B pathway. The CNV gain in this family provides a new candidate genetic marker for future familial endometriosis studies. Additional longitudinal studies of affected families must confirm any associations between this rare CNV gain and genes involved in the NF-kappaβ pathway in predisposition to endometriosis.

Novel pathogenic variants and quantitative phenotypic analyses of Robinow syndrome: WNT signaling perturbation and phenotypic variability.

Robinow syndrome (RS) is a genetically heterogeneous disorder with six genes that converge on the WNT/planar cell polarity (PCP) signaling pathway implicated (DVL1, DVL3, FZD2, NXN, ROR2, and WNT5A). RS is characterized by skeletal dysplasia and distinctive facial and physical characteristics. To further explore the genetic heterogeneity, paralog contribution, and phenotypic variability of RS, we investigated a cohort of 22 individuals clinically diagnosed with RS from 18 unrelated families. Pathogenic or likely pathogenic variants in genes associated with RS or RS phenocopies were identified in all 22 individuals, including the first variant to be reported in DVL2. We retrospectively collected medical records of 16 individuals from this cohort and extracted clinical descriptions from 52 previously published cases. We performed Human Phenotype Ontology (HPO) based quantitative phenotypic analyses to dissect allele-specific phenotypic differences. Individuals with FZD2 variants clustered into two groups with demonstrable phenotypic differences between those with missense and truncating alleles. Probands with biallelic NXN variants clustered together with the majority of probands carrying DVL1, DVL2, and DVL3 variants, demonstrating no phenotypic distinction between the NXN-autosomal recessive and dominant forms of RS. While phenotypically similar diseases on the RS differential matched through HPO analysis, clustering using phenotype similarity score placed RS-associated phenotypes in a unique cluster containing WNT5A, FZD2, and ROR2 apart from non-RS-associated paralogs. Through human phenotype analyses of this RS cohort and OMIM clinical synopses of Mendelian disease, this study begins to tease apart specific biologic roles for non-canonical WNT-pathway proteins.

Smoking-induced chromosomal segregation anomalies identified by FISH analysis of sperm.

BACKGROUND: Numerical chromosome aberrations in gametes are directly related to infertility and aneuploid embryos. Previous studies have shown that toxic substances from cigarette smoke induce structural and numerical chromosomal aberrations in vitro and could potentially increase levels of aneusomy in sperm. Moreover, increased levels of aneusomy in sperm are correlated with low implantation rates, spontaneous abortions and fetal losses. Studies of chromosome 3 in sperm suggest it may be more prone to segregation anomalies than other autosomes, but there has been no systematic investigation of the incidence of disomy for chromosome 3 in sperm derived from donor male smokers. The objective of this study was to use FISH to evaluate the influence of smoking on the levels of disomy for chromosomes X and Y, and to determine whether disomy levels for chromosome 3 were elevated in sperm derived from male smokers. RESULTS: FISH analysis was used to evaluate the frequency of disomies of chromosomes 3, X, and Y in sperm of 10 smokers, compared to a control group of 7 non-smoking fertile men. All the subjects presented a normal somatic karyotype. There was a significant increase in the overall frequency of disomies in sperm derived from the smoking group (P< 0.0001). When each chromosome pair was analyzed individually, disomy of chromosome 3 in smokers was found to be more than twice that observed in the matched non-smoker control group. In addition we observed a higher frequencies of disomy of the X and Y chromosomes, indicating elevated levels of diploidy in the sperm from the smoking group. CONCLUSIONS: In this study we have shown that chromosome 3 may be susceptible to smoking-related segregation anomalies. Our results also suggest that errors can occur in both meiosis I and II, confirming the emerging literature that the male meiotic process may generally be affected by the genotoxic damage from tobacco use. Collectively, these findings provide additional evidence for enhancing tobacco control measures, and suggest that FISH analysis of chromosome 3 in sperm may be useful for monitoring smoking-induced segregation damage as part of the evaluation of infertile males.

Deregulation of LOXL1 and HTRA1 gene expression in endometriosis.

Endometriosis is a gynecologic disease characterized by the presence of endometrial tissue outside the uterine cavity. Although 15% of the female population in reproductive age is affected by endometriosis, its pathogenesis remains unclear. According to the most accepted pathogenesis hypothesis, endometrial fragments from the menstrual phase are transported through the uterine tubes to the peritoneal cavity, where they undergo implantation and growth, invading adjacent tissues. However, the establishment of the disease requires that endometrial cells present molecular characteristics favoring the onset and progression of ectopic implantation. In this investigation, we analyzed the differential gene expression profiles of peritoneal and ovarian endometriotic lesions compared to the endometrial tissue of nonaffected women using rapid subtraction hybridization (RaSH). In our study, this method was applied to samples of endometriotic lesions from affected women and to biopsies of endometrium of healthy women without endometriosis, where we could identify 126 deregulated genes. To evaluate the expression of genes found by RaSH method, we measured LOXL1, HTRA1, and SPARC genes by real-time polymerase chain reaction. Significant different expression was obtained for HTRA1 and LOXL1, upregulated in the ectopic endometrium, suggesting that these genes are involved in the physiopathology of endometriosis and may favor the viability of endometrial cells at ectopic sites.

Differentially expressed genes in eutopic and ectopic endometrium of women with endometriosis.

OBJECTIVE: To elucidate the potential mechanisms involved in the physiopathology of endometriosis. We analyzed the differential gene expression profiles of eutopic and ectopic tissues from women with endometriosis. DESIGN: Prospective laboratory study. SETTING: University hospital. PATIENT(S): Seventeen patients in whom endometriosis was diagnosed and 11 healthy fertile women. INTERVENTION(S): Endometrial biopsy specimens from the endometrium of healthy women without endometriosis and from the eutopic and ectopic endometrium tissues of patients with endometriosis were obtained in the early proliferative phase of the menstrual cycle. MAIN OUTCOME MEASURE(S): Six paired samples of eutopic and ectopic tissue were analyzed by subtractive hybridization. To evaluate the expression of genes found by rapid subtraction hybridization methods, we measured CTGF, SPARC, MYC, MMP, and IGFBP1 genes by real-time polymerase chain reaction in all samples. RESULT(S): This study identified 291 deregulated genes in the endometriotic lesions. Significant expression differences were obtained for SPARC, MYC, and IGFBP1 in the peritoneal lesions and for MMP3 in the ovarian endometriomas. Additionally, significant differences were obtained for SPARC and IGFBP1 between the peritoneal and ovarian lesions. No significant differences were found for the studied genes between the control and the eutopic endometrium. CONCLUSION(S): This study identified 291 genes with differential expression in endometriotic lesions. The deregulation of the SPARC, MYC, MMP3, and IGFBPI genes may be responsible for the loss of cellular homeostasis in endometriotic lesions.

Association between MUC1 gene polymorphism and expected progeny differences in Nelore cattle (Bos primigenius indicus)

MUC1 is a heavily glycosylated mammalian transmembrane protein expressed by mucosal secretory tissues for both protection against microbial infection and lubrication. An important characteristic of MUC1 is its variable number of tandem repeats (VNTR) containing several sites for O-glycosylation. VNTR length has been associated with many human diseases and with certain economically important traits in domestic ruminants. The aim of the present study was to correlate the length of MUC1 gene VNTR with expected progeny differences (EPDs) obtained for growth, fertility and carcass traits. Five alleles were identified, with alleles containing short VNTRs being more frequent than those with long, thereby demonstrating that Brazilian Nelore cattle are characterized by high frequencies in short MUC1 VNTRs. Statistical analyses revealed there to be no significant association between VNTR length and EPDs for weight at 120 days (W120), scrotal circumference at 365 (SC365) and 450 (SC450) days, age at first calving (AFC), and rib eye area (REA).

A retrospective model of oocyte competence: global mRNA and housekeeping transcripts are not associated with in vitro developmental outcome.

SummaryOocyte developmental competence depends on maternal stores that support development throughout a transcriptionally silent period during early embryogenesis. Previous attempts to investigate transcripts associated with oocyte competence have relied on prospective models, which are mostly based on morphological criteria. Using a retrospective model, we quantitatively compared mRNA among oocytes with different embryo development competence. A cytoplasm biopsy was removed from in vitro matured oocytes to perform comparative analysis of amounts of global polyadenylated (polyA) mRNA and housekeeping gene transcripts. After parthenogenetic activation of biopsied oocytes, presumptive zygotes were cultured individually in vitro and oocytes were classified according to embryo development: (i) blocked before the 8-cell stage; (ii) blocked between the 8-cell and morulae stages; or (iii) developed to the blastocyst stage. Sham-manipulated controls confirmed that biopsies did not alter development outcome. Total polyA mRNA amounts correlate with oocyte diameter but not with the ability to develop to the 8-cell and blastocyst stages. The last was also confirmed by relative quantification of GAPDH, H2A and Hprt1 transcripts. In conclusion, we describe a novel retrospective model to identify putative markers of development competence in single oocytes and demonstrate that global mRNA amounts at the metaphase II stage do not correlate with embryo development in vitro.

Glycodelin expression in the endometrium of healthy women and in the eutopic and ectopic endometrium of women with endometriosis.

OBJECTIVE: To analyze the expression of the glycodelin gene to better understand the molecular environment of endometriotic lesions and to elucidate the potential mechanisms that underlie the complex physiopathology of endometriosis. DESIGN: Prospective laboratory study. SETTING: University hospital. PATIENT(S): Eleven healthy fertile women and 17 patients with endometriosis in the early proliferative phase of the menstrual cycle. INTERVENTION(S): Endometrial biopsy specimens were obtained from the endometrium of healthy women without endometriosis (controls) and from eutopic and ectopic endometrium tissues (pelvic and ovarian endometriotic implants) of endometriosis patients. MAIN OUTCOME MEASURE(S): The glycodelin relative expression level by real-time polymerase chain reaction (PCR) analysis. RESULT(S): The glycodelin down-regulation found in the endometriotic lesions was 332.26 and 123.17-fold lower, respectively, when compared with the eutopic tissue and the control endometrium. CONCLUSION(S): Glycodelin may be one of the molecules that contributes to the loss of cellular homeostasis in endometriotic lesions.

Widening the clinical spectrum of Pitt-Rogers-Danks/Wolf-Hirschhorn syndromes

Chromosomal rearrangements involving partial deletion of the short arm of chromosome 4 and partial duplication of the short arm of chromosome 8 have been described both in Pitt-Rogers-Danks syndrome (PRDS) and Wolf-Hirschhorn syndrome (WHS), the former being considered a milder phenotype of the latter. We describe a patient with partial deletion of chromosome 4 and partial duplication of chromosome 8 documented by array-comparative genomic hybridization (Array-CGH). In addition to the typical features of PRDS, the patient exhibited some clinical signs (genital hypoplasia, radioulnar synostosis and mesomelic limb shortness) infrequently, or never previously, reported in PRDS. These findings broaden the spectrum of anomalies generally associated with these syndromes.

Mitochondrial DNA single nucleotide polymorphism associated with weight estimated breeding values in Nelore cattle (Bos indicus)

We sampled 119 Nelore cattle (Bos indicus), 69 harboring B. indicus mtDNA plus 50 carrying Bos taurus mtDNA, to estimate the frequencies of putative mtDNA single nucleotide polymorphisms (SNPs) and investigate their association with Nelore weight and scrotal circumference estimated breeding values (EBVs). The PCR restriction fragment length polymorphism (PCR-RFLP) method was used to detect polymorphisms in the mitochondrial asparagine, cysteine, glycine, leucine and proline transporter RNA (tRNA) genes (tRNAasn, tRNAcys, tRNAgly, tRNAleu and tRNApro). The 50 cattle carrying B. taurus mtDNA were monomorphic for all the tRNA gene SNPs analyzed, suggesting that they are specific to mtDNA from B. indicus cattle. No tRNAcys or tRNAgly polymorphisms were detected in any of the cattle but we did detect polymorphic SNPs in the tRNAasn, tRNAleu and tRNApro genes in the cattle harboring B. indicus mtDNA, with the same allele observed in the B. taurus sequence being present in the following percentage of cattle harboring B. indicus mtDNA: 72.46 percent for tRNAasn, 95.23 percent for tRNAleu and 90.62 percent for tRNApro. Analyses of variance using the tRNAasn SNP as the independent variable and EBVs as the dependent variable showed that the G -> T SNP was significantly associated (p < 0.05) with maternal EBVs for weight at 120 and 210 days (p < 0.05) and animal's EBVs for weight at 210, 365 and 455 days. There was no association of the tRNAasn SNP with the scrotal circumference EBVs. These results confirm that mtDNA can affect weight and that mtDNA polymorphisms can be a source of genetic variation for quantitative traits.

Down syndrome and precocious menopause.

PURPOSE: To determine whether women who had children with Down syndrome (DS) had precocious menopause. METHODS: We selected 104 mothers of children with DS and 121 normal women who had children with no genetic problems. We conducted an interview and compared their mean age at menopause. RESULTS: We did not detect a statistically significant difference in mean age at menopause. When we stratified into women who had conceived when younger or older than 35 years of age, we found a statistically significant difference only for women who had conceived at the age of 35 years or older. We observed three cases of previous unilateral ovarian surgery in the study group and one case in the control group. CONCLUSIONS: We cannot conclude that mothers of children with DS will have precocious menopause. Nevertheless, our findings do not exclude the theory of reduced ovarian reserve as a primordial factor in the genesis of DS.

Analysis of restriction fragment length polymorphism in the kappa-casein gene related to weight expected progeny difference in Nellore cattle

Restriction fragment length polymorphism (RFLP) has been detected at the bovine kappa-casein locus. The polymorphism has been analyzed for its effects in cattle production, mostly for milk traits and even for maternal effect on pre-weaning weights. We used polymerase chain reaction - restriction fragment length polymorphism (PCR-RFLP) to genotype 408 Nellore animals for the non-silent mutation (Thr/Ile136 and Asp/Ala148) that characterizes the A and B variants of the polymorphism and compared expected progeny difference (EPD) for a maternal effect on 120 and 210 days weights and direct EPD for 120, 210, 450 and 550 day weight between AA and AB animals. The EPD values were obtained from the University of São Paulo (Brazil) Nellore Cattle Breeding Program, which evaluated 266,272 animals in 2001. Analysis of Variance was used to compare weight expected progeny differences (EPDs) between animals genotyped as AA and AB. The A allele frequency was 0.911. Although the AA animals had higher weight EPDs than AB animals the differences were not statistically significant (p > 0.05).

Use of the TSPY gene for sexing cattle

The Y-encoded, testis-specific protein (TSPY) is a Y-specific gene. The copy numbers of TSPY range from 20 to 60 in men and up to 200 in bulls. In this study, we examined the possibility of using the TSPY gene to sex cattle. DNA from blood samples of 100 Nelore cattle (50 males and 50 females) from the Nelore Cattle Breeding Program (PMGRN) was screened for TSPY by PCR using TSPY-specific primers. The assay was highly specific since all male samples were TSPY-positive and all female samples were negative. Positive results were also obtained at low DNA concentrations (less than 1 rhog/muL). These results showed that TSPY was a good male-specific marker, the usefulness of which was enhanced by the high copy number of the gene. This is the first report to demonstrate the applicability of TSPY for sexing cattle.

Trissomia 13 e pré-eclâmpsia / Trisomy 13 and preeclampsia

Um total de quatro recém-nascidos com trissomia 13 (80 por cento do total de casos) foi estudado retrospectivamente, assim como suas genitoras, no que concerne à associaçao entre trissomia 13 e pré-eclâmpsia. Os parâmetros analisados referiram-se aos dados de pré-natal concernentes aos níveis pressóricos, intercorrências clínico-obstétricas e idade materna. Para cada criança com trissomia 13, foram escolhidas, aleatoriamente, quatro crianças com cariótipo normal. A incidência de trissomias 13 mostrou-se discretamente elevada no nosso meio (1,4/10.000 recém-nascidos vivos). A idade materna dos recém-nascidos trissômicos mostrou-se significativamente mais, elevada que a do grupo-controle, condizente com os achados vigentes da literatura, onde a idade materna avançada é fator importante na etiologia das trissomias. A análise da série revelou associaçao entre gestaçoes com trissomia 13 e pré-eclâmpsia (p

Síndrome da insensibilidade androgênica: estudo comparativo entre as formas completa e incompleta / Androgen insensitivity syndrome: a comparative study between the complete and the incomplete forms

Com o objetivo de apresentar o perfil hormonal, a avaliaçäo diagnóstica e a conduta terapêutica na Síndrome da Insensibilidade Androgênica (SIA), realizamos um estudo de 15 pacientes, incluindo 10 com a forma completa e 5 com a forma incompleta. O diagnóstico foi firmado durante ou após a puberdade em 60 por cento dos casos. Gonadectomia bilateral foi realizada à época do diagnóstico na forma incompleta, e após a puberdade na forma completa. Estrogenioterapia foi mantida na puberdade após gonadectomia. Considerando o perfil hormonal, os níveis de LH estavam elevados em ambos os grupos, o FSH somente na forma incompleta e o E2, T e PRL. Após gonadectomia, o LH e FSH elevaram-se e a T diminuiu, significativamente.

46, XX, r (13)/46, XX, iso psu dic (13)? mosaicism

Os autores descrevem um caso raro de mosaicismo cromossômico 46,XX, r (13)/46, XX, iso psu dic (13)? em uma paciente que apresentava sinais clínicos semelhantes aos observados na síndrome do anel do cromossomo 13 (categoria 2a de Niebuhr, 1977). Discutem a hipótese de uma origem comum para as duas linhagens celulares e comparam os achados clínicos e citogenéticos aos descritos na literatura