Lúcia Martelli

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.

Clinical characterization of autosomal dominant and recessive variants of Robinow syndrome

Robinow syndrome is a genetically heterogeneous condition characterized by mesomelic limb shortening associated with facial and genital anomalies that can be inherited in an autosomal dominant or recessive mode. We characterized these two variants clinically, with the aim of establishing clinical criteria to enhance the differential diagnosis between them or other similar conditions. The frequencies of clinical signs considered important for the discrimination of the dominant or recessive variants were estimated in a sample consisting of 38 patients personally examined by the authors and of 50 affected subjects from the literature. Using the presence of rib fusions as diagnostic of the recessive variant, and also based on the inheritance pattern in familial cases, we classified 37 patients as having the recessive form and other 51 as having the dominant form. The clinical signs present in more than 75% of patients with either form, and therefore the most important for the characterization of this syndrome were hypertelorism, nasal features (large nasal bridge, short upturned nose, and anteverted nares), midface hypoplasia, mesomelic limb shortening, brachydactyly, clinodactyly, micropenis, and short stature. Hemivertebrae and scoliosis were present in more than 75% of patients with the recessive form, but in less than 25% of patients with the dominant form. Umbilical hernia (32.3%) and supernumerary teeth (10.3%) were found exclusively in patients with the dominant form.

SRY-negative true hermaphrodites and an XX male in two generations of the same family

Two 46,XX true hermaphrodites and one XX male without genital ambiguities are reported. They coexist in two generations of the same pedigree, with paternal transmission and in the absence of SRY (sex-determining region, Y chromosome). These familial cases provide evidence to support the hypothesis that these disorders are alternative manifestations of the same genetic defect, probably an autosomal dominant mutation (with incomplete penetrance) or an X-linked mutation (limited by the presence of the Y chromosome).

Recurrent 22q11.2 deletion in a sibship suggestive of parental germline mosaicism in velocardiofacial syndrome.

Deletions of chromosome 22q11.2 are recognized as the main cause of a number of clinical phenotypes, including velocardiofacial syndrome (VCFS) and DiGeorge syndrome (DGS). Velocardiofacial syndrome is a relatively common developmental disorder that is characterized by craniofacial anomalies and conotruncal heart defects. Most 22q11.2 deletions occur sporadically, although the deletion may be transmitted in some cases. The present performed a molecular analysis in one family including a patient with clinical diagnosis of VCFS and his sister with a suggestive phenotype. Six polymorphic 22q11.2 markers (i.e. D22S420, D22S264, D22S941, D22S306, D22S425 and D22S257) were used for genotype analysis of the DNA from the patients and unaffected relatives. The results revealed a 22q11.2 deletion in the patient and his sister from one of six markers (i.e. D22S941). Genotype analysis demonstrated that the deletion in this sib was of maternal origin. The results suggest that the mother probably has gonadal mosaicism. The other relatives present normal DNA profiles for all markers. These results have implications for genetic counseling because of a risk of transmission by germ cells carrying the deletion, even when parents present with a normal DNA profile in their blood cells.

Global poly(A) mRNA expression profile measured in individual bovine oocytes and cleavage embryos.

The objective of this article was to estimate quantitative differences for GAPDH transcripts and poly(A) mRNA: (i) between oocytes collected from cumulus-oocyte complexes (COCs) qualified morphologically as grades A and B; (ii) between grade A oocytes before and after in vitro maturation (IVM); and (iii) among in vitro-produced embryos at different developmental stages. To achieve this objective a new approach was developed to estimate differences between poly(A) mRNA when using small samples. The approach consisted of full-length cDNA amplification (acDNA) monitored by real-time PCR, in which the cDNA from half of an oocyte or embryo was used as a template. The GAPDH gene was amplified as a reverse transcription control and samples that were not positive for GAPDH transcripts were discarded. The fold differences between two samples were estimated using delta Ct and statistical analysis and were obtained using the pairwise fixed reallocation randomization test. It was found that the oocytes recovered from grade B COCs had quantitatively less poly(A) mRNA (p<0.01) transcripts compared with grade A COCs (1 arbitrary unit expression rate). In the comparison with immature oocytes (1 arbitrary unit expression rate), the quantity of poly(A) mRNA did not change during IVM, but declined following IVF and varied with embryo culture (p<0.05). Amplification of cDNA by real-time PCR was an efficient method to estimate differences in the amount of poly(A) mRNA between oocytes and embryos. The results obtained from individual oocytes suggested an association between poly(A) mRNA abundance and different morphological qualities of oocytes from COCs. In addition, a poly(A) mRNA profile was characterized from oocytes undergoing IVM, fertilization and blastocyst heating.

Three new cases of spondylocarpotarsal synostosis syndrome: clinical and radiographic studies.

Spondylocarpotarsal synostosis syndrome (SSS) or congenital synspondylism is a recently delineated clinical entity. At least 15 patients have been reported. We present 3 new patients, 2 of whom were sibs born to first-cousin parents. All of our patients had multiple synostoses involving cervical, thoracic and/or lumbar vertebral bodies and carpal/tarsal bones, scoliosis/lordosis, and short stature. Sensorineural deafness was found in 2 of the 3 patients. Analysis of clinical manifestations suggests clinical variability and genetic heterogeneity in SSS. Of a total of 18 SSS patients, 10 were five pairs of sibs from five families, with first-cousin consanguinity of parents in 3, indicating that at least one type of SS is an autosomal-recessive disorder.

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.

Typical phenotypic spectrum of velocardiofacial syndrome occurs independently of deletion size in chromosome 22q11.2

Velocardiofacial syndrome (VCFS) is a relatively common developmental disorder characterized by craniofacial anomalies and conotruncal heart defects. Many VCFS patients present hemizygous deletions on part of chromosome 22q11.2; suggestive that haploinsufficiency in this region is responsible for this etiology. Most 22q11.2 deletions occur sporadically, although in some cases the deletion may be transmitted. A total of 29 VCFS patients and their parents were genotyped using six consecutive polymorphic markers (STS) of the chromosome 22q11.2: D22S420, D22S941, D22S264, D22S306, D22S425, and D22S257. The results revealed that 72% (21/29) of the patients harbored a deletion involving the polymorphic markers D22S420, D22S941, and/or D22S264. Haplotype analysis showed that among the patients studied, the deletions were either of maternal or paternal origin. Our findings demonstrated that independently of their size, any deletion occurring in the VCFS critical region is enough to confer the patient phenotype.

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 rg/mL). 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.

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.