Dicephalus dipus dibrachius twins: report of an autopsy case

Dicephalus dipus dibrachius twins are a rare form of conjoined twins. An autopsy of conjoined dicephalus twins is reported. The diagnosis was performed during the pregnancy and the family received a judicial authorization for termination of pregnancy. The preterm newborn progressed to cardiac arrest and died following a court order to terminate the pregnancy. The conceptus presented two arms, two legs, a trunk, two heads, and a single umbilical cord. The two spinal columns presented vertebral fusion in the sacral region. The heart exhibited complex malformations. The external genitalia were female; and cytogenetic analysis confirmed female sex (46, XX). This analysis also corroborated the etiopathogenic hypotheses described for this abnormality, which proposes failures in embryonic formation rather than specific chromosomal alterations. Current identification of cases by ultrasound permits medical management and multidisciplinary action with the family, enabling the legal termination of pregnancy

Evidence of epigenetic regulation of the tumor suppressor gene cluster flanking RASSF1 in breast cancer cell lines

Epigenetic mechanisms are frequently deregulated in cancer cells and can lead to the silencing of genes with tumor suppressor activities. The isoform A of the Ras-association domain family member 1 (RASSF1A) gene is one of the most frequently silenced transcripts in human tumors; however, few studies have simultaneously investigated epigenetic abnormalities associated with the 3p21.3 tumor suppressor gene cluster flanking RASSF1 (i.e., SEMA3B, HYAL3, HYAL2, HYAL1, TUSC2, RASSF1, ZMYND10, NPRL2, TMEM115 and CACNA2D2). This study aimed to investigate the role of epigenetic changes to these genes in 17 breast cancer cell lines and in three non-tumorigenic epithelial breast cell lines (184A1, 184B5 and MCF 10A) and to evaluate the effect on gene expression of treatment with the demethylating agent 5-Aza -2'-deoxycytidine and/or Trichostatin A (TSA), a histone deacetylase inhibitor. We report that, although the RASSF1A isoform was determined to be epigenetically silenced in 15 of the 17 breast cancer cell lines, all the cell lines expressed the RASSF1C isoform. Five breast cancer cell lines overexpressed RASSF1C when compared with the normal epithelial cell line 184A1. Furthermore, the genes HYAL1 and CACNA2D2 were significantly overexpressed after the treatments. After the combined treatment, RASSF1A re-expression was accompanied by an increase in expression levels of the flanking genes. The Spearman's correlation coefficient indicated a positive co-regulation of the following gene pairs: RASSF1 and TUSC2 (r = 0.64, p = 0.002), RASSF1 and ZMYND10 (r = 0.58, p = 0.07), RASSF1 and NPRL2 (r = 0.48, p = 0.03), ZMYND10 and NPRL2 (r = 0.71; p = 0.0004) and NPRL2 and TMEM115 (r = 0.66, p = 0.001). Interestingly, the genes TUSC2, NPRL2 and TMEM115 were found to be unmethylated in each of the untreated cell lines. Chromatin immunoprecipitation using antibodies against the acetylated and trimethylated lysine 9 of histone H3 demonstrated low levels of histone methylation in these genes, which are located closest to RASSF1. These results provide evidence that epigenetic repression is involved in the downregulation of multiple genes at 3p21.3 in breast cancer cells.Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES

Epstein-barr virus infection and single nucleotide Polymorphisms in the promoter region of interleukin 10 gene in patients with Hodgkin lymphoma

Context. - Hodgkin lymphoma is a neoplastic disease in which the immune system plays a major role in its pathogenesis. Interleukin 10 ( IL-10), an immunosuppressive cytokine actively produced in patients with Hodgkin lymphomas, favors the survival of the Hodgkin/Reed-Sternberg cells. Individual variations in IL-10 levels may be due, in part, to the presence of single nucleotide polymorphisms in the IL10 gene promoter.Objective. - To evaluate whether particular single nucleotide polymorphisms in the IL10 gene are found more frequently in Hodgkin lymphoma cases associated with Epstein-Barr virus infection.Design. - the identification of single nucleotide polymorphisms at positions -1082 and -819/-592 in the IL10 gene was performed by polymerase chain reaction and restriction length fragment polymorphisms analysis in 65 cases of Hodgkin lymphoma and 50 cases of reactive benign follicular lymphoid hyperplasia ( non-Hodgkin lymphoma control group).Results. - the frequency of the genotype GG at position -1082 was found to be significantly higher in patients with Epstein-Barr virus-positive Hodgkin lymphoma compared with Epstein-Barr virus-negative cases.Conclusions. - the results suggest that the presence of specific single nucleotide polymorphisms in the IL10 gene, notably those associated with high IL-10 production, may play a role in the susceptibility to Epstein-Barr virus -positive Hodgkin lymphoma development

DNA methylation patterns of steroid receptor genes ESR1, ESR2 and PGR in deep endometriosis compromising the rectum

Endometriosis is characterized by the presence of endometrial-like tissue located outside the uterine cavity. Recent evidence suggests that endometriosis may be an epigenetic disease, as well as an estrogen-dependent disease. Based on the unique steroid hormone receptor expression profile observed in endometriotic lesions as compared to eutopic endometrium, the present study aimed to gain further insight into the DNA methylation patterns of alternative promoters of the steroid receptor genes ESR1, ESR2 and PGR in intestinal deep endometriosis, one of the most aggressive forms of endometriosis. The DNA methylation patterns were evaluated by methylation-specific polymerase chain reaction (MS-PCR) after bisulfite modification in 44 endometriotic tissues as well as in 7 matched eutopic endometrium. No differences in the DNA methylation were observed for the ESR1 and ESR2 genes. Methylation of the PGR gene was observed in 39% (17 out of 44) and 19% (7 out of 37) of the cases in the promoter regions B (PGRB) and A (PGRA), respectively. Both PGR promoter regions were methylated in 3 cases. PGRB methylated alleles were detected exclusively in the endometriotic lesions when compared to the eutopic endometrium obtained from the same patient. The effect of DNA methylation in inhibiting the PGR gene expression was corroborated by immunostaining for PgR protein in a subset of tissue samples. The present study demonstrated that epigenetic changes occur in both promoter regions of the PGR gene in intestinal endometriosis. Since eutopic and ectopic tissues do not respond sufficiently to progesterone in women with endometriosis, further study is necessary to evaluate the effect of epigenetic alterations in progesterone-resistance in this enigmatic disease.Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES

Genetic polymorphisms associated with steroids metabolism and insulin action in polycystic ovary syndrome

Polycystic ovary syndrome (PCOS) is an endocrinopathy associated with infertility, diabetes and cardiovascular events. This study aimed to correlate polymorphisms of genes involved in the biosynthesis and metabolism of steroids and insulin action (CYP17A1, CYP19A1, AR, ESR1, ESR2, INSR, IGF2 and PAI1) with clinical and biochemical parameters of PCOS. DNA was extracted from peripheral blood samples obtained from 117 PCOS and 105 healthy women. The PAI1 insertion/deletion (-675 ins/delG) polymorphism (rs1799768) was genotyped by PCR-SSCP. CYP19A1 [TTTA](n), AR [CAG](n), ESR1 [TA](n), and ESR2 [CA](n) genes were evaluated by PCR-based GeneScan analysis, while CYP17A1 5'UTR (rs743572), INSR 1058 CT (rs1799817), and IGF2 3'UTR GA (rs680) polymorphisms were evaluated by PCR-RFLP. The results showed a prevalence of PAI1 4G5G+4G4G genotypes in PCOS (p=0.025). Younger PCOS women showed a predominance of CT+TT, GA+AA and 4G5G+4G4G genotypes of INSR, IGF2, and PAI1 (p=0.0499, p=0.0300, p=0.0350, respectively). AR shorter alleles (<= 20 repeats) were significantly associated with higher serum levels of total testosterone (TT, p=0.0086). In conclusion, PAI1 polymorphism seems to be associated with the risk of PCOS development. Younger PCOS women had specific genotypes of INSR, IGF2 and PAI1 genes. AR shorter alleles can be associated with higher serum levels of TT in PCOS patients.Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq

Defining suitable reference genes for RT-qPCR analysis on human sertoli cells after 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) exposure

Quantitative real-time RT-PCR (qPCR) has proven to be a valuable molecular technique to quantify gene expression. There are few studies in the literature that describe suitable reference genes to normalize gene expression data. Studies of transcriptionally disruptive toxins, like tetrachlorodibenzo-p-dioxin (TCDD), require careful consideration of reference genes. The present study was designed to validate potential reference genes in human Sertoli cells after exposure to TCDD. 32 candidate reference genes were analyzed to determine their applicability. geNorm and NormFinder softwares were used to obtain an estimation of the expression stability of the 32 genes and to identify the most suitable genes for qPCR data normalization.Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP

PSA and androgen-related gene (AR, CYP17, and CYP19) polymorphisms and the risk of adenocarcinoma at prostate biopsy

The aim of the present study was to examine the impact of polymorphisms in prostate-specific antigen (PSA) and androgen-related genes (AR, CYP17, and CYP19) on prostate cancer (PCa) risk in selected high-risk patients who underwent prostate biopsy. Blood samples and prostate tissues were obtained for DNA analysis. Single-nucleotide polymorphisms in the 50-untranslated regions (UTRs) of the PSA (substitution A > G at position -158) and CYP17 (substitution T > C at 50-UTR) genes were detected by polymerase chain reaction (PCR)-restriction fragment length polymorphism assays. The CAG and TTTA repeats in the AR and CYP19 genes, respectively, were genotyped by PCR-based GeneScan analysis. Patients with the GG genotype of the PSA gene had a higher risk of PCa than those with the AG or AA genotype (OR = 3.79, p = 0.00138). The AA genotype was associated with lower PSA levels (6.44 +/- 1.64 ng/mL) compared with genotypes having at least one G allele (10.44 +/- 10.06 ng/mL) (p = 0.0687, 95% CI - 0.3146 to 8.315, unpaired t-test). The multivariate analysis confirmed the association between PSA levels and PSA genotypes (AA vs. AG+GG; chi(2) = 0.0482) and CYP19 (short alleles homozygous vs. at least one long allele; chi(2) = 0.0110) genotypes. Genetic instability at the AR locus leading to somatic mosaicism was detected in one PCa patient by comparing the length of AR CAG repeats in matched peripheral blood and prostate biopsy cores. Taken together, these findings suggest that the PSA genotype should be a clinically relevant biomarker to predict the PCa risk

Shorter CAG repeat in the AR gene is associated with atypical hyperplasia and breast carcinoma

Background: Previous reports into the role of [CAG]n repeat lengths in the androgen receptor (AR) gene indicate that these may play an important part in the development and progression of breast cancer, however, knowledge regarding benign breast lesions is limited. Patients and Methods: PCR-based GeneScan analysis was used to investigate the [CAG]n repeat length at exon 1 of the AR gene in 59 benign breast lesions (27 fibroadenomas, 18 atypical hyperplasias, and 14 hyperplasias without atypia) and 54 ductal breast carcinomas. Seventy-two cancer-free women were used as a control group. In addition, [CAG]n repeats were evaluated for the presence of loss of heterozygosity (LOH) and microsatellite instability (MSI) in a subset of these samples (27 fibroadenomas, 14 hyperplasias without atypia and 22 breast carcinomas). Results: Shorter [CAG]n repeat lengths were strongly correlated with atypical hyperplasias (p=0.0209) and carcinomas (p<0.0001). LOH was found in 1/12 and 4/20 informative cases of hyperplasias without atypia and breast carcinomas, respectively. Three patients with breast carcinoma who had previously presented atypical hyperplasia showed a reduction in the [CAG]n repeat length in their carcinomas. Conclusion: Short [CAG]n repeat length (≤20) polymorphisms are strongly associated with breast carcinomas and atypical hyperplasias. Although non-significant, a subgroup of patients with breast carcinoma and genotype SS showed an association with parameters of worse outcome