Identification and in silico analyses of novel TGFBR1 and TGFBR2 mutations in Marfan syndrome-related disorders

Very recently, heterozygous mutations in the genes encoding transforming growth factor beta receptors I (TGFBR1) and II (TGFBR2) have been reported in Loeys-Dietz aortic aneurysm syndrome (LDS). In addition, dominant TGFBR2 mutations have been identified in Marfan syndrome type 2 (MFS2) and familial thoracic aortic aneurysms and dissections (TAAD). In the past, mutations of these genes were associated with atherosclerosis and several human cancers. Here, we report a total of nine novel and one known heterozygous sequence variants in the TGFBR1 and TGFBR2 genes in nine of 70 unrelated individuals with MFS-like phenotypes who previously tested negative for mutations in the gene encoding the extracellular matrix protein fibrillin-1 (FBN1). To assess the pathogenic impact of these sequence variants, in silico analyses were performed by the PolyPhen, SIFT, and Fold-X algorithms and by means of a 3D homology model of the TGFBR2 kinase domain. Our results showed that in all but one of the patients the pathogenic effect of at least one sequence variant is highly probable (c.722C > T, c.799A > C, and c.1460G > A in TGFBR1 and c.773T > G, c.1106G > T, c.1159G > A, c.1181G > A, and c.1561T > C in TGFBR2). These deleterious alleles occurred de novo or segregated with the disease in the families, indicating a causative association between the sequence variants and clinical phenotypes. Since TGFBR2 mutations found in patients with MFS-related disorders cannot be distinguished from heterozygous TGFBR2 mutations reported in tumor samples, we emphasize the importance of segregation analysis in affected families. In order to be able to find the mutation that is indeed responsible for a MFS-related phenotype, we also propose that genetic testing for sequence alterations in TGFBR1 and TGFBR2 should be complemented by mutation screening of the FBN1 gene.     

Sonographic Findings and Clinical Outcomes in Women With Massive Subchorionic Hematoma Detected in the Second Trimester

BackgroundSmall hematomas on the placental surface, termed subamniotic hemorrhage, are a common finding either at the routine 18- to 20-week anatomy ultrasound or at subsequent assessments of fetal growth and well-being. Hemorrhage beneath or at the edge of the placenta, or behind an isolated area of the fetal membranes, is of greater concern.The CasesWe describe the ultrasound findings and clinical outcomes in two women with a diagnosis of massive intrauterine hematoma arising from the fetal membranes. Both required blood transfusion because of the extent of concealed and revealed bleeding. In each case the initial placental appearances and uterine artery Doppler studies were normal. Both hematomas resolved with growth of the fetus and amniotic sac. Each neonate survived the perinatal period favourably. One was born vaginally at 32 weeks’ gestation following premature preterm rupture of the membranes, and the second was born by emergency Caesarean section at 37 weeks because of a recurrence of antepartum hemorrhage.ConclusionsLarge intrauterine hematomas may be acutely detrimental to maternal health in the second trimester. Ultrasound assessment of the placenta is useful to define the perinatal prognosis and may demonstrate gradual resolution. Despite a dramatic initial presentation, this finding may be compatible with a favourable outcome.     

THE EFFECT OF CULTURAL FACTORS ON DAILY COPING AND INVOLUNTARY RESPONSES TO STRESS AMONG LOW‐INCOME LATINO ADOLESCENTS

This study used daily diary methodology to examine associations between cultural factors and daily coping and responses to stress among predominantly low‐income Latino adolescents. A total of 58 middle school students (53% male, mean age = 13.31, 95% Latino) completed baseline measures assessing demographic characteristics, familism, ethnic identity, and family ethnic socialization. They subsequently completed 7 consecutive daily diaries assessing daily stress, coping, and involuntary stress responses. Results yielded main effects of stress, gender, familism, and ethnic identity on adolescents’ coping and involuntary stress responses. In addition, interactions between stress and familism, ethnic identity, and family ethnic socialization emerged. Results suggest that familism may promote adaptive responses to stress, while adolescents who report more family ethnic socialization may rely more on maladaptive responses at high levels of stress. Findings related to ethnic identity were mixed and varied depending on levels of ethnic identity exploration versus commitment.     

High frequency of precancerous lesions of gastric cancer associated with Helicobacter pylori and response to treatment,in Chiapas,Mexico

Stomach cancer is the second cause of death in Mexico in patients with malignant tumors. This disease represents a public health problem. A strong association has been described between chronic infection with Helicobacter pylori and gastric cancer. This malignancy is preceded by a series of preneoplastic conditions, including chronic atrophic gastritis (CAG), intestinal metaplasia (IM), and dysplasia. The objective of this study was to establish the prevalence of preneoplastic conditions associated with infection of Helicobacter pylori in the state of Chiapas and its eradication with antibiotics. Persons infected with Helicobacter pylori and with CAG were identified by serology against CagA protein and serologic levels of gastrin. An endoscopy with biopsy was performed at the beginning of the study, and at 6 weeks and 1 year thereafter. A total of 281 people were enrolled and randomly assigned to treatment or placebo group. CAG was found in 59%, IM in 51%, and dysplasia in 13%. In intent-to-treat and per-protocol analysis, Helicobacter pylori was eliminated in 70 and 76%, respectively. These results indicate high frequency of preneoplastic conditions associated with Helicobacter pylori and an excellent eradication rate. They also offer a possible alternative for preventing gastric cancer.     

Evaluation and application of denaturing HPLC for mutation detection in Marfan syndrome: Identification of 20 novel mutations and two novel polymorphisms in the FBN1 gene

Mutations in the human fibrillin 1 gene (FBN1) cause the Marfan syndrome (MFS), an autosomal dominant connective tissue disorder. Knowledge about FBN1 mutations is important for early diagnosis, management, and genetic counseling. However, mutation detection in FBN1 is a challenge because the gene is very large in size ( approximately 200 kb) and the approximately 350 mutations detected so far are scattered over 65 exons. Conventional methods for large-scale detection of mutations are expensive, technically demanding, or time consuming. Recently, a high-capacity low-cost mutation detection method was introduced based on denaturing high-performance liquid chromatography (DHPLC). To assess the sensitivity and specificity of this method, we blindly screened 64 DNA samples of known FBN1 genotype exon-by-exon using exon-specific DHPLC conditions. Analysis of 682 PCR amplicons correctly identified 62 out of 64 known sequence variants. In three MFS patients of unknown FBN1 genotype, we detected two mutations and eight polymorphisms. Overall, 20 mutations and two polymorphisms are described here for the first time. Our results demonstrate 1) that DHPLC is a highly sensitive (89-99%, P = 0.05) method for FBN1 mutation detection; but 2) that chromatograms with moderate and weak pattern abnormalities also show false positive signals (in all 45-59%, P = 0.05); 3) that the difference in the chromatograms of heterozygous and homozygous amplicons is mostly independent of the type of sequence change; and 4) that DHPLC column conditions, additional base changes, and the amounts of injected PCR products influence significantly the shape of chromatograms. A strategy for FBN1 mutation screening is discussed.     

Towards defining roles and relationships for tenascin-C and TGFbeta-1 in the normal and neoplastic urinary bladder

Tenascin-C (TN-C) is an extracellular matrix glycoprotein expressed along epithelial/stromal boundaries during tissue remodelling events, such as those that occur during morphogenesis, wound healing, and tumour invasion. Using clinical specimens and a range of in vitro models that simulate homeostasis, wound healing, and malignant progression, this study sought to establish the patterns of TN-C expression in normal and neoplastic bladder and to determine the role of exogenous transforming growth factor beta-1 (TGFbeta-1), interleukin-4 (IL-4), basic fibroblast growth factor (bFGF), tumour necrosis factor alpha (TNFalpha), and interferon gamma (IFNgamma) in the induction of TN-C expression by bladder uro-epithelial cells. The findings indicate that normal urothelial cells may express TN-C, with both TGFbeta-1 and IL-4 able to induce expression. TN-C was not expressed in neoplastic urothelium, although both TN-C and TGFbeta-1 may be involved in tissue remodelling during papillary tumour formation and invasion. Furthermore, the urothelium of high-grade papillary tumours and carcinoma in situ specimens exhibited little TGFbeta-1 immunoreactivity, compared with the urothelium of low-grade tumours and normal specimens, suggesting an association between TGFbeta-1 expression and urothelial differentiation. A tumour invasion model, in which established bladder cancer cell lines were seeded onto a normal bladder stroma, corroborated the evidence from the clinical specimens and demonstrated that TN-C was strongly expressed around foci of stromal invasion. Thus, TN-C immunoreactivity may provide an additional tool in the assessment of early stromal invasion in bladder cancer.