The Werner Syndrome Protein Suppresses Telomeric Instability Caused by Chromium (VI) Induced DNA Replication Stress

Telomeres protect the chromosome ends and consist of guanine-rich repeats coated by specialized proteins. Critically short telomeres are associated with disease, aging and cancer. Defects in telomere replication can lead to telomere loss, which can be prevented by telomerase-mediated telomere elongation or activities of the Werner syndrome helicase/exonuclease protein (WRN). Both telomerase and WRN attenuate cytotoxicity induced by the environmental carcinogen hexavalent chromium (Cr(VI)), which promotes replication stress and DNA polymerase arrest. However, it is not known whether Cr(VI)-induced replication stress impacts telomere integrity. Here we report that Cr(VI) exposure of human fibroblasts induced telomeric damage as indicated by phosphorylated H2AX (γH2AX) at telomeric foci. The induced γH2AX foci occurred in S-phase cells, which is indicative of replication fork stalling or collapse. Telomere fluorescence in situ hybridization (FISH) of metaphase chromosomes revealed that Cr(VI) exposure induced an increase in telomere loss and sister chromatid fusions that were rescued by telomerase activity. Human cells depleted for WRN protein exhibited a delayed reduction in telomeric and non-telomeric damage, indicated by γH2AX foci, during recovery from Cr(VI) exposure, consistent with WRN roles in repairing damaged replication forks. Telomere FISH of chromosome spreads revealed that WRN protects against Cr(VI)-induced telomere loss and downstream chromosome fusions, but does not prevent chromosome fusions that retain telomere sequence at the fusion point. Our studies indicate that environmentally induced replication stress leads to telomere loss and aberrations that are suppressed by telomerase-mediated telomere elongation or WRN functions in replication fork restoration

Impact of PGL-I Seropositivity on the Protective Effect of BCG Vaccination among Leprosy Contacts: A Cohort Study

Although leprosy has become a neglected disease, it is an important cause of disability, and 250,000 new cases are still diagnosed worldwide every year. The current study was carried out in Brazil, where almost 40,000 new cases of leprosy are diagnosed every year. The study targeted contacts of leprosy patients, who are at the highest risk of contracting the disease. We studied 2,135 contacts who were diagnosed at the Leprosy Outpatient Clinic at the Oswaldo Cruz Foundation in Rio de Janeiro, RJ, Brazil, between 1987 and 2007. The presence of antibodies against a specific Mycobacterium leprae antigen (PGL-I) at the first examination and BCG vaccination status were evaluated. PGL-I-positive contacts had a higher risk of developing leprosy than PGL-I-negative contacts. Among the former, vaccinated contacts were at higher risk than unvaccinated contacts. Our results indicate that contact examination combined with PGL-I testing and BCG vaccination appears to justify the targeting of PGL-I-positive individuals for enhanced surveillance. Furthermore, it is highly recommended that PGL-I-positive contacts and contacts with a high familial bacterial index (i.e., the sum of results from index and co-prevalent cases), regardless of serological response, should be monitored. This group could be considered as a target for chemoprophylaxis

Diminished Telomeric 3′ Overhangs Are Associated with Telomere Dysfunction in Hoyeraal-Hreidarsson Syndrome

BACKGROUND:Eukaryotic chromosomes end with telomeres, which in most organisms are composed of tandem DNA repeats associated with telomeric proteins. These DNA repeats are synthesized by the enzyme telomerase, whose activity in most human tissues is tightly regulated, leading to gradual telomere shortening with cell divisions. Shortening beyond a critical length causes telomere uncapping, manifested by the activation of a DNA damage response (DDR) and consequently cell cycle arrest. Thus, telomere length limits the number of cell divisions and provides a tumor-suppressing mechanism. However, not only telomere shortening, but also damaged telomere structure, can cause telomere uncapping. Dyskeratosis Congenita (DC) and its severe form Hoyeraal-Hreidarsson Syndrome (HHS) are genetic disorders mainly characterized by telomerase deficiency, accelerated telomere shortening, impaired cell proliferation, bone marrow failure, and immunodeficiency. METHODOLOGY/PRINCIPAL FINDINGS:We studied the telomere phenotypes in a family affected with HHS, in which the genes implicated in other cases of DC and HHS have been excluded, and telomerase expression and activity appears to be normal. Telomeres in blood leukocytes derived from the patients were severely short, but in primary fibroblasts they were normal in length. Nevertheless, a significant fraction of telomeres in these fibroblasts activated DDR, an indication of their uncapped state. In addition, the telomeric 3' overhangs are diminished in blood cells and fibroblasts derived from the patients, consistent with a defect in telomere structure common to both cell types. CONCLUSIONS/SIGNIFICANCE:Altogether, these results suggest that the primary defect in these patients lies in the telomere structure, rather than length. We postulate that this defect hinders the access of telomerase to telomeres, thus causing accelerated telomere shortening in blood cells that rely on telomerase to replenish their telomeres. In addition, it activates the DDR and impairs cell proliferation, even in cells with normal telomere length such as fibroblasts. This work demonstrates a telomere length-independent pathway that contributes to a telomere dysfunction disease

Speech therapy in food transition from probe to breast in newborn in kangaroo method

Objetivo: Verificar a relação entre idade gestacional e tempo de intervenção fonoaudiológica para início da alimentação via oral, quando utilizada a técnica de transição alimentar da sonda direta para o peito. Métodos: Trata-se de um estudo do prontuário médico/fonoaudiológico de 38 recém-nascidos de risco em Unidade Canguru. Foram coletados os seguintes dados: idade gestacional ao nascimento e corrigida, dias de vida, peso ao nascimento e atual, tipo e duração da intervenção fonoaudiológica, volume de dieta por sonda. Utilizou-se o tempo de uso de antibióticos e o suporte ventilatório como critérios de divisão dos recém-nascidos em dois grupos (G1 e G2). Na análise estatística, aplicou-se o teste não paramétrico de Mann-Whitney e o coeficiente de correlação de Pearson. Resultados: O tempo de intervenção para os recém-nascidos que receberem alta fonoaudiológica não apresentou resultados significativos entre os grupos (G1= 9,35 dias e G2= 10,12 dias), embora a hipótese inicial deste estudo fosse a de que os recém-nascidos do G1 necessitariam de menor período de atendimento fonoaudiológico que os do G2. Houve diferença estatisticamente significativa para o peso ao nascimento, entre G1 (1563,53 g) e G2 (1409,62 g). Conclusão: Quando utilizada a técnica de transição alimentar da sonda direta para o peito, em recém-nascidos de risco com média de idade gestacional semelhante e mesmo tempo de intervenção fonoaudiológica, os bebês demonstraram aptidão para coordenar os movimentos de sucção/respiração/deglutição, e consequentemente, a amamentação efetiva em seio materno exclusivo. _________________________________________________________________________________________ ABSTRACT: Purpose: Verify the relationship between gestational age and duration of speech therapy to start oral feeding, when used the technique of feeding transition of the probe directly to the chest. Methods: This is a study of newborn medical/speech records of 38 risk in Kangaroo unit. Were collected: gestational age at birth and corrected days of life, birth weight and current, type and duration of speech therapy, enteral feeding volume. We used time use of antibiotics and ventilatory support as criteria for division of newborns into two groups (G1 and G2). At the statistical analysis was applied the nonparametric test of Mann-Whitney and the Pearson’s correlation coefficient. Results: Intervention time for newborns were discharged speech showed no significant results between groups (G1 = 9.35 days and G2 = 10.12 days), although the initial hypothesis of this study was that the newborn G1 would require fewer days of speech therapy than the G2. There was statistically significant difference in birth weight between G1 (1563.53 g) and G2 (1409.62 g). Conclusion: It was observed that both groups started speech therapy and oral feeding practically medium of similar gestational ages and both speech intervention, demonstrating ability to coordinate sucking movements/breathing/swallowing, and consequently the effective and exclusive breastfeeding

A Non-Canonical Function of Zebrafish Telomerase Reverse Transcriptase Is Required for Developmental Hematopoiesis

Although it is clear that telomerase expression is crucial for the maintenance of telomere homeostasis, there is increasing evidence that the TERT protein can have physiological roles that are independent of this central function. To further examine the role of telomerase during vertebrate development, the zebrafish telomerase reverse transcriptase (zTERT) was functionally characterized. Upon zTERT knockdown, zebrafish embryos show reduced telomerase activity and are viable, but develop pancytopenia resulting from aberrant hematopoiesis. The blood cell counts in TERT-depleted zebrafish embryos are markedly decreased and hematopoietic cell differentiation is impaired, whereas other somatic lineages remain morphologically unaffected. Although both primitive and definitive hematopoiesis is disrupted by zTERT knockdown, the telomere lengths are not significantly altered throughout early development. Induced p53 deficiency, as well as overexpression of the anti-apoptotic proteins Bcl-2 and E1B-19K, significantly relieves the decreased blood cells numbers caused by zTERT knockdown, but not the impaired blood cell differentiation. Surprisingly, only the reverse transcriptase motifs of zTERT are crucial, but the telomerase RNA-binding domain of zTERT is not required, for rescuing complete hematopoiesis. This is therefore the first demonstration of a non-canonical catalytic activity of TERT, which is different from “authentic” telomerase activity, is required for during vertebrate hematopoiesis. On the other hand, zTERT deficiency induced a defect in hematopoiesis through a potent and specific effect on the gene expression of key regulators in the absence of telomere dysfunction. These results suggest that TERT non-canonically functions in hematopoietic cell differentiation and survival in vertebrates, independently of its role in telomere homeostasis. The data also provide insights into a non-canonical pathway by which TERT functions to modulate specification of hematopoietic stem/progenitor cells during vertebrate development. (276 words

Prevalence of Age-Related Macular Degeneration in Europe: The Past and the Future

Purpose Age-related macular degeneration (AMD) is a frequent, complex disorder in elderly of European ancestry. Risk profiles and treatment options have changed considerably over the years, which may have affected disease prevalence and outcome. We determined the prevalence of early and late AMD in Europe from 1990 to 2013 using the European Eye Epidemiology (E3) consortium, and made projections for the future. Design Meta-analysis of prevalence data. Participants A total of 42 080 individuals 40 years of age and older participating in 14 population-based cohorts from 10 countries in Europe. Methods AMD was diagnosed based on fundus photographs using the Rotterdam Classification. Prevalence of early and late AMD was calculated using random-effects meta-analysis stratified for age, birth cohort, gender, geographic region, and time period of the study. Best-corrected visual acuity (BCVA) was compared between late AMD subtypes; geographic atrophy (GA) and choroidal neovascularization (CNV). Main Outcome Measures Prevalence of early and late AMD, BCVA, and number of AMD cases. Results Prevalence of early AMD increased from 3.5% (95% confidence interval [CI] 2.1%–5.0%) in those aged 55–59 years to 17.6% (95% CI 13.6%–21.5%) in those aged ≥85 years; for late AMD these figures were 0.1% (95% CI 0.04%–0.3%) and 9.8% (95% CI 6.3%–13.3%), respectively. We observed a decreasing prevalence of late AMD after 2006, which became most prominent after age 70. Prevalences were similar for gender across all age groups except for late AMD in the oldest age category, and a trend was found showing a higher prevalence of CNV in Northern Europe. After 2006, fewer eyes and fewer ≥80-year-old subjects with CNV were visually impaired (P = 0.016). Projections of AMD showed an almost doubling of affected persons despite a decreasing prevalence. By 2040, the number of individuals in Europe with early AMD will range between 14.9 and 21.5 million, and for late AMD between 3.9 and 4.8 million. Conclusion We observed a decreasing prevalence of AMD and an improvement in visual acuity in CNV occuring over the past 2 decades in Europe. Healthier lifestyles and implementation of anti–vascular endothelial growth factor treatment are the most likely explanations. Nevertheless, the numbers of affected subjects will increase considerably in the next 2 decades. AMD continues to remain a significant public health problem among Europeans

Nucleo-cytoplasmic transport of proteins and RNA in plants

Merkle T. Nucleo-cytoplasmic transport of proteins and RNA in plants. Plant Cell Reports. 2011;30(2):153-176.Transport of macromolecules between the nucleus and the cytoplasm is an essential necessity in eukaryotic cells, since the nuclear envelope separates transcription from translation. In the past few years, an increasing number of components of the plant nuclear transport machinery have been characterised. This progress, although far from being completed, confirmed that the general characteristics of nuclear transport are conserved between plants and other organisms. However, plant-specific components were also identified. Interestingly, several mutants in genes encoding components of the plant nuclear transport machinery were investigated, revealing differential sensitivity of plant-specific pathways to impaired nuclear transport. These findings attracted attention towards plant-specific cargoes that are transported over the nuclear envelope, unravelling connections between nuclear transport and components of signalling and developmental pathways. The current state of research in plants is summarised in comparison to yeast and vertebrate systems, and special emphasis is given to plant nuclear transport mutants

Diagnosis and management of gastroesophageal reflux disease

Gastroesophageal reflux disease (GERD) is probably one of the most prevalent diseases in the world that also compromises the quality of life of the affected significantly. Its incidence in Brazil is 12%, corresponding to 20 million individuals. OBJECTIVE: To update the GERD management and the new trends on diagnosis and treatment, reviewing the international and Brazilian experience on it. METHOD: The literature review was based on papers published on Medline/Pubmed, SciELO, Lilacs, Embase and Cochrane crossing the following headings: gastroesophageal reflux disease, diagnosis, clinical treatment, surgery, fundoplication. RESULTS: Various factors are involved on GERD physiopathology, the most important being the transient lower esophageal sphincter relaxation. Clinical manifestations are heartburn, regurgitation (typical symptoms), cough, chest pain, asthma, hoarseness and throat clearing (atypical symptoms), which may be followed or not by typical symptoms. GERD patients may present complications such as peptic stenosis, hemorrhage, and Barrett's esophagus, which is the most important predisposing factor to adenocarcinoma. The GERD diagnosis must be based on the anamnesis and the symptoms must be evaluated in terms of duration, intensity, frequency, triggering and relief factors, pattern of evolution and impact on the patient's quality of life. The diagnosis requires confirmation with different exams. The goal of the clinical treatment is to relieve the symptoms and surgical treatment is indicated for patients who require continued drug use, with intolerance to prolonged clinical treatment and with GERD complications. CONCLUSION: GERD is a major digestive health problem and affect 12% of Brazilian people. The anamnesis is fundamental for the diagnosis of GERD, with special analysis of the typical and atypical symptoms (duration, intensity, frequency, triggering and relief factors, evolution and impact on the life quality). High digestive endoscopy and esophageal pHmetry are the most sensitive diagnosctic methods. The clinical treatment is useful in controlling the symptoms; however, the great problem is keeping the patients asymptomatic over time. Surgical treatment is indicated for patients who required continued drug use, intolerant to the drugs and with complicated forms of GERD.A doença do refluxo gastroesofágico (DRGE) é, provavelmente, uma das doenças mais prevalentes no mundo que compromete significativamente a qualidade de vida. Sua incidência no Brasil é de 12%, o que corresponde a 20 milhões de indivíduos. OBJETIVO: Atualizar o manuseio da DRGE e as novas tendências no diagnóstico e tratamento, revendo as experiências internacional e brasileira sobre o tema. MÉTODO: Foi realizada revisão da literatura baseada em artigos publicados no Medline/Pubmed, SciELO, Lilacs, Embase e Cochrane cruzando os seguintes descritores: doença do refluxo gastroesofágico, diagnóstico, tratamento clínico, cirurgia, fundoplicatura. RESULTADOS: Vários fatores estão envolvidos na fisiopatologia da DRGE, sendo o mais importante o relaxamento transitório do esfíncter inferior do esôfago. As manifestações clínicas são azia, regurgitação (sintomas típicos), tosse, dor torácica, asma, rouquidão e pigarro (sintomas atípicos), que podem ser seguidos ou não de sintomas típicos. Pacientes com DRGE podem apresentar complicações como estenose péptica, hemorragia e esôfago de Barrett, que é o fator predisponente mais importante para adenocarcinoma. O diagnóstico deve ser baseado na anamnese e os sintomas devem ser avaliados em termos de duração, intensidade, frequência, fatores precipitantes e relevância, padrão de evolução e impacto na qualidade de vida do paciente. O diagnóstico exige confirmação com exames diferentes. O objetivo do tratamento clínico é aliviar os sintomas e o tratamento cirúrgico é indicado para os que necessitam de uso contínuo de drogas, com intolerância ao tratamento clínico prolongado e com complicações. CONCLUSÃO: A anamnese é fundamental para o diagnóstico de DRGE, com análise especial dos sintomas típicos e atípicos. Endoscopia digestiva alta e pHmetria esofágica são os métodos diagnósticos mais sensíveis. O tratamento clínico é útil no controle dos sintomas; no entanto, o grande problema é manter os pacientes assintomáticos ao longo do tempo. O tratamento cirúrgico é indicado para pacientes que necessitaram o uso contínuo de drogas, intolerantes às drogas e com formas complicadas da DRGE.Universidade Estadual Paulista Júlio de Mesquita Filho, Departamento de Cirurgia e Ortopedia, Faculdade de Medicina de Botucatu, Botucatu, Anexo Verde, Rubião Junior s/n, CEP 18605-970, SP, BrasilUniversidade Estadual Paulista Júlio de Mesquita Filho, Departamento de Cirurgia e Ortopedia, Faculdade de Medicina de Botucatu, Botucatu, Anexo Verde, Rubião Junior s/n, CEP 18605-970, SP, Brasi

Metabolomics and Age-Related Macular Degeneration

Age-related macular degeneration (AMD) leads to irreversible visual loss, therefore, early intervention is desirable, but due to its multifactorial nature, diagnosis of early disease might be challenging. Identification of early markers for disease development and progression is key for disease diagnosis. Suitable biomarkers can potentially provide opportunities for clinical intervention at a stage of the disease when irreversible changes are yet to take place. One of the most metabolically active tissues in the human body is the retina, making the use of hypothesis-free techniques, like metabolomics, to measure molecular changes in AMD appealing. Indeed, there is increasing evidence that metabolic dysfunction has an important role in the development and progression of AMD. Therefore, metabolomics appears to be an appropriate platform to investigate disease-associated biomarkers. In this review, we explored what is known about metabolic changes in the retina, in conjunction with the emerging literature in AMD metabolomics research. Methods for metabolic biomarker identification in the eye have also been discussed, including the use of tears, vitreous, and aqueous humor, as well as imaging methods, like fluorescence lifetime imaging, that could be translated into a clinical diagnostic tool with molecular level resolution