Thermodynamic behavior of short oligonucleotides in microarray hybridizations can be described using Gibbs free energy in a nearest-neighbor model

While designing oligonucleotide-based microarrays, cross-hybridization between surface-bound oligos and non-intended labeled targets is probably the most difficult parameter to predict. Although literature describes rules-of-thumb concerning oligo length, overall similarity, and continuous stretches, the final behavior is difficult to predict. The aim of this study was to investigate the effect of well-defined mismatches on hybridization specificity using CodeLink Activated Slides, and to study quantitatively the relation between hybridization intensity and Gibbs free energy (Delta G), taking the mismatches into account. Our data clearly showed a correlation between the hybridization intensity and Delta G of the oligos over three orders of magnitude for the hybridization intensity, which could be described by the Langmuir model. As Delta G was calculated according to the nearest-neighbor model, using values related to DNA hybridizations in solution, this study clearly shows that target-probe hybridizations on microarrays with a three-dimensional coating are in quantitative agreement with the corresponding reaction in solution. These results can be interesting for some practical applications. The correlation between intensity and Delta G can be used in quality control of microarray hybridizations by designing probes and corresponding RNA spikes with a range of Delta G values. Furthermore, this correlation might be of use to fine-tune oligonucleotide design algorithms in a way to improve the prediction of the influence of mismatching targets on microarray hybridizations.Comment: 32 pages on a single pdf fil

Earning and related variables of the otorhinolaryngologist registered by SBORL (ORL profile 2002)

After the 2002 Brazilian Otorhinolaryngologists' Cense conducted by the Brazilian Society of Otorhinolaryngology, the otorhinolaryngologists' profile was performed with several data that will be analyzed separately. AIM: To analyze the Otorhinolaryngologists' earning and related variables. STUDY DESIGN: Epidemiologist. MATERIAL AND METHOD: 1.636 answered questionnaires were analyzed from a total of 5.830 posted to Brazilian otorhinolaryngologists. EpiInfo was used as database and to perform statistical tests. The chi-square test was used to compare proportions, adjustment by stratification, and Maentel-Haenszel procedure, multivariate logistic regression RESULTS: Proportion of answered questionnaires and total of otorhinolaryngologysts grouped by geographical regions are similar (p=0,040). From 13 variables analyzed 12 were related to earning in univariate analysis. Multivariate analysis evidence five independent variables related to earning. CONCLUSION: The otorhinolaryngologyst has a higher chance of earning 10 thousand reais if is male, a member of university service, has surgery ability, owns some equipment and has some years of graduation.Após a realização do Censo 2002 do Otorrinolaringologista (ORL) Brasileiro pela Sociedade Brasileira de Otorrinolaringologia (SBORL), realizou-se o perfil do ORL obtendo-se uma série de dados que serão analisados separadamente. OBJETIVO: Estimar o rendimento do Otorrinolaringologista Brasileiro e verificar as variáveis associadas. FORMA DE ESTUDO: Epidemiológico. MATÉRIAL E MÉTODO: Foram analisados 1.636 questionários respondidos de um total de 5.830 enviados para os ORL. Foi utilizado o programa EpiInfo para armazenar e analisar os dados. Testes estatísticos usados: qui-quadrado para comparar proporções, ajustamento por estratificação e procedimento de Mantel-Haenszel, regressão logística multivariada. RESULTADOS: A proporção dos questionários e o número total de ORL agrupados por região geográfica são semelhantes (p=0,040). Inicialmente foram avaliadas 13 variáveis sendo 12 associadas com o rendimento. Análise multivariada mostrou que cinco variáveis estão associadas de maneira independente. CONCLUSÕES: O ORL que for do sexo masculino, tiver vínculo com serviço universitário, tiver boa habilidade cirúrgica em mais de uma área, possuir aparelhos e tiver mais tempo de formado tem mais chance de ter rendimento mensal acima de 10 mil reais.Universidade Federal de Minas Gerais Faculdade de Medicina Departamento de Oftalmo, Otorrino e FonoaudiologiaUniversidade Federal de São Paulo (UNIFESP), Escola Paulista de Medicina (EPM) Departamento de ORL PediatriaUniversidade Federal de Minas Gerais Hospital das ClínicasUNIFESP, EPM, Depto. de ORL PediatriaSciEL

Comparative Study of the Standard Fluorescent Antibody to Membrane Antigen (FAMA) Assay and a Flow Cytometry-Adapted FAMA Assay To Assess Immunity to Varicella-Zoster Virus

A flow cytometry-adapted fluorescent antibody to membrane antigen (FAMA) assay to detect IgG antibodies against varicella-zoster virus (VZV) was developed and tested in 62 serum samples, showing 90.32% accuracy obtained from a receiver operating characteristic (ROC) curve with a 0.9125 (95% confidence interval [CI], 0.829 to 1.00) area below the curve compared to the result with standard FAMA.Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)Fed Univ São Paulo UNIFESP EPM, Div Pediat Infect Dis, BR-04040000 São Paulo, BrazilColumbia Univ, Dept Pediat, Coll Phys & Surg, New York, NY 10027 USAFed Univ São Paulo UNIFESP EPM, Div Pediat Infect Dis, BR-04040000 São Paulo, BrazilCAPES: 0108-08-1Web of Scienc

Epithelium and stroma from nasal polyp mucosa exhibits inverse expression of TGF- beta(1) as compared with healthy nasal mucosa

Objective: To evaluate TGF-beta(1) expression in polypoid mucosa (epithelium and stroma) of patients with chronic rhinosinusitis with nasal polyposis (CRSwNP).Methods: Cross-sectional study with two groups: 17 patients with nasal polyposis and 11 controls. Polyps and normal nasal mucosa were processed by immunohistochemical methods for TGF-beta 1 visualization. Then, the percentage of TGF-beta 1 expression in stroma and epithelium was objectively quantified using UT Morph software.Results: A lower percentage of positive expression was found in the epithelium of CRSwNP patients (32.44%) versus normal controls (55.91%) (p < 0.05), and a higher percentage of positive expression in the stroma of CRSwNP patients (23.24%) versus controls (5.88%) (p < 0.05).Conclusion: the lower percentage of TGF-beta(1) expression in the nasal epithelium of CRSwNP patients may have an impact on epithelium-directed topical treatments employed in this patient population.Universidade Federal de São Paulo, Dept Otolaryngol Head & Neck Surg, São Paulo, BrazilUniv Ghent, Ghent Univ Hosp, Dept Otorhinolaryngol, Upper Airway Res Lab, B-9000 Ghent, BelgiumUniversidade Federal de São Paulo, Dept Pathol, São Paulo, BrazilUniversidade Federal de São Paulo, Dept Otolaryngol Head & Neck Surg, São Paulo, BrazilUniversidade Federal de São Paulo, Dept Pathol, São Paulo, BrazilWeb of Scienc

Efficacy of celecoxib in treating symptoms of viral pharyngitis: A double-blind, randomized study of celecoxib versus diclofenac

This study compared the efficacy and safety of the cyclooxygenase-2 specific inhibitor celecoxib with the conventional non-steroidal anti-inflammatory drug diclofenac in the symptomatic treatment of viral pharyngitis. Adult patients from 27 study centers in Latin America were treated with oral doses of celecoxib 200 mg once daily or 200 mg twice daily, or diclofenac 75 mg twice daily for 5 days in a double-blind, randomized study. The primary efficacy assessment was 'Throat Pain on Swallowing' on day 3. In addition, secondary quality-of-life assessments were performed on days 3 and 5. All adverse events and treatment-emergent signs and symptoms were recorded. Data from 313 patients were evaluable for efficacy (105 celecoxib 200 mg once daily, 107 celecoxib 200 mg twice daily, 101 diclofenac 75 mg twice daily). The upper 95% confidence limits for the visual analog scale of 'Throat Pain on Swallowing' on day 3 for celecoxib 200 mg once daily relative to diclofenac 75 mg twice daily, and celecoxib 200 mg twice daily relative to diclofenac 75 mg twice daily were 9.26 and 7.83, respectively. All secondary efficacy and quality-of-life measures were clinically similar for the three treatment groups, and no statistically significant differences were detected. The incidences of treatment-emergent adverse events and withdrawals due to adverse events were similar for all groups, but numerically higher among patients taking diclofenac than celecoxib. More patients in the diclofenac group reported gastrointestinal complaints (7.3%) compared with those in the celecoxib groups (4.3% in the celecoxib 200 mg once-daily group and 3.4% in the celecoxib 200 mg twice-daily group). In conclusion, 5 days of treatment with celecoxib 200 mg once daily is as effective as diclofenac 75 mg twice daily in the symptomatic treatment of viral pharyngitis. Celecoxib 200 mg once daily is also as effective as celecoxib 200 mg twice daily in this condition

Mycobacterium leprae is identified in the oral mucosa from paucibacillary and multibacillary leprosy patients

In leprosy, the nasal mucosa is considered as the principal route of transmission for the bacillus Mycobacterium leprae. the objective of this study was to identify M.leprae in the oral mucosa of 50 untreated leprosy patients, including 21 paucibacillary (PB) and 29 multibacillary (MB) patients, using immunohistochemistry (IHC), with antibodies against bacillus Calmette-Guerin (BCG) and phenolic glycolipid antigen-1 (PGL-1), and polymerase chain reaction (PCR), with MntH-specific primers for M.leprae, and to compare the results. the material was represented by 163 paraffin blocks containing biopsy samples obtained from clinically normal sites (including the tongue, buccal mucosa and soft palate) and visible lesions anywhere in the oral mucosa. All patients and 158 available samples were included for IHC study. Among the 161 available samples for PCR, 110 had viable DNA. There was viable DNA in at least one area of the oral mucosa for 47 patients. M.leprae was detected in 70% and 78% of patients using IHC and PCR, respectively, and in 94% of the patients by at least one of the two diagnostic methods. There were no differences in detection of M.leprae between MB and PB patients. Similar results were obtained using anti-BCG and anti-PGL-1 antibodies, and immunoreactivity occurred predominantly on free-living bacteria on the epithelial surface, with a predilection for the tongue. Conversely, there was no area of predilection according to the PCR results. M.leprae is present in the oral mucosa at a high frequency, implicating this site as a potential means of leprosy transmission.Univ Oeste Paulista, Presidente Prudente Reg Hosp, Dermatol Unit, Presidente Prudente, BrazilUniversidade Federal de São Paulo, Dept Dermatol, São Paulo, BrazilUniv São Paulo, Multiuser Mol Biol Lab, Div Dermatol, Dept Clin Med,Ribeirao Preto Coll Med, BR-14049 Ribeirao Preto, BrazilAdolfo Lutz Inst, Dept Pathol, São Paulo, BrazilUniversidade Federal de São Paulo, Dept Dermatol, São Paulo, BrazilWeb of Scienc

Nonequilibrium effects in DNA microarrays: a multiplatform study

It has recently been shown that in some DNA microarrays the time needed to reach thermal equilibrium may largely exceed the typical experimental time, which is about 15h in standard protocols (Hooyberghs et al. Phys. Rev. E 81, 012901 (2010)). In this paper we discuss how this breakdown of thermodynamic equilibrium could be detected in microarray experiments without resorting to real time hybridization data, which are difficult to implement in standard experimental conditions. The method is based on the analysis of the distribution of fluorescence intensities I from different spots for probes carrying base mismatches. In thermal equilibrium and at sufficiently low concentrations, log I is expected to be linearly related to the hybridization free energy $\Delta G$ with a slope equal to $1/RT_{exp}$, where $T_{exp}$ is the experimental temperature and R is the gas constant. The breakdown of equilibrium results in the deviation from this law. A model for hybridization kinetics explaining the observed experimental behavior is discussed, the so-called 3-state model. It predicts that deviations from equilibrium yield a proportionality of $\log I$ to $\Delta G/RT_{eff}$. Here, $T_{eff}$ is an effective temperature, higher than the experimental one. This behavior is indeed observed in some experiments on Agilent arrays. We analyze experimental data from two other microarray platforms and discuss, on the basis of the results, the attainment of equilibrium in these cases. Interestingly, the same 3-state model predicts a (dynamical) saturation of the signal at values below the expected one at equilibrium.Comment: 27 pages, 9 figures, 1 tabl

Squamous cell carcinoma of the lip: assessment of prognostic factors

Among lip cancers, 90% to 95% of them affect the lower lip and squamous cell carcinoma is the most frequent type. The TNM classification synthesizes the clinical characteristics of the tumor that allows prognosis and makes possible the comparisons of the results. Three parameters have to be considered: size of the tumor (T), propagation to regional lymphatic ganglion (N) and metastasis (M); however, patterns starting from 2cm are established by the TNM classification. In the case of squamous cell carcinoma of the lips, 2cm lesions are considered extremely large. AIM: The objective of this study was to verify the correlation among epidemiologic, clinical, evolutionary and histopathological characteristics of squamous cell carcinoma of the lips, having as parameter lesions as small as 0.5cm. MATERIAL AND METHODS: In the period 1993-2000 in São Paulo, Brazil, a transversal retrospective study was performed with patients exhibiting squamous cell carcinoma of the lips. The characteristics of the tumor were investigated through the analysis of patients' medical charts, original reports of the histopathological exams and tumors' samples. The tumors were classified from 0.5 to 2.5cm. In addition, type, grade of histologic differentiation, the presence of desmoplasia, muscular, neural and vascular infiltration, and type of inflammatory infiltrate were investigated. RESULTS: The statistical analyses indicated that metastasis and recurrences do not depend on patients' gender and race. The independence of the tumor's localization, either in the upper or lower lip, and the incidence of metastasis and recurrence were demonstrated. A correlation between lesions as large as 0.5cm and the occurrence of metastasis and recurrence was verified. Furthermore, it was observed that the size of lesion determines the infiltration in other tissues. Lymphoplasmocytarian is the type of inflammatory infiltrate that was found in each and every lesion. On the other hand, in some of the lesions, the inflammatory infiltrate was associated with eosinophils without correlation to size of tumor. CONCLUSION: Smaller tumors than 2cm, from a histopathological and clinical point of view, may present a distinct evolution behavior. Most of the lesions are ulcerative, though the ulcerative-vegetative is the one that presents the most metastasis and recurrence. The ulcerative-vegetative and vegetative types are associated with the largest lesions. The size of the tumor is related, in a similar way, to grades II and III, in which higher rates of metastasis and recurrence were observed. In decreasing order of frequency, the tumor invades muscles, nerves and blood vessels, which can be related to the size of the lesion. Thus, in order to have metastasis, it is necessary to have infiltration of muscles. Yet, blood vessels can only be involved when there is concomitant infiltration of nerves. As a result, desmoplasia is directly related to size of the lesion as well as to occurrence of metastasis.Dentre os cânceres do lábio de 90% a 95% dos casos afetam o lábio inferior, sendo o carcinoma espinocelular o mais freqüente. A classificação TNM sintetiza as características clínicas do tumor, permitindo realizar um prognóstico e possibilitando comparações dos resultados. Relaciona três parâmetros: tamanho do tumor (T), propagação aos gânglios linfáticos regionais (N) e metástases à distância (M), mas estabelece padrões a partir de 2cm. Para o carcinoma espinocelular do lábio lesões com 2cm são extremamente grandes. OBJETIVO: O objetivo deste estudo é verificar a relação entre as características epidemiológicas, clínicas, evolutivas e histopatológicas do carcinoma espinocelular do lábio tendo como parâmetro lesões de tamanhos a partir de 0,5cm. CASUÍSTICA E MÉTODO: Foi elaborado um estudo retrospectivo transversal em pacientes com carcinoma espinocelular do lábio, no período 1993-2000, em São Paulo, Brasil. Estudou-se prontuários, laudos originais dos exames histopatológicos e lâminas de tumores de pacientes com carcinoma espinocelular do lábio. Os tumores foram classificados de 0.5 em 0.5cm, sendo verificado o tipo, o grau de diferenciação histológica, a presença de desmoplasia, as invasões muscular, neural e vascular, e o tipo de infiltrado inflamatório. RESULTADOS: A análise estatística mostrou que metástases e recidivas não dependem da cor de pele ou do sexo dos pacientes e que há independência entre a localização do tumor, no lábio superior ou inferior, e a incidência de metástases e recidiva. Houve correlação entre o tamanho da lesão a partir de 0,5cm e a ocorrência de metástases e recidiva. Verificou-se que o tamanho da lesão determina a invasão em outros tecidos. O infiltrado inflamatório verificado em todas as lesões era linfoplasmocitário e, em algumas, associado com eosinófilos sem relação com o tamanho do tumor. CONCLUSÃO: Tumores menores que 2cm podem apresentar comportamentos evolutivos distintos, sob o ponto de vista clínico e histopatológico. O tipo mais prevalente de lesão é o ulcerativo e o que mais metastatiza e recidiva é o úlcero-vegetante. Os tipos úlcero-vegetante e vegetante estão ligados a lesões de maior tamanho. O tamanho do tumor se relaciona, de forma semelhante, com os graus II e III, nos quais ocorrem os maiores índices de metástases e recidivas. O tumor invade em ordem decrescente de freqüência músculos, nervos e vasos sanguíneos, e esta pode ser prevista pelo tamanho da lesão. É necessária a invasão dos músculos para a ocorrência de metástases, sendo que os vasos sanguíneos somente podem estar implicados quando há invasão concomitante dos nervos. A desmoplasia está diretamente relacionada ao tamanho da lesão e à ocorrência de metástases.UNIFESP-EPMUNIFESP-EPM setor de EstomalogiaUNIFESP-EPM Departamento de DermatologiaUNIFESP, EPM, setor de EstomalogiaUNIFESP, EPM Depto. de DermatologiaSciEL

The prevalence of hepatitis A antibodies in HIV exposed and/or infected children and adolescents

OBJECTIVE: To evaluate the prevalence of hepatitis A virus antibodies in HIV-exposed and/or HIV-infected children and adolescents. METHODS: Between September 1996 and August 2002, 352 patients (200 exposed, but not HIV-infected and 152 HIV exposed and infected) were included in this study. These children and adolescents (age ranged between 1 and 14 years) were all followed up at the Pediatric AIDS Clinic of the Federal University of São Paulo (UNIFESP) and had anti-HAV antibodies determined by a commercially available ELISA method (tests for total anti-HAV antibodies and specific IgM antibodies) (Dia Sorin and Radim). Statistical analyses were done with chi-squared and t test. RESULTS: The prevalence of hepatitis A virus antibodies in HIV-infected and HIV-exposed, but uninfected patients was 34% and 19.7%, respectively. We noticed that in the age range between 2 years and 10 years, the group of HIV-infected children presented a higher prevalence of hepatitis A virus antibodies (35.5%) than the group of uninfected children (16.7%) (p = 0.005). In the HIV infected group, children from B and C categories had a prevalence of hepatitis A virus antibodies (40.5%) higher than N and A categories (24.1%) (p = 0.042). Mean age did not differ when children from B and C categories were compared with N and A categories (5.18 and 5.66 years, respectively) (p = 0.617). CONCLUSIONS: The prevalence of hepatitis A virus antibodies in HIV exposed and/or infected children and adolescents between 1 and 14 years old was 26%. Considering the possibility of HIV infection aggravation when associated with hepatitis A virus infection, we suggest that hepatitis A virus inactivated vaccine should be administered to these patients.OBJETIVO: Avaliar a prevalência de anticorpos contra o vírus da hepatite A em crianças e adolescentes expostos e/ou infectados pelo HIV. MÉTODOS: Entre setembro de 1996 e agosto de 2002, foram incluídos neste estudo 352 crianças e adolescentes, filhos de mães soropositivas para o HIV (200 expostos e não-infectados pelo HIV, e 152 expostos e infectados pelo HIV). Essas crianças e adolescentes, com idade entre 1 e 14 anos, acompanhados no Ambulatório de AIDS Pediátrica da Universidade Federal de São Paulo (UNIFESP), fizeram teste sorológico contra hepatite A como parte da avaliação de rotina. A dosagem de anticorpos anti-HAV (anticorpos totais e IgM) foi realizada através do método ELISA (Dia Sorin e Radim). A comparação das faixas etárias entre os grupos foi feita utilizando o teste do qui-quadrado e, para comparar as médias de idade das categorias clínicas entre as crianças infectadas, utilizou-se o teste t. RESULTADOS: A prevalência de anticorpos contra o vírus da hepatite A foi de 34% nos pacientes infectados e expostos ao HIV e 19,7% no grupo de soro-revertidos (expostos ao HIV e não-infectados). Estratificando a amostra por faixa etária, observamos que, para as crianças de 2 a 10 anos, o grupo de infectados pelo HIV apresentou prevalência de anticorpos para o vírus hepatite A (35,5%) maior do que o grupo de soro-revertidos (16,7%) (p = 0,005). Dentro do grupo de infectados pelo HIV, estratificando a amostra em relação à categoria clínica da infecção pelo HIV, observamos que as crianças pertencentes às categorias B e C apresentaram prevalência de anticorpos para o vírus da hepatite A maior (40,5%) do que aquelas pertencentes às categorias N e A (24,1%) (p = 0,042), apesar de apresentarem média de idade sem diferença estatística: 5,66 anos para as categorias N e A e 5,18 anos para as categorias B e C (p = 0,617). CONCLUSÕES: A prevalência de anticorpos contra o vírus da hepatite A na população de crianças e adolescentes infectados e/ou expostos ao HIV na faixa etária de 1 a 14 anos foi de 26%. Considerando-se a possibilidade de agravamento da infecção pelo HIV quando associada à infecção pelo vírus da hepatite A, sugerimos a profilaxia vacinal nesse grupo de indivíduos.Universidade Federal de São Paulo (UNIFESP) Ambulatório de Infectologia PediátricaUniversidade Federal de São Paulo (UNIFESP) Laboratório de Infectologia PediátricaUniversidade Federal de São Paulo (UNIFESP) Departamento de PediatriaUniversidade Federal de São Paulo (UNIFESP) Centro de Referência para Imunobiológicos EspeciaisUNIFESP Ambulatório da Disciplina de Infectologia PediátricaUNIFESP, Ambulatório de Infectologia PediátricaUNIFESP, Laboratório de Infectologia PediátricaUNIFESP, Depto. de PediatriaUNIFESP, Centro de Referência para Imunobiológicos EspeciaisUNIFESP, Ambulatório da Disciplina de Infectologia PediátricaSciEL

Genes for hereditary sensory and autonomic neuropathies: a genotype–phenotype correlation

Hereditary sensory and autonomic neuropathies (HSAN) are clinically and genetically heterogeneous disorders characterized by axonal atrophy and degeneration, exclusively or predominantly affecting the sensory and autonomic neurons. So far, disease-associated mutations have been identified in seven genes: two genes for autosomal dominant (SPTLC1 and RAB7) and five genes for autosomal recessive forms of HSAN (WNK1/HSN2, NTRK1, NGFB, CCT5 and IKBKAP). We performed a systematic mutation screening of the coding sequences of six of these genes on a cohort of 100 familial and isolated patients diagnosed with HSAN. In addition, we screened the functional candidate gene NGFR (p75/NTR) encoding the nerve growth factor receptor. We identified disease-causing mutations in SPTLC1, RAB7, WNK1/HSN2 and NTRK1 in 19 patients, of which three mutations have not previously been reported. The phenotypes associated with mutations in NTRK1 and WNK1/HSN2 typically consisted of congenital insensitivity to pain and anhidrosis, and early-onset ulcero-mutilating sensory neuropathy, respectively. RAB7 mutations were only found in patients with a Charcot-Marie-Tooth type 2B (CMT2B) phenotype, an axonal sensory-motor neuropathy with pronounced ulcero-mutilations. In SPTLC1, we detected a novel mutation (S331F) corresponding to a previously unknown severe and early-onset HSAN phenotype. No mutations were found in NGFB, CCT5 and NGFR. Overall disease-associated mutations were found in 19% of the studied patient group, suggesting that additional genes are associated with HSAN. Our genotype–phenotype correlation study broadens the spectrum of HSAN and provides additional insights for molecular and clinical diagnosis