Identification of a Novel Human E-Cadherin Splice Variant and Assessment of Its Effects Upon EMT-Related Events.

Epithelial Cadherin (E-cadherin) is involved in calcium-dependent cell-cell adhesion and signal transduction. The E-cadherin decrease/loss is a hallmark of Epithelial to Mesenchymal Transition (EMT), a key event in tumor progression. The underlying molecular mechanisms that trigger E-cadherin loss and consequent EMT have not been completely elucidated. This study reports the identification of a novel human E-cadherin variant mRNA produced by alternative splicing. A bioinformatics evaluation of the novel mRNA sequence and biochemical verifications suggest its regulation by Nonsense-Mediated mRNA Decay (NMD). The novel E-cadherin variant was detected in 29/42 (69%) human tumor cell lines, expressed at variable levels (E-cadherin variant expression relative to the wild type mRNA = 0.05-11.6%). Stable transfection of the novel E-cadherin variant in MCF-7 cells (MCF7Ecadvar) resulted in downregulation of wild type E-cadherin expression (transcript/protein) and EMT-related changes, among them acquisition of a fibroblastic-like cell phenotype, increased expression of Twist, Snail, Zeb1, and Slug transcriptional repressors and decreased expression of ESRP1 and ESRP2 RNA binding proteins. Moreover, loss of cytokeratins and gain of vimentin, N-cadherin and Dysadherin/FXYD5 proteins was observed. Dramatic changes in cell behavior were found in MCF7Ecadvar, as judged by the decreased cell-cell adhesion (Hanging-drop assay), increased cell motility (Wound Healing) and increased cell migration (Transwell) and invasion (Transwell w/Matrigel). Some changes were found in MCF-7 cells incubated with culture medium supplemented with conditioned medium from HEK-293 cells transfected with the E-cadherin variant mRNA. Further characterization of the novel E-cadherin variant will help understanding the molecular basis of tumor progression and improve cancer diagnosis. J. Cell. Physiol. 232: 1368-1386, 2017. © 2016 Wiley Periodicals, Inc.

Spatial distribution and temporal trends of AIDS in Brazil and regions between 2005 and 2020.

OBJECTIVE: To analyze the spatial distribution and the temporal trend of the AIDS incidence rate in Brazil from 2005 to 2020. METHODS: This is an ecological, temporal, and spatial study on AIDS cases in Brazil. Data from the Notifiable Diseases Information System were stratified by year of diagnosis, region of the country/municipalities of residence, and age group (over 13 years). Incidence rates were calculated for temporal estimation using the Joinpoint model, as well as Spatial Empirical Bayes (SEB) for spatial distribution, using the Kernel density estimator. RESULTS: The incidence rate in Brazil, in 2020, was 17.69 cases per 100 thousand inhabitants. The general trend (2005-2020) was decrease in Brazil (Annual Percent Change - APC=-2.0%), in the Southeast (APC=-4.4%) and South (APC=-3.0%) regions. The North (APC=2.3%) showed an increase trend, whereas the Southeast and Midwest regions were stationary (p>0.05). Brazil, Southeast, South, and Midwest regions showed a decrease trend in most age groups. The Northeast and North regions showed an increase in the age groups of 13-29 years and 13-24 years, respectively. The Kernel estimator showed clusters with SEB above 30/10 thousand inhabitants in the states of Paraíba, Sergipe, Alagoas, Pernambuco, São Paulo, Minas Gerais, Pará, Rio Grande do Sul, and Santa Catarina. CONCLUSION: Brazil, the Southeast, and South regions showed a decrease in the incidence rate, whereas the North region increased and the Northeast and Midwest regions were stationary. The Southeast, South, and Northeast regions presented the largest clusters of SEB.

Factors Associated with Spontaneous Clearance of Recently Acquired Hepatitis C Virus among HIV-Positive Men in Brazil.

INTRODUCTION: The objective of the present study was to describe the clinical and epidemiological aspects of recently acquired hepatitis C virus (HCV) infection and the frequency of its spontaneous clearance in a people living with the human immunodeficiency virus (PLWH) cohort. METHODS: We reviewed the medical records from all PLWH at the human immunodeficiency virus (HIV) outpatient reference clinic affiliated with the University of São Paulo, Brazil, and identified, by immunoassays and RNA-PCR individuals who acquired HCV infection between January 2015 and December 2017. The factors associated with subsequent spontaneous clearance of the infection in this group were identified and analyzed. RESULTS: Among 3143 PLWH individuals, 362 (11.5%) were coinfected with HCV. Forty-eight (13.2%) of these subjects first became HCV-positive between January 2015 and December 2017. Spontaneous HCV clearance was documented in 23 individuals (47.9%). The majority of this latter group were male (83.3%), and the median age was 31 years (23-39). The main risk group for HCV acquisition was men who had sex with men (MSM) (89.5%). In a multivariate analysis, only an elevated CD4+ T lymphocyte count at the time of seroconversion was found to be associated with subsequent HCV clearance (p = 0.025). CONCLUSIONS: In HIV-infected individuals in Sao Paulo, Brazil, most cases of recent HCV transmission were by sexual exposure. In PLWH, particularly in MSM, the individual's CD4+ T lymphocyte count is a determinant of whether an acquired HCV infection will be prolonged or will spontaneously clear.

The Changing Epidemiology of Hepatitis C Virus Acquisition Among HIV-Infected Individuals in Brazil.

Identification of mechanisms of hepatitis C virus (HCV) acquisition among HIV-infected people is critical for prevention guidance. The aim of this study was to investigate risk factors for HCV infection and variations in HCV genotype distribution in a cohort of HIV-HCV coinfected patients in Brazil. This was a cross-sectional observational epidemiological study of a cohort of HIV-HCV coinfected individuals seen at a referral center for HIV-infected patients in the city of São Paulo between January and December 2017. The time of HCV acquisition, as determined by chart review, was categorized as before 2000, between 2000 and 2009, and from 2010 onward. HCV genotypes were determined by gene amplification and analysis. Among 3,143 HIV-infected individuals analyzed, 362 (11.5%) were HCV-HIV coinfected. Overall, the reported modes of HCV acquisition were sexual exposure in 172 (47.5%), injection drug use (IDU) in 86 (23.8%), use of inhaled drugs in 67 (18.5%) and blood transfusion in 10 (2.8%) individuals. All individuals who acquired HCV after IDU became infected before 2010. HCV acquisition by sexual contact was reported by 26.4%, 65.9%, and 63.8% of patients before 2000, between 2000 and 2009, and from 2010, respectively. There was an increase (p < .001) in the proportion of cases due to sexual transmission from the period before 2000 (26.4%) to between 2000 and 2009 (65.9%). There was no corresponding increase from 2000 and 2009 to after 2010 (p = .751). HCV genotype 1 was most prevalent at all time periods. The genotype 3 frequency decreased over time (test for trend p < .001), whereas genotype 4, extremely uncommon before 2010, became the second most prevalent genotype from 2010 onward. In HIV-infected individuals in Sao Paulo, Brazil, sexual transmission has replaced IDU as the most frequent mode of HCV acquisition.

Corrigendum: FXYD5/Dysadherin, a biomarker of endometrial cancer myometrial invasion and aggressiveness: its relationship with TGF-ß1 and NF-κB pathways.

[This corrects the article DOI: 10.3389/fonc.2019.01306.].

Expression analysis of epithelial cadherin and related proteins in IBH-6 and IBH-4 human breast cancer cell lines.

Epithelial cadherin (E-cadherin) is a 120 kDa cell-cell adhesion molecule involved in the establishment of epithelial adherens junctions. It is connected to the actin cytoskeleton by adaptor proteins such as beta-catenin. Loss of E-cadherin expression/function has been related to tumor progression and metastasis. Several molecules associated with down-regulation of E-cadherin have been described, within them neural cadherin, Twist and dysadherin. Human breast cancer cell lines IBH-6 and IBH-4 were developed from ductal primary tumors and show characteristic features of malignant epithelial cells. In this study expression of E-cadherin and related proteins in IBH-6 and IBH-4 cell lines was evaluated. In IBH-6 and IBH-4 cell extracts, only an 89 kDa E-cadherin form (Ecad89) was detected, which is truncated at the C-terminus and is present at low levels. Moreover, no accumulation of the 86 kDa E-cadherin ectodomain and of the 38 kDa CTF1 fragment was observed. IBH-6 and IBH-4 cells showed an intracellular scattered E-cadherin localization. beta-catenin accompanied E-cadherin localization, and actin stress fibers were identified in both cell types. E-cadherin mRNA levels were remarkably low in IBH-6 and IBH-4 cells. The E-cadherin mRNA and genomic sequence encoding exons 14-16 could not be amplified in either cell line. Neither the mRNA nor the protein of neural cadherin and dysadherin were detected. Up-regulation of Twist mRNA was found in both cell lines. In conclusion, IBH-6 and IBH-4 breast cancer cells show down-regulation of E-cadherin expression with aberrant protein localization, and up-regulation of Twist; these features can be related to their invasive/metastatic characteristics.

Comparative randomised trial of high and conventional doses of praziquantel in the treatment of schistosomiasis mansoni.

The efficacy of oral praziquantel in the treatment of schistosomiasis has been considered low by most public health institutions. In this paper, we compared the efficacy of two dosages of praziquantel (80 mg/kg vs. 50 mg/kg) in patients with chronic schistosomiasis mansoni. Two hundred eighty-eight patients with schistosomiasis from a community in Brazil were randomly divided into two groups: 145 patients (Group 1) received 80 mg/kg body weight of oral praziquantel divided in two equal doses with 1 h interval and 143 patients (Group 2) received 50 mg/kg body weight of oral praziquantel. To keep the study masked, patients in Group 2 received placebo 1 h after the first dose. All patients were subjected to clinical and ultrasonographic examination. Cure assessment was performed by repeating two stool examinations, by a quantitative method, at 30, 90 and 180 days after treatment. The morbidity of schistosomiasis was low, with a few cases of light periportal thickening and 16 cases of mild splenomegaly. The cure rates were 89.7% for Group 1 and 83.9% for Group 2. There was no difference in the efficacy of both therapeutic dosages of praziquantel assayed. The adverse reactions were more frequent with higher dosage.

Characterization of the molecular changes associated with the overexpression of a novel epithelial cadherin splice variant mRNA in a breast cancer model using proteomics and bioinformatics approaches: identification of changes in cell metabolism and an increased expression of lactate dehydrogenase B.

BACKGROUND: Breast cancer (BC) is the most common female cancer and the leading cause of cancer death in women worldwide. Alterations in epithelial cadherin (E-cadherin) expression and functions are associated to BC, but the underlying molecular mechanisms have not been fully elucidated. We have previously reported a novel human E-cadherin splice variant (E-cadherin variant) mRNA. Stable transfectants in MCF-7 human BC cells (MCF7Ecadvar) depicted fibroblast-like cell morphology, E-cadherin wild-type downregulation, and other molecular changes characteristic of the epithelial-to-mesenchymal transition process, reduced cell-cell adhesion, and increased cell migration and invasion. In this study, a two-dimensional differential gel electrophoresis (2D-DIGE) combined with mass spectrometry (MS) protein identification and bioinformatics analyses were done to characterize biological processes and canonical pathways affected by E-cadherin variant expression. RESULTS: By 2D-DIGE and MS analysis, 50 proteins were found differentially expressed (≥ Δ1.5) in MCF7Ecadvar compared to control cells. Validation of transcript expression was done in the ten most overexpressed and underexpressed proteins. Bioinformatics analyses revealed that 39 of the 50 proteins identified had been previously associated to BC. Moreover, metabolic processes were the most affected, and glycolysis the canonical pathway most altered. The lactate dehydrogenase B (LDHB) was the highest overexpressed protein, and transcript levels were higher in MCF7Ecadvar than in control cells. In agreement with these findings, MCF7Ecadvar conditioned media had lower glucose and higher lactate levels than control cells. MCF7Ecadvar cell treatment with 5 mM of the glycolytic inhibitor 2-deoxy-glucose led to decreased cell viability, and modulation of LDHB expression in MCF7Ecadvar cells with a specific small interfering RNA resulted in decreased cell proliferation. Finally, a positive association between expression levels of the E-cadherin variant and LDHB transcripts was demonstrated in 21 human breast tumor tissues, and breast tumor samples with higher Ki67 expression showed higher LDHB mRNA levels. CONCLUSIONS: Results from this investigation contributed to further characterize molecular changes associated to the novel E-cadherin splice variant expression in BC cells. They also revealed an association between expression of the novel variant and changes related to BC progression and aggressiveness, in particular those associated to cell metabolism.

Characteristics of a hepatitis C patient cohort at a specialized tertiary care facility: Identifying criteria to improve the allocation of public health resources.

OBJECTIVES: Our objective was to analyze, in a population treated for hepatitis C infection at a tertiary care treatment unit, the prevalence of comorbidities and extrahepatic manifestations, the range and degree of the clinical complexity and the associations between advanced liver disease and clinical variables. METHODS: Medical records from chronically infected hepatitis C patients seen at a dedicated treatment facility for complex cases in the Infectious Diseases Division of Hospital das Clínicas in Brazil were analyzed. Clinical complexity was defined as the presence of one or more of the following conditions: advanced liver disease (Metavir score F3 or F4 and/or clinical manifestations or ultrasound/endoscopy findings consistent with cirrhosis) or hepatocellular carcinoma and/or 3 or more extrahepatic manifestations and/or comorbidities concomitantly. RESULTS: Among the 1574 patients analyzed, only 41% met the definition of being clinically complex. Cirrhosis or hepatocarcinoma was identified in 22.2% and 1.8% of patients, respectively. According to multiple logistic regression analysis, male sex (p=0.007), age>40 years (p<0.001) and the presence of metabolic syndrome (p=0.008) were independently associated with advanced liver disease. CONCLUSION: The majority of patients did not meet the criteria for admittance to this specialized tertiary service, reinforcing the need to reevaluate public health policies. Enhanced utilization of existing basic and intermediate complexity units for the management of less complex hepatitis C cases could improve care and lower costs.

FXYD5/Dysadherin, a Biomarker of Endometrial Cancer Myometrial Invasion and Aggressiveness: Its Relationship With TGF-ß1 and NF-κB Pathways.

Objective: Endometrial cancer (EC) is the second most common gynecological cancer worldwide. Myometrial invasion (MI) is a key event in EC dissemination. This study aimed to evaluate FXYD5/dysadherin (FXYD5/Dys) expression in EC tissue and uterine aspirate (UA) biopsies and to assess molecular/functional changes associated with its expression in cellular models. Methods: FXYD5/Dys messenger RNA (mRNA) levels were determined in EC tissue and UA biopsies. FXYD5/Dys expression was evaluated in EC RNAseq data from The Cancer Genome Atlas (TCGA) and GENEVESTIGATOR tools. FXYD5/Dys impact on E-cadherin expression and cell behavior was assessed in EC Hec1a cells treated with transforming growth factor (TGF)-ß1, stably transfected with ETV5, and transiently transfected with FXYD5/Dys small interfering RNA (siRNA) or pcDNA3-FXYD5/Dys plasmid. Results: FXYD5/Dys was associated with EC aggressiveness, finding high mRNA levels in tumors depicting MI > 50%, Grade 3, and intermediate/high risk of recurrence. FXYD5/Dys was highly expressed at the tumor invasive front compared to the superficial area. Most results were recapitulated in UA biopsies. FXYD5/Dys modulation in Hec1a cells altered cell migration/adhesion and E-cadherin expression. TGF-ß1 treatment of Hec1a cells induced FXYD5/Dys expression. TCGA-UCEC RNAseq analysis revealed a positive correlation between FXYD5/Dys, TGF-ß1, and plasminogen activator inhibitor (PAI)-1 mRNA levels. FXYD5/Dys induced nuclear factor (NF)-κB pathway activation in Hec1a cells. FXYD5/Dys mRNA levels positively correlated with transcriptional activation of NF-κB p65-regulated genes. Survival analysis revealed patient segregation into low- and high-risk groups, the latter depicting the highest FXYD5/Dys, PAI-1, tumor necrosis factor (TNF)-α, and TGF-ß1 mRNA levels and shorter survival rates. Conclusion: FXYD5/Dys is a novel biomarker of EC progression related to TGF-ß1 and NF-κB pathways that collectively promote tumor dissemination and result in poor patient prognosis.

Spatial distribution and temporal trends of AIDS in Brazil and regions between 2005 and 2020 / Distribuição espacial e tendência temporal da AIDS no Brasil e regiões entre 2005 e 2020

ABSTRACT: Objective: To analyze the spatial distribution and the temporal trend of the AIDS incidence rate in Brazil from 2005 to 2020. Methods: This is an ecological, temporal, and spatial study on AIDS cases in Brazil. Data from the Notifiable Diseases Information System were stratified by year of diagnosis, region of the country/municipalities of residence, and age group (over 13 years). Incidence rates were calculated for temporal estimation using the Joinpoint model, as well as Spatial Empirical Bayes (SEB) for spatial distribution, using the Kernel density estimator. Results: The incidence rate in Brazil, in 2020, was 17.69 cases per 100 thousand inhabitants. The general trend (2005-2020) was decrease in Brazil (Annual Percent Change - APC=-2.0%), in the Southeast (APC=-4.4%) and South (APC=-3.0%) regions. The North (APC=2.3%) showed an increase trend, whereas the Southeast and Midwest regions were stationary (p>0.05). Brazil, Southeast, South, and Midwest regions showed a decrease trend in most age groups. The Northeast and North regions showed an increase in the age groups of 13-29 years and 13-24 years, respectively. The Kernel estimator showed clusters with SEB above 30/10 thousand inhabitants in the states of Paraíba, Sergipe, Alagoas, Pernambuco, São Paulo, Minas Gerais, Pará, Rio Grande do Sul, and Santa Catarina. Conclusion: Brazil, the Southeast, and South regions showed a decrease in the incidence rate, whereas the North region increased and the Northeast and Midwest regions were stationary. The Southeast, South, and Northeast regions presented the largest clusters of SEB.

RESUMO: Objetivo: Analisar a distribuição espacial e a tendência temporal da taxa de incidência de AIDS no Brasil no período de 2005 a 2020. Métodos: Estudo ecológico, temporal e espacial sobre os casos de AIDS no Brasil. Dados provenientes do Sistema de Informação de Agravos de Notificação do Departamento de Informática do Sistema Único de Saúde foram estratificados por ano do diagnóstico, região do país/municípios de residência e faixa etária (acima de 13 anos). Foram calculadas as taxas de incidência (TI) para a estimativa temporal por meio do modelo de joinpoint, bem como as taxas bayesianas empíricas espaciais (TBEE) para a distribuição espacial pelo estimador de densidade de Kernel. Resultados: A TI no Brasil no ano de 2020 foi de 17,69 casos para cada 100 mil habitantes. A tendência geral (2005-2020) foi de diminuição no Brasil (variação percentual anual — VPA=-2,0%), Sudeste (VPA=-4,4%) e Sul (VPA=-3,0%). O Norte (VPA=2,3%) demonstrou aumento, enquanto o Sudeste e Centro-oeste foram estacionários (p>0,05). O Brasil, Sudeste, Sul e Centro-oeste apresentaram tendência de diminuição na maioria das faixas etárias. O Nordeste e Norte apresentaram aumento nas faixas etárias de 13 a 29 anos e 13 a 24 anos, respectivamente. O estimador de Kernel demonstrou conglomerados com TBEE acima de 30/10 mil habitantes nos estados de Paraíba, Sergipe, Alagoas, Pernambuco, São Paulo, Minas Gerais, Pará, Rio Grande do Sul e Santa Catarina. Conclusão: O Brasil e as Regiões Sudeste e Sul apresentaram diminuição da TI, enquanto o Norte aumentou e o Nordeste e Centro-oeste foram estacionários. As Regiões Sudeste, Sul e Nordeste apresentaram os maiores conglomerados das TBEE.

E-cadherin: A determinant molecule associated with ovarian cancer progression, dissemination and aggressiveness.

Ovarian cancer (OC) is the fifth cancer death cause in women worldwide. The malignant nature of this disease stems from its unique dissemination pattern. Epithelial-to-mesenchymal transition (EMT) has been reported in OC and downregulation of Epithelial cadherin (E-cadherin) is a hallmark of this process. However, findings on the relationship between E-cadherin levels and OC progression, dissemination and aggressiveness are controversial. In this study, the evaluation of E-cadherin expression in an OC tissue microarray revealed its prognostic value to discriminate between advanced- and early-stage tumors, as well as serous tumors from other histologies. Moreover, E-cadherin, Neural cadherin (N-cadherin), cytokeratins and vimentin expression was assessed in TOV-112, SKOV-3, OAW-42 and OV-90 OC cell lines grown in monolayers and under anchorage-independent conditions to mimic ovarian tumor cell dissemination, and results were associated with cell aggressiveness. According to these EMT-related markers, cell lines were classified as mesenchymal (M; TOV-112), intermediate mesenchymal (IM; SKOV-3), intermediate epithelial (IE; OAW-42) and epithelial (E; OV-90). M- and IM-cells depicted the highest migration capacity when grown in monolayers, and aggregates derived from M- and IM-cell lines showed lower cell death, higher adhesion to extracellular matrices and higher invasion capacity than E- and IE-aggregates. The analysis of E-cadherin, N-cadherin, cytokeratin 19 and vimentin mRNA levels in 20 advanced-stage high-grade serous human OC ascites showed an IM phenotype in all cases, characterized by higher proportions of N- to E-cadherin and vimentin to cytokeratin 19. In particular, higher E-cadherin mRNA levels were associated with cancer antigen 125 levels more than 500 U/mL and platinum-free intervals less than 6 months. Altogether, E-cadherin expression levels were found relevant for the assessment of OC progression and aggressiveness.

Characteristics of a hepatitis C patient cohort at a specialized tertiary care facility: Identifying criteria to improve the allocation of public health resources

OBJECTIVES: Our objective was to analyze, in a population treated for hepatitis C infection at a tertiary care treatment unit, the prevalence of comorbidities and extrahepatic manifestations, the range and degree of the clinical complexity and the associations between advanced liver disease and clinical variables. METHODS: Medical records from chronically infected hepatitis C patients seen at a dedicated treatment facility for complex cases in the Infectious Diseases Division of Hospital das Clínicas in Brazil were analyzed. Clinical complexity was defined as the presence of one or more of the following conditions: advanced liver disease (Metavir score F3 or F4 and/or clinical manifestations or ultrasound/endoscopy findings consistent with cirrhosis) or hepatocellular carcinoma and/or 3 or more extrahepatic manifestations and/or comorbidities concomitantly. RESULTS: Among the 1574 patients analyzed, only 41% met the definition of being clinically complex. Cirrhosis or hepatocarcinoma was identified in 22.2% and 1.8% of patients, respectively. According to multiple logistic regression analysis, male sex (p=0.007), age>40 years (p<0.001) and the presence of metabolic syndrome (p=0.008) were independently associated with advanced liver disease. CONCLUSION: The majority of patients did not meet the criteria for admittance to this specialized tertiary service, reinforcing the need to reevaluate public health policies. Enhanced utilization of existing basic and intermediate complexity units for the management of less complex hepatitis C cases could improve care and lower costs.

High rate of virologic suppression with darunavir/ritonavir plus optimized background therapy among highly antiretroviral-experienced HIV-infected patients: results of a prospective cohort study in São Paulo, Brazil.

OBJECTIVES: To assess the virologic and immunological response of darunavir/ritonavir plus optimized background therapy in highly antiretroviral-experienced HIV-infected patients in Brazil. METHODS: Prospective cohort study carried out in a tertiary center in Sao Paulo, Brazil. Three-class antiretroviral-experienced patients with confirmed virologic failure began darunavir/ritonavir plus optimized background therapy (nucleoside/tide reverse transcriptase inhibitors ± raltegravir ± enfuvirtide ± maraviroc) after performing a genotypic resistance assay. Clinical evaluation and laboratory tests were collected at baseline and at weeks 12, 24, and 48. Multivariate analysis was performed to identify predictors of virologic response at 48 weeks. RESULTS: Ninety-two patients were included. The median of darunavir resistant mutation was 1 (range 0-6). The median genotypic sensitivity score in the optimized background therapy was 2 (interquartile range 1-2). At week 48, 83% (95% CI: 75-90%) had an HIV RNA level <50 copies/mL and the median CD4 cell count was 301 (interquartile range 224-445) cells/mm(3). Baseline HIV RNA >100000 copies/mL was inversely associated with virologic success at week 48 (HR: 0.22, 95% CI: 0.06-0.85, p=0.028). CONCLUSIONS: Darunavir/ritonavir plus optimized background therapy was a highly effective salvage regimen under clinical routine conditions in a referral center in Brazil, which is similar to the reported in high-income countries.

Expression of dysadherin in the human male reproductive tract and in spermatozoa.

OBJECTIVE: To study expression of dysadherin in human testis, epididymis, and spermatozoa. DESIGN: Prospective study. SETTING: Basic research laboratory. PATIENT(S): Testis, epididymis, and testicular spermatozoa from patients under treatment and semen from volunteer donors. INTERVENTION(S): Reverse transcription-polymerase chain reaction, immunohistochemistry, immunocytochemistry, and Western immunoblotting. MAIN OUTCOME MEASURE(S): Dysadherin messenger RNA (mRNA) analysis in testis, epididymis, and ejaculated spermatozoa, immunohistochemistry of both tissues, Western immunoblotting of tissue/cell extracts, and immunocytochemistry of spermatozoa. RESULT(S): Dysadherin mRNA was found in testis, epididymis, and ejaculated spermatozoa. Whereas testis and spermatozoa exhibited a distinctive 91-kDa protein form, the epididymis showed a 50-kDa moiety, also found in MDA-MB-231 breast cancer cells. Nucleotide sequence analysis revealed >99% homology between testicular and somatic cell mRNA, suggesting differential protein glycosylation. Dysadherin was immunodetected in round spermatids and testicular/ejaculated spermatozoa. It localizes to the acrosomal region and flagellum and colocalized with E-cadherin in the head and with the Na(+),K(+)-ATPase α4 subunit in the flagellum. CONCLUSION(S): This is the first report on expression of dysadherin in the male gonad and in spermatozoa. Its colocalization with E-cadherin and Na(+),K(+)-ATPase leads us to postulate a role for dysadherin as a modulator of sperm function.

High rate of virologic suppression with darunavir/ritonavir plus optimized background therapy among highly antiretroviral-experienced HIV-infected patients: results of a prospective cohort study in São Paulo, Brazil

OBJECTIVES: To assess the virologic and immunological response of darunavir/ritonavir plus optimized background therapy in highly antiretroviral-experienced HIV-infected patients in Brazil. METHODS: Prospective cohort study carried out in a tertiary center in Sao Paulo, Brazil. Three-class antiretroviral-experienced patients with confirmed virologic failure began darunavir/ritonavir plus optimized background therapy (nucleoside/tide reverse transcriptase inhibitors ± raltegravir ± enfuvirtide ± maraviroc) after performing a genotypic resistance assay. Clinical evaluation and laboratory tests were collected at baseline and at weeks 12, 24, and 48. Multivariate analysis was performed to identify predictors of virologic response at 48 weeks. RESULTS: Ninety-two patients were included. The median of darunavir resistant mutation was 1 (range 0-6). The median genotypic sensitivity score in the optimized background therapy was 2 (interquartile range 1-2). At week 48, 83% (95% CI: 75-90%) had an HIV RNA level <50 copies/mL and the median CD4 cell count was 301 (interquartile range 224-445) cells/mm³. Baseline HIV RNA >100 000 copies/mL was inversely associated with virologic success at week 48 (HR: 0.22, 95% CI: 0.06-0.85, p = 0.028). CONCLUSIONS: Darunavir/ritonavir plus optimized background therapy was a highly effective salvage regimen under clinical routine conditions in a referral center in Brazil, which is similar to the reported in high-income countries.

Comparative randomised trial of high and conventional doses of praziquantel in the treatment of schistosomiasis mansoni

The efficacy of oral praziquantel in the treatment of schistosomiasis has been considered low by most public health institutions. In this paper, we compared the efficacy of two dosages of praziquantel (80 mg/kg vs. 50 mg/kg) in patients with chronic schistosomiasis mansoni. Two hundred eighty-eight patients with schistosomiasis from a community in Brazil were randomly divided into two groups: 145 patients (Group 1) received 80 mg/kg body weight of oral praziquantel divided in two equal doses with 1 h interval and 143 patients (Group 2) received 50 mg/kg body weight of oral praziquantel. To keep the study masked, patients in Group 2 received placebo 1 h after the first dose. All patients were subjected to clinical and ultrasonographic examination. Cure assessment was performed by repeating two stool examinations, by a quantitative method, at 30, 90 and 180 days after treatment. The morbidity of schistosomiasis was low, with a few cases of light periportal thickening and 16 cases of mild splenomegaly. The cure rates were 89.7 percent for Group 1 and 83.9 percent for Group 2. There was no difference in the efficacy of both therapeutic dosages of praziquantel assayed. The adverse reactions were more frequent with higher dosage.