Prevalência de Influenza A, vírus sincicial respiratório eSARS-CoV-2 em pacientes com síndrome respiratóriaaguda grave em Passo Fundo, Rio Grande do Sul / Prevalence of Influenza A, respiratory syncytial virus andSARS-CoV-2 in patients with severe acute respiratorysyndrome in Passo Fundo, Rio Grande do Sul

A síndrome respiratória aguda grave (SRAG) é caracterizada por sintomas de febre alta, tosse e dispneia, e, na maioria dos casos, relacionada a uma quantidade reduzida de agentes infecciosos. O objetivo foi avaliar a prevalência dos vírus respiratórios Influenza A (FluA), vírus sincicial respiratório (RSV) e do novo coronavírus (SARS-CoV-2) em pacientes com internação hospitalar por SRAG. Estudo transversal, com pacientes em internação hospitalar com SRAG entre novembro de 2021 e maio de 2022. Dados sociodemográficos e clínicos e amostras da nasofaringe foram coletados/as, as quais foram submetidas à extração de RNA e testadas quanto à positividade para Influenza A, RSV e SARS-CoV-2 por meio da técnica de PCR em tempo real pelo método SYBR Green. Foram incluídos 42 pacientes, sendo 59,5% do sexo feminino, 57,1% idosos, 54,8% com ensino fundamental. A maior parte dos pacientes reportou hábito tabagista prévio ou atual (54,8%), não etilista (73,8%) e 83,3% deles apresentavam alguma comorbidade, sendo hipertensão arterial sistêmica e diabetes mellitus tipo 2 as mais prevalentes. Um total de 10,5% dos pacientes testou positivo para FluA, nenhuma amostra positiva para RSV e 76,3% positivos para SARS-CoV-2. Na população estudada, SRAG com agravo hospitalar foi observado em maior proporção, em mulheres, idosos e pessoas com comorbidades, embora sem significância estatística, sendo o novo coronavírus o agente etiológico mais relacionado, o que evidencia a patogenicidade desse agente e suas consequências ainda são evidentes após quase 2 anos de período pandêmico.

Severe acute respiratory syndrome (SARS) is characterized by symptoms of high fever, cough and dyspnea, and is in most cases related to a reduced amount of infectious agents. The objective was to assess the prevalence of respiratory viruses Influenza A (FluA), respiratory syncytial virus (RSV) and the new coronavirus (SARS-CoV-2) in patients hospitalized for SARS. Cross-sectional study, with patients hospitalized with SARS between November 2021 and May 2022. Sociodemographic and clinical data and nasopharyngeal samples were collected, which were subjected to RNA extraction and tested for positivity for Influenza A, RSV and SARS-CoV-2 using the real-time PCR technique using the SYBR Green method. 42 patients were included, 59.5% female, 57.1% elderly, 54.8% with primary education. Most patients reported previous or current smoking habits (54.8%), non-drinkers (73.8) and 83.3% of them had some comorbidity, with systemic arterial hypertension and type 2 diabetes mellitus being the most prevalent. A total of 10.5% of patients tested positive for FluA, no samples positive for RSV, and 76.3% positive for SARS-CoV-2. In the studied population, SARS with hospital injury was observed more frequently in women and the elderly, with associated comorbidities, with the new coronavirus being the most related etiological agent, which shows, although not statistically significant, that the pathogenicity of this agent and its consequences are still evident after almost 2 years of period pandemic.

The Impact of Helicobacter pylori on Laparoscopic Sleeve Gastrectomy Postoperative Complications: a Systematic Review and Meta-analysis.

We aimed to assess the impact of Helicobacter pylori infection on postoperative outcomes following laparoscopic sleeve gastrectomy (LSG). We searched Cochrane, Scopus, and PubMed databases, reviewed 1026 studies, and thoroughly analyzed 42 of them. Our final analysis included 13 studies comprising 6199 patients. We found that H. pylori infection was correlated with higher rates of risk of overall postoperative complications (OR 1.56; 95% CI 1.13, 2.16; P = 0.007) and staple line leak (OR 1.89; 95% CI 1.05, 3.41; P = 0.03). There were no significant differences in hospital length of stay or postoperative bleeding rates. Despite observed correlations between H. pylori positivity in gastric specimen and postoperative complications in LSG, definitive causation remains elusive, emphasizing the need for prospective randomized studies evaluating the effect of preoperative H. pylori screening and eradication.

P16 gene promoter methylation is associated with oncogenesis and progression of gastric carcinomas: A systematic review and meta-analysis.

INTRODUCTION: Gastric cancer is the fourth leading cause of neoplastic morbidity worldwide, and its pathogenesis has been related to genetic and epigenetic alterations in cell cycle regulatory genes, such as p16. OBJECTIVES: This systematic review was developed to investigate the association of the p16 gene promoter methylation with oncogenesis and the progression of gastric carcinomas. METHODOLOGY: MEDLINE and Scopus databases were searched for relevant subsidiary studies, with the publication until July 2021 and without language restriction. The complete protocol was registered in the PROSPERO platform under the identification 308218. The Newcastle-Ottawa scale (NOS) was used to assess the methodological quality of the included manuscripts. The meta-analysis was conducted using RevMan 5.4 ® software. The random effects model was used, Odds Ratio (OR) was calculated with 95% confidence intervals (95% CI). Heterogeneity and inferential significance were measured. RESULTS: 48 articles were aggregated in the qualitative synthesis and 47 in the meta-analysis, totaling 6599 gastric specimens evaluated. Associations of p16 methylation with the following outcomes were observed: gastric oncogenesis (p < 0.00001); intestinal metaplasia (p = 0.002); poor histological differentiation (p = 0.03); local invasion (p = 0.001); lymph node dissemination (p = 0.03); more advanced TNM staging (p = 0.01); and Epstein Barr virus infection (p < 0.00001). In contrast, no association of p16 methylation was found with Lauren's histological classification (p = 0.62); distant metastasis (p = 0.71); or Helicobacter pylori infection (p = 0.79). CONCLUSIONS: the findings described provide empirical evidence for the categorization of p16 methylation as a substantial biomolecular step in gastric carcinogenesis, and reveal a crucial role of Epstein Barr virus in triggering this epigenetic alteration.

Genital Mycoplasmas and Biomarkers of Inflammation and Their Association With Spontaneous Preterm Birth and Preterm Prelabor Rupture of Membranes: A Systematic Review and Meta-Analysis.

Genital mycoplasmas (GM), such as Mycoplasma hominis, Mycoplasma genitalium, Ureaplasma parvum, and Ureaplasma urealyticum are commonly associated with spontaneous preterm labor (SPTL), spontaneous preterm birth (PTB), and preterm prelabor rupture of membranes (PPROM). This study determined the association between GM and such adverse pregnancy outcomes. We searched for studies published 1980-2019 in MEDLINE, EMBASE, and Web of Science. Studies were eligible when GM was detected during pregnancy. We included 93 and 51 studies in determining the prevalence and the inflammatory biomarkers associated with GM, respectively, using the "metafor" package within R. The protocol was registered with PROSPERO (registration no. CRD42016047297). Women with the studied adverse pregnancy outcomes had significantly higher odds of presence with GM compared to women who delivered at term. For PTB, the odds ratios were: M. hominis (OR: 2.25; CI: 1.35-3.75; I 2: 44%), M. genitalium (OR: 2.04; CIL 1.18-3.53; I 2: 20%), U. parvum (OR: 1.75; CI: 1.47-2.07; I 2: 0%), U. urealyticum (OR: 1.50; CI: 1.08-2.07; I 2: 58%). SPTL had significantly higher odds with M. hominis (OR: 1.96; CI: 1.19-3.23; I 2: 1%) or U. urealyticum (OR: 2.37; CI: 1.20-4.70; I 2: 76%) compared to women without SPTL. Women with PPROM had significantly higher odds with M. hominis (OR: 2.09; CI: 1.42-3.08; I 2: 0%) than women without PPROM. However, our subgroup analysis based on the diagnostic test and the sample used for detecting GM showed a higher prevalence of GM in maternal samples than in fetal samples. GM presence of the cervix and vagina was associated with lower odds of PTB and preterm labor (PTL). In contrast, GM presence in the AF, fetal membrane, and placenta was associated with increased odds of PTB and PTL. However, genital mycoplasmas may not elicit the massive inflammation required to trigger PTB. In conclusion, GM presence in the fetal tissues was associated with significantly increased odds of PTB and PTL.

Cervicovaginal levels of human beta defensins during bacterial vaginosis.

AIMS: To compare the cervicovaginal levels of human beta defensin (hBD)-1, 2 and 3 of women according to the status of Nugent-defined bacterial vaginosis (BV). METHODS: A total of 634 women of reproductive age were included in the study. Participants were equally distributed in two groups: according to the classification of vaginal smears according to Nugent criteria in normal (scores 0 to 3) and BV (scores ≥7). Cervicovaginal fluid samples were used for measurements of hBDs1, 2 and 3 levels by enzyme-linked immunosorbent assay (ELISA). Levels of each hBD were compared between the two study groups using Mann-Whitney test, with p-value <0.05 considered as significant. Odds ratio (OR) and 95% confidence interval (95% CI) were calculated for sociodemographic variables and hBD1-3 levels associated with BV a multivariable analysis. Correlation between Nugent score and measured levels of hBDs1-3 were calculated using Spearman's test. RESULTS: Cervicovaginal fluids from women with BV showed lower levels of hBD-1 [median 2,400.00 pg/mL (0-27,800.00); p<0.0001], hBD-2 [5,600.00 pg/mL (0-45,800.00); p<0.0001] and hBD-3 [1,600.00 pg/mL (0-81,700.00); p = 0.012] when compared to optimal microbiota [hBD-1: [median 3,400.00 pg/mL (0-35,600.00), hBD-2: 12,300.00 pg/mL (0-95,300.00) and hBD-3: 3,000.00 pg/mL (0-64,300.00), respectively]. Multivariable analysis showed that lower levels of hBD-1 (OR: 2.05; 95% CI: 1.46-2.87), hBD-2 (OR: 1.85; 95% CI: 1.32-2.60) and hBD-3 (OR: 1.90; 95% CI: 1.37-2.64) were independently associated BV. Significant negative correlations were observed between Nugent scores and cervicovaginal levels of hBD-1 (Spearman's rho = -0.2118; p = 0.0001) and hBD-2 (*Spearman's rho = -0.2117; p = 0.0001). CONCLUSIONS: Bacterial vaginosis is associated with lower cervicovaginal levels of hBDs1-3 in reproductive-aged women.

Telomere-Related Disorders in Fetal Membranes Associated With Birth and Adverse Pregnancy Outcomes.

Telomere disorders have been associated with aging-related diseases, including diabetes, vascular, and neurodegenerative diseases. The main consequence of altered telomere is the induction of the state of irreversible cell cycle arrest. Though several mechanisms responsible for the activation of senescence have been identified, it is still unclear how a cell is indeed induced to become irreversibly arrested. Most tissues in the body will experience senescence throughout its lifespan, but intrinsic and extrinsic stressors, such as chemicals, pollution, oxidative stress (OS), and inflammation accelerate the process. Pregnancy is a state of OS, as the higher metabolic demand of the growing fetus results in increased reactive oxygen species production. As a temporary organ in the mother, senescence in fetal membranes and placenta is expected and linked to term parturition (>37 weeks of gestation). However, a persistent, overwhelming, or premature OS affects placental antioxidant capacity, with consequent accumulation of OS causing damage to lipids, proteins, and DNA in the placental tissues. Therefore, senescence and its main inducer, telomere length (TL) reduction, have been associated with pregnancy complications, including stillbirth, preeclampsia, intrauterine growth restriction, and prematurity. Fetal membranes have a notable role in preterm births, which continue to be a major health issue associated with increased risk of neo and perinatal adverse outcomes and/or predisposition to disease in later life; however, the ability to mediate a delay in parturition during such cases is limited, because the pathophysiology of preterm births and physiological mechanisms of term births are not yet fully elucidated. Here, we review the current knowledge regarding the regulation of telomere-related senescence mechanisms in fetal membranes, highlighting the role of inflammation, methylation, and telomerase activity. Moreover, we present the evidences of TL reduction and senescence in gestational tissues by the time of term parturition. In conclusion, we verified that telomere regulation in fetal membranes requires a more complete understanding, in order to support the development of successful effective interventions of the molecular mechanisms that triggers parturition, including telomere signals, which may vary throughout placental tissues.

Novel pathways of inflammation in human fetal membranes associated with preterm birth and preterm pre-labor rupture of the membranes.

Spontaneous preterm birth (PTB) and preterm pre-labor rupture of the membranes (pPROM) are major pregnancy complications. Although PTB and pPROM have common etiologies, they arise from distinct pathophysiologic pathways. Inflammation is a common underlying mechanism in both conditions. Balanced inflammation is required for fetoplacental growth; however, overwhelming inflammation (physiologic at term and pathologic at preterm) can lead to term and preterm parturition. A lack of effective strategies to control inflammation and reduce the risk of PTB and pPROM suggests that there are several modes of the generation of inflammation which may be dependent on the type of uterine tissue. The avascular fetal membrane (amniochorion), which provides structure, support, and protection to the intrauterine cavity, is one of the key contributors of inflammation. Localized membrane inflammation helps tissue remodeling during pregnancy. Two unique mechanisms that generate balanced inflammation are the progressive development of senescence (aging) and cyclic cellular transitions: epithelial to mesenchymal (EMT) and mesenchymal to epithelial (MET). The intrauterine build-up of oxidative stress at term or in response to risk factors (preterm) can accelerate senescence and promote a terminal state of EMT, resulting in the accumulation of inflammation. Inflammation degrades the matrix and destabilizes membrane function. Inflammatory mediators from damaged membranes are propagated via extracellular vesicles (EV) to maternal uterine tissues and transition quiescent maternal uterine tissues into an active state of labor. Membrane inflammation and its propagation are fetal signals that may promote parturition. This review summarizes the mechanisms of fetal membrane cellular senescence, transitions, and the generation of inflammation that contributes to term and preterm parturitions.

Aleitamento materno e seu conhecimento por alunos de enfermagem / Breastfeeding and their knowledge by nursing students / Lactancia materna y su conocimiento por los estudiantes de enfermeira

Objetivo: Investigar o conhecimento dos alunos de enfermagem sobre o aleitamento materno. Métodos: Abordagem transversal, qualitativa e quantitativa de campo desenvolvido em Universidade particular do interior de São Paulo. A coleta de dados foi realizada utilizando-se questionário, com 15 perguntas sobre amamentação, distribuídas em três partes. Foram incluídas perguntas sobre manejo na amamentação, aspectos legais na proteção ao aleitamento materno e benefícios da amamentação para o binômio. Analisou-se os dados por meio da descritiva simples e para as comparações foi utilizado a análise de variância. Resultados: Os alunos de enfermagem do oitavo semestre obtiveram desempenho geral de 81,2% nas respostas corretas, valor estatisticamente superior a uma das turmas do sétimo período, mas semelhante à outra turma. Assim a turma sétimo A e oitavo tiveram resultados significativos na pesquisa. Conclusão: Apesar do bom conhecimento sobre as vantagens do aleitamento, evidenciou-se conhecimento inferior ao esperado no que se refere ao processo fisiológico do aleitamento materno.(AU)

Objective: To investigate the knowledge of nursing students about breastfeeding. Methods: Cross-sectional, qualitative and quantitative approach developed at a private university in an inner city in São Paulo State. Data collection was performed using a questionnaire comprising 15 questions about breastfeeding, distributed in three parts. Questions about breastfeeding management, legal aspects of breastfeeding protection and breastfeeding benefits were included. Data were analyzed using the simple descriptive and variance analysis was performed for comparisons. Results: Nursing students in the eighth semester achieved an overall performance of 81.2% in the correct answers, a value statistically superior to one of the classes in the seventh period, but similar to the other. Conclusion: Despite the good knowledge about the advantages of breastfeeding, knowledge was shown to be lower than expected with regard to the physiological process of breastfeeding.(AU)

Objetivo: Investigar el conocimiento de los estudiantes de enfermería sobre la lactancia materna. Métodos: enfoque transversal, cualitativo y cuantitativo desarrollado en una universidad privada en una ciudad del interior del estado de São Paulo. La recolección de datos se realizó mediante un cuestionario que comprende 15 preguntas sobre lactancia materna, distribuidas en tres partes. Se incluyeron preguntas sobre el manejo de la lactancia materna, los aspectos legales de la protección y los beneficios de la lactancia materna. Los datos se analizaron mediante el análisis descriptivo simple y se realizó un análisis de varianza para las comparaciones. Resultados: Los estudiantes de enfermería en el octavo semestre lograron un rendimiento general del 81.2% en las respuestas correctas, un valor estadísticamente superior a una de las clases en el séptimo período, pero similar al otro. Conclusión: a pesar del buen conocimiento sobre las ventajas de la lactancia materna, se demostró que el conocimiento es menor de lo esperado con respecto al proceso fisiológico de la lactancia materna.(AU)

Profile of pro-inflammatory cytokines in colostrum of nursing mothers at the extremes of reproductive age.

Gestations at the extremes of reproductive age are characterized as high-risk pregnancies, conditions that might influence colostrum composition. This first milk secretion contains nutrients necessary for the development and immunity of the newborn; therefore, this study aims to compare adolescent and advanced maternal age mothers regarding sociodemographic, gestational, and perinatal characteristics and the colostrum levels of pro-inflammatory cytokines in these groups of study. This cross-sectional study has compared sociodemographic, gestational and perinatal data from adolescent mothers (between 10 and 24 years old) (n = 117), advanced maternal age mothers (over 35 years of age) (n = 39) and mothers considered a control group (25 to 34 years old) (n = 58). Additionally, colostrum samples were obtained from the studied and control group subjects by manual milking, between 48 and 72 hours postpartum, and the samples were analyzed for cytokine concentrations by enzyme-linked immunosorbent assay (ELISA). The majority of the studied mothers reported living a stable union, and 81.2% of the adolescent mothers did not carry out any paid activity. Mothers with advanced maternal age mainly delivered by cesarean section and presented a higher body mass index (BMI). Neonatal weight and Apgar score were not different between the groups. The concentrations of interleukin (IL)-1ß and IL-6 were higher in the colostrum of mothers with advanced age compared to adolescent mothers, but did not differ from the control group. The concentrations of IL-8 and tumor necrosis factor alpha did not differ between the three groups. Therefore, our data demonstrated that maternal age influenced the sociodemographic and gestational characteristics as well as the composition of colostrum cytokines.

Telomere Length and Telomerase Activity in Foetal Membranes from Term and Spontaneous Preterm Births.

The reduction of telomere length, the protective cap structures of chromosomes, is one of the biomarkers of senescence (a mechanism of ageing), and ageing of foetal gestational tissues is associated with both term and preterm parturition. A mechanism regulating telomere length is the activity of telomerase, an enzyme that adds telomere fragments during DNA replication and cell division; however, its role in regulating telomere length is not well studied in gestational tissues. The objective of this study is to correlate telomere length and telomerase activity in foetal membranes from term and spontaneous preterm births. Foetal membrane samples were collected from pregnant women experiencing term labour (TL), term not in labour (TNL), preterm premature rupture of membranes (pPROM) and spontaneous preterm labour (PTL) with intact membranes (n = 20/group). Telomere length and telomerase activity were analyzed by relative quantification (T/S), real-time PCR and PCR-based fluorometric detection, respectively. Data were analyzed by ANOVA or the Kruskal-Wallis test. Demographic variables were not statistically different among the groups. Foetal membranes from the TL group showed telomere length reduction compared with those from the others (p < 0.0002). Telomerase activity did not change in foetal membranes irrespective of pregnancy outcome. Telomere shortening in foetal membranes is suggestive of senescence associated with triggering of labour at term; however, this is likely independent of telomerase activity, while prematurity may be associated with senescence, but due to other mechanisms than telomere length reduction in foetal membranes.

Determination of antimicrobial susceptibility and biofilm production in Staphylococcus aureus isolated from white coats of health university students.

This study aimed at detecting Staphylococcus aureus from white coats of college students and characterizing antimicrobial susceptibility and biofilm production. Bacterial samples (n = 300) were obtained from white coats of 100 college students from August 2015 to March 2017 S. aureus was isolated and it´s resistance profile was assessed by antimicrobial disk-diffusion technique, screening for methicillin-resistant Staphylococcus aureus (MRSA), detection of mecA gene by PCR, and determination of staphylococcal cassette chromosome mec (SCCmec) by multiplex PCR. Congo red agar (CRA) and icaA and icaD genes by PCR were used for biofilm characterization. S. aureus was identified in 45.0% of samples. Resistance of S. aureus sample to antimicrobial was seen for penicillin (72.59%), erythromycin (51.85%), cefoxitin (20.74%), oxacillin (17.04%), clindamycin (14.81%) and levofloxacin (5.18%). MRSA was detected in 53.3% of the samples with SCCmec I (52.8%), SCCmec III (25%) and SCCmec IV (11.1%). Biofilm production was observed in 94.0% S. aureus samples. These data show that biosafety measures need to be enhanced in order to prevent dissemination of multiresistant and highly adhesive bacteria across other university settings, relatives, and close persons.

Maternal age influence in human colostrum acidity composition in a Maternity in an country side city of São Paulo State / Impacto da idade materna na acidez do colostro de nutrizes em maternidade do interior do Estado de São Paulo

INTRODUCTION: OBJECTIVE: To analyze sociodemographic and gestational characteristics and to compare colostrum acidity content in adolescent and advanced maternal age mothers METHODS: This cross-sectional study included 98 adolescents (up to 18 year-old) and 33 advanced maternal age (more than 35 year-old) mothers admitted at the maternity of Presidente Prudente Regional Hospital, Brazil. An approximated volume of 10 mL of colostrum was obtained by manual expression of the breasts within 72h postpartum and stored at -20ºC up to processing. Acidity of colostrum was evaluated in Dornic degrees. Statistical analysis was performed using Mann-Whitney test or X² RESULTS: The median age adolescents and advanced maternal age mothers were 20 years (12-24) 37 years (35-45), respectively. The majority of the adolescents (66.3%) were primiparous and C-section delivery had occurred in 66.7% of advanced age mothers. Gestational body mass index (BMI) was statically higher in advanced maternal age women (32.17±5.6 vs 26.09±3.6, p< 0.0001). Acidity measurement was similar between the studied groups CONCLUSIONS: Sociodemographic and gestational characteristics of adolescents and advanced age mothers showed population heterogeneity, thus, specific follow up and orientations should be applied at each one. Colostrum constitution was constant regarding casein and proteins content, independently of maternal age

INTRODUÇÃO: OBJETIVO: Analisar as características sociodemográficas e gestacionais e comparar a acidez do colostro de nutrizes adolescentes e idade materna avançada MÉTODO: Estudo prospectivo transversal, que incluiu 98 nutrizes adolescentes (até 18 anos) e 33 nutrizes com idade materna avançada (maior que 35) atendidas na Maternidade do Hospital Regional de Presidente Prudente. Amostras de aproximadamente 10 mL de colostro foram obtidas por meio da ordenha manual entre 48 a 72h após o parto, e armazenadas a -20ºC até o processamento. A acidez foi verificada por meio da titulação e calculada em graus Dornic. A análise estatística foi realizada utilizando-se o teste de Mann-Whitney ou X² RESULTADOS: A mediana da idade materna das nutrizes adolescentes foi de 20 (12-24) anos, e das nutrizes com idade materna avançada foi de 37 (35-45) anos. A maioria das nutrizes adolescentes (66,3%) eram primíparas e a via de parto cesárea ocorreu em 66,7% das nutrizes em idade materna avançada. O índice de massa corpórea (IMC) gestacional foi estatisticamente maior em nutrizes com idade materna avançada (31,3±5,6 vs 26,06±3,6, p< 0,0001). A acidez do colostro das nutrizes nos extremos da idade reprodutiva não apresentou diferença significativa entre os grupos de nutrizes CONCLUSÃO: As características sociodemográficas e gestacionais de adolescentes e mães com idade materna avançada evidenciam heterogeneidade dessas populações e consequentemente, atendimento e condutas específicas devem ser aplicados a cada uma. A constituição do colostro em caiseina e proteinas apresentou-se constante, independente da idade materna

Oxidative stress induces senescence and sterile inflammation in murine amniotic cavity.

OBJECTIVE: A physiologic increase of reactive oxygen species (ROS) is observed through pregnancy. ROS-induced damage to major cellular elements, specifically protein peroxidation, can lead to fetal and placental tissue senescence and inflammation often associated with normal parturition. The purpose of this study was to examine the effects of oxidative stress (OS) in inducing changes in proteins, senescence, and sterile inflammation in pregnant mice. METHODS: CD-1 mice (n = 5/group) on day 14 of gestation were subjected to minilaparotomy and the uterine horn between gestational sacs was injected with the following: saline (control), cigarette smoke extract (CSE) CSE diluted in saline and CSE + SB 203580 (SB) (a p38 mitogen-activated protein kinase (MAPK) inhibitor). Mice were sacrificed on day 18, and amniotic sacs, placentas and amniotic fluid (AF) were collected. Protein damage was evaluated by immunostaining for 3-Nitrotyrosine modified proteins (3-NT). Activation of prosenescence p38MAPK was evaluated by western blot. Senescence features, ß-galactosidase (SA-ß-Gal) and AF inflammatory cytokines were analyzed by immunostaining and multiplex luminex-based immunoassays, respectively. The data were analyzed by ANOVA and Tukey's test, p < .05 was used for significance. RESULTS: Amniotic sac from CSE-treated animals showed significant protein peroxidation compared to control as indicated by 3-NT staining. CSE activated p38MAPK phosphorylation in amniotic sac but not in placenta. Membrane p38MAPK activation was reduced after treatment with SB. CSE increased fetal membrane senescence (staining for SA-ß-Gal) and increased AF concentrations of all evaluated cytokines. High inflammation correlated with pup loss and a decrease in placental weight. Treatment with p38MAPK inhibitor (SB) minimized damages, senescence and sterile inflammation. CONCLUSION: OS induction by cigarette smoke extract cause fetal tissue protein damage, p38MAPK activation, senescence and sterile inflammation in the amniotic cavity of mouse. Prevention of p38MAPK activation can be a novel approach to prevention of adverse pregnancy outcomes related to OS induced premature senescence.

Polybacterial stimulation suggests discrete IL-6/IL-6R signaling in human fetal membranes: Potential implications on IL-6 bioactivity.

The polybacterial invasion of the amniotic cavity and risk of preterm birth is often due to cervicovaginal bacteria such as genital mycoplasmas (Mycoplasma hominis and Ureaplasma urealyticum) and Gardnerella vaginalis. The most studied biomarker associated with preterm birth is interleukin-6 (IL-6), a pleiotropic cytokine that performs different functions based on classical or trans-signaling mechanisms. This study evaluated the changes in IL-6 and IL-6 function associated accessory molecules by human fetal membranes to determine the functional availability of IL-6 assessment in an in vitro model of polybacterial infection. Fetal membranes were treated with LPS or heat-inactivated genital mycoplasmas and G. vaginalis alone or in combination. IL-6 and its soluble receptors (sgp130, sIL-6R) were assessed in conditioned medium by immunoassays and membrane-bound receptors were evaluated in the tissue using immunohistochemistry and RT-PCR. Data from protein and gene expression were evaluated using linear mixed effects models. Data from immunohistochemistry were evaluated using one-way analysis of variance followed by the Tukey test. Genital mycoplasmas alone, or in combination, inhibited IL-6 trans-signaling with increased sgp130 production. G. vaginalis activated the classical IL-6 signaling pathway, as did LPS. Polybacterial treatment resulted in a balanced response with neither pathway being favored. The increase in IL-6 production by fetal membranes in response to infection is likely a non-specific innate response and not an indicator of a functional mediator of any labor-inducing pathways. This suggests that correlating the risk of adverse pregnancy outcomes and designing interventions based on IL-6 levels without considering soluble receptors may be an ineffective strategy.

Real-time PCR for traceability and quantification of genetically modified seeds in lots of non-transgenic soybean / PCR em tempo real para rastreabilidade e quantificação de sementes geneticamente modificados em lotes de soja não transgênica

The constant presence of genetically modified (GM) soybean in conventional seed lots has become a growing problem for international seed trade. In this context, seed companies have prompted the development of routine tests for accurate genetically modified soybean seeds detection. In this study, a quantitative PCR-based method was standardized in order to detect and quantify mixtures of seeds (i.e. certified seed) or GM grains (i.e. seeds came from field) into samples of non-GM soybean, in a way that soybean lots can be assessed within the standards established by legislation. The method involved the use of p35S-f2/petu-r1 primers targeting CP-4 enolpyruvylshikimate-3-phosphate synthase (cp4-epsps) gene (i.e. that confers herbicide tolerance in Roundup ReadyTM (RR)) for real-time PCR detection and quantification through mericon Quant GMO Detection Assay. The results revealed the method efficiency to detect and quantify the presence of even one soybean seed in batch used for routine evaluation of GM seeds. In addition, it was possible to detect of up to 0.1% of transgenic DNA relative to the soybean grains content. Thus, the sensitive GMO quantitative approach described in this study will provide support in supervising activities, and facilitate the process and control of GM soybean.

A constante presença da soja geneticamente modificada (GM) em lotes de sementes convencionais têm se tornado um grande problema para o comércio internacional de sementes. Neste contexto, as empresas de sementes estão em busca de testes de rotina extremamente precisos para a detecção de sementes de soja geneticamente modificadas. Neste estudo, um método baseado em PCR quantitativo foi padronizado para detectar e quantificar misturas de sementes (i.e. sementes certificadas) ou grãos geneticamente modificados (i.e. sementes oriundas do campo) dentro de lotes de soja não transgênica, de um modo que os lotes de soja possam ser avaliados dentro dos parâmetros estabelecidos pela legislação. O método envolveu o uso dos iniciadores p35S-f2/petu-r1 alvejando o gene CP-4 5-nolpiruvil-shikimato-3-fosfato sintase (cp4-epsps) (i.e. que confere a tolerância ao herbicida Roundup Ready® (RR)) para detecção e quantificação em PCR de tempo real via Ensaio de detecção Mericon Quant GMO. Os resultados revelaram um método eficiente para detectar e quantificar a presença de até mesmo uma única semente de soja no lote usado para a avaliação de rotina de sementes geneticamente modificadas. Adicionalmente, foi possível detectar até 0,1% de DNA transgênico relativo ao conteúdo de grãos de soja. Dessa forma, uma abordagem quantitativa sensível à soja geneticamente modificada foi descrita nesse estudo e poderá fornecer suporte em atividades de supervisão, além de facilitar o processo de controle da soja geneticamente modificada.

Pigment epithelial-derived factor in human fetal membranes.

OBJECTIVE: Our main objective was to document, pigment epithelial-derived factor (PEDF), a secreted serine protease inhibitor with anti-angiogenic, anti-inflammatory, and anti-oxidant properties, expression in human fetal membranes from preterm prelabor rupture of the membranes (pPROM) and in in vitro cultures stimulated with cigarette smoke extract (CSE) or lipopolysaccharides (LPS), two major risk factors for pPROM (behavioral and bacterial, respectively). METHOD: We documented PEDF mRNA expression in clinical samples of fetal membranes from patients with pPROM using quantitative RT-PCR. Also, mRNA and protein levels were documented in fetal membranes (from normal term cesarean sections [not in labor]) in an organ explant system stimulated with CSE or lipopolysaccharide (LPS). Immunohistochemistry (IHC) was used to localize PEDF in fetal membranes. RESULTS: We report no changes in PEDF mRNA expression in pPROM compared to term births (p = .59) or after treatment with CSE or LPS. However, by adding sulforaphane the PEDF mRNA expression increased significantly p < .000032. PEDF was localized to both amnion and chorion layers, but no difference was seen in staining intensities after CSE or LPS treatment compared to control. CONCLUSIONS: PEDF, a product of fetal membrane cells, is unaltered in pPROM or after exposure to risk factors of pPROM. The antioxidant stimulating substance sulforaphane contribute to an increase in PEDF mRNA in fetal membranes.

Histologic chorioamnionitis does not modulate the oxidative stress and antioxidant status in pregnancies complicated by spontaneous preterm delivery.

BACKGROUND: Infection induced-inflammation and other risk factors for spontaneous preterm birth (PTB) and preterm premature rupture of membranes (pPROM) may cause a redox imbalance, increasing the release of free radicals and consuming antioxidant defenses. Oxidative stress, in turn, can initiate intracellular signaling cascades that increase the production of pro-inflammatory mediators. The objective of this study was to evaluate the oxidative damage to proteins and antioxidant capacity profiles in amniochorion membranes from preterm birth (PTB) and preterm premature rupture of membranes (pPROM) and to determine the role of histologic chorioamnionitis in this scenario. METHODS: We included 27 pregnant women with PTB, 27 pPROM and 30 at term. Protein oxidative damage was assayed by 3-nitrotyrosine (3-NT) and carbonyl levels, using enzyme-linked immunosorbent assay (ELISA) and modified dinitrophenylhydrazine assay (DNPH), respectively. Total antioxidant capacity (TAC) was measured by ELISA. RESULTS: Protein oxidative damage determined by carbonyl levels was lower in PTB group than pPROM and term groups (p < 0.001). PTB group presented higher TAC compared with pPROM and term groups (p = 0.002). Histologic chorioamnionitis did not change either protein oxidative damage or TAC regardless of gestational outcome. CONCLUSION: These results corroborates previous reports that pPROM and term birth exhibit similarities in oxidative stress- induced senescence and histologic chorioamnionitis does not modulate oxidative stress or antioxidant status.

Cervicovaginal Levels of Human ß-Defensin 1, 2, 3, and 4 of Reproductive-Aged Women With Chlamydia trachomatis Infection.

OBJECTIVE: This study included women attending primary health care units in Botucatu, São Paulo, Brazil, to assess the cervicovaginal levels of human ß-defensin (hBD) 1, 2, 3, and 4 during Chlamydia trachomatis infection. PATIENTS AND METHODS: Cervicovaginal samples were collected for Pap testing and assessing the presence of infection by C. trachomatis, human papillomavirus, Neisseria gonorrhoeae, and Trichomonas vaginalis. Vaginal smears were taken to evaluate local microbiota. Human ß-defensin levels were determined using enzyme-linked immunosorbent assay in cervicovaginal fluid samples. Seventy-four women with normal vaginal microbiota and no evidence of infection were included in hBD quantification assays; 37 tested positive for C. trachomatis and 37 were negative. Statistical analysis was performed using Mann-Whitney U test. RESULTS: Women positive for C. trachomatis had significantly lower cervicovaginal hBD-1, hBD-2, and hBD-3 compared with those who tested negative (hBD-1: 0 pg/mL [0-2.1] vs 1.6 pg/mL [0-2.4], p < .0001; hBD-2: 0 pg/mL [0-3.9] vs 0.61 pg/mL [0-8.9], p = .0097; and hBD-3: 0 pg/mL [0-4.3] vs 0.28 pg/mL [0-8.4], p = .0076). Human ß-defensin 4 was not detected. CONCLUSIONS: Lower levels of hBD-1, hBD-2, and hBD-3 in cervicovaginal fluid were detected in the presence of C. trachomatis infection.

Biomarkers of spontaneous preterm birth: a systematic review of studies using multiplex analysis.

OBJECTIVE: Despite decades of research on risk indicators of spontaneous preterm birth (PTB), reliable biomarkers are still not available to screen or diagnose high-risk pregnancies. Several biomarkers in maternal and fetal compartments have been mechanistically linked to PTB, but none of them are reliable predictors of pregnancy outcome. This systematic review was conducted to synthesize the knowledge on PTB biomarkers identified using multiplex analysis. MATERIALS AND METHODS: Three electronic databases (PubMed, EMBASE and Web of Science) were searched for studies in any language reporting the use of multiplex assays for maternal biomarkers associated with PTB published from January 2005 to March 2014. RESULTS: Retrieved citations (3631) were screened, and relevant studies (33) were selected for full-text reading. Ten studies were included in the review. Forty-two PTB-related proteins were reported, and RANTES and IL-10 (three studies) followed by MIP-1ß, GM-CSF, Eotaxin, and TNF-RI (two studies) were reported more than once in maternal serum. However, results could not be combined due to heterogeneity in type of sample, study population, assay, and analysis methods. CONCLUSION: By this systematic review, we conclude that multiplex assays are a potential technological advancement for identifying biomarkers of PTB, although no single or combination of biomarkers could be identified to predict PTB risk.