Estudo do papel da proteína multifuncional APE1/Ref-1 sobre a resposta inflamatória na meningite bacteriana

Despite advances in antibiotic therapy, bacterial meningitis (BM) remains with high mortality and morbidity rates in worldwide. One important mechanism associated to sequels during disease is the intense inflammatory response which promotes an oxidative burst and release of reactive oxygen species, consequently leading to cell death. Activation of DNA repair enzymes during oxidative stress has been demonstrated in several neurological disorders. APE1/Ref-1 is a multifunctional protein involved in DNA repair and plays a redox function on transcription factors such as NFkB and AP-1.The aim of this study was assess the role of APE1/Ref-1 on inflammatory response and the possibility of its modulation to reduce the sequels of the disease. Firstly it was performed an assay to measure cytokine in cerebrospinal fluid of patients with BM due to Streptococcus pneumoniae and Neisseriae meningitides. Further, a cellular model of inflammation was used to observe the effect of the inhibition of the endonuclease and redox activity of APE1/Ref-1 on cytokine levels. Additionally, APE1/Ref-1 expression in cortex and hippocampus of rat with MB after vitamin B6 treatment was evaluated. Altogether, results showed a similar profile of cytokines in the cerebrospinal fluid of patients from both pathogens, although IFNy showed higher expression in patients with BM caused by S. pneumoniae. On the other hand, inhibitors of APE1/Ref-1 reduced cytokine levels, mainly TNF-α. Reduction of oxidative stress markers was also observed after introduction of inhibitors in the LPS-stimulated cell. In the animal model, BM increased the expression of the protein APE1/Ref-1, while vitamin B6 promoted reduction. Thereby, this data rise important factors to be considered in pathogenesis of BM, e.g., IFNy can be used as prognostic factor during corticosteroid therapy, APE1/Ref-1 can be an important target to modulate the level of inflammation and VIII oxidative stress, and vitamin B6 seems modulates several proteins related to cell death. So, this study highlights a new understanding on the role of APE1/Ref-1 on the inflammation and the oxidative stress during inflammation condition

Análise do efeito de polimorfismos não-sinônimos em genes de reparo de DNA da via BER na resposta inflamatória da meningite

In vitro and in animal models, APE1, OGG1, and PARP-1 have been proposed as being involved with inflammatory response. In this work, we have investigated if the SNPs APE1 Asn148Glu, OGG1 Ser326Cys, and PARP-1 Val762Ala are associated to meningitis and also developed a system to enable the functional analysis of polymorphic proteins. Patients with bacterial meningitis (BM), aseptic meningitis (AM) and controls (non-infected) genotypes were investigated by PIRA-PCR or PCR-RFLP. DNA damages were detected in genomic DNA by Fpg treatment. IgG and IgA were measured from plasma and the cytokines and chemokines were measured from cerebrospinal fluid samples using Bio-Plex assays. The levels of NF-κB and c-Jun were measured in CSF by dot blot assays. A significant (P<0.05) increase in the frequency of APE1 148Glu allele in BM and AM patients was observed. A significant increase in the genotypes Asn/Asn in control group and Asn/Glu in BM group was also found. For the SNP OGG1 Ser326Cys, the genotype Cys/Cys was more frequent (P<0.05) in BM group. The frequency of PARP-1 Val/Val genotype was higher in control group (P<0.05). The occurrence of combined SNPs increased significantly in BM patients, indicating that these SNPs may be associated to the disease. Increasing in sensitive sites to Fpg was observed in carriers of APE1 148Glu allele or OGG1 326Cys allele, suggesting that SNPs affect DNA repair activity. Alterations in IgG production were observed in the presence of SNPs APE1Asn148Glu, OGG1Ser326Cys or PARP-1Val762Ala. Reductions in the levels ofIL-6, IL-1Ra, MCP-1/CCL2and IL-8/CXCL8 were observed in the presence of APE1148Glu allele in BM patients, however no differences were observed in the levels of NF-κB and c-Jun considering genotypes and analyzed groups. Using APE1 as model, a system to enable the analysis of cellular effects and functional characterization of polymorphic proteins was developed using strategies of cloning APE1 cDNA in pIRES2-EGFP vector, cellular transfection of the construction obtained, siRNA for endogenous APE1 and cellular cultures genotyping. In conclusion, we obtained evidences of an effect of SNPs in DNA repair genes on the regulation of immune response. This is a pioneering work in the field that shows association of BER variant enzymes with an infectious disease in human patients, suggesting that the SNPs analyzed may affect immune response and damage by oxidative stress level during brain infection. Considering these data, new approaches of functional characterization must be developed to better analysis and interactions of polymorphic proteins in response to this context

Marcadores genéticos associados com a resposta inflamatória na meningite bacteriana

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior

Atuação de técnicos de enfermagem junto ao recém-nascido com dor em uma unidade de terapia intensiva neonatal

The pain is a sensuous and emotional experience unpleasant associated or related to real injury or potencial of the tissues. It is considered an individual and subjective experience generally has been described in the literature about in the neonatal stage a lot. This study has descriptive and exploratory character with a qualitative approach. The study has with objectives to analyze the performance of the nursing technicians working with newborns admitted in the ITUN, seeking to describe the perception of the nursing technicians about the pain, identify the parameters used for the detection and evaluation of pain in them, trying to describe the ons of this team about the pain in the newborns in ITUN. The subjects are nine nursing technicians of the ITU of the Parenting School Januário Cicco in Natal-RN, engaged in direct assistance to newborns in the ITU, on the turn of the morning, which was prepared to participate in the search. The collection of the data was conducted through a structured interview with tree questions; through a non-participatory observation with a structured roadmap and were used to record and pass on call was also as a way of obtaining data. The start of the collection made after the assent of the Ethics Committee / UFRN in November, 2007. The speakings have been transcribed and data read extensively to obtain categories.The analysis of the content made in terms of Bardin. Emerged three main categories of significance: Perceptioning of pain in newborns; Caring for the newborns with pain; Registering the pain in the newborns. A nursing technicians identifies the pain in the newborns, for the most part, so empirical, using signs of behavioral or physiological changes in isolation, giving little emphasis to the environment and to respect that the newborns is inserted. It was found that the attitudes cited by subjects of the search before the newborns with pain, are for the most part non-pharmacological actions such as sucking nutrient not, a proper positioning and measures of comfort, however pharmacological actions have also been reported.These is also the absence of records of nursing records in the report of pain and actions to minimize them and, in records and for the passage of call. With this study we understand the role of the nursing technicians, and seek to contribute to subsidies for the practice of professionals involved in caring for this age group, and also in the search for a humane assistance to the newborns

Análise de polimorfismos dos genes da rota quinureniana em pacientes com meningite bacteriana.

Bacterial meningitis (BM) is still an important infectious disease causing death and disability. Invasive bacterial infections of the central nervous systems (CNS) generate some of the most powerful inflammatory responses known, which contributes to neuronal damage. The DNA microarray technology showed alterations in the kynurenine (KYN) pathway that is induced in BM and other diseases associated with inflammation, leading to brain injury. Our main aim was to search SNPs previously described in the KYN path enzymes to investigate a putative association of this SNPs with imbalanced in this pathway in patients with BM. The patients included in this study were 33 males and 24 females, with ages varying from 02 months to 68 years. SNPs were located inside of the domain conserved in KYNU, IDO, KATI and KATII. Primers were designed for analysis of SNPs already described by PIRA-PCR followed by RFLP. The analysis of KYNU+715G/A SNP found a heterozygous frequency of 0.033. We did not found the variant allele of SNP KYNU+693G/A, KATI+164T/C, KATII+650C/T and IDO+434T/G. Despite of previews studies showing the importance of KYN pathway we did not found one association of these SNPs analyzed with susceptibility or severity of MB in study population.

Associação entre polimorfismos em enzimas de reparo de DNA e ocorrência de meningite bacteriana.

Despite advances in vaccine development and therapy, bacterial meningitis (BM) remains a major cause of death and long-term neurological disabilities. As part of the host inflammatory response to the invading pathogen, factors such as reactive oxygen species are generated, which may damage DNA and trigger the overactivation of DNA repair mechanisms. It is conceivable that the individual susceptibility and outcome of BM may be in part determined by non synonymous polymorphisms that may alter the function of crucial BER DNA repair enzymes as PARP-1, OGG-1 and APE-1. These enzymes, in addition to their important DNA repair function, also perform role of inflammatory regulators. In this work was investigated the non synonymous SNPs APE-1 Asn148Glu, OGG-1 Ser326Cys,PARP-1 Val762Ala, PARP-1 Pro882Leu and PARP-1 Cys908Tyr in patients with bacterial meningitis (BM), chronic meningitis (CM), aseptic meningitis (AM) and not infected (controls). As results we found increased frequency of variant alleles of PARP-1 Val762Ala (P = 0.005) and APE-1 Asn148Glu (P=0.018) in BM patients, APE-1 Asn148Glu in AM patients (P = 0.012) and decrease in the frequency of the variant allele OGG-1 Ser326Cys in patients with CM (P = 0.013), regarding the allelic frequencies in the controls. A major incidence of individuals heterozygous and/ or polymorphic homozygous in BM for PARP-1 Val762Ala (P= 0.0399, OD 4.2, 95% IC 1.213 -14.545) and PARP-1 Val762Ala/ APE-1 Asn148Glu (P = 0.0238, OD 11.111, 95% IC 1.274 - 96.914) was observed related to what was expected in a not infected population. It was also observed a major incidence of combined SNPs in the BM patients compared with the control group (P=0.0281), giving evidences that SNPs can cause some susceptibility to the disease. This combined effect of SNPs seems to regulate the principal cytokines and other factors related to BM inflammatory response and point the importance of DNA repair not only to repair activity when DNA is damaged, but to others essential functions to human organism balance.

Ativação local da rota da quinurenina por moduladores inflamatórios durante a meningite bacteriana.

Activation of the kynurenine (KYN) pathway (KP) by modulators of immune system has been observed during several neurological diseases. Here we assessed the association of chemo-/cytokine levels with the concentration of KP metabolites in cerebrospinal fluid (CSF) and plasma samples from patients with bacterial meningitis (BM). All samples were collected from 42 patients diagnosed with acute bacterial meningitis (ABM), aseptic meningitis, tuberculous meningitis and patients without infection neurological disorders. CSF and plasma concentration of metabolites from the KP was assessed by high pressure liquid chromatography (HPLC) and cytokines and chemokines by Bio-plex 200 suspension array system. Concentrations of the KP metabolites KYN and kynurenic acid (KYNA) were significantly higher in CSF of patients with ABM compared to other groups. Tryptophan (TRP), anthranilic acid (AA), 3-hydroxykynurenine (3HK) and 3-hydroxyanthranilic acid (3HAA) did not show statistical significance, although some of them presented a good accumulation during ABM. The expression of TNF-alpha, IL-6, IL-1beta, IFN-gamma, IL-10, IL-1 receptor antagonist (IL-1Ra), MIP-1alpha, MIP-1beta, MCP-1 and G-CSF was about 100-fold higher in CSF from ABM patients than other infected groups. In all CSF and plasma samples, the concentration of IL-2, IL-12(p70), IL-4, IL-8 and GM-CSF was not significant. ABM still showed significant concentrations of IL-6, IL-10, IL-1Ra and MCP-1 in plasma samples. Based on the comparison of KP metabolites concentrations between plasma and CSF samples we conclude that the activation of the tryptophan pathway upon BM occurs within the brain. This increase in KP metabolites is most due to activation of the KP by molecules as IFN-gamma and TNF-alpha in response to infection.

Desigualdade regional da mortalidade neonatal no Brasil

In Brazil, despite the decline in infant mortality in recent decades it still has high rates going against recommended by WHO. Being the largest percentage of infant mortality rate composed of neonatal deaths. Objective: A study was conducted to analyze the spatial distribution of neonatal mortality and its correlation with the biological, socioeconomic and maternal and child health care in the Brazilian states in the period from 2006 to 2010. Method: The study made thematic maps and correlation (LISA) for verification of spatial dependence and multiple linear regression models. Results: Was found that there is no spatial autocorrelation for neonatal mortality in the Brazilian states (R = 0.002, p = 0.48). Most of variables were correlated (r> 0.3, p <0.05) with neonatal mortality, forming clusters in the North and Northeast, with the highest rates of teenage mothers, low household income per capita, lower prenatal appointments and beds of Neonatal Intensive Care Unit. The number of Neonatal UCI beds remained independent effect after regression analysis. Conclusion: The study concludes that regional inequalities in living conditions and especially the access to maternal and child health services contribute to the unequal distribution of neonatal mortality in Brazil

Aleitamento materno e alimentação complementar em crianças menores de um ano: chamada neonatal, RN, 2010

Avaliar os dados de aleitamento materno e alimentação complementar de crianças menores de um ano, do Rio Grande do Norte (RN), de acordo com o que é preconizado pelas políticas e ações de alimentação e nutrição. Métodos: Foi analisado o banco de dados da Chamada Neonatal do RN, pesquisa realizada pelo Ministério da Saúde em 2010. A amostra analisada foi de 837 pares mãe/filho que responderam ao questionário da pesquisa nos municípios investigados. Foram analisadas a prevalência de dados de aleitamento materno exclusivo (AME), na primeira hora de vida, parcial (AMP) e total (AMT), assim como dos alimentos ingeridos, pelas crianças, nas ultimas 24 horas anterior a entrevista. As frequências e médias foram feitas pelo comando Complex samples, no SPSS® 20.0, com IC95%. Foram estimadas as medianas de tempo de AME e AMT, assim como a mediana de tempo de introdução dos grupos de alimentos consumidos, em relação a idade da criança pelo método de probitos. Foi feita associação das probabilidades de tempo de AME e AMT com as variáveis sociodemográficas e de pré-natal (p<0.05). Resultados: Foram encontradas médias de idades de 5,28 ± 3,4 meses e 25,9 ± 6,4 anos, para crianças e mães, respectivamente. A prática de aleitamento na primeira hora de vida foi considerada boa (66,6%) e o percentual de AME (20%) razoável,segundo a Organização Mundial da Saúde, 2008. Mais da metade das crianças (55,1%) estavam em AMP. No total 60% estavam sendo amamentadas (AMT) ao final do primeiro ano de vida. O AME teve mediana de 63 dias e AMT de 358 dias. Estes dados não se diferenciaram muito entre a capital e os municípios do interior. A maioria das mães entrevistadas (73,8%) referiu ter tido orientação sobre aleitamento no pré- natal, tendo associação (p=0,03) com a probabilidade de tempo de AME, porém com pouca explicabilidade (R2= 0,011). Água ou chá, alimentos lácteos, frutas, legumes e verduras foram introduzidos precocemente com medianas menores que 180 dias. O aleitamento tende a diminuir e os alimentos tendem a aumentar o consumo de acordo com a idade da criança, com aumento exponencial do grupo calorias vazias . Conclusões: Conclui-se que mesmo com maioria das crianças amamentadas até um ano de vida, poucas estavam em AME e introduziram alimentos precocemente, não tendo resultado satisfatório frente ao preconizado pelas políticas públicas de alimentação e nutrição

Atenção à criança no período neonatal: chamada neonatal, Rio Grande do Norte, 2010

The neonatal period, which includes the first 27 da ys postpartum, is a vulnerability phase in child health, making it necessary for a greater mon itoring by health professional through actions that add value to the binomial mother/child and comprehensive care to the newborn. To this end, this study aimed to evaluate the care actions the neonato from the strategies recommended by the Ministry of Health. This is a cr oss-sectional study carried out from the database of the national survey of population base entitled "Call Neonatal: evaluation of prenatal care and to children younger than one year old in the North and Northeast regions". It used as the sample unit the mothers and children yo unger than 1-year-old, costal residents of Rio Grande do Norte, Natal, Brazil, who attended th e vaccination campaign on June 12, 2010 in nine municipality’s priority for the Pact to Red uce Infant and Neonatal Mortality. To compose the study variables were selected issues/ac tions regarding the neonatal period and socio-demographic factors, followed by a descriptiv e and inferential analysis. A sample of 837 mother/child pairs was obtained, being 57.6% in capital and 42.4% in the whole from the interiors, which was weighted to represent the muni cipalities of the State. It was predominated by mothers aged between 20-29 years, complete high school, not entitled to income transfer program and male children (51.2%). The frequency of the actions of the hospital ranged from 35% to 96% and those performed at the Basic Health Unit (BHU) from 57% to 91.2%. Most actions had an association with hospitals and publi c nature of the state capital (p<0.05). The results for most of the actions are recommended in the care programs and policies for children, and reveal the regional inequities in hea lth and the need for the involvement of services and professionals in search of comprehensi ve care for enabling better care through humanized practices during this increased vulnerabi lity period.

Reanimação neonatal: capacitação de profissionais de saúde com base na aprendizagem baseada em problemas

In whole world, including in Brazil, there is an absence of professionals who are able to be present at the moment of birth and give to the newborn the cares that they need to because either an absence of opportunity or inappropriate training to those professionals. This master´s thesis describes a construction and application of a neonatal resuscitation course that uses the problem based learning (PBL) methodology. The course has done in two meetings, one for the tutorial session, and another for practice session. The students were divided in groups of eight students each, under supervision of two teachers with experience in PBL methodology. The experience was considered successfully because there were students involvement and motivation. Some course aspects were rebuilt for its upgrading, like the correct use of methodology and building of custom educational material for students learning necessity. It suggests that the course can be used by the medical and nursing schools and perhaps other kind of health courses.

Segurança do paciente em terapia intensiva neonatal: identificação e análise de eventos adversos

The patient safety is a major concern in health services for its global dimension, as evidenced by the fragility of care processes that predispose an occurrence of adverse events. These events in a neonatal intensive care unit are considered serious and hazardous to lives of newborns. The present study aimed to identify and analyze adverse events in a neonatal intensive care unit based in Trigger Tool. It is an epidemiological, cross-sectional , exploratory, retrospective study with quantitative, descriptive and analytical approach, performed in 2015 at a school hospital. The sample was not probabilistic, involving 116 newborns who met the eligibility criteria. Data collection was performed by retrospective review of medical records, using a specific kind of "trigger" instrument, composed of sentinel events in neonatology, adapted from the American model used by the Vermont-Oxford Network. Data were analyzed using descriptive and inferential statistics. The chi-square test for linear trend was used to assess the associations between the variables of interest. The research received a favorable agreement from Ethics Committee of the Federal University of Rio Grande do Norte, under number 1055533, and Presentation Certificate for Ethics Assessment 43894515.6.0000.5537. The results show among investigated newborns, 110 experienced at least one adverse event during their stay, with a total of 391 medical records analyzed and rate of 3.37 events per patient. Prevailed the preterm newborns with low birth weight, from mother who had hypertensive diseases during pregnancy and urinary tract infection. The average hospitalization time was 25 days, associated with hospital-acquired infections events (p = 0.01). Among the identified adverse events stood out the events related to thermoregulation disorders (39.0%), with prevalence of hypothermia (26.0%), followed by health care-related infections (16.4%) and blood glucose disorders, hypoglycemia (9.00%) and hyperglycemia (6.64%). Most of these incidents were classified in categories E and F, which represents that there was damage small proportion. Due to these damages come from the care practice with newborn, 78% were classified as avoidable. There was statistically significant association between the variable birth weight with infections (p = 0.006) as well as peri/intraventricular bleeding (p = 0.02), hypoglycemia (p = 0.021), hyperglycemia (p = 0.001), hyperthermia (p = 0.39) and death (p=0,02). Gestational age was associated with seizures (p = 0.002), hyperglycemia (p=0.017) e hyperthermia (p=0.027). The security institution culture was reported by the health workers as intermediate, even though the number of adverse events found in only one unit of service indicates that there is much to be done. Thus the high rate of adverse events identified in the neonatal intensive care unit reinforces the necessity to elaborate specific preventive strategies for this risk environment.

Segurança do paciente em terapia intensiva neonatal: identificação e análise de eventos adversos

The patient safety is a major concern in health services for its global dimension, as evidenced by the fragility of care processes that predispose an occurrence of adverse events. These events in a neonatal intensive care unit are considered serious and hazardous to lives of newborns. The present study aimed to identify and analyze adverse events in a neonatal intensive care unit based in Trigger Tool. It is an epidemiological, cross-sectional , exploratory, retrospective study with quantitative, descriptive and analytical approach, performed in 2015 at a school hospital. The sample was not probabilistic, involving 116 newborns who met the eligibility criteria. Data collection was performed by retrospective review of medical records, using a specific kind of "trigger" instrument, composed of sentinel events in neonatology, adapted from the American model used by the Vermont-Oxford Network. Data were analyzed using descriptive and inferential statistics. The chi-square test for linear trend was used to assess the associations between the variables of interest. The research received a favorable agreement from Ethics Committee of the Federal University of Rio Grande do Norte, under number 1055533, and Presentation Certificate for Ethics Assessment 43894515.6.0000.5537. The results show among investigated newborns, 110 experienced at least one adverse event during their stay, with a total of 391 medical records analyzed and rate of 3.37 events per patient. Prevailed the preterm newborns with low birth weight, from mother who had hypertensive diseases during pregnancy and urinary tract infection. The average hospitalization time was 25 days, associated with hospital-acquired infections events (p = 0.01). Among the identified adverse events stood out the events related to thermoregulation disorders (39.0%), with prevalence of hypothermia (26.0%), followed by health care-related infections (16.4%) and blood glucose disorders, hypoglycemia (9.00%) and hyperglycemia (6.64%). Most of these incidents were classified in categories E and F, which represents that there was damage small proportion. Due to these damages come from the care practice with newborn, 78% were classified as avoidable. There was statistically significant association between the variable birth weight with infections (p = 0.006) as well as peri/intraventricular bleeding (p = 0.02), hypoglycemia (p = 0.021), hyperglycemia (p = 0.001), hyperthermia (p = 0.39) and death (p=0,02). Gestational age was associated with seizures (p = 0.002), hyperglycemia (p=0.017) e hyperthermia (p=0.027). The security institution culture was reported by the health workers as intermediate, even though the number of adverse events found in only one unit of service indicates that there is much to be done. Thus the high rate of adverse events identified in the neonatal intensive care unit reinforces the necessity to elaborate specific preventive strategies for this risk environment.

Length of stay in a neonatal intensive care unit and its association with low rates of exclusive breastfeeding in very low birth weight infants

OBJECTIVE: To identify the inpatient maternal and neonatal factors associated to the weaning of very low birth weight (VLBW) infants. METHODS: One hundred nineteen VLBW (<1500 g) infants were monitored from July 2005 through August 2006, from birth to the first ambulatory visit after maternity discharge. This maternity unit uses the Kangaroo Method and the Baby Friendly Hospital Initiative. Out of 119 VLBW infants monitored until discharge, 88 (75%) returned to the facility, 22 (25%) were on exclusive breastfeeding (EB), and 66 (75%) were weaned (partial breastfeeding or formula feeding). RESULTS: Univariate analysis found an association between weaning and lower birth weight, longer stays in the neonatal intensive care unit (NICU), and longer hospitalization times, in addition to more prolonged enteral feeding and birth weight recovery period. Logistic regression showed length of NICU stay as being the main determinant of weaning. CONCLUSION: The negative repercussion on EB of an extended stay in the NICU is a significant challenge for health professionals to provide more adequate nutrition to VLBW infants.

Length of stay in a neonatal intensive care unit and its association with low rates of exclusive breastfeeding in very low birth weight infants

OBJECTIVE: To identify the inpatient maternal and neonatal factors associated to the weaning of very low birth weight (VLBW) infants. METHODS: One hundred nineteen VLBW (<1500 g) infants were monitored from July 2005 through August 2006, from birth to the first ambulatory visit after maternity discharge. This maternity unit uses the Kangaroo Method and the Baby Friendly Hospital Initiative. Out of 119 VLBW infants monitored until discharge, 88 (75%) returned to the facility, 22 (25%) were on exclusive breastfeeding (EB), and 66 (75%) were weaned (partial breastfeeding or formula feeding). RESULTS: Univariate analysis found an association between weaning and lower birth weight, longer stays in the neonatal intensive care unit (NICU), and longer hospitalization times, in addition to more prolonged enteral feeding and birth weight recovery period. Logistic regression showed length of NICU stay as being the main determinant of weaning. CONCLUSION: The negative repercussion on EB of an extended stay in the NICU is a significant challenge for health professionals to provide more adequate nutrition to VLBW infants.