Novel Method Probe-based Real-Time PCR to Detect 2 Single-Nucleotide Polymorphisms Close to Each Other: HFE Hemochromatosis Gene Model.

Hereditary hemochromatosis is known as the most common genetic disorder among individuals of European genetic background. It is possible to find 2 mutations closely placed in the HFE gene (H63D and S65C) and this proximity can cause errors when genotyped by real-time polymerase chain reaction (PCR) genotyping assay. The aim of this study was to develop a hydrolysis probe-based PCR assay for detection of the H63D and S65C mutations without interference from on each other. Herein the study involved the standardization of an improvement of the real-time PCR 5' nuclease assay to detect the desired mutations close placed using a same probe system. The assay analytical properties performances were tested, including the primers selectivity and detection limits. Also, the interexaminer reproducibility and repeatability of assay were estimated in 30 blood samples. Others 153 results of samples were compared with reference method (PCR_RFLP) and the accordance of the results evaluated by Fleiss' κ method. The results of variation of interexaminer reproducibility and repeatability of assay were not statistically relevant (P<0.001). The comparison between the 2 methods by Fleiss' κ analysis showed that 5' nuclease assay identified the H63D and S65C haplotype as well as the reference method in all 153 tested samples. Our results showed that novel method probe-based real-time PCR were capable to detect 2 adjacent polymorphisms without errors in genotyping.

Epidemiologic profile, survival, and maternal prognosis factors among women at an obstetric intensive care unit.

OBJECTIVE: To identify the epidemiologic profile, maternal survival, and prognosis factors that might affect survival rates in the obstetric intensive care unit (ICU). METHODS: A prospective cohort study was conducted between January 2007 and February 2009 in a tertiary referral ICU, Belo Horizonte, Brazil. Critical patients during pregnancy and puerperium were followed from admission until discharge or death. Maternal survival was assessed in association with the cause of ICU admission, grouped into direct or indirect obstetric causes, by Kaplan-Meier curves and log-rank tests. RESULTS: Among 298 patients admitted to the ICU during the study period, mortality was 4.7% (n=14). Hypertensive disorders (46.0%), hemorrhage (15.9%), sepsis (14.2%), and heart disease (5.7%) were the main causes of admission. Half of the patients who died were admitted for direct obstetric reasons (n=7). Survival was statistically linked to the cause of admission: most survivors were admitted for a direct obstetric cause (75.5%; P=0.044). Maternal survival rates of patients admitted for indirect obstetric causes were lower than those admitted for direct obstetric causes (27.8 and 19.6 days, respectively; P=0.019). CONCLUSION: The main cause of admission was a decisive factor for maternal survival in the obstetric ICU. Direct obstetric complications had a better prognosis.

Variation of endothelial function during the menstrual cycle evaluated by flow-mediated dilatation of brachial artery.

OBJECTIVE: It is well defined that ovarian steroids play a role in the cardiovascular system, acting as vasoactive compounds. The aim of this study is to assess the endothelial function during the menstrual cycle using flow-mediated dilation of the brachial artery. METHODS: A total of 21 healthy premenopausal women, with regular menstrual cycles, were included in this observational, longitudinal, and prospective study. The endothelium function was assessed by ultrasound during four phases of the menstrual cycle: early follicular phase (EFP), late follicular phase (LFP), early luteal phase (ELP) and late luteal phase (LLP). RESULTS: We observed a significant variation among the vasodilatation response during the menstrual cycle phases (P<0.001). The result was higher during LFP than during ELP (P<0.001) or LLP (P<0.001). Late luteal phase had higher response than observed during ELP (P=0.003) and EFP was higher than LLP (P=0.003). There were no significant differences between the results observed during EFP and LFP (P=1.0), or EFP and ELP (P=0.137). CONCLUSION: Our results suggest that the ovarian steroids may play an important role in modulating endothelial function.

Presymptomatic prediction of preeclampsia with angiogenic factors, in high risk pregnant women.

INTRODUCTION: The aim of this study was to investigate the value of placental growth factor (PLGF), soluble fms-like tyrosine kinase-1 (sFlt-1) and sFlt-1/PLGF ratio, in predicting symptomatic preeclampsia (PE). METHODS: A prospective longitudinal study was carried out on 71 high risk preeclamptic women cohort. All of them had normal blood pressure level (≤140/90 mmHg) at the time of enrolment, 26.8 ± 1.5 weeks. Maternal blood was collected and plasma was stored in a freezer at -80 °C. PE was defined according to the National High Blood Pressure Education Program Working Group Criteria. Accuracy of angiogenic factors in predicting PE was evaluated using Receiver-operating characteristics. RESULTS: Maternal plasma concentrations of PLGF and sFlt-1 were able to predict PE (0.90, p < 001; 0.78, p = 0.003, area under the curve, respectively) but the sFlt-1/PLGF ratio presented the best prediction potential over the others (0.95, area under the curve, p < 0.001). CONCLUSION: All angiogenesis factors were effective biomarkers in predicting PE during the second trimester, before the clinical onset of PE.

Echocardiographic findings of congenital cardiopathies among fetuses of diabetic pregnant women and their relationship with plasma fructosamine levels.

OBJECTIVE: To study the occurrence of congenital cardiopathies at echocardiography (CCE) in fetuses whose mothers had preexisting diabetes mellitus (PGDM) and to study the potential of using fructosamine level as a late marker (beyond the first trimester) for CCE. METHODS: A register study covering 91 pregnant women that underwent routine fetal echocardiography ordered due to PGDM. The first dosage of plasma fructosamine found in 65 medical records was analyzed during prenatal care (20.4 ± 8.0 weeks of gestation). The presence or absence of structural or functional CCE was associated with fructosamine levels by logistic regression. We assessed the effect modification odds ratio by maternal age and insulin usage. RESULTS: Thirty-four fetuses (52.3% of 65 fetuses) presented CCE. Twenty of them had functional CCE and 14 presented structural CCE. The mean maternal plasma fructosamine level was higher among pregnant women whose fetuses presented CCE than in those whose fetuses did not (2.86 ± 0.73 mmol/l, 2.22 ± 0.54 mmol/l, respectively, p < 0.0001). Crude OR for CCE and abnormal plasma fructosamine (>2.68 mmol/l) was 9.6 (2.8-33.7, 95% CI, p < 0.0001). Adjusted OR by maternal age and insulin usage was 10.9 (2.7-45.2, 95% CI p < 0.0001). CONCLUSIONS: An abnormal plasma fructosamine level increases the chances of CCE occurring among referral pregnant women with PGDM.

Prognostic factors in prenatally-detected posterior urethral valves: a multivariate analysis.

To identify prognostic factors associated with chronic renal insufficiency in children with posterior urethral valves (PUV), 22 children with PUV were submitted to a systematic protocol and prospectively followed. Prognostic factors associated with fetal echography and clinical and laboratory findings were studied on admission. Median follow-up was 76 months. The analysis was conducted in two steps: in univariate analysis, variables associated with adverse outcome were identified by the Kaplan-Meier method. The variables that were significantly associated with adverse outcome were then included in a multivariate analysis using the Cox model. Eleven patients (50%) developed chronic renal failure (CRF) during follow-up. After adjustment by the multivariate model, four factors were identified as independent predictors of adverse outcome: oligohydramnios (relative risk [RR] = 10.6, 95% CI = 2.7 - 77, P = 0.02), ventilatory support (RR = 6, 95% CI = 2 - 24, P = 0.01), urea higher than 40 mg/dl (RR = 3.7, 95% CI = 0.92 - 15.0, P = 0.06), and bilateral vesicoureteral reflux (VUR) (RR = 6.1, 95% CI = 1.25 - 30, P = 0.02). On the other hand, the presence of unilateral VUR was a protective factor against the development of CRF or death during follow-up (RR = 0.92, 95% CI = 0.87 - 0.98, P = 0.05).

Curso clínico da válvula de uretra posterior detectada intra-útero: seguimento a longo prazo / Clinical course of posterior urethra detected intra-uterus : long follow-up

Com o objetivo de avaliar o curso clínico da VUP identificada na vida fetal, foram estudadas 15 crianças prospectivamente com o objetivo de avaliar o curso clínico da válvula de uretra posterior (VUP). Pacientes com hidronefrose detectada no pré-natal foram avaliados no período pós-natal, submetidos a um protocolo sistemático e seguidos prospectivamente. Após início daquimioprofilaxia, a avaliaçäo de imagens consistiu de ultrasonografia (US), uretrocistografia miccional (UCM) e cintilografia estática por DMSA. Nos casos confirmados de VUP, as crianças foram submetidas a vesicostomia e seguidas com avaliaçöes clínicas e laboratoriais periódicas. Quinze meninos foram identificados como portadores de VUP entre 1989 e 1998. A média de idade do diagnóstico pré-natal foi de 30 meses. O tempo médio de seguimento foi de 40 meses. Das 15 crianças acompanhadas, 7 (47 porcento) evoluíram com isuficiência renal crônica (IRC) e 6 (40 porcento) faleceram. O óbito esteve associado à perda da funçäo renal (RR = 7, p=0,001). Apesar do diagnóstico precoce, a VUP continua a representar um desafio para a área da nefrologia pediátrica, e os riscos de perda da funçäo renal e de óbito se mantêm elevado