Oxidative stress as common trait of endothelial dysfunction in chorionic arteries from fetuses with IUGR and LGA.

INTRODUCTION: Fetal macrosomia and intrauterine growth restriction (IUGR) associate with increased morbidity in the neonate. Placental vascular relaxation is impaired in fetal macrosomia, as well as in IUGR, and this could result from increased oxidative stress present in both conditions. We determined the role of pro- and anti-oxidants on NOS dependent relaxation in placental chorionic arteries from pregnancies with LGA babies from overweight and/or obese mothers (LOOM) and IUGR fetuses from normal BMI women. METHODS: Chorionic arteries were mounted in a wire-myograph, where responses to the NOS-dependent agent CGRP in presence or absence of the antioxidant N-acetyl cysteine (NAC), the pro-oxidant SIN-1, the SOD inhibitor DDC, and the GPx inhibitor MS were determined. Additionally the presence of pro- and antioxidant enzymes (NOX-4, SOD-1, SOD-2 and GPx-1) and eNOS in chorionic and umbilical vessels were addressed by immunohistochemistry. RESULTS: Maximal CGRP-induced relaxation was comparable to controls but presented a reduced potency in chorionic arteries from LOOM placentae, whilst in IUGR vessels both maximal response and potency were reduced. NAC increased maximal relaxation in controls, IUGR and LOOM arteries, whilst SIN-1 completely abolished the CGRP-induced relaxation only in IUGR and LOOM samples, the later effect was paralleled by SOD or GPx inhibition. These responses associated with the presence of NOX-4, SOD-1 and GPx-1 in the endothelium and vascular wall of chorionic and umbilical arteries in the different groups studied. DISCUSSION: These data suggest that NOS dependent relaxation in placental vessels from IUGR and LOOM pregnancies present a higher sensitivity to oxidative stress.

Small-angle X-ray scattering and in silico modeling approaches for the accurate functional annotation of an LysR-type transcriptional regulator.

Xylella fastidiosa is a xylem-limited, Gram-negative phytopathogen responsible for economically relevant crop diseases. Its genome was thus sequenced in an effort to characterize and understand its metabolism and pathogenic mechanisms. However, the assignment of the proper functions to the identified open reading frames (ORFs) of this pathogen was impaired due to a lack of sequence similarity in the databases. In the present work, we used small-angle X-ray scattering and in silico modeling approaches to characterize and assign a function to a predicted LysR-type transcriptional regulator in the X. fastidiosa (XfLysRL) genome. XfLysRL was predicted to be a homologue of BenM, which is a transcriptional regulator involved in the degradation pathway of aromatic compounds. Further functional assays confirmed the structural prediction because we observed that XfLysRL interacts with benzoate and cis,cis-muconic acid (also known as 2E,4E-hexa-2,4-dienedioic acid; hereafter named muconate), both of which are co-factors of BenM. In addition, we showed that the XfLysRL protein is differentially expressed during the different stages of X. fastidiosa biofilm formation and planktonic cell growth, which indicates that its expression responds to a cellular signal that is likely related to the aromatic compound degradation pathway. The assignment of the proper function to a protein is a key step toward understanding the cellular metabolic pathways and pathogenic mechanisms. In the context of X. fastidiosa, the characterization of the predicted ORFs may lead to a better understanding of the cellular pathways that are linked to its bacterial pathogenicity.

Prevalence of Chagas disease in pregnant women and incidence of congenital transmission in Santa Cruz de la Sierra, Bolivia.

Congenital transmission of Chagas disease stand out as a major public health problem since the vector control was performed in all endemic areas and has shown its effectiveness. An epidemiological study was performed in three maternity hospitals of the city of Santa Cruz de la Sierra, Bolivia from 2006 to 2008. The serological screening for Trypanosoma cruzi infection was carried out in 15,767 pregnant women. Chagas infection was detected in 3725 women (23.6%), who gave birth to 125 newborns infected by T. cruzi at birth, representing an incidence of 790 per 100,000 births during a period of 16 months and a vertical transmission rate by 3.4%. There was a significant difference between hospitals that might be explained by socio-economic origins of mothers and diagnostic constraints.

OS059. Blockade of the bradykinin B2 receptor in early pregnancy reduces fetal growth and trophoblast invasion in guinea-pigs.

INTRODUCTION: Research in preeclampsia (PE) is hampered by the difficulty of sampling the placental bed in early pregnancies followed to delivery to be defined as normal or preeclamptic. Thus, animal models contribute to the understanding of its physiopathology. The guinea-pig shares with humans extensive vascular remodelling, a hemomonochorial placenta [1] and a vasodilator and angiogenic utero-placental repertoire [2]. In pregnancy it expresses bradykinin (BK) B1R and B2R receptors in cells related to invasion, angiogenesis and vasodilatation. In addition, in HTR-8/SVneo cells, BK induces a B2R-mediated increase in migration and invasion [3]. OBJECTIVES: To test whether blocking the B2R with a rodent-selective non-peptide antagonist Bradyzide (BDZ) from days 20 to 34 of an ≈65 day gestation - period of maximal trophoblast invasion and placental development - induces PE-like morphological and functional alterations. METHODS: Virgin Pirbright guinea-pigs (Cavia Porcellus) after mating and echographic confirmation of pregnancy, were allocated in gestational day 20 to to subcutaneous implantation of Alzet pumps that delivered for 14 days saline (Control; n=7), BDZ0,875mg/kg/day (BDZ0,87; n=6) and BDZ 1,2mg/kg/day (BDZ1,2; n=7). Systolic pressure was acquired in the right hindlimb with a Power Lab 8 SP and analyzed with Labchart at day 34. On that day dams were sacrificed, vesical urine was extracted for protein determination, the fetuses and corresponding placentas weighed and the cephalo-caudal length measured. The placentas were studied by HE and immunohistochemistry for cytokeratin to identify trophoblasts. Results are expressed as means±SE. Statistical analysis was performed with Graphpad Prism 5.1, using one-way ANOVA, the recommended post hoc tests and χ2 test. RESULTS: Maternal systolic pressure tended to increase in BDZ0,875 and BDZ1,2 versus controls (63±567±6 versus 56±2,mm Hg respectively; NS). Proteinuria was not observed in any group. The number of viable fetuses tended to be reduced in both BZD treated groups (NS). The fetal weight (% maternal weight) was reduced in animals treated with BDZ0,875 and BDZ1,2 (042±002 and 045±001 versus 054±0,04 in controls; p (P<0.01 and 0.05 respectively). The cephalo-caudal length was reduced in BDZ0,875 and BDZ1,2 (373±24 and 371±07 versus 422±21mm in controls; P<0.01). No differences were observed in placental weight. Spiral arteries surrounded by trophoblasts (%) were reduced in BDZ 0875 and BDZ1,2 versus controls (63 and 66.6 versus 100%, P=0.02). Invaded spiral arteries (%) were also reduced in the treated groups (80 and 43 versus 100%; P<0.002) No differences were observed in the depth of trophoblast decidual invasion. CONCLUSION: This study demonstrates that blocking the B2R in early pregnancy impairs fetal growth and transformation of the spiral arteries, and supports the role of the B2R in the local physiological adaptation. Further studies are needed to elucidate whether the early impairment translates to hypertension and proteinuria in the last third of pregnancy; if so, the guinea-pig would provide a model to understand the physiopathology of the syndrome. Study financed by Fondecyt 1080228.

Characterization of an oxidative stress response regulator, homologous to Escherichia coli OxyR, from the phytopathogen Xylella fastidiosa.

The OxyR oxidative stress transcriptional regulator is a DNA-binding protein that belongs to the LysR-type transcriptional regulators (LTTR) family. It has the ability to sense oxidative species inside the cell and to trigger the cell's response, activating the transcription of genes involved in scavenging oxidative species. In the present study, we have overexpressed, purified and characterized the predicted OxyR homologue (orf xf1273) of the phytopathogen Xylella fastidiosa. This bacterium is the causal agent of citrus variegated chlorosis (CVC) disease caused by the 9a5c strain, resulting in economic and social losses. The secondary structure of the recombinant protein was analyzed by circular dichroism. Gel filtration showed that XfoxyR is a dimer in solution. Gel shift assays indicated that it does bind to its own predicted promoter under in vitro conditions. However, considering our control experiment we cannot state that this interaction occurs in vivo. Functional complementation assays indicated that xfoxyR is able to restore the oxidative stress response in an oxyr knockout Escherichia coli strain. These results show that the predicted orfxf1273 codes for a transcriptional regulator, homologous to E. coli OxyR, involved in the oxidative stress response. This may be important for X. fastidiosa to overcome the defense mechanisms of its host during the infection and colonization processes.

Overexpression and purification of PWL2D, a mutant of the effector protein PWL2 from Magnaporthe grisea.

The rice blast disease caused by the ascomycete Magnaporthe grisea continues to cause a tremendous impact in rice (Oryza sativa) cultures around the world. Elucidating the molecular basis of the fungus interactions with its host might help increase the general understanding of the pathogen-host relationship. At the moment of invasion, the fungus secretes effectors that modify host defenses and cellular processes as they successively invade living rice cells. PWL2, an effector protein, is a known AVR (avirulence) gene product. The PWL2 gene prevents the fungus from infecting weeping lovegrass (Eragrostis curvula). In this study, we identified a PWL2 allele gene (which we termed PWL2D) in a strain of M. grisea. The sequence of PWL2D has only two bases different from that of PWL2, producing alterations in residue 90 and residue 142. However, the alteration of residue 90 (from D(90) to N(90)) is critical to gene function. Here, we cloned the gene PWL2D in a pET System vector, expressed the gene product in Escherichia coli and evaluated by spectroscopic techniques some aspects of the PWL2D structure. While TRX-tagged PWL2D is prone to aggregation, the solubility of PWL2D is improved when it is overexpressed without its original signal peptide. Expression and purification procedures for these constructs are described. Finally, we found out that the protein seems to be an intrinsically disordered protein. Results from these studies will facilitate structural analysis of PWL2D and might contribute to understanding the gene's function and of fungal/plant interactions.

[Screening for congenital infection by Trypanosoma cruzi in France]. / Le dépistage de l'infection congénitale par Trypanosoma cruzi en France métropolitaine.

The importance of congenital transmission of Chagas' disease increases with its emergence in communities infected with Trypanosoma cruzi, but where vector transmission has never existed or is fully controlled through vector control campaigns. In both endemic and non-endemic areas, the rates of mother-to-child transmission (MTCT) could be the same, by 5%, generating a constant source of new cases of the disease. Risk factors for vertical transmission are not fully elucidated, but the effectiveness of the adaptive immune response and the genetic susceptibility of both the mother and the child are suspected. Besides the risk of miscarriage or premature birth, neonatal infection by T. cruzi causes an acute form of Chagas disease, which may be accompanied by a severe infectious syndrome that can causes death if not treated early. This form of the disease is a real public health priority because it is frequent, severe, identifiable and curable. Indeed, almost all newborns diagnosed and treated before the end of their first year of life will be definitely cured. In all non-endemic areas, detection of cases of congenital Chagas disease is hampered by a very low prevalence of the disease in the general population of pregnant women, the lack of symptoms in most infected women and the disregard of these problems from health personnel in charge of monitoring pregnancy. Secondary prevention firstly consists in identifying infected women (with history of exposure and positive serology for Chagas disease) and secondly to look for the parasite in newborns from infected mothers. No primary prevention is indeed possible during pregnancy, since the only two drugs are toxic and possibly teratogenic. However, after birth, treatment could be offered to all infected women in order to prevent late complications of the disease and to make an attempt at breaking the chain of MTCT in future pregnancies.

Níveis glicêmicos de pacientes diabéticos segundo estudo comparativo entre duas técnicas. / Blood glucose levels of diabetic patients: comparative study of two measuring techniques

O estudo teve como objetivo identificar e comparar os resultados de glicemia de pacientes adultos com diabetes tipo 2 por meio de duas técnicas, laboratorial e capilar. Estudo comparativo foi desenvolvido no período de setembro a dezembro de 2007. A população foi constituída de 36 pacientes adultos residentes no município de Planaltina do Paraná, PR. A pesquisa foi realizada no laboratório de análises clínicas municipal. Para o exame laboratorial foi usado o sangue venoso e para o teste capilar o sangue da ponta do dedo. A maioria dos pacientes (63,9%) era do gênero feminino, idade média de 62,4 anos. Quanto à escolaridade, 41,6% dos pacientes possuíam menos de 8 anos de estudo. Em relação aos níveis glicêmicos, o p-valor do teste Kappa foi significativo em todas as categorias (p< 0,001, p=0,005 e p=0,001), mostrando concordância. Os dados indicam elevada precisão e acurácia dos resultados de glicemia obtidos por meio do glicosímetro, quando comparada ao teste padrão obtido em laboratório.

The goal of this research was to measure the blood glucose levels of type 2 diabetic adult patients and compare the results obtained by two techniques, the standard laboratory (venous) and portable glucometer (capillary) tests. A comparative longitudinal study was conducted from September to December 2007. The study population consisted of 36 resident adult patients in the town of Planaltina do Paraná in upstate Paraná, Brazil. The tests were performed in the local health authority clinical laboratory. For the lab test, venous blood from the arm was used, and for the glucometer test, capillary blood from the fingertip. Most of the patients (63.9%) were women and the average age was 62.4 years. Regarding education, 41.6% of the patients had studied for less than 8 years. Comparing the glucose measurements, the results of the Kappa test were significant in all categories (p<0.001, p=0.005 and p=0.001), showing very good agreement between the methods. The findings indicate a high level of precision and accuracy for the blood glucose readings obtained with the glucometer, compared with the standard test carried out in the laboratory.

Congenital Chagas disease: diagnostic and clinical aspects in an area without vectorial transmission, Bermejo, Bolivia.

The authors carried out a 1-year study of a population of pregnant women delivering at Bermejo hospital, South Bolivia. In this area, vectorial transmission of Trypanosoma cruzi is negligible and women infect themselves during displacements in close endemic areas. The prevalence of T. cruzi in 508 pregnant women, diagnosed by several serological tests, was 33.9%. In eight infants, we observed T. cruzi in the umbilical cord (congenital transmission rate of 5.2%). The means of birth weights, lengths and hemoglobin rates were similar in the children from both seronegative and seropositive women, and in children infected or not by T. cruzi. This study could confirm a less severity of the congenital disease of Chagas in the absence of re-infestation of the mother during pregnancy. Serological screening of pregnant women by rapid diagnostic tests and examination of babies born from seropositive mothers by microhematocrit method at birth is a suitable strategy to detect and prevent congenital Chagas disease in non-endemic areas.

Hip axis length variation: its correlation with anthropometric measurements in women from three ethnic groups.

UNLABELLED: We compared hip axis length (HAL) in 157 non-Hispanic white women, 292 African-American women, and 210 Mexican-American women. After adjusting for maximal hip girth, there were no residual differences in HAL by ethnicity. Differences in hip fracture risk seen between these groups cannot be explained by ethnic differences in HAL. INTRODUCTION: Hip axis length (HAL) has been reported to be an independent predictor of hip fracture. Significant ethnic differences in HAL have been noted, but no direct comparison has been made between African-American, Mexican-American, and non-Hispanic white women using the same protocol. METHODS: We compared 157 non-Hispanic white women from the Rancho Bernardo Study, 292 women from the Health Assessment Study of African-American Women, and 210 women from the Skeletal Health of Mexican-American Women Project. A standardized questionnaire was used to obtain medical history; height, weight, waist girth, and hip girth were measured; and percentage body fat and HAL were obtained using dual energy X-ray absorptiometry. All HAL comparisons were adjusted for maximum hip girth to control for differences in size magnification by fan-beam absorptiometry. RESULTS: Though there were ethnic differences in the unadjusted HAL measurement, after adjusting for hip circumference, there were no residual differences in HAL with regard to ethnicity: 10.7 cm in Mexican-American women vs. 10.8 in non-Hispanic white women and African-American women (p = 0.61). CONCLUSIONS: There were no ethnic differences in HAL in women from the three ethnic groups. Differences in fracture risk among these groups cannot be explained by ethnic differences in HAL.

Risk factors and consequences of congenital Chagas disease in Yacuiba, south Bolivia.

OBJECTIVE: To determine the risk factors of congenital Chagas disease and the consequences of the disease in newborns. METHODS: Study of 2712 pregnant women and 2742 newborns in Yacuiba, south Bolivia. Chagas infection was determined serologically in mothers and parasitologically in newborns. Consequences of congenital Chagas disease were assessed clinically. RESULTS: The prevalence of Chagas disease in pregnant women was 42.2%. Congenital transmission was estimated at 6% of infected mothers leading to an incidence rate of 2.6% among newborns. Main risk factors of congenital transmission were mothers' seropositivity and maternal Trypanosoma cruzi parasitaemia. Parity was higher in infected than in non-infected mothers, but it was not associated with the risk of congenital transmission. The rate of congenital infection was significantly higher in newborns from multiple pregnancies than in singletons. However, we did not observe statistically significant consequences of Chagas disease in newborns from single pregnancies or among twins. CONCLUSIONS: The main risk factors for congenital transmission were infection and parasitaemia of mothers. Consequences of the disease seemed mild in newborns from single pregnancies and perhaps more important in multiple births.

Evidence of congenital transmission of Trypanosoma cruzi in a vector-free area of Bolivia.

A seroprevalence survey of Trypanosoma cruzi was carried out in two areas of South Bolivia. Triatoma infestans, the main vector of Tryp. cruzi, was abundant in the first area, but absent in the second one. Titration of Tryp. cruzi antibodies was carried out in children aged 6-24 months and their mothers. The seroprevalence of Chagas' disease was significantly higher in the area with the vector, but also high in the second area. Chagas' infection in children under 2 years old could be linked to congenital transmission of parasites during pregnancy and/or delivery, despite active vector control in both areas.

Hypercalciuria in children with renal glycosuria: evidence of dual renal tubular reabsorptive defects.

During the past 5 years, we have identified idiopathic hypercalciuria in five of seven patients referred for evaluation of renal glycosuria between 1985 and 1991. The children, all boys, ranged in age from 6 to 12 years. Endocrine function was normal, and none of the patients had hyperparathyroidism, hypercalcemia, renal tubular acidosis, or other secondary causes of hypercalciuria. The calcium/creatinine ratio in a fasting urine specimen was elevated in all five children who had hypercalciuria, with a mean value (+/- SD) of 0.34 +/- 0.06 (normal, < 0.2). In one child who had renal colic with spontaneous passage of gravel-like material, the idiopathic hypercalciuria persisted after 1 week on a diet containing 2000 mg of sodium and 300 mg of calcium. On the basis of studies that examined the site along the nephron responsible for hypercalciuria in rats with streptozocin-induced diabetes, we speculate that in children with renal glycosuria, there is defective reabsorption of glucose and calcium in the straight portion of the proximal tubule or in the collecting duct. It is likely that a similar mechanism accounts for the idiopathic hypercalciuria in children with diabetes mellitus.

Therapeutic risk-assessment model for identifying patients with adverse drug reactions.

The association between factors that place patients at risk for adverse drug reactions (ADRs) and the occurrence of ADRs was examined, and a therapeutic risk-assessment model was developed. Theoretical risk factors for ADRs to digoxin and theophylline were identified through the literature by researchers at a private tertiary-care hospital. Data were then collected from two groups of 67 patient charts each during a 15-month period. One group of charts represented patients who had experienced an ADR to digoxin or theophylline. The other group represented matched control patients who had not experienced an ADR to either drug. ICD-9-CM (International Classification of Diseases, 9th Revision, Clinical Modifications) ADR codes were assigned by medical records department personnel, and the ADRs were verified by using the Naranjo algorithm. Seven risk factors for each drug were found to be significantly associated with ADRs. A serum digoxin concentration greater than 2.5 ng/mL and elevated blood urea nitrogen were the two best predictors of an ADR to digoxin. The probability of experiencing an ADR to digoxin was 94.1% for a patient with both of these risk factors. A serum theophylline concentration greater than 25 micrograms/mL was the greatest predictor of an ADR to theophylline; the probability of experiencing an ADR to theophylline was 85.2% if a patient had that risk factor. The sensitivity and specificity of the therapeutic risk-assessment model were 92.9% and 61.8%, respectively, for digoxin and 95.8% and 84.0%, respectively, for theophylline. Several laboratory-based screening criteria demonstrated an ability to predict ADRs to digoxin and theophylline.