Normative spatiotemporal fetal brain maturation with satisfactory development at 2 years.

Maturation of the human fetal brain should follow precisely scheduled structural growth and folding of the cerebral cortex for optimal postnatal function1. We present a normative digital atlas of fetal brain maturation based on a prospective international cohort of healthy pregnant women2, selected using World Health Organization recommendations for growth standards3. Their fetuses were accurately dated in the first trimester, with satisfactory growth and neurodevelopment from early pregnancy to 2 years of age4,5. The atlas was produced using 1,059 optimal quality, three-dimensional ultrasound brain volumes from 899 of the fetuses and an automated analysis pipeline6-8. The atlas corresponds structurally to published magnetic resonance images9, but with finer anatomical details in deep grey matter. The between-study site variability represented less than 8.0% of the total variance of all brain measures, supporting pooling data from the eight study sites to produce patterns of normative maturation. We have thereby generated an average representation of each cerebral hemisphere between 14 and 31 weeks' gestation with quantification of intracranial volume variability and growth patterns. Emergent asymmetries were detectable from as early as 14 weeks, with peak asymmetries in regions associated with language development and functional lateralization between 20 and 26 weeks' gestation. These patterns were validated in 1,487 three-dimensional brain volumes from 1,295 different fetuses in the same cohort. We provide a unique spatiotemporal benchmark of fetal brain maturation from a large cohort with normative postnatal growth and neurodevelopment.

Pregnancy outcomes and vaccine effectiveness during the period of omicron as the variant of concern, INTERCOVID-2022: a multinational, observational study.

BACKGROUND: In 2021, we showed an increased risk associated with COVID-19 in pregnancy. Since then, the SARS-CoV-2 virus has undergone genetic mutations. We aimed to examine the effects on maternal and perinatal outcomes of COVID-19 during pregnancy, and evaluate vaccine effectiveness, when omicron (B.1.1.529) was the variant of concern. METHODS: INTERCOVID-2022 is a large, prospective, observational study, involving 41 hospitals across 18 countries. Each woman with real-time PCR or rapid test, laboratory-confirmed COVID-19 in pregnancy was compared with two unmatched women without a COVID-19 diagnosis who were recruited concomitantly and consecutively in pregnancy or at delivery. Mother and neonate dyads were followed until hospital discharge. Primary outcomes were maternal morbidity and mortality index (MMMI), severe neonatal morbidity index (SNMI), and severe perinatal morbidity and mortality index (SPMMI). Vaccine effectiveness was estimated, adjusted by maternal risk profile. FINDINGS: We enrolled 4618 pregnant women from Nov 27, 2021 (the day after WHO declared omicron a variant of concern), to June 30, 2022: 1545 (33%) women had a COVID-19 diagnosis (median gestation 36·7 weeks [IQR 29·0-38·9]) and 3073 (67%) women, with similar demographic characteristics, did not have a COVID-19 diagnosis. Overall, women with a diagnosis had an increased risk for MMMI (relative risk [RR] 1·16 [95% CI 1·03-1·31]) and SPMMI (RR 1·21 [95% CI 1·00-1·46]). Women with a diagnosis, compared with those without a diagnosis, also had increased risks of SNMI (RR 1·23 [95% CI 0·88-1·71]), although the lower bounds of the 95% CI crossed unity. Unvaccinated women with a COVID-19 diagnosis had a greater risk of MMMI (RR 1·36 [95% CI 1·12-1·65]). Severe COVID-19 symptoms in the total sample increased the risk of severe maternal complications (RR 2·51 [95% CI 1·84-3·43]), perinatal complications (RR 1·84 [95% CI 1·02-3·34]), and referral, intensive care unit (ICU) admission, or death (RR 11·83 [95% CI 6·67-20·97]). Severe COVID-19 symptoms in unvaccinated women increased the risk of MMMI (RR 2·88 [95% CI 2·02-4·12]) and referral, ICU admission, or death (RR 20·82 [95% CI 10·44-41·54]). 2886 (63%) of 4618 total participants had at least a single dose of any vaccine, and 2476 (54%) of 4618 had either complete or booster doses. Vaccine effectiveness (all vaccines combined) for severe complications of COVID-19 for all women with a complete regimen was 48% (95% CI 22-65) and 76% (47-89) after a booster dose. For women with a COVID-19 diagnosis, vaccine effectiveness of all vaccines combined for women with a complete regimen was 74% (95% CI 48-87) and 91% (65-98) after a booster dose. INTERPRETATION: COVID-19 in pregnancy, during the first 6 months of omicron as the variant of concern, was associated with increased risk of severe maternal morbidity and mortality, especially among symptomatic and unvaccinated women. Women with complete or boosted vaccine doses had reduced risk for severe symptoms, complications, and death. Vaccination coverage among pregnant women remains a priority. FUNDING: None.

Searching for drug leads targeted to the hydrophobic cleft of dengue virus capsid protein.

We synthesised and screened 18 aromatic derivatives of guanylhydrazones and oximes aromatic for their capacity to bind to dengue virus capsid protein (DENVC). The intended therapeutic target was the hydrophobic cleft of DENVC, which is a region responsible for its anchoring in lipid droplets in the infected cells. The inhibition of this process completely suppresses virus infectivity. Using NMR, we describe five compounds able to bind to the α1-α2 interface in the hydrophobic cleft. Saturation transfer difference experiments showed that the aromatic protons of the ligands are important for the interaction with DENVC. Fluorescence binding isotherms indicated that the selected compounds bind at micromolar affinities, possibly leading to binding-induced conformational changes. NMR-derived docking calculations of ligands showed that they position similarly in the hydrophobic cleft. Cytotoxicity experiments and calculations of in silico drug properties suggest that these compounds may be promising candidates in the search for antivirals targeting DENVC.

The Evolution and Distribution of Pneumococcal Serotypes in Adults Hospitalized With Community-Acquired Pneumonia in Spain Using a Serotype-Specific Urinary Antigen Detection Test: The CAPA Study, 2011-2018.

BACKGROUND: Spain introduced the 13-valent pneumococcal conjugate vaccine (PCV13) in the childhood National Immunization Program in 2015-2016 with coverage of 3 doses of 94.8% in 2018. We assessed the evolution of all pneumococcal, PCV13 vaccine type (VT), and experimental PCV20-VT (PCV13 + serotypes 8, 10A, 11A, 12F, 15B, 22F, 33F) hospitalized community-acquired pneumonia (CAP) in adults in Spain from 2011-2018. METHODS: A prospective observational study of immunocompetent adults (≥18 years) admitted to 4 Spanish hospitals with chest X-ray-confirmed CAP between November 2011 and November 2018. Microbiological confirmation was obtained using the Pfizer serotype-specific urinary antigen detection tests (UAD1/UAD2), BinaxNow test for urine, and conventional cultures of blood, pleural fluid, and high-quality sputum. RESULTS: Of 3107 adults hospitalized with CAP, 1943 were ≥65 years. Underlying conditions were present in 87% (n = 2704) of the participants. Among all patients, 895 (28.8%) had pneumococcal CAP and 439 (14.1%) had PCV13-VT CAP, decreasing from 17.9% (n = 77) to 13.2% (n = 68) from 2011-2012 to 2017-2018 (P = .049). PCV20-VT CAP occurred in 243 (23.8%) of those included in 2016-2018. The most identified serotypes were 3 and 8. Serotype 3 accounted for 6.9% (n = 215) of CAP cases, remaining stable during the study period, and was associated with disease severity. CONCLUSIONS: PCV13-VT caused a substantial proportion of CAP in Spanish immunocompetent adults 8 years after introduction of childhood PCV13 immunization. Improving direct PCV13 coverage of targeted adult populations could further reduce PCV13-VT burden, a benefit that could be increased further if PCV20 is licensed and implemented.

International gestational age-specific centiles for blood pressure in pregnancy from the INTERGROWTH-21st Project in 8 countries: A longitudinal cohort study.

BACKGROUND: Gestational hypertensive and acute hypotensive disorders are associated with maternal morbidity and mortality worldwide. However, physiological blood pressure changes in pregnancy are insufficiently defined. We describe blood pressure changes across healthy pregnancies from the International Fetal and Newborn Growth Consortium for the 21st Century (INTERGROWTH-21st) Fetal Growth Longitudinal Study (FGLS) to produce international, gestational age-specific, smoothed centiles (third, 10th, 50th, 90th, and 97th) for blood pressure. METHODS AND FINDINGS: Secondary analysis of a prospective, longitudinal, observational cohort study (2009 to 2016) was conducted across 8 diverse urban areas in Brazil, China, India, Italy, Kenya, Oman, the United Kingdom, and the United States of America. We enrolled healthy women at low risk of pregnancy complications. We measured blood pressure using standardised methodology and validated equipment at enrolment at <14 weeks, then every 5 ± 1 weeks until delivery. We enrolled 4,607 (35%) women of 13,108 screened. The mean maternal age was 28·4 (standard deviation [SD] 3.9) years; 97% (4,204/4,321) of women were married or living with a partner, and 68% (2,955/4,321) were nulliparous. Their mean body mass index (BMI) was 23.3 (SD 3.0) kg/m2. Systolic blood pressure was lowest at 12 weeks: Median was 111.5 (95% CI 111.3 to 111.8) mmHg, rising to a median maximum of 119.6 (95% CI 118.9 to 120.3) mmHg at 40 weeks' gestation, a difference of 8.1 (95% CI 7.4 to 8.8) mmHg. Median diastolic blood pressure decreased from 12 weeks: 69.1 (95% CI 68.9 to 69.3) mmHg to a minimum of 68.5 (95% CI 68.3 to 68.7) mmHg at 19+5 weeks' gestation, a change of -0·6 (95% CI -0.8 to -0.4) mmHg. Diastolic blood pressure subsequently increased to a maximum of 76.3 (95% CI 75.9 to 76.8) mmHg at 40 weeks' gestation. Systolic blood pressure fell by >14 mmHg or diastolic blood pressure by >11 mmHg in fewer than 10% of women at any gestational age. Fewer than 10% of women increased their systolic blood pressure by >24 mmHg or diastolic blood pressure by >18 mmHg at any gestational age. The study's main limitations were the unavailability of prepregnancy blood pressure values and inability to explore circadian effects because time of day was not recorded for the blood pressure measurements. CONCLUSIONS: Our findings provide international, gestational age-specific centiles and limits of acceptable change to facilitate earlier recognition of deteriorating health in pregnant women. These centiles challenge the idea of a clinically significant midpregnancy drop in blood pressure.

Evaluation of Cardiotonic Steroid Modulation of Cellular Cholesterol and Phospholipid.

We have previously shown that 21-benzylidene digoxin (21-BD) increases the total cholesterol and phospholipid content on the membrane of HeLa cells. Lipid modulation caused by cardiotonic steroids (CTS) is still unexplored. Therefore, the aim of the present study was to evaluate the cholesterol and phospholipid modulation of the cell membrane caused by ouabain and 21-BD and the possible involvement of the caveolae on this modulation. For this, one cell line containing caveolae (HeLa) and other not containing (Caco-2) were used. The modulation of the lipid profile was evaluated by total cholesterol and phospholipids measurements, and identification of membrane phospholipids by HPTLC. The cholesterol distribution was evaluated by filipin staining. The caveolin-1 expression was evaluated by Western Blotting. Ouabain had no effect on the total membrane lipid content in both cell lines. However, 21-BD increased total membrane phospholipid content and had no effect on the membrane cholesterol content in Caco-2 cells. CTS were not able to alter the specific phospholipids content. In the filipin experiments, 21-BD provoked a remarkable redistribution of cholesterol to the perinuclear region of HeLa cells. In Caco-2 cells, it was observed only a slight increase in cholesterol, especially as intracellular vesicles. The caveolin-1 expression was not altered by any of the compounds. Our data mainly show different effects of two cardiotonic steroids. Ouabain had no effect on the lipid profile of cells, whereas 21-BD causes important changes in cholesterol and phospholipid content. Therefore, the modulation of cholesterol content in the plasma membrane of HeLa cells is not correlated with the expression of caveolin-1.

Implications of Synthetic Modifications of the Cardiotonic Steroid Lactone Ring on Cytotoxicity.

Na,K-ATPase (NKA) and cardiotonic steroids (CTS) have shown potent cytotoxic and anticancer effects. Here, we have synthesized a series of CTS digoxin derivatives (γ-benzylidene) with substitutions in the lactone ring and evaluated the cytotoxicity caused by digoxin derivatives in tumor and non-tumor cells lines, as well as their effects on NKA. The cytotoxicity assay was determined in HeLa, A549, and WI-26 VA4 after they were treated for 48 h with increased concentrations of CTS. The effects of CTS on NKA activity and immunoblotting of α1 and ß1 isoforms were evaluated at IC50 concentrations in A549 cell membrane. NKA activity from mouse brain cortex was also measured. The majority of CTS exhibited low cytotoxicity in tumor and non-tumor cells, presenting IC50 values at micromolar concentrations, while digoxin showed cytotoxicity at nanomolar concentrations. BD-15 presented the lowest IC50 value (8 µM) in A549 and reduced its NKA activity in 28%. In contrast, BD-7 was the compound that most inhibited NKA (56% inhibition) and presented high IC50 value for A549. In mouse cortex, only BD-15 modulated the enzyme activity in a concentration-dependent inhibition curve. These results demonstrate that the cytotoxicity of these compounds is not related to NKA inhibition. The substitutions in the lactone ring of digoxin led to an increase in the cytotoxic concentration in tumor cells, which may not be interesting for cancer, but it has the advantage of increasing the therapeutic margin of these molecules when compared to classic CTS, and can be used safely in research for other diseases.

The γ-Benzylidene Digoxin Derivative BD-15 Increases the α3-Na, K-ATPase Activity in Rat Hippocampus and Prefrontal Cortex and no Change on Heart.

Our study aimed to investigate the effects of the new cardiotonic steroid BD-15 (γ-benzylidene derivatives) in the behavioral parameters, oxidative stress and the Na, K-ATPase activity in the hippocampus, prefrontal cortex and heart from rats to verify the safety and possible utilization in brain disorders. For this study, groups of male Wistar rats were used after intraperitoneal injection of 20, 100 and 200 µg/Kg with BD-15. The groups were treated for three consecutive days and the control group received 0.9% saline. BD-15 did not alter behavior of rats treated with different doses. An increase in the specific α2,3-Na, K-ATPase activity was observed for all doses of BD-15 tested in the hippocampus. However, in the prefrontal cortex, only the dose of 100 µg/Kg increased the activity of all Na, K-ATPase isoforms. BD-15 did not cause alteration in the lipid peroxidation levels in the hippocampus, but in the prefrontal cortex, a decrease of lipid peroxidation (~ 25%) was observed. In the hippocampus, GSH levels increased with all doses tested, while in the prefrontal cortex no changes were found. Subsequently, when the effect of BD-15 on cardiac tissue was analyzed, no changes were observed in the tested parameters. BD-15 at a dosage of 100 µg/Kg proved to be promising because it is considered therapeutic for brain disorders, since it increases the activity of the α3-Na, K-ATPase in the hippocampus and prefrontal cortex, as well as decreasing the oxidative stress in these brain regions. In addition, this drug did not cause changes in the tissues of the heart and kidneys, preferentially demonstrating specificity for the brain.

Fetal growth velocity standards from the Fetal Growth Longitudinal Study of the INTERGROWTH-21st Project.

BACKGROUND: Human growth is susceptible to damage from insults, particularly during periods of rapid growth. Identifying those periods and the normative limits that are compatible with adequate growth and development are the first key steps toward preventing impaired growth. OBJECTIVE: This study aimed to construct international fetal growth velocity increment and conditional velocity standards from 14 to 40 weeks' gestation based on the same cohort that contributed to the INTERGROWTH-21st Fetal Growth Standards. STUDY DESIGN: This study was a prospective, longitudinal study of 4321 low-risk pregnancies from 8 geographically diverse populations in the INTERGROWTH-21st Project with rigorous standardization of all study procedures, equipment, and measurements that were performed by trained ultrasonographers. Gestational age was accurately determined clinically and confirmed by ultrasound measurement of crown-rump length at <14 weeks' gestation. Thereafter, the ultrasonographers, who were masked to the values, measured the fetal head circumference, biparietal diameter, occipitofrontal diameter, abdominal circumference, and femur length in triplicate every 5 weeks (within 1 week either side) using identical ultrasound equipment at each site (4-7 scans per pregnancy). Velocity increments across a range of intervals between measures were modeled using fractional polynomial regression. RESULTS: Peak velocity was observed at a similar gestational age: 16 and 17 weeks' gestation for head circumference (12.2 mm/wk), and 16 weeks' gestation for abdominal circumference (11.8 mm/wk) and femur length (3.2 mm/wk). However, velocity growth slowed down rapidly for head circumference, biparietal diameter, occipitofrontal diameter, and femur length, with an almost linear reduction toward term that was more marked for femur length. Conversely, abdominal circumference velocity remained relatively steady throughout pregnancy. The change in velocity with gestational age was more evident for head circumference, biparietal diameter, occipitofrontal diameter, and femur length than for abdominal circumference when the change was expressed as a percentage of fetal size at 40 weeks' gestation. We have also shown how to obtain accurate conditional fetal velocity based on our previous methodological work. CONCLUSION: The fetal skeleton and abdomen have different velocity growth patterns during intrauterine life. Accordingly, we have produced international Fetal Growth Velocity Increment Standards to complement the INTERGROWTH-21st Fetal Growth Standards so as to monitor fetal well-being comprehensively worldwide. Fetal growth velocity curves may be valuable if one wants to study the pathophysiology of fetal growth. We provide an application that can be used easily in clinical practice to evaluate changes in fetal size as conditional velocity for a more refined assessment of fetal growth than is possible at present (https://lxiao5.shinyapps.io/fetal_growth/). The application is freely available with the other INTERGROWTH-21st tools at https://intergrowth21.tghn.org/standards-tools/.

Alkaloid and phenolic compounds of Xylopia aromatica inhibits tumor growth by down-regulating matrix metalloproteinase-2 (MMP-2) expression.

Annonacea species have been reported to possess antitumor properties. However, the in vitro and in vivo antitumor activities of Xylopia aromatica (Annonacea) have not yet been elucidated. This study aimed to investigate the effects of Xylopia aromatica leaves hexane fraction (XaHF) on Ehrlich ascites carcinoma cells lines (EAC), both in vitro and in vivo. In vitro assays revealed a significant cytotoxic effect with the two lower XaHF concentrations (62.5 and 32.3mg/mL). EAC (2.5x106 cells) were inoculated in the right flank of Swiss mice, and the animals were treated intraperitoneally with 32.3mg kg-1 of XaHF daily, for 20 days. Our findings indicate that XaHF suppressed the growth of EAC in vivo, with a significant decrease (46%) in tumor volume. There was also a decrease in the necrosis area (71%), inflammatory infiltrate, and MMP-2 expression. High-Performance Liquid Chromatography with Diode Array Detector (HPLC-DAD) identified secondary metabolites possibly related to phenolic acids, flavonoids, and alkaloids. Thus, the results confirmed the antitumoral activity that may be related to the presence of the identified metabolites in XaHF extract.

International gestational age-specific centiles for umbilical artery Doppler indices: a longitudinal prospective cohort study of the INTERGROWTH-21st Project.

BACKGROUND: Reference values for umbilical artery Doppler indices are used clinically to assess fetal well-being. However, many studies that have produced reference charts have important methodologic limitations, and these result in significant heterogeneity of reported reference ranges. OBJECTIVES: To produce international gestational age-specific centiles for umbilical artery Doppler indices based on longitudinal data and the same rigorous methodology used in the original Fetal Growth Longitudinal Study of the INTERGROWTH-21st Project. STUDY DESIGN: In Phase II of the INTERGROWTH-21st Project (the INTERBIO-21st Study), we prospectively continued enrolling pregnant women according to the same protocol from 3 of the original populations in Pelotas (Brazil), Nairobi (Kenya), and Oxford (United Kingdom) that had participated in the Fetal Growth Longitudinal Study. Women with a singleton pregnancy were recruited at <14 weeks' gestation, confirmed by ultrasound measurement of crown-rump length, and then underwent standardized ultrasound every 5±1 weeks until delivery. From 22 weeks of gestation umbilical artery indices (pulsatility index, resistance index, and systolic/diastolic ratio) were measured in a blinded fashion, using identical equipment and a rigorously standardized protocol. Newborn size at birth was assessed using the international INTERGROWTH-21st Standards, and infants had detailed assessment of growth, nutrition, morbidity, and motor development at 1 and 2 years of age. The appropriateness of pooling data from the 3 study sites was assessed using variance component analysis and standardized site differences. Umbilical artery indices were modeled as functions of the gestational age using an exponential, normal distribution with second-degree fractional polynomial smoothing; goodness of fit for the overall models was assessed. RESULTS: Of the women enrolled at the 3 sites, 1629 were eligible for this study; 431 (27%) met the entry criteria for the construction of normative centiles, similar to the proportion seen in the original fetal growth longitudinal study. They contributed a total of 1243 Doppler measures to the analysis; 74% had 3 measures or more. The healthy low-risk status of the population was confirmed by the low rates of preterm birth (4.9%) and preeclampsia (0.7%). There were no neonatal deaths and satisfactory growth, health, and motor development of the infants at 1 and 2 years of age were documented. Only a very small proportion (2.8%-6.5%) of the variance of Doppler indices was due to between-site differences; in addition, standardized site difference estimates were marginally outside this threshold in only 1 of 27 comparisons, and this supported the decision to pool data from the 3 study sites. All 3 Doppler indices decreased with advancing gestational age. The 3rd, 5th 10th, 50th, 90th, 95th, and 97th centiles according to gestational age for each of the 3 indices are provided, as well as equations to allow calculation of any value as a centile and z scores. The mean pulsatility index according to gestational age = 1.02944 + 77.7456*(gestational age)-2 - 0.000004455*gestational age3. CONCLUSION: We present here international gestational age-specific normative centiles for umbilical artery Doppler indices produced by studying healthy, low-risk pregnant women living in environments with minimal constraints on fetal growth. The centiles complement the existing INTERGROWTH-21st Standards for assessment of fetal well-being.

Complexity in Biological Organization: Deconstruction (and Subsequent Restating) of Key Concepts.

The "magic" word complexity evokes a multitude of meanings that obscure its real sense. Here we try and generate a bottom-up reconstruction of the deep sense of complexity by looking at the convergence of different features shared by complex systems. We specifically focus on complexity in biology but stressing the similarities with analogous features encountered in inanimate and artefactual systems in order to track an integrative path toward a new "mainstream" of science overcoming the actual fragmentation of scientific culture.

Hemicranial Cough-Induced Headache as a First Symptom of a Carotid-Cavernous Fistula-Case Report.

Background and objectives: Spontaneous carotid-cavernous fistulas (CCFs) are rare, and they may be caused by an aneurysm rupture. Materials and Methods: A case of a man hospitalized for high-intensity hemicranial headache with sudden cough onset as part of an upper respiratory tract infection is presented. The pain was of a pulsating character, localized on the right, behind the eye, followed by nausea and vomiting. Neurological finding registered a wider rima oculi to the right and slight neck rigidity. Laboratory findings detected a mild leukocytosis with neutrophil predominance, while cytobiochemical findings of CSF and a computerized tomography (CT) scan of the endocranium were normal. Results: Magnetic resonance imaging (MRI) angiography indicated the presence of a carotid cavernous fistula with a pseudoaneurysm to the right. Digital subtraction angiography (DSA) was performed to confirm the existence of the fistula. The planned artificial embolization was not performed because a complete occlusion of the fistula occurred during angiographic examination. Patient was discharged without subjective complaints and with normal neurological findings. Conclusions: Hemicranial cough-induced headache may be the first sign of carotid cavernous fistula, which was resolved by a spontaneous thrombosis in preparation for artificial embolization.

Evaluation of neuroprotective activity of digoxin and semisynthetic derivatives against partial chemical ischemia.

Recently, cardiotonic steroids (CTS) have been shown to lead to the activation of Na,K-ATPase at low concentrations in brain, promoting neuroprotection against ischemia. We report here the results of the use of digoxin and its semisynthetic derivatives BD-14, BD-15, and BD-16 against partial chemical ischemic induction followed by reperfusion in murine neuroblastoma cells neuro-2a (N2a). For chemical ischemic induction, sodium azide (5 mM) was used for 5 hours, and then reperfusion was induced for 24 hours. Na,K-ATPase activity and protein levels were analyzed in membrane preparation of N2a cells pretreated with the compounds (150 nM), in the controls and in induced chemical ischemia. In the Na,K-ATPase activity and protein levels assays, the steroids digoxin and BD-15 demonstrated a capacity to modulate the activity of the enzyme directly, increasing its levels of expression and activity. Oxidative parameters, such as superoxide dismutase (SOD) activity, lipid peroxidation (thiobarbituric acid reactive substance), glutathione peroxidase (GPx), glutathione (GSH) levels, hydrogen peroxide content, and the amount of free radicals (reactive oxygen species) during induced chemical ischemia were also evaluated. Regarding the redox state, lipid peroxidation, hydrogen peroxide content, and GPx activity, we have observed an increase in the chemical ischemic group, and a reduction in the groups treated with CTS. SOD activity increased in all treated groups when compared to control and GSH levels decreased when treated with sodium azide and did not change with CTS treatments. Regarding the lipid profile, we saw a decrease in the content of phospholipids and cholesterol in the chemical ischemic group, and an increase in the groups treated with CTS. In conclusion, the compounds used in this study demonstrate promising results, since they appear to promote neuroprotection in cells exposed to chemical ischemia.

Expression, purification and spectrophotometric analysis of nucleoside hydrolase from Leishmania chagasi (LcNH).

Leishmaniasis represents an important public health problem in several countries. The main target in this study is the nucleoside hydrolase Leishmania chagasi (LcNH) that is responsible for causing visceral leishmaniasis, principally in Brazil. Nucleoside hydrolase enzymes are members of this pathway, hydrolyzing the N-glycosidic bond of ribonucleosides for the synthesis of nucleic acids. We present here for the first time, the expression and purification protocols to obtain the enzymes LcNH1 and LcNH2 that can be employed to explore novel strategies to produce nucleoside hydrolase inhibitors for use in chemotherapy. Protein integrity was also confirmed by SDS-PAGE gel, mass spectrometry and enzymatic activity.

Synthesis of New Quinoline-Piperonal Hybrids as Potential Drugs against Alzheimer's Disease.

Six quinoline-piperonal hybrids were synthesized and evaluated as potential drugs against Alzheimer's disease (AD). Theoretical analysis of the pharmacokinetic and toxicological properties of the compounds suggest that they present good oral bio-availability and are also capable of penetrating the blood-brain barrier, qualifying as leads for new drugs against AD. Evaluation of their inhibitory capacity against acetyl- and butyrilcholinesterases (AChE and BChE) through Ellmann's test showed that three compounds present promising results with one of them being capable of inhibiting both enzymes. Further docking studies of the six compounds synthesized helped to elucidate the main interactions that may be responsible for the inhibitory activities observed.

Involvement of Src signaling in the synergistic effect between cisplatin and digoxin on cancer cell viability.

Cisplatin and other platinum-containing drugs have played a crucial role in anticancer treatments for over 30 years. However, treatment with cisplatin may cause serious side effects, such as myelosuppression, nausea, ototoxicity, nephrotoxicity, and cell resistance processes. In addition, cardiotonic steroids, particularly digoxin, have recently been suggested to exert potent anticancer effects. Therefore, it is possible that the combined treatment of HeLa cells with cisplatin and digoxin can ameliorate the cytotoxic effects and decrease the side effects of cisplatin. In this study, we demonstrated that the interaction between cisplatin and digoxin had a synergistic effect on cervical cancer cells and a significantly positive cytotoxic and antiproliferative effect on this cell line compared to the control and single cisplatin treatments. Although a decrease in the Na,K-ATPase α1 subunit expression was observed in total extracts, its expression remains unchanged in the membrane, as does the Na,K-ATPase activity. The antiproliferative effect of the synergistic treatment appears to depend on Src kinase activation, indicating the possible involvement of the Scr-EGFR-ERK1/2 pathway in the antitumor effect. The inhibition of ERK1/2 provoked the same synergism with 1 µM cisplatin as that observed with 1 nM digoxin plus 1 µM cisplatin but not with 1 nM digoxin. Pretreatment with PP2 during combined treatment abolished the synergistic effect on the antiproliferative activity. Cisplatin and digoxin are already used in the clinical setting; therefore, this study opens possibilities for future clinical trials of combined treatments to improve treatment outcomes with a lower incidence of toxicity and side effects.

The INTERGROWTH-21st fetal growth standards: toward the global integration of pregnancy and pediatric care.

The purpose of the INTERGROWTH-21st project was to develop international, prescriptive standards for fetal growth assessed by ultrasound and fundal height, preterm postnatal growth, newborn size and body composition, maternal weight gain, and infant development at the age of 2 years. Hence, we have produced, based on World Health Organization recommendations, the first comprehensive set of international standards of optimal fetal and newborn growth that perfectly match the existing World Health Organization child growth standards. Uniquely, the same population was followed up longitudinally from 9 weeks of fetal life to 2 years of age, with growth, health, and nutritional status assessment at 2 years supporting the appropriateness of the population for construction of growth standards. The resulting package of clinical tools allows, for the first time, growth and development to be monitored from early pregnancy to infancy. The INTERGROWTH-21st fetal growth standards, which are based on observing >4500 healthy pregnancies, nested in a study of >59,000 pregnancies from populations with low rates of adverse perinatal outcomes, show how fetuses should grow-rather than the more limited objective of past references, which describe how they have grown at specific times and locations. Our work has confirmed the fundamental biological principle that variation in human growth across different populations is mostly dependent on environmental, nutritional, and socioeconomic factors. We found that when mothers' nutritional and health needs are met and there are few environmental constraints on growth, <3.5% of the total variability of skeletal growth was due to differences between populations. We propose that not recognizing the concept of optimal growth could deprive the most vulnerable mothers and their babies of optimal care, because local growth charts normalize those at highest risk for growth restriction and overweight, and can be valuable for policymakers to ensure rigorous evaluation and effective resource allocation. We strongly encourage colleagues to join efforts to provide integrated, evidence-based growth monitoring to pregnant women and their infants worldwide. Presently, there are 23.3 million infants born small for gestational age in low- to middle-income countries according to the INTERGROWTH-21st newborn size standards. We suggest that misclassification of these infants by using local charts could affect the delivery of optimal health care.

The satisfactory growth and development at 2 years of age of the INTERGROWTH-21st Fetal Growth Standards cohort support its appropriateness for constructing international standards.

BACKGROUND: The World Health Organization recommends that human growth should be monitored with the use of international standards. However, in obstetric practice, we continue to monitor fetal growth using numerous local charts or equations that are based on different populations for each body structure. Consistent with World Health Organization recommendations, the INTERGROWTH-21st Project has produced the first set of international standards to date pregnancies; to monitor fetal growth, estimated fetal weight, Doppler measures, and brain structures; to measure uterine growth, maternal nutrition, newborn infant size, and body composition; and to assess the postnatal growth of preterm babies. All these standards are based on the same healthy pregnancy cohort. Recognizing the importance of demonstrating that, postnatally, this cohort still adhered to the World Health Organization prescriptive approach, we followed their growth and development to the key milestone of 2 years of age. OBJECTIVE: The purpose of this study was to determine whether the babies in the INTERGROWTH-21st Project maintained optimal growth and development in childhood. STUDY DESIGN: In the Infant Follow-up Study of the INTERGROWTH-21st Project, we evaluated postnatal growth, nutrition, morbidity, and motor development up to 2 years of age in the children who contributed data to the construction of the international fetal growth, newborn infant size and body composition at birth, and preterm postnatal growth standards. Clinical care, feeding practices, anthropometric measures, and assessment of morbidity were standardized across study sites and documented at 1 and 2 years of age. Weight, length, and head circumference age- and sex-specific z-scores and percentiles and motor development milestones were estimated with the use of the World Health Organization Child Growth Standards and World Health Organization milestone distributions, respectively. For the preterm infants, corrected age was used. Variance components analysis was used to estimate the percentage variability among individuals within a study site compared with that among study sites. RESULTS: There were 3711 eligible singleton live births; 3042 children (82%) were evaluated at 2 years of age. There were no substantive differences between the included group and the lost-to-follow up group. Infant mortality rate was 3 per 1000; neonatal mortality rate was 1.6 per 1000. At the 2-year visit, the children included in the INTERGROWTH-21st Fetal Growth Standards were at the 49th percentile for length, 50th percentile for head circumference, and 58th percentile for weight of the World Health Organization Child Growth Standards. Similar results were seen for the preterm subgroup that was included in the INTERGROWTH-21st Preterm Postnatal Growth Standards. The cohort overlapped between the 3rd and 97th percentiles of the World Health Organization motor development milestones. We estimated that the variance among study sites explains only 5.5% of the total variability in the length of the children between birth and 2 years of age, although the variance among individuals within a study site explains 42.9% (ie, 8 times the amount explained by the variation among sites). An increase of 8.9 cm in adult height over mean parental height is estimated to occur in the cohort from low-middle income countries, provided that children continue to have adequate health, environmental, and nutritional conditions. CONCLUSION: The cohort enrolled in the INTERGROWTH-21st standards remained healthy with adequate growth and motor development up to 2 years of age, which supports its appropriateness for the construction of international fetal and preterm postnatal growth standards.

Differences of lipid membrane modulation and oxidative stress by digoxin and 21-benzylidene digoxin.

Cardiotonic steroids (CTS) are compounds which bind to the Na,K-ATPase, leading to its inhibition and in some cases initiating signaling cascades. Long utilized as a treatment for congestive heart disease, CTS have more recently been observed to inhibit proliferation and cause apoptosis in several cancer cell lines. A synthetic derivative of the CTS digoxin, called 21-benzylidene digoxin (21-BD), activates the Na,K-ATPase rather than cause its inhibition, as its parent compound does. Here, the mechanism behind the unique effects of 21-BD are further explored. In HeLa cancer cells, low (5µM) and high (50µM) doses of 21-BD activated and inhibited the Na,K-ATPase, respectively, without altering the membrane expression of the Na,K-ATPase. While digoxin did not affect HeLa membrane cholesterol or phospholipid content, 50µM 21-BD increased both lipids via a mechanism reliant on an intact cell. Afterwards, the direct action of 21-BD was evaluated on erythrocyte membranes; however, no effect was observed. As CTS may generate reactive oxygen species (ROS) which can affect plasma membrane fluidity and therefore Na,K-ATPase activity, several markers involved in ROS generation were analyzed such as, lipid peroxidation (TBARS), reduced glutathione (GSH), catalase (CAT) and superoxide dismutase (SOD). GSH content and catalase activity were unaffected by digoxin or 21-BD. Surprisingly, TBARS and SOD activity was decreased with digoxin and with 50µM 21-BD. Thus, 21-BD and digoxin altered components involved in ROS generation and inhibition in a similar fashion. This study suggests alterations to the Na,K-ATPase and membrane lipids by 21-BD is not reliant on ROS generation.