Rare PECAM1 variants in three families with lymphedema.

PECAM1 is a member of the immunoglobulin superfamily and is expressed in monocytes, neutrophils, macrophages and other types of immune cells as well as in endothelial cells. PECAM1 function is crucial for the development and maturation of B lymphocytes. The aim of this study was to link rare PECAM1 variants found in lymphedema patients with the development of lymphatic system malformations. Using NGS, we previously tested 246 Italian lymphedema patients for variants in 29 lymphedema-associated genes and obtained 235 negative results. We then tested these patients for variants in the PECAM1 gene. We found three probands with rare variants in PECAM1. All variants were heterozygous missense variants. In Family 1, the unaffected mother and brother of the proband were found to carry the same variant as the proband. Lymphoscintigraphy was performed to determine possible lymphatic malformations and showed that in both cases a bilateral slight reduction in the speed and lymphatic clearance of the lower limbs. PECAM1 function is important for lymphatic vasculature formation. We found variants in PECAM1 that may be associated with susceptibility to lymphedema.

Biophysical mechanism for zinc as an anticancer agent.

The experimental observation that an increase in calcium above micromolar concentrations results in a slowing or stopping of anaphase-A motion is evidence for an electrostatic mechanism for poleward mitotic chromosome motions. Specifically, higher concentrations of doubly-charged calcium ions screen negative charges at microtubule free "plus" ends at kinetochores and at centrosomes. These structures normally interact with positive charges at kinetochores and positively charged microtubule free ends vicinal to centrosomes to generate poleward force. As with calcium ions, doubly-charged zinc cations can also shield these negative charges, thereby interfering with force generation for anaphase-A chromosome motion, aborting mitosis. Experimental evidence reveals that dysregulation of free cytosolic zinc homeostasis contributes to cancerous transformation. Treatment of cancers by increasing zinc concentration has unknowingly been accomplished by utilizing zinc ionophores to facilitate zinc transport across the plasma membrane, revealing an inverse relationship between malaria incidence - and malaria treatment with zinc ionophores - and cancer mortality. Here we hypothesize a biophysical mechanism for cancer therapy employing zinc supplementation enhanced by zinc ionophores.

CYP26B1 and its implications in lymphangiogenesis: Literature review and study of rare variants in two families.

CYP26B1 is a member of the cytochrome P450 family and is responsible for the break-down of retinoic acid for which appropriate levels are important for normal development of the cardiovascular and lymphatic systems. In a cohort of 235 patients with lymphatic malformations, we performed genetic testing for the CYP26B1 gene. These probands had previously tested negative for known lymphedema genes. We identified two heterozygous missense CY-P26B1 variants in two patients. Our bioinformatic study suggested that alterations caused by these variants have no major effect on the overall stability of CYP26B1 protein structure. Balanced levels of retinoic acid maintained by CYP26B1 are crucial for the lymphatic system. We identified that CYP26B1 could be involved in predisposition for lymphedema. We propose that CYP26B1 be further explored as a new candidate gene for genetic testing of lymphedema patients.

NWChem: Past, present, and future.

Specialized computational chemistry packages have permanently reshaped the landscape of chemical and materials science by providing tools to support and guide experimental efforts and for the prediction of atomistic and electronic properties. In this regard, electronic structure packages have played a special role by using first-principle-driven methodologies to model complex chemical and materials processes. Over the past few decades, the rapid development of computing technologies and the tremendous increase in computational power have offered a unique chance to study complex transformations using sophisticated and predictive many-body techniques that describe correlated behavior of electrons in molecular and condensed phase systems at different levels of theory. In enabling these simulations, novel parallel algorithms have been able to take advantage of computational resources to address the polynomial scaling of electronic structure methods. In this paper, we briefly review the NWChem computational chemistry suite, including its history, design principles, parallel tools, current capabilities, outreach, and outlook.

Maternal anxiety, depression and sleep disorders before and during pregnancy, and preschool ADHD symptoms in the NINFEA birth cohort study.

AIMS.: Maternal mental disorders have been associated with the risk of attention-deficit/hyperactivity disorder (ADHD) in children. Within the context of a mother-child cohort, we examined whether maternal anxiety, depression and sleep disorders are associated with pre-school ADHD symptoms. METHODS.: The study included 3634 singletons from the Italian NINFEA (Nascita e INFanzia: gli Effetti dell'Ambiente') cohort. Maternal doctor-diagnosed anxiety, depression and sleep disorders before and during pregnancy were assessed from the questionnaires completed during pregnancy and 6 months after delivery. Mothers rated child ADHD symptoms at 4 years of age, according to the Diagnostic and Statistical Manual of Mental Disorders. Hyperactive-impulsive (ADHD-H), inattentive (ADHD-I) and total ADHD scores were analysed in the models adjusted for child's gender, first-born status, maternal age, education, alcohol consumption and smoking during pregnancy. RESULTS.: The total ADHD score at age 4 was associated with maternal lifetime anxiety (17.1% percentage difference in score compared with never; 95% CI 7.3-27.9%), sleep disorders (35.7%; 95% CI 10.7-66.5%) and depression (17.5%; 95% CI 3.2-33.8%). Similar positive associations were observed also for ADHD-H and ADHD-I traits, with slightly attenuated associations between maternal sleep disorders and child ADHD-I score, and maternal depression and both ADHD scores. All the estimates were enhanced when the disorders were active during pregnancy and attenuated for disorders active only during the pre-pregnancy period. CONCLUSIONS.: Maternal anxiety, depression and sleep disorders are associated with a relative increase in the number of ADHD-H, ADHD-I and total ADHD symptoms in preschoolers.

High throughput method to characterize acid-base properties of insoluble drug candidates in water.

In drug design experimental characterization of acidic groups in candidate molecules is one of the more important steps prior to the in-vivo studies. Potentiometry combined with Yasuda-Shedlovsky extrapolation is one of the more important strategy to study drug candidates with low solubility in water, although, it requires a large number of sequences to determine pKa values at different solvent-mixture compositions to, finally, obtain the pKa in water (pwwKa) by extrapolation. We have recently proposed a method which requires only two sequences of additions to study the effect of organic solvent content in liquid chromatography mobile phases on the acidity of the buffer compounds usually dissolved in it along wide ranges of compositions. In this work we propose to apply this method to study thermodynamic pwwKa of drug candidates with low solubilities in pure water. Using methanol/water solvent mixtures we study six pharmaceutical drugs at 25 °C. Four of them: ibuprofen, salicylic acid, atenolol and labetalol, were chosen as members of carboxylic, amine and phenol families, respectively. Since these compounds have known pwwKa values, they were used to validate the procedure, the accuracy of Yasuda-Shedlovsky and other empirical models to fit the behaviors, and to obtain pwwKa by extrapolation. Finally, the method is applied to determine unknown thermodynamic pwwKa values of two pharmaceutical drugs: atorvastatin calcium and the two dissociation constants of ethambutol. The procedure proved to be simple, very fast and accurate in all of the studied cases.

Erratum to: Electrostatic forces drive poleward chromosome motions at kinetochores.

[This corrects the article DOI: 10.1186/s13008-016-0026-1.].

Perinatal health services organization for preterm births: a multinational comparison.

OBJECTIVE: To explore population characteristics, organization of health services and comparability of available information for very low birth weight or very preterm neonates born before 32 weeks' gestation in 11 high-income countries contributing data to the International Network for Evaluating Outcomes of Neonates (iNeo). STUDY DESIGN: We obtained population characteristics from public domain sources, conducted a survey of organization of maternal and neonatal health services and evaluated the comparability of data contributed to the iNeo collaboration from Australia, Canada, Finland, Israel, Italy, Japan, New Zealand, Spain, Sweden, Switzerland and UK. RESULTS: All countries have nationally funded maternal/neonatal health care with >90% of women receiving prenatal care. Preterm birth rate, maternal age, and neonatal and infant mortality rates were relatively similar across countries. Most (50 to >95%) between-hospital transports of neonates born at non-tertiary units were conducted by designated transport teams; 72% (8/11 countries) had designated transfer and 63% (7/11 countries) mandate the presence of a physician. The capacity of 'step-down' units varied between countries, with capacity for respiratory care available in <10% to >75% of units. Heterogeneity in data collection processes for benchmarking and quality improvement activities were identified. CONCLUSIONS: Comparability of healthcare outcomes for very preterm low birth weight neonates between countries requires an evaluation of differences in population coverage, healthcare services and meta-data.

Electrostatic forces drive poleward chromosome motions at kinetochores.

BACKGROUND: Recent experiments regarding Ndc80/Hec1 in force generation at kinetochores for chromosome motions have prompted speculation about possible models for interactions between positively charged molecules at kinetochores and negative charge at and near the plus ends of microtubules. DISCUSSION: A clear picture of how kinetochores and centrosomes establish and maintain a dynamic coupling to microtubules for force generation during the complex motions of mitosis remains elusive. The current paradigm of molecular cell biology requires that specific molecules, or molecular geometries, for force generation be identified. However, it is possible to explain several different mitotic motions-including poleward force production at kinetochores-within a classical electrostatics approach in terms of experimentally known charge distributions, modeled as surface and volume bound charges interacting over nanometer distances. CONCLUSION: We propose here that implicating Ndc80/Hec1 as a bound volume positive charge distribution in electrostatic generation of poleward force at kinetochores is most consistent with a wide range of experimental observations on mitotic motions, including polar production of poleward force and chromosome congression.

Size at birth by gestational age and hospital mortality in very preterm infants: results of the area-based ACTION project.

BACKGROUND: Size at birth is an important predictor of neonatal outcomes, but there are inconsistencies on the definitions and optimal cut-offs. AIMS: The aim of this study is to compute birth size percentiles for Italian very preterm singleton infants and assess relationship with hospital mortality. STUDY DESIGN: Prospective area-based cohort study. SUBJECTS: All singleton Italian infants with gestational age 22-31 weeks admitted to neonatal care in 6 Italian regions (Friuli Venezia-Giulia, Lombardia, Marche, Tuscany, Lazio and Calabria) (n. 1605). OUTCOME MEASURE: Hospital mortality. METHODS: Anthropometric reference charts were derived, separately for males and females, using the lambda (λ) mu (µ) and sigma (σ) method (LMS). Logistic regression analysis was used to estimate mortality rates by gestational age and birth weight centile class, adjusting for sex, congenital anomalies and region. RESULTS: At any gestational age, mortality decreased as birth weight centile increased, with lowest values observed between the 50th and the 89th centiles interval. Using the 75th-89th centile class as reference, adjusted mortality odds ratios were 7.94 (95% CI 4.18-15.08) below 10th centile; 3.04 (95% CI 1.63-5.65) between the 10th and 24th; 1.96 (95% CI 1.07-3.62) between the 25th and the 49th; 1.25 (95% CI 0.68-2.30) between the 50(h) and the 74th; and 2.07 (95% CI 1.01-4.25) at the 90th and above. CONCLUSIONS: Compared to the reference, we found significantly increasing adjusted risk of death up to the 49th centile, challenging the usual 10th centile criterion as risk indicator. Continuous measures such as the birthweight z-score may be more appropriate to explore the relationship between growth retardation and adverse perinatal outcomes.

Role of BRAF molecular analysis in the management of papillary thyroid carcinoma: analysis of cytological and histological samples.

BACKGROUND: Although fine needle aspiration (FNA) is the standard diagnostic test for the characterization of a suspicious thyroid nodule, in some cases cytological evaluation is inconclusive. The aim of this study was to determine the role of BRAF mutation in aiding diagnosis and to verify whether archival cytological samples could be suitable for molecular analysis. METHODS: Eighty-five patients with suspicious (Thy4) or follicular (Thy3) lesions on cytology were resubmitted to a second FNA for BRAF mutation analysis. Of these, 56 subsequently underwent surgery. The usefulness of archival samples for molecular analysis was also studied in a second cohort of 42 patients with a confirmed diagnosis of papillary thyroid carcinoma for whom both archived paraffin-embedded histological samples and cytological smears were available. A further 15 patients with paired fresh FNA and archived cytological and histological samples were recruited. RESULTS: BRAF mutation was found in the fresh FNA samples from 10 of 56 patients who had surgery with previous inconclusive cytology (4/45, 9%, Thy3 and 6/11, 55%, Thy4). The BRAF test showed a specificity and positive predictive value of 100% (26/26 and 10/10, respectively), sensitivity of 33% (10/30) and negative predictive value of 57% (26/46). There was absolute concordance between the BRAF results obtained with 42 histological and cytological archived samples. BRAF analysis on 15 archived cytological samples showed absolute concordance with histology, whereas there was one false negative on the matched fresh FNA. CONCLUSION: BRAF analysis is a highly specific test that can facilitate cytological diagnosis in some cases and can also be performed on archived cytological samples.

Automatized sspKa measurements of dihydrogen phosphate and Tris(hydroxymethyl) aminomethane in acetonitrile/water mixtures from 20 to 60°C.

We measured pKa values of Tris(hydroxymethyl)aminomethane and dihydrogen phosphate; both are commonly used to prepare buffers for reverse-phase liquid chromatography (RPLC), in acetonitrile/water mixtures from 0% to 70% (v/v) (64.6% (w/w)) acetonitrile and at 20, 30, 40, 50, and 60°C. The procedure is based on potentiometric measurements of pH of buffer solutions of variable solvent compositions using a glass electrode and a novel automated system. The method consists in the controlled additions of small volumes of a thermostated solution from an automatic buret into another isothermal solution containing exactly the same buffer-component concentrations, but a different solvent composition. The continuous changes in the solvent composition induce changes in the potentials. Thus, only two sequences of additions are needed: increasing the amount of acetonitrile from pure water and decreasing the content of acetonitrile from 70% (v/v) (64.6% (w/w)). In the procedure with homemade apparatus, times for additions, stirring, homogenization, and data acquisition are entirely controlled by software programmed for this specific routine. This rapid, fully automated method was applied to acquire more than 40 potential data covering the whole composition range (at each temperature) in about two hours and allowed a systematic study of the effect of temperature and acetonitrile composition on acid-base equilibria of two widely used substances to control pH close to 7. The experimental pKa results were fitted to empirical functions between pKa and temperature and acetonitrile composition. These equations allowed predictions of pKa to estimate the pH of mixtures at any composition and temperature, which would be very useful, for instance, during chromatographic method development.

Chromosome congression explained by nanoscale electrostatics.

Nanoscale electrostatic microtubule disassembly forces between positively charged molecules in kinetochores and negative charges on plus ends of microtubules have been implicated in poleward chromosome motions and may also contribute to antipoleward chromosome movements. We propose that chromosome congression can be understood in terms of antipoleward nanoscale electrostatic microtubule assembly forces between negatively charged microtubule plus ends and like-charged chromosome arms, acting in conjunction with poleward microtubule disassembly forces. Several other aspects of post-attachment prometaphase chromosome motions, as well as metaphase oscillations, are consistently explained within this framework.

Is intracellular pH a clock for mitosis?

Experiments have shown that the intracellular pH of many cells rises to a maximum at the onset of mitosis, subsequently decreasing 0.3 to 0.5 pH units by the end of mitosis. This result, and observations that tubulin net charge depends strongly on pH, may be critical for microtubule (MT) dynamics during mitosis. In vivo studies demonstrate that MT dynamics is sensitive to pH, with MT growth favored by higher pH values. Therefore it seems likely that the shift from the dominance of microtubule growth during prophase, and to a lesser extent during prometaphase, to a parity between MT polymerization and depolymerization during metaphase chromosome oscillations is a consequence of gradually decreasing intracellular pH during mitosis. Thus the timing and sequencing of prophase, prometaphase, and metaphase chromosome motions may be understood as an increase in the MT disassembly to assembly probability ratio resulting from a continuously declining intracellular pH.

Can molecular cell biology explain chromosome motions?

BACKGROUND: Mitotic chromosome motions have recently been correlated with electrostatic forces, but a lingering "molecular cell biology" paradigm persists, proposing binding and release proteins or molecular geometries for force generation. RESULTS: Pole-facing kinetochore plates manifest positive charges and interact with negatively charged microtubule ends providing the motive force for poleward chromosome motions by classical electrostatics. This conceptual scheme explains dynamic tracking/coupling of kinetochores to microtubules and the simultaneous depolymerization of kinetochore microtubules as poleward force is generated. CONCLUSION: We question here why cells would prefer complex molecular mechanisms to move chromosomes when direct electrostatic interactions between known bound charge distributions can accomplish the same task much more simply.

Effect of carbohydrate counting and medical nutritional therapy on glycaemic control in Type 1 diabetic subjects: a pilot study.

AIMS: The effect of a balanced, carbohydrate-counting diet on glycaemic control in Type 1 diabetic subjects is unclear. Our aim was to determine its effect in a small, pilot trial. METHODS: We randomized 256 Type 1 diabetic subjects to a Nutritional Education Programme (group A) or not (group B). Weight, body mass index, glycated haemoglobin (HbA1c), lipid profile, urate, creatinine, microalbuminuria and daily insulin requirements were measured at baseline and at the end of the study (9 months). During the study, the number of hypoglycaemic events (blood glucose<3.9 mmol/l) was also measured. RESULTS: Compared with group B, group A showed: (i) a reduction in HbA1c (group A: 7.8+/-1.3-7.4+/-0.9%; group B: 7.5+/-0.8-7.5+/-1.1%; P<0.01); (ii) less hypoglycaemic events (4% vs. 7%; P<0.05); (iii) a reduction in dose of rapid insulin analogues (23.5+/-10.9 vs. 27.7+/-17.1 IU/24 h; P=0.03). No other between-group changes were observed. CONCLUSIONS: This study shows the importance of medical nutritional therapy on glycaemic control in Type 1 diabetic subjects.

On the analysis of the Cr-Cr multiple bond in several classes of dichromium compounds.

Since the discovery of a formal quintuple bond in Ar'CrCrAr' (CrCr = 1.835 A) by Power and co-workers in 2005, many efforts have been dedicated to isolating dichromium species featuring quintuple-bond character. In the present study we investigate the electronic configuration of several, recently synthesized dichromium species with ligands using nitrogen to coordinate the metal centers. The bimetallic bond distances of Power's compound and Cr(2)-diazadiene (1) (CrCr = 1.803 A) are compared to those found for Cr(2)(mu-eta(2)-ArNC(R)NAr)(2) (2) (CrCr = 1.746 A; R = H, Ar = 2,6-Et(2)C(6)H(3)), Cr(2)(mu-eta(2)-Ar(Xyl)NC(H)NAr(Xyl))(3) (3) (CrCr = 1.740(reduced)/1.817(neutral) A; Ar(Xyl)= 2,6-C(6)H(3)-(CH(3))(2)), Cr(2)(mu-eta(2)-TippPyNMes)(2) (4) (CrCr = 1.749 A; TippPyNMes = 6-(2,4,6-triisopropylphenyl)pyridin-2-yl (2,4,6-trimethylphenyl)amide), and Cr(2)(mu-eta(2)-DippNC(NMe(2))N-Dipp)(2) (5) (CrCr = 1.729 A, Dipp = 2,6-i-Pr(2)C(6)H(3)). We show that the correlation between the CrCr bond length and the effective bond order (EBO) is strongly affected by the nature of the ligand, as well as by the steric hindrance due to the ligand structure (e.g., the nature of the coordinating nitrogen). A linear correlation between the EBO and CrCr bond distance is established within the same group of ligands. As a result, the CrCr species based on the amidinate, aminopyridinate, and guanidinate ligands have bond patterns similar to the Ar'CrCrAr' compound. Unlike these latter species, the dichromium diazadiene complex is characterized by a different bonding pattern involving Cr-Npi interactions, resulting in a lower bond order associated with the short metal-metal bond distance. In this case the short CrCr distance is most probably the result of the constraints imposed by the diazadiene ligand, implying a Cr(2)N(4) core with a closer CrCr interaction.

Screening for subclinical Cushing's syndrome in type 2 diabetes mellitus: low false-positive rates with nocturnal salivary cortisol.

The diagnosis of subclinical Cushing's syndrome (SCS) is important, but its relative rarity amongst patients with common metabolic disorders requires a simple test with a low false-positive rate. Using nocturnal salivary cortisol (NSC), which we first validated in patients with suspected and proven Cushing's syndrome, we screened 106 overweight patients with type 2 diabetes mellitus, a group at high risk of SCS and nontumoral hypothalamic-pituitary-adrenal axis perturbations. Our hypothesis was that a lower false-positive rate with NSC was likely, compared with that reported with the dexamethasone suppression test (DST) (10-20%), currently the foundation of diagnosis of SCS. No participant had clinically apparent Cushing's syndrome. Three participants had an elevated NSC but further testing excluded SCS. In this study, NSC had a lower false-positive rate (3%) than previously reported for the DST. Given the reported excellent performance of NSC in detection of hypercortisolism, the low false-positive rate in SCS suggests NSC may be superior to the DST for SCS screening. The NSC and DST should be compared directly in metabolic disorder patients; although our data suggest the patient group will need to be substantially larger to definitively determine the optimal screening test.

Identification and quantitation of vitamins K1 and K3 in cosmetic products for facial skin protection.

A simple and rapid analytical method was developed for the determination of vitamins K1 and K3 in facial anti-rash creams. The procedure is based on an ultrasonic extraction of the cosmetic sample with dimethylacetamide, in the presence of an internal standard, followed by HPLC separation. HPLC was performed using a C18 column and spectrophotometric detection at 333 nm. A linear gradient elution was carried out starting with 50% acetonitrile-methanol (75:25 v/v) and water up to 100% acetonitrile-methanol for 5 min. Linearity was established over the concentration range from 0.2 to 1.0 mg/ml for vitamin K1 and from 0.02 to 0.1 mg/ml for vitamin K3, with LOD values of 100 ng and 20 ng injected, respectively. The accuracy was verified by spiking experiments on model cosmetic samples. The proposed method has been successfully applied for the analysis of commercial samples of creams.