Prolonged treatment with vitamin D in postmenopausal women with primary hyperparathyroidism.
Vitamin D deficiency further increases circulating parathyroid hormone (PTH) levels in patients with primary hyperparathyroidism (pHPT), with potential detrimental effects on bone mass. This was an observational clinical study in consecutive conservatively treated postmenopausal women (n=40) with pHPT and coexistent 25-hydroxyvitamin D deficiency (25OHD ≤50 nmol/l (≤20 ng/ml)). Patients who showed an increase in serum 25OHD above the threshold of vitamin D deficiency (>50 nmol/l; n=28) using treatment with various commonly prescribed vitamin D preparations were, for the purposes of statistical analyses, allocated to the treatment group. Patients who were retrospectively identified as having received no treatment with vitamin D and/or remained vitamin D deficient were considered as non-responders/controls (n=12). Adjusted calcium (adjCa), PTH and 25OHD concentrations were monitored in all subjects up to 54 months (mean observation period of 18±2 months). Prolonged increased vitamin D intake, regardless of the source (serum 25OHD, increase from 32.2±1.7 nmol/l at baseline to 136.4±11.6 nmol/l, P<0.0001), significantly reduced serum PTH (13.3±1.1 vs 10.5±1.0 pmol/l, P=0.0001), with no adverse effects on adjCa levels (2.60±0.03 vs 2.60±0.02 mmol/l, P=0.77) and renal function tests (P>0.73). In contrast, serum PTH remained unchanged (15.8±2.6 vs 16.3±1.9 pmol/l, P=0.64) in patients who remained vitamin D deficient, with a significant difference between groups in changes of PTH (P=0.0003). Intrapartial correlation analyses showed an independent negative correlation of changes in 25OHD with PTH levels (r ic=-0.41, P=0.014). Prolonged treatment with vitamin D in various commonly prescribed preparations appeared to be safe and significantly reduced PTH levels by 21%.


Novel Striatal GABAergic Interneuron Populations Labeled in the 5HT3a(EGFP) Mouse.
Histological and morphological studies indicate that approximately 5% of striatal neurons are cholinergic or γ-aminobutyric acidergic (GABAergic) interneurons (gINs). However, the number of striatal neurons expressing known interneuron markers is too small to account for the entire interneuron population. We therefore studied the serotonin (5HT) receptor 3a-enhanced green fluorescent protein (5HT3a(EGFP)) mouse, in which we found that a large number of striatal gINs are labeled. Roughly 20% of 5HT3a(EGFP)-positive cells co-express parvalbumin and exhibit fast-spiking (FS) electrophysiological properties. However, the majority of labeled neurons do not overlap with known molecular interneuron markers. Intrinsic electrical properties reveal at least 2 distinct novel subtypes: a late-spiking (LS) neuropeptide-Y (NPY)-negative neurogliaform (NGF) interneuron, and a large heterogeneous population with several features resembling low-threshold-spiking (LTS) interneurons that do not express somatostatin, NPY, or neuronal nitric oxide synthase. Although the 5HT3a(EGFP) NGF and LTS-like interneurons have electrophysiological properties similar to previously described populations, they are pharmacologically distinct. In direct contrast to previously described NPY(+) LTS and NGF cells, LTS-like 5HT3a(EGFP) cells show robust responses to nicotine administration, while the 5HT3a(EGFP) NGF cell type shows little or no response. By constructing a molecular map of the overlap between these novel populations and existing interneuron populations, we are able to reconcile the morphological and molecular estimates of striatal interneuron numbers.


Racial and Ethnic Inequities in Cesarean Birth and Maternal Morbidity in a Low-Risk, Nulliparous Cohort.
To evaluate race and ethnicity differences in cesarean birth and maternal morbidity in low-risk nulliparous people at term. We conducted a secondary analysis of a randomized trial of expectant management compared with induction of labor in low-risk nulliparous people at term. The primary outcome was cesarean birth. Secondary outcome was maternal morbidity, defined as: transfusion of 4 or more units of red blood cells, any transfusion of other products, postpartum infection, intensive care unit admission, hysterectomy, venous thromboembolism, or maternal death. Multivariable modified Poisson regression was used to evaluate associations between race and ethnicity, cesarean birth, and maternal morbidity. Indication for cesarean birth was assessed using multivariable multinomial logistic regression. A mediation model was used to estimate the portion of maternal morbidity attributable to cesarean birth by race and ethnicity. Of 5,759 included participants, 1,158 (20.1%) underwent cesarean birth; 1,404 (24.3%) identified as non-Hispanic Black, 1,670 (29.0%) as Hispanic, and 2,685 (46.6%) as non-Hispanic White. Adjusted models showed increased relative risk of cesarean birth among non-Hispanic Black (adjusted relative risk [aRR] 1.21, 95% CI 1.03-1.42) and Hispanic (aRR 1.26, 95% CI 1.08-1.46) people compared with non-Hispanic White people. Maternal morbidity affected 132 (2.3%) individuals, and was increased among non-Hispanic Black (aRR 2.05, 95% CI 1.21-3.47) and Hispanic (aRR 1.92, 95% CI 1.17-3.14) people compared with non-Hispanic White people. Cesarean birth accounted for an estimated 15.8% (95% CI 2.1-48.7%) and 16.5% (95% CI 4.0-44.0%) of excess maternal morbidity among non-Hispanic Black and Hispanic people, respectively. Non-Hispanic Black and Hispanic nulliparous people who are low-risk at term undergo cesarean birth more frequently than low-risk non-Hispanic White nulliparous people. This difference accounts for a modest portion of excess maternal morbidity.


Regulation of a transcription factor network by Cdk1 coordinates late cell cycle gene expression.
To maintain genome stability, regulators of chromosome segregation must be expressed in coordination with mitotic events. Expression of these late cell cycle genes is regulated by cyclin-dependent kinase (Cdk1), which phosphorylates a network of conserved transcription factors (TFs). However, the effects of Cdk1 phosphorylation on many key TFs are not known. We find that elimination of Cdk1-mediated phosphorylation of four S-phase TFs decreases expression of many late cell cycle genes, delays mitotic progression, and reduces fitness in budding yeast. Blocking phosphorylation impairs degradation of all four TFs. Consequently, phosphorylation-deficient mutants of the repressors Yox1 and Yhp1 exhibit increased promoter occupancy and decreased expression of their target genes. Interestingly, although phosphorylation of the transcriptional activator Hcm1 on its N-terminus promotes its degradation, phosphorylation on its C-terminus is required for its activity, indicating that Cdk1 both activates and inhibits a single TF. We conclude that Cdk1 promotes gene expression by both activating transcriptional activators and inactivating transcriptional repressors. Furthermore, our data suggest that coordinated regulation of the TF network by Cdk1 is necessary for faithful cell division.


Automated quantitative analysis of 3D morphology and mean corpuscular hemoglobin in human red blood cells stored in different periods.
Quantitative phase (QP) images of red blood cells (RBCs), which are obtained by off-axis digital holographic microscopy, can provide quantitative information about three-dimensional (3D) morphology of human RBCs and the characteristic properties such as mean corpuscular hemoglobin (MCH) and MCH surface density (MCHSD). In this paper, we investigate modifications of the 3D morphology and MCH in RBCs induced by the period of storage time for the purpose of classification of RBCs with different periods of storage by using off-axis digital holographic microscopy. The classification of RBCs based on the duration of storage is highly relevant because a long storage of blood before transfusion may alter the functionality of RBCs and, therefore, cause complications in patients. To analyze any changes in the 3D morphology and MCH of RBCs due to storage, we use data sets from RBC samples stored for 8, 13, 16, 23, 27, 30, 34, 37, 40, 47, and 57 days, respectively. The data sets consist of more than 3,300 blood cells in eleven classes, with more than 300 blood cells per class. The classes indicate the storage period of RBCs and are listed in chronological order. Using the RBCs donated by healthy persons, the off-axis digital holographic microscopy reconstructs several quantitative phase images of RBC samples stored for eleven different periods. We employ marker-controlled watershed transform to remove the background in the RBC quantitative phase images obtained by the off-axis digital holographic microscopy. More than 300 single RBCs are extracted from the segmented quantitative phase images for each class. Such a large number of RBC samples enable us to obtain statistical distributions of the characteristic properties of RBCs after a specific period of storage. Experimental results show that the 3D morphology of the RBCs, in contrast to MCH, is essentially related to the aging of the RBCs.


Complete determination of the orientation of NV centers with radially polarized beams.
In this paper, we report on the precise determination of the orientation of NV centers by imaging with a radially polarized beam. Vectorial Debye theory is applied to the field in the focus of radially polarized beams to generate emission profiles of two orthogonal optical dipoles. By comparing features of the measured emission intensity patterns with simulated results, complete orientation determination of the NV axis is achieved. Results are corroborated by using established methods requiring the polarization rotation of a linearly polarized excitation source, and by analysis of optically detected magnetic resonance spectra. These results lay new ground for any application where the knowledge of the orientation of the NV centers is prerequisite.


Palladium-catalyzed tetrakis(dimethylamino)ethylene-promoted reductive coupling of aryl halides.
Tetrakis(dimethylamino)ethylene (TDAE)/cat. PdCl(2)(PhCN)(2)-promoted reductive coupling of aryl bromides having either electron-donating or electron-withdrawing groups on their para- and/or meta-position proceeded smoothly to afford the corresponding biaryls in good to excellent yields. Notably, TDAE is such a mild reductant that easily reducible groups, such as carbonyl and nitro groups, are tolerate. A similar reductive coupling of ortho-substituted aryl bromides did not occur at all. The proper choice of palladium catalysts is essential for the reductive coupling; thus, PdCl(2)(PhCN)(2), PdCl(2)(MeCN)(2), Pd(hfacac)(2), Pd(2)(dba)(3), PdCl(2), and Pd(OAc)(2) were used successively for this reaction, but phosphine-ligated palladium catalysts such as Pd(PPh(3))(4), PdCl(2)(PPh(3))(2), and Pd(dppp) did not promote the reaction. The reductive coupling did not occur with nickel catalysts such as NiBr(2), NiCl(2)(bpy), and Ni(acac)(2). The TDAE/cat. palladium-promoted reductive coupling of aryl halides having electron-withdrawing groups took place more efficiently than that of aryl halides substituted with electron-donating groups. A plausible mechanism of TDAE/cat. palladium-promoted reaction is discussed.


Genome Sequence of Oxalobacter formigenes Strain OXCC13.
The lack of Oxalobacter formigenes colonization in the human gut is generally acknowledged as a risk factor for kidney stone formation since this microorganism can play an important role in oxalate homeostasis. Here, we present the genome sequence of OXCC13, a human strain isolated from an individual residing in Germany.


Association Between Weight Status and Executive Function in Young Adults.
Background and objectives: To explore the association between weight status and executive function in young adults. Materials and Methods: Ninety-seven young males (age 17-26 years) underwent adiposity and body composition measurements using body composition analyzer. Inhibitory control and working memory were measured using the Cambridge Neuropsychological Test Battery (CANTAB). Results: Multiple linear regression using both unadjusted and adjusted analyses revealed no association between adiposity and body composition variables with executive tasks, apart from a significant association between skeletal muscle mass (SMM) and mean reaction time on go trial (standardized B = -0.28; p = 0.02). Multivariate analysis of covariance (MANCOVA) revealed that underweight participants presented inferior working memory compared to their normal weight (p = 0.001) or overweight peers (p = 0.008). However, according to the percentage fat quartiles (Q) participants with the highest quartile (Q4) were inferior in inhibitory control than their peers with Q2 (p = 0.04), and participants with the lowest quartile (Q1) were inferior in working memory compared with their peers with Q2 (p = 0.01) or Q3 (p = 0.02). A worse inhibitory control was observed for participants with the highest fat/SMM (Q4) compared to participants in Q3 (p = 0.03), and in contrast worse working memory was observed for participants with the lowest fat/SMM (Q1) compared to participants in Q2 (p = 0.04) or Q3 (p = 0.009). Conclusions: Low adiposity is associated with worse working memory, whereas high adiposity is associated with worse inhibitory control. Therefore, our findings show that normal adiposity, but greater SMM may have a positive impact on executive function in young adults.


Femoral Morphology in the Dysplastic Hip: Three-dimensional Characterizations With CT.
Hip dysplasia represents a spectrum of complex deformities on both sides of the joint. Although many studies have described the acetabular side of the deformity, to our knowledge, little is known about the three-dimensional (3-D) head and neck offset differences of the femora of dysplastic hips. A thorough knowledge of proximal femoral anatomy is important to prevent potential impingement and improve results after acetabular reorientation. (1) Are there common proximal femoral characteristics in patients with symptomatic hip dysplasia undergoing periacetabular osteotomy (PAO)? (2) Where is the location of maximal femoral head and neck offset deformity in hip dysplasia? (3) Do certain subgroups of dysplastic hips more commonly have cam-type femoral morphology? (4) Is there a relationship between hip ROM as well as impingement testing and 3-D head and neck offset deformity? Using our hip preservation database, 153 hips (148 patients) underwent PAO from October 2013 to July 2015. We identified 103 hips in 100 patients with acetabular dysplasia (lateral center-edge angle [LCEA] < 20°) and who had a Tönnis grade of 0 or 1. Eighty-six patients (86%) underwent preoperative low-dose pelvic CT scans at our institution as part of the preoperative planning for PAO. It is currently our standard to obtain preoperative low-dose pelvic CT scans (0.75-1.25 mSv, equivalent to three to five AP pelvis radiographs) on all patients before they undergo PAO unless a prior CT scan is performed at an outside institution. Hips with a history of a neuromuscular disorder, prior trauma, prior surgery, radiographic evidence of joint degeneration, ischemic necrosis, or Perthes-like deformities were excluded. Fifty hips in 50 patients met inclusion criteria and had CT scans available for review. Hips were analyzed with Dyonics Plan software and characterized with regard to version, neck-shaft angle, femoral head diameter, head and neck offset, femoral neck length, femoral offset, head center height, trochanteric height, and alpha angle. The maximum head and neck offset deformity was assessed using an entire clockface and an alpha angle ≥ 55° defined coexisting cam morphology. Subgroups included severity of lateral dysplasia: mild (LCEA 15°-20°) and moderate/severe (LCEA < 15°). Femoral version subgroups were defined as normal (5°-20°), decreased (≤ 5°), or increased (> 20°). The senior author (JCC) performed all physical examination testing. The mean LCEA was 14° (±4°), whereas the mean femoral anteversion was 19° (±12°). Eight hips (16%) demonstrated relative femoral retroversion (≤ 5°), whereas 26 (52%) showed excessive femoral anteversion (> 20°). Four hips (8%) had ≥ 35° of femoral anteversion. The mean neck-shaft angle was 136° (±5°). The mean maximum alpha location was 2:00 o'clock (±45 minutes) and the mean maximum alpha angle was 52° (±6°). Minimum head-neck offset ratio was located at 1:30 with a mean of 0.14 (±0.03). An anterior head-neck offset ratio of ≤ 0.17 or an alpha angle ≥ 55° was found in 43 (86%) of hips. Twenty-one dysplastic hips (42%) had an alpha angle ≥ 55°. Mildly dysplastic hips had decreased femoral head and neck offset (9 ± 1) and head and neck offset ratio (0.20 ± 0.03) at 12 o'clock compared with moderate/severe dysplastic hips (10 ± 1 and 0.22 ± 0.03, respectively; p = 0.04 and p = 0.01). With the numbers available, we found that hips with excessive femoral anteversion (> 20°) had no difference in the alpha angle at 3 o'clock (42 ± 7) compared with hips with relative femoral retroversion (≤ 5°; 48 ± 4; p = 0.06). No other differences in femoral morphology were found between hips with mild or moderate/severe dysplasia or in the femoral version subgroups with the numbers available. Anterior impingement test was positive in 76% of hips with an alpha angle ≥ 55° and 83% of the hips with an alpha angle ≤ 55°. No correlation was found between proximal femoral morphology and preoperative ROM. In this subset of dysplastic hips, cam deformity of the femoral head and neck was present in 42% of hips with maximal head-neck deformity at 2 o'clock, and 82% had reduced head-neck offset at the 1:30 point. We conclude that cam-type deformities and decreased head-neck offset in developmental dysplasia of the hip are common. Patients should be closely assessed for need of a head and neck osteochondroplasty, especially after acetabular correction. Future prospective studies should evaluate the influence of proximal femoral anatomy on surgical results of PAO for dysplastic hips. Level IV, prognostic study.


Analysis of a child who developed abnormal neuropsychiatric symptoms after administration of oseltamivir: a case report.
Neuropsychiatric side effects of oseltamivir occur occasionally, especially in infants and young patients, but nothing is known about possible contributory factors. We report a case of a 15-year-old Japanese female with influenza infection who developed abnormal psychiatric symptoms after administration of standard doses of oseltamivir. She had no history of neurological illness, had never previously taken oseltamivir, and had not developed psychiatric reactions during previous influenza infection. Her delirium-like symptoms, including insomnia, visual hallucinations, and a long-term memory deficit, disappeared after cessation of oseltamivir and administration of benzodiazepine. Detailed assessment was performed, including neurological examination (electroencephalogram, brain magnetic resonance imaging, single photon emission computed tomography with 99mTc-ethyl cysteinate dimer and with (123)I-iomazenil, cerebrospinal fluid analysis and glutamate receptor autoantibodies), drug level determination and simulation, and genetic assessment (OAT1, OAT3, CES1, Neu2). Abnormal slowing in the electroencephalogram, which is characteristic of influenza-associated encephalopathy, was not observed in repeated recordings. The serum level determination of active metabolite Ro 64-0802 determined at 154 h after final dosing of oseltamivir was higher than the expected value, suggesting delayed elimination of Ro 64-0802. Thus, abnormal exposure to Ro 64-0802 might have contributed, at least in part, to the development of neuropsychiatric symptoms in this patient. The score on Naranjo's adverse drug reaction probability scale was 6. Mutation of c.122G > A (R41Q) in the sialidase Neu2 gene, increased CSF glutamate receptor autoantibodies, and limbic GABAergic dysfunction indicated by SPECT with (123)I-iomazenil were found as possible contributory factors to the CNS side effects.


Preoperative chemotherapy for stage IIIA non-small cell lung cancer.
Recent studies have shown that preoperative chemotherapy is feasible and advisable for patients with stage IIIA mediastinoscopically confirmed N2 involvement in non-small cell lung cancer. Such studies confirm the crucial pillar of attaining complete resection, and the importance of downstaging N2 disease as a useful surrogate for long-term survival. Resectability rate is about 70% in almost all studies examined. Better means of predicting resectability and relapse are addressed, such as the potential role of positron emission tomography and some hints for the management of replication error-positive non-small cell lung cancer patients. A major issue for the role of preoperative chemotherapy is how not to confuse apples and oranges. Although the role of surgery alone or radiotherapy is considered suboptimal for treatment of stage IIIA N2, whether surgery is required remains to be clarified. Furthermore, large randomized studies of chemoradiation followed by surgery versus chemoradiation alone are needed as well as modification of the current tumor, node, metastasis classification in order to better select those patients for whom preoperative chemotherapy would be beneficial. The surgeon's judgment and skill are essential in the identification of eligible patients and multimodality treatment research is also spreading toward early disease and some subsets of T4 non-small cell lung cancers.


Association between stress and breast cancer in women: a meta-analysis.
The objective of the current meta-analysis was to verify the association between stressful life events and primary breast cancer incidence in women. A total of 618 studies from 1982-2007 were found in the PubMed, LILACS, and Cochrane Library databases. Methodological quality was evaluated according to the Downs & Black criteria. Eight studies were selected (six case-controls and two cohorts). The studies were grouped in three analyses, two of which based on the categories widowhood and divorce and the other based on self-rated intensity and frequency of stressful events. Relative risks were: widowhood 1.04 (95%CI: 0.75-1.44; p = 0.800); divorce 1.03 (95%: 0.72-1.48; p = 0.850); and intensity/frequency of stress 1.73 (95%CI: 0.98-3.05; p = 0.059). We conclude that stressful life events as a whole are not associated with risk of breast cancer in women. However, it is not possible to rule out high-intensity stress as a risk factor for breast cancer.


Metabolism-mediated interaction potential of standardized extract of Tinospora cordifolia through rat and human liver microsomes.
Tinospora cordifolia is used for treatment of several diseases in Indian system of medicine. In the present study, the inhibition potential of T. cordifolia extracts and its constituent tinosporaside to cause herb-drug interactions through rat and human liver cytochrome enzymes was evaluated. Bioactive compound was quantified through reverse phase high-performance liquid chromatography, to standardize the plant extracts and interaction potential of standardized extract. Interaction potential of the test sample was evaluated through cytochrome P450-carbon monoxide complex (CYP450-CO) assay with pooled rat liver microsome. Influence on individual recombinant human liver microsomes such as CYP3A4, CYP2D6, CYP2C9, and CYP1A2 isozymes was analyzed through fluorescence microplate assay, and respective IC50 values were determined. The content of tinosporaside was found to be 1.64% (w/w) in T. cordifolia extract. Concentration-dependent inhibition was observed through T. cordifolia extract. Observed IC50 (μg/ml) value was 136.45 (CYP3A4), 144.37 (CYP2D6), 127.55 (CYP2C9), and 141.82 (CYP1A2). Tinosporaside and extract showed higher IC50 (μg/ml) value than the known inhibitors. T. cordifolia extract showed significantly less interaction potential and indicates that the selected plant has not significant herb-drug interactions relating to the inhibition of major CYP450 isozymes. Plant extract showed significantly higher IC50 value than respective positive inhibitors against CYP3A4, 2D6, 2C9, and 1A2 isozymes. Consumption of T. cordifolia may not cause any adverse effects when consumed along with other xenobiotics.


Comparison of azithromycin and erythromycin in the treatment of mycoplasma pneumonia in children.
To study and compare the clinical effects of azithromycin and erythromycin on children with mycoplasma pneumonia. Total 132 children with mycoplasma pneumonia who were admitted to our hospital between November 2017 and September 2018 were selected as the research subjects. All the children were divided into an observation group and a control group according to random number table, 66 each. The observation group was treated with azithromycin, while the control group was treated with erythromycin. The therapeutic effect, incidence of adverse reactions and disappearance time of clinical symptoms were compared between the two groups. The total efficacy of the observation group was 98.04%, and that of the control group was 74.51%; there was a significant difference (X2=7.184, P=0.007). The incidence of adverse reactions in the observation group was 15.69%, significantly lower than that in the control group (41.18%) (X2=6.376, P=0.002). The disappearance of fever, cough, rale and X ray shadow of the observation group was significantly earlier than that of the control group, and the difference was statistically significant (P<0.05). Compared with erythromycin, azithromycin is more effective in the treatment of mycoplasma pneumonia in children. Azithromycin can further shorten the improvement time of clinical symptoms and signs and has few adverse reactions and high safety. It is worth clinical application.


Arginine and asthma.
Recent studies suggest that alterations of the arginine metabolome and a dysregulation of nitric oxide (NO) homeostasis play a role in the pathogenesis of asthma. L-Arginine, a semi-essential amino acid, is a common substrate for both the arginases and NO synthase (NOS) enzyme families. NO is an important vasodilator of the bronchial circulation, with both bronchodilatory and anti-inflammatory properties, and is synthesized from oxidation of its obligate substrate L-arginine, which is catalyzed by a family of NOS enzymes. Arginase is an essential enzyme in the urea cycle, responsible for the conversion of arginine to ornithine and urea. The NOS and arginase enzymes can be expressed simultaneously under a wide variety of inflammatory conditions, resulting in competition for their common substrate. Although much attention has been directed towards measurements of exhaled NO in asthma, accumulating data show that low bioavailability of L-arginine also contributes to inflammation, hyperresponsiveness and remodeling of the asthmatic airway. Aberrant arginine catabolism represents a novel asthma paradigm that involves excess arginase activity, elevated levels of asymmetric dimethyl arginine, altered intracellular arginine transport, and NOS dysfunction. Addressing the alterations in arginine metabolism may result in new strategies for treatment of asthma.


Meta-analysis using a novel database, miRStress, reveals miRNAs that are frequently associated with the radiation and hypoxia stress-responses.
Organisms are often exposed to environmental pressures that affect homeostasis, so it is important to understand the biological basis of stress-response. Various biological mechanisms have evolved to help cells cope with potentially cytotoxic changes in their environment. miRNAs are small non-coding RNAs which are able to regulate mRNA stability. It has been suggested that miRNAs may tip the balance between continued cytorepair and induction of apoptosis in response to stress. There is a wealth of data in the literature showing the effect of environmental stress on miRNAs, but it is scattered in a large number of disparate publications. Meta-analyses of this data would produce added insight into the molecular mechanisms of stress-response. To facilitate this we created and manually curated the miRStress database, which describes the changes in miRNA levels following an array of stress types in eukaryotic cells. Here we describe this database and validate the miRStress tool for analysing miRNAs that are regulated by stress. To validate the database we performed a cross-species analysis to identify miRNAs that respond to radiation. The analysis tool confirms miR-21 and miR-34a as frequently deregulated in response to radiation, but also identifies novel candidates as potentially important players in this stress response, including miR-15b, miR-19b, and miR-106a. Similarly, we used the miRStress tool to analyse hypoxia-responsive miRNAs. The most frequently deregulated miRNAs were miR-210 and miR-21, as expected. Several other miRNAs were also found to be associated with hypoxia, including miR-181b, miR-26a/b, miR-106a, miR-213 and miR-192. Therefore the miRStress tool has identified miRNAs with hitherto unknown or under-appreciated roles in the response to specific stress types. The miRStress tool, which can be used to uncover new insight into the biological roles of miRNAs, and also has the potential to unearth potential biomarkers for therapeutic response, is freely available at http://mudshark.brookes.ac.uk/MirStress.


Usefulness of sonographic guidance during percutaneous biopsy of mesenteric masses.
The objective of our study was to determine the usefulness of sonographic guidance for biopsy of mesenteric masses. Twenty-five sonographically guided percutaneous biopsies of mesenteric masses were performed in 23 patients. Biopsies were performed with an 18-, 20-, or 22-gauge self-aspirating needle or core biopsy device. Final pathology results and patient medical records were reviewed for biopsy accuracy and complications. A biopsy was considered successful if a specific benign or malignant diagnosis was rendered by the pathologist or if surgical-pathologic confirmation was obtained. Open surgical biopsy was performed after sonographically guided biopsy in 13 patients and led to 12 concordant diagnoses (nine true-positives and three true-negatives) and one discordant diagnosis (false-negative). Specific pathologic diagnosis was rendered for the 10 percutaneous biopsies that were not confirmed by surgical biopsy: five biopsies matched known primary malignancies, consistent with metastases; four biopsies revealed primary tumors, and one biopsy revealed chronic inflammation (nine true-positives and one true-negative). Two biopsies were nondiagnostic because of insufficient material (n = 1) and necrotic tumor (n = 1). In the biopsies with diagnostic tissue specimens, sonographically guided biopsy achieved a sensitivity of 95% (18/19) and specificity of 100% (4/4) for allowing neoplastic tissue to be distinguished from nonneoplastic tissue. Complications included a mesenteric hematoma and abdominal wall cellulitis. Percutaneous biopsy of mesenteric masses is a useful and safe procedure.


A personalised medicine approach for ponatinib-resistant chronic myeloid leukaemia.
Chronic myeloid leukaemia (CML) is characterised by the presence of a fusion driver oncogene, BCR-ABL1, which is a constitutive tyrosine kinase. Tyrosine kinase inhibitors (TKIs) are the central treatment strategy for CML patients and have significantly improved survival rates, but the T315I mutation in the kinase domain of BCR-ABL1 confers resistance to all clinically approved TKIs, except ponatinib. However, compound mutations can mediate resistance even to ponatinib and remain a clinical challenge in CML therapy. Here, we investigated a ponatinib-resistant CML patient through whole-genome sequencing (WGS) to identify the cause of resistance and to find alternative therapeutic targets. We carried out WGS on a ponatinib-resistant CML patient and demonstrated an effective combination therapy against the primary CML cells derived from this patient in vitro. Our findings demonstrate the emergence of compound mutations in the BCR-ABL1 kinase domain following ponatinib treatment, and chromosomal structural variation data predicted amplification of BCL2. The primary CD34(+) CML cells from this patient showed increased sensitivity to the combination of ponatinib and ABT-263, a BCL2 inhibitor with a negligible effect against the normal CD34(+) cells. Our results show the potential of personalised medicine approaches in TKI-resistant CML patients and provide a strategy that could improve clinical outcomes for these patients.


Selection of probiotic bacteria for prevention of allergic diseases: immunomodulation of neonatal dendritic cells.
Modification of intestinal microbiota early in life by administration of probiotic bacteria may be a potential approach to prevent allergic disease. To select probiotic bacteria for in vivo purposes, we investigated the capacity of probiotic bacteria to interact with neonatal dendritic cells (DC) and studied the ensuing T cell polarizing effect. Immature DC were generated from cord blood-derived monocytes and maturation was induced by maturation factors (MF), lipopolysaccharide (LPS) plus MF and Bifidobacterium bifidum, B. infantis, Lactobacillus salivarius, Lactococcus lactis alone or combined with MF. After 12 days of co-culture with DC and Staphylococcus aureus enterotoxin B (SEB) as antigenic stimulus, cytokine production by autologous T cells was determined by intracellular cytokine staining. Additionally, cells were stimulated with CD3 and CD28 monoclonal antibodies and cytokines were measured in supernatants by multiplex assay. The probiotic strains induced partial maturation of DC. Full maturation of DC was induced for all strains tested when MF was added. The percentage of interleukin (IL)-4 producing T cells was lower in T cell cultures stimulated with B. bifidum matured DC compared to MF and LPS matured DC, which coincided with a higher percentage of interferon (IFN)-gamma-producing T cells. Furthermore, T cells stimulated by B. bifidum matured DC produced significantly more IL-10 compared to MF matured DC. Selected species of the Bifidobacterium genus prime in vitro cultured neonatal DC to polarize T cell responses and may therefore be candidates to use in primary prevention of allergic diseases.


Expanding the repertoire of L-DOPA's actions: A comprehensive review of its functional neurochemistry.
Though a multi-facetted disorder, Parkinson's disease is prototypically characterized by neurodegeneration of nigrostriatal dopaminergic neurons of the substantia nigra pars compacta, leading to a severe disruption of motor function. Accordingly, L-DOPA, the metabolic precursor of dopamine (DA), is well-established as a treatment for the motor deficits of Parkinson's disease despite long-term complications such as dyskinesia and psychiatric side-effects. Paradoxically, however, despite the traditional assumption that L-DOPA is transformed in residual striatal dopaminergic neurons into DA, the mechanism of action of L-DOPA is neither simple nor entirely clear. Herein, focussing on its influence upon extracellular DA and other neuromodulators in intact animals and experimental models of Parkinson's disease, we highlight effects other than striatal generation of DA in the functional profile of L-DOPA. While not excluding a minor role for glial cells, L-DOPA is principally transformed into DA in neurons yet, interestingly, with a more important role for serotonergic than dopaminergic projections. Moreover, in addition to the striatum, L-DOPA evokes marked increases in extracellular DA in frontal cortex, nucleus accumbens, the subthalamic nucleus and additional extra-striatal regions. In considering its functional profile, it is also important to bear in mind the marked (probably indirect) influence of L-DOPA upon cholinergic, GABAergic and glutamatergic neurons in the basal ganglia and/or cortex, while anomalous serotonergic transmission is incriminated in the emergence of L-DOPA elicited dyskinesia and psychosis. Finally, L-DOPA may exert intrinsic receptor-mediated actions independently of DA neurotransmission and can be processed into bioactive metabolites. In conclusion, L-DOPA exerts a surprisingly complex pattern of neurochemical effects of much greater scope that mere striatal transformation into DA in spared dopaminergic neurons. Their further experimental and clinical clarification should help improve both L-DOPA-based and novel strategies for controlling the motor and other symptoms of Parkinson's disease.


Smoke-Free Policies in the Workplace and in the Home among American Indians.
American Indians are more likely to smoke, less likely to have smoke-free homes, and potentially less likely to have worksite smoke-free policies. We examined correlates of smoke-free policies at home and work among a community-based sample of American Indians in the Midwest. We examined correlates of smoke-free policies at home and work in a sample of American Indians in the Midwest using a community-based participatory research approach. 66.7% were nonsmokers, 15.6% smoked on some days, and 17.6% smoked every day. The majority (72.4%) had complete smoke-free home policies, 13.1% had partial restrictions, and 14.5% had no rules. Moreover, 62.7% had complete smoke-free worksite policies, 27.9% had partial policies, and 9.4% had no worksite smoke-free policies. Factors associated with having a complete smoke-free home policy included being a college graduate (p=.005) and a nonsmoker versus a nondaily (p=.006) or a daily smoker (p<.001). Correlates of having a complete smoke-free worksite policy included being female (p=.005) and a nonsmoker versus a nondaily (p=.03) or a daily smoker (p<.001). Having complete worksite policies was associated with having smoke-free homes (p<.001). Having complete worksite policies was related to having smoke-free home policies; both were associated with being a nonsmoker.


The relationship between prior night's sleep and measures of infant imitation.
We examined whether sleep quality during the night and naps during the day preceding a learning event are related to memory encoding in human infants. Twenty-four 6- and twenty-four 12-month-old infants' natural sleeping behavior was monitored for 24 hr using actigraphy. After the recording period, encoding was assessed using an imitation paradigm. In an initial baseline phase, infants were allowed to interact with the stimulus to assess spontaneous production of any target actions. Infants then watched an experimenter demonstrate a sequence of three target actions and were immediately given the opportunity to reproduce the demonstrated target actions to assess memory encoding. Analyses revealed significant correlations between nighttime sleep quality variables (sleep efficiency, sleep fragmentation) and immediate imitation in 6-month-olds, but not in 12-month-olds. High sleep quality in the preceding night was thus positively associated with next day's memory encoding in 6-month-old infants.


Immune checkpoint blockade as a potential therapeutic target: surveying CNS malignancies.
Expression of programmed cell death protein 1 (PD-1)/programmed death ligand 1 (PD-L1) across glioma grades is undocumented, and their interactions with commonly expressed genetic and epigenetic alterations are undefined but nonetheless highly relevant to combinatorial treatments. Patients with CNS malignancies were profiled by Caris Life Sciences from 2009 to 2016. Immunohistochemistry findings for PD-1 on tumor-infiltrating lymphocytes (TIL) and PD-L1 on tumor cells were available for 347 cases. Next-generation sequencing, pyrosequencing, immunohistochemistry, fragment analysis, and fluorescence in situ hybridization were used to determine isocitrate dehydrogenase 1 (IDH1), phosphatase and tensin homolog (PTEN), and tumor protein 53 mutational status, O(6)-DNA methylguanine-methyltransferase promoter methylation (MGMT-Me) status, PTEN expression, plus epidermal growth factor receptor variant III and 1p/19q codeletion status. PD-1+ TIL expression and grade IV gliomas were significantly positively correlated (odds ratio [OR]: 6.363; 95% CI: 1.263, 96.236)-especially in gliosarcomas compared with glioblastoma multiforme (P = .014). PD-L1 expression was significantly correlated with tumor grade with all PD-L1+ cases (n = 21) being associated with grade IV gliomas. PD-1+ TIL expression and PD-L1 expression were significantly correlated (OR: 5.209; 95% CI: 1.555, 20.144). Mutations of PTEN, tumor protein 53, BRAF, IDH1, and epidermal growth factor receptor or MGMT-Me did not associate with increased intratumoral expression of either PD-1+ TIL or PD-L1 in glioblastoma multiforme even before false discovery rate correction for multiple comparison. Targeting immune checkpoints in combination with other therapeutics based on positive biomarker selection will require screening of large patient cohorts.


Hypothermia versus Normothermia after Out-of-Hospital Cardiac Arrest.
Targeted temperature management is recommended for patients after cardiac arrest, but the supporting evidence is of low certainty. In an open-label trial with blinded assessment of outcomes, we randomly assigned 1900 adults with coma who had had an out-of-hospital cardiac arrest of presumed cardiac or unknown cause to undergo targeted hypothermia at 33°C, followed by controlled rewarming, or targeted normothermia with early treatment of fever (body temperature, ≥37.8°C). The primary outcome was death from any cause at 6 months. Secondary outcomes included functional outcome at 6 months as assessed with the modified Rankin scale. Prespecified subgroups were defined according to sex, age, initial cardiac rhythm, time to return of spontaneous circulation, and presence or absence of shock on admission. Prespecified adverse events were pneumonia, sepsis, bleeding, arrhythmia resulting in hemodynamic compromise, and skin complications related to the temperature management device. A total of 1850 patients were evaluated for the primary outcome. At 6 months, 465 of 925 patients (50%) in the hypothermia group had died, as compared with 446 of 925 (48%) in the normothermia group (relative risk with hypothermia, 1.04; 95% confidence interval [CI], 0.94 to 1.14; P = 0.37). Of the 1747 patients in whom the functional outcome was assessed, 488 of 881 (55%) in the hypothermia group had moderately severe disability or worse (modified Rankin scale score ≥4), as compared with 479 of 866 (55%) in the normothermia group (relative risk with hypothermia, 1.00; 95% CI, 0.92 to 1.09). Outcomes were consistent in the prespecified subgroups. Arrhythmia resulting in hemodynamic compromise was more common in the hypothermia group than in the normothermia group (24% vs. 17%, P<0.001). The incidence of other adverse events did not differ significantly between the two groups. In patients with coma after out-of-hospital cardiac arrest, targeted hypothermia did not lead to a lower incidence of death by 6 months than targeted normothermia. (Funded by the Swedish Research Council and others; TTM2 ClinicalTrials.gov number, NCT02908308.).


Macrophage pro-inflammatory cytokine secretion is enhanced following interaction with autologous platelets.
Macrophages are the dominant phagocyte at sites of wound healing and inflammation, and the cellular and acellular debris encountered by macrophages can have profound effects on their inflammatory profile. Following interaction with apoptotic cells, macrophages are known to switch to an anti-inflammatory phenotype. Activated platelets, however, are also a major component of inflammatory lesions and have been proposed to be pro-inflammatory mediators. In the present study, we tested the hypothesis that macrophage interaction with activated platelets results in an inflammatory response that differs from the response following phagocytosis of apoptotic cells. Human monocyte-derived macrophages (hMDMs) were co-incubated with autologous activated platelets (AAPs) and the platelet-macrophage interaction was examined by electron microscopy and flow cytometry. The cytokines TNF-α, IL-6, and IL-23 were also measured during LPS-activated hMDM co-incubation with AAPs, which was compared to co-incubation with apoptotic lymphocytes. Cytokine secretion was also compared to platelets pre-treated with the gluococorticoid dexamethasone. Macrophages trapped and phagocytized AAPs utilizing a mechanism that was significantly inhibited by the scavenger receptor ligand fucoidan. LPS-induced macrophage secretion of TNF-α, IL-6, and IL-23 was inhibited by co-incubation with apoptotic cells, but enhanced by co-incubation with AAPs. The platelet-dependent enhancement of LPS-induced cytokines could be reversed by pre-loading the platelets with the glucocorticoid dexamethasone. The interaction of human macrophages with autologous platelets results in scavenger-receptor-mediated platelet uptake and enhancement of LPS-induced cytokines. Therefore, the presence of activated platelets at sites of inflammation may exacerbate pro-inflammatory macrophage activation. The possibility of reversing macrophage activation with dexamethasone-loaded platelets is a promising therapeutic approach to treating unresolved inflammation.


Role of small GTPases and alphavbeta5 integrin in Pseudomonas aeruginosa-induced increase in lung endothelial permeability.
Pseudomonas aeruginosa is an opportunistic pathogen that can cause severe pneumonia associated with airspace flooding with protein-rich edema in critically ill patients. The type III secretion system is a major virulence factor and contributes to dissemination of P. aeruginosa. However, it is still unknown which particular bacterial toxin and which cellular pathways are responsible for the increase in lung endothelial permeability induced by P. aeruginosa. Thus, the first objective of this study was to determine the mechanisms by which this species causes an increase in lung endothelial permeability. The results showed that ExoS and ExoT, two of the four known P. aeruginosa type III cytotoxins, were primarily responsible for bacterium-induced increases in protein permeability across the lung endothelium via an inhibition of Rac1 and an activation of the RhoA signaling pathway. In addition, inhibition of the alphavbeta5 integrin, a central regulator of lung vascular permeability, prevented these P. aeruginosa-mediated increases in albumin flux due to endothelial permeability. Finally, prior activation of the stress protein response or adenoviral gene transfer of the inducible heat shock protein Hsp72 also inhibited the damaging effects of P. aeruginosa on the barrier function of lung endothelium. Taken together, these results demonstrate the critical role of the RhoA/alphavbeta5 integrin pathway in mediating P. aeruginosa-induced lung vascular permeability. In addition, activation of the stress protein response with pharmacologic inhibitors of Hsp90 may protect lungs against P. aeruginosa-induced permeability changes.


The ontogeny of mu opiate tolerance and dependence in the rat: antinociceptive and biochemical studies.
The ontogeny of tolerance to mu opiate antinociception and the behavioral and endocrine profiles of the opiate withdrawal syndrome were studied in rats. Animals were treated with saline or an increasing dose regimen of morphine for 5 days (5-25 mg/kg b.i.d. s.c.) and were tested 36 hr later for morphine or sufentanil antinociception in the hot-plate paw-lift test, or withdrawal was precipitated with 5 mg/kg of naloxone 12 hr after the last chronic morphine dose. Twenty-seven-, 20- and 15-day-old rats all developed tolerance as indicated by a rightward shift of the dose-response curve after chronic morphine. Animals treated on days 4 to 8 and tested on day 10 did not develop tolerance to the same chronic dose regimen used in older animals. In contrast to the observed developmental difference in tolerance, both neonatal and weanling rats developed physical dependence after morphine treatment, as evidenced by the presence of withdrawal symptoms after naloxone administration. Withdrawal in weanling rats was characterized by ptosis, piloerection, abnormal posture, forepaw treading, vocalization on touch and mastication. In addition, both serum corticosterone and adrenocorticotropic hormone secretion were increased during passive withdrawal. The behaviors constituting the withdrawal syndrome precipitated in neonates were distinct from those in weanling rats. Spontaneous vocalization, wall climbing, tremor, mouthing and increased locomotion were all observed. As in the older animals, both serum corticosterone and adrenocorticotropic hormone secretion were elevated during passive withdrawal. Tremor also was induced in opiate naive neonates when naloxone (5 mg/kg) was administered 2 hr after a single 25-mg/kg morphine injection. Brain and serum morphine levels and the time course of antinociception were not altered by chronic morphine treatment at any age. Saturation binding assays in brain homogenates indicated that chronic morphine did not produce changes in receptor number or affinity for the antagonist. The shift induced by the guanine nucleotide Gpp(NH)p (10 microM) from high to low affinity on days 27 and 10, respectively, was not altered by chronic morphine treatment. These data indicate that 10-day-old rats are refractory to developing tolerance relative to older animals, and that changes in receptor number or coupling to guanine nucleotide proteins do not accompany tolerance to this regimen.


CD4+ CD56+ lineage negative malignancies: a new entity developed from malignant early plasmacytoid dendritic cells.
The CD4+ CD56+ lin- immunophenotype characterizes rare malignancies, so far considered as arising from the transformation of NK progenitors, and therefore classified as blastic NK-cell leukemia/lymphoma by the WHO committee. Recently it was formally demonstrated that such malignancies do, in fact, develop from plasmacytoid dendritic cells (pDC), according to immunophenotypic and functional criteria. The clinico-biological features of this neoplasm were moreover recently summarized from a large series of 23 patients. The main symptoms at presentation were cutaneous lesions and bone marrow failure, due to invasion by blastic cells, all of which were EBV negative and agranular. Most patients were initially sensitive to chemotherapy regimens, but they rapidly relapsed and died within 3 years. Only 2 allotransplanted patients were long survivors. Recurrent chromosomal aberrations involving chromosomes 5q, 6q, 12p, 13q, 15q and 9 were described and it was characteristic that these were associated in the same cell. In the present review we compared these findings to those in the literature. The concordant characteristics led us to confirm that this neoplasm actually represents a new entity, that we propose to rename early pDC leukemia/lymphoma. The diagnosis is primarily based on a characteristic immunophenotypic profile i.e. CD4+ CD56+ CD3- CD13- CD33- CD19-. Complementary analyses assessing the expression of more specific pDC-related markers showed the cells to be HLA-DR+, CD123high, CD116low, CD45RA+, BDCA-2+ or BDCA-4+. Such complementary investigations are necessary only in the case of an atypical phenotype, in order to confirm a pDC origin and exclude another hematologic disease. This presently regards the expression of CD33 or cytoplasmic CD3e (cyCD3e) and the negativity of CD56.


Adherence to international follow-up guidelines in type 2 diabetes: a longitudinal cohort study in Luxembourg.
Type 2 diabetes is associated with severe micro- and macro-vascular complications. Physicians' and patients' adherence to follow-up guidelines permits postponing or reducing these complications. The objectives were to assess the level of adherence to fundamental follow-up guidelines and determine patients' characteristics associated with this level of adherence in the context of Luxembourg, where no guidelines were implemented. The exhaustive residing population treated for type 2 diabetes in Luxembourg during the 2000-2006 period (N = 21,068). Seven fundamental criteria were extracted from international guidelines (consultation with the treating physician, HbA1c tests, electrocardiogram, retinal, dental, lipid and renal check-ups). The factors associated with the level of adherence to those criteria were identified using a partial proportional odds model. In 2006, despite 90% of the patients consulted at least 4 times their treating physician, only 0.6% completed all criteria; 55.0% had no HbA1c test (-8.6 points since 2000) and 31.1% had a renal check-up (+21.6 points). The sex (OR(male): 0.87 [95%CI, 0.83-0.92]), the nationality (OR(NonEU): 0.64 [0.52-0.78]), the type of antidiabetic treatment (ORoral: 1.48 [1.35-1.63], OR(mixed): 1.35 [1.20-1.52]) and the type of treating physician (ORG-ID: 0.47 [0.42-0.53]) were the main factors associated with the level of adherence in 2006 (3 or more criteria). A large percentage of patients were not provided with a systematic annual follow-up between 2000 and 2006. This study highlighted the necessity to promote guidelines in Luxembourg, education for physicians and to launch a national discussion on a disease management program for diabetic patients.


Anisotropic Alginate Hydrogels Promote Axonal Growth across Chronic Spinal Cord Transections after Scar Removal.
We have previously reported that cell-seeded alginate hydrogels (AHs) with anisotropic capillaries can restore the continuity of the spinal cord and support axonal regeneration in a rat model of acute partial spinal cord transection. Whether similar effects can be found after transplantation into sites of complete chronic spinal cord transections without additional growth-promoting stimuli has not been investigated. We therefore implanted AHs into the cavity of a chronic thoracic transection following scar resection (SR) 4 weeks postinjury and examined electrophysiological and functional recovery as well as regeneration of descending and ascending projections within and beyond the AH scaffold up to 3 months after engraftment. Our results indicate that both electrophysiological conductivity and locomotor function are significantly improved after AH engraftment. SR transiently impairs locomotor function immediately after surgery but does not affect long-term outcomes. Histological analysis shows numerous host cells migrating into the scaffold channels and a reduction of fibroglial scaring around the lesion by AH grafts. In contrast to corticospinal axons, raphaespinal and propriospinal descending axons and ascending sensory axons regenerate throughout the scaffolds and extend into the distal host parenchyma. These results further support the pro-regenerative properties of AHs and their therapeutic potential for chronic SCI in combination with other strategies to improve functional outcomes after spinal cord injury.


Nerve growth factor stimulates clonal growth of human lung cancer cell lines and a human glioblastoma cell line expressing high-affinity nerve growth factor binding sites involving tyrosine kinase signaling.
The growth of a panel of 22 different human tumor, leukemia, and lymphoma cell lines was examined in a human tumor cloning assay in agar or methylcellulose and a tritiated thymidine uptake assay. The cultures were performed in the absence or presence of increasing concentrations (0.5-500 ng/ml) of nerve growth factor (NGF). The growth of 17 of the 22 cell lines was not significantly and reproducibly affected by NGF. There was minor (1.2-fold) but reproducible stimulation of clonal growth in one glioblastoma cell line (86-HG-39) by NGF, but in this cell line NGF induced no growth modulation in a tritiated thymidine uptake assay. However, clonal growth of another glioblastoma cell line (87-HG-31) and all three lung cancer cell lines tested (HTB 119, HTB 120, CCL 185) could be stimulated up to 3-fold by NGF with a dose-response relationship for the growth factor. Growth stimulation by NGF could be completely reversed by neutralizing anti-NGF antibody and by the tyrosine kinase inhibitor genistein. Evaluation of secondary plating efficiency revealed the stimulation of colony formation as representing self-renewal and not terminal differentiation. Reverse transcriptase-PCR experiments in the five responding cell lines showed expression of both low-affinity NGF receptor (glycoprotein 75) and c-trk transcripts on the mRNA level. Of the five responding cell lines, only 86-HG-39, the cell line with the lowest responsiveness, revealed low-affinity NGF receptor on the protein level; the other four cell lines with high responsiveness, including the three lung cancer cell lines, expressed no low-affinity NGF receptor as shown by fluorescence-activated cell sorter analysis and immunoprecipitation using the ME 20.4 antibody. Immunoprecipitation using anti-trk antibodies was negative in all five responding cell lines. However, binding studies with iodinated NGF showed only low-affinity binding on the 86-HG-39 cell line and only high-affinity binding on the high-responder cell lines CCL 185 and 87-HG-31. In summary, our data suggest that NGF can be operative in stimulation of clonal growth of malignant tumor cells. High-affinity but not low-affinity binding sites mediate signal transduction for clonal growth and signaling involves tyrosine kinase activity.


In Situ Focused Beam Reflectance Measurement (FBRM), Attenuated Total Reflectance Fourier Transform Infrared (ATR-FTIR) and Raman Characterization of the Polymorphic Transformation of Carbamazepine.
The objective of this work was to study the polymorphic transformation of carbamazepine from Form II to Form III in 1-propanol during seeded isothermal batch crystallization. First, the pure Form II and Form III were obtained and characterized. Then their solubilities and metastable zone limits were measured by in-situ attenuated total reflectance Fourier transform infrared (ATR-FTIR) spectroscopy and focused beam reflectance measurement (FBRM). A transition temperature at about 34.2 °C was deduced suggesting the enantiotropic nature of this compound over the studied temperature range. To quantify the polymorph ratio during the transformation process, a new in-situ quantitative method was developed to measure the fraction of Form II by Raman spectroscopy. Successful tracking of the nucleation of the stable form and the transformation from Form II to Form III during isothermal crystallization was achieved by Raman spectroscopy and FBRM. The results from these three in-situ techniques, FBRM, FTIR and Raman were consistent with each other. The results showed a strong dependency on the amount of seeds added during isothermal crystallization.


Crystal structure of the Mycobacterium tuberculosis enoyl-ACP reductase, InhA, in complex with NAD+ and a C16 fatty acyl substrate.
Enoyl-ACP reductases participate in fatty acid biosynthesis by utilizing NADH to reduce the trans double bond between positions C2 and C3 of a fatty acyl chain linked to the acyl carrier protein. The enoyl-ACP reductase from Mycobacterium tuberculosis, known as InhA, is a member of an unusual FAS-II system that prefers longer chain fatty acyl substrates for the purpose of synthesizing mycolic acids, a major component of mycobacterial cell walls. The crystal structure of InhA in complex with NAD+ and a C16 fatty acyl substrate, trans-2-hexadecenoyl-(N-acetylcysteamine)-thioester, reveals that the substrate binds in a general "U-shaped" conformation, with the trans double bond positioned directly adjacent to the nicotinamide ring of NAD+. The side chain of Tyr158 directly interacts with the thioester carbonyl oxygen of the C16 fatty acyl substrate and therefore could help stabilize the enolate intermediate, proposed to form during substrate catalysis. Hydrophobic residues, primarily from the substrate binding loop (residues 196-219), engulf the fatty acyl chain portion of the substrate. The substrate binding loop of InhA is longer than that of other enoyl-ACP reductases and creates a deeper substrate binding crevice, consistent with the ability of InhA to recognize longer chain fatty acyl substrates.


Climate change adaptation as a global public good: implications for financing.
Beginning as an afterthought in the UN Framework Convention on Climate Change, adaptation as an agenda has come a long way since 1992. With no ambitious mitigation, recent years have witnessed an increasing frequency of extreme climate events, including cross-border or borderless climate risks. Accordingly, the Paris Agreement frames adaptation as a global goal and global responsibility. However, financing for adaptation continues to remain extremely poor, relative to the estimated needs, even though the regime has obligatory provisions for support by developed countries. Why is this so? Why should the majority of the countries, with an insignificant contribution to causing the problem, suffer from increasing climate impacts? How can adaptation finance be enhanced at scale? As a response to these queries, the paper substantiates three claims: (1) that poor funding can be attributed to the territorial framing under the regime that conceptualizes adaptation largely as a local or national public good and, hence, the inefficacy of market mechanisms, (2) that it makes conceptual and political sense to consider adaptation as a global public good, and (3) that such a reframing should make a difference in boosting adaptation finance. In a multi-polar world with different views on adaptation finance, multilateral agencies should lead in promoting the proposed framing.


Generation of FX-/- and Gmds-/- CHOZN host cell lines for the production of afucosylated therapeutic antibodies.
Antibody-dependent cellular cytotoxicity (ADCC) is the primary mechanism of actions for several marketed therapeutic antibodies (mAbs) and for many more in clinical trials. The ADCC efficacy is highly dependent on the ability of therapeutic mAbs to recruit effector cells such as natural killer cells, which induce the apoptosis of targeted cells. The recruitment of effector cells by mAbs is negatively affected by fucose modification of N-Glycans on the Fc; thus, utilization of afucosylated mAbs has been a trend for enhanced ADCC therapeutics. Most of afucosylated mAbs in clinical or commercial manufacturing were produced from Fut8-/- Chinese hamster ovary cells (CHO) host cells, generally generating low yields compared to wildtype CHO host. This study details the generation and characterization of two engineered CHOZN® cell lines, in which the enzyme involved in guanosine diphosphate (GDP)-fucose synthesis, GDP mannose-4,6-dehydratase (Gmds) and GDP-L-fucose synthase (FX), was knocked out. The top host cell lines for each of the knockouts, FX-/- and Gmds-/-, were selected based on growth robustness, bulk MSX selection tolerance, production titer, fucosylation level, and cell stability. We tested the production of two proprietary IgG1 mAbs in the engineered host cells, and found that the titers were comparable to CHOZN® cells. The mAbs generated from either KO cell line exhibited loss of fucose modification, leading to significantly boosted FcγRIIIa binding and ADCC effects. Our data demonstrated that both FX-/- and Gmds-/- host cells could replace Fut8-/- CHO cells for clinical manufacturing of antibody therapeutics.


Computer-automated tinnitus assessment: noise-band matching, maskability, and residual inhibition.
Psychoacoustic measures of tinnitus typically include loudness and pitch match, minimum masking level (MML), and residual inhibition (RI). We previously developed and documented a computer-automated tinnitus evaluation system (TES) capable of subject-guided loudness and pitch matching. The TES was further developed to conduct computer-aided, subject-guided testing for noise-band matching (NBM), MML, and RI. The purpose of the present study was to document the capability of the upgraded TES to obtain measures of NBM, MML, and RI, and to determine the test-retest reliability of the responses obtained. Three subject-guided, computer-automated testing protocols were developed to conduct NBM. For MML and RI testing, a 2-12 kHz band of noise was used. All testing was repeated during a second session. Subjects meeting study criteria were selected from those who had previously been tested for loudness and pitch matching in our laboratory. A total of 21 subjects completed testing, including seven females and 14 males. The upgraded TES was found to be fairly time efficient. Subjects were generally reliable, both within and between sessions, with respect to the type of stimulus they chose as the best match to their tinnitus. Matching to bandwidth was more variable between measurements, with greater consistency seen for subjects reporting tonal tinnitus or wide-band noisy tinnitus than intermediate types. Between-session repeated MMLs were within 10 dB of each other for all but three of the subjects. Subjects who experienced RI during Session 1 tended to be those who experienced it during Session 2. This study may represent the first time that NBM, MML, and RI audiometric testing results have been obtained entirely through a self-contained, computer-automated system designed specifically for use in the clinic. Future plans include refinements to achieve greater testing efficiency.


A variant octamer motif in a Xenopus H2B histone gene promoter is not required for transcription in frog oocytes.
Xenopus oocytes, arrested in G2 before the first meiotic division, accumulate histone mRNA and protein in the absence of chromosomal DNA replication and therefore represent an attractive biological system in which to examine histone gene expression uncoupled from the cell cycle. Previous studies have shown that sequences necessary for maximal levels of transcription in oocytes are present within 200 bp at the 5' end of the transcription initiation site for genes encoding each of the five major Xenopus histone classes. We have defined by site-directed mutagenesis individual regulatory sequences and characterized DNA-binding proteins required for histone H2B gene transcription in injected oocytes. The Xenopus H2B gene has a relatively simple promoter containing several transcriptional regulatory elements, including TFIID, CBP, and ATF/CREB binding sites, required for maximal transcription. A sequence (CTTTACAT) in the H2B promoter resembling the conserved octamer motif (ATTTGCAT), the target for cell-cycle regulation of a human H2B gene, is not required for transcription in oocytes. Nonetheless, substitution of a consensus octamer motif for the variant octamer element activates H2B transcription. Oocyte factors, presumably including the ubiquitous Oct-1 factor, specifically bind to the consensus octamer motif but not to the variant sequence. Our results demonstrate that a transcriptional regulatory element involved in lymphoid-specific expression of immunoglobulin genes and in S-phase-specific activation of mammalian H2B histone genes can activate transcription in nondividing amphibian oocytes.


Early Psychosis and Trauma-Related Disorders: Clinical Practice Guidelines and Future Directions.
Despite high rates of trauma-related disorders among individuals with early psychosis, no clinical practice guidelines for the treatment of comorbid early psychosis and trauma-related disorders exist to date. Indeed, the routine exclusion of individuals with past and current psychosis from participation in trauma research and practice has limited the accumulation of research that could support such clinical practice guidelines. While preliminary research evidence suggests that traditional, evidence-based treatments for trauma-related disorders can be safely and effectively employed to reduce symptoms of posttraumatic stress and chronic psychosis, it remains unclear whether such treatments are appropriate for individuals in the early stages of psychotic illness. Clinical experts (N = 118) representing 121 early psychosis programs across 28 states were surveyed using the expert consensus method. Forty-nine clinical experts responded and reached consensus on 46 of 49 expert consensus items related to the treatment of comorbid early psychosis and trauma-related disorders. Conjoint or family therapy and individual therapy were rated as treatment approaches of choice. Anxiety or stress management and psychoeducation were rated as treatment interventions of choice for addressing both trauma symptoms and psychotic symptoms. In addition, case management was rated as a treatment intervention of choice for addressing psychotic symptoms. No consensus was reached on expert consensus items regarding the appropriateness of a parallel treatment approach exposure interventions for addressing psychotic symptoms, or sensorimotor or movement interventions for addressing trauma symptoms. In areas where expert consensus exists and is supported by current research, preliminary clinical practice guidelines for the treatment of comorbid early psychosis and trauma-related disorders are offered. In areas where expert consensus does not exist, recommendations for future research are offered. The results of this study are intended to serve as a launching point for scientists and practitioners interested in advancing appropriate treatment for high-risk and underserved individuals with comorbid early psychosis and trauma-related disorders.


Transcriptome meta-analysis of three follicular compartments and its correlation with ovarian follicle maturity and oocyte developmental competence in cows.
Oocyte developmental competence in superstimulated cows is dependent in part on the duration of the FSH coasting. FSH coasting refers to superstimulation with FSH (2 days of endogenous FSH following follicle ablation and 3 days of FSH injections) followed by no FSH for a specific duration. The optimal duration varies among individuals. FSH coasting appears to modulate the transcriptome of different follicular compartments, which cooperate as a single functional unit. However, the integrative effects of FSH coasting on different follicular compartments remain ambiguous. Meta-analysis of three independent transcriptome studies, each focused on a single cell type (granulosa, cumulus, and oocyte) during FSH coasting, allowed the identification of 12 gene clusters with similar time-course expression patterns in all three compartments. Network analysis identified HNF4A (involved in metabolic functions) and ELAVL1 (an RNA-binding protein) as hub genes regulated respectively upward and downward in the clusters enriched at the optimal coasting time, and APP (involved in mitochondrial functions) and COPS5 (a member of the COP9 signalosome) as hub genes regulated respectively upwards and downwards in the clusters enriched progressively throughout the coasting period. We confirmed the effects on HNF4A downstream targets (TTR, PPL) and other hub genes (ELAVL1, APP, MYC, and PGR) in 30 cows with RT-quantitative PCR. The correlation of hub gene expression levels with FSH coasting indicated that a combination of these genes could predict oocyte competence with 83% sensitivity, suggesting that they are potential biomarkers of follicle differentiation. These findings could be used to optimize FSH coasting on an individual basis.


Longitudinal Variations in Antibody Responses against SARS-CoV-2 Spike Epitopes upon Serial Vaccinations.
The COVID-19 pandemic caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) impacted healthcare, the workforce, and worldwide socioeconomics. Multi-dose mono- or bivalent mRNA vaccine regimens have shown high efficacy in protection against SARS-CoV-2 and its emerging variants with varying degrees of efficacy. Amino acid changes, primarily in the receptor-binding domain (RBD), result in selection for viral infectivity, disease severity, and immune evasion. Therefore, many studies have centered around neutralizing antibodies that target the RBD and their generation achieved through infection or vaccination. Here, we conducted a unique longitudinal study, analyzing the effects of a three-dose mRNA vaccine regimen exclusively using the monovalent BNT162b2 (Pfizer/BioNTech) vaccine, systematically administered to nine previously uninfected (naïve) individuals. We compare changes in humoral antibody responses across the entire SARS-CoV-2 spike glycoprotein (S) using a high-throughput phage display technique (VirScan). Our data demonstrate that two doses of vaccination alone can achieve the broadest and highest magnitudes of anti-S response. Moreover, we present evidence of novel highly boosted non-RBD epitopes that strongly correlate with neutralization and recapitulate independent findings. These vaccine-boosted epitopes could facilitate multi-valent vaccine development and drug discovery.


Mars: The Origin of the 3.58- and 3.69-Micron Minima in the Infrared Spectra.
The 3- to 4-micron spectra of Mars, recorded with the 200-inch telescope in 1958, were reexamined in order to ascertain whether the minima at 3.58 and 3.69 microns are due to telluric HDO molecules. Solar spectra obtained at Denver and water vapor abundances derived from radiosonde flights during the observing period were used. There seems to be a correlation between the intensities of the 3.58- and 3.69-micron features and the amount of telluric water vapor in the optical path. An important corollary is that there is no evidence for attributing these spectral features to Mars.


Biochemical Characterization of Mouse Retina of an Alzheimer's Disease Model by Raman Spectroscopy.
The presence of biomarkers characteristic for Alzheimer's disease in the retina is a controversial topic. Raman spectroscopy offers information on the biochemical composition of tissues. Thus, it could give valuable insight into the diagnostic value of retinal analysis. Within the present study, retinas of a double transgenic mouse model, that expresses a chimeric mouse/human amyloid precursor protein and a mutant form of human presenilin 1, and corresponding control group were subjected to ex vivo Raman imaging. The Raman data recorded on cross sections of whole eyes highlight the layered structure of the retina in a label-free manner. Based on the Raman information obtained from en face mounted retina samples, a discrimination between healthy and Alzheimer's disease retinal tissue can be done with an accuracy of 85.9%. For this a partial least squares-linear discriminant analysis was applied. Therefore, although no macromolecular changes in form of, i.e., amyloid beta plaques, can be noticed based on Raman spectroscopy, subtle biochemical changes happening in the retina could lead to Alzheimer's disease identification.


Pseudotumours associated with metal-on-metal hip resurfacings.
We report 17 patients (20 hips) in whom metal-on-metal resurfacing had been performed and who presented with various symptoms and a soft-tissue mass which we termed a pseudotumour. Each patient underwent plain radiography and in some, CT, MRI and ultrasonography were also performed. In addition, histological examination of available samples was undertaken. All the patients were women and their presentation was variable. The most common symptom was discomfort in the region of the hip. Other symptoms included spontaneous dislocation, nerve palsy, a noticeable mass or a rash. The common histological features were extensive necrosis and lymphocytic infiltration. To date, 13 of the 20 hips have required revision to a conventional hip replacement. Two are awaiting revision. We estimate that approximately 1% of patients who have a metal-on-metal resurfacing develop a pseudotumour within five years. The cause is unknown and is probably multifactorial. There may be a toxic reaction to an excess of particulate metal wear debris or a hypersensitivity reaction to a normal amount of metal debris. We are concerned that with time the incidence of these pseudotumours may increase. Further investigation is required to define their cause.


Antiepileptogenic Effect of Subchronic Palmitoylethanolamide Treatment in a Mouse Model of Acute Epilepsy.
Research on the antiepileptic effects of (endo-)cannabinoids has remarkably progressed in the years following the discovery of fundamental role of the endocannabinoid (eCB) system in controlling neural excitability. Moreover, an increasing number of well-documented cases of epilepsy patients exhibiting multi-drug resistance report beneficial effects of cannabis use. Pre-clinical and clinical research has increasingly focused on the antiepileptic effectiveness of exogenous administration of cannabinoids and/or pharmacologically induced increase of eCBs such as anandamide (also known as arachidonoylethanolamide [AEA]). Concomitant research has uncovered the contribution of neuroinflammatory processes and peripheral immunity to the onset and progression of epilepsy. Accordingly, modulation of inflammatory pathways such as cyclooxygenase-2 (COX-2) was pursued as alternative therapeutic strategy for epilepsy. Palmitoylethanolamide (PEA) is an endogenous fatty acid amide related to the centrally and peripherally present eCB AEA, and is a naturally occurring nutrient that has long been recognized for its analgesic and anti-inflammatory properties. Neuroprotective and anti-hyperalgesic properties of PEA were evidenced in neurodegenerative diseases, and antiepileptic effects in pentylenetetrazol (PTZ), maximal electroshock (MES) and amygdaloid kindling models of epileptic seizures. Moreover, numerous clinical trials in chronic pain revealed that PEA treatment is devoid of addiction potential, dose limiting side effects and psychoactive effects, rendering PEA an appealing candidate as antiepileptic compound or adjuvant. In the present study, we aimed at assessing antiepileptic properties of PEA in a mouse model of acute epileptic seizures induced by systemic administration of kainic acid (KA). KA-induced epilepsy in rodents is assumed to resemble to different extents human temporal lobe epilepsy (TLE) depending on the route of KA administration; intracerebral (i.c.) injection was recently shown to most closely mimic human TLE, while systemic KA administration causes more widespread pathological damage, both in brain and periphery. To explore the potential of PEA to exert therapeutic effects both in brain and periphery, acute and subchronic administration of PEA by intraperitoneal (i.p.) injection was assessed on mice with systemically administered KA. Specifically, we investigated: (i) neuroprotective and anticonvulsant properties of acute and subchronic PEA treatment in KA-induced seizure models, and (ii) temporal dynamics of eCB and eicosanoid (eiC) levels in hippocampus and plasma over 180 min post seizure induction in PEA-treated and non-treated KA-injected mice vs. vehicle injected mice. Finally, we compared the systemic PEA treatment with, and in combination with, pharmacological blockade of fatty acid amide hydrolase (FAAH) in brain and periphery, in terms of anticonvulsant properties and modulation of eCBs and eiCs. Here, we demonstrate that subchronic administration of PEA significantly alleviates seizure intensity, promotes neuroprotection and induces modulation of the plasma and hippocampal eCB and eiC levels in systemic KA-injected mice.


A life where concealment and exposure coexist-The antiretroviral therapy adherence of people living with HIV infection: A phenomenological study.
The aim of this study is to identify the nature and structure of the real world experiences of people living with HIV infection and adhering to antiretroviral therapy (ART). Giorgi's phenomenological method was applied. Data collection was conducted from 1 November 2016-1 September 2017. A total of six participants participated in the study. The Consolidated Criteria for Reporting Qualitative Research (COREQ) was used to report this study. The findings show that the structure of the experience of people living with HIV and adhering to ART is composed of six essential common constituents: (a) ART adherence has become a prerequisite for a normal life; (b) Endless adjustments responding to a sensitive body; (c) Dosing Time becomes the framework of the day; (d) Dosing time needing to coincide with the availability of confidential dosing space; (e) Classification of Relationships-those who know of my taking antiretroviral drugs and those who do not; and (f) Recognition of the multiple relationships affecting ART. Adhering to ART is experienced by people infected with HIV not only as a process they want to hide, but also a critical lifeline that stabilizes their lives. That is, their day-to-day living is split into a double life reflecting their coexisting need for concealment and exposure. Although much existing literature focuses on the adherence to medication, this study reveals the meaning of ART in the context of viewing the patient as an independent subject. Contrary with what people living with HIV want, they are at risk of external exposure during the process of treatment. This study highlights the need for nurses to communicate with patients about the strategies they need to meet the challenges they face.


Prognostic significance of the treadmill exercise test performance 6 months after myocardial infarction.
A submaximal treadmill exercise test performed before hospital discharge after an uncomplicated myocardial infarction is often utilized to estimate prognosis and guide management, but there is little experience with a maximal exercise test performed 6 months after infarction to identify prognosis later in the convalescent period. The performance characteristics during an exercise test 6 months after myocardial infarction were related to the development of death, recurrent nonfatal myocardial infarction and coronary artery bypass surgery in the subsequent 12 months (that is, 6 to 18 months after infarction) in 473 patients. Mortality was significantly greater in patients who exhibited any of the following: inability to perform the exercise test because of cardiac limitations, the development of ST segment elevation of 1 mm or greater during the exercise test, an inadequate blood pressure response during exercise, the development of any ventricular premature depolarizations during exercise or the recovery period and inability to exercise beyond stage I of the modified Bruce protocol. By utilizing a combination of four high risk prognostic features from the exercise test, it was possible to stratify patients in terms of risk of mortality, from 1% if none of these features were present to 17% if three or four were present. Recurrent nonfatal myocardial infarction was predicted by an inability to perform the exercise test because of cardiac limitations, but not by any characteristics of exercise test performance. Coronary artery bypass surgery was associated with the development of ST segment depression of 1 mm or greater during the exercise test. Although clinical evidence of angina and heart failure 6 months after infarction was predictive of subsequent mortality among all survivors, among the low risk group without severely limiting cardiac disease, the exercise test provided unique prognostic information not available from clinical assessment alone. Therefore, a maximal exercise test performed 6 months after myocardial infarction is a valuable, noninvasive tool to evaluate prognosis. It provides information that is independent of and additive to clinical evaluation performed at the same time.


Relax, you're soaking in it: sources of information about infant formula.
Although the advertising of infant and follow-on formula products in Australia is prohibited by the Marketing in Australia of Infant Formulas: Manufacturers and Importers Agreement (1992), toddler milk is advertised without restriction. Recent research suggests that Australian mothers perceive advertisements for toddler milk to also be advertisements for infant formula. Furthermore, they tend to accept the messages they encounter in these advertisements uncritically. This study used established qualitative market research strategies to investigate what mothers, and those who influence mothers, know about formula milk products. This included exploration of commonly used sources of information, how toddler milk advertisements are interpreted and how the claims made in these advertisements are evaluated. Eight interviews were conducted: two individual interviews (involving a General Practitioner and a Community Dietitian) and six discussion groups (two groups of two mothers, two groups of two grandmothers, one group of two child and family health nurses and one group of five Child and Family Health Nurses). The results suggest that mothers seek advice about formula milk products from health professionals, their friends and their own mothers. The responses also suggest that all of these groups understand toddler milk advertisements to be advertising formula milk products and tend to rely on the messages contained in them to inform their decisions or advice about infant feeding products. It may therefore be difficult for mothers to access independent information upon which to base their decisions about infant feeding products. Further research is needed on whether sufficient independent information about infant feeding products is available to health professionals and mothers.


Upregulation of PD-1 expression on HIV-specific CD8+ T cells leads to reversible immune dysfunction.
The engagement of programmed death 1 (PD-1) to its ligands, PD-L1 and PD-L2, inhibits proliferation and cytokine production mediated by antibodies to CD3 (refs. 5,6,7). Blocking the PD-1-PD-L1 pathway in mice chronically infected with lymphocytic choriomeningitis virus restores the capacity of exhausted CD8(+) T cells to undergo proliferation, cytokine production and cytotoxic activity and, consequently, results in reduced viral load. During chronic HIV infection, HIV-specific CD8(+) T cells are functionally impaired, showing a reduced capacity to produce cytokines and effector molecules as well as an impaired capacity to proliferate. Here, we found that PD-1 was upregulated on HIV-specific CD8(+) T cells; PD-1 expression levels were significantly correlated both with viral load and with the reduced capacity for cytokine production and proliferation of HIV-specific CD8(+) T cells. Notably, cytomegalovirus (CMV)-specific CD8(+) T cells from the same donors did not upregulate PD-1 and maintained the production of high levels of cytokines. Blocking PD-1 engagement to its ligand (PD-L1) enhanced the capacity of HIV-specific CD8(+) T cells to survive and proliferate and led to an increased production of cytokines and cytotoxic molecules in response to cognate antigen. The accumulation of HIV-specific dysfunctional CD8(+) T cells in the infected host could prevent the renewal of a functionally competent HIV-specific CD8(+) repertoire.


Effects of antisocial behaviour on opponent's anger, attention, and performance.
Sledging, which is verbal antisocial behaviour in sport, aims to impair an opponent's performance. Previously, variations in performance have been attributed to changes in emotion and cognition. To improve our understanding of sledging, the current experiment examined the effects of verbal antisocial behaviour on anger, attention and performance. Participants performed a competitive basketball free-throw shooting task under insult (verbal behaviour designed to offend and upset the performer), distraction (verbal behaviour designed to draw attention away from the task), or control (neutral verbal behaviour) conditions. Performance was assessed by the number of successful baskets and a points-based scoring system, while anger and attention were measured post-task. The insult condition provoked more anger than the control and distraction conditions, whereas the insult and distraction conditions increased distraction and reduced self-focus compared to the control condition. Although verbal antisocial behaviour had no overall direct effect on performance, mediation analysis showed that anger indirectly impaired performance via distraction. Implications for the antisocial behaviour-performance relationship are discussed.


Factors associated with the development of self-harm amongst a socio-economically deprived cohort of adolescents in Santiago, Chile.
Studies carried out in the West indicate that the incidence of self-harm (SH) is particularly high amongst adolescents, but few studies have investigated its incidence and aetiology in low-income countries. The purpose of this study was to investigate risk factors associated with new onset episodes of SH, amongst Chilean adolescents from low socio-economic backgrounds. Prospective cohort study nested within a cluster randomised controlled trial. A 6-month follow-up for 2,042 adolescents, median age 14 years, from socio-economically deprived areas of Santiago, Chile. The lifetime prevalence of SH was 23%. The incidence rate of SH at 6 months was 14% amongst those reporting no SH at baseline. In multivariable analyses, risk factors for incident SH include depressive symptoms, suicidal thoughts, poor problem-solving skills and cannabis misuse. The prevalence and incidence of SH in this socio-economically deprived sample differed highly according to gender. Poor problem-solving skills, suicidal thoughts, and cannabis misuse were associated with onset of SH.


Understanding the effect of magnesium ion concentration on the catalytic activity of ribonuclease H through computation: does a third metal binding site modulate endonuclease catalysis?
Ribonuclease H (RNase H) belongs to the nucleotidyl-transferase superfamily and hydrolyzes the phosphodiester linkage on the RNA strand of a DNA/RNA hybrid duplex. Due to its activity in HIV reverse transcription, it represents a promising target for anti-HIV drug design. While crystallographic data have located two ions in the catalytic site, there is ongoing debate concerning just how many metal ions bound at the active site are optimal for catalysis. Indeed, experiments have shown a dependency of the catalytic activity on the Mg(2+) concentration. Moreover, in RNase H, the glutamate residue E188 has been shown to be essential for full enzymatic activation, regardless of the Mg(2+) concentration. The catalytic center is known to contain two Mg(2+) ions, and E188 is not one of the primary metal ligands. Herein, classical molecular dynamics (MD) simulations are employed to study the metal-ligand coordination in RNase H at different concentration of Mg(2+). Importantly, the presence of a third Mg(2+) ion, bound to the second-shell ligand E188, is a persistent feature of the MD simulations. Free energy calculations have identified two distinct conformations, depending on the concentration of Mg(2+). At standard concentration, a third Mg(2+) is found in the catalytic pocket, but it does not perturb the optimal RNase H active conformation. However, at higher concentration, the third Mg(2+) ion heavily perturbs the nucleophilic water and thereby influences the catalytic efficiency of RNase H. In addition, the E188A mutant shows no ability to engage additional Mg(2+) ions near the catalytic pocket. This finding likely explains the decrease in catalytic activity of E188A and also supports the key role of E188 in localizing the third Mg(2+) ion at the active site. Glutamate residues are commonly found surrounding the metal center in the endonuclease family, which suggests that this structural motif may be an important feature to enhance catalytic activity. The present MD calculations support the hypothesis that RNase H can accommodate three divalent metal ions in its catalytic pocket and provide an in-depth understanding of their dynamic role for catalysis.


The structure of the TRAPP subunit TPC6 suggests a model for a TRAPP subcomplex.
The TRAPP (transport protein particle) complexes are tethering complexes that have an important role at the different steps of vesicle transport. Recently, the crystal structures of the TRAPP subunits SEDL and BET3 have been determined, and we present here the 1.7 Angstroms crystal structure of human TPC6, a third TRAPP subunit. The protein adopts an alpha/beta-plait topology and forms a dimer. In spite of low sequence similarity, the structure of TPC6 strikingly resembles that of BET3. The similarity is especially prominent at the dimerization interfaces of the proteins. This suggests heterodimerization of TPC6 and BET3, which is shown by in vitro and in vivo association studies. Together with TPC5, another TRAPP subunit, TPC6 and BET3 are supposed to constitute a family of paralogous proteins with closely similar three-dimensional structures but little sequence similarity among its members.


Simple Hyper-Heuristics Control the Neighbourhood Size of Randomised Local Search Optimally for LeadingOnes.
Selection hyper-heuristics (HHs) are randomised search methodologies which choose and execute heuristics during the optimisation process from a set of low-level heuristics. A machine learning mechanism is generally used to decide which low-level heuristic should be applied in each decision step. In this article, we analyse whether sophisticated learning mechanisms are always necessary for HHs to perform well. To this end we consider the most simple HHs from the literature and rigorously analyse their performance for the LeadingOnes benchmark function. Our analysis shows that the standard Simple Random, Permutation, Greedy, and Random Gradient HHs show no signs of learning. While the former HHs do not attempt to learn from the past performance of low-level heuristics, the idea behind the Random Gradient HH is to continue to exploit the currently selected heuristic as long as it is successful. Hence, it is embedded with a reinforcement learning mechanism with the shortest possible memory. However, the probability that a promising heuristic is successful in the next step is relatively low when perturbing a reasonable solution to a combinatorial optimisation problem. We generalise the "simple" Random Gradient HH so success can be measured over a fixed period of time τ, instead of a single iteration. For LeadingOnes we prove that the Generalised Random Gradient (GRG) HH can learn to adapt the neighbourhood size of Randomised Local Search to optimality during the run. As a result, we prove it has the best possible performance achievable with the low-level heuristics (Randomised Local Search with different neighbourhood sizes), up to lower-order terms. We also prove that the performance of the HH improves as the number of low-level local search heuristics to choose from increases. In particular, with access to k low-level local search heuristics, it outperforms the best-possible algorithm using any subset of the k heuristics. Finally, we show that the advantages of GRG over Randomised Local Search and Evolutionary Algorithms using standard bit mutation increase if the anytime performance is considered (i.e., the performance gap is larger if approximate solutions are sought rather than exact ones). Experimental analyses confirm these results for different problem sizes (up to n=108) and shed some light on the best choices for the parameter τ in various situations.


Altered fatty acid membrane composition modifies lymphocyte localization in vivo.
In the present paper, we report the results of a study on the in vivo localization of 51Cr-labeled lymphocytes with an altered lipid bilayer. In vitro treatment of lymphocytes with fatty acids (arachidic and linolenic acids) modifies the relative composition of plasma membrane fatty acids. Phospholipids of the plasma membrane of lymphocytes incubated with arachidic acid show a preferential increase of fatty acids with chain length between C:12 and C:16. Cells incubated with linolenic acid show an increase percentage of fatty acids C:16 to C:20 and the relative amount of the fatty acids with chain length superior to C:20 is higher in cells treated with linolenic than with arachidic acid. We have found that these alterations in plasma membrane fatty acid composition can modify the normal pattern of lymphocyte localization in vivo after iv transfer into syngeneic hosts. The possible role of factors such as cell to cell adhesion and/or fluidity of plasma membranes in the control of lymphocyte migration are discussed.


Epigenetic mechanisms underlying the dynamic expression of cancer-testis genes, PAGE2, -2B and SPANX-B, during mesenchymal-to-epithelial transition.
Cancer-testis (CT) genes are expressed in various cancers but not in normal tissues other than in cells of the germline. Although DNA demethylation of promoter-proximal CpGs of CT genes is linked to their expression in cancer, the mechanisms leading to demethylation are unknown. To elucidate such mechanisms we chose to study the Caco-2 colorectal cancer cell line during the course of its spontaneous differentiation in vitro, as we found CT genes, in particular PAGE2, -2B and SPANX-B, to be up-regulated during this process. Differentiation of these cells resulted in a mesenchymal-to-epithelial transition (MET) as evidenced by the gain of epithelial markers CDX2, Claudin-4 and E-cadherin, and a concomitant loss of mesenchymal markers Vimentin, Fibronectin-1 and Transgelin. PAGE2 and SPAN-X up-regulation was accompanied by an increase in Ten-eleven translocation-2 (TET2) expression and cytosine 5-hydroxymethylation as well as the disassociation of heterochromatin protein 1 and the polycomb repressive complex 2 protein EZH2 from promoter-proximal regions of these genes. Reversal of differentiation resulted in down-regulation of PAGE2, -2B and SPANX-B, and induction of epithelial-to-mesenchymal transition (EMT) markers, demonstrating the dynamic nature of CT gene regulation in this model.