Effect of ABT-888 on the apoptosis, motility and invasiveness of BRAFi-resistant melanoma cells.
Melanoma is a molecularly heterogeneous disease with many genetic mutations and altered signaling pathways. Activating mutations in the BRAF oncogene are observed in approximately 50% of cutaneous melanomas and the use of BRAF inhibitor (BRAFi) compounds has been reported to improve the outcome of patients with BRAF-mutated metastatic melanoma. However, the majority of these patients develop resistance within 6-8 months following the initiation of BRAFi treatment. In this study, we examined the possible use of the poly(ADP-ribose) polymerase 1 (PARP1) inhibitor, ABT-888 (veliparib), as a novel molecule that may be successfully employed in the treatment of BRAFi-resistant melanoma cells. Sensitive and resistant to BRAFi dabrafenib A375 cells were exposed to increasing concentrations of ABT-888. Cell viability and apoptosis were assessed by MTT assay and Annexin V-FITC analysis, respectively. The cell migratory and invasive ability was investigated using the xCELLigence technology and Boyden chamber assays, respectively. ABT-888 was found to reduce cell viability and exhibited pro-apoptotic activity in melanoma cell lines, independently from the BRAF/NRAS mutation status, in a dose-dependent manner, with the maximal effect being reached in the 25-50 µM concentration range. Moreover, ABT-888 promoted apoptosis in both the sensitive and resistant A375 cells, suggesting that ABT-888 may be useful in the treatment of BRAFi-resistant subsets of melanoma cells. Finally, in accordance with the involvement of PARP1 in actin cytoskeletal machinery, we found that the cytoskeletal organization, motility and invasive capability of both the A375 and A375R cells decreased upon exposure to 5 µM ABT-888 for 24 h. On the whole, the findings of this study highlight the pivotal role of PARP1 in the migration and invasion of melanoma cells, suggesting that ABT-888 may indeed be effective, not only as a pro-apoptotic drug for use in the treatment of BRAFi-resistant melanoma cells, but also in suppressing their migratory and invasive activities.


Systemic effects of missense mutations on SARS-CoV-2 spike glycoprotein stability and receptor-binding affinity.
The spike (S) glycoprotein of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is responsible for the binding to the permissive cells. The receptor-binding domain (RBD) of SARS-CoV-2 S protein directly interacts with the human angiotensin-converting enzyme 2 (ACE2) on the host cell membrane. In this study, we used computational saturation mutagenesis approaches, including structure-based energy calculations and sequence-based pathogenicity predictions, to quantify the systemic effects of missense mutations on SARS-CoV-2 S protein structure and function. A total of 18 354 mutations in S protein were analyzed, and we discovered that most of these mutations could destabilize the entire S protein and its RBD. Specifically, residues G431 and S514 in SARS-CoV-2 RBD are important for S protein stability. We analyzed 384 experimentally verified S missense variations and revealed that the dominant pandemic form, D614G, can stabilize the entire S protein. Moreover, many mutations in N-linked glycosylation sites can increase the stability of the S protein. In addition, we investigated 3705 mutations in SARS-CoV-2 RBD and 11 324 mutations in human ACE2 and found that SARS-CoV-2 neighbor residues G496 and F497 and ACE2 residues D355 and Y41 are critical for the RBD-ACE2 interaction. The findings comprehensively provide potential target sites in the development of drugs and vaccines against COVID-19.


Infliximab in severe ulcerative colitis: short-term results of different infusion regimens and long-term follow-up.
Severe ulcerative colitis is a life-threatening disorder, despite i.v. glucocorticoids treatment. Infliximab has been proposed as a safe rescue therapy. To evaluate short- and long-term effectiveness and safety of infliximab in severe refractory ulcerative colitis. Eighty-three patients with severe ulcerative colitis (i.v. glucocorticoids treatment-refractory) were treated with infliximab in 10 Italian Gastroenterology Units. Patients underwent one or more infusions according to the choice of treating physicians. Short-term outcome was colectomy/death 2 months after the first infusion. Long-term outcome was survival free from colectomy. Safety data were recorded. Twelve patients (15%) underwent colectomy within 2 months. One died of Legionella pneumophila infection 12 days after infliximab. Early colectomy rates were higher in patients receiving one infusion (9/26), compared with those receiving two/more infusions (3/57, P = 0.001, OR = 9.53). Seventy patients who survived colectomy and did not experience any fatal complications were followed-up for a median time of 23 months; 58 patients avoided colectomy during the follow-up. Forty-two patients were maintained on immunosuppressive drugs. No clinical features were associated with outcomes. Infliximab is an effective and relatively safe therapy to avoid colectomy and maintain long-term remission for patients with severe refractory ulcerative colitis. In the short term, two or more infusions seem to be more effective than one single infusion.


Regulation of ob gene expression and leptin secretion by insulin and dexamethasone in rat adipocytes.
Leptin, the ob gene product, is produced by adipocytes, and it acts to decrease caloric intake and increase energy expenditure. To better understand the molecular mechanisms of hormone-regulated leptin synthesis and secretion, we assessed the ability of insulin and dexamethasone to acutely modulate ob gene expression and leptin secretion in rat adipocytes. Incubation of rat adipocytes with 100 nmol/l insulin for 2 h had no effect on ob mRNA levels, but it stimulated a twofold increase in leptin secretion. Dexamethasone (100 nmol/l) stimulated both a two- to fourfold increase in ob mRNA and a twofold increase in leptin secretion. Consonant with a posttranscriptional and transcriptional regulatory mechanism for insulin- and dexamethasone-stimulated leptin secretion, respectively, actinomycin D blocked dexamethasone-stimulated leptin secretion but did not affect insulin-stimulated leptin secretion. Cycloheximide treatment did not significantly affect ob mRNA accumulation, but it reduced total secreted leptin. Interestingly, however, insulin was still able to stimulate a twofold increase in leptin secretion. These data suggest that insulin, but not dexamethasone, is able to stimulate leptin secretion from a preexisting intracellular pool, although de novo protein synthesis is required for the full insulin-stimulated effect. Signaling pathways involved in leptin synthesis/secretion were also evaluated. The phosphatidylinositol 3-kinase inhibitor LY294002, the Map/Erk kinase inhibitor PD98059, and the immunosuppressant rapamycin had no effect on basal levels of leptin secretion. However, all three inhibitors markedly decreased both insulin- and dexamethasone-stimulated leptin secretion. These findings suggest a complex set of signaling pathways involved in mediating insulin- and dexamethasone-stimulated leptin synthesis and secretion.


Effect of a quadrivalent meningococcal ACWY glycoconjugate or a serogroup B meningococcal vaccine on meningococcal carriage: an observer-blind, phase 3 randomised clinical trial.
Meningococcal conjugate vaccines protect individuals directly, but can also confer herd protection by interrupting carriage transmission. We assessed the effects of meningococcal quadrivalent glycoconjugate (MenACWY-CRM) or serogroup B (4CMenB) vaccination on meningococcal carriage rates in 18-24-year-olds. In this phase 3, observer-blind, randomised controlled trial, university students aged 18-24 years from ten sites in England were randomly assigned (1:1:1, block size of three) to receive two doses 1 month apart of Japanese Encephalitis vaccine (controls), 4CMenB, or one dose of MenACWY-CRM then placebo. Participants were randomised with a validated computer-generated random allocation list. Participants and outcome-assessors were masked to the treatment group. Meningococci were isolated from oropharyngeal swabs collected before vaccination and at five scheduled intervals over 1 year. Primary outcomes were cross-sectional carriage 1 month after each vaccine course. Secondary outcomes included comparisons of carriage at any timepoint after primary analysis until study termination. Reactogenicity and adverse events were monitored throughout the study. Analysis was done on the modified intention-to-treat population, which included all enrolled participants who received a study vaccination and provided at least one assessable swab after baseline. This trial is registered with ClinicalTrials.gov, registration number NCT01214850. Between Sept 21 and Dec 21, 2010, 2954 participants were randomly assigned (987 assigned to control [984 analysed], 979 assigned to 4CMenB [974 analysed], 988 assigned to MenACWY-CRM [983 analysed]); 33% of the 4CMenB group, 34% of the MenACWY-CRM group, and 31% of the control group were positive for meningococcal carriage at study entry. By 1 month, there was no significant difference in carriage between controls and 4CMenB (odds ratio 1·2, 95% CI 0·8-1·7) or MenACWY-CRM (0·9, [0·6-1·3]) groups. From 3 months after dose two, 4CMenB vaccination resulted in significantly lower carriage of any meningococcal strain (18·2% [95% CI 3·4-30·8] carriage reduction), capsular groups BCWY (26·6% [10·5-39·9] carriage reduction), capsular groups CWY (29·6% [8·1-46·0] carriage reduction), and serogroups CWY (28·5% [2·8-47·5] carriage reduction) compared with control vaccination. Significantly lower carriage rates were also noted in the MenACWY-CRM group compared with controls: 39·0% (95% CI 17·3-55·0) carriage reduction for serogroup Y and 36·2% (15·6-51·7) carriage reduction for serogroup CWY. Study vaccines were generally well tolerated, with increased rates of transient local injection pain and myalgia in the 4CMenB group. No safety concerns were identified. Although we detected no significant difference between groups at 1 month after vaccine course, MenACWY-CRM and 4CMenB vaccines reduced meningococcal carriage rates during 12 months after vaccination and therefore might affect transmission when widely implemented. Novartis Vaccines.


Ultrafiltration for decompensated heart failure: renal implications.
The negative prognostic impact of worsening renal function in patients with decompensated heart failure has been widely recognised. As diuretics are thought to contribute to deterioration of kidney function in this setting, attempts have been made to either spare or suppress the diuretic-related pathophysiological mechanisms involved in this phenomenon. In this regard, extracorporeal ultrafiltration represents a novel therapy for patients with heart failure, lacking the adverse impacts of diuretics on kidney function (eg, activation of tubuloglomerular feedback). Consequently, besides its other positive clinical outcomes, there has been much hope for ultrafiltration therapy to play a protective role against negative effects of diuretics in patients with decompensated heart failure. However, the potential biological advantage has not been confirmed by clinical studies; currently available data from recent clinical trials have so far failed to demonstrate such expected positive results possibly due to counterbalance of the potential negative effects and other not well-known mechanisms. This paper briefly reviews the relevant pathophysiological mechanisms as well as existing evidence in this area and emphasises on the need for further studies specifically designed to explore the impact of ultrafiltration on kidney function in patients with decompensated heart failure.


Fruits of Black Chokeberry Aronia melanocarpa in the Prevention of Chronic Diseases.
In recent years, growing attention has been focused on the utilization of natural sources of antioxidants in the prevention of chronic diseases. Black chokeberry (Aronia melanocarpa) represents a lesser known fruit species utilized mainly as juices, purees, jams, jellies and wine, as important food colorants or nutritional supplements. The fruit is valued as a great source of antioxidants, especially polyphenols, such as phenolic acids (neochlorogenic and chlorogenic acids) and flavonoids (anthocyanins, proanthocyanidins, flavanols and flavonols), particularly cyanidin-3-galactoside and cyanidin-3-arabinoside, as well as (-)-epicatechin units. The berries of A. melanocarpa, due to the presence and the high content of these bioactive components, exhibit a wide range of positive effects, such as strong antioxidant activity and potential medicinal and therapeutic benefits (gastroprotective, hepatoprotective, antiproliferative or anti-inflammatory activities). They could be also contributory toward the prevention of chronic diseases including metabolic disorders, diabetes and cardiovascular diseases, because of supportive impacts on lipid profiles, fasting plasma glucose and blood pressure levels.


Family history as a risk factor for peripheral arterial disease.
The association of a family history of peripheral arterial disease (PAD) with the presence of PAD is largely unknown. We conducted a case-control study of 2,296 patients with PAD (69 ± 10 years, 64% men) and 4,390 controls (66 ± 11 years, 62% men) identified from noninvasive vascular and stress testing laboratories at Mayo Clinic, Rochester, Minnesota, from October 2006 through June 2012. PAD was defined as an ankle brachial index of ≤ 0.9 at rest and/or after exercise, a history of lower extremity revascularization, or having poorly compressible leg arteries. Controls were patients with normal ankle brachial index or without a history of PAD. Family history of PAD was defined as having at least 1 first-degree relative who had undergone revascularization or stent placement for PAD before the age of 65 years. Logistic regression analyses were used to evaluate whether a family history of PAD was associated with the presence of PAD, independent of conventional risk factors. A family history of PAD was present more often in patients with PAD than in controls, with a resulting odds ratio (OR) of 2.20 (95% confidence interval [CI] 1.82 to 2.67). The association remained significant after adjustment for conventional risk factors (OR 1.97, 95% CI 1.60 to 2.42). The association was stronger in younger subjects (age <68 years; adjusted OR 2.46, 95% CI 1.79 to 3.38) than in older subjects (adjusted OR 1.61, 95% CI 1.22 to 2.12). A greater number of affected relatives with PAD was also associated with greater odds of presence of PAD (adjusted OR 1.86, 95% CI 1.48 to 2.33 and adjusted OR 2.56, 95% CI 1.60 to 4.11 for patients with 1 and ≥ 2 affected relatives with PAD, respectively). In conclusion, individuals with a family history of PAD have nearly double the odds of having PAD relative to those without such a history.


Abnormal attentional modulation of retinotopic cortex in parietal patients with spatial neglect.
Brain regions beyond visual cortex are thought to be responsible for attention-related modulation of visual processing [1, 2], but most evidence is indirect. Here, we applied functional magnetic resonance imaging (fMRI), including retinotopic mapping of visual areas, to patients with focal right-parietal lesions and left spatial neglect [3, 4]. When attentional load at fixation was minimal, retinotopic areas in right visual cortex showed preserved responses to task-irrelevant checkerboards in the contralateral left hemifield, analogously to left visual cortex for right-hemifield checkerboards, indicating a "symmetric" pattern in both hemispheres with respect to contralateral stimulation under these conditions. But when attentional load at fixation was increased, a functional asymmetry emerged for visual cortex, with contralateral responses in right visual areas being pathologically reduced (even eliminated for right V4/TEO), whereas left visual areas showed no such reduction in their contralateral response. These results reveal attention-dependent abnormalities in visual cortex after lesions in distant (parietal) regions. This may explain otherwise puzzling aspects of neglect [5, 6], as confirmed here by additional behavioral testing.


Evolution of the gene lineage encoding the carbon dioxide receptor in insects.
A heterodimer of the insect chemoreceptors Gr21a and Gr63a has been shown to be the carbon dioxide receptor in Drosophila melanogaster (Meigen) (Diptera: Drosophilidae). Comparison of the genes encoding these two proteins across the 12 available drosophilid fly genomes allows refined definition of their N-termini. These genes are highly conserved, along with a paralog of Gr21a, in the Anopheles gambiae, Aedes aegypti, and Culex pipiens mosquitoes, as well as in the silk moth Bombyx mori and the red flour beetle Tribolium castaneum. In the latter four species we name these three proteins Gr1, Gr2, and Gr3. Intron evolution within this distinctive three gene lineage is considerable, with at least 13 inferred gains and 39 losses. Surprisingly, this entire ancient gene lineage is absent from all other available more basal insect and related arthropod genomes, specifically the honey bee, parasitoid wasp, human louse, pea aphid, waterflea, and blacklegged tick genomes. At least two of these species can detect carbon dioxide, suggesting that they evolved other means to do so.


Magnetophoretic transistors in a tri-axial magnetic field.
The ability to direct and sort individual biological and non-biological particles into spatially addressable locations is fundamentally important to the emerging field of single cell biology. Towards this goal, we demonstrate a new class of magnetophoretic transistors, which can switch single magnetically labeled cells and magnetic beads between different paths in a microfluidic chamber. Compared with prior work on magnetophoretic transistors driven by a two-dimensional in-plane rotating field, the addition of a vertical magnetic field bias provides significant advantages in preventing the formation of particle clumps and in better replicating the operating principles of circuits in general. However, the three-dimensional driving field requires a complete redesign of the magnetic track geometry and switching electrodes. We have solved this problem by developing several types of transistor geometries which can switch particles between two different tracks by either presenting a local energy barrier or by repelling magnetic objects away from a given track, hereby denoted as "barrier" and "repulsion" transistors, respectively. For both types of transistors, we observe complete switching of magnetic objects with currents of ∼40 mA, which is consistent over a range of particle sizes (8-15 μm). The switching efficiency was also tested at various magnetic field strengths (50-90 Oe) and driving frequencies (0.1-0.6 Hz); however, we again found that the device performance only weakly depended on these parameters. These findings support the use of these novel transistor geometries to form circuit architectures in which cells can be placed in defined locations and retrieved on demand.


Protein-protein interactions and the spatiotemporal dynamics of bacterial outer membrane proteins.
It has until recently been unclear whether outer membrane proteins (OMPs) of Gram-negative bacteria are organized or distributed randomly. Studies now suggest promiscuous protein-protein interactions (PPIs) between β-barrel OMPs in Escherichia coli govern their local and global dynamics, engender spatiotemporal patterning of the outer membrane into micro-domains and are the basis of β-barrel protein turnover. We contextualize these latest advances, speculate on areas of bacterial cell biology that might be influenced by the organization of OMPs into supramolecular assemblies, and highlight the new questions and controversies this revised view of the bacterial outer membrane raises.


Role of lipooligosaccharides in experimental dermal lesions caused by Haemophilus ducreyi.
The mouse and rabbit intradermal injection models have been used to define factors that may be important in Haemophilus ducreyi pathogenesis. We used H. ducreyi strains with diverse geographic origins and phenotypic characteristics to evaluate the experimental models. Injection of live and heat-killed bacteria caused skin abscesses in both models. Semiquantitative cultures of skin injected with live bacteria showed that H. ducreyi failed to replicate in animal tissue. These data suggested that the experimental lesions were caused by a heat-stable substance such as lipooligosaccharide (LOS). In mice, injection of H. ducreyi and Haemophilus influenzae LOS and Escherichia coli lipopolysaccharide caused mild to moderate inflammation. In rabbits, injection of H. ducreyi LOS caused intradermal abscesses that were histologically similar to those caused by live and heat-killed bacteria. H. ducreyi and Neisseria gonorrhoeae LOS caused significantly larger lesions than equivalent amounts of H. influenzae LOS and E. coli lipopolysaccharide in the rabbit model. We conclude that the intradermal injection models are not valid models to study the growth of H. ducreyi in vivo. However, these data indicate that H. ducreyi LOS may play an important role in the pathogenesis of chancroid and that the rabbit model should be useful in studying H. ducreyi LOS toxicity at the cellular level.


The ratio of monomeric to aggregated forms of Abeta40 and Abeta42 is an important determinant of amyloid-beta aggregation, fibrillogenesis, and toxicity.
Aggregation and fibril formation of amyloid-beta (Abeta) peptides Abeta40 and Abeta42 are central events in the pathogenesis of Alzheimer disease. Previous studies have established the ratio of Abeta40 to Abeta42 as an important factor in determining the fibrillogenesis, toxicity, and pathological distribution of Abeta. To better understand the molecular basis underlying the pathologic consequences associated with alterations in the ratio of Abeta40 to Abeta42, we probed the concentration- and ratio-dependent interactions between well defined states of the two peptides at different stages of aggregation along the amyloid formation pathway. We report that monomeric Abeta40 alters the kinetic stability, solubility, and morphological properties of Abeta42 aggregates and prevents their conversion into mature fibrils. Abeta40, at approximately equimolar ratios (Abeta40/Abeta42 approximately 0.5-1), inhibits (> 50%) fibril formation by monomeric Abeta42, whereas inhibition of protofibrillar Abeta42 fibrillogenesis is achieved at lower, substoichiometric ratios (Abeta40/Abeta42 approximately 0.1). The inhibitory effect of Abeta40 on Abeta42 fibrillogenesis is reversed by the introduction of excess Abeta42 monomer. Additionally, monomeric Abeta42 and Abeta40 are constantly recycled and compete for binding to the ends of protofibrillar and fibrillar Abeta aggregates. Whereas the fibrillogenesis of both monomeric species can be seeded by fibrils composed of either peptide, Abeta42 protofibrils selectively seed the fibrillogenesis of monomeric Abeta42 but not monomeric Abeta40. Finally, we also show that the amyloidogenic propensities of different individual and mixed Abeta species correlates with their relative neuronal toxicities. These findings, which highlight specific points in the amyloid peptide equilibrium that are highly sensitive to the ratio of Abeta40 to Abeta42, carry important implications for the pathogenesis and current therapeutic strategies of Alzheimer disease.


Identification of 8-methyladenosine as the modification catalyzed by the radical SAM methyltransferase Cfr that confers antibiotic resistance in bacteria.
The Cfr methyltransferase confers combined resistance to five different classes of antibiotics that bind to the peptidyl transferase center of bacterial ribosomes. The Cfr-mediated modification has previously been shown to occur on nucleotide A2503 of 23S rRNA and has a mass corresponding to an additional methyl group, but its specific identity and position remained to be elucidated. A novel tandem mass spectrometry approach has been developed to further characterize the Cfr-catalyzed modification. Comparison of nucleoside fragmentation patterns of A2503 from Escherichia coli cfr+ and cfr- strains with those of a chemically synthesized nucleoside standard shows that Cfr catalyzes formation of 8-methyladenosine. In addition, analysis of RNA derived from E. coli strains lacking the m(2)A2503 methyltransferase reveals that Cfr also has the ability to catalyze methylation at position 2 to form 2,8-dimethyladenosine. The mutation of single conserved cysteine residues in the radical SAM motif CxxxCxxC of Cfr abolishes its activity, lending support to the notion that the Cfr modification reaction occurs via a radical-based mechanism. Antibiotic susceptibility data confirm that the antibiotic resistance conferred by Cfr is provided by methylation at the 8 position and is independent of methylation at the 2 position of A2503. This investigation is, to our knowledge, the first instance where the 8-methyladenosine modification has been described in natural RNA molecules.


Network Alterations in Comorbid Chronic Pain and Opioid Addiction: An Exploratory Approach.
The comorbidity of chronic pain and opioid addiction is a serious problem that has been growing with the practice of prescribing opioids for chronic pain. Neuroimaging research has shown that chronic pain and opioid dependence both affect brain structure and function, but this is the first study to evaluate the neurophysiological alterations in patients with comorbid chronic pain and addiction. Eighteen participants with chronic low back pain and opioid addiction were compared with eighteen age- and sex-matched healthy individuals in a pain-induction fMRI task. Unified structural equation modeling (SEM) with Lagrange multiplier (LM) testing yielded a network model of pain processing for patient and control groups based on 19 a priori defined regions. Tests of differences between groups on specific regression parameters were determined on a path-by-path basis using z-tests corrected for the number of comparisons. Patients with the chronic pain and addiction comorbidity had increased connection strengths; many of these connections were interhemispheric and spanned regions involved in sensory, affective, and cognitive processes. The affected regions included those that are commonly altered in chronic pain or addiction alone, indicating that this comorbidity manifests with neurological symptoms of both disorders. Understanding the neural mechanisms involved in the comorbidity is crucial to finding a comprehensive treatment, rather than treating the symptoms individually.


Persistent iodine deficiency in a cohort of Tasmanian school children: associations with socio-economic status, geographical location and dietary factors.
To determine the adequacy of iodine nutrition of Tasmanian primary school-aged children and to examine possible associations with socio-economic status (SES), location and dietary factors. Urinary iodine levels and measures of SES, geographical information and dietary habits were surveyed in a population-based sample of 170 children (4 to 12 years) at baseline (1998/99) and at follow-up (2000/01). Median urinary iodine concentration in 1989--99 and 2000--01 were 75 microg/L (range 15 microg/L to 240 microg/L) and 76 microg/L (range 18 microg/L to 480 microg/L) respectively. No significant associations with SES or geographical location were found. More frequent or recent intake of foods that are likely to be dietary sources of iodine tended to be associated with greater prevalence of adequate urinary iodine, particularly consumption of yoghurt and 'fruche' (p=0.04). After several decades of iodine sufficiency, Tasmanian primary school-aged children are again mildly iodine-deficient by WHO criteria. Despite reduction in iodophor use by the dairy industry in the past decade, consumption of dairy products continues to be associated with higher levels of iodine nutrition. The lack of association of iodine levels with SES and geographical location within Tasmania found in our study, and the results of studies of iodine levels in Melbourne and Sydney, suggest that inadequate iodine nutrition is a widespread problem in south-eastern Australia. Our study suggests that milk-containing products continue to be an important source of iodine for children.


Elevated sialic acid, but not CRP, predicts features of the metabolic syndrome independently of BMI in women.
C-reactive protein (CRP) is a predictor of many diseases including type II diabetes and cardiovascular disease. Fewer studies have similarly shown sialic acid (SA) to be a predictor of obesity-related diseases, but importantly SA shows less intra-individual variability than CRP and acts as an integrated marker of the activity of a number of acute-phase proteins. This study examines the association between both CRP and SA with individual and combined features of the metabolic syndrome. In all, 257 women with a body mass index (BMI) ranging from 25.1 to 54.5 kg/m2 (geometric mean 33.1+/-5.8 kg/m2) and aged 19-71 y (mean 45.6+/-12.1 y) were studied. Subjects had no symptoms of intercurrent infection, known diabetes, treated dyslipidaemia, a chronic inflammatory condition, liver disease or malignancy. Linear regression demonstrates that both CRP and SA were positively associated with weight, BMI, insulin resistance, dyslipidaemia and hypertension. There was a highly significant (P<0.0001) positive association of both SA and CRP with none, one, two, three or four features of the metabolic syndrome. For a 1 s.d. (4.0 mg/l) increase in CRP, there was a significant increased risk when comparing the odds of having metabolic syndrome (defined as three or more individual features) compared with the remainder of the population (odds ratio=1.7, P<0.0001), but this was not significant after adjustment for BMI. However, for a 1 s.d. (0.34 mmol/l) increase in SA, the odds of having metabolic syndrome compared with those without metabolic syndrome was 2.5 (P<0.0001), and persisted after additional adjustment for BMI (adjusted odds ratio=1.9, P<0.0001). While SA and CRP are both univariately associated with individual features of the metabolic syndrome, SA, but not CRP, is significantly associated with the metabolic syndrome, independent of BMI. We conclude that SA identifies a subgroup of overweight individuals with an inflammatory phenotype, who are at the greatest risk of metabolic syndrome.


GNAS(R201H) and Kras(G12D) cooperate to promote murine pancreatic tumorigenesis recapitulating human intraductal papillary mucinous neoplasm.
Intraductal papillary mucinous neoplasm (IPMN), the most common pancreatic cystic neoplasm, is known to progress to invasive ductal adenocarcinoma. IPMNs commonly harbor activating somatic mutations in GNAS and KRAS, primarily GNAS(R201H) and KRAS(G12D). GNAS encodes the stimulatory G-protein α subunit (Gsα) that mediates a stimulatory signal to adenylyl cyclase to produce cyclic adenosine monophosphate (cAMP), subsequently activating cAMP-dependent protein kinase A. The GNAS(R201H) mutation results in constitutive activation of Gsα. To study the potential role of GNAS in pancreatic tumorigenesis in vivo, we generated lines of transgenic mice in which the transgene consisted of Lox-STOP-Lox (LSL)-GNAS(R201H) under the control of the CAG promoter (Tg(CAG-LSL-GNAS)). These mice were crossed with pancreatic transcription factor 1a (Ptf1a)-Cre mice (Ptf1a(Cre/+)), generating Tg(CAG-LSL-GNAS);Ptf1a(Cre/+) mice. This mouse line showed elevated cAMP levels, small dilated tubular complex formation, loss of acinar cells and fibrosis in the pancreas; however, no macroscopic tumorigenesis was apparent by 2 months of age. We then crossed Tg(CAG-LSL-GNAS);Ptf1a(Cre/+) mice with LSL-Kras(G12D) mice, generating Tg(CAG-LSL-GNAS);LSL-Kras(G12D);Ptf1a(Cre/+) mice. We used these mice to investigate a possible cooperative effect of GNAS(R201H) and Kras(G12D) in pancreatic tumorigenesis. Within 5 weeks, Tg(CAG-LSL-GNAS);LSL-Kras(G12D);Ptf1a(Cre/+) mice developed a cystic tumor consisting of marked dilated ducts lined with papillary dysplastic epithelia in the pancreas, which closely mimicked the human IPMN. Our data strongly suggest that activating mutations in GNAS and Kras cooperatively promote murine pancreatic tumorigenesis, which recapitulates IPMN. Our mouse model may serve as a unique in vivo platform to find biomarkers and effective drugs for diseases associated with GNAS mutations.


[Decline in homicide rates in São Paulo, Brasil: a descriptive analysis].
To describe homicide mortality in the municipality of São Paulo according to type of weapon, sex, race or skin color, age, and areas of socioeconomic inequalities, between 1996 and 2008. For this ecological time-series study, data about deaths in the municipality of São Paulo were collected from the municipal program for improvement of mortality information, using International Classification of Diseases, 10th revision (ICD-10) codes. Homicide mortality rates (HMR) were calculated for the overall population and specifically for each sex, race or skin color, age range, type of weapon, and occurrence in social deprivation/affluence areas. HMR were adjusted for age using the direct method. The percentage age of variation in HMR was calculated for the study period. For areas of socioeconomic inequalities, the relative risk of death from homicide was calculated. HMR fell 73.7% between 2001 and 2008. A reduction in HMR was observed in all groups, especially males (-74.5%), young men between 15 and 24 years of age (-78.0%), and residents in areas of extreme socioeconomic deprivation (-79.3%). The reduction occurred mostly in firearm homicide rates (-74.1%). The relative risk of death from homicide in areas of extreme socioeconomic deprivation, as compared to areas with some degree of socioeconomic deprivation, was 2.77 in 1996, 3.9 in 2001, and 2.13 in 2008. In areas of high socioeconomic deprivation, the relative risk was 2.07 in 1996 and 1.96 in 2008. To understand the reduction in homicide rates in the municipality of São Paulo, it is important to take into consideration macrodeterminants that affect the entire municipality and all population subgroups, as well as micro/local determinants that have special impact on homicides committed with firearms and on subgroups such as the young, males, and residents of areas of high socioeconomic deprivation.


Effects of leptin on human breast cancer cell lines in relationship to estrogen receptor and HER2 status.
Obesity is a risk factor for postmenopausal breast cancer and is associated with poor prognosis. Leptin, a cytokine synthesized in adipose tissue, has been implicated as a link between obesity and breast cancer. In the present study, the effects of leptin on cell proliferation and proteins associated with leptin signaling and/or breast cell growth were investigated in ER-positive, MCF-7, T47-D and MDA-MB-361, and ER-negative, MDA-MB-231 and SK-BR-3, breast cancer cell lines. MDA-MB-361 and SK-BR-3 also overexpress HER2/neu. For proliferation assays, 96-well plates were used and for protein determinations cells were synchronized in 6-well plates for 18-24 h in serum-free medium. Leptin was added at 0, 5, 10, 25, 50 and 100 ng/ml for 24 and 48 h. For Western blot analyses, protein extracts were probed for Ob-Rb, Ob-R, leptin, Jak2, PI3K, Stat3, p-Stat3, PCNA, cyclin D1, Cox-2, VEGF, Bcl-2, Bcl-xL, Bax, insulin, IGF-I, IGFBP3, IGF-IRalpha, aromatase, CYP1A1 and CYP1B1. Overall, except for MCF-7 cells, leptin stimulated proliferation in all lines. MCF-7 cells expressed higher levels of Ob-Rb, Jak2, PI3K, Stat3 and p-Stat3 in a dose-dependent manner to 50 ng/ml at 24 h; and IGF-IRalpha increased at 24 h. Cyclin D1 and Cox-2 levels increased with leptin treatment. Higher CYP1B1 expression was observed at both 24 and 48 h. In MDA-MB-231 cells, p-Stat3 and Bcl-xL were increased at 48 h; whereas PCNA and cyclin D1 expression increased in leptin-treated cells at 24 and 48 h. In T47-D cells, Jak2 and Stat3 were elevated at higher leptin concentrations at 24 and 48 h. However, p-Stat3 and PCNA demonstrated an increase only in 48-h leptin-treated cells. Furthermore, cyclin D1 exhibited higher expression at both 24 and 48 h, while Bcl-xL expression was lower with increasing concentrations of leptin at 48 h. In MDA-MB-361 cells, Ob-Rb and VEGF increased at 24 and 48 h; whereas PI3K, Stat3, PCNA and insulin levels increased in leptin-treated MDA-MB-361 cells after 48 h. Bcl-2 and IGF-IRalpha were decreased at 24 h and a dose-dependent increase at 48 h was noted. Higher expression of CYP1B1 was observed with leptin for 24 h. In SK-BR-3 cells, Ob-R increased at both 24 and 48 h. A similar trend was found for IGF-I and IGFBP3 expression. Higher levels of Jak2 and PI3K were observed after 24 h. Interestingly, there was a gradual increase of leptin expression at 24 h, but a gradual decrease at 48 h in relation to the dose of leptin. In contrast, PCNA and IGF-IRalpha showed a decline at 24 h and an increase at 48 h. Elevated levels of cyclin D1, VEGF and Bax were detected at 48 h in cells and increased Cox-2 expression was observed at 24 h. These data indicate that leptin may influence breast cancer development in relation to ER status as well as to the presence or absence of HER2. Continued study on leptin may be helpful for a better understanding of breast cancer development in obese women.


Attitudes toward early palliative care in cancer patients and caregivers: a Korean nationwide survey.
Integrated early palliative care (EPC) improves quality of life and reduces psychological distress in adult patients with cancer and caregivers, but attitudes toward EPC have been poorly studied. We aimed to investigate attitudes toward EPC in a nationwide survey of patients with cancer and caregivers. From July to October 2016, we administered nationwide questionnaires examining attitudes toward EPC in patients with cancer (n = 1001) and their families (n = 1006) from 12 Korean hospitals. When an individual considered EPC unnecessary, the reasons were collected and analyzed. Factors associated with perception of EPC were examined. A majority of patients (84.5%) and caregivers (89.5%) had positive attitudes toward EPC. The most common reasons for deeming EPC unnecessary were that EPC may be an obstacle to cancer treatment (patients: 37%; caregivers: 23%; respectively) or that they were not sure if EPC is beneficial (patients: 21%; caregivers: 24%; respectively). Financial burden as a reason was more evident in caregivers (23%) than in patients (17%). Male gender, age <50, early stage, intensive care unit admission, and not believing that dying people should prepare to practice charity were associated with patients' negative attitudes. In caregivers, opposition to EPC was associated with not thinking death should be feared, not thinking people should be remembered, and lower educational level. Our findings showed that significant numbers of patients with advanced cancer and family caregivers showed positive attitudes toward EPC. However, more than 10% of participants did not consider EPC necessary. Physicians' communication with patients and caregivers and financial support could help overcome the barriers of EPC.


Auditory temporal coding in dyslexia.
Developmental dyslexia is generally believed to result from impaired linguistic processing rather than from deficits in low-level sensory function. Challenging this view, we studied the perception of non-verbal acoustic stimuli and low-level auditory evoked potentials in dyslexic adults. Compared with matched controls, dyslexics were selectively impaired in tasks (frequency discrimination and binaural unmasking) which rely on decoding neural discharges phase-locked to the fine structure of the stimulus. Furthermore, this ability to use phase-locking was related to reading ability. In addition, the evoked potential reflecting phase-locked discharges was significantly smaller in dyslexics. These results demonstrate a low-level auditory impairment in dyslexia traceable to the brainstem nuclei.


Treatments of internet gaming disorder: a systematic review of the evidence.
Introduction: The American Psychiatric Association included Internet gaming disorder (IGD) in the 5th Edition of the Diagnostic and Statistical Manual of Mental Disorders, and the World Health Organization included gaming disorder in the 11th revision of the International Classification of Diseases. These recent updates suggest significant concern related to the harms of excessive gaming.Areas covered: This systematic review provides an updated summary of the scientific literature on treatments for IGD. Inclusion criteria were that studies: 1) evaluate the effectiveness of an intervention for IGD or excessive gaming; 2) use an experimental design (i.e. multi-armed [randomized or nonrandomized] or pretest-posttest); 3) include at least 10 participants per group; and 4) include an outcome measure of IGD symptoms or gaming duration. The review identified 22 studies evaluating treatments for IGD: 8 evaluating medication, 7 evaluating cognitive behavioral psychotherapy, and 7 evaluating other interventions and psychosocial treatments.Expert opinion: Even with the recent uptick in publication of such clinical trials, methodological flaws prevent strong conclusions about the efficacy of any treatment for IGD. Additional well-designed clinical trials using common metrics for assessing IGD symptoms are needed to advance the field.


Expression of a homeobox gene product in normal and mutant zebrafish embryos: evolution of the tetrapod body plan.
An antibody was used to detect antigens in zebrafish that appear to be homologous to the frog homeodomain-containing protein XlHbox 1. These antigens show a restricted expression in the anteroposterior axis and an anteroposterior gradient in the pectoral fin bud, consistent with the distribution of XlHbox 1 protein in frog and mouse embryos. In the somitic mesoderm, a sharp anterior limit of expression coincides exactly with the boundary between somites 4 and 5, and the protein level fades out posteriorly. A similar, graded expression of the antigen is seen within the series of Rohon-Beard sensory neurons of the CNS. We also immunostained the mutant spt-1 ('spadetail'), in which the trunk mesoderm is greatly depleted and disorganized in the region of XlHbox 1 expression. The defects stem from misdirected cell movements during gastrulation, but nervertheless, newly recruited cells that partially refill the trunk mesoderm express the antigen within the normal span of the anteroposterior axis. This finding suggests that the mutation does not delete positional information required for activation of the XlHbox 1 gene.


[A case of recurrent pleural effusion and positive cytology of advanced gastric cancer treated by TS-1 plus weekly taxane as second-line chemotherapy].
A second-line chemotherapy for advanced gastric cancer has not been established. We report a case of good response in a 39-year-old woman who had recurrent pleural effusion and positive cytology of type 4 gastric cancer and was treated with TS-1, a DPD inhibitory fluoropyrimidine, in combination with weekly taxane. After a partial response for type 4 gastric cancer from the treatment with 2 courses of TS-1 plus low-dose cisplatinum (CDDP), followed by outpatient chemotherapy with TS-1 alone or TS-1 plus weekly CDDP, left pleural effusion appeared and CA19-9 increased during the 7th course of the chemotherapy. Cytology of the effusion was class IV. The patient was treated with a course of TS-1 (120 mg/day, day 1-21) plus paclitaxel (50 mg/m2, day 1, 8) followed by 2 week washout. In the following courses, paclitaxel was replaced with docetaxel (30 mg/m2, day 1 and 8) and the course was continued in the outpatient setting. After 2 courses, the left pleural effusion disappeared and remained absent after 6 courses. Gastric biopsy showed no cancer cells and abdominal CT showed no recurrence. Serum CA19-9 doubled 1 week after taxane treatment and decreased gradually week by week during the course. This case suggests that a combination of TS-1 and taxane is effective against recurrent pleural effusion of advanced gastric cancer and useful as a second-line chemotherapy.


Using mixed methods evaluation to assess the feasibility of online clinical training in evidence based interventions: a case study of cognitive behavioural treatment for low back pain.
Cognitive behavioural (CB) approaches are effective in the management of non-specific low back pain (LBP). We developed the CB Back Skills Training programme (BeST) and previously provided evidence of clinical and cost effectiveness in a large pragmatic trial. However, practice change is challenged by a lack of treatment guidance and training for clinicians. We aimed to explore the feasibility and acceptability of an online programme (iBeST) for providing training in a CB approach. This mixed methods study comprised an individually randomised controlled trial of 35 physiotherapists and an interview study of 8 physiotherapists. Participants were recruited from 8 National Health Service departments in England and allocated by a computer generated randomisation list to receive iBeST (n = 16) or a face-to-face workshop (n = 19). Knowledge (of a CB approach), clinical skills (unblinded assessment of CB skills in practice), self-efficacy (reported confidence in using new skills), attitudes (towards LBP management), and satisfaction were assessed after training. Engagement with iBeST was assessed with user analytics. Interviews explored acceptability and experiences with iBeST. Data sets were analysed independently and jointly interpreted. Fifteen (94 %) participants in the iBeST group and 16 (84 %) participants in the workshop group provided data immediately after training. We observed similar scores on knowledge (MD (95 % CI): 0.97 (-1.33, 3.26)), and self-efficacy to deliver the majority of the programme (MD (95 % CI) 0.25 (-1.7; 0.7)). However, the workshop group showed greater reduction in biomedical attitudes to LBP management (MD (95 % CI): -7.43 (-10.97, -3.89)). Clinical skills were assessed in 5 (33 %) iBeST participants and 7 (38 %) workshop participants within 6 months of training and were similar between groups (MD (95 % CI): 0.17(-0.2; 0.54)). Interviews highlighted that while initially sceptical, participants found iBeST acceptable. A number of strategies were identified to enhance future versions of iBeST such as including more skills practice. Combined quantitative and qualitative data indicated that online training was an acceptable and promising method for providing training in an evidence based complex intervention. With future enhancement, the potential reach of this training method may facilitate evidence-based practice through large scale upskilling of the workforce. Current Controlled Trials ISRCTN82203145 (registered prospectively on 03.09.2012).


Rotary-actuated folding polyhedrons for midwater investigation of delicate marine organisms.
Self-folding polyhedra have emerged as a viable design strategy for a wide range of applications, with advances largely made through modeling and experimentation at the micro- and millimeter scale. Translating these concepts to larger scales for practical purposes is an obvious next step; however, the size, weight, and method of actuation present a new set of problems to overcome. We have developed large-scale folding polyhedra to rapidly and noninvasively enclose marine organisms in the water column. The design is based on an axisymmetric dodecahedron net that is folded by an external assembly linkage. Requiring only a single rotary actuator to fold, the device is suited for remote operation onboard underwater vehicles and has been field-tested to encapsulate a variety of delicate deep-sea organisms. Our work validates the use of self-folding polyhedra for marine biological applications that require minimal actuation to achieve complex motion. The device was tested to 700 m, but the system was designed to withstand full ocean depth (11 km) pressures. We envision broader terrestrial applications of rotary-actuated folding polyhedra, ranging from large-scale deployable habitats and satellite solar arrays to small-scale functional origami microelectromechanical systems.


[Dexmedetomidine for neurocognitive testing in awake craniotomy: case report.].
Tumor resections in the speech areas of the brain are more safely done using cognitive tests to determine their exact location. Patients must be awake, comfortable, and cooperative for the precise identification of the areas to be preserved. The objective of this report is to present a surgical procedure done with the patient awake, without endotracheal intubation, using sevofluorane initially, followed by dexmedetomidine. This technique allowed the realization of motor and speech evaluation tests. Twenty-seven years old male patient, physical status ASA I, with a brain tumor. In the operating room, without pre-anesthetic medication, midazolam (1 mg) was administered, and general anesthesia was induced with propofol (80 mg). Maintenance was done with O2, N2O, and sevofluorane, with a mask, for catheterization of the right radial artery, introduction of a vesical catheter, and infiltration of the surgical site. This phase lasted around 20 minutes, and the infusion of dexmedetomidine was initiated in the last 10 minutes to maintain a level of sedation Ramsay score 2. Cortical mapping followed (75 minutes). Afterwards, tumor resection was done while the patient remained sedated with higher doses of dexmedetomidine. Hemodynamic and respiratory parameters remained stable, and the procedure was performed without complications, lasting a total of five hours. After the surgical procedure the patient was transferred to the ICU. He did not develop any neurological changes, being discharged to a regular ward the following day. Awake craniotomy with the proper mapping of speech and motor cortical areas was successfully done with the continuous infusion of dexmedetomidine. Both the patient and the surgical team were pleased with the technique.


Role of cytochrome P450 3A4 in human metabolism of MK-639, a potent human immunodeficiency virus protease inhibitor.
MK-639 (L-735,524) is a potent human immunodeficiency virus protease inhibitor under investigation in the treatment of acquired immunodeficiency syndrome. Five in vitro approaches have been used to identify the cytochrome P450 isoform(s) responsible for the human microsomal oxidative metabolism of MK-639. These approaches are: 1) chemical inhibition; 2) immunochemical inhibition; 3) metabolism by cDNA-expressed human cytochrome P450 enzymes; 4) a correlation analysis; and 5) competitive inhibition of marker activities. Ketoconazole and troleandomycin, both selective inhibitors for cytochrome P450 3A4 (CYP3A4), markedly inhibited the formation of all oxidative metabolites of MK-639; whereas other inhibitors (furafylline, sulfaphenazole, quinidine, S-mephenytoin, and diethyldithiocarbamate) had little effect on MK-639 metabolism. This suggested the involvement of CYP3A4 in MK-639 metabolism. Consistent with this, an anti-rat CYP3A1 rabbit polyclonal antibody, which shows a cross-reactive inhibition of CYP3A4-dependent testosterone 6beta-hydroxylation in human liver microsomes, completely inhibited MK-639 metabolism. Human recombinant CYP3A4 showed a high metabolic activity to form all MK-639 metabolites found in native human liver microsomes. In addition, the formation of individual MK-639 metabolites correlated well with each other and with testosterone 6beta-hydroxylation in 12 different human liver microsomes, whereas no correlation was observed between MK-639 metabolite formation and bufuralol 1'-hydroxylation (or tolbutamide methyl hydroxylation). Furthermore, MK-639 strongly inhibited testosterone 6beta-hydroxylation in a concentration-dependent manner. Kinetic analysis showed that MK-639 is a very potent competitive inhibitor for testosterone 6beta-hydroxylation, with a Ki value of approximately 0.5 mu M. Collectively, these results consistently indicate that CYP3A4 is the isoform responsible for the oxidative metabolism of MK-639 in human liver microsomes.


ComM is a hexameric helicase that promotes branch migration during natural transformation in diverse Gram-negative species.
Acquisition of foreign DNA by natural transformation is an important mechanism of adaptation and evolution in diverse microbial species. Here, we characterize the mechanism of ComM, a broadly conserved AAA+ protein previously implicated in homologous recombination of transforming DNA (tDNA) in naturally competent Gram-negative bacterial species. In vivo, we found that ComM was required for efficient comigration of linked genetic markers in Vibrio cholerae and Acinetobacter baylyi, which is consistent with a role in branch migration. Also, ComM was particularly important for integration of tDNA with increased sequence heterology, suggesting that its activity promotes the acquisition of novel DNA sequences. In vitro, we showed that purified ComM binds ssDNA, oligomerizes into a hexameric ring, and has bidirectional helicase and branch migration activity. Based on these data, we propose a model for tDNA integration during natural transformation. This study provides mechanistic insight into the enigmatic steps involved in tDNA integration and uncovers the function of a protein required for this conserved mechanism of horizontal gene transfer.


Expression of E-cadherin and alpha- and beta-catenins in thymoma.
Expression of the cell adhesion molecules including E-cadherin and its cytosolic binding proteins, alpha- and beta-catenins, has been widely studied in a variety of tumours, but not, to date, in thymic epithelial tumours. To observe the expression pattern of these adhesion molecules, immunohistochemical stains for E-cadherin (E-CD) and alpha- and beta-catenins were performed on 89 cases of thymoma which were classified as cortical (57 cases), mixed (18 cases), and medullary (14 cases), based on the classification of Marino and Müller-Hermelink. The majority of cortical thymomas showed diffuse and homogenous membrane immunoreactivity for these molecules (88 per cent for E-CD; 86 per cent for alpha-catenin; 91 per cent for beta-catenin) and the remaining cases showed heterogeneous immunoreactivity, whereas almost all mixed and medullary thymomas revealed decreased expression or were negative. In each histological subtype of thymoma, the expression did not correlate with invasion or with the presence of myasthenia gravis. These results indicate that the expression of E-CD and alpha- and beta-catenins is more closely associated with the histological subtypes of thymoma than with their biological behaviour.


Carcinogenesis Bioassay of 2,3,7,8-Tetrachlorodibenzo-p-dioxin (CAS No. 1746-01-6) in Osborne-Mendel Rats and B6C3F1 Mice (Gavage Study).
A carcinogenesis bioassay of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), a contaminant in several phenoxy herbicides, was conducted by administering TCDD by gavage to Osborne-Mendel rats and B6C3F1 mice for 104 weeks. Fifty rats and mice of each sex were given TCDD suspended in a vehicle of 9:1 corn oil-acetone 2 days per week for 104 weeks at doses of 0.01, 0.05, or 0.5 &mgr;g/kg/wk for rats and male mice and 0.04, 0.2, or 2.0 &mgr;g/kg/wk for female mice. Seventy-five rats and 75 mice of each sex served as vehicle controls. One untreated control group containing 25 rats and 25 mice of each sex was present in the TCDD treatment room, and one untreated control group containing 25 rats and 25 mice of each sex was present in the vehicle-control room. All surviving animals were killed at 105 to 108 weeks. In rats, a dose-related depression in mean body weight gain was observed in the males after week 55 of the bioassay and in the females after week 45. In mice, the mean body weight gain in the dosed groups was comparable to that of the vehicle-control groups. In male rats, increased incidences of follicular-cell adenomas in the thyroid were dose related and were significantly (P=0.001) higher in the high-dose group than in the vehicle controls (1/69, 1%; 5/48, 10%; 6/50, 12%; 10/50, 20%). Similarly in the female rats, an increase (though not statistically significant) was seen in the high-dose group (3/73, 4%; 2/45, 4%; 1/49, 2%; 6/47, 13%). In female rats, the incidence of neoplastic nodules of the liver in the high-dose group was significantly (P=0.006) higher than that in the vehicle control group (5/75, 7%; 1/49, 2%; 3/50, 6%; 12/49, 24%). In male and female mice, incidences of hepatocellular carcinomas were dose related and the incidences in the high-dose groups were significantly (P=0.002 and 0.014, respectively) higher than those in the corresponding vehicle control groups (males: 8/73, 11%; 9/49, 18%; 9/49, 16%; 17/50, 34%; females: 1/73, 1%; 2/50, 4%; 2/48, 4%; 6/47, 13%). In female mice, follicular-cell adenomas in the thyroid occurred at dose-related incidences, and were significantly (P=0.009) higher in the high-dose groups than those in the vehicle controls (0/69, 0%; 3/50, 6%; 1/47, 2%; 5/46, 11%). Increased incidences of toxic hepatitis related to the administration of the test chemical were detected among high-dose rats and high-dose mice of each sex. Under the conditions of this bioassay, 2,3,7,8-tetrachlorodibenzo-p-dioxin was carcinogenic for Osborne-Mendel rats, including follicular-cell thyroid adenomas in males and neoplastic nodules of the liver in females. TCDD was also carcinogenic for B6C3F1 mice, including hepatocellular carcinomas in male and females and follicular-cell thyroid adenomas in females. Levels of Evidence of Carcinogenicity: Male Rats: Positive Female Rats: Positive Male Mice: Positive Female Mice: Positive Synonyms: 2,3,7,8-TCDD; TCDD


Reproductive outcome of women with 21-hydroxylase-deficient nonclassic adrenal hyperplasia.
Because many women with 21-hydroxylase (21-OH)-deficient nonclassic adrenal hyperplasia (NCAH) carry at least one allele affected by a severe mutation of CYP21, they are at risk for giving birth to infants with classic adrenal hyperplasia (CAH). Our objective was to determine the frequency of CAH and NCAH infants born to mothers with 21-OH-deficient NCAH. We conducted an international multicenter retrospective/prospective study. The outcome of 203 pregnancies among 101 women with 21-OH-deficient NCAH was reviewed. The diagnosis of 21-OH-deficient NCAH was established by a basal or post-ACTH-stimulation 17-hydroxyprogesterone level of more than 10 ng/ml (30.3 nmol/liter). When possible, genotype analyses were performed to confirm CAH or NCAH in the offspring. Of the 203 pregnancies, 138 (68%) occurred before the mother's diagnosis of NCAH and 65 (32%) after the diagnosis. Spontaneous miscarriages occurred in 35 of 138 pregnancies (25.4%) before the maternal diagnosis of NCAH, and in only four of 65 pregnancies (6.2%) after the diagnosis (P < 0.002). Four (2.5%; 95% confidence interval, 0.7-6.2%) of the 162 live births were diagnosed with CAH. To date, 24 (14.8%; 95% confidence interval, 9.0-20.6%) children, 13 girls and 11 boys, have been diagnosed with NCAH. The distribution of NCAH children and their mothers varied significantly by ethnicity (P < 0.0001, for both). The risk of a mother with 21-OH-deficient NCAH for giving birth to a child affected with CAH is 2.5%; at least 14.8% of children born to these mothers have NCAH.


Targeting liver stage malaria with metformin.
Despite an unprecedented 2 decades of success, the combat against malaria - the mosquito-transmitted disease caused by Plasmodium parasites - is no longer progressing. Efforts toward eradication are threatened by the lack of an effective vaccine and a rise in antiparasite drug resistance. Alternative approaches are urgently needed. Repurposing of available, approved drugs with distinct modes of action are being considered as viable and immediate adjuncts to standard antimicrobial treatment. Such strategies may be well suited to the obligatory and clinically silent first phase of Plasmodium infection, where massive parasite replication occurs within hepatocytes in the liver. Here, we report that the widely used antidiabetic drug, metformin, impairs parasite liver stage development of both rodent-infecting Plasmodium berghei and human-infecting P. falciparum parasites. Prophylactic treatment with metformin curtails parasite intracellular growth in vitro. An additional effect was observed in mice with a decrease in the numbers of infected hepatocytes. Moreover, metformin provided in combination with conventional liver- or blood-acting antimalarial drugs further reduced the total burden of P. berghei infection and substantially lessened disease severity in mice. Together, our findings indicate that repurposing of metformin in a prophylactic regimen could be considered for malaria chemoprevention.


DNA carryover in milk samples from routine milk recording used for PCR-based diagnosis of bovine Staphylococcus aureus mastitis.
Real-time PCR techniques are increasingly used to detect udder pathogens from milk samples collected non-aseptically at routine milk recording. The objectives of this study were (1) to estimate the statistical associations between cycle threshold (Ct) values for Staphylococcus aureus in non-aseptically collected composite samples taken at routine milk recording from cows milked consecutively with the same milking unit and milk meter; and (2) to formulate practical and plausible guidelines for understanding the diagnostic implications of PCR testing for Staph. aureus intramammary infection at routine milk recording. The study included 4 herds with conventional milking parlors and repeatedly low Ct-values for Staph. aureus (representing a high DNA load) in bulk tank milk. Composite milk samples were collected from all cows at all milking units during routine milk recording using the Tru-Test electronic milk meter (Tru-Test Group, Auckland, New Zealand) and analyzed using the PathoProof PCR (Thermo Fisher Scientific, Vantaa, Finland) assay. Milking clock times were retrieved at each milk meter to establish the milking order of the cows at each unit. A multinomial logistic regression was applied to estimate the association between Ct-values from cows milked consecutively with the same milking unit and milk meter. The following groups were selected based on Ct-values: (1) 0-31.3, (2) 31.4-33.9, (3) 34.0-37, (4) 37.1-39.9, and (5) 40 (negative result). The association between groups from cows milked consecutively with the same milking unit and milk meter was statistically significant. Approximately 60% of cows were in Ct group 5 if the antecedent cow was also in Ct group 5, but only 20% of cows were in Ct group 5 if the antecedent cow was in Ct group 1. The probability of cows being in Ct group 1 was not markedly influenced by the group of the antecedent cow. Statistical relationships in the intermediate range gave a plausible indication of a dose-response relationship. Carryover of bacterial DNA via the milking unit and milk meter is very likely to affect PCR results for Staph. aureus. Therefore, information about milking order must be considered in mastitis control efforts. We suggest a practical interpretation of PCR results: cows with a Ct-value <32 can be labeled "very likely to be infected with Staph. aureus," but cows with Ct-values of >37 and 32-37 can be labeled "very likely to be negative for Staph. aureus" and "uncertain Staph. aureus status," respectively.


Are increases in mammographic screening still a valid explanation for trends in breast cancer incidence in the United States?
A number of studies have attributed much of the sharp increase in breast cancer incidence in the United States during the 1980s to the increased detection through mammography. The most recent breast cancer data from the US National Cancer Institute's Surveillance, Epidemiology, and End Results (SEER) program show that the incidence trend has slowed, while results from the National Health Interview Survey (NHIS) of 1987 and 1990 indicate that the percentage of women receiving mammograms continues to increase. This phenomenon suggested the need to reassess the relationship between increasingly early detection of breast cancer and overall incidence trends. A polynomial age-cohort model was used to establish the secular trend in incidence rates excluding the impact of recent increases in detection due to the rising use of mammography. Based on the model, the incidence trend in the youngest age group (40 to 49 years) would peak and then begin to decline in the early 1980s. This pattern would manifest itself later in successively older age groups as these younger cohorts age. Breast cancer trends are seen to be generally consistent with the impact of the increased use of mammography when its effect is superimposed upon the background of declining or slowing secular trends. These results support previous reports linking incidence rates with the increase in screening-mammography.


Abusive head trauma at a tertiary care children's hospital in Mexico City. A preliminary study.
Determine the prevalence, clinical signs and symptoms, and demographic and family characteristics of children attending a tertiary care hospital in Mexico City, Mexico, to illustrate the characteristics of abusive head trauma among this population. This is a cross-sectional descriptive study of infants and children under 5, who suffered head trauma and were admitted to the National Pediatrics Institute in Mexico City, a tertiary care referral center. We reviewed medical records and extracted data on clinical and neurological signs and symptoms, fundus, radiological (long bones, thorax, CAT scan), and laboratory tests. We administered a standardized questionnaire assessing child abuse and neglect to the parents of the children included in the study. One hundred and twenty children, under 5 presenting with head trauma, were recruited, 13 (11%) were considered abusive head trauma (AbHT) and 107 (89%) were diagnosed as accidental head injury (AcHI). The AbHT group comprised younger infants (mean age 8 months) and the AcHI group included toddlers about an average of 25 months. To account for this significant age difference, we performed a comparison of age matched cases. The children in the AbHT were more likely to be female, the result of the first unintended pregnancy and the children of younger mothers (17-19). Mothers in this group had attended fewer than 5 prenatal care visits and fathers had a history of alcohol abuse. Five (38%) of the 13 AbHT children did not survive their injuries and overall showed greater neurological and respiratory compromise, increased prothrombin time (PT), and lower hematocrit values. The most common intracranial injuries suffered by children in the AbHT group were subdural/epidural hematoma and parenchymal/subarachnoid hemorrhage. Retinal hemorrhage was the most frequent ocular injury. In a tertiary care children's hospital, 11% of the children presenting with head trauma, were considered of abusive origin. Unintended pregnancy among teen mothers and substance abuse in the father were associated with abusive head trauma in this descriptive study.


Brucella evades macrophage killing via VirB-dependent sustained interactions with the endoplasmic reticulum.
The intracellular pathogen Brucella is the causative agent of brucellosis, a worldwide zoonosis that affects mammals, including humans. Essential to Brucella virulence is its ability to survive and replicate inside host macrophages, yet the underlying mechanisms and the nature of the replicative compartment remain unclear. Here we show in a model of Brucella abortus infection of murine bone marrow-derived macrophages that a fraction of the bacteria that survive an initial macrophage killing proceed to replicate in a compartment segregated from the endocytic pathway. The maturation of the Brucella-containing vacuole involves sustained interactions and fusion with the endoplasmic reticulum (ER), which creates a replicative compartment with ER-like properties. The acquisition of ER membranes by replicating Brucella is independent of ER-Golgi COPI-dependent vesicular transport. A mutant of the VirB type IV secretion system, which is necessary for intracellular survival, was unable to sustain interactions and fuse with the ER, and was killed via eventual fusion with lysosomes. Thus, we demonstrate that live intracellular Brucella evade macrophage killing through VirB-dependent sustained interactions with the ER. Moreover, we assign an intracellular function to the VirB system, as being required for late maturation events necessary for the biogenesis of an ER-derived replicative organelle.


Toll-like receptor 7 protects from atherosclerosis by constraining "inflammatory" macrophage activation.
Toll-like receptors (TLRs) have long been considered to be major culprits in the development of atherosclerosis, contributing both to its progression and clinical complications. However, evidence for most TLRs beyond TLR2 and TLR4 is lacking. We used experimental mouse models, human atheroma cultures, and well-established human biobanks to investigate the role of TLR7 in atherosclerosis. We report the unexpected finding that TLR7, a receptor recognizing self-nucleic acid complexes, is protective in atherosclerosis. In Apoe(-/-) mice, functional inactivation of TLR7 resulted in accelerated lesion development, increased stenosis, and enhanced plaque vulnerability as revealed by Doppler ultrasound and/or histopathology. Mechanistically, TLR7 interfered with macrophage proinflammatory responses to TLR2 and TLR4 ligands, reduced monocyte chemoattractant protein-1 production, and prevented expansion of Ly6C(hi) inflammatory monocytes and accumulation of inflammatory M1 macrophages into developing atherosclerotic lesions. In human carotid endarterectomy specimens TLR7 levels were consistently associated with an M2 anti-inflammatory macrophage signature (interleukin [IL]-10, IL-1RA, CD163, scavenger and C-type lectin receptors) and collagen genes, whereas they were inversely related or unrelated to proinflammatory mediators (IL-12/IL-23, interferon beta, interferon gamma, CD40L) and platelet markers. Moreover, in human atheroma cultures, TLR7 activation selectively suppressed the production of key proatherogenic factors such as monocyte chemoattractant protein-1 and tumor necrosis factor without affecting IL-10. These findings provide evidence for a beneficial role of TLR7 in atherosclerosis by constraining inflammatory macrophage activation and cytokine production. This challenges the prevailing concept that all TLRs are pathogenic and supports the exploitation of the TLR7 pathway for therapy.


The population of users of upper limb prostheses attending the Oxford Limb Fitting Service.
There is no central database that records the changing provision of prostheses in the United Kingdom. Experience suggests there have been some shifts in the population, particularly in the past decade. Because the detailed records of these changes are contained in the patients' medical records it is difficult to assess the substance of these data except on an individual basis; the larger picture the sifting of a centre's or many centres' data. This paper describes the analysis of one such set of records at the Oxford Limb Fitting Centre. It relates the profile of the population that attends the centre to the general population, and compares the information with that obtainable from other sources. The possible causes for the findings are discussed.


Radiographic evaluation of different techniques for ridge preservation after tooth extraction: a randomized controlled clinical trial.
To evaluate the radiographic changes of the alveolar ridge following application of different ridge preservation techniques 6 months after tooth extraction. Four treatment modalities were randomly assigned in 40 patients: β-tricalcium-phosphate-particles with polylactid coating (β-TCP), demineralized bovine bone mineral with 10% collagen covered with a collagen matrix (DBBM-C/CM), DBBM-C covered with an autogenous soft-tissue graft (DBBM-C/PG) and spontaneous healing (control). Cone-beam computed tomography scans were performed after treatment and 6 months later. After 6 months, the vertical changes ranged between -0.6 mm (-10.2%) for control and a gain of 0.3 mm (5.6%) for DBBM-C/PG on the lingual side, and between -2.0 mm (-20.9%) for β-TCP and a gain of 1.2 mm (8.1%) for DBBM-C/PG on the buccal side. The most accentuated ridge width changes were recorded 1 mm below the crest: -3.3 mm (-43.3%, C), -6.1 mm (-77.5%, β-TCP), -1.2 mm (-17.4%, DBBM-C/CM) and -1.4 mm (-18.1%, DBBM-C/PG). At all three levels, DBBM-C with either CM or PG was not significantly differing (p > 0.05), while most other differences between the groups reached statistical significance (p < 0.05). Application of DBBM-C, covered with CM or PG, resulted in less vertical and horizontal changes of the alveolar ridge as compared with controls 6 months after extraction.


Cultural adaptation and validation of the painDETECT scale into Spanish.
The aim of this study was to culturally adapt into Spanish and validate the painDETECT questionnaire, a brief self-administered instrument designed to screen the presence of a neuropathic pain component in usual clinical practice. The original English painDETECT questionnaire was culturally adapted into Spanish (Spain) by 2 independent translators under supervision of an expert panel. The LANNS, and a pain visual analog scale were administered along with the painDETECT questionnaire to a sample of 252 patients with neuropathic, nociceptive, or mixed pain for at least 3 months. Patients were classified by experienced specialists under normal conditions of clinical practice. A retest measure after 24 to 48 hours was also carried out. Reliability, construct validity, convergent validity, criterion, and discriminant validity were assessed. An effective sample of 221 patients with chronic pain was recruited, 32% diagnosed of neuropathic origin, 32% of nociceptive, and remaining 36% presented mixed pain. The average age was 57.8 years (SD=14.2) and 59% were women. Cronbach alpha attained a value of 0.86, and the intraclass correlation coefficient with the retest was 0.93. The factor structure was coherent with the one informed for the original instrument. Pearson correlation with the LANSS scale was 0.88. Area under the receiver operating characteristic curve was 0.88 when comparing neuropathic and nociceptive groups. Using the suggested cutoff value for pain presence of 19 points or higher, the following discriminant values are obtained: sensitivity=75%, specificity=84%, Youden Index=0.595, positive predictive value=92%, and negative predictive value=60%. The absence cutoff value of 12 points or bellow raised the following values: sensitivity=93%, specificity=68%, Youden Index=0.61, positive predictive value=87%, and negative predictive value=80%. When mixed pain patients are included in the group with a neuropathic component, discriminant values are slightly reduced, as expected. The culturally adapted version of the painDETECT presents good psychometric properties and shows to be a valid patient-reported outcome for measuring the presence of a neuropathic component in patients with chronic pain.


Effect of plant oils on Candida albicans.
Candida species, notably Candida albicans, is the major fungal pathogen in humans. It is a dimorphic fungus capable of causing superficial mucosal infections, as well as systemic infections, in immunocompromised individuals. The factors responsible for its pathogenesis are still not fully understood and increasing resistance to commonly used antifungal agents necessitates the search for new formulations. The inhibitory effect of 30 different plant oils on Candida albicans isolated from clinical samples was evaluated. The antifungal agent fluconazole was used as a positive control. Plant oils were tested at concentrations from 0.03% to 3% (v/v) to determine the minimum inhibitory concentration and minimum fungicidal concentration (MFC) using agar dilution and macro broth dilution assays. Of the 30 plant oils tested, 18 were found to be effective and 12 were ineffective. Based on their MFCs, effective oils were placed into three categories: most effective, moderately effective and least effective. Eucalyptus and peppermint oils were most effective, with MFC values of 0.12% and 0.15% (v/v), respectively. The significant antifungal activity of these oils suggests that they could serve as a source of compounds with therapeutic potential against Candida-related infections.


Motivations for intimate partner violence in men and women arrested for domestic violence and court referred to batterer intervention programs.
Research has attempted to elucidate men and women's proximal motivations for perpetrating intimate partner violence (IPV). However, previous research has yet to clarify and resolve contention regarding whether motives for IPV are gender-neutral or gender specific. Thus, the purpose of this present study was to compare motives for physical IPV perpetration among a sample of men (n =90) and women (n =87) arrested for domestic violence and court-referred to batterer intervention programs. Results demonstrated that the most frequently endorsed motives for IPV by both men and women were self-defense, expression of negative emotions, and communication difficulties. With the exception of expression of negative emotions and retaliation, with women endorsing these motives more often than men, there were no significant differences between men and women's self-reported reasons for perpetrating physical aggression. The implications of these findings for future research and intervention programs are discussed.


Forgoing medical care because of cost: assessing disparities in healthcare access among cancer survivors living in the United States.
Many US cancer survivors live years after diagnosis, which emphasizes the importance of healthcare access for survivors. It is not known whether having cancer has an impact on disparities in healthcare access that are present in the general population. The objective of this study was to examine the prevalence of forgoing care because of financial concerns in a representative sample of US adults to determine whether cancer history and race/ethnicity are associated with the likelihood of forgoing medical care. Data from the US National Health Interview Survey (NHIS) from 2003 to 2006 were used to identify 6602 adult cancer survivors and 104,364 individuals who had no history of cancer. Self-reports of forgoing medical care services because of cost were analyzed according to cancer history and race/ethnicity using multivariate logistic regression. The prevalence of forgoing care because of cost among cancer survivors was 7.8% for medical care, 9.9% for prescription medications, 11.3% for dental care, and 2.7% for mental healthcare. Cancer survivors aged<65 years were more likely to delay or forego all types of medical care compared with adults who did not have a history of cancer. Hispanic and black cancer survivors were more likely to forego prescription medications and dental care than white survivors. Disparities among cancer survivors largely were reflective of those in the general adult population. More than 2 million US cancer survivors did not get 1 or more needed medical services because of financial concerns during the studied period. Future research needs to examine the impact of forgoing care on survivors' quality of life and survival.


Analysis of the posttranslational modifications of the influenza virus M2 protein.
The sites of posttranslational modifications of the influenza A virus M2 protein were examined, and the effect of these modifications on the M2 protein ion channel activity was analyzed. Cysteine residues 17 and 19 in the M2 protein ectodomain form disulfide bonds. The cytoplasmic tail is posttranslationally modified by palmitoylation, and mutagenic studies support the view that cysteine residue 50 is the site for fatty acylation. In addition, the cytoplasmic tail of the M2 protein was found to be posttranslationally modified by the addition of phosphate to specific serine residues. Site-directed mutagenesis of serine residues in the M2 protein cytoplasmic tail, combined with phosphoamino acid analysis, indicated that serine residue 64 is the predominant site for phosphorylation but that serine residues 82, 89, and 93 were also phosphorylated but to much lesser extents. Disulfide-bond formation, palmitoylation, and phosphorylation occurred on M2 protein expressed in mammalian cells infected with influenza virus, in mammalian cells in which the M2 protein was expressed from DNA expression vectors, and when the M2 protein was expressed in oocytes of Xenopus laevis. The membrane currents of oocytes of Xenopus laevis expressing wild-type and site-specifically altered forms of the M2 protein, to ablate posttranslational modifications, indicated that none of the posttranslational modifications significantly affected the ion channel activity of the M2 protein in oocytes. Therefore, these data do not indicate a functional role for posttranslational modifications of the M2 protein in its ion channel activity.


[Right ventricular outflow tract reconstruction using monocusp valved outflow patch for pulmonary atresia with ventricular septal defect: influence of the presence of major aorto-pulmonary collateral arteries].
We have preferably utilized monocusp valved outflow patch (MVOP) for right ventricular outflow tract (RVOT) reconstruction in pulmonary atresia with ventricular septal defect (PA + VSD). The purpose of this study was to evaluate the influence of the presence of major aorto-pulmonary collateral arteries (MAPCAs) on probability of MVOP reconstruction and development of RVOT restenosis in midterm. 49 patients underwent complete repair (either MVOP reconstruction or Rastelli procedure) of PA + VSD in our service. These patients were divided into 2 groups: group 1; 21 patients with MAPCAs, group 2; 28 patients without MAPCAs. There was one operative death (group 1). The probably of MVOP reconstruction was similar between group 1 and group 2 (71 vs 79%, p = 0.57, chi 2 test). Follow-up was completed for 48 survivors with the period ranged 3-108 months (mean 47 months). In group 1, one patient died suddenly at home 10 months after surgery. For 47 long-term patients, the ratio of freedom from RVOT restenosis was 72% (95% CI: 52-92%, Kaplan-Meier method) at 5 year. There was no difference between 2 groups (group 1; 73%, 95% CI: 45-100%, group 2; 74%, 95% CI: 48-99%, respectively, p = 0.85 by Log-Rank test). The presence of MAPCAs in PA + VSD was not a risk factor for either the probably of MVOP reconstruction or development of RVOT restenosis in midterm.


Mammalian target of rapamycin inhibition and alloantigen-specific regulatory T cells synergize to promote long-term graft survival in immunocompetent recipients.
Minimization of immunosuppression and donor-specific tolerance to MHC-mismatched organ grafts are important clinical goals. The therapeutic potential of regulatory T cells (Tregs) has been demonstrated, but conditions for optimizing their in vivo function posttransplant in nonlymphocyte-depleted hosts remain undefined. In this study, we address mechanisms through which inhibition of the mammalian target of rapamycin (Rapa) synergizes with alloantigen-specific Treg (AAsTreg) to permit long-term, donor-specific heart graft survival in immunocompetent hosts. Crucially, immature allogeneic dendritic cells allowed AAsTreg selection in vitro, with minimal expansion of unwanted (Th17) cells. The rendered Treg potently inhibited T cell proliferation in an Ag-specific manner. However, these AAsTreg remained unable to control T cells stimulated by allogeneic mature dendritic cells, a phenomenon dependent on the release of proinflammatory cytokines. In vivo, Rapa administration reduced danger-associated IL-6 production, T cell proliferation, and graft infiltration. Based on these observations, AAsTreg were administered posttransplant (day 7) in combination with a short course of Rapa and rendered >80% long-term (>150 d) graft survival, a result superior to that achieved with polyclonal Treg. Moreover, graft protection was alloantigen-specific. Significantly, long-term graft survival was associated with alloreactive T cell anergy. These findings delineate combination of transient mammalian target of Rapa inhibition with appropriate AAsTreg selection as an effective approach to promote long-term organ graft survival.


Starch-Derived Nanographene Oxide Paves the Way for Electrospinnable and Bioactive Starch Scaffolds for Bone Tissue Engineering.
A straightforward process that enabled electrospinning of bioactive starch-based nanofiber scaffolds was developed by utilizing starch derived nano graphene oxide (nGO) as a property enhancer and formic acid as a solvent and esterification reagent. The reaction mechanism and process were followed by detailed spectroscopic investigation. Furthermore, the incorporation of nGO as a "green bioactive additive" endorsed starch nanofibrous scaffolds several advantageous functionalities including improved electrospinnability and thermal stability, good cytocompatibility, osteo-bioactivity, and retained biodegradability. The biodegradable starch/nGO nanofibers underwent simultaneous degradation and mineralization process during 1 week of cell culture and mineralization test, thus, mimicking the structure and function of extracellular matrices (ECMs) and indicating promise for bone tissue engineering applications.


Super magnetic nanoparticles NiFe2O4, coated with aluminum-nickel oxide sol-gel lattices to safe, sensitive and selective purification of his-tagged proteins.
Super magnetic nanoparticle NiFe2O4 with high magnetization, physical and chemical stability was introduced as a core particle which exhibits high thermal stability (>97%) during the harsh coating process. Instead of multi-stage process for coating, the magnetic nanoparticles was mineralized via one step coating by a cheap, safe, stable and recyclable alumina sol-gel lattice (from bohemite source) saturated by nickel ions. The TEM, SEM, VSM and XRD imaging and BET analysis confirmed the structural potential of NiFe2O4@NiAl2O4 core-shell magnetic nanoparticles for selective and sensitive purification of His-tagged protein, in one step. The functionality and validity of the nickel magnetic nanoparticles were attested by purification of three different bioactive His-tagged recombinant fusion proteins including hIGF-1, GM-CSF and bFGF. The bonding capacity of the nickel magnetics nanoparticles was studied by Bradford assay and was equal to 250 ± 84 μg Protein/mg MNP base on protein size. Since the metal ion leakage is the most toxicity source for purification by nickel magnetic nanoparticles, therefor the nickel leakage in purified final protein was determined by atomic absorption spectroscopy and biological activity of final purified protein was confirmed in comparison with reference. Also, in vitro cytotoxicity of nickel magnetic nanoparticles and trace metal ions were investigated by MTS assay analysis. The results confirmed that the synthesized nickel magnetic nanoparticles did not show metal ion toxicity and not affected on protein folding.


A companionship between strangers – learning from fellow people with cancer in oncology wards.
An identification and discussion of learning from experience among hospitalized people with cancer. A good deal of literature focuses on the fact that the needs of patients for information about their disease and treatment should be met by healthcare professionals. Less attention is given to the kind of information that can be provided by personal experience. Patient-patient interaction as a learning situation has not been identified in current research. The study methodology was qualitative and influenced by ethnography. A qualitative approach with participant observation and qualitative interviews was used. From a total of 85 observed people with cancer, ten men and ten women were interviewed in 2010-2011. Data were analysed using qualitative data analysis inspired by Ian Dey and structured using NVivo9. The hospitalized patients learnt about life with cancer from sharing information and personal experiences with fellow patients. Information from fellow patients was complementary to information given by healthcare professionals. Sharing personal experiences led patients to oscillate between four general response stages: the response of fighting, the response of keeping hope, the response of non-acceptance and the response of capitulation. Learning from exchanges of experiences with fellow patients provided a better understanding about the disease and was valued because it was first-hand knowledge. Patients' personal experience of disease is an underused resource in nursing.


Multiple vitrification-warming and biopsy procedures on human embryos: clinical outcome and neonatal follow-up of children.
Does double vitrification and warming of human blastocysts having undergone biopsy once or twice have an impact on the clinical outcome? The clinical pregnancy rate obtained with double vitrification single biopsy blastocysts was comparable to that obtained with single vitrification single biopsy blastocysts in our center in the same time period (46%; 2016-2018), whereas that obtained with double-vitrified double-biopsied blastocysts seemed lower and will need further study. Genetic testing on cryopreserved unbiopsied embryos involves two cryopreservation procedures. Retesting of failed/inconclusive-diagnosed blastocysts inevitably involves a second round of biopsy and a second round of vitrification as well. To what extent this practice impacts on the developmental potential of blastocysts has been studied to a limited extent so far and holds controversy. Additionally, the obstetrical/perinatal outcome after the transfer of double-vitrified/single or double-biopsied blastocysts is poorly documented. This retrospective observational study included 97 cycles of trophectoderm biopsy and preimplantation genetic testing (PGT) on vitrified-warmed embryos followed by a second round of vitrification between March 2015 and December 2019. In 36 warming cycles, no biopsy was performed on the embryos before the first vitrification (single biopsy group). In 61 warming cycles, the embryos had been biopsied on Day 3 (n = 4) or on Day 5/6 (n = 57) before the first vitrification (double biopsy group). A second biopsy was mostly indicated in cycles of failed or inconclusive diagnosis at the first biopsy. Two cycles involved a more specific mutation test for X-linked diseases on male embryos and one cycle involved testing for a second monogenic indication supplementary to a previously tested reciprocal translocation. Post-warming suitability for biopsy, availability of genetically transferable embryos and clinical outcome of subsequent frozen-thawed embryo transfer (FET) cycles were reported. Neonatal follow-up of the children was included. In total, 91 cleavage-stage embryos and 154 blastocysts were warmed, of which 34 (37.4%) and 126 (81.8%), respectively, were of sufficient quality to undergo trophectoderm biopsy and were subsequently vitrified for a second time. Out of these, 92 underwent biopsy for the first time (single biopsy), whereas 68 underwent a second biopsy (double biopsy). After diagnosis, 77 blastocysts (48.1%) were revealed to be genetically transferable (44 in the single biopsy group and 33 in the double biopsy group). In 46 warming cycles, 51 blastocysts were warmed and 49 survived this second warming procedure (96.0%). Subsequently, there were 45 FET cycles resulting in 27 biochemical pregnancies and 18 clinical pregnancies with fetal heartbeat (40.0% per FET cycle: 44.0% in the single biopsy group and 35.0% in the double biopsy group, P = 0.54). Thirteen singletons were born (eight in the single biopsy group and five in the double biopsy group), while three pregnancies were ongoing. A total of 26 embryos (13 in each group) remain vitrified and have the potential to increase the final clinical pregnancy rate. The neonatal follow-up of the children born so far is reassuring. This is a small retrospective cohort, thus, the implantation potential of double vitrification double biopsy blastocysts, as compared to double vitrification single biopsy blastocysts and standard PGT (single vitrification, single biopsy), certainly needs further investigation. Although one could speculate on birthweight being affected by the number of biopsies performed, the numbers in this study are too small to compare birthweight standard deviation scores in singletons born after single or double biopsy. PGT on vitrified-warmed embryos, including a second vitrification-warming step, results in healthy live birth deliveries, for both single- and double-biopsied embryos. The neonatal follow-up of the 13 children born so far did not indicate any adverse effect. The present study is important in order to provide proper counseling to couples on their chance of a live birth per initial warming cycle planned and concerning the safety issue of rebiopsy and double vitrification. None. N/A.


Temporal and spatial evolution of grey matter atrophy in primary progressive multiple sclerosis.
Grey matter (GM) atrophy occurs early in primary progressive MS (PPMS), but it is unknown whether its progression involves different brain regions at different rates, as is seen in other neurodegenerative diseases. We aimed to investigate the temporal and regional evolution of GM volume loss over 5years and its relationship with disability progression in early PPMS. We studied 36 patients with PPMS within five years from onset and 19 age and gender-matched healthy controls with clinical and imaging assessments at study entry and yearly for 3years and then at 5years. Patients were scored on the expanded disability status scale (EDSS) and MS Functional Composite (MSFC) at each time-point. An unbiased longitudinal voxel-based morphometry approach, based on high-dimensional spatial alignment within-subject, was applied to the serial imaging data. The rate of local (voxel-wise) volume change per year was compared between groups and its relationship with clinical outcomes was assessed. Patients deteriorated significantly during the five years follow-up. Patients showed a greater decline of GM volume (p<0.05, FWE-corrected) bilaterally in the cingulate cortex, thalamus, putamen, precentral gyrus, insula and cerebellum when compared to healthy controls over five years, although the rate of volume loss varied across the brain, and was the fastest in the cingulate cortex. Significant (p<0.05, FWE-corrected) volume loss was detected in the left insula, left precuneus, and right cingulate cortex in patients at three years, as compared to baseline, whilst the bilateral putamen and the left superior temporal gyrus showed volume loss at five years. In patients, there was a relationship between a higher rate of volume loss in the bilateral cingulate cortex and greater clinical disability, as measured by the MSFC, at five years (Pearson's r=0.49, p=0.003). Longitudinal VBM demonstrated that the progression of GM atrophy in PPMS occurs at different rates in different regions across the brain. The involvement of the cingulate cortex occurs early in the disease course, continues at a steady rate throughout the follow-up period and is associated with patient outcome. These findings provide new insights into the characteristics of GM atrophy across the brain in MS, and have potential consequences for the selection of brain atrophy as an outcome measure in neuroprotective clinical trials.


Association between antihypertensive drugs and the elderly's oral health- related quality of life: Results of Amirkola cohort study.
Hypertension is one of the most common chronic diseases in the world. The prevalence of hypertension in the elderly is increasing. Patients with high blood pressure have to take medication throughout their lives. In this study, the relationship between antihypertensive drugs and oral health-related quality of life in the elderly was evaluated. This modified cross-sectional study, which is the part of the second phase of the Amirkola Health and Ageing Project (AHAP), was performed on 900 elderly people. Participants included 300 people with hypertension under medical treatment, 300 people with hypertension without medication and 300 people with normal blood pressure. All patients' blood pressure was recorded, and the standard xerostomia questionnaire and GOHAI questionnaire (Geriatric Oral Health Assessment Index) was completed for all participants. Then, the obtained data were analyzed by SPSS 17, whereby student's t-test, ANOVA and chi square, Pearson correlation coefficient and logistic regression model were used. A p<0.05 was considered statistically significant. The mean GOHAI score in the three studied groups: hypertensive under medication treatment, hypertensive without medication treatment and normal blood pressure (51.1±7.4, 51.7±7.3, 51.1±7.5, respectively) did not differ significantly (P=0.533).The frequency of xerostomia was significantly different in the three groups (P=0.008). Among the antihypertensive drugs, the highest rate of xerostomia was due to the use of calcium channel blockers (31.1%) and diuretics (26.8%). In our study, although antihypertensive medications were associated with xerostomia, they did not decrease the oral health-related quality of life.