Determinants of the variability of aflatoxin-albumin adduct levels in Ghanaians.
Hepatocellular carcinoma (HCC) is a multifactorial disease with various host and environmental factors involved in its etiology. Of these, aflatoxin exposure has been established as an important risk factor in the development of HCC; the presence of aflatoxin-albumin (AA) adducts in the blood serves as a valuable biomarker of human exposure. In this study, the relationship between a variety of different HCC host factors and the incidence of AA adduct levels was examined in a Ghanaian population at high risk for HCC. These factors included age, gender, hepatitis virus B (HVB) and hepatitis C virus (HCV) status, and genetic polymorphisms in both microsomal epoxide hydrolase (mEH) and glutathione S-transferases (GSTs). Blood samples were analyzed for AA adducts and HBV and HCV status. GSTM1 and GSTT1 deletion polymorphisms and mEH exon 3 and exon 4 single-nucleotide polymorphisms (SNPs) were determined from urine samples. In univariate analysis, age, HBV and HVC status, and GSTT1 and mEH exon 3 genotypes were not associated with AA adduct levels. However, mean adduct levels were significantly higher in both females and individuals typed heterozygous for mEH exon 4 (vs. wild types). Stratification analysis also showed that gender along with mEH exon 4 genotype and HBV status had a significant effect on adduct levels. Both females typed HBsAg+ and males with mEH exon 4 heterozygote genotypes showed significantly higher adduct levels as compared to the HBsAg- and wild types, respectively. Understanding the relationships between these host factors and the variability in aflatoxin-adduct levels may help in identifying susceptible populations in developing countries and for targeting specific public health interventions for the prevention of aflatoxicoses in populations with HCC and chronic liver diseases.


Hereditary angioedema: the use of fresh frozen plasma for prophylaxis in patients undergoing oral surgery.
Six patients with hereditary anagioedema (HAE) undergoing 7 episodes of dental surgery received transfusions with fresh frozen plasma one day before surgery. Although the morbidity observed in these patients following similar procedures had been high, no significant complications of surgery were noted with this therapy. Thus, fresh frozen plasma infusion appears to provide a safe and effective method of prophylaxis in patients with HAE. Following infusion of fresh frozen plasma, serum levels of C4 esterase inhibitor (C1EI) rose transiently, and then fell to preinfusion levels within 1 to 12 days. In all but one patient the rise in C4 was greater than could be accounted for by the amount of C4 infused. In no patient did the level of C1EI or C4 rise to within the normal range. The data raise the question of the role of C1EI in the pathogenesis of angioedema in these patients.


Carpal tunnel syndrome.
Carpal tunnel syndrome is the most common compressive neuropathy of the upper extremity. As a result of median nerve compression, the patient reports pain, weakness, and paresthesias in the hand and digits. The etiology of this condition is multifactorial; anatomic, systemic, and occupational factors have all been implicated. The diagnosis is based on the patient history and physical examination and is confirmed by electrodiagnostic testing. Treatment methods range from observation and splinting, to cortisone injection and splinting, to surgical intervention. Both nonsurgical and surgical management provide symptom relief in most patients. The results of open and endoscopic surgery essentially are equivalent at 3 months; the superiority of one technique over the other has yet to be established.


Facilitating sensory responses in developing mouse somatosensory barrel cortex.
The sensory responses in the barrel cortex of mice aged postnatal day (P)7-P12 evoked by a single whisker deflection are smaller in amplitude and spread over a smaller area than those measured in P13-P21 mice. However, repetitive 10-Hz stimulation or paired pulse whisker stimulation in P7-P12 mice evoked facilitating sensory responses, contrasting with the depressing sensory responses observed in P13-P21 mice. This facilitation occurred during an interval ranging 300-1,000 ms after the first stimulus and was measured using whole cell recordings, voltage-sensitive dye imaging, and calcium-sensitive dye imaging. The facilitated responses were not only larger in amplitude but also propagated over a larger cortical area. The facilitation could be blocked by local application of pharmacological agents reducing cortical excitability. Local cortical microstimulation could substitute for the first whisker stimulus to produce a facilitated sensory response. The enhanced sensory responses evoked by repetitive sensory stimuli in P7-P12 mice may contribute to the activity-dependent specification of the developing cortical circuits. In addition, the facilitating sensory responses allow long integration times for sensory processing compatible with the slow behavior of mice during early postnatal development.


Integration of mass spectrometry into early-phase discovery and development of central nervous system agents.
The early-phase discovery and development of useful central nervous system (CNS) agents present ample opportunities to exploit mass spectrometry and provide detailed compound/mixture characterization, or to make the process faster and/or more economic. Neuropeptide FF antagonists and centrally active thyrotropin-releasing hormone analogues were used as specific examples in this work. We evaluated the characterization of focused libraries of peptide derivatives by electrospray ionization, tandem mass spectrometry and liquid chromatography/tandem mass spectrometry on a quadrupole ion trap and nanoelectrospray on a Fourier transform ion cyclotron resonance mass spectrometer. Immobilized artificial-membrane chromatography was employed as a model to predict/rank new agents against lead compounds for their potential to reach the central nervous system in pharmacologically significant amounts. Measuring brain concentrations in rodents after the intravenous administration of test compounds was used as an in vivo approach, and we took advantage of microdialysis sampling that furnished samples without interfering tissue matrix and afforded the estimation of extracellular concentrations in a localized part of the brain. Overall, making atmospheric-pressure ionization mass spectrometry an integral part of the process has played a major role in increasing throughput, selectivity, specificity and detection sensitivity and thereby providing useful information about the extent or mechanism of transport and metabolic activation/inactivation in early-phase discovery and development of CNS agents.


HSP90 inhibition overcomes EGFR amplification-induced resistance to third-generation EGFR-TKIs.
Patients with non-small cell lung cancer (NSCLC) harboring activating EGFR mutations are sensitive to epidermal growth factor receptor-tyrosine kinase inhibitors (EGFR-TKIs) but inevitably develop resistance to the inhibitors mostly through acquisition of the secondary T790M mutation. Although third-generation EGFR-TKIs overcome this resistance by selectively inhibiting EGFR with EGFR-TKI-sensitizing and T790M mutations, acquired resistance to third-generation EGFR-TKIs invariably develops. Next-generation sequencing (NGS) and fluorescence in situ hybridization (FISH) analysis were performed in an EGFR T790M-mutated NSCLC patient who had progressed after a third-generation EGFR-TKI, TAS-121. EGFR-mutated cell lines were subjected to a cell proliferation assay and western blotting analysis with EGFR-TKIs and a heat shock protein 90 (HSP90) inhibitor. NGS and FISH analysis revealed EGFR amplification in the resistant cancer cells. While EGFR L858R/T90M-mutated cell line was sensitive to osimertinib or TAS-121 in vitro, EGFR-overexpressing cell lines displayed resistance to these EGFR-TKIs. Western blot analysis showed that EGFR phosphorylation and overexpression of EGFR in cell lines was not suppressed by third-generation EGFR-TKIs. In contrast, an HSP90 inhibitor reduced total and phosphorylated EGFR and inhibited the proliferation of resistant cell lines. EGFR amplification confers resistance to third-generation EGFR-TKIs which can be overcome by HSP90 inhibition. The results provide a preclinical rationale for the use of HSP90 inhibitors to overcome EGFR amplification-mediated resistance.


Increased cell proliferation and neurogenesis in the hippocampal dentate gyrus of old GFAP(-/-)Vim(-/-) mice.
In response to central nervous system (CNS) injury, and more discretely so also during aging, astrocytes become reactive and increase their expression of the intermediate filament proteins glial fibrillary acidic protein (GFAP) and vimentin. Studies of mice deficient in astrocytic intermediate filaments have provided insights into the function of reactive gliosis. Recently we demonstrated robust integration of retinal transplants (1) and increased posttraumatic synaptic regeneration (2) in GFAP(-/-)Vim(-/-) mice, suggesting that modulation of astrocyte activity affects the permissiveness of the CNS environment for regeneration. Neurogenesis in the adult mammalian CNS is restricted to essentially two regions, the hippocampus and the subventricular zone. Here, we assessed neurogenesis in the hippocampus of 18-month-old GFAP(-/-)Vim(-/-) mice. In the granular layer of the dentate gyrus, cell proliferation/survival was 34% higher and neurogenesis 36% higher in GFAP(-/-)Vim(-/-) mice than in wildtype controls. These findings suggest that the adult hippocampal neurogenesis in healthy old mice can be increased by modulating astrocyte reactivity.


In vivo electrochemical studies of dopamine clearance in subregions of rat nucleus accumbens: differential properties of the core and shell.
The dopamine (DA) uptake/clearance properties of the DA transporter (DAT) in the core and shell of the nucleus accumbens were measured using in vivo electrochemical recordings. Calibrated amounts of a DA solution were pressure-ejected from a micropipette/electrode assembly placed in the core or shell of the nucleus accumbens in anesthetized male Fischer 344 rats. Initial studies in the two brain regions revealed that the core and shell have different DA clearance properties as measured by the extracellular DA signal amplitudes, clearance times, and clearance rates. Although the same number of picomoles of DA were applied, DA clearance signals recorded in shell had significantly greater amplitudes but faster clearance rates than those recorded in the core. Systemic administration of 20 mg/kg cocaine, a monoamine transporter inhibitor, greatly increased the signal amplitude from the locally applied DA in both the core and shell. Signal amplitudes were increased to a greater extent in the shell, compared with the core, after cocaine administration. However, cocaine affected the clearance time of DA only in the core and the DA clearance rate only in the shell. Taken together with previously reported data, these studies further support differential activity of the DAT in the core versus shell subregions of the nucleus accumbens. In addition, these data indicate that DATs are more sensitive to the effects of psychomotor stimulants, such as cocaine, in the shell of the nucleus accumbens.


Understanding dissociation and insight in the treatment of shortness of breath with hypnosis: a case study.
Training in hypnosis is particularly valuable for the physician seeking to better appreciate the interplay between mind and body. Through such experiences the physician can learn that presentation of symptoms often is affected by patients' psychological states, and that symptoms sometimes serve as solutions for patients' psychological dilemmas. The presented case study demonstrates how an 11-year-old's complaint of shortness of breath becomes an opportunity for an appropriately trained physician to provide treatment by helping the patient to engage his inner resources. The case illustrates the strength of hypnosis for accessing resources outside of conscious awareness and use of dissociative language to both support and alter the patient's defenses. We discuss the role of hypnosis when working psychodynamically with a patient, and whether and when insight is important or necessary for change of behavior.


The definition of IVM is clear-variations need defining.
Oocyte in vitro maturation (IVM) is currently defined as the maturation in vitro of immature cumulus-oocyte complexes collected from antral follicles. This is the original definition as first described by Pincus and Enzmann and then by Edwards many decades ago, and this clear and unambiguous definition has served us well ever since. In an attempt to clarify apparent differences among clinicians, the following revised definition of IVM was recently proposed: 'The retrieval of oocytes from small and intermediate sized follicles in an ovary before the largest follicle has surpassed 13 mm in mean diameter'. As such, this proposed definition should encompass the use of hCG triggering. To change the clear and long-serving definition of IVM to fit varying clinical practices requires a compelling justification based on solid scientific and clinical grounds. We are of the opinion that the proposed revised definition of IVM is counterintuitive as it includes protocols that are intended to mature oocytes in vivo The proposed definitions are cumbersome and indeed further complicate the situation. It is not scientifically rational to base the definition on follicular size, and the definition ignores the vast corporate knowledge acquired from the many decades and >6000 publications in animal research that universally practices IVM as per the existing definition. Furthermore, such a definition can lead to false results in interpreting the follow-up of children conceived using IVM. Hence, we see no rationale to change the existing definition of IVM. However, we agree that variations on IVM require alternative nomenclature-these definitions need to be intuitive and need to clearly distinguish themselves from the existing long-standing definition of IVM. This would help to clarify the recent confusion within the clinical ART community as to what is and what is not, IVM.


The effects of pre-intervention mindset induction on a brief intervention to increase risk perception and reduce alcohol use among university students: A pilot randomized controlled trial.
Brief interventions based on personalized feedback have shown promising results in reducing risky alcohol use among university students. We investigated the effects of activating deliberative (predecisional) or implemental (postdecisional) mindsets on the effectiveness of a standardized brief intervention, the ASSIST-linked Brief Intervention. This intervention comprises a personalized feedback and a decisional balance exercise. We hypothesized that participants in a deliberative mindset should show better outcomes related to risk perception and behavior than participants in an implemental mindset. A sample of 257 students provided baseline measures on risk perception, readiness to change, and alcohol use. Of those, 64 students with risky alcohol use were randomly allocated to one of two mindset induction conditions-deliberative or implemental mindset. Thereafter, they received the ASSIST-linked Brief Intervention and completed self-report questionnaires on changes in risk perception, alcohol use, and readiness to change at post-intervention and four-week follow-up. In contrast to our hypotheses, the four-weeks follow-up revealed that participants in the implemental mindset consumed significantly less alcohol than participants in a deliberative mindset did. The former decreased and the latter increased their alcohol intake; resistance to the brief intervention was stronger in the latter condition. However, neither deliberative nor implemental mindset participants showed any changes in risk perceptions or in their readiness to change alcohol consumption. These findings suggest that mindset induction is a powerful moderator of the effects of the ASSIST-linked Brief Intervention. We argue that systematic research on mindset effects on brief intervention techniques aimed to reduce risky alcohol use is highly needed in order to identify the processes involved with commitment and resistance being the main candidates.


Dynamic Arterial Elastance Is Associated With the Vascular Waterfall in Patients Treated With Norepinephrine: An Observational Study.
Introduction: It has been suggested that dynamic arterial elastance (Eadyn) can predict decreases in arterial pressure in response to changing norepinephrine levels. The objective of this study was to determine whether Eadyn is correlated with determinants of the vascular waterfall [critical closing pressure (CCP) and systemic arterial resistance (SARi)] in patients treated with norepinephrine. Materials and Methods: Patients treated with norepinephrine for vasoplegia following cardiac surgery were studied. Vascular and flow parameters were recorded immediately before the norepinephrine infusion and then again once hemodynamic parameters had been stable for 15 min. The primary outcomes were Eadyn and its associations with CCP and SARi. The secondary outcomes were the associations between Eadyn and vascular/flow parameters. Results: At baseline, all patients were hypotensive with Eadyn of 0.93 [0.47;1.27]. Norepinephrine increased the arterial blood pressure, cardiac index, CCP, total peripheral resistance (TPRi), arterial elastance, and ventricular elastance and decreased Eadyn [0.40 (0.30;0.60)] and SARi. Eadyn was significantly associated with arterial compliance (CA), CCP, and TPRi (p < 0.05). Conclusion: In patients with vasoplegic syndrome, Eadyn was correlated with determinants of the vascular waterfall. Eadyn is an easy-to-read functional index of arterial load that can be used to assess the patient's macro/microcirculatory status. Clinical Trial Registration: ClinicalTrials.gov #NCT03478709.


Biochemical Characterization of Kat1: a Domesticated hAT-Transposase that Induces DNA Hairpin Formation and MAT-Switching.
Kluyveromyces lactis hAT-transposase 1 (Kat1) generates hairpin-capped DNA double strand breaks leading to MAT-switching (MATa to MATα). Using purified Kat1, we demonstrate the importance of terminal inverted repeats and subterminal repeats for its endonuclease activity. Kat1 promoted joining of the transposon end into a target DNA molecule in vitro, a biochemical feature that ties Kat1 to transposases. Gas-phase Electrophoretic Mobility Macromolecule analysis revealed that Kat1 can form hexamers when complexed with DNA. Kat1 point mutants were generated in conserved positions to explore structure-function relationships. Mutants of predicted catalytic residues abolished both DNA cleavage and strand-transfer. Interestingly, W576A predicted to be impaired for hairpin formation, was active for DNA cleavage and supported wild type levels of mating-type switching. In contrast, the conserved CXXH motif was critical for hairpin formation because Kat1 C402A/H405A completely blocked hairpinning and switching, but still generated nicks in the DNA. Mutations in the BED zinc-finger domain (C130A/C133A) resulted in an unspecific nuclease activity, presumably due to nonspecific DNA interaction. Kat1 mutants that were defective for cleavage in vitro were also defective for mating-type switching. Collectively, this study reveals Kat1 sharing extensive biochemical similarities with cut and paste transposons despite being domesticated and evolutionary diverged from active transposons.


Experimental investigation on the effect of drill quality on the performance of bone drilling.
Bone drilling is a well-known process in operative fracture treatment and reconstructive surgery. The cutting ability of the drill is lost when used for multiple times. In this study, the effect of different levels of drill wear on bone temperature, drilling force, torque, delamination around the drilling region and surface roughness of the hole was investigated using a series of experiments. Experimental results demonstrated that the wear of the drill is strongly related to the drilling force, torque, temperature and surface roughness of the drilled hole. Statistical analysis was performed to find the effect of various factors on multiple response variables in the bone drilling process. The favorable conditions for bone drilling are obtained when feed rate, drill speed and the roughness of the cutting edge of the drill were fixed at 30 mm, 2000 rpm and up to 2 mm, respectively. Further, analysis of variance (ANOVA) was performed to determine the factor with a significant impact on the response variables. F-test and p-value indicated that the feed rate had the highest effect on grey relational grade followed by the roughness of the drill. This study suggests that the sharp drill along with controlled drilling speed and feed rate may be used for safe and efficient surgical drilling in bone.


Reconstitution of KCNE1 into lipid bilayers: comparing the structural, dynamic, and activity differences in micelle and vesicle environments.
KCNE1 (minK), found in the human heart and cochlea, is a transmembrane protein that modulates the voltage-gated potassium KCNQ1 channel. While KCNE1 has previously been the subject of extensive structural studies in lyso-phospholipid detergent micelles, key observations have yet to be confirmed and refined in lipid bilayers. In this study, a reliable method for reconstituting KCNE1 into lipid bilayer vesicles composed of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) and 1-palmitoyl-2-oleoyl-sn-glycero-3-phospho(1'-rac-glycerol) (sodium salt) (POPG) was developed. Microinjection of the proteoliposomes into Xenopus oocytes expressing the human KCNQ1 (K(V)7.1) voltage-gated potassium channel led to nativelike modulation of the channel. Circular dichroism spectroscopy demonstrated that the percent helicity of KCNE1 is significantly higher for the protein reconstituted in lipid vesicles than for the previously described structure in 1.0% 1-myristoyl-2-hydroxy-sn-glycero-3-phospho(1'-rac-glycerol) (sodium salt) (LMPG) micelles. SDSL electron paramagnetic resonance spectroscopic techniques were used to probe the local structure and environment of Ser28, Phe54, Phe57, Leu59, and Ser64 of KCNE1 in both POPC/POPG vesicles and LMPG micelles. Spin-labeled KCNE1 cysteine mutants at Phe54, Phe57, Leu59, and Ser64 were found to be located inside POPC/POPG vesicles, whereas Ser28 was found to be located outside the membrane. Ser64 was shown to be water inaccessible in vesicles but found to be water accessible in LMPG micelle solutions. These results suggest that key components of the micelle-derived structure of KCNE1 extend to the structure of this protein in lipid bilayers but also demonstrate the need to refine this structure using data derived from the bilayer-reconstituted protein to more accurately define its native structure. This work establishes the basis for such future studies.


Reclassification of Anaerobaculum mobile, Anaerobaculum thermoterrenum, Anaerobaculum hydrogeniformans as Acetomicrobium mobile comb. nov., Acetomicrobium thermoterrenum comb. nov. and Acetomicrobium hydrogeniformans comb. nov., respectively, and emendation of the genus Acetomicrobium.
Taking into account their 16S rRNA gene sequences, it appears that Acetomicrobium flavidum and the three species of the genus Anaerobaculum described so far belong to the same phylogenetic clade with high levels (>95 %) of similarity. In this respect, these three Anaerobaculum species should be reclassified within the genus Acetomicrobium, which has priority over the genus Anaerobaculum, which was validated since the genus Acetomicrobium. The DNA G+C content of Acetomicrobium flavidum is 47.1 mol%, which is of the same order as that of the three Anaerobaculum species. All these bacteria have in common iso-C15 : 0 as their main fatty acid. Based on further phylogenetic, genetic and chemotaxonomic studies, we propose that Anaerobaculum mobile ( = DSM 13181T = JCM 12221T), Anaerobaculum thermoterrenum ( = DSM 13490T = ACM 5076T) and Anaerobaculum hydrogeniformans ( = DSM 22491T = ATCC BAA-1850T) be reclassified as Acetomicrobium mobile comb. nov., Acetomicrobium thermoterrenum comb. nov. and Acetomicrobium hydrogeniformans comb. nov., respectively. The four bacterial species belong to the phylum Synergistetes.


Evidence for two tumor suppressor loci associated with proximal chromosome 9p to q and distal chromosome 9q in bladder cancer and the initial screening for GAS1 and PTC mutations.
The most common genetic alteration identified to date in bladder cancer is loss of heterozygosity (LOH) of chromosome 9, suggesting the presence of possible tumor suppressor genes on this chromosome. We attempted to map the location of these genes by analyzing 69 primary transitional cell carcinomas of the bladder with a panel of microsatellite markers for LOH on chromosome 9. Monosomy 9 (defined by LOH of all informative markers analyzed on 9p and 9q) was detected in 26 of 69 (38%) tumors, and 22 of 69 (32%) tumors showed subchromosomal deletions. Twelve tumors (17%) demonstrated partial LOH of chromosome 9 and indicated two distinct regions of LOH. Eight tumors showed distal allelic loss of 9q with a minimal region of common deletion flanked proximally by marker GSN on 9q33. Six tumors showed proximal allelic loss of 9p and 9q with a minimal area of common deletion flanked by markers D9S970 on 9p12 and D9S283 on 9q21. Two tumors showed loss of both the distal region of 9q and the proximal region of 9p and 9q, which were separated by a possible 6-44 cM of retained genetic material. The proximal minimal area of common deletion excluded 9q22.3-q31 to where two putative tumor suppressor genes, the nevoid basal cell carcinoma syndrome and multiple self-healing squamous epithelioma (ESS1) genes, have been mapped. The growth arrest-specific gene (GAS1), a candidate tumor suppressor gene, was included within the proximal minimal region. We evaluated the GAS1 gene for its potential role in bladder cancer using single-strand conformational polymorphism to screen for mutations in GAS1 in 10 bladder cancer cell lines and 14 primary bladder tumors. A polymorphism at codon 88 was noted in one primary bladder tumor, but no other abnormalities were found, suggesting that another potential tumor suppressor gene important to bladder cancer resides in these minimally deleted regions. Because the nevoid basal cell carcinoma syndrome gene has long been speculated to be a putative tumor suppressor gene in bladder cancer and this gene has recently been characterized as the human homologue of the Drosophila patched gene (PTC), 20 primary bladder tumors with chromosome 9q LOH were screened for mutations in PTC using single-strand conformational polymorphism and heteroduplex analysis. No alterations were found in any of the samples analyzed. Furthermore, 4 of 37 noninvasive papillary (Ta) tumors demonstrated loss of all 9q markers with retention of 9p, whereas no Ta tumor showed loss of 9p with retention of all 9q markers, suggesting that LOH of 9q is the earlier event in bladder tumorigenesis. In summary, our results indicate two tumor suppressor loci associated with proximal chromosome 9p to q and distal chromosome 9q that may be important in bladder cancer. GAS1 and PTC do not seem to be frequently mutated in bladder cancer.


17th Century Variola Virus Reveals the Recent History of Smallpox.
Smallpox holds a unique position in the history of medicine. It was the first disease for which a vaccine was developed and remains the only human disease eradicated by vaccination. Although there have been claims of smallpox in Egypt, India, and China dating back millennia [1-4], the timescale of emergence of the causative agent, variola virus (VARV), and how it evolved in the context of increasingly widespread immunization, have proven controversial [4-9]. In particular, some molecular-clock-based studies have suggested that key events in VARV evolution only occurred during the last two centuries [4-6] and hence in apparent conflict with anecdotal historical reports, although it is difficult to distinguish smallpox from other pustular rashes by description alone. To address these issues, we captured, sequenced, and reconstructed a draft genome of an ancient strain of VARV, sampled from a Lithuanian child mummy dating between 1643 and 1665 and close to the time of several documented European epidemics [1, 2, 10]. When compared to vaccinia virus, this archival strain contained the same pattern of gene degradation as 20th century VARVs, indicating that such loss of gene function had occurred before ca. 1650. Strikingly, the mummy sequence fell basal to all currently sequenced strains of VARV on phylogenetic trees. Molecular-clock analyses revealed a strong clock-like structure and that the timescale of smallpox evolution is more recent than often supposed, with the diversification of major viral lineages only occurring within the 18th and 19th centuries, concomitant with the development of modern vaccination.


Network clustering revealed the systemic alterations of mitochondrial protein expression.
The mitochondrial protein repertoire varies depending on the cellular state. Protein component modifications caused by mitochondrial DNA (mtDNA) depletion are related to a wide range of human diseases; however, little is known about how nuclear-encoded mitochondrial proteins (mt proteome) changes under such dysfunctional states. In this study, we investigated the systemic alterations of mtDNA-depleted (ρ(0)) mitochondria by using network analysis of gene expression data. By modularizing the quantified proteomics data into protein functional networks, systemic properties of mitochondrial dysfunction were analyzed. We discovered that up-regulated and down-regulated proteins were organized into two predominant subnetworks that exhibited distinct biological processes. The down-regulated network modules are involved in typical mitochondrial functions, while up-regulated proteins are responsible for mtDNA repair and regulation of mt protein expression and transport. Furthermore, comparisons of proteome and transcriptome data revealed that ρ(0) cells attempted to compensate for mtDNA depletion by modulating the coordinated expression/transport of mt proteins. Our results demonstrate that mt protein composition changed to remodel the functional organization of mitochondrial protein networks in response to dysfunctional cellular states. Human mt protein functional networks provide a framework for understanding how cells respond to mitochondrial dysfunctions.


A novel approach for autism spectrum condition patients with eating disorders: Analysis of treatment cost-savings.
In the current economic context, it is critical to ensure that eating disorder (ED) treatments are both effective and cost-effective. We describe the impact of a novel clinical pathway developed to better meet the needs of autistic patients with EDs on the length and cost of hospital admissions. The pathway was based on the Institute for Healthcare's Model of Improvement methodology, using an iterative Plan, Do, Study, Act format to introduce change and to co-produce the work with people with lived experience and with healthcare professionals. We explored the change in length and cost of admissions before and after the pathway was introduced. Preliminary results suggest that the treatment innovations associated with this pathway have led to reduced lengths of admission for patients with the comorbidity, which were not seen for patients without the comorbidity. Estimated cost-savings were approximately £22,837 per patient and approximately £275,000 per year for the service as a whole. Going forward, our aim is to continue to evaluate the effectiveness and cost-effectiveness of investment in the pathway to determine whether the pathway improves the quality of care for patients with a comorbid ED and autism and is good value for money.


Prematurity affects cortical maturation in early childhood.
Cortical development in the first years of age for children with very low birth weight is not well characterized. We obtained high-resolution structural magnetic resonance images from children aged 18-22 months (16 very low birth weight/7 term) and 3-4 years (12 very low birth weight/8 term). Cortical surface area and thickness of the brain were assessed using the FreeSurfer data analysis program, and manually inspected for accuracy. For children with very low birth weight, a negative correlation was evident between birth weight and cortical thickness at 18-22 months (P = 0.04), and a positive correlation with cortical surface area at 3-4 years (P = 0.02). Between groups, children with very low birth weight demonstrated a consistent trend for thicker cortices and reduced surface area, compared with control term children (18-22 month surface area, P = 0.08; thickness, P = 0.11; 3-4 year surface area, P = 0.73; thickness, P = 0.14). The normal processes of cortical thinning and surface area expansion in the first several years of age may be delayed by premature delivery, a potentially more prominent effect with greater degrees of prematurity.


Selecting elephant grass families and progenies to produce bioenergy through mixed models (REML/BLUP).
Brazil has great potential to produce bioenergy since it is located in a tropical region that receives high incidence of solar energy and presents favorable climatic conditions for such purpose. However, the use of bioenergy in the country is below its productivity potential. The aim of the current study was to select full-sib progenies and families of elephant grass (Pennisetum purpureum S.) to optimize phenotypes relevant to bioenergy production through mixed models (REML/BLUP). The circulating diallel-based crossing of ten elephant grass genotypes was performed. An experimental design using the randomized block methodology, with three repetitions, was set to assess both the hybrids and the parents. Each plot comprised 14-m rows, 1.40 m spacing between rows, and 1.40 m spacing between plants. The number of tillers, plant height, culm diameter, fresh biomass production, dry biomass rate, and the dry biomass production were assessed. Genetic-statistical analyses were performed through mixed models (REML/BLUP). The genetic variance in the assessed families was explained through additive genetic effects and dominance genetic effects; the dominance variance was prevalent. Families such as Capim Cana D'África x Guaçu/I.Z.2, Cameroon x Cuba-115, CPAC x Cuba-115, Cameroon x Guaçu/I.Z.2, and IAC-Campinas x CPAC showed the highest dry biomass production. The family derived from the crossing between Cana D'África and Guaçu/I.Z.2 showed the largest number of potential individuals for traits such as plant height, culm diameter, fresh biomass production, dry biomass production, and dry biomass rate. The individual 5 in the family Cana D'África x Guaçu/I.Z.2, planted in blocks 1 and 2, showed the highest dry biomass production.


Multi-Regression based supervised sample selection for predicting baby connectome evolution trajectory from neonatal timepoint.
The connectional map of the baby brain undergoes dramatic changes over the first year of postnatal development, which makes its mapping a challenging task, let alone learning how to predict its evolution. Currently, learning models for predicting brain connectomic developmental trajectories remain broadly absent despite their great potential in spotting atypical neurodevelopmental disorders early. This is most likely due to the scarcity and often incompleteness of longitudinal infant neuroimaging studies for training such models. In this paper, we propose the first approach for progressively predicting longitudinal development of brain networks during the postnatal period solely from a baseline connectome around birth. To this end, a supervised multi-regression sample selection strategy is designed to learn how to identify the best set of neighbors of a testing baseline connectome to eventually predict its evolution trajectory at follow-up timepoints. However, given that the training dataset may have missing samples (connectomes) at certain timepoints, this may affect the training of the predictive model. To overcome this problem, we perform a low-rank tensor completion based on a robust principal component analysis to impute the missing training connectomes by linearly approximating similar complete training networks. In the prediction step, our sample selection strategy aims to preserve spatiotemporal relationships between consecutive timepoints. Therefore, the proposed method learns how to identify the set of the local closest neighbors to a target network by training an ensemble of bidirectional regressors leveraging temporal dependency between consecutive timepoints with a recall to the baseline observations to progressively predict the evolution of a testing network over time. Our method achieves the best prediction results and better captures the dynamic changes of each brain connectome over time in comparison to its ablated versions using leave-one-out cross-validation strategy.


Tumor tropic delivery of doxorubicin-polymer conjugates using mesenchymal stem cells for glioma therapy.
Mesenchymal stem cells (MSCs) possess inherent tropism to malignant cells and home to disseminated tumor foci, making MSCs an ideal candidate as targeting vehicle for drug delivery. Our previously synthesized doxorubicin (DOX)-polymer conjugates (PPCD) modified by RGD (RGD-PPCD) harness tremendous potential with active targeting and sustained release. To further enhance the conjugates penetrability and tumor tracking capability, MSCs-mediated RGD-/PPCD delivery system was created and constructed by spontaneous incorporation. The present study investigated the anti-tumor activities of MSCs loaded with RGD-/PPCD (MSCsRGD-/PPCD), compared to their corresponding RGD-/PPCD and MSCs loaded with DOX (MSCsDOX), especially. MTT assay demonstrated that although RGD-/PPCD could inhibit MSCs proliferation, there was no direct cytotoxicity to MSCs. After optimization, maximum drug loading amount was obtained by incubating MSCs for 12 h at the drug concentration of 50 μg/mL (DOX-equiv.). Further drug release and intracellular retention proved that, compared to MSCsDOX, MSCsRGD-/PPCD delayed the release peak to 12-48 h with continuous drug liberation more than 5 days, declaring the time dependent release behavior and lasting stability of conjugates in MSCs for cell-directed tumoritropic delivery. For MSCsRGD-/PPCD, intrinsic cell functions without compromise were verified by cell cycle analysis, transmigration assay and tumor penetration, while MSCsDOX showed observable but not significant suppression. As for in vivo studies, among all groups, mice intracranially administrated with MSCsPPCD and MSCsRGD-PPCD acquired the longest survival time with median being 41 and 45 days, and increased life span with 34% and 46.8% versus RGD-PPCD treatment group, respectively. Additionally, MSCsRGD-/PPCD exhibited more extensive penetration and enhanced tumor cell apoptosis than MSCsDOX. As proof of the concept, without any genetic manipulation and chemical modification, a novel modality integrating MSCs and DOX-polymer conjugates could vigorously ship therapeutics for efficient glioma therapy.


Neural mechanisms mediating optimism bias.
Humans expect positive events in the future even when there is no evidence to support such expectations. For example, people expect to live longer and be healthier than average, they underestimate their likelihood of getting a divorce, and overestimate their prospects for success on the job market. We examined how the brain generates this pervasive optimism bias. Here we report that this tendency was related specifically to enhanced activation in the amygdala and in the rostral anterior cingulate cortex when imagining positive future events relative to negative ones, suggesting a key role for areas involved in monitoring emotional salience in mediating the optimism bias. These are the same regions that show irregularities in depression, which has been related to pessimism. Across individuals, activity in the rostral anterior cingulate cortex was correlated with trait optimism. The current study highlights how the brain may generate the tendency to engage in the projection of positive future events, suggesting that the effective integration and regulation of emotional and autobiographical information supports the projection of positive future events in healthy individuals, and is related to optimism.


Formation of hydroxyl radicals contributes to the bactericidal activity of ciprofloxacin against Pseudomonas aeruginosa biofilms.
Antibiotic-tolerant, biofilm-forming Pseudomonas aeruginosa has long been recognized as a major cause of chronic lung infections of cystic fibrosis patients. The mechanisms involved in the activity of antibiotics on biofilm are not completely clear. We have investigated whether the proposed induction of cytotoxic hydroxyl radicals (OH˙) during antibiotic treatment of planktonically grown cells may contribute to action of the commonly used antibiotic ciprofloxacin on P. aeruginosa biofilms. For this purpose, WT PAO1, a catalase deficient ΔkatA and a ciprofloxacin resistant mutant of PAO1 (gyrA), were grown as biofilms in microtiter plates and treated with ciprofloxacin. Formation of OH˙ and total amount of reactive oxygen species (ROS) was measured and viability was estimated. Formation of OH˙ and total ROS in PAO1 biofilms treated with ciprofloxacin was shown but higher levels were measured in ΔkatA biofilms, and no ROS production was seen in the gyrA biofilms. Treatment with ciprofloxacin decreased the viability of PAO1 and ΔkatA biofilms but not of gyrA biofilms. Addition of thiourea, a OH˙ scavenger, decreased the OH˙ levels and killing of PAO1 biofilm. Our study shows that OH˙ is produced by P. aeruginosa biofilms treated with ciprofloxacin, which may contribute to the killing of biofilm subpopulations.


Mutations in APC, CTNNB1 and K-ras genes and expression of hMLH1 in sporadic colorectal carcinomas from the Netherlands Cohort Study.
The early to intermediate stages of the majority of colorectal tumours are thought to be driven by aberrations in the Wnt (APC, CTNNB1) and Ras (K-ras) pathways. A smaller proportion of cancers shows mismatch repair deficiency. The aim of this study was to analyse the co-occurrence of these genetic alterations in relation to tumour and patient characteristics. In a group of 656 unselected sporadic colorectal cancer patients, aberrations in the APC, K-ras, CTNNB1 genes, and expression of hMLH1 were investigated. Additionally, tumours were divided in groups based on molecular features and compared with respect to patient's age at diagnosis, sex, family history of colorectal cancer, tumour sub-localisation, Dukes' stage and differentiation. Mutations at the phosphorylation sites (codons 31, 33, 37, and 45) in the CTNNB1 gene were observed in tumours from only 5/464 patients. Tumours with truncating APC mutations and activating K-ras mutations in codons 12 and 13 occurred at similar frequencies (37% (245/656) and 36% (235/656), respectively). Seventeen percent of tumours harboured both an APC and a K-ras mutation (109/656). Nine percent of all tumours (58/656) lacked hMLH1 expression. Patients harbouring a tumour with absent hMLH1 expression were older, more often women, more often had proximal colon tumours that showed poorer differentiation when compared to patients harbouring tumours with an APC and/or K-ras mutation. CTNNB1 mutations seem to be of minor importance in sporadic colorectal cancer. The main differences in tumour and patient characteristics are found between groups of patients based on mismatch repair deficiency.


Prospective screening study of 0.5 Tesla dedicated magnetic resonance imaging for the detection of breast cancer in young, high-risk women.
Evidence-based screening guidelines are needed for women under 40 with a family history of breast cancer, a BRCA1 or BRCA2 mutation, or other risk factors. An accurate assessment of breast cancer risk is required to balance the benefits and risks of surveillance, yet published studies have used narrow risk assessment schemata for enrollment. Breast density limits the sensitivity of film-screen mammography but is not thought to pose a limitation to MRI, however the utility of MRI surveillance has not been specifically examined before in women with dense breasts. Also, all MRI surveillance studies yet reported have used high strength magnets that may not be practical for dedicated imaging in many breast centers. Medium strength 0.5 Tesla MRI may provide an alternative economic option for surveillance. We conducted a prospective, nonrandomized pilot study of 30 women age 25-49 years with dense breasts evaluating the addition of 0.5 Tesla MRI to conventional screening. All participants had a high quantitative breast cancer risk, defined as > or = 3.5% over the next 5 years per the Gail or BRCAPRO models, and/or a known BRCA1 or BRCA2 germline mutation. The average age at enrollment was 41.4 years and the average 5-year risk was 4.8%. Twenty-two subjects had BIRADS category 1 or 2 breast MRIs (negative or probably benign), whereas no category 4 or 5 MRIs (possibly or probably malignant) were observed. Eight subjects had BIRADS 3 results, identifying lesions that were "probably benign", yet prompting further evaluation. One of these subjects was diagnosed with a stage T1aN0M0 invasive ductal carcinoma, and later determined to be a BRCA1 mutation carrier. Using medium-strength MRI we were able to detect 1 early breast tumor that was mammographically undetectable among 30 young high-risk women with dense breasts. These results support the concept that breast MRI can enhance surveillance for young high-risk women with dense breasts, and further suggest that a medium-strength instrument is sufficient for this application. For the first time, we demonstrate the use of quantitative breast cancer risk assessment via a combination of the Gail and BRCAPRO models for enrollment in a screening trial.


Artificial sweeteners and metabolic dysregulation: Lessons learned from agriculture and the laboratory.
Escalating rates of obesity and public health messages to reduce excessive sugar intake have fuelled the consumption of artificial sweeteners in a wide range of products from breakfast cereals to snack foods and beverages. Artificial sweeteners impart a sweet taste without the associated energy and have been widely recommended by medical professionals since they are considered safe. However, associations observed in long-term prospective studies raise the concern that regular consumption of artificial sweeteners might actually contribute to development of metabolic derangements that lead to obesity, type 2 diabetes and cardiovascular disease. Obtaining mechanistic data on artificial sweetener use in humans in relation to metabolic dysfunction is difficult due to the long time frames over which dietary factors might exert their effects on health and the large number of confounding variables that need to be considered. Thus, mechanistic data from animal models can be highly useful because they permit greater experimental control. Results from animal studies in both the agricultural sector and the laboratory indicate that artificial sweeteners may not only promote food intake and weight gain but can also induce metabolic alterations in a wide range of animal species. As a result, simple substitution of artificial sweeteners for sugars in humans may not produce the intended consequences. Instead consumption of artificial sweeteners might contribute to increases in risks for obesity or its attendant negative health outcomes. As a result, it is critical that the impacts of artificial sweeteners on health and disease continue to be more thoroughly evaluated in humans.


Factors predicting vitamin D response variation in non-Hispanic white postmenopausal women.
It is well documented that there is wide variation in the response of serum 25-hydroxyvitamin D [25(OH)D] to a given dose of vitamin D supplementation. Understanding factors affecting the response variation is important for identifying subjects who are susceptible to vitamin D deficiency or toxicity. This study aimed to evaluate potential predictors for vitamin D response variation. A total of 1179 non-Hispanic white postmenopausal women were enrolled into a 4-yr calcium and vitamin D (1100 IU/d) clinical trial. Among them, serum 25(OH)D level of 1063 subjects were measured at both baseline and after 12 months treatment. Vitamin D response was computed for these 1063 subjects as the difference in levels of serum 25(OH)D concentration at the end of a 12-month vitamin D treatment compared with baseline. Stepwise linear regression was used to identify predictors of vitamin D response variation. Increase in vitamin D intake, baseline serum 25(OH)D level, baseline blood collection season, baseline serum calcium level, and baseline body mass index were predictors of vitamin D response variation. These five factors explained 46.8% of the vitamin D response variation in the 1063 subjects. The first three factors [increase in vitamin D intake, baseline serum 25(OH)D level, baseline blood collection season] remained as predictors in the 392 subjects with trial vitamin D supplementation. For the first time, our study indicated that season is an important prediction factor for vitamin D response variation. Subjects who started vitamin D treatment in a cold season (autumn and winter) achieved a significantly higher serum 25(OH)D increase than those started in a hot season (summer) (P < 0.001). Our study suggests that the increase in vitamin D supplementation, baseline serum 25(OH)D level, and the season when initiating the vitamin D supplementation can partially predict vitamin D response variation in non-Hispanic postmenopausal women.


Determinants of preterm birth among mothers who gave birth in East Africa: systematic review and meta-analysis.
Preterm birth (PTB) can be caused by different factors. The factors can be classified into different categories: socio demographic, obstetric, reproductive health, medical, behavioral and nutritional related. The objective of this review was identifying determinants of PTB among mothers who gave birth in East African countries. We have searched the following electronic bibliographic databases: PubMed, Google scholar, Cochrane library, AJOL (African journal online). Cross sectional, case control and cohort study published in English were included. There was no restriction on publication period. Studies with no abstracts and or full texts, editorials, and qualitative in design were excluded. Funnel plot was used to check publication bias. I-squared statistic was used to check heterogeneity. Pooled analysis was done by using fixed and random effect model. The Joanna Briggs Critical Appraisal Tools for review and meta-analysis was used to check the study quality. A total of 58 studies with 134,801 participants were used to identify determinants of PTB. On pooled analysis, PTB was associated with age < 20 years (AOR 1.76, 95% CI: 1.33-2.32), birth interval less than 24 months (AOR 2.03, 95% CI 1.57-2.62), multiple pregnancy (AOR 3.44,95% CI: 3.02-3.91), < 4 antenatal care (ANC) visits (AOR 5.52, 95% CI: 4.32-7.05), and absence of ANC (AOR 5.77, 95% CI: 4.27-7.79). Other determinants of PTB included: Antepartum hemorrhage (APH) (AOR 4.90, 95% CI: 3.48-6.89), pregnancy induced hypertension (PIH) (AOR 3.10, 95% CI: 2.34-4.09), premature rupture of membrane (PROM) (AOR 5.90, 95% CI: 4.39-7.93), history of PTB (AOR 3.45, 95% CI: 2.72-4.38), and history of still birth/abortion (AOR 3.93, 95% CI: 2.70-5.70). Furthermore, Anemia (AOR 4.58, 95% CI: 2.63-7.96), HIV infection (AOR 2.59, 95% CI: 1.84-3.66), urinary tract infection (UTI) (AOR 5.27, 95% CI: 2.98-9.31), presence of vaginal discharge (AOR 5.33, 95% CI: 3.19-8.92), and malaria (AOR 3.08, 95% CI: 2.32-4.10) were significantly associated with PTB. There are many determinants of PTB in East Africa. This review could provide policy makers, clinicians, and program officers to design intervention on preventing occurrence of PTB.


TCRαβ/CD3 disruption enables CD3-specific antileukemic T cell immunotherapy.
T cells engineered to express chimeric antigen receptors (CARs) against B cell antigens are being investigated as cellular immunotherapies. Similar approaches designed to target T cell malignancies have been hampered by the critical issue of T-on-T cytotoxicity, whereby fratricide or self-destruction of healthy T cells prohibits cell product manufacture. To date, there have been no reports of T cells engineered to target the definitive T cell marker, CD3 (3CAR). Recent improvements in gene editing now provide access to efficient disruption of such molecules on T cells, and this has provided a route to generation of 3CAR, CD3-specific CAR T cells. T cells were transduced with a lentiviral vector incorporating an anti-CD3ε CAR derived from OKT3, either before or after TALEN-mediated disruption of the endogenous TCRαβ/CD3 complex. Only transduction after disrupting assembly of TCRαβ/CD3 yielded viable 3CAR T cells, and these cultures were found to undergo self-enrichment for 3CAR+TCR-CD3- T cells without any further processing. Specific cytotoxicity against CD3ε was demonstrated against primary T cells and against childhood T cell acute lymphoblastic leukemia (T-ALL). 3CAR T cells mediated potent antileukemic effects in a human/murine chimeric model, supporting the application of cellular immunotherapy strategies against T cell malignancies. 3CAR provides a bridging strategy to achieve T cell eradication and leukemic remission ahead of conditioned allogeneic stem cell transplantation.


New Wnt/β-catenin target genes promote experimental metastasis and migration of colorectal cancer cells through different signals.
We have previously identified a 115-gene signature that characterises the metastatic potential of human primary colon cancers. The signature included the canonical Wnt target gene BAMBI, which promoted experimental metastasis in mice. Here, we identified three new direct Wnt target genes from the signature, and studied their functions in epithelial-mesenchymal transition (EMT), cell migration and experimental metastasis. We examined experimental liver metastases following injection of selected tumour cells into spleens of NOD/SCID mice. Molecular and cellular techniques were used to identify direct transcription target genes of Wnt/β-catenin signals. Microarray analyses and experiments that interfered with cell migration through inhibitors were performed to characterise downstream signalling systems. Three new genes from the colorectal cancer (CRC) metastasis signature, BOP1, CKS2 and NFIL3, were identified as direct transcription targets of β-catenin/TCF4. Overexpression and knocking down of these genes in CRC cells promoted and inhibited, respectively, experimental metastasis in mice, EMT and cell motility in culture. Cell migration was repressed by interfering with distinct signalling systems through inhibitors of PI3K, JNK, p38 mitogen-activated protein kinase and/or mTOR. Gene expression profiling identified a series of migration-promoting genes, which were induced by BOP1, CKS2 and NFIL3, and could be repressed by inhibitors that are specific to these pathways. We identified new direct Wnt/β-catenin target genes, BOP1, CKS2 and NFIL3, which induced EMT, cell migration and experimental metastasis of CRC cells. These genes crosstalk with different downstream signalling systems, and activate migration-promoting genes. These pathways and downstream genes may serve as therapeutic targets in the treatment of CRC metastasis.


Opportunities to improve cardiovascular health in the new American workplace.
Adult working-class Americans spend on average 50% of their workday awake time at their jobs. The vast majority of these jobs involve mostly physically inactive tasks and frequent exposure to unhealthy food options. Traditionally, the workplace has been a challenging environment for cardiovascular prevention, where cardiovascular guidelines have had limited implementation. Despite the impact that unhealthy lifestyles at the workplace may have on the cardiovascular health of U.S. workers, there is currently no policy in place aimed at improving this. In this review, we discuss recent evidence on the prevalence of physical inactivity among Americans, with a special focus on the time spent at the workplace; and the invaluable opportunity that workplace-based lifestyle interventions may represent for improving the prevention of cardiovascular disease. We describe the current regulatory context, the key stakeholders involved, and present specific, guideline-inspired initiatives to be considered by both Congress and employers to improve the "cardiovascular safety" of US jobs. Additionally, we discuss how the COVID-19 pandemic has forever altered the workplace, and what lessons can be taken from this experience and applied to cardiovascular disease prevention in the new American workplace. For many Americans, long sitting hours at their job represent a risk to their cardiovascular health. We discuss how a paradigm shift in how we approach cardiovascular health, from focusing on leisure time to also focusing on work time, may help curtail the epidemic of cardiovascular disease in this country.


Contact between good friends: what limiting dilution analysis taught us.
The demonstration in the 1960s that T cells collaborated with B cells in enabling antibody responses to simple protein antigens opened up the challenge of how to investigate the interactions of two rare antigen-specific lymphocytes within a vast population. One idea was that T cells made soluble factors that could activate B cells at a distance; the other was that rare T cells and B cells came into contact. Using limiting dilution analysis, we asked the question 'How many B cells could a single T cell collaborate with in the short term?' Unequivocally, the answer was just one. This implied a need for cell contact, and coupled with the observation for genetic restriction in T-cell/B-cell co-operation, seemed to make a watertight case for direct coupling. Claims of diffusible antigen-specific factors undermined the importance of our findings at that time. Remarkably, those claims have not yet been retracted and our original findings that collaborating T cells and B cells must come into contact have never achieved the recognition that they deserved.


A critical overview of the current myofascial pain literature - March 2017.
After two years of having contributed to this overview series of articles, we sadly say goodbye to Dr. Rob Grieve. We would like to thank Dr. Grieve for his insightful contributions and analyses of the myofascial pain literature. Dr. Grieve would have preferred to continue, but his many university and research responsibilities had to take priority. We are looking forward to reviewing his future research endeavors in this article. We are pleased that Dr. Li-Wei Chou, MD, PhD has agreed to replace Dr. Grieve and join our team. Dr. Chou is Assistant Professor at China Medical University in Taichung, Taiwan and he has an impressive publication record with many research studies and book chapters. In this edition of the overview article, we once again have included articles from around the world with a combination of basic research and clinical studies and case reports. The majority of papers deal with dry needling, but there are also several more basic research studies and manual therapy papers.


Use of solid-state nanopores for sensing co-translocational deformation of nano-liposomes.
Membrane deformation of nano-vesicles is crucial in many cellular processes such as virus entry into the host cell, membrane fusion, and endo- and exocytosis; however, studying the deformation of sub-100 nm soft vesicles is very challenging using the conventional techniques. In this paper, we report detecting co-translocational deformation of individual 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC) nano-liposomes using solid-state nanopores. Electrokinetic translocation through the nanopore caused the soft DOPC liposomes (85 nm diameter) to change their shape, which we attribute to the strong electric field strength and physical confinement inside the pore. The experiments were performed at varying transmembrane voltages and the deformation was observed to mount up with increasing applied voltage and followed an exponential trend. Numerical simulations were performed to simulate the concentrated electric field strength inside the nanopore and a field strength of 14 kV cm(-1) (at 600 mV applied voltage) was achieved at the pore center. The electric field strength inside the nanopore is much higher than the field strength known to cause deformation of 15-30 μm giant membrane vesicles. As a control, we also performed experiments with rigid polystyrene beads that did not show any deformation during translocation events, which further established our hypothesis of co-translocational deformation of liposomes. Our technique presents an innovative and high throughput means for investigating deformation behavior of soft nano-vesicles.


A Bacterial Expression Platform for Production of Therapeutic Proteins Containing Human-like O-Linked Glycans.
We have developed an Escherichia coli strain for the in vivo production of O-glycosylated proteins. This was achieved using a dual plasmid approach: one encoding a therapeutic protein target, and a second encoding the enzymatic machinery required for O-glycosylation. The latter plasmid encodes human polypeptide N-acetylgalactosaminyl transferase as well as a β1,3-galactosyl transferase and UDP-Glc(NAc)-4-epimerase, both from Campylobacter jejuni, and a disulfide bond isomerase of bacterial or human origin. The effectiveness of this two-plasmid synthetic operon system has been tested on three proteins with therapeutic potential: the native and an engineered version of the naturally O-glycosylated human interferon α-2b, as well as human growth hormone with one engineered site of glycosylation. Having established proof of principle for the addition of the core-1 glycan onto proteins, we are now developing this system as a platform for producing and modifying human protein therapeutics with more complex O-glycan structures in E. coli.


Mathematical modelling of energy expenditure during tissue deposition.
Predicting the magnitude and rate of weight gain for a given increase of energy intake requires a model of whole-body energy expenditure that includes the energy cost of tissue deposition. Here, I introduce a mathematical framework for modelling energy expenditure that elucidates conceptual problems with the classical Kielanowski method for estimating the efficiencies of body fat and protein deposition. An alternative approach uses the theoretical biochemical efficiencies for protein and fat synthesis in combination with models of energy expenditure that include body fat and protein turnover costs. I illustrate this alternative approach using a simple mathematical model applied to previously published data from growing rats and human infants and compare the simple model results with the classical Kielanowski model. While both models fit the data reasonably well (R2>0.87 in rats and R2>0.67 in infants), the Kielanowski method resulted in parameter estimates that varied widely across experiments, had poor precision, and occasionally produced efficiency estimates greater than 1. In contrast, the new method provided precise parameter values and revealed consistencies across different experiments. The proposed mathematical framework has implications for interpreting studies of animal nutrition as well as providing a roadmap for future modelling efforts.


Muscle Mass and Gait Characteristics in Older Women Fallers vs. Non-Fallers.
Falling is a major public health concern of elderly people. We aimed to determine if lean mass and spatiotemporal gait parameters could predict the risk of falling in elderly women and also study the relationships between lean mass and gait characteristics. Twenty-four community women were prospectively recruited (mean age, 72.30 ± 5.31 years). Lean mass was measured using dual-energy fan-beam X-ray absorptiometry. Gait characteristics were assessed using spatiotemporal analysis. Fall risks were assessed using the Berg Balance Scale (BBS) and the Falls Efficacy Scale-International. Fall histories were recorded. Appropriate statistical analyses were applied to determine lean mass and gait characteristics in predicting the risk of fall and the associations between lean mass and gait characteristics. There were 14 participants (58.33%) with fall histories. Patients with fall histories had a significantly narrower base of support and lower BBS score. However, only the base of support was significantly associated with fall risk (odds ratio, 0.415; p = 0.022). Lean mass was significantly negatively associated with proportion of swing phase and positively associated with proportions of stance and double-support phases. Fall risk among elderly women can be predicted using base of support, where a narrower base predicts a greater fall risk. Although the lean mass was not related to risk of fall, lean mass is still related to some gait characteristics.


The construction of the eukaryotic expression plasmid pcDNA3.1/azurin and the increased apoptosis of U2OS cells transfected with it.
In our previous study, we demonstrated that azurin could selectively trigger apoptosis in human osteosarcoma cell line U2OS cells. However, the rate of apoptosis (35.8 +/- 3.2%) is not very high, and azurin is too expensive to obtain readily. To solve these problems, we constructed a eukaryotic expression plasmid containing the azurin gene with an influenza virus haemagglutinin 9 peptide HA epitope tag, and transfected the recombinant plasmid pcDNA3.1(+)/azurin into U2OS cells. RT-PCR and Western blot analysis validated the successful transfection and the expression of the azurin-HA protein. Conspicuous apoptosis of the transfected cells was detected by flow cytometry (FCM) and the DNA ladder test. The apoptosis rate reached 64.3 +/- 13.1%. The transcriptional levels of the Bax and p53 genes increased significantly in U2OS cells transfected with pcDNA3.1(+)/azurin, but the Bcl-2 mRNA level decreased. There was no difference in the levels of Bcl-xl mRNA and Survivin mRNA. We propose that the transfection of the recombinant plasmid pcDNA3.1(+)/azurin can significantly induce apoptosis in U2OS cells. This is closely associated with the up-regulation of the transcriptional level of the Bax and p53 genes, and the down-regulation of that of the Bcl-2 gene.


Worse prognosis of osteosarcoma patients expressing IGF-1 on a tissue microarray.
It is hardly possible to define osteosarcoma (OS) patients at greatest risk for non-response to chemotherapy, metastasis and short survival times. Our goal was the investigation of local expression of insulin-like growth factor (IGF-1) with regard to survival time of OS patients using a tissue microarray (TMA). Tumor tissue specimens from surgical primary tumor resections were collected from patients with OS. A TMA was composed, sections were stained with rabbit anti-IGF-1 and grading was performed. Statistics involved Kaplan-Meier curves and the log-rank test. We analyzed immunohistochemical expression of local IGF-1 on a TMA based on surgical primary tumor resections of 67 OS patients. The mean clinical follow-up time was 98 months. Twenty-two (33%) OS patients stained negatively and 44 (66%) OS patients stained positively for IGF-1. Significantly shorter survival was detected with expression of IGF-1 (p=0.007). The 5-year survival rate for patients expressing IGF-1 was 63% compared to 92% in patients without expression of IGF-1. Non-responders to chemotherapy and patients with metastasis, who also stained positively for IGF-1 manifested a significantly (p=0.002 and p<0.0001, respectively) shorter survival. Expression of local IGF-1 in primary tumor tissue appears to significantly affect the aggressiveness of OS, may predict survival time and, above all, may discriminate patients with non-response to chemotherapy and metastasis. This represents the basis for successful patient selection with regard to the decision process for or against chemotherapy and the choice of the most effective therapeutic drug. It may be a more important marker of tumor progression and indicator of prognosis than serum IGF-1. Novel tumor markers and therapeutic agents targeting the local IGF-1 pathway may increase the likelihood of therapeutic success.


Fallopian tube tumorigenesis and clinical implications for ovarian cancer risk-reduction.
Ovarian cancer remains the leading cause of gynecologic cancer death among American women. Prevention is the only proven approach to reduce the incidence of the disease. Oral contraception, tubal ligation, and risk-reducing salpingo-oophorectomy (rrBSO) for high-risk groups are all established risk reduction strategies. This paradigm is changing as recent biologic studies suggest that many ovarian cancers, especially high-grade serous ovarian cancers, originate in the distal end of the fallopian tube rather than the ovarian surface epithelium. A putative precursor lesion has been identified called the serous tubal intraepithelial carcinoma (STIC). Theoretically, removal of the fallopian tubes alone may prevent these lesions and prevent overt disease. Opportunistic salpingectomy during benign gynecologic surgery appears to be safe and may offer some protection from ovarian cancer without compromising ovarian endocrine function. Despite a lack of evidence for efficacy, several professional societies now recommend this approach for average-risk women. Whether salpingectomy can also serve as a temporizing measure to delay risk-reducing oophorectomy in women with a genetic predisposition to ovarian cancer remains to be seen. Several ongoing non-randomized clinical trials will test the feasibility of this approach. Therefore, the societal impact of increasing salpingectomy rates on ovarian cancer incidence will be an area of intense focus for the next 10-20 years.


The treatment of allergies using Sanfujiu: A method of applying Chinese herbal medicine paste to acupoints on three peak summer days.
Sanfujiu is a method of applying herbal drug paste onto acupoints Fengmen and Feishu during the 3 hottest summer days, to treat patients with allergies using traditional Chinese medicine. The objectives of this study were to describe the immediate reactions (adverse effects and perceived efficacy) to the Sanfujiu treatment, and examine the variations in the prevalence of the reactions to and perceived efficacy of Sanfujiu among different sub-groups, based on patient age and diagnosis. The study subjects included 119 patients who completed Sanfujiu treatment at a regional hospital in Taipei. One week after treatment, trained interviewers conducted telephone interviews with the patients. More than 80% of the patients reported having reactive symptoms after using the Sanfujiu treatment. Younger subjects (< or = 16 years of age) were more likely to have reactive symptoms. Patients with rhinitis were more likely to have runny noses and nasal congestion after the treatment. Patients with allergic eczema were more likely to have skin itching all over the body. Overall, 44.6% of patients perceived the treatment as being effective, while 52.1% of patients did not notice any effects, either good or bad, from the treatment. The perceived efficacy of Sanfujiu treatment was not related to patient age, sex, or diagnosis. This study demonstrated that Sanfujiu treatment was moderately effective, as perceived by the patients, in treating their allergic symptoms immediately after the treatment. Symptoms reactive to the treatment were common but are usually mild. This information is essential for patient consultation, and serves as a reference for clinicians.


The thylakoid FtsH protease plays a role in the light-induced turnover of the photosystem II D1 protein.
The photosystem II reaction center D1 protein is known to turn over frequently. This protein is prone to irreversible damage caused by reactive oxygen species that are formed in the light; the damaged, nonfunctional D1 protein is degraded and replaced by a new copy. However, the proteases responsible for D1 protein degradation remain unknown. In this study, we investigate the possible role of the FtsH protease, an ATP-dependent zinc metalloprotease, during this process. The primary light-induced cleavage product of the D1 protein, a 23-kD fragment, was found to be degraded in isolated thylakoids in the dark during a process dependent on ATP hydrolysis and divalent metal ions, suggesting the involvement of FtsH. Purified FtsH degraded the 23-kD D1 fragment present in isolated photosystem II core complexes, as well as that in thylakoid membranes depleted of endogenous FtsH. In this study, we definitively identify the chloroplast protease acting on the D1 protein during its light-induced turnover. Unlike previously identified membrane-bound substrates for FtsH in bacteria and mitochondria, the 23-kD D1 fragment represents a novel class of FtsH substrate-functionally assembled proteins that have undergone irreversible photooxidative damage and cleavage.


Inflammatory bowel disease-attributable costs and cost-effective strategies in the United States: a review.
The United States spends more for healthcare than any other country in the world. With the rising prevalence of both Crohn's disease and ulcerative colitis, inflammatory bowel disease (IBD) represents the leading chronic gastrointestinal disease with increasing healthcare expenditures in the US. IBD costs have shifted from inpatient to outpatient care since the introduction of biologic therapies as the standard of care. Gastroenterologists need to be aware of the national cost burden of IBD and clinical practices that optimize cost-efficiency. This investigation offers a systematic review of the economics of IBD and evidence-based strategies for cost-effective management.


Effect of plasma [K+] on the DC potential and on ion distributions between CSF and blood.
Keeping the arterial pH at 7.4 and PaCO2 at 40 mmHg in eight anesthetized dogs, we acutely raised plasma potassium concentration from 3.4 to 8.2 meq/1, then allowed it to decay back to control levels. The cerebrospinal fluid (CSF)-blood electrical potential difference (pd) increased 13.2 mV per 10-fold increase in plasma [K+]. Again keeping arterial pH at 7.4 and PaCO2 at 40 mmHg, we elevated plasma [K+] in four dogs from 3.3 to 8.0 meq/1 and maintained this level for 6 h. We found 1) that the PD increased from a control value of +1.3 to +8.9mV, showing no tendency to decay over the 6 h; and 2) that the change in PD did not affect the distribution of Na+, K+, H+, Cl-, or HCO3- between blood and CSF over the 6 h. These results suggest that under these conditions the PD between CSF and blood may play no effective role in determining the distributions of these charged species by 6 h. These results are contrasted with recent findings which suggest that H+ and HCO3- are distributed according to passive forces between CSF and blood.


Seawater desalination using pillared graphene as a novel nano-membrane in reverse osmosis process: nonequilibrium MD simulation study.
Herein, the applicability and efficiency of two types of pillared graphene nanostructures, namely, (6,6)@G and (7,7)@G, were investigated as membranes in reverse osmosis seawater desalination using extensive nonequilibrium molecular dynamics simulations. The water permeability for (6,6)@G and (7,7)@G membranes was estimated at 4.2 and 6.6 L h-1 cm-2 MPa-1, respectively. According to the results, a complete (100%) and pressure-independent salt rejection was estimated for both membranes. In addition, the mechanism of seawater desalination through the pillared graphene membranes was investigated via the density distribution profile of water molecules inside the pillar channels. Furthermore, a series of steered MD simulations were performed to construct the potential of mean force (PMF) profile of water molecules and salt ions passing through the membranes channels. The passing free energy barriers of Na+ and Cl- ions and water molecules are 0.86, 0.62, and 0.22 eV, respectively, for the (6,6)@G membrane. The corresponding quantities for the (7,7)@G membrane are 0.71, 0.44, and 0.11 eV, respectively.


Alternative versus conventional treatment strategy in uncomplicated acute otitis media in children: a prospective, open, controlled parallel-group comparison.
Evidence from clinical trials questions the benefit-risk ratio of first-line antibiotic treatment for uncomplicated acute otitis media in childhood. Alternative treatment strategies are very popular but have not been the subject of larger controlled clinical trials. This trial compares an alternative with a conventional treatment strategy for acute otitis media. 390 children aged 1-10 years presenting with uncomplicated acute otitis media participated in a prospective, open, non-randomized, controlled, parallel-group study. According to self-assignment of investigators, children were treated either conventionally (free combinations of decongestant nose drops, mucolytics, analgesics and antibiotics) or alternatively with Otovowen (fixed combination of plant-based tinctures and homeopathic potencies), supplemented by conventional medications when considered necessary. Alternatively treated patients (n = 192) had significantly less severe otoscopic findings and clinical symptom ratings at baseline than children treated in conventional centers (n = 193). Patients cared for by conventional therapists took more antibiotics (80.5% vs. 14.4%; chi2-test, p < 0.001) and analgesics (66.8% vs. 53.2%; chi2-test, p = 0.007). Times to recovery were 5.3 +/- 2.4 and 5.1 +/- 2.2 days for alternative and conventional treatment, respectively. Odds ratios (OR) with a lower limit of 1-sided 97.5% confidence interval (CI) were 0.98 (0.76), 0.95 (0.73) and 0.88 (0.69) for results adjusted to baseline otoscopy, pain and symptom score, respectively (Cox-Mantel test). Absence from school or preschool nursery was 1.7 days in both groups; ORs (CI) were 1.00 (0.76), 0.96 (0.73) and 1.04 (0.80). Noninferiority of alternative treatment (CI limit of OR above 0.696) was not proven for pain resolution (-5.2 vs. -5.8 score points); OR (CI) were 0.87 (0.68), 1.15 (0.87) and 0.74 (0.58). Alternative treatment was judged both by doctors (Mann-Whitney estimator with 2-sided 95% CI 0.41 (0.35-0.47)) and parents (0.42 (0.36-0.48)), to be significantly better tolerated than conventional treatment. In primary care management of uncomplicated acute otitis media in childhood, an alternative treatment strategy based on the natural medicine, Otovowen may substantially reduce the use of antibiotics without disadvantage to the clinical outcome.


Physical Exercise Keeps the Brain Connected: Biking Increases White Matter Integrity in Patients With Schizophrenia and Healthy Controls.
It has been shown that learning a new skill leads to structural changes in the brain. However, it is unclear whether it is the acquisition or continuous practicing of the skill that causes this effect and whether brain connectivity of patients with schizophrenia can benefit from such practice. We examined the effect of 6 months exercise on a stationary bicycle on the brain in patients with schizophrenia and healthy controls. Biking is an endemic skill in the Netherlands and thus offers an ideal situation to disentangle the effects of learning vs practice. The 33 participating patients with schizophrenia and 48 healthy individuals were assigned to either one of two conditions, ie, physical exercise or life-as-usual, balanced for diagnosis. Diffusion tensor imaging brain scans were made prior to and after intervention. We demonstrate that irrespective of diagnosis regular physical exercise of an overlearned skill, such as bicycling, significantly increases the integrity, especially of motor functioning related, white matter fiber tracts whereas life-as-usual leads to a decrease in fiber integrity. Our findings imply that exercise of an overlearned physical skill improves brain connectivity in patients and healthy individuals. This has important implications for understanding the effect of fitness programs on the brain in both healthy subjects and patients with schizophrenia. Moreover, the outcome may even apply to the nonphysical realm.


Increased blood pressure and hyperdynamic cardiovascular responses in carriers of a common hyperfunctional variant of adenylyl cyclase 6.
Adenylyl cyclase (ADCY) is a critical regulator of metabolic and cardiovascular function. We have identified a genetic variant (A674S) in ADCY isoform 6 (ADCY6). Subsequent studies demonstrated that the expression of this ADCY6 variant paralleled an increase in adenylyl cyclase-mediated functions. However, the impact of this hyperfunctional variant on cardiovascular function is unknown. Therefore, we evaluated the hemodynamic profile of carriers of ADCY6 A674S. The association of ADCY6 A674S with anthropometric and hemodynamic parameters was assessed in 364 healthy white subjects. The allele encoding this variant was present in 6.9% of the subjects, and those individuals had increased blood pressure. To determine the hemodynamic basis for increased blood pressure in carriers of ADCY6 A674S, we assessed forearm blood flow (FBF) and cardiac output at rest, during handgrip exercise (to test vasodilator responsiveness), and with lower body negative pressure [to test forearm vasoconstrictor and heart rate (HR) responsiveness] in a subsample of 21 subjects. At rest, cardiac output and blood pressure were higher in carriers of ADCY6 A674S. This was paralleled by an increase in plasma renin activity, but not in plasma norepinephrine. During handgrip exercise, FBF and vasodilator responses were greater in carriers of ADCY6 A674S. Responses to reactive hyperemia were not different between genotypes. With lower body negative pressure, the HR response to this orthostatic stress was markedly higher in carriers of ADCY6 A674S. These data indicate that the relatively common hyperfunctional ADCY6 A674S variant underlies a hyperdynamic cardiovascular response and increased blood pressure.


Postoperative urinary extravasation does not impact anterior urethroplasty surgical outcomes: a Latin American large cohort study.
To determine the prevalence of postoperative urinary extravasation (POUE) following anterior urethroplasty, to analyze factors associated with its occurrence, and to study the impact of POUE on surgical success. Retrospective cohort study including all male patients who have undergone a urethroplasty at our center between 2011 and 2018. Subjects with posterior location stricture, those who did not undergo routine radiographic follow-up, or patients with inadequate follow-up were excluded. Urinary extravasation was defined as presence of evident contrast extravasation on the postoperative voiding cystourethrogram (VCUG). Impact was determined as "need-for-reoperation". Uni- and multivariate analysis were performed to determine clinical and demographic variables associated with occurrence of extravasation and postoperative stricture. A total of 783 men underwent a urethroplasty and 630 fulfilled inclusion criteria. Urinary extravasation prevalence was 12.2%, and there was a "need-for-reoperation" in 1.1% of cases. On uni- and multivariate analysis, greatest stricture length (HR: 1.07 (1-1.2), p = 0.05) and penile urethral location (HR: 2.29 (1.1-4.6), p = 0.021) showed to be POUE predictors. POUE did not show to be a risk factor for postoperative stricture (HR: 1.57, 95% CI (0.8-3), p = 0.173). However, reoperation group showed to be a risk factor (HR: 6.6, 95% CI 1.4-31, p = 0.019). Prevalence of POUE was 12.2%. Stricture length and penile urethral strictures were POUE predictors. POUE occurrence with successful conservative management did not appear to have impact on urethroplasty outcomes as it did not predict re-stricture. POUE was reoperation cause in 1.1% of total cases.


Challenging identity: development of a measure of veterinary career motivations.
While little is known about the motivations underpinning veterinary work, previous literature has suggested that the main influences on veterinary career choice are early/formative exposure to animals or veterinary role models. The aim of this study was to develop and provisionally validate a veterinary career motivations questionnaire to assess the strength of various types of career motivations in graduating and experienced veterinarians. A cross-sectional sample of experienced veterinarians (n=305) and a smaller cohort of newly graduated veterinarians (n=53) were surveyed online using a long-form questionnaire. Exploratory factor analysis (EFA) was used to iteratively derive a final, short-form questionnaire for survey of a second cross-sectional sample of experienced veterinarians (n=751). EFA derived a final questionnaire with 22 items loading onto six factors (social purpose, animal orientation, vocational identity, challenge and learning, career affordances, and people orientation). While motivations based on animal orientation were predictably strong, those based on vocational identity were not universal and were weaker in younger and graduate veterinarians; both of these motivations were rated lower by male veterinarians. Motivations based on challenge and learning emerged as some of the strongest, most universal and most influential; people orientation and social purpose were also important, particularly for older veterinarians. The major motivations for pursuing a veterinary career may best be represented as an intrinsic passion for animal care and for learning through solving varied challenges. These motivations are largely intrinsically oriented and autonomously regulated, thus likely to be supportive of work satisfaction and wellbeing.


Basal ryanodine receptor activity suppresses autophagic flux.
The inositol 1,4,5-trisphosphate receptors (IP3Rs) and intracellular Ca2+ signaling are critically involved in regulating different steps of autophagy, a lysosomal degradation pathway. The ryanodine receptors (RyR), intracellular Ca2+-release channels mainly expressed in excitable cell types including muscle and neurons, have however not yet been extensively studied in relation to autophagy. Yet, aberrant expression and excessive activity of RyRs in these tissues has been implicated in the onset of several diseases including Alzheimer's disease, where impaired autophagy regulation contributes to the pathology. In this study, we determined whether pharmacological RyR inhibition could modulate autophagic flux in ectopic RyR-expressing models, like HEK293 cells and in cell types that endogenously express RyRs, like C2C12 myoblasts and primary hippocampal neurons. Importantly, RyR3 overexpression in HEK293 cells impaired the autophagic flux. Conversely, in all cell models tested, pharmacological inhibition of endogenous or ectopically expressed RyRs, using dantrolene or ryanodine, augmented autophagic flux by increasing lysosomal turn-over (number of autophagosomes and autolysosomes measured as mCherry-LC3 punctae/cell increased from 70.37±7.81 in control HEK RyR3 cells to 111.18±7.72 and 98.14±7.31 after dantrolene and ryanodine treatments, respectively). Moreover, in differentiated C2C12 cells, transmission electron microscopy demonstrated that dantrolene treatment decreased the number of early autophagic vacuoles from 5.9±2.97 to 1.8±1.03 per cellular cross section. The modulation of the autophagic flux could be linked to the functional inhibition of RyR channels as both RyR inhibitors efficiently diminished the number of cells showing spontaneous RyR3 activity in the HEK293 cell model (from 41.14%±2.12 in control cells to 18.70%±2.25 and 9.74%±2.67 after dantrolene and ryanodine treatments, respectively). In conclusion, basal RyR-mediated Ca2+-release events suppress autophagic flux at the level of the lysosomes.


Long-term clinical and immunological effects of probiotic and peanut oral immunotherapy after treatment cessation: 4-year follow-up of a randomised, double-blind, placebo-controlled trial.
Oral immunotherapy has attracted much interest as a potential treatment for food allergy, yet little is known about its long-term effects. We aimed to assess long-term outcomes in participants who completed a randomised, double-blind, placebo-controlled trial of combined probiotic and peanut oral immunotherapy (PPOIT), which was previously shown to induce desensitisation and 2-week sustained unresponsiveness. All participants who completed the PPOIT randomised trial were eligible to participate in this follow-up study 4 years after treatment cessation. Peanut intake and adverse reactions to peanut in the 4 years after treatment cessation were systematically documented with a structured questionnaire administered by allergy nurses. Additionally, participants were invited to undergo peanut skin prick tests, measurement of peanut sIgE and sIgG4 concentrations, and double-blind placebo-controlled peanut challenge to assess 8-week sustained unresponsiveness. 48 (86%) of 56 eligible participants were enrolled in the follow-up study. Mean time since stopping treatment was 4·2 years in both PPOIT (SD 0·6) and placebo (SD 0·7) participants. Participants from the PPOIT group were significantly more likely than those from the placebo group to have continued eating peanut (16 [67%] of 24 vs one [4%] of 24; absolute difference 63% [95% CI 42-83], p=0·001; number needed to treat 1·6 [95% CI 1·2-2·4]). Four PPOIT-treated participants and six placebo participants reported allergic reactions to peanut after intentional or accidental intake since stopping treatment, but none had anaphylaxis. PPOIT-treated participants had smaller wheals in peanut skin prick test (mean 8·1 mm [SD 7·7] vs 13·3 mm [7·6]; absolute difference -5·2 mm [95% CI -10·3 to 0·0]; age-adjusted and sex-adjusted p=0·035) and significantly higher peanut sIgG4:sIgE ratios than placebo participants (geometric mean 67·3 [95% CI 10·3-440·0] vs 5·2 [1·2-21·8]; p=0·031). Seven (58%) of 12 participants from the PPOIT group attained 8-week sustained unresponsiveness, compared with one (7%) of 15 participants from the placebo group (absolute difference 52% [95% CI 21-82), p=0·012; number needed to treat 1·9 [95% CI 1·2-4·8]). PPOIT provides long-lasting clinical benefit and persistent suppression of the allergic immune response to peanut. Murdoch Childrens Research Institute and Australian Food Allergy Foundation.


Predatory fish select for coordinated collective motion in virtual prey.
Movement in animal groups is highly varied and ranges from seemingly disordered motion in swarms to coordinated aligned motion in flocks and schools. These social interactions are often thought to reduce risk from predators, despite a lack of direct evidence. We investigated risk-related selection for collective motion by allowing real predators (bluegill sunfish) to hunt mobile virtual prey. By fusing simulated and real animal behavior, we isolated predator effects while controlling for confounding factors. Prey with a tendency to be attracted toward, and to align direction of travel with, near neighbors tended to form mobile coordinated groups and were rarely attacked. These results demonstrate that collective motion could evolve as a response to predation, without prey being able to detect and respond to predators.


Intensification of cattle ranching production systems: socioeconomic and environmental synergies and risks in Brazil.
Intensification of Brazilian cattle ranching systems has attracted both national and international attention due to its direct relation with Amazon deforestation on the one hand and increasing demand of the global population for meat on the other. Since Brazilian cattle ranching is predominantly pasture-based, we particularly focus on pasture management. We summarize the most recurrent opportunities and risks associated with pasture intensification that are brought up within scientific and political dialogues, and discuss them within the Brazilian context. We argue that sustainable intensification of pasturelands in Brazil is a viable way to increase agricultural output while simultaneously sparing land for nature. Since environmental degradation is often associated with low-yield extensive systems in Brazil, it is possible to obtain higher yields, while reversing degradation, by adopting practices like rotational grazing, incorporation of legumes and integrated crop-livestock-forestry systems. Technical assistance is however essential, particularly for small- and medium-scale farmers. Sound complementary policies and good governance must accompany these measures so that a 'rebound effect' does not lead to increased deforestation and other adverse social and environmental impacts. It is also important that animal welfare is not compromised. Although the discussion is presented with respect to Brazil, some aspects are relevant to other developing countries.


Integrated strategy for the production of therapeutic retroviral vectors.
The broad application of retroviral vectors for gene delivery is still hampered by the difficulty to reproducibly establish high vector producer cell lines generating sufficient amounts of highly concentrated virus vector preparations of high quality. To enhance the process for producing clinically relevant retroviral vector preparations for therapeutic applications, we have integrated novel and state-of-the-art technologies in a process that allows rapid access to high-efficiency vector-producing cells and consistent production, purification, and storage of retroviral vectors. The process has been designed for various types of retroviral vectors for clinical application and to support a high-throughput process. New modular helper cell lines that permit rapid insertion of DNA encoding the therapeutic vector of interest at predetermined, optimal chromosomal loci were developed to facilitate stable and high vector production levels. Packaging cell lines, cultivation methods, and improved medium composition were coupled with vector purification and storage process strategies that yield maximal vector infectivity and stability. To facilitate GMP-grade vector production, standard of operation protocols were established. These processes were validated by production of retroviral vector lots that drive the expression of type VII collagen (Col7) for the treatment of a skin genetic disease, dystrophic epidermolysis bullosa. The potential efficacy of the Col7-expressing vectors was finally proven with newly developed systems, in particular in target primary keratinocyte cultures and three-dimensional skin tissues in organ culture.


Colloidal particle lens arrays-assisted nano-patterning by harmonics of a femtosecond laser.
We consider nanopatterning of dielectric substrates by harmonics of single powerful femtosecond pulses from a Ti:Sapphire laser. The nanopatterning is mediated by closely packed monolayers of polystyrene microspheres that act as microlenses at the surface. Observed modification of the material proceeds via ionization. By our theory, the second harmonic is more effective in multi-photon ionization and is better focused than the fundamental frequency which is effective in multiplying of the amount of free electrons via impact ionization. Experiments show that conversion of a part of the pulse energy into the second harmonic decreases the modification threshold and improves the localization of the structures. Optimization of the time offset between the harmonics could further improve the efficiency and quality of nanostructuring.


Application of a Q Method Study to Understanding Nurses' Perspective of Adopting Evidence-Based Nursing.
This study applied the Q method to identify and describe the various types of nurse perceptions that are crucially associated with their engagement in evidence-based nursing (EBN). The study participants were nurses at a medical center and a regional teaching hospital. A series of Q sorts was performed by nurses to subjectively rank the Q statements. Q statements were constructed based on the literature related to EBN adoption by nurses and face-to-face interviews. A total of 60 participants were invited to rank 44 Q statements related to EBN. Factor analysis was conducted on the rankings of the Q statements. The following are the five prominent shared perspectives: (1) emphasized the obstacles to evidence searching and reading ability; (2) emphasized the organizational promotive strategies; (3) emphasized the available supportive resources; (4) emphasized the significance of EBN; and (5) emphasized the evidence-searching ability and external incentives. The five identified groups of perspectives can enhance hospital administrators to acknowledge the barriers and incentives associated with EBN practices. The exploration of clustering nurses' perceptions may facilitate the development of customized strategies to enable more appropriate training.