Mitochondrial Membrane Potential Identifies Cells with Enhanced Stemness for Cellular Therapy.
Long-term survival and antitumor immunity of adoptively transferred CD8(+) T cells is dependent on their metabolic fitness, but approaches to isolate therapeutic T cells based on metabolic features are not well established. Here we utilized a lipophilic cationic dye tetramethylrhodamine methyl ester (TMRM) to identify and isolate metabolically robust T cells based on their mitochondrial membrane potential (ΔΨm). Comprehensive metabolomic and gene expression profiling demonstrated global features of improved metabolic fitness in low-ΔΨm-sorted CD8(+) T cells. Transfer of these low-ΔΨm T cells was associated with superior long-term in vivo persistence and an enhanced capacity to eradicate established tumors compared with high-ΔΨm cells. Use of ΔΨm-based sorting to enrich for cells with superior metabolic features was observed in CD8(+), CD4(+) T cell subsets, and long-term hematopoietic stem cells. This metabolism-based approach to cell selection may be broadly applicable to therapies involving the transfer of HSC or lymphocytes for the treatment of viral-associated illnesses and cancer.


Impact of life history traits on gene flow: A multispecies systematic review across oceanographic barriers in the Mediterranean Sea.
Marine species can demonstrate strong genetic differentiation and population structure despite the hypothesis of open seas and high connectivity. Some suggested drivers causing the genetic breaks are oceanographic barriers and the species' biology. We assessed the relevance of seven major oceanographic fronts on species connectivity while considering their dispersal capacity and life strategy. We systematically reviewed the scientific articles reporting population genetic differentiation along the Mediterranean Sea and across the Atlantic-Mediterranean transition. We retained those considering at least one sampling locality at each side of an oceanographic front, and at least two localities with no-front between them to correctly assess the effect of the front. To estimate the impact of life history characteristics affecting connectivity we considered the planktonic larval duration (PLD) and adult life strategy. Oceanographic barriers in the Mediterranean Sea seem to reduce gene flow globally; however, this effect is not homogeneous considering the life history traits of the species. The effect of the oceanographic fronts reduces gene flow in highly mobile species with PLD larger than 2-4 weeks. Benthic sessile species and/or with short PLD (< 2 weeks) have more significant genetic breaks between localities than species with higher motility; however, genetic differentiation occurs independently of the presence of a front. Genetic connectivity is important for populations to recover from anthropogenic or natural impacts. We show that species with low mobility, mostly habitat-formers, have high genetic differentiation but low gene flow reduction mediated by the front, therefore, considering the importance of these species, we emphasize the vulnerability of the Mediterranean ecosystems and the necessity of protection strategies based on the whole ecosystem.


Calcification of primary human osteoblast cultures under flow conditions using polycaprolactone scaffolds for intravascular applications.
Total atherosclerotic occlusion is a leading cause of death. Recent animal models of this disease are devoid of cell-mediated calcification and arteries are often not occluded gradually. This study is part of a project with the objective of developing a new model featuring the above two characteristics, using a tissue-engineering scaffold. The amount and distribution of calcium deposits in primary human osteoblast (HOB) cultures on polycaprolactone (PCL) scaffolds under flow conditions were investigated. HOBs were cultured on PCL scaffolds with TGF-β1 loadings of 0 (control), 5 and 50 ng. HOB-PCL constructs were cultured in spinner flasks. Under flow conditions, cell numbers present in HOB cultures on PCL scaffolds increased from day 7 to day 14, and most calcification was induced at day 21. TGF-β1 loadings of 5 and 50 ng did not show a significant difference in ALP activity, cell numbers and amount of calcium deposited in HOB cultures, but calcium staining showed that 50 ng TGF-β1 had higher calcium deposited on both days 21 and 28 under flow conditions compared with 5 ng of loading. Amount of calcium deposited by HOBs on day 28 showed a decrease from their levels on day 21. PCL degradation may be a factor contributing to this loss. The results indicate that cell-induced calcification can be achieved on PCL scaffolds under flow conditions. In conclusion, TGFβ1-HOB loaded PCL can be applied to create a model for total atherosclerotic occlusion with cell-deposited calcium in animal arteries.


Membrane fusion.
Common themes are emerging from the study of viral, cell-cell, intracellular, and liposome fusion. Viral and cellular membrane fusion events are mediated by fusion proteins or fusion machines. Viral fusion proteins share important characteristics, notably a fusion peptide within a transmembrane-anchored polypeptide chain. At least one protein involved in a cell-cell fusion reaction resembles viral fusion proteins. Components of intracellular fusion machines are utilized in multiple membrane trafficking events and are conserved through evolution. Fusion pores develop during and intracellular fusion events suggesting similar mechanisms for many, if not all, fusion events.


EAU guidelines on renal cell carcinoma: 2014 update.
The European Association of Urology Guideline Panel for Renal Cell Carcinoma (RCC) has prepared evidence-based guidelines and recommendations for RCC management. To provide an update of the 2010 RCC guideline based on a standardised methodology that is robust, transparent, reproducible, and reliable. For the 2014 update, the panel prioritised the following topics: percutaneous biopsy of renal masses, treatment of localised RCC (including surgical and nonsurgical management), lymph node dissection, management of venous thrombus, systemic therapy, and local treatment of metastases, for which evidence synthesis was undertaken based on systematic reviews adhering to Preferred Reporting Items for Systematic Reviews and Meta-analyses (PRISMA) guidelines. Relevant databases (Medline, Cochrane Library, trial registries, conference proceedings) were searched (January 2000 to November 2013) including randomised controlled trials (RCTs) and retrospective or controlled studies with a comparator arm. Risk of bias (RoB) assessment and qualitative and quantitative synthesis of the evidence were performed. The remaining sections of the document were updated following a structured literature assessment. All chapters of the RCC guideline were updated. For the various systematic reviews, the search identified a total of 10,862 articles. A total of 151 studies reporting on 78,792 patients were eligible for inclusion; where applicable, data from RCTs were included and meta-analyses were performed. For RCTs, there was low RoB across studies; however, clinical and methodological heterogeneity prevented data pooling for most studies. The majority of studies included were retrospective with matched or unmatched cohorts based on single or multi-institutional data or national registries. The exception was for systemic treatment of metastatic RCC, in which several RCTs have been performed, resulting in recommendations based on higher levels of evidence. The 2014 guideline has been updated by a multidisciplinary panel using the highest methodological standards, and provides the best and most reliable contemporary evidence base for RCC management. The European Association of Urology Guideline Panel for Renal Cell Carcinoma has thoroughly evaluated available research data on kidney cancer to establish international standards for the care of kidney cancer patients.


Plasmodium malariae and P. ovale genomes provide insights into malaria parasite evolution.
Elucidation of the evolutionary history and interrelatedness of Plasmodium species that infect humans has been hampered by a lack of genetic information for three human-infective species: P. malariae and two P. ovale species (P. o. curtisi and P. o. wallikeri). These species are prevalent across most regions in which malaria is endemic and are often undetectable by light microscopy, rendering their study in human populations difficult. The exact evolutionary relationship of these species to the other human-infective species has been contested. Using a new reference genome for P. malariae and a manually curated draft P. o. curtisi genome, we are now able to accurately place these species within the Plasmodium phylogeny. Sequencing of a P. malariae relative that infects chimpanzees reveals similar signatures of selection in the P. malariae lineage to another Plasmodium lineage shown to be capable of colonization of both human and chimpanzee hosts. Molecular dating suggests that these host adaptations occurred over similar evolutionary timescales. In addition to the core genome that is conserved between species, differences in gene content can be linked to their specific biology. The genome suggests that P. malariae expresses a family of heterodimeric proteins on its surface that have structural similarities to a protein crucial for invasion of red blood cells. The data presented here provide insight into the evolution of the Plasmodium genus as a whole.


Surface science and catalysis.
During the last 20 years, surface scientists have developed a variety of techniques that make it possible to study, on the molecular level, the structure, composition, and chemical bonding in the surface monolayer. Both single-crystal model catalysts and real catalyst systems that are active in important chemical processes have been investigated by a combination of ultrahigh-vacuum surface science techniques and high-pressure kinetic techniques in an effort to determine the relation between the structure and composition of the surface and the rates of reaction and selectivities. The structure of the atomic surface, a strongly adsorbed overlayer, and the oxidation states of surface atoms are the important molecular features of an active catalyst. As a consequence of modern surface science, the design and preparation of catalysts has developed from an art into a high technology.


Genome-scale rates of evolutionary change in bacteria.
Estimating the rates at which bacterial genomes evolve is critical to understanding major evolutionary and ecological processes such as disease emergence, long-term host-pathogen associations and short-term transmission patterns. The surge in bacterial genomic data sets provides a new opportunity to estimate these rates and reveal the factors that shape bacterial evolutionary dynamics. For many organisms estimates of evolutionary rate display an inverse association with the time-scale over which the data are sampled. However, this relationship remains unexplored in bacteria due to the difficulty in estimating genome-wide evolutionary rates, which are impacted by the extent of temporal structure in the data and the prevalence of recombination. We collected 36 whole genome sequence data sets from 16 species of bacterial pathogens to systematically estimate and compare their evolutionary rates and assess the extent of temporal structure in the absence of recombination. The majority (28/36) of data sets possessed sufficient clock-like structure to robustly estimate evolutionary rates. However, in some species reliable estimates were not possible even with 'ancient DNA' data sampled over many centuries, suggesting that they evolve very slowly or that they display extensive rate variation among lineages. The robustly estimated evolutionary rates spanned several orders of magnitude, from approximately 10-5 to 10-8 nucleotide substitutions per site year-1. This variation was negatively associated with sampling time, with this relationship best described by an exponential decay curve. To avoid potential estimation biases, such time-dependency should be considered when inferring evolutionary time-scales in bacteria.


Antibody-unfolding and metastable-state binding in force spectroscopy and recognition imaging.
Force spectroscopy and recognition imaging are important techniques for characterizing and mapping molecular interactions. In both cases, an antibody is pulled away from its target in times that are much less than the normal residence time of the antibody on its target. The distribution of pulling lengths in force spectroscopy shows the development of additional peaks at high loading rates, indicating that part of the antibody frequently unfolds. This propensity to unfold is reversible, indicating that exposure to high loading rates induces a structural transition to a metastable state. Weakened interactions of the antibody in this metastable state could account for reduced specificity in recognition imaging where the loading rates are always high. The much weaker interaction between the partially unfolded antibody and target, while still specific (as shown by control experiments), results in unbinding on millisecond timescales, giving rise to rapid switching noise in the recognition images. At the lower loading rates used in force spectroscopy, we still find discrepancies between the binding kinetics determined by force spectroscopy and those determined by surface plasmon resonance-possibly a consequence of the short tethers used in recognition imaging. Recognition imaging is nonetheless a powerful tool for interpreting complex atomic force microscopy images, so long as specificity is calibrated in situ, and not inferred from equilibrium binding kinetics.


COVID-19: Preliminary Data on the Impact of Social Distancing on Loneliness and Mental Health.
The COVID-19 pandemic and its need for social distancing as a response have the potential to increase the experience of loneliness in the population, with an associated increase in symptoms of mental disorders. As the world has largely adapted to remote platforms for employment, socializing, and health service delivery, the degree to which virtual opportunities for social engagement may offset the impact of limited in-person interactions on mental health functioning is unknown. This column offers preliminary data from an ongoing experience sampling study of the prevalence, course, and impact of loneliness on mental health in a community adult sample living under social distancing requirements during the COVID-19 pandemic. Initial findings aggregated across all experience sampling reports showed negative associations between virtual social contact (via text, phone, or videoconferencing) and feelings of loneliness, while in-person interactions appeared to have no impact on loneliness. In addition, respondents reported frequent instances of negative effects on physical and mental health, including disruptions in sleep and recurring suicidal ideation. While further analyses are needed, these findings are consistent with similar emerging reports showing increased rates of mental health concerns during periods of social distancing. Possible avenues for addressing these concerns using remote interventions are explored.


Gastric alkalinization after major trauma.
Gastric function in trauma patients is poorly understood. In animals, shock causes gastric luminal alkalinization and bile reflux. In trauma patients, studies of stress gastritis prophylaxis demonstrated with continuous gastric pH monitoring that the stomach became alkaline even without antisecretory therapy. Therefore, we hypothesized that trauma patients have an alkaline gastric environment that may be because of bile reflux. A prospective observational study at an urban Level I trauma center was performed. All major torso trauma patients (severe head injuries excluded) who met the criteria for standardized shock resuscitation were eligible for inclusion. A 12.5 Fr silastic pH probe (Sandhill Scientific) was placed in the stomach and the gastric pH continuously monitored for 7 days. Patients received no stress gastritis prophylaxis. Gastric samples were obtained each day and assayed for total bile acids and pH. Twelve patients were entered into the study. Mean age was 31 years +/- 4 years, 67% men, 75% blunt mechanism of injury, and mean Injury Severity Score 28 +/- 3. Three patients (25%) developed multiple organ failure and four acquired ventilator-associated pneumonia. During the first day of continuous pH monitoring, 9 of 12 patients had a gastric pH >4 for the majority of the day with 7 patients having essentially no acid production. During subsequent days, gastric pH began to drop and by the 4th day the majority of each day was spent at a pH <4. Additionally, gastric pH of patients with ventilator-associated pneumonia or multiple organ failure tended to be more alkaline. Bile acid was present in the gastric fluid of all patients in varying amounts. However, there was no significant correlation between gastric pH and bile acid concentration. Traumatic injury causes gastric luminal alkalinization that may be related, only in part, to bile acid reflux. Other alkalinizing factors remain to be elucidated.


Structure-Based Optimization of Nonquaternary Reactivators of Acetylcholinesterase Inhibited by Organophosphorus Nerve Agents.
Acetylcholinesterase (AChE), a key enzyme in the central and peripheral nervous systems, is the principal target of organophosphorus nerve agents. Quaternary oximes can regenerate AChE activity by displacing the phosphyl group of the nerve agent from the active site, but they are poorly distributed in the central nervous system. A promising reactivator based on tetrahydroacridine linked to a nonquaternary oxime is also an undesired submicromolar reversible inhibitor of AChE. X-ray structures and molecular docking indicate that structural modification of the tetrahydroacridine might decrease inhibition without affecting reactivation. The chlorinated derivative was synthesized and, in line with the prediction, displayed a 10-fold decrease in inhibition but no significant decrease in reactivation efficiency. X-ray structures with the derivative rationalize this outcome. We thus show that rational design based on structural studies permits the refinement of new-generation pyridine aldoxime reactivators that may be more effective in the treatment of nerve agent intoxication.


Meniscal tear in knees without surgery and the development of radiographic osteoarthritis among middle-aged and elderly persons: The Multicenter Osteoarthritis Study.
Although partial meniscectomy is a risk factor for the development of knee osteoarthritis (OA), there is a lack of evidence that meniscal damage that is not treated with surgery would also lead to OA, suggesting that surgery itself may cause joint damage. Furthermore, meniscal damage is common. The aim of this study was to evaluate the association between meniscal damage in knees without surgery and the development of radiographic tibiofemoral OA. We conducted a prospective case-control study nested within the observational Multicenter Osteoarthritis Study, which included a sample of men and women ages 50-79 years at high risk of knee OA who were recruited from the community. Patients who had no baseline radiographic knee OA but in whom tibiofemoral OA developed during the 30-month followup period were cases (n = 121). Control subjects (n = 294) were drawn randomly from the same source population as cases but had no knee OA after 30 months of followup. Individuals whose knees had previously undergone surgery were excluded. Meniscal damage was defined as the presence of any medial or lateral meniscal tearing, maceration, or destruction. Meniscal damage at baseline was more common in case knees than in control knees (54% versus 18%; P < 0.001). The model comparing any meniscal damage with no meniscal damage (adjusted for baseline age, sex, body mass index, physical activity, and mechanical knee alignment) yielded an odds ratio of 5.7 (95% confidence interval 3.4-9.4). In knees without surgery, meniscal damage is a potent risk factor for the development of radiographic OA. These results highlight the need for better understanding, prevention, and treatment of meniscal damage.


Immunologic timeline of Ebola virus disease and recovery in humans.
A complete understanding of human immune responses to Ebola virus infection is limited by the availability of specimens and the requirement for biosafety level 4 (BSL-4) containment. In an effort to bridge this gap, we evaluated cryopreserved PBMCs from 4 patients who survived Ebola virus disease (EVD) using an established mass cytometry antibody panel to characterize various cell populations during both the acute and convalescent phases. Acute loss of nonclassical monocytes and myeloid DCs, especially CD1c+ DCs, was noted. Classical monocyte proliferation and CD38 upregulation on plasmacytoid DCs coincided with declining viral load. Unsupervised analysis of cell abundance demonstrated acute declines in monocytic, NK, and T cell populations, but some populations, many of myeloid origin, increased in abundance during the acute phase, suggesting emergency hematopoiesis. Despite cell losses during the acute phase, upregulation of Ki-67 correlated with recovery of cell populations over time. These data provide insights into the human immune response during EVD.


Nonlinear quantum optomechanics via individual intrinsic two-level defects.
We propose to use the intrinsic two-level system (TLS) defect states found naturally in integrated optomechanical devices for exploring cavity QED-like phenomena with localized phonons. The Jaynes-Cummings-type interaction between TLS and mechanics can reach the strong coupling regime for existing nano-optomechanical systems, observable via clear signatures in the optomechanical output spectrum. These signatures persist even at finite temperature, and we derive an explicit expression for the temperature at which they vanish. Further, the ability to drive the defect with a microwave field allows for realization of phonon blockade, and the available controls are sufficient to deterministically prepare non-classical states of the mechanical resonator.


Facilitators and Barriers to Implementing Healthy School Canteen Intervention among Malaysian Adolescents: A Qualitative Study.
This study evaluated implementing a school-based intervention to promote healthier dietary habits in the school environment among Malaysian adolescents using qualitative methods. This qualitative study was conducted in four secondary schools in Perak and Selangor (two urban and two rural schools) that received the intervention (either training or training and food subsidy). A total of eight focus groups (68 students aged 15 years old) and 16 in-depth interviews were conducted with canteen operators, school convenience shop operators, school teachers and school principals in each school. Thematic analysis was used to analyse the qualitative data to identify suitable themes. We found several initiatives and changes by the schools' stakeholders to change to a healthy school canteen programme. The stakeholders also noticed the students' food preferences that influence healthy food intake in canteens and convenience shops. The food vendors and school administrators also found that subsidising healthy meals might encourage healthy eating. Among barriers to implementing healthy school initiatives were the student's perception of healthy food and their eating habits, which also affect the food vendors' profit if they want to implement a healthy canteen. The school-based intervention has the potential to promotes healthier eating among school adolescents. Continuous training and monitoring of canteen operators and convenience shops are needed, including building partnerships and educating the students on healthy eating to cultivate healthy eating habits.


Brownian dynamics of confined rigid bodies.
We introduce numerical methods for simulating the diffusive motion of rigid bodies of arbitrary shape immersed in a viscous fluid. We parameterize the orientation of the bodies using normalized quaternions, which are numerically robust, space efficient, and easy to accumulate. We construct a system of overdamped Langevin equations in the quaternion representation that accounts for hydrodynamic effects, preserves the unit-norm constraint on the quaternion, and is time reversible with respect to the Gibbs-Boltzmann distribution at equilibrium. We introduce two schemes for temporal integration of the overdamped Langevin equations of motion, one based on the Fixman midpoint method and the other based on a random finite difference approach, both of which ensure that the correct stochastic drift term is captured in a computationally efficient way. We study several examples of rigid colloidal particles diffusing near a no-slip boundary and demonstrate the importance of the choice of tracking point on the measured translational mean square displacement (MSD). We examine the average short-time as well as the long-time quasi-two-dimensional diffusion coefficient of a rigid particle sedimented near a bottom wall due to gravity. For several particle shapes, we find a choice of tracking point that makes the MSD essentially linear with time, allowing us to estimate the long-time diffusion coefficient efficiently using a Monte Carlo method. However, in general, such a special choice of tracking point does not exist, and numerical techniques for simulating long trajectories, such as the ones we introduce here, are necessary to study diffusion on long time scales.


Population genetic analysis of shotgun assemblies of genomic sequences from multiple individuals.
We introduce a simple, broadly applicable method for obtaining estimates of nucleotide diversity from genomic shotgun sequencing data. The method takes into account the special nature of these data: random sampling of genomic segments from one or more individuals and a relatively high error rate for individual reads. Applying this method to data from the Celera human genome sequencing and SNP discovery project, we obtain estimates of nucleotide diversity in windows spanning the human genome and show that the diversity to divergence ratio is reduced in regions of low recombination. Furthermore, we show that the elevated diversity in telomeric regions is mainly due to elevated mutation rates and not due to decreased levels of background selection. However, we find indications that telomeres as well as centromeres experience greater impact from natural selection than intrachromosomal regions. Finally, we identify a number of genomic regions with increased or reduced diversity compared with the local level of human-chimpanzee divergence and the local recombination rate.


Sequences within the coding regions of clotting factor VIII and CFTR block transcriptional elongation.
The clotting factor VIII (FVIII) and cystic fibrosis transmembrane conductance regulator (CFTR) cDNAs have dramatically reduced levels of expression compared to clotting factor IX (FIX) and other cDNAs (100 and 1,000-fold lower, respectively), when produced in cells by using an expression vector. Part of the inhibitory signal in the FVIII cDNA has been localized to a 1.2-kb inhibitory sequence (FVIII INS), which decreased steady-state RNA levels from a retroviral vector by 30- to 100-fold. An analysis of RNA degradation indicated that the FVIII INS vector RNA is relatively stable. Nuclear run-on experiments with the FVIII INS vector demonstrated a low signal for FVIII, in contrast to the high signal for a FIX vector. The low signal for FVIII INS was not due to a decrease in transcriptional initiation. Thus, FVIII expression is reduced through a block to transcriptional elongation, as has been found in c-myc and other genes. We show that the inhibitory effect of FVIII INS is orientation dependent with regard to the promoter. In addition, the inhibitory effect is position dependent, because expression of FVIII INS sequence increased when it was moved 1 kb further from the promoter in a retroviral vector. Similar results were observed by using a retroviral vector for expression of the CFTR cDNA. The CFTR retroviral vector produced 1,000-fold decreased steady-state RNA levels, compared to the parent vector. Nuclear run-on analysis with the CFTR vector revealed a block to transcriptional elongation within the CFTR cDNA. The presence of blocks to transcriptional elongation within the FVIII and CFTR cDNAs complicates efforts to produce high levels of these proteins for therapeutic purposes and to develop high-titer retroviral expression vectors for human gene therapy.


Depressive symptomatology in obese adults with type II diabetes.
This study compared depressive symptomatology in 32 obese subjects with type II (non-insulin-dependent) diabetes (16 men, 16 women) and their obese nondiabetic spouses. All subjects completed the Beck Depression Inventory (BDI) before participation in a behavioral weight-loss program. Diabetic subjects reported significantly more depressive symptomatology than their overweight nondiabetic spouses (10.6 +/- 6.4 vs. 7.5 +/- 6.2, P less than 0.04). Diabetic subjects scored higher than their spouses on 15 of 20 BDI items, with significant differences in feelings of being punished, perceived appearance, and interest in sex. Mean BDI score in the diabetic spouses was similar to that observed in the first 123 diabetic subjects to enter the weight-loss program (BDI 11.2 +/- 6.9). Further studies are needed to determine whether diabetic subjects differ from age-, sex-, and weight-matched nondiabetic individuals in clinical depression and depressive symptomatology.


Surgical treatment of single-suture craniosynostosis: an argument for quantitative methods to evaluate cosmetic outcomes.
The traditional reasons for surgical intervention in children with single-suture craniosynostosis (SSC) are cosmetic improvement and the avoidance/treatment of intracranial hypertension, which has been thought to contribute to neurocognitive deficits. Despite considerable work on the topic, the exact prevalence of intracranial hypertension in the population of patients with SSC is unknown, although it appears to be present in only a minority. Additionally, recent neuropsychological and anatomical literature suggests that the subtle neurocognitive deficits identified in children with a history of SSC may not result from external compression. They may instead reflect an underlying developmental condition that includes disordered primary CNS development and early suture fusion. This implies that current surgical techniques are unlikely to prevent neurocognitive deficits in patients with SSC. As such, the most common indication for surgical treatment in SSC is cosmetic, and most patients benefit from considerable subjective cosmetic normalization following surgery. Pediatric craniofacial surgeons have not, however, agreed upon objective means to assess postoperative cranial morphological improvement. We should therefore endeavor to agree upon objective craniometric tools for the assessment of operative outcomes, allowing us to accurately compare the various surgical techniques that are currently available.


Multiscale modelling and nonlinear simulation of vascular tumour growth.
In this article, we present a new multiscale mathematical model for solid tumour growth which couples an improved model of tumour invasion with a model of tumour-induced angiogenesis. We perform nonlinear simulations of the multi-scale model that demonstrate the importance of the coupling between the development and remodeling of the vascular network, the blood flow through the network and the tumour progression. Consistent with clinical observations, the hydrostatic stress generated by tumour cell proliferation shuts down large portions of the vascular network dramatically affecting the flow, the subsequent network remodeling, the delivery of nutrients to the tumour and the subsequent tumour progression. In addition, extracellular matrix degradation by tumour cells is seen to have a dramatic affect on both the development of the vascular network and the growth response of the tumour. In particular, the newly developing vessels tend to encapsulate, rather than penetrate, the tumour and are thus less effective in delivering nutrients.


Transmission properties of atypical Creutzfeldt-Jakob disease: a clue to disease etiology?
The genotype at polymorphic codon 129 of the PRNP gene has a profound influence on both phenotypic expression and prion strain susceptibility in humans. For example, while the most common sporadic Creutzfeldt-Jakob disease (CJD) subtype, sporadic CJD-MM1 (M1 strain), induces a single phenotype after experimental transmission regardless of the codon 129 genotype of the recipient animal, the phenotype elicited by sporadic CJD-VV2 (V2 strain), the second most common subtype, varies according to the host codon 129 genotype. In particular, the propagation of the V2 strain in codon 129 methionine homozygotes has been linked only to acquired forms of CJD such as plaque-type dura mater graft-associated CJD (dCJD), a subgroup of iatrogenic CJD with distinctive phenotypic features, but has never been observed in sporadic CJD cases. In the present report, we describe atypical CJD cases carrying codon 129 methionine homozygosity, in a neurosurgeon and in a patient with a medical history of neurosurgery without dural grafting, showing the distinctive phenotypic features and transmission properties of plaque-type dCJD. These findings raise the possibility that the two cases, previously thought to represent sporadic CJD, might actually represent acquired CJD caused by infection with the V2 strain. Thus, careful analyses of phenotypic features and transmission properties in atypical cases may be useful to distinguish acquired from sporadic cases of CJD. Susceptibility to and phenotypic expression of Creutzfeldt-Jakob disease (CJD) depend on both the prion strain and genotype at polymorphic codon 129 of the PRNP gene. For example, propagation of the second most common sporadic CJD strain (V2 strain) into codon 129 methionine homozygotes has been linked to plaque-type dura mater graft-associated CJD (dCJD), a subgroup of iatrogenic CJD with distinctive phenotypic features, but has never been observed in sporadic CJD. In the present report, we describe atypical CJD cases in a neurosurgeon and in a patient with a medical history of neurosurgery without dural grafting, showing the distinctive phenotypic features and transmission properties of plaque-type dCJD. These findings raise the possibility that the two cases, previously considered to represent sporadic CJD, might actually represent acquired CJD caused by infection with the V2 strain.


Anticipatory grief in patients with cancer.
Patients and their loved ones often experience anticipatory grief when learning of a diagnosis of advanced or terminal cancer. Anticipatory grief can be a response to threats of loss of ability to function independently, loss of identity, and changes in role definition, which underlie fear of death. Dealing with multiple losses is a primary task that the dying patient must face. When an oncologist delivers bad news, the patient and family members often hear the same discussion through different filters, which can lead to conflict and dysfunction. By providing a supportive and safe environment, oncology nurses can help patients and their loved ones understand that their feelings are common and are experienced by others in similar situations and assist them with developing coping strategies and in redefining their roles within the family and in the outside world. In addition, an important goal at this time is to help the patients reframe "hope" realistically so they may have the opportunity for personal growth as well as reconciliation of primary relationships toward the end of life.


Improvements in Essential Newborn Care and Newborn Resuscitation Services Following a Capacity Building and Quality Improvement Program in Three Districts of Uttar Pradesh, India.
Neonatal death remains a global challenge contributing to 45% of underfive deaths. With rising institutional delivery, to accelerate decline in neonatal mortality rate (NMR) improvement in the quality of perinatal care requires attention. This implementation research targeted improving service delivery readiness for quality of newborn care at public health facilities in three districts of Uttar Pradesh, India, with high NMR. This before-after study assessed the facility readiness and quality of newborn services at 42 health facilities. The changes in 26 signal functions for routine and emergency obstetric and newborn care were tracked. There was marked improvement in newborn service availability: skilled birth attendants (51%), resuscitation (30%), and kangaroo mother care (27%) at these facilities. A multifold rise in newborn resuscitation efforts and documentation (n = 4431 vs. n = 144 in preintervention period) with high success rate (98.6%) was observed. There was also improvement in obstetric care services including partograph use (31%) and active management of third stage of labor (46%). However, several infrastructural indicators (electricity, water supply, toilets, and sanitation) remained unchanged. Overall improvements were observed in the majority of the signal functions for perinatal care and newborn resuscitation efforts. There was a limited impact on the infrastructural and supervision components.


Carbohydrate metabolism and its response to catecholamines as modified in alkalotic rat.
Metabolic activities and their responses to catecholamines were studied in fasted rats exposed to simulated altitudes. Analysis of hepatic levels of gluconeogenic intermediates revealed the inhibition of gluconeogenesis between glyceric acid 3-P and fructose 6-P associated with a rise of the ratios of redox pairs such as lactate to pyruvate in livers of alkalotic rats. Inhibition of gluconeogenesis was indicated also by the suppressed incorporation of glutamate 14C into blood glucose. Since no activation was detected on glycolytic pathway of skeletal muscles, marked hyperlactacidemia during alkalosis appeared to result from the suppression of hepatic gluconeogenesis. Most of metabolic actions of epinephrine and isoproterenal known to be mediated via the beta receptor were significantly reduced but not completely abolished during alkalosis. Exceptionally, hyperinsulinemia induced by isoproterenol was completely reversed and replaced by hypoinsulinemia during alkalosis. Despite hypoinsulinemia, hyperglycemia induced by glucose load decreased more rapidly in alkalotic than in normal rats. In view of the fact that the adrenergic alpha receptor is involved in theinhibition of insulin secretion, the observed irregular modifications of catecholamine actions could be explained on the basis of a postulate that the adrenergic alpha-receptor functions are potentiated in alkalosis.


Estimating microtubule distributions from 2D immunofluorescence microscopy images reveals differences among human cultured cell lines.
Microtubules are filamentous structures that are involved in several important cellular processes, including cell division, cellular structure and mechanics, and intracellular transportation. Little is known about potential differences in microtubule distributions within and across cell lines. Here we describe a method to estimate information pertaining to 3D microtubule distributions from 2D fluorescence images. Our method allows for quantitative comparisons of microtubule distribution parameters (number of microtubules, mean length) between different cell lines. Among eleven cell lines compared, some showed differences that could be accounted for by differences in the total amount of tubulin per cell while others showed statistically significant differences in the balance between number and length of microtubules. We also observed that some cell lines that visually appear different in their microtubule distributions are quite similar when the model parameters are considered. The method is expected to be generally useful for comparing microtubule distributions between cell lines and for a given cell line after various perturbations. The results are also expected to enable analysis of the differences in gene expression underlying the observed differences in microtubule distributions among cell types.


Phylogenetic analyses to uncover the evolutionary relationship of a newly sequenced mitochondrial genome from an Eastern spinebill (Acanthorhynchus tenuirostris).
The Eastern spinebill (Acanthorhynchus tenuirostris), a passerine bird in the family Meliphagidae (honeyeaters), a dominant group of birds in Australia and New Guinea. The aim of this study was to sequence the complete mitochondrial genome of the Eastern spinebill and use its sequence to better define the phylogeny of this species. The complete mitogenome sequence of A. tenuirostris was circular and 16,614 bp in length, and its architecture was conserved in comparison to other mitogenome sequences under the family Meliphagidae. The Eastern spinebill mitogenome had the highest sequence identity with mitogenome sequences of two other honeyeaters, the white eared honeyeater, Nesoptilotis leucotis, (84.9%) and the white-plumed honeyeater, Ptilotula penicillata (85.5%). The maximum-likelihood topology distinctly discriminated the Eastern spinebill sequence against all other species of the Meliphagidae with significant bootstrap supports. We suggest the widespread sampling and complete mitogenome sequencing would be valuable in establishing the most accurate phylogenetic taxonomy of the family Meliphagidae.


Sun protection behaviors among African Americans.
The anatomic distribution of some skin cancers suggests that sun exposure may be an etiologic factor for skin cancer among African Americans. Yet little is known about sun protection behaviors among African Americans. We analyzed data from the 1992 National Health Interview Survey (N = 1,583) to determine the prevalence of sun protection behaviors and sun sensitivity. About 6% of African Americans reported being extremely sensitive to the sun and severe sunburning, and 9% reported mild burns. Overall, 53% of respondents (47% of men and 57% of women) reported that they were very likely to wear protective clothing, seek shade, or use sunscreen lotion. Women were more likely than men to report seeking shade and using sunscreen. Sun protection behaviors were more frequently reported by those who sunburn more easily and were positively associated with age. Use of sunscreen was positively associated with income and education. Education about sun protection and early detection may help reduce the morbidity and mortality of skin cancer among African Americans.


Response characteristics of the phase Doppler particle analyzer for sizing spherical particles larger than the light wavelength.
A theoretical model, based on the geometrical optics approach, has been developed to simulate various aspects of the phase Doppler particle analyzer (PDPA). The model has taken into consideration the nonuniform (Gaussian) illumination of the particles as they pass through the measurement probe volume. Instrument response curves have been generated for various scattering angles by performing spatial and temporal integration of the scattered intensity distribution over the receiver surface. Experimental and theoretical investigations have established the applicability of this instrument to both forward scattered and backscattered angles.


Second-line anti-tuberculosis drug concentrations for susceptibility testing in the MODS assay.
Multidrug-resistant tuberculosis (TB) threatens TB control worldwide. The microscopic observation drug susceptibility (MODS) assay is a low-cost, high-performance TB diagnostic tool for rapid liquid culture and direct isoniazid and rifampicin drug susceptibility testing (DST). The objective of this study was to explore the potential for extending the MODS assay to rapid second-line DST and to identify critical concentrations of candidate drugs for prospective testing. Sputum samples from 94 TB culture-positive patients receiving second-line TB agents were cultured following standardised MODS protocols, with a range of titrations of antimicrobial drugs added. Critical concentrations were determined using a modified Kaplan-Meier survival curve analysis. Candidate critical concentrations were determined for capreomycin (10 μg·mL(-1)), ciprofloxacin (1.25 μg·mL(-1)), cycloserine (40 μg·mL(-1)), ethambutol (10 μg·mL(-1)), ethionamide (5 μg·mL(-1)), kanamycin (5 μg·mL(-1)), para-aminosalicylic acid (10 μg·mL(-1)) and streptomycin (10 μg·mL(-1)). No cut-off point was identified for the other second-line drugs or for pyrazinamide. At particular concentrations of some second-line TB drugs this novel Kaplan-Meier analysis clearly differentiated populations that were susceptible or resistant. These candidate critical concentrations should now be tested in a range of epidemiological settings to define the performance of direct, second-line TB DST with MODS, offering potential low-cost second-line TB DST capacity.


Voltage amplification of thermopower waves via current crowding at high resistances in self-propagating combustion waves.
Combustion wave propagation in micro/nanostructured materials generates a chemical-thermal-electrical energy conversion, which enables the creation of an unusual source of electrical energy, called a thermopower wave. In this paper, we report that high electrical resistance regimes would significantly amplify the output voltage of thermopower waves, because the current crowding creates a narrow path for charge carrier transport. We show that the structurally defective regions in the hybrid composites of chemical fuels and carbon nanotube (CNT) arrays determine both the resistance levels of the hybrid composites and the corresponding output voltage of thermopower waves. A sudden acceleration of the crowded charges would be induced by the moving reaction front of the combustion wave when the supplied driving force overcomes the potential barrier to cause charge carrier transport over the defective region. This property is investigated experimentally for the locally manipulated defective areas using diverse methods. In this study, thermopower waves in CNT-based hybrid composites are able to control the peak voltages in the range of 10-1000 mV by manipulating the resistance from 10 Ω to 100 kΩ. This controllable voltage generation from thermopower waves may enable applications using the combustion waves in micro/nanostructured materials and better understanding of the underlying physics.


Functional and Esthetic Indication for Dental Implant Treatment and Immediate Loading (2) Case Report and Considerations: Typical Attitudes of Dentists (and their Unions) toward Tooth Extractions and the Prevention of Early, Effective, and Helpful Dental Implant Treatment in the European Union.
The technology of the Strategic Implant® facilitates dental implant treatment almost regardless of the available bone. This explains its increasing use to resolve problems with the dentition of the middle-aged patient in a radical manner, simply by extracting the residual dentition and placing implants. While a radical change in the implant treatment paradigm has taken place in some countries, in other countries, dentists try hard to keep their work area free of implants or to deliver them only as a last resort. Liberating patients from the burdens of their own teeth are not advocated by dental universities, as their main field of teaching deals with the repair of teeth and conventional ways of replacing them. This case report shows a standard treatment with Strategic Implants®, discussing the topic from the point of view of practitioners, universities, and other parties involved in decision-making. The authors conclude that the interests of the patients are not respected in many cases because the parties involved have vested interests in other treatment modalities.


Clinical and Immunologic Profiles in Incomplete Lupus Erythematosus and Improvement with Hydroxychloroquine Treatment.
Objective. The study goals were to evaluate performance of SLE classification criteria, to define patients with incomplete lupus erythematosus (ILE), and to probe for features in these patients that might be useful as indicators of disease status and hydroxychloroquine response. Methods. Patients with ILE (N = 70) and SLE (N = 32) defined by the 1997 American College of Rheumatology criteria were reclassified using the 2012 Systemic Lupus International Collaborating Clinics criteria. Disease activity, patient reported outcomes, and levels of Type I interferon- (IFN-) inducible genes, autoantibodies, and cytokines were measured. Subgroups treated with hydroxychloroquine (HCQ) were compared to patients not on this drug. Results. The classification sets were correlated (R2 = 0.87). ILE patients were older (P = 0.0043) with lower disease activity scores (P < 0.001) and greater dissatisfaction with health status (P = 0.034) than SLE patients. ILE was associated with lower levels of macrophage-derived cytokines and levels of expressed Type I IFN-inducible genes. Treatment of ILE with HCQ was associated with better self-reported health status scores and lower expression levels of Type I IFN-inducible genes than ILE patients not on HCQ. Conclusion. The 2012 SLICC SLE classification criteria will be useful to define ILE in trials. Patients with ILE have better health status and immune profiles when treated with HCQ.


Anxious women do not show the expected decrease in cardiovascular stress responsiveness as pregnancy advances.
Altered stress responsiveness is a risk factor for mental and physical illness. In non-pregnant populations, it is well-known that anxiety can alter the physiological regulation of stress reactivity. Characterization of corresponding risks for pregnant women and their offspring requires greater understanding of how stress reactivity and recovery are influenced by pregnancy and women's anxiety feelings. In the current study, women were presented repeatedly with mental arithmetic stress tasks in the first and third pregnancy trimester and reported their trait anxiety using the state trait anxiety inventory. Cardiovascular stress reactivity in late pregnancy was lower than reactivity in the first pregnancy trimester (heart rate (HR): t(197)=4.98, p<.001; high frequency heart rate variability (HF HRV): t(196)=-2.09, p=.04). Less attenuation of stress reactivity occurred in more anxious women (HR: b=0.15, SE=0.06, p=.008; HF HRV: b=-10.97, SE=4.79, p=.02). The study design did not allow the influence of habituation to repeated stress task exposure to be assessed separately from the influence of pregnancy progression. Although this is a limitation, the clear differences between anxious and non-anxious pregnant women are important, regardless of the extent to which differing habituation between the groups is responsible. Less dampened stress reactivity through pregnancy may pose long-term risks for anxious women and their offspring. Follow-up studies are required to determine these risks.


Is ethnic density associated with health in a context of social disadvantage? Findings from the Born in Bradford cohort.
In this study we aimed to test the associations between area-level ethnic density and health for Pakistani and White British residents of Bradford, England. The sample consisted of 8610 mothers and infant taking part in the Born in Bradford cohort. Ethnic density was measured as the percentage of Pakistani, White British or South Asian residents living in a Lower Super Output Area. Health outcomes included birth weight, preterm birth and smoking during pregnancy. Associations between ethnic density and health were tested in multilevel regression models, adjusted for individual covariates and area deprivation. In the Pakistani sample, higher own ethnic density was associated with lower birth weight (β = -0.82, 95% CI: -1.63, -0.02), and higher South Asian density was associated with a lower probability of smoking during pregnancy (OR = 0.99, 95% CI: 0.98, 1.00). Pakistani women in areas with 50-70% South Asian residents were less likely to smoke than those living in areas with less than 10% South Asian residents (OR = 0.39, 95% CI: 0.16, 0.97). In the White British sample, neither birth weight nor preterm birth was associated with own ethnic density. The probability of smoking during pregnancy was lower in areas with 10-29.99% compared to <10% South Asian density (OR = 0.79, 95% CI: 0.64, 0.98). In this sample, ethnic density was associated with lower odds of smoking during pregnancy but not with higher birth weight or lower odds of preterm birth. Possibly, high levels of social disadvantage inhibit positive effects of ethnic density on health.


Effect of tetrahydrobiopterin on endothelial function in canine middle cerebral arteries.
Tetrahydrobiopterin is an essential cofactor required for activation of NO synthase. However, in intact arteries, the exact role of tetrahydrobiopterin in the regulation of NO synthase activity is not fully understood. The present study was designed to determine the effect of increasing intracellular tetrahydrobiopterin levels on endothelial function in isolated canine middle cerebral arteries. The arterial segments were incubated in MEM for 24 hours at 37 degrees C in the presence or absence of a tetrahydrobiopterin precursor, sepiapterin (10(-4) mol/L), and/or superoxide dismutase (150 U/mL). The rings were suspended for isometric tension recording. Tetrahydrobiopterin levels were assayed by high-performance liquid chromatography. Production of cGMP was measured by radioimmunoassay. Incubation with sepiapterin markedly increased intracellular tetrahydrobiopterin levels. In sepiapterin-treated arteries, endothelium-dependent relaxations to calcium ionophore A23187 and intracellular cGMP levels were significantly reduced. Superoxide dismutase alone did not affect either relaxation to A23187 or production of cGMP. However, when arteries were incubated with superoxide dismutase plus sepiapterin, endothelium-dependent relaxations to A23187, as well as cGMP production, were significantly augmented. The augmentation of cGMP was observed in rings with (but not without) endothelium. Incubation of arteries in calcium-free medium almost abolished the synergistic effect of tetrahydrobiopterin and superoxide dismutase on cGMP production. These results demonstrate that increased availability of tetrahydrobiopterin may activate endothelial NO synthase. This effect appears to be critically dependent on the presence of superoxide dismutase.


Characterization of piezocrystals for practical configurations with temperature- and pressure-dependent electrical impedance spectroscopy.
Piezoelectric single crystal materials such as (x)Pb(Mg(1/3)Nb(2/3))O(3-)(1-x)PbTiO(3) (PMN-PT) have, by some measures, significantly better performance than established piezoelectric ceramics for ultrasound applications. However, they are also subject to phase transitions affecting their behavior at temperatures and pressures encountered in underwater sonar and actuator applications and in non-destructive testing at elevated temperatures. Materials with modified compositions to reduce these problems are now under development, but application-oriented characterization techniques need further attention. Characterization with temperature variation has been reported extensively, but the range of parameters measured is often limited and the effects of pressure variation have received almost no attention. Furthermore, variation in properties between samples is now rarely reported. The focus of this paper is an experimental system set up with commercially available equipment and software to carry out characterization of piezoelectric single crystals with variation in temperature, pressure, and electrical bias fields found in typical practical use. We illustrate its use with data from bulk thickness-mode PMN-29%PT samples, demonstrating variation among nominally identical samples and showing not only the commonly reported changes in permittivity with temperature for bulk material but also significant and complicated changes with pressure and bias field and additional ultrasonic modes which are attributed to material phase changes. The insight this provides may allow the transducer engineer to accelerate new material adoption in devices.


Pressure-stabilized polymerization of nitrogen in alkaline-earth-metal strontium nitrides.
High-pressure technology can help us to obtain excellent materials. We have explored alkaline-earth-metal strontium nitrides under different pressures, theoretically. A variety of stable Sr-N structures were predicted by the structure searching method using CALYPSO code. Six new stoichiometries, SrN, Sr2N3, SrN2, SrN3, SrN4, and SrN5, were predicted. And our calculation proved that all these compounds were stable existing under ambient pressure up to 100 GPa. A rich variety of poly-nitrogen forms appeared in the newly predicted SrNx compounds, including four nitrogen polymerization forms: ranging from N2, N3, N4, and N5 molecules, to zig-zag nitrogen chains and extended chains connected by puckered "N6" rings. Significantly, the 1D extended polymeric chain of puckered "N6" rings was firstly identified in the P1[combining macron]-SrN3 structure at 60 GPa. Another N-rich C2/c-SrN4 was stable only under the relatively high-pressure of 20 GPa, but this phase can be quenched under atmospheric pressure. The N-rich phase SrN5 maintained structural stability when the pressure reached 50-70 GPa. The delocalization of π electrons from N atoms was the principal cause for its metallicity in SrN5. In this paper, our calculated results indicated that the energetic poly-nitrides in alkaline-earth-metal nitrides can be obtained by the high-pressure method.


Long-term effects of 6-week whole-body vibration on balance recovery and activities of daily living in the postacute phase of stroke: a randomized, controlled trial.
The long-term effects of 6-weeks whole-body vibration, as a novel method of somatosensory stimulation, on postural control and activities of daily living were compared with those of 6 weeks of exercise therapy on music of the same intensity in the postacute phase of stroke. Fifty-three patients with moderate to severe functional disabilities were randomized within 6 weeks poststroke and within 3 days after admission to a rehabilitation center to either whole-body vibration or exercise therapy on music in addition to a regular inpatient rehabilitation program. The whole-body vibration group received 4x45-second stimulation on the Galileo 900 (30-Hz frontal plane oscillations of 3-mm amplitude) for 5 days per week during 6 weeks. The exercise therapy on music group received the same amount of exercise therapy on music. Outcome variables included the Berg Balance Scale, Trunk Control Test, Rivermead Mobility Index, Barthel Index, Functional Ambulation Categories, Motricity Index, and somatosensory threshold at 0, 6, and 12 weeks follow up. At baseline, both groups were comparable in terms of prognostic factors and outcome measures. Both at 6 and 12 weeks follow up, no clinically relevant or statistical differences in outcome were observed between the groups. No side effects were reported. Daily sessions of whole-body vibration during 6 weeks are not more effective in terms of recovery of balance and activities of daily living than the same amount of exercise therapy on music in the postacute phase of stroke.


Dystrophic chicken muscle: altered synaptic acetylcholinesterase.
Individual motor endplates in the skeletal muscles of chickens genetically homozygous for muscular dystrophy have been compared with those in normal chickens. Measurements were made there, by specific autoradiographic techniques, of the numbers of total cholinesterase-like molecules and of acetylcholinesterase molecules. The acetylcholinesterase is distinctly decreased at the endplates in dystrophic muscles. The various data available on these muscles are compatible with the concept that a neural factor which determines the synaptic acetylcholinesterase, along with a number of other characters in the muscle cell, is defective in this disorder.


Same-day computed tomography during arterial portography and delayed high-dose iodine computed tomography: an efficient approach to imaging potentially resectable liver tumours.
Preoperative imaging in patients with potentially resectable liver tumours has traditionally been performed in specialized hepatobiliary centres. To assess the feasibility and value of establishing a same-day, pre-resectional hepatic imaging service, computed tomography during arterial portography (CTAP) and delayed high-dose iodine computed tomography (DICT) were used to examine patients considered suitable for curative hepatic resection. The study group comprised 27 patients (14 from hospitals not affiliated with a university and 13 from two university-affiliated hospitals) for whom pre-referral imaging showed tumour distribution amenable to resection. Among the patients from centres not affiliated with a university, seven had undergone ultrasonography (US) and equilibrium-phase CT, four had undergone US and unenhanced CT, two had undergone equilibrium-phase CT only and one had undergone bolus dynamic incremented CT (BDCT) only. For 12 of the patients coming from the university-affiliated centres, the pre-referral imaging had consisted of US and BDCT; the other had undergone US only. After CTAP and DICT, 20 (74%) of the 27 patients were reclassified as having unresectable disease: 7 (54%) of the 13 patients who had undergone BDCT before CTAP and DICT and 12 (92%) of the 13 patients who had undergone unenhanced or equilibrium-phase CT, as well as the patient who had undergone US only. By averting laparotomy in 20 of the patients, CTAP and DICT resulted in short-term health care savings of about $160,000. These findings suggest that BDCT was underused in the hospitals not affiliated with university centres.(ABSTRACT TRUNCATED AT 250 WORDS)


Loss of TIMP-3 promotes tumor invasion via elevated IL-6 production and predicts poor survival and relapse in HPV-infected non-small cell lung cancer.
Human papillomavirus (HPV) 16/18 E6 oncoprotein is expressed in lung tumors and is associated with p53 inactivation. The tissue inhibitor of metalloproteinase 3 (TIMP-3) is essential for limiting inflammation; therefore, we expected that TIMP-3 loss might induce chronic inflammation, thereby promoting tumor malignancy as well as poor survival and relapse in patients with HPV-infected non-small cell lung cancer. In this study, the loss of TIMP-3 by loss of heterozygosity and/or promoter hypermethylation was more frequent in HPV16/18 E6-positive tumors than in E6-negative tumors. To explore the possible underlying mechanism, E6-negative TL4 and CL1-0 cells were transfected with an E6 cDNA plasmid. A marked decrease in TIMP-3 expression was caused by promoter hypermethylation via increased DNA (cytosine-5-)-methyltransferase 1 (DNMT1) expression. Mechanistic studies indicated that TIMP-3 loss promoted interleukin-6 (IL-6) production, which led to cell invasion and anchorage-independent growth on soft agar plates. Kaplan-Meier and Cox regression models showed that patients with low-TIMP-3/high-IL-6 tumors had shorter overall survival and relapse-free survival periods when compared with patients with high-TIMP-3/low-IL-6 tumors. In summary, loss of TIMP-3 may increase IL-6 production via the tumor necrosis factor α/nuclear factor κB axis, thereby promoting tumor malignancy and subsequent relapse and poor survival in patients with HPV-infected non-small cell lung cancer.


A meta-analysis of the role of smoking in inflammatory bowel disease.
A relationship between not smoking and ulcerative colitis has been examined in all English reports. This paper evaluates the ulcerative colitis/nonsmoking and the Crohn's disease/smoking association by meta-analysis and against causality criterion for chronic diseases. A review of the literature, meta-analysis of selected studies, and assessment of causality criterion all suggest that not smoking and ulcerative colitis and smoking and Crohn's disease are consistent with a causal relationship. It is not inconceivable that tobacco may contain some substance beneficial to ulcerative colitis patients. Identification of the specific product in tobacco producing a beneficial effect would be a prudent next step in the study of this association. In spite of the findings of this review and analysis, the author finds no justification for health care providers to change the Surgeon General's recommendations on smoking or tobacco use for ulcerative colitis patients specifically.


Community health centers tackle rising demands and expectations.
As key providers of preventive and primary care for underserved people, including the uninsured, community health centers (CHCs) are the backbone of the U.S. health care safety net. Despite significant federal funding increases, community health centers are struggling to meet rising demand for care, particularly for specialty medical, dental and mental health services, according to findings from the Center for Studying Health System Change's (HSC) 2007 site visits to 12 nationally representative metropolitan communities. Health centers are responding to these pressures by expanding capacity and adding services but confront staffing, resource and other constraints. At the same time, CHCs are facing other demands, including increased quality reporting expectations, addressing racial and ethnic disparities, developing electronic medical records, and preparing for public health emergencies.


Antibacterial performance of bovine lactoferrin-fish gelatine electrospun membranes.
The increase of antibiotic resistant microorganisms urged the development and synthesis of novel antimicrobial biomaterials to be employed in a broad range of applications, ranging from food packaging to medical devices. This work describes the production and characterization of a protein-based electrospun fibrous membranes bearing antimicrobial properties. Its composition is exclusively comprised of proteins, with fish gelatine as structural matrix and bovine lactoferrin (bLF) as the active antimicrobial agent. The bLF bactericidal effect was determined against clinical isolates of Escherichia coli and Staphylococcus aureus through microdilution assays. Two distinctive methods were used to incorporate bLF into the fish gelatine nanofibres: (i) as a filler in the electrospinning formulation with concentrations of 2, 5 and 10 (wt%), and cross-linked with glutaraldehyde vapour, in order to achieve stability in aqueous solution; and (ii) through adsorption in a solution with 40mgmL(-1) bLF. Fourier transform infrared spectroscopy analysis showed that the structure of both proteins remained intact through the electrospinning blending and cross-linking procedure. Remarkable antibacterial properties were obtained with membranes containing 5% and 10% bLF with a bacterial reduction of approximately 90% and 100%, respectively.


Fibroblastic subpopulations in uninjured and wounded rabbit oral mucosa.
Fibroblast cultures derived from uninjured and reparative rabbit buccal mucosa were compared in terms of extracellular glycosaminoglycan (GAG) content and cellular response to interleukin-1 (IL-1). Under identical growth conditions, proliferation of both cell lines was the same. Both lines incorporated [3H]-glucosamine into GAG in cellular, pericellular, and medium fractions, with the majority of incorporated label residing in the medium. Dermatan sulfate (DS) was the predominant GAG in the medium fraction of both normal and wound fibroblast cultures; however, the two cell lines differed in the identity of the medium fraction's secondary GAG: chondroitin sulfate (CS) for normal fibroblasts and hyaluronic acid (HA) for wound-derived cells. The GAG content of the pericellular matrix for all cultures was the same regardless of the tissue of origin: heparan sulfate (HS) accompanied by a very small amount of CS. Exposure to IL-1 produced limited but highly specific effects: It was not mitogenic for either cell line but did cause a quantitative change (increase) in overall incorporation into GAG for medium and pericellular fractions for both cell lines. Further, IL-1 induced a qualitative change in GAG composition for normal mucosal fibroblastic medium fractions by causing the synthesis/release of heparan sulfate (HS) and a variant form of DS. These data support the hypothesis that different fibroblastic substrains can populate a given oral site as a function of variables such as injury and/or healing status.


Robust polymer microfluidic device fabrication via contact liquid photolithographic polymerization (CLiPP).
Microfluidic devices are commonly fabricated in silicon or glass using micromachining technology or elastomers using soft lithography methods; however, invariable bulk material properties, limited surface modification methods and difficulty in fabricating high aspect ratio devices prevent these materials from being utilized in numerous applications and/or lead to high fabrication costs. Contact Liquid Photolithographic Polymerization (CLiPP) was developed as an alternative microfabrication approach that uniquely exploits living radical photopolymerization chemistry to facilitate surface modification of device components, fabrication of high aspect ratio structures from many different materials with numerous covalently-adhered layers and facile construction of three-dimensional devices. This contribution describes CLiPP and demonstrates unique advantages of this new technology for microfabrication of polymeric microdevices. Specifically, the procedure for fabricating devices with CLiPP is presented, the living radical photopolymerization chemistry which enables this technology is described, and examples of devices made using CLiPP are shown.


Pyrazine odour makes visually conspicuous prey aversive.
Unpalatable insects frequently adopt multimodal signals to ward off predators, incorporating sounds and odours into their colourful displays. Pyrazine is an odour commonly used in insect warning displays, and has previously been shown to elicit unlearned biases against common warning colours, e.g. yellow and red in naive predators. We designed two experiments to test for similar effects of pyrazine on the conspicuousness of prey, perhaps the most ubiquitous aspect of aposematic coloration. In the first experiment, we offered predators (Gallus gallus domesticus) a choice between conspicuous crumbs and cryptic crumbs in the presence or absence of pyrazine. In the second experiment, we manipulated the birds' experience of conspicuous prey during an initial training phase. Only in the presence of pyrazine did birds show a bias against conspicuously coloured food, and this occurred whether or not they had previously experienced food that contrasted with the background. This emergent behaviour relied upon the visual and odorous signal components being presented together. These unlearned, yet hidden, responses against conspicuousness demonstrate that there are initial benefits to prey being conspicuous when the multimodal nature of warning signals is accounted for.


Cannabinoid Attenuation of Intestinal Inflammation in Chronic SIV-Infected Rhesus Macaques Involves T Cell Modulation and Differential Expression of Micro-RNAs and Pro-inflammatory Genes.
Cannabis use is frequent in HIV-infected individuals for its appetite stimulation and anti-inflammatory effects. To identify the underlying molecular mechanisms associated with these effects, we simultaneously profiled micro-RNA (miRNA) and mRNA expression in the colon of chronically simian immunodeficiency virus (SIV)-infected rhesus macaques administered either vehicle (VEH/SIV; n = 9) or Δ9-tetrahydrocannabinol (Δ9-THC; THC/SIV; n = 8). Pro-inflammatory miR-130a, miR-222, and miR-29b, lipopolysaccharide-responsive miR-146b-5p and SIV-induced miR-190b were significantly upregulated in VEH/SIV rhesus macaques. Compared to VEH/SIV rhesus macaques, 10 miRNAs were significantly upregulated in THC/SIV rhesus macaques, among which miR-204 was confirmed to directly target MMP8, an extracellular matrix-degrading collagenase that was significantly downregulated in THC/SIV rhesus macaques. Moreover, THC/SIV rhesus macaques failed to upregulate pro-inflammatory miR-21, miR-141 and miR-222, and alpha/beta-defensins, suggesting attenuated intestinal inflammation. Further, THC/SIV rhesus macaques showed higher expression of tight junction proteins (occludin, claudin-3), anti-inflammatory MUC13, keratin-8 (stress protection), PROM1 (epithelial proliferation), and anti-HIV CCL5. Gomori one-step trichrome staining detected significant collagen deposition (fibrosis) in the paracortex and B cell follicular zones of axillary lymph nodes from all VEH/SIV but not in THC/SIV rhesus macaques, thus demonstrating the ability of Δ9-THC to prevent lymph node fibrosis, a serious irreversible consequence of HIV induced chronic inflammation. Furthermore, using flow cytometry, we showed that Δ9-THC suppressed intestinal T cell proliferation/activation (Ki67/HLA-DR) and PD-1 expression and increased the percentages of anti-inflammatory CD163+ macrophages. Finally, while Δ9-THC did not affect the levels of CD4+ T cells, it significantly reduced absolute CD8+ T cell numbers in peripheral blood at 14 and 150 days post-SIV infection. These translational findings strongly support a role for differential miRNA/gene induction and T cell activation in Δ9-THC-mediated suppression of intestinal inflammation in HIV/SIV and potentially other chronic inflammatory diseases of the intestine.


Prevalence and Determinants of Fasted Training in Endurance Athletes: A Survey Analysis.
Athletes may choose to perform exercise in the overnight-fasted state for a variety of reasons related to convenience, gut comfort, or augmenting the training response, but it is unclear how many endurance athletes use this strategy. We investigated the prevalence and determinants of exercise performed in the overnight-fasted state among endurance athletes using an online survey and examined differences based on sex, competitive level, and habitual dietary pattern. The survey was completed by 1,950 endurance athletes (51.0% female, mean age 40.9 ± 11.1 years). The use of fasted training was reported by 62.9% of athletes, with significant effects of sex (p < .001, Cramer's V [φc] = 0.18, 90% CI [0.14, 0.22]), competitive level (p < .001, φc = 0.09, 90% CI [0.5, 0.13]), and habitual dietary pattern noted (p < .001, φc = 0.26, 90% CI [0.22, 0.29]). Males, nonprofessional athletes, and athletes following a low-carbohydrate, high-fat diet were most likely to perform fasted training. The most common reasons for doing so were related to utilizing fat as a fuel source (42.9%), gut comfort (35.5%), and time constraints/convenience (31.4%), whereas the most common reasons athletes avoided fasted training were that it does not help their training (47.0%), performance was worse during fasted training (34.7%), or greater hunger (34.6%). Overall, some athletes perform fasted training because they think it helps their training, whereas others avoid it because they think it is detrimental to their training goals, highlighting a need for future research. These findings offer insights into the beliefs and practices related to fasted-state endurance training.


Preference pulses and the win-stay, fix-and-sample model of choice.
Two groups of six rats each were trained to respond to two levers for a food reinforcer. One group was trained on concurrent variable-ratio 20 extinction schedules of reinforcement. The second group was trained on a concurrent variable-interval 27-s extinction schedule. In both groups, lever-schedule assignments changed randomly following reinforcement; a light cued the lever providing the next reinforcer. In the next condition, the light cue was removed and reinforcer assignment strictly alternated between levers. The next two conditions redetermined, in order, the first two conditions. Preference pulses, defined as a tendency for relative response rate to decline to the just-reinforced alternative with time since reinforcement, only appeared during the extinction schedule. Although the pulse's functional form was well described by a reinforcer-induction equation, there was a large residual between actual data and a pulse-as-artifact simulation (McLean, Grace, Pitts, & Hughes, 2014) used to discern reinforcer-dependent contributions to pulsing. However, if that simulation was modified to include a win-stay tendency (a propensity to stay on the just-reinforced alternative), the residual was greatly reduced. Additional modifications of the parameter values of the pulse-as-artifact simulation enabled it to accommodate the present results as well as those it originally accommodated. In its revised form, this simulation was used to create a model that describes response runs to the preferred alternative as terminating probabilistically, and runs to the unpreferred alternative as punctate with occasional perseverative response runs. After reinforcement, choices are modeled as returning briefly to the lever location that had been just reinforced. This win-stay propensity is hypothesized as due to reinforcer induction.


Velocity changes, long runs, and reversals in the Chromatium minus swimming response.
The velocity, run time, path curvature, and reorientation angle of Chromatium minus were measured as a function of light intensity, temperature, viscosity, osmotic pressure, and hydrogen sulfide concentration. C. minus changed both velocity and run time. Velocity decreased with increasing light intensity in sulfide-depleted cultures and increased in sulfide-replete cultures. The addition of sulfide to cultures grown at low light intensity (10 microeinsteins m-2 s-1) caused mean run times to increase from 10.5 to 20.6 s. The addition of sulfide to cultures grown at high light intensity (100 microeinsteins m-2 s-1) caused mean run times to decrease from 15.3 to 7.7 s. These changes were maintained for up to an hour and indicate that at least some members of the family Chromatiaceae simultaneously modulate velocity and turning frequency for extended periods as part of normal taxis.


The relations between joint action and theory of mind: a neuropsychological analysis.
We examined the relations between joint action and Theory of Mind (ToM) in neurological patients with impairments in ToM, in control patients (who passed ToM tasks) and non-lesioned controls. In two experiments, joint action was assessed in a "social Simon" procedure where spatial compatibility effects were tested under two-alternative forced-choice and under go/nogo conditions, which participants performed in isolation or alongside another participant (the joint action condition). In Experiment 1, patients with impaired ToM showed evidence of increased spatial compatibility effects under standard (two-alternative forced-choice) conditions but, unlike the control participants, these effects disappeared in the joint action condition. In Experiment 2, the ToM patients were asked to pay particular attention to their co-actor. With these instructions, ToM patients with lesions of posterior parietal cortex now showed a sustained spatial compatibility effect in the joint action condition, while ToM patients with lesions primarily involving frontal regions showed an initial effect of spatial compatibility that decreased across trials. The data suggest common processes involved in ToM processing and joint action effects, related to either the ability to attend to appropriate social cues (affected in posterior parietal patients) or the ability to recruit sufficient resources to code another's actions (affected in frontal patients).


Systematic review of cost effectiveness studies of telemedicine interventions.
To systematically review cost benefit studies of telemedicine. Systematic review of English language, peer reviewed journal articles. Searches of Medline, Embase, ISI citation indexes, and database of Telemedicine Information Exchange. STUDIES SELECTED: 55 of 612 identified articles that presented actual cost benefit data. Scientific quality of reports assessed by use of an established instrument for adjudicating on the quality of economic analyses. 557 articles without cost data categorised by topic. 55 articles with data initially categorised by cost variables employed in the study and conclusions. Only 24/55 (44%) studies met quality criteria justifying inclusion in a quality review. 20/24 (83%) restricted to simple cost comparisons. No study used cost utility analysis, the conventional means of establishing the "value for money" that a therapeutic intervention represents. Only 7/24 (29%) studies attempted to explore the level of utilisation that would be needed for telemedicine services to compare favourably with traditionally organised health care. None addressed this question in sufficient detail to adequately answer it. 15/24 (62.5%) of articles reviewed here provided no details of sensitivity analysis, a method all economic analyses should incorporate. There is no good evidence that telemedicine is a cost effective means of delivering health care.


Photoacoustic Sounds from Meteors.
Concurrent sound associated with very bright meteors manifests as popping, hissing, and faint rustling sounds occurring simultaneously with the arrival of light from meteors. Numerous instances have been documented with -11 to -13 brightness. These sounds cannot be attributed to direct acoustic propagation from the upper atmosphere for which travel time would be several minutes. Concurrent sounds must be associated with some form of electromagnetic energy generated by the meteor, propagated to the vicinity of the observer, and transduced into acoustic waves. Previously, energy propagated from meteors was assumed to be RF emissions. This has not been well validated experimentally. Herein we describe experimental results and numerical models in support of photoacoustic coupling as the mechanism. Recent photometric measurements of fireballs reveal strong millisecond flares and significant brightness oscillations at frequencies ≥40 Hz. Strongly modulated light at these frequencies with sufficient intensity can create concurrent sounds through radiative heating of common dielectric materials like hair, clothing, and leaves. This heating produces small pressure oscillations in the air contacting the absorbers. Calculations show that -12 brightness meteors can generate audible sound at ~25 dB SPL. The photoacoustic hypothesis provides an alternative explanation for this longstanding mystery about generation of concurrent sounds by fireballs.


An active play intervention to improve physical activity and fundamental movement skills in children of low socio-economic status: feasibility cluster randomised controlled trial.
Active play is a novel approach to addressing low physical activity levels and fundamental movement skills (FMS) in childhood and new interventions must be developed and evaluated. This study aimed to determine the feasibility of a 10-week school-based 'active play' intervention, and present preliminary findings on four outcomes: physical activity levels, FMS, inhibition, and maths fluency. This was a feasibility cluster RCT in which eight schools (one primary three class per school) were paired and randomly allocated to either the 10-week intervention (n = 4) or waiting-list control (n = 4). The active play intervention consisted of a 1-h outdoor physical activity session per week, incorporating 30 min of facilitated games and 30 min of free play. Feasibility measures were gathered using appropriate methods and physical activity was measured using an ActiGraph GT3X accelerometer, FMS were assessed using the Test of Gross Motor Development-2 (TGMD-2), inhibition was measured using a Flanker Test and maths fluency was assessed using the One Minute Basic Number Facts Test. Sixty-six percent of eligible children (n = 137) agreed to participate in the research. No schools withdrew from the study and three participants were lost to follow-up. Compliance to the intervention was high-none of the participants missed more than two of the ten scheduled active play sessions. Data lost to follow-up were minimal; most were lost (14%) for school day physical activity. Active play sessions were shorter than planned on average by 10 min, and participants spent a mean of 39.4% (14.2) of the session time in moderate-to-vigorous intensity physical activity (MVPA). There was preliminary evidence of a small intervention effect on MVPA (d = 0.3), FMS score (d = 0.4), inhibition (fish trial: d = 0.1, arrow trial d = 0.1) and maths fluency (addition: d = 0.3, subtraction: d = 0.1). The active play intervention was feasible and benefitted from a relatively high MVPA content; however, preliminary findings suggest the intervention had a small effect on the outcomes. Having more active play sessions per week and/or extending the duration of the intervention may increase the effects and these should be tested before a future definitive cluster RCT is undertaken. This trial was registered on the International Standardised Randomised Controlled Trials Number register (ISRCTN) in August 2017 (ISRCTN11607781).


A Biochemical Nickel(I) State Supports Nucleophilic Alkyl Addition: A Roadmap for Methyl Reactivity in Acetyl Coenzyme A Synthase.
Nickel-containing enzymes such as methyl coenzyme M reductase (MCR) and carbon monoxide dehydrogenase/acetyl coenzyme A synthase (CODH/ACS) play a critical role in global energy conversion reactions, with significant contributions to carbon-centered processes. These enzymes are implied to cycle through a series of nickel-based organometallic intermediates during catalysis, though identification of these intermediates remains challenging. In this work, we have developed and characterized a nickel-containing metalloprotein that models the methyl-bound organometallic intermediates proposed in the native enzymes. Using a nickel(I)-substituted azurin mutant, we demonstrate that alkyl binding occurs via nucleophilic addition of methyl iodide as a methyl donor. The paramagnetic NiIII-CH3 species initially generated can be rapidly reduced to a high-spin NiII-CH3 species in the presence of exogenous reducing agent, following a reaction sequence analogous to that proposed for ACS. These two distinct bioorganometallic species have been characterized by optical, EPR, XAS, and MCD spectroscopy, and the overall mechanism describing methyl reactivity with nickel azurin has been quantitatively modeled using global kinetic simulations. A comparison between the nickel azurin protein system and existing ACS model compounds is presented. NiIII-CH3 Az is only the second example of two-electron addition of methyl iodide to a NiI center to give an isolable species and the first to be formed in a biologically relevant system. These results highlight the divergent reactivity of nickel across the two intermediates, with implications for likely reaction mechanisms and catalytically relevant states in the native ACS enzyme.


Extracting phosphorous from incinerated sewage sludge ash rich in iron or aluminum.
Ashes from mono-incineration of sewage sludge (ISSA) generally contain high concentrations of phosphorous (P) and can be regarded as secondary P resources. ISSA has no direct value as fertilizer as P is not plant available. The present paper experimentally compares P extraction in acid from two different ISSAs; one rich in Al (67g/kg) and the other in Fe (58g/kg). The difference related to P precipitation at the waste water treatment facilities. Another major difference between the ashes was that flue gas purification products were mixed into the first ash and it contained about 5% activated carbon. The Al rich ash had a significantly higher buffering capacity and required more acid for extraction of P. When acid extraction of P from ISSA is the method for recovery, it is thus beneficial to go back to the waste water treatment facility and e.g. choose Fe for P precipitation rather than Al. Formation of a high amount of gypsum crystals in both ashes after extraction in H2SO4 was seen by SEM-EDX. H2SO4 is the cheapest mineral ash, but the gypsum formation must be taken into account when either finding possibility for using the remaining ash in e.g. construction materials or if the choice is deposition, as the gypsum increases the volume significantly.


Genome-wide association analysis on five isolated populations identifies variants of the HLA-DOA gene associated with white wine liking.
Wine is the most popular alcoholic beverage around the world and because of its importance in society has been widely studied. Understanding what drives its flavor has been a quest for decades but much is still unknown and will be determined at least in part by individual taste preferences. Recently studies in the genetics of taste have uncovered the role of different genes in the determination of food preferences giving new insight on its physiology. In this context we have performed a genome-wide association study on red and white wine liking using three isolated populations collected in Italy, and replicated our results on two additional populations coming from the Netherland and Central Asia for a total of 3885 samples. We have found a significant association (P=2.1 × 10(-8)) between white wine liking and rs9276975:C>T a polymorphism in the HLA-DOA gene encoding a non-canonical MHC II molecule, which regulates other MHC II molecules. The same association was also found with red wine liking (P=8.3 × 10(-6)). Sex-separated analysis have also revealed that the effect of HLA-DOA is twice as large in women as compared to men suggesting an interaction between this polymorphism and gender. Our results are one of the first examples of genome-wide association between liking of a commonly consumed food and gene variants. Moreover, our results suggest a role of the MHC system in the determination of food preferences opening new insight in this field in general.


Moving toward personalized medicine in the methadone maintenance treatment program: a pilot study on the evaluation of treatment responses in Taiwan.
This pilot study simultaneously evaluated the effects of various factors, including genetic variations of CYP2B6, CYP2C19, and ABCB1, demographic characteristics, disease states, methadone-drug interactions (MDIs), and poly-substance use, on the treatment responses among non-HIV patients in the methadone maintenance treatment program (MMTP) in Taiwan. A total of 178 patients were recruited from two major hospitals that provided MMTP services in southern Taiwan, and information regarding concomitant medications and diseases was acquired from the National Health Insurance (NHI) program. The results demonstrated that the methadone maintenance dose, CYP2B6 785G allele, and ABCB1 2677T allele have positive effects on the methadone plasma concentration. In contrast, patients with HCV coinfection, alcohol problems, and psychiatric diseases may have a negative response to treatment. Thus, a comprehensive evaluation of treatment responses in the MMTP should include not only genetic polymorphisms in methadone metabolism and transporter proteins, but also concomitant diseases, MDIs, and poly-substance use. The results also suggest that personalized medicine may be indispensable for a better outcome of the MMTP.


The Role of CD4 and CD8 T Cells in Human Cutaneous Leishmaniasis.
Leishmaniasis, caused by infection with parasites of the Leishmania genus, affects millions of individuals worldwide. This disease displays distinct clinical manifestations ranging from self-healing skin lesions to severe tissue damage. The control of Leishmania infection is dependent on cellular immune mechanisms, and evidence has shown that CD4 and CD8 T lymphocytes play different roles in the outcome of leishmaniasis. Although the presence of CD4 T cells is important for controlling parasite growth, the results in the literature suggest that the inflammatory response elicited by these cells could contribute to the pathogenesis of lesions. However, recent studies on CD8 T lymphocytes show that these cells are mainly involved in tissue damage through cytotoxic mechanisms. In this review, we focus on the recent advances in the study of the human adaptive immunological response in the pathogenesis of tegumentary leishmaniasis.