Relative virulence of three isolates of Piscirickettsia salmonis for coho salmon Oncorhynchus kisutch.
Piscirickettsia salmonis was first recognized as the cause of mortality among pen-reared coho salmon Oncorhynchus kisutch in Chile. Since the initial isolation of this intracellular Gram-negative bacterium in 1989, similar organisms have been described from several areas of the world, but the associated outbreaks were not reported to be as serious as those that occurred in Chile. To determine if this was due to differences in virulence among isolates of P. salmonis, we conducted an experiment comparing isolates from Chile, British Columbia, Canada, and Norway (LF-89, ATL-4-91 and NOR-92, respectively). For each of the isolates, 3 replicates of 30 coho salmon were injected intraperitoneally with each of 3 concentrations of the bacterium. Negative control fish were injected with MEM-10. Mortalities were collected daily for 41 d post-injection. Piscirickettsiosis was observed in fish injected with each of the 3 isolates, and for each isolate, cumulative mortality was directly related to the concentration of bacterial cells administered. The LF-89 isolate was the most virulent, with losses reaching 97% in the 3 replicates injected with 10(5.0) TCID50, 91% in the replicates injected with 10(4.0) TCID50, and 57% in the fish injected with 10(3.0) TCID50. The ATL-4-91 isolate caused losses of 92% in the 3 replicates injected with 10(5.0) TCID50, 76% in the fish injected with 10(4.0) TCID50, and 32% in those injected with 10(3.0) TCID50. The NOR-92 isolate was the least virulent, causing 41% mortality in the replicates injected with 10(4.6) TCID50. At 41 d post-injection, 6% of the fish injected with 10(3.6) TCID50 NOR-92 had died. Mortality was only 2% in the fish injected with 10(2.6) TCID50 NOR-92, which was the same as the negative control group. Because the group injected with the highest concentration (10(4.6) TCID50) of NOR-92 was still experiencing mortality at 41 d, it was held for an additional 46 d. At 87 d post-injection, the cumulative mortality in this group had reached 70%. These differences in virulence among the isolates were statistically significant (p < 0.0001), and are important for the management of affected stocks of fish.


Craniopharyngioma or its surgery induces diabetes mellitus. Case report.
A 23-year-old non-diabetic woman presented to our emergency room with progressive headache. She was diagnosed with craniopharyngioma and received tumor resection. Her blood glucose level was within the normal limit before surgery, and she had no family history of diabetes. Three months after the surgery, acute hyperosmolar hyperglycemic state developed. After 4 months of follow-up, her diabetes persisted but improved with oral antidiabetic drugs. This is the third case report of diabetes developing several months after craniopharyngioma tumor resection. The possible mechanisms of cranipharyngioma or its surgery inducing diabetes mellitus are hypothalamic obesity or hypothalamic damage. The degree of hypothalamic damage before the operation is predictive of diabetes development, and blood glucose level monitoring is important for these patients.


Akt promotes survival of cardiomyocytes in vitro and protects against ischemia-reperfusion injury in mouse heart.
IGF-1 has been shown to protect myocardium against death in animal models of infarct and ischemia-reperfusion injury. In the present study, we investigated the role of the IGF-1-regulated protein kinase Akt in cardiac myocyte survival in vitro and in vivo. IGF-1 promoted survival of cultured cardiomyocytes under conditions of serum deprivation in a dose-dependent manner but had no effect on cardiac fibroblast survival. The cytoprotective effect of IGF-1 on cardiomyocytes was abrogated by the phosphatidylinositol 3-kinase (PI 3-kinase) inhibitor wortmannin. Wortmannin had no effect on cardiomyocyte viability in the absence of IGF-1. IGF-1-mediated cytoprotection correlated with the wortmannin-sensitive induction of Akt protein kinase activity. To examine the functional consequences of Akt activation in cardiomyocyte survival, replication-defective adenoviral constructs expressing wild-type, dominant-negative, and constitutively active Akt genes were constructed. Transduction of dominant-negative Akt blocked IGF-1-induced survival but had no effect on cardiomyocyte survival in the absence of IGF-1. In contrast, transduction of wild-type Akt enhanced cardiomyocyte survival at subsaturating levels of IGF-1, whereas constitutively active Akt protected cardiomyocytes from apoptosis in the absence of IGF-1. After transduction into the mouse heart in vivo, constitutively active Akt protected against myocyte apoptosis in response to ischemia-reperfusion injury. These data are the first documentation that Akt functions to promote cellular survival in vivo, and they indicate that the activation of this pathway may be useful in promoting myocyte survival in the diseased heart.


Classifying the wandering mind: revealing the affective content of thoughts during task-free rest periods.
Many powerful human emotional thoughts are generated in the absence of a precipitating event in the environment. Here, we tested whether we can decode the valence of internally driven, self-generated thoughts during task-free rest based on neural similarities with task-related affective mental states. We acquired functional magnetic resonance imaging (fMRI) data while participants generated positive and negative thoughts as part of an attribution task (Session A) and while they reported the occurrence of comparable mental states during task-free rest periods (Session B). With the use of multivariate pattern analyses (MVPA), we identified response patterns in the medial orbitofrontal cortex (mOFC) that encode the affective content of thoughts that are generated in response to an external experimental cue. Importantly, these task driven response patterns reliably predicted the occurrence of affective thoughts generated during unconstrained rest periods recorded one week apart. This demonstrates that at least certain elements of task-cued and task-free affective experiences rely on a common neural code. Furthermore, our findings reveal the role that the mOFC plays in determining the affective tone of unconstrained thoughts. More generally, our results suggest that MVPA is an important methodological tool for attempts to understand unguided subject driven mental states such as mind-wandering and daydreaming based on neural similarities with task-based experiences.


Association Between Childhood Consumption of Ultraprocessed Food and Adiposity Trajectories in the Avon Longitudinal Study of Parents and Children Birth Cohort.
Reports of associations between higher consumption of ultraprocessed foods (UPF) and elevated risks of obesity, noncommunicable diseases, and mortality in adults are increasing. However, associations of UPF consumption with long-term adiposity trajectories have never been investigated in children. To assess longitudinal associations between UPF consumption and adiposity trajectories from childhood to early adulthood. This prospective birth cohort study included children who participated in the Avon Longitudinal Study of Parents and Children (ALSPAC) in Avon County, southwest England. Children were followed up from 7 to 24 years of age during the study period from September 1, 1998, to October 31, 2017. Data were analyzed from March 1, 2020, to January 31, 2021. Baseline dietary intake data were collected using 3-day food diaries. Consumption of UPF (applying the NOVA food classification system) was computed as a percentage of weight contribution in the total daily food intake for each participant and categorized into quintiles. Repeated recordings of objectively assessed anthropometrics (body mass index [BMI; calculated as weight in kilograms divided by height in meters squared], weight, and waist circumference) and dual-energy x-ray absorptiometry measurements (fat and lean mass indexes [calculated as fat and lean mass, respectively, divided by height in meters squared] and body fat percentage). Associations were evaluated using linear growth curve models and were adjusted for study covariates. A total of 9025 children (4481 [49.7%] female and 4544 [50.3%] male) were followed up for a median of 10.2 (interquartile range, 5.2-16.4) years. The mean (SD) UPF consumption at baseline was 23.2% (5.0%) in quintile 1, 34.7% (2.5%) in quintile 2, 43.4% (2.5%) in quintile 3, 52.7% (2.8%) in quintile 4, and 67.8% (8.1%) in quintile 5. Among those in the highest quintile of UPF consumption compared with their lowest quintile counterpart, trajectories of BMI increased by an additional 0.06 (95% CI, 0.04-0.08) per year; fat mass index, by an additional 0.03 (95% CI, 0.01-0.05) per year; weight, by an additional 0.20 (95% CI, 0.11-0.28) kg per year; and waist circumference, by an additional 0.17 (95% CI, 0.11-0.22) cm per year. These findings suggest that higher UPF consumption is associated with greater increases in adiposity from childhood to early adulthood. Robust public health measures that promote minimally processed foods and discourage UPF consumption among children are urgently needed to reduce obesity in England and globally.


The major DNA repair pathway after both proton and carbon-ion radiation is NHEJ, but the HR pathway is more relevant in carbon ions.
The purpose of this study was to identify the roles of non-homologous end-joining (NHEJ) or homologous recombination (HR) pathways in repairing DNA double-strand breaks (DSBs) induced by exposure to high-energy protons and carbon ions (C ions) versus gamma rays in Chinese hamster cells. Two Chinese hamster cell lines, ovary AA8 and lung fibroblast V79, as well as various mutant sublines lacking DNA-PKcs (V3), X-ray repair cross-complementing protein-4 [XRCC4 (XR1), XRCC3 (irs1SF) and XRCC2 (irs1)] were exposed to gamma rays ((137)Cs), protons (200 MeV; 2.2 keV/μm) and C ions (290 MeV; 50 keV/μm). V3 and XR1 cells lack the NHEJ pathway, whereas irs1 and irs1SF cells lack the HR pathway. After each exposure, survival was measured using a clonogenic survival assay, in situ DSB induction was evaluated by immunocytochemical analysis of histone H2AX phosphorylation at serine 139 (γ-H2AX foci) and chromosome aberrations were examined using solid staining. The findings from this study showed that clonogenic survival clearly depended on the NHEJ and HR pathway statuses, and that the DNA-PKcs(-/-) cells (V3) were the most sensitive to all radiation types. While protons and γ rays yielded almost the same biological effects, C-ion exposure greatly enhanced the sensitivity of wild-type and HR-deficient cells. However, no significant enhancement of sensitivity in cell killing was seen after C-ion irradiation of NHEJ deficient cells. Decreases in the number of γ-H2AX foci after irradiation occurred more slowly in the NHEJ deficient cells. In particular, V3 cells had the highest number of residual γ-H2AX foci at 24 h after C-ion irradiation. Chromosomal aberrations were significantly higher in both the NHEJ- and HR-deficient cell lines than in wild-type cell lines in response to all radiation types. Protons and gamma rays induced the same aberration levels in each cell line, whereas C ions introduced higher but not significantly different aberration levels. Our results suggest that the NHEJ pathway plays an important role in repairing DSBs induced by both clinical proton and C-ion beams. Furthermore, in C ions the HR pathway appears to be involved in the repair of DSBs to a greater extent compared to gamma rays and protons.


Network cloning using DNA barcodes.
The connections between neurons determine the computations performed by both artificial and biological neural networks. Recently, we have proposed SYNSeq, a method for converting the connectivity of a biological network into a form that can exploit the tremendous efficiencies of high-throughput DNA sequencing. In SYNSeq, each neuron is tagged with a random sequence of DNA-a "barcode"-and synapses are represented as barcode pairs. SYNSeq addresses the analysis problem, reducing a network into a suspension of barcode pairs. Here, we formulate a complementary synthesis problem: How can the suspension of barcode pairs be used to "clone" or copy the network back into an uninitialized tabula rasa network? Although this synthesis problem might be expected to be computationally intractable, we find that, surprisingly, this problem can be solved efficiently, using only neuron-local information. We present the "one-barcode-one-cell" (OBOC) algorithm, which forces all barcodes of a given sequence to coalesce into the same neuron, and show that it converges in a number of steps that is a power law of the network size. Rapid and reliable network cloning with single-synapse precision is thus theoretically possible.


Analysis of a multiple reflective translucent photocathode.
In order to evaluate methods of enhancing the response of state-of-the-art photocathodes, such as the S-20, we have used a mathematical absorption-diffusion model to study the effects of skewed incidence of light on the quantum efficiency. The phenomenon of increased optical absorption due to multiple reflections of light within the emitting layer-glass substrate combination is treated as a function of the angle of incidence. When the quantum efficiency generated for skewed incidence is compared with the quantum yield generated for normal incidence, a gain of 3.5 is predicted. The gain is greatest for the smallest absorption coefficient-photocathode thickness product.


Extraction of dynamic features from hand drawn data for the identification of children with handwriting difficulty.
Studies have shown that differences between children with and without handwriting difficulties lie not only in the written product (static data) but also in dynamic data of handwriting process. Since writing system varies among countries and individuals, this study was conducted to determine the feasibility of using quantitative outcome measures of children's drawing to identify children who are at risk of handwriting difficulties. A sample of 143 first graders of a normal primary school was investigated regarding their handwriting ability. The children were divided into two groups: test and control. Ten children from test group and 40 children from control group were individually tested for their Visual Motor Integration skills. Analysis on dynamic data indicated significant differences between the two groups in temporal and spatial measures of the drawing task performance. Thus, kinematic analysis of children's drawing is feasible to provide performance characteristic of handwriting ability, supporting its use in screening for handwriting difficulty.


Predeployment mass casualty and clinical trauma training for US Army forward surgical teams.
Since the beginning of the program in 2002, 84 Forward Surgical Teams (FSTs) have rotated through the Army Trauma Training Center (ATTC) at the University of Miami/Ryder Trauma Center including all those deployed to Iraq and Afghanistan. The purpose of this study was to provide the latest updates of our experience with FSTs at the ATTC. Before deployment, each FST participates in a 2-week training rotation at the ATTC. The rotation is divided into 3 phases. Phase 1 is to refresh FST knowledge regarding the initial evaluation and management of the trauma patient. Phase 2 is the clinical phase and is conducted entirely at the Ryder Trauma Center. The training rotation culminates in phase 3, the Capstone exercise. During the Capstone portion of their training, the entire 20-person FST remains at the Ryder Trauma Center and is primarily responsible for the evaluation and resuscitation of all patients arriving over a 24-hour period. Subject awareness concerning their role within the team improved from 71% to 95%, indicating that functioning as a team in the context of the mass casualty training exercise along with clinical codes was beneficial. The clinical component of the rotation was considered by 47% to be the most valuable aspect of the training. Our experience strongly suggests that a multimodality approach is beneficial for preparing a team of individuals with minimal combat (or trauma) experience for the rigors of medical care and triage on the battlefield. The data provided by participants rotating through the ATTC show that through clinical exposure and simulation over a 2-week period, FST performance is optimized by defining provider roles and improving communication. The mass casualty training exercise is a vital component of predeployment training that participants feel is valuable in preparing them for the challenges that lay ahead.


Fat and fibrosis as confounding cofactors in viscoelastic measurements of the liver.
Elastography provides significant information on staging of fibrosis in patients with liver disease and may be of some value in assessing steatosis. However, there remain questions as to the role of steatosis and fibrosis as cofactors influencing the viscoelastic measurements of liver tissues, particularly shear wave speed (SWS) and shear wave attenuation (SWA). In this study, by employing the theory of composite elastic media as well as two independent experimental measurements on oil-in-gelatin phantoms and also finite element simulations, it is consistently shown that fat and fibrosis jointly influence the SWS and SWA measurements. At a constant level of fat, fibrosis stages can influence the SWA by factors of 2-4. Moreover, the rate of increase in SWA with increasing fat is strongly influenced by the stages of fibrosis; softer background cases (low fibrosis stages) have higher rate of SWA increase with fat than those with stiffer moduli (higher fibrosis stages). Meanwhile, SWS results are influenced by the presence of fat, however the degree of variability is more subtle. The results indicate the importance of jointly considering fat and fibrosis as contributors to SWS and SWA measurements in complex liver tissues and in the design and interpretation of clinical trials.


Steady-state kinetic characterization of RB69 DNA polymerase mutants that affect dNTP incorporation.
The function of six highly conserved residues (Arg482, Lys483, Lys486, Lys560, Asn564, and Tyr567) in the fingers domain of bacteriophage RB69 DNA polymerase (RB69 gp43) were analyzed by kinetic studies with mutants in which each of these residues was replaced with Ala. Our results suggest that Arg482, Lys486, Lys560, and Asn564 contact the incoming dNTP during the nucleotidyl transfer reaction as judged by variations in apparent Km and kcat values for dNTP incorporation by these mutants compared to those for the exonuclease deficient parental polymerase under steady-state conditions. On the basis of our studies, as well as on the basis of the crystal structure of RB69 gp43, we propose that a conformational change in the fingers domain, which presumably occurs prior to polymerization, brings the side chains of Arg482, Lys486, Lys560, and Asn564 into the vicinity of the primer-template terminus where they can contact the triphosphate moiety of the incoming dNTP. In particular, on the basis of structural studies reported for the "closed" forms of two other DNA polymerases and from the kinetic studies reported here, we suggest that (i) Lys560 and Asn564 contact the nonbonding oxygens of the alpha and beta phosphates, respectively, and (ii) both Arg482 and Lys486 contact the gamma phosphate oxygens of the incoming dNTP of RB69 gp43 prior to the nucleotidyl transfer reaction. We also found that Ala substitutions at each of these four RB69 gp43 sites could incorporate dGDP as a substrate, although with markedly reduced efficiency compared to that with dGTP. In contrast in the parental exo- background, the K483A and Y567A substituted enzymes could not use dGDP as a substrate for primer extension. These results, taken together, are consistent with the putative roles of the four conserved residues in RB69 gp43 as stated above.


Bone marrow fibrosis in primary myelofibrosis: pathogenic mechanisms and the role of TGF-β.
Primary myelofibrosis (PMF) is a Philadelphia chromosome negative myeloproliferative neoplasm (MPN) with adverse prognosis and is associated with bone marrow fibrosis and extramedullary hematopoiesis. Even though the discovery of the Janus kinase 2 (JAK2), thrombopoietin receptor (MPL) and calreticulin (CALR) mutations have brought new insights into the complex pathogenesis of MPNs, the etiology of fibrosis is not well understood. Furthermore, since JAK2 inhibitors do not lead to reversal of fibrosis further understanding of the biology of fibrotic process is needed for future therapeutic discovery. Transforming growth factor beta (TGF-β) is implicated as an important cytokine in pathogenesis of bone marrow fibrosis. Various mouse models have been developed and have established the role of TGF-β in the pathogenesis of fibrosis. Understanding the molecular alterations that lead to TGF-β mediated effects on bone marrow microenvironment can uncover newer therapeutic targets against myelofibrosis. Inhibition of the TGF-β pathway in conjunction with other therapies might prove useful in the reversal of bone marrow fibrosis in PMF.


Lumbar Facet Joint Kinematics and Load Effects During Dynamic Lifting.
To examine the lumbar facet joint kinematics in vivo during dynamic lifting and the effects of the load lifted. Although extensive efforts have been dedicated to investigating the risk factors of low back pain (LBP) associated with load handling in the workplace, the biomechanics of lumbar facet joints during such activities is not well understood. Fourteen healthy participants performed a load-lifting task while a dynamic stereo-radiography system captured their lumbar motion continuously. Data from 11 participants were included for subsequent analysis. A randomized block design was employed to study the load effect (4.5 kg, 9.0 kg, and 13.5 kg) on bilateral facet joint motions at approximately 60°, 40°, 20°, and 0° trunk-flexion postures. The facet orientations were also examined. Significant load effects were found for the flexion and lateral bending and superior-inferior translation of the facet joints. The L5-S1 displayed greater lateral bending and twisting, which was due to its more posterolateral orientation than the L2-L3, L3-L4, and L4-L5 facet joints. The left-right asymmetry in facet orientation was observed, most prominently at L3-L4 and L5-S1 facet joints. The lumbar facet joint kinematics are affected by the magnitude of the lifted load and are dependent on the orientations of articulating adjacent facets. This study provided new insights into the role of lumbar facet joints in vivo during lifting. Alterations in the facet joint kinematics due to vigorous functional demand can be one of the primary but overlooked mechanical factors in the causation of LBP.


Effect of carbon monoxide on rabbit cerebral arteries.
Carbon monoxide produces relaxation in some peripheral arteries. Recently it has been suggested that carbon monoxide may be generated in brain tissue. In the present study we examined the hypothesis that carbon monoxide directly relaxes cerebral blood vessels. The aorta and basilar and middle cerebral arteries were removed from New Zealand White rabbits and mounted for tension recording in vitro. Canine basilar arteries were also studied. After precontraction, cumulative relaxation concentration-response curves to carbon monoxide, nitric oxide, sodium nitroprusside, acetylcholine (rabbit arteries), and ATP (dog basilar artery) were obtained. Maximum relaxation and the concentration of agonists that induced half-maximal relaxation (ED50) were determined. Carbon monoxide (10(-6) to 3 x 10(-4) mol/L) did not affect tension in rabbit or dog cerebral arteries. In rabbit aorta, carbon monoxide induced 29 +/- 4% (mean +/- SEM) relaxation at the highest concentration used (3 x 10(-4) mol/L). In contrast, nitric oxide produced 80% to 100% relaxation of all arteries, with ED50 values ranging from 7.1 to 7.4 -log mol/L. Nitroprusside, acetylcholine, and ATP also produced 80% to 100% relaxation of the arteries. Carbon monoxide does not appear to have significant effect on tone in cerebral arteries. In contrast, at high concentrations carbon monoxide produces concentration-dependent relaxation in rabbit aorta. Factors that account for this regional heterogeneity are not clear. Although neurons may produce both nitric oxide and carbon monoxide, our findings suggest that only nitric oxide has direct effects on cerebral vascular tone.


Prevalence of viscoelastic relaxation after the 2011 Tohoku-oki earthquake.
After a large subduction earthquake, crustal deformation continues to occur, with a complex pattern of evolution. This postseismic deformation is due primarily to viscoelastic relaxation of stresses induced by the earthquake rupture and continuing slip (afterslip) or relocking of different parts of the fault. When postseismic geodetic observations are used to study Earth's rheology and fault behaviour, it is commonly assumed that short-term (a few years) deformation near the rupture zone is caused mainly by afterslip, and that viscoelasticity is important only for longer-term deformation. However, it is difficult to test the validity of this assumption against conventional geodetic data. Here we show that new seafloor GPS (Global Positioning System) observations immediately after the great Tohoku-oki earthquake provide unambiguous evidence for the dominant role of viscoelastic relaxation in short-term postseismic deformation. These data reveal fast landward motion of the trench area, opposing the seaward motion of GPS sites on land. Using numerical models of transient viscoelastic mantle rheology, we demonstrate that the landward motion is a consequence of relaxation of stresses induced by the asymmetric rupture of the thrust earthquake, a process previously unknown because of the lack of near-field observations. Our findings indicate that previous models assuming an elastic Earth will have substantially overestimated afterslip downdip of the rupture zone, and underestimated afterslip updip of the rupture zone; our knowledge of fault friction based on these estimates therefore needs to be revised.


Respiratory viruses in exacerbations of chronic obstructive pulmonary disease requiring hospitalisation: a case-control study.
Acute exacerbations of chronic obstructive pulmonary disease (AE-COPD) are a common cause of hospital admission. Many exacerbations are believed to be due to upper and/or lower respiratory tract viral infections, but the incidence of these infections in patients with COPD is still undetermined. Respiratory syncytial virus (RSV), influenza A and B, parainfluenza 3, and picornaviruses were detected by nested reverse transcription polymerase chain reaction (RT-PCR) in upper (nasal lavage) and lower respiratory tract specimens (induced sputum). In a 2:1 case-control set up, 85 hospitalised patients with AE-COPD and 42 patients with stable COPD admitted for other medical reasons were studied. Respiratory viruses were found more often in sputum and nasal lavage of patients with AE-COPD (48/85, 56%) than in patients with stable COPD (8/42, 19%, p<0.01). The most common viruses were picornaviruses (21/59, 36%), influenza A (15/59, 25%), and RSV (13/59, 22%). When specimens were analysed separately, this difference was seen in induced sputum (exacerbation 40/85 (47%) v stable 4/42 (10%), p<0.01) but was not significant in nasal lavage (exacerbation 26/85 (31%) v stable 7/42 (17%), p=0.14). In patients with AE-COPD, fever was more frequent in those in whom viruses were detected (12/48, 25%) than in those in whom viruses were not detected (2/37, 5%, p=0.03). Viral respiratory pathogens are found more often in respiratory specimens of hospitalised patients with AE-COPD than in control patients. Induced sputum detects respiratory viruses more frequently than nasal lavage in these patients. These data indicate that nasal lavage probably has no additional diagnostic value to induced sputum in cross-sectional studies on hospitalised patients with AE-COPD and that the role of viral infection in these patients is still underestimated.


"Isolated" elevation of alkaline phosphatase: significance in hospitalized patients.
The significance of alkaline phosphatase (AP) elevation with otherwise normal transaminase and bilirubin values remains unclear. We evaluated the clinical outcome of hospitalized patients with an "isolated" AP elevation. Eighty-seven inpatients with isolated AP elevation were identified during February 1984 and followed for 1 year. Forty-five of 87 patients had normalization of AP during the follow-up period, usually within 1-3 months. The most common diagnoses in this group were congestive heart failure in nine, benign bone disease in six, and treatable malignancy in three patients. Twelve patients had no apparent explanation for the transient rise of AP. Persistent AP elevations were noted in 42 patients--14 of whom had terminal malignancies. Clinically obvious life-threatening diagnoses were made in 24 of the patients with persistent AP elevation. The etiology of AP elevation remained enigmatic in seven patients: two died, four are stable during 1 1/2-3 years of follow-up, and one patient was found to have metastatic carcinoma 17 months later. If the initial AP was greater than 1 1/2 times normal, there was a higher likelihood of persistent elevation (68% vs. 41%, p less than 0.05). Isolated elevations of AP in inpatients may be associated with a variety of medical illnesses and often normalize within months. If the AP elevation is persistent, there is usually a clinically obvious diagnosis. A reasonable approach to such patients is a careful history, physical exam, and routine lab studies to detect obvious diagnoses, followed by repeat enzyme determination at 1-3 months.


Individual Control and Quantification of 3D Spheroids in a High-Density Microfluidic Droplet Array.
As three-dimensional cell culture formats gain in popularity, there emerges a need for tools that produce vast amounts of data on individual cells within the spheroids or organoids. Here, we present a microfluidic platform that provides access to such data by parallelizing the manipulation of individual spheroids within anchored droplets. Different conditions can be applied in a single device by triggering the merging of new droplets with the spheroid-containing drops. This allows cell-cell interactions to be initiated for building microtissues, studying stem cells' self-organization, or observing antagonistic interactions. It also allows the spheroids' physical or chemical environment to be modulated, as we show by applying a drug over a large range of concentrations in a single parallelized experiment. This convergence of microfluidics and image acquisition leads to a data-driven approach that allows the heterogeneity of 3D culture behavior to be addressed across the scales, bridging single-cell measurements with population measurements.


Cost and cost-effectiveness of switching from d4T or AZT to a TDF-based first-line regimen in a resource-limited setting in rural Lesotho.
Latest World Health Organization guidelines recommend shifting away from Stavudine (d4T)-based regimens due to severe side effects. However, widespread replacement of d4T by Tenofovir (TDF) or Zidovudine (AZT) is hampered by cost concerns. We established the cost-effectiveness of alternative first-line regimens using primary utilization, cost, and outcome data from a program in a rural district in Lesotho. We calculated cost per patient-year, incremental costs, and incremental cost-effectiveness ratios per life year, and per Quality Adjusted Life Year gained. Uncertainty was assessed using multiway and probabilistic sensitivity analyses. Our study included 1260 patients representing 1635 patient-years on antiretroviral therapy (ART). Six hundred eight patients were on TDF, 290 were on AZT, and 362 were on d4T. Patients on d4T experienced more toxicities; toxicities with the biggest impact on quality of life were moderate neuropathy and severe lipodystrophy. The cost per patient-year ranged from US $266 on d4T to US $353 on TDF. Inpatient care and essential drug costs were higher for patients on d4T than on AZT or TDF. Incremental cost-effectiveness ratio results suggest that AZT-based ART is weakly dominated by a combination of d4T- and TDF-based ART. This is one of the first analyses to investigate the cost-effectiveness of TDF using primary data in a resource-poor setting. Although TDF-based first-line ART is more costly than d4T, it is also more effective. Political pressure should be exerted to encourage further price reductions and additional generic manufacturing for TDF and partner drugs such as Efavirenz. This should be met by a commitment from donors and implementers to ensure that supply is met by a clear demand.


Differential response to IV carfentanil in chronic cocaine users and healthy controls.
Chronic cocaine exposure in both rodents and humans increases regional brain mu-opioid receptor (mOR) binding potential, suggesting that cocaine users might have an altered response to mOR agonists. We evaluated the response to IV carfentanil (a selective mOR agonist) in 23 cocaine users [mean (standard deviation) age 33.8 (4.0) years, 83% men] who underwent positron emission tomography (PET) scanning with [C-11]-carfentanil [44.7 (19.5) ng/kg] while housed on a closed research ward and 15 healthy non-drug-using controls [43.9 (14.2) years, 80% men] scanned [49.5 (12.6) ng/kg] as outpatients. Cocaine users had used for 8.7 (4.3) years and on 73 (22)% of days in the two weeks prior to PET scanning. Common adverse effects associated with mOR agonists (nausea, dizziness, headache, vomiting, itchiness) were assessed by self-report (five-point Likert scales) during and for 90 minutes after the scans. Cocaine users were significantly less likely than controls to report any symptom (30.4% versus 60%) and had fewer total symptoms [0.43 (0.73) versus 1.1 (1.0)] during scans, even after statistically controlling for age and carfentanil dose. These differences were also present after the scans and at repeat scans performed after about one week or 12 weeks of monitored cocaine abstinence. In a larger group of cocaine users and separate controls, there was no significant group difference in carfentanil half-life, suggesting that the observed difference was pharmacodynamically, rather than pharmacokinetically, based. These findings suggest that cocaine users are less responsive than healthy controls to mOR agonist adverse effects despite having increased regional brain mOR binding potential.


Neutralization of HIV-1 subtypes: implications for vaccine formulations.
More than 20.8 million people are infected with HIV in sub-Saharan Africa, with South Africa having one of the fastest growing HIV-1 epidemics, where an estimated 2.4 million people were infected. Thirty-two sera from 25 patients were tested for their ability to neutralize HTLV-IIIB (IIIB) and four primary isolates representing subtypes B, C, D, and a recombinant gag C/env B type. A CEM-SS cell line-based assay was used and the neutralizing titer was defined as the reciprocal of the highest dilution giving a 50% reduction in p24 antigen production. All isolates were neutralized better by subtype-specific sera, except for the C4714 strain, which was neutralized by both subtype B and C sera. C4714 was neutralized by 18/25 (72%) sera, IIIB by 19/32 (59%) sera, D482 by 7/31(23%) sera, B3245 by 6/29 (21%) sera, and the recombinant B/C1491 isolate by 4/25 (16%) sera. Five sera were unable to neutralize any of the isolates. The V3 region of the isolates used in the neutralization assay was amplified by PCR, directly sequenced, and analyzed to reveal variability between the consensus HIV-1 sequences and the isolates. HIV-1 strain C4714 was neutralized more effectively with the sera tested than the IIIIB laboratory strain. Variability in the amino acid sequence of the V3 region, which can alter the conformation of the V3 loop secondary structure, can influence the neutralization of a particular viral isolate. Vaccine formulations should be broadened to include multiple subtypes, especially C subtypes, which is rapidly spreading worldwide.


CD4+ T Cells are Exhausted and Show Functional Defects in Chronic Lymphocytic Leukemia.
Chronic lymphocytic leukemia (CLL) is the most common adult leukemia in the western world. This health problem is caused due to the accumulation of mature B-lymphocytes in the peripheral blood and bone marrow. In the course of cancer, CD4+ T cells become "exhausted" and characterized with poor effector functions and the expression of multiple inhibitory receptors. To investigate the frequency and functional properties of exhausted CD4+ T lymphocytes in patients with CLL. Peripheral blood mononuclear cells were obtained from 25 untreated CLL patients and 15 healthy volunteers. CLL patients were clinically classified according to the Rai staging system. The frequency of CD4+/Tim-3+/PD-1+ cells was obtained by flow cytometry. To evaluate cell proliferation and cytokine production, CD4+ T cells were isolated and stimulated with phytohemagglutinin and PMA/ionomycin. Concentrations of IL-2, IFN-γ, TNF-α, and IL-10 were measured in the culture supernatants of stimulated cells by the ELISA technique. The percentage of CD4+/Tim-3+/PD-1+ cells was significantly higher in CLL patients than that of healthy controls. CD4+ T cells from CLL patients showed lower proliferative responses, a lower production of IL-2, IFN-γ, and TNF-α, and a higher production of IL-10, compared to healthy controls. CD4+ T cells from CLL patients in advanced clinical stages showed more exhaustion features than those of early stages. Given that the exhaustion phase of T cells can be reversible, targeted blocking of immune inhibitory molecules could be a promising tool to restore the host immune responses against leukemic cells in CLL.


The Effects of Steroids on Coagulation Dysfunction Induced by Cardiopulmonary Bypass: A Steroids in Cardiac Surgery (SIRS) Trial Substudy.
Cardiopulmonary bypass (CPB) surgery, despite heparin administration, elicits activation of coagulation system resulting in coagulopathy. Anti-inflammatory effects of steroid treatment have been demonstrated, but its effects on coagulation system are unknown. The primary objective of this study is to assess the effects of methylprednisolone on coagulation function by evaluating thrombin generation, fibrinolysis, and platelet activation in high-risk patients undergoing cardiac surgery with CPB. The Steroids In caRdiac Surgery study is a double-blind, randomized, controlled trial performed on 7507 patients worldwide who were randomized to receive either intravenous methylprednisolone, 250 mg at anesthetic induction and 250 mg at initiation of CPB (n = 3755), or placebo (n = 3752). A substudy was conducted in 2 sites to collect blood samples perioperatively to measure prothrombin fragment 1.2 (PF1+2, thrombin generation), plasmin-antiplasmin complex (PAP, fibrinolysis), platelet factor 4 (PF4 platelet activation), and fibrinogen. Eighty-one patients were enrolled in the substudy (37 placebo vs 44 in treatment group). No difference in clinical outcome was detected, including postoperative bleeding and need for blood products transfusion. All patients showed changes of all plasma biomarkers with greater values than baseline in both groups. This reaction was attenuated significantly in the treatment group for PF1.2 (P = 0.040) and PAP (P = 0.042) values at the first intraoperative measurement. No difference between groups was detected for PF4. Methylprednisolone treatment attenuates activation of coagulation system in high-risk patients undergoing CPB surgery. Reduction of thrombin generation and fibrinolysis activation may lead to reduced blood loss after surgery.


How can a work rehabilitation process be improved?--a qualitative study from the perspective of social insurance officers.
The aim of this study is to describe social insurance officers' experiences of a work rehabilitation process at a rehabilitation centre in the northern parts of Sweden. In Sweden the social insurance company has a key role to coordinate all efforts concerning work rehabilitation planning between all rehabilitation actors. Ten social insurance officers at the social insurance company in a city in northern Sweden were interviewed using a narrative approach about their experiences of work rehabilitation processes in general. The interviews were analysed by qualitative content analysis with a focus on their experiences of goals, content and results of a work rehabilitation process. The social insurance officers' experiences of how a work rehabilitation process could be improved were described in six categories; (1) Early identification of problems, needs and interventions (2) clear goal formulations, (3) a focus on psychosocial factors (4) a greater variety of possible interventions, (5) activating employers to a higher extent in work rehabilitation and (6) a closer cooperation and information exchange with other rehabilitation actors. It is possible to improve a work rehabilitation process by focusing on early identification of problems, needs and interventions, with a variety of interventions to choose between and with clear goal formulations and recognizing psychosocial factors in the process. To activate employers to a higher extent in work rehabilitation and to make the information exchange between rehabilitation actors more frequent may also improve work rehabilitation processes.


Elastin-associated microfibrils (10 nm) in a three-dimensional fibroblast culture.
The purpose of this study is to present a three-dimensional dermal fibroblast model. Skin fibroblasts cultured in this system deposit large amounts of collagen and microfibrils. Fibroblasts were seeded onto a nylon filtration mesh and incubated in the presence or absence of ascorbic acid. Collagen fibril formation was found in the presence of ascorbic acid whereas microfibril formation was seen independent of ascorbic acid supplementation. Immunoelectron microscopy revealed that microfibrils were labeled with fibrillin at 67 nm periodicity. Isolated microfibrils studied by rotary shadowing had a beaded appearance consisting of beads linked to each other by a filamentous structure. The spaces between the beads ranged from 10.00-33.33 nm, suggesting that these microfibrils may have an extension-contraction mechanism. Furthermore, the size and spacing of the beads were similar to that seen in microfibrils from tissues (measured after rotary shadowing). Fibroblasts cultured in a three-dimensional mesh represent an effective in vitro model with which to study microfibril formation.


Chronic thromboembolic pulmonary hypertension: emerging endovascular therapy.
Chronic thromboembolic pulmonary hypertension (CTEPH) is a debilitating but potentially reversible complication of chronic pulmonary thromboembolic disease characterized by progressive right heart dysfunction secondary to pulmonary arterial stenosis or occlusion. Balloon pulmonary angioplasty (BPA) has recently emerged as an alternative intervention for non-surgical candidates with CTEPH. Modern reperfusion angioplasty techniques relieve sequela of chronic pulmonary hypertension, ameliorate right ventricular failure, and improve functional status. This article will discuss the diagnosis and treatment of patients with CTEPH and the current state of endovascular management with BPA.


HLA alleles influence the clinical signature of amoxicillin-clavulanate hepatotoxicity.
The genotype-phenotype interaction in drug-induced liver injury (DILI) is a subject of growing interest. Previous studies have linked amoxicillin-clavulanate (AC) hepatotoxicity susceptibility to specific HLA alleles. In this study we aimed to examine potential associations between HLA class I and II alleles and AC DILI with regards to phenotypic characteristics, severity and time to onset in Spanish AC hepatotoxicity cases. High resolution genotyping of HLA loci A, B, C, DRB1 and DQB1 was performed in 75 AC DILI cases and 885 controls. The distributions of class I alleles A*3002 (P/Pc = 2.6E-6/5E-5, OR 6.7) and B*1801 (P/Pc = 0.008/0.22, OR 2.9) were more frequently found in hepatocellular injury cases compared to controls. In addition, the presence of the class II allele combination DRB1*1501-DQB1*0602 (P/Pc = 5.1E-4/0.014, OR 3.0) was significantly increased in cholestatic/mixed cases. The A*3002 and/or B*1801 carriers were found to be younger (54 vs 65 years, P = 0.019) and were more frequently hospitalized than the DRB1*1501-DQB1*0602 carriers. No additional alleles outside those associated with liver injury patterns were found to affect potential severity as measured by Hy's Law criteria. The phenotype frequencies of B*1801 (P/Pc = 0.015/0.42, OR 5.2) and DRB1*0301-DQB1*0201 (P/Pc = 0.0026/0.07, OR 15) were increased in AC DILI cases with delayed onset compared to those corresponding to patients without delayed onset, while the opposite applied to DRB1*1302-DQB1*0604 (P/Pc = 0.005/0.13, OR 0.07). HLA class I and II alleles influence the AC DILI signature with regards to phenotypic expression, latency presentation and severity in Spanish patients.


Short stature and pubertal delay in Duchenne muscular dystrophy.
Children with Duchenne muscular dystrophy (DMD) are shorter than their healthy peers. The introduction of corticosteroid (CS) has beneficial effects on muscle function but slows growth further and is associated with pubertal delay. In contrast to CS usage in most children and adolescents, weaning glucocorticoid is not a key objective of management in DMD. As the outlook for these young people improves, one of the main challenges is to reduce or offset the detrimental effects of CS on growth and development. This is a review of the aetiology and prevalence of short stature and delayed puberty in DMD, a summary of the treatments available and suggestions for areas of further research.


Endocrine disrupting pesticides impair the neuroendocrine regulation of reproductive behaviors and secondary sexual characters of red munia (Amandava amandava).
The exposure effects of two endocrine disrupting pesticides (EDPs), mancozeb/MCZ and imidacloprid/IMI of the group dithiocarbamate and neonicotinoid respectively, on reproductive behaviors and secondary sexual characters have been studied in a seasonally breeding wildlife bird, red munia (Amandava amandava). Adult male birds were exposed to both the pesticides individually (0.25% LD50 of each) as well as co-exposed (MIX-I: 0.25% LD50 of each and MIX-II: 0.5% LD50 of each) through food for 30d in preparatory (July-August) and breeding (September-October) phase of reproductive cycle. Singing and pairing patterns started decreasing from 2nd week to complete disappearance during 4th week of pesticides exposures at both the phases of reproductive cycles. Similar trend was observed in the disappearance of spots on the plumage as well as color of both plumage and beak which turned black/gray from red. Pesticides caused impairment of the lactotropic as well as hypothalamic-pituitary-testicular (HPT) axes as there was increased plasma PRL and decreased LH, FSH and testosterone levels. Testicular expressions of GnRH and androgen receptor/AR were significantly decreased but that of GnIH significantly increased as compared to control. Significant differences among individually- and co-exposed groups were also present. Abnormalities in sexual behaviors and secondary sexual characteristics could be linked to inhibition of HPT axis and/or direct toxicity at the level of hypothalamus, pituitary and testis. In addition, pesticide-induced hyperprolactinemia as well as impaired thyroid hormones might have also affected maintenance of reproductive behaviors. On co-exposures, the more distinct impairments might be due to cumulative toxicity of pesticides.


Sequential Protein Capture in Multiplex Single Molecule Arrays: A Strategy for Eliminating Assay Cross-Reactivity.
Measurements of multiple biomolecules within the same biological sample are important for many clinical applications to enable accurate disease diagnosis or classification. These disease-related biomarkers often exist at very low levels in biological fluids, necessitating ultrasensitive measurement methods. Single-molecule arrays (Simoa), a bead-based digital enzyme-linked immunosorbent assay, is the current state of the art for ultrasensitive protein detection and can detect sub-femtomolar protein concentrations, but its ability to achieve high-order multiplexing without cross-reactivity remains a challenge. Here, a sequential protein capture approach for multiplex Simoa assays is implemented to eliminate cross-reactivity between binding reagents by sequentially capturing each protein analyte and then incubating each capture bead with only its corresponding detection antibody. This strategy not only reduces cross-reactivity to background levels and significantly improves measurement accuracies, but also enables higher-order multiplexing. As a proof of concept, the sequential multiplex Simoa assay is used to measure five different cytokines in plasma samples from Coronavirus Disease 2019 (COVID-19) patients. The ultrasensitive sequential multiplex Simoa assays will enable the simultaneous measurements of multiple low-abundance analytes in a time- and cost-effective manner and will prove especially critical in many cases where sample volumes are limited.


Cohort analysis of child abuse and neglect cases treated during the initial 2 years of a programme to support hospital-based child protection work in Austria.
To describe characteristics of suspected child abuse and neglect (CAN) cases associated with the decision of paediatric departments (PDs) in Vienna, Austria, to involve services of a regional tertiary child protection service programme (Forensische Kinder- und JugendUntersuchungsStelle, FOKUS). Retrospective cohort analysis of a regional data collection of CAN cases over the first 2 years of FOKUS's operational period (1 July 2015-30 June 2017). All CAN cases reported to the PDs of six public hospitals in Vienna. Five of these public hospitals were secondary heath care centres and one was a tertiary healthcare centre. Overall, 231 cases (59.1%) were treated without and 160 (40.9%) with additional involvement of the FOKUS service programme. The odds of a case to be treated without FOKUS involvement were higher if neglect was suspected (OR 3.233, 95% CI 2.024 to 5.279). In contrast, when sexual abuse was suspected, the odds for involvement with FOKUS were significantly higher (OR 7.577, 95% CI 4.580 to 12.879). The odds of being managed with FOKUS services nearly doubled when multiple forms of abuse were suspected (OR 1.926, 95% CI 1.136 to 3.285). The odds for additional FOKUS involvement were significantly lower for patients treated as inpatients (OR 0.239, 95% CI 0.151 to 0.373). CAN patients managed with FOKUS involvement were significantly more often reported to law enforcement (LE) (OR 3.234, 95% CI 2.078 to 5.002). Concurrently, suspected sexual abuse cases and cases reported to LE were more frequently treated in the PD of the tertiary centre than in other PDs (χ2 p<0.001). CAN case characteristics significantly influenced if PDs involved a tertiary child protection programme. Suspected sexual abuse, if more than one form of CAN was suspected and cases reported to LE required additional specialist expertise. For suspected neglect involvement of tertiary services seemed less important.


Paramagnetic ions enable tuning of nuclear relaxation rates and provide long-range structural restraints in solid-state NMR of proteins.
Magic-angle-spinning solid-state nuclear magnetic resonance (SSNMR) studies of natively diamagnetic uniformly (13)C,(15)N-enriched proteins, intentionally modified with side chains containing paramagnetic ions, are presented, with the aim of using the concomitant nuclear paramagnetic relaxation enhancements (PREs) as a source of long-range structural information. The paramagnetic ions are incorporated at selected sites in the protein as EDTA-metal complexes by introducing a solvent-exposed cysteine residue using site-directed mutagenesis, followed by modification with a thiol-specific reagent, N-[S-(2-pyridylthio)cysteaminyl]EDTA-metal. Here, this approach is demonstrated for the K28C and T53C mutants of B1 immunoglobulin-binding domain of protein G (GB1), modified with EDTA-Mn(2+) and EDTA-Cu(2+) side chains. It is shown that incorporation of paramagnetic moieties, exhibiting different relaxation times and spin quantum numbers, facilitates the convenient modulation of longitudinal (R(1)) and transverse (R(2), R(1rho)) relaxation rates of the protein (1)H, (13)C, and (15)N nuclei. Specifically, the EDTA-Mn(2+) side chain generates large distance-dependent transverse relaxation enhancements, analogous to those observed previously in the presence of nitroxide spin labels, while this phenomenon is significantly attenuated for the Cu(2+) center. Both Mn(2+) and Cu(2+) ions cause considerable longitudinal nuclear PREs. The combination of negligible transverse and substantial longitudinal relaxation enhancements obtained with the EDTA-Cu(2+) side chain is especially advantageous, because it enables structural restraints for most sites in the protein to be readily accessed via quantitative, site-resolved measurements of nuclear R(1) rate constants by multidimensional SSNMR methods. This is demonstrated here for backbone amide (15)N nuclei, using methods based on 2D (15)N-(13)C chemical shift correlation spectroscopy. The measured longitudinal PREs are found to be highly correlated with the proximity of the Cu(2+) ion to (15)N spins, with significant effects observed for nuclei up to approximately 20 A away, thereby providing important information about protein structure on length scales that are inaccessible to traditional SSNMR techniques.


Effect of fermented milk containing Lactobacillus casei strain Shirota on constipation-related symptoms and haemorrhoids in women during puerperium.
Constipation and haemorrhoids are common complaints after childbirth. The objective of this pilot study was to evaluate impact of fermented milk containing Lactobacillus casei strain Shirota (LcS) on stool consistency and frequency, constipation-related symptoms and quality of life, and incidence of haemorrhoids in women during puerperium. Forty women who had natural childbirth were randomised to group consuming either one bottle/day of fermented milk containing at least 6.5×109 cfu of LcS, or placebo, for 6 weeks after childbirth. Subjects filled in a diary on their bowel habits including number of bowel movement, stool consistency and incidence of haemorrhoids, and answered questionnaires on constipation-related symptoms (PAC-SYM) and quality of life (PAC-QOL) during the study period. The probiotic group showed the better scores on overall PAC-SYM (P=0.013), PAC-SYM subscales of abdominal symptoms (P=0.043) and rectal symptoms (P=0.031), and PAC-QOL satisfaction subscale (P=0.037) in comparison with the placebo group. In the probiotic group, two to four subjects experienced haemorrhoids during the first 3 weeks of treatment. The number decreased in week 4 and no one had haemorrhoids on most days in week 5-6. In the placebo group, on average four subjects had haemorrhoids from the beginning, and no obvious change was observed until week 6. No statistically significant effect was observed on stool consistency and frequency. The study products did not cause any adverse event in the subjects. Results of this study indicate that continuous consumption of fermented milk containing LcS might alleviate constipation-related symptoms, provide satisfactory bowel habit and result in earlier recovery from haemorrhoids in women during puerperium. Nonetheless, there are several limitations in interpretation of the results attributed to the study design, including lack of baseline data. Further study is required in order to confirm the efficacy.


Chronic Dyrk1 Inhibition Delays the Onset of AD-Like Pathology in 3xTg-AD Mice.
There is a critical need for new treatment approaches that can slow or prevent the progression of Alzheimer's disease (AD). Targets that act simultaneously on multiple relevant pathways could have significant therapeutic potential. Dual-specificity tyrosine phosphorylation-regulated kinase 1A (Dyrk1a) phosphorylates both amyloid precursor protein (APP) and tau. Dyrk1a is upregulated in post-mortem brains of AD patients, and such elevated expression is associated with cognitive deficits. We previously demonstrated that small molecule inhibition of Dyrk1 is well-tolerated and reduces amyloid plaques and pathological forms of tau in 3xTg-AD mice if administered after formation of these pathologies. However, while insoluble forms of hyperphosphorylated tau were reduced by Dyrk1 inhibition, overt neurofibrillary tangle (NFT) pathology remained unchanged. Herein, we specifically test the hypothesis that inhibition of Dyrk1 prior to NFT formation will delay the onset of pathology. 3xTg-AD mice were treated chronically, beginning at 6 months of age, prior to NFT pathology. Mice were dosed daily for either 3 or 6 months and amyloid and tau pathology were assessed. We show that chronic Dyrk1 inhibition reduces insoluble forms of amyloid beta peptides (Aβ) and hyper-phosphorylated tau long-term and that these reductions are associated with dramatic delay in the onset of both amyloid plaques and NFTs. In addition, we show that DYR219, a potent and selective small molecule Dyrk1 inhibitor, induces degradation of Dyrk1a protein, likely contributing to the efficacy of this small molecule approach in vivo. Collectively, these results suggest that therapeutic strategies targeting tau phosphorylation will show the greatest effect if administered very early in the pathogenesis of AD.


Virus specific cell-mediated immunity may play a role in controlling reactivated human herpesvirus 6B in patients under measles induced immunosuppression.
For a better understanding of the cellular immune responses to reactivated HHV 6B the lymphoproliferative response to human herpesvirus 6B (HHV 6B) antigen was measured in three consecutive specimens obtained biweekly from 22 young children and infants suffering from acute measles, and in 19 influenza patients and nine healthy control subjects. HHV 6B DNA in peripheral blood mononuclear cells (PBMCs) was detected in 18 of 22 subjects with measles, but not in the influenza patients or the healthy population. A novel reactivation profile of HHV 6B was found in patients with measles in the milder form of immunosuppression than in patients with organ transplantation. HHV 6B specific lymphoproliferation activities increased correspondingly with reactivation of HHV 6B assessed by detecting HHV 6B DNA in PBMCs in patients with measles, but no significant change in either the antibody response to HHV 6B or DNAemia occurred in serial specimens obtained either from patients with influenza or healthy subjects. This novel form of HHV 6B reactivation without antibody response was observed in patients with measles. The dynamic fluctuations in lymphoproliferative responses in measles may represent the balance between HHV 6B reactivation and its suppression by the host immune system.


Ecklonia Cava Extract Attenuates Endothelial Cell Dysfunction by Modulation of Inflammation and Brown Adipocyte Function in Perivascular Fat Tissue.
It is well known that perivascular fat tissue (PVAT) dysfunction can induce endothelial cell (EC) dysfunction, an event which is related with various cardiovascular diseases. In this study, we evaluated whether Ecklonia cava extract (ECE) and pyrogallol-phloroglucinol-6,6-bieckol (PPB), one component of ECE, could attenuate EC dysfunction by modulating diet-induced PVAT dysfunction mediated by inflammation and ER stress. A high fat diet (HFD) led to an increase in the number and size of white adipocytes in PVAT; PPB and ECE attenuated those increases. Additionally, ECE and PPB attenuated: (i) an increase in the number of M1 macrophages and the expression level of monocyte chemoattractant protein-1 (MCP-1), both of which are related to increases in macrophage infiltration and induction of inflammation in PVAT, and (ii) the expression of pro-inflammatory cytokines (e.g., tumor necrosis factor-α (TNF-α) and interleukin (IL)-6, chemerin) in PVAT which led to vasoconstriction. Furthermore, ECE and PPB: (i) enhanced the expression of adiponectin and IL-10 which had anti-inflammatory and vasodilator effects, (ii) decreased HFD-induced endoplasmic reticulum (ER) stress and (iii) attenuated the ER stress mediated reduction in sirtuin type 1 (Sirt1) and peroxisome proliferator-activated receptor γ (PPARγ) expression. Protective effects against decreased Sirt1 and PPARγ expression led to the restoration of uncoupling protein -1 (UCP-1) expression and the browning process in PVAT. PPB or ECE attenuated endothelial dysfunction by enhancing the pAMPK-PI3K-peNOS pathway and reducing the expression of endothelin-1 (ET-1). In conclusion, PPB and ECE attenuated PVAT dysfunction and subsequent endothelial dysfunction by: (i) decreasing inflammation and ER stress, and (ii) modulating brown adipocyte function.


Risk factors for child mental health problems in Lithuania: The role of parental nationality.
We compare the mental health status of children who reside in Lithuania with parents who are either Lithuanian nationals or non-Lithuanian nationals. Data were drawn from the School Child Mental Health Europe survey (SCMHE), a cross-sectional survey of school children aged 6-11years. A total of 1152 Lithuanian children participated, among them 11.7% from a non-Lithuanian family. Child mental health was assessed using the Dominique Interactive (DI) and the parent- and teacher Strength and Difficulties Questionnaire (SDQ). Parental attitudes were evaluated, and socio-demographics were collected. Overall 26.7% of non-Lithuanian versus 17.2% of Lithuanian children reported having an internalizing disorder (p=0.01) mainly due to separation anxiety (16.4% versus 10.2%, p=0.04). Odds ratio (OR) for child-reported internalizing disorders was 1.86 (95% CI=1.17-2.96) once adjusted for other factors including being a girl, to be younger, parental unemployment and low caring and low autonomy parental attitudes which were associated with greater odds of internalizing disorders. In addition, 31.9% of non-Lithuanian reported suicidal thoughts versus 22.0% of Lithuanian children p=.02); OR=1.60 (95% CI=1.04-2.46) once adjusted for single parent, parental unemployment, parental alcohol problems and overreactivity attitude. Being a non-national minority in Lithuania is a risk factor for child mental health. These findings suggest that further studies are needed to inform local policy-makers on targeted prevention and intervention programs in these children.


A provoking choice--Swedish women's experiences of reactions to their plans to give birth at home.
The home birth rate in Sweden is less than 1 in 1000, and home birth is not included within the health care system. This study describes women's experiences concerning reactions to their decision to give birth at home. A nationwide survey (SHE--Swedish Homebirth Experience) in Sweden was conducted between 1992 and 2005 whereas 735 women had given birth to 1038 children. Of 1038 questionnaires 1025 were returned. In the questionnaires an open-ended question asked women to report their experience of reactions to their decision to give birth at home The question was answered by 594 women, and data were analysed using content analysis. The analysis yielded one overarching theme; "To be faced with fear for life and death" including being exposed to reactions about risks. This describes attitudes of professionals and family towards life and death and suggests perceptions of risk and fear of unexpected events. Four main categories were identified; Seen as an irresponsible person, Met with emotional arguments, Exposed to persuasion and Alienation. Women who plan for a home birth were confronted with negative attitudes and persuasion to make them change their mind. This made them feel alienated, and they searched for support among like-minded. Negative attitudes from health care professionals may erode their confidence in conventional health services and turn them towards other options. Women who want to give birth at home should be given evidence-based information about risks and benefits. Enhanced knowledge among public and professionals about home births would improve the options for respectful encounters.


Axillary recurrence rate in breast cancer patients with negative sentinel lymph node (SLN) or SLN micrometastases: prospective analysis of 150 patients after SLN biopsy.
To assess the axillary recurrence rate in breast cancer patients with negative sentinel lymph node (SLN) or SLN micrometastases (>0.2 mm to <or=2.0 mm) after breast surgery and SLN procedure without formal axillary lymph node dissection (ALND). Under controlled study conditions, the SLN procedure proved to be a reliable method for the evaluation of the axillary nodal status in patients with early-stage invasive breast cancer. Axillary dissection of levels I and II can thus be omitted if the SLN is free of macrometastases. The prognostic value and potential therapeutic consequences of SLN micrometastases, however, remain a matter of great debate. We present the follow-up data of our prospective SLN study, particularly focusing on the axillary recurrence rate in patients with negative SLN and SLN micrometastases. In this prospective study, 236 SLN procedures were performed in 234 patients with early-stage breast cancer between April 1998 and September 2002. The SLN were marked and identified with 99m technetium-labeled colloid and blue dye (Isosulfanblue 1%). The excised SLNs were examined by step sectioning and stained with hematoxylin and eosin and immunohistochemistry (cytokeratin antibodies Lu-5 or CK 22). Only patients with SLN macrometastases received formal ALND of levels I and II, while patients with negative SLN or SLN micrometastases did not undergo further axillary surgery. The SLN identification rate was 95% (224/236). SLN macrometastases were found in 33% (74/224) and micrometastases (>0.2 mm to <or=2 mm) in 12% (27/224) of patients. Adjuvant therapy did not differ between the group of SLN-negative patients and those with SLN micrometastases. After a median follow-up of 42 months (range 12-64 months), 99% (222/224) of evaluable patients were reassessed. While 1 patient with a negative SLN developed axillary recurrence (0.7%, 1/122), all 27 patients with SLN micrometastases were disease-free at the last follow-up control. Axillary recurrences in patients with negative SLN or SLN micrometastases did not occur more frequently after SLN biopsy alone compared with results from the recent literature regarding breast cancer patients undergoing formal ALND. Based on a median follow-up of 42 months-one of the longest so far in the literature-the present investigation does not provide evidence that the presence of SLN micrometastases leads to axillary recurrence or distant disease and supports the theory that formal ALND may be omitted in these patients.


Amelioration of tamoxifen-induced liver injury in rats by grape seed extract, black seed extract and curcumin.
Liver injury was induced in female rats using tamoxifen (TAM). Grape seeds (Vitis vinifera) extract (GSE), black seed (Nigella sativa) extract (NSE), curcumin (CUR) or silymarin (SYL) were orally administered to TAM-intoxicated rats. Liver histopathology of TAM-intoxicated:rats showed pathological changes. TAM-intoxication elicited declines in liver antioxidant enzymes levels (glutathione peroxidase, glutathione reductase, superoxide dismutase and catalase), reduced glutathione (GSH) and GSH/GSSG ratio plus the hepatic elevations in lipid peroxides, oxidized glutathione (GSSG), tumor necrosis factor-alpha (TNF-alpha) and serum liver enzymes; alanine transaminase, aspartate transaminase, alkaline phosphatase, lactate dehydrogenase and gamma glutamyl transferase levels. Oral intake of NSE, GSE, CUR or SYL to TAM-intoxicated rats, attenuated histopathological changes and corrected all parameters mentioned above. Improvements were prominent in case of NSE (similarly SYL) > CUR > GSE. Data indicated that NSE, GSE or CUR act as free radicals scavengers and protect TAM-induced liver injury in rats.


Protective Effect of Hericium erinaceus on Alcohol Induced Hepatotoxicity in Mice.
We investigated the effects of Hericium erinaceus (HEM) on liver injury induced by acute alcohol administration in mice. Mice received ethanol (5 g/kg BW) by gavage every 12 hrs for a total of 3 doses. HEM (200 mg/kg BW) was gavage before ethanol administration. Subsequent serum alanine aminotransferase (ALT) level, aspartate aminotransaminase (AST) level, Maleic dialdehyde (MDA) level, hepatic total antioxidant status (TAOS), and activated nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) were determined by ELISA and immunohistochemistry, respectively. HEM administration markedly (P < 0.05) decreased serum ALT, AST, and MDA levels. The hepatic histopathological observations showed that HEM had a relatively significant role in mice model, which had alcoholic liver damage. In conclusion, we observed that HEM (200 mg/kg BW) supplementation could restrain the hepatic damage caused by acute alcohol exposure.


Development of MILIONCAST, an Improved Model for Predicting Downy Mildew Sporulation on Onions.
The effects of temperature and relative humidity on Peronospora destructor sporulation on onion (Allium cepa) leaves were studied under controlled environmental conditions. Sporangia were produced most rapidly at 8 to 12°C after 5 h of high humidity during dark periods. The greatest number of sporangia was produced at 100% relative humidity (RH), and sporulation decreased to almost nil when humidity decreased to 93% RH. A model, named MILIONCAST (an acronym for MILdew on onION foreCAST), was developed based on the data from these controlled environment studies to predict the rate of sporulation in relation to temperature and relative humidity. The accuracy of prediction of sporulation was evaluated by comparing predictions with observations of sporulation on infected plants in pots outdoors. The accuracy of MILIONCAST was compared with the accuracy of existing models based on DOWNCAST. MILIONCAST gave more correct predictions of sporulation than the DOWNCAST models and a random model. All models based on DOWNCAST were more accurate than the random model when compared on the basis of all predictions (including positive and negative predictions), but they gave fewer correct predictions of sporulation than the random model. De Visser's DOWNCAST and ONIMIL improved their accuracy of prediction of sporulation events when the threshold humidity for sporulation was reduced to 92% RH. The temporal pattern of predicted sporulation by MILIONCAST generally corresponded well to the pattern of sporulation observed on the outdoor potted plants at Wellesbourne, UK.


Benefit of intravenous antibiotic therapy in patients referred for treatment of neurologic Lyme disease.
We have shown previously that extended intravenous antibiotic therapy is associated with low morbidity and no mortality in patients referred for treatment of neurologic Lyme disease. In this study, we evaluated the benefit of extended intravenous antibiotic therapy in patients with symptoms of neurologic Lyme disease. Patients with significant neurologic symptoms and positive testing for Borrelia burgdorferi were treated with intravenous antibiotics, and biweekly evaluation of symptom severity was performed using a six-level ordinal scale. Four symptoms were selected a priori as primary outcome measures in the study, ie, fatigue, cognition, myalgias, and arthralgias. Patients were placed into five groups according to time on treatment (1-4, 5-8, 9-12, 13-24, and 25-52 weeks), and changes in the primary symptoms as a function of time on treatment were analyzed using a mixed-effects proportional odds model. Among 158 patients with more than one follow-up visit who were monitored for up to 1 year, there were on average 6.7 visits per person (median 5, range 2-24). The last follow-up day was on average 96 days after enrollment (median 69, range 7-354 days), corresponding to the length of antibiotic therapy. Each primary symptom was significantly improved at one or more time points during the study. For cognition, fatigue, and myalgias, the greatest improvement occurred in patients on the longest courses of treatment (25-52 weeks) with odds ratios (OR) for improvement of 1.97 (P = 0.02), 2.22 (P < 0.01), and 2.08 (P = 0.01), respectively. In contrast, arthralgias were only significantly improved during the initial 1-4 weeks of therapy (OR: 1.57, P = 0.04), and the beneficial effect of longer treatment did not reach statistical significance for this symptom. Prolonged intravenous antibiotic therapy is associated with improved cognition, fatigue, and myalgias in patients referred for treatment of neurologic Lyme disease. Treatment for 25-52 weeks may be necessary to obtain symptomatic improvement in these patients.


Unrecognised joint penetration in treatment of femoral neck fractures.
Screw devices used to treat fractures of the femoral neck are usually positioned under image-intensifier control, using anteroposterior and lateral views. The volume projected by these views is over 27% larger than the femoral head; the tips of screws so placed may be outside the femoral head. This can be avoided by placement within the central two-thirds of the head: we have designed a template which can confirm safe placement.


Biological sources and sinks of nitrous oxide and strategies to mitigate emissions.
Nitrous oxide (N(2)O) is a powerful atmospheric greenhouse gas and cause of ozone layer depletion. Global emissions continue to rise. More than two-thirds of these emissions arise from bacterial and fungal denitrification and nitrification processes in soils, largely as a result of the application of nitrogenous fertilizers. This article summarizes the outcomes of an interdisciplinary meeting, 'Nitrous oxide (N(2)O) the forgotten greenhouse gas', held at the Kavli Royal Society International Centre, from 23 to 24 May 2011. It provides an introduction and background to the nature of the problem, and summarizes the conclusions reached regarding the biological sources and sinks of N(2)O in oceans, soils and wastewaters, and discusses the genetic regulation and molecular details of the enzymes responsible. Techniques for providing global and local N(2)O budgets are discussed. The findings of the meeting are drawn together in a review of strategies for mitigating N(2)O emissions, under three headings, namely: (i) managing soil chemistry and microbiology, (ii) engineering crop plants to fix nitrogen, and (iii) sustainable agricultural intensification.


Obesity and metabolic surgery in type 1 diabetes mellitus.
Obesity surgery is an effective method for treating obesity and diabetes mellitus type 2. This type of diabetes can be completely resolved in 78.1% of diabetic patients and can be improved or resolved in 86.6% of diabetic patients. But little is known about bariatric surgery in type 1 diabetes mellitus. We report of 6 female obese patients with diabetes mellitus type 1 who had bariatric surgery. Two of them underwent Roux-en Y gastric bypass (RNYGB), one of them had sleeve gastrectomy and the remaining three had biliopancreatic diversion with duodenal-switch (BPD-DS). Our results showed a remarkable weight reduction as well as an improvement in their blood glucose control and the insulin requirement in the followup years after surgery. Pre-surgery the BMI of our 6 patients ranged between 37.3-46.0 kg/m2 and improved to 25.8-29.0 kg/m2 one year after surgery. HbA1c decreased from 6.7-9.8% pre-surgery to 5.7-8.5% after one year post-surgery. The total amount of daily insulin requirement was reduced from 62-150 IU/day pre-surgery to 15- 54 IU/day after one year. The results are impressive and show an improvement in insulin sensitivity following obesity surgery. However, an optimal blood glucose control still remains very important in the therapy of diabetes mellitus type 1 to avoid long-term-complications.


Matrix metalloproteinase 9 plays a key role in lyme arthritis but not in dissemination of Borrelia burgdorferi.
Borrelia burgdorferi, the causative agent of Lyme arthritis, does not produce any exported proteases capable of degrading extracellular matrix despite the fact that it is able to disseminate from a skin insertion site to infect multiple organs. Prior studies have shown that B. burgdorferi induces the host protease, matrix metalloproteinase 9 (MMP-9), and suggested that the induction of MMP-9 may allow the organism to disseminate and produce local tissue destruction. We examined the role of MMP-9 in dissemination of B. burgdorferi and pathogenesis of Lyme arthritis. In a MMP-9(-/-) mouse model, MMP-9 was not required for the dissemination of the spirochete to distant sites. However, MMP-9(-/-) exhibited significantly decreased arthritis compared to wild-type mice. The decrease in arthritis was not due to an inability to control infection since the spirochete numbers in the joints were identical. Levels of inflammatory chemokines and cytokines were also similar in MMP-9(-/-) and wild-type mice. We examined whether decreased inflammation in MMP-9(-/-) mice may be the result of decreased production of neoattractants by MMP-9-dependent cleavage of collagen. MMP-9 cleavage of type I collagen results in increased monocyte chemoattraction. MMP-9 plays an important role in regulating inflammation in Lyme arthritis, potentially through the cleavage of type I collagen.


Assessment of family functioning: evaluation of the General Functioning Scale in a Swedish Bariatric Sample.
The General Functioning Scale (GFS) was developed to assess self-perceived overall family functioning. The scale has satisfactory psychometric properties, is internationally recognised and has been used in different contexts. However, no validated Swedish version is available. Healthy family functioning can support patients and help them adhere to treatment regimens. Moreover, it maintains the physical and emotional health and that of the family as a unit. Yet, there is limited information regarding family functioning postgastric bypass surgery. Thus, it is important to use validated instruments to understand family functioning in bariatric contexts. To evaluate aspects of reliability and validity in GFS in a Swedish bariatric sample, focusing on factor structure. The Swedish version of the GFS (S-GFS) was administered on two occasions to 163 participants who had undergone gastric bypass surgery 6-8 weeks prior to testing. Internal consistency, temporal stability and construct validity were assessed. Data were positively skewed. The S-GFS showed good internal consistency (ordinal α = 0.92) with a sufficient overall mean interitem correlation (0.500) and adequate temporal stability (intraclass correlation coefficient = 0.833). After modifying response alternatives, confirmatory factor analysis indicated acceptable fit for a one-factor model. The scale is a promising tool for assessing family functioning in bariatric settings. The S-GFS showed satisfactory reliability - consistent with prior research - and acceptable validity in the study sample. This study contributes to the limited research on the scale's validity. However, the S-GFS needs to be evaluated in different cultural and clinical contexts, focusing on various aspects of validity and responsiveness (sensitivity to detect significant change over time) in different samples.


GABAergic/glutamatergic imbalance relative to excessive neuroinflammation in autism spectrum disorders.
Autism spectrum disorder (ASD) is characterized by three core behavioral domains: social deficits, impaired communication, and repetitive behaviors. Glutamatergic/GABAergic imbalance has been found in various preclinical models of ASD. Additionally, autoimmunity immune dysfunction, and neuroinflammation are also considered as etiological mechanisms of this disorder. This study aimed to elucidate the relationship between glutamatergic/ GABAergic imbalance and neuroinflammation as two recently-discovered autism-related etiological mechanisms. Twenty autistic patients aged 3 to 15 years and 19 age- and gender-matched healthy controls were included in this study. The plasma levels of glutamate, GABA and glutamate/GABA ratio as markers of excitotoxicity together with TNF-α, IL-6, IFN-γ and IFI16 as markers of neuroinflammation were determined in both groups. Autistic patients exhibited glutamate excitotoxicity based on a much higher glutamate concentration in the autistic patients than in the control subjects. Unexpectedly higher GABA and lower glutamate/GABA levels were recorded in autistic patients compared to control subjects. TNF-α and IL-6 were significantly lower, whereas IFN-γ and IFI16 were remarkably higher in the autistic patients than in the control subjects. Multiple regression analysis revealed associations between reduced GABA level, neuroinflammation and glutamate excitotoxicity. This study indicates that autism is a developmental synaptic disorder showing imbalance in GABAergic and glutamatergic synapses as a consequence of neuroinflammation.


The living conditions of elderly Americans.
This article profiles the housing settings of frail elderly individuals, whether their homes are facilitating or impeding their ability to live in the community, and the change in disability and housing status before and after passage of the 1990 Americans with Disabilities Act. The analysis relies primarily on statistical analysis of the 1995 national American Housing Survey (AHS), with supplementary analysis of the 1978 AHS. In 1995, roughly 14% of elderly individuals had a "housing-related disability," 49% had at least one dwelling modification, and 23% had an unmet need for modifications. Because half those with dwelling modification also reported unmet need, the match between disabling condition and modification, not the presence of modifications, is key. Multivariate results indicate that although unmet need is greater among the poor, lack of modifications is not. Prevalence of modifications nearly doubled between 1978 and 1995. Overall unmet need declined, but some needs were less likely to be met in 1995 than 1978. The analysis highlights the importance of information about housing for understanding the care and service needs of elderly individuals and provides a compelling argument for a minimum dataset on their housing and neighborhood environments.


Cyclooxygenase-2 induction and transforming growth factor beta growth inhibition in rat intestinal epithelial cells.
Rat intestinal epithelial cells (RIE-1) permanently transfected with the prostaglandin endoperoxide synthase 2 (also referred to as cyclooxygenase-2; COX-2) gene exhibit decreased cyclin D1 levels, decreased cdk4-associated kinase activity, and delayed G1 cell cycle progression, which represents a phenotype similar to that which follows transforming growth factor beta (TGF-beta) treatment. In the current study, we have found that addition of TGF-beta 1 to the parental RIE-1 cells (designated RIE-P) caused a rapid induction of COX-2 mRNA and protein. COX-2 protein levels progressively increased and reached peak levels 6 h after TGF-beta 1 addition. Cyclin D1 was decreased by 74% at 6 h and was undetectable 24 h after addition of TGF-beta 1. In RIE cells transfected with the COX-2 antisense expression vector (RIE-AS cells), TGF-beta 1 induction of COX-2 protein was reduced greater than 90%. Addition of TGF-beta 1 did not reduce the abundant cyclin D1 protein expression in the RIE-AS cells, unlike the effect in RIE-P cells. TGF-beta 1 treatment reduced peak [3H]thymidine incorporation by 60% and delayed G1/S-phase transition by at least 4 h in the RIE-P cells. In contrast, S-phase entry occurred at 16 h in RIE-AS cells and was not altered by TGF-beta 1 treatment. Restoration of cyclin D1 expression by transfection of the cyclin D1 cDNA under transcriptional control of the cytomegalovirus promoter/enhancer in the COX-2-overexpressing (RIE-S) cells decreased the time required for S-phase entry by at least 4 h and increased the peak level of [3H]thymidine incorporation. Taken together, the results demonstrate that TGF-beta 1 strongly induces COX-2 at both the mRNA and protein levels and suggest that this induction of COX-2 is involved in the down-regulation of cyclin D1 and inhibition of cell growth caused by TGF-beta 1 in rat intestinal epithelial cells.


Challenges in modelling the reaction chemistry of interstellar dust.
Studies aiming to understand the physicochemical properties of interstellar dust and the chemical reactions that occur on and in it have traditionally been the preserve of astronomical observation and experimental attempts to mimic astronomically relevant conditions in the laboratory. Increasingly, computational modelling in its various guises is establishing a complementary third pillar of support to this endeavour by providing detailed insights into the complexities of interstellar dust chemistry. Inherently, the basis of computational modelling is to be found in the details (e.g. atomic structure/composition, reaction barriers) that are difficult to probe accurately from observation and experiment. This bottom-up atom-based theoretical approach, often itself based on deeper quantum mechanical principles, although extremely powerful, also has limitations when systems become too large or complex. In this Perspective, after first providing a general background to the current state of observational-based knowledge, we introduce a number of computational modelling methods with reference to recent state-of-the-art studies, in order to highlight the capabilities of such approaches in this field. Specifically, we first outline the use of computational chemistry methods for dust nucleation, structure, and individual reactions on bare and icy dust surfaces. Later, we review kinetic modelling of networks of reactions relevant to dust chemistry and how to take into account quantum tunnelling effects in the low temperature reactions in the interstellar medium. Finally, we point to the future challenges that need to be overcome for computational modelling to provide even more detailed and encompassing perspectives on the nature and reaction chemistry of interstellar dust.


The incidence of JAK2 V617F mutation in bcr/abl-negative chronic myeloproliferative disorders: assessment by two different detection methods.
A recurrent specific JAK2 V617F mutation has been reported in bcr/abl-negative chronic myeloproliferative diseases (cMPD), including polycythemia vera (PV), essential thrombocythemia (ET) and idiopathic myelofibrosis (IMF). The mutation is detectable in a variable proportion of neoplastic clones, depending on the molecular methods employed. In this study, we attempted to establish the JAK2 V617F mutation frequency in two partially overlapping cMPD patient series by two different PCR-based techniques. Using an allele-specific polymerase chain reaction assay (AS-PCR), the JAK2 V617F mutation was detected in 124/158 (78.5%) cMPD patients; in particular, 90.2, 72.1, 63.2 and 50% of PV, ET, IMF and unclassified (U)-MPD cases, respectively, showed the mutation. Employing a semiquantitative 5' fluorogenic TaqMan assay, the JAK2 V617F mutation was identified in a much larger percentage of cMPDs patients (118/127: 92.9%). Rates of mutation in PV, ET, IMF and U-MPD cases were 95.9, 85.7, 91.7 and 92.9%, respectively. Furthermore, a statistically higher percentage of JAK2 mutated alleles was detected by TaqMan assay in PV (68+/-3.5, mean value+/-SEM) and IMF (64+/-9.3) cases as compared with ET (35+/-5.4). Finally, a significant correlation between JAK2 V617F mutational status and hematocrit (Ht), white blood cell and platelet counts in PV patients, and Ht values in ET cases, was observed by AS-PCR. Overall, these data indicate that TaqMan technology significantly improved sensitivity in detecting the JAK2 mutation in cMPD patients and may be worth of further evaluations as a clinically useful tool for detection of small amounts of mutated clones.


Oxidation of gamma II-crystallin solutions yields dimers with a high phase separation temperature.
Aqueous solutions of the bovine eye lens protein gamma II (or gamma B)-crystallin at neutral pH show a gradual increase in phase separation temperature, Tph, when allowed to stand for several weeks at room temperature without reducing agents. In a typical experiment, the Tph of the protein solution (218 mg/ml) increases from 2.5 +/- 1 degree C to 32.5 +/- 1 degree C after 21 days, and a new protein species, gamma IIH, is formed. The Tph of pure gamma IIH is at least 40 degrees C higher than that of pure gamma II. The average apparent hydrodynamic radius is 36 A for gamma IIH compared to 26 A for gamma II. The molecular mass of gamma IIH is approximately 41.5 kDa compared to 20 kDa for native gamma II. Therefore, gamma IIH is probably a dimer of gamma II crystallin. gamma IIH has a lower thiol content than gamma II and is not formed in the presence of dithiothreitol. We conclude that gamma IIH is a thiol oxidation product of gamma II-crystallin and is a dimer containing an intermolecular disulfide crosslink. Thus, some oxidative modifications of protein thiol groups lead to an increase in net attractive interactions between proteins. As a result, Tph increases and protein aggregates are formed. These two microscopic changes produce the increased light scattering associated with lens opacification.


Risk factor analysis for long-term unfavorable ocular outcomes in children treated for retinopathy of prematurity.
The aim in this study was to report long-term ocular outcomes of neonates treated for retinopathy of prematurity (ROP) and potential risk factors for unfavorable ocular outcomes. The study consisted of neonates treated for ROP between March 1999 and November 2009. Data relating baseline characteristics and late structural, functional and refractive ocular outcomes were recorded. The association between the unfavorable ocular outcomes and ROP-related risk factors was evaluated by regression analysis. Forty-eight children were included for assessment. Average chronological age at the time of followup was 3.11±0.73 years. The rates of unfavorable structural and functional outcomes were 12% and 15.3%, respectively. Ocular deviation was common (27.1%), and mostly esotropic (12/13). A clear myopic tendency was observed (51.2%), and the mean spherical equivalent per eye was -0.72±2.9 diopters. Regression analyses for unfavorable ocular outcomes revealed intraventricular hemorrhage as a core independent risk factor. In conclusion, ROP treatment has shown promising results in both structure and function. Because of the high risk of developing an unfavorable outcome, a more intense follow-up is required in neonates with a history of intraventricular hemorrhage in the neonatal period. Further studies from other centers are needed to develop a national database, which may validate this observation.


A History of Dystonia: Ancient to Modern.
Before 1911, when Hermann Oppenheim introduced the term dystonia, this movement disorder lacked a unifying descriptor. While words like epilepsy, apoplexy, and palsy have had their meanings since antiquity, references to dystonia are much harder to identify in historical documents. Torticollis is an exception, although there is difficulty distinguishing dystonic torticollis from congenital muscular torticollis. There are, nevertheless, possible representations of dystonia in literature and visual art from the pre-modern world. Eighteenth century systematic nosologists such as Linnaeus, de Sauvages, and Cullen had attempted to classify some spasmodic conditions, including torticollis. But only after Charcot's contributions to clinical neuroscience were the various forms of generalized and focal dystonia clearly delineated. They were categorized as névroses: Charcot's term for conditions without an identifiable neuroanatomical cause. For a time thereafter, psychoanalytic models of dystonia based on Freud's ideas about unconscious conflicts transduced into physical symptoms were ascendant, although there was always a dissenting "organic" school. With the rise of subspecialization in movement disorders during the 1970s, the pendulum swung strongly back toward organic causation. David Marsden's clinical and electrophysiological research on the adult-onset focal dystonias was particularly important in establishing a physical basis for these disorders. We are still in a period of "living history" of dystonia, with much yet to be understood about pathophysiology. Rigidly dualistic models have crumbled in the face of evidence of electrophysiological and psychopathological overlap between organic and functional dystonia. More flexible biopsychosocial frameworks may address the demand for new diagnostic and therapeutic rationales.


Mitochondrial DNA of ancient Cumanians: culturally Asian steppe nomadic immigrants with substantially more western Eurasian mitochondrial DNA lineages.
The Cumanians were originally Asian pastoral nomads who in the 13th century migrated to Hungary. We have examined mitochondrial DNA from members of the earliest Cumanian population in Hungary from two archeologically well-documented excavations and from 74 modern Hungarians from different rural locations in Hungary. Haplogroups were defined based on HVS I sequences and examinations of haplogroup-associated polymorphic sites of the protein coding region and of HVS II. To exclude contamination, some ancient DNA samples were cloned. A database was created from previously published mtDNA HVS I sequences (representing 2,615 individuals from different Asian and European populations) and 74 modem Hungarian sequences from the present study. This database was used to determine the relationships between the ancient Cumanians, modern Hungarians, and Eurasian populations and to estimate the genetic distances between these populations. We attempted to deduce the genetic trace of the migration of Cumanians. This study is the first ancient DNA characterization of an eastern pastoral nomad population that migrated into Europe. The results indicate that, while still possessing a Central Asian steppe culture, the Cumanians received a large admixture of maternal genes from more westerly populations before arriving in Hungary. A similar dilution of genetic, but not cultural, factors may have accompanied the settlement of other Asian nomads in Europe.


Pediatric Acute Lymphoblastic Leukemia, Version 2.2020, NCCN Clinical Practice Guidelines in Oncology.
Acute lymphoblastic leukemia (ALL) is the most common pediatric malignancy. Advancements in technology that enhance our understanding of the biology of the disease, risk-adapted therapy, and enhanced supportive care have contributed to improved survival rates. However, additional clinical management is needed to improve outcomes for patients classified as high risk at presentation (eg, T-ALL, infant ALL) and who experience relapse. The NCCN Clinical Practice Guidelines in Oncology (NCCN Guidelines) for pediatric ALL provide recommendations on the workup, diagnostic evaluation, and treatment of the disease, including guidance on supportive care, hematopoietic stem cell transplantation, and pharmacogenomics. This portion of the NCCN Guidelines focuses on the frontline and relapsed/refractory management of pediatric ALL.


Pediatrician attitudes, clinical activities, and knowledge of environmental health in Wisconsin.
Pediatricians can reduce exposures to environmental hazards but most have little training in environmental health. To assess whether Wisconsin pediatricians perceive a relative lack of self-efficacy for common environmental exposures and diseases of environmental origin, we assessed their attitudes and beliefs about the role of the environment in children's health. A 4-page survey was sent to the membership of the Wisconsin Chapter of the American Academy of Pediatrics. We obtained a 35.4% response rate after 1 follow-up mailing. Respondents agreed that the role of the environment in children's health is significant (mean 4.28 +/- .78 on 1-5 Likert scale). They expressed high confidence in dealing with lead exposure (means 4.22-4.27 +/- 1.01-1.09), but confidence in their skills for pesticide, mercury, and mold was much lower (means 2.49-3.09 +/- 1.06-1.26; P<.001). Of those surveyed, 88.6% would refer patients to a clinic "where pediatricians could refer patients for clinical evaluation and treatment of their environmental health concerns." These findings indicate that Wisconsin pediatricians agree that children are suffering preventable illnesses of environmental origin, but feel ill equipped to educate families about many common exposures. Significant demand exists for centers that can evaluate environmental health concerns, as well as for educational opportunities.


Fully Convolutional Neural Networks to Detect Clinical Dermoscopic Features.
The presence of certain clinical dermoscopic features within a skin lesion may indicate melanoma, and automatically detecting these features may lead to more quantitative and reproducible diagnoses. We reformulate the task of classifying clinical dermoscopic features within superpixels as a segmentation problem, and propose a fully convolutional neural network to detect clinical dermoscopic features from dermoscopy skin lesion images. Our neural network architecture uses interpolated feature maps from several intermediate network layers, and addresses imbalanced labels by minimizing a negative multilabel Dice-F 1 score, where the score is computed across the minibatch for each label. Our approach ranked first place in the 2017 ISIC-ISBI Part 2: Dermoscopic Feature Classification Task, challenge over both the provided validation and test datasets, achieving a 0.895% area under the receiver operator characteristic curve score. We show how simple baseline models can outrank state-of-the-art approaches when using the official metrics of the challenge, and propose to use a fuzzy Jaccard Index that ignores the empty set (i.e., masks devoid of positive pixels) when ranking models. Our results suggest that the classification of clinical dermoscopic features can be effectively approached as a segmentation problem, and the current metrics used to rank models may not well capture the efficacy of the model. We plan to make our trained model and code publicly available.