Idiopathic hypercalciuria: association with isolated hematuria and risk for urolithiasis in children. The Southwest Pediatric Nephrology Study Group.
A prospective multicenter study was designed to determine the frequency and prognostic importance of hypercalciuria in children with hematuria. Urinary calcium excretion was examined in 215 patients with unexplained isolated hematuria (no proteinuria, urolithiasis, infection or systemic disorder). Hypercalciuria (urinary calcium excretion greater than 4 mg/kg/day) was identified in 76 patients (35%). Compared to patients with normal urinary calcium excretion, children with hematuria and hypercalciuria were characterized by male preponderance, white race, family history of urolithiasis, gross hematuria and calcium oxalate crystals. Renal biopsies were performed in 10 patients with urinary calcium excretion 0.4 to 2.5 mg/kg/day; three had IgA glomerulonephritis, three had glomerular basement membrane thinning, one had proliferative glomerulonephritis and three were normal. Renal biopsies in three patients with hypercalciuria showed focal segmental glomerulosclerosis, hereditary nephritis or no abnormalities. Oral calcium loading tests showed renal hypercalciuria in 26 patients, absorptive hypercalciuria in 15 patients and were not diagnostic in 35 patients. Serum parathyroid hormone, bicarbonate and phosphorus and urinary cyclic adenosine monophosphate concentrations were similar in the three groups of hypercalciuric patients. Urinary calcium excretion after one week of dietary calcium restriction was higher (5.8 mg/kg/day) in renal hypercalciuria than in other hypercalciuric patients (3.4 mg/kg/day), P less than 0.01. One to four years follow-up was available for 184 patients. Eight of 60 hypercalciuric patients developed urolithiasis or renal colic compared to 2 of 124 patients with normal urinary calcium excretion (P less than 0.001). Hypercalciuria is commonly associated with isolated hematuria and represents a risk factor for future urolithiasis in children with hematuria.(ABSTRACT TRUNCATED AT 250 WORDS)


Phase-resolved acoustic radiation force optical coherence elastography.
Many diseases involve changes in the biomechanical properties of tissue, and there is a close correlation between tissue elasticity and pathology. We report on the development of a phase-resolved acoustic radiation force optical coherence elastography method (ARF-OCE) to evaluate the elastic properties of tissue. This method utilizes chirped acoustic radiation force to produce excitation along the sample's axial direction, and it uses phase-resolved optical coherence tomography (OCT) to measure the vibration of the sample. Under 500-Hz square wave modulated ARF signal excitation, phase change maps of tissue mimicking phantoms are generated by the ARF-OCE method, and the resulting Young's modulus ratio is correlated with a standard compression test. The results verify that this technique could efficiently measure sample elastic properties accurately and quantitatively. Furthermore, a three-dimensional ARF-OCE image of the human atherosclerotic coronary artery is obtained. The result indicates that our dynamic phase-resolved ARF-OCE method can delineate tissues with different mechanical properties.


Exercise-induced refractoriness in asthmatic subjects involves leukotriene and prostaglandin interdependent mechanisms.
Exercise-induced bronchoconstriction is caused, in part, by leukotriene (LT)D4 release in asthmatic airways. Asthmatics become refractory to exercise bronchoconstriction with repeated challenges, due to inhibitory prostaglandin release. The purpose of this study was to test the hypothesis that exercise refractoriness is caused by LTD4-induced inhibitory prostaglandin release. Fourteen stable asthmatic subjects with exercise-induced bronchoconstriction were studied. On the first 2 days, subjects underwent two challenges, 1 h apart, with either exercise or inhaled LTD4. Eight subjects then took part in three double-blind, randomized, placebo-controlled, crossover studies with flurbiprofen, a prostaglandin synthetase inhibitor, to determine whether cross refractoriness occurs between exercise and LTD4, whether flurbiprofen attenuates this effect, and whether flurbiprofen attenuates LTD4 tachyphylaxis. There was a reduction in the intensity of bronchoconstriction to the second challenge both with exercise (refractoriness) and with LTD4 (tachyphylaxis). The degrees of refractoriness and tachyphylaxis were correlated (r = 0.72, p = 0.005). Flurbiprofen attenuated LTD4 tachyphylaxis. Cross refractoriness occurred between exercise and LTD4, and flurbiprofen treatment also attenuated this effect. One hour after LTD4 challenge, the mean fall in FEV1 after exercise was 12.3% (%SEM 2.3) on placebo and 17.1% (%SEM 3.8) on flurbiprofen (p = 0.027). Similarly, 1 h after exercise, the LTD4 PC20 increased to 0.73 (%SEM 1.4) microgram/ml on placebo and 0.30 (%SEM 1.8) microgram/ml on flurbiprofen (p = 0.026). These results suggest that LTD4 released in asthmatic airways as a result of exercise stimulates inhibitory prostaglandin release, resulting in exercise refractoriness.


The DMRT3 'Gait keeper' mutation affects performance of Nordic and Standardbred trotters.
In a previous study it was shown that a nonsense mutation in the DMRT3 gene alters the pattern of locomotion in horses and that this mutation has a strong positive impact on trotting performance of Standardbreds. One aim of this study was to test if racing performance and trotting technique in the Nordic (Coldblood) trotters are also influenced by the DMRT3 genotype. Another aim was to further investigate the effect of the mutation on performance in Standardbreds, by using a within-family analysis and genotype-phenotype correlations in a larger horse material than in the previous study. We genotyped 427 Nordic trotters and 621 Standardbreds for the DMRT3 nonsense mutation and a SNP in strong linkage disequilibrium with it. In Nordic trotters, we show that horses homozygous for the DMRT3 mutation (A) had significantly higher EBV for trotting performance traits than heterozygous (CA) or homozygous wild-type (CC) horses (P = 0.001). Furthermore, AA homozygotes had a higher proportion of victories and top 3 placings than horses heterozygous or homozygous wild-type, when analyzing performance data for the period 3 to 6 yr of age (P = 0.06 and P = 0.05, respectively). Another finding in the Nordic trotters was that the DMRT3 mutation influenced trotting technique (P = 2.1 × 10(-8)). Standardbred horses homozygous AA had significantly higher EBV for all traits than horses with at least 1 wild-type allele (CA and CC; P = 1.6 × 10(-16)). In a within-family analysis of Standardbreds, we found significant differences in several traits (e.g., earnings, P = 0.002; number of entered races, P = 0.004; and fraction of offspring that entered races, P = 0.002) among paternal half-sibs with genotype AA or CA sired by a CA stallion. For most traits, we found significant differences at young ages. For Nordic trotters, most of the results were significant at 3 yr of age but not for the older ages, and for the Standardbreds most of the results for the ages 3 to 5 were significant. For Nordic trotters, the proportion of victories and placings were the only traits that were significant for other ages than 3 yr.


RS-1 Gene Delivery to an Adult Rs1h Knockout Mouse Model Restores ERG b-Wave with Reversal of the Electronegative Waveform of X-Linked Retinoschisis.
To create and evaluate a mouse model of human X-linked juvenile retinoschisis (XLRS) and then investigate whether supplementing with the retinoschisin protein by gene delivery can reverse the abnormal "electronegative" electroretinogram (ERG) retinal response. An X-linked retinoschisis mouse (Rs1h-KO) model was created by substituting a neomycin resistance cassette for exon 1 and 1.6 kb of intron 1 of Rs1h, the murine orthologue of the human RS-1 gene. RS protein was evaluated by immunohistochemistry and Western blot analysis with a polyclonal RS N-terminus antibody. Retinal function was evaluated by conventional, full-field flash ERG recordings. RS protein supplementation therapy was evaluated by gene transfer with an AAV(2/2)-CMV-Rs1h vector containing C57BL/6J Rs1h cDNA under the regulation of a CMV promoter, and ERG functional analysis was performed. No RS protein was detected by Western blot analysis or immunohistochemistry in the Rs1h-KO mouse. Dark-adapted ERG responses showed an electronegative configuration, with b-wave reduction in both Rs1h(-/Y) and Rs1h-/- mice, typical of XLRS in humans. Histologic examination of Rs1h-KO mice showed disorganization of multiple retinal layers, including duplication and mislocalization of ganglion cells, laminar dissection through the inner plexiform layer, disorganization of the outer plexiform layer, loss of regularity of the outer nuclear layer, and shortening of the inner/outer segments with mislocalization of photoreceptor nuclei into this layer. After intraocular administration of AAV(2/2)-CMV-Rs1h, immunohistochemistry showed retinoschisin expression in all retinal layers of Rs1h(-/Y) mice, and ERG recordings showed reversal of the electronegative waveform and restoration of the normal positive b-wave. The RS-KO mouse mimics structural features of human X-linked juvenile retinoschisis with dissection through, and disorganization of, multiple retinal layers. The Rs1h-KO functional deficit results in an electronegative ERG waveform that is characteristic of human retinoschisis disease and that implicates a synaptic transmission deficit in the absence of retinoschisin protein. Replacement therapy by supplementing normal Rs1h protein in the adult Rs1h-KO mouse restored the normal ERG configuration. This indicates that gene therapy is a viable strategy of therapeutic intervention even in the postdevelopmental adult stage of XLRS disease.


[Increasing prevalence of fosfomycin resistance in extended-spectrum-beta-lactamase-producing Escherichia coli urinary isolates (2005-2009-2011)].
Escherichia coli is the most important uropathogen. The appearance of extended- spectrum beta-lactamase (ESBL)-producing E.coli in urinary tract infections (UTI) constitutes an important therapeutic challenge that requires the study of its evolution throughout time in order to establish a suitable empirical treatment. Our aim was to determine the prevalence of ESBL-producing E. coli urinary isolates in 2005, 2009 and 2011. We also determined the antimicrobial coresistance to several agents, including fosfomycin. We analyzed 5053, 6324 and 6644 E. coli isolates obtained from urine cultures in 2005, 2009 and 2011 respectively. Duplicate isolates were excluded. Antimicrobial susceptibility was determined by the Wider microdilution system (Soria Melguizo S.A.) and the phenotypic pattern of resistance that indicated a BLEE-producing E.coli was selected (CLSI 2009). 3.9% of strains (198) were ESBL producers in 2005, 7.3% (463) in 2009 and 8.7% (584) in 2011. Resistance to carbapenems was detected in 2009, they inhibited more than the 95% of strains in 2011. Among the non-beta-lactams, colistin was the most active antibiotic followed by nitrofurantoin. Ciprofloxacin and sulfamethoxazole-trimethoprim were not effective with 80% and 60% resistant isolates, respectively. An increasing resistance trend, from 0% to 9.3% in 2009 and 14.4% in 2011 was observed for fosfomycin. From 2005 our institution had an increasing prevalence of ESBL-producing E. coli rising to 8.7% in 2011. Carbapenems are still the most active agents. The increase of resistance was significant for fosfomycin.


Wastewater sequencing reveals early cryptic SARS-CoV-2 variant transmission.
As SARS-CoV-2 continues to spread and evolve, detecting emerging variants early is critical for public health interventions. Inferring lineage prevalence by clinical testing is infeasible at scale, especially in areas with limited resources, participation, or testing and/or sequencing capacity, which can also introduce biases1-3. SARS-CoV-2 RNA concentration in wastewater successfully tracks regional infection dynamics and provides less biased abundance estimates than clinical testing4,5. Tracking virus genomic sequences in wastewater would improve community prevalence estimates and detect emerging variants. However, two factors limit wastewater-based genomic surveillance: low-quality sequence data and inability to estimate relative lineage abundance in mixed samples. Here we resolve these critical issues to perform a high-resolution, 295-day wastewater and clinical sequencing effort, in the controlled environment of a large university campus and the broader context of the surrounding county. We developed and deployed improved virus concentration protocols and deconvolution software that fully resolve multiple virus strains from wastewater. We detected emerging variants of concern up to 14 days earlier in wastewater samples, and identified multiple instances of virus spread not captured by clinical genomic surveillance. Our study provides a scalable solution for wastewater genomic surveillance that allows early detection of SARS-CoV-2 variants and identification of cryptic transmission.


Identification of seven new prostate cancer susceptibility loci through a genome-wide association study.
Prostate cancer (PrCa) is the most frequently diagnosed cancer in males in developed countries. To identify common PrCa susceptibility alleles, we previously conducted a genome-wide association study in which 541,129 SNPs were genotyped in 1,854 PrCa cases with clinically detected disease and in 1,894 controls. We have now extended the study to evaluate promising associations in a second stage in which we genotyped 43,671 SNPs in 3,650 PrCa cases and 3,940 controls and in a third stage involving an additional 16,229 cases and 14,821 controls from 21 studies. In addition to replicating previous associations, we identified seven new prostate cancer susceptibility loci on chromosomes 2, 4, 8, 11 and 22 (with P = 1.6 x 10(-8) to P = 2.7 x 10(-33)).


Serum biopterin--a novel marker for immune activation during pre-diabetes in the BB rat.
Tetrahydrobiopterin (BH4) is a pteridine product which is released by rodent macrophages on activation by cytokines. We have used serial pancreatic biopsy, and measurement of serum biopterin at 30, 60, 90 and 120 days in the BB/S rat to relate histological change to macrophage activation during the course of pre-diabetes. Using immunohistochemistry, and an arbitrary scoring system read blind and standardised against day 30, we found that pancreatic MHC class I, MHC class II and infiltrating macrophage staining were up-regulated in the BB/S diabetes-prone rats (n = 17) at day 60, markedly so at day 90, and less so at day 120. Staining for resident pancreatic macrophages remained unchanged throughout in diabetes prone, diabetes resistant and Wistar (n = 28) control animals. Serum biopterin fell progressively and identically with age in BB diabetes resistant rats (n = 11) and Wistar controls. No change in weight gain or biopterin levels was observed in the biopsied animals. Mean serum biopterin levels in diabetes prone rats (of which 13 of 17 became diabetic at median 85 days) were the same as in diabetes resistant and Wistar rats at days 30, 60 and 120, but showed a striking and highly significant elevation (p < 0.001) at day 90. Although macrophages infiltrate the islet early in pre-diabetes, the timing of their activation is unknown. The rise in biopterin we observed is a potentially important immunological event which occurred late in the progression of pre-diabetes. This acute terminal event has not been reported previously, and may modify current concepts concerning the tempo of cell destruction during pre-diabetes.


Effects of Qibaipingfei capsules on pulmonary vascular relaxation through KATP channel activation by the NO/cGMP signaling pathway.
This research explores the effects of Qibaipingfei (QBPF) capsules on pulmonary vascular relaxation in vitro and the relationship of the ATP-sensitive K+ (KATP) channel and nitric oxide (NO) pathway. Vasodilator effects of QBPF (0.125-2 g/kg) on rat pulmonary artery rings were observed using a multi-wire myograph system. The maximum relaxation (Emax) of QBPF was detected following treatment involving endothelial denudation, Nω-nitro-L-arginine methyl ester (L-NAME), 1H-[1,2,4] oxadiazolo[4,3-α]quinoxalin-1-one (ODQ), or glyburide (GLYB). Furthermore, rat models of phlegm and blood stasis syndrome combined with chronic obstructive pulmonary disease (COPD) were established using compound factors. KIR6.1 and SUR2B protein expression was analyzed by western blotting. After 9,11-dideoxy-11-α],9-α]-epoxy-methanoprostaglandinF2α (U46619) was used to pre-constrict endothelium-intact pulmonary artery rings, QBPF induced the effects of concentration-dependent relaxation at a concentration for 50% of maximal effect (EC50) of 0.56 g/L and Emax of 84.30% ± 6.27%. After the endothelium was denuded, the vasodilator effects reduced significantly (P<0.01). QBPF-induced relaxation was inhibited by L-NAME, ODQ, and GLYB (P<0.01). The vasodilator effect was also attenuated in the model group (Emax=62.63%±10.02, EC50 = 0.72 g/L, P<0.01). In comparison with expression in the control group, SUR2B protein expression was down-regulated in the model group (P<0.01) but no significant difference was detected in KIR6.1 protein expression between the groups (P>0.05). QBPF and nicorandil (Nic) treatment up-regulated SUR2B KATP channel expression (P<0.05). QBPF induces endothelial-dependent relaxation in pulmonary artery rings in vitro, through a mechanism that potentially activates the KATP channel in pulmonary vascular smooth muscles via the NO-cyclic GMP (cGMP)-dependent pathway.


Inhibition of transmitter release shortens the duration of the excitatory synaptic current at a calyceal synapse.
1. We investigated the effect of reducing transmitter release on the time course of multiquantal, evoked synaptic currents to test for transmitter "cross talk" between neighboring synaptic release sites within a calyceal synapse. By using a brain slice preparation, neurons in the chick nucleus magnocellularis (nMAG) were voltage clamped and individual presynaptic axons were stimulated to evoke excitatory postsynaptic currents (EPSCs). 2. Application of 100-microM baclofen or 50-microM GABA in the presence of a gamma-aminobutyric acid-A (GABAA) receptor antagonist produced an 85% reduction of EPSCs, consistent with the activation of presynaptic gamma-aminobutyric acid-B (GABAB) receptors. In parallel with the reduction in the amplitude of the EPSC by GABAB receptor activation, the normally strong paired pulse depression (PPD) of the EPSC was converted to facilitation. The reduction in EPSC amplitude by gamma-aminobutyric acid (GABA) or baclofen was accompanied by a 20% reduction in the exponential time constant of decay of the EPSC. Weaker effects on the EPSC time course were observed for synapses with the least PPD. 3. Cd2+ (5 microM), which inhibits presynaptic calcium current, also reduced EPSC amplitude by 85% and converted PPD to facilitation. EPSCs were narrower in Cd2+, though less so than in baclofen. 4. The time course of the EPSC was longer than that of miniature synaptic currents, even after significant block by baclofen, GABA or Cd2+, indicating that dispersion of quantal release may help shape the synaptic waveform. However, the narrowing of the EPSC by baclofen, GABA, and Cd2+ suggests that high levels of quantal release at the calyceal synapse may delay the removal of transmitter, further slowing the EPSC.


A novel long non-coding RNA in the rheumatoid arthritis risk locus TRAF1-C5 influences C5 mRNA levels.
Long non-coding RNAs (lncRNAs) can regulate the transcript levels of genes in the same genomic region. These locally acting lncRNAs have been found deregulated in human disease and some have been shown to harbour quantitative trait loci (eQTLs) in autoimmune diseases. However, lncRNAs linked to the transcription of candidate risk genes in loci associated to rheumatoid arthritis (RA) have not yet been identified. The TRAF1 and C5 risk locus shows evidence of multiple eQTLs and transcription of intergenic non-coding sequences. Here, we identified a non-coding transcript (C5T1lncRNA) starting in the 3' untranslated region (UTR) of C5. RA-relevant cell types express C5T1lncRNA and RNA levels are further enhanced by specific immune stimuli. C5T1lncRNA is expressed predominantly in the nucleus and its expression correlates positively with C5 mRNA in various tissues (P=0.001) and in peripheral blood mononuclear cells (P=0.02) indicating transcriptional co-regulation. Knockdown results in a concurrent decrease in C5 mRNA levels but not of other neighbouring genes. Overall, our data show the identification of a novel lncRNA C5T1lncRNA that is fully located in the associated region and influences transcript levels of C5, a gene previously linked to RA pathogenesis.


Spatial distribution of the risk of dengue fever in southeast Brazil, 2006-2007.
Many factors have been associated with circulation of the dengue fever virus and vector, although the dynamics of transmission are not yet fully understood. The aim of this work is to estimate the spatial distribution of the risk of dengue fever in an area of continuous dengue occurrence. This is a spatial population-based case-control study that analyzed 538 cases and 727 controls in one district of the municipality of Campinas, São Paulo, Brazil, from 2006-2007, considering socio-demographic, ecological, case severity, and household infestation variables. Information was collected by in-home interviews and inspection of living conditions in and around the homes studied. Cases were classified as mild or severe according to clinical data, and they were compared with controls through a multinomial logistic model. A generalized additive model was used in order to include space in a non-parametric fashion with cubic smoothing splines. Variables associated with increased incidence of all dengue cases in the multiple binomial regression model were: higher larval density (odds ratio (OR) = 2.3 (95%CI: 2.0-2.7)), reports of mosquito bites during the day (OR = 1.8 (95%CI: 1.4-2.4)), the practice of water storage at home (OR = 2.5 (95%CI: 1.4, 4.3)), low frequency of garbage collection (OR = 2.6 (95%CI: 1.6-4.5)) and lack of basic sanitation (OR = 2.9 (95%CI: 1.8-4.9)). Staying at home during the day was protective against the disease (OR = 0.5 (95%CI: 0.3-0.6)). When cases were analyzed by categories (mild and severe) in the multinomial model, age and number of breeding sites more than 10 were significant only for the occurrence of severe cases (OR = 0.97, (95%CI: 0.96-0.99) and OR = 2.1 (95%CI: 1.2-3.5), respectively. Spatial distribution of risks of mild and severe dengue fever differed from each other in the 2006/2007 epidemic, in the study area. Age and presence of more than 10 breeding sites were significant only for severe cases. Other predictors of mild and severe cases were similar in the multiple models. The analyses of multinomial models and spatial distribution maps of dengue fever probabilities suggest an area-specific epidemic with varying clinical and demographic characteristics.


Phenotypic and molecular characterisation of multiresistant monophasic Salmonella Typhimurium (1,4,[5],12:i:-) in Greece, 2006 to 2011.
Recently, multiresistant Salmonella enterica serovar1,4,[5],12:i:-, a monophasic variant of S. Typhimurium(1,4,[5],12:i:1,2) emerged, and is now among the most common serovars isolated from humans in many countries.In Greece, monophasic Typhimurium which was recorded for the first time in human isolates in 2007(0.3% of total isolates), increased sharply thereafter,and since 2009 is the third most frequent serovar. In the present study, 119 S. enterica 1,4,[5],12:i:- strains of human, animal and food origin, isolated during the period between 2006 and 2011, were examined.Strains verified as monophasic Typhimurium variants by polymerase chain reaction (PCR) (97 strains), were further characterised by phenotypic (antibiotic resistance and phage typing) and molecular (pulsed-field gel electrophoresis – PFGE) methods. The results indicate that multiple clones of multiresistant monophasic Typhimurium are circulating in Greece. The most frequently encountered clone in humans and pigs was that of phage type DT120, R-type ASSuTSpTm and PFGEprofile STYMXB.0010, while in poultry other clones were detected. The data indicate that pigs may be a reservoir of this clone in Greece.


An investigative study into the influence of a commercially available carbohydrate-protein-electrolyte beverage on short term repeated exercise performance.
The purpose of this study was to undertake an independent investigation into the effects of ingesting a carbohydrate-protein-electrolyte (CPE) beverage on repeated submaximal and time-trial cycling performance. Sixteen recreationally trained males (height: 1.76 ± 0.07 m; weight: 70.05 ± 7.90 kg; VO2max: 49.69 ± 4.19 ml.kg-1.min-1) performed two exercise trials separated by 7 days. Each trial comprised two bouts of 90 minutes exercise separated by a 2 hour recovery period. Each bout comprised 45 minutes exercise on a cycle-ergometer at 60%VO2max (ST), followed immediately by a 45 minute performance test (PT). Participants were randomly assigned an 8% CPE beverage or colour/taste matched placebo (PL) prior to each trial. Participants consumed 100 ml of the assigned beverage every 10 minutes during each ST, and 500 ml at 0 and 60 minutes into recovery (total caloric delivery per trial: 617.6 kcal for CPE and12.8 kcal for PL). Mean power output (W), speed (km.hr-1) and distance covered (km) were assessed throughout both trials. Expired air was sampled at 10 minute intervals throughout ST. Blood glucose and lactate were assessed during ST and recovery. Distance covered during ST was significantly reduced with PL by 9.12% (20.18 ± 0.28 km in ST1 v 18.34 ± 0.36 km in ST2; P = 0.0001). With CPE, distance covered, power output and average speed were maintained between ST1 and ST2. Oxygen uptake was not significantly different between ST1 and ST2, or conditions. Respiratory exchange ratio (RER) values decreased from 0.98 ± 0.02 in ST1 to 0.91 ± 0.02 in ST2 for PL (P = 0.003), supporting reduced total carbohydrate oxidation rates (P = 0.007). Mean blood glucose was maintained in CPE across ST trials, and was significantly greater than PL in ST2 (4.77 ± 0.09 mmol.L-1 for CPE compared with 4.18 ± 0.06 mmol.L-1 for PL, P < 0.001). Mean distance during PT2 was 2.96 km (or 17.1%) further with CPE than PL (P = 0.003). Mean power significantly decreased across PT with PL (134.21 ± 4.79 W and 106.90 ± 3.25 W, respectively; P < 0.04). The use of a CPE beverage improves short-term repeated exercise and subsequent performance compared to PL. Higher rates of carbohydrate oxidation, maintenance of plasma glucose, and decreased levels of fatigue may be beneficial for secondary bouts of performance and faster recovery turnover.


Bone mesenchymal stem cells transplanted into rabbit intervertebral discs can increase proteoglycans.
We sought to determine whether transplanted allogeneic bone mesenchymal stem cells can survive and increase the amount of proteoglycans in intervertebral discs. We used the rabbit intervertebral disc as a model, creating three groups: an uninjected control group, a group injected with saline, and a group injected with 1 x 10(5) of bone mesenchymal stem cells containing trace marker gene LacZ from young rabbits. At 1, 3, and 6 months, X-gal staining and DNA-polymerase chain reaction of the neomycin-resistant gene were used to ascertain cell location. Reverse transcription-polymerase chain reaction and enzyme-linked immunosorbant assay analysis were done to evaluate the effect on the disc matrix. Transplanted mesenchymal stem cells were located and identified in the group injected with mesenchymal stem cells, whereas we detected none in the saline and control groups. The amount of mRNA and protein of proteoglycan and collagen Type II in the mesenchymal stem cells group was increased, whereas the amount of collagen Type I did not change. We found no changes in the saline group. Our data suggest transplanted allogenic bone mesenchymal stem cells can survive and increase proteoglycan amount, supporting its potential use as a treatment of intervertebral disc degeneration.


Multifocal demyelinating motor neuropathy and hamartoma syndrome associated with a de novo PTEN mutation.
To describe a patient with a multifocal demyelinating motor neuropathy with onset in childhood and a mutation in phosphatase and tensin homolog (PTEN), a tumor suppressor gene associated with inherited tumor susceptibility conditions, macrocephaly, autism, ataxia, tremor, and epilepsy. Functional implications of this protein have been investigated in Parkinson and Alzheimer diseases. We performed whole-exome sequencing in the patient's genomic DNA validated by Sanger sequencing. Immunoblotting, in vitro enzymatic assay, and label-free shotgun proteomic profiling were performed in the patient's fibroblasts. The predominant clinical presentation of the patient was a childhood onset, asymmetric progressive multifocal motor neuropathy. In addition, he presented with macrocephaly, autism spectrum disorder, and skin hamartomas, considered as clinical criteria for PTEN-related hamartoma tumor syndrome. Extensive tumor screening did not detect any malignancies. We detected a novel de novo heterozygous c.269T>C, p.(Phe90Ser) PTEN variant, which was absent in both parents. The pathogenicity of the variant is supported by altered expression of several PTEN-associated proteins involved in tumorigenesis. Moreover, fibroblasts showed a defect in catalytic activity of PTEN against the secondary substrate, phosphatidylinositol 3,4-trisphosphate. In support of our findings, focal hypermyelination leading to peripheral neuropathy has been reported in PTEN-deficient mice. We describe a novel phenotype, PTEN-associated multifocal demyelinating motor neuropathy with a skin hamartoma syndrome. A similar mechanism may potentially underlie other forms of Charcot-Marie-Tooth disease with involvement of the phosphatidylinositol pathway.


Crystal structure of a cytokine-binding region of gp130.
The structure of the cytokine-binding homology region of the cell surface receptor gp130 has been determined by X-ray crystallography at 2.0 A resolution. The beta sandwich structure of the two domains conforms to the topology of the cytokine receptor superfamily. This first structure of an uncomplexed receptor exhibits a similar L-shaped quaternary structure to that of ligand-bound family members and suggests a limited flexibility in relative domain orientation of some 3 degrees. The putative ligand-binding loops are relatively rigid, with a phenylalanine side chain similarly positioned to exposed aromatic residues implicated in ligand binding for other such receptors. The positioning and structure of the N-terminal portion of the polypeptide chain have implications for the structure and function of cytokine receptors, such as gp130, which contain an additional N-terminal immunoglobulin-like domain.


Chemical modification of proteins at cysteine: opportunities in chemistry and biology.
Chemical modification of proteins is a rapidly expanding area in chemical biology. Selective installation of biochemical probes has led to a better understanding of natural protein modification and macromolecular function. In other cases such chemical alterations have changed the protein function entirely. Additionally, tethering therapeutic cargo to proteins has proven invaluable in campaigns against disease. For controlled, selective access to such modified proteins, a unique chemical handle is required. Cysteine, with its unique reactivity, has long been used for such modifications. Cysteine has enjoyed widespread use in selective protein modification, yet new applications and even new reactions continue to emerge. This Focus Review highlights the enduring utility of cysteine in protein modification with special focus on recent innovations in chemistry and biology associated with such modifications.


Lateral supraorbital approach applied to anterior clinoidal meningiomas: experience with 73 consecutive patients.
Anterior clinoidal meningiomas (ACMs) are a subgroup of meningiomas accounting for less than 10% of supratentorial meningiomas. To assess the reliability and safeness of the lateral supraorbital approach (LSO) to remove ACMs. Between September 1997 and October 2009, a total of 73 ACM patients were operated on at the Department of Neurosurgery, Helsinki University Central Hospital, by the senior author (J.H.). We retrospectively analyzed the clinical data, radiological findings, surgical treatment, histology, and outcome of patients, and discuss the operative technique. Seventy-three patients were operated on by applying the LSO approach. Apparently complete removal was achieved in 57 patients (78%). Anterior clinoidectomy was performed in 21 cases. Preexisting visual deficit improved in 11 of 39 patients and worsened in 4; 3 had de novo visual deficit. At 3 months after discharge, 60 (82%) patients had a good recovery, 9 (12%) patients were moderately disabled, 1 presented with severe disability, and 3 (4%) patients died of surgery-related causes. Sixteen (22%) patients had residual tumors, 6 of which required reoperation. During the median follow-up of 36 months (range, 3-146), tumor recurred in 3 patients: 2 were followed-up and 1 was reoperated on. ACMs can be removed via the LSO approach with relatively low morbidity and mortality. Anterior clinoidectomy is required only in selected cases, and we prefer the intradural approach during the LSO approach. High-power coagulation should be avoided in proximity of the optic nerve.


On the simulation of vibrationally resolved electronic spectra of medium-size molecules: the case of styryl substituted BODIPYs.
BODIPY dyes are used in a variety of applications because of their peculiar spectroscopic and photo-physical properties that vary depending on the stereochemistry of the functional groups attached to the boron-dipyrromethene core structure. In this work, we have applied several computational methods, adapted for semi-rigid molecules based on the Franck-Condon principle, for the study of the optical properties of BODIPY systems and for the understanding of the influence of functional groups on their spectroscopic features. We have analyzed the electronic spectra of two styryl substituted BODIPY molecules of technological interest, properly taking into account the vibronic contribution. For comparison with recently recorded experimental data in methanol, the vibrationally resolved electronic spectra of these systems were computed using both Time-Independent (TI) and Time-Dependent (TD) formalisms. The first step toward the analysis of optical properties of the styryl modified BODIPYs was a benchmark of several density functionals, to select the most appropriate one. We have found that all benchmarked functionals provide good results in terms of band shape but some of them show strong discrepancies in terms of band position. Beyond the issue of the electronic structure calculation method, different levels of sophistication can be adopted for the calculation of vibronic transitions. In this study, the effect of mode couplings and the influence of the Herzberg-Teller terms on the theoretical spectra has been investigated. It has been found that all levels of theory considered give reproducible results for the investigated systems: band positions and shapes are similar at all levels and little improvements have been found in terms of band shape with the inclusion of Herzberg-Teller effect. Inclusion of temperature effects proved to be challenging due to the important impact of large amplitude motions. Better agreement can be achieved by adopting a suitable set of coordinates coupled with a reduced-dimensionality scheme.


Reliability, validity, and responsiveness of the six-minute walk test in patients with heart failure.
Our purpose was to evaluate the reliability, validity, and responsiveness of the 6-minute walk test (6MWT) in patients with heart failure (HF) enrolled in the Randomized Evaluation of Strategies for Left Ventricular Dysfunction (RESOLVD) pilot study. A total of 768 patients was enrolled in a multicenter randomized clinical trial evaluating the effect of candesartan, enalapril, and metoprolol on left ventricular ejection fraction (LVEF), 6MWT distance, neurohormones, and quality of life. The 6MWT was performed once at screening and twice at baseline, 18 weeks, and 43 weeks by a standardized method. Test-retest reliability at baseline (intraclass correlation coefficient [ICC] = 0.90), 18 weeks (ICC = 0.88), and 43 weeks (ICC = 0.91) was very good. Baseline 6MWT distance was weakly inversely correlated to the quality-of-life cumulative score (r = -0.26, P =.0001) and moderately inversely correlated to the New York Heart Association functional classification (NYHA-FC) (r = -0.43, P =.001). In the RESOLVD study, the 6MWT was not responsive to change when effect sizes and standardized response means were used. Disease-specific quality of life was responsive to change in patients treated with candesartan and enalapril and NYHA-FC was responsive to change in the candesartan and enalapril combination and for enalapril alone with small effect sizes. The 6MWT, NYHA-FC, and quality of life were not responsive to change during the metoprolol or placebo phase. The 6MWT is highly reproducible in patients with symptoms of HF. It is somewhat correlated to NYHA-FC and quality of life. Overall, quality of life was most responsive to change, whereas 6MWT and NYHA-FC were comparable but less responsive to change in the RESOLVD study.


Quantification of protein expression in cells and cellular subcompartments on immunohistochemical sections using a computer supported image analysis system.
Quantification of protein expression based on immunohistochemistry (IHC) is an important step for translational research and clinical routine. Several manual ('eyeballing') scoring systems are used in order to semi-quantify protein expression based on chromogenic intensities and distribution patterns. However, manual scoring systems are time-consuming and subject to significant intra- and interobserver variability. The aim of our study was to explore, whether new image analysis software proves to be sufficient as an alternative tool to quantify protein expression. For IHC experiments, one nucleus specific marker (i.e., ERG antibody), one cytoplasmic specific marker (i.e., SLC45A3 antibody), and one marker expressed in both compartments (i.e., TMPRSS2 antibody) were chosen. Stainings were applied on TMAs, containing tumor material of 630 prostate cancer patients. A pathologist visually quantified all IHC stainings in a blinded manner, applying a four-step scoring system. For digital quantification, image analysis software (Tissue Studio v.2.1, Definiens AG, Munich, Germany) was applied to obtain a continuous spectrum of average staining intensity. For each of the three antibodies we found a strong correlation of the manual protein expression score and the score of the image analysis software. Spearman's rank correlation coefficient was 0.94, 0.92, and 0.90 for ERG, SLC45A3, and TMPRSS2, respectively (p⟨0.01). Our data suggest that the image analysis software Tissue Studio is a powerful tool for quantification of protein expression in IHC stainings. Further, since the digital analysis is precise and reproducible, computer supported protein quantification might help to overcome intra- and interobserver variability and increase objectivity of IHC based protein assessment.


Cerebrospinal Fluid from Alzheimer's Disease Patients Contains Fungal Proteins and DNA.
The identification of biomarkers for Alzheimer's disease is important for patient management and to assess the effectiveness of clinical intervention. Cerebrospinal fluid (CSF) biomarkers constitute a powerful tool for diagnosis and monitoring disease progression. We have analyzed the presence of fungal proteins and DNA in CSF from AD patients. Our findings reveal that fungal proteins can be detected in CSF with different anti-fungal antibodies using a slot-blot assay. Additionally, amplification of fungal DNA by PCR followed by sequencing distinguished several fungal species. The possibility that these fungal macromolecules could represent AD biomarkers is discussed.


Manual therapy for cervicogenic dizziness: Long-term outcomes of a randomised trial.
Manual therapy is effective for reducing cervicogenic dizziness, a disabling and persistent problem, in the short term. This study investigated the effects of sustained natural apophyseal glides (SNAGs) and passive joint mobilisations (PJMs) on cervicogenic dizziness compared to a placebo at 12 months post-treatment. Eighty-six participants (mean age 62 years, standard deviation (SD) 12.7) with chronic cervicogenic dizziness were randomised to receive SNAGs with self-SNAGs (n = 29), PJMs with range-of-motion (ROM) exercises (n = 29), or a placebo (n = 28) for 2-6 sessions over 6 weeks. Outcome measures were dizziness intensity, dizziness frequency (rated between 0 [none] and 5 [>once/day]), the Dizziness Handicap Inventory (DHI), pain intensity, head repositioning accuracy (HRA), cervical spine ROM, balance, and global perceived effect (GPE). At 12 months both manual therapy groups had less dizziness frequency (mean difference SNAGs vs placebo -0.7, 95% confidence interval (CI) -1.3, -0.2, p = 0.01; PJMs vs placebo -0.7, -1.2, -0.1, p = 0.02), lower DHI scores (mean difference SNAGs vs placebo -8.9, 95% CI -16.3, -1.6, p = 0.02; PJMs vs placebo -13.6, -20.8, -6.4, p < 0.001) and higher GPE compared to placebo, whereas there were no between-group differences in dizziness intensity, pain intensity or HRA. There was greater ROM in all six directions for the SNAG group and in four directions for the PJM group compared to placebo, and small improvements in balance for the SNAG group compared to placebo. There were no adverse effects. These results provide evidence that both forms of manual therapy have long-term beneficial effects in the treatment of chronic cervicogenic dizziness.


Impact of hemodialysis on liver stiffness measured with real-time two-dimensional shear wave elastography.
The impact of hemodialysis on liver stiffness is still unclear. The aim of the study was to assess liver fibrosis by real-time two-dimensional shear wave elastography (RT 2D-SWE) and to quantify the influence of net fluid withdrawal on liver stiffness during one hemodialysis session. The second aim was to investigate the influence of systolic blood pressure and time spent on dialysis (in years) on liver stiffness measurements. This before/after hemodialysis (HD) study in a group of end stage renal disease (ESRD) patients was carried out with patients on regular HD. Measurements of liver stiffness were done using RT 2D-SWE directly before and after a hemodialysis session. In this study 27 patients with mean age 69.4 ± 14.75 years were included. Mean net fluid withdrawal volume per session was 2874.07 ± 778.35 ml. Mean pre-HD and post-HD liver stiffness measurements were 8.15 kPa (95% confidence interval, CI 7.61-8.68) and 6.70 kPa (95% CI 6.10-7.30 kPa), respectively. Mean liver stiffness reduction was 1.448 ± 1.14 kPa. The amount of fluid removed correlated with the decline in liver stiffness values after HD (ρ = 0.523, P = 0.003). There was a positive but statistically not significant correlation between time spent in HD and liver stiffness (ρ = 0.151, P = 0.623) CONCLUSION: Liver stiffness significantly declined after one session of HD. The change in liver stiffness was strongly correlated with the amount of net fluid withdrawal. Random liver stiffness measurements (LSM) by RT 2D-SWE does not precisely show the degree of fibrosis, Furthermore, it is presumed that postdialysis liver stiffness values likely reflect the real degree of liver fibrosis.


Prognostic value of an abnormal signal-averaged electrocardiogram in patients with nonischemic congestive cardiomyopathy.
An abnormal signal-averaged ECG (SAECG) has predictive value for arrhythmic events in patients with coronary artery disease. The purpose of this study was to investigate whether an abnormal SAECG could provide prognostic information in patients with nonischemic dilated cardiomyopathy. We prospectively obtained SAECGs in 114 patients with dilated nonischemic cardiomyopathy. Twelve-lead ECGs, left ventricular ejection fractions, hemodynamic measurements, and peak exercise oxygen consumption (VO2) also were measured. An SAECG was defined as abnormal by any one of the three following criteria: filtered QRS duration > 120 msec, root-mean-square voltage in the last 40 msec < 20 microV, or duration < 40 microV > 38 msec at 40 Hz. Sixty-six patients had a normal SAECG, 20 patients had an abnormal SAECG, and 28 patients had bundle branch block (BBB). Mean follow-up was 10 +/- 5 months. Age, ejection fraction, peak VO2, pulmonary capillary wedge pressure, and cardiac index were not statistically different among the three groups. Use of antiarrhythmic drugs was similar among the three groups, although patients with BBB had more implantable defibrillators (p < 0.05). The incidence of previous atrial arrhythmias was similar for the three groups. Patients with abnormal SAECG or BBB had more past episodes of sustained ventricular tachycardia and/or sudden death episodes (n = 9) than patients with normal SAECG (n = 1) (p < 0.01). Prospectively, none of the 66 patients with normal SAECG died suddenly or had sustained ventricular arrhythmias. Two deaths occurred from progressive heart failure, and three patients required urgent transplant. In the 20 patients with an abnormal SAECG, four patients had sustained ventricular tachycardia, five patients died suddenly, two patients died from progressive heart failure, and one patient required urgent transplant. In the patients with BBB, four patients had sustained ventricular tachycardia, and four patients required urgent transplant. One-year event-free survival, i.e., absence of ventricular tachycardia and/or death, was 95% in patients with normal SAECG, 88% in patients with BBB, and only 39% in patients with an abnormal SAECG (p < 0.001). Multivariate analysis demonstrated that SAECG and New York Heart Association classification were independent predictors of survival. Patients with an abnormal SAECG had a statistically significant increase in sustained ventricular arrhythmias and/or death than did patients with a normal SAECG or BBB. This study demonstrates that an abnormal SAECG is a marker of past and future arrhythmic events in patients with nonischemic dilated cardiomyopathy. In contrast, patients with a dilated cardiomyopathy with a normal SAECG have an excellent prognosis with adverse outcome only from progressive heart failure.


Is virtual reality training superior to conventional treatment in improving lower extremity motor function in chronic hemiplegic patients?
This study aims to examine the effect of virtual reality (VR) training, frequently included in rehabilitation programs, on lower extremity functional status, mobility, balance, and walking speed in chronic stroke patients. This randomized, controlled study was conducted with 60 chronic stroke patients (26 males, 34 females; mean age: 64.0 years; range, 33 to 80 years) who presented to the physical therapy and rehabilitation outpatient clinic of the Kütahya Health Sciences University Evliya Çelebi Training and Research Hospital between February 2019 and February 2020. The participants were randomized to the VR group and the control group by simple randomization with 1:1 allocation. The VR group received 30 min of VR training and 30 min of conventional physiotherapy, while the control group received 60 min of conventional physiotherapy. The patients were evaluated before and after treatment using the Fugl-Meyer Assessment-Lower Extremity (FMA-LE), Rivermead Mobility Index (RMI), 10-m walk test (10MWT), and Berg Balance Scale (BBS). The FMA-LE, RMI, 10MWT, and BBS scores significantly improved in both groups after treatment (p<0.001). The post-treatment change in the FMA-LE score was significantly higher in the VR group than in the control group (Z=-3.560, p<0.001). Similarly, the change in the BBS score was significantly higher in the VR group (Z=-3.769, p<0.001). Post-treatment changes in the RMI and 10MWT were not significant (p>0.05). Virtual reality training combined with conventional physiotherapy was found to be superior to conventional physiotherapy alone in improving lower extremity functional status in chronic stroke patients; therefore, adding a VR component to rehabilitation programs will have a favorable impact on treatment outcomes.


Palynological origin, phenolic content, and antioxidant properties of honeybee-collected pollen from Bahia, Brazil.
The aim of this study was to determine the palynological origin, phenolic and flavonoid content, and antioxidant properties of twenty-five samples of bee pollen harvested during a nine-month period (February-November) from the Canavieiras municipality (northeastern Brazil). Of the 25 samples analyzed, only two (February 01 and 02) were heterofloral. The predominant pollens in the samples analyzed during that month were: Cecropia, Eucalyptus, Elaeis, Mimosa pudica, Eupatorium, and Scoparia. Ethyl acetate fractions were analyzed by HPLC-DAD. The flavonoids isoquercetin, myricetin, tricetin, quercetin, luteolin, selagin, kaempferol, and isorhamnetin were detected. The flavonoid present in all 22 samples was isolated and identified as isorhamnetin 3-O-b-neohesperidoside. The total phenolic contents determined using the Folin-Ciocalteu reagent ranged from 41.5 to 213.2 mg GAE/g. Antioxidant activities based on the 1,1-diphenyl-2-picryl hydrazyl (DPPH), 2,2-azinobis 3-ethylbenzothiozoline-6-sulfonic acid (ABTS), and Fe2+ ion chelating activity assays were observed for all extracts, and correlated with the total phenolic content.


Overuse of screening colonoscopy in the Medicare population.
All relevant authorities recommend an interval of 10 years between normal screening colonoscopies. We assessed the timing of repeated colonoscopies after a negative screening colonoscopy finding in a population-based sample of Medicare patients. A 5% national sample of Medicare enrollees from 2000 through 2008 was used to identify average-risk patients undergoing screening colonoscopy between 2001 and 2003. Colonoscopy was classified as a negative screening examination finding if no indication other than screening were in the claims and if no biopsy, fulguration, or polypectomy was performed. Time to repeated colonoscopy was calculated. Among 24,071 Medicare patients who had a negative screening colonoscopy finding in 2001 through 2003, 46.2% underwent a repeated examination in fewer than 7 years. In 42.5% of these patients (23.5% of the overall sample), there was no clear indication for the early repeated examination. In patients aged 75 to 79 years or 80 years or older at the time of the initial negative screening colonoscopy result, 45.6% and 32.9%, respectively, received a repeated examination within 7 years. In multivariable analyses, male sex, more comorbidities, and colonoscopy by a high-volume colonoscopist or in an office setting were associated with higher rates of early repeated colonoscopy without clear indication, while those 80 years or older had a reduced risk. There were also marked geographic variations, from less than 5% in some health referral regions to greater than 50% in others. A large proportion of Medicare patients who undergo screening colonoscopy do so more frequently than recommended. Current Medicare regulations intending to limit reimbursement for screening colonoscopy to every 10 years would not appear to be effective.


Wayfinding and Glaucoma: A Virtual Reality Experiment.
Wayfinding, the process of determining and following a route between an origin and a destination, is an integral part of everyday tasks. The purpose of this study was to investigate the impact of glaucomatous visual field loss on wayfinding behavior using an immersive virtual reality (VR) environment. This cross-sectional study included 31 glaucomatous patients and 20 healthy subjects without evidence of overall cognitive impairment. Wayfinding experiments were modeled after the Morris water maze navigation task and conducted in an immersive VR environment. Two rooms were built varying only in the complexity of the visual scene in order to promote allocentric-based (room A, with multiple visual cues) versus egocentric-based (room B, with single visual cue) spatial representations of the environment. Wayfinding tasks in each room consisted of revisiting previously visible targets that subsequently became invisible. For room A, glaucoma patients spent on average 35.0 seconds to perform the wayfinding task, whereas healthy subjects spent an average of 24.4 seconds (P = 0.001). For room B, no statistically significant difference was seen on average time to complete the task (26.2 seconds versus 23.4 seconds, respectively; P = 0.514). For room A, each 1-dB worse binocular mean sensitivity was associated with 3.4% (P = 0.001) increase in time to complete the task. Glaucoma patients performed significantly worse on allocentric-based wayfinding tasks conducted in a VR environment, suggesting visual field loss may affect the construction of spatial cognitive maps relevant to successful wayfinding. VR environments may represent a useful approach for assessing functional vision endpoints for clinical trials of emerging therapies in ophthalmology.


[Evaluation of oral health conditions among the elderly in a Brazilian city].
This study assessed the oral health of individuals 60 years or older in the city of Araraquara, São Paulo, Brazil, in 1998. Of the 194 people who participated in the study, 91 of them were institutionalized and had an average age of 73.6 years, and 103 were not institutionalized and had an average age of 69.3 years. The study participants were examined by a previously trained oral surgeon who determined the prevalence of the most common oral health problems. The results revealed a large number of edentulous individuals (72% of those institutionalized and 60% of the noninstitutionalized participants) and many persons with extracted teeth (93% and 90%, respectively), as well as a high frequency of periodontal pockets (57% and 75%, respectively) and of inadequate dentures (80% and 61%, respectively). Our results show reduced quality of life for a large proportion of these older individuals, and also indicate that public health services should pay greater attention to this population group.


Psychometric Properties of an Indian Translation of the Vision-related Activity Limitation Item Bank in Cataract.
The Indian translated and culturally adapted version of the vision-related activity limitation (VRAL) item bank is a validated instrument to assess the difficulty in performing daily activities by cataract patients and can also be used to capture self-reported changes in ability to perform daily activities after cataract surgery. The purpose of this study was to document (a) translation, cross-cultural adaptation of VRAL item bank into an Indian language, and (b) its validation using Rasch analysis in a South Indian cataract population. At the first stage, a translated Indian version of VRAL item bank was produced using recommended procedures. At the second stage, Rasch analysis was performed to investigate its psychometric properties in 787 cataract patients (mean age, 58.2 years; mean ± SD visual acuity [logMAR], 1.19 ± 0.96 at baseline in eye for surgery) including comparison with the original version. Post-translation equivalence of meaning was achieved, but some English phrases required cross-cultural adaptation. Subsequently, all items were appropriate for the Indian culture, and VRAL item bank demonstrated excellent measurement precision (7.39). Dimensionality assessment suggested that VRAL construct may contain other dimensions such as self-care and visual search, and mobility. Self-care and visual search formed a unidimensional measure but was highly correlated with main VRAL dimension, and the removal of its items weakened precision of the main VRAL dimension measurement. Taken together, evidence favored retaining self-care and visual search items in a larger VRAL item bank. Mobility subscale lacked adequate measurement precision, so it was not examined further; again, items were retained in VRAL scale because they strengthened its measurement properties. Majority of items (99%) did not demonstrate notable differential item functioning (>1.0 logit) by presenting visual acuity (median, 0.20 logMAR) in the better-seeing eye. Items in the translated Indian VRAL item bank measure the same construct as the English version and fulfilled the psychometric requirements for use in cataract patients.


Theoretical prediction of spin-crossover temperatures in ligand-driven light-induced spin change systems.
Spin-crossover compounds exhibit two alternative spin states with distinctive chemical and physical properties, a particular feature that makes them promising materials for nanotechnological applications as memory or display devices. A key parameter that characterizes these compounds is the spin-crossover temperature, T(1/2), defined as the temperature with equal populations of high and low-spin species. In this study, a theoretical/computational approach is described for the calculation of T(1/2) for the trans-[Fe(styrylpyridine)(4)(NCX)(2)] (X = S, Se, and BH(3), styrylpyridine in the trans configuration) ligand driven light-induced spin change (LD-LISC) complexes. In all cases, the present calculations provide an accurate description of both structural and electronic properties of the LD-LISC complexes and, importantly, predict spin-crossover temperatures in good agreement with the corresponding experimental data. Fundamental insights into the dependence of T(1/2) on the nature of the axial ligands are obtained from the direct analysis of the underlying electronic structure in terms of the relevant molecular orbitals.


An international survey of predictive genetic testing in children for adult onset conditions.
Predictive genetic testing is offered to asymptomatic adults even when there is no effective prophylaxis or treatment. Testing of young people in similar circumstances is controversial, and guidelines recommend against it. We sought to document descriptive examples of the occurrence of genetic testing in young people for nonmedical reasons, in the countries where guidelines exist. Clinical geneticists in the USA, Canada, UK, Australia, and New Zealand were surveyed about the occurrence and outcomes of testing in asymptomatic young people for conditions where no prophylaxis or treatment exists and onset is usually in adulthood. Of 301 responses, details were provided of 49 cases where such testing had occurred. The most common condition tested for was Huntington Disease. In 22 cases (45%), the young person tested was immature, defined as under the age of 14 years. Results were disclosed to only two immature minors and in three cases parents experienced clinically significant anxiety related to how they would pass on information to their gene positive child. In 27 cases (55%), the young person tested was mature. Results were disclosed to 26 mature minors and it was reported that two individuals experienced an adverse event. Consistent follow-up did not take place and findings represent the minimum frequency of adverse events. The majority of respondents agree with existing guidelines but many believe each case must be considered individually. Clinicians agree with existing guidelines regarding predictive testing in young people, but choose to provide tests for nonmedical reasons in specific cases.


Optical microangiography of retina and choroid and measurement of total retinal blood flow in mice.
We present a novel application of optical microangiography (OMAG) imaging technique for visualization of depth-resolved vascular network within retina and choroid as well as measurement of total retinal blood flow in mice. A fast speed spectral domain OCT imaging system at 820nm with a line scan rate of 140 kHz was developed to image the posterior segment of eyes in mice. By applying an OMAG algorithm to extract the moving blood flow signals out of the background tissue, we are able to provide true capillary level imaging of the retinal and choroidal vasculature. The microvascular patterns within different retinal layers are presented. An en face Doppler OCT approach [Srinivasan et al., Opt Express 18, 2477 (2010)] was adopted for retinal blood flow measurement. The flow is calculated by integrating the axial blood flow velocity over the vessel area measured in an en face plane without knowing the blood vessel angle. Total retinal blood flow can be measured from both retinal arteries and veins. The results indicate that OMAG has the potential for qualitative and quantitative evaluation of the microcirculation in posterior eye compartments in mouse models of retinopathy and neovascularization.


Susceptibility of different populations of Colombian Aotus monkeys to the FCB-1 strain of Plasmodium falciparum.
p6udies of susceptibility to the FCB-1 strain of Plasmodium falciparum from Colombia were conducted in three different groups of Aotus monkeys. The results indicated that Aotus trivirgatus griseimembra from the north of the Magdalena Valley were the most susceptible host. A. t. griseimembra from the central Magdalena Valley appeared to be less susceptible to infection, while Aotus monkeys from the eastern Andes were even more resistant to infection. Serum electrophoretic studies were not useful as indicators of resistance or susceptibility to infection. However, the presence of 58 chromosomes in monkeys from the east Andes, a karyotype not previously reported, may be an indicator for the resistance of these animals to the FCB-1 strain. Based on these results, it is possible to assume that Aotus monkeys from the eastern Andes region of Colombia may represent a new species of Aotus; further studies are needed to clarify its taxonomic status.


Development of a scale to measure stigma related to podoconiosis in Southern Ethiopia.
Health-related stigma adds to the physical and economic burdens experienced by people suffering from neglected tropical diseases (NTDs). Previous research into the NTD podoconiosis showed significant stigma towards those with the disease, yet no formal instrument exists by which to assess stigma or interventions to reduce stigma. We aimed to develop, pilot and validate scales to measure the extent of stigma towards podoconiosis among patients and in podoconiosis-endemic communities. Indicators of stigma were drawn from existing qualitative podoconiosis research and a literature review on measuring leprosy stigma. These were then formulated into items for questioning and evaluated through a Delphi process in which irrelevant items were discounted. The final items formed four scales measuring two distinct forms of stigma (felt stigma and enacted stigma) for those with podoconiosis and those without the disease. The scales were formatted as two questionnaires, one for podoconiosis patients and one for unaffected community members. 150 podoconiosis patients and 500 unaffected community members from Wolaita zone, Southern Ethiopia were selected through multistage random sampling to complete the questionnaires which were interview-administered. The scales were evaluated through reliability assessment, content and construct validity analysis of the items, factor analysis and internal consistency analysis. All scales had Cronbach's alpha over 0.7, indicating good consistency. The content and construct validity of the scales were satisfactory with modest correlation between items. There was significant correlation between the felt and enacted stigma scales among patients (Spearman's r = 0.892; p < 0.001) and within the community (Spearman's r = 0.794; p < 0.001). We report the development and testing of the first standardised measures of podoconiosis stigma. Although further research is needed to validate the scales in other contexts, we anticipate they will be useful in situational analysis and in designing, monitoring and evaluating interventions. The scales will enable an evidence-based approach to mitigating stigma which will enable implementation of more effective disease control and help break the cycle of poverty and NTDs.


Atmospheric correction algorithm for hyperspectral remote sensing of ocean color from space.
Existing atmospheric correction algorithms for multichannel remote sensing of ocean color from space were designed for retrieving water-leaving radiances in the visible over clear deep ocean areas and cannot easily be modified for retrievals over turbid coastal waters. We have developed an atmospheric correction algorithm for hyperspectral remote sensing of ocean color with the near-future Coastal Ocean Imaging Spectrometer. The algorithm uses look-up tables generated with a vector radiative transfer code. Aerosol parameters are determined by a spectrum-matching technique that uses channels located at wavelengths longer than 0.86 mum. The aerosol information is extracted back to the visible based on aerosol models during the retrieval of water-leaving radiances. Quite reasonable water-leaving radiances have been obtained when our algorithm was applied to process hyperspectral imaging data acquired with an airborne imaging spectrometer.


Glutamatergic drive facilitates synaptic inhibition of dorsal vagal motor neurons after experimentally induced diabetes in mice.
The role of central regulatory circuits in modulating diabetes-associated glucose dysregulation has only recently been under rigorous investigation. One brain region of interest is the dorsal motor nucleus of the vagus (DMV), which contains preganglionic parasympathetic motor neurons that regulate subdiaphragmatic visceral function. Previous research has demonstrated that glutamatergic and GABAergic neurotransmission are independently remodeled after chronic hyperglycemia/hypoinsulinemia. However, glutamatergic circuitry within the dorsal brain stem impinges on GABAergic regulation of the DMV. The present study investigated the role of glutamatergic neurotransmission in synaptic GABAergic control of DMV neurons after streptozotocin (STZ)-induced hyperglycemia/hypoinsulinemia by using electrophysiological recordings in vitro. The frequency of spontaneous inhibitory postsynaptic currents (sIPSCs) was elevated in DMV neurons from STZ-treated mice. The effect was abolished in the presence of the ionotropic glutamate receptor blocker kynurenic acid or the sodium channel blocker tetrodotoxin, suggesting that after STZ-induced hyperglycemia/hypoinsulinemia, increased glutamatergic receptor activity occurs at a soma-dendritic location on local GABA neurons projecting to the DMV. Although sIPSCs in DMV neurons normally demonstrated considerable amplitude variability, this variability was significantly increased after STZ-induced hyperglycemia/hypoinsulinemia. The elevated amplitude variability was not related to changes in quantal release, but rather correlated with significantly elevated frequency of sIPSCs in these mice. Taken together, these findings suggest that GABAergic regulation of central vagal circuitry responsible for the regulation of energy homeostasis undergoes complex functional reorganization after several days of hyperglycemia/hypoinsulinemia, including both glutamate-dependent and -independent forms of plasticity.


Genome puzzle master (GPM): an integrated pipeline for building and editing pseudomolecules from fragmented sequences.
Next generation sequencing technologies have revolutionized our ability to rapidly and affordably generate vast quantities of sequence data. Once generated, raw sequences are assembled into contigs or scaffolds. However, these assemblies are mostly fragmented and inaccurate at the whole genome scale, largely due to the inability to integrate additional informative datasets (e.g. physical, optical and genetic maps). To address this problem, we developed a semi-automated software tool-Genome Puzzle Master (GPM)-that enables the integration of additional genomic signposts to edit and build 'new-gen-assemblies' that result in high-quality 'annotation-ready' pseudomolecules. With GPM, loaded datasets can be connected to each other via their logical relationships which accomplishes tasks to 'group,' 'merge,' 'order and orient' sequences in a draft assembly. Manual editing can also be performed with a user-friendly graphical interface. Final pseudomolecules reflect a user's total data package and are available for long-term project management. GPM is a web-based pipeline and an important part of a Laboratory Information Management System (LIMS) which can be easily deployed on local servers for any genome research laboratory. The GPM (with LIMS) package is available at https://github.com/Jianwei-Zhang/LIMS CONTACTS: email@example.com or firstname.lastname@example.com information: Supplementary data are available at Bioinformatics online.


Characteristics and outcomes of pregnant women with type 1 or type 2 diabetes: a 5-year national population-based cohort study.
Diabetes in pregnancy is associated with preterm delivery, birthweight extremes, and increased rates of congenital anomaly, stillbirth, and neonatal death. We aimed to identify and compare modifiable risk factors associated with adverse pregnancy outcomes in women with type 1 diabetes and those with type 2 diabetes and to identify effective maternity clinics. In this national population-based cohort study, we used data for pregnancies among women with type 1 or type 2 diabetes collected in the first 5 years of the National Pregnancy in Diabetes audit across 172 maternity clinics in England, Wales, and the Isle of Man, UK. Data for obstetric complications (eg, preterm delivery [<37 weeks' gestation], large for gestational age [LGA] birthweight [>90th percentile]) and adverse pregnancy outcomes (congenital anomaly, stillbirth, neonatal death) were obtained for pregnancies completed between Jan 1, 2014, and Dec 31, 2018. We assessed associations between modifiable (eg, HbA1c, BMI, pre-pregnancy care, maternity clinic) and non-modifiable risk factors (eg, age, ethnicity, deprivation, duration of type 1 diabetes) with pregnancy outcomes in women with type 1 diabetes compared with those with type 2 diabetes. We calculated associations between maternal factors and perinatal deaths using a regression model, including diabetes type and duration, maternal age, BMI, deprivation quintile, first trimester HbA1c, preconception folic acid, potentially harmful medications, and third trimester HbA1c. Our dataset included 17 375 pregnancy outcomes in 15 290 pregnant women. 8690 (50·0%) of 17 375 pregnancies were in women with type 1 diabetes (median age at delivery 30 years [10-90th percentile 22-37], median duration of diabetes 13 years [3-25]) and 8685 (50·0%) were in women with type 2 diabetes (median age at delivery 34 years [27-41], median duration of diabetes 3 years [0-10]). The rates of preterm delivery (3325 [42·5%] of 7825 pregnancies among women with type 1 diabetes, 1825 [23·4%] of 7815 with type 2 diabetes; p<0·0001), and LGA birthweight (4095 [52·2%] of 7845 with type 1 diabetes, 2065 [26·2%] of 7885 with type 2 diabetes; p<0·0001) were higher in type 1 diabetes. The prevalence of congenital anomaly (among women with type 1 diabetes: 44·8 per 1000 livebirths, terminations, and fetal losses; among women with type 2 diabetes: 40·5 per 1000 livebirths, terminations, and fetal losses; p=0·17) and stillbirth (type 1 diabetes: 10·4 per 1000 livebirths and stillbirths; type 2 diabetes: 13·5 per 1000 livebirths and stillbirths; p=0·072) did not significantly differ between diabetes types, but rates of neonatal death were higher in mothers with type 2 diabetes than in those with type 1 diabetes (type 1 diabetes: 7·4 per 1000 livebirths; type 2 diabetes 11·2 per 1000 livebirths; p=0·013). Across the whole study population, independent risk factors for perinatal death (ie, stillbirth or neonatal death) were third trimester HbA1c of 6·5% (48 mmol/mol) or higher (odds ratio 3·06 [95% CI 2·16-4·33] vs HbA1c <6·5%), being in the highest deprivation quintile (2·29 [1·16-4·52] vs the lowest quintile), and having type 2 diabetes (1·65 [1·18-2·31] vs type 1 diabetes). Variations in HbA1c and LGA birthweight were associated with maternal characteristics (age, diabetes duration, deprivation, BMI) without substantial differences between maternity clinics. Our data highlight persistent adverse pregnancy outcomes in women with type 1 or type 2 diabetes. Maternal glycaemia and BMI are the key modifiable risk factors. No maternity clinics were had appreciably better outcomes than any others, suggesting that health-care system changes are needed across all clinics. None.


Consistent size-independent harvest selection on fish body shape in two recreationally exploited marine species.
Harvesting wild animals may exert size-independent selection pressures on a range of morphological, life history, and behavioral traits. Most work so far has focused on selection pressures on life history traits and body size as morphological trait. We studied here how recreational fishing selects for morphological traits related to body shape, which may correlate with underlying swimming behavior. Using landmark-based geometric morphometrics, we found consistent recreational fishing-induced selection pressures on body shape in two recreationally exploited marine fish species. We show that individuals with larger-sized mouths and more streamlined and elongated bodies were more vulnerable to passively operated hook-and-line fishing independent of the individual's body size or condition. While the greater vulnerability of individuals with larger mouth gapes can be explained by the direct physical interaction with hooks, selection against streamlined and elongated individuals could either involve a specific foraging mode or relate to underlying elevated swimming behavior. Harvesting using passive gear is common around the globe, and thus, size-independent selection on body shape is expected to be widespread potentially leaving behind individuals with smaller oral gapes and more compact bodies. This might have repercussions for food webs by altering foraging and predation.


Mindfulness-Based Stress Reduction vs Escitalopram for the Treatment of Adults With Anxiety Disorders: A Randomized Clinical Trial.
Anxiety disorders are common, highly distressing, and impairing conditions. Effective treatments exist, but many patients do not access or respond to them. Mindfulness-based interventions, such as mindfulness-based stress reduction (MBSR) are popular and can decrease anxiety, but it is unknown how they compare to standard first-line treatments. To determine whether MBSR is noninferior to escitalopram, a commonly used first-line psychopharmacological treatment for anxiety disorders. This randomized clinical trial (Treatments for Anxiety: Meditation and Escitalopram [TAME]) included a noninferiority design with a prespecified noninferiority margin. Patients were recruited between June 2018 and February 2020. The outcome assessments were performed by blinded clinical interviewer at baseline, week 8 end point, and follow-up visits at 12 and 24 weeks. Of 430 individuals assessed for inclusion, 276 adults with a diagnosed anxiety disorder from 3 urban academic medical centers in the US were recruited for the trial, and 208 completed the trial. Participants were 1:1 randomized to 8 weeks of the weekly MBSR course or the antidepressant escitalopram, flexibly dosed from 10 to 20 mg. The primary outcome measure was anxiety levels as assessed with the Clinical Global Impression of Severity scale (CGI-S), with a predetermined noninferiority margin of -0.495 points. The primary noninferiority sample consisted of 208 patients (102 in MBSR and 106 in escitalopram), with a mean (SD) age of 33 (13) years; 156 participants (75%) were female; 32 participants (15%) were African American, 41 (20%) were Asian, 18 (9%) were Hispanic/Latino, 122 (59%) were White, and 13 (6%) were of another race or ethnicity (including Native American or Alaska Native, more than one race, or other, consolidated owing to low numbers). Baseline mean (SD) CGI-S score was 4.44 (0.79) for the MBSR group and 4.51 (0.78) for the escitalopram group in the per-protocol sample and 4.49 (0.77) vs 4.54 (0.83), respectively, in the randomized sample. At end point, the mean (SD) CGI-S score was reduced by 1.35 (1.06) for MBSR and 1.43 (1.17) for escitalopram. The difference between groups was -0.07 (0.16; 95% CI, -0.38 to 0.23; P = .65), where the lower bound of the interval fell within the predefined noninferiority margin of -0.495, indicating noninferiority of MBSR compared with escitalopram. Secondary intent-to-treat analyses using imputed data also showed the noninferiority of MBSR compared with escitalopram based on the improvement in CGI-S score. Of patients who started treatment, 10 (8%) dropped out of the escitalopram group and none from the MBSR group due to adverse events. At least 1 study-related adverse event occurred for 110 participants randomized to escitalopram (78.6%) and 21 participants randomized to MBSR (15.4%). The results from this randomized clinical trial comparing a standardized evidence-based mindfulness-based intervention with pharmacotherapy for the treatment of anxiety disorders found that MBSR was noninferior to escitalopram. ClinicalTrials.gov Identifier: NCT03522844.


Mechanisms underlying the neurotoxicity induced by glyphosate-based herbicide in immature rat hippocampus: involvement of glutamate excitotoxicity.
Previous studies demonstrate that glyphosate exposure is associated with oxidative damage and neurotoxicity. Therefore, the mechanism of glyphosate-induced neurotoxic effects needs to be determined. The aim of this study was to investigate whether Roundup(®) (a glyphosate-based herbicide) leads to neurotoxicity in hippocampus of immature rats following acute (30min) and chronic (pregnancy and lactation) pesticide exposure. Maternal exposure to pesticide was undertaken by treating dams orally with 1% Roundup(®) (0.38% glyphosate) during pregnancy and lactation (till 15-day-old). Hippocampal slices from 15 day old rats were acutely exposed to Roundup(®) (0.00005-0.1%) during 30min and experiments were carried out to determine whether glyphosate affects (45)Ca(2+) influx and cell viability. Moreover, we investigated the pesticide effects on oxidative stress parameters, (14)C-α-methyl-amino-isobutyric acid ((14)C-MeAIB) accumulation, as well as glutamate uptake, release and metabolism. Results showed that acute exposure to Roundup(®) (30min) increases (45)Ca(2+) influx by activating NMDA receptors and voltage-dependent Ca(2+) channels, leading to oxidative stress and neural cell death. The mechanisms underlying Roundup(®)-induced neurotoxicity also involve the activation of CaMKII and ERK. Moreover, acute exposure to Roundup(®) increased (3)H-glutamate released into the synaptic cleft, decreased GSH content and increased the lipoperoxidation, characterizing excitotoxicity and oxidative damage. We also observed that both acute and chronic exposure to Roundup(®) decreased (3)H-glutamate uptake and metabolism, while induced (45)Ca(2+) uptake and (14)C-MeAIB accumulation in immature rat hippocampus. Taken together, these results demonstrated that Roundup(®) might lead to excessive extracellular glutamate levels and consequently to glutamate excitotoxicity and oxidative stress in rat hippocampus.


A community-wide hepatitis A outbreak in the Shoalhaven region, New South Wales.
We investigated a community-wide outbreak of hepatitis A virus infection in the Shoalhaven region of the New South Wales south coast. Epidemiological features of the outbreak suggested that transmission was predominantly person-to-person. These included: the prolonged course of the epidemic, the bimodal age-specific attack rate, the lower socioeconomic risk groups affected, and the large proportion of cases who reported prior contact with another case. Although widespread use of post-exposure immunoprophylaxis appeared to be effective in preventing symptomatic infection in individuals, it did not rapidly halt the outbreak. We review methods of mass intervention for community-wide outbreaks of hepatitis A virus infection, and define priorities for investigation of future outbreaks.


Machine learning-based classification of viewing behavior using a wide range of statistical oculomotor features.
Since the seminal work of Yarbus, multiple studies have demonstrated the influence of task-set on oculomotor behavior and the current cognitive state. In more recent years, this field of research has expanded by evaluating the costs of abruptly switching between such different tasks. At the same time, the field of classifying oculomotor behavior has been moving toward more advanced, data-driven methods of decoding data. For the current study, we used a large dataset compiled over multiple experiments and implemented separate state-of-the-art machine learning methods for decoding both cognitive state and task-switching. We found that, by extracting a wide range of oculomotor features, we were able to implement robust classifier models for decoding both cognitive state and task-switching. Our decoding performance highlights the feasibility of this approach, even invariant of image statistics. Additionally, we present a feature ranking for both models, indicating the relative magnitude of different oculomotor features for both classifiers. These rankings indicate a separate set of important predictors for decoding each task, respectively. Finally, we discuss the implications of the current approach related to interpreting the decoding results.


The social/sexual environment of gay men residing in a rural frontier state: implications for the development of HIV prevention programs.
The incidence and prevalence of HIV/AIDS is increasing among rural men who have sex with men (MSM). Yet little is known about the social/ sexual environment of rural frontier areas. The purpose of this study was to assess the social/sexual environment of gay men living in rural areas and how this environment contributes to the development of HIV/AIDS prevention programs. A qualitative study was conducted in Wyoming. In-depth guided interviews were conducted with 39 self-identified gay men. Data were analyzed for emergent themes using constant comparative analysis. Four broadly related themes emerged. Participants perceive that they live in a hostile social environment in which the potential for becoming a target of violence is present. In order to cope with this social reality, men adopt strategies to assimilate into the predominant heterosexual culture and to look for sex partners. These, in turn, are related to their attitudes about HIV/AIDS and prevention activities. Notably, the Internet was discussed by participants as a means for men to connect to a larger gay society and look for sex partners and as a potential venue to HIV/AIDS prevention programs. Data provided a number of implications for developing HIV/AIDS prevention programs targeting rural MSM. Especially apparent was the need for programs to be mindful of the desire to keep one's sexual preferences shielded from public knowledge and the effect this may have on recruiting rural MSM to participate in prevention activities. The Internet, because men can access it privately, might provide a venue for prevention projects.


A glycosylation-dependent CD45RB epitope defines previously unacknowledged CD27⁻IgM(high) B cell subpopulations enriched in young children and after hematopoietic stem cell transplantation.
The immune system is dysfunctional for years after hematopoietic stem cell transplantation (HSCT). A potential cause is an intrinsic B cell deficiency. In a cohort of pediatric HSCT patients few CD27(+) B cells formed after transplantation with the number of CD27(+)IgM(high) cells more affected than class-switched ones. A previously unacknowledged population of CD27(-)IgM(high) cells made up the majority of B cells and this population was also enlarged in healthy children compared to adults. Only a minority of these CD27(-)IgM(high) B cells expressed markers typical for transitional B cells, and the non-transitional CD27(-)IgM(high) cells could be further divided into subpopulations based on their ability to extrude the dye Rhodamine 123 and their expression of CD45RB(MEM55), a glycosylation-dependent epitope. Thus, we define several novel human CD27(-)IgM(high) B cell subpopulations in blood, all of which are present in higher frequencies and numbers in young children and after HSCT than in adults.


Domain structure of a NHEJ DNA repair ligase from Mycobacterium tuberculosis.
A prokaryotic non-homologous end-joining (NHEJ) system for the repair of DNA double-strand breaks (DSBs), composed of a Ku homodimer (Mt-Ku) and a multidomain multifunctional ATP-dependent DNA ligase (Mt-Lig), has been described recently in Mycobacterium tuberculosis. Mt-Lig exhibits polymerase and nuclease activity in addition to DNA ligation activity. These functions were ascribed to putative polymerase, nuclease and ligase domains that together constitute a monomeric protein. Here, the separate polymerase, nuclease and ligase domains of Mt-Lig were cloned individually, over-expressed and the soluble proteins purified to homogeneity. The polymerase domain demonstrated DNA-dependent RNA primase activity, catalysing the synthesis of unprimed oligoribonucleotides on single-stranded DNA templates. The polymerase domain can also extend DNA in a template-dependent manner. This activity was eliminated when the catalytic aspartate residues were replaced with alanine. The ligase domain catalysed the sealing of nicked double-stranded DNA designed to mimic a DSB, consistent with the role of Mt-Lig in NHEJ. Deletion of the active-site lysine residue prevented the formation of an adenylated ligase complex and consequently thwarted ligation. The nuclease domain did not function independently as a 3'-5' exonuclease. DNA-binding assays revealed that both the polymerase and ligase domains bind DNA in vitro, the latter with considerably higher affinity. Mt-Ku directly stimulated the polymerase and nuclease activities of Mt-Lig. The polymerase domain bound Mt-Ku in vitro, suggesting it may recruit Mt-Lig to Ku-bound DNA in vivo. Consistent with these data, Mt-Ku stimulated the primer extension activity of the polymerase domain, suggestive of a functional interaction relevant to NHEJ-mediated DSB repair processes.


Acute gastric changes in patients with acute stroke. Part 1: with reference to gastroendoscopic findings.
A gastroendoscopic study was performed on 177 patients with acute stroke. Gastric changes were found in 92 among them (52%), including ten with acute ulcer. A high frequency of gastric changes was found in patients with serious stroke and/or in patients whose cerebral lesions were located close to the hypothalamus or its centrifugal tract. The mortality in patients with gastric changes was high, especially in patients with acute ulcers, multiple erosions and petechiae. The frequency of brown gastric juice and coffee grounds at gastroendoscopic examination (gastric hemorrhage) was three times as high as that of melena and/or hematoemesis. On the other hand, the mortality of patients with gastric hemorrhage differed little from that of patients with melena and/or hematoemesis.


Specific binding of soluble fibrin to macrophages.
Guinea pig peritoneal macrophages were demonstrated to bind selectively soluble 125I-fibrin and fibrin/fibrinogen complexes as compared with fibrinogen, fibrinogen degradation products, and fibrin degradation products. Cellular uptake was considered to be surface receptor binding on the basis of removal of bound 125I-fibrin by trypsin and because uptake occurred in the presence of metabolic inhibitors. 125I-fibrin uptake could be blocked by nonradioactive fibrin but not by IgG or immune complexes. Binding was uneffected by prior treatment with plasmin or trypsin but was calcium dependent. Only limited reversibility of binding could be demonstrated after prolonged incubation. Scatchard plots permitted an estimate of the number of bound molecules. At saturation 6.92 X 10(6) 125I-fibrin molecules were bound per cell. Similar binding of fibrin was noted in polymorphonuclear leukocytes, but not lymphocytes or fibroblasts. Soluble fibrin binding may be a host defense mechanism whereby the reticuloendothelial system can remove fibrin from the blood before the development of microthrombi.


The spatiotemporal dynamics of Tribolium castaneum (Herbst): adult flight and gene flow.
Tribolium castaneum (Herbst) has been used as a model organism to develop and test important ecological and evolutionary concepts and is also a major pest of grain and grain products globally. This beetle species is assumed to be a good colonizer of grain storages through anthropogenic movement of grain, and active dispersal by flight is considered unlikely. Studies using T. castaneum have therefore used confined or walking insects. We combine an ecological study of dispersal with an analysis of gene flow using microsatellites to investigate the spatiotemporal dynamics and adult flight of T. castaneum in an ecological landscape in eastern Australia. Flying beetles were caught in traps at grain storages and in fields at least 1 km from the nearest stored grain at regular intervals for an entire year. Significantly more beetles were trapped at storages than in fields, and almost no beetles were caught in native vegetation reserves many kilometres from the nearest stored grain. Genetic differentiation between beetles caught at storages and in fields was low, indicating that although T. castaneum is predominantly aggregated around grain storages, active dispersal takes place to the extent that significant gene flow occurs between them, mitigating founder effects and genetic drift. By combining ecological and molecular techniques, we reveal much higher levels of active dispersal through adult flight in T. castaneum than previously thought. We conclude that the implications of adult flight to previous and future studies on this model organism warrant consideration.


TMC125 displays a high genetic barrier to the development of resistance: evidence from in vitro selection experiments.
TMC125 is a potent new investigational nonnucleoside reverse transcriptase inhibitor (NNRTI) that is active against human immunodeficiency virus type 1 (HIV-1) with resistance to currently licensed NNRTIs. Sequential passage experiments with both wild-type virus and NNRTI-resistant virus were performed to identify mutations selected by TMC125 in vitro. In addition to "classic" selection experiments at a low multiplicity of infection (MOI) with increasing concentrations of inhibitors, experiments at a high MOI with fixed concentrations of inhibitors were performed to ensure a standardized comparison between TMC125 and current NNRTIs. Both low- and high-MOI experiments demonstrated that the development of resistance to TMC125 required multiple mutations which frequently conferred cross-resistance to efavirenz and nevirapine. In high-MOI experiments, 1 muM TMC125 completely inhibited the breakthrough of resistant virus from wild-type and NNRTI-resistant HIV-1, in contrast to efavirenz and nevirapine. Furthermore, breakthrough of virus from site-directed mutant (SDM) SDM-K103N/Y181C occurred at the same time or later with TMC125 as breakthrough from wild-type HIV-1 with efavirenz or nevirapine. The selection experiments identified mutations selected by TMC125 that included known NNRTI-associated mutations L100I, Y181C, G190E, M230L, and Y318F and the novel mutations V179I and V179F. Testing the antiviral activity of TMC125 against a panel of SDMs indicated that the impact of these individual mutations on resistance was highly dependent upon the presence and identity of coexisting mutations. These results demonstrate that TMC125 has a unique profile of activity against NNRTI-resistant virus and possesses a high genetic barrier to the development of resistance in vitro.


Proposal of a novel MELD exception point system for hepatocellular carcinoma based on tumor characteristics and dynamics.
Patients listed with exception points for hepatocellular carcinoma (HCC) have been more likely to be transplanted than those listed for chronic liver failure (LF) based on the model for end-stage liver disease (MELD) score. The aim of this study was to determine outcomes in the 5-year experience of a scoring system designed to reflect heterogeneity of tumor load of patients listed for HCC. A novel MELD exception point system based on size and number of HCC was implemented in July 2009. This system allows stratification of patients based on risk of dropping out from the waiting list according to Milan criteria. LF patients were listed according to biological MELD sodium score; HCC patients were reassigned points every three months upon repeat imaging. Among 624 patients listed for liver transplant (LT), 505 were eligible. 94 (18.6%) were assigned MELD HCC points. Only 24.7% required changes in allocated points over time. Transplantation rates (HCC 83% vs. LF 73%, p=0.04) and waiting time in days (HCC 258 vs. LF 325; p=0.07) were similar. The method of competing risk analysis revealed that HCC patients were more likely to be transplanted than LF during the 5-year period preceding implementation, whereas transplant rates became equivalent for HCC and non-HCC in 2009-2014. One- and two-year survivals were similar between the two groups. Our study demonstrates that a novel MELD point system for HCC, taking into account dynamics in tumor size and number, allows for equitable liver allocation without compromising graft and patient survival. It has historically been difficult to achieve equitable liver allocation for liver cancer and chronic liver failure with the allocation systems currently in place in many countries worldwide. We designed a new system to help improve access to organs for liver failure patients in Québec, Canada. Our 5-year experience demonstrates that this unique system renders access to transplant similar for both liver cancer and liver failure indications.


Numerical correction of detector channel cross-talk using full-spectrum fluorescence correlation spectroscopy.
Fluorescence correlation spectroscopy (FCS) uses fluctuations in the fluorescence collected from a small illuminated volume to measure dynamic processes of fluorophores. In traditional FCS, spectral overlap produces cross-talk in dedicated detector channels, undermining the accuracy of measurements of molecular interactions. Here, the experimental realization of full-spectrum fluorescence correlation spectroscopy is described and coupled with multivariate data analysis to numerically correct detector cross-talk, isolating spectra and fluctuation traces of mixture components in spite of overlap. Application of this methodology is illustrated using the measurement of the diffusion constant of labeled polystyrene in hydroxypropyl cellulose in the presence of a persistent dye. Additionally, the results show that full-spectrum FCS with multivariate analysis can isolate and characterize signals from unanticipated sample components.


Dispersion-free pulse propagation in a negative-index material.
The possibility of controlling the spectral position of the zero group-velocity dispersion point of a negative-index material can be exploited by varying the ratio between the electric and the magnetic plasma frequency to obtain dispersion-free propagation in spectral regions otherwise inaccessible using conventional positive-index materials. Our predictions are confirmed by pulse propagation simulations where all the orders of the complex dispersion of the material are taken into account.


Down-Regulation of miRNA-128 Contributes to Neuropathic Pain Following Spinal Cord Injury via Activation of P38.
BACKGROUND Neuropathic pain (NPP) arises from a lesion or dysfunction of the somatosensory nervous system. Recent studies have demonstrated multiple microRNAs (miRNAs) play key roles in NPP development. This study aimed to investigate the effects of miR-128 on microglial cells. MATERIAL AND METHODS We established a compressive spinal cord injury (SCI) model and collected the spinal cord segment-derived conditioned medium (CM). We then measured the expression of miR-128 in the murine microglial cell line BV2 treated with CM-SCI or CM obtained from control (CM-NC). Furthermore, lentivirus production of miR-128 and scrambled control were transfected into BV2 cells, which were first treated with CM-SCI or CM-NC. Moreover, the effects of miR-128 on cell viability, M1/M2 microglial gene expression, inflammatory cytokines concentration, and the protein expression of P38 and phosphorylated P38 (P-P38) were investigated. RESULTS The expression of miR-128 was downregulated in murine microglial BV2 cells treated with CM-SCI. Overexpression of miR-128 markedly promoted the viability of murine microglial cells. In addition, miR-128 overexpression significantly decreased the expression levels of microglial M1 phenotypic markers CD86 and CD32, and increased the expression levels of M2 phenotypic markers Arg1 and CD206. Furthermore, miR-128 overexpression obviously decreased the concentration of TNF-α, IL-1β, and IL-6. We found that miR-128 overexpression significantly downregulated the expression levels of P38 andP-P38. CONCLUSIONS Our findings indicate that down-regulation of miR-128 in murine microglial cells may contribute to the development of NPP following SCI via activation of P38. MiR-128 may be a potential intervention target for NPP.