Robot-assisted low anterior resection for situs inversus totalis: a novel technical approach for an uncommon condition.
Situs inversus totalis (SIT) is an uncommon condition, with an incidence of 1 in 10,000. Surgery for SIT patients is more difficult because of the uncommon anatomy. Experience in laparoscopic surgery for patients with SIT is very limited. Only a few cases of laparoscopic colorectal resections have been reported in the literature. We present the first robot-assisted low anterior resection for rectal cancer in a patient with SIT. A 70-year-old woman with SIT who presented with rectal bleeding underwent a colonoscopy and barium enema. This workup revealed a rectal cancer 10 cm from the anal verge. The magnetic resonance imaging scan revealed a T3/4 tumor in the rectum with perirectal lymph node involvement, whereas the computed tomography positron emission tomography scan did not reveal any distal metastasis. She underwent neoadjuvant chemoradiotherapy 6 weeks before surgery. Postoperatively, she made an uneventful recovery and was discharged on day 6. After laparoscopic examination and displacement of the small bowel, 4 robot trocars were inserted into 4 quadrants of the abdomen. A fifth port was inserted and used by the assistant. The robot cart was docked from the right side with arms 1, 2, and 3 in the right upper quadrant (Cadiere grasper), left lower quadrant (bipolar Maryland grasper), and left upper quadrant (monopolar scissors), respectively, for colonic mobilization without splenic flexure takedown. For pelvic dissection, arms 1 and 3 were moved to the right upper quadrant and right lower quadrant, respectively. After adequate pelvic dissection, the robot cart was undocked, and a laparoscopic articulating linear stapler was used to transect the rectum from the left lower quadrant port. Bowel continuity was restored with a circular stapler. A loop ileostomy was created through the extraction site in the right lower quadrant. Robot-assisted low anterior resection for SIT patients can be performed safely and confers the benefits of laparoscopic low anterior resection with additional advantages unique to the da Vinci system.


MPV17 encodes an inner mitochondrial membrane protein and is mutated in infantile hepatic mitochondrial DNA depletion.
The mitochondrial (mt) DNA depletion syndromes (MDDS) are genetic disorders characterized by a severe, tissue-specific decrease of mtDNA copy number, leading to organ failure. There are two main clinical presentations: myopathic (OMIM 609560) and hepatocerebral (OMIM 251880). Known mutant genes, including TK2, SUCLA2, DGUOK and POLG, account for only a fraction of MDDS cases. We found a new locus for hepatocerebral MDDS on chromosome 2p21-23 and prioritized the genes on this locus using a new integrative genomics strategy. One of the top-scoring candidates was the human ortholog of the mouse kidney disease gene Mpv17. We found disease-segregating mutations in three families with hepatocerebral MDDS and demonstrated that, contrary to the alleged peroxisomal localization of the MPV17 gene product, MPV17 is a mitochondrial inner membrane protein, and its absence or malfunction causes oxidative phosphorylation (OXPHOS) failure and mtDNA depletion, not only in affected individuals but also in Mpv17-/- mice.


Investigating the time-lapsed effects of rigid cervical collars on the dimensions of the internal jugular vein.
Protocol advocates the use of rigid cervical collars (RCCs) in head trauma patients as they are at risk of concomitant cervical spine injury. Literature has shown RCCs to be a potential cause of venous outflow obstruction, changing internal jugular vein (IJV) cross-sectional area (CSA), and raising intracranial pressure (ICP). This study aims to investigate the effects of applying a RCC, for a period of four hours, on the dimensions of the IJV, in healthy participants. Seventeen participants (nine male, eight female) took part in this study. Circumference and CSAs of the IJV were measured bilaterally by a single observer using a GE LOGIQ e ultrasound system. Measurements were taken pre-RCC application, immediately after, every hour over four hours, and five minutes postcollar removal. The CSA of the IJV was 8.3 ±6.0 mm2 pre-RCC application. The CSA of the IJV doubled (18.92 ±10.55 mm2 ) after four hours and decreased back to 9.36 ±6.8 mm2 five minutes postcollar removal. The circumference of the IJV was 17.29 ±6.03 mm pre-RCC application, increasing to 20.34 ±5.59 mm by the end of the fourth hour and returning to 16.14 ±5.16 mm five minutes postcollar removal. Related-samples Friedman's ANOVA test showed statistically significant differences for both left and right CSAs and circumferences of the IJV measured across the four hours (P-value<0.05). Ultrasound assessment of CSA of the IJV may correlate with changes in ICP. Further studies may provide insight into the effects of collar design, and guide future trauma protocol to minimize intracranial pressure fluctuations. Clin. Anat. 32:196-200, 2019. © 2018 Wiley Periodicals, Inc.


Evidence for complex genic interactions between conspecific chromosomes underlying hybrid female sterility in the Drosophila simulans clade.
F1 hybrid females between the sibling species Drosophila simulans, Drosophila mauritiana and Drosophila sechellia are completely fertile. However, we have found that female sterility can be observed in F2 backcross females who are homozygous for D. simulans X chromosomes and homozygous for autosomal regions from either D. mauritiana or D. sechellia. Our results indicate that neither D. mauritiana autosome (2 or 3) can cause complete female sterility in a D. simulans background. The simultaneous presence of homozygous regions from both the second and third chromosomes of D. mauritiana, however, causes nearly complete female sterility which cannot be accounted for by their individual effects. The two autosomes of D. sechellia may show a similar pattern. From the same crosses, we also obtained evidence against a role for cytoplasmic or maternal effects in causing hybrid male sterility between these species. Taken with the results presented elsewhere, these observations suggest that epistatic interactions between conspecific genes in a hybrid background may be the prevalent mode of hybrid sterility between recently diverged species.


Positive association between hepatitis C infection and oral cavity cancer: a nationwide population-based cohort study in Taiwan.
The association between viral hepatitis (B and C) and oral cavity cancer has been widely debated. This nationwide, population-based cohort study assessed the subsequent risk of oral cavity cancer among patients with chronic viral hepatitis infection. Data were retrieved from insurance claims data of 1,000,000 randomly sampled individuals covered under the Taiwan National Health Insurance system. We identified a total of 21,199 adults with chronic viral hepatitis infection (12,369 with HBV alone, 5,311 with HCV alone, and 3,519 with HBV/HCV dual infections) from 2000-2005. Comparison group comprised 84,796 sex- and age-matched subjects without viral hepatitis during the same study period. Incidence and risk of subsequent oral cavity cancer were measured until 2008. The incidence of oral cavity cancers was 2.28-fold higher among patients with HCV alone than non-viral hepatitis group (6.15 versus 2.69 per 10,000 person-years). After adjusting for sociodemographic covariates, HCV alone was significantly associated with an increased risk for oral cavity cancer (hazard ratio (HR) = 1.90, 95% confidence interval (CI) = 1.20-3.02). This positive association was highest among individuals in the 40-49-year age group (HR = 2.57, 95% CI = 1.21-5.46). However, there were no significant associations between HBV alone or HBV/HCV dual infections and risk for oral cavity cancer. Our data suggest that HCV but not HBV infection is a risk factor for oral cavity cancer. In addition, subjects with HCV infection tend to be at early onset risk for oral cavity cancer. This finding needs to be replicated in further studies.


Atmospheric carbon dioxide, the southern oscillation, and the weak 1975 el nino.
The observed rate of change of the atmospheric carbon dioxide concentration at the South Pole, Fanning Island, Hawaii, and ocean weather station P correlates with an index of the southern oscillation and with El Niño occurrences. There are changes at all four stations that seem to be in response to the weak 1975 El Niño. Thus, even poorly developed El Niño events may affect the atmospheric carbon dioxide concentration.


Genetic and fitness changes accompanying adaptation of an arbovirus to vertebrate and invertebrate cells.
The alternating host cycle and persistent vector infection may constrain the evolution of arboviruses. To test this hypothesis, eastern equine encephalitis virus was passaged in BHK or mosquito cells, as well as in alternating (both) host cell passages. High and low multiplicities were used to examine the effect of defective interfering particles. Clonal BHK and persistent mosquito cell infections were also evaluated. Fitness was measured with one-step growth curves and competition assays, and mutations were evaluated by nucleotide sequencing and RNA fingerprinting. All passages and assays were done at 32 degrees C to eliminate temperature as a selection factor. Viruses passaged in either cell type alone exhibited fitness declines in the bypassed cells, while high-multiplicity and clonal passages caused fitness declines in both types of cells. Bypassed cell fitness losses were mosquito and vertebrate specific and were not restricted to individual cell lines. Fitness increases occurred in the cell line used for single-host-adaptation passages and in both cells for alternately passaged viruses. Surprisingly, single-host-cell passage increased fitness in that cell type no more than alternating passages. However, single-host-cell adaptation resulted in more mutations than alternating cell passages. Mosquito cell adaptation invariably resulted in replacement of the stop codon in nsP3 with arginine or cysteine. In one case, BHK cell adaptation resulted in a 238-nucleotide deletion in the 3' untranslated region. Many nonsynonymous substitutions were shared among more than one BHK or mosquito cell passage series, suggesting positive Darwinian selection. Our results suggest that alternating host transmission cycles constrain the evolutionary rates of arboviruses but not their fitness for either host alone.


Vaccination with the immediate-early protein ICP47 of herpes simplex virus-type 1 (HSV-1) induces virus-specific lymphoproliferation, but fails to protect against lethal challenge.
Assessing the immunobiological function of the individual proteins of herpes simplex virus-type 1 (HSV-1) continues to be important in elucidating virus-host interactions and for the rational design of subunit vaccines. In this report, the non-structural, immediate-early protein ICP47 of HSV-1 was examined for its ability to induce virus-specific immune responses. The ICP47 protein, when expressed from a recombinant vaccinia virus or when produced by cell-free, in vitro translation, induced a vigorous HSV-1-specific lymphoproliferative response. However, other common parameters of immunity such as neutralizing antibody, delayed-type hypersensitivity, and class I major histocompatibility complex (MHC)-restricted cytotoxic T lymphocytes (CTL) were not induced by ICP47. Moreover, mice immunized with vaccinia-expressed ICP47 were unable to survive lethal challenge with virulent HSV, indicating that in spite of its ability to induce significant HSV-1-specific lymphoproliferation, ICP47 appears unable to afford protective immunity in vivo. Possible reasons for this failure and the implications of these results in terms of vaccine design are discussed.


Proteolytic processing of the replicase ORF1a protein of equine arteritis virus.
To study the proteolytic processing of the equine arteritis virus (EAV) replicase open reading frame 1a (ORF1a) protein, specific antisera were raised in rabbits, with six synthetic peptides and a bacterial fusion protein as antigens. The processing of the EAV ORF1a product in infected cells was analyzed with Western blot (immunoblot) and immunoprecipitation techniques. Additional information was obtained from transient expression of ORF1a cDNA constructs. The 187-kDa ORF1a protein was found to be subject to at least five proteolytic cleavages. The processing scheme, which covers the entire ORF1a protein, results in cleavage products of approximately 29, 61, 22, 31, 41, and 3 kDa, which were named nonstructural proteins (nsps) 1 through 6, respectively. Pulse-chase experiments revealed that the cleavages at the nsp1/2 and nsp2/3 junctions are the most rapid processing steps. The remaining nsp3456 precursor is first cleaved at the nsp4/5 site. Final processing of the nsp34 and nsp56 intermediates is extremely slow. As predicted from previous in vitro translation experiments (E. J. Snijder, A. L. M. Wassenaar, and W. J. M. Spaan, J. Virol. 66:7040-7048, 1992), a cysteine protease domain in nsp1 was shown to be responsible for the nsp1/2 cleavage. The other processing steps are carried out by the putative EAV serine protease in nsp4 and by a third protease, which remains to be identified.


Ca2+-dependent and Ca2+-independent degradation of phosphatidylinositol in rabbit vas deferens.
The effects of Ca2+ and acetylcholine on the degradation and synthesis of phosphatidylinositol in rabbit vas deferens was studied in vitro by a pulse--chase technique and by measuring the content of the phospholipid in the tissue. Ca2+-dependent degradation of phosphatidylinositol was found in slices and homogenates prelabelled with myo-[2-3H]inositol. The phosphatidylinositol content of the slices also decreased by a Ca2+-dependent mechanism. On the other hand, removal of intracellular Ca2+ with the ionophore A23187 and EGTA increased the amount of phosphatidylinositol. These results indicate that the intracellular Ca2+ concentration has an important role in regulating the phosphatidylinositol content of the tissue. Increasing the extracellular K+ concentration, which causes an increase in plasma-membrane Ca2+ permeability, did not enhance phosphatidylinositol breakdown nor decrease its tissue content. However, phosphatidylinositol synthesis was clearly inhibited. After stimulation of the smooth muscle with acetylcholine, degradation of phosphatidylinositol was enhanced. Furthermore, the content of phosphatidylinositol in the tissue also decreased. These phenomena were evident even in the absence of Ca2+. The acetylcholine-induced degradation of phosphatidylinositol was blocked by the muscarinic antagonist atropine, but not by the nicotinic antagonist (+)-tubocurarine. The acetylcholine-induced decrease in the phosphatidylinositol content of the tissue led to the compensatory synthesis of phosphatidylinositol. Synthesis was separated from degradation in the same tissue. Compensatory synthesis was inhibited by acetylcholine. The degradation of phosphatidylinositol induced by acetylcholine was not inhibited by 8-bromoguanosine 3':5'-cyclic monophosphate, indicating that the degradative process was not mediated by an increase in the cyclic nucleotide.


Structural features of the rice chromosome 4 centromere.
A complete sequence of a chromosome centromere is necessary for fully understanding centromere function. We reported the sequence structures of the first complete rice chromosome centromere through sequencing a large insert bacterial artificial chromosome clone-based contig, which covered the rice chromosome 4 centromere. Complete sequencing of the 124-kb rice chromosome 4 centromere revealed that it consisted of 18 tracts of 379 tandemly arrayed repeats known as CentO and a total of 19 centromeric retroelements (CRs) but no unique sequences were detected. Four tracts, composed of 65 CentO repeats, were located in the opposite orientation, and 18 CentO tracts were flanked by 19 retroelements. The CRs were classified into four types, and the type I retroelements appeared to be more specific to rice centromeres. The preferential insert of the CRs among CentO repeats indicated that the centromere-specific retroelements may contribute to centromere expansion during evolution. The presence of three intact retrotransposons in the centromere suggests that they may be responsible for functional centromere initiation through a transcription-mediated mechanism.


Standard guidelines for the chromosome-centric human proteome project.
The objective of the international Chromosome-Centric Human Proteome Project (C-HPP) is to map and annotate all proteins encoded by the genes on each human chromosome. The C-HPP consortium was established to organize a collaborative network among the research teams responsible for protein mapping of individual chromosomes and to identify compelling biological and genetic mechanisms influencing colocated genes and their protein products. The C-HPP aims to foster the development of proteome analysis and integration of the findings from related molecular -omics technology platforms through collaborations among universities, industries, and private research groups. The C-HPP consortium leadership has elicited broad input for standard guidelines to manage these international efforts more efficiently by mobilizing existing resources and collaborative networks. The C-HPP guidelines set out the collaborative consensus of the C-HPP teams, introduce topics associated with experimental approaches, data production, quality control, treatment, and transparency of data, governance of the consortium, and collaborative benefits. A companion approach for the Biology and Disease-Driven HPP (B/D-HPP) component of the Human Proteome Project is currently being organized, building upon the Human Proteome Organization's organ-based and biofluid-based initiatives (www.hupo.org/research). The common application of these guidelines in the participating laboratories is expected to facilitate the goal of a comprehensive analysis of the human proteome.


Laser desorption ionization mass spectrometry of protein tryptic digests on nanostructured silicon plates.
We report on the simple application of a new nanostructured silicon (NanoSi) substrate as laser desorption/ionization (LDI)-promoting surface for high-throughput identification of protein tryptic digests by a rapid MS profiling and subsequent MS/MS analysis. The NanoSi substrate is easily prepared by chemical etching of crystalline silicon in NH(4)F/HNO(3)/AgNO(3) aqueous solution. To assess the LDI performances in terms of sensitivity, repeatability and robustness, the detection of small synthetic peptides (380-1700Da) was investigated. Moreover, peptide sequencing was tackled. Various tryptic synthetic peptide mixtures were first characterized in MS and MS/MS experiments carried out on a single deposit. Having illustrated the capability to achieve peptide detection and sequencing on these ionizing surfaces in the same run, protein tryptic digests from Cytochrome C, β-Casein, BSA and Fibrinogen were then analyzed in the femtomolar range (from 50 fmol for Cytochrome C down to 2 fmol for Fibrinogen). Comparison of the NanoSi MS and MS/MS data with those obtained with sample conditioned in organic matrix demonstrated a great behavior for low mass responses. We demonstrated the capability of LDI on NanoSi to be a complementary method to MALDI peptide mass fingerprinting ensuring determination of peptide molecular weights and sequences for more efficient protein database searches.


A molecular classifier for predicting future graft loss in late kidney transplant biopsies.
Kidney transplant recipients that develop signs of renal dysfunction or proteinuria one or more years after transplantation are at considerable risk for progression to renal failure. To assess the kidney at this time, a "for-cause" biopsy is performed, but this provides little indication as to which recipients will go on to organ failure. In an attempt to identify molecules that could provide this information, we used microarrays to analyze gene expression in 105 for-cause biopsies taken between 1 and 31 years after transplantation. Using supervised principal components analysis, we derived a molecular classifier to predict graft loss. The genes associated with graft failure were related to tissue injury, epithelial dedifferentiation, matrix remodeling, and TGF-beta effects and showed little overlap with rejection-associated genes. We assigned a prognostic molecular risk score to each patient, identifying those at high or low risk for graft loss. The molecular risk score was correlated with interstitial fibrosis, tubular atrophy, tubulitis, interstitial inflammation, proteinuria, and glomerular filtration rate. In multivariate analysis, molecular risk score, peritubular capillary basement membrane multilayering, arteriolar hyalinosis, and proteinuria were independent predictors of graft loss. In an independent validation set, the molecular risk score was the only predictor of graft loss. Thus, the molecular risk score reflects active injury and is superior to either scarring or function in predicting graft failure.


Refined structure of the nicotinic acetylcholine receptor at 4A resolution.
We present a refined model of the membrane-associated Torpedo acetylcholine (ACh) receptor at 4A resolution. An improved experimental density map was obtained from 342 electron images of helical tubes, and the refined structure was derived to an R-factor of 36.7% (R(free) 37.9%) by standard crystallographic methods, after placing the densities corresponding to a single molecule into an artificial unit cell. The agreement between experimental and calculated phases along the helical layer-lines was used to monitor progress in the refinement and to give an independent measure of the accuracy. The atomic model allowed a detailed description of the whole receptor in the closed-channel form, including the ligand-binding and intracellular domains, which have not previously been interpreted at a chemical level. We confirm that the two ligand-binding alpha subunits have a different extended conformation from the three other subunits in the closed channel, and identify several interactions on both pairs of subunit interfaces, and within the alpha subunits, which may be responsible for their "distorted" structures. The ACh-coordinating amino acid side-chains of the alpha subunits are far apart in the closed channel, indicating that a localised rearrangement, involving closure of loops B and C around the bound ACh molecule, occurs upon activation. A comparison of the structure of the alpha subunit with that of AChBP having ligand present, suggests how the localised rearrangement overcomes the distortions and initiates the rotational movements associated with opening of the channel. Both vestibules of the channel are strongly electronegative, providing a cation-stabilising environment at either entrance of the membrane pore. Access to the pore on the intracellular side is further influenced by narrow lateral windows, which would be expected to screen out electrostatically ions of the wrong charge and size.


Fish muscle energy metabolism measured by in vivo 31P-NMR during anoxia and recovery.
By means of in vivo 31P nuclear magnetic resonance (NMR) we measured energy stores and intracellular pH at 10-min intervals in the myotome of unanesthetized carp and goldfish before, during, and after a period of anoxia (1 h for carp and 4 h for goldfish). The fish were mounted in a modified bioprobe, and their gills were irrigated with a constant flow of aerated or anoxic water. Anoxia caused a steep decline of phosphocreatine and intracellular pH in carp muscle. After the phosphocreatine stores had been exhausted by greater than 85%, [ATP] fell, whereas IMP and phosphodiesters accumulated. In goldfish muscle, initial changes followed the same pattern, but after 20 min a steady state of high-energy phosphates was reached and the development of acidosis was dampened. The resistance of goldfish to anoxia is due to metabolic suppression and a switch from lactate to ethanol and CO2 as the anaerobic end products. In both species, recovery was complete within 3 h. The fast pH recovery seems to be mainly caused by H+ and lactic acid efflux.


Pancreatic eukaryotic initiation factor-2alpha kinase (PEK) homologues in humans, Drosophila melanogaster and Caenorhabditis elegans that mediate translational control in response to endoplasmic reticulum stress.
In response to different cellular stresses, a family of protein kinases regulates translation by phosphorylation of the alpha subunit of eukaryotic initiation factor-2 (eIF-2alpha). Recently, we identified a new family member, pancreatic eIF-2alpha kinase (PEK) from rat pancreas. PEK, also referred to as RNA-dependent protein kinase (PKR)-like endoplasmic reticulum (ER) kinase (PERK) is a transmembrane protein implicated in translational control in response to stresses that impair protein folding in the ER. In this study, we identified and characterized PEK homologues from humans, Drosophila melanogaster and Caenorhabditis elegans. Expression of human PEK mRNA was found in over 50 different tissues examined, with highest levels in secretory tissues. In mammalian cells subjected to ER stress, we found that elevated eIF-2alpha phosphorylation was coincident with increased PEK autophosphorylation and eIF-2alpha kinase activity. Activation of PEK was abolished by deletion of PEK N-terminal sequences located in the ER lumen. To address the role of C. elegans PEK in translational control, we expressed this kinase in yeast and found that it inhibits growth by hyperphosphorylation of eIF-2alpha and inhibition of eIF-2B. Furthermore, we found that vaccinia virus K3L protein, an inhibitor of the eIF-2alpha kinase PKR involved in an anti-viral defence pathway, also reduced PEK activity. These results suggest that decreased translation initiation by PEK during ER stress may provide the cell with an opportunity to remedy the folding problem prior to introducing newly synthesized proteins into the secretory pathway.


The Ski proto-oncogene regulates body composition and suppresses lipogenesis.
The Ski gene regulates skeletal muscle differentiation in vitro and and in vivo. In the c-Ski overexpression mouse model there occurs marked skeletal muscle hypertrophy with decreased adipose tissue mass. In this study, we have investigated the underlying molecular mechanisms responsible for the increased skeletal muscle and decreased adipose tissue mass in the c-Ski mouse. Growth and body composition analysis (tissue weights and dual energy X-ray absorptiometry) coupled with skeletal muscle and white adipose gene expression and metabolic phenotyping in c-Ski mice and wild-type (WT) littermate controls was performed. The growth and body composition studies confirmed the early onset of accelerated body growth, with increased lean mass and decreased fat mass in the c-Ski mice. Gene expression analysis in skeletal muscle from c-Ski mice compared with WT mice showed significant differences in myogenic and lipogenic gene expressions that are consistent with the body composition phenotype. Skeletal muscle of c-Ski mice had significantly repressed Smad1, 4, 7 and myostatin gene expression and elevated myogenin, myocyte enhancer factor 2, insulin-like growth factor-1 receptor and insulin-like growth factor-2 expression. Strikingly, expression of the mRNAs encoding the master lipogenic regulators, sterol-regulatory enhancer binding protein 1c (SREBP1c), and the nuclear receptor liver X-receptor-alpha, and their downstream target genes, SCD-1 and FAS, were suppressed in skeletal muscle of c-Ski mice, as were the expressions of other nuclear receptors involved in adipogenesis and metabolism, such as peroxisome proliferator-activated receptor-gamma, glucocorticoid receptor and retinoic acid receptor-related orphan receptor-alpha. Transfection analysis demonstrated Ski repressed the SREBP1c promoter. Moreover, palmitate oxidation and oxidative enzyme activity was increased in skeletal muscle of c-Ski mice. These results suggest that the Ski phenotype involves attenuated lipogenesis, decreased myostatin signalling, coupled to increased myogenesis and fatty acid oxidation. Ski regulates several genetic programs and signalling pathways that regulate skeletal muscle and adipose mass to influence body composition development, suggesting that Ski may have a role in risk for obesity and metabolic disease.


Case report: Benefits and challenges of long-term eculizumab in atypical hemolytic uremic syndrome.
Atypical hemolytic uremic syndrome (aHUS) is caused by dysregulation of the complement system, leading to complement overactivation. A humanized anti-C5 monoclonal antibody, eculizumab, has been available for the treatment of aHUS since 2011. The long-term safety and efficacy of this novel drug in the pediatric population remain under review. We present a child with a hybrid CFH/CFHR3 gene who, having had multiple disease relapses despite optimal treatment with plasma exchange, commenced eculizumab therapy in August 2010. She remains relapse free in follow-up at 52 months, and treatment has been well tolerated. The risk of meningococcal disease during this treatment is recognized. Despite vaccination against meningococcal disease and appropriate antibiotic prophylaxis, our patient developed meningococcal bacteremia 30 months into treatment. She presented with nonspecific symptoms but recovered without sequelae with appropriate treatment. We recommend that children be vaccinated against invasive meningococcal infection before beginning eculizumab therapy and take appropriate antibiotic prophylaxis during treatment, and we suggest that vaccine responses should be checked and followed annually. Clinicians need to maintain a high index of suspicion for invasive meningococcal disease. Neither vaccination nor antibiotic prophylaxis provides complete protection in patients on eculizumab therapy. The appropriate dosage of eculizumab needed to achieve remission in aHUS in the pediatric population is unknown. Having achieved remission in our patient, we monitor eculizumab and CH50 levels to evaluate ongoing blockade of the terminal complement cascade. Such information may help guide dosing intervals in the future.


Primary care specialty choices of United States medical graduates, 1997-2006.
To describe trends in specialty choice and to identify predictors of primary care specialty choices among graduates of U.S. MD-granting medical schools. A longitudinal study evaluated 1997-2006 medical school graduates who completed the Association of American Medical Colleges' Matriculating Student Questionnaire and Graduation Questionnaire. Multivariate logistic regression identified significant predictors of graduates' choice of primary care specialty (general internal medicine, general pediatrics, internal medicine subspecialties, pediatrics subspecialties, family medicine, and obstetrics-gynecology) or "no-board-certification specialty," compared with all other specialties (reference). The sample included 102,673 graduates (64.9% of all 1997-2006 graduates). General internal medicine, family medicine, general pediatrics, and obstetrics-gynecology choice decreased, whereas internal medicine subspecialties, pediatrics subspecialties, and no-board-certification specialty choice increased over time (each: P < .001). Female graduates and those who planned to practice in underserved communities, espoused more-altruistic beliefs about health care, and ascribed greater importance to social responsibility in their choice of medicine at matriculation were more likely to choose general internal medicine, general pediatrics, family medicine, or obstetrics-gynecology, whereas graduates who had a physician parent and who planned full-time academic medicine careers were less likely to do so (each: P < .01). Graduates with higher debt were less likely to choose internal medicine and pediatrics specialties (each: P < .001) and more likely to choose obstetrics-gynecology (P = .001). Generalist-primary care specialty choices declined since 1997, whereas primary care subspecialty and no-board-certification specialty choices increased. Associations between primary care specialty choices and demographic, attitudinal, and career intention variables can inform the design of interventions to address expected primary care workforce shortages.


Maximizing and stabilizing luminescence from halide perovskites with potassium passivation.
Metal halide perovskites are of great interest for various high-performance optoelectronic applications. The ability to tune the perovskite bandgap continuously by modifying the chemical composition opens up applications for perovskites as coloured emitters, in building-integrated photovoltaics, and as components of tandem photovoltaics to increase the power conversion efficiency. Nevertheless, performance is limited by non-radiative losses, with luminescence yields in state-of-the-art perovskite solar cells still far from 100 per cent under standard solar illumination conditions. Furthermore, in mixed halide perovskite systems designed for continuous bandgap tunability (bandgaps of approximately 1.7 to 1.9 electronvolts), photoinduced ion segregation leads to bandgap instabilities. Here we demonstrate substantial mitigation of both non-radiative losses and photoinduced ion migration in perovskite films and interfaces by decorating the surfaces and grain boundaries with passivating potassium halide layers. We demonstrate external photoluminescence quantum yields of 66 per cent, which translate to internal yields that exceed 95 per cent. The high luminescence yields are achieved while maintaining high mobilities of more than 40 square centimetres per volt per second, providing the elusive combination of both high luminescence and excellent charge transport. When interfaced with electrodes in a solar cell device stack, the external luminescence yield-a quantity that must be maximized to obtain high efficiency-remains as high as 15 per cent, indicating very clean interfaces. We also demonstrate the inhibition of transient photoinduced ion-migration processes across a wide range of mixed halide perovskite bandgaps in materials that exhibit bandgap instabilities when unpassivated. We validate these results in fully operating solar cells. Our work represents an important advance in the construction of tunable metal halide perovskite films and interfaces that can approach the efficiency limits in tandem solar cells, coloured-light-emitting diodes and other optoelectronic applications.


Pathway analysis by randomization incorporating structure-PARIS: an update.
We present an update to the pathway enrichment analysis tool 'Pathway Analysis by Randomization Incorporating Structure (PARIS)' that determines aggregated association signals generated from genome-wide association study results. Pathway-based analyses highlight biological pathways associated with phenotypes. PARIS uses a unique permutation strategy to evaluate the genomic structure of interrogated pathways, through permutation testing of genomic features, thus eliminating many of the over-testing concerns arising with other pathway analysis approaches. We have updated PARIS to incorporate expanded pathway definitions through the incorporation of new expert knowledge from multiple database sources, through customized user provided pathways, and other improvements in user flexibility and functionality. PARIS is freely available to all users at https://ritchielab.psu.edu/software/paris-download email@example.com Supplementary data are available at Bioinformatics online.


Infrastructuring Multicultural Healthcare Information Systems.
This paper stresses the need for more research in the field of Participatory Design (PD) and in particular into how to design Health Information Technology (HIT) together with care providers and -receivers in multicultural settings. We contribute to this research by describing a case study, the 'Health-Cultures' project, in which we designed HIT for the context of home care of older people with a migration background. The Health-Cultures project is located in the city of Genk, Belgium, which is known for its multicultural population, formed by three historical migration waves of people coming to work in the nowadays closed coal mines. Via a PD approach, we studied existing means of dialogue and designed HIT that both care receivers and care providers in Genk can use in their daily exchanges between cultures in home care contexts. In discussing relevant literature as well as the results of this study, we point to the need and the ways of taking spatio-historical aspects of a specific healthcare situation into account in the PD of HIT to support multicultural perspectives on healthcare.


Maternal stress and psychological distress preconception: association with offspring atopic eczema at age 12 months.
Perinatal maternal stress and low mood have been linked to offspring atopic eczema. To examine the relation of maternal stress/mood with atopic eczema in the offspring, focusing particularly on stress/psychological distress preconception. At recruitment in the UK Southampton Women's Survey, preconception maternal reports of perceived stress in daily living and the effect of stress on health were recorded; in a subsample, psychological distress was assessed (12-item General Health Questionnaire). Infants were followed up at ages 6 (n = 2956) and 12 (n = 2872) months and atopic eczema ascertained (based on UK Working Party Criteria for the Definition of Atopic Dermatitis). At 6 months post-partum, mothers were asked if they had experienced symptoms of low mood since childbirth and completed the Edinburgh Postnatal Depression Scale. Preconception perceived stress affecting health [OR 1.21 (95% CI 1.08-1.35), P = 0.001] and stress in daily living [OR 1.16 (1.03-1.30), P = 0.014] were associated with an increased risk of offspring atopic eczema at age 12 months but not at 6 months, robust to adjustment for potentially confounding variables. Findings were similar for maternal psychological distress preconception. Low maternal mood between delivery and 6 months post-partum was associated with an increased risk of infantile atopic eczema at age 12 months, but no significant association between post-natal mood and atopic eczema was seen after taking account of preconception stress. Our data provide novel evidence linking maternal stress at preconception to atopic eczema risk, supporting a developmental contribution to the aetiology of atopic eczema and pointing to potentially modifiable influences.


Germinal centers output clonally diverse plasma cell populations expressing high- and low-affinity antibodies.
Germinal centers (GCs) form in lymph nodes after immunization or infection to facilitate antibody affinity maturation and memory and plasma cell (PC) development. PC differentiation is thought to involve stringent selection for GC B cells expressing the highest-affinity antigen receptors, but how this plays out during complex polyclonal responses is unclear. We combine temporal lineage tracing with antibody characterization to gain a snapshot of PCs developing during influenza infection. GCs co-mature B cell clones with antibody affinities spanning multiple orders of magnitude; however, each generates PCs with similar efficiencies, including weak binders. Within lineages, PC selection is not restricted to variants with the highest-affinity antibodies. Differentiation is commonly associated with proliferative expansion to produce "nodes" of identical PCs. Immunization-induced GCs generate fewer PCs but still of low- and high-antibody affinities. We propose that generating low-affinity antibody PCs reflects an evolutionary compromise to facilitate diverse serum antibody responses.


Accountable interventional pain management: a collaboration among practitioners, patients, payers, and government.
The prevalence, costs, and disability associated with chronic pain continue to escalate. So too, the numerous modalities of treatments applied in managing these patients continue to increase as well. In the period from 2000 to 2011 interventional techniques increased 228%. In addition, analysis of utilization trends and expenditures for spinal interventional techniques alone from 2000 to 2008 illustrated an increase in Medicare fee-for-service expenditures of 240% in terms of dollars spent in the United States. The Office of Inspector General (OIG) of the Department of Health and Human Services showed an increase in facet joint and transforaminal epidural injections, with a significant proportion of these services did not meet the medical necessity criteria.The increasing utilization of interventional techniques is also associated with significant variations among specialty groups and regional variations among states. Overall procedures have increased by 173%, with rate of 130% per 100,000 Medicare beneficiaries for epidural injections; 383%, with a rate of 308% for facet joint interventions; and overall 410%, or a rate of 331% for sacroiliac joint interventions. Certain high volume interventions such as lumbar transforaminal epidural injections and lumbar facet joint neurolysis have actually increased a staggering 806% and 662%.Coverage policies across ambulatory settings and by multiple payers are highly variable. Apart from variability in the development of coverage policies, payments also substantially vary by site of service. In general, amongst the various ambulatory settings the highest payments are made to hospital outpatient departments (HOPDs) the lowest to in-office procedures, and payment to ambulatory surgery centers (ASCs) falling somewhere in the middle.This manuscript describes the many differences that exist between the various settings, and includes suggestions for accountable interventional pain management with coverage for techniques with evidence, addressing excessive use of specific techniques, and equalizing payments across multiple ambulatory settings.


Insulin resistance is associated with impaired nitric oxide synthase activity in skeletal muscle of type 2 diabetic subjects.
Type 2 diabetes is an insulin-resistant state characterized by hyperinsulinemia and accelerated atherosclerosis. In vitro and in vivo studies in rodents have suggested that nitric oxide generation plays an important role in glucose transport and insulin action. We determined nitric oxide synthase (NOS) activity in skeletal muscle of 10 type 2 diabetic (hemoglobin A(1C) = 6.8 +/- 0.1%) and 11 control subjects under basal conditions and during an 80 mU/m(2).min euglycemic insulin clamp performed with vastus lateralis muscle biopsies before and after 4 h of insulin. In diabetics, insulin-stimulated glucose disposal (Rd) was reduced by 50%, compared with controls (5.4 +/- 0.3 vs. 10.4 +/- 0.5 mg/kg.min, P < 0.01). Basal NOS activity was markedly reduced in the diabetic group (101 +/- 33 vs. 457 +/- 164 pmol/min.mg protein, P < 0.05). In response to insulin, NOS activity increased 2.5-fold in controls after 4 h (934 +/- 282 pmol/min.mg protein, P < 0.05 vs. basal), whereas insulin failed to stimulate NOS activity in diabetics (86 +/- 28 pmol/min.mg protein, P = NS from basal). Basal NOS protein content in muscle was similar in controls and diabetics and did not change following insulin. In controls, insulin-stimulated NOS activity correlated inversely with fasting plasma insulin concentration (r = -0.58, P = 0.05) and positively with Rd (r = 0.71, P = 0.03). In control and diabetic groups collectively, Rd correlated with insulin-stimulated NOS activity (r = 0.52, P = 0.02). We conclude that basal and insulin-stimulated muscle NOS activity is impaired in well-controlled type 2 diabetic subjects, and the defect in insulin-stimulated NOS activity correlates closely with the severity of insulin resistance. These results suggest that impaired NOS activity may play an important role in the insulin resistance in type 2 diabetic individuals.


Religation of the T cell receptor after primary activation of mature T cells inhibits proliferation and induces apoptotic cell death.
The proliferative response of murine splenic T cells, initially activated by cross-linking the TCR complex with either antibodies, mitogenic lectin, or alloantigen was severely inhibited when the activated cells were recovered and given an additional activation signal by recross-linking the TCR complex, or by adding Ca2+ ionophore and phorbol ester. Under the same conditions, cross-linking other T cell surface determinants such as CD4, CD8, or class I MHC on preactivated T cells had no effect. Assessment of cell viability using vital dye exclusion together with the detection of DNA fragmentation revealed that the reduction of the proliferative responses was associated with an induction of apoptotic-like cell death in the activated T cell population and not due to a blockade of cell division. Accumulation of eosin stained (dead) cells did not occur immediately upon replating the activated cells, but began after a lag period during which at least two cell divisions occurred. In addition, perturbation of T cell proliferation after activation depended on how the cells were initially activated. Only T cells activated in the presence of additional cells found in spleen and lymph node were susceptible to inhibition; T cells activated after nylon wool purification were not susceptible. These results have potential implications for understanding self-tolerance and immunoregulation.


A unique semiconductor-carbon-metal hybrid structure design as a counter electrode in dye-sensitized solar cells.
The catalytic activity of counter electrodes (CEs) severely restricts the photovoltaic conversion efficiency of dye-sensitized solar cells. However, electrons trapped by bulk defects greatly reduce the catalytic activity of the CE. In this study, we report a novel In2S3-C-Au hybrid structure designed by simply decorating Au particles on the surface of carbon-coated hierarchical In2S3 flower-like architectures, which could avoid the abovementioned problems. This effect can be attributed to the unique contribution of indium sulfide, carbon, and Au from the hybrid structure, as well as to their synergy. Electrochemical measurements revealed that the hybrid structure possessed high catalytic activity and electrochemical stability for the interconversion of the redox couple I3-/I-. Moreover, this superior performance can be incorporated into the dye-sensitized solar cells system. We used this hybrid structure as a counter electrode by casting it on an FTO substrate to form a film, which displayed better photovoltaic conversion efficiency (8.91%) than the commercial Pt counterpart (7.67%).


AdE-1, a new inotropic Na(+) channel toxin from Aiptasia diaphana, is similar to, yet distinct from, known anemone Na(+) channel toxins.
Heart failure is one of the most prevalent causes of death in the western world. Sea anemone contains a myriad of short peptide neurotoxins affecting many pharmacological targets, several of which possess cardiotonic activity. In the present study we describe the isolation and characterization of AdE-1 (ion channel modifier), a novel cardiotonic peptide from the sea anemone Aiptasia diaphana, which differs from other cnidarian toxins. Although AdE-1 has the same cysteine residue arrangement as sea anemone type 1 and 2 Na(+) channel toxins, its sequence contains many substitutions in conserved and essential sites and its overall homology to other toxins identified to date is low (<36%). Physiologically, AdE-1 increases the amplitude of cardiomyocyte contraction and slows the late phase of the twitch relaxation velocity with no induction of spontaneous twitching. It increases action potential duration of cardiomyocytes with no effect on its threshold and on the cell's resting potential. Similar to other sea anemone Na(+) channel toxins such as Av2 (Anemonia viridis toxin II), AdE-1 markedly inhibits Na(+) current inactivation with no significant effect on current activation, suggesting a similar mechanism of action. However, its effects on twitch relaxation velocity, action potential amplitude and on the time to peak suggest that this novel toxin affects cardiomyocyte function via a more complex mechanism. Additionally, Av2's characteristic delayed and early after-depolarizations were not observed. Despite its structural differences, AdE-1 physiologic effectiveness is comparable with Av2 with a similar ED(50) value to blowfly larvae. This finding raises questions regarding the extent of the universality of structure-function in sea anemone Na(+) channel toxins.


Incidence of adverse drug events and potential adverse drug events. Implications for prevention. ADE Prevention Study Group.
To assess incidence and preventability of adverse drug events (ADEs) and potential ADEs. To analyze preventable events to develop prevention strategies. Prospective cohort study. All 4031 adult admissions to a stratified random sample of 11 medical and surgical units in two tertiary care hospitals over a 6-month period. Units included two medical and three surgical intensive care units and four medical and two surgical general care units. Adverse drug events and potential ADEs. Incidents were detected by stimulated self-report by nurses and pharmacists and by daily review of all charts by nurse investigators. Incidents were subsequently classified by two independent reviewers as to whether they represented ADEs or potential ADEs and as to severity and preventability. Over 6 months, 247 ADEs and 194 potential ADEs were identified. Extrapolated event rates were 6.5 ADEs and 5.5 potential ADEs per 100 nonobstetrical admissions, for mean numbers per hospital per year of approximately 1900 ADEs and 1600 potential ADEs. Of all ADEs, 1% were fatal (none preventable), 12% life-threatening, 30% serious, and 57% significant. Twenty-eight percent were judged preventable. Of the life-threatening and serious ADEs, 42% were preventable, compared with 18% of significant ADEs. Errors resulting in preventable ADEs occurred most often at the stages of ordering (56%) and administration (34%); transcription (6%) and dispensing errors (4%) were less common. Errors were much more likely to be intercepted if the error occurred earlier in the process: 48% at the ordering stage vs 0% at the administration stage. Adverse drug events were common and often preventable; serious ADEs were more likely to be preventable. Most resulted from errors at the ordering stage, but many also occurred at the administration stage. Prevention strategies should target both stages of the drug delivery process.


Inflammatory stimuli induce inhibitory S-nitrosylation of the deacetylase SIRT1 to increase acetylation and activation of p53 and p65.
Inflammation increases the abundance of inducible nitric oxide synthase (iNOS), leading to enhanced production of nitric oxide (NO), which can modify proteins by S-nitrosylation. Enhanced NO production increases the activities of the transcription factors p53 and nuclear factor κB (NF-κB) in several models of disease-associated inflammation. S-nitrosylation inhibits the activity of the protein deacetylase SIRT1. SIRT1 limits apoptosis and inflammation by deacetylating p53 and p65 (also known as RelA), a subunit of NF-κB. We showed in multiple cultured mammalian cell lines that NO donors or inflammatory stimuli induced S-nitrosylation of SIRT1 within CXXC motifs, which inhibited SIRT1 by disrupting its ability to bind zinc. Inhibition of SIRT1 reduced deacetylation and promoted activation of p53 and p65, leading to apoptosis and increased expression of proinflammatory genes. In rodent models of systemic inflammation, Parkinson's disease, or aging-related muscular atrophy, S-nitrosylation of SIRT1 correlated with increased acetylation of p53 and p65 and activation of p53 and NF-κB target genes, suggesting that S-nitrosylation of SIRT1 may represent a proinflammatory switch common to many diseases and aging.


The effect of the long-term ingestion of konjac-glucomannan on glucose tolerance and immunoreactive insulin values of baboons.
It is uncertain whether the beneficial effects of the high-fibre, high-carbohydrate diets generally recommended for improved glucose control in diabetic patients are in fact due to the fibre. In this study the effect of the long-term ingestion of the dietary fibre, konjac - glucomannan (K-GM), on glucose tolerance and immunoreactive insulin (IRI) concentrations was investigated in normal baboons and compared with the short-term effect of the concurrent feeding of K-GM and glucose. Eighteen young baboons were divided in a control and a K-GM group. Glucose tolerance tests ( GTTs ) performed at baseline showed that the two groups were strictly comparable. The K-GM group then received the same diet as the control group plus 5% K-GM. After a period of 12 - 14 weeks on the diets, the K-GM baboons had significantly higher fasting blood glucose and serum IRI values. The results of GTTs showed, however, that long-term feeding of K-GM improved glucose tolerance significantly and decreased the area under the IRI response. These long-term effects were more pronounced than the short-term effects. It is concluded that the long-term ingestion of K-GM per se has beneficial effects on glucose homeostasis in normal baboons and that the possible mechanisms by which these effects are established warrant further investigation.


Role of abnormalities of carbohydrate and lipoprotein metabolism in the pathogenesis and clinical course of hypertension.
Patients with untreated hypertension have been shown to be resistant to insulin-stimulated glucose uptake. They are also hyperinsulinemic and hypertriglyceridemic when compared with matched control groups with normal blood pressure. In addition, insulin resistance, hyperinsulinemia, and hypertriglyceridemia have been demonstrated in rat models of hypertension, including spontaneous hypertensive rats and Sprague-Dawley rats fed a fructose-enriched diet. The defect in insulin-stimulated glucose uptake in these experimental models can also be shown at the cellular level. Furthermore, experimental interventions that prevent insulin resistance or hyperinsulinemia from developing in fructose-fed rats also greatly attenuate increases in blood pressure. Endogenous hyperinsulinemia and hypertriglyceridemia have been identified as factors that increase the risk of coronary artery disease (CAD), and they may contribute to the increased prevalence of ischemic heart disease in patients with high blood pressure. The fact that past antihypertensive treatment has not focused on these metabolic abnormalities, and indeed may have exacerbated them, could help explain why it has been difficult to show that lowering blood pressure decreases the risk of CAD. These observations raise the possibility that abnormalities of carbohydrate and lipoprotein metabolism may play a role in both the etiology and the clinical course of hypertension.


The effect of a ketogenic diet and synergy with rapamycin in a mouse model of breast cancer.
The effects of diet in cancer, in general, and breast cancer in particular, are not well understood. Insulin inhibition in ketogenic, high fat diets, modulate downstream signaling molecules and are postulated to have therapeutic benefits. Obesity and diabetes have been associated with higher incidence of breast cancer. Addition of anti-cancer drugs together with diet is also not well studied. Two diets, one ketogenic, the other standard mouse chow, were tested in a spontaneous breast cancer model in 34 mice. Subgroups of 3-9 mice were assigned, in which the diet were implemented either with or without added rapamycin, an mTOR inhibitor and potential anti-cancer drug. Blood glucose and insulin concentrations in mice ingesting the ketogenic diet (KD) were significantly lower, whereas beta hydroxybutyrate (BHB) levels were significantly higher, respectively, than in mice on the standard diet (SD). Growth of primary breast tumors and lung metastases were inhibited, and lifespans were longer in the KD mice compared to mice on the SD (p<0.005). Rapamycin improved survival in both mouse diet groups, but when combined with the KD was more effective than when combined with the SD. The study provides proof of principle that a ketogenic diet a) results in serum insulin reduction and ketosis in a spontaneous breast cancer mouse model; b) can serve as a therapeutic anti-cancer agent; and c) can enhance the effects of rapamycin, an anti-cancer drug, permitting dose reduction for comparable effect. Further, the ketogenic diet in this model produces superior cancer control than standard mouse chow whether with or without added rapamycin.


Wnt5a cloning, expression, and up-regulation in human primary breast cancers.
Wnt genes are involved in mouse mammary cancer, but their role in human cancer is unknown. Human Wnt5a was cloned from a placental cDNA library and used to assess expression by ribonuclease protection and in situ hybridization in human breast cell lines and in normal, benign, and malignant breast tissues. Human Wnt5a shows over 99% homology at amino acid level with mouse Wnt5a, and 90% with Xenopus Wnt5a. It was expressed only at low levels in breast cell lines and normal breast tissue. Benign proliferations and invasive cancer respectively showed 10-fold and 4-fold higher Wnt5a than normal breast tissues. The greater up-regulation in benign conditions suggests a role in aberrant differentiation. In situ hybridization localized the signal to the epithelial component. Wnt5a is the first member of the Wnt family to demonstrate overexpression in human breast cancer. It was not associated with factors known to affect breast cancer prognosis such as lymph node status or epidermal growth factor receptor status.


Instabilities of a Bose-Einstein condensate in a periodic potential: an experimental investigation.
By accelerating a Bose-Einstein condensate in a controlled way across the edge of the Brillouin zone of a 1D optical lattice, we investigate the stability of the condensate in the vicinity of the zone edge. Through an analysis of the visibility of the interference pattern after a time-of-flight and the widths of the interference peaks, we characterize the onset of instability as the acceleration of the lattice is decreased. We briefly discuss the significance of our results with respect to recent theoretical work.


Formulation and implementation of a phase-resolved fluorescence technique for latent-fingerprint imaging: theoretical and experimental analysis.
A theoretical and experimental study of the imaging of latent fingerprints by a phase-resolved fluorescence technique along with associated signal-processing analysis is described. The system configuration is optimized by incorporation of a novel approach of homodyne-assisted even-step phase shifting in a signal-processing concept. The excitation laser source and gain of the detection device, which are modulated at megahertz frequency followed by sensitive signal-processing concepts, are employed to separate the fingerprint fluorescence from background fluorescence. Experiments are carried out with fingerprints deposited upon different types of substrate surfaces. Later, a quantitative image-quality assessment is carried out, which confirms the improvement in the quality of the phase-resolved fingerprint image. Imaging of older fingerprints with better contrast is also carried out with the proposed novel technique.


Human fibroblast-macrophage tissue spheroids demonstrate ratio-dependent fibrotic activity for in vitro fibrogenesis model development.
Fibrosis is a pathological accumulation of excessive collagen that underlies many of the most common diseases, representing dysfunction of the essential processes of normal tissue healing. Fibrosis research aims to limit this response without ameliorating the essential role of fibrogenesis in organ function. However, the absence of a realistic in vitro model has hindered investigation into mechanisms and potential interventions because the standard 2D monolayer culture of fibroblasts has limited applicability. We sought to develop and optimize fibrosis spheroids: a scaffold-free three-dimensional human fibroblast-macrophage spheroid system representing an improved benchtop model of human fibrosis. We created, characterized and optimized human fibroblast-only spheroids, demonstrating increased collagen deposition compared to monolayer fibroblasts, while spheroids larger than 300 μm suffered from progressively increasing apoptosis. Next, we improved the spheroid system with the addition of human macrophages to more precisely recapitulate the environment during fibrogenesis, creating a hybrid spheroid system with different ratios of fibroblasts and macrophages ranging from 2 : 1 to 64 : 1. We found that in the hybrid spheroids (particularly the 16 : 1 [F16] ratio) more fibroblasts were activated, with greater macrophage polarization towards a pro-inflammatory M1 phenotype. Hybrid spheroids containing higher ratios of macrophages showed greater macrophage heterogeneity and less fibrogenesis, while low macrophage ratios limited macrophage-induced effects and yielded less collagen deposition. The F16 group also had the highest expression levels of fibrosis-related genes (Col-1a1, Col-3a1 and TGF-β) and inflammation-related genes (TNF, IL1β and IL6). IF staining demonstrated that F16 spheroids had the highest levels of αSMA, collagen-1 and collagen-3 deposition among all groups as well as formation of a dense collagen rim surrounding the spheroid. Future studies exploring the greater fibrotic activity of F16 spheroids may provide new mechanistic insights into diseases involving excessive fibrotic activity. Microtissue fibrosis models capable of achieving greater clinical fidelity have the potential to combine the relevance of animal models with the scale, cost and throughput of in vitro testing.


Fragment-specific volar hook plate for volar marginal rim fractures.
The volar lunate facet/volar marginal rim fragment of distal radius fractures is a challenging problem for current volar plating systems to adequately maintain internal fixation. Fragment-specific wireforms and other constructs can be used to capture these difficult fragments but are difficult to place and have limited resistance to bending and axial load. Fixed-angle volar plates have limited distal translation without increasing the risk of flexor tendon rupture. Failure to reduce and maintain reduction of these volar ulnar intra-articular distal marginal rim fractures result in disastrous volar carpal subluxation. We present an alternative surgical technique using a low-profile volar hook plate designed specifically to address this fragment. The first 6 cases are presented here with no observed surgical complications to date.


Joint Consensus Statement of the American Academy of Sleep Medicine and Sleep Research Society on the Recommended Amount of Sleep for a Healthy Adult: Methodology and Discussion.
The American Academy of Sleep Medicine and Sleep Research Society recently released a Consensus Statement regarding the recommended amount of sleep to promote optimal health in adults. This paper describes the methodology, background literature, voting process, and voting results for the consensus statement. In addition, we address important assumptions and challenges encountered during the consensus process. Finally, we outline future directions that will advance our understanding of sleep need and place sleep duration in the broader context of sleep health.


Dyadic coping and salivary interleukin-6 responses to interpersonal stress.
Dysregulated immune responses to stress are a potential pathway linking close relationship processes to health, and couples' abilities to cope with stress together (dyadic coping) likely impact such immune responses. Most stress research has focused on immune reactivity, whereas knowledge of immune recovery remains limited. The present study examined how acute interpersonal stress affects immune reactivity and recovery, as well as whether dyadic coping moderates these effects. Healthy couples (N = 24) completed the Dyadic Coping Inventory and provided saliva samples 4 times each day for 5 days, including 2 days before a laboratory dyadic stressor (discussing an area of disagreement), the day of, and 2 days after. Four additional saliva samples were taken throughout the laboratory stressor. Saliva samples were assayed for interleukin (IL)-6. Multilevel models that adjusted for demographic and health variables indicated that partners low in dyadic coping showed immune reactivity to the stressor whereas partners high in dyadic coping did not. Dyadic coping did not moderate immune recovery, which had occurred by 5 hr poststressor across all participants. Results suggest that partners low in dyadic coping show increased reactivity of immune responses to interpersonal stress. Enhancing dyadic coping in couples may impact not only their mental health and relationship quality, but also their risk of stress-related immune disorders. (PsycINFO Database Record


Therapeutic studies in NZB/W mice. III. Relationship between renal status and efficacy of immunosuppressive drug therapy.
Female NZB/W mice develop a disease closely resembling human systemic lupus and serve as an animal model for therapeutic studies. Several previous studies have demonstrated the efficacy of different immunosuppressive drug regimens in the therapy of glomerulonephritis in NZB/W mice. After the onset of immune complex deposition, treatment with intermittent high doses of cyclophosphamide or daily low doses of the combination of cyclophosphamide, azathioprine, and methylprednisolone has been effective. The present study was designed to compare such effective regimens in mice early in the course of their renal disease versus mice late in the course of glomerulonephritis. One to three injections of high-dose cyclophosphamide during active immune complex deposition and early histologic changes were significantly effective in prolonging survival, whereas treatment late in the course of glomerulonephritis was less effective. Even more striking was the result of low-dose combination therapy. Daily treatment with cyclophosphamide, azathiprine, and methylprednisolone (C + A + M) effectively prolonged survival when started in mice 5 months old, but was of no benefit when started in those 8 months of age. In a concluding experiment, older mice were selected on the basis of degree of renal disease and studied with regard to proteinuria and survival. Those with mild renal disease responded to daily treatment for 6 months with C + A + M at 1 mg/kg of each drug, whereas those with advanced renal disease at the onset of therapy did not benefit.


Uptake of HIV testing and treatment in an isolated population with access to free and universal healthcare.
This Western Australian study analysed clinical information from 344 people with AIDS, and experiential data from a sample of 36 people with AIDS to investigate 2 relevant issues: the timing of HIV testing and uptake and experiences of HIV drug treatment. We found that the proportion of people presenting with AIDS before being diagnosed with HIV infection ('AIDS-presenters') fell from 47% in 1985-1987 to 20% in 1991-1993. Since antiretroviral therapy (ART) became available in 1987, two-thirds of Western Australian people with AIDS who knew of their HIV infection prior to the development of AIDS have consistently elected to take recommended HIV treatment. Four phases of the treatment experience were identified. It is concluded that 'AIDS-presentation' is unlikely to reduce further and that the timing of testing and issues of treatment acceptance are only understood when the experiences of people with illness are examined using a patient-centred approach.


The nucleotide binding dynamics of human MSH2-MSH3 are lesion dependent.
Here we report that the human DNA mismatch complex MSH2-MSH3 recognizes small loops by a mechanism different from that of MSH2-MSH6 for single-base mismatches. The subunits MSH2 and MSH3 can bind either ADP or ATP with similar affinities. Upon binding to a DNA loop, however, MSH2-MSH3 adopts a single 'nucleotide signature', in which the MSH2 subunit is occupied by an ADP molecule and the MSH3 subunit is empty. Subsequent ATP binding and hydrolysis in the MSH3 subunit promote ADP-ATP exchange in the MSH2 subunit to yield a hydrolysis-independent ATP-MSH2-MSH3-ADP intermediate. Human MSH2-MSH3 and yeast Msh2-Msh6 both undergo ADP-ATP exchange in the Msh2 subunit but, apparently, have opposite requirements for ATP hydrolysis: ADP release from DNA-bound Msh2-Msh6 requires ATP stabilization in the Msh6 subunit, whereas ADP release from DNA-bound MSH2-MSH3 requires ATP hydrolysis in the MSH3 subunit. We propose a model in which lesion binding converts MSH2-MSH3 into a distinct nucleotide-bound form that is poised to be a molecular sensor for lesion specificity.


Light adaptation in cat retinal rods.
It has long been an open question whether individual rod receptors in the mammalian retina show any light adaptation. The prevailing evidence so far has suggested that these cells, unlike those in lower vertebrates, adapt little if at all. The experiments on cat rods reported here, however, indicate that this is not really true. Since the cone system in the cat retina has a fairly high light threshold, the rods also need to adapt so that they do not saturate with light before the cones fully take over vision at higher light intensities. In similar experiments, adaptation was found in rods of other mammalian species, including primates.


Phage-type RNA polymerase RPOTmp performs gene-specific transcription in mitochondria of Arabidopsis thaliana.
Transcription of mitochondrial genes in animals, fungi, and plants relies on the activity of T3/T7 phage-type RNA polymerases. Two such enzymes, RPOTm and RPOTmp, are present in the mitochondria of eudicotyledonous plants; RPOTmp is additionally found in plastids. We have characterized the transcriptional role of the dual-targeted RNA polymerase in mitochondria of Arabidopsis thaliana. Examination of mitochondrial transcripts in rpoTmp mutants revealed major differences in transcript abundances between wild-type and rpoTmp plants. Decreased levels of specific transcripts were correlated with reduced abundances of the respiratory chain complexes I and IV. Altered transcript levels in rpoTmp were found to result from gene-specific transcriptional changes, establishing that RPOTmp functions in distinct transcriptional processes within mitochondria. Decreased transcription of specific genes in rpoTmp was not associated with changes in promoter utilization; therefore, RPOTmp function is not promoter specific but gene specific. This implies that additional gene-specific elements direct the transcription of a subset of mitochondrial genes by RPOTmp.


Mechanism of action of KIAA1456 gene on the proliferation and apoptosis of alveolar epithelial cells.
To explore the mechanism by which KIAA1456 acts on alveolar epithelial cells through lentiviral transfection. After constructing a KIAA1456 gene vector, 293T cells were co-transfected with lentiviral vectors and after incubation cells were examined by fluorescence microscopy. CCL-149 cells were transfected with LV-KIAA1456 and were examined by fluorescence microscopy. The proliferation capacity of transfected CCL-149 cells was evaluated using flow cytometry. The effect of KIAA1456 overexpression on CCL-149 cells proliferation was studied using the CCK-8 method. The expression level of KIAA1456 in the LV- KIAA1456 group was significantly higher compared with the LV-Con group and the blank group. Compared with the LV-Con and the blank groups, the proportion of responding cells in G2/M phase showed statistically significant differences. Viable cells had adarker color and higher OD value measured by ELISA. Compared with the control and the blank groups, the growth and proliferation in the CCL-149 transfection group were significantly slower. KIAA1456 gene inhibited the proliferation of CCL-149 cells by negative regulation of the G2/M cell cycle. We suggest that it can be used as a specific target for the treatment of alveolar epithelium.


Food and Drug Administration warning on anesthesia and brain development: implications for obstetric and fetal surgery.
There has been growing concern about the detrimental effects of certain anesthetic agents on the developing brain. Preclinical studies in small animal models as well as nonhuman primates suggested loss or death of brain cells and consequent impaired neurocognitive function following anesthetic exposure in neonates and late gestation fetuses. Human studies in this area are limited and currently inconclusive. On Dec. 14, 2016, the US Food and Drug Administration issued a warning regarding impaired brain development in children following exposure to certain anesthetic agents used for general anesthesia, namely the inhalational anesthetics isoflurane, sevoflurane, and desflurane, and the intravenous agents propofol and midazolam, in the third trimester of pregnancy. Furthermore, this warning recommends that health care professionals should balance the benefits of appropriate anesthesia in young children and pregnant women against potential risks, especially for procedures that may last >3 hours or if multiple procedures are required in children <3 years old. The objective of this article is to highlight how the Food and Drug Administration warning may impact the anesthetic and surgical management of the obstetric patient. Neuraxial anesthesia (epidural or spinal anesthesia) is more commonly administered for cesarean delivery than general anesthesia. The short duration of fetal exposure to general anesthesia during cesarean delivery has not been associated with learning disabilities. However, the fetus can also be exposed to both intravenous and inhalation anesthetics during nonobstetric or fetal surgery in the second and third trimester; this exposure is typically longer than that for cesarean delivery. Very few studies address the effect of anesthetic exposure on the fetus in the second trimester when most nonobstetric and fetal surgical procedures are performed. It is also unclear how the plasticity of the fetal brain at this stage of development will modulate the consequences of anesthetic exposure. Strategies that may circumvent possible untoward long-term neurologic effects of anesthesia in the baby include: (1) use of nonimplicated (nongamma-aminobutyric acid agonist) agents for sedation such as opioids (remifentanil, fentanyl) or the alpha-2 agonist, dexmedetomidine, when appropriate; (2) minimizing the duration of exposure to inhalational anesthetics for fetal, obstetric, and nonobstetric procedures in the pregnant patient, as much as possible within safe limits; and (3) commencing surgery promptly and limiting the interval between induction of anesthesia and surgery start time will help decrease patient exposure to inhalational agents. While the Food and Drug Administration warning was based on duration and repetitive nature of exposure rather than concentration of inhalational agents, intravenous tocolytics can be considered for intraoperative use, to provide uterine relaxation for fetal surgery, in lieu of high concentrations of inhalational anesthetic agents. Practitioners should consider the type of anesthesia that will be administered and the potential risks when scheduling patients for nonobstetric and fetal surgery.


Holographic interferometry using two-wavelength holography for the measurement of large deformations.
Visible holographic interferometry is generally too sensitive for the measurement of large deformations. We present a holographic method that permits an increase in the range of measurable deformations. It requires the use of two different wavelengths, λ(1) and λ(2), and two holograms in series. We develop the theoretical basis of a method that permits the obtention of an interferogram as if a longer equivalent wavelength, λ(eq) = λ(1)λ(2)/|λ(1) -λ(2)|, were used. The method is experimentally tested by use of a setup that can be easily converted into a classical single-wavelength holographic interferometer, permitting comparison of the interferograms of the same deformation produced with both methods. Significant results are presented.


Applying a sunburst visualization to summarize user navigation sequences.
For many Web-based applications, it's important to be able to analyze the paths users have taken through a site--for example, to understand how they're discovering engaging content. These paths are difficult to summarize visually because of the underlying data's complexity. A Google researcher applied a sunburst visualization to this problem, after simplifying the data into a hierarchical format. The resulting visualization was successful in YouTube and is widely referenced and accessed. The code for the visualization is available as open source.


A novel angiogenic peptide, ΔADT: A truncated adrenotensin peptide revealed by secretory peptidome analysis of human retinal pericytes.
Retinal pericytes play an important role in the maintenance of retinal microvascular homeostasis. We performed a secretory peptidome of primary human retinal pericytes. Using liquid chromatography-tandem mass spectrometry analysis in the culture medium of retinal pericytes, we identified 256 peptides derived from 114 proteins, and identified a novel partial fragment Leu163-His183 (termed ΔADT) of adrenotensin (ADT). To elucidate the role of ΔADT as a soluble mediator of pericyte-endothelial cell interactions, we investigated the bioactivity of ΔADT in human retinal microvascular endothelial cells (HRMVECs). The cell proliferation assay indicated that the proliferation of HRMVECs was promoted by ADT or ΔADT. Moreover, ΔADT had a greater growth promoting effect than ADT in HRMVECs and induced migration and tube formation of HRMVECs. We also observed actin reorganization and that the levels of phosphorylated focal adhesion kinase in ΔADT stimulated HRMVECs. These results showed that ΔADT induces profound actin reorganization and increases the levels of phosphorylated focal adhesion kinase. Collectively, our study showed that ΔADT has an angiogenic activity, and suggested that ΔADT is a novel angiogenic peptide.


Alteration in serum and bone component findings induced in streptozotocin-diabetic rats is restored by zinc acexamate.
The effect of zinc acexamate in streptozotocin (STZ)-induced diabetic rats was investigated. Rats received a single subcutaneous administration of STZ (6.0 mg/100 g body weight), and the animals were orally administered once daily for 14 days with zinc acexamate (2.5, 5 or 10 mg/100 g body weight). The administration of STZ caused a significant increase in serum glucose, triglyceride and calcium levels and a significant decrease in body weight, serum zinc and inorganic phosphorus levels, indicating diabetic condition. Moreover, calcium content, alkaline phosphatase activity and deoxyribonucleic acid (DNA) content in the femoral-diaphyseal and -metaphyseal tissues were significantly reduced in STZ-diabetic rats. The change in these serum and bone components of STZ-diabetic rats was significantly restored by the oral administration of zinc acexamate (2.5, 5 or 10 mg Zn/100 g body weight). The restoration of bone components was not seen by the oral administration of zinc sulfate (2.5 mg Zn/100 g) for 14 days. Moreover, when the femoral-diaphyseal and -metaphyseal tissues obtained at 14 days after STZ administration were cultured for 48 h in a medium containing either vehicle or zinc acexamate (10(-5) M), the femoral calcium content and alkaline phosphatase activity were significantly increased in vitro. The effect of zinc acexamate was completely abolished in the presence of cycloheximide (10(-6) M), an inhibitor of protein synthesis. The present study demonstrates that the oral administration of zinc acexamate has a preventive effect on STZ-induced diabetic condition in rats, and that it can restorate bone loss of STZ-induced diabetes in vivo.


Effects of growth temperature, 47-megadalton plasmid, and calcium deficiency on the outer membrane protein porin and lipopolysaccharide composition of Yersinia pestis EV76.
The expression of several virulence determinants of Yersinia pestis is known to be dependent on the in vitro growth temperature. One of these, calcium dependence, is associated with the presence of a 47-megadalton plasmid. We have examined the effects of incubation temperature, calcium in the growth medium, the presence of the 47-megadalton plasmid on the outer membrane protein, and the lipopolysaccharide composition of Y. pestis EV76. When cells were grown at 37 degrees C as opposed to 26 degrees C, a change in lipopolysaccharide composition and a decrease in the amount of an outer membrane protein (protein E) were observed. The lipopolysaccharide obtained from cells incubated at 37 degrees C had a lower proportion of 2 keto-3-deoxyoctanate, a lower phosphate to 2-keto-3-deoxyoctanate ratio, and an increased gel mobility upon sodium dodecyl sulfate-polyacrylamide gel electrophoresis when compared with lipopolysaccharide obtained from cells grown at 26 degrees C. Because of its growth temperature-related abundance, we investigated the nature of protein E. This protein had physical properties similar to those of other enterobacterial porins, including apparent formation of an oligomer on sodium dodecyl sulfate-polyacrylamide gels when solubilized at low temperature, acidic isoelectric point, and strong noncovalent association with the peptidoglycan. Protein E was purified and shown to form an aqueous channel in planar lipid membranes with a conductance of 1.1 nS in 1 M KCl. In addition to growth temperature-related alterations in the lipopolysaccharide and porin components of the outer membrane, the amount of three spots in two-dimensional polyacrylamide gels was shown to be related to the temperature or the presence of calcium during growth. One of these spots was shown to contain residual unmodified portions of two major heat-modifiable proteins which failed to shift to their heat-modified positions on gels, despite solubilization at 100 degrees C for 10 min before electrophoresis. The other two spots were the heat-modified and unmodified forms of another outer membrane protein (J) which did not appear in the isoelectric focusing gel of cells grown at 37 degrees C. It is proposed that the appearance of these spots in two-dimensional analyses is related to the lipopolysaccharide composition of the cells from which the outer membrane is derived and reflects lipopolysaccharide-protein interactions or calcium-protein interactions.


Genomic and epigenomic insights into the origin, pathogenesis, and clinical behavior of mantle cell lymphoma subtypes.
Mantle cell lymphoma (MCL) is a mature B-cell neoplasm initially driven by CCND1 rearrangement with 2 molecular subtypes, conventional MCL (cMCL) and leukemic non-nodal MCL (nnMCL), that differ in their clinicobiological behavior. To identify the genetic and epigenetic alterations determining this diversity, we used whole-genome (n = 61) and exome (n = 21) sequencing (74% cMCL, 26% nnMCL) combined with transcriptome and DNA methylation profiles in the context of 5 MCL reference epigenomes. We identified that open and active chromatin at the major translocation cluster locus might facilitate the t(11;14)(q13;32), which modifies the 3-dimensional structure of the involved regions. This translocation is mainly acquired in precursor B cells mediated by recombination-activating genes in both MCL subtypes, whereas in 8% of cases the translocation occurs in mature B cells mediated by activation-induced cytidine deaminase. We identified novel recurrent MCL drivers, including CDKN1B, SAMHD1, BCOR, SYNE1, HNRNPH1, SMARCB1, and DAZAP1. Complex structural alterations emerge as a relevant early oncogenic mechanism in MCL, targeting key driver genes. Breakage-fusion-bridge cycles and translocations activated oncogenes (BMI1, MIR17HG, TERT, MYC, and MYCN), generating gene amplifications and remodeling regulatory regions. cMCL carried significant higher numbers of structural variants, copy number alterations, and driver changes than nnMCL, with exclusive alterations of ATM in cMCL, whereas TP53 and TERT alterations were slightly enriched in nnMCL. Several drivers had prognostic impact, but only TP53 and MYC aberrations added value independently of genomic complexity. An increasing genomic complexity, together with the presence of breakage-fusion-bridge cycles and high DNA methylation changes related to the proliferative cell history, defines patients with different clinical evolution.


Information processing during NREM sleep and sleep quality in insomnia.
Insomnia sufferers (INS) are cortically hyperaroused during sleep, which seems to translate into altered information processing during nighttime. While information processing, as measured by event-related potentials (ERPs), during wake appears to be associated with sleep quality of the preceding night, the existence of such an association during nighttime has never been investigated. This study aims to investigate nighttime information processing among good sleepers (GS) and INS while considering concomitant sleep quality. Following a multistep clinical evaluation, INS and GS participants underwent 4 consecutive nights of PSG recordings in the sleep laboratory. Thirty nine GS (mean age 34.56±9.02) and twenty nine INS (mean age 43.03±9.12) were included in the study. ERPs (N1, P2, N350) were recorded all night on Night 4 (oddball paradigm) during NREM sleep. Regardless of sleep quality, INS presented a larger N350 amplitude during SWS (p=0.042) while GS showed a larger N350 amplitude during late-night stage 2 sleep (p=0.004). Regardless of diagnosis, those who slept objectively well showed a smaller N350 amplitude (p=0.020) while those who slept subjectively well showed a smaller P2 (p<0.001) and N350 amplitude (p=0.006). Also, those who reported an objectively bad night as good showed smaller P2 (p< 0.001) and N350 (p=0.010) amplitudes. Information processing seems to be associated with concomitant subjective and objective sleep quality for both GS and INS. However, INS show an alteration in information processing during sleep, especially for inhibition processes, regardless of their sleep quality.


Comparison of lead bioavailability in F344 rats fed lead acetate, lead oxide, lead sulfide, or lead ore concentrate from Skagway, Alaska.
An animal model using rats was developed to initiate investigations on the bioavailability of different sources of environmental lead. Lead must be absorbed and transported to target organs like brain, liver, kidney, and bone, before susceptible cells can be harmed. The bioavailability and therefore the toxicity of lead are dependent upon the route of exposure, dose, chemical structure, solubility, particle size, matrix incorporation, and other physiological and physicochemical factors. In the present study male F344 rats were fed < or = 38 microns size particles of lead sulfide, lead oxide, lead acetate, and a lead ore concentrate from Skagway, Alaska, mixed into the diet at doses of 0, 10, 30, and 100 ppm as lead for 30 d. No mortality or overt symptoms of lead toxicity were observed during the course of the study. Maximum blood lead concentrations attained in the 100 ppm groups were approximately 80 micrograms/dl in rats fed lead acetate and lead oxide, and were approximately 10 micrograms/dl in those fed lead sulfide and lead ore concentrate. Maximum bone lead levels in rats fed soluble lead oxide and lead acetate were much higher (approximately 200 micrograms/g) than those seen in rats fed the less soluble lead sulfide and lead ore (approximately 10 micrograms); kidney lead concentrations were also about 10-fold greater in rats fed the more soluble compared to the less soluble lead compounds. However, strong correlations between dose and tissue lead concentrations were observed in rats fed each of the four different lead compounds. Kidney lesions graded as minimal occurred in 7/10 rats fed 30 ppm and in 10/10 rats fed 100 ppm lead acetate, but not at lower doses or from other lead compounds. Similarly, urinary aminolevulinic acid excretion, a biomarker for lead toxicity, was increased in rats fed 100 ppm lead acetate or lead oxide, but was unaffected at lower doses or by the less soluble lead compounds. Although the histological and biochemical responses to lead toxicity were restricted to the more soluble lead compounds in this study, lead from Skagway lead ore concentrate and lead sulfide was also bioavailable, and accumulated in proportion to dose in vulnerable target organs such as bone and kidney. Longer-term studies with different mining materials are being conducted to determine if tissue lead continues to increase, and whether the levels attained are toxic. Data from such studies can be used to compare the toxicity and bioavailability of lead from different sources in the environment.


Reorientational dynamics of highly asymmetric binary non-polymeric mixtures - a dielectric spectroscopy study.
We present an analysis of dielectric spectra measured for a specially designed non-polymeric asymmetric binary glass former characterized by a large difference of the component's Tg (ΔTg = 216 K). We cover the whole additive concentration range from 4% up to 90% (by mass). Two main relaxations α1 and α2 are identified, which are characterized by well separated time scales and are attributed to the dynamics associated with the high-Tg component (α1) and the low-Tg component (α2). Frequency-temperature superposition does not apply. To cope with the extraordinary spectral broadening, we introduce a model consisting of a generalized Cole-Davidson (α1) and a Havriliak-Negami function with a low frequency truncation (α2). Whereas the α1-relaxation reflects essentially homogeneous dynamics and its spectra mainly broaden on the high-frequency flank of the relaxation peak, the α2-relaxation becomes broader on the low-frequency side reflecting pronounced dynamic heterogeneity in a more or less arrested matrix of high-Tg molecules. From the extracted time constants, two glass transition temperatures Tg1 and Tg2 can be derived, showing a non-trivial concentration dependence for Tg2. Supplementary, we find a β-relaxation. The total relaxation strength Δε strongly deviates from ideal mixing, and therefore care has to be taken interpreting the corresponding Δεαi as representation of molecular populations.


Global coronal decompensation and adult spinal deformity surgery: comparison of upper-thoracic versus lower-thoracic proximal fixation for long fusions.
Deterioration of global coronal alignment (GCA) may be associated with worse outcomes after adult spinal deformity (ASD) surgery. The impact of fusion length and upper instrumented vertebra (UIV) selection on patients with this complication is unclear. The authors' objective was to compare outcomes between long sacropelvic fusion with upper-thoracic (UT) UIV and those with lower-thoracic (LT) UIV in patients with worsening GCA ≥ 1 cm. This was a retrospective analysis of a prospective multicenter database of consecutive ASD patients. Index operations involved instrumented fusion from sacropelvis to thoracic spine. Global coronal deterioration was defined as worsening GCA ≥ 1 cm from preoperation to 2-year follow-up. Of 875 potentially eligible patients, 560 (64%) had complete 2-year follow-up data, of which 144 (25.7%) demonstrated worse GCA at 2-year postoperative follow-up (35.4% of UT patients vs 64.6% of LT patients). At baseline, UT patients were younger (61.6 ± 9.9 vs 64.5 ± 8.6 years, p = 0.008), a greater percentage of UT patients had osteoporosis (35.3% vs 16.1%, p = 0.009), and UT patients had worse scoliosis (51.9° ± 22.5° vs 32.5° ± 16.3°, p < 0.001). Index operations were comparable, except UT patients had longer fusions (16.4 ± 0.9 vs 9.7 ± 1.2 levels, p < 0.001) and operative duration (8.6 ± 3.2 vs 7.6 ± 3.0 hours, p = 0.023). At 2-year follow-up, global coronal deterioration averaged 2.7 ± 1.4 cm (1.9 to 4.6 cm, p < 0.001), scoliosis improved (39.3° ± 20.8° to 18.0° ± 14.8°, p < 0.001), and sagittal spinopelvic alignment improved significantly in all patients. UT patients maintained smaller positive C7 sagittal vertical axis (2.7 ± 5.7 vs 4.7 ± 5.7 cm, p = 0.014). Postoperative 2-year health-related quality of life (HRQL) significantly improved from baseline for all patients. HRQL comparisons demonstrated that UT patients had worse Scoliosis Research Society-22r (SRS-22r) Activity (3.2 ± 1.0 vs 3.6 ± 0.8, p = 0.040) and SRS-22r Satisfaction (3.9 ± 1.1 vs 4.3 ± 0.8, p = 0.021) scores. Also, fewer UT patients improved by ≥ 1 minimal clinically important difference in numerical rating scale scores for leg pain (41.3% vs 62.7%, p = 0.020). Comparable percentages of UT and LT patients had complications (208 total, including 53 reoperations, 77 major complications, and 78 minor complications), but the percentage of reoperated patients was higher among UT patients (35.3% vs 18.3%, p = 0.023). UT patients had higher reoperation rates of rod fracture (13.7% vs 2.2%, p = 0.006) and pseudarthrosis (7.8% vs 1.1%, p = 0.006) but not proximal junctional kyphosis (9.8% vs 8.6%, p = 0.810). In ASD patients with worse 2-year GCA after long sacropelvic fusion, UT UIV was associated with worse 2-year HRQL compared with LT UIV. This may suggest that residual global coronal malalignment is clinically less tolerated in ASD patients with longer fusion to the proximal thoracic spine. These results may inform operative planning and improve patient counseling.


A Lys644Glu substitution in fibroblast growth factor receptor 3 (FGFR3) causes dwarfism in mice by activation of STATs and ink4 cell cycle inhibitors.
Missense mutations of human fibroblast growth factor receptor 3 (FGFR3) result in several skeletal dysplasias, including hypochondroplasia, achondroplasia and thanatophoric dysplasia. To study the function of FGFR3 in bone growth and to create animal models for the FGFR3-related inherited skeletal disorders, we introduced a point mutation (Lys644Glu) into the murine FGFR3 genome using a knock-in approach. We found that the Lys644Glu mutation resulted in retarded endochondral bone growth with severity directly linked to the expression level of the mutated Fgfr3. Mice heterozygous for the mutation ( Fgfr3(TD/+) ) expressed the mutant allele at approximately 20% of the wild-type level and exhibited a mild bone dysplasia. However, when the copy number of the mutant allele increased from one (Fgfr3(TD/+) to two (Fgfr3(TD/TD), the retardation of bone growth became more severe and showed phenotypes resembling those of achondroplasia patients, characterized by a dramatically reduced proliferation of growth plate cartilage, macrocephaly and shortening of the long bones, which was most pronounced in the femur. Molecular analysis revealed that expression of the mutant receptor caused the activation of Stat1, Stat5a and Stat5b, and the up-regulation of p16, p18 and p19 cell cycle inhibitors, leading to dramatic expansion of the resting zone of chondrocytes at the expense of the proliferating chondrocytes. The mutant growth plates consequently were in a less active state and generated fewer maturing and hypertrophic chondrocytes. These data provide direct genetic evidence that the point mutations in FGFR3 cause human skeletal dysplasias and uncover a mechanism through which the FGFR3 signals regulate bone growth by modulating expression of Stats and ink4 cell cycle inhibitors.


Direct analysis by time-of-flight secondary ion mass spectrometry reveals action of bacterial laccase-mediator systems on both hardwood and softwood samples.
The modification and degradation of lignin play a vital role in carbon cycling as well as production of biofuels and bioproducts. The possibility of using bacterial laccases for the oxidation of lignin offers a route to utilize existing industrial protein expression techniques. However, bacterial laccases are most frequently studied on small model compounds that do not capture the complexity of lignocellulosic materials. This work studied the action of laccases from Bacillus subtilis and Salmonella typhimurium (EC 184.108.40.206) on ground wood samples from yellow birch (Betula alleghaniensis) and red spruce (Picea rubens). The ability of bacterial laccases to modify wood can be facilitated by small molecule mediators. Herein, 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid) (ABTS), gallic acid and sinapic acid mediators were tested. Direct analysis of the wood samples was achieved by time-of-flight secondary ion mass spectrometry (ToF-SIMS), a surface sensitive mass spectrometry technique that has characteristic peaks for H, G and S lignin. The action of the bacterial laccases on both wood samples was demonstrated and revealed a strong mediator influence. The ABTS mediator led to delignification, evident in an overall increase of polysaccharide peaks in the residual solid, along with equal loss of G and S-lignin peaks. The gallic acid mediator demonstrated minimal laccase activity. Meanwhile, the sinapic acid mediator altered the S/G peak ratio consistent with mediator attaching to the wood solids. The current investigation demonstrates the action of bacterial laccase-mediator systems directly on woody materials, and the potential of using ToF-SIMS to uncover the fundamental and applied role of bacterial enzymes in lignocellulose conversion.


Circulation of bluetongue virus in goats in the Karamoja region of Uganda.
The presence of bluetongue virus (BTV) in indigenous goats from the Karamoja region of northern Uganda was investigated. A total of 300 goats were sampled (serum and whole blood) from five districts within the Karamoja region. The samples were analysed for the presence of bluetongue (BT) antibodies using a commercial Enzyme-linked immunosorbent assay (ELISA) and for the presence of BTV viral RNA by real-time Reverse transcription polymerase chain reaction (RT-PCR), because BTV is an RNA virus. Of the 300 goats tested, 269 (90%) were positive for BTV antibodies, indicating high levels of BTV circulation within the region. Out of the 150 whole blood samples tested for the presence of the virus by real-time RT-PCR, 84 (56%) were positive for BTV RNA. This study, which is the first of its kind in Uganda, showed a high seroprevalence of BT antibodies and active circulation of BTV in a high proportion of goats in the Karamoja region.