Patient- and Surgeon-Level Variation in Patient-Reported Sexual Function Outcomes Following Radical Prostatectomy Over 2 Years: Results From a Statewide Surgical Improvement Collaborative.
Of patient-reported outcomes for individuals undergoing radical prostatectomy, sexual function outcomes are among the most reported and the most detrimental to quality of life. Understanding variations at the patient and surgeon level may inform collaborative quality improvement. To describe patient- and surgeon-level sexual function outcomes for patients undergoing radical prostatectomy in the Michigan Urological Surgery Improvement Collaborative (MUSIC) and to examine the correlation between surgeon case volume and sexual function outcomes. This is a prospective cohort study using the MUSIC registry and patient-reported sexual function outcome data. Patient- and surgeon-level variation in sexual function outcomes were examined among patients undergoing radical prostatectomy from May 2014 to August 2019. Sexual function outcome data were collected using validated questionnaires, which were completed before surgery and at 3, 6, 12, and 24 months' follow-up following surgery. All participants were male. Race and ethnicity data were self-reported and were included to examine potential variation in outcomes by race and/or ethnicity. Data were analyzed from January 2021 to March 2021. There were 4 outcomes in this study, including the 26-item Expanded Prostate Cancer Index Composite (EPIC-26) sexual function scores at 3, 6, 12, and 24 months' follow-up; patient-level sexual function recovery at 12- and 24-month follow-up; surgeon-level variation in sexual function outcomes at 12- and 24-month follow-up; and correlation between surgeon case volume and sexual function outcomes. A total of 1426 male patients met inclusion criteria for this study. The median (IQR) age was 64 (58-68) years. A total of 115 participants (8%) were Black, 1197 (84%) were White, 25 (2%) were of another race or ethnicity (consolidated owing to low numbers), and 89 (6%) were of unknown race or ethnicity. Among patients undergoing bilateral nerve-sparing radical prostatectomy, mean (SD) EPIC-26 sexual function scores at 12- and 24-month follow-up (12 months, 39 [28]; 24 months, 63 [29]) did not return to baseline levels. There was wide variation in EPIC-26 sexual function scores at both 12-month follow-up (range, 23-69; P < .001) and 24-month follow-up (range, 27-64; P < .001). Similar variations were found in EPIC-26 sexual function scores and recovery of sexual function by surgeon. Recovery rates ranged from 0% to 40% of patients at 12-month follow-up (18 surgeons; P < .001) and 3% to 44% of patients at 24-month follow-up (12 surgeons; P < .001). Surgeon case volume and sexual function outcomes were not significantly correlated. On multivariable analysis, the following variables were associated with better recovery at 24-month follow-up: younger age (P < .001), lower baseline EPIC-26 sexual function score (P < .001), lower Gleason score (P = .05), and nonobesity (P = .03). In this study, there was significant patient- and surgeon-level variation in sexual function recovery over 2 years following radical prostatectomy. Variation in surgeon-level sexual function outcomes presents an opportunity and model for surgical collaborative quality improvement.


Cilia functions in development.
Recent advances in developmental genetics and human disease gene cloning have highlighted the essential roles played by cilia in developmental cell fate decisions, left-right asymmetry, and the pathology of human congenital disorders. Hedgehog signaling in sensory cilia illustrates the importance of trafficking receptors to the cilia membrane (Patched and Smoothened) and the concept of cilia 'gatekeepers' that restrict entry and egress of cilia proteins (Suppressor of fused: Gli complexes). Cilia-driven fluid flow in the embryonic node highlights the role of motile cilia in both generation and detection of mechanical signals in development. In this brief review I select examples of recent studies that have clarified and consolidated our understanding of the role of cilia in development.


Are major bleeding events from falls more likely in patients on warfarin?
There is no evidence of increased risk for major bleeding as a result of falls in hospitalized patients taking warfarin (strength of recommendation [SOR]: B, based on retrospective cohort studies). In the average patient taking warfarin for atrial fibrillation, the risk of intracranial hemorrhage from a fall is much smaller than the benefit gained from reducing risk of stroke (SOR: A, based on decision analysis of systematic reviews with sensitivity analysis).


Surgical resection of large incompletely treated intracranial arteriovenous malformations following stereotactic radiosurgery.
Although radiosurgery is effective in obliterating small arteriovenous malformations (AVMs), it has a lower success rate for thrombosing larger AVMs. The authors surgically resected AVMs from 33 patients ranging in age from 7 to 64 years (mean 30.4 years) 1 to 11 years after radiosurgery. Initial AVM volumes were 0.8 to 117 cm3 (mean 21.6 cm3), and doses ranged from 4.6 to 45 GyE (mean 21.2 GyE). Of 27 AVMs in eloquent or critical areas, 10 were located in language, motor, sensory, or visual cortex, 11 in the basal ganglia/thalamus, one each in the brainstem, hypothalamus, and cerebellum, and three in the corpus callosum. Venous drainage was deep in 13, superficial in 12, or both in eight lesions. Spetzler-Martin grades were II in one, III in 12, IV in 16, and V in four patients. Eight patients experienced rebleeding after radiosurgery but prior to surgery. Three patients developed radiation necrosis and 25 underwent endovascular embolization prior to surgery. At surgery the AVMs were found to be markedly less vascular, partially thrombosed, and more easily resected, compared to those seen in patients who had not undergone radiosurgery. Pathological investigation showed endothelial proliferation with hyaline and calcium in vessel walls. There was partial or complete thrombosis of some AVM vessels and evidence of vessel and brain necrosis in many cases. Complete resection was achieved in 28 patients and partial resection in five. Clinical outcome was excellent or good in 31 cases, and two patients died of rebleeding from residual AVM. Four patients' conditions worsened following microsurgical resection. Final clinical outcome was largely related to the pretreatment grade. Radiosurgery several years prior to open microsurgery may prove to be a useful adjunct in treating unusually large and complex AVMs.


Identification of transplanted human cells in animal tissues.
The potential of cell and gene therapy has generated extensive interest over the past several years. More recently, identification of stem cells of various types, especially embryonic stem cells, reinforced this interest. Systematic studies are now being launched to define the biology of various stem cells, including after transplantation of cells in mmunodeficient animals. This requires robust and unequivocal means to identify transplanted cells. Ideally, it should be possible to screen animal tissues for human cells with relatively simpler methods, followed by more precise localization of transplanted cells. We describe the application of conserved primate Charcot-Marie-Tooth disease type 1A repeat element for polymerase chain reaction-based screening of animal tissues for human cells. Similarly, direct polymerase chain reaction labeling of pancentromeric human alphoid sequences with digoxigenin-UTP generates in situ hybridization probes for identifying transplanted human cells. This pancentromeric probe identifies human cells irrespective of the original tissue source and can be combined with additional in situ methods to analyze cell differentiation. Incorporation of these strategies will facilitate translational studies aimed at understanding mechanisms concerning the trafficking, engraftment, proliferation, differentiation and function of human stem cells in animals.


[Viability and efects of the probiotic lactobacillus aracaseissp aracasei in Chilean low-fat Gauda cheese].
The objective of this study is to infer the survival of the probiotic Lactobacillus paracasei ssp paracasei added during the processing of low-fat Gouda cheese, during the maturation (21 days) and the commercialization (14 days), in order to see the influence that this organism has on the quality of the product. The treatments were: TI (control): Gouda with normal fat content; T2: Low fat Gouda cheese (QGS) T3: QGS, with additional probiotic added with the initial culture; T4: QGS, with the probiotic added in the cooking of the curd. For the count of the probiotic, the methodology was used set forth by the American Public Health Association, (APHA), proteolysis by the method of soluble tyrosine in trichloroacetic acid (TCA), and the physical and chemical analysis using the methods standardized by the International Dairy Federation, and Chilean normative. The counts obtained were from 10(8) ufc/g y 10(7) ufc/g in the cheeses that had the probiotic additive (T3 and T4, respectively) close to the level of innocuous (10(8) ufc/g) during the study period of 35 days. The proteolysis incremented normally, and was the same in all of the treatments during the 35 days studied. The treatments with fat-reduction presented approximately, 31% less fat than the control treatment, and also higher moistness. Within the treatments, there was no evidence of taste and general feel; in turn the cheeses with the reduction of fat resulted firmer, less cohesive, than the control, and with similar elasticity.


Unexpected Phenotype Reversion and Survival in a Zebrafish Model of Multiple Sulfatase Deficiency.
Multiple sulfatase deficiency (MSD) is a rare recessively inherited Mendelian disorder that manifests with developmental delay, neurodegeneration, skeletal deformities, facial dysmorphism, congenital growth retardation, and other clinical signs. The disorder is caused by mutations in the SUMF1 gene, which encodes the formylglycine-generating enzyme (FGE), and responsible for the activation of sulfatases. Mutations in SUMF1 result in reduced or absent FGE function with consequent compromised activities of its client sulfatases. This leads to an accumulation of enzyme substrates, such as glycosaminoglycans and sulfolipids, within lysosomes and subsequently impaired lysosome function and cellular pathology. Currently, there are no disease modifying therapeutic options for MSD patients, hence the need for more suitable animal models to investigate the disorder. Here, we describe the characterisation of a sumf1 null zebrafish model, which has negligible sulfatase activity. Our sumf1 -/- zebrafish model successfully recapitulates the pathology of MSD such as cranial malformation, altered bone development, an enlarged population of microglia, and growth retardation during early development but lacks early lethality of mouse Sumf1 -/- models. Notably, we provide evidence of recovery in MSD pathology during later developmental stages, resulting in homozygous mutants that are viable. Hence, our data suggest the possibility of a unique compensatory mechanism that allows the sumf1 -/- null zebrafish to survive better than human MSD patients and mouse Sumf1 -/- models.


Topical Application of Hyaluronic Acid-RGD Peptide-Coated Gelatin/Epigallocatechin-3 Gallate (EGCG) Nanoparticles Inhibits Corneal Neovascularization Via Inhibition of VEGF Production.
Neovascularization (NV) of the cornea disrupts vision which leads to blindness. Investigation of antiangiogenic, slow-release and biocompatible approaches for treating corneal NV is of great importance. We designed an eye drop formulation containing gelatin/epigallocatechin-3-gallate (EGCG) nanoparticles (NPs) for targeted therapy in corneal NV. Gelatin-EGCG self-assembled NPs with hyaluronic acid (HA) coating on its surface (named GEH) and hyaluronic acid conjugated with arginine-glycine-aspartic acid (RGD) (GEH-RGD) were synthesized. Human umbilical vein endothelial cells (HUVECs) were used to evaluate the antiangiogenic effect of GEH-RGD NPs in vitro. Moreover, a mouse model of chemical corneal cauterization was employed to evaluate the antiangiogenic effects of GEH-RGD NPs in vivo. GEH-RGD NP treatment significantly reduced endothelial cell tube formation and inhibited metalloproteinase (MMP)-2 and MMP-9 activity in HUVECs in vitro. Topical application of GEH-RGD NPs (once daily for a week) significantly attenuated the formation of pathological vessels in the mouse cornea after chemical cauterization. Reduction in both vascular endothelial growth factor (VEGF) and MMP-9 protein in the GEH-RGD NP-treated cauterized corneas was observed. These results confirm the molecular mechanism of the antiangiogenic effect of GEH-RGD NPs in suppressing pathological corneal NV.


External quality control performance in clinical chemistry: experience in Kenya.
Analysis of eleven biochemical laboratory tests was done during an International External Quality Assessment Scheme (IEQAS) in which the clinical chemistry laboratory at Kenyatta National Hospital participated. Technicon SMA II continuous flow system was used in the biochemical analyses apart from glucose which was assayed manually by the glucose oxidase method. Using the standard deviation index (SDI), twenty six percent of the results were found to be outside the two standard deviation (2SD) limit. However, when variance index score (VIS) was used, 42% of the results were found to be outliers. Overall, our laboratory performed poorly compared to other laboratories in both the IEQAS and the United Kingdom External Quality Assurance Scheme (UKEQAS). This poor performance is attributed to the use of improper equipment which is not regularly maintained, lack of diagnostic reagents, lack of quality control (QC) materials and inadequate staff training in the field of quality control.


Carbon-Based Polymer Nanocomposite for High-Performance Energy Storage Applications.
In recent years, numerous discoveries and investigations have been remarked for the development of carbon-based polymer nanocomposites. Carbon-based materials and their composites hold encouraging employment in a broad array of fields, for example, energy storage devices, fuel cells, membranes sensors, actuators, and electromagnetic shielding. Carbon and its derivatives exhibit some remarkable features such as high conductivity, high surface area, excellent chemical endurance, and good mechanical durability. On the other hand, characteristics such as docility, lower price, and high environmental resistance are some of the unique properties of conducting polymers (CPs). To enhance the properties and performance, polymeric electrode materials can be modified suitably by metal oxides and carbon materials resulting in a composite that helps in the collection and accumulation of charges due to large surface area. The carbon-polymer nanocomposites assist in overcoming the difficulties arising in achieving the high performance of polymeric compounds and deliver high-performance composites that can be used in electrochemical energy storage devices. Carbon-based polymer nanocomposites have both advantages and disadvantages, so in this review, attempts are made to understand their synergistic behavior and resulting performance. The three electrochemical energy storage systems and the type of electrode materials used for them have been studied here in this article and some aspects for example morphology, exterior area, temperature, and approaches have been observed to influence the activity of electrochemical methods. This review article evaluates and compiles reported data to present a significant and extensive summary of the state of the art.


Should the definition of micropenis vary according to ethnicity?
We determined whether the existing reference values for the diagnosis of micropenis are appropriate for optimal care of neonates in a multiethnic environment like Vancouver. The stretched penile length and width were measured in 105 full-term newborn males of Caucasian (n = 40), Chinese (n = 40) and East-Indian origin (n = 25). Mean length -2.5 SD was used for the definition of micropenis and was 2.6, 2.5 and 2.3 cm for Caucasian, East-Indian and Chinese babies, respectively (p < 0.05). This is close to the widely accepted recommendation that a penile length of 2.4- 2.5 cm be considered as the lowest limit for the definition of micropenis. Mean penile length and diameter are slightly but significantly smaller in newborns of Chinese origin compared to newborns of Caucasian and East-Indian origins.


Modeling and simulation of genetic regulatory systems: a literature review.
In order to understand the functioning of organisms on the molecular level, we need to know which genes are expressed, when and where in the organism, and to which extent. The regulation of gene expression is achieved through genetic regulatory systems structured by networks of interactions between DNA, RNA, proteins, and small molecules. As most genetic regulatory networks of interest involve many components connected through interlocking positive and negative feedback loops, an intuitive understanding of their dynamics is hard to obtain. As a consequence, formal methods and computer tools for the modeling and simulation of genetic regulatory networks will be indispensable. This paper reviews formalisms that have been employed in mathematical biology and bioinformatics to describe genetic regulatory systems, in particular directed graphs, Bayesian networks, Boolean networks and their generalizations, ordinary and partial differential equations, qualitative differential equations, stochastic equations, and rule-based formalisms. In addition, the paper discusses how these formalisms have been used in the simulation of the behavior of actual regulatory systems.


An algorithm that identifies coronary and heart failure events in the electronic health record.
The advent of universal health care coverage in the United States and the use of electronic health records can make the medical record a disease surveillance tool. The objective of our study was to identify criteria that accurately categorize acute coronary and heart failure events by using electronic health record data exclusively so that the medical record can be used for surveillance without manual record review. We serially compared 3 computer algorithms to manual record review. The first 2 algorithms relied on ICD-9-CM (International Classification of Diseases, 9th Revision, Clinical Modification) codes, troponin levels, electrocardiogram (ECG) data, and echocardiograph data. The third algorithm relied on a detailed coding system, Intelligent Medical Objects, Inc., (IMO) interface terminology, troponin levels, and echocardiograph data. Cohen's κ for the initial algorithm was 0.47 (95% confidence interval [CI], 0.41-0.54). Cohen's κ was 0.61 (95% CI, 0.55-0.68) for the second algorithm. Cohen's κ for the third algorithm was 0.99 (95% CI, 0.98-1.00). Electronic medical record data are sufficient to categorize coronary heart disease and heart failure events without manual record review. However, only moderate agreement with medical record review can be achieved when the classification is based on 4-digit ICD-9-CM codes because ICD-9-CM 410.9 includes myocardial infarction with elevation of the ST segment on ECG (STEMI) and myocardial infarction without elevation of the ST segment on ECG (nSTEMI). Nearly perfect agreement can be achieved using IMO interface terminology, a more detailed coding system that tracks to ICD9, ICD10 (International Classification of Diseases, Tenth Revision, Clinical Modification), and SnoMED-CT (Systematized Nomenclature of Medicine - Clinical Terms).


Nitric oxide-dependent generation of reactive species in sickle cell disease. Actin tyrosine induces defective cytoskeletal polymerization.
The intermittent vascular occlusion occurring in sickle cell disease (SCD) leads to ischemia-reperfusion injury and activation of inflammatory processes including enhanced production of reactive oxygen species and increased expression of inducible nitric-oxide synthase (NOS2). Appreciating that impaired nitric oxide-dependent vascular function and the concomitant formation of oxidizing and nitrating species occur in concert with increased rates of tissue reactive oxygen species production, liver and kidney NOS2 expression, tissue 3-nitrotyrosine (NO(2)Tyr) formation and apoptosis were evaluated in human SCD tissues and a murine model of SCD. Liver and kidney NOS2 expression and NO(2)Tyr immunoreactivity were significantly increased in SCD mice and humans, but not in nondiseased tissues. TdT-mediated nick end-label (TUNEL) staining showed apoptotic cells in regions expressing elevated levels of NOS2 and NO(2)Tyr in all SCD tissues. Gas chromatography mass spectrometry analysis revealed increased plasma protein NO(2)Tyr content and increased levels of hepatic and renal protein NO(2)Tyr derivatives in SCD (21.4 +/- 2.6 and 37.5 +/- 7.8 ng/mg) versus wild type mice (8.2 +/- 2.2 and 10 +/- 1.2 ng/mg), respectively. Western blot analysis and immunoprecipitation of SCD mouse liver and kidney proteins revealed one principal NO(2)Tyr-containing protein of 42 kDa, compared with controls. Enzymatic in-gel digestion and MALDI-TOF mass spectrometry identified this nitrated protein as actin. Electrospray ionization and fragment analysis by tandem mass spectrometry revealed that 3 of 15 actin tyrosine residues are nitrated (Tyr(91), Tyr(198), and Tyr(240)) at positions that significantly modify actin assembly. Confocal microscopy of SCD human and mouse tissues revealed that nitration led to morphologically distinct disorganization of filamentous actin. In aggregate, we have observed that the hemoglobin point mutation of sickle cell disease that mediates hemoglobin polymerization defects is translated, via inflammatory oxidant reactions, into defective cytoskeletal polymerization.


Direct momentum-resolved observation of one-dimensional confinement of externally doped electrons within a single subnanometer-scale wire.
Cutting-edge research in the band engineering of nanowires at the ultimate fine scale is related to the minimum scale of nanowire-based devices. The fundamental issue at the subnanometer scale is whether angle-resolved photoemission spectroscopy (ARPES) can be used to directly measure the momentum-resolved electronic structure of a single wire because of the difficulty associated with assembling single wire into an ordered array for such measurements. Here, we demonstrated that the one-dimensional (1D) confinement of electrons, which are transferred from external dopants, within a single subnanometer-scale wire (subnanowire) could be directly measured using ARPES. Convincing evidence of 1D electron confinement was obtained using two different gold subnanowires with characteristic single metallic bands that were alternately and spontaneously ordered on a stepped silicon template, Si(553). Noble metal atoms were adsorbed at room temperature onto the gold subnanowires while the overall structure of the wires was maintained. Only one type of gold subnanowire could be controlled using external noble metal dopants without transforming the metallic band of the other type of gold subnanowires. This result was confirmed by scanning tunnelling microscopy experiments and first-principles calculations. The selective control clearly showed that externally doped electrons could be confined within a single gold subnanowire. This experimental evidence was used to further investigate the effects of the disorder induced by external dopants on a single subnanowire using ARPES.


Vegetable Nitrate Intakes Are Associated with Reduced Self-Reported Cardiovascular-Related Complications within a Representative Sample of Middle-Aged Australian Women, Prospectively Followed up for 15 Years.
Nitric oxide (NO) facilitates anti-atherosclerotic effects. Vegetables are a major source of dietary nitrate. Experimental data indicates that dietary nitrate can significantly reduce major risk factors for atherosclerosis and subsequent cardiovascular disease (CVD), as nitrate can be metabolized to produce NO via the nitrate-nitrite-NO pathway. The purpose of this study was to prospectively investigate the association between habitual dietary nitrate intakes and the incidence of self-reported CVD-related complications within a representative sample of middle-aged Australian women (1946⁻1951 cohort of the Australian Longitudinal Study on Women's Health). Women free from disease at baseline who had completed the food frequency questionnaire data were included. Generalized estimating equations were used to estimate odds ratios (OR) and 95% confidence intervals (95% CI) across quartiles for nitrate intakes. Of the 5324 women included for analysis, there were 1951 new cases of CVD-related complications over 15-years of follow-up. Women reporting higher total dietary nitrate intakes (Q4 > 78.2 mg/day) and vegetable nitrate intakes (Q4 > 64.4 mg/day) were 25% and 27% reduced risk of developing CVD-related complications respectively, compared with women reporting low total (Q1 < 45.5 mg/day) and vegetable nitrate intakes (Q1 < 34.8 mg/day). Our findings were consistent with other observational data indicating that dietary nitrate may explain some of the cardiovascular benefits of vegetable consumption.


Analysis of the subunit stoichiometries in viral entry.
Virions of the Human Immunodeficiency Virus (HIV) infect cells by first attaching with their surface spikes to the CD4 receptor on target cells. This leads to conformational changes in the viral spikes, enabling the virus to engage a coreceptor, commonly CCR5 or CXCR4, and consecutively to insert the fusion peptide into the cellular membrane. Finally, the viral and the cellular membranes fuse. The HIV spike is a trimer consisting of three identical heterodimers composed of the gp120 and gp41 envelope proteins. Each of the gp120 proteins in the trimer is capable of attaching to the CD4 receptor and the coreceptor, and each of the three gp41 units harbors a fusion domain. It is still under debate how many of the envelope subunits within a given trimer have to bind to the CD4 receptors and to the coreceptors, and how many gp41 protein fusion domains are required for fusion. These numbers are referred to as subunit stoichiometries. We present a mathematical framework for estimating these parameters individually by analyzing infectivity assays with pseudotyped viruses. We find that the number of spikes that are engaged in mediating cell entry and the distribution of the spike number play important roles for the estimation of the subunit stoichiometries. Our model framework also shows why it is important to subdivide the question of the number of functional subunits within one trimer into the three different subunit stoichiometries. In a second step, we extend our models to study whether the subunits within one trimer cooperate during receptor binding and fusion. As an example for how our models can be applied, we reanalyze a data set on subunit stoichiometries. We find that two envelope proteins have to engage with CD4-receptors and coreceptors and that two fusion proteins must be revealed within one trimer for viral entry. Our study is motivated by the mechanism of HIV entry but the experimental technique and the model framework can be extended to other viral systems as well.


Characterization of platelet-monocyte complexes in HIV-1-infected individuals: possible role in HIV-associated neuroinflammation.
HIV-1-associated neuroinflammation persists even with effective combined antiretroviral therapy, and it is associated with the presence of activated monocytes/macrophages within the CNS. To infiltrate the CNS, monocytes transmigrate across the selectively permeable blood-brain barrier, which is compromised during HIV-1 infection. Interestingly, platelet-derived excess soluble CD40 ligand found in the plasma and cerebrospinal fluid of HIV-1-infected individuals with cognitive impairment has previously been implicated in increased blood-brain barrier permeability. In this study we show that soluble CD40 ligand also promotes the formation of complexes between inflammatory monocytes and activated platelets (PMCs), which are detected by flow cytometry as monocytes that express excess of CD61, a platelet marker, and that these complexes are increased in individuals with HIV-1 infection. PMCs exhibit an enhanced ability to adhere to human brain microvascular endothelial cells as compared with monocytes alone, and they migrate across the transendothelial barrier. These complexes can be found marginalized in the lumen of postcapillary venules in postmortem brain tissue derived from cases of HIV-1-associated encephalitis. The extravasation of monocytes across the brain endothelium may exacerbate neuroinflammation, indicating that enhancing this event via platelet interaction may be a contributing factor in the development of cognitive impairment. Thus, dampening platelet activation, and in turn PMC formation, with antiplatelet agents may prove beneficial in developing adjunctive therapies for use in combination with combined antiretroviral therapy in an effort to reduce HIV-1-associated neurologic deficit.


The mathematical modelling of cell kinetics in corneal epithelial wound healing.
This paper considers the comparison of experimental spatial and temporal data of mitotic rates measured during corneal epithelial wound healing (CEWH) of a rat model with the predictions of a computer modelling framework. We begin by briefly showing that previous models, used in the study of corneal epithelial wound healing speeds, are inadequate for the study of cell kinetics. We proceed to formulate a new modelling framework more suited to such a study. This framework is simulated in its simplest form, and the results from this motivate a new realisation of the modelling framework, including a caricature of age structuring. Finally, a model with a simple representation of juxtacrine signalling is considered. The final model captures many, though not all, of the trends of the experimental data. This paper thus lays a foundation for the modelling of the cell kinetics of corneal epithelial wound healing, and yields valuable insight regarding the important mechanisms a model should consider in order to reproduce the observed experimental trends.


High-dimensional NMR spectra for structural studies of biomolecules.
Recent developments in the acquisition and processing of NMR data sets facilitate the recording of ultra-high-resolution NMR spectra in a reasonable time. The new experiments allow easy resonance assignment for folded and unfolded proteins, as well as the precise determination of spectral parameters, for example, chemical shifts, NOE contacts, coupling constants or cross-correlated relaxation rates. Owing to exceptional resolution of 4D-6D spectroscopy, detailed studies of biomolecules of unprecedented complexity are now possible. Herein, the principles of acquisition and processing methods are presented. The main applications of high-dimensional NMR experiments, including backbone and side-chain resonance assignment in proteins, as well as heteronuclear edited NOE techniques are reviewed.


Structure and sequence determinants required for the RNA editing of ADAR2 substrates.
ADAR2 is a double-stranded RNA-specific adenosine deaminase involved in the editing of mammalian RNAs by the site-specific conversion of adenosine to inosine. We have demonstrated previously that ADAR2 can modify its own pre-mRNA, leading to the creation of a proximal 3'-splice junction containing a non-canonical adenosine-inosine (A-I) dinucleotide. Alternative splicing to this proximal acceptor shifts the reading frame of the mature mRNA transcript, resulting in the loss of functional ADAR2 expression. Both evolutionary sequence conservation and mutational analysis support the existence of an extended RNA duplex within the ADAR2 pre-mRNA formed by base-pairing interactions between regions approximately 1.3-kilobases apart in intron 4 and exon 5. Characterization of ADAR2 pre-mRNA transcripts isolated from adult rat brain identified 16 editing sites within this duplex region, and sites preferentially modified by ADAR1 and ADAR2 have been defined using both tissue culture and in vitro editing systems. Statistical analysis of nucleotide sequences surrounding edited and non-edited adenosine residues have identified a nucleotide sequence bias correlating with ADAR2 site preference and editing efficiency. Among a mixed population of ADAR substrates, ADAR2 preferentially favors its own transcript, yet mutation of a poor substrate to conform to the defined nucleotide bias increases the ability of that substrate to be modified by ADAR2. These data suggest that both sequence and structural elements are required to define adenosine moieties targeted for specific ADAR2-mediated deamination.


Functional assembly of aptamer binding sites by single-molecule cut-and-paste.
Bottom up assembly of functional molecular ensembles with novel properties emerging from composition and arrangement of its constituents is a prime goal of nanotechnology. By single-molecule cut-and-paste we assembled binding sites for malachite green in a molecule-by-molecule assembly process from the two halves of a split aptamer. We show that only a perfectly joined binding site immobilizes the fluorophore and enhances the fluorescence quantum yield by several orders of magnitude. To corroborate the robustness of this approach we produced a micrometer-sized structure consisting of more than 500 reconstituted binding sites. To the best of our knowledge, this is the first demonstration of one by one bottom up functional biomolecular assembly.


Potential costs of acclimatization to a warmer climate: growth of a reef coral with heat tolerant vs. sensitive symbiont types.
One of the principle ways in which reef building corals are likely to cope with a warmer climate is by changing to more thermally tolerant endosymbiotic algae (zooxanthellae) genotypes. It is highly likely that hosting a more heat-tolerant algal genotype will be accompanied by tradeoffs in the physiology of the coral. To better understand one of these tradeoffs, growth was investigated in the Indo-Pacific reef-building coral Acropora millepora in both the laboratory and the field. In the Keppel Islands in the southern Great Barrier Reef this species naturally harbors nrDNA ITS1 thermally sensitive type C2 or thermally tolerant type D zooxanthellae of the genus Symbiodinium and can change dominant type following bleaching. We show that under controlled conditions, corals with type D symbionts grow 29% slower than those with type C2 symbionts. In the field, type D colonies grew 38% slower than C2 colonies. These results demonstrate the magnitude of trade-offs likely to be experienced by this species as they acclimatize to warmer conditions by changing to more thermally tolerant type D zooxanthellae. Irrespective of symbiont genotype, corals were affected to an even greater degree by the stress of a bleaching event which reduced growth by more than 50% for up to 18 months compared to pre-bleaching rates. The processes of symbiont change and acute thermal stress are likely to act in concert on coral growth as reefs acclimatize to more stressful warmer conditions, further compromising their regeneration capacity following climate change.


Do honeybees (Apis mellifera) differentiate between different pollen types?
Bees receive nectar and pollen as reward for pollinating plants. Pollen of different plant species varies widely in nutritional composition. In order to select pollen of appropriate nutritional quality, bees would benefit if they could distinguish different pollen types. Whether they rely on visual, olfactory and/or chemotactile cues to distinguish between different pollen types, has however been little studied. In this study, we examined whether and how Apis mellifera workers differentiate between almond and apple pollen. We used differential proboscis extension response conditioning with olfactory and chemotactile stimulation, in light and darkness, and in summer and winter bees. We found that honeybees were only able to differentiate between different pollen types, when they could use both chemotactile and olfactory cues. Visual cues further improved learning performance. Summer bees learned faster than winter bees. Our results thus highlight the importance of multisensory information for pollen discrimination.


Epigenetic dysregulation of Insulin-like growth factor (IGF)-related genes and adverse pregnancy outcomes: a systematic review.
Preterm birth (PTB), low birth weight (LBW) and small for gestational age (SGA) are leading causes of neonatal mortality and morbidity around the world. Epigenetic alterations of the human genome may be involved in the causal chain of adverse pregnancy outcomes. In this systematic review we investigated whether PTB, LBW and SGA are associated with epigenetic dysregulation of insulin-like growth factor-related genes (IGF). We searched MEDLINE and EMBASE for peer-reviewed articles about IGF and PTB, LBW and SGA published up to February 2015. Two independent reviewers selected original, controlled, human studies published in any language and graded them using the Newcastle-Ottawa Quality Assessment Scale. Disagreements were resolved by consensus with a third reviewer. Eighteen observational studies of low-to-moderate quality met the eligibility criteria out of 210 unique studies. There was substantial heterogeneity across studies. Most studies reported no, limited or borderline association between epigenetic changes (methylation or imprinting) of IGF-related genes and LBW or SGA. There were no IGF-related epigenetic studies of PTB. Overall, evidence of an association between epigenetic abnormalities of IGF-related genes and LBW or SGA was weak and inconsistent. Methodological concerns limited results validity.


Temporal recruitment of transcription factors and SWI/SNF chromatin-remodeling enzymes during adipogenic induction of the peroxisome proliferator-activated receptor gamma nuclear hormone receptor.
The peroxisome proliferator-activated receptor gamma (PPARgamma) regulates adipogenesis, lipid metabolism, and glucose homeostasis, and roles have emerged for this receptor in the pathogenesis and treatment of diabetes, atherosclerosis, and cancer. We report here that induction of the PPARgamma activator and adipogenesis forced by overexpression of adipogenic regulatory proteins is blocked upon expression of dominant-negative BRG1 or hBRM, the ATPase subunits of distinct SWI/SNF chromatin-remodeling enzymes. We demonstrate that histone hyperacetylation and the binding of C/EBP activators, polymerase II (Pol II), and general transcription factors (GTFs) initially occurred at the inducible PPARgamma2 promoter in the absence of SWI/SNF function. However, the polymerase and GTFs were subsequently lost from the promoter in cells expressing dominant-negative SWI/SNF, explaining the inhibition of PPARgamma2 expression. To corroborate these data, we analyzed interactions at the PPARgamma2 promoter in differentiating preadipocytes. Changes in promoter structure, histone hyperacetylation, and binding of C/EBP activators, Pol II, and most GTFs preceded the interaction of SWI/SNF enzymes with the PPARgamma2 promoter. However, transcription of the PPARgamma2 gene occurred only upon subsequent association of SWI/SNF and TFIIH with the promoter. Thus, induction of the PPARgamma nuclear hormone receptor during adipogenesis requires SWI/SNF enzymes to facilitate preinitiation complex function.


Angioedema due to angiotensin-converting enzyme inhibitors.
Angiotensin-converting enzyme (ACE) inhibitor associated angioedema was detected in 39 subjects (17%) of 231 consecutive patients examined in the last 5 years at our out-patient clinic for symptoms of angioedema without urticaria. In these patients, angioedema was most commonly localized to the face. The duration of ACE-inhibitor treatment at the onset of angioedema ranged from 1 day to 8 years with a median of 6 months. The time elapsed between onset of angioedema and withdrawal of ACE-inhibitor ranged from 1 day to 10 years with a median of 10 months. Delayed diagnosis is explained by the unusual characteristics of this adverse reaction: angioedema may start years after beginning the treatment and then it recurs irregularly. In fact, ACE-inhibitors seem to facilitate angioedema in predisposed subjects, rather than causing it with an allergic or idiosyncratic mechanism. Thus, while Cl-inhibitor levels are usually normal in subjects developing ACE-inhibitor-dependent angioedema, we found that ACE-inhibitors caused angioedema in Cl-inhibitor-deficient patients. Because the main inactivator of bradykinin is kininase II, which is identical with ACE, it is believed that bradykinin mediates ACE-inhibitor-dependent angioedema. We had the possibility to examine the plasma bradykinin levels in one ACE-inhibitor-treated patient during an angioedema attack and we found very high levels, but we did not find an increase of break-down products of high-molecular-weight-kininogen as observed during acute attacks in hereditary angioedema. Bradykinin fell to normal levels during remission after withdrawal of the drug. These observations indicate that in ACE-inhibitor-induced angioedema, contrary to hereditary angioedema, the reduction of bradykinin catabolic rate plays a predominant role.


The potential for predicted climate shifts to impact genetic landscapes of lizards in the South African Cape Floristic Region.
The Cape Floristic Region (CFR) is well-known for its floral diversity, yet also contains a rich herpetofauna with >180 species, 28% of which are endemic. Recent studies conducted on CFR lizards indicated that phylogeographic patterns show some congruency, and that the western CFR shows higher overall diversity in the form of population and/or clade turnover. Here, we combine mitochondrial sequence data from two published (Bradypodion spp. and Agama atra) and one new dataset (Pedioplanis burchelli) to investigate whether geographic patterns of genetic diversity could be influenced by predicted climatic changes. We utilised Bayesian methodology and spatial genetic landscapes to establish broad-scale patterns and show that the western CFR is a contact zone for several clades in all three taxa, supporting the hypothesis of phylogeographic congruence. Current levels of gene flow are virtually zero between the western and eastern CFR. In the east, gene flow between populations is negligible at present but was probably stronger in the past given the present lack of strong genetic structure. Bioclimatic modelling predicted that climatically suitable areas within the CFR will decline for Bradypodion spp. and P. burchelli, with areas high in clade turnover loosing more climatically suitable areas than areas with low clade turnover. The models also predict that loss of climatic suitability may result in highly fragmented and patchy distributions, resulting in a greater loss of connectivity. In contrast, A. atra does not show significant climatic suitability losses overall, although it may experience localised losses (and gains). This species is not predicted to loose suitability in areas of high clade turnover. Thus, the incorporation of genetic data into climatic models has extended our knowledge on the vulnerability of these species given the predicted threat of landscape change.


Analysis of tanshinone IIA induced cellular apoptosis in leukemia cells by genome-wide expression profiling.
Tanshinone IIA (Tan IIA) is a diterpene quinone extracted from the root of Salvia miltiorrhiza, a Chinese traditional herb. Although previous studies have reported the anti-tumor effects of Tan IIA on various human cancer cells, the underlying mechanisms are not clear. The current study was undertaken to investigate the molecular mechanisms of Tan IIA's apoptotic effects on leukemia cells in vitro. The cytotoxicity of Tan IIA on different types of leukemia cell lines was evaluated by the 3-[4,5-dimethylthiazol-2,5]-diphenyl tetrazolium bromide (MTT) assay on cells treated without or with Tan IIA at different concentrations for different time periods. Cellular apoptosis progression with and without Tan IIA treatment was analyzed by Annexin V and Caspase 3 assays. Gene expression profiling was used to identify the genes regulated after Tan IIA treatment and those differentially expressed among the five cell lines. Confirmation of these expression regulations was carried out using real-time quantitative PCR and ELISA. The antagonizing effect of a PXR inhibitor L-SFN on Tan IIA treatment was tested using Colony Forming Unit Assay. Our results revealed that Tan IIA had different cytotoxic activities on five types of leukemia cells, with the highest toxicity on U-937 cells. Tan IIA inhibited the growth of U-937 cells in a time- and dose-dependent manner. Annexin V and Caspase-3 assays showed that Tan IIA induced apoptosis in U-937 cells. Using gene expression profiling, 366 genes were found to be significantly regulated after Tan IIA treatment and differentially expressed among the five cell lines. Among these genes, CCL2 was highly expressed in untreated U-937 cells and down-regulated significantly after Tan IIA treatment in a dose-dependent manner. RT-qPCR analyses validated the expression regulation of 80% of genes. Addition of L-sulforaphane (L-SFN), an inhibitor of Pregnane×receptor (PXR) significantly attenuated Tan IIA's effects using colony forming assays. Tan IIA has significant growth inhibition effects on U-937 cells through the induction of apoptosis. And Tan IIA-induced apoptosis might result from the activation of PXR, which suppresses the activity of NF-κB and lead to the down-regulation of CCL2 expression.


Assessment of Moral Injury in Veterans and Active Duty Military Personnel With PTSD: A Review.
Background: Moral injury (MI) involves distress over having transgressed or violated core moral boundaries, accompanied by feelings of guilt, shame, self-condemnation, loss of trust, loss of meaning, and spiritual struggles. MI is often found in Veterans and Active Duty Military personnel with posttraumatic stress disorder (PTSD). MI is widespread among those with PTSD symptoms, adversely affects mental health, and may increase risk of suicide; however, MI is often ignored and neglected by mental health professionals who focus their attention on PTSD only. Methods: A review of the literature between 1980 and 2018 conducted in 2018 is presented here to identify scales used to assess MI. Databases used in this review were PsychInfo, PubMed (Medline), and Google Scholar. Search terms were "moral injury," "measuring," "screening," "Veterans," and "Active Duty Military." Inclusion criteria were quantitative measurement of MI and health outcomes, Veteran or Active Duty Military status, and peer-review publication. Excluded were literature reviews, dissertations, book chapters, case reports, and qualitative studies. Results: Of the 730 studies identified, most did not meet eligibility criteria, leaving 118 full text articles that were reviewed, of which 42 did not meet eligibility criteria. Of the remaining 76 studies, 34 were duplicates leaving 42 studies, most published in 2013 or later. Of 22 studies that assessed MI, five used scales assessing multiple dimensions, and 17 assessed only one or two aspects (e.g., guilt, shame, or forgiveness). The remaining 20 studies used one of the scales reported in the first 22. Of the five scales assessing multiple dimensions of MI, two assess both morally injurious events and symptoms and the remaining three assess symptoms only. All studies were cross-sectional, except three that tested interventions. Conclusions: MI in the military setting is widespread and associated with PTSD symptom severity, anxiety, depression, and risk of suicide in current or former military personnel. Numerous measures exist to assess various dimensions of MI, including five multidimensional scales, although future research is needed to identify cutoff scores and clinically significant change scores. Three multidimensional measures assess MI symptoms alone (not events) and may be useful for determining if treatments directed at MI may both reduce symptoms and impact other mental health outcomes including PTSD.


Effect of Quercus infectoria and Rosa damascena on lipid profile and atherosclerotic plaque formation in rabbit model of hyperlipidemia.
Hyperlipidemia is the cause of many complications in the human societies. In this study, the effect of methanol extracts of Quercus infectoria (QI) galls and Rosa damascena (RD) Mill flower were studied on lipid profile and atherosclerotic plaques formation in hyperlipidemic rabbits. Thirty-six New Zeland white rabbits randomly divided into 6 groups as control (I), hyperlipidemic (II), hyperlipidemic+QI (III), hyperlipidemic+RD (IV), +Atorvastolin (V) and hyperlipidemic+Orlistat (VI) and were fed with high fat diet (0.5% cholesterol and 16% hydrogenated vegetable oil) for 45 days. At the end of the study period, lipid profile and plaque formation were assessed. Total Cholesterol (TC), Low Density Lipoprotein (LDL) and Triglyceride (TG) levels were significantly increased in hyperlipidemic group compared with control group (p < 0.001). Methanol extract consumption of Quercus infectoria significantly decreased plasma levels of TC, TG and LDL (p < 0.001). It also decreased plaques formation in semi lunar valve and thoracic aorta. Rosa damascena mill flower methanol extract moderately decreased the levels of TC, TG, LDL and plaques formation but it was not significant. HDL levels and weight of animals did not show significant difference among groups. Based on the doses used in this study, our finding indicated that QI but no RD methanol extract has anti atherogenic and hypolipidemic activities.


Type-2 diabetes mellitus: does prenatal care affect outcomes?
To determine if prenatal care affects adverse perinatal outcomes in pregnant women with Type-2 diabetes mellitus (T2DM). This was a retrospective cohort study of pregnant women with pregestational diabetes mellitus pregnancies in the state of California between 1997 and 2006, using vital statistics data linked to birth certificates. Women were stratified by time of presentation to care and we compared those who presented in the first trimester, third trimester, and those who had no prenatal care prior to delivery. Perinatal outcomes looked at included: preeclampsia, macrosomia, preterm delivery, cesarean delivery, and intrauterine fetal demise (IUFD). The two groups were compared with chi-squared testing to determine statistical significance. In women with pregestational diabetes those who presented at time of delivery had an 11.3% risk of IUFD compared to 0.9% in those who presented in the first trimester. There was also an increased rate of preterm birth in the late presentation cohort (29.4% at time of delivery versus 21.0% in the first trimester). After adjusting for possible confounding variables using logistic regression models, rates of IUFD and preterm delivery were still found to be statistically significant with adjusted odds ratios of 11.37 (95% CI: 6.10-21.16) and 1.55 (95% CI: 1.03-2.32), respectively. There were no differences in rates of macrosomia or preeclampsia between the three cohorts. Treatment of T2DM throughout pregnancy leads to improved maternal and neonatal outcomes.


Morphological and morphometric studies of the dysmyelinating mutant, the Long Evans shaker rat.
The Long Evans shaker (les) rat is a recently identified CNS myelin mutant with an autosomal recessive mode of inheritance. Although scattered myelin sheaths are present in some areas of the CNS, most notably the ventral spinal cord in the young neonatal rat, this myelin is gradually lost, and 8-12 weeks little myelin is present throughout the CNS. Despite this severe myelin deficiency, some mutants may live beyond one year of age. Rare, thin myelin sheaths that are present early in development lack myelin basic protein (MBP) and on ultrastructural examination are poorly compacted and lack a major dense line. Many oligodendrocytes develop an accumulation of vesicles and membranous bodies, but no abnormal cell death is observed. In the optic nerve, cell kinetic studies show an increase in proliferation at early time points in les, while total glial cell counts are also increased in les from 2 months of age. In situ hybridization studies demonstrate that the numbers of mature oligodendrocytes are similar to controls early in life and increase with time compared to controls. There is both a progressive astrocyte hypertrophy and microgliosis. While les has a mutation in the myelin basic protein (mbp) gene, it is dissimilar in both genotype and phenotype to the previously described mbp mouse mutants, shiverer (shi) and shiverer(mld). Unlike shi and its allele, where myelin increases with time and oligodendrocytes become ultrastructurally normal, les oligodendrocytes are permanently disabled, continue to demonstrate cytoplasmic abnormalities, and fail to produce myelin beyond the first weeks of life.


Fracture healing is delayed in the absence of gasdermin-interleukin-1 signaling.
Amino-terminal fragments from proteolytically cleaved gasdermins (GSDMs) form plasma membrane pores that enable the secretion of interleukin-1β (IL-1β) and IL-18. Excessive GSDM-mediated pore formation can compromise the integrity of the plasma membrane thereby causing the lytic inflammatory cell death, pyroptosis. We found that GSDMD and GSDME were the only GSDMs that were readily expressed in bone microenvironment. Therefore, we tested the hypothesis that GSDMD and GSDME are implicated in fracture healing owing to their role in the obligatory inflammatory response following injury. We found that bone callus volume and biomechanical properties of injured bones were significantly reduced in mice lacking either GSDM compared with wild-type (WT) mice, indicating that fracture healing was compromised in mutant mice. However, compound loss of GSDMD and GSDME did not exacerbate the outcomes, suggesting shared actions of both GSDMs in fracture healing. Mechanistically, bone injury induced IL-1β and IL-18 secretion in vivo, a response that was mimicked in vitro by bone debris and ATP, which function as inflammatory danger signals. Importantly, the secretion of these cytokines was attenuated in conditions of GSDMD deficiency. Finally, deletion of IL-1 receptor reproduced the phenotype of Gsdmd or Gsdme deficient mice, implying that inflammatory responses induced by the GSDM-IL-1 axis promote bone healing after fracture.


Studies on the genetic linkage of bilirubin and androsterone UDP-glucuronyltransferases by cross-breeding of two mutant rat strains.
Gunn rats, which have defects in bilirubin and 4-nitrophenol UDP-glucuronyltransferases (GT), were crossed with LA Wistar rats with a defect in androsterone GT. The F1 hybrids showed normal GT activities towards androsterone, bilirubin and 4-nitrophenol, demonstrating that Gunn and LA ('low activity') Wistar rats inherit a homozygous dominant trait for androsterone GT and bilirubin GT respectively. The F2 progeny showed four different combinations of bilirubin and androsterone GT activities: defects in both GT activities, a single defect in bilirubin GT activity, a single defect in androsterone GT activity and two normal GT activities. They were segregated in the approximate ratio of 1:3:3:9, which is compatible with Mendel's Principle of Independent Assortment. These results provide evidence that androsterone GT and bilirubin GT are located on different chromosomes. In the F2 generation, defective bilirubin and 4-nitrophenol GT activities were not segregated, indicating that these two mutant genes are closely linked on the same chromosome.


Activation of multiple signaling pathways by terminal complement complexes involved in myocellular sodium homeostasis.
Soluble C5b-9 complexes (SC5b-9), hemolytically inactive end-products of complement activation have long been considered to be irrelevant. Recent investigations, however, have demonstrated that SC5b-9 induces numerous biological effects via a series of intracellular signal transduction events. We have previously demonstrated that SC5b-9 enriched sera increased intracellular Na+ in rat skeletal muscles. This study was purposed to determine if the protein kinase C (PKC) or mitogen-activated protein kinase (MAPK) signaling pathway mediates the effects of SC5b-9. Fast-twitch extensor digitorum longus (EDL) muscles isolated from infant rats were incubated at 30 degrees C for 60 minutes with 10% zymosan-activated rat sera (ZARS) as a source of complement. Heat-inactivated rat sera (HIRS) were used as a control. The muscles were also incubated with ZARS or HIRS in the presence of specific inhibitors against PKC (GF109203X) or MAPK (PD98059 and SB202190). Intracellular Na+ and K+ contents were then measured. ZARS significantly increased intracellular Na+ and the Na+/K+ ratio in EDL muscles as compared to HIRS. GF109203X, PD98059 and SB202190 markedly attenuated increase in myocellular Na+ induced by ZARS, respectively. We concluded that SC5b-9 enriched sera alter myocellular Na+ homeostasis, at least in part, via the mechanisms linked to PKC and MAPK signal transduction pathways.


How does the medical graduates' self-assessment of their clinical competency differ from experts' assessment?
The assessment of the performance of medical school graduates during their first postgraduate years provides an early indicator of the quality of the undergraduate curriculum and educational process. The objective of this study was to assess the clinical competency of medical graduates, as perceived by the graduates themselves and by the experts. This is a hospital based cross-sectional study. It covered 105 medical graduates and 63 experts selected by convenient sampling method. A self-administered questionnaire covering the different areas of clinical competency constructed on a five-point Likert scale was used for data collection. Data processing and analysis were performed using the Statistical Package for Social Science (SPSS) 16.0. The mean, frequency distribution, and percentage of the variables were calculated. A non-parametric Kruskal Wallis test was applied to verify whether the graduates' and experts' assessments were influenced by the graduates' variables such as age, gender, experience, type of hospital, specialty and location of work at a (p ≤ 0.05) level of significance. The overall mean scores for experts' and graduates' assessments were 3.40 and 3.63, respectively (p= 0.035). Almost 87% of the graduates perceived their competency as good and very good in comparison with only 67.7% by experts. Female and male graduates who rated themselves as very good were 33.8% and 25% respectively. More than 19% of the graduates in the age group > 30 years perceived their clinical competency as inadequate in contrast with only 6.2% of the graduates in the youngest age group. Experts rated 40% of the female graduates as inadequate versus 20% of males, (p= 0.04). More than 40% of the graduates in younger age group were rated by experts as inadequate, versus 9.7% of the higher age group >30 years (p = 0.03). There was a wide discrepancy between the graduates' self-assessment and experts' assessment, particularly in the level of inadequate performance. Graduates in general, and those of younger age groups in particular, tend to overestimate their clinical skills and competency.


Crystal structure of the DENR-MCT-1 complex revealed zinc-binding site essential for heterodimer formation.
The density-regulated protein (DENR) and the malignant T cell-amplified sequence 1 (MCT-1/MCTS1) oncoprotein support noncanonical translation initiation, promote translation reinitiation on a specific set of mRNAs with short upstream reading frames, and regulate ribosome recycling. DENR and MCT-1 form a heterodimer, which binds to the ribosome. We determined the crystal structure of the heterodimer formed by human MCT-1 and the N-terminal domain of DENR at 2.0-Å resolution. The structure of the heterodimer reveals atomic details of the mechanism of DENR and MCT-1 interaction. Four conserved cysteine residues of DENR (C34, C37, C44, C53) form a classical tetrahedral zinc ion-binding site, which preserves the structure of the DENR's MCT-1-binding interface that is essential for the dimerization. Substitution of all four cysteines by alanine abolished a heterodimer formation. Our findings elucidate further the mechanism of regulation of DENR-MCT-1 activities in unconventional translation initiation, reinitiation, and recycling.


Targeting proteins to membranes: structure of the signal recognition particle.
In all three kingdoms of life, co-translational targeting of secretory and membrane proteins to the prokaryotic plasma membrane or eukaryotic endoplasmic reticulum is mediated by a ribonucleoprotein complex, the signal recognition particle (SRP), and its membrane-associated receptor (SR). SRP binds to signal sequences of nascent proteins as they emerge from the exit tunnel of the ribosome. The resulting targeting complex, composed of the SRP and the ribosome-nascent chain complex (RNC), then docks with the SR in a GTP-dependent manner. Passing through a complex series of conformational states, SRP and SR deliver the RNC to the translocon, which in turn mediates protein translocation across or integration into the membrane. The core structural and mechanistic principles of SRP-dependent protein targeting are universally conserved. Recent structural investigations combining X-ray crystallography and cryo-electron microscopy have provided new insights into three essentials steps of the SRP-dependent protein targeting cycle: the assembly and interaction of the SRP ribonucleoprotein core, the GTP-dependent SRP-SR association, and the interaction between SRP and the ribosome.


Structure of a proteasome Pba1-Pba2 complex: implications for proteasome assembly, activation, and biological function.
The 20S proteasome is an essential, 28-subunit protease that sequesters proteolytic sites within a central chamber, thereby repressing substrate degradation until proteasome activators open the entrance/exit gate. Two established activators, Blm10 and PAN/19S, induce gate opening by binding to the pockets between proteasome α-subunits using C-terminal HbYX (hydrophobic-tyrosine-any residue) motifs. Equivalent HbYX motifs have been identified in Pba1 and Pba2, which function in proteasome assembly. Here, we demonstrate that Pba1-Pba2 proteins form a stable heterodimer that utilizes its HbYX motifs to bind mature 20S proteasomes in vitro and that the Pba1-Pba2 HbYX motifs are important for a physiological function of proteasomes, the maintenance of mitochondrial function. Other factors that contribute to proteasome assembly or function also act in the maintenance of mitochondrial function and display complex genetic interactions with one another, possibly revealing an unexpected pathway of mitochondrial regulation involving the Pba1-Pba2 proteasome interaction. Our determination of a proteasome Pba1-Pba2 crystal structure reveals a Pba1 HbYX interaction that is superimposable with those of known activators, a Pba2 HbYX interaction that is different from those reported previously, and a gate structure that is disrupted but not sufficiently open to allow entry of even small peptides. These findings extend understanding of proteasome interactions with HbYX motifs and suggest multiple roles for Pba1-Pba2 interactions throughout proteasome assembly and function.


Faculty perceptions and knowledge of career development of trainees in biomedical science: What do we (think we) know?
The Broadening Experiences in Scientific Training (BEST) program is an NIH-funded effort testing the impact of career development interventions (e.g. internships, workshops, classes) on biomedical trainees (graduate students and postdoctoral fellows). BEST Programs seek to increase trainees' knowledge, skills and confidence to explore and pursue expanded career options, as well as to increase training in new skills that enable multiple career pathways. Faculty mentors are vital to a trainee's professional development, but data about how faculty members of biomedical trainees view the value of, and the time spent on, career development are lacking. Seven BEST institutions investigated this issue by conducting faculty surveys during their BEST experiment. The survey intent was to understand faculty perceptions around professional and career development for their trainees. Two different, complementary surveys were employed, one designed by Michigan State University (MSU) and the other by Vanderbilt University. Faculty (592) across five institutions responded to the MSU survey; 225 faculty members from two institutions responded to the Vanderbilt University survey. Participating faculty were largely tenure track and male; approximately 1/3 had spent time in a professional position outside of academia. Respondents felt a sense of urgency in introducing broad career activities for trainees given a recognized shortage of tenure track positions. They reported believing career development needs are different between a graduate student and postdoctoral fellow, and they indicated that they actively mentor trainees in career development. However, faculty were uncertain as to whether they actually have the knowledge or training to do so effectively. Faculty perceived that trainees themselves lack a knowledge base of skills that are of interest to non-academic employers. Thus, there is a need for exposure and training in such skills. Faculty stated unequivocally that institutional support for career development is important and needed. BEST Programs were considered beneficial to trainees, but the awareness of local BEST Programs and the national BEST Consortium was low at the time surveys were employed at some institutions. It is our hope that the work presented here will increase the awareness of the BEST national effort and the need for further career development for biomedical trainees.


Evaluation of several microcrystalline celluloses obtained from agricultural by-products.
Microcrystalline cellulose (MCCI) has been widely used as an excipient for direct compression due to its good flowability, compressibility, and compactibility. In this study, MCCI was obtained from agricultural by-products, such as corn cob, sugar cane bagasse, rice husk, and cotton by pursuing acid hydrolysis, neutralization, clarification, and drying steps. Further, infrared spectroscopy (IR), X-ray diffraction (XRD), optical microscopy, degree of polymerization (DP), and powder and tableting properties were evaluated and compared to those of Avicel PH101, Avicel PH102, and Avicel PH200. Except for the commercial products, all materials showed a DP from 55 to 97. Particles of commercial products and corn cob had an irregular shape, whereas bagasse particles were elongated and thick. Rice and cotton particles exhibited a flake-like and fiber-like shape, respectively. MCCI as obtained from rice husk and cotton was the most densified material, while that produced from corn cob and bagasse was bulky, porous, and more compressible. All products had a moisture content of less than 10% and yields from 7.4% to 60.4%. MCCI as obtained from bagasse was the most porous and compressible material among all materials. This product also showed the best tableting properties along with Avicel products. Likewise, all MCCI products obtained from the above-mentioned sources showed a more rapid disintegration time than that of Avicel products. These materials can be used as a potential source of MCCI in the production of solid dosage forms.


Co-culture of osteoblasts with immature dural cells causes an increased rate and degree of osteoblast differentiation.
For decades surgeons have exploited the ability of infants to reossify large calvarial defects. To demonstrate the role of dura mater-osteoblast communication during the process of calvarial reossification, the authors used a novel in vitro system that recapitulates the in vivo anatomic relationship of these cell populations. Primary cultures of osteoblast cells from 2-day-old Sprague-Dawley rat pups were grown on six-well plates, and cultures of immature, non-suture-associated dura mater cells from 6-day-old Sprague-Dawley rat pups were grown on Transwell inserts. When the osteoblast and dura mater cell cultures reached confluence, they were combined. This Transwell co-culture system permitted the two cell populations to grow together in the same well, but it prevented direct cell-to-cell contact. Therefore, the authors were able to determine, for the first time, whether paracrine signaling from immature, non-suture-associated dura mater could influence the biologic activity of osteoblasts. Osteoblasts co-cultured with dural cells proliferated significantly faster after 2 days (2.1 x 10(5) +/- 2.4 x 10(4) versus 1.4 x 10(5) +/- 2.2 x 10(4), p < or = 0.05) and 4 days (3.1 x 10(5) +/- 5 x 10(4) versus 2.2 x 10(5) +/- 4.0 x 10(4), p < or = 0.01) than did osteoblasts cultured alone. After 20 days, co-cultured osteoblasts expressed greater amounts of mRNA for several markers of osteoblast differentiation, including collagen I alpha I (4-fold), alkaline phosphatase (2.5-fold), osteopontin (3-fold), and osteocalcin (4-fold), than did osteoblasts cultured alone. After 30 days, co-cultured osteoblasts produced bone nodules that were significantly greater both in number (324 +/- 29 nodules versus 252 +/- 29 nodules per well, p , < or = 0.04) and total area of nodules (65 +/- 11 mm(2) versus 24 +/- 1.6 mm(2), p < or = 0.003) than osteoblasts cultured alone. To begin to understand how dural cells effect changes in osteoblast gene expression, the authors compared the expression of candidate genes, transforming growth factor beta 1 and fibroblast growth factor 2, in dural cells and osteoblasts before and after 5 days of culture. Interestingly, the dura mater produced marked amounts of these osteogenic cytokines compared with osteoblasts.The described co-culture system demonstrated that co-cultured osteoblasts proliferated more rapidly and experienced an increased rate and degree of cellular maturation than did osteoblasts cultured alone. The authors hypothesize that this effect was due to paracrine signaling (e.g., transforming growth factor beta 1 and fibroblast growth factor 2) from the dura mater, and they are investigating those mechanisms in ongoing experiments. Collectively these data verify that immature, non-suture-associated dura mater can influence the biologic activity of osteoblasts. Moreover, the production of cytokines derived from the dura mater (e.g., transforming growth factor beta 1 and fibroblast growth factor 2), and they may begin to explain why immature animals and infants with intact dura mater can reossify large calvarial defects.


Thalassaemia of intermediate severity resulting from the interaction between alpha- and beta-thalassaemia.
A Sicilian family is described in which the alpha-thalassaemia gene is interacting in several members with beta-thalassaemia resulting in a balanced alpha/beta chain production ratio. In one patient, affected by homozygous beta-thalassaemia, the presence of alpha-thalassaemia resulted in a less severe clinical expression of the disease, less marked imbalance in the alpha/non-alpha ratio, and a lower level of HbF. Further studies of haemoglobin synthesis are needed to clarify the complex genetic picture that results from the interaction of different forms of thalassaemia.


Hepatocellular carcinoma in Port Harcourt, Nigeria: clinicopathologic study of 75 cases.
Hepatocellular carcinoma (HCC) is a complication of liver cirrhosis caused by hepatitis B viral infection, fungal infection and genetic factors. This disease though having a silent course is associated with variable morbidity and mortality in this environment. Previously prepared histologic slides were retrieved and re-evaluated to confirm the diagnosis. Clinical data such as the age, sex, clinical presentations, histologic grading and cause of death were extracted from the case notes, histology request forms and autopsy protocol forms. Seventy five cases were used for the study during the period under review. The youngest was a 14 year old male while the oldest was a 66 years old male. Male to female ratio was 2:1. Majority (28%) occurred in the age group 40-49 years. Upper abdominal mass was the most common clinical presentation (30.7%) and well differentiated hepatocellular carcinoma was the most common (64.0%) histologic grade. Trabecular inusoidal pattern (49.3%) was the most common histologic type (pattern) while upper gastro-intestinal bleeding (38.7%) was the most common cause of death at autopsy. Hepatocellular carcinoma is a notable malignancy of the liver associated with variable morbidities and mortality as it has an insidious onset and very aggressive course.


Development and psychometric testing of the childhood obesity perceptions (COP) survey among African American caregivers: A tool for obesity prevention program planning.
Currently, public health practitioners are analyzing the role that caregivers play in childhood obesity efforts. Assessing African American caregiver's perceptions of childhood obesity in rural communities is an important prevention effort. This article's objective is to describe the development and psychometric testing of a survey tool to assess childhood obesity perceptions among African American caregivers in a rural setting, which can be used for obesity prevention program development or evaluation. The Childhood Obesity Perceptions (COP) survey was developed to reflect the multidimensional nature of childhood obesity including risk factors, health complications, weight status, built environment, and obesity prevention strategies. A 97-item survey was pretested and piloted with the priority population. After pretesting and piloting, the survey was reduced to 59-items and administered to 135 African American caregivers. An exploratory factor analysis (EFA) and confirmatory factor analysis (CFA) was conducted to test how well the survey items represented the number of Social Cognitive Theory constructs. Twenty items were removed from the original 59-item survey and acceptable internal consistency of the six factors (α=0.70-0.85) was documented for all scales in the final COP instrument. CFA resulted in a less than adequate fit; however, a multivariate Lagrange multiplier test identified modifications to improve the model fit. The COP survey represents a promising approach as a potentially comprehensive assessment for implementation or evaluation of childhood obesity programs.


Platelet apoptosis in paediatric immune thrombocytopenia is ameliorated by intravenous immunoglobulin.
To evaluate the role of intravenous immunoglobulin (IVIg) in platelet apoptosis in paediatric immune thrombocytopenia, we investigated the platelets of 20 paediatric patients with acute immune thrombocytopenia (ITP), before and after IVIg treatment. Healthy children with platelet counts in the normal range and children with thrombocytopenia due to chemotherapy were enrolled as controls. All ITP patients presented with platelet counts <20 × 10(9) /l and bleeding symptoms. Markers of apoptosis, including activated caspase-3, -8 and -9, phosphatidylserine (PS) exposure, mitochondrial inner membrane potential (ΔΨm), as well as platelet-derived microparticle formation, were analysed by flow cytometry. After IVIg treatment, platelet counts increased to >20 × 10(9) /l in all patients. ITP patients had significantly increased proportions of platelets with activated caspase-3, -8 and -9, with PS exposure, and with decreased ΔΨm, and demonstrated increased microparticle formation. Except for ΔΨm, these markers for apoptosis were reduced by IVIg treatment. Platelets of children with thrombocytopenia after chemotherapy also demonstrated increased microparticle formation and decreased ΔΨm, but no activation of caspases 3, 8 and 9 or PS exposure. In conclusion, in acute paediatric ITP, enhanced platelet apoptosis is seen at diagnosis that normalizes after IVIg treatment.


Peroral ciprofloxacin therapy impairs the generation of a protective immune response in a mouse model for Salmonella enterica serovar Typhimurium diarrhea, while parenteral ceftriaxone therapy does not.
Nontyphoidal Salmonella (NTS) species cause self-limiting diarrhea and sometimes severe disease. Antibiotic treatment is considered only in severe cases and immune-compromised patients. The beneficial effects of antibiotic therapy and the consequences for adaptive immune responses are not well understood. We used a mouse model for Salmonella diarrhea to assess the effects of per os treatment with ciprofloxacin (15 mg/kg of body weight intragastrically 2 times/day, 5 days) or parenteral ceftriaxone (50 mg/kg intraperitoneally, 5 days), two common drugs used in human patients. The therapeutic and adverse effects were assessed with respect to generation of a protective adaptive immune response, fecal pathogen excretion, and the emergence of nonsymptomatic excreters. In the mouse model, both therapies reduced disease severity and reduced the level of fecal shedding. In line with clinical data, in most animals, a rebound of pathogen gut colonization/fecal shedding was observed 2 to 12 days after the end of the treatment. Yet, levels of pathogen shedding and frequency of appearance of nonsymptomatic excreters did not differ from those for untreated controls. Moreover, mice treated intraperitoneally with ceftriaxone developed an adaptive immunity protecting the mice from enteropathy in wild-type Salmonella enterica serovar Typhimurium challenge infections. In contrast, the mice treated intragastrically with ciprofloxacin were not protected. Thus, antibiotic treatment regimens can disrupt the adaptive immune response, but treatment regimens may be optimized in order to preserve the generation of protective immunity. It might be of interest to determine whether this also pertains to human patients. In this case, the mouse model might be a tool for further mechanistic studies.


Effects of gender and depressive symptoms on quality of life among colorectal and lung cancer patients and their family caregivers.
Cancer patients and their family caregivers often report elevated levels of depressive symptoms, along with poorer mental and physical health (quality of life: QOL). Although the mutuality in distress between patients and their caregivers is relatively well known, unknown are the degree to which caregivers' depressive symptoms independently predict their patient's QOL and vice versa, and whether the relations vary by cancer type or gender. Colorectal or lung cancer patients and their caregivers (398 dyads) provided complete data for study variables (212 colorectal cancer patient dyads, 186 lung cancer patient dyads; 257 male patient dyads, 141 female patient dyads). Patients' depressive symptoms and QOL were measured approximately 4 and 12 months post-diagnosis; caregivers' depressive symptoms and QOL were measured approximately 5 months post-diagnosis. The actor-partner interdependence model confirmed that each person's depressive symptom level was uniquely associated with his/her own concurrent QOL. Female patients' depressive symptoms were also related to their caregivers' poorer physical and better mental health, particularly when the pair's depressive symptoms were at similarly elevated level. On the other hand, male patients' elevated depressive symptoms were related to their caregivers' poorer mental health. Findings suggest that QOL among patients and their family caregivers is interdependent. In light of this interdependency, psychosocial interventions for managing depressive symptoms should target both patients and their family caregivers, from which both may benefit by not only alleviating depressive symptoms but also improving QOL.


Brain temperature in volunteers subjected to intranasal cooling.
Intranasal cooling can be used to initiate therapeutic hypothermia. However, direct measurement of brain temperature is difficult and the intra-cerebral distribution of temperature changes with cooling is unknown. The purpose of this study was to measure the brain temperature of human volunteers subjected to intranasal cooling using non-invasive magnetic resonance (MR) methods. Intranasal balloons catheters circulated with saline at 20°C were applied for 60 min in ten awake volunteers. No sedation was used. Brain temperature changes were measured and mapped using MR spectroscopic imaging (MRSI) and phase-mapping techniques. Heart rate and blood pressure were monitored throughout the experiment. Rectal temperature was measured before and after the cooling. Mini Mental State Examination (MMSE) test and nasal inspection were done before and after the cooling. Questionnaires about the subjects' personal experience were completed after the experiment. Brain temperature decrease measured by MRSI was -1.7 ± 0.8°C and by phase-mapping -1.8 ± 0.9°C (n = 9) at the end of cooling. Spatial distribution of temperature changes was relatively uniform. Rectal temperature decreased by -0.5 ± 0.3°C (n = 5). The physiological parameters were stable and no shivering was reported. The volunteers remained alert during cooling and no cognitive dysfunctions were apparent in the MMSE test. Postcooling nasal examination detected increased nasal secretion in nine of the ten volunteers. Volunteers' acceptance of the method was good. Both MR techniques revealed brain temperature reductions after 60 min of intranasal cooling with balloons circulated with saline at 20°C in awake, unsedated volunteers.


Primary fibrosarcoma of bone. Outcome after primary surgical treatment.
To investigate outcome and evaluate prognostic factors in primary fibrosarcoma of bone, all patients at the authors' institution who had surgical treatment for primary fibrosarcoma of bone from 1910 to 1995 were studied. Medical records, surgical reports, radiographs, and histologic slides of 92 patients (51 males, 41 females; mean age, 38 years; range, 8-84 years) were reviewed. The most common tumor locations were the femur (28 patients), tibia (21 patients), and pelvis (14 patients). Sixty-one tumors (66%) were Enneking Stage IIB. Twenty-nine patients (31.5%) had adjuvant therapy: 16 had radiation, nine had chemotherapy alone, and four had radiation and chemotherapy. Amputation or disarticulation was performed in 61 patients (66%), wide excision was performed in 13 (14%), marginal excision was performed in 15 (16%), and intralesional excision was performed in three (3.3%). Local recurrence occurred in 14 patients (15%) at a median time of 7 months (range, 3-21 months). Metastases developed in 58 of the 85 patients (68%) with Stage I or II tumors at presentation, at a median of 9 months (range, 1-51 months). Survivorship analysis showed that the overall probability of survival was 33.4% at 5 years after surgery. Multivariate analysis showed that the main prognostic risk factors affecting overall survival included age older than 40 years, tumor location in the axial skeleton, and high-grade tumor (Grade 3 or 4). With the high incidence of systemic failure after surgical treatment, perioperative adjuvant treatment modalities should be considered.


Structural mechanics of DNA wrapping in the nucleosome.
Experimental X-ray crystal structures and a database of calculated structural parameters of DNA octamers were used in combination to analyse the mechanics of DNA bending in the nucleosome core complex. The 1kx5 X-ray crystal structure of the nucleosome core complex was used to determine the relationship between local structure at the base-step level and the global superhelical conformation observed for nucleosome-bound DNA. The superhelix is characterised by a large curvature (597 degrees) in one plane and very little curvature (10 degrees) in the orthogonal plane. Analysis of the curvature at the level of 10-step segments shows that there is a uniform curvature of 30 degrees per helical turn throughout most of the structure but that there are two sharper kinks of 50 degrees at +/-2 helical turns from the central dyad base pair. The curvature is due almost entirely to the base-step parameter roll. There are large periodic variations in roll, which are in phase with the helical twist and account for 500 degrees of the total curvature. Although variations in the other base-step parameters perturb the local path of the DNA, they make minimal contributions to the total curvature. This implies that DNA bending in the nucleosome is achieved using the roll-slide-twist degree of freedom previously identified as the major degree of freedom in naked DNA oligomers. The energetics of bending into a nucleosome-bound conformation were therefore analysed using a database of structural parameters that we have previously developed for naked DNA oligomers. The minimum energy roll, the roll flexibility force constant and the maximum and minimum accessible roll values were obtained for each base step in the relevant octanucleotide context to account for the effects of conformational coupling that vary with sequence context. The distribution of base-step roll values and corresponding strain energy required to bend DNA into the nucleosome-bound conformation defined by the 1kx5 structure were obtained by applying a constant bending moment. When a single bending moment was applied to the entire sequence, the local details of the calculated structure did not match the experiment. However, when local 10-step bending moments were applied separately, the calculated structure showed excellent agreement with experiment. This implies that the protein applies variable bending forces along the DNA to maintain the superhelical path required for nucleosome wrapping. In particular, the 50 degrees kinks are constraints imposed by the protein rather than a feature of the 1kx5 DNA sequence. The kinks coincide with a relatively flexible region of the sequence, and this is probably a prerequisite for high-affinity nucleosome binding, but the bending strain energy is significantly higher at these points than for the rest of the sequence. In the most rigid regions of the sequence, a higher strain energy is also required to achieve the standard 30 degrees curvature per helical turn. We conclude that matching of the DNA sequence to the local roll periodicity required to achieve bending, together with the increased flexibility required at the kinks, determines the sequence selectivity of DNA wrapping in the nucleosome.


Effect of electrical parameters on ECT convulsive threshold and duration.
Optimal stimulus parameters for electroconvulsive therapy (ECT) are unclear. Pulse duration and frequency related to convulsive threshold and seizure duration in the first ECT treatment in a series were evaluated. Convulsive threshold was estimated for all patients (N = 550) receiving ECT over 27 months. Thresholds were estimated using different brief pulse stimulators, starting with a dose of approximately 25 mC per pulse train for right unilateral (RUL) stimulation (50 mC for bilateral [BL] stimulation). The charge was applied in 25-mC serial increments (approximately doubling for BL stimulation) up to approximately 100 mC (>200 mC for BL stimulation) to a generalized motor seizure endpoint. Patients lacking seizure response at 100 mC (200 mC for BL stimulation) received >500 mC. Convulsive threshold increased with age, African American identity, diagnosis other than depression, and female sex, and decreased with RUL electrode placement, low frequency (30 Hz), and brief pulse width (0.5 msec). RUL stimulation and lower anesthetic medication doses promoted longer seizures. Younger patients had longer seizures than older patients. Pulse width and frequency did not affect seizure duration. Lower charge yielded longer seizures. ECT efficiency appears to be achieved by lower frequency and briefer pulse duration stimulation. Randomized trials are needed for corroboration of these findings.


Cellular requirements for LARK in the Drosophila circadian system.
RNA-binding proteins mediate posttranscriptional functions in the circadian systems of multiple species. A conserved RNA recognition motif (RRM) protein encoded by the lark gene is postulated to serve circadian output and molecular oscillator functions in Drosophila and mammals, respectively. In no species, however, has LARK been eliminated, in vivo, to determine the consequences for circadian timing. The present study utilized RNA interference (RNAi) techniques in Drosophila to decrease LARK levels in clock neurons and other cell types in order to evaluate the circadian functions of the protein. Knockdown of LARK in timeless (TIM)- or pigment dispersing factor (PDF)-containing clock cells caused a significant number of flies to exhibit arrhythmic locomotor activity, demonstrating a requirement for the protein in pacemaker cells. There was no obvious effect on PER protein cycling in lark interference (RNAi) flies, but a knockdown within the PDF neurons was associated with increased PDF immunoreactivity at the dorsal termini of the small ventral lateral neuronal (s-LNv) projections, suggesting an effect on neuropeptide release. The expression of lark RNAi in multiple neurosecretory cell populations demonstrated that LARK is required within pacemaker and nonpacemaker cells for the manifestation of normal locomotor activity rhythms. Interestingly, decreased LARK function in the prothoracic gland (PG), a peripheral organ containing a clock required for the circadian control of eclosion, was associated with weak population eclosion rhythms or arrhythmicity.


Gene Expression in Mouse Thyrotrope Adenoma: Transcription Elongation Factor Stimulates Proliferation.
Thyrotrope hyperplasia and hypertrophy are common responses to primary hypothyroidism. To understand the genetic regulation of these processes, we studied gene expression changes in the pituitaries of Cga(-/-) mice, which are deficient in the common α-subunit of TSH, LH, and FSH. These mice have thyrotrope hypertrophy and hyperplasia and develop thyrotrope adenoma. We report that cell proliferation is increased, but the expression of most stem cell markers is unchanged. The α-subunit is required for secretion of the glycoprotein hormone β-subunits, and mutants exhibit elevated expression of many genes involved in the unfolded protein response, consistent with dilation and stress of the endoplasmic reticulum. Mutants have elevated expression of transcription factors that are important in thyrotrope function, such as Gata2 and Islet 1, and those that stimulate proliferation, including Nupr1, E2f1, and Etv5. We characterized the expression and function of a novel, overexpressed gene, transcription elongation factor A (SII)-like 5 (Tceal5). Stable expression of Tceal5 in a pituitary progenitor cell line is sufficient to increase cell proliferation. Thus, Tceal5 may act as a proto-oncogene. This study provides a rich resource for comparing pituitary transcriptomes and an analysis of gene expression networks.


The burden and quality of life of caregivers of sickle cell anemia patients taking hydroxyurea versus those not taking hydroxyurea.
To assess the burden and quality of life of caregivers of patients with sickle cell anemia taking hydroxyurea versus those of patients not taking hydroxyurea. A cross-sectional study was performed of caregivers of outpatients with sickle cell anemia in two public hospitals in Campo Grande, MS, from January through June 2010. The World Health Organization Quality of Life-BREF Scale and the Caregiver Burden Scale were used. Of the 37 caregivers in this study, 81.1% were women, 73.0% were mothers, 59.5% were married, 54.1%were mulattos, 48.6% were housewives, 54.1% had family incomes of up to one minimum wage and 75.7% had onlycompleted elementary education. The mean duration of care provided (time after diagnosis) was 16.08 ± 9.88 yearsand 89.2% reported that they provided 24-hour care. Regarding health, 27.0% of study participants reported having physical and 13.5% emotional problems. There were no significant relationships between these variables either with the different domains or the total score of the WHOQOL-BREF comparing caregivers of patients taking hydroxyurea versusthose of patients not taking hydroxyurea. There was a moderate negative linear correlation between the WHOQOL-BREF and the Caregiver Burden Scale scores (linear correlation test of Pearson: p-value = 0.003, r = -0.477). The burden of caregivers of patients who did not take hydroxyurea was significantly higher than those of patients who took the medication in terms of general tension, disappointment, environment and total score (student t-test: p-value < 0.05). In the perception of the caregiver, looking after sickle cell anemia patients represents a moderate negative burden.


An anaerobic methane-oxidizing community of ANME-1b archaea in hypersaline Gulf of Mexico sediments.
Sediments overlying a brine pool methane seep in the Gulf of Mexico (Green Canyon 205) were analyzed using molecular and geochemical approaches to identify geochemical controls on microbial community composition and stratification. 16S rRNA gene and rRNA clone libraries, as well as mcrA gene clone libraries, showed that the archaeal community consists predominantly of ANME-1b methane oxidizers; no archaea of other ANME subgroups were found with general and group-specific PCR primers. The ANME-1b community was found in the sulfate-methane interface, where undersaturated methane concentrations of ca. 100 to 250 microM coexist with sulfate concentrations around 10 mM. Clone libraries of dsrAB genes and bacterial 16S rRNA genes show diversified sulfate-reducing communities within and above the sulfate-methane interface. Their phylogenetic profiles and occurrence patterns are not linked to ANME-1b populations, indicating that electron donors other than methane, perhaps petroleum-derived hydrocarbons, drive sulfate reduction. The archaeal component of anaerobic oxidation of methane is comprised of an active population of mainly ANME-1b in this hypersaline sediment.


Complications After Vulvar and Perineal Reconstruction With a Lotus Petal Flap.
Lotus petal flaps are often used to reconstruct defects in the vulvoperineal area. Wound complications occur more often after surgery in the vulvoperineal area than in other areas of the body. Postoperative complications have a great impact on patients' well-being, length of hospital stay, care needed, and return to participation in daily life. Therefore, it is important to determine which patients are more prone to develop complications. All lotus petal flap reconstructions in our center between January 2004 and August 2014 were retrospectively reviewed. Complications were categorized by the Clavien-Dindo classification. Ninety-three patients with 137 flaps were reviewed. The complication rates did not differ significantly between the reconstruction areas. In 30.1% of subjects, no complications were reported. A total of 51.7% of the subjects experienced Clavien-Dindo grade I or II complications, which required no or minor intervention. In 18.3% of subjects, intervention was required under general anesthesia (Clavien-Dindo grade IIIb complications), usually for debridement (64.7%). Donor site morbidity occurred in 14.0% of the subjects. The age group younger than 60 years showed a significantly higher rate of complications compared with the group older than 60 years (P = 0.015; odds ratio, 0.235; 95% confidence interval, 0.073-0.754). This is the largest study to date on the complications of the lotus petal flap reconstruction. Complication rates did not differ between vulvar, perineal, and vulvoperineal reconstruction. Young patients seem to have a higher risk of developing complications.


[The Efficacy of Neoadjuvant Chemoradiation Therapy for Borderline Resectable Pancreatic Cancer].
Surgical outcomes for locally advanced pancreatic cancer with severe vascular invasion are generally poor. The aim of this study was to evaluate whether neoadjuvant chemoradiation therapy (NACRT) could improve the survival of patients with locally advanced pancreatic cancer without distant metastasis. Between 2010 and 2014, a total of 20 consecutive patients with locally advanced pancreatic cancer without distant metastasis were treated with NACRT. These patients included borderline resectable (BR) and unresectable (UR) pancreatic cancer patients according to the NCCN guidelines. All patients were treated with 2 courses of low-dose gemcitabine plus S-1 combination (GS) chemotherapy and 50.4 Gy of radiation therapy. Restaging was performed 4-6 weeks after completion of NACRT, and curative resection was performed if local control was achieved and distant metastasis was absent. Of the 20 patients, 7 (35.0%) underwent curative resection after NACRT. R0 resections were achieved in 6 patients (85.6%), and R1 resection was achieved in only 1 patient. Curative operation could not be performed in 13 patients (65.0%), because 3 patients refused surgery, 3 patients were diagnosed with dissemination on probe laparotomy or laparoscopy, and 7 patients had progressive disease with liver metastasis or dissemination on CT or MRI. Patients with a curative operation had a median disease-free survival (DFS) of 16.3 months and a median overall survival (OS) of 25.2 months. All of the patients without curative operation were treated with full-dose GS chemotherapy after NACRT. These patients had a median OS of 11.3 months. Our strategy for treating BR or UR pancreatic cancer patients seems to be effective and there might be survival benefits if curative operations can be performed after NACRT.


Circadian light-input pathways in Drosophila.
Light is the most important environmental cue to entrain the circadian clock in most animals. In the fruit fly Drosophila melanogaster, the light entrainment mechanisms of the clock have been well-studied. The Drosophila brain contains approximately 150 neurons that rhythmically express circadian clock genes. These neurons are called "clock neurons" and control behavioral activity rhythms. Many clock neurons express the Cryptochrome (CRY) protein, which is sensitive to UV and blue light, and thus enables clock neurons deep in the brain to directly perceive light. In addition to the CRY protein, external photoreceptors in the Drosophila eyes play an important role in circadian light-input pathways. Recent studies have provided new insights into the mechanisms that integrate these light inputs into the circadian network of the brain. In this review, we will summarize the current knowledge on the light entrainment pathways in the Drosophila circadian clock.