Analysis of micro-contact printed protein patterns by SPR imaging with a LED light source.
We demonstrate the characterization of mu-contact printed protein patterns and analysis of protein-protein interactions by two-dimensional (2-D) surface plasmon resonance imaging (SPRi). Advancements in SPRi image quality from employing a light emitting diode (LED) as the light source are described. We show that a LED offers an ideal point source that can eliminate interference artifacts and speckles found when using a laser source. The attainable thickness resolution in fixed-angle imaging is comparable to that of a monochromatic source, providing a solid foundation for quantitative analysis with the system. The SPR imaging technique reported here affords sub-nanometer thickness sensitivity and micrometer lateral resolution, allowing for convenient studies of biomolecular interactions and surface morphologies of ultrathin films. Spatially well-defined protein patterns of bacterial toxins were obtained by microcontact printing using a polydimethylsiloxane (PDMS) stamp on a functionalized self-assembled monolayer on Au. The influence of protein concentration in the inking solution on transfer efficiency was investigated, and a nonlinear correlation was observed between the solution concentration and the amount of protein immobilized on the surface. Quantitative analysis of protein interaction was performed with toxin-specific antibody, showing a concentration-dependent relationship that verifies the retention of biological activity of the protein after printing. The study demonstrates the feasibility and effectiveness of using LEDs as light sources in SPR imaging, opening doors for developing compact SPR instruments for direct, sensitive, and label-free detection of biohazardous molecules.


Neural retina limits the nonviral gene transfer to retinal pigment epithelium in an in vitro bovine eye model.
We investigated the permeation of liposomal and polymeric gene delivery systems through neural retina into retinal pigment epithelium (RPE) and determined the roles of various factors in permeation and subsequent uptake of the delivery systems by RPE. Anterior parts and vitreous of fresh bovine eyes were removed. Retina was left intact or peeled away. Complexes of ethidium monoazide (EMA)-labeled plasmid DNA and cationic carriers (polyethyleneimine, poly-L-lysine, DOTAP liposomes) were pipetted on the retina or RPE. Two hours later the neural retina was removed, if present, and the RPE cells were detached. Contaminants were removed by sucrose centrifugation, and the RPE cells were analyzed for DNA uptake by flow cytometry. Cellular uptake of FITC-dextrans (molecular weight [mw] 20,000, 500,000 and 2,000,000), FITC-poly-L-lysine (mw 20,000), FITC-labeled oligonucleotide (15-mer), and naked EMA-labeled plasmid DNA was determined after pipetting the solutions on the RPE or neural retina. Location of the fluorescent materials in the retina was visualized with fluorescence microscopy. Neural retina decreased the cellular uptake of DNA complexes by an order of magnitude, the uptake of FITC-dextrans slightly, whereas delivery of polycationic FITC-poly-L-lysine to RPE was almost completely inhibited. Neural retina decreased the cellular uptake of FITC-oligonucleotides, while the uptake of uncomplexed plasmid was always negligible. Conclusions from FACS and fluorescence microscopy were similar: delivery of polymeric and liposomal DNA complexes into RPE are limited by the neural retina. This is due to the size and positive charge of the complexes.


Surgical planing of the skin; evaluation of a method for reducing scars and other defects.
Skin defects such as pitted acne scars, unsightly moles and birthmarks, and other disfiguring skin deformities can be effectively corrected with good cosmetic results. This is accomplished as an office procedure by abrading or planing the skin with a rapidly rotating wire brush using a local anesthetic. The treatment does not require hospitalization and is relatively simple and painless.


Impact of thrombus length on recanalization and clinical outcome following mechanical thrombectomy in acute ischemic stroke.
The impact of thrombus length on recanalization in IV thrombolysis for acute intracranial artery occlusion has been well studied. Here we analyzed the influence of thrombus length on the number of thrombectomy maneuvers needed for recanalization, intraprocedural complications, recanalization success, and clinical outcome after mechanical thrombectomy. We retrospectively analyzed angiographic and clinical data from 72 consecutive patients with acute occlusion of the M1 segment of the middle cerebral artery who were treated with mechanical thrombectomy using stent retrievers. Successful recanalization was defined as a Thrombolysis in Cerebral Infarction score of 2b or 3. Good neurological outcome was defined as a modified Rankin Scale score of ≤2 at 90 days after stroke onset. Mean thrombus length was 13.4±5.2 mm. Univariate binary logistic regression did not show an association of thrombus length with the probability of a good clinical outcome (OR 0.95, 95% CI 0.84 to 1.03, p=0.176) or successful recanalization (OR 0.92, 95% CI 0.81 to 1.05, p=0.225). There was no significant correlation between thrombus length and the number of thrombectomy maneuvers needed for recanalization (p=0.112). Furthermore, thrombus length was not correlated with the probability of intraprocedural complications (p=0.813), including embolization in a new territory (n=3). In this study, thrombus length had no relevant impact on recanalization, neurological outcome, or intraprocedural complications following mechanical thrombectomy of middle cerebral artery occlusions. Therefore, mechanical thrombectomy with stent retrievers can be attempted with large clots.


Methylphenidate in hyperkinetic children: differences in dose effects on learning and social behavior.
Methylphenidate (Ritalin) is widely prescribed for hyperkinetic children. This study showed a peak enchancement of learning in children after being given a dose of 0.3 milligram per kilogram of body weight, and a decrement in learning in those given larger doses; social behavior showed the most improvement in children given 1.0 milligram per kilogram. These results had been hypothesized from theoretical dose-response curves which indicate different target behaviors would improve at different doses.


The genetic linkage map of the human X chromosome.
A database useful for mapping the human X chromosome has been established. The data consist of the genotypic characterizations obtained at more than 20 DNA marker loci from a set of 38 selected families. Multilocus linkage analysis has provided an initial genetic map completely spanning the distance from the distal short arm to the distal long arm of the chromosome, for a total genetic length of at least 185 recombination units. Analysis of the recombinational behavior of fully marked chromosomes suggests that the number of recombination events on the X chromosome may be nonrandom. Linkage studies of six families that carry the mutation which causes Duchenne muscular dystrophy were combined with linkage data from a large number of normal families. This permitted mapping of the locus for Duchenne muscular dystrophy with greater precision and statistical confidence than studies in which disease families alone provided the genotypic database. This observation suggests that the normal linkage map of this chromosome should be especially valuable in the mapping of rare X-linked diseases.


The Importance of Parent-Teacher Informant Discrepancy in Characterizing Autistic Youth: A Replication Latent Profile Analysis.
Research about autism spectrum disorder (ASD) supports variation in symptom presentations across settings, and there is a growing literature that explicates how this variability may improve characterization of the autism phenotype. Capitalizing on a well-established literature on informant discrepancy as an index of contextual variability, research suggests that differing parent and teacher perceptions may impact treatment or education-related outcomes. A prior investigation by Lerner and colleagues suggests that parent-teacher discrepancies in ASD symptom ratings define discrete and clinically meaningful subgroups. However, replication in a larger sample is important to support the validity and utility of the subgroups for use in research and practice. The present paper used latent profile analysis (LPA) to (1) replicate the previous study by Lerner and colleagues in a larger sample of 514 clinic-referred autistic youth (aged 6-18, 83.2% male, 90.4% White, IQ 19-140) and (2) determine if parent-teacher informant discrepancies relate to clinical and functional correlates. We hypothesized that parent-teacher discrepancies in ASD symptom severity ratings would validly characterize ASD subgroups and predict clinical and functional correlates. The results of the LPA supported a 4-profile solution made up of two parent-teacher agreement groups (high parent-teacher, 21.2%, and low parent-teacher, 34.2%) and two parent-teacher discrepancy groups (high parent-low teacher, 18.1%, and moderate parent-high teacher, 26.5%), replicating findings from Lerner and colleagues. Latent profile membership differentially predicted IQ, age, and educational outcomes of participants. Unique, clinically useful information about the taxonomy and impact of ASD is obtained by considering informant discrepancies in symptom severity ratings, which underscores the importance of considering contextual variability assessed through multiple informants.


Thoracic aortic distensibility and thoracic aortic calcium (from the Multi-Ethnic Study of Atherosclerosis [MESA]).
Decreased arterial distensibility is an early manifestation of adverse structural and functional changes within the vessel wall. Its correlation with thoracic aortic calcium (TAC), a marker of atherosclerosis, has not been well demonstrated. We tested the hypothesis that decreasing aortic compliance and increasing arterial stiffness would be independently associated with increased TAC. We included 3,540 subjects (61 +/- 10 years, 46% men) from the Multi-ethnic Study of Atherosclerosis who had undergone an aortic distensibility (AD) assessment using magnetic resonance imaging. TAC was calculated using a modified Agatston algorithm on noncontrast cardiac computed tomographic scans. Multivariate regression models were calculated for the presence of TAC. Overall, 861 subjects (24%) had detectable TAC. Lower AD was observed among those with versus without TAC (2.02 +/- 1.34 vs 1.28 +/- 0.74, p <0.0001). The prevalence of TAC increased significantly across decreasing quartiles of AD (7%, 17%, 31%, and 42%, p <0.0001). Using multivariate analysis, TAC was independently associated with AD after adjusting for age, gender, ethnicity, and other covariates. In conclusion, our analysis has demonstrated that increased arterial stiffness is associated with increased TAC, independent of ethnicity and other atherosclerotic risk factors.


Genotype and phenotype of NAT2 and the occurrence of adverse drug reactions in Mexican individuals to an isoniazid-based prophylactic chemotherapy for tuberculosis.
Isoniazid (INH) is a drug extensively used as a prophylactic and therapeutic agent for human tuberculosis (TB). INH is metabolized by the enzymatic activity of N-acetyltransferase 2 (NAT2). Human NAT2, encoded by a highly polymorphic gene, is involved in the biotransformation of xenobiotics, including drugs and certain chemical carcinogens. Numerous studies have established the correlation between the acetylator phenotype and the NAT2 genotype in several populations; however, little is known regarding Latin-American populations and the pharmacogenetics of NAT2. Here, we report the molecular genotyping of the NAT2 gene, the acetylator phenotype, and the incidence of INH-related adverse reactions in a group of 25 Mexican individuals enrolled in a prophylactic protocol for TB. Using both the NAT2 genotyping and acetylation phenotyping approach, we found a ratio of 69.2 and 30.8% of slow and fast acetylators, respectively. Concordance of the NAT2 genotype and phenotype classification was 88% in the bimodal model. Regarding INH-related adverse reactions, only 2 individuals (8%) exhibited declared gastric intolerance. In our study group, we found an association between the NAT2 genotype and acetylator phenotype (OR=7.78, 95% CI, 0.87-87.98, Fisher's exact test, p<0.05), but did not find any genotype or phenotype association with the incidence of INH-related adverse reactions (Fisher's exact test, p>0.05).


The independent nurse practitioner. Alive and well and living in rural Saskatchewan.
At this point, it is impossible to express analysis of the data collected in concrete terms as the project will not be finalized until October, 1976. It is possible, however, to make some personal observations. Each nurse practitioner has been truly independent in carrying out her role in the community, influenced to a degree by the communities' felt need and the attitudes of other health professionals. All began practice with a heavy emphasis on diagnosis and cure rather than comprehensive care. As time evolves and the nurses feel more confident in their new role, some are continuing a cure orientation, and others are beinning to find that they have a unique dimension to add to health care services. Those who were professional "nurses" in the true sense of the word before the project began have not stopped being nurses, but have only become more competent professionals by recognizing and responding to the health needs of today.


Comparative population analysis of metallothionein promoter alleles suggests stress-induced microevolution in the field.
We investigate a model system for microevolution of transcriptional regulation: metallothionein expression in springtails. A previous survey of the metallothionein promoter in Orchesella cincta (Collembola) revealed nine alleles with differential basal activities and responses to cadmium and oxidative stress. In this study, 23 woodlands, with a divergent degree of pollution, were sampled, and heavy metals were measured. When grouped to their contamination degree, they were discriminated best on the pmtD2 metallothionein promoter allele frequency, which was higher in populations from heavily polluted sites. Taken together with previous work showing high inducibility of the pmtD2 promoter allele by Cd in a reporter assay, this suggests a fitness advantage of the pmtD2 allele in polluted sites. Redundancy analysis revealed associations between allele frequencies and specific metals in the environment, resulting in a subdivision between pollution associated alleles and others. A positive relationship between the pmtD2 allele frequency and the Cd content of the soil as well as between pmtE and Ni in the litter emerged. An increase of genetic diversity was observed with increasing Pb in the soil, reached through substitution of the pmtA1 allele, suggesting balancing selection. Our results illustrate that environmental factors can exert selection on promoter polymorphisms and cause adaptation through altered transcriptional regulation.


The microRNA159-regulated GAMYB-like genes inhibit growth and promote programmed cell death in Arabidopsis.
The microRNA159 (miR159) family represses the conserved GAMYB-like genes that encode R2R3 MYB domain transcription factors that have been implicated in gibberellin (GA) signaling in anthers and germinating seeds. In Arabidopsis (Arabidopsis thaliana), the two major miR159 family members, miR159a and miR159b, are functionally specific for two GAMYB-like genes, MYB33 and MYB65. These transcription factors have been shown to be involved in anther development, but there are differing reports about their role in the promotion of flowering and little is known about their function in seed germination. To understand the function of this pathway, we identified the genes and processes controlled by these GAMYB-like genes. First, we demonstrate that miR159 completely represses MYB33 and MYB65 in vegetative tissues. We show that GA does not release this repression and that these transcription factors are not required for flowering or growth. By contrast, in the absence of miR159, the deregulation of MYB33 and MYB65 in vegetative tissues up-regulates genes that are highly expressed in the aleurone and GA induced during seed germination. Confirming that these genes are GAMYB-like regulated, their expression was reduced in myb33.myb65.myb101 seeds. Aleurone vacuolation, a GA-mediated programmed cell death process required for germination, was impaired in these seeds. Finally, the deregulation of MYB33 and MYB65 in vegetative tissues inhibits growth by reducing cell proliferation. Therefore, we conclude that miR159 acts as a molecular switch, only permitting the expression of GAMYB-like genes in anthers and seeds. In seeds, these transcription factors participate in GA-induced pathways required for aleurone development and death.


Efficient bioconjugation of protein capture agents to biosensor surfaces using aniline-catalyzed hydrazone ligation.
Aniline-catalyzed hydrazone ligation between surface-immobilized hydrazines and aldehyde-modified antibodies is shown to be an efficient method for attaching protein capture agents to model oxide-coated biosensor substrates. Silicon photonic microring resonators are used to directly evaluate the efficiency of this surface bioconjugate reaction at various pHs and in the presence or absence of aniline as a nucleophilic catalyst. It is found that aniline significantly increases the net antibody loading for surfaces functionalized over a pH range from 4.5 to 7.4, allowing derivatization of substrates with reduced incubation time and sample consumption. This increase in antibody loading directly results in more sensitive antigen detection when functionalized microrings are employed in a label-free immunoassay. Furthermore, these experiments also reveal an interesting pH-dependent noncovalent binding trend that plays an important role in dictating the amount of antibody attached onto the substrate, highlighting the competing contributions of the bioconjugate reaction rate and the dynamic interactions that control opportunities for a solution-phase biomolecule to react with a substrate-bound reagent.


Successful Endoscopic Removal of Anisakis Larva that Deeply Invaded the Gastric Mucosa Using Jumbo Forceps.
Anisakiasis is a parasitic disease caused by the ingestion of raw or uncooked seafood infected with third-stage larvae of anisakid nematodes. Generally, the larvae parasites live at the surface of the mucosa, but in this case, the larva deeply invaded its head into the gastric mucosa and was not removable with conventional biopsy forceps. In our case, we demonstrated the usefulness of jumbo forceps to remove the Anisakis larva in such a situation.


Maintenance of genomic integrity in hematopoietic stem cells.
Hematopoietic stem cells (HSCs) maintain hematopoietic homeostasis throughout a mammal's lifespan through self-renewal and differentiation into mature blood cells. Within a bone marrow niche, HSCs adopt a quiescent state and remain in the non-dividing, G0 phase of the cell cycle. It was recently shown that maintenance of genomic integrity is crucial for the preservation of self-renewal capacity of HSCs. In this review, we focus on progress in elucidating the roles of reactive oxygen species (ROS) and DNA damage responses (DDR) in maintaining genomic integrity, and thus HSC function. Several studies have demonstrated that inappropriate ROS levels arising from disruption of the Atm, PI3K-Akt, or Mdm2-p53 pathways impair HSC function in vivo. Intriguing evidence that stem cells use specific DDR mechanisms is also accumulating. Although murine HSCs are more resistant than progenitor cells to mild DNA damage in vivo, the surviving HSCs frequently acquire genetic aberrations that can lead to leukemogenesis. Indeed, non-dividing HSCs employ the error-prone non-homologous end-joining pathway of DNA repair to fix DNA breaks, whereas progenitors undergo apoptosis; proliferating HSCs employ the high-fidelity homologous recombination mechanism. Dissection of HSC-specific mechanisms for the maintenance of genomic integrity may provide valuable insights into the biology of both HSCs and leukemia stem cells.


Therapeutic interventions and the length of hospital stay for pediatric patients with COVID-19: a multicenter cohort study.
The evidence for pediatric patients with COVID-19 was very limited, which was attributed to the small number of the cases as well as the rare incidence of severe pneumonia in this population. This retrospective cohort study aimed to identify the characteristics of pediatric patients with COVID-19 in the early period of the pandemic by analyzing Diagnosis Procedure Combination (DPC) data in Japan. This retrospective cohort analysis of Japanese multicenter research on COVID-19 using DPC data compared the outcomes and costs of treatment for pediatric patients with COVID-19. Of 4700 patients with COVID-19, 186 pediatric patients were included in this study. Among the included pediatric patients, 17 received therapeutic drugs specifically for COVID-19, while the remaining 169 pediatric patients received only symptomatic therapy. There were no significant differences in the length of hospital stay (9 vs. 8 days, p = 0.96), and medical cost (97,585 vs. 73,291 JPY) for the intervention and control groups, respectively by multiple regression analysis. This is the first epidemiological study to use DPC data to summarize the pathophysiology of pediatric patients in the early period of COVID-19 pandemic. There was no significant difference in length of hospital stay or medical cost by intervention.


An Overview of Genes From Cyberlindnera americana, a Symbiont Yeast Isolated From the Gut of the Bark Beetle Dendroctonus rhizophagus (Curculionidae: Scolytinae), Involved in the Detoxification Process Using Genome and Transcriptome Data.
Bark beetles from Dendroctonus genus promote ecological succession and nutrient cycling in coniferous forests. However, they can trigger outbreaks leading to important economic losses in the forest industry. Conifers have evolved resistance mechanisms that can be toxic to insects but at the same time, bark beetles are capable of overcoming tree barriers and colonize these habitats. In this sense, symbiont yeasts present in the gut of bark beetles have been suggested to play a role in the detoxification process of tree defensive chemicals. In the present study, genes related to this process were identified and their response to a terpene highly toxic to bark beetles and their symbionts was analyzed in the Cyberlindnera americana yeast. The genome and transcriptome of C. americana (ChDrAdgY46) isolated from the gut of Dendroctonus rhizophagus were presented. Genome analysis identified 5752 protein-coding genes and diverse gene families associated with the detoxification process. The most abundant belonged to the Aldo-Keto Reductase Superfamily, ATP-binding cassette Superfamily, and the Major Facilitator Superfamily transporters. The transcriptome analysis of non-α-pinene stimulated and α-pinene stimulated yeasts showed a significant expression of genes belonging to these families. The activities demonstrated by the genes identified as Aryl-alcohol dehydrogenase and ABC transporter under (+)-α-pinene suggest that they are responsible, that C. americana is a dominant symbiont that resists high amounts of monoterpenes inside the gut of bark beetles.


Use of menstrual cups among school girls: longitudinal observations nested in a randomised controlled feasibility study in rural western Kenya.
A menstrual cup can be a good solution for menstrual hygiene management in economically challenged settings. As part of a pilot study we assessed uptake and maintenance of cup use among young school girls in Kenya. A total of 192 girls between 14 to 16 years were enrolled in 10 schools in Nyanza Province, Western Kenya; these schools were assigned menstrual cups as part of the cluster-randomized pilot study. Girls were provided with menstrual cups in addition to training and guidance on use, puberty education, and instructions for menstrual hygiene. During repeated individual visits with nurses, girls reported use of the menstrual cup and nurses recorded colour change of the cup. Girls were able to keep their cups in good condition, with only 12 cups (6.3%) lost (dropped in toilet, lost or destroyed). Verbally reported cup use increased from 84% in the first 3 months (n = 143) to 96% after 9 months (n = 74). Colour change of the cup, as 'uptake' indicator of use, was detected in 70.8% of 192 participants, with a median time of 5 months (range 1-14 months). Uptake differed by school and was significantly higher among girls who experienced menarche within the past year (adjusted risk ratio 1.29, 95% CI 1.04-1.60), and was faster among girls enrolled in the second study year (hazard ratio 3.93, 95% CI 2.09-7.38). The kappa score comparing self-report and cup colour observation was 0.044 (p = 0.028), indicating that agreement was only slightly higher than by random chance. Objective evidence through cup colour change suggests school girls in rural Africa can use menstrual cups, with uptake improving with peer group education and over time. ISRCTN17486946 . Retrospectively registered 09 December 2014.


Midwives perceptions of managing pregnancies complicated by obesity: A mixed methods study.
The growing prevalence of obesity is a concern for midwives. In Canada, the absence of regulatory standards, varying protocols and consultant preferences shape clinical decision making for the midwife and may lead to inconsistent practice. Our aim was to understand the barriers, enablers, and knowledge gaps that influenced experiences of midwives in Ontario, Canada when providing care to clients impacted by obesity. Mixed methods design using a sequential, explanatory approach. Surveys conducted with midwives were administered using an online platform, followed by semi-structured interviews to understand the perspectives elicited in the survey in greater detail. Interviews were audio recorded and transcribed verbatim. Survey data were analyzed using descriptive statistics, and thematic analysis was used for generating codes, categories and themes from the interview data. 144 midwives completed the survey and 20 participated in an interview. The participants described their clinical management when caring for those with obesity which included considerations regarding additional tests/investigations, consultation and transfer of care, and place of birth. Up to 93% of surveyed midwives believed that clients with obesity were appropriate for midwifery-led care however there was less certainty about suitability as BMI increased to higher ranges such as > 45). The care management was influenced by beliefs and attitudes, knowledge, and system-level factors. Midwives experienced barriers such as inconsistent practices and role confusion, and felt ill equipped to care for pregnancies affected by obesity due to unclear guidelines. Overall, midwives believe clients with obesity are suitable for midwifery-led care due to its individualized, non-judgmental approach to care. Additional training for midwives and other obstetric care providers would be beneficial to help overcome barriers in providing effective care to pregnancies affected by obesity.


Review of current protocols for protontherapy in USA.
The basis for interest in proton beams by clinical radiation oncologists lies in reduction in treatment volume. The yields from employing a smaller treatment volume are the increase of tumor control probability and the reduction of normal tissues complication probability. The clinical use of proton therapy began in 1954 at Uppsala University in Sweden and in 1961 at Harvard Cyclotron Laboratory in Boston, USA. So far, the total number of worldwide patients treated by protons is about 20,000. In this paper attention will be given to the treatment of patients at the Massachusetts General Hospital-Massachusetts Eye and Ear Infirmary-Harvard Cyclotron Laboratory, and at the Loma Linda University Medical Center. In particular, a review of the literature about the techniques and the results of treatment of skull base and cervical spine chordoma and low-grade chondrosarcoma, skull base meningioma, pituitary tumors, paranasal sinus carcinoma, glioblastoma multiforme, artero-venous malformations, uveal melanoma, macular degeneration, retinoblastoma, thoracic spine-sacrum tumors, and prostate carcinoma is presented. In order to verify and improve the clinical results, the conduct of prospective trials on an inter-institutional basis is essential. To facilitate the conduct of such studies the US National Cancer Institute and the American College of Radiology have established the Proton Therapy Oncology Group (PROG). Several phase III and some phase I-II trials are active at the Massachusetts General Hospital, Harvard Cyclotron Laboratory, and at the Loma Linda University Medical Center.


Hypervariable intronic region in NCX1 is enriched in short insertion-deletion polymorphisms and showed association with cardiovascular traits.
Conserved non-coding regions (CNR) have been shown to harbor gene expression regulatory elements. Genetic variations in these regions may potentially contribute to complex disease susceptibility. We targeted CNRs of cardiovascular disease (CVD) candidate gene, Na(+)-Ca(2+) exchanger (NCX1) with polymorphism screening among CVD patients (n = 46) using DHPLC technology. The flanking region (348 bp) of the 14 bp indel in intron 2 was further genotyped by DGGE assay in two Eastern-European CVD samples: essential hypertension (HYPEST; 470 cases, 652 controls) and coronary artery disease, CAD (CADCZ; 257 cases, controls 413). Genotype-phenotype associations were tested by regression analysis implemented in PLINK. Alignments of primate sequences were performed by ClustalW2. Nine of the identified NCX1 variants were either singletons or targeted by commercial platforms. The 14 bp intronic indel (rs11274804) was represented with substantial frequency in HYPEST (6.82%) and CADCZ (14.58%). Genotyping in Eastern-Europeans (n = 1792) revealed hypervariable nature of this locus, represented by seven alternative alleles. The alignments of human-chimpanzee-macaque sequences showed that the major human variant (allele frequency 90.45%) was actually a human-specific deletion compared to other primates. In humans, this deletion was surrounded by other short (5-43 bp) deletion variants and a duplication (40 bp) polymorphism possessing overlapping breakpoints. This indicates a potential indel hotspot, triggered by the initial deletion in human lineage. An association was detected between the carrier status of 14 bp indel ancestral allele and CAD (P = 0.0016, OR = 2.02; Bonferroni significance level alpha = 0.0045), but not with hypertension. The risk for the CAD development was even higher among the patients additionally diagnosed with metabolic syndrome (P = 0.0014, OR = 2.34). Consistent with the effect on metabolic processes, suggestive evidence for the association with heart rate, serum triglyceride and LDL levels was detected (P = 0.04). Compared to SNPs targeted by large number of locus-specific and genome-wide assays, considerably less attention has been paid to short indel variants in the human genome. The data of genome dynamics, mutation rate and population genetics of short indels, as well as their impact on gene expressional profile and human disease susceptibility is limited. The characterization of NCX1 intronic hypervariable non-coding region enriched in human-specific indel variants contributes to this gap of knowledge.


Dwarfism in insular sloths: biogeography, selection, and evolutionary rate.
The islands of Bocas del Toro, Panama, were sequentially separated from the adjacent mainland by rising sea levels during the past 10,000 years. Three-toed sloths (Bradypus) from five islands are smaller than their mainland counterparts, and the insular populations themselves vary in mean body size. We first examine relationships between body size and physical characteristics of the islands, testing hypotheses regarding optimal body size, evolutionary equilibria, and the presence of dispersal in this system. To do so, we conduct linear regressions of body size onto island area, distance from the mainland, and island age. Second, we retroactively calculate two measures of the evolutionary rate of change in body size (haldanes and darwins) and the standardized linear selection differential, or selection intensity (i). We also test the observed morphological changes against models of evolution by genetic drift. The results indicate that mean body size decreases linearly with island age, explaining up to 97% of the variation among population means. Neither island area nor distance from the mainland is significant in multiple regressions that include island age. Thus, we find no evidence for differential optimal body size among islands, or for dispersal in the system. In contrast, the dependence of body size on island age suggests uniform directional selection for small body size in the insular populations. Although genetic drift cannot be discounted as the cause for this evolution in body size, the probability is small given the consistent direction of evolution (repeated dwarfism). The insular sloths show a sustained rate of evolution similar to those measured in haldanes over tens of generations, appearing to unite micro- and macroevolutionary time scales. Furthermore, the magnitude and rate of this example of rapid differentiation fall within predictions of theoretical models from population genetics. However, the linearity of the relationship between body size and island age is not predicted, suggesting that either more factors are involved than those considered here, or that theoretical advances are necessary to explain constant evolutionary rates over long time spans in new selective environments.


The outcome of patients with unclassified hypogammaglobulinemia in early childhood.
Symptomatic hypogammaglobulinemia in childhood may be the initial finding of primary immunodeficiency (PID) or may be due to delay in maturation of immunoglobulin synthesis. The aim of this study was to review the clinical and laboratory records of patients with unclassified hypogammaglobulinemia and to evaluate whether these children experience changes in serum immunoglobulin concentrations during long-term followup and have an exact diagnosis in natural course of disease. We reviewed the data of 412 patients who were diagnosed as PID with symptomatic hypogammaglobulinemia. Thirty-seven patients with hypogammaglobulinemia [19 males (51.4%) and 18 females (48.6%), with a followup of 34.1 +/- 22.0 months] who were not classified according to European Society for Immunodeficiencies diagnostic criteria were included in this study. The mean age at the beginning of the symptoms was 21.4 +/- 20.6 months and the mean age at admission was 51.5 +/- 25.8 months. The commonest clinical presentations were recurrent upper (94.6%) and/or lower (40.5%) respiratory infections, urinary infection (27%) and gastroenteritis (10.8%). Percentage of consanguinity was 8%. Of the initial 37 patients, 18 (48.6%) spontaneously corrected their immunoglobulin abnormalities during followup. Clinical symptoms of these patients were also improved. IgG, IgA and IgM levels reached to normal levels at ages 62.5 +/- 21.8, 72.0 +/- 11.2, 55.2 +/- 7.8 months, respectively. In remaining 19 patients with undefined/unclassified hypogammaglobulinemia, three partial IgA deficiency, seven IgG subclass deficiency, two selective IgM deficiency and two common variable immunodeficiency (CVID) were diagnosed by long-term monitoring of immunoglobulin levels. Five (13.5%) of the 37 unclassified patients could not be exactly diagnosed while two of them might have a T-cell defect and three of them still had low IgG and IgA levels but adequate antibody responses against vaccine antigens. In conclusion, it is important to monitor symptomatic patients with hypogammaglobulinemia periodically. Some children may spontaneously correct their immunoglobulin abnormalities not in the first 30 months of age, but during the first decade of life and some of them may have a severe PID like CVID.


Notes from an epicenter: navigating behavioral clinical trials on autism spectrum disorder amid the COVID-19 pandemic in the Bronx.
The COVID-19 pandemic impacted nearly all facets of our daily lives, and clinical research was no exception. Here, we discuss the impact of the pandemic on our ongoing, three-arm randomized controlled trial (RCT) Sensory Integration Therapy (SIT) in Autism: Mechanisms and Effectiveness (NCT02536365), which investigates the immediate and sustained utility of SIT to strengthen functional daily-living skills and minimize the presence of maladaptive sensory behaviors in autistic children. In this text, we detail how we navigated the unique challenges that the pandemic brought forth between the years 2020 and 2021, including the need to rapidly adjust our study protocol, recruitment strategy, and in-person assessment battery to allow for virtual recruitment and data collection. We further detail how we triaged participants and allocated limited resources to best preserve our primary outcome measures while prioritizing the safety of our participants and study team. We specifically note the importance of open and consistent communication with all participating families throughout the pandemic in ensuring all our protocol adjustments were successfully implemented. Though the COVID-19 pandemic resulted in an unprecedented interruption to in-person clinical research, clinical trials have always been and will continue to be at risk for unforeseen interruptions, whether from world events or participants' personal circumstances. By presenting our steps to preserving this RCT throughout the pandemic, we offer suggestions for successfully managing unexpected interruptions to research. Ideally, by taking these into account, future RCTs may be increasingly prepared to minimize the impact of these potential interruptions to research.


Evidence of free tropospheric and long-range transport of microplastic at Pic du Midi Observatory.
The emerging threat of atmospheric microplastic pollution has prompted researchers to study areas previously considered beyond the reach of plastic. Investigating the range of atmospheric microplastic transport is key to understanding the global extent of this problem. While atmospheric microplastics have been discovered in the planetary boundary layer, their occurrence in the free troposphere is relatively unexplored. Confronting this is important because their presence in the free troposphere would facilitate transport over greater distances and thus the potential to reach more distal and remote parts of the planet. Here we show evidence of 0.09-0.66 microplastics particles/m3 over 4 summer months from the Pic du Midi Observatory at 2877 meters above sea level. These results exhibit true free tropospheric transport of microplastic, and high altitude microplastic particles <50 µm (aerodynamic diameter). Analysis of air/particle history modelling shows intercontinental and trans-oceanic transport of microplastics illustrating the potential for global aerosol microplastic transport.


Circulating CD14+CD204+ cells predict postoperative recurrence in non-small-cell lung cancer patients.
The expression of CD204 on macrophages in the stroma of the primary tumor is reportedly correlated with an unfavorable prognosis for lung cancer. The purpose of this study is to investigate the correlation among the number of CD204 tumor-associated macrophages infiltrating the stroma of the primary tumor, the number of circulating CD14CD204 cells from the pulmonary vein (PV), and recurrence-free probability in non-small-cell lung cancer patients. Human mononuclear cells were isolated from the PV of resected lungs. We examined the expressions of CD14 and CD204 on these cells by flow cytometry. Immunohistochemical staining for CD204 was performed in the resected specimens. The number of CD14CD204 cells from the PV was found to be correlated with the number of CD204 tumor-associated macrophages identified in the stroma of the tumor. Significantly more cases with high levels of CD14CD204 cells from the PV were found to have developed early recurrences. CD14CD204 cells, which were polarized to the tumor-promoting phenotype cultured in lung cancer cell line-conditioned medium, facilitated the lung metastasis of cancer cells more effectively than CD14CD204 cells in our in vivo mouse model. In multivariate analysis, only the high number of CD14CD204 cells from the PV was found to be a statistically significant independent risk factor for early recurrence. Our results showed the possibility that circulating CD14CD204 cells contribute to the metastasis of cancer cells. The blockage of circulating CD14CD204 cells activity may prevent postoperative recurrence in resected non-small-cell lung cancer patients.


Effect of the COVID-19 pandemic on life expectancy in Australia, 2020-22.
Australia provides a valuable international case study of life expectancy during the pandemic. In contrast to many other countries, it experienced relatively stringent restrictions and low COVID-19 mortality during 2020-21, followed by relaxation of these restrictions when high vaccination rates were achieved. This study measures Australia's life expectancy trends and the contributions of age group and causes of death, during the pandemic. Trends in life expectancy at birth in Australia and its states and territories were measured from 2020 to 2022. The contributions of age group and cause of death to these trends were measured using decomposition methods. Life expectancy was compared with other high-income countries. Australia's life expectancy fell by more than half a year in 2022, following a sharp increase in 2020 and moderate decline in 2021. For the 3 years 2020 to 2022, life expectancy was 0.13 years (95% confidence interval 0.07-0.19) higher for males and 0.09 years (0.03-0.14) higher for females versus 2017-19. Australia's life expectancy increase in 2020 was larger than that in the vast majority of other high-income countries, but its decline in 2022 was greater than in other countries whose life expectancy rose in the first year of the pandemic. The small negative contribution of COVID-19 deaths to life expectancy in Australia was more than offset by lower non-communicable disease mortality. There were only small differences in life expectancy change between the states with the most stringent restrictions (Victoria and New South Wales) and the rest of Australia. Australia's life expectancy trends during 2020-22 were relatively favourable compared with other high-income countries, with the exception of its sharp decline in 2022 once restrictions were loosened.


Potential of Snoezelen room multisensory stimulation to improve balance in individuals with dementia: a feasibility randomized controlled trial.
To investigate the influence of multisensory stimulations in a Snoezelen room on the balance of individuals with dementia. Randomized controlled trial. Canadian long-term care home. Twenty-four residents (average age 86 years), in a long-term care home diagnosed with dementia, were assigned randomly to intervention and control groups. Nineteen participants completed the study. Nine intervention group participants completed 30-minute Snoezelen room sessions twice a week for six weeks. Sessions were guided by participants' preferences for stimulation. Interactions with tactile, visual and proprioceptive sensations were encouraged. Ten control group participants received an equal amount of volunteer visits. The Functional Reach Test, the eyes-open Sharpened Romberg and the Timed Up and Go Test with and without dual task, assessed static and dynamic balance at baseline and after the intervention. Falls frequencies were recorded six weeks before, during and after intervention. A journal was kept of observations in Snoezelen room. Split-plot MANOVA analyses revealed no significant effects of unstructured Snoezelen room sessions on participants' balance. There were no multivariate effects of time (F(4,14) = 1.13, P = 0.38) or group (F(4,14) = 0.63, P = 0.65). Group membership did not alter falls frequency. However, observations of participants' interactions with elements of the Snoezelen room, such as imagery-induced head and eye movements, vibrating sensations and kicking activities, captured events that can be used to create specific multisensory balance-enhancing stimulations. Although the null hypothesis was not rejected, further investigation of a potential to influence balance in individuals with dementia through Snoezelen room intervention in long-term care homes is warranted.


Perinatal Mortality in the United States, 2020-2021.
Perinatal mortality(late fetal deaths at 28 completed weeks of gestation or more and early neonatal deaths younger than age 7 days) can be an indicator of the quality of health care before, during, and after delivery, and of the health status of the nation (1,2). The U.S. perinatal mortality rate declined 30% from 1990 through 2011, was stable from 2011 through 2016, and declined 4% from 2017 through 2019 (1,3-5). This report describes changes in perinatal mortality, as well as its components, late fetal and early neonatal mortality, from 2020 to 2021, during the COVID-19 pandemic. Also shown are perinatal mortality rates by mother's age, the three largest race and Hispanic-origin groups, and state for 2021 compared with 2020.


Preliminary experience of the Cannulok revision hip prosthesis in late periprosthetic fracture management.
We report our preliminary experience of a cannulated, distally locked revision femoral component to manage peri-prosthetic fractures in the elderly. The majority of cases in our series were further complicated by osteolysis secondary to aseptic loosening, the original hip arthroplasties having been in situ for several years. Sixteen patients with an average age of 77 underwent treatment of periprosthetic fractures with the Cannulok Hip. The device combines techniques of revision hip surgery with locked intramedullary nailing. We describe our initial evaluation of this modular prosthesis which has enabled us to satisfactorily treat periprosthetic fractures with early mobilisation and discharge from hospital. In all of our surviving patients the fractures have united within four to six months. All cases were in addition treated with proximal femoral impaction allografting leading to improved local bone stock. We believe this implant may provide a satisfactory alternative for the management of periprosthetic fracture in the presence of a loose femoral component. (Hip International 2002; 1: 1-10).


Nutritional Implications of Trade-Offs Between Fresh and Processed Potato Products in the United Kingdom (UK).
The UK ranks eleven among world potato producing countries with annual per capita production of about 102 kg. Since 2007, the price of potatoes has increased by 44 per cent and UK households have shown a decreasing trend on their purchases of potatoes. At the same time, retailers and manufacturers have been introducing processed potato products, which also has affected the demand for fresh potatoes. This has shifted demand from fresh potatoes to processed potatoes suggesting that consumers substitute fresh potatoes for processed ones. However, the extent to which this affect individual weekly nutritional composition is unknown. The objective of this study is to estimate the nutritional trade-offs between fresh and processed potatoes consumed in the UK using home scanner panel dataset for Great Britain in 2018. Price and expenditure elasticities were estimated using the linearized version of the Exact Affine Stone Index (EASI) Demand System. Using estimated elasticities, we analyzed the implications of substituting fresh potatoes for processed potatoes on nutrient intake. The results, in terms of the degree of substitution between fresh potatoes and processed potato products, suggest that consumers consider new potatoes baby and baking potatoes as substitutes for mashed potatoes. Maris piper potatoes and new potatoes baby are substitutes for frozen chips and other potatoes whilst white old potatoes and other vegetables and salads are complements to frozen chips and other potatoes. Finally, price reductions in the processed potatoes will increase average weekly caloric intake as well as the intakes of saturated fat and sodium. The latter has implications for public health as they are the major causes of cardiovascular diseases and certain cancers.


Impact of exposure to tobacco smoke, arsenic, and phthalates on locally advanced cervical cancer treatment-preliminary results.
Cancer research is a national and international priority, with the efficiency and effectiveness of current anti-tumor therapies being one of the major challenges with which physicians are faced. To assess the impact of exposure to tobacco smoke, arsenic, and phthalates on cervical cancer treatment. We investigated 37 patients with locally advanced cervical carcinoma who underwent chemotherapy and radiotherapy. We determined cotinine and five phthalate metabolites in urine samples collected prior to cancer treatment, by gas chromatography coupled to mass spectrometry, and urinary total arsenic by atomic absorption spectrometry with hydride generation. We used linear regression to evaluate the effects of cotinine, arsenic, and phthalates on the change in tumor size after treatment, adjusted for confounding variables. We detected no significant associations between urinary cotinine, arsenic, or phthalate monoesters on change in tumor size after treatment, adjusted for urine creatinine, age, baseline tumor size, and cotinine (for arsenic and phthalates). However, higher %mono-ethylhexyl phthalate (%MEHP), a putative indicator of phthalate diester metabolism, was associated with a larger change in tumor size (β = 0.015, 95% CI [0.003-0.03], P = 0.019). We found no statistically significant association between the urinary levels of arsenic, cotinine, and phthalates metabolites and the response to cervical cancer treatment as measured by the change in tumor size. Still, our results suggested that phthalates metabolism may be associated with response to treatment for locally advanced cervical cancer. However, these observations are preliminary and will require confirmation in a larger, more definitive investigation.


Language barrier leads to the unnecessary intubation of trauma patients.
Airway evaluation in trauma patients is performed immediately upon patient contact, with communication being a vital component to this exam. Language and communication barriers may lead to the unnecessary placement of an artificial airway with resultant patient risk and elevation of health care costs. The objective of our study was to evaluate potentially preventable intubations in Spanish-speaking patients. A 9-year retrospective review was performed using the National Trauma Registry for The American College of Surgeons (NTRACS) database. We evaluated patients intubated on arrival to the trauma center and remaining intubated for less than 48 hours. Deaths were excluded. Patients who typically speak English were compared with patients who typically speak Spanish. Mechanism of injury (MOI), hypotension during resuscitation (HDR), illicit substance use, alcohol use, mean Glasgow Coma Score (GCS), mean Injury Severity Score (ISS), payer source, and hospital cost were compared. Forty-nine per cent and 38 per cent of Spanish and English speaking individuals, respectively, were intubated for less than 48 hours (P = 0.072). MOI, HDR, ISS, illicit substance use, alcohol use, and payer source were similar. GCS was statistically higher in the Spanish-speaking group (14 vs 12; P = 0.004). Language and communication barriers lead to potentially preventable intubations in trauma patients.


A novel immune checkpoint-related signature for prognosis and immune analysis in breast cancer.
Breast cancer is one of the most prevailing forms of cancer globally. Immunotherapy has demonstrated efficacy in improving the overall survival of breast cancer. The aim of us was to formulate a novel signature predicated on immune checkpoint-related genes (ICGs) that could anticipate the prognosis and further analyze the immune status of patients with breast cancer. After acquiring data, we pinpointed the definitive ICGs for constructing the prognostic model of breast cancer. We constructed a novel prognostic model and created a fresh risk score called Immune Checkpoint-related Risk Score in breast cancer (ICRSBC). The nomogram was constructed to evaluate the accuracy of the model, and the new web-based tool was created to be more intuitive for predicting prognosis. We also investigated immunotherapy responsiveness and analyzed the tumor mutational burden (TMB) in ICRSBC subgroups. The ICRSBC was found to have significant correlations with the immune environment, immunotherapy responsiveness, and TMB. The expression levels of the 9 ICGs that construct the prognostic model and their promoter methylation levels are significantly different between breast cancer and normal tissues. Furthermore, the mutation profiles, the copy number alterations, and the levels of protein expression also exhibit marked disparities among the 9 ICGs. We have identified and validated a novel signature related to ICGs that is strongly associated with breast cancer progression. This signature enables us to create a risk score for prognosticating the survival and assessing the immune status of individuals affected by breast cancer.


The role of repetition and reinforcement in school-based oral health education-a cluster randomized controlled trial.
Repetition and reinforcement have been shown to play a crucial role in the sustainability of the effect of Oral Health Education (OHE) programs. However, its relevance to school-based OHE imparted by different personnel is not depicted by the existing dental literature. The present study was undertaken to determine the effectiveness of the repeated and reinforced OHE (RR-OHE) compared to one-time OHE intervention and to assess its role in school-based OHE imparted by dentist, teachers and peers. The study was a cluster randomized controlled trial that involved 935 adolescents aged 10-11 years. Twenty four boys' and girls' schools selected at random in two towns of Karachi, Pakistan were randomly assigned to three groups to receive OHE by dentist (DL), teachers (TL) and peer-leaders (PL). The groups received a single OHE session and were evaluated post-intervention and 6 months after. The three groups were then exposed to OHE for 6 months followed by 1 year of no OHE activity. Two further evaluations at 6-month and 12-month intervals were conducted. The data were collected by a self-administered questionnaire preceded by a structured interview and followed by oral examination of participants. The adolescents' oral health knowledge (OHK) in the DL and PL groups increased significantly by a single OHE session compared to their baseline knowledge (p < 0.05) and the increase was sustained over 6 months. Although one-time OHE resulted in a significant improvement in adolescents' oral health behavior (OHB) related to the prevention of gingivitis in the two groups (p < 0.05), no significant change was observed in their behavior towards prevention of oral cancer. One-time teacher-led OHE was ineffective in improving adolescents' OHK and OHB. The oral hygiene status (OHS) of the participants in all three groups did not change statistically after one-time OHE. The OHK, OHB and OHS indices increased significantly 6 months after RR-OHE than the initial scores (p < 0.001) irrespective of OHE strategy. Although the OHK scores of the DL and PL groups decreased significantly at 12-month evaluation of RR-OHE (p < 0.05), the said score of the TL group; and OHB and OHS scores of all three groups remained statistically unchanged during this period. The repetition and reinforcement play a key role in school-based OHE irrespective of educators. The trained teachers and peers can play a complementary role in RR-OHE.


A case report and literature study on Alport syndrome featuring nephrotic syndrome as its primary manifestation.
Historically, due to the lack of distinct clinical symptoms, Alport syndrome, a hereditary kidney disease prevalent in children and a leading cause of kidney failure, has often been misdiagnosed as other kidney conditions. This article presents a comprehensive review and analysis of clinical data concerning a child diagnosed with Alport syndrome, where nephrotic syndrome served as the primary manifestation. The male child in this case exhibited symptoms starting at the age of 6, initially diagnosed as nephrotic syndrome. Consequently, oral steroid medication was administered, proving ineffective. Due to persistent proteinuria and microscopic hematuria, a renal biopsy was performed. Immunofluorescence staining revealed no abnormal expression of the α3, α4, and α5 chains of type IV collagen. Notably, electron microscopy revealed the basement membrane to be partially torn and arachnoid. Genetic testing indicated a hemizygous COL4A5 acceptor-splice-site mutation c.4707-1(IVS50)G > A, inherited from his mother. This specific mutated locus, being the first of its kind reported, adds valuable information to the existing gene mutation spectrum of Alport syndrome. Consequently, it emphasizes the importance for clinicians to deepen their understanding of rare kidney diseases, contributing to enhanced diagnostic accuracy and improved patient care.


Transient expression of hemagglutinin antigen from low pathogenic avian influenza A (H7N7) in Nicotiana benthamiana.
The influenza A virus is of global concern for the poultry industry, especially the H5 and H7 subtypes as they have the potential to become highly pathogenic for poultry. In this study, the hemagglutinin (HA) of a low pathogenic avian influenza virus of the H7N7 subtype isolated from a Swedish mallard Anas platyrhynchos was sequenced, characterized and transiently expressed in Nicotiana benthamiana. Recently, plant expression systems have gained interest as an alternative for the production of vaccine antigens. To examine the possibility of expressing the HA protein in N. benthamiana, a cDNA fragment encoding the HA gene was synthesized de novo, modified with a Kozak sequence, a PR1a signal peptide, a C-terminal hexahistidine (6×His) tag, and an endoplasmic retention signal (SEKDEL). The construct was cloned into a Cowpea mosaic virus (CPMV)-based vector (pEAQ-HT) and the resulting pEAQ-HT-HA plasmid, along with a vector (pJL3:p19) containing the viral gene-silencing suppressor p19 from Tomato bushy stunt virus, was agro-infiltrated into N. benthamiana. The highest gene expression of recombinant plant-produced, uncleaved HA (rHA0), as measured by quantitative real-time PCR was detected at 6 days post infiltration (dpi). Guided by the gene expression profile, rHA0 protein was extracted at 6 dpi and subsequently purified utilizing the 6×His tag and immobilized metal ion adsorption chromatography. The yield was 0.2 g purified protein per kg fresh weight of leaves. Further molecular characterizations showed that the purified rHA0 protein was N-glycosylated and its identity confirmed by liquid chromatography-tandem mass spectrometry. In addition, the purified rHA0 exhibited hemagglutination and hemagglutination inhibition activity indicating that the rHA0 shares structural and functional properties with native HA protein of H7 influenza virus. Our results indicate that rHA0 maintained its native antigenicity and specificity, providing a good source of vaccine antigen to induce immune response in poultry species.


Hemodynamic characteristics of large unruptured internal carotid artery aneurysms prior to rupture: a case control study.
Post-ruptured intracranial aneurysm geometry models have been widely used in computational fluid dynamic studies to assess hemodynamic parameters associated with aneurysm rupture. However, their results may not be valid due to the morphological changes of the aneurysm after rupture. Our aim was to identify the hemodynamic features of aneurysms prior to rupture in comparison with unruptured aneurysms. We retrospectively identified three large unruptured internal carotid artery (ICA) aneurysms (pre-ruptured group) with adequate image quality just before rupture. Matched with the same location and similar size, eight unruptured aneurysms (unruptured group) were selected as controls during the same time period. Flow simulations for these aneurysms were performed to compare differences in hemodynamics. Compared with unruptured aneurysms, pre-ruptured aneurysms had a significantly more irregular aneurysm shape, a higher aspect ratio, and lower aneurysm averaged wall shear stress (WSS) (p=0.024, p=0.048, and p=0.048, respectively). Although pre-ruptured aneurysms had a lower low WSS area and higher Oscillatory Shear Index, these were not statistically significant. For large unruptured ICA aneurysms, low WSS, higher aspect ratio, and irregular shape were indicators of fatal rupture. Early treatment for such lesions with flow diverter and coils may be the best therapeutic option.


Estimating directional epistasis.
Epistasis, i.e., the fact that gene effects depend on the genetic background, is a direct consequence of the complexity of genetic architectures. Despite this, most of the models used in evolutionary and quantitative genetics pay scant attention to genetic interactions. For instance, the traditional decomposition of genetic effects models epistasis as noise around the evolutionarily-relevant additive effects. Such an approach is only valid if it is assumed that there is no general pattern among interactions-a highly speculative scenario. Systematic interactions generate directional epistasis, which has major evolutionary consequences. In spite of its importance, directional epistasis is rarely measured or reported by quantitative geneticists, not only because its relevance is generally ignored, but also due to the lack of simple, operational, and accessible methods for its estimation. This paper describes conceptual and statistical tools that can be used to estimate directional epistasis from various kinds of data, including QTL mapping results, phenotype measurements in mutants, and artificial selection responses. As an illustration, I measured directional epistasis from a real-life example. I then discuss the interpretation of the estimates, showing how they can be used to draw meaningful biological inferences.


Lymphoma diagnosis and plasma Epstein-Barr virus load during vicriviroc therapy: results of the AIDS Clinical Trials Group A5211.
Lack of functional CCR5 increases the severity of certain viral infections, including West Nile virus and tickborne encephalitis. In a phase II trial of the investigational CCR5 antagonist vicriviroc (AIDS Clinical Trials Group protocol A5211), 4 lymphomas occurred in study patients who received vicriviroc. Because of the known association between unregulated Epstein-Barr virus (EBV) replication and lymphoma in immunocompromised patients, we evaluated whether vicriviroc exposure was associated with lymphoma EBV antigen positivity and/or had an effect on plasma levels of EBV DNA. Clinical findings for all 4 patients enrolled in the A5211 study who developed lymphoma (2 Hodgkin and 2 non-Hodgkin) were reviewed, and tumor specimens were assessed for evidence of ongoing EBV replication. Longitudinal plasma samples from 116 patients in the A5211 study were analyzed, and EBV DNA was quantified by real-time polymerase chain reaction. Plasma EBV DNA was not detected in the 2 patients with non-Hodgkin lymphoma; both patients with Hodgkin lymphoma who had samples tested had EBV DNA levels <3200 copies/mL. One patient with Hodgkin lymphoma had a lymph node core biopsy specimen that was strongly positive for EBV; the other 3 lymphomas were histochemically EBV negative. None of the 116 patients with available samples experienced sustained increases in plasma EBV levels. CCR5 antagonism by vicriviroc treatment in treatment-experienced patients was not associated with reactivation of EBV infection.


Isolation, growth, and characterization of a gluconeogenic strain of renal cells.
LLC-PK1 cells, derived from pig kidney, retain several properties of the proximal tubule, but are incapable of gluconeogenesis, due to the lack of fructose-1,6-bisphosphatase (FBPase) [Am. J. Physiol. 248 (Cell Physiol. 17): C181-185, 1985]. Cells incapable of gluconeogenesis require a hexose, pentose, or nucleoside to provide ribose-5-phosphate for RNA biosynthesis. To induce or select cells that express FBPase activity, we cultured LLC-PK1 cells in glucose-free medium. We obtained cells (designated LLC-PK1-FBPase+) that express FBPase activity and are capable of growing in the complete absence of sugars or nucleosides. The cells have apical membrane enzyme activities that differ from those of wildtype cells. Tests of metabolic flow through the gluconeogenic pathway, using 3-mercaptopicolinic acid, a specific inhibitor of phosphoenolpyruvate carboxykinase, confirmed that the cells are gluconeogenic. LLC-PK1-FBPase+ cells grown in medium containing 5 mM glucose for five weekly passages continued to express FBPase activity and apical membrane enzyme activities characteristic of the FBPase+ strain. When switched back to glucose-free medium, they proliferated well. The strain appears to be stable. It should provide a model for studying the relationship between gluconeogenesis and other proximal tubule functions. An incidental finding is that in both strains, the activity of lactate dehydrogenase varied directly with the concentration of glucose in the growth medium, indicating that the expression of lactate dehydrogenase may be regulated by glucose or a metabolite of glucose.


An amino-terminal fragment of the Friend murine leukemia virus envelope glycoprotein binds the ecotropic receptor.
Retrovirus entry into cells is mediated by specific binding of the envelope glycoprotein to a cell membrane receptor. Constitutive envelope gene expression prevents infection by interfering with the binding of viruses which recognize the same receptor. We have used this property to investigate the receptor binding capacities of deleted or truncated murine leukemia virus ecotropic envelope glycoproteins. Friend murine leukemia virus envelope glycoproteins bearing internal amino-terminal deletions, or a soluble 245-amino-acid gp70 amino-terminal fragment, were expressed in NIH 3T3 cells. The susceptibility of these cells to ecotropic and amphotropic virus infection was determined. We observed that both membrane-bound and soluble forms of the gp70 245-amino-acid amino-terminal domain induced resistance to ecotropic virus, indicating that this fragment binds the ecotropic receptor. Binding occurs both at the cell surface and in the endoplasmic reticulum, as shown by the use of soluble envelope fragments either secreted in the culture supernatants or retained in the endoplasmic reticulum lumen by a KDEL sequence. These results suggest that the gp70 amino-terminal domain folds into a structure which recognizes the ecotropic receptor regardless of the carboxy-terminal part of the molecule.


Alkaline ceramidase 2 (ACER2) and its product dihydrosphingosine mediate the cytotoxicity of N-(4-hydroxyphenyl)retinamide in tumor cells.
Increased generation of dihydrosphingosine (DHS), a bioactive sphingolipid, has been implicated in the cytotoxicity of the synthetic retinoid N-(4-hydroxyphenyl)retinamide (4-HPR) in tumor cells. However, how 4-HPR increases DHS remains unclear. Here we demonstrate that 4-HPR increases the expression of ACER2, which catalyzes the hydrolysis of dihydroceramides to generate DHS, and that ACER2 up-regulation plays a key role in mediating the 4-HPR-induced generation of DHS as well as the cytotoxicity of 4-HPR in tumor cells. Treatment with 4-HPR induced the accumulation of dihydroceramides (DHCs) in tumor cells by inhibiting dihydroceramide desaturase (DES) activity, which catalyzes the conversion of DHCs to ceramides. Treatment with 4-HPR also increased ACER2 expression through a retinoic acid receptor-independent and caspase-dependent manner. Overexpression of ACER2 augmented the 4-HPR-induced generation of DHS as well as 4-HPR cytotoxicity, and 4-HPR-induced death in tumor cells, whereas knocking down ACER2 had the opposite effects. ACER2 overexpression, along with treatment with GT11, another DES inhibitor, markedly increased cellular DHS, leading to tumor cell death, whereas ACER2 overexpression or GT11 treatment alone failed to do so, suggesting that both ACER2 up-regulation and DES inhibition are necessary and sufficient to mediate 4-HPR-induced DHS accumulation, cytotoxicity, and death in tumor cells. Taken together, these results suggest that up-regulation of the ACER2/DHS pathway mediates the cytotoxicity of 4-HPR in tumor cells and that up-regulating or activating ACER2 may improve the anti-cancer activity of 4-HRR and other DHC-inducing agents.


Ena/VASP proteins enhance actin polymerization in the presence of barbed end capping proteins.
Ena/VASP proteins influence the organization of actin filament networks within lamellipodia and filopodia of migrating cells and in actin comet tails. The molecular mechanisms by which Ena/VASP proteins control actin dynamics are unknown. We investigated how Ena/VASP proteins regulate actin polymerization at actin filament barbed ends in vitro in the presence and absence of barbed end capping proteins. Recombinant His-tagged VASP increased the rate of actin polymerization in the presence of the barbed end cappers, heterodimeric capping protein (CP), CapG, and gelsolin-actin complex. Profilin enhanced the ability of VASP to protect barbed ends from capping by CP, and this required interactions of profilin with G-actin and VASP. The VASP EVH2 domain was sufficient to protect barbed ends from capping, and the F-actin and G-actin binding motifs within EVH2 were required. Phosphorylation by protein kinase A at sites within the VASP EVH2 domain regulated anti-capping and F-actin bundling by VASP. We propose that Ena/VASP proteins associate at or near actin filament barbed ends, promote actin assembly, and restrict the access of barbed end capping proteins.


The clinical implications of molecular monitoring and analyses of inherited retinal diseases.
Retinal dystrophies (RDs) are the most common cause of inherited blindness and one of the most genetically heterogeneous human diseases. RDs arise from mutations in genes involved in development and function of photoreceptors or other retinal cells. Identification of the genetic defect causing RD allows accurate diagnosis, prognosis, and counseling in affected patients. Molecular diagnosis is a tremendous challenge in RDs due to their locus and phenotypic heterogeneity. As conventional DNA sequencing approaches are impractical in such situation, Next Generation Sequencing (NGS)-based protocols are needed to identify RD-causing mutations. This is being accomplished by sequencing RD gene panels or by whole exome or whole genome sequencing approaches. Areas covered: This review discusses the current strategies for molecular diagnosis in RDs including their advantages and limitations, as well as their utility in diagnosis of non-syndromic versus syndromic RDs. Results of ongoing gene therapy protocols in RDs are also presented. Expert commentary: Molecular diagnosis in RD improves the medical management of patients. Importantly, demand for molecular screening for RDs is greatly expanding not only as a result of increasing development and availability of NGS technologies, but also of the growing number of gene-based clinical trials offering a potential treatment to patients.


Responding to the deaf in disasters: establishing the need for systematic training for state-level emergency management agencies and community organizations.
Deaf and hard-of-hearing (Deaf/HH) individuals have been underserved before and during emergencies. This paper will assess Deaf/HH related emergency preparedness training needs for state emergency management agencies and deaf-serving community-based organizations (CBOs). Four approaches were used: 1) a literature review; 2) results from 50 key informant (KI) interviews from state and territorial-level emergency management and public health agencies; 3) results from 14 KI interviews with deaf-serving CBOs in the San Francisco Bay Area; and 4) a pilot program evaluation of an emergency responder training serving the Deaf/HH in one urban community. Results from literature review and state and territorial level KIs indicate that there is a substantive gap in emergency preparedness training on serving Deaf/HH provided by state agencies. In addition, local KI interviews with 14 deaf-serving CBOs found gaps in training within deaf-serving CBOs. These gaps have implications for preparing for and responding to all-hazards emergencies including weather-related or earthquake-related natural disasters, terrorist attacks, and nuclear-chemical disasters. Emergency preparedness trainings specific to responding to or promoting preparedness of the Deaf/HH is rare, even for state agency personnel, and frequently lack standardization, evaluation, or institutionalization in emergency management infrastructure. This has significant policy and research implications. Similarly, CBOs are not adequately trained to serve the needs of their constituents.


Glossy15, an APETALA2-like gene from maize that regulates leaf epidermal cell identity.
Vegetative development in many plants progresses through distinct juvenile and adult phases. In maize, the transition from juvenile to adult shoot development affects a variety of leaf epidermal cell traits. These include epicuticular waxes, leaf hairs, and cell wall characteristics. Previous genetic and phenotypic analyses have shown that the maize Glossy15 (Gl15) gene is required for the expression of juvenile epidermal traits after leaf 2. We report here the molecular cloning of the Gl15 gene using a defective Suppressor-Mutator (dSpm) element insertion as a transposon-tag. Consistent with the gl15 mutant phenotype, the pattern of Gl15 mRNA expression was correlated with a juvenile leaf epidermal cell identity and was regulated by upstream factors such as Corngrass1. The Gl15 gene encodes a putative transcription factor with significant sequence similarity to the Arabidopsis regulatory genes APETALA2 and AINTEGUMENTA, which act primarily to regulate floral organ identity and ovule development. This finding expands the known functions of APETALA2-related genes to include the control of both vegetative and reproductive lateral organ identity and provides molecular support for the hypothesis that leaves and floral organs are related structures derived from a common growth plan.


[Epstein-Barr virus (EBV) and Kaposi's sarcoma-associated herpesvirus (KSHV, HHV-8)].
Epstein-Barr virus (EBV) and Kaposi's sarcoma-associated herpesvirus (KSHV or human herpesvirus 8, HHV-8) are members of gamma-herpes virus family. Both viruses infect to B cells and cause malignancies such as lymphoma. Since EBV and HHV-8 are so-called 'oncovirus', their oncogenecities have been focused in the researches on EBV and KSHV for a long time. EBV was discovered in 1964, whereas KSHV was identified in 1994. However, KSHV was analyzed rapidly in these fifteen years. One of the recent progresses in the research on EBV and KSHV is that virus-encoded small RNAs were identified in their genomes and characterized. EBV is the first human virus in whose genome microRNA was identified. The oncogenecity of EBV and KSHV remains unclear. Here, I discuss the pathogenesis by EBV and KSHV with special reference to recent progress in this field.


RBF neural network center selection based on Fisher ratio class separability measure.
For classification applications, the role of hidden layer neurons of a radial basis function (RBF) neural network can be interpreted as a function which maps input patterns from a nonlinear separable space to a linear separable space. In the new space, the responses of the hidden layer neurons form new feature vectors. The discriminative power is then determined by RBF centers. In the present study, we propose to choose RBF centers based on Fisher ratio class separability measure with the objective of achieving maximum discriminative power. We implement this idea using a multistep procedure that combines Fisher ratio, an orthogonal transform, and a forward selection search method. Our motivation of employing the orthogonal transform is to decouple the correlations among the responses of the hidden layer neurons so that the class separability provided by individual RBF neurons can be evaluated independently. The strengths of our method are double fold. First, our method selects a parsimonious network architecture. Second, this method selects centers that provide large class separation.


Anopheles gambiae on remote islands in the Indian Ocean: origins and prospects for malaria elimination by genetic modification of extant populations.
The mosquito Anopheles gambiae s.s. is a primary malaria vector throughout sub-Saharan Africa including the islands of the Comoros archipelago (Anjouan, Grande Comore, Mayotte and Mohéli). These islands are located at the northern end of the Mozambique Channel in eastern Africa. Previous studies have shown a relatively high degree of genetic isolation between the Comoros islands and mainland populations of A. gambiae, but the origin of the island populations remains unclear. Here, we analyzed phylogenetic relationships among island and mainland populations using complete mitochondrial genome sequences of individual A. gambiae specimens. This work augments earlier studies based on analysis of the nuclear genome. We investigated the source population of A. gambiae for each island, estimated the number of introductions, when they occurred and explored evidence for contemporary gene flow between island and mainland populations. These studies are relevant to understanding historical patterns in the dispersal of this important malaria vector and provide information critical to assessing their potential for the exploration of genetic-based vector control methods to eliminate this disease. Phylogenetic analysis and haplotype networks were constructed from mitogenome sequences of 258 A. gambiae from the four islands. In addition, 112 individuals from seven countries across sub-Saharan Africa and Madagascar were included to identify potential source populations. Our results suggest that introduction events of A. gambiae into the Comoros archipelago were rare and recent events and support earlier claims that gene flow between the mainland and these islands is limited. This study is concordant with earlier work suggesting the suitability of these oceanic islands as appropriate sites for conducting field trial releases of genetically engineered mosquitoes (GEMs).


Treatment of posterolateral tibial plateau fractures through fibular osteotomy approach.
The best approach for treating posterolateral tibial plateau fractures remains controversial. The aim of the present study was to evaluate the outcomes of patients with posterolateral tibial plateau fractures after open reduction and internal fixation with the fibular osteotomy approach. Between January 2009 and July 2012, twelve patients with posterolateral tibial plateau fracture were treated using the fibular osteotomy approach with a proximal tibia locking compression plate. The epidemiological data, operation details, and clinical outcomes over 25.6 ± 2.1 months (range, 22-38 months) of follow-up were prospectively collected and analyzed. The average surgical duration was 92.5 ± 14.5 min (range, 83-107 min). An anatomical reduction rate of 91.7% (11/12) was observed, although one patient with a lateral comminuted fracture and dislocation had a 2-mm joint surface depression postoperatively. The average fracture healing time was 11.6 ± 3.1 weeks (range, 8-15 weeks), with an average hospital for special surgery knee score of 94.1 ± 3.2 points (range, 80-100 points), an average knee flexion of 118.6° ± 18.1° (range, 96°-138°) and an average knee extension of 1.9° ± 2.2° (range, 0°-6.3°). No complications were found. The fibular osteotomy approach with a proximal tibial compression plate can produce a satisfactory clinical outcome for patients with posterolateral tibial plateau fractures.


A Positively Charged Surface Triggers Coagulation Activation Through Factor VII Activating Protease (FSAP).
Contact between biomedical materials and blood often initiates undesirable pro-coagulant and pro-inflammatory processes. On negatively charged materials, blood coagulation is known to be triggered through autoactivation of Factor XII, while activation on cationic surfaces follows a distinct and so far enigmatic mechanism. Because Factor VII activating protease (FSAP) is known to be activated on positively and on negatively charged macromolecules in plasma, we have investigated its interaction with charged biomaterials and its consequences for coagulation. Several activation processes in blood and plasma were characterized after contact with material surfaces with varied charge. FSAP was found to be exclusively activated by the positively charged surfaces polyethylenimine (PEI) and poly-l-lysine (PLL), not by the negatively charged glass or self-assembled monolayer with carboxyl group termination (SAM-COOH), as well as uncharged (Teflon AF) surfaces. Whole blood incubation on PEI showed that this activation was concomitant with coagulation as determined by thrombin and fibrin formation, which was high for glass (F1+2, 138 nM) and PEI (F1+2, 44 nM) but low for Teflon AF (F1+2, 3.3 nM) and SAM COOH (F1+2, 5.8 nM). Contact phase inhibitor diminished coagulation to background levels for all surfaces except PEI (F1+2: ^PEI 43 to 25 nM; glass, 58 to 1.5 nM) indicating that coagulation activation is not dependent on FXII activation on the PEI surface. A decisive role of endogenous FSAP for coagulation however was confirmed with the use of FSAP inhibitory antibodies which showed no influence on Teflon AF, glass and SAM COOH but diminished F1+2 on PEI to less than 50%. We propose that FSAP activation could be a novel mechanism of surface-driven coagulation. An inhibition of this protease might improve hemocompatibility of cationic surfaces and therefore facilitate the application of polycationic surfaces in blood.


Simple hysterectomy in the presence of invasive cervical cancer in Iran.
The objective of this study was to determine the reasons leading to an inappropriate simple hysterectomy in the presence of invasive cervical cancer. During 1997-2001, 62 cases of invasive cervical carcinoma that had been treated by simple hysterectomy were referred to the gynecology oncology service in Vali-e-Asr hospital, Tehran, Iran. Five had microinvasive carcinoma. The remaining 57 women had either adenocarcinomas or squamous cell carcinoma (SCC). Medical records were reviewed retrospectively to determine the reasons for inappropriate hysterectomy. Reasons for inappropriate hysterectomy were as follow: lack of preoperative Pap smear (P/S) (29%), deliberate hysterectomy for biopsy-proven cancer (25.8%), negative P/S (6.5%), inadequate evaluation of abnormal P/S (6.5%), failure to perform an indicated conization (3.2%), and emergent operation because of uterine perforation (1.6%). Reasons for inappropriate hysterectomy in the remainder of patients (27.4%) were not found because of lack of sufficient information. Although 45.2% of these patients had complained for vaginal bleeding, only four of them had preoperative endocervical and endometrial sampling. Despite the increasing emphasis on performing cervical cancer screening before hysterectomy, only 18 (29%) referral patients had preoperative P/S. We conclude that by close adherence to the cervical cancer screening guidelines and appropriate evaluation of the presenting symptoms, we may avoid inappropriate management of cervical carcinomas with simple hysterectomy. Many cases of simple hysterectomy in the presence of biopsy-proven squamous cell carcinoma of the cervix necessitates some reconsideration of gynecology oncology postgraduate courses for general gynecologists.


Concussion occurrence and recognition in professional boxing and MMA matches: toward a concussion protocol in combat sports.
Objectives: Determine, through video reviews, how often concussions occur in combat sport matches, what influence they have on the outcome, and how well non-physician personnel can be trained to recognize concussions.Methods: This is a retrospective video analysis by an 8-person panel of 60 professional fights (30 boxing and 30 mixed martial arts). Through video review, physician and non-physician personnel recorded details about each probable concussion and determined if and when they would have stopped the fight compared to the official stoppage time.Results: A concussion was recorded in 47/60 fights. The mean number of concussions per minute of fight time was 0.061 (0.047 for boxers and 0.085 for MMA). When stratifying by outcome of the bout, the mean number of concussions per minute for the winner was 0.010 compared to the loser at 0.111 concussions per minute. The fighter that sustained the first concussion ultimately lost 98% of the time. The physician and non-physician raters had high agreement regarding the number of concussions that occurred to each fighter per match. The physician raters judged that 24 of the 60 fights (11 boxing [37%]; 13 MMA [43%]) should have been stopped sooner than what occurred.Conclusion: Recognizing that the concussions often occur in combat sport matches, that the losing fighter almost always is concussed first and tends to sustain more concussions during the fight, along with the demonstration that non-physician personnel can be taught to recognize concussion, may guide policy changes that improve brain health in combat sports.


Inactivation of avirulent Yersinia pestis in beef bologna by gamma irradiation.
Yersinia pestis, a psychrotrophic pathogen capable of growth at refrigeration temperatures, can cause pharyngeal and gastrointestinal plague in humans that consume contaminated foods. Because Y. pestis is listed as a select agent for food safety and defense, evaluation of food safety intervention technologies for inactivation of this pathogen is needed. Ionizing (gamma) radiation is a safe and effective intervention technology that can inactivate pathogens in raw and processed meats, produce, and seafood. In this study, we investigated the effect of temperature on the ability of ionizing radiation to inactivate avirulent Y. pestis in beef bologna. The mean (±standard error of the mean) radiation D(10)-values (the radiation dose needed to inactivate 1 log unit or 90% of the population of a microorganism) for avirulent Y. pestis suspended in beef bologna samples were 0.20 (±0.01), 0.22 (±0.01), 0.25 (±0.02), 0.31 (±0.01), 0.35 (±0.01), and 0.37 (±0.01) kGy at temperatures of 5, 0, -5, -10, -15, and -20°C, respectively. When incorporated into a three-dimensional mesh, the predictive model followed a parabolic fit (R(2) = 0.84), where the log reduction = -0.264 - (0.039 × temp) - (3.833 × dose) - (0.0013 × temp(2)) - (0.728 × dose(2)). These results indicate that ionizing radiation would be an effective technology for control of Y. pestis in ready-to-eat fine emulsion sausage products.


Gun-carrying restrictions and gun-related mortality, Colombia: a difference-in-difference design with fixed effects.
To assess the effect of a permanent gun-carrying restriction on gun-related mortality in Colombia between 2008 and 2014, and determine differences in the effect of the restriction by place of death and sex. In 2012, Bogotá and Medellín introduced a permanent gun-carrying restriction. We compared gun-related mortality rates in these cities (intervention cities) with the rates in all other Colombian cities with more than 500 000 inhabitants (control cities). We used data from the Colombian National Department of Statistics to calculate monthly gun-related mortality rates between 2008 and 2014 for intervention and control cities. We used a differences-in-differences method with fixed effects to assess differences in gun-related mortality in intervention and control cities before and after the introduction of the gun-carrying restriction. We stratified effects by place of death (public area or residence) and sex. We made robustness checks to test the assumptions of the models. Gun-related deaths in the control and intervention cities decreased between 2008 and 2014; however, the decrease was greater in the intervention cities (from 20.29 to 14.93 per 100 000 population; 26.4%) than in the control cities (from 37.88 to 34.56 per 100 000 population; 8.8%). The restriction led to a 22.3% reduction in the monthly gun-related mortality rate in Bogotá and Medellín. The reduction was greater in public areas and for males. Robustness checks supported the assumptions of the models. The permanent restriction on carrying guns reduced gun-related deaths. This policy could be used to reduce gun-related injuries in urban centres of other countries with large numbers of gun-related deaths.


Reactivity studies of the tyrosyl radical in ribonucleotide reductase from Mycobacterium tuberculosis and Arabidopsis thaliana--comparison with Escherichia coli and mouse.
Ribonucleotide reductase (RNR) is a key enzyme for DNA synthesis since it provides cells with deoxyribonucleotides, the DNA precursors. Class I alpha2beta2 RNRs contain a dinuclear iron center and an essential tyrosyl radical in the beta2 component (protein R2). This is also true for the purified protein R2 of Mycobacterium tuberculosis RNR, as shown by iron analysis, light absorption and EPR spectroscopy. EPR spectroscopy at 286 GHz revealed a high g(x) value, suggesting that the radical is not hydrogen bonded, as in other prokaryotic R2s and in contrast with eukaryotic R2s (from Arabidopsis thaliana and mouse). Furthermore, it proved to be very resistant to scavenging by a variety of phenols and thiols and by hydroxyurea, similar to the Escherichia coli radical. By comparison, the plant and mouse radicals are very sensitive to drugs such as resveratrol and 2-thiophenthiol. The radical from M. tuberculosis RNR does not seem to be an appropriate target for new antituberculous agents.


Combination of serum histidine and plasma tryptophan as a potential biomarker to detect clear cell renal cell carcinoma.
In previous work, we showed that serum-free amino acid (SFAA) profiles were different between kidney cancer patients and age and sex matched controls. The goals of the current study are to: (1) confirm our initial observation on an independent sample set; (2) examine if there were similar differences in plasma-free amino acids (PFAA); and (3) determine if removal of tumors changed SFAA and PFAA profiles. SFAA and PFAA profiles were measured in 484 samples taken from 124 healthy controls and 56 clear cell renal cell carcinoma (ccRCC) patients both before and after resection of renal tumors. SFAA and PFAA profiles taken from identical blood samples were remarkably different, with the mean individual amino acid concentrations being 40% less in plasma compared to serum. Both SFAA and PFAA profiles differed significantly between ccRCC patients and controls, but the individual amino acids that differed the most, and the direction of the changes, were quite different between the two blood components. Removal of the tumor had almost no effect on either the SFAA or PFAA profiles. A logistic regression model using serum histidine and plasma tryptophan correctly classified 85.5% of control and 84.7% of case samples. Our findings show that that tumor mass is not directly linked to alterations in blood amino acid levels, and that a combination of serum histidine and plasma tryptophan may be useful as a biomarker to detect ccRCC.


[Experience of the Baby Friendly Hospital initiative].
In the study is analyzed and described the initiative called "Initiative Baby Friendly Hospitals", a program which started in Brazil, 1992. This initiative intends to support, to protect and to promote the breastfeeding as proposed in a meeting in 1990 in Florence, Italy, which was promoted by WHO and UNICEF. The basic goal of this initiative is to mobilize health professionals and hospital or maternity workers for changing their routines and conducts aiming to prevent the early wean. The health establishments are evaluated based on the "ten steps for success of breastfeeding, a group of goals created in the same meeting. In Brazil, the evaluation is coordinated by the Federal Government through the PNIAM (Programa Nacional de Incentivo ao Aleitamento Materno). A baby friendly hospital, if approved, receives from the Minister of Health, a Federal Governmental Agency (SUS) a differential payment for childbirth assistance and prenatal accompaniment, 10% and 40%, more respectively. Until 1998 year there were 103 baby friendly hospitals in Brazil, with the majority of them located in the northeast area (68.1%). However, taking in accounting the number of 5650 hospitals linked to SUS in the country, less than 2.0% are baby friendly hospitals. On the basis of the experience and according with PNIAM data the implementation of the ten steps and the incentive to breastfeeding through baby friendly hospitals have resulted in a significant increase of breastfeeding incidence and duration in Brazil.


Characterisation of hepatitis B virus X protein mutants in tumour and non-tumour liver cells using laser capture microdissection.
The analysis of hepatitis B virus (HBV) X protein genetic variability and is correlation with liver disease severity have only been addressed, so far, on whole liver extracts. We have studied, therefore, the HBV X protein (HBx) gene sequence in morphologically well-characterised tumour and non-tumour liver cells from patients with HBV-related hepatocellular carcinoma. Using laser capture microdissection (LCM), we picked up six to eight groups of tumour and non-tumour hepatocytes in serial frozen sections from six patients. After global DNA preamplification followed by HBx-specific polymerase chain reaction, the HBx gene was sequenced in each group of microdissected cells. We also validated the quantification of HBV-DNA in microdissected hepatocytes using HBV Amplicor. Heterogeneous mutations in HBx gene were found in distinct cirrhotic nodules and tumour areas from the same patient. Mutations at aa 127, 130 and 131 were frequently detected but there was no distinct point mutation profile between tumour and non-tumour samples. In contrast, deletions in HBx gene, which were found in five/six patients, were more frequent in tumour-derived sequences (6/18) than in non-tumour-derived sequences (1/20). We have shown that LCM provides a direct insight of intrahepatic HBV infection. Using this technique, we demonstrated the persistence of distinct HBx encoding sequences in clonally expanding cells, thus supporting the hypothesis that HBx deletions may be implicated in liver carcinogenesis.


Medial branch neurotomy in low back pain.
This study aimed to assess the effectiveness of pulsed radiofrequency medial branch dorsal ramus neurotomy in patients with facet joint syndrome. From January 2008 to April 2010, 92 patients with facet joint syndrome diagnosed by strict inclusion criteria and controlled diagnostic blocks undergone medial branch neurotomy. We did not exclude patients with failed back surgery syndrome (FBSS). Electrodes (20G) with 5-mm active tip were placed under fluoroscopy guide parallel to medial branch. Patients were followed up by physical examination and by Visual Analog Scale and Oswestry Disability Index at 1, 6, and 12 months. In all cases, pain improvement was statistically significant and so quality of life. Three non-FBSS patients had to undergo a second neurotomy because of non-satisfactory pain decrease. Complications were reported in no case. Medial branch radiofrequency neurotomy has confirmed its well-established effectiveness in pain and quality of life improvement as long as strict inclusion criteria be fulfilled and nerve ablation be accomplished by parallel electrode positioning. This statement can be extended also to FBSS patients.


Teamwork as an essential component of high-reliability organizations.
Organizations are increasingly becoming dynamic and unstable. This evolution has given rise to greater reliance on teams and increased complexity in terms of team composition, skills required, and degree of risk involved. High-reliability organizations (HROs) are those that exist in such hazardous environments where the consequences of errors are high, but the occurrence of error is extremely low. In this article, we argue that teamwork is an essential component of achieving high reliability particularly in health care organizations. We describe the fundamental characteristics of teams, review strategies in team training, demonstrate the criticality of teamwork in HROs and finally, identify specific challenges the health care community must address to improve teamwork and enhance reliability.