Evaluation of biological protein-based collagen scaffolds in cartilage and musculoskeletal tissue engineering--a systematic review of the literature.
The term tissue engineering is the technology that combines cells, engineering and biological/synthetic material in order to repair, replace or regenerate biological tissues such as bone, muscle, tendons and cartilage. The major human applications of tissue engineering are: skin, bone, cartilage, corneas, blood vessels, left mainstem bronchus and urinary structures. In this systematic review several criteria were identified as the most desirable characteristics of an ideal scaffold. These state that an ideal scaffolds needs to be biodegradable, possess mechanical strength, be highly porous, biocompatible, non-cytotoxic, non antigentic, stuitable for cell attachment, proliferation and differentiation, flexible and elastic, three dimensional, osteoconductive and support the transport of nutrients and metabolic waste. Subsequently, studies reporting on the various advantages and disadvantages of using collagen based scaffolds in musculoskeletal and cartilage tissue engineering were identified. The purpose of this review is to 1) provide a list of ideal characteristics of a scaffold as identified in the literature 2) identify different types of biological protein-based collagen scaffolds used in musculoskeletal and cartilage tissue engineering 3) assess how many of the criteria each scaffold type meets 4) weigh different scaffolds against each other according to their relative properties and shortcomings. The rationale behind this approach is that the ideal scaffold material has not yet been identified. Hence, this review will define how many of the identified ideal characteristics are fulfilled by natural collagen-based scaffolds and address the shortcomings of its use as found in the literature.


Forage choice in pasturelands: Influence on cattle foraging behavior and performance.
We determined if combinations of adjacent pastures of 3 forage species led to complementary relationships that influenced animal behavior and performance over monocultures. Grazing bouts, behavioral levels of activity, blood urea N (BUN), chemical composition of feces, BW, and herbage biomass before and after grazing were monitored when beef calves strip-grazed 3 replications of 4 treatments from June 14 through August 23, 2013 (9 animals/treatment). Animals grazed monocultures of: 1) tall fescue (TF), 2) alfalfa (ALF), 3) sainfoin (SAN), or 4) a choice of strips of forages TF, ALF, and SAN (CHOICE). The lowest and greatest incidence of foraging bouts occurred for cattle in CHOICE and SAN, respectively (P < 0.01). Animals in CHOICE grazed SAN > ALF > TF (P < 0.01). Animals on TF and CHOICE took greater number of steps than animals grazing a monocultures of either legume (P = 0.01). Calves in TF had lower BUN (P < 0.01) and fecal CP concentration (P < 0.01) than calves grazing the remaining treatments, whereas animals in SAN showed the greatest concentrations of fecal CP (P < 0.01). Fecal NDF concentration was the greatest for animals grazing TF and the lowest for animals grazing SAN (P < 0.01), whereas fecal ADF concentration was greater for animals grazing TF and SAN than for animals grazing CHOICE and ALF (P = 0.02). Calcium, Mg, and Zn concentrations were the lowest in feces from calves grazing TF and the greatest for calves grazing a monoculture of either legume (P < 0.05). When averaging both periods, animals grazing SAN, ALF, or CHOICE gained more BW than animals grazing TF (P < 0.01). Thus, calves in CHOICE incorporated tall fescue into their diets, were more active, and displayed a lower number of grazing bouts than calves grazing monoculture of either legume. Herbage diversity may lead to levels of ADG comparable to legume monocultures with the potential benefit of maintaining plant species diversity in pasturelands.


HaplotypeCN: copy number haplotype inference with Hidden Markov Model and localized haplotype clustering.
Copy number variation (CNV) has been reported to be associated with disease and various cancers. Hence, identifying the accurate position and the type of CNV is currently a critical issue. There are many tools targeting on detecting CNV regions, constructing haplotype phases on CNV regions, or estimating the numerical copy numbers. However, none of them can do all of the three tasks at the same time. This paper presents a method based on Hidden Markov Model to detect parent specific copy number change on both chromosomes with signals from SNP arrays. A haplotype tree is constructed with dynamic branch merging to model the transition of the copy number status of the two alleles assessed at each SNP locus. The emission models are constructed for the genotypes formed with the two haplotypes. The proposed method can provide the segmentation points of the CNV regions as well as the haplotype phasing for the allelic status on each chromosome. The estimated copy numbers are provided as fractional numbers, which can accommodate the somatic mutation in cancer specimens that usually consist of heterogeneous cell populations. The algorithm is evaluated on simulated data and the previously published regions of CNV of the 270 HapMap individuals. The results were compared with five popular methods: PennCNV, genoCN, COKGEN, QuantiSNP and cnvHap. The application on oral cancer samples demonstrates how the proposed method can facilitate clinical association studies. The proposed algorithm exhibits comparable sensitivity of the CNV regions to the best algorithm in our genome-wide study and demonstrates the highest detection rate in SNP dense regions. In addition, we provide better haplotype phasing accuracy than similar approaches. The clinical association carried out with our fractional estimate of copy numbers in the cancer samples provides better detection power than that with integer copy number states.


Molecular evidence for multiple introductions of garlic mustard (Alliaria petiolata, Brassicaceae) to North America.
Invasive species offer excellent model systems for studying rapid evolutionary change. In this context, molecular markers play an important role because they provide information about pathways of introduction, the amount of genetic variation introduced, and the extent to which founder effects and inbreeding after population bottlenecks may have contributed to evolutionary change. Here, we studied microsatellite variation in eight polymorphic loci among and within 27 native and 26 introduced populations of garlic mustard (Alliaria petiolata), a European herb which is a current serious invader in North American deciduous forests. Overall, introduced populations were genetically less diverse. However, considerable variability was present and when compared to the probable source regions, no bottleneck was evident. Observed heterozygosity was very low and resulted in high inbreeding coefficients, which did not differ significantly between native and introduced populations. Thus, selfing seems to be equally dominant in both ranges. Consequently, there was strong population differentiation in the native (F(ST) = 0.704) and the introduced (F(ST) = 0.789) ranges. The high allelic diversity in the introduced range strongly suggests multiple introductions of Alliaria petiolata to North America. Out of six European regions, the British Isles, northern Europe, and central Europe had significantly higher proportions of alleles, which are common to the introduced range, and are therefore the most probable source regions. The genetic diversity established by multiple introductions, and the lack of inbreeding depression in this highly selfing species, may have contributed to the invasion success of Alliaria petiolata.


Addition of Rye Bran and Pea Fiber to Pork Meatballs Enhances Subjective Satiety in Healthy Men, but Does Not Change Glycemic or Hormonal Responses: A Randomized Crossover Meal Test Study.
Background: The development of high-protein, fiber-rich foods targeting appetite control could be an efficient tool in obesity prevention.Objectives: We investigated whether ad libitum energy intake (EI), appetite, and metabolic markers in a meal context were affected by 1) fiber addition (rye bran and pea fiber) to pork meatballs, 2) the food matrix of the fiber (fiber meatballs compared with fiber bread), or 3) the protein source (animal compared with vegetable protein patties).Methods: In a crossover design, 40 healthy men [mean ± SD: body mass index (BMI; in kg/m2), 22.2 ± 1.9; age, 23.3 ± 2.9 y] consumed 4 test meals: a low-fiber meal consisting of pork meatballs plus wheat bread (LF meal); pork meatballs plus fiber bread; fiber meatballs plus wheat bread, and vegetable patties with a natural fiber content plus wheat bread (∼3000 kJ; protein ∼18% of energy, carbohydrate ∼50% of energy, fat ∼30% of energy; 13 g fiber in the fiber meals). Ad libitum EI after 4 h was the primary endpoint. Moreover, appetite sensations and postprandial responses of glucose, insulin, glucagon-like peptide-1, peptide YY 3-36, and plasma amino acids were measured.Results: Ad libitum EI did not differ significantly between the meals. Satiety and fullness increased 11% and 13%, respectively, and hunger and prospective intake decreased 17% and 15%, respectively, after the meal of fiber meatballs plus wheat bread compared with the LF meal (P < 0.01). Hormonal and metabolic responses did not differ between the meals. In general, plasma amino acid concentrations were higher after the fiber-rich meals than after the LF meal.Conclusions: Meals based on meatballs and bread with differences in the fiber content, food matrix of fiber, and protein source had similar effects on ad libitum EI in healthy men. However, fiber addition to pork meatballs favorably affected appetite sensations but without changes in hormonal and metabolic responses. Moreover, animal- and vegetable-protein-based, fiber-matched meals had similar effects on appetite regulation. This trial was registered at clinicaltrials.gov as NCT02521805.


Distribution function of transit times in the human pulmonary circulation.
The distribution function of pulmonary transit times (fPTT) defines contact time between blood and vascular bed, which affects gas exchange and endothelial metabolic functions. This study was undertaken to assess effects of abnormal pulmonary inflow (PPA) and outflow pressures (PLA) on fPTT. Three groups were studied: five patients with elevated PLA and passive pulmonary hypertension (LVD-Ab), eight with normal PLA (LVD-Nl), and six with pulmonary disease and various levels of PPA (PD). Empirical complex exponential functions were convoluted on right and left ventricular indicator-dilution curves to derive fPTT; mean transit time (Mo1), standard deviation (Sm2), and cube root of the third moment about Mo1 (Sm3) were calculated by standard equations. A single linear regression of Sm3 and Sm2 on Mo1 was observed for all patients, regardless of disease process. Inverse relations between Mo1, Sm2, and Sm3 and blood flow were highly significant, but dispersion volumes (DV = Mo1 x flow) were higher in patients with elevated PPA. Significant linear regressions of fPTT parameters on PPA, derived in LVD-Nl and LVD-Ab patients, failed to predict Mo1, Sm2, and Sm3 for the PD group, whereas linear regressions on PLA accurately predicted Mo1, Sm2, and Sm3 in the PD group. Relations between fPTT parameters and PLA were equally well fit by exponential equations in all 19 patients, consistent with an asymptotic pressure-volume relation of distensible vessels. Microvascular pressure (PMV), combining PPA and PLA, was not a better predictor of fPTT parameters in LVD-NL and LVD-Ab patients but provided a slightly closer estimate of relative dispersion and skewness in PD patients.(ABSTRACT TRUNCATED AT 250 WORDS)


Antihistamines are superior to topical steroids in managing human immunodeficiency virus (HIV)-associated papular pruritic eruption.
Papular pruritic eruption (PPE) is a frequent HIV-comorbidity in tropical countries. Because of constant itching and social stigma, effective treatment is highly valued. In our HIV cohort in rural Tanzania with 12% prevalence of PPE, we have retrospectively analyzed responses to available treatments. Oral promethazine improved itching (P < 0.0058) and clinical scores (P < 0.032) significantly more than topical steroids. Disease activity did not correlate with CD4+ and CD8+ T cell counts and was independent of anti-retroviral medication. Therefore, oral antihistamines are an effective first-line treatment for PPE.


Retinal nerve fiber layer and macular thinning in systemic lupus erythematosus: an optical coherence tomography study comparing SLE and neuropsychiatric SLE.
Due to the lack of reliable biomarkers in diagnosing and monitoring neuropsychiatric systemic lupus erythematosus (NPSLE), the aim of this study was to examine the utility of measurements obtained through spectral domain optical coherence tomography (SD-OCT) as a biomarker for NP involvement in SLE. Retinal nerve fiber layer (RNFL) and macula scans were performed using SD-OCT on 15 NPSLE patients, 16 SLE patients without NP symptoms (non-NP SLE), and 16 healthy controls. Macular volume and thickness of the central macula and peripapillary RNFL were compared between the groups and to scores on two validated cognitive tests. NPSLE patients did not differ significantly from non-NP SLE patients in retinal thickness or macular volume. However, SLE patients as a whole showed significant RNFL and macular thinning compared to controls. Scores on the Trail Making Test B, a test of complex attention, showed significant correlation to temporal superior and temporal inferior RNFL thickness. Our results demonstrate RNFL thinning in SLE, and confirm the previous finding of high incidence of abnormal brain scans in SLE. These findings suggest that OCT measurements may be indicative of neurodegeneration in SLE and may be a useful biomarker for early cognitive impairment in SLE.


Autophagy Requirements for Eye Lens Differentiation and Transparency.
Recent evidence points to autophagy as an essential cellular requirement for achieving the mature structure, homeostasis, and transparency of the lens. Collective evidence from multiple laboratories using chick, mouse, primate, and human model systems provides evidence that classic autophagy structures, ranging from double-membrane autophagosomes to single-membrane autolysosomes, are found throughout the lens in both undifferentiated lens epithelial cells and maturing lens fiber cells. Recently, key autophagy signaling pathways have been identified to initiate critical steps in the lens differentiation program, including the elimination of organelles to form the core lens organelle-free zone. Other recent studies using ex vivo lens culture demonstrate that the low oxygen environment of the lens drives HIF1a-induced autophagy via upregulation of essential mitophagy components to direct the specific elimination of the mitochondria, endoplasmic reticulum, and Golgi apparatus during lens fiber cell differentiation. Pioneering studies on the structural requirements for the elimination of nuclei during lens differentiation reveal the presence of an entirely novel structure associated with degrading lens nuclei termed the nuclear excisosome. Considerable evidence also indicates that autophagy is a requirement for lens homeostasis, differentiation, and transparency, since the mutation of key autophagy proteins results in human cataract formation.


How does age-related macular degeneration affect real-world visual ability and quality of life? A systematic review.
To review systematically the evidence of age-related macular degeneration (AMD) affecting real-world visual ability and quality of life (QoL). To explore trends in specific topics within this body of the literature. Systematic review. A systematic literature search was carried out using MEDLINE, EMBASE, CINAHL, PsycINFO, PsychARTICLES and Health and Psychosocial Instruments for articles published up to January 2015 for studies including people diagnosed with AMD, assessing real-world visual ability or QoL as an outcome. Two researchers screened studies for eligibility. Details of eligible studies including study design, characteristics of study population and outcomes measured were recorded in a data extraction table. All included studies underwent quality appraisal using the Mixed Methods Appraisal Tool 2011 Version (MMAT). From 5284 studies, 123 were eligible for inclusion. A range of approaches were identified, including performance-based methods, quantitative and qualitative patient-reported outcome measures (PROMs). AMD negatively affects tasks including mobility, face recognition, perception of scenes, computer use, meal preparation, shopping, cleaning, watching TV, reading, driving and, in some cases, self-care. There is evidence for higher rates of depression among people with AMD than among community dwelling elderly. A number of adaptation strategies have been associated with AMD of varying duration. Much of the research fails to report the type of AMD studied (59% of included studies) or the duration of disease in participants (74%). Of those that do report type studied, the breakdown is as follows: wet AMD 20%, dry AMD 4% and both types 17%. There are many publications highlighting the negative effects of AMD in various domains of life. Future research should focus on delivering some of this research knowledge into patient management and clinical trials and differentiating between the types of AMD.


Nursing student attitudes towards older people: validity and reliability of the Italian version of the Kogan Attitudes towards Older People scale.
This paper reports a study testing the validity and reliability of the Italian version of the Kogan Attitude towards Older People scale. Nursing students' attitudes towards older people can affect their preference for working with them. One of the most common instruments used to assess these attitudes is the Kogan Attitude towards Older People scale. Previous validation studies performed on college students internationally have demonstrated good reliability and validity. The psychometric properties of the Italian version have not yet been tested. A cross-sectional, descriptive study. The study was conducted from March-June 2010 with a convenience sample of nursing students attending two Italian universities. Reliability was evaluated using internal consistency and item-to-total correlation. Content and construct validity were evaluated using a content validity index and principal factor analysis. A total of 1637 nursing students participated in the study. Two factors were extracted from the factor analysis: prejudice describing a negative disposition and appreciation indicating a favourable feeling and opinion towards older people. The Kogan Attitude towards Older People scale applied to Italian nursing students exhibited good internal consistency. The two-dimensional construct of the scale was congruent with some, but not all, results of previous studies due to methodological differences among the studies. Based on the study findings, the scale can be recommended for future research studies in Italy including use as pre/post-test measurement associated with nursing education programmes.


Guanine nucleotide-dependent assembly of FtsZ into filaments.
FtsZ is an essential cell division protein that is localized to the leading edge of the bacterial septum in a cytokinetic ring. It contains the tubulin signature motif and is a GTP binding protein with a GTPase activity. Further comparison of FtsZ with eukaryotic tubulins revealed some additional sequence similarities, perhaps indicating a similar GTP binding site. Examination of FtsZ incubated in vitro by electron microscopy revealed a guanine nucleotide-dependent assembly into protein filaments, supporting the hypothesis that the FtsZ ring is formed through self-assembly. FtsZ3, which is unable to bind GTP, does not polymerize, whereas FtsZ2, which binds GTP but is deficient in GTP hydrolysis, is capable of polymerization.


Nano-FTIR absorption spectroscopy of molecular fingerprints at 20 nm spatial resolution.
We demonstrate Fourier transform infrared nanospectroscopy (nano-FTIR) based on a scattering-type scanning near-field optical microscope (s-SNOM) equipped with a coherent-continuum infrared light source. We show that the method can straightforwardly determine the infrared absorption spectrum of organic samples with a spatial resolution of 20 nm, corresponding to a probed volume as small as 10 zeptoliter (10(-20) L). Corroborated by theory, the nano-FTIR absorption spectra correlate well with conventional FTIR absorption spectra, as experimentally demonstrated with poly(methyl methacrylate) (PMMA) samples. Nano-FTIR can thus make use of standard infrared databases of molecular vibrations to identify organic materials in ultrasmall quantities and at ultrahigh spatial resolution. As an application example we demonstrate the identification of a nanoscale PDMS contamination on a PMMA sample.


Early adopters of electronic prescribing struggle to make meaningful use of formulary checks and medication history documentation.
Use of electronic prescribing (e-prescribing) can improve safety and reduce costs of care by alerting prescribers to drug-drug interactions, patient nonadherence to therapies, and insurance coverage information. Deriving these benefits will require clinical decision support based on presentation of accurate and complete formulary and benefit (F&B) and medication history (RxH) data to prescribers, but relatively little is known about how this information is used in primary care. This is a multimethod comparative case study of 8 practices, which were selected to ensure practice size and physician specialty variation, implementing a stand-alone e-prescribing program. Field researchers observed prescription workflow and interviewed physicians and office staff. Before implementation, few prescribers reported using F&B references when making medication choices; all used paper-based methods for tracking medication history. After implementation, some prescribers reported using F&B data to inform medication choices but missing information reduced confidence in these resources. Low confidence in RxH data led to paper-based workarounds. Challenges experienced with formulary checks and RxH documentation led to prescriber distrust and unwillingness to rely on e-prescribing-based information. Greater data accuracy and completeness must be assured if e-prescribing is to meet meaningful use objectives to improve the efficiency and safety of prescribing in primary care settings.


TNFR2 gene polymorphism in coronary artery disease.
Recently atherosclerosis and coronary artery disease (CAD) are considered to be inflammatory diseases. The genetic polymorphism in inflammatory markers has been well studied and found to be associated with development of CAD. To study the association of biallelic polymorphism at position 196 in exon 6 of tumor necrosis factor 2 (TNFR2) gene and coronary artery disease. The study design was a prospective case control study conducted at a tertiary referral center mainly catering to the north Indian population. One hundred and fifty angiographically proven patients with coronary artery disease and one hundred and fifty age matched controls were genotyped for TNFR2 gene by polymerase chain reaction followed by analysis of restriction fragment length polymorphism. Genotype frequencies were compared in patients and controls by Chi-square test. Binary logistic regression analysis was used to examine the relationship between genotypes and disease, incorporating other variables into the model. The incidence of CAD in those with MM genotype was 65% and in those with RM genotype was 42%. Genotype frequency shows significant association of MM genotype with development of CAD (P < 0.001; odds ratio-2.585; 95% confidence interval 1.533-4.359). The association of TNFR2 genotype with CAD persisted on logistic regression analysis. MM genotype of TNFR2 gene is associated with development of CAD and RM genotype appears to be protective.


Frequency distribution of thiopurine S-methyltransferase alleles in a polish population.
Thiopurine S-methyltransferase (TPMT) is an enzyme that catalyzes the S-methylation of thiopurine drugs such as 6-mercaptopurine, 6-thioguanine, and azathioprine. TPMT activity exhibits an interindividual variability mainly a result of genetic polymorphism. Patients with intermediate or deficient TPMT activity are at risk for toxicity after receiving standard doses of thiopurine drugs. It has previously been reported that 3 variant alleles:TPMT*2, *3A, and *3C are responsible for over 95% cases of lower enzyme activity. The purpose of this study was to determine the frequency of TPMT variant alleles in a Polish population. DNA samples were obtained from 358 unrelated healthy Polish subjects of white origin, and TPMT genetic polymorphism was determined using PCR-RFLP and allele-specific PCR methods. The results showed that allelic frequencies were 0.4% for TPMT*2, 2.7% for TPMT*3A, and 0.1% for TPMT*3C, respectively. A TPMT*3B allele was not found in the studied population. The general pattern of TPMT allele disposition in the Polish population is similar to those determined for other white populations, but the frequency of total variant alleles is lower than in other European populations studied to date.


Clonality analysis of intraductal proliferative lesions using the human androgen receptor assay.
Recently, it is accepted that invasive breast carcinoma is of monoclonal origin. Ductal intraepithelial neoplasia (DIN) may progress toward invasive carcinoma with an increased risk. However, it is not fully understood whether DIN is polyclonal or monoclonal. In this current study, we detected clonal origin of DIN using x-inactivation at the human androgen receptor (HUMARA) locus. Lesional and normal breast gland cells were microdissected from paraffin-embedded tissues using a laser capture microdissection system. Genomic DNA was extracted. After digestion by restriction enzyme Hpa II, the HUMARA exon1 was amplified by a fluorescent nested-PCR procedure and the PCR products were separated on DNA sequencer and analyzed the fluorescent intensity of the two HUMARA alleles. DNA from 88 of 101(87%) patients was able to be amplified at the HUMARA locus and 68 of them (77.3%) were heterozygous and informative. 9/12 usual ductal hyperplasia (UDH) and 5/18 DIN 1A showed a polyclonal inactivation. 3/12 UDH, 13/18 DIN 1A, 28/28 DIN 1B, 10/10 carcinoma in situ are of monoclonal origin. Taken together, DIN 1A, 1B and carcinoma in situ, are monoclonal and DIN 1, but not UDH, represents the obligate and direct precursor of DCIS.


The Aspergillus Genome Database: multispecies curation and incorporation of RNA-Seq data to improve structural gene annotations.
The Aspergillus Genome Database (AspGD; http://www.aspgd.org) is a freely available web-based resource that was designed for Aspergillus researchers and is also a valuable source of information for the entire fungal research community. In addition to being a repository and central point of access to genome, transcriptome and polymorphism data, AspGD hosts a comprehensive comparative genomics toolbox that facilitates the exploration of precomputed orthologs among the 20 currently available Aspergillus genomes. AspGD curators perform gene product annotation based on review of the literature for four key Aspergillus species: Aspergillus nidulans, Aspergillus oryzae, Aspergillus fumigatus and Aspergillus niger. We have iteratively improved the structural annotation of Aspergillus genomes through the analysis of publicly available transcription data, mostly expressed sequenced tags, as described in a previous NAR Database article (Arnaud et al. 2012). In this update, we report substantive structural annotation improvements for A. nidulans, A. oryzae and A. fumigatus genomes based on recently available RNA-Seq data. Over 26 000 loci were updated across these species; although those primarily comprise the addition and extension of untranslated regions (UTRs), the new analysis also enabled over 1000 modifications affecting the coding sequence of genes in each target genome.


Exploring the virulence gene interactome with CRISPR/dCas9 in the human malaria parasite.
Mutually exclusive expression of the var multigene family is key to immune evasion and pathogenesis in Plasmodium falciparum, but few factors have been shown to play a direct role. We adapted a CRISPR-based proteomics approach to identify novel factors associated with var genes in their natural chromatin context. Catalytically inactive Cas9 ("dCas9") was targeted to var gene regulatory elements, immunoprecipitated, and analyzed with mass spectrometry. Known and novel factors were enriched including structural proteins, DNA helicases, and chromatin remodelers. Functional characterization of PfISWI, an evolutionarily divergent putative chromatin remodeler enriched at the var gene promoter, revealed a role in transcriptional activation. Proteomics of PfISWI identified several proteins enriched at the var gene promoter such as acetyl-CoA synthetase, a putative MORC protein, and an ApiAP2 transcription factor. These findings validate the CRISPR/dCas9 proteomics method and define a new var gene-associated chromatin complex. This study establishes a tool for targeted chromatin purification of unaltered genomic loci and identifies novel chromatin-associated factors potentially involved in transcriptional control and/or chromatin organization of virulence genes in the human malaria parasite.


Conformational changes of DNA induced by a trans-azobenzene derivative via non-covalent interactions.
In biological environments and in aqueous solution, DNA generally adopts the canonical B conformation. Recently, an azobenzene photoswitch containing a polyamine chain with three positive charges was shown to induce a reversible conformational transition between the A and B forms of DNA, the transition being triggered by trans-cis isomerization of the photoswitch upon non-covalent intercalation. It was proposed that, in its trans conformation, azobenzene stabilizes the A form of DNA. The structural details and the mechanism upon which trans-azobenzene induces the B-to-A DNA transition remain, however, unclear. In the present work, two possible intercalating modes of trans-azobenzene, from the minor groove and from the major groove, were investigated with all-atom molecular-dynamics simulations. Intercalation from the major groove was found to be the most probable binding mode due to favorable electrostatic and π-π stacking interactions. The free-energy profile associated with the B-to-A conformational transition reveals that intercalation from the major groove leads to a conformational change of DNA, showing a slight tendency to interconvert from B- to A-DNA, in agreement with the CD spectrum obtained from the experiment. However, the presence of only one interacting azobenzene is not sufficient to lead to a global conformational change to A-DNA. The present results are expected to serve in the design of DNA switches, which can induce reversible DNA conformational changes.


Smoking Cessation Advisory Intervention in Patients with Cardiovascular Disease.
Several studies have examined the efficacy of smoking cessation therapies in the general population. However little is known about the efficacy of these advisory methods in cardiovascular patients. The aim of the study is to determine the prevalence and the characteristics of smoking abstinence in cardiovascular patients, after a smoking intervention during hospitalization. The study involved 442 patients, smokers admitted for cardiovascular disease to the Department of Cardiology. During hospitalization patient's data were collected and all patients were subjected to a 30-minutes long advisory session with drug administration in selected cases (varenicycline, bupropione, nocitine replacement therapy), according to standard protocol. After the discharge patients were asked about smoking abstinence at time intervals of 24 hours, 1 month, 3, 6 and 12 months. After hospital discharge 11 patients (2.49%) could not be contacted after several attempts and 19 patients (4.3%) were died during follow up period. A total of 412 patients (218 men and 194 women, mean age 56.49+10.57 years) made up the final study population. Twenty four hours after hospital discharge 364 patients (88.35%) had quitted smoking. At 1, 3, 6 and 12 months the abstinence rates were 70.87%, 64.8%, 55.82% and 47.83% respectively. Patients with ischaemic cardiovascular diseases (angina - infarction) had a significantly higher probability of quitting smoking at 12 months (Hazard ratio: 0.64 - p=0.01). A smoking cessation program in cardiovascular patients during hospitalization was unlikely to result in success. These patients might benefit by following programs promoting smoking cessation in experienced specialized centers, involving a group of health professionals, such as psychologists and/or trained nurses.


Development and Risk Factors of Type 2 Diabetes in a Nationwide Population of Women With Polycystic Ovary Syndrome.
Polycystic ovary syndrome (PCOS) is associated with insulin resistance and obesity. Prospective population-based data regarding development and possible predictors of type 2 diabetes (T2D) in PCOS are limited. National Patient Register-based study. Patients with PCOS [PCOS Denmark and embedded cohort, PCOS Odense University Hospital (OUH)] and a control population with no previous diagnosis of T2D. PCOS OUH (N = 1,162) included premenopausal women with PCOS and standardized clinical and biochemical examination. PCOS Denmark (N = 18,477) included women with PCOS in the Danish National Patient Register. Three age-matched controls were included per patient (N = 54,680). T2D events according to diagnosis codes and filled medicine prescriptions. The median (quartiles) follow-up was 11.1 (6.9 to 16.0) years. The hazard ratio (HR) with 95% confidence interval (CI) for development of T2D in PCOS Denmark was HR = 4.0 (95% CI, 3.7 to 4.3; P < 0.001), and the total event rate of T2D was 8.0 per 1000 person years in PCOS Denmark vs 2.0 per 1000 person years in controls (P < 0.001). The median age at diagnosis of T2D was 31 (26 to 37) years in PCOS Denmark vs 35 (27 to 44) years in controls (P < 0.001). In multiple regression analyses, body mass index, glycated hemoglobin, fasting blood glucose, 2-hour blood glucose, homeostasis model assessment of insulin resistance, and triglycerides were positively associated with development of T2D, whereas higher number of births was negatively associated with development of T2D. The event rate of T2D was higher in PCOS compared with controls, and T2D was diagnosed at a younger age.


Response-guided ABVD chemotherapy plus involved-field radiation therapy for intermediate-stage Hodgkin lymphoma in the pre-positron emission tomography era: a Gruppo Italiano Studio Linfomi (GISL) prospective trial.
In the pre-positron emission tomography era, the Gruppo Italiano Studio Linfomi (GISL) investigated the feasibility and efficacy of a treatment based on a response-tailored number of doxorubicin/bleomycin/vinblastine/dacarbazine (ABVD) courses in 218 intermediate-stage Hodgkin lymphoma patients. Patients with stage I/II showing at least one adverse prognostic factor and stage IIIA without adverse prognostic factors were recruited. Treatment included a first step of 3 ABVD courses, followed by an early-restaging. Patients in CR/CRu received 1 additional ABVD cycle, patients in PR received 3 more ABVD, and nonresponder patients went off study. Involved-field radiation therapy (RT) was recommended on chemotherapy completion. The median age was 30 years (range, 15-68 years) and 131 patients (61%) were female. Seven percent of patients were in stage I, 78% in stage II, and 15% in stage III; B-symptoms, bulky tumor and erythrocyte sedimentation rate > 30 were recorded in 20%, 26%, and 43% of cases, respectively. The CR/CRu rate was 62% at early restaging, 72% at the end of chemotherapy, and 95% following RT. With a median follow-up of 74 months (range, 6-193 months), 7-year overall survival, relapse-free survival, and freedom from treatment failure were 91.8% (95% CI, 86%-95.5%), 89.2% (95% CI, 82.8%-93.3%), and 81.8% (95% CI, 75.2%-86.7%), respectively. Patients in CR/CRu on early restaging, receiving 4 ABVD, had an excellent outcome with 7-year RFS and cause-specific survival similar to those of the late responders treated with 6 ABVD (RFS, 87.5% vs. 90.5% and CSS, 96.6% vs. 92.7%, respectively). The response-guided ABVD program we report, based on standard clinical staging procedures, proved to be feasible and safe in patients with intermediate-stage Hodgkin lymphoma.


Laminin polymer treatment accelerates repair of the crushed peripheral nerve in adult rats.
Promoting axon growth after peripheral nerve injury may support recovery. Soluble laminin polymers formed at pH 4 (aLam) accelerate axon growth from adult dorsal root ganglion neurons in vitro. We used an adult rat model of a peripheral (peroneal) nerve crush to investigate whether an injection of aLam enhances axon growth and functional recovery in vivo. Rats that received an injection of aLam into the crush at 2 days post-injury show significant improvements in hind limb motor function at 2 and 5 weeks after injury compared with control rats that received phosphate-buffered saline. Functional improvement was not associated with changes in sensitivity to thermal or mechanical stimuli. Treatment with aLam decreased the occurrence of autophagia and abolished non-compliance with treadmill walking. Rats treated with aLam showed increased axon presence in the crush site at 2 weeks post-injury and larger axon diameter at 10 weeks post-injury compared with controls. Treatment with aLam did not affect Schwann cell presence or axon myelination. Our results demonstrated that aLam accelerates axon growth and maturity in a crushed peroneal nerve associated with expedited hind limb motor function recovery. Our data support the therapeutic potential of injectable aLam polymers for treatment of peripheral nerve crush injuries. STATEMENT OF SIGNIFICANCE: Incidence of peripheral nerve injury has been estimated to be as high as 5% of all cases entering a Level 1 trauma center and the majority of cases are young males. Peripheral nerves have some endogenous repair capabilities, but overall recovery of function remains limited, which typically has devastating effects on the individual, family, and society, as wages are lost and rehabilitation is extended until the nerves can repair. We report here that laminin polymers injected into a crush accelerated repair and recovery, had no adverse effects on sensory function, obliterated non-compliance for walking tests, and decreased the occurrence of autophagia. These data support the use of laminin polymers for safe and effective recovery after peripheral nerve injury.


Reproductive Outcomes After Gestational Trophoblastic Neoplasia. A Comparison Between Single-Agent and Multiagent Chemotherapy: Retrospective Analysis From the MITO-9 Group.
Gestational trophoblastic neoplasia affects women of reproductive age and is usually treated by chemotherapy. Major concerns related to chemotherapy in young women are the possible infertility, risk of early menopause, and teratogenic effects on subsequent pregnancies. The study's aim was to analyze menstrual and reproductive outcomes of women treated with single-agent versus multiagent chemotherapy for gestational trophoblastic neoplasia. One-hundred fifty-one patients were treated. Seventy-six patients older than 45 years, with a placental site or epithelioid trophoblastic tumor, undergoing hysterectomy for patient choice, or undergoing human chorionic gonadotropin follow-up at the time of the analysis were excluded. Seventy-five patients were divided into subgroups according to International Federation of Gynecology and Obstetrics score: patients scoring less than 7, receiving single-agent chemotherapy (group A, n = 42); patients scoring 7 or greater, receiving combination treatment (group B, n = 33). Patients' outcomes were compared by univariate and multivariate analyses. Temporary amenorrhea occurred in 33% of group A patients and 66.7% of group B (P = 0.01). Premature menopause occurred in 3 patients in group B (0% vs 9%, P = 0.02). Ten patients in group B underwent salvage hysterectomy. Pregnancy desire did not differ between the 2 groups (P = 0.555). In group A, 57.1% became pregnant; in group B, 36.4% did (P = 0.060). Instead, pregnancy rate was 52.2% among high-risk patients not undergoing hysterectomy (57.1% vs 52.2%, P = 0.449). There was no difference in miscarriage (P = 0.479) and premature birth (P = 0.615) rates. In a multivariate analysis that included age, International Federation of Gynecology and Obstetrics score, chemotherapy type, use of assisted reproductive technologies, previous pregnancies, and pregnancy desire, only age (P = 0.006) and pregnancy desire (P = 0.002) had a significant impact on the probability to have subsequent pregnancies. Except for the risk of premature ovarian failure, a rare adverse effect of combined treatments, both single-agent and multiagent chemotherapy can be safely administered to patients with a desire for childbearing. High-risk patients have worse reproductive outcomes because they undergo hysterectomy more frequently than low-risk patients.


Human leukocyte antigen A1-B8-DR3-DQ2-DPB1*0401 extended haplotype in autoimmune hepatitis.
Genetic susceptibility to autoimmune hepatitis is associated with the human leukocyte antigen haplotype A1-B8-DR3 and DR4. To date, only one study in Japan has considered the human leukocyte antigen DP locus in this disease, and no studies have been reported in whites. In this study we used a series of sequence-specific oligonucleotide probes to determine human leukocyte antigen DPB1 genotypes in 101 unrelated white northern European patients and 105 racially and geographically matched controls. The aims of the study were twofold: first, to determine the degree of DPB-encoded susceptibility to autoimmune hepatitis, and, second, to establish whether susceptibility can be extended to include human leukocyte antigen DPB. None of 17 DPB1 alleles was significantly associated with the susceptibility to autoimmune hepatitis. Although one particular seven-locus haplotype A1-B8-DRB3*0101-DRB1*0301-DQA1*0501-DQB1*0201-++ +DPB1*0401 was significantly associated with the disease (27% vs. 7%, relative risk = 5.14, p < 0.0005), the association with this haplotype was weaker than that for the six-locus haplotype excluding DPB (40% vs. 11%, RR = 5.52, p < 0.0005). When the patients first seen at ages younger than 16 yr (pediatric patients) were considered separately, the greatest relative risk was for the seven-locus haplotype (41% vs. 7%; relative risk = 9.60, p < 0.0005). The results of this study further confirm that major histocompatibility complex-encoded susceptibility to autoimmune hepatitis is located at or close to the human leukocyte antigen DR locus; however, the A1-B8-DR3-DQ2-DPB1*0401 extended haplotype may be important in determining the age of onset and severity of disease.


[Can Patients with History of Malignancy Become Organ Donors?]
Cancer transmission from solid organ donors to recipients is a known risk factor in transplantation. The Italian National Network for Transplantation (CNT) has adopted specific guidelines to evaluate the suitability of donors with history of malignancy. CNT also provides a Second Opinion service to assess oncological cases with a potential risk of neoplastic transmission to the recipient. CNT aims to minimize the risk of disease transmission from donors to recipients. According to CNT guidelines, "standard" donors are defined as individuals with no signs of active malignancy and no history of cancer at the time of organ procurement. Unsuitable donors, defined as those with an "unacceptable risk", are those patients with evidence of malignancy at the time of donation or in their medical history that carries an unacceptably high risk of disease transmission. Between these two categories, a broad spectrum of "non-standard" donors exists, where the risk of transmission is not entirely absent, but remains low enough to consider organ utilization. Malignancy should not be considered an absolute contraindication for organ donation. CNT has also adopted a specific repository for adverse events (AE) after transplantation. Since 2012, with 10.493 donors and 34.193 performed transplants, 283 AE have been recorded, occurring in approximately 3% of donation processes and 1% of performed transplants. Oncological AE represented 13% of all reports. In the majority of cases, oncological AE resulted from missed diagnosis during organ procurement, benchwork, or transplantation surgery. CNT guidelines, the oncological second opinion service, and the repository helped minimize the risk of cancer transmission with transplantation.


Honokiol-induced apoptosis and autophagy in glioblastoma multiforme cells.
Honokiol, a hydroxylated biphenyl compound isolated from the Chinese herb Magnolia officinalis, has been reported to have anticancer activities in a variety of cancer cell lines. The present study aimed to evaluate the anticancer effect and possible molecular mechanisms of honokiol in a glioblastoma multiforme (GBM) cell line. The anticancer activities of honokiol were investigated in the DBTRG-05MG GBM cell line. The effect of honokiol on cell growth was determined using a sulforhodamine B assay. Flow cytometry and immunoblotting were used to measure honokiol-induced apoptosis (programmed cell death type I) and autophagy (programmed cell death type II). Honokiol was observed to reduce DBTRG-05MG cell viability in a dose-dependent manner. At a dose of 50 μM, honokiol markedly decreased the expression of Rb protein and led to the cleavage of poly(ADP-ribose) polymerase and Bcl-xL to promote apoptosis in the cancer cells. In addition, markers of autophagy, including Beclin-1 and LC3-II, were also significantly increased. In addition to apoptosis, honokiol was also able to induce autophagy in the DBTRG-05MG cells. The mechanisms that are responsible for the correlation between honokiol-induced apoptosis and autophagy require further investigation. Such efforts may provide a potential strategy for improving the clinical outcome of GBM treatment.


Functions of microtubules in the Saccharomyces cerevisiae cell cycle.
We used the inhibitor nocodazole in conjunction with immunofluorescence and electron microscopy to investigate microtubule function in the yeast cell cycle. Under appropriate conditions, this drug produced a rapid and essentially complete disassembly of cytoplasmic and intranuclear microtubules, accompanied by a rapid and essentially complete block of cellular and nuclear division. These effects were similar to, but more profound than, the effects of the related drug methyl benzimidazole carbamate (MBC). In the nocodazole-treated cells, the selection of nonrandom budding sites, the formation of chitin rings and rings of 10-nm filaments at those sites, bud emergence, differential bud enlargement, and apical bud growth appeared to proceed normally, and the intracellular distribution of actin was not detectably perturbed. Thus, the cytoplasmic microtubules are apparently not essential for the establishment of cell polarity and the localization of cell-surface growth. In contrast, nocodazole profoundly affected the behavior of the nucleus. Although spindle-pole bodies (SPBs) could duplicate in the absence of microtubules, SPB separation was blocked. Moreover, complete spindles present at the beginning of drug treatment appeared to collapse, drawing the opposed SPBs and associated nuclear envelope close together. Nuclei did not migrate to the mother-bud necks in nocodazole-treated cells, although nuclei that had reached the necks before drug treatment remained there. Moreover, the double SPBs in arrested cells were often not oriented toward the budding sites, in contrast to the situation in normal cells. Thus, microtubules (cytoplasmic, intranuclear, or both) appear to be necessary for the migration and proper orientation of the nucleus, as well as for SPB separation, spindle function, and nuclear division.


Enhanced responses of spinothalamic tract neurons to excitatory amino acids accompany capsaicin-induced sensitization in the monkey.
Sensitization of the responses of dorsal horn neurons to mechanical stimulation may play a role in the generation of hyperalgesia. Intradermal injection of capsaicin (CAP) provides a model of experimental hyperalgesia that possesses a component of allodynia. This hyperalgesia is produced by chemical stimulation of C-fibers, leading to sensitization of dorsal horn neurons, including spinothalamic tract (STT) cells. The changes in the physiological responses of STT neurons following intradermal CAP in monkeys parallel the acute pain and hyperalgesia produced by intradermal CAP in humans. The present study addresses the role that excitatory amino acids (EAAs) may play in the sensitization of STT neurons by intradermal CAP. Our results show that the background discharge rate and the responses of STT cells to mechanical stimulation increase following intradermal CAP. In addition, the responses of the sensitized cells to one or more iontophoretically released EAA agonists, including NMDA, glutamate, aspartate, kainate, DL-alpha-amino-3-hydroxy-5-methyl-isoxazoleproprionic acid, and/or quisqualate, increase following intradermal CAP. It is proposed that an increase in the responses of STT neurons to EAAs contributes to the hyperalgesia produced by this noxious chemical stimulus.


Phylogenetic structure of unusual aquatic microbial formations in Nullarbor caves, Australia.
The nature of unusual aquatic microbial formations in flooded passages of cave systems in the Nullarbor region of Australia was investigated using electron microscopy and DNA analysis. The caves are located in a semiarid region but intersect the watertable at depths of approximately 100 m below the surface. Throughout submerged portions of the caves divers have noted the presence of unusual microbial formations. These 'microbial mantles' comprise sheets or tongues of mucoid material in which small crystals are embedded. Examination of the biomass revealed it to be primarily composed of densely packed, unbranched filaments, together with spherical-, rod- and spiral-shaped cells, and microcrystals of calcite in a mucoid matrix. Molecular phylogenetic analysis of the community structure revealed approximately 12% of clones showed high similarity to autotrophic nitrite-oxidizing bacteria (Nitrospira moscoviensis). The remainder of the clones exhibited a high proportion of phylogenetically novel sequence types. Chemical analysis of water samples revealed high levels of sulphate and nitrate together with significant nitrite. The community structure, the presence of nitrite in the water, and the apparent absence of aquatic macrofauna, suggest these microbial structures may represent biochemically novel, chemoautotrophic communities dependent on nitrite oxidation.


Generation and analysis of blueberry transcriptome sequences from leaves, developing fruit, and flower buds from cold acclimation through deacclimation.
There has been increased consumption of blueberries in recent years fueled in part because of their many recognized health benefits. Blueberry fruit is very high in anthocyanins, which have been linked to improved night vision, prevention of macular degeneration, anti-cancer activity, and reduced risk of heart disease. Very few genomic resources have been available for blueberry, however. Further development of genomic resources like expressed sequence tags (ESTs), molecular markers, and genetic linkage maps could lead to more rapid genetic improvement. Marker-assisted selection could be used to combine traits for climatic adaptation with fruit and nutritional quality traits. Efforts to sequence the transcriptome of the commercial highbush blueberry (Vaccinium corymbosum) cultivar Bluecrop and use the sequences to identify genes associated with cold acclimation and fruit development and develop SSR markers for mapping studies are presented here. Transcriptome sequences were generated from blueberry fruit at different stages of development, flower buds at different stages of cold acclimation, and leaves by next-generation Roche 454 sequencing. Over 600,000 reads were assembled into approximately 15,000 contigs and 124,000 singletons. The assembled sequences were annotated and functionally mapped to Gene Ontology (GO) terms. Frequency of the most abundant sequences in each of the libraries was compared across all libraries to identify genes that are potentially differentially expressed during cold acclimation and fruit development. Real-time PCR was performed to confirm their differential expression patterns. Overall, 14 out of 17 of the genes examined had differential expression patterns similar to what was predicted from their reads alone. The assembled sequences were also mined for SSRs. From these sequences, 15,886 blueberry EST-SSR loci were identified. Primers were designed from 7,705 of the SSR-containing sequences with adequate flanking sequence. One hundred primer pairs were tested for amplification and polymorphism among parents of two blueberry populations currently being used for genetic linkage map construction. The tetraploid mapping population was based on a cross between the highbush cultivars Draper and Jewel (V. darrowii is also in the background of 'Jewel'). The diploid mapping population was based on a cross between an F1 hybrid of V. darrowii and diploid V. corymbosum and another diploid V. corymbosum. The overall amplification rate of the SSR primers was 68% and the polymorphism rate was 43%. These results indicate that this large collection of 454 ESTs will be a valuable resource for identifying genes that are potentially differentially expressed and play important roles in flower bud development, cold acclimation, chilling unit accumulation, and fruit development in blueberry and related species. In addition, the ESTs have already proved useful for the development of SSR and EST-PCR markers, and are currently being used for construction of genetic linkage maps in blueberry.


Tissue-specific profiling of the Arabidopsis thaliana auxin metabolome.
The plant hormone auxin is believed to influence almost every aspect of plant growth and development. Auxin transport, biosynthesis and degradation combine to form gradients of the hormone that influence a range of key developmental and environmental response processes. There is abundant genetic evidence for the existence of multiple pathways for auxin biosynthesis and degradation. The complexity of these pathways makes it difficult to obtain a clear picture of the relative importance of specific metabolic pathways during development. We have developed a sensitive mass spectrometry-based method to simultaneously profile the majority of known auxin precursors and conjugates/catabolites in small amounts of Arabidopsis tissue. The method includes a new derivatization technique for quantification of the most labile of the auxin precursors. We validated the method by profiling the auxin metabolome in root and shoot tissues from various Arabidopsis thaliana ecotypes and auxin over-producing mutant lines. Substantial differences were shown in metabolite patterns between the lines and tissues. We also found differences of several orders of magnitude in the abundance of auxin metabolites, potentially indicating the relative importance of these compounds in the maintenance of auxin levels and activity. The method that we have established will enable researchers to obtain a better understanding of the dynamics of auxin metabolism and activity during plant growth and development.


EGFR-HIF1α signaling positively regulates the differentiation of IL-9 producing T helper cells.
Interleukin 9 (IL-9)-producing helper T (Th9) cells are essential for inducing anti-tumor immunity and inflammation in allergic and autoimmune diseases. Although transcription factors that are essential for Th9 cell differentiation have been identified, other signaling pathways that are required for their generation and functions are yet to be explored. Here, we identify that Epidermal Growth Factor Receptor (EGFR) is essential for IL-9 induction in helper T (Th) cells. Moreover, amphiregulin (Areg), an EGFR ligand, is critical for the amplification of Th9 cells induced by TGF-β1 and IL-4. Furthermore, our data show that Areg-EGFR signaling induces HIF1α, which binds and transactivates IL-9 and NOS2 promoters in Th9 cells. Loss of EGFR or HIF1α abrogates Th9 cell differentiation and suppresses their anti-tumor functions. Moreover, in line with its reliance on HIF1α expression, metabolomics profiling of Th9 cells revealed that Succinate, a TCA cycle metabolite, promotes Th9 cell differentiation and Th9 cell-mediated tumor regression.


Validation of a multivariate model predicting presence and size of varices.
A model developed by our group identified low platelets and advanced Child-Pugh class (CPC) as being associated with large varices. To validate a defined cut-off of platelets < or =80,000/microL in CPC-A for large varices and platelets < or =90,000/microL in CPC-B/C for any varices. Validation cohorts consisted of patients with cirrhosis undergoing screening for varices from Oregon Health and Science University (n=152), Indiana University (n=252), and Genoa, Italy (n=101). Similar clinical and laboratory data were collected as for the original cohort. To assess the ability of these cut-offs to predict presence of large and any varices, sensitivity, specificity, positive and negative predictive values, likelihood ratios, and the c-statistic were measured. The validation cohorts were statistically different from the original cohort with regards to CPC and prevalence of large varices. Combining the original (n=301) and validation cohorts resulted in a negative predictive value of 92.1% for platelets < or =80,000/microL in CPC-A for large varices and positive predictive value of 80.1% for platelets < or =90,000/microL in CPC-B/C for any varices. Combining the 4 cohorts yielded a c-statistic of 0.67 (95% confidence interval: 063-0.72). No other factors such as splenomegaly and Model for End-Stage Liver Disease score were identified as significant. This study confirms the validity of a previous model identifying low platelets and advanced CPC class as predictors of large varices. Despite combining the cohorts, no other risk factors were identified.


Through-Focus Energy Efficiency and Longitudinal Chromatic Aberration of Three Presbyopia-Correcting Intraocular Lenses.
To compare the chromatic performance of the Bausch & Lomb Versario 3F trifocal intraocular lens (IOL) with the PhysIOL FineVision MicroF trifocal IOL and the Johnson & Johnson Vision TECNIS Symfony ZXR00 extended range of vision (ERV) IOL. The through-focus energy efficiency (TF-EE) was measured in vitro with red (R), green (G), and blue (B) wavelengths and was used to obtain the focus powers and longitudinal chromatic aberrations (LCAs) for each IOL. Other metrics, derived from the RGB TF-EE curves, were assessed for a more complete description of the chromatic performance of the IOLs. Both of the trifocal IOLs, although not specifically designed to tackle chromatic aberrations, showed acceptable LCA (≤0.50 D) in all foci with more balanced R and B efficiencies of their foci. Despite having the lowest TF-EE value at all foci, the Versario 3F demonstrated the most balanced chromatic performance with the smoothest energy transition among all foci and the smallest chromatic span. The Symfony lens effectively reduced LCA at distance and intermediate foci (≤0.36 D), despite the unbalanced and asymmetric R and B efficiencies at its foci. To fully describe the chromatic performance of an IOL it is necessary to take into account not only the LCA but also the RGB TF-EE and chromatic span. This comprehensive analysis suggests that, in comparison with the other IOLs under study, the Versario 3F lens might contribute to further mitigating the impact of chromatic aberration. The in vitro bench testing of the optical properties of modern presbyopia-correcting intraocular lenses (more specifically in this work, the polychromatic through-focus energy efficiency and longitudinal chromatic aberration) provides objective and complementary information that helps to interpret the visual quality outcomes of pseudophakic patients obtained in clinics.


Muscle power in "high-risk" preterm infants at 12 and 24 weeks corrected age: a measure for early detection.
A group of 72 "high-risk" preterm infants was studied at the corrected ages of 12 and 24 wk. Only infants with a high risk for developmental deviance with gestational ages below 32 wk and/or birthweights of less than 1500 g were included in the study. In addition, the infants were categorized according to their medical history, as confirmed by the "Neonatal Medical Index" (NMI I to V), with category I describing infants with few medical problems and V characterizing those with the most serious complications. In this study we included only "high-risk" infants as categorized in NMI III to V, since infants with a "low risk" have been described earlier. Apart from the standard paediatric follow-up, an age-adequate neurological assessment was made, with special emphasis on the relationship between active and passive muscle power. When development is optimal, these two components of muscle power should be in balance in order to create a stable posture and fluent motility. We compared muscle power at the corrected ages of 12 and 24 wk to determine whether the method employed to assess muscle power could enhance early detection of deviant development. At 12 wk of age, only 5 infants showed overall optimal muscle power, while at 24 wk this figure had increased to 27. Significantly more infants in NMI III had optimal outcomes at 24 wk of age. When muscle power in the different parts of the body was studied separately at 24 wk, outcomes in shoulders and trunk still showed significant discrepancies in all NMI groups. At this age, fewer asymmetries were found compared to outcomes at 12 wk. Our method of assessing muscle power is useful in detecting preterm infants at risk for pathological development.


Migraine and possible etiologic heterogeneity for hormone-receptor-negative breast cancer.
Migraine headache is often timed with the menstrual cycle. Some studies have reported reduced risk of breast cancer in migraineurs but most of those did not distinguish menstrually-related from non-menstrually-related migraine. To examine the possible associations between breast cancer and migraine overall and between cancer subcategories and the two migraine subtypes, we used a cohort study of 50,884 women whose sister had breast cancer and a sister-matched case-control study including 1,418 young-onset (<50 years) breast cancer cases. We analyzed the two studies individually and also in tandem via a hybrid Cox model, examining subcategories of breast cancer in relation to menstrually-related and non-menstrually-related migraine. History of migraine was not associated with breast cancer overall. Migraine showed an inverse association with ductal carcinoma in situ (HR = 0.77; 95% CI (0.62,0.96)). Also, women with non-menstrually-related migraine had increased risk (HR = 1.30, 95% CI (0.93,1.81)) while women with menstrually-related migraine had decreased risk (HR = 0.63, 95% CI (0.42,0.96)) of hormone-receptor-negative (ER-/PR-) cancer, with a significant contrast in estimated effects (P = 0.005). While replication of these subset-based findings will be needed, effect specificity could suggest that while migraine has little overall association with breast cancer, menstrual migraine may be associated with reduced risk of ER-/PR- breast cancer.


Cortical region-specific engraftment of embryonic stem cell-derived neural progenitor cells restores axonal sprouting to a subcortical target and achieves motor functional recovery in a mouse model of neonatal hypoxic-ischemic brain injury.
Hypoxic-ischemic encephalopathy (HIE) at birth could cause cerebral palsy (CP), mental retardation, and epilepsy, which last throughout the individual's lifetime. However, few restorative treatments for ischemic tissue are currently available. Cell replacement therapy offers the potential to rescue brain damage caused by HI and to restore motor function. In the present study, we evaluated the ability of embryonic stem cell-derived neural progenitor cells (ES-NPCs) to become cortical deep layer neurons, to restore the neural network, and to repair brain damage in an HIE mouse model. ES cells stably expressing the reporter gene GFP are induced to a neural precursor state by stromal cell co-culture. Forty-hours after the induction of HIE, animals were grafted with ES-NPCs targeting the deep layer of the motor cortex in the ischemic brain. Motor function was evaluated 3 weeks after transplantation. Immunohistochemistry and neuroanatomical tracing with GFP were used to analyze neuronal differentiation and axonal sprouting. ES-NPCs could differentiate to cortical neurons with pyramidal morphology and expressed the deep layer-specific marker, Ctip2. The graft showed good survival and an appropriate innervation pattern via axonal sprouting from engrafted cells in the ischemic brain. The motor functions of the transplanted HIE mice also improved significantly compared to the sham-transplanted group. These findings suggest that cortical region specific engraftment of preconditioned cortical precursor cells could support motor functional recovery in the HIE model. It is not clear whether this is a direct effect of the engrafted cells or due to neurotrophic factors produced by these cells. These results suggest that cortical region-specific NPC engraftment is a promising therapeutic approach for brain repair.


Hydrogen-deuterium exchange reveals long-range dynamical allostery in soybean lipoxygenase.
In lipoxygenases, the topologically conserved C-terminal domain catalyzes the oxidation of polyunsaturated fatty acids, generating an assortment of biologically relevant signaling mediators. Plant and animal lipoxygenases also contain a 100-150-amino acid N-terminal C2-like domain that has been implicated in interactions with isolated fatty acids and at the phospholipid bilayer. These interactions may lead to increased substrate availability and contribute to the regulation of active-site catalysis. Because of a lack of structural information, a molecular understanding of this lipid-protein interaction remains unresolved. Herein, we employed hydrogen-deuterium exchange MS (HDXMS) to spatially resolve changes in protein conformation upon interaction of soybean lipoxygenase with a fatty acid surrogate, oleyl sulfate (OS), previously shown to act at a site separate from the substrate-binding site. Specific, OS-induced conformational changes are detected both at the N-terminal domain and within the substrate portal nearly 30 Å away. Combining previously measured kinetic properties in the presence of OS with its impact on the Kd for linoleic acid substrate binding, we conclude that OS binding brings about an increase in rate constants for both the ingress and egress of substrate. We discuss the role of OS-induced changes in protein flexibility in the context of changes in the mechanism of substrate acquisition.


Detection of markers predictive of macrolide and fluoroquinolone resistance in Mycoplasma genitalium from patients attending sexual health services.
This study sought to provide data on the prevalence of macrolide (23S rRNA) and fluoroquinolone (parC) resistance-associated mutations seen in Mycoplasma genitalium-positive specimens received in the UK national reference laboratory. In total, 2580 clinical specimens from patients with suspected or confirmed M. genitalium infection were received at the national reference laboratory between September 2017 and November 2018. M. genitalium-positive clinical specimens were identified using a reverse transcription-PCR targeting two M. genitalium genes: MgPa and gap. Resistance-associated single nucleotide poylmorphisms were sought in all positive specimens by sequence analysis of the 23S rRNA and parC genes. Eighteen per cent (458 of 2580) of clinical specimens were positive for M. genitalium and 389 had sequence data for both macrolide and fluoroquinolone resistance markers. Of these, 71% (275 of 389) had macrolide resistance-associated mutations, 8% (31 of 389) had fluoroquinolone resistance-associated mutations (S83I/R and D87Y/N) and 7% (26 of 389) had mutations associated with resistance to both antimicrobials. Only 28% (108 of 389) had no mutations associated with resistance to either class of antibiotic. Five specimens had mutations of unknown clinical significance in the parC gene (eg, G81C and S83N). Mutations associated with resistance to macrolides were very frequent. By contrast, susceptibility to the second-line treatment, moxifloxacin (a fluoroquinolone), was estimated at 92% based on the absence of resistance-associated mutations. The few specimens with mutations of unknown clinical significance in the parC gene were excluded from the analysis and so the actual level of fluoroquinolone susceptibility may be slightly lower than that reported here. Surveillance of antimicrobial resistance in M. genitalium is imperative for this to remain a treatable infection.


The targeting of the atToc159 preprotein receptor to the chloroplast outer membrane is mediated by its GTPase domain and is regulated by GTP.
The multimeric translocon at the outer envelope membrane of chloroplasts (Toc) initiates the recognition and import of nuclear-encoded preproteins into chloroplasts. Two Toc GTPases, Toc159 and Toc33/34, mediate preprotein recognition and regulate preprotein translocation. Although these two proteins account for the requirement of GTP hydrolysis for import, the functional significance of GTP binding and hydrolysis by either GTPase has not been defined. A recent study indicates that Toc159 is equally distributed between a soluble cytoplasmic form and a membrane-inserted form, raising the possibility that it might cycle between the cytoplasm and chloroplast as a soluble preprotein receptor. In the present study, we examined the mechanism of targeting and insertion of the Arabidopsis thaliana orthologue of Toc159, atToc159, to chloroplasts. Targeting of atToc159 to the outer envelope membrane is strictly dependent only on guanine nucleotides. Although GTP is not required for initial binding, the productive insertion and assembly of atToc159 into the Toc complex requires its intrinsic GTPase activity. Targeting is mediated by direct binding between the GTPase domain of atToc159 and the homologous GTPase domain of atToc33, the Arabidopsis Toc33/34 orthologue. Our findings demonstrate a role for the coordinate action of the Toc GTPases in assembly of the functional Toc complex at the chloroplast outer envelope membrane.


Social and behavioral outcomes: pre-injury to six months following childhood traumatic brain injury.
This study aimed to extend the limited research investigating social and behavioral outcomes following childhood traumatic brain injury (TBI). The study compared pre-and post-injury measures of these skills and investigated the role of pre-injury child status and pre-injury family functioning in the prediction of outcome at six months post-injury. A secondary aim was to compare rates of impairment at six months post-injury between children post-TBI and a typically developing (TD) control group. This study comprised 140 children, 97 survivors of TBI (67 males) and 43 TD children (24 males), matched for age, sex, and socio-economic status. All participants were ascertained between 2007 and 2010, and were between ages 5.5 and 15.0 years. Children with TBI represented consecutive hospital admissions and were recruited at time of injury into a longitudinal study. TD children were recruited from the community, through local schools chosen to provide a range of socio-economic backgrounds. Findings indicated a deterioration of social participation skills post-injury, particularly for those sustaining a more severe injury, and a consistently higher rate of impairment in social and behavioral outcomes in the TBI group. Pre-injury function, injury severity and restrictions to social participation (e.g., reduced sport activities) as recommended by clinicians contributed significantly to outcome. Difficulties are evident in the short-term post-childhood TBI in social and behavioral domains. It is essential to monitor children long-term, particularly as societal expectations and demands increase.


Retained asymptomatic third molars and risk for second molar pathology.
Prophylactic extraction of unerupted asymptomatic third molars is the most common oral surgery procedure in the United States. However, limited evidence exists to justify its costs and associated morbidity. We analyzed data collected over 25 years from 416 adult men enrolled in the Veterans Affairs Dental Longitudinal Study to evaluate the association of retained asymptomatic third molars with risk of adjacent second molar pathology (caries and/or periodontitis), based on third molar status (i.e., absent, erupted, or unerupted). Unerupted molars were further categorized as either "soft tissue" or "bony" impacted. We found that the lowest prevalence and incidence of second molar pathology occurred when the adjacent third molar was absent. The presence of a third molar that was soft tissue impacted increased the risk of incident second molar pathology 4.88-fold (95% confidence interval: 2.62, 9.08). Having an erupted or "bony" impacted third molar increased the risk of incident second molar pathology by 1.74 (95% confidence interval: 1.34, 2.25) and 2.16 (95% confidence interval: 1.56, 2.99), respectively. The retention of third molars is associated with increased risk of second molar pathology in middle-aged and older adult men.


[A Future Perspective on the Involvement of n-3 Polyunsaturated Fatty Acid in the Development of Nonalcoholic Fatty Liver Disease/Nonalcoholic Steatohepatitis].
Nonalcoholic fatty liver disease (NAFLD) is characterized by the pathological accumulation of fat in the liver in the absence of any other disease related to liver steatosis, which includes a wide spectrum of liver diseases ranging from mild asymptomatic fatty liver to nonalcoholic steatohepatitis (NASH) and cirrhosis. Recently, it was reported that NAFLD is characterized by the impaired bioavailability of liver n-6 and n-3 long-chain polyunsaturated fatty acids (PUFAs). That is, compared with healthy individuals, steatosis and steatohepatitis patients have higher n-6/n-3 PUFA ratios. Furthermore, per recent research, decreasing the intake of total fats and increasing the intake of n-3 PUFAs may be beneficial in the treatment of NAFLD. In contrast, some reports describe that NASH patients have more metabolic abnormalities than NAFLD patients; however, these are not influenced by dietary fatty acids. Thus, at present, various opinions exist regarding the efficacy of n-3 PUFA in the treatment of NAFLD. In this review, we discuss the considerable interest n-3 PUFA has attracted as a potential treatment for NAFLD.


OX40 is a potent immune-stimulating target in late-stage cancer patients.
OX40 is a potent costimulatory receptor that can potentiate T-cell receptor signaling on the surface of T lymphocytes, leading to their activation by a specifically recognized antigen. In particular, OX40 engagement by ligands present on dendritic cells dramatically increases the proliferation, effector function, and survival of T cells. Preclinical studies have shown that OX40 agonists increase antitumor immunity and improve tumor-free survival. In this study, we performed a phase I clinical trial using a mouse monoclonal antibody (mAb) that agonizes human OX40 signaling in patients with advanced cancer. Patients treated with one course of the anti-OX40 mAb showed an acceptable toxicity profile and regression of at least one metastatic lesion in 12 of 30 patients. Mechanistically, this treatment increased T and B cell responses to reporter antigen immunizations, led to preferential upregulation of OX40 on CD4(+) FoxP3(+) regulatory T cells in tumor-infiltrating lymphocytes, and increased the antitumor reactivity of T and B cells in patients with melanoma. Our findings clinically validate OX40 as a potent immune-stimulating target for treatment in patients with cancer, providing a generalizable tool to favorably influence the antitumor properties of circulating T cells, B cells, and intratumoral regulatory T cells.


Involvement of phosphatidylinositol 4,5-bisphosphate in RNA polymerase I transcription.
RNA polymerase I (Pol I) transcription is essential for the cell cycle, growth and protein synthesis in eukaryotes. In the present study, we found that phosphatidylinositol 4,5-bisphosphate (PIP2) is a part of the protein complex on the active ribosomal promoter during transcription. PIP2 makes a complex with Pol I and the Pol I transcription factor UBF in the nucleolus. PIP2 depletion reduces Pol I transcription, which can be rescued by the addition of exogenous PIP2. In addition, PIP2 also binds directly to the pre-rRNA processing factor fibrillarin (Fib), and co-localizes with nascent transcripts in the nucleolus. PIP2 binding to UBF and Fib modulates their binding to DNA and RNA, respectively. In conclusion, PIP2 interacts with a subset of Pol I transcription machinery, and promotes Pol I transcription.


Shared signatures of social stress and aging in peripheral blood mononuclear cell gene expression profiles.
Chronic social stress is a predictor of both aging-related disease and mortality risk. Hence, chronic stress has been hypothesized to directly exacerbate the process of physiological aging. Here, we evaluated this hypothesis at the level of gene regulation. We compared two data sets of genome-wide gene expression levels in peripheral blood mononuclear cells (PBMCs): one that captured aging effects and another that focused on chronic social stress. Overall, we found that the direction, although not necessarily the magnitude, of significant gene expression changes tends to be shared between the two data sets. This overlap was observable at three levels: (i) individual genes; (ii) general functional categories of genes; and (iii) molecular pathways implicated in aging. However, we also found evidence that heterogeneity in PBMC composition limits the power to detect more extensive similarities, suggesting that our findings reflect an underestimate of the degree to which age and social stress influence gene regulation in parallel. Cell type-specific data on gene regulation will be important to overcome this limitation in the future studies.


Effect of percutaneous endoscopic gastrostomy on gastric emptying in clinically normal cats.
To assess the effect of percutaneous endoscopic gastrostomy (PEG) tube placement on gastric emptying in clinically normal cats. 8 healthy adult 3- to 5-year-old cats. Cats were accommodated to the diet for 2 weeks prior to scintigraphy. Caloric needs were divided into 3 feedings/d. Food was withheld for 24 hours after tube placement, then was fed as a third of the caloric needs on day 1, two-thirds on day 2, and full caloric requirements thereafter. Gastric emptying was measured via nuclear scintigraphy. Labeled meals contained 111 MBq (3 mCi) of 99mTc-labeled disofenin. Sixty-second ventral scintigraphic images were acquired immediately, every 20 minutes for the first hour, then every 30 minutes for 4 hours after feeding. Each cat was evaluated 3 times prior to PEG tube placement. Cats were anesthetized, and 16-F mushroom-tipped Pezzar gastrostomy tubes were placed, using a video endoscope. Scintigraphy was repeated on days 1, 4, 7, 11, 14, and 21 after PEG tube placement. Gastric emptying was faster with a PEG tube in place. Percentage of retained gastric activity was significantly lower after PEG for 150, 180, 210, and 240 minutes versus time before PEG tube placement. Placement of a PEG tube does not delay gastric emptying in clinically normal cats. Gastric retention of food, vomiting, and aspiration pneumonia after PEG tube placement may not be related to delayed gastric emptying.


Factor IX is activated in vivo by the tissue factor mechanism.
Despite significant progress in elucidating the biochemistry of the hemostatic mechanism, the process of blood coagulation in vivo remains poorly understood. Factor IX is a vitamin K-dependent glycoprotein that can be activated by factor XIa or the factor VII-tissue factor complex in vitro. To investigate the role of these two pathways in factor IX activation in humans, we have developed a sensitive procedure for quantifying the peptide that is liberated with the generation of factor IXa. The antibody population used for the immunoassay was raised in rabbits and chromatographed on a factor IX-agarose immunoadsorbent to obtain antibody populations with minimal intrinsic reactivity toward factor IX. We determined that the mean level of the factor IX activation peptide (FIXP) in normal individuals under the age of 40 years was 203 pmol/L and that levels increased significantly with advancing age. The mean concentration of FIXP was markedly reduced to 22.7 pmol/L in nine patients with hereditary factor VII deficiency (factor VII coagulant activity less than 7%) but was not significantly different from normal controls in nine subjects with factor XI deficiency (factor XI coagulant activity less than 8%). These data indicate that factor IXa generation in vivo results mainly from the activity of the tissue factor mechanism rather than the contact system (factor XII, prekallikrein, high molecular-weight kininogen, factor XI). Our results may also help to explain the absence of a bleeding diathesis in many patients with deficiencies of the contact factors of coagulation.


Methods and reliability of echocardiographic assessment of left atrial size and mechanical function in horses.
To assess the feasibility, describe the techniques, and determine the reliability of transthoracic echocardiography for characterization of left atrial (LA) size and LA mechanical function in horses. 6 healthy adult horses. Repeated echocardiographic examinations were performed independently by 2 observers in standing, unsedated horses by use of 2-dimensional echocardiography, pulsed-wave flow Doppler, and tissue Doppler imaging (TDI) techniques. Test reliability was determined by estimating measurement variability, within-day interobserver variability, and between-day inter- and intraobserver variability of all echocardiographic variables. Variability was expressed as the coefficient of variation (CV) and the absolute value below which the difference between 2 measurements will lie with 95% probability. Most echocardiographic variables of LA size had low overall variability (CV, < 15%). Among the 2-dimensional indices of LA mechanical function, area-based and volume-based ejection phase indices had moderate between-day variability (CV usually < 25%). Transmitral Doppler flow indices were characterized by low to high between-day variability (CV, 6% to 35%). The TDI wall motion velocities had high between-day variability (CV, > 25%), whereas most TDI-derived time intervals had low variability (CV, < 15%). LA size and mechanical function can be reliably assessed in standing, unsedated horses by use of 2-dimensional echocardiography, transmitral blood flow velocity profiles, and analyses of LA wall motion by use of TDI. These results may provide useful recommendations for echocardiographic assessment of LA size and function in horses.


Integration of HIV infant testing into immunization programmes: a systematic review.
Integration of HIV infant testing into immunization sessions is one of the strategies designed to increase coverage of early infant diagnosis. To determine the evidence on the outcomes of such integration. A systematic review of peer-reviewed and grey literature was undertaken from electronic sources such as MEDLINE, Google Scholar, websites of international agencies, past conferences and ministries of health reports published between year 2002 and 2013. Randomized controlled trials, observational and qualitative studies were searched and those meeting selection criteria were selected and relevant information extracted using structured tool. Statistical pooling was not possible owing to the heterogeneity of the study designs and outcome measures. Of the nine articles which met the selection criteria, none used a randomized controlled design. Of these, five articles measured mother's acceptability of their infants being tested for HIV during its first pentavalent or DPT vaccination visit, and 89·5-100% accepted. Four articles reported the proportion of mothers who returned for HIV test results, ranging from 56·8% to 86·0%. Increased uptake of HIV testing following integration was confirmed by two articles. Only one study in Tanzania determined the uptake of vaccinations following integration, with urban facilities showing stable or slight increase of monthly vaccine uptake while decreases were observed across the rural sites. In two articles, stigma was perceived by service-providers and mothers as the potential risk following integration, particularly in rural settings. Despite the limited number of articles, the findings in this systematic review suggest that HIV testing during immunization clinic visits is acceptable and feasible as a possible model for service delivery. However, the impact on vaccination uptake needs further study.


Sida rhomboidea.Roxb aqueous extract down-regulates in vivo expression of vascular cell adhesion molecules in atherogenic rats and inhibits in vitro macrophage differentiation and foam cell formation.
The present study evaluates efficacy of Sida rhomboidea.Roxb (SR) leaves extract in ameliorating experimental atherosclerosis using in vitro and in vivo experimental models. Atherogenic (ATH) diet fed rats recorded significant increment in the serum total cholesterol (TC), triglycerides (TG), low-density lipoprotein (LDL), very LDL (VLDL), autoantibody against oxidized LDL (Ox-LDL), markers of LDL oxidation and decrement in high-density lipoprotein (HDL) along with increment in aortic TC and TG. The ex vivo LDL oxidation assay revealed an increased susceptibility of LDL isolated from ATH rats to undergo copper mediated oxidation. These set of changes were minimized by simultaneous co-supplementation of SR extract to ATH diet fed rats. Histopathology of aorta and immunolocalization studies recorded pronounced atheromatous plaque formation, vascular calcification, significant elastin derangements and higher expression of macrophage surface marker (F4/80), vascular cell adhesion molecule-1 (VCAM-1) and p-selectin in ATH rats. Whereas, ATH+SR rats depicted minimal evidence of atheromatous plaque formation, calcium deposition, distortion/defragmentation of elastin and accumulation of macrophages along with lowered expression of VCAM-1 and P-selectin compared to ATH rats. Further, monocyte to macrophage differentiation and in vitro foam cell formation were significantly attenuated in presence of SR extract. In conclusion, SR extract has the potency of controlling experimental atherosclerosis and can be used as promising herbal supplement in combating atherosclerosis.


Strain effects on the work function of an organic semiconductor.
Establishing fundamental relationships between strain and work function (WF) in organic semiconductors is important not only for understanding electrical properties of organic thin films, which are subject to both intrinsic and extrinsic strains, but also for developing flexible electronic devices. Here we investigate tensile and compressive strain effects on the WF of rubrene single crystals. Mechanical strain induced by thermal expansion mismatch between the substrate and rubrene is quantified by X-ray diffraction. The corresponding WF change is measured by scanning Kelvin probe microscopy. The WF of rubrene increases (decreases) significantly with in-plane tensile (compressive) strain, which agrees qualitatively with density functional theory calculations. An elastic-to-plastic transition, characterized by a steep rise of the WF, occurs at ∼0.05% tensile strain along the rubrene π-stacking direction. The results provide the first concrete link between mechanical strain and WF of an organic semiconductor and have important implications for understanding the connection between structural and electronic disorder in soft organic electronic materials.


New selectable marker for manipulating the simple genomes of Mycoplasma species.
Over the past several years, significant advances have been made in the molecular genetics of the Mollicutes (the simplest cells that can be grown in axenic culture). Nevertheless, a number of basic molecular tools are still required before genetic manipulations become routine. Here we describe the development of a new dominant selectable marker based on the enzyme puromycin-N-acetyltransferase from Streptomyces alboniger. Puromycin is an antibiotic that mimics the 3'-terminal end of aminoacylated tRNAs and attaches to the carboxyl terminus of growing protein chains. This stops protein synthesis. Because puromycin conscripts rRNA recognition elements that are used by all of the various tRNAs in a cell, it is unlikely that spontaneous antibiotic resistance can be acquired via a simple point mutation--an annoying issue with existing mycoplasma markers. Our codon-optimized cassette confers pronounced puromycin resistance on all five of the mycoplasma species we have tested so far. The resistance cassette was also designed to function in Escherichia coli, which simplifies the construction of shuttle vectors and makes it trivial to produce the large quantities of DNA generally necessary for mycoplasma transformation. Due to these and other features, we expect the puromycin marker to be a widely applicable tool for studying these simple cells and pathogens.


Sanitary Transportation of Human and Animal Food. Final rule.
The Food and Drug Administration (FDA or we) is issuing a final rule to establish requirements for shippers, loaders, carriers by motor vehicle and rail vehicle, and receivers engaged in the transportation of food, including food for animals, to use sanitary transportation practices to ensure the safety of the food they transport. This action is part of our larger effort to focus on prevention of food safety problems throughout the food chain and is part of our implementation of the Sanitary Food Transportation Act of 2005 (2005 SFTA) and the Food Safety Modernization Act of 2011 (FSMA).


Premenstrual dysphoric disorder: evidence for a new category for DSM-5.
Premenstrual dysphoric disorder, which affects 2%–5% of premenopausal women, was included in Appendix B of DSMIV, "Criterion Sets and Axes Provided for Further Study." Since then, aided by the inclusion of specific and rigorous criteria in DSM-IV, there has been an explosion of research on the epidemiology, phenomenology, pathogenesis, and treatment of the disorder. In 2009, the Mood Disorders Work Group for DSM-5 convened a group of experts to examine the literature on premenstrual dysphoric disorder and provide recommendations regarding the appropriate criteria and placement for the disorder in DSM-5. Based on thorough review and lengthy discussion, the work group proposed that the information on the diagnosis, treatment, and validation of the disorder has matured sufficiently for it to qualify as a full category in DSM-5. A move to the position of category, rather than a criterion set in need of further study, will provide greater legitimacy for the disorder and encourage the growth of evidence-based research, ultimately leading to new treatments.


Cancer risk in the reindeer breeding Saami population of Sweden, 1961-1997.
The Saami people are the natives of northern Scandinavia and the Kola Peninsula. In a cohort of 2033 Swedish reindeer breeding Saamis, the cancer risks between 1961 and 1997 were studied. In total, 193 cases of cancer were observed versus 322 expected in the general Swedish population and 249 in a geographically matched reference population of non-Saamis. In comparison to non-Saamis living in the same area, the reindeer breeding Saamis showed a statistically significant lower risk of developing cancer of the prostate and of malignant lymphoma, whereas the risk of stomach cancer was significantly higher. Although there were no statistically significant changes of cancer risks over time, temporal trends were indicated towards a decreased risk of cancer in the stomach and the prostate. The results suggest that the explanations of the low cancer risk of the reindeer breeding Saamis, in relation to the non-Saamis in the same environment, are to be found among lifestyle and/or genetic factors.


Selective digestive tract decontamination to prevent healthcare associated infections in critically ill children: the PICNIC multicentre randomised pilot clinical trial.
Healthcare-associated infections (HCAIs) are a major cause of morbidity and mortality in critically ill children. Data from adult studies suggest Selective Decontamination of the Digestive tract (SDD) may reduce the incidence of HCAIs and improve survival. There are no data from randomised clinical trials in the paediatric setting. An open label, parallel group pilot cRCT and mixed-methods perspectives study was conducted in six paediatric intensive care units (PICUs) in England. Participants were children (> 37 weeks corrected gestational age, up to 16 years) requiring mechanical ventilation expected to last for at least 48 h. Sites undertook standard care for a period of 9 weeks and were randomised into 3 sites which continued standard care and 3 where SDD was incorporated into infection control practice for eligible children. Interviews and focus groups were conducted for parents and staff working in PICU. 434 children fulfilled eligibility criteria, of whom 368 (85%) were enrolled. This included 207 in the baseline phase (Period One) and 161 in the intervention period (Period Two). In sites delivering SDD, the majority (98%) of children received at least one dose of SDD and of these, 68% commenced within the first 6 h. Whilst admission swabs were collected in 91% of enrolled children, consent for the collection of additional swabs was low (44%). Recruited children were representative of the wider PICU population. Overall, 3.6 children/site/week were recruited compared with the potential recruitment rate for a definitive cRCT of 3 children/site/week, based on data from all UK PICUs. Parents (n = 65) and staff (n = 44) were supportive of the aims of the study, suggesting adaptations for a larger definitive trial including formulation and administration of SDD paste, approaches to consent and ecology monitoring. Stakeholders identified preferred clinical outcomes, focusing on complications of critical illness and quality-of-life. A definitive cRCT in SDD to prevent HCAIs in critically ill children is feasible but should include adaptations to ecology monitoring along with the dosing schedule and packaging into a paediatric specific format. A definitive study is supported by the findings with adaptations to ecology monitoring and SDD administration.Trial Registration: ISRCTN40310490 Registered 30/10/2020.


Compensatory effects in the PI3K/PTEN/AKT signaling network following receptor tyrosine kinase inhibition.
Overcoming de novo and acquired resistance to anticancer drugs that target signaling networks is a formidable challenge for drug design and effective cancer therapy. Understanding the mechanisms by which this resistance arises may offer a route to addressing the insensitivity of signaling networks to drug intervention and restore the efficacy of anticancer therapy. Extending our recent work identifying PTEN as a key regulator of Herceptin sensitivity, we present an integrated theoretical and experimental approach to study the compensatory mechanisms within the PI3K/PTEN/AKT signaling network that afford resistance to receptor tyrosine kinase (RTK) inhibition by anti-HER2 monoclonal antibodies. In a computational model representing the dynamics of the signaling network, we define a single control parameter that encapsulates the balance of activities of the enzymes involved in the PI3K/PTEN/AKT cycle. By varying this control parameter we are able to demonstrate both distinct dynamic regimes of behavior of the signaling network and the transitions between those regimes. We demonstrate resistance, sensitivity, and suppression of RTK signals by the signaling network. Through model analysis we link the sensitivity-to-resistance transition to specific compensatory mechanisms within the signaling network. We study this transition in detail theoretically by variation of activities of PTEN, PI3K, AKT enzymes, and use the results to inform experiments that perturb the signaling network using combinatorial inhibition of RTK, PTEN, and PI3K enzymes in human ovarian carcinoma cell lines. We find good alignment between theoretical predictions and experimental results. We discuss the application of the results to the challenges of hypersensitivity of the signaling network to RTK signals, suppression of drug resistance, and efficacy of drug combinations in anticancer therapy.