A Multicentered Study on Epidemiologic and Clinical Characteristics of 37 Neonates With Community-acquired COVID-19.
Coronavirus disease 2019 (COVID-19) primarily affects adults and spares children, whereas very little is known about neonates. We tried to define the clinical characteristics, risk factors, laboratory, and imagining results of neonates with community-acquired COVID-19. This prospective multicentered cohort study included 24 neonatal intensive care units around Turkey, wherein outpatient neonates with COVID-19 were registered in an online national database. Full-term and premature neonates diagnosed with COVID-19 were included in the study, whether hospitalized or followed up as ambulatory patients. Neonates without severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) via reverse transcriptase-polymerase chain reaction testing or whose mothers had been diagnosed with COVID-19 during pregnancy were excluded. Thirty-seven symptomatic neonates were included. The most frequent findings were fever, hypoxemia, and cough (49%, 41%, 27%, respectively). Oxygen administration (41%) and noninvasive ventilation (16%) were frequently required; however, mechanical ventilation (3%) was rarely needed. Median hospitalization was 11 days (1-35 days). One patient with Down syndrome and congenital cardiovascular disorders died in the study period. C-reactive protein (CRP) and prothrombin time (PT) levels were found to be higher in patients who needed supplemental oxygen (0.9 [0.1-8.6] vs. 5.8 [0.3-69.2] p = 0.002, 11.9 [10.1-17.2] vs. 15.2 [11.7-18.0] p = 0.01, respectively) or who were severe/critical (1.0 [0.01-8.6] vs. 4.5 [0.1-69.2] p = 0.01, 11.7 [10.1-13.9] vs. 15.0 [11.7-18.0] p = 0.001, respectively). Symptomatic neonates with COVID-19 had high rates of respiratory support requirements. High CRP levels or a greater PT should alert the physician to more severe disease.


Growth hormone receptor C-terminal domains required for growth hormone-induced intracellular free Ca2+ oscillations and gene transcription.
The biological effects of growth hormone (GH) are initiated by its binding to the GH receptor (GHR) followed by association and activation of the tyrosine kinase JAK2. Here we report that GH can stimulate an increase in intracellular free Ca2+ concentration ([Ca2+]i) in cells expressing wild-type GHRs and receptor mutants lacking up to 132 amino acids of the C terminus, whereas GHRs lacking a further 52 amino acids in the C terminus are unable to induce Ca2+ signaling. The GH-induced rise in [Ca2+]i was dependent upon extracellular Ca2+ and the response consisted of GH-induced Ca2+ oscillations of varying frequency and amplitude. GH-induced transcription of the serine protease inhibitor 2.1 gene required the same C-terminal 52-amino acid domain of the receptor as for Ca2+ signaling. Mutation of the four proline residues in the conserved box 1 region of the GHR, which is responsible for binding and activation of JAK2 kinase, completely abolished GH-induced gene transcription but did not affect the GH-induced rise in [Ca2+]i. The Ca2+ channel blocker verapamil prevented GH-induced Ca2+ signaling as well as GH-induced gene transcription in cells expressing endogenous GHRs. These findings indicate that the GHR can initiate two independent signaling pathways, one requiring the box 1 region and the other requiring the region between amino acids 454 and 506, and suggest that both of these pathways are required for GH-induced gene transcription.


Normal response to large posteroanterior lumbar loads--a case study approach.
Characterization of the responses to spinal posteroanterior loading has involved measuring the gradient of the linear region of the force-displacement relationship between 30 N and 100 N of applied force. However, forces applied during manual assessment of stiffness may reach 300 N. This study investigated whether the force-displacement relation remains linear above 100 N. A single-case-study design with replication was used. Two male subjects, with no history of low back pain requiring treatment over the preceding twelve months, participated in this study. Posteroanterior loads of 275 N were applied to the 1.3 vertebra of both subjects using a mechanical device. The force-displacement responses in these subjects were measured and analyzed to examine the linearity of the data. The responses from the two subjects were similar. In both cases, the responses were approximately linear at all levels of load up to 275 N. Because of the small degree of nonlinearity, there was a tendency for the stiffness to show moderate increases as the load increased. Stiffness was approximately 25% greater when calculated between 30 N and 275 N than when calculated between 30 N and 100 N. In these two subjects, the force-displacement relation was approximately linear for all three force intervals considered, although there was some increasing stiffness with increasing force.


SOX10 expression in malignant melanoma, carcinoma, and normal tissues.
Sry-related HMg-Box gene 10 (SOX10) is a nuclear transcription factor that plays an important role in melanocytic cell differentiation. It has been shown to be a sensitive marker of melanoma including spindle and desmoplastic subtypes. We assessed its frequency of expression in melanoma, carcinoma, benign nevi, and non-neoplastic tissues with routine immunohistochemistry for SOX10. The 109 primary melanoma included 49 epithelioid, 19 spindle cell, 22 desmoplastic, and 19 mixed spindle cell/desmoplastic melanoma. All primary, except 8 desmoplastic melanoma, and 11 metastatic melanoma were strongly and diffusely nuclear SOX10-positive. Six desmoplastic melanoma had ≤10% cells positive, and 2 were <50% positive, all of 3+ intensity. Eighteen of 149 (12%) breast carcinoma were SOX10-positive. All 24 ovarian, 23 endometrial, 26 lung, and 25 colon carcinoma were SOX10-negative. All 43 benign nevi, 18 dysplastic nevi, 68 non-neoplastic and benign skins, and all 56 non-neoplastic breast tissue were SOX10-positive. The sensitivity and specificity for SOX10 in the diagnosis of melanoma are 1.0 and 0.93, respectively; the positive and negative predictive values are 0.87 and 1.0, respectively. SOX10 is a sensitive, specific marker for melanoma. As benign nevi also express SOX10, it cannot be used to differentiate between benign and malignant pigmented skin lesions. Only a small number of breast carcinoma (12%), and breast lobules, express SOX10; no carcinoma of the ovary, endometrium, lung, or colon expressed SOX10.


Mixtures of aromatic compounds induce ligninolytic gene expression in the wood-rotting fungus Dichomitus squalens.
Heterologous production of fungal ligninolytic cocktails is challenging due to the low yields of catalytically active lignin modifying peroxidases. Production using a natural system, such as a wood-rotting fungus, is a promising alternative if specific or preferential induction of the ligninolytic activities could be achieved. Using transcriptomics, gene expression of the white-rot Dichomitus squalens during growth on mixtures of aromatic compounds, with ring structures representing the two major lignin sub-units, was compared to a wood substrate. Most of the genes encoding lignin modifying enzymes (laccases and peroxidases) categorised as highly or moderately expressed on wood were expressed similarly on aromatic compounds. Higher expression levels of a subset of manganese and versatile peroxidases was observed on di- compared to mono-methoxylated aromatics. The expression of polysaccharide degrading enzymes was lower on aromatic compounds compared to wood, demonstrating that the induction of lignin modifying enzymes became more specific. This study suggests potential for aromatic waste streams, e.g. from lignocellulose pretreatment, to produce a lignin-specific enzyme cocktail from D. squalens or other white-rot fungi.


Weighted filtered backprojection for quantitative fluorescence optical projection tomography.
Reconstructing images from a set of fluorescence optical projection tomography (OPT) projections is a relatively new problem. Several physical aspects of fluorescence OPT necessitate a different treatment of the inverse problem to that required for non-fluorescence tomography. Given a fluorophore within the depth of field of the imaging system, the power received by the optical system, and therefore the CCD detector, is related to the distance of the fluorophore from the objective entrance pupil. Additionally, due to the slight blurring of images of sources positioned off the focal plane, the CCD image of a fluorophore off the focal plane is lower in intensity than the CCD image of an identical fluorophore positioned on the focal plane. The filtered backprojection (FBP) algorithm does not take these effects into account and so cannot be expected to yield truly quantitative results. A full model of image formation is introduced which takes into account the effects of isotropic emission and defocus. The model is used to obtain a weighting function which is used in a variation of the FBP algorithm called weighted filtered backprojection (WFBP). This new algorithm is tested with simulated data and with experimental data from a phantom consisting of fluorescent microspheres embedded in an agarose gel.


Web-based randomised controlled trials in orthodontics.
Randomised controlled trials (RCT) are considered the best source of scientific evidence--the gold standard--when evaluating the efficacy of orthodontic treatments. Frequently, RCT are planned as multicentre trials, with the intention of increasing statistical power and raising the precision of outcome estimates. The management of large-scale RCT, however, requires even more thorough organisation than conventional RCT. Indeed, the need for high accuracy and standardisation in data collection, research aids, secretarial skills, staff and patient training, and organisational meetings, make these studies time-consuming, expensive and, in general, relatively complex to carry out well. A website was developed to support a large scale-orthodontic RCT which aimed to evaluate the efficacy of a functional appliance(www.ortodonzia.unina.it). Websites such as this can increase the quality of data collection, simplify the randomisation process, speed up data collection, and improve trial monitoring. Web-based RCT have the potential to help globalise orthodontic research and also increase our rate of acquisition of evidence in orthodontics.


Investigating the pretreatment miRNA expression patterns of advanced hepatocellular carcinoma patients in association with response to TACE treatment.
Hepatocellular carcinoma (HCC) is a lethal malignancy with poor prognosis and limited treatment options. Transarterial chemoembolization (TACE) using chemotherapy agents-doxorubicin and cisplatin-is an accepted treatment option for locally advanced hepatocellular carcinoma. In the current study, we analyzed the expression pattern of a selected panel of 94 miRNAs in archival samples that were collected prior to treatment from 15 Egyptian patients diagnosed with advanced hepatocelleular carcinoma. We observed an overall increase in miRNA expression in HCC samples compared with normal subjects. Out of 94 examined miRNAs, 53 were significantly upregulated while 3 miRNAs were downregulated in HCC samples compared to normal liver samples. Comparing the pretreatment miRNA expression profiles in HCC patients and the patients response to TACE treatment resulted in the identification of a set of 12 miRNAs that are significantly upregulated in nonresponders group. This miRNA panel includes miR-10a-1, miR-23a-1, miR-24, miR-26a, miR-27a, miR-30c, miR-30e, miR-106b, miR-133b, miR-199a, miR-199-3p, and miR-200b. Furthermore, we observed that a panel of 10 miRNAs was significantly associated with patients' survival status at 1 year. These results highlight the potential implications of pretreatment miRNAs expression profiling in prediction of the patients' response to TACE treatment in liver cancer.


Identification of different types of spinal afferent nerve endings that encode noxious and innocuous stimuli in the large intestine using a novel anterograde tracing technique.
In mammals, sensory stimuli in visceral organs, including those that underlie pain perception, are detected by spinal afferent neurons, whose cell bodies lie in dorsal root ganglia (DRG). One of the major challenges in visceral organs has been how to identify the different types of nerve endings of spinal afferents that transduce sensory stimuli into action potentials. The reason why spinal afferent nerve endings have been so challenging to identify is because no techniques have been available, until now, that can selectively label only spinal afferents, in high resolution. We have utilized an anterograde tracing technique, recently developed in our laboratory, which facilitates selective labeling of only spinal afferent axons and their nerve endings in visceral organs. Mice were anesthetized, lumbosacral DRGs surgically exposed, then injected with dextran-amine. Seven days post-surgery, the large intestine was removed. The characteristics of thirteen types of spinal afferent nerve endings were identified in detail. The greatest proportion of nerve endings was in submucosa (32%), circular muscle (25%) and myenteric ganglia (22%). Two morphologically distinct classes innervated myenteric ganglia. These were most commonly a novel class of intraganglionic varicose endings (IGVEs) and occasionally rectal intraganglionic laminar endings (rIGLEs). Three distinct classes of varicose nerve endings were found to innervate the submucosa and circular muscle, while one class innervated internodal strands, blood vessels, crypts of lieberkuhn, the mucosa and the longitudinal muscle. Distinct populations of sensory endings were CGRP-positive. We present the first complete characterization of the different types of spinal afferent nerve endings in a mammalian visceral organ. The findings reveal an unexpectedly complex array of different types of primary afferent endings that innervate specific layers of the large intestine. Some of the novel classes of nerve endings identified must underlie the transduction of noxious and/or innocuous stimuli from the large intestine.


Optical detection of buccal epithelial nanoarchitectural alterations in patients harboring lung cancer: implications for screening.
We have recently developed a novel optical technology, partial wave spectroscopic (PWS) microscopy, which is exquisitely sensitive to the nanoarchitectural manifestation of the genetic/epigenetic alterations of field carcinogenesis. Our approach was to screen for lung cancer by assessing the cheek cells based on emerging genetic/epigenetic data which suggests that the buccal epithelium is altered in lung field carcinogenesis. We performed PWS analysis from microscopically normal buccal epithelial brushings from smokers with and without lung cancer (n = 135). The PWS parameter, disorder strength of cell nanoarchitecture (L(d)), was markedly (>50%) elevated in patients harboring lung cancer compared with neoplasia-free smokers. The performance characteristic was excellent with an area under the receiver operator characteristic curve of >0.80 and was equivalent for both disease stage (early versus late) and histologies (small cell versus non-small cell lung cancers). An independent data set validated the findings with only a minimal degradation of performance characteristics. Our results offer proof of concept that buccal PWS may potentially herald a minimally intrusive prescreening test that could be integral to the success of lung cancer population screening programs.


European Association of Urology Guidelines on Renal Cell Carcinoma: The 2019 Update.
The European Association of Urology Renal Cell Carcinoma (RCC) Guideline Panel has prepared evidence-based guidelines and recommendations for the management of RCC. To provide an updated RCC guideline based on standardised methodology including systematic reviews, which is robust, transparent, reproducible, and reliable. For the 2019 update, evidence synthesis was undertaken based on a comprehensive and structured literature assessment for new and relevant data. Where necessary, formal systematic reviews adhering to the Preferred Reporting Items for Systematic Reviews and Meta-analyses (PRISMA) guidelines were undertaken. Relevant databases (Medline, Cochrane Libraries, trial registries, conference proceedings) were searched until June 2018, including randomised controlled trials (RCTs) and retrospective or controlled studies with a comparator arm, systematic reviews, and meta-analyses. Where relevant, risk of bias (RoB) assessment, and qualitative and quantitative syntheses of the evidence were performed. The remaining sections of the document were updated following a structured literature assessment. Clinical practice recommendations were developed and issued based on the modified GRADE framework. All chapters of the RCC guidelines were updated based on a structured literature assessment, for prioritised topics based on the availability of robust data. For RCTs, RoB was low across studies. For most non-RCTs, clinical and methodological heterogeneity prevented pooling of data. The majority of included studies were retrospective with matched or unmatched cohorts, based on single- or multi-institutional data or national registries. The exception was for the treatment of metastatic RCC, for which there were several large RCTs, resulting in recommendations based on higher levels of evidence. The 2019 RCC guidelines have been updated by the multidisciplinary panel using the highest methodological standards. These guidelines provide the most reliable contemporary evidence base for the management of RCC in 2019. The European Association of Urology Renal Cell Carcinoma Guideline Panel has thoroughly evaluated the available research data on kidney cancer to establish international standards for the care of kidney cancer patients.


Associations between Physical Activity and Cognitive Functioning among Middle-Aged and Older Adults.
To describe aerobic physical activity among middle-aged and older adults by their self-reported cognitive decline and their receipt of informal care for declines in cognitive functioning and most common type of physical activity. Cross-sectional study using data from the 2011 Behavioral Risk Factor Surveillance System. Landline and cellular telephone survey. 93,082 respondents aged 45 years and older from 21 US states in 2011. Subjective cognitive decline (SCD) was defined as experiencing confusion or memory loss that was happening more often or getting worse during the past 12 months. Regular care was defined as always, usually, or sometimes receiving care from family or friends because of SCD. Using the 2008 Physical Activity Guidelines for Americans, respondents were classified as being inactive, insufficiently active, or sufficiently active based on their reported aerobic exercise. We calculated weighted proportions and used chi-square tests for differences across categories by SCD status and receipt of care. We estimated the prevalence ratio (PR) for being inactive, insufficiently active, and sufficiently active using separate log-binomial regression models, adjusting for covariates. 12.3% of respondents reported SCD and 23.1% of those with SCD received regular care. 29.6% (95%CI: 28.9-30.4) of respondents without SCD were inactive compared to 37.1% (95%CI: 34.7-39.5) of those with SCD who did not receive regular care and 50.2% (95%CI: 45.2-55.1) of those with SCD who received regular care. 52.4% (95%CI: 51.6-53.2) of respondents without SCD were sufficiently active compared to 46.4% (95%CI: 43.8-49.0) of respondents with SCD and received no regular care and 30.6% (95%CI: 26.1-35.6) of respondents with SCD who received regular care. After adjusting for demographic and health status differences, people receiving regular care for SCD had a significantly lower prevalence of meeting aerobic guidelines compared to people without SCD (PR=0.80, 95%CI: 0.69-0.93, p=0.005). The most prevalent physical activity was walking for adults aged ≥ 45 years old (41-52%) regardless of SCD status or receipt of care. Overall, the prevalence of inactivity was high, especially among people with SCD. These findings suggest a need to increase activity among middle-aged and older adults, particularly those with SCD who receive care. Examining ways to increase walking, potentially by involving informal caregivers, could be a promising way for people with SCD to reduce inactivity and gain the health benefits associated with meeting physical activity guidelines.


Identification of the molecular make-up of the Potato virus Y strain PVY(Z): genetic typing of PVY(Z)-NTN.
Potato virus Y (PVY) strains were originally defined by interactions with different resistance genes in standard potato cultivars. Five distinct strain groups are defined that cause local or systemic hypersensitive responses (HRs) in genetic background with a corresponding N gene: PVY(O), PVY(N), PVY(C), PVY(Z), and PVY(E). The nucleotide sequences of multiple isolates of PVY(O) and PVY(N) differ from each other by ≈8% along their genomes. Additionally, complete genome sequences of multiple recombinant isolates are composed of segments of parental PVY(O) and PVY(N) sequences. Here, we report that recombinant isolate PVY-L26 induces an HR in potato 'Maris Bard' carrying the putative Nz gene, and is not recognized by two other resistance genes, Nc and Ny(tbr). These genetic responses in potato, combined with the inability of PVY-L26 to induce vein necrosis in tobacco, clearly define it as an isolate from the PVY(Z) strain group and provide the first information on genome structure and sequence of PVY(Z). The genome of PVY-L26 displays typical features of European NTN-type isolates with three recombinant junctions (PVY(EU-NTN)), and the PVY-L26 is named PVY(Z)-NTN. Three typical PVY(NTN) isolates and two PVY(N) isolates, all inducing vein necrosis in tobacco, were compared with PVY-L26. One PVY(NTN) isolate elicited HR reactions in Maris Bard, similar to PVY-L26, while two induced a severe systemic HR-like reaction quite different from the quasi-symptomless reaction induced by two PVY(N) isolates. 'Yukon Gold' potato from North America produced HR against several PVY(NTN) isolates, including PVY-L26, but only late and limited systemic necrosis against one PVY(N) isolate. Consequently, according to symptoms in potato indicators, both PVY(Z) and PVY(NTN) isolates appeared biologically very close and clearly distinct from PVY(O) and PVY(N) strain groups.


Computational biology approaches for mapping transcriptional regulatory networks.
Transcriptional Regulatory Networks (TRNs) are mainly responsible for the cell-type- or cell-state-specific expression of gene sets from the same DNA sequence. However, so far there are no precise maps of TRNs available for each cell-type or cell-state, and no ideal tool to map those networks clearly and in full from biological samples. In this review, major approaches and tools to map TRNs from high-throughput data are presented, depending on the type of methods or data used to infer them, and their advantages and limitations are discussed. After summarizing the main principles defining the topology and structure–function relationships in TRNs, an overview of the extensive work done to map TRNs from bulk transcriptomic data will be presented by type of methodological approach. Most recent modellings of TRNs using other types of molecular data or integrating different data types, including single-cell RNA-sequencing and chromatin information, will then be discussed, before briefly concluding with improvements expected to come in the field.


Testing multiple levels of influence in the intergenerational transmission of alcohol disorders from a developmental perspective: the example of alcohol use promoting peers and μ-opioid receptor M1 variation.
This study examined the interplay between the influence of peers who promote alcohol use and μ-opioid receptor M1 (OPRM1) genetic variation in the intergenerational transmission of alcohol use disorder (AUD) symptoms while separating the "traitlike" components of AUD symptoms from their age-specific manifestations at three ages from emerging adulthood (17-23 years) to adulthood (29-40 years). The results for males were consistent with genetically influenced peer selection mechanisms as mediators of parent alcoholism effects. Male children of alcoholics were less likely to be carriers of the G allele in single nucleotide polymorphism A118G (rs1799971), and those who were homozygous for the A allele were more likely to affiliate with alcohol use promoting peers who increased the risk for AUD symptoms at all ages. There was evidence for women of an interaction between OPRM1 variation and peer affiliations but only at the earliest age band. Peer influences had stronger effects among women who were G-carriers. These results illustrate the complex ways in which the interplay between influences at multiple levels of analysis can underlie the intergenerational transmission of alcohol disorders as well as the importance of considering age and gender differences in these pathways.


Evaluation of antibiotic prescribing for adult inpatients at Sultan Qaboos University Hospital, Sultanate of Oman.
Little is known into the prudent use of antibiotics in hospitals in Oman. This study is to evaluate antibiotic prescribing by measuring the overall compliance with the local antibiotic prescribing guidelines. An observational study involving 366 patients' admission episodes as determined by power analysis on patients (≥18 years) on oral and/or parenteral antibiotic during admission, in the period of 10 weeks (1 February-15 April, 2014). The adapted audit tool of the Barking, Havering and Redbridge University Hospitals NHS Trust was used for this study. Analyses were performed using descriptive statistics. Main outcome measures: antibiotic prescribing compliance with the local guidelines as well as the overall restricted antibiotic policy adherence at Sultan Qaboos University Hospital (SQUH). The number of prescribed and audited antibiotics totalled 825, compliance with local guidelines was suboptimal at 63% (n=520), and of 211 restricted antibiotics prescribed, the overall adherence to restricted antibiotic policy was inadequate at 46% (n=98). The majority of the antibiotics prescribed were broad spectrum at 90% (n=739), mainly penicillins at 31% (n=256) and cephalosporins at 17% (n=139). The study has provided valuable baseline details of antibiotic prescribing patterns in SQUH. The diagnosis was documented in 89% (n=327) of the admission episodes. However, the compliance with SQUH antibiotic prescribing guidelines was suboptimal, and the overall compliance with SQUH restricted antibiotic guidelines was in 46% of the prescriptions. Further studies are required to address the reasons behind the non-compliance with local guidelines.


3D CT imaging method for measuring temporal bone aeration.
3D volume reconstruction of CT images can be used to measure temporal bone aeration. This study evaluates the technique with respect to reproducibility and acquisition parameters. Helical CT images acquired from patients with radiographically normal temporal bones using standard clinical protocols were retrospectively analyzed. 3D image reconstruction was performed to measure the volume of air within the temporal bone. The appropriate threshold values for air were determined from reconstruction of a phantom with a known air volume imaged using the same clinical protocols. The appropriate air threshold values were applied to the clinical material. Air volume was measured according to an acquisition algorithm. The average volume in the temporal bone CT group was 5.56 ml, compared to 5.19 ml in the head CT group (p = 0.59). The correlation coefficient between examiners was > 0.92. There was a wide range of aeration volumes among individual ears (0.76-18.84 ml); however, paired temporal bones differed by an average of just 1.11 ml. The method of volume measurement from 3D reconstruction reported here is widely available, easy to perform and produces consistent results among examiners. Application of the technique to archival CT data is possible using corrections for air segmentation thresholds according to acquisition parameters.


Influence of cryopreservation on the cytosine methylation state of potato genomic NDA.
Shoot tips of Solanum tuberosum (Désirée) were successfully cryopreserved by the DMSO droplet method and stored for almost 7 years, while control material was maintained in vitro for the same period of time. To analyse potential epigenetic changes, the DNA methylation status was assayed by methylation-sensitive amplified polymorphism (MSAP) analysis using restriction endonucleases MspI and HpaII. An amount of 93.6% of the analysed MSAP signals were stable among all cryopreserved and in vitro maintained samples tested, indicating extensive stability of DNA methylation. Only 0.9 % of MSAP signals showed results that differed between the two treatments and at the same time matched for all three biological replications within each treatment. These can be seen as indicating directed effects of the two treatments on the DNA methylation. Cryopreserved samples displayed in comparison to in vitro stored samples consistent hypomethylation for 0.6 % (3 of 469) of MSAP signals (Table 4, pattern 4) and consistent hypermethylation for 0.2% (1 of 469), respectively. For 5.6% of all MSAP signals, inconsistent results were observed among the three biological replications at least for one of the two treatments. These were interpreted as resulting from stochastic DNA methylation changes in individual samples. As results for two biological replications were identical and different from the result for the third biological replication, the direction of methylation change could be determined in those cases. Cases of stochastic loss of CG methylation in cryopreserved samples were most frequent among them, adding up to 3.4% of MSAP signals. Stochastic loss of CG methylation was also found in material maintained in vitro, only for 0.6% of all MSAP signals. In conclusion, methylation changes occurred in long-term cryopreservation of potato, in a random rather than directed fashion. Hence, cryopreservation and long-term in vitro maintenance both induce limited changes of DNA methylation status. The order of magnitude of methylation changes observed was consistent with other studies, where similar rates of DNA methylation changes have been found.


Localization of laminin B1 mRNA in retinal ganglion cells by in situ hybridization.
In the nervous system, neuronal migration and axonal growth are dependent on specific interactions with extracellular matrix proteins. During development of the vertebrate retina, ganglion cell axons extend along the internal limiting (basement) membrane and form the optic nerve. Laminin, a major component of basement membranes, is known to be present in the internal limiting membrane, and might be involved in the growth of ganglion cell axons. The identity of the cells that produce retinal laminin, however, has not been established. In the present study, we have used in situ hybridization to localize the sites of laminin B1 mRNA synthesis in the developing mouse retina. Our results show that there are at least two principal sites of laminin B1 mRNA synthesis: (a) the hyaloid vessels and the lens during the period of major axonal outgrowth, and (b) the retinal ganglion cells at later development stages. Müller (glial) cells, the major class of nonneuronal cells in the retina, do not appear to express laminin B1 mRNA either during development or in the adult retina. In Northern blots, we found a single transcript of approximately 6-kb size that encodes the laminin B1 chain in the retina. Moreover, laminin B1 mRNA level was four- to fivefold higher in the postnatal retina compared to that in the adult. Our results show that in addition to nonneuronal cells, retinal ganglion cells also synthesize laminin. The function of laminin in postnatal retinas, however, remains to be elucidated. Nevertheless, our findings raise the possibility that neurons in other parts of the nervous system might also synthesize extracellular matrix proteins.


Increased biomass yield of Lactococcus lactis during energetically limited growth and respiratory conditions.
Lactococcus lactis is known to be capable of respiration under aerobic conditions in the presence of haemin. In the present study the effect of respiration on ATP production during growth on different sugars was examined. With glucose as the sole carbon source, respiratory conditions in L. lactis MG1363 resulted in only a minor increase, 21%, in biomass yield. Since ATP production through substrate-level phosphorylation was essentially identical with and without respiration, the increased biomass yield was a result of energy-saving under respiratory conditions estimated to be 0.4 mol of ATP/mol of glucose. With maltose as the energy source, the increase in biomass yield amounted to 51% compared with an aerobic culture that lacked haemin. This higher ATP yield was obtained by redirecting pyruvate metabolism from lactate to acetate production, and from savings through respiration. However, even after subtracting these contributions, approx. 0.3 mol of ATP/mol of glucose remained unaccounted for. A similar response to respiratory conditions (0.2 mol of ATP/mol of glucose) was observed in a mutant that had a decreased glucose uptake rate during growth on glucose caused by disruption of the PTS(mannose) (glucose/mannose-specific phosphotransferase system). Amino acid catabolism could be excluded as the source of the additional ATP. Since mutants without a functional H+-ATPase produced less ATP under sugar starvation and respiratory conditions, the additional ATP yield appears to come partly from energy saved on proton pumping through the H+-ATPase due to respiration and partly from a reversed function of the H+-ATPase towards oxidative phosphorylation. These results may contribute to the design and implementation of carbon-efficient high-cell-density cultures of this industrially important species of bacterium.


Characterization of the angR gene of Vibrio anguillarum: essential role in virulence.
The ability to utilize the iron bound by high-affinity iron-binding proteins in the vertebrate host is an important virulence factor for the marine fish pathogen Vibrio anguillarum. Virulence in septicemic infections is due to the presence of a highly efficient plasmid-encoded iron transport system. AngR, a 110-kDa protein component of this system, appears to play a role in both regulation of the expression of the iron transport genes fatDCBA and the production of the siderophore anguibactin. Therefore, study of the expression of the angR gene and the properties of its product, the AngR protein, may contribute to the understanding of the mechanisms of virulence of this pathogen. In this work, we present genetic and molecular evidence from transposition mutagenesis experiments and RNA analysis that angR, which maps immediately downstream of the fatA gene, is part of a polycistronic transcript that also includes the iron transport genes fatDCBA and angT, a gene located downstream of angR which showed domain homology to certain thioesterases involved in nonribosomal peptide synthesis of siderophores and antibiotics. In order to dissect the specific domains of AngR associated with regulation of iron transport gene expression, anguibactin production, and virulence, we also generated a panel of site-directed angR mutants, as well as deletion derivatives. Both virulence and anguibactin production were dramatically affected by each one of the angR modifications. In contrast to the need for an intact AngR molecule for anguibactin production and virulence, the regulation of iron transport gene expression does not require the entire AngR molecule, since truncation of the carboxy terminus carrying the nonribosomal peptide synthetase cores, as well as the site-directed mutations, resulted in derivatives that retained their ability to regulate gene expression which was only abolished after truncation of amino-terminal sequences containing helix-turn-helix and leucine zipper motifs and a specialized heterocyclization and condensation domain found in certain nonribosomal peptide synthetases. The evidence, while not rigorously eliminating the possibility that a separate regulatory polypeptide exists and is encoded somewhere within the 5'-end region of the angR gene, strongly supports the idea that AngR is a bifunctional protein and that it plays an essential role in the virulence mechanisms of V. anguillarum. We also show in this study that the angT gene, found downstream of angR, intervenes in the mechanism of anguibactin production but is not essential for virulence or iron transport gene expression.


Photoinduced Force Mapping of Plasmonic Nanostructures.
The ability to image the optical near-fields of nanoscale structures, map their morphology, and concurrently obtain spectroscopic information, all with high spatiotemporal resolution, is a highly sought-after technique in nanophotonics. As a step toward this goal, we demonstrate the mapping of electromagnetic forces between a nanoscale tip and an optically excited sample consisting of plasmonic nanostructures with an imaging platform based on atomic force microscopy. We present the first detailed joint experimental-theoretical study of this type of photoinduced force microscopy. We show that the enhancement of near-field optical forces in gold disk dimers and nanorods follows the expected plasmonic field enhancements with strong polarization sensitivity. We then introduce a new way to evaluate optically induced tip-sample forces by simulating realistic geometries of the tip and sample. We decompose the calculated forces into in-plane and out-of-plane components and compare the calculated and measured force enhancements in the fabricated plasmonic structures. Finally, we show the usefulness of photoinduced force mapping for characterizing the heterogeneity of near-field enhancements in precisely e-beam fabricated nominally alike nanostructures - a capability of widespread interest for precise nanomanufacturing, SERS, and photocatalysis applications.


Breast cancer-related lymphedema.
Every year in the United States, breast cancer is diagnosed in more than 200,000 women. Because of the prevalence of breast cancer, treatment-related sequelae are of Importance to many survivors of the disease. One such sequela is upper extremity lymphedema, which occurs when fluid accumulates in the Interstitial space and causes enlargement and usually a feeling of heaviness in the limb. Axillary surgery contributes considerably to the incidence of lymphedema, with the incidence and severity of swelling related to the number of lymph nodes removed. Lymphedema after standard axillary lymph node dissection can occur in up to approximately 50% of patients. However, the risk of lymphedema is decreased substantially with newer sentinel lymph node sampling procedures. Adjuvant radiotherapy to the breast or lymph nodes increases the risk of lymphedema, which has been reported in 9% to 40% of these patients. Management of lymphedema requires a multidisciplinary approach to minimize the effect on the patient's quality of life. This review presents an overview of the pathophysiology, diagnosis, prevention, and treatment of breast cancer-related lymphedema.


A multi-center screening trial of rasagiline in patients with amyotrophic lateral sclerosis: Possible mitochondrial biomarker target engagement.
Rasagiline, a monoamine oxidase B inhibitor, slowed disease progression in the SOD1 mouse, and in a case series of patients with amyotrophic lateral sclerosis (ALS). Here we determine whether rasagiline is safe and effective in ALS compared to historical placebo controls, and whether it alters mitochondrial biomarkers. We performed a prospective open-label, multicenter screening trial of 36 ALS patients treated with 2 mg oral rasagiline daily for 12 months. Outcomes included the slope of deterioration of the revised ALS Functional Rating Scale (ALSFRS-R), adverse event monitoring, time to treatment failure, and exploratory biomarkers. Participants experienced no serious drug-related adverse events, and the most common adverse event was nausea (11.1%). Rasagiline did not improve the rate of decline in the ALSFRS-R; however, differences in symptom duration compared to historical placebo controls differentially affected ALSFRS-R slope estimates. Rasagiline changed biomarkers over 12 months, such that the mitochondrial membrane potential increased (JC-1 red/green fluorescent ratio 1.92, p = 0.0001) and apoptosis markers decreased (Bcl-2/Bax ratio 0.24, p < 0.0001). In conclusion, engagement of exploratory biomarkers and questions about comparability of baseline characteristics lead us to recommend a further placebo-controlled trial.


Air travel and pneumothorax.
The number of medical emergencies onboard aircraft is increasing as commercial air traffic increases and the general population ages, becomes more mobile, and includes individuals with serious medical conditions. Travelers with respiratory diseases are at particular risk for in-flight events because exposure to lower atmospheric pressure in a pressurized cabin at cruising altitude may result in not only hypoxemia but also pneumothorax due to gas expansion within enclosed pulmonary parenchymal spaces based on Boyle's law. Risks of pneumothorax during air travel pertain particularly to those patients with cystic lung diseases, recent pneumothorax or thoracic surgery, and chronic pneumothorax. Currently available guidelines are admittedly based on sparse data and include recommendations to delay air travel for 1 to 3 weeks after thoracic surgery or resolution of the pneumothorax. One of these guidelines declares existing pneumothorax to be an absolute contraindication to air travel although there are reports of uneventful air travel for those with chronic stable pneumothorax. In this article, we review the available data regarding pneumothorax and air travel that consist mostly of case reports and retrospective surveys. There is clearly a need for additional data that will inform decisions regarding air travel for patients at risk for pneumothorax, including those with recent thoracic surgery and transthoracic needle biopsy.


Shear stress induces ATP-independent transient nitric oxide release from vascular endothelial cells, measured directly with a porphyrinic microsensor.
Shear stress causes the vascular endothelium to release nitric oxide (NO), which is an important regulator of vascular tone. However, direct measurement of NO release after the imposition of laminar flow has not been previously accomplished because of chemical (oxidative degradation) and physical (diffusion, convection, and washout) complications. Consequently, the mechanism, time course, kinetics, and Ca2+ dependence of NO release due to shear stress remain incompletely understood. In this study, we characterized these parameters by using fura 2 fluorescence and a polymeric porphyrin/Nafion-coated carbon fiber microsensor (detection limit, 5 nmol/L; response time, 1 millisecond) to directly measure changes in [Ca2+]i and NO release due to shear stress or agonist (ATP or brominated Ca2+ ionophore [Br-A23187]) from bovine aortic endothelial cells. The cells were grown to confluence on glass coverslips, loaded with fura 2-AM, and mounted in a parallel-plate flow chamber (volume, 25 microL). The microsensor was positioned approximately 100 microns above the cells with its long axis parallel to the direction of flow. Laminar flow of perfusate was maintained from 0.04 to 1.90 mL/min, which produced shear stresses of 0.2 to 10 dyne/cm2. Shear stress caused transient NO release 3 to 5 seconds after the initiation of flow and 1 to 3 seconds after the rise in [Ca2+]i, which reached a plateau after 35 to 70 seconds. Although the amount (peak rate) of NO release increased as a function of the shear stress (0.08 to 3.80 pmol/s), because of the concomitant increase in the flow rate, the peak NO concentration (133 +/- 9 nmol/L) remained constant. Maintenance of flow resulted in additional transient NO release, with peak-to-peak intervals of 15.5 +/- 2.5 minutes. During this 13- to 18-minute period, when the cells were unresponsive to shear stress, exogenous ATP (10 mumol/L) or Br-A23187 (10 mumol/L) evoked NO release. Prior incubation of the cells with exogenous NO or the removal and EGTA (100 mumol/L) chelation of extracellular Ca2+ blocked shear stress but not ATP-dependent NO release. The kinetics of shear stress-induced NO release (2.23 +/- 0.07 nmol/L per second) closely resembled the kinetics of Ca2+ flux but differed markedly from the kinetics of ATP-induced NO release (5.64 +/- 0.32 nmol/L per second). These data argue that shear stress causes a Ca(2+)-mediated ATP-independent transient release of NO, where the peak rate of release but not the peak concentration depends on the level of shear stress.(ABSTRACT TRUNCATED AT 400 WORDS)


A three-year-old boy with X-linked adrenoleukodystrophy and congenital pulmonary adenomatoid malformation: a case report.
X-linked adrenoleukodystrophy leads to demyelination of the nervous system, adrenal insufficiency, and accumulation of long-chain fatty acids. Most young patients with X-linked adrenoleukodystrophy develop seizures and progressive neurologic deficits, and die within the first two decades of life. Congenital or acquired disorders of the respiratory system have not been previously described in patients with X-linked adrenoleukodystrophy. A 3-year-old Arabic boy from Yemen presented with discoloration of the mucous membranes and nail beds, which were considered cyanoses due to methemoglobinemia. He also had shortness of breath, fatigue, emesis and dehydration episodes for which he was admitted to our hospital. Chest radiograph and chest computed tomography scans showed congenital pulmonary adenomatoid malformation. A few weeks before the removal of the malformation, he had a significant episode of hypotension and hypoglycemia. This development required further in-hospital evaluation that led to the diagnosis of adrenal insufficiency and the initiation of treatment with corticosteroids. One year later, he developed seizures and loss of consciousness. Magnetic resonance imaging of his head showed diffuse demyelination secondary to X-linked adrenoleukodystrophy. He was treated with anti-seizure and anti-oxidants, and was referred for bone marrow transplant evaluation. The presence of adrenal insufficiency, neurologic deficits and seizures are common manifestations of X-linked adrenoleukodystrophy. The association of congenital lung disease with X-linked adrenoleukodystrophy or Addison's disease has not been described previously.


New Insights Into Cryptococcus Spp. Biology and Cryptococcal Meningitis.
Defective cell-mediated immunity is a major risk factor for cryptococcosis, a fatal disease if untreated. Cryptococcal meningitis (CM), the main presentation of disseminated disease, occurs through hematogenous spread to the brain from primary pulmonary foci, facilitated by yeast virulence factors. We revisit remarkable recent improvements in the prevention, diagnosis and management of CM. Cryptococcal antigen (CrAg), main capsular polysaccharide of Cryptococcus spp. is detectable in blood and cerebrospinal fluid of infected patients with point of care lateral flow assays. Recent World Health Organization guidelines recommend 7-day amphotericin B plus flucytosine, then 7-day high dose (1200 mg/day) fluconazole for induction treatment of HIV-associated CM. Management of raised intracranial pressure, a consequence of CM, should rely mainly on daily therapeutic lumbar punctures until normalisation. In HIV-associated CM, following introduction of antifungal therapy, (re)initiation of antiretroviral therapy should be delayed by 4-6 weeks to prevent immune reconstitution inflammatory syndrome, common in CM. CM is a fatal disease whose diagnosis has recently been simplified. Treatment should always include antifungal combination therapy and management of raised intracranial pressure. Screening for immune deficiency should be mandatory in all patients with cryptococcosis.


Cancer incidence and mortality among flight personnel: a meta-analysis.
Increased cancer risk among flight personnel have previously been noted, including breast cancer among flight attendants and acute myeloid leukemia among pilots. Exposure to cosmic radiation and other physical or chemical agents may pose health risks for flight personnel. We performed an exhaustive search for published and unpublished cohort studies of flight personnel from 1986-98. We combined relative risks (RR) for selected causes from four mortality and/or incidence studies of pilots and two incidence studies of flight attendants, using standard meta-analytic methods. Heterogeneity among the combined studies was explored and adjustments were made for possible confounding by socioeconomic status (SES), where indicated, using correction factors from published studies. SES-adjusted combined RRs were elevated (>1.2) among male pilots for mortality from melanoma 11.97 (95%, CI: 1.02-3.82)] and brain cancer [1.49 (0.89-2.20)], and for cancer incidence of the prostate [1.65 (1.19-2.29)] and the brain [1.74 (0.87-3.30)]. Among female flight attendants, increases were seen for incidence of all cancers [1.29 (0.98-1.70)], melanoma [11.54 (0.83-2.87)], and breast cancer [1.35 (1.00-1.83)]. Flight personnel appear to be at increased risk for several types of cancer. Both occupational exposures and well-established non-occupational risk factors may contribute to this increased risk. To better control for confounding factors and to identify exposures potentially amenable to preventive measures, future studies should compare risks within cohorts by flight routes, work history, and exposure to cosmic and UV radiation, electromagnetic fields, and chemical substances.


To Infuse or Ingest in Human Laboratory Alcohol Research.
Human alcohol laboratory studies use two routes of alcohol administration: ingestion and infusion. The goal of this paper was to compare and contrast these alcohol administration methods. The work summarized in this report was the basis of a 2019 Research Society on Alcoholism Roundtable, "To Ingest or Infuse: A Comparison of Oral and Intravenous Alcohol Administration Methods for Human Alcohol Laboratory Designs." We review the methodological approaches of each and highlight strengths and weaknesses pertaining to different research questions. We summarize methodological considerations to aid researchers in choosing the most appropriate method for their inquiry, considering exposure variability, alcohol expectancy effects, safety, bandwidth, technical skills, documentation of alcohol exposure, experimental variety, ecological validity, and cost. Ingestion of alcohol remains a common and often a preferable, methodological practice in alcohol research. Nonetheless, the main problem with ingestion is that even the most careful calculation of dose and control of dosing procedures yields substantial and uncontrollable variability in the participants' brain exposures to alcohol. Infusion methodologies provide precise exposure control but are technically complex and may be limited in ecological validity. We suggest that alcohol ingestion research may not be the same thing as alcohol exposure research; investigators should be aware of the advantages and disadvantages that the choice between ingestion and infusion of alcohol invokes.


Maternal inflammation at midgestation impairs subsequent fetal myoblast function and skeletal muscle growth in rats, resulting in intrauterine growth restriction at term.
Maternal inflammation induces intrauterine growth restriction (MI-IUGR) of the fetus, which compromises metabolic health in human offspring and reduces value in livestock. The objective of this study was to determine the effect of maternal inflammation at midgestation on fetal skeletal muscle growth and myoblast profiles at term. Pregnant Sprague-Dawley rats were injected daily with bacterial endotoxin (MI-IUGR) or saline (controls) from the 9th to the 11th day of gestational age (dGA; term = 21 dGA). At necropsy on dGA 20, average fetal mass and upper hindlimb cross-sectional areas were reduced (P < 0.05) in MI-IUGR fetuses compared with controls. MyoD+ and myf5+ myoblasts were less abundant (P < 0.05), and myogenin+ myoblasts were more abundant (P < 0.05) in MI-IUGR hindlimb skeletal muscle compared with controls, indicating precocious myoblast differentiation. Type I and Type II hindlimb muscle fibers were smaller (P < 0.05) in MI-IUGR fetuses than in controls, but fiber type proportions did not differ between experimental groups. Fetal blood plasma TNFα concentrations were below detectable amounts in both experimental groups, but skeletal muscle gene expression for the cytokine receptors TNFR1, IL6R, and FN14 was greater (P < 0.05) in MI-IUGR fetuses than controls, perhaps indicating enhanced sensitivity to these cytokines. Maternal blood glucose concentrations at term did not differ between experimental groups, but MI-IUGR fetal blood contained less (P < 0.05) glucose, cholesterol, and triglycerides. Fetal-to-maternal blood glucose ratios were also reduced (P < 0.05), which is indicative of placental insufficiency. Indicators of protein catabolism, including blood plasma urea nitrogen and creatine kinase, were greater (P < 0.05) in MI-IUGR fetuses than in controls. From these findings, we conclude that maternal inflammation at midgestation causes muscle-centric fetal programming that impairs myoblast function, increases protein catabolism, and reduces skeletal muscle growth near term. Fetal muscle sensitivity to inflammatory cytokines appeared to be enhanced after maternal inflammation, which may represent a mechanistic target for improving these outcomes in MI-IUGR fetuses.


A bidirectional YAC walk from the Norrie disease (NDP) locus.
The region of Xp between DXS7 and the centromere contains the gene for Norrie disease in addition to the genes for several other ophthalmic disorders. A 650-kb YAC containing the loci MAOA, MAOB, and NDP has been used as the starting point for a bidirectional chromosomal walk. A contig of 16 YACs covering between 2 and 3 Mb has been developed in which the following markers/genes are located (in physical order): Xpter--DXS1201 (256ze5)--DXS6668--DXS228--DXS77--MAOA--++ +MAOB--FR12 (pseudogene)--NDP--DXS6670--RRM2P3--DXS6671--DXS742 --Xcen. Seven new STSs are described both for end clones and for internal Alu PCR products from the contig. The contig contains the breakpoint of the t75-2ma-1b (t75) translocation, close to the 5' end of the MAOB gene.


Applying species--energy theory to conservation: a case study for North American birds.
Ecosystem energy is now recognized as a primary correlate and potential driver of global patterns of species richness. The increasingly well-tested species-energy relationship is now ripe for application to conservation, and recent advances in satellite technology make this more feasible. While the correlates for the species-energy relationship have been addressed many times previously, this study is among the first to apply species-energy theory to conservation. Our objectives were to: (1) determine the strongest model of bird richness across North America; (2) determine whether the slope of the best species-energy model varied with varying energy levels; and (3) identify the spatial patterns with similar or dissimilar slopes to draw inference for conservation. Model selection techniques were used to evaluate relationships between Moderate Resolution Imaging Spectroradiometer (MODIS) measures of ecosystem energy and species richness of native land birds using the USGS Breeding Bird Survey (BBS) data. Linear, polynomial, and break point regression techniques were used to evaluate the shape of the relationships with correction for spatial autocorrelation. Spatial analyses were used to determine regions where slopes of the relationship differed. We found that annual gross primary production (GPP) was the strongest correlate of richness (adjusted R2 = 0.55), with a quadratic model being the strongest model. The negative slope of the model was confirmed significantly negative at the highest energy levels. This finding demonstrates that there are three different slopes to the species-energy relationship across the energy gradient of North America: positive, flat, and negative. If energy has a causal relationship with richness, then species-energy theory implies that energy causes richness to increase in low-energy areas, energy has little effect in intermediate-energy areas, and energy depresses richness in the highest-energy areas. This information provides a basis for potential applications for more effective conservation. For example, in low-energy areas, increased nutrients could improve vegetation productivity and increase species richness. In high-energy areas where competitive dominance of vegetation might reduce species richness, vegetation manipulation could increase species richness. These strategies will likely be most effective if tailored to the local energy gradient.


Prevention of postpartum haemorrhage by community-based auxiliary midwives in hard-to-reach areas of Myanmar: a qualitative inquiry into acceptability and feasibility of task shifting.
In Myanmar, postpartum haemorrhage is the leading cause of maternal mortality and contributes to around 30% of all maternal deaths. The World Health Organization recommends training and supporting auxiliary midwives to administer oral misoprostol for prevention of postpartum haemorrhage in resource-limited settings. However, use of misoprostol by auxiliary midwives has not formally been approved in Myanmar. Our study aimed to explore community and provider perspectives on the roles of auxiliary midwives and community-level provision of oral misoprostol by auxiliary midwives. A qualitative inquiry was conducted in Ngape Township, Myanmar. A total of 15 focus group discussions with midwives, auxiliary midwives, community members and mothers with children under the age of three were conducted. Ten key informant interviews were performed with national, district and township level health planners and implementers of maternal and child health services. All audio recordings were transcribed verbatim in Myanmar language. Transcripts of focus group discussions were fully translated into English before coding, while key informants' data were coded in Myanmar language. Thematic analysis was done using ATLAS.ti software. Home births are common and auxiliary midwives were perceived as an essential care provider during childbirth in hard-to-reach areas. Main reasons provided were that auxiliary midwives are more accessible than midwives, live in the hard-to-reach areas, and are integrated in the community and well connected with midwives. Auxiliary midwives generally reported that their training involved instruction on active management of the third stage of labour, including use of misoprostol, but not all auxiliary midwives reported using misoprostol in practice. Supportive reasons for task-shifting administration of oral misoprostol to auxiliary midwives included discussions around the good relationship and trust between auxiliary midwives and midwives, whereby midwives felt confident distributing misoprostol to auxiliary midwives. However, the lack of clear government-level written permission to distribute the drug was perceived as a barrier to task shifting. This study highlights the acceptability of misoprostol use by auxiliary midwives to prevent postpartum haemorrhage, and findings suggest that it should be considered as a promising intervention for task shifting in Myanmar.


Global regulation of H2A.Z localization by the INO80 chromatin-remodeling enzyme is essential for genome integrity.
INO80 is an evolutionarily conserved, ATP-dependent chromatin-remodeling enzyme that plays roles in transcription, DNA repair, and replication. Here, we show that yeast INO80 facilitates these diverse processes at least in part by controlling genome-wide distribution of the histone variant H2A.Z. In the absence of INO80, H2A.Z nucleosomes are mislocalized, and H2A.Z levels at promoters show reduced responsiveness to transcriptional changes, suggesting that INO80 controls H2A.Z dynamics. Additionally, we demonstrate that INO80 has a histone-exchange activity in which the enzyme can replace nucleosomal H2A.Z/H2B with free H2A/H2B dimers. Genetic interactions between ino80 and htz1 support a model in which INO80 catalyzes the removal of unacetylated H2A.Z from chromatin as a mechanism to promote genome stability.


Research ethics and the medical profession. Report of the Advisory Committee on Human Radiation Experiments.
The Advisory Committee on Human Radiation Experiments was convened by President Clinton in January 1994 in response to allegations of unethical practices in radiation experiments involving human subjects that were sponsored by the US government between 1944 and 1974. The committee's Final Report was released in October 1995. In addition to analyzing the history of the ethics of medical research involving human subjects, the committee reviewed current federal policies and procedures for protection of human subjects. In this article, the committee's findings are discussed as they relate to the patient-physician relationship, the issue of trust, and the specific role of the physician-investigator in all types of human experimentation. The committee found evidence of discussion of the conduct of human research at the highest levels of the government and within the medical profession, particularly with regard to risk, during the 1940s and 1950s. However, in both federal policy and professional practice, requirements for consent were more likely to apply to "healthy volunteers" than to patient-subjects (ie, those with disease or illness). Today, consensus exists that duties to obtain informed consent apply to all human subjects, whether healthy or sick, regardless of the risk or potential for medical benefit from participation in the research and regardless of the nature of sponsorship or funding (eg, federal, military, or private). Based on a finding of serious deficiencies in the current system of protections for human subjects, the committee offers a number of recommendations, including changes in institutional review boards; in the interpretation of ethics rules and policies; in oversight, accountability, and sanctions for ethics violations; and in compensation for research injuries. More than public policy changes, however, the committee recommends that the medical profession intensify its commitment to the ethics of research involving human subjects.


Effect of the histamine (H2) inhibitor metiamide on histamine-stimulated bile flow in dogs.
The effects of the histamine H2-receptor inhibitor metiamide on histamine-stimulated canine bile flow and gastric hydrogen ion output were evaluated. Histamine was found to stimulate bile volume in doses comparable to those that stimulated gastric hydrogen ion output; both responses appeared to have the same maximal response dose, 150 mug/kg per h. Metiamide alone did not alter hepatic bile flow. Administration of metiamide, 2 mg/kg per h, along with various doses of histamine demonstrated that the H2-receptor antagonist decreased bile volume and gastric hydrogen ion output from values obtained with histamine administration alone. The D50 of histamine for bile flow was 16.3 mug/kg per h and the D50 for hydrogen ion output was 44.2 mug/kg per h, Kinetic analysis suggests that the decrease in histamine-stimulated hydrogen ion output produced by metiamide is the result of competitive inhibition; the decrease in histamine-stimulated bile volume by metiamide which is different from the hydrogen ion inhibition, suggests noncompetitive inhibition. These data indicate that the mechanism of histamine choleresis is different from the mechanism of histamine-stimulated gastric acid output and that histamine-stimulated bile flow may not be the result of direct hormone-receptor interaction.


Associations between Repeated Measures of Maternal Urinary Phthalate Metabolites and Thyroid Hormone Parameters during Pregnancy.
Maintaining thyroid homeostasis during pregnancy is essential for normal fetal growth and development. Growing evidence suggests that phthalates interfere with normal thyroid function. Few human studies have investigated the degree to which phthalates may affect thyroid hormone levels in particularly susceptible populations such as pregnant women. We examined the associations between repeated measures of urinary phthalate metabolites and plasma thyroid hormone levels in samples collected at up to four time points per subject in pregnancy. Additionally, we investigated the potential windows of susceptibility to thyroid hormone disturbances related to study visit of sample collection. Data were obtained from pregnant women (n = 439) participating in a nested case-control study of preterm birth with 116 cases and 323 controls. We measured 9 phthalate metabolite concentrations in urine samples collected at up to four study visits per subject during pregnancy (median = 10, 18, 26, and 35 weeks of gestation, respectively). We also measured a panel of thyroid function markers in plasma collected at the same four time points per subject during pregnancy. Although our results were generally null, in repeated measures analyses we observed that phthalate metabolites were largely inversely associated with thyrotropin and positively associated with free and total thyroid hormones. Cross-sectional analyses by study visit revealed that the magnitude and/or direction of these relationships varied by timing of exposure during gestation. These results support previous reports showing the potential for environmental phthalate exposure to alter circulating levels of thyroid hormones in pregnant women. Citation: Johns LE, Ferguson KK, McElrath TF, Mukherjee B, Meeker JD. 2016. Associations between repeated measures of maternal urinary phthalate metabolites and thyroid hormone parameters during pregnancy. Environ Health Perspect 124:1808-1815; http://dx.doi.org/10.1289/EHP170.


Programmed death 1 protects from fatal circulatory failure during systemic virus infection of mice.
The inhibitory programmed death 1 (PD-1)-programmed death ligand 1 (PD-L1) pathway contributes to the functional down-regulation of T cell responses during persistent systemic and local virus infections. The blockade of PD-1-PD-L1-mediated inhibition is considered as a therapeutic approach to reinvigorate antiviral T cell responses. Yet previous studies reported that PD-L1-deficient mice develop fatal pathology during early systemic lymphocytic choriomeningitis virus (LCMV) infection, suggesting a host protective role of T cell down-regulation. As the exact mechanisms of pathology development remained unclear, we set out to delineate in detail the underlying pathogenesis. Mice deficient in PD-1-PD-L1 signaling or lacking PD-1 signaling in CD8 T cells succumbed to fatal CD8 T cell-mediated immunopathology early after systemic LCMV infection. In the absence of regulation via PD-1, CD8 T cells killed infected vascular endothelial cells via perforin-mediated cytolysis, thereby severely compromising vascular integrity. This resulted in systemic vascular leakage and a consequential collapse of the circulatory system. Our results indicate that the PD-1-PD-L1 pathway protects the vascular system from severe CD8 T cell-mediated damage during early systemic LCMV infection, highlighting a pivotal physiological role of T cell down-regulation and suggesting the potential development of immunopathological side effects when interfering with the PD-1-PD-L1 pathway during systemic virus infections.


Community-integrated omics links dominance of a microbial generalist to fine-tuned resource usage.
Microbial communities are complex and dynamic systems that are primarily structured according to their members' ecological niches. To investigate how niche breadth (generalist versus specialist lifestyle strategies) relates to ecological success, we develop and apply an integrative workflow for the multi-omic analysis of oleaginous mixed microbial communities from a biological wastewater treatment plant. Time- and space-resolved coupled metabolomic and taxonomic analyses demonstrate that the community-wide lipid accumulation phenotype is associated with the dominance of the generalist bacterium Candidatus Microthrix spp. By integrating population-level genomic reconstructions (reflecting fundamental niches) with transcriptomic and proteomic data (realised niches), we identify finely tuned gene expression governing resource usage by Candidatus Microthrix parvicella over time. Moreover, our results indicate that the fluctuating environmental conditions constrain the accumulation of genetic variation in Candidatus Microthrix parvicella likely due to fitness trade-offs. Based on our observations, niche breadth has to be considered as an important factor for understanding the evolutionary processes governing (microbial) population sizes and structures in situ.


The Accuracy of Incident Vertebral Fracture Detection in Children Using Targeted Case-Finding Approaches.
Vertebral fractures are clinically important sequelae of a wide array of pediatric diseases. In this study, we examined the accuracy of case-finding strategies for detecting incident vertebral fractures (IVF) over 2 years in glucocorticoid-treated children (n = 343) with leukemia, rheumatic disorders, or nephrotic syndrome. Two clinical situations were addressed: the prevalent vertebral fracture (PVF) scenario (when baseline PVF status was known), which assessed the utility of PVF and low lumbar spine bone mineral density (LS BMD; Z-score <-1.4), and the non-PVF scenario (when PVF status was unknown), which evaluated low LS BMD and back pain. LS BMD was measured by dual-energy X-ray absorptiometry, vertebral fractures were quantified on spine radiographs using the modified Genant semiquantitative method, and back pain was assessed by patient report. Forty-four patients (12.8%) had IVF. In the PVF scenario, both low LS BMD and PVF were significant predictors of IVF. Using PVF to determine which patients should have radiographs, 11% would undergo radiography (95% confidence interval [CI] 8-15) with 46% of IVF (95% CI 30-61) detected. Sensitivity would be higher with a strategy of PVF or low LS BMD at baseline (73%; 95% CI 57-85) but would require radiographs in 37% of children (95% CI 32-42). In the non-PVF scenario, the strategy of low LS BMD and back pain produced the highest specificity of any non-PVF model at 87% (95% CI 83-91), the greatest overall accuracy at 82% (95% CI 78-86), and the lowest radiography rate at 17% (95% CI 14-22). Low LS BMD or back pain in the non-PVF scenario produced the highest sensitivity at 82% (95% CI 67-92), but required radiographs in 65% (95% CI 60-70). These results provide guidance for targeting spine radiography in children at risk for IVF. © 2021 American Society for Bone and Mineral Research (ASBMR).


GLIF3: a representation format for sharable computer-interpretable clinical practice guidelines.
The Guideline Interchange Format (GLIF) is a model for representation of sharable computer-interpretable guidelines. The current version of GLIF (GLIF3) is a substantial update and enhancement of the model since the previous version (GLIF2). GLIF3 enables encoding of a guideline at three levels: a conceptual flowchart, a computable specification that can be verified for logical consistency and completeness, and an implementable specification that is intended to be incorporated into particular institutional information systems. The representation has been tested on a wide variety of guidelines that are typical of the range of guidelines in clinical use. It builds upon GLIF2 by adding several constructs that enable interpretation of encoded guidelines in computer-based decision-support systems. GLIF3 leverages standards being developed in Health Level 7 in order to allow integration of guidelines with clinical information systems. The GLIF3 specification consists of an extensible object-oriented model and a structured syntax based on the resource description framework (RDF). Empirical validation of the ability to generate appropriate recommendations using GLIF3 has been tested by executing encoded guidelines against actual patient data. GLIF3 is accordingly ready for broader experimentation and prototype use by organizations that wish to evaluate its ability to capture the logic of clinical guidelines, to implement them in clinical systems, and thereby to provide integrated decision support to assist clinicians.


POTRA: a conserved domain in the FtsQ family and a class of beta-barrel outer membrane proteins.
POTRA (for polypeptide-transport-associated domain) is a novel domain identified in proteins of the ShlB, Toc75, D15 and FtsQ/DivIB families. In most cases, the POTRA domain is associated with a beta-barrel outer membrane domain and its function has been experimentally related to polypeptide transport in Toc75 (Tic-Toc protein import system in chloroplast) and ShlB families. In addition to potential key roles in protein transport across the outer membrane and in bacterial septation, the POTRA domain has attractive features for vaccine development in diseases such as cholera, meningitis, gonorrhoea and syphilis.


Sleep and sleepiness: impact of entering or leaving shiftwork--a prospective study.
Very little is known about the effects on sleep and sleepiness of entering or exiting shiftwork. The present study used a longitudinal database (n = 3637). Participants completed a questionnaire on work hours, sleep, and work environment at the start and end of a 5-yr period. Changes in shift/day work status were related to change in a number of subjective sleep variables using logistic regression analysis. The analyses were adjusted for age, sex, and differences in socioeconomic status, work demands, work control, physical workload, marriage status, and number of children. In comparison with constant day work, entering shiftwork (with or without night shifts) from day work increased the risk of difficulties in falling asleep, and leaving shiftwork reduced this risk (odds ratio [OR] = 2.8 [confidence interval, CI = 1.8-4.5]). Also falling asleep at work showed a consistent pattern; an increased risk of falling asleep for those with shiftwork on both occasions, and for those with night work on both occasions. Also entering night work was associated with a strongly increased risk of falling asleep at work (OR = 2.9 [CI = 1.3-6.7]). These results suggest that entering and leaving shiftwork has a considerable impact on sleep and alertness. However, there is a need for large and more extended longitudinal studies to support our findings.


Evaluation of 3 approaches for assessing adherence to vaginal gel application in clinical trials.
Accurate measurement of adherence to product use is an ongoing challenge in microbicide trials. We compared adherence estimates using 2 applicator tests (a dye stain assay [DSA] and an ultraviolet light assay [UVA]), the Wisebag (an applicator container that electronically tracks container openings), and self-reported adherence (ability, frequency, and percent missed doses). Healthy, HIV-negative, nonpregnant US women aged 23 to 45 years received a Wisebag and 32 applicators filled with placebo gel were instructed to insert 1 applicator daily for 30 days, returned the Wisebag and all applicators, and completed an exit interview. Emptied applicators were tested by UVA and then DSA, and scored by 2 blinded readers. Positive and negative controls were randomly included in applicator batches. Among 42 women enrolled, 39 completed the study. Both DSA and UVA yielded similar sensitivity (97% and 95%) and specificity (79% and 79%). Two participants had fully inoperable Wisebags, and 9 had partially inoperable Wisebags. The proportion of participants considered to have high adherence (≥80%) varied: 43% (Wisebag), 46% (UVA), 49% (DSA), and 62% to 82% (self-reports). For estimating high adherence, Wisebag had a sensitivity of 76% (95% confidence interval, 50%-93%) and a specificity of 85% (95% confidence interval, 62%-97%) compared with DSA. Although 28% of participants reported forgetting to open the Wisebag daily, 59% said that it helped them remember gel use. Dye stain assay and UVA performed similarly. Compared with these tests, self-reports overestimated and Wisebag underestimated adherence. Although Wisebag may encourage gel use, the applicator tests currently seem more useful for measuring use in clinical trials.


Heritability of interpack aggression in a wild pedigreed population of North American grey wolves.
Aggression is a quantitative trait deeply entwined with individual fitness. Mapping the genomic architecture underlying such traits is complicated by complex inheritance patterns, social structure, pedigree information and gene pleiotropy. Here, we leveraged the pedigree of a reintroduced population of grey wolves (Canis lupus) in Yellowstone National Park, Wyoming, USA, to examine the heritability of and the genetic variation associated with aggression. Since their reintroduction, many ecological and behavioural aspects have been documented, providing unmatched records of aggressive behaviour across multiple generations of a wild population of wolves. Using a linear mixed model, a robust genetic relationship matrix, 12,288 single nucleotide polymorphisms (SNPs) and 111 wolves, we estimated the SNP-based heritability of aggression to be 37% and an additional 14% of the phenotypic variation explained by shared environmental exposures. We identified 598 SNP genotypes from 425 grey wolves to resolve a consensus pedigree that was included in a heritability analysis of 141 individuals with SNP genotype, metadata and aggression data. The pedigree-based heritability estimate for aggression is 14%, and an additional 16% of the phenotypic variation was explained by shared environmental exposures. We find strong effects of breeding status and relative pack size on aggression. Through an integrative approach, these results provide a framework for understanding the genetic architecture of a complex trait that influences individual fitness, with linkages to reproduction, in a social carnivore. Along with a few other studies, we show here the incredible utility of a pedigreed natural population for dissecting a complex, fitness-related behavioural trait.


Patient safety in the office-based setting.
After studying this article, the participant should be able to: 1. Discern the importance of the physician's office administrative capacity. 2. Recognize the necessity of a system for quality assessment. 3. Assess which procedures are safe in the office-based setting. 4. Know the basic steps to properly evaluate patients for office-based plastic surgery. At least 44,000 Americans die annually as a result of preventable medical errors. Medical mistakes are the eighth leading cause of death in the United States, costing between $54.6 billion and $79 billion, or 6 percent of total annual national health care expenditures. Office-based procedures comprise a 10-fold increase in risk for serious injury or death as compared with an ambulatory surgical facility. This article reviews the literature on office-based patient safety issues. It places special emphasis on the statements and advisories published by the American Society of Plastic Surgeons' convened Task Force on Patient Safety in Office-Based Settings. This article stresses areas of increased patient safety concern, such as deep vein thrombosis prophylaxis and liposuction surgery. The article divides patient safety in health care delivery into three broad categories. First, patient safety starts with emphasis at the administrative level. The physician or independent governing body must develop a system of quality assessment that functions to minimize preventable errors and report outcomes and errors. Second, the clinical aspects of patient safety require that the physician evaluate whether the procedure(s) and the patient are proper for the office setting. Finally, this article gives special attention to liposuction, the most frequently performed office-based plastic surgery procedure. Patient safety must be every physician's highest priority, as reflected in the Hippocratic Oath: primum non nocere ("first, do no harm"). In the office setting, this priority requires both administrative and clinical emphasis. The physician who gives the healing touch of quality care must always have patient safety as the foremost priority.


Interstitial myofibroblasts: predictors of progression in membranous nephropathy.
To determine the role of interstitial myofibroblasts in the progression of membranous nephropathy; and to assess the predictive value of quantifying myofibroblasts in determining long term renal outcome. All cases of membranous nephropathy, diagnosed by renal biopsy at University Hospital of South Manchester between 1984 and 1987, were studied retrospectively. The biopsy specimens (n = 26) were reviewed and analysed morphometrically to measure interstitial volume as a proportion of the total volume of renal cortex, and numbers of interstitial myofibroblasts (cells positive for alpha-smooth muscle actin within the interstitium). Clinical data, with a follow up of seven to eight years, was available for 24 patients, and renal outcome was correlated with pathological changes in the initial diagnostic biopsy specimen. The number of myofibroblasts and interstitial volume were inversely correlated with creatinine clearance at the initial biopsy, and at the end of follow up. Percentage sclerosed glomeruli or stage of glomerular disease, assessed by electron microscopy, did not correlate with renal function at initial biopsy or during follow up. The number of myofibroblasts, but not interstitial volume, correlated with severity of proteinuria at initial biopsy. Of 15 biopsy specimens showing no or mild interstitial fibrosis, four showed a notable increase in the number of interstitial myofibroblasts. All of these patients developed chronic renal failure, compared with three of 11 patients whose specimens showed no or a mild increase in myofibroblast numbers. Interstitial myofibroblasts play a role in the development of interstitial fibrosis and progressive renal failure in membranous nephropathy. Increased numbers of myofibroblasts in biopsy specimens showing only mild fibrosis may predict subsequent chronic renal failure.


The use of massive bone allograft with intramedullary free fibular flap for limb salvage in a pediatric and adolescent population.
Long segmental bony defects after tumor extirpation can pose difficult problems for the reconstructive surgeon. Capanna and colleagues have described a technique that places a free fibular flap within the intramedullary canal of an allograft for reconstruction of large intercalary bony defects. This article describes the authors' long-term follow-up with this technique for the treatment of large segmental bone defects in a pediatric population. Over a 6-year period, seven patients underwent bony reconstruction with an allograft and vascularized fibular construct. All reconstructions were performed for lower extremity salvage after tumor extirpation. Grafts were evaluated for viability with bone scans 10 days postoperatively. Radiologic and clinical evaluations were performed on all patients. Time to union was recorded through evaluation of plain radiographs. Patients' charts were evaluated for postoperative complications. There were two female and five male patients with an average age of 10.5 years. The average follow-up time was 36 months (range, up to 72 months). Limb salvage was 100 percent, with all bone scans positive at 10 days. Two nonunions at the allograft interface were treated successfully with a secondary bone graft. The average time to complete bony union of the fibula and allograft to the native bone was 9 months. There were no allograft fractures and no infections. One patient developed nonunion at the donor leg syndesmosis site. Average final knee motion was 110 degrees. All patients returned to ambulation. Intramedullary free fibular flaps in combination with massive bony allografts provide an excellent option in the pediatric population for reconstruction of large bony defects after tumor extirpation.


Adoptive transfer of neonatal T lymphocytes rescues immunoglobulin production in mice with severe combined immune deficiency.
Mice with the autosomal recessive severe combined immune deficiency (scid) mutation lack mature lymphocytes because of defective joining of T cell receptor and immunoglobulin (Ig) gene segments. Penetrance of this mutation is incomplete since 10-25% of SCID mice produce some T or B lymphocytes. This "leaky" phenotype could be due to a reversion of the mutation in some mice or to a constant, low frequency of functional lymphocytes generated in all SCID mice with variable survival of such cells. We report here that all SCID mice can be stimulated to produce functional B cells by the transfer of normal neonatal, but not adult, T cells. T cell-induced rescue of C.B-17scid B cells results in high levels of Ig expressing the Ighb allotype of the SCID recipient. These results show that all SCID mice generate some functional B cells, the majority of which do not survive in the absence of a subset of T cells present in high frequency in the neonate.


Attitudes and behaviours of maternal health care providers in interactions with clients: a systematic review.
High maternal mortality and morbidity persist, in large part due to inadequate access to timely and quality health care. Attitudes and behaviours of maternal health care providers (MHCPs) influence health care seeking and quality of care. Five electronic databases were searched for studies from January 1990 to December 2014. Included studies report on types or impacts of MHCP attitudes and behaviours towards their clients, or the factors influencing these attitudes and behaviours. Attitudes and behaviours mentioned in relation to HIV infection, and studies of health providers outside the formal health system, such as traditional birth attendants, were excluded. Of 967 titles and 412 abstracts screened, 125 full-text papers were reviewed and 81 included. Around two-thirds used qualitative methods and over half studied public-sector facilities. Most studies were in Africa (n = 55), followed by Asia and the Pacific (n = 17). Fifty-eight studies covered only negative attitudes or behaviours, with a minority describing positive provider behaviours, such as being caring, respectful, sympathetic and helpful. Negative attitudes and behaviours commonly entailed verbal abuse (n = 45), rudeness such as ignoring or ridiculing patients (n = 35), or neglect (n = 32). Studies also documented physical abuse towards women, absenteeism or unavailability of providers, corruption, lack of regard for privacy, poor communication, unwillingness to accommodate traditional practices, and authoritarian or frightening attitudes. These behaviours were influenced by provider workload, patients' attitudes and behaviours, provider beliefs and prejudices, and feelings of superiority among MHCPs. Overall, negative attitudes and behaviours undermined health care seeking and affected patient well-being. The review documented a broad range of negative MHCP attitudes and behaviours affecting patient well-being, satisfaction with care and care seeking. Reported negative patient interactions far outweigh positive ones. The nature of the factors which influence health worker attitudes and behaviours suggests that strengthening health systems, and workforce development, including in communication and counselling skills, are important. Greater attention is required to the attitudes and behaviours of MHCPs within efforts to improve maternal health, for the sake of both women and health care providers.


Collembola at three alpine subarctic sites resistant to twenty years of experimental warming.
This study examined the effects of micro-scale, site and 19 and 21 years of experimental warming on Collembola in three contrasting alpine subarctic plant communities (poor heath, rich meadow, wet meadow). Unexpectedly, experimental long-term warming had no significant effect on species richness, effective number of species, total abundance or abundance of any Collembola species. There were micro-scale effects on species richness, total abundance, and abundance of 10 of 35 species identified. Site had significant effect on effective number of species, and abundance of six species, with abundance patterns differing between sites. Site and long-term warming gave non-significant trends in species richness. The highest species richness was observed in poor heath, but mean species richness tended to be highest in rich meadow and lowest in wet meadow. Warming showed a tendency for a negative impact on species richness. This long-term warming experiment across three contrasting sites revealed that Collembola is capable of high resistance to climate change. We demonstrated that micro-scale and site effects are the main controlling factors for Collembola abundance in high alpine subarctic environments. Thus local heterogeneity is likely important for soil fauna composition and may play a crucial role in buffering Collembola against future climate change.


Exploring the binding site structure of the PPAR gamma ligand-binding domain by computational solvent mapping.
Solvent mapping moves molecular probes, small organic molecules containing various functional groups, around the protein surface, finds favorable positions, clusters the conformations, and ranks the clusters based on the average free energy. Using at least six different solvents as probes, the probes cluster in major pockets of the functional site, providing detailed and reliable information on the amino acid residues that are important for ligand binding. Solvent mapping was applied to 12 structures of the peroxisome proliferator activated receptor gamma (PPARgamma) ligand-binding domain (LBD), including 2 structures without a ligand, 2 structures with a partial agonist, and 8 structures with a PPAR agonist bound. The analysis revealed 10 binding "hot spots", 4 in the ligand-binding pocket, 2 in the coactivator-binding region, 1 in the dimerization domain, 2 around the ligand entrance site, and 1 minor site without a known function. Mapping is a major source of information on the role and cooperativity of these sites. It shows that large portions of the ligand-binding site are already formed in the PPARgamma apostructure, but an important pocket near the AF-2 transactivation domain becomes accessible only in structures that are cocrystallized with strong agonists. Conformational changes were seen in several other sites, including one involved in the stabilization of the LBD and two others at the region of the coactivator binding. The number of probe clusters retained by these sites depends on the properties of the bound agonist, providing information on the origin of correlations between ligand and coactivator binding.


Differential regulation of protein tyrosine kinase signalling by Dock and the PTP61F variants.
Tyrosine phosphorylation-dependent signalling is coordinated by the opposing actions of protein tyrosine kinases (PTKs) and protein tyrosine phosphatases (PTPs). There is a growing list of adaptor proteins that interact with PTPs and facilitate the dephosphorylation of substrates. The extent to which any given adaptor confers selectivity for any given substrate in vivo remains unclear. Here we have taken advantage of Drosophila melanogaster as a model organism to explore the influence of the SH3/SH2 adaptor protein Dock on the abilities of the membrane (PTP61Fm)- and nuclear (PTP61Fn)-targeted variants of PTP61F (the Drosophila othologue of the mammalian enzymes PTP1B and TCPTP respectively) to repress PTK signalling pathways in vivo. PTP61Fn effectively repressed the eye overgrowth associated with activation of the epidermal growth factor receptor (EGFR), PTK, or the expression of the platelet-derived growth factor/vascular endothelial growth factor receptor (PVR) or insulin receptor (InR) PTKs. PTP61Fn repressed EGFR and PVR-induced mitogen-activated protein kinase signalling and attenuated PVR-induced STAT92E signalling. By contrast, PTP61Fm effectively repressed EGFR- and PVR-, but not InR-induced tissue overgrowth. Importantly, coexpression of Dock with PTP61F allowed for the efficient repression of the InR-induced eye overgrowth, but did not enhance the PTP61Fm-mediated inhibition of EGFR and PVR-induced signalling. Instead, Dock expression increased, and PTP61Fm coexpression further exacerbated the PVR-induced eye overgrowth. These results demonstrate that Dock selectively enhances the PTP61Fm-mediated attenuation of InR signalling and underscores the specificity of PTPs and the importance of adaptor proteins in regulating PTP function in vivo.


Composite spirometric-computed tomography outcome measure in early cystic fibrosis lung disease.
With the advent of therapies aimed at young patients with cystic fibrosis, who have mildly reduced pulmonary function, the need for improved outcome measures that discriminate treatment effects has become important. Pulmonary function measurements or chest high-resolution computed tomography (HRCT) scores have been separately used to assess interventions. We evaluated these modalities separately and together during a treatment study to develop a more sensitive outcome measure. In a 1-year trial, 25 children randomized either to daily Pulmozyme or to normal saline aerosol were evaluated at randomization and at 3 and 12 months. Outcome variables were pulmonary function test (PFT) results, a global HRCT score, and a composite score incorporating PFTs and HRCT scoring. Regression analyses with generalized estimating equations permitted estimation of the difference in treatment effect between groups over time for each outcome. The largest difference in treatment effects observed at 12 months, measured by the percentage change from baseline, were with the composite total and maximal CT/PFT scores (35.4 and 30.4%), compared with mean forced expiratory flow during the middle half of the FVC (FEF25-75%) (13.0%) and total and maximal global HRCT scores (6.2%, 7.2%). The composite total and maximal CT/PFT scores were the most sensitive outcome measures for discriminating a treatment effect in children with cystic fibrosis with normal or mildly reduced pulmonary function during a 1-year trial of Pulmozyme.


The contribution of cohesin-SA1 to gene expression and chromatin architecture in two murine tissues.
Cohesin, which in somatic vertebrate cells consists of SMC1, SMC3, RAD21 and either SA1 or SA2, mediates higher-order chromatin organization. To determine how cohesin contributes to the establishment of tissue-specific transcriptional programs, we compared genome-wide cohesin distribution, gene expression and chromatin architecture in cerebral cortex and pancreas from adult mice. More than one third of cohesin binding sites differ between the two tissues and these show reduced overlap with CCCTC-binding factor (CTCF) and are enriched at the regulatory regions of tissue-specific genes. Cohesin/CTCF sites at active enhancers and promoters contain, at least, cohesin-SA1. Analyses of chromatin contacts at the Protocadherin (Pcdh) and Regenerating islet-derived (Reg) gene clusters, mostly expressed in brain and pancreas, respectively, revealed remarkable differences that correlate with the presence of cohesin. We could not detect significant changes in the chromatin contacts at the Pcdh locus when comparing brains from wild-type and SA1 null embryos. In contrast, reduced dosage of SA1 altered the architecture of the Reg locus and decreased the expression of Reg genes in the pancreas of SA1 heterozygous mice. Given the role of Reg proteins in inflammation, such reduction may contribute to the increased incidence of pancreatic cancer observed in these animals.


Supernumerary Testis Discovered Upon Workup of a Paratesticular Mass.
A supernumerary testis is a rare finding with limited description in the literature. Presentations of supernumerary testes vary and often involve initial discovery intraoperatively or in association with a separate condition. In this case report, we describe our experience of a patient with ultrasound imaging concerning for a left paratesticular mass. The patient was taken to the operating room where the mass was found to be an atrophic, supernumerary testis. We also address the specific anatomy encountered in our case and discuss both the intraoperative and post-operative management that resulted, in addition to reviewing relevant literature.


The "syncytial variant" of nodular sclerosing Hodgkin's disease.
The histologic and immunologic features of an unusual morphologic expression of nodular sclerosing Hodgkin's disease, which ahs been termed the "syncytial variant," are described. In biopsy material from 18 cases, numerous Reed-Sternberg cell variants were observed in sheets and cohesive clusters, and at least focal evidence of nodular sclerosis was present in each case. The granulocyte antibody anti-Leu M1 reacted with antigenic determinants in Reed-Sternberg cells and atypical variants thereof in 13 of the 18 cases; the lack of staining with antibodies reactive with the leukocyte common (T200) antigen (PD7/26), keratin (AE1), and S100 protein (polyclonal anti-S100) was helpful in excluding non-Hodgkin's lymphoma, carcinoma, and melanoma, respectively. This unusual form of nodular sclerosing Hodgkin's disease is important to recognize, since it may simulate metastatic neoplasms, thymoma, and non-Hodgkin's lymphoma.