Construction of a cloned library of the EcoRI fragments from the human cytomegalovirus genome (strain AD169).
The DNA genome of human cytomegalovirus (HCMV) strain AD169 is 158 x 10(6) Mr. Cleavage of the HCMV DNA with the restriction endonuclease EcoRI yields 35 major fragments ranging in size from 0.54 x 10(6) Mr. We have constructed a cloned library of the EcoRI fragments of this strain of HCMV, using the plasmid pACYC184 and the recipient bacterium Escherichia coli strain HB101 RecA-. The viral origin of the cloned inserts was determined by hybridization to viral DNA. The fragments were characterized further by digestion with other restriction enzymes. Several clones were obtained which contained sequences spanning the junction between the long (L) and short (S) components of the viral DNA sequences. These clones differed in molecular weight by multiples of 0.3 x 10(6) to 0.4 x 10(6) Mr. The variability found in the clones was also reflected in the genome. Each clone containing a junction sequence hybridized to a series of bands on Southern filters of EcoRI-digested HCMV DNA. This "ladder effect" provided evidence for a region of heterogeneity within the L-S junction.


Arginase inhibitory properties of flavonoid compounds from the leaves of Mulberry (Morus alba, Moraceae).
We aimed to isolate and identify bioactive molecules from Morus alba (Moraceae) leaves having arginase inhibitory activity towards the combat of clinical outcomes related to endothelial dysfunction. Extraction and isolation were carried out by successive macerations, prepurification by using a Solid Phase Extraction (SPE) and separation using preparative PLC. The structures of the isolated components were established and confirmed by spectroscopic analyses, including the ESI-HRMS and NMR spectroscopic investigations. Biological evaluation was performed by using an in vitro assay with liver bovine purified arginase and by an ex vivo aortic ring study. We demonstrated that a phenolic extract from the leaves of M. alba possesses mammalian arginase inhibitory capacities. Investigation of the chemical constituents of its leaves results in the isolation and identification of ten compounds investigated in vitro for their arginase inhibitory capacities. Four compounds showed significant inhibition of arginase, with percentage inhibition ranging from 54% to 83% at 100 µm. In isolated rat aortic rings incubated with NO synthase inhibitor, Luteolin-7-diglucoside compound (2) was able to increase acetylcholine-induced relaxation. These results demonstrated the attractive ability of M. alba to be a potential source for the discovery of new active products on vascular system.


[Right atrial diverticulum in an adult patient].
Right atrial diverticula are rare clinical findings and thus this possibility is usually omitted in the differential diagnosis of right atrial pathologies. Only 105 cases of right atrial anomalies were reported in medical journals in years 1955-1998, of which only 13 cases referred to a single diverticulum of the right atrium. The presented case is a 65-year-old patient with a right atrial diverticulum whose presence was confirmed intra-operatively.


[Factors predicting failure of adjuvant hormonotherapy of breast carcinoma. A study in tamoxifen treated patients].
The latest clinical trials indicate better performance of aromatase inhibitors, compared to tamoxifen, in adjuvant hormonotherapy of breast carcinoma. The identification of molecular markers, predicting resistance to tamoxifen, could help to identify patients, which are most likely to benefit from aromatase inhibitors in up-front adjuvant hormonotherapy. Tissue microarrays were constructed from archival paraffin blocks of primary tumors of 179 patients with estrogen receptor positive operable breast carcinoma in stage I-III, subsequently treated with tamoxifen for five years or until relapse, with at least 7 years follow up available. The amplifications of Her-2 and cyclin D1 genes were evaluated by fluorescence in-situ hybridization. The level of progesterone receptor (PR) and Ki67 were estimated by immunohistochemistry. 54 of above patients recurred during follow up. In univariate analysis of disease free survival, the presence of more than three nodal metastases (RR=4,5 p<0,001), grade 3 (RR=2,3 p=0,035), cyclin D1 (RR=3,06 p<0,001) and Her-2 (RR=3,06 p<0,001) amplifications were identified as significant risk factors, together with the negativity of PR (RR=2,1 p=0,013). In multivariate analysis, only clinical stage III (RR=2,6 p=0,003), cyclin D1 (RR=2,7 p=0,001) and Her-2 (RR=2,1 p=0,014) amplifications proved significant. In 77 patients who received adjuvant chemotherapy no statistically significant risk factor was identified. In multivariate analysis of 102 patients without adjuvant chemotherapy only stage III (RR=6,9 p=0,001) and Her-2 amplification (RR=4,5 p=0,001) were confirmed. The advanced clinical stage, cyclin D1 and Her-2 gene amplifications represent factors, predicting the failure of adjuvant tamoxifen treatment, but their predictive value is much lower in patients receiving adjuvant chemotherapy. This fact indicates, they can reflect the common biological aggressiveness of tumor and need not to be tamoxifen specific.


Ancestral sequence reconstruction produces thermally stable enzymes with mesophilic enzyme-like catalytic properties.
Enzymes have high catalytic efficiency and low environmental impact, and are therefore potentially useful tools for various industrial processes. Crucially, however, natural enzymes do not always have the properties required for specific processes. It may be necessary, therefore, to design, engineer, and evolve enzymes with properties that are not found in natural enzymes. In particular, the creation of enzymes that are thermally stable and catalytically active at low temperature is desirable for processes involving both high and low temperatures. In the current study, we designed two ancestral sequences of 3-isopropylmalate dehydrogenase by an ancestral sequence reconstruction technique based on a phylogenetic analysis of extant homologous amino acid sequences. Genes encoding the designed sequences were artificially synthesized and expressed in Escherichia coli. The reconstructed enzymes were found to be slightly more thermally stable than the extant thermophilic homologue from Thermus thermophilus. Moreover, they had considerably higher low-temperature catalytic activity as compared with the T. thermophilus enzyme. Detailed analyses of their temperature-dependent specific activities and kinetic properties showed that the reconstructed enzymes have catalytic properties similar to those of mesophilic homologues. Collectively, our study demonstrates that ancestral sequence reconstruction can produce a thermally stable enzyme with catalytic properties adapted to low-temperature reactions.


Chondrosarcoma of the base of the skull: a clinicopathologic study of 200 cases with emphasis on its distinction from chordoma.
Conventional chondrosarcoma (CSA) of the skull base is an uncommon neoplasm that can resemble chordoma, and indeed it is misdiagnosed frequently as such. This has important clinical implications, because when treated with similar aggressive treatment strategies, CSA has a much better prognosis than chordoma. In an effort to identify those morphologic and immunohistochemical features that help to identify conventional skull base CSA correctly and to understand its prognosis better, particularly compared with chordoma, when treated with surgery and proton beam irradiation, the authors performed a clinicopathologic analysis of 200 CSAs. The patients ranged in age from 10 to 79 years (mean, 39 years), 87 patients were male and 113 patients were female, and most presented with symptoms related to the central nervous system. Approximately 6% of the tumors arose in the sphenoethmoid complex, 28% originated in the clivus, and 66% developed in the temperooccipital junction. Histologically, 15 tumors (7.5%) were classified as hyaline CSA, 59 (29.5%) as myxoid CSA, and 126 (63%) as mixed hyaline and myxoid CSA. A total of 101 (50.5%) tumors were grade 1, 57 (28.5%) had areas of grades 1 and 2, and 42 (21%) were pure grade 2 neoplasms. The vast majority of patients originated from referring hospitals, and the diagnosis was changed prospectively at our institution to CSA from chordoma in 74 patients (37%). Of the tumors studied immunohistochemically, 96 of 97 (98.9%) stained for S-100 protein, 0 of 97 (0%) stained for keratin, and faint staining for epithelial membrane antigen was seen in 7 of 88 tumors (7.95%). All patients underwent high-dose postoperative fractionated precision conformal radiation therapy with a dose that ranged from 64.2 to 79.6 Cobalt-Gray-equivalents (median, 72.1 Cobalt-Gray-equivalents, given in 38 fractions. The 200 patients had a median follow-up of 63 months (range, 2.1 mos - 18.5 yrs). Tumor control was defined as lack of progression by clinical and radiographic assessment. Based on this definition, there were three local recurrences, and two of these patients died of tumor-related complications. The 5- and 10-year local control rates were 99% and 98% respectively, and the 5- and 10-year disease-specific survival rates were both 99%. In contrast to CSA, the 5- and 10-year survival rates of chordoma have been reported to be approximately 51 % and 35% respectively, and in our institution intensive treatment has resulted in 5- and 10-year progression-free survival rates of 70% and 45% respectively. CSA of the skull base can be distinguished reliably from chordoma, and this distinction is important because skull base CSA has an excellent prognosis when treated with surgery and proton beam irradiation, whereas chordomas have a substantially poorer clinical course despite similar aggressive management.


Hypoglycemia during treatment with the ketogenic diet in a child with refractory epilepsy-results of continuous glucose monitoring.
The ketogenic diet (KD) is an effective treatment for intractable epilepsy in children. Hypoglycemia can be one of its side-effects, which is considered to be present mainly during the introductory phase of KD. Continuous glucose monitoring in a 6-year old non-diabetic child treated with KD for more than 8 months revealed long periods of asymptomatic hypoglycemia (8.9% of the total time under 2.5 mmol/l, 10.6% of the total time in the range between 2.5-3.0, 29.1% in the range of 3.0-3.6 mmol/l). The episodes of serious hypoglycemia were associated with a fasting state. The amount of sacharides in KD was increased with substantial glycemic profile improvement.


Empowering Stories: Transportation into Narratives with Strong Protagonists Increases Self-Related Control Beliefs.
Several studies have shown that narratives can influence readers' beliefs about themselves. In the present study, our goal was to investigate whether stories portraying a strong protagonist can positively influence recipients' beliefs of being in control of events in their own lives (self-related control beliefs). Experiment 1 showed that narratives in both written text and video form with protagonists displaying high versus low self-efficacy can, at least temporarily, affect recipients' own self-related control beliefs when they experience strong transportation into the stories. In addition, the results suggest that recipients' perceived ability to generate vivid mental imagery affects their transportation into and identification with characters in texts versus films. Experiment 2 manipulated transportation and identification experimentally and showed that the effect of this manipulation on self-related control beliefs was indeed mediated by experienced transportation and identification. The results are discussed within the framework of the Transportation Imagery Model of narrative persuasion.


Patients with celiac disease have an increased risk for pancreatitis.
Patients with celiac disease have been reported to be at increased risk for pancreatitis and pancreatic insufficiency, but the risk might have been overestimated because of patient selection and limited numbers of patients for analysis. Furthermore, no distinction has been made between patients with gallstone-related and non-gallstone-related pancreatitis. We performed a nationwide study to determine the risk for any pancreatitis or subtype of pancreatitis among patients with biopsy-verified celiac disease. We analyzed data from patients in Sweden with celiac disease (n = 28,908) who were identified on the basis of small intestinal biopsy records from 28 pathology departments (those with villous atrophy, Marsh 3). Biopsies were performed from 1969 to 2008, and biopsy report data were collected from 2006 to 2008. Patients with pancreatitis were identified on the basis of diagnostic codes in the Swedish Patient Register and records of pancreatic enzyme use in the Swedish Prescribed Drug Register. Data were matched with those from 143,746 individuals in the general population; Cox regression was used to estimate hazard ratios (HRs) for pancreatitis. We identified 406 patients with celiac disease who were later diagnosed with pancreatitis (and 143 with expected pancreatitis) (HR, 2.85; 95% confidence interval [CI], 2.53-3.21). The absolute risk of any pancreatitis among patients with celiac disease was 126/100,000 person-years, with an excess risk of 81/100,000 person-years. The HR for gallstone-related acute pancreatitis was 1.59 (95% CI, 1.06-2.40), for non-gallstone-related acute pancreatitis HR was 1.86 (95% CI, 1.52-2.26), for chronic pancreatitis HR was 3.33 (95% CI, 2.33-4.76), and for supplementation with pancreatic enzymes HR was 5.34 (95% CI, 2.99-9.53). The risk of any pancreatitis within 5 years of diagnosis was 2.76 (95% CI, 2.36-3.22). Based on an analysis of medical records from Sweden, patients with celiac disease have an almost 3-fold increase in risk of developing pancreatitis, compared with the general population.


Achievement of European standards by CROB-IRCCS.
The Organisation of European Cancer Institutes (OECI) has recently endorsed a program for the accreditation of Italian cancer institutes. Any cancer center that aims to provide research, education, and care services to cancer patients should undergo an evaluation process in order to become OECI accredited. On a center basis, the task turned out to be challenging, and required commitment and increased workload. The timing is an adjunctive constraint, especially when dealing with bureaucracy. Once undertaken, the accreditation process goes through preparation and completion of the self-evaluation, peer review, report, and final designation. This process constitutes an unrepeatable opportunity for improvement. It is required to implement the necessary changes in order to improve policies, procedures, and employee training. Sharing the highlighted general remarks, strengths, and opportunities provided by the different audit teams (on a cancer center basis) will constitute a significant instrument to enhance cancer care.


[Lysosome activity in mice during rejection of a heterotopic heart implant].
Allotransplantation of the heart from newborn mice C57B1/6 into the pinna of adult CBA mice on the 3rd day after operation led to increased acid phosphatase activity and activation of the Golgi apparatus. Numerous lysosomes whose membrane became disrupted on the 7th-9th day from the time of implantation appeared later. At the same time histochemical study demonstrated local loss of acid phosphatase activity and a diffuse staining (brown) of the cytoplasm. No destruction of the lysosome membranes or release of the enzymes into the cytoplasm occurred in case of heart isografts (CBA-GBA).


Downregulation of kinin B1 receptor function by B2 receptor heterodimerization and signaling.
Signaling through the G protein-coupled kinin receptors B1 (kB1R) and B2 (kB2R) plays a critical role in inflammatory responses mediated by activation of the kallikrein-kinin system. The kB2R is constitutively expressed and rapidly desensitized in response to agonist whereas kB1R expression is upregulated by inflammatory stimuli and it is resistant to internalization and desensitization. Here we show that the kB1R heterodimerizes with kB2Rs in co-transfected HEK293 cells and natively expressing endothelial cells, resulting in significant internalization and desensitization of the kB1R response in cells pre-treated with kB2R agonist. However, pre-treatment of cells with kB1R agonist did not affect subsequent kB2R responses. Agonists of other G protein-coupled receptors (thrombin, lysophosphatidic acid) had no effect on a subsequent kB1R response. The loss of kB1R response after pretreatment with kB2R agonist was partially reversed with kB2R mutant Y129S, which blocks kB2R signaling without affecting endocytosis, or T342A, which signals like wild type but is not endocytosed. Co-endocytosis of the kB1R with kB2R was dependent on β-arrestin and clathrin-coated pits but not caveolae. The sorting pathway of kB1R and kB2R after endocytosis differed as recycling of kB1R to the cell surface was much slower than that of kB2R. In cytokine-treated human lung microvascular endothelial cells, pre-treatment with kB2R agonist inhibited kB1R-mediated increase in transendothelial electrical resistance (TER) caused by kB1R stimulation (to generate nitric oxide) and blocked the profound drop in TER caused by kB1R activation in the presence of pyrogallol (a superoxide generator). Thus, kB1R function can be downregulated by kB2R co-endocytosis and signaling, suggesting new approaches to control kB1R signaling in pathological conditions.


Bipolar II disorder: symptoms, course, and response to treatment.
The authors provide an overview of the diagnosis, course, and treatment of bipolar II disorder, a distinct subtype that is often misdiagnosed as unipolar depression or bipolar I disorder. They discuss research suggesting that underdiagnosis of bipolar II disorder reflects a failure to identify subthreshold expression of mania (hypomania). The course of bipolar II disorder is different from that of bipolar I disorder or unipolar depression, with distinct differences in rates of recovery, clinical features, and number of episodes. The risk of suicide appears to be particularly elevated. High rates of comorbid disorders have been reported, including substance abuse or dependence, anxiety disorders, and personality disorders. Few definitive studies exist on which to base conclusions about the differential efficacy of various treatment strategies in bipolar II disorder and bipolar I disorder. Preliminary studies suggest that the newer anticonvulsants may be of benefit for patients with bipolar II disorder, while other data suggest that there may be a greater role for antidepressant medications.


Development and validation of a spectroscopic method to predict wheat protein digestibility.
The CP digestibility is traditionally measured by chemical analyses of CP and marker concentration in digesta and diets. Potentially, CP digestibility can also be predicted by marker concentrations and spectral analyses of digesta and diet. Spectroscopy is a rapid, nondestructive method to ascertain qualitative and quantitative chemical information. Based on Beer's law, a spectroscopic method was developed to predict in vivo CP digestibility. To validate, samples of digesta and diet of wheat grain with predetermined apparent ileal digestibility (AID) of CP were scanned on a Fourier transform midinfrared (FTIR) instrument with a single-reflection attenuated total reflectance attachment. The AID of CP was calculated from peak intensities of spectra and measured marker concentrations in digesta and diet and then compared with in vivo AID of CP. The AID of CP of a wheat-based diet was predicted accurately with a deviation of 0.68 ± 0.86% from in vivo AID of CP ranging from 60.4 to 87.8%. Functional group digestibility based on the peak at 1,643 cm(-1) or the Amide I region was strongly correlated (r ≥ 0.99; P < 0.001) with in vivo AID of CP. In conclusion, instead of predictions based on calibrations, CP digestibility can also be potentially predicted directly from FTIR spectra.


Association Between Stillbirth at 23 Weeks of Gestation or Greater and Severe Maternal Morbidity.
To estimate whether stillbirth at 23 weeks of gestation or more is associated with increased risk of severe maternal morbidity compared with live birth, when stratified by maternal comorbidities. This retrospective cohort study used International Classification of Diseases, 9th Revision, Clinical Modification (ICD-9-CM) diagnosis and procedure codes within the Healthcare Cost and Utilization Project's Florida State Inpatient Database. The first delivery of female Florida residents aged 13-54 years old from 2005 to 2014 was included. The exposure was an ICD-9-CM code of stillbirth at 23 weeks of gestation or more; the control was an ICD-9-CM code of singleton live birth. Deliveries were stratified by the presence of 1 or more conditions within a well-validated maternal morbidity composite using ICD-9-CM codes during delivery hospitalization. The primary outcome was an ICD-9-CM diagnosis or procedure code during delivery hospitalization of any indices within the Centers for Disease Control and Prevention's severe maternal morbidity composite. Multivariable analyses adjusted for maternal sociodemographic factors and delivery mode to compare outcomes after stillbirth with live-birth delivery. Nine thousand five hundred twenty-three women who delivered stillborn fetuses and 1,353,044 with liveborn neonates were included. Among 6,590 stillbirths and 935,913 live births without maternal comorbidities, severe maternal morbidity was significantly more common during stillbirth delivery (n=345 [5.2%]), corresponding to a seven-fold increased risk compared with live birth (n=8,318 [0.9]; adjusted odds ratio [aOR] 7.05 [95% CI 6.27-7.93]). Among 2,933 stillbirths and 417,131 live births with maternal comorbidities, severe maternal morbidity was significantly more common during stillbirth delivery (n=390 [13.3%]): the risk was more than six-fold higher comparatively (n=11,122 [2.7%]; aOR 6.21 [95% CI 5.54-6.96]). Most maternal comorbidities were individually associated with higher risk of severe maternal morbidity during stillbirth compared with live-birth delivery. Though severe maternal morbidity is overall uncommon, delivering a stillborn fetus 23 weeks of gestation or greater is associated with increased likelihood of severe maternal morbidity, particularly among women with comorbidities, suggesting health care providers must be vigilant about severe maternal morbidity during stillbirth delivery.


Gastric analysis in selected populations. Improvement of its clinical usefulness.
Little doubt exists concerning the limited usefulness of gastric analysis as a screening test in unselected populations. Reliable tests, such as those for glycemia, are also of limited value when performed blindly on the general population. When, however, gastric analysis is performed on an adequate preselected population, it has a valuable role in the diagnosis and management of several gastroduodenal diseases. Highly satisfactory results can be obtained when the proper population is selected because errors are minimized due to an increased prevalence of diseases. We have verified the reliability of two types of gastric analyses: the conventional method of gastric analysis based on measurement of acid output alone (CGA) and the so-called gastroionogram (GIG), in 100 consecutive gastric analyses using the method proposed by Kronborg. Gastroionogram increases specificity (even at the risk of decreasing sensitivity) and provides an excellent predictive value of the test. With the same conditions CGA is less useful because of the overlapping among normal and diseased groups. It is useful, however, for separating patients into three groups of normal secretors, hyposecretors and hypersecretors.


Hemodynamic Effect of a Fontan Assist Device on a Numerical Fontan Circulatory Model under Various Medication Scenarios.
Patients with single-ventricle heart disease and failing Fontan circulation represent the largest and most rapidly growing subgroup of adults with congenital heart disease referred for transplant assessment. Few clinical therapies are available for improving Fontan hemodynamics. Mechanical circulatory support devices have been used successfully in the clinical setting to assist the single ventricle, but no device is currently available to support the subpulmonary circulation. A subpulmonary pump could be used to support patients with failing Fontan circulation by mitigating chronic venous hypertension and restoring normal physiology. Our group designed a Fontan assist device (FAD) to augment right-heart (subpulmonary) flow and decrease venous pressures. To ensure that our FAD could achieve target hemodynamic parameters, we developed a numerical Fontan circulatory model to evaluate the interaction between the cardiovascular system and the FAD. To ensure that the circulatory model can mimic real-world clinical conditions, we investigated the effects of various medications in the FAD loop. Results showed that the FAD can significantly increase cardiac output in Fontan patients and can create a pressure difference between the pulmonary arteries and venae cavae. Further, the systemic venous pressure can be significantly reduced by using the FAD plus diuretic treatment. The downstream pulmonary artery pressure can be increased by augmenting the FAD with vasodilator treatment, diuretic treatment, or both.Clinical Relevance- This work supports FAD development by providing a method for studying human cardiovascular effects under various hemodynamic scenarios.


Integrative analysis of large scale expression profiles reveals core transcriptional response and coordination between multiple cellular processes in a cyanobacterium.
Cyanobacteria are the only known prokaryotes capable of oxygenic photosynthesis. They play significant roles in global biogeochemical cycles and carbon sequestration, and have recently been recognized as potential vehicles for production of renewable biofuels. Synechocystis sp. PCC 6803 has been extensively used as a model organism for cyanobacterial studies. DNA microarray studies in Synechocystis have shown varying degrees of transcriptome reprogramming under altered environmental conditions. However, it is not clear from published work how transcriptome reprogramming affects pre-existing networks of fine-tuned cellular processes. We have integrated 163 transcriptome data sets generated in response to numerous environmental and genetic perturbations in Synechocystis. Our analyses show that a large number of genes, defined as the core transcriptional response (CTR), are commonly regulated under most perturbations. The CTR contains nearly 12% of Synechocystis genes found on its chromosome. The majority of genes in the CTR are involved in photosynthesis, translation, energy metabolism and stress protection. Our results indicate that a large number of differentially regulated genes identified in most reported studies in Synechocystis under different perturbations are associated with the general stress response. We also find that a majority of genes in the CTR are coregulated with 25 regulatory genes. Some of these regulatory genes have been implicated in cellular responses to oxidative stress, suggesting that reactive oxygen species are involved in the regulation of the CTR. A Bayesian network, based on the regulation of various KEGG pathways determined from the expression patterns of their associated genes, has revealed new insights into the coordination between different cellular processes. We provide here the first integrative analysis of transcriptome data sets generated in a cyanobacterium. This compilation of data sets is a valuable resource to researchers for all cyanobacterial gene expression related queries. Importantly, our analysis provides a global description of transcriptional reprogramming under different perturbations and a basic framework to understand the strategies of cellular adaptations in Synechocystis.


A Stochastic Model for Block Segmentation of Images Based on the Quadtree and the Bayes Code for It.
In information theory, lossless compression of general data is based on an explicit assumption of a stochastic generative model on target data. However, in lossless image compression, researchers have mainly focused on the coding procedure that outputs the coded sequence from the input image, and the assumption of the stochastic generative model is implicit. In these studies, there is a difficulty in discussing the difference between the expected code length and the entropy of the stochastic generative model. We solve this difficulty for a class of images, in which they have non-stationarity among segments. In this paper, we propose a novel stochastic generative model of images by redefining the implicit stochastic generative model in a previous coding procedure. Our model is based on the quadtree so that it effectively represents the variable block size segmentation of images. Then, we construct the Bayes code optimal for the proposed stochastic generative model. It requires the summation of all possible quadtrees weighted by their posterior. In general, its computational cost increases exponentially for the image size. However, we introduce an efficient algorithm to calculate it in the polynomial order of the image size without loss of optimality. As a result, the derived algorithm has a better average coding rate than that of JBIG.


Induction of morphological changes in death-induced cancer cells monitored by holographic microscopy.
We are using the label-free technique of holographic microscopy to analyze cellular parameters including cell number, confluence, cellular volume and area directly in the cell culture environment. We show that death-induced cells can be distinguished from untreated counterparts by the use of holographic microscopy, and we demonstrate its capability for cell death assessment. Morphological analysis of two representative cell lines (L929 and DU145) was performed in the culture flasks without any prior cell detachment. The two cell lines were treated with the anti-tumour agent etoposide for 1-3days. Measurements by holographic microscopy showed significant differences in average cell number, confluence, volume and area when comparing etoposide-treated with untreated cells. The cell volume of the treated cell lines was initially increased at early time-points. By time, cells decreased in volume, especially when treated with high doses of etoposide. In conclusion, we have shown that holographic microscopy allows label-free and completely non-invasive morphological measurements of cell growth, viability and death. Future applications could include real-time monitoring of these holographic microscopy parameters in cells in response to clinically relevant compounds.


Prospective, Head-to-Head Study of Three Computerized Neurocognitive Assessment Tools (CNTs): Reliability and Validity for the Assessment of Sport-Related Concussion.
Limited data exist comparing the performance of computerized neurocognitive tests (CNTs) for assessing sport-related concussion. We evaluated the reliability and validity of three CNTs-ANAM, Axon Sports/Cogstate Sport, and ImPACT-in a common sample. High school and collegiate athletes completed two CNTs each at baseline. Concussed (n=165) and matched non-injured control (n=166) subjects repeated testing within 24 hr and at 8, 15, and 45 days post-injury. Roughly a quarter of each CNT's indices had stability coefficients (M=198 day interval) over .70. Group differences in performance were mostly moderate to large at 24 hr and small by day 8. The sensitivity of reliable change indices (RCIs) was best at 24 hr (67.8%, 60.3%, and 47.6% with one or more significant RCIs for ImPACT, Axon, and ANAM, respectively) but diminished to near the false positive rates thereafter. Across time, the CNTs' sensitivities were highest in those athletes who became asymptomatic within 1 day before neurocognitive testing but was similar to the tests' false positive rates when including athletes who became asymptomatic several days earlier. Test-retest reliability was similar among these three CNTs and below optimal standards for clinical use on many subtests. Analyses of group effect sizes, discrimination, and sensitivity and specificity suggested that the CNTs may add incrementally (beyond symptom scores) to the identification of clinical impairment within 24 hr of injury or within a short time period after symptom resolution but do not add significant value over symptom assessment later. The rapid clinical recovery course from concussion and modest stability probably jointly contribute to limited signal detection capabilities of neurocognitive tests outside a brief post-injury window. (JINS, 2016, 22, 24-37).


Generation and Validation of the iKp1289 Metabolic Model for Klebsiella pneumoniae KPPR1.
Klebsiella pneumoniae has a reputation for causing a wide range of infectious conditions, with numerous highly virulent and antibiotic-resistant strains. Metabolic models have the potential to provide insights into the growth behavior, nutrient requirements, essential genes, and candidate drug targets in these strains. Here we develop a metabolic model for KPPR1, a highly virulent strain of K. pneumoniae. We apply a combination of Biolog phenotype data and fitness data to validate and refine our KPPR1 model. The final model displays a predictive accuracy of 75% in identifying potential carbon and nitrogen sources for K. pneumoniae and of 99% in predicting nonessential genes in rich media. We demonstrate how this model is useful in studying the differences in the metabolic capabilities of the low-virulence MGH 78578 strain and the highly virulent KPPR1 strain. For example, we demonstrate that these strains differ in carbohydrate metabolism, including the ability to metabolize dulcitol as a primary carbon source. Our model makes numerous other predictions for follow-up verification and analysis.


Method for obtaining structure and interactions from oriented lipid bilayers.
Precise calculations are made of the scattering intensity I(q) from an oriented stack of lipid bilayers using a realistic model of fluctuations. The quantities of interest include the bilayer bending modulus Kc, the interbilayer interaction modulus B, and bilayer structure through the form factor F(qz). It is shown how Kc and B may be obtained from data at large q(z) where fluctuations dominate. Good estimates of F(qz) can be made over wide ranges of q(z) by using I(q) in q regions away from the peaks and for q(r) not equal0 where details of the scattering domains play little role. Rough estimates of domain sizes can also be made from smaller q(z) data. Results are presented for data taken on fully hydrated, oriented DOPC bilayers in the L(alpha) phase. These results illustrate the advantages of oriented samples compared to powder samples.


Salivary proteins rescue within-session suppression and conditioned avoidance in response to an intragastric quinine infusion.
A subset of salivary proteins (SPs) upregulates in response to a quinine-containing diet. The presence of these SPs then results in decreased bitter taste responding and taste nerve signaling. Bitter taste receptors in the oral cavity are also found in the stomach and intestines and contribute to behaviors that are influenced by post-oral signaling. It has been previously demonstrated that after several pairings of post-orally infused bitter stimuli and a neutral flavor, animals learn to avoid the flavor that was paired with gastric bitter, this is referred to as conditioned avoidance. Furthermore, animals will decrease licking of a neutral solution within a test session, when licking is paired with an intragastric bitter infusion; this has been described as within-session suppression. We used these paradigms to test the role of SPs in behaviors influenced by post-oral signaling. In both paradigms, the animal is given a test solution directly into the stomach (with or without quinine, and with or without SPs), and the infusions are self-administered by licking to a neutral solution (Kool-Aid). Quinine successfully conditioned a flavor avoidance, but, in a separate trial, we were unable to detect conditioning in the presence of SPs from donor animals. Likewise, quinine was able to suppress licking within the conditioned suppression paradigm, but the effect of the bitter was blocked in the presence of saliva containing SPs. Together, these data suggest that behaviors driven by post-oral signaling can be altered by SPs. Author contributions: Experiments were designed by A-MT, LEM and VA-G, Data were collected by VA-G and LEM, Data were analyzed by VA-G, LEM and A-MT, Manuscript was prepared by A-MT, VA-G and LEM.


Infection control in intravenous therapy.
The insertion, maintenance, and surveillance of intravenous systems requires close attention by both physicians and nurses. Improvements are needed. With the knowledge presently available, each nurse should strive to improve individual and institutional techniques. Further research into all areas of intravenous therapy should be encouraged and supported by actively involved clinicians. Further improvements and refinements in equipment may also be appropriate. Only with the combined efforts of all concerned will the hazards of intravenous therapy be reduced.


Analysis of SCARECROW expression using a rapid system for assessing transgene expression in Arabidopsis roots.
Although the introduction of foreign genes into Arabidopsis has become routine, the production of transgenic Arabidopsis plants still requires several months. A transgene expression system (TES) has been developed that allows characterization of gene expression patterns and the effects of foreign genes in the Arabidopsis root in 2-4 weeks. The method is based on regeneration of stably transformed roots directly from callus tissue. TES has been used to study the expression of the SCARECROW gene, which is involved in establishing radial patterning in the root. The 2.5 kb region directly upstream of the SCARECROW coding region was found to be sufficient to confer cell-type specific expression. Furthermore, this promoter is active in the scr mutant background, indicating that factors essential for cell-type specific expression are present even in the absence of correct radial patterning. Finally, TES was used to demonstrate that the SCARECROW gene under control of this promoter complements the root organization defect of the scr mutant. These experiments demonstrate the utility of the TES system for studying gene expression in roots in wild-type and mutant backgrounds and for molecular complementation of root mutant phenotypes. It is possible that the method will also be applicable to other organs.


Electron-Transfer Route in the Early Oxidation States of the Mn4CaO5 Cluster in Photosystem II.
The electron transfer from the oxygen-evolving Mn4CaO5 cluster to the electron acceptor D1-Tyr161 (TyrZ) is a prerequisite for water oxidation and O2 evolution. Here, we analyzed the electronic coupling in the rate-limiting electron-transfer transitions using a combined quantum mechanical/molecular mechanical/polarizable continuum model approach. In the S0 to S1 transition, the electronic coupling between the electron-donor Mn3(III) and TyrZ is small (2 meV). In contrast, the electronic coupling between the dangling Mn4(III) and TyrZ is significantly large (172 meV), which suggests that the electron transfer proceeds from Mn3(III) to TyrZ via Mn4(III). In the S1 to S2 transition, the electronic coupling between Mn4(III) and TyrZ is also larger (124 meV) than that between Mn1(III) and TyrZ (1 meV), which favors the formation of the open-cubane S2 conformation with Mn4(IV) over the formation of the closed-cubane S2 conformation with Mn1(IV). In the S0 to S1 and S1 to S2 transitions, the Mn4 d-orbital and the TyrZ π-orbital are hybridized via D1-Asp170, which suggests that D1-Asp170 commonly provides a dominant electron-transfer route.


Respiratory allergy to the indoor ant (Monomorium pharaonis) not related to sting allergy.
Many studies are available on systemic reactions to ant sting, but few have described the direct role of ants in respiratory allergy. The nonstinging house ant, Monomorium pharaonis (pharaoh ant), is a highly infesting species in indoor environments. To determine whether the pharaoh ant is an indoor source of aeroallergens. Two patients with asthma who lived in homes with ant infestation were enrolled. Pharaoh ants were collected at the patients' homes, and crude extracts were prepared. Skin prick tests with ant extracts were performed. Specific IgE to pharaoh ant was measured by enzyme-linked immunosorbent assay (ELISA), and the allergenic components were determined by using immunoblot analysis. Cross-reactivity among pharaoh ant, imported fire ant, Pachycondyla chinensis ant, and other indoor allergens was evaluated by ELISA inhibition tests. Specific bronchial challenge testing was performed using pharaoh ant extracts. Both patients had positive skin test reactions to pharaoh ant extract and high levels of specific IgE antibodies to pharaoh ant. The ELISA inhibition test results demonstrated significant inhibition by pharaoh ant; however, P. chinensis, cockroach, and house dust mite showed no inhibition of the IgE binding to pharaoh ant. Two important IgE-binding components, 9.4 and 34 kDa, were identified by using immunoblot analysis. Pharaoh ant bronchial challenge test results showed typical early asthmatic reactions in 1 patient and dual asthmatic reactions in the other patient. Ants can induce IgE-mediated bronchoconstriction regardless of sting in sensitized patients. Ants should be taken into consideration as a cause of respiratory allergy in patients living in homes with visual evidence of infestation.


Treatments for androgenetic alopecia and alopecia areata: current options and future prospects.
Androgenetic alopecia and alopecia areata are common disorders of the hair follicle which may heavily influence self esteem and self image. Androgenetic alopecia is caused by the heightened sensitivity of scalp follicles to dihydro- testosterone whereas alopecia areata is induced by an autoimmune reaction. Current drug treatment approaches include the use of regrowth stimulators such as topical minoxidil and oral finasteride for androgenetic alopecia, as well as topical minoxidil, dithranol (anthralin), corticosteroids, contact sensitisers, and psoralen plus ultraviolet A irradiation (PUVA) therapy for alopecia areata. Combination regimens are also proposed. However, extreme cases of either type of alopecia do not generally respond well to these existing treatments. For this reason, new therapeutic strategies are directed towards both improving the targeting of existing agents, as well as the development of novel hypertrichotic modalities.


Isolation and characterization of a macrophage-derived heparin-binding growth factor.
Human mononuclear cells were plated in culture, and the conditioned media of these cells were analyzed by heparin-Sepharose affinity chromatography. The fractions were tested for growth factor activity as measured by the stimulation of DNA synthesis in BALB/c 3T3 cells. After 2 d in culture, two peaks of heparin-binding growth factor (HBGF) activity were detected, one eluting with 0.5 M NaCl, which could be shown to be platelet-derived growth factor (PDGF)-like, and the other eluting with 1.0 M NaCl. After 7-11 d in culture, when monocytes had clearly differentiated into macrophages, greater than 95% of the HBGF activity in conditioned medium consisted of the 1.0 M NaCl elution peak. This activity, which was designated macrophage-derived HBGF (MD-HBGF), was found to be a cationic heat-resistant polypeptide with a molecular weight in the range of 14-25 kDa. Analysis using Western blots and specific neutralizing antisera, as well as comparative heparin affinity analysis, indicated that MD-HBGF was not identical to other heparin-binding 3T3 cell growth factors known to be produced by macrophages, such as PDGF (AB, AA, and BB forms), acidic fibroblast growth factor, and basic fibroblast growth factor. In addition to stimulating mitogenesis in 3T3 cells, MD-HBGF also stimulated the proliferation of vascular smooth muscle cells, but did not stimulate the proliferation of vascular endothelial cells.


The effect of insulin and of anti-insulin serum on handling of sodium by the isolated, perfused kidney of the streptozotocin-diabetic rat.
We studied the effects of insulin and of anti-insulin serum (AIS) on sodium excretion in isolated diabetic and normal, fed, rat kidneys perfused at a constant pressure of 100 mm Hg with Krebs-Ringer bicarbonate buffer containing 7.5% bovine serum albumin and 5 mM glucose and gassed with 95% O2 and 5% CO2. The addition of insulin produced antinatriuresis in kidneys from diabetic rats but not in those from normal rats. Moreover, before the addition of insulin, the baseline rate of sodium excretion was greater in the diabetic rat kidney than in the normal rat kidney, while no differences in glomerular filtration rate (GFR) were observed. The addition of AIS (enough to bind 1.4 mU of insulin per milliliter of perfusate) produced a brisk natriuresis in kidneys from normal rats, and this natriuresis could be reversed by the addition of saturating amounts of insulin. By contrast, the addition of normal guinea pig serum to the perfusate of normal kidneys or AIS to the perfusate of diabetic kidneys was unassociated with natriuresis. We conclude that (a) insulin promoted antinatriuresis in diabetic but not in normal rat kidneys, (b) AIS promoted natriuresis in normal kidneys, and (c) this natriuresis was a specific effect of AIS and was reversed by the addition of insulin. Our data support the view that insulin-induced antinatriuresis depends on endogenous levels of circulating insulin and on the availability of insulin-binding sites in the kidney.


Cervical carcinoma: dynamic MR imaging with a turbo-FLASH technique.
The purpose of this study was to evaluate the characteristic findings of cervical carcinoma in dynamic MR imaging with a turbo-FLASH technique and to investigate the cause of the hyperintense rim surrounding the tumor on dynamic MR images by comparing them with the pathological findings in the resected specimens. Sixty consecutive patients with cervical carcinoma were included in this study. Dynamic MR imaging was performed with the rapid administration of Gd-DTPA using the turbo-FLASH technique. After dynamic MR imaging, T1-weighted spin echo (SE) images were obtained. Dynamic patterns of the lesions were investigated from images acquired in the early and late phases of dynamic MR imaging. The causes of the hyperintense rim were investigated by comparing dynamic MR images with the pathological findings. Most of the cervical carcinomas showed higher signal intensity than normal cervical stroma in the early phase of dynamic MR imaging. The pathological findings of the hyperintense rim on dynamic MR images differed according to the time of appearance of the hyperintense rim. Dynamic MR imaging with the turbo-FLASH technique, which shows high contrast between the lesion and normal cervical stroma, should be performed before obtaining contrast-enhanced T1-weighted SE images in patients with cervical carcinoma.


Effect of sampling frequency on estimates of cumulative nitrous oxide emissions.
It is generally recognized that soil N(2)O emissions can exhibit pronounced day-to-day variations; however, measurements of soil N(2)O flux with soil chambers typically are done only at discrete points in time. This study evaluated the impact of sampling frequency on the precision of cumulative N(2)O flux estimates calculated from field measurements. Automated chambers were deployed in a corn/soybean field and used to measure soil N(2)O fluxes every 6 h from 25 Feb. 2006 through 11 Oct. 2006. The chambers were located in two positions relative to the fertilizer bands-directly over a band or between fertilizer bands. Sampling frequency effects on cumulative N(2)O-N flux estimation were assessed using a jackknife technique where populations of N(2)O fluxes were constructed from the average daily fluxes measured in each chamber. These test populations were generated by selecting measured flux values at regular time intervals ranging from 1 to 21 d. It was observed that as sampling interval increased from 7 to 21 d, variances associated with cumulative flux estimates increased. At relatively frequent sampling intensities (i.e., once every 3 d) N(2)O-N flux estimates were within +/-10% of the expected value at both sampling positions. As the time interval between sampling was increased, the deviation in estimated cumulative N(2)O flux increased, such that sampling once every 21 d yielded estimates within +60% and -40% of the actual cumulative N(2)O flux. The variance of potential fluxes associated with the between-band positions was less than the over-band position, indicating that the underlying temporal variability impacts the efficacy of a given sampling protocol.


"Progress in Medicine Is Slower to Happen": Qualitative Insights Into How Trans and Gender Nonconforming Medical Students Navigate Cisnormative Medical Cultures at Canadian Training Programs.
Trans and gender nonconforming (TGNC) people face significant health disparities compared with their cisgender (nontrans) counterparts. Physician-level factors play a role in these disparities, and increasing the participation of individuals from sexuality and gender minority (SGM) communities in medical training has been proposed as one way of addressing this issue; however, very little is known about the experiences of TGNC medical students. This study aimed to understand the experiences of TGNC medical students in Canada. Between April 2017 and April 2018, 7 TGNC participants either currently enrolled in or recently graduated from a Canadian medical school completed audiorecorded semistructured interviews. Interviewers asked about experiences with admissions; academic, clinical, and social environments; and interactions with administration. The authors analyzed interviews using a constructivist grounded theory approach. The authors developed 5 overarching themes: navigating cisnormative medical culture; balancing authenticity, professionalism, and safety; negotiating privilege and power differentials; advocating for patients and curricular change; and seeking mentorship in improving access and quality of care to TGNC patients. This article focuses on the first theme, with associated subthemes of culture and context; interactions with classmates, curriculum, policy, and administration; and gendered spaces. The results of this study delineate heterogeneous experiences of medical cultures with a shared underlying pattern of erasure of TGNC people as both patients and clinicians. Findings were largely consistent with previously published recommendations for improving academic medical institutional climates for SGM people, though the need for access to appropriate gendered spaces beyond washrooms was highlighted.


The benefits of antimicrobial photodynamic therapy with RLP068 in the management of diabetic foot ulcers.
This article is the first part of a literature review concerning diabetic foot ulcers and the use of antimicrobial photodynamic therapy (PDT). Diabetic foot ulcers are associated with high morbidity, mortality, and healthcare costs. Natural healing is often delayed by microbial infection or colonisation, which might lead to serious complications, such as amputation. Furthermore, antibiotic treatment could have limited success because of the development of bacterial resistance and severely limited drug delivery to the ulcer due to vascular damage. PDT has antimicrobial effects and has been used to reduce the total and pathogenic microbial load in diabetic ulcers without inducing bacterial resistance. It is safe and can be used to improve outcomes. A clinical trial demonstrated that PDT with RLP068 reduced the microbial load of diabetic ulcers in 62 patients. This article reports previously published evidence and presents four, unpublished, clinical cases treated in the real-life setting.


Backbone Cyclization and Dimerization of LL-37-Derived Peptides Enhance Antimicrobial Activity and Proteolytic Stability.
Can antimicrobial activity and peptide stability of alpha-helical peptides be increased by making them into dimers and macrocycles? Here, we explore that concept by using KR-12 as the starting point for peptide engineering. KR-12 has previously been determined as the minimalized antimicrobial fragment of the human host defense peptide LL-37. Backbone-cyclized KR-12 dimers, tethered by linkers of two to four amino acid residues, were synthesized and their antimicrobial activity, proteolytic stability and structures characterized. A modified KR-12 sequence, with substitutions at previously identified key residues, were also included in the screening panel. The backbone cyclized KR-12 dimers showed improved antimicrobial activity and increased stability compared to monomeric KR-12. The most active cyclic dimer displayed 16-fold higher antibacterial activity compared to KR-12 against Pseudomonas aeruginosa and Staphylococcus aureus, and 8-fold increased fungicidal activity against Candida albicans. It also showed increased hemolytic and cytotoxic activity. Enhanced antimicrobial activity coincided with increased membrane permeabilization of liposomes with one distinct discrepancy: monomeric KR-12 was much less disruptive of liposomes with bacterial lipid composition compared to liposomes from fungal lipid extract. Circular dichroism showed that the four-residue linked most active cyclic dimer had 65% helical content when bound to lyso-phosphatidylglycerol micelles, indicating that the helical propensity of the parent peptide is maintained in the new macrocyclic form. In conclusion, the current work on KR-12 suggests that dimerization together with backbone cyclization is an effective strategy for improving both potency and stability of linear antimicrobial peptides.


Role of the lung microbiome in HIV pathogenesis.
The purpose of this review is to summarize recent findings on the lung microbiome in HIV-infected patients and associated pulmonary diseases, and the relationship of airway microbial communities to metabolic and immune signatures within this patient population. The lung microbiome in HIV infection is a relatively new and rapidly developing field; early studies in the field produced inconclusive evidence as to whether HIV-infection changes the lower airway microbiome. More recent microbiome investigations have addressed these inconsistencies by incorporating systems biology approaches and laboratory models. Several investigations have now identified enrichment of Prevotella, Veillonella, and Streptococcus in the lower airways as consistent correlates of advanced HIV-infection and HIV-associated pulmonary diseases. These bacteria are associated with specific metabolic and immune profiles within the lung and circulation, providing the first indication that the lung microbiome may play a functional role in the pathogenesis of HIV-infection and HIV-associated pulmonary disease. This review summarizes knowledge to date on the lung microbiome in HIV infection, as well as challenges and accomplishments in the field within the last 2 years. Although the lung microbiome in HIV infection is still an emerging field, recent studies have formed a framework for future functional analysis of microbes in HIV pathogenesis.


Functional module identification in protein interaction networks by interaction patterns.
Identifying functional modules in protein-protein interaction (PPI) networks may shed light on cellular functional organization and thereafter underlying cellular mechanisms. Many existing module identification algorithms aim to detect densely connected groups of proteins as potential modules. However, based on this simple topological criterion of 'higher than expected connectivity', those algorithms may miss biologically meaningful modules of functional significance, in which proteins have similar interaction patterns to other proteins in networks but may not be densely connected to each other. A few blockmodel module identification algorithms have been proposed to address the problem but the lack of global optimum guarantee and the prohibitive computational complexity have been the bottleneck of their applications in real-world large-scale PPI networks. In this article, we propose a novel optimization formulation LCP(2) (low two-hop conductance sets) using the concept of Markov random walk on graphs, which enables simultaneous identification of both dense and sparse modules based on protein interaction patterns in given networks through searching for LCP(2) by random walk. A spectral approximate algorithm SLCP(2) is derived to identify non-overlapping functional modules. Based on a bottom-up greedy strategy, we further extend LCP(2) to a new algorithm (greedy algorithm for LCP(2)) GLCP(2) to identify overlapping functional modules. We compare SLCP(2) and GLCP(2) with a range of state-of-the-art algorithms on synthetic networks and real-world PPI networks. The performance evaluation based on several criteria with respect to protein complex prediction, high level Gene Ontology term prediction and especially sparse module detection, has demonstrated that our algorithms based on searching for LCP(2) outperform all other compared algorithms. All data and code are available at http://www.cse.usf.edu/~xqian/fmi/slcp2hop/.


Mortality inequalities measured by socioeconomic indicators in Brazil: a scoping review.
Summarize the literature on the relationship between composite socioeconomic indicators and mortality in different geographical areas of Brazil. This scoping review included articles published between January 1, 2000, and August 31, 2020, retrieved by means of a bibliographic search carried out in the Medline, Scopus, Web of Science, and Lilacs databases. Studies reporting on the association between composite socioeconomic indicators and all-cause, or specific cause of death in any age group in different geographical areas were selected. The review summarized the measures constructed, their associations with the outcomes, and potential study limitations. Of the 77 full texts that met the inclusion criteria, the study reviewed 24. The area level of composite socioeconomic indicators analyzed comprised municipalities (n = 6), districts (n = 5), census tracts (n = 4), state (n = 2), country (n = 2), and other areas (n = 5). Six studies used composite socioeconomic indicators such as the Human Development Index, Gross Domestic Product, and the Gini Index; the remaining 18 papers created their own socioeconomic measures based on sociodemographic and health indicators. Socioeconomic status was inversely associated with higher rates of all-cause mortality, external cause mortality, suicide, homicide, fetal and infant mortality, respiratory and circulatory diseases, stroke, infectious and parasitic diseases, malnutrition, gastroenteritis, and oropharyngeal cancer. Higher mortality rates due to colorectal cancer, leukemia, a general group of neoplasms, traffic accident, and suicide, in turn, were observed in less deprived areas and/or those with more significant socioeconomic development. Underreporting of death and differences in mortality coverage in Brazilian areas were cited as the main limitation. Studies analyzed mortality inequalities in different geographical areas by means of composite socioeconomic indicators, showing that the association directions vary according to the mortality outcome. But studies on all-cause mortality and at the census tract level remain scarce. The results may guide the development of new composite socioeconomic indicators for use in mortality inequality analysis.


Clofarabine Dosing in a Patient With Acute Myeloid Leukemia on Intermittent Hemodialysis: Case Report and Review of the Literature.
Clofarabine containing chemotherapeutic regimens have demonstrated efficacy in the treatment of relapsed refractory acute myeloid leukemia. Nonetheless, there are limited data on the use of clofarabine in patients with renal failure. The present report describes the use of clofarabine in a patient with renal failure undergoing intermittent dialysis. We describe our rationale for dosing, clofarabine plasma levels obtained, and discuss our findings in the context of other available literature. Consistent with previous findings, intermittent hemodialysis was not found to be a reliable method of removing clofarabine in patients with renal insufficiency.


[Drug-induced cardiovascular complications].
Many drugs, including those prescribed in cardiology, can induce adverse cardiovascular side effects. Most of the anti-arrhythmia drugs have a negative inotropic action and proarrhythmic effects. Estroprogestative contraceptive drugs favor thromboembolitic events and increase the incidence of hypertension. Due to the risk of coronary insufficiency, substitutive hormonotherapy in hypothyroidism must be introduced gradually. Tricyclic anti-depressive drugs and neuroleptics can induce orthostatic hypotension. However, all these adverse effects can usually be avoided if every drug-specific contra-indication is respected, if doses are individually adjusted to the patient's age and physical condition and if the patient is closely monitored.


An Accessible Proteogenomics Informatics Resource for Cancer Researchers.
Proteogenomics has emerged as a valuable approach in cancer research, which integrates genomic and transcriptomic data with mass spectrometry-based proteomics data to directly identify expressed, variant protein sequences that may have functional roles in cancer. This approach is computationally intensive, requiring integration of disparate software tools into sophisticated workflows, challenging its adoption by nonexpert, bench scientists. To address this need, we have developed an extensible, Galaxy-based resource aimed at providing more researchers access to, and training in, proteogenomic informatics. Our resource brings together software from several leading research groups to address two foundational aspects of proteogenomics: (i) generation of customized, annotated protein sequence databases from RNA-Seq data; and (ii) accurate matching of tandem mass spectrometry data to putative variants, followed by filtering to confirm their novelty. Directions for accessing software tools and workflows, along with instructional documentation, can be found at z.umn.edu/canresgithub. Cancer Res; 77(21); e43-46. ©2017 AACR.


Unbiased classification of the elderly human brain proteome resolves distinct clinical and pathophysiological subtypes of cognitive impairment.
Cognitive impairment in the elderly features complex molecular pathophysiology extending beyond the hallmark pathologies of traditional disease classification. Molecular subtyping using large-scale -omic strategies can help resolve this biological heterogeneity. Using quantitative mass spectrometry, we measured ∼8000 proteins across >600 dorsolateral prefrontal cortex tissues with clinical diagnoses of no cognitive impairment (NCI), mild cognitive impairment (MCI), and Alzheimer's disease (AD) dementia. Unbiased classification of MCI and AD cases based on individual proteomic profiles resolved three classes with expression differences across numerous cell types and biological ontologies. Two classes displayed molecular signatures atypical of AD neurodegeneration, such as elevated synaptic and decreased inflammatory markers. In one class, these atypical proteomic features were associated with clinical and pathological hallmarks of cognitive resilience. We were able to replicate these classes and their clinicopathological phenotypes across two additional tissue cohorts. These results promise to better define the molecular heterogeneity of cognitive impairment and meaningfully impact its diagnostic and therapeutic precision.


Effect of cryosolvents and subzero temperatures on the hydrolysis of L-leucine-p-nitroanilide by porcine kidney leucine aminopeptidase.
The hydrolysis of L-leucine-p-nitroanilide by porcine kidney leucine aminopeptidase in aqueous mixed-solvent systems containing methanol, ethanol, dimethyl sulfoxide, and dimethylformamide has been investigated in the -30 to -23 degrees C temperature range. At 23 degrees C and pH* values in the 8-10 range, the enzyme is stable for over 25 h in solutions containing 50% v/v of any of these four cosolvents. Measurements of the tryptophan fluorescence of the enzyme at pH* 9.0 confirm that the enzyme is not denatured under these conditions. KM increases exponentially and kcat decreases linearly with increasing cosolvent concentration. Methanol, in particular, has a very small effect on KM. Ultrafiltration experiments demonstrate that there is no dissociation of monomers of the enzyme brought about by the presence of 50% v/v methanol or dimethyl sulfoxide. Preliminary tests with the partition method provide no evidence for an acyl-enzyme intermediate. The effect of pH* on kcat and KM in 50% v/v methanol is very similar to the effect of pH on these kinetic constants in aqueous solution. Lowering the temperature from 23 to 0 degree C does not alter the shape of the pH* profile obtained in 50% v/v methanol. The Arrhenius plot obtained in 50% v/v methanol is linear over the -30 to -23 degrees C temperature range, and the calculated energy of activation, 8.2 +/- 0.8 kcal/mol, is in good agreement with the value of 7.4 +/- 0.7 kcal/mol found for the reaction in aqueous solution. Collectively, these data indicate that methanol is the best cosolvent for cryoenzymological studies, that ethanol and dimethyl sulfoxide are also suitable cosolvents, and that the presence of any of these cosolvents at either ambient or subzero temperatures does not perturb the catalytic pathway.


Immunomodulatory activity of orphanin FQ/nociceptin on traumatic rats.
To explore the neuro-immune modulatory effect of orphanin FQ/nociceptin (OFQ) and opioid receptor like 1 (ORL1) receptor on the traumatic rats. The quantitative method of immuno-cytochemistry and i n situ hybridization combined with cytokine bioassay were used to detect the expression of endogenous OFQ and ORL1 and the production of interleukin-1 (IL-1) and tumor necrosis factor-alpha (TNF-alpha) from peritoneal macrophage. Strong signals for both OFQ immuno-reactive cells and ORL1 mRNA were detected in cerebral cortex, hippocampus, and hypothalamus in normal condition, whereas they were significantly reduced after trauma (P<0.05). However, the production of IL-1 and TNF-alpha from peritoneal macrophage was increased, when expressed as percentage of enhancement, the increment attained to 233 % and 521 % (sample dilution 1:4), 195 % and 566 % (1:8), 233 % and 757 % ( 1:16), 214 % and 622 % (1:32), respectively, after trauma. After icv injection of OFQ at doses of 0.055 nmol, 0.55 nmol, and 2.75 nmol, the units of IL-1 and TNF-alpha were reversed (P<0.05); however, the action of OFQ (0.55 nmol) was blocked by ORL1 selective antagonist [phe1psi(CH2-NH)Gly2]nociceptin-(1-13)-NH2. OFQ and ORL1, the new opioid peptide system, are involved in the immune response elicited by traumatic stress.


Discriminating stress from rest based on resting-state connectivity of the human brain: A supervised machine learning study.
Acute stress induces large-scale neural reorganization with relevance to stress-related psychopathology. Here, we applied a novel supervised machine learning method, combining the strengths of a priori theoretical insights with a data-driven approach, to identify which connectivity changes are most prominently associated with a state of acute stress and individual differences therein. Resting-state functional magnetic resonance imaging scans were taken from 334 healthy participants (79 females) before and after a formal stress induction. For each individual scan, mean time-series were extracted from 46 functional parcels of three major brain networks previously shown to be potentially sensitive to stress effects (default mode network (DMN), salience network (SN), and executive control networks). A data-driven approach was then used to obtain discriminative spatial linear filters that classified the pre- and post-stress scans. To assess potential relevance for understanding individual differences, probability of classification using the most discriminative filters was linked to individual cortisol stress responses. Our model correctly classified pre- versus post-stress states with highly significant accuracy (above 75%; leave-one-out validation relative to chance performance). Discrimination between pre- and post-stress states was mainly based on connectivity changes in regions from the SN and DMN, including the dorsal anterior cingulate cortex, amygdala, posterior cingulate cortex, and precuneus. Interestingly, the probability of classification using these connectivity changes were associated with individual cortisol increases. Our results confirm the involvement of DMN and SN using a data-driven approach, and specifically single out key regions that might receive additional attention in future studies for their relevance also for individual differences.


Magnetic resonance imaging of knee hyaline cartilage and intraarticular pathology.
Injuries to the hyaline cartilage of the knee joint are difficult to diagnose without invasive techniques. Even though these defects may be the most important prognostic factors in assessing knee joint injury, they are usually not diagnosed until arthrotomy or arthroscopy. Once injuries to hyaline cartilage are found and/or treated, no technique exists to follow these over time. Plain radiographs, arthrograms, and even computed tomography fail to detail most hyaline cartilage defects. We used magnetic resonance imaging (MRI) to evaluate five fresh frozen cadaver limbs and 10 patients whose pathology was known from arthrotomy or arthroscopic examination. Using a 0.35 Tesla superconducting magnet and spin-echo imaging technique with a head coil, we found that intraarticular fluid or air helped to delineate hyaline cartilage pathology. The multiplane capability of MRI proved to be excellent in detailing small (3 mm or more) defects on the femoral condyles and patellar surface. Cruciate ligaments were best visualized on sagittal oblique projections while meniscal pathology was best seen on true sagittal and coronal projections. MRI shows great promise in providing a noninvasive technique of evaluating hyaline cartilage defects, their response to treatment, and detailed anatomical information about cruciate ligaments and menisci.


Substantia nigra vulnerability after a single moderate diffuse brain injury in the rat.
Dementia and parkinsonism are late-onset symptoms associated with repetitive head injury, as documented in multiple contact-sport athletes. Clinical symptomatology is the likely phenotype of chronic degeneration and circuit disruption in the substantia nigra (SN). To investigate the initiating neuropathology, we hypothesize that a single diffuse brain injury is sufficient to initiate SN neuropathology including neuronal loss, vascular disruption and microglial activation, contributing to neurodegeneration and altered dopamine regulation. Adult, male Sprague-Dawley rats were subjected to sham or moderate midline fluid percussion brain injury. Stereological estimates indicated a significant 44% loss of the estimated total neuron number in the SN at 28-days post-injury, without atrophy of neuronal nuclear volumes, including 25% loss of tyrosine hydroxylase positive neurons by 28-days post-injury. Multi-focal vascular compromise occurred 1-2 days post-injury, with ensuing microglial activation (significant 40% increase at 4-days). Neurodegeneration (silver-stain technique) encompassed on average 21% of the SN by 7-days post-injury and increased to 29% by 28-days compared to sham (1%). Whole tissue SN, but not striatum, dopamine metabolism was altered at 28-days post-injury, without appreciable gene or protein changes in dopamine synthesis or regulation elements. Together, single moderate diffuse brain injury resulted in SN neurovascular pathology potentially associated with neuroinflammation or dopamine dysregulation. Compensatory mechanisms may preserve dopamine signaling acutely, but subsequent SN damage with aging or additional injury may expose clinical symptomatology of motor ataxias and dementia.


Variety of endogenous proviruses in the genomes of chickens of different breeds.
We have compared endogenous proviruses in DNA of chickens of 11 breeds by means of Southern's technique. Many of the endogenous virus loci found were missing from the genomes of the extensively studied white leghorn chickens, although some of the proviruses and especially ev-1 appeared to be widely spread among different chickens. Most of the proviruses were similar to Rous-associated virus (RAV-O) as judged by the EcoRI digestion patterns. Several genetically different proviruses were found in the DNA of 28 brown leghorns. They contain neither ev-1 provirus in their genome, nor any other one common for all individuals. All four Italian partridge-coloured chickens examined appeared to be free from ALV-related sequences in their DNA showing the possibility of normal life without known endogenous proviruses. Possible causes of inter-breed differences of chicken endogenous proviruses are discussed.


Co-catabolism of arginine and succinate drives symbiotic nitrogen fixation.
Biological nitrogen fixation emerging from the symbiosis between bacteria and crop plants holds promise to increase the sustainability of agriculture. One of the biggest hurdles for the engineering of nitrogen-fixing organisms is an incomplete knowledge of metabolic interactions between microbe and plant. In contrast to the previously assumed supply of only succinate, we describe here the CATCH-N cycle as a novel metabolic pathway that co-catabolizes plant-provided arginine and succinate to drive the energy-demanding process of symbiotic nitrogen fixation in endosymbiotic rhizobia. Using systems biology, isotope labeling studies and transposon sequencing in conjunction with biochemical characterization, we uncovered highly redundant network components of the CATCH-N cycle including transaminases that interlink the co-catabolism of arginine and succinate. The CATCH-N cycle uses N2 as an additional sink for reductant and therefore delivers up to 25% higher yields of nitrogen than classical arginine catabolism-two alanines and three ammonium ions are secreted for each input of arginine and succinate. We argue that the CATCH-N cycle has evolved as part of a synergistic interaction to sustain bacterial metabolism in the microoxic and highly acid environment of symbiosomes. Thus, the CATCH-N cycle entangles the metabolism of both partners to promote symbiosis. Our results provide a theoretical framework and metabolic blueprint for the rational design of plants and plant-associated organisms with new properties to improve nitrogen fixation.


Stable gastric pentadecapeptide BPC 157: novel therapy in gastrointestinal tract.
Stable gastric pentadecapeptide BPC 157 is an anti-ulcer peptidergic agent, safe in inflammatory bowel disease clinical trials (GEPPPGKPADDAGLV, M.W. 1419, PL 14736) and wound healing, stable in human gastric juice and has no reported toxicity. We focused on BPC 157 as a therapy in peridontitis, esophagus, stomach, duodenum, intestine, liver and pancreas lesions. Particularly, it has a prominent effect on alcohol-lesions (i.e., acute, chronic) and NSAIDs-lesions (interestingly, BPC 157 both prevents and reverses adjuvant arthritis). In rat esophagitis and failed function of both lower esophageal sphincter (LES) and pyloric sphincters (PS), BPC 157 increased pressure in both sphincters till normal and reduced esophagitis. However, in healthy rats, it may decrease (PS) or increase (LES) the pressure in sphincters. It has strong angiogenic potential, it acts protectively on endothelium, prevents and reverses thrombus formation after abdominal aorta anastomosis, affects many central disturbances (i.e., dopamine and 5-HT system), the NO-system (either L-arginine and L-NAME effects), endothelin, acts as a free radical scavenger (counteracting CCl4-, paracetamol-, diclofenac-injuries) and exhibits neuroprotective properties. BPC 157 successfully heals the intestinal anastomosis, gastrocutaneous, duodenocutaneous and colocutaneous fistulas in rats, as well as interacting with the NO-system. Interestingly, the fistula closure was achieved even when the BPC 157 therapy was postponed for one month. In short-bowel syndrome escalating throughout 4 weeks, the constant weight gain above preoperative values started immediately with peroral and parental BPC 157 therapy and the villus height, crypth depth and muscle thickness (inner (circular) muscular layer) additionally increased. Thus, BPC 157 may improve gastrointestinal tract therapy.


[Regulation of various isoenzymes of glutathione transferases of protein kinase A and cAMP].
Activation of purified glutathione transferase (CE 18.104.22.168) from the rabbit liver by cAMP-dependent protein kinase (protein kinase A) and activation of glutathione transferase from the rat liver and heart by cAMP preparations have been studied. A comparison of glutathione transferase activation on different substrates and the results of the inhibitor analysis of the activation phenomenon have shown that the second (mu) family of glutathione transferase isoenzymes (subunits, 3, 4, 6) is the most probable object of regulation. The first (alpha) family (subunits 1 and 2) and isoenzyme 5-5 are not probably regulated.


Fibrinogen-fibrin degradation products in menstrual blood from women with normal and excessive menstrual losses.
Fibrinogen-fibrin degradation products, FDP, in menstrual blood during the first three days of menstrual blood during the first three days of menstruation have been investigated. Two groups of women were studied, those with normal menstrual blood loss (15 women, mean loss 30 ml, range 8-60 ml) and those with menorrhagia (14 women, mean loss 222 ml, range 107-729 ml). The following results were obtained: 1). The FDP concentrations decreased during menstruation in both groups. 2). The FDP concentrations in the two were compared for each day of the menstrual period. No differences were found between the two groups. A possible explanation of the results is given: there is a higher rate of coagulation and fibrinolysis in the endometrium of women with menorrhagia compared with women with normal blood losses. The hypothesis is supported by results of studies in which tranexamic is supported by results of studies in which tranexamic acid, an inhibitor of fibrinolysis, was given to reduce the menstrual blood loss. 3). The FDP excretion patterns differed from women to women. There were no consistent differences between the menorrhagic and the control group.


Intranasal immunotherapy and polymerized grass pollen allergens.
The effects of immunotherapy by intranasal insufflation of Lolium perenne allergens modified with glutaraldehyde (Glu-Rye) was tested by a stratified placebo-controlled trial in 16 adult subjects with hay fever. The purpose of this pilot study was to determine whether the treatment could reduce nasal sensitivity to allergen in provocation tests. Modification of the allergen was shown to include polymerization and reduced allergenicity, but retained immunogenicity. Nasal sensitivity to unmodified L. perenne allergens was measured by threshold-type quantitative nasal challenge and posterior rhinometry for measurement of nasal airway resistance. In challenge tests after a 10-week course of intranasal preseasonal treatment with Glu-Rye, nasal sensitivity was reduced, but this effect was not maintained when testing was repeated 9 months after treatment. Post-seasonal serum levels of IgE antibodies to L. perenne were also not significantly affected by the treatment.


Disseminated infection caused by urease-negative Cryptococcus neoformans.
We report a case of fungemia and disseminated disease caused by a urease-negative strain of Cryptococcus neoformans in a patient with the acquired immune deficiency syndrome. Except for failure to hydrolyze urea, the microbiological characteristics of the isolate were typical of C. neoformans. Laboratory specialists should be aware of the occurrence of atypical strains of C. neoformans, particularly those recovered from patients with the acquired immune deficiency syndrome.


Molecular modeling and experimental approaches toward designing a minimalist protein having Fc-binding activity of Staphylococcal protein A.
Protein A (PA), a cell wall constituent of Staphylococcus aureus, has got the unique property of binding with the Fc fragment of IgG from various species. The sequence data indicate five highly homologous Fc-binding regions in protein A. Computer sequence analysis provided the tryptic and chymotryptic fragments of IgG-binding domains of protein A. Molecular modeling in conjunction with molecular mechanical calculation has been used to search for the smallest possible proteolytic fragments of PA, still retaining Fc-binding activity. A 20-residue peptide (typtic fragment) and a 16-residue peptide (chymotryptic fragment) have been indicated, by molecular modeling studies, to possess IgG-binding affinity comparable to that of the B domain of Protein A. Binding of a 20-residue peptide has been substantiated experimentally by immunoprecipitation, capillary electrophoresis, and circular dichroism spectroscopic analyses.


Evidence concerning the mechanism of insulin-receptor interaction and the structure of the insulin receptor from biological properties of covalently linked insulin dimers.
Covalently linked insulin dimers have been prepared by cross-linking two insulin monomers with a flexible suberoyl chain at either the B1 phenylalanine or the B29 lysine residue. Binding potencies of dimers determined by inhibition of binding of 125I-insulin to isolated rat liver plasma membranes or adipocytes were 2.5-7-fold greater than their abilities to stimulate lipogenesis in adipocytes. Rates of liver plasma-membrane-associated degradation of labelled insulin and dimers, measured by gel filtration, were similar at 37 degrees C. Binding and lipogenesis potencies of dimers prepared by substitution of each monomeric half of an asymmetrical dimer with desoctapeptide insulin, an almost inactive derivative, implicated the B1-cross-linked monomeric half as predominantly interacting with the insulin receptor. These results suggest that (1) dimers bind univalently to a bivalent insulin-receptor complex, in which the two individual binding subunits are arranged with anti-parallel symmetry and (2) the mechanism by which insulin binds and initiates its biological responses requires a conformational change within the insulin-receptor complex and/or in the insulin molecule for full biological expression.


Modest reversal of metabolic syndrome manifestations with vitamin D status correction: a 12-month prospective study.
Numerous cross-sectional studies have noted significant negative associations between circulating levels of 25-hydroxyvitamin D and cardiometabolic risk factors, highlighting potential extraskeletal functions of this sterol hormone. Prospective studies, however, have been limited; and hence, no cause-and-effect relations can be inferred. This study aims to determine whether vitamin D status correction can reverse already established manifestations of the metabolic syndrome (MetS). A total of 59 adult nondiabetic, overweight, and obese Saudis (31 male, 28 female) were prospectively enrolled in this 1-year interventional study. Anthropometry and biochemical evaluation were performed, including determination of serum 25-hydroxyvitamin D, calcium, and phosphorous concentrations, as well as fasting blood glucose and lipid profile. Subjects were advised to regularly expose themselves to sunlight and increase intake of vitamin D-rich foods. All measurements were repeated 6 and 12 months later. At the initial baseline visit, the prevalence of both low high-density lipoprotein cholesterol and hypertension was significantly increased among patients with 25-vitamin D deficiency (P < .05), even after adjusting for sex and body mass index. Overall prevalence of MetS patients by the modified National Health and Nutrition Examination Survey Adult Treatment Panel III definition decreased from 25.2% to 13.0%; and this was largely due to a parallel decrease in the prevalence of low high-density lipoprotein cholesterol, triglycerides, and hypertension. Optimization of vitamin D status through sun exposure and increased intake of a vitamin D-rich diet can lead to an improved cardiometabolic profile, offering a promising nonpharmacologic approach in the prevention of MetS manifestations.


Muscle ultrasound: A useful tool in newborn screening for infantile onset pompe disease.
Our study aimed to evaluate the utility of muscle ultrasound in newborn screening of infantile-onset Pompe disease (IOPD) and to establish a system of severity grading. We retrospectively selected 35 patients with initial low acid alpha-glucosidase (GAA) activity and collected data including muscle ultrasound features, GAA gene mutation, activity/performance, and pathological and laboratory findings. The echogenicity of 6 muscles (the bilateral vastus intermedius, rectus femoris, and sartorius muscles) was compared to that of epimysium on ultrasound and rated either 1 (normal), 2 (mildly increased), or 3 (obviously increased). These grades were used to divide patients into 3 groups. IOPD was present in none of the grade-1 patients, 5 of 9 grade-2 patients, and 5 of 5 grade-3 patients (P < .001). Comparing grade-2 plus grade-3 patients to grade-1 patients, muscle ultrasound detected IOPD with a sensitivity and specificity of 100.0% (95% confidence interval [CI]: 69.2%-100%) and 84.0% (95% CI: 63.9%-95.5%), respectively. The mean number of affected muscles was larger in grade-3 patients than in grade-2 patients (4.2 vs. 2.0, P = .005). Mean alanine transaminase (ALT), aspartate transaminase (AST), creatine kinase (CK), and lactate dehydrogenase (LDH) levels were differed significantly different between grade-3 and grade-1 patients (P < .001). Because it permits direct visualization of injured muscles, muscle ultrasound can be used to screen for IOPD. Our echogenicity grades of muscle injury also correlate well with serum levels of muscle-injury biochemical markers.


[Amniocentesis: psychological stress and coping by the pregnant female].
Anxiety as induced by aminocentesis and consequential strategies for coping there with, are the focus of this study. A group of 32 women were selected for AC due to age, and a second group of 30 women as a result of risk factors in the case history. Anxiety related to AC was self-assessed by the patients and coping strategies were rated according to the "Bernese Coping Modes" prospectively and retrospectively. Results indicate, that AC is anxiety-inducing, but does not vary with parity, indication or the degree of information, but solely with the fact of preexisting experience or absence of such with AC. 50% of the patients would have been willing to interrupt the pregnancy, had the result indicated a high risk of malformation. The coping with the waiting time until the result of AC was known was such, that the "age" group showed a great deal of optimism, whereas the risk group used several coping modes, which are more complex and also potentially problematic. This does not make the somatic risk group a psychological hazard group, but nevertheless suggests, that professionals should stand by to intervene on a psychological plane.


Functional health literacy and understanding of medications at discharge.
The objective of this study was to evaluate patient knowledge of newly prescribed medication after hospital discharge. We reviewed the charts of 172 patients who were discharged from February 1, 2006, through April 25, 2006, from the internal medicine residency service at a community-based teaching hospital with prescriptions for 1 or more new medications. Between 4 and 18 days after discharge, patients were contacted by telephone and asked about the name, number, dosages, schedule, purpose, and adverse effects of the new medication(s) and whether they could name their medical contact person. We recorded the number of correct answers, patient age, and years of education. Of the survey respondents, 86% were aware that they had been prescribed new medications, but fewer could identify the name (64%) or number (74%) of new medications or their dosages (56%), schedule (68%), or purpose (64%). Only 11% could recall being told of any adverse effects, and only 22% could name at least 1 adverse effect. Older patients tended to answer fewer questions correctly (P=.02). We observed no association between the number of correctly answered questions and years of education (P=.57), time between discharge and survey (P=.17), or number of new medications (P=.65). Overall, we found that patients had limited knowledge about their medications after discharge from an internal medicine residency service, with age but not years of education significantly associated with level of knowledge.


Service evaluation of the GOALS family-based childhood obesity treatment intervention during the first 3 years of implementation.
To evaluate the impact of the GOALS (Getting Our Active Lifestyles Started) family-based childhood obesity treatment intervention during the first 3 years of implementation. Single-group repeated measures with qualitative questionnaires. Community venues in a socioeconomically deprived, urban location in the North-West of England. 70 overweight or obese children (mean age 10.5 years, 46% boys) and their parents/carers who completed GOALS between September 2006 and March 2009. GOALS was a childhood obesity treatment intervention that drew on social cognitive theory to promote whole family lifestyle change. Sessions covered physical activity (PA), diet and behaviour change over 18 2 h weekly group sessions (lasting approximately 6 months). A Template for Intervention Description and Replication (TIDieR) checklist of intervention components is provided. The primary outcome measure was child body mass index (BMI) z-score, collected at baseline, post-intervention and 12 months. Secondary outcome measures were child self-perceptions, parent/carer BMI and qualitative changes in family diet and PA (parent/carer questionnaire). Child BMI z-score reduced by 0.07 from baseline to post-intervention (p<0.001) and was maintained at 12 months (p<0.05). There was no change in parent/carer BMI or child self-perceptions, other than an increase in perceived social acceptance from baseline to post-intervention (p<0.05). Parents/carers reported positive changes to family PA and dietary behaviours after completing GOALS. GOALS completion was associated with small improvements in child BMI z-score and improved family PA and dietary behaviours. Several intervention modifications were necessary during the implementation period and it is suggested childhood obesity treatment interventions need time to embed before a definitive evaluation is conducted. Researchers are urged to use the TIDieR checklist to ensure transparent reporting of interventions and facilitate the translation of evidence to practice.


The effect of spirituality on health and healing: a critical review for athletic trainers.
To provide a comprehensive overview of the published literature regarding the effect of spirituality on health and healing. I searched MEDLINE from 1976 to 1999 using the terms "spirituality," "religion," "faith," "healing," and "health." Strong scientific evidence suggests that individuals who regularly participate in spiritual worship services or related activities and who feel strongly that spirituality or the presence of a higher being or power are sources of strength and comfort to them are healthier and possess greater healing capabilities. Numerous research investigations have reported positive correlations between spirituality and decreased rates of stroke, cancer, cardiovascular disease, hypertension, drug abuse, suicide, and general mortality. It has been suggested that faith is beneficial for health and healing because it helps people avoid unhealthy behaviors such as smoking and excessive drinking. However, studies designed to statistically control for such factors also report positive associations between spirituality and health in individuals with unhealthy behaviors. The impact of spirituality on health and healing is a topic that has been virtually ignored in the disciplines of athletic training and sports medicine. Because of their lack of exposure to this topic, most athletic trainers are unaware of the many positive associations that exist between spirituality and health and healing. The available literature base regarding this topic is quite large; its findings need to be explored and integrated into our profession.


Drosophila Gyf/GRB10 interacting GYF protein is an autophagy regulator that controls neuron and muscle homeostasis.
Autophagy is an essential process for eliminating ubiquitinated protein aggregates and dysfunctional organelles. Defective autophagy is associated with various degenerative diseases such as Parkinson disease. Through a genetic screening in Drosophila, we identified CG11148, whose product is orthologous to GIGYF1 (GRB10-interacting GYF protein 1) and GIGYF2 in mammals, as a new autophagy regulator; we hereafter refer to this gene as Gyf. Silencing of Gyf completely suppressed the effect of Atg1-Atg13 activation in stimulating autophagic flux and inducing autophagic eye degeneration. Although Gyf silencing did not affect Atg1-induced Atg13 phosphorylation or Atg6-Pi3K59F (class III PtdIns3K)-dependent Fyve puncta formation, it inhibited formation of Atg13 puncta, suggesting that Gyf controls autophagy through regulating subcellular localization of the Atg1-Atg13 complex. Gyf silencing also inhibited Atg1-Atg13-induced formation of Atg9 puncta, which is accumulated upon active membrane trafficking into autophagosomes. Gyf-null mutants also exhibited substantial defects in developmental or starvation-induced accumulation of autophagosomes and autolysosomes in the larval fat body. Furthermore, heads and thoraxes from Gyf-null adults exhibited strongly reduced expression of autophagosome-associated Atg8a-II compared to wild-type (WT) tissues. The decrease in Atg8a-II was directly correlated with an increased accumulation of ubiquitinated proteins and dysfunctional mitochondria in neuron and muscle, which together led to severe locomotor defects and early mortality. These results suggest that Gyf-mediated autophagy regulation is important for maintaining neuromuscular homeostasis and preventing degenerative pathologies of the tissues. Since human mutations in the GIGYF2 locus were reported to be associated with a type of familial Parkinson disease, the homeostatic role of Gyf-family proteins is likely to be evolutionarily conserved.


Functional analysis of a mosquito short-chain dehydrogenase cluster.
The short-chain dehydrogenases (SDR) constitute one of the oldest and largest families of enzymes with over 46,000 members in sequence databases. About 25% of all known dehydrogenases belong to the SDR family. SDR enzymes have critical roles in lipid, amino acid, carbohydrate, hormone, and xenobiotic metabolism as well as in redox sensor mechanisms. This family is present in archaea, bacteria, and eukaryota, emphasizing their versatility and fundamental importance for metabolic processes. We identified a cluster of eight SDRs in the mosquito Aedes aegypti (AaSDRs). Members of the cluster differ in tissue specificity and developmental expression. Heterologous expression produced recombinant proteins that had diverse substrate specificities, but distinct from the conventional insect alcohol (ethanol) dehydrogenases. They are all NADP⁺-dependent and they have S-enantioselectivity and preference for secondary alcohols with 8-15 carbons. Homology modeling was used to build the structure of AaSDR1 and two additional cluster members. The computational study helped explain the selectivity toward the (10S)-isomers as well as the reduced activity of AaSDR4 and AaSDR9 for longer isoprenoid substrates. Similar clusters of SDRs are present in other species of insects, suggesting similar selection mechanisms causing duplication and diversification of this family of enzymes.


Regulation of inositol 1,4,5-trisphosphate metabolism in insulin-secreting RINm5F cells.
Factors underlying the transience of inositol 1,4,5-trisphosphate [Ins(1,4,5)P3] accumulation following muscarinic stimulation of RINm5F cells were examined. Transience was not due to a protein kinase C-mediated stimulation of Ins(1,4,5)P3 dephosphorylation, since pretreatment of cells with tetradecanoyl-phorbol acetate (TPA) did not alter the rate of this conversion. However, preincubation with TPA did inhibit carbamoylcholine-stimulated Ins(1,4,5)P3 formation. In permeabilized cells, the conversion of Ins(1,4,5)P3 to inositol 1,3,4,5-tetrakisphosphate [Ins(1,3,4,5)P4] was slightly enhanced in the presence of TPA or cyclic AMP, but much more markedly by raising the Ca2+ concentration from 10(-7) M to 10(-6) or 10(-5) M. In intact cells the most rapid rate of accumulation of Ins(1,4,5)P3 and Ins(1,3,4,5)P4 occurred in the first 2 s following stimulation, whereas the levels of inositol 1,4-bisphosphate were not increased until after 5 s. This suggests that Ins(1,4,5)P3 kinase is chiefly responsible for the early disposal of Ins(1,4,5)P3 following cellular stimulation. The results are consistent with the proposal that the transient accumulation of Ins(1,4,5)P3 is due both to its enhanced metabolism via the Ca2+-calmodulin-sensitive Ins(1,4,5)P3 kinase, as well as a down-regulation of phosphatidylinositol 4,5-bisphosphate hydrolysis.