kappa-Opioid receptors in marrow stroma mediate the hematopoietic effects of melatonin-induced opioid cytokines.
Melatonin exerts colony-stimulating activity and rescues myeloid progenitors from apoptosis, induced either in vivo or in vitro by cancer chemotherapy compounds in tumor-bearing mice. These effects are mediated mainly by T-helper cell-derived opioid cytokines with an apparent molecular mass of 15 kDa and 67 kDa that are recognized both by anti-interleukin-4 and anti-dynorphin B antibodies. These putative new cytokines were named melatonin-induced-opioid (MIO). The most active and naltrexone-sensitive MIO was the smaller molecule, which was called MIO15 and found to act on an opioid-binding site present in adherent bone marrow cells. However, the hematopoietic action of MIO15 was dependent on the presence of colony-stimulating factors (CSF). To investigate this point, we studied the ability of melatonin to rescue granulocyte/macrophage colony-forming units (GM-CFU) in the bone marrow of tumor-free animals treated with cancer chemotherapeutic compounds. We found that melatonin not only is unable to protect bone marrow GM-CFU unless the mice are transplanted with Lewis lung carcinoma (LLC), but also that melatonin seems to increase the myelotoxicity of cyclophosphamide in tumor-free mice. In both tumor-bearing or healthy mice, the effect of melatonin is negated by naltrexone, indicating the involvement of MIO15. Competition studies classified the target opioid-binding site as a kappa-opioid receptor with low affinity in tumor-free mice and high affinity in LLC-implanted mice. LLC is known to release CSF. Consistently, addition of CSF in the form of lung-conditioned medium (LCM) to adherent bone marrow cells increased the affinity of the kappa-opioid receptor. Addition of antigranulocyte/macrophage colony-stimulating factor (GM-CSF) mAbs neutralized the effect of LCM. In conclusion, the affinity state of the kappa-opioid receptors in stromal bone marrow cells seems to modulate the hematopoietic effect of melatonin and/or MIO15.


Visuomotor cerebellum in human and nonhuman primates.
In this paper, we will review the anatomical components of the visuomotor cerebellum in human and, where possible, in non-human primates and discuss their function in relation to those of extracerebellar visuomotor regions with which they are connected. The floccular lobe, the dorsal paraflocculus, the oculomotor vermis, the uvula-nodulus, and the ansiform lobule are more or less independent components of the visuomotor cerebellum that are involved in different corticocerebellar and/or brain stem olivocerebellar loops. The floccular lobe and the oculomotor vermis share different mossy fiber inputs from the brain stem; the dorsal paraflocculus and the ansiform lobule receive corticopontine mossy fibers from postrolandic visual areas and the frontal eye fields, respectively. Of the visuomotor functions of the cerebellum, the vestibulo-ocular reflex is controlled by the floccular lobe; saccadic eye movements are controlled by the oculomotor vermis and ansiform lobule, while control of smooth pursuit involves all these cerebellar visuomotor regions. Functional imaging studies in humans further emphasize cerebellar involvement in visual reflexive eye movements and are discussed.


Hepatitis C Virus Vaccine Research: Time to Put Up or Shut Up.
Unless urgently needed to prevent a pandemic, the development of a viral vaccine should follow a rigorous scientific approach. Each vaccine candidate should be designed considering the in-depth knowledge of protective immunity, followed by preclinical studies to assess immunogenicity and safety, and lastly, the evaluation of selected vaccines in human clinical trials. The recently concluded first phase II clinical trial of a human hepatitis C virus (HCV) vaccine followed this approach. Still, despite promising preclinical results, it failed to protect against chronic infection, raising grave concerns about our understanding of protective immunity. This setback, combined with the lack of HCV animal models and availability of new highly effective antivirals, has fueled ongoing discussions of using a controlled human infection model (CHIM) to test new HCV vaccine candidates. Before taking on such an approach, however, we must carefully weigh all the ethical and health consequences of human infection in the absence of a complete understanding of HCV immunity and pathogenesis. We know that there are significant gaps in our knowledge of adaptive immunity necessary to prevent chronic HCV infection. This review discusses our current understanding of HCV immunity and the critical gaps that should be filled before embarking upon new HCV vaccine trials. We discuss the importance of T cells, neutralizing antibodies, and HCV genetic diversity. We address if and how the animal HCV-like viruses can be used for conceptualizing effective HCV vaccines and what we have learned so far from these HCV surrogates. Finally, we propose a logical but narrow path forward for HCV vaccine development.


Voltammetric methods for electrochemical characterization and quantification of artemether-based antimalarials.
Every year substandard and falsified (SF) artemisinin derivative-based antimalarials are responsible for the loss of 450 000 deaths and billions of GBP. The lack of infrastructure and funds to support pharmaceutical quality control in many low-and-middle-income countries contributes to this problem. This work assesses fitness for purpose of voltammetric methods for identification and quantification of artemether in the presence of excipients. Electrochemical characterization of artemether using cyclic voltammetry shows that the reduction of artemether is chemically irreversible within the potential range of -0.4 V to -1.4 V. A chronocoulometric quantification algorithm for artemether is created and tested with pure artemether, as well as filtered and unfiltered Riamet® tablets. Filtration of Riamet® tablets provides no additional benefit for the quantification of artemether in Riamet®. In addition, artemether's response to pH indicates possible protonation and coupled homogeneous chemistry. Finally, sodium sulfite is an effective means of removing dissolved oxygen and improving artemether signal resolution in air-equilibrated PBS. This concludes that electrochemical analysis is a promising method for artemether identification and quantification.


Spin-dependent quantum interference in nonlocal graphene spin valves.
Up to date, all spin transport experiments on graphene were done in a semiclassical regime, disregarding quantum transport properties such as phase coherence and interference. Here we show that in a quantum coherent graphene nanostructure the nonlocal voltage is strongly modulated. Using nonlocal measurements, we separate the signal in spin-dependent and spin-independent contributions. We show that the spin-dependent contribution is about 2 orders of magnitude larger than the spin-independent one, when corrected for the finite polarization of the electrodes. The nonlocal spin signal is not only strongly modulated but also changes polarity as a function of the applied gate voltage. By locally tuning the carrier density in the constriction via a side gate electrode we show that the constriction plays a major role in this effect. Our results show the potential of quantum coherent graphene nanostructures for the use in future spintronic devices.


Clinical and cytogenetic survey of 39 individuals with Prader-Labhart-Willi syndrome.
In a clinical and cytogenetic survey of 39 individuals with Prader-Labhart-Willi syndrome (PLWS) (23 males and 16 females ranging in age from 2 weeks to 39 years), an interstitial deletion of chromosome 15 (breakpoints q11 and q13) was identified in 21 cases and apparently normal chromosomes in the remainder. Studies of parental chromosome 15 variants showed that the del[15q] was paternal in origin, although chromosomes of both parents were normal. All chromosome deletions were de novo events. Possible causes for the chromosome deletion and the role of chromosome rearrangements in individuals with PLWS are discussed. Clinical characteristics of the deletion and nondeletion groups were recorded and compared with 124 individuals reported in the literature. Individuals with the chromosome deletion were found to have lighter hair, eye, and skin color, greater sun sensitivity, and higher intelligence scores than individuals with normal chromosomes. Correlation studies of metacarpophalangeal pattern profile variables and dermatoglyphic findings indicate apparent homogeneity of the deletion group and heterogeneity of individuals with PLWS and normal chromosomes.


Effects of asbestos and smoking on the levels and rates of change of lung function in a crocidolite exposed cohort in Western Australia.
Increased rates of death from asbestos related diseases have been reported in former workers and residents exposed to crocidolite (blue asbestos) at Wittenoom, Western Australia. Exposure to asbestos is associated with reduced static lung volumes, gas transfer and lung compliance, and a restrictive ventilatory abnormality. The effects of crocidolite exposure and smoking history on levels and rates of change of lung function were evaluated using a linear mixed model. Lung function was measured as forced expiratory volume in 1 second (FEV1), forced vital capacity (FVC), and FEV1/FVC. Cumulative doses of asbestos and the presence of radiographic asbestosis were associated with lower levels of FEV1 and FVC and a steeper decline during the period of observation. Subjects exposed to asbestos at a younger age had lower levels of FEV1 and FVC. Current smokers had lower levels and a steeper decline in lung function than never smokers. No significant interactions between crocidolite exposure and smoking on the levels or rates of change of lung function were found. The deleterious effects of crocidolite exposure on lung function persist in this population, despite asbestos exposure having ceased more than 30 years ago. No significant interactions were found in this population between asbestos and smoking at the first visit or longitudinally.


Characterization and quantification of anthocyanins in grape juices obtained from the grapes cultivated in Korea by HPLC/DAD, HPLC/MS, and HPLC/MS/MS.
The characterization and quantification of anthocyanins in grape cultivars of Oll-Meoru (Vitis coignetiaexVitis labrusca), Neut-Meoru (Vitis coignetiaexVitis labrusca), Muscal Bailey A. (Vitis labruscana), and Campbell Early (Vitis labruscaxV. vinifera) cultivated in Korea were carried out by partial purification through XAD-7 column chromatography followed by C-18 HPLC/diode array detector (DAD), HPLC/MS, and HPLC/MS/MS analyses. The column oven temperature during the reverse phase C-18 HPLC greatly affected the separation of individual anthocyanins. The result showed that the optimum column oven temperature was 35 degrees C. Sixteen different anthocyanins (11 nonacylated and 5 acylated anthocyanins) were identified in the grape juices. Oll-Meoru, Neut-Meoru, and Muscat Bailey A (MBA) grape juices contained only nonacylated anthocyanins. Oll-Meoru and Neut-Meoru grape juices had same anthocyanins, but their proportions were considerably different. Peonidin 3,5-diglucoside and malvidin 3,5-diglucoside were the major anthocyanins in Oll-Meoru grape juice. Delphinidin 3-glucoside was, however, the major anthocyanin in Neut-Meoru grape juice. Peonidin 3-glucoside and malvidin 3-glucoside were the most abundant anthocyanins in Muscal Bailey A grape juice. Campbell Early grape juice contained both nonacylated and acylated anthocyanins. Cyanidin 3-(p-coumaroyl)glucoside-5-glucoside and peonidin 3-(p-coumaroyl)glucoside-5-glucoside were the most abundant anthocyanins in Campbell Early grape juice. Total anthocyanin contents were greatly different in different grape jucies, with the highest in Neut-Meoru juice (1043.5 microg/mL), followed by Oll-Meoru (997.7 microg/mL), MBA (390.2 microg/mL), and Campbell Early (183.9 microg/mL) juices. The total anthocyanin content in Neut-Meoru grape juice was 5.67 times higher than that in Campbell Early grape juice. This represents the 1st report on the systematic characterization and quantification of anthocyanins in the juices of these grapes cultivated in Korea.


Sensitivity and performance time in MRI dephasing artifact reduction methods.
Although shimming can improve static field inhomogeneity, local field imperfections induced by tissue susceptibility differences cannot be completely corrected and can cause substantial signal loss in gradient echo images through intravoxel dephasing. Dephasing increases with voxel size so that one simple method of reducing the effect is to use thin slices. Signal-to-noise ratio (SNR) can then be increased by averaging over the subslices to form the final, thick slice. We call this method subslice averaging or SSAVE. Alternatively, a range of different amplitude slice select rephase gradients can be used to compensate for different susceptibility induced gradient offsets. The final image can then be formed by combining individual images in a variety of ways: summation, summation of the squares of the images, forming the maximum intensity projection of the image set, and Fourier transformation followed by summation. We show here that, contrary to previous claims, the theoretical sensitivity (i.e., SNR divided by the square root of the imaging time) of all these alternative methods is very similar. However, performance time (i.e., minimum-imaging time) of the simplest method, SSAVE, is much shorter than that of alternatives. This is confirmed experimentally on phantoms and anesthetized mice. Magn Reson Med 45:470-476, 2001.


Crohn's Disease Exclusion Diet Plus Partial Enteral Nutrition Induces Sustained Remission in a Randomized Controlled Trial.
Exclusive enteral nutrition (EEN) is recommended for children with mild to moderate Crohn's disease (CD), but implementation is challenging. We compared EEN with the CD exclusion diet (CDED), a whole-food diet coupled with partial enteral nutrition (PEN), designed to reduce exposure to dietary components that have adverse effects on the microbiome and intestinal barrier. We performed a 12-week prospective trial of children with mild to moderate CD. The children were randomly assigned to a group that received CDED plus 50% of calories from formula (Modulen, Nestlé) for 6 weeks (stage 1) followed by CDED with 25% PEN from weeks 7 to 12 (stage 2) (n = 40, group 1) or a group that received EEN for 6 weeks followed by a free diet with 25% PEN from weeks 7 to 12 (n = 38, group 2). Patients were evaluated at baseline and weeks 3, 6, and 12 and laboratory tests were performed; 16S ribosomal RNA gene (V4V5) sequencing was performed on stool samples. The primary endpoint was dietary tolerance. Secondary endpoints were intention to treat (ITT) remission at week 6 (pediatric CD activity index score below 10) and corticosteroid-free ITT sustained remission at week 12. Four patients withdrew from the study because of intolerance by 48 hours, 74 patients (mean age 14.2 ± 2.7 years) were included for remission analysis. The combination of CDED and PEN was tolerated in 39 children (97.5%), whereas EEN was tolerated by 28 children (73.6%) (P = .002; odds ratio for tolerance of CDED and PEN, 13.92; 95% confidence interval [CI] 1.68-115.14). At week 6, 30 (75%) of 40 children given CDED plus PEN were in corticosteroid-free remission vs 20 (59%) of 34 children given EEN (P = .38). At week 12, 28 (75.6%) of 37 children given CDED plus PEN were in corticosteroid-free remission compared with 14 (45.1%) of 31 children given EEN and then PEN (P = .01; odds ratio for remission in children given CDED and PEN, 3.77; CI 1.34-10.59). In children given CDED plus PEN, corticosteroid-free remission was associated with sustained reductions in inflammation (based on serum level of C-reactive protein and fecal level of calprotectin) and fecal Proteobacteria. CDED plus PEN was better tolerated than EEN in children with mild to moderate CD. Both diets were effective in inducing remission by week 6. The combination CDED plus PEN induced sustained remission in a significantly higher proportion of patients than EEN, and produced changes in the fecal microbiome associated with remission. These data support use of CDED plus PEN to induce remission in children with CD. Clinicaltrials.gov no: NCT01728870.


Guiding and cooling of atoms in an interference field composed of two hollow beams.
We propose a new scheme to guide and cool three-level alkali-metal atoms in a blue-detuned interference field composed of two counter-propagating doughnut hollow beams, and analyze the intensity distribution of the interference field of the two hollow beams and its intensity gradient one. Our study shows that the high intensity gradient of the interference field is desirable to realize intensity-gradient cooling for the guided atoms, and the minimum optical potential at the nodes of the interference field is high enough to guide almost all atoms released from a standard magneto-optical trap. We also perform Monte-Carlo simulations for dynamic process of the intensity-gradient cooling, and show that an (87)Rb atomic sample with a temperature of 120 muK can be directly cooled to a final equilibrium temperature of 4.71 muK in our guiding scheme.


Pulmonary function tests in extremely low gestational age infants at one year of age.
Identifying neonatal and post-discharge exposures among extremely low gestational age newborns (ELGANs) that drive increased pulmonary morbidity and abnormal lung function at 1 year of age proves challenging. The NIH-sponsored Prematurity and Respiratory Outcomes Program (PROP), evaluated infant pulmonary function tests (iPFTs) at 1 year corrected age to determine which demographic and clinical factors are associated with abnormal lung function. iPFTs were performed on a PROP subcohort of 135 participants following Institutional Review Board (IRB)-approved written consent. Demographic data, Neonatal Intensive Care Unit (NICU) clinical care, and post-NICU exposures were analyzed for association with iPFTs. A significant decrease in forced expiratory volume at 0.5 s (FEV0.5 ) and/or forced expiratory flows at 75% of forced vital capacity (FEF75 ), were associated with male sex and African American race. Clinical factors including longer duration of ventilatory support, exposure to systemic steroids, and weight less than the 10th percentile at 36 weeks postmenstrual age were also associated with airflow obstruction, whereas supplemental oxygen requirement and bronchopulmonary dysplasia were not. Additionally, the need for respiratory medications, technology, or hospitalizations during the first year, ascertained by a quarterly survey, were the only post-NICU factors associated with decreased FEV0.5 and FEF75 . Only 7% of infants had reversible airflow obstruction. Neonatal demographic factors, respiratory support in the NICU, and a history of greater post-NICU medical utilization for respiratory disease had the strongest association with lower lung function at 1 year in ELGANs.


The histiocytoses of infancy.
The histiocytoses comprise a rare and incompletely understood group of diseases that are characterized by an abnormal proliferation of histiocytes, mononuclear phagocytic, and antigen-presenting cells belonging to the reticuloendothelial system. The two major types of histiocytoses, which often present during infancy and require significant medical attention, are Langerhans cell histiocytosis and primary hemophagocytic lymphohistiocytosis. Their clinical manifestations frequently overlap, complicating their distinction, but a prompt and accurate diagnosis is essential to deliver the optimal treatment and maximize the chances for a favorable outcome. For Langerhans cell histiocytosis, careful risk stratification is critical for the appropriate administration of therapy. Patients with good prognostic factors may need only observation as their disease spontaneously regresses or minimal intervention. Poor prognostic factors mandate more intensive treatment. Patients with primary hemophagocytic lymphohistiocytosis require chemotherapeutic induction of their disease into a state of remission followed by stem cell transplantation, which currently offers the only known cure. Improvements on current therapies for the histiocytoses will depend on continued advances in the understanding of these enigmatic diseases.


A generalized discrete model linking rippling pattern formation and individual cell reversal statistics in colonies of myxobacteria.
Self-organization processes in multicellular aggregates of bacteria and amoebae offer fascinating insights into the evolution of cooperation and differentiation of cells. During myxobacterial development a variety of spatio-temporal patterns emerges such as counterpropagating waves of cell density that are known as rippling. Recently, several models have been introduced that qualitatively reproduce these patterns. All models include active motion and a collision-triggered reversal of individual bacteria. Here, we present a systematic study of a generalized discrete model that is based on similar assumptions as the continuous model by Igoshin et al (2001 Proc. Natl Acad. Sci. USA 98 14913). We find counterpropagating as well as unidirectional rippling waves in extended regions of the parameter space. If the interaction strength and the degree of cooperativity are large enough, rippling patterns appear even in the absence of a refractory period. We show for the first time that the experimentally observed double peak in the reversal statistics of bacteria in rippling colonies (Welch and Kaiser 2001 Proc. Natl Acad. Sci. USA 98 14907) can be reproduced in simulations of counterpropagating rippling waves which are dominant in experiments. In addition, the reversal statistics in the pre-rippling phase is correctly reproduced.


Transformation of subcutaneous nitric oxide into nitrate in the rat.
Following its addition to arterialized blood in vitro, nitric oxide (NO) is transformed into nitrate in the erythrocytes. Inhaled NO is similarly transformed into nitrate in the blood in vivo. These observations suggest that nitrate is a universal end-metabolite of NO, i.e. of endogenously formed NO as well. However, endogenous NO may also be inactivated in tissues, i.e. outside the vascular lumen. To study the fate of NO metabolized with delayed access to the blood, rats were given subcutaneous injections of 15NO or K15NO3, and the plasma concentrations of 15NO3(-) were followed for 450 min after injection. The values for the distribution volume and plasma decay (t12) of 15NO3(-) did not differ between rats given 15N-labelled NO and NO3(-). The area under the plasma decay curve for rats given 15NO amounted to 89% of the corresponding area for animals given K15NO3. This demonstrates that 15NO, when given extravascularly in millimolar concentrations, is mainly transformed into 15N-labelled nitrate. Other rats were kept in an atmosphere containing a mixture of 16O2 and 18O2. Nitrate residues containing either one or two 18O atoms were isolated from the blood, indicating that inhaled oxygen was incorporated during both the formation of NO and the subsequent transformation of NO into nitrate. The fraction of nitrate residues containing two 18O atoms was larger than that containing one 18O atom. We propose that nitrate is a major stable metabolite of endogenous NO that does not primarily diffuse into the vascular lumen following formation. Hence nitrate seems to be the quantitatively most important end-product of the metabolism of endogenous NO. The transformation of endogenous NO into nitrate involves the incorporation of inhaled oxygen.


Metabolic studies in a child with a pancreatic insulinoma.
An 8-year-old boy with a convulsive disorder for 3 1/2 years remined seizure free for 20 months while being treated with phenytoin (diphenylhydantoin) sodium, and then he had a relapse. He first demonstrated hypoglycemia when he fasted prior to being placed on a ketogenic diet. An oral glucose tolerance test indicated fasting and postglucose hypoglycemia and substantial hyperinsulinemia. Somatostatin infusion resulted in a modest increase in plasma glucose levels and a decrease in serum insulin concentrations. A discrete pancreatic mass was demonstrated preoperatively by celiac angiography that on surgical extirpation, proved to be a benign intrapancreatic insulinoma. Evaluation for islet cell tumors is of importance in children with seizure disorders unresponsive to anticonvulsant medication. Furthermore, somatostatin may be useful preoperatively in maintaining normal blood glucose concentrations in patients with islet cell adenomas.


Use of in vitro mutagenesis to analyze the molecular basis of the difference in Adh expression associated with the allozyme polymorphism in Drosophila melanogaster.
In natural populations of Drosophila melanogaster, the alcohol dehydrogenase (Adh) locus is polymorphic for two allozymes, designated Slow and Fast. Fast homozygotes generally have a two- to threefold higher ADH activity level than Slow homozygotes for two reasons: they have a higher concentration of ADH protein and the Fast protein has a higher catalytic efficiency. DNA sequencing studies have shown that the two allozymes generally differ by only a single amino acid at residue 192, which must therefore be the cause of the catalytic efficiency difference. A previous P element-transformation experiment mapped the difference in ADH protein level to a 2.3-kb HpaI/ClaI restriction fragment; which contains all of the Adh coding sequences but excludes all of the 5' flanking region of the distal transcriptional unit. Here we report the results of a site-directed in vitro mutagenesis experiment designed to investigate the effects of the amino acid replacement. This replacement has the expected effect on catalytic efficiency, but there is no detectable effect on the concentration of ADH protein estimated immunologically. This result shows that the average difference in ADH protein level between the allozymic classes is due to linkage disequilibrium between the amino acid replacement and one or more other polymorphisms within the HpaI/ClaI fragment. Sequence analysis of several Fast and Slow alleles suggested that the other polymorphism might be a silent substitution at nucleotide 1443, but another in vitro mutagenesis experiment reported here shows that this is not the case. Therefore, the molecular basis of the difference in ADH protein concentration between the allozymic classes remains an open question.


Patterns of Crystallin Gene Expression in Differentiation State Specific Regions of the Embryonic Chicken Lens.
Transition from lens epithelial cells to lens fiber cell is accompanied by numerous changes in gene expression critical for lens transparency. We identify expression patterns of highly prevalent genes including ubiquitous and enzyme crystallins in the embryonic day 13 chicken lens. Embryonic day 13 chicken lenses were dissected into central epithelial cell (EC), equatorial epithelial cell (EQ), cortical fiber cell (FP), and nuclear fiber cell (FC) compartments. Total RNA was prepared, subjected to high-throughput unidirectional mRNA sequencing, analyzed, mapped to the chicken genome, and functionally grouped. A total of 77,097 gene-specific transcripts covering 17,450 genes were expressed, of which 10,345 differed between two or more lens subregions. Ubiquitous crystallin gene expression increased from EC to EQ and was similar in FP and FC. Highly expressed crystallin genes fell into three coordinately expressed groups with R2 ≥ 0.93: CRYAA, CRYBB2, CRYAB, and CRYBA2; CRYBB1, CRYBA4, CRYGN, ASL1, and ASL; and CRYBB3 and CRYBA1. The highly expressed transcription factors YBX1, YBX3, PNRC1, and BASP1 were coordinately expressed with the second group of crystallins (r2 > 0.88). Although it is well known that lens crystallin gene expression changes during the epithelial to fiber cell transition, these data identify for the first time three distinct patterns of expression for specific subsets of crystallin genes, each highly correlated with expression of specific transcription factors. The results provide a quantitative basis for designing functional experiments pinpointing the mechanisms governing the landscape of crystallin expression during fiber cell differentiation to attain lens transparency.


Usefulness of field potential as a marker of embryonic stem cell-derived cardiomyocytes, and endpoint analysis of embryonic stem cell test.
The embryonic stem cell test (EST) is a validated method and a useful screening tool for drug discovery. EST requires microscopic observation of beating cells to be considered cardiomyocytes as an endpoint assay. However, this procedure is time-consuming and limits the throughput performance. Instead of microscopic observation, we previously established a novel assay method based on cardiac field potential as an endpoint. However, cardiac specificity of this field potential is not yet clarified, because beating cells have not been rigorously evaluated as skeletal or cardiomyocyte. Here, we investigated the relationships between field potential, beating, and cardiac troponin T (cTnT) expression, selected as a cardiomyocyte-specific marker, and evaluated suitability of the field potential as a marker for cardiomyocyte in vehicle or 5-fluorouracil treated embryo bodies. Embryoid bodies of mouse embryonic stem cells (D3) were differentiated in a chamber with multi-electrode array for 5 days, and field potential and beating were measured at the end of differentiation. In addition, these chambers were immunohistochemically stained with anti-cTnT antibody, and the correlation between field potential, beating, and cTnT expression was examined. These results indicated the area of field potential or beating mainly coincided with that of cTnT expression. 5-fluorouracil treatment decreased not only the number of field potential detecting electrodes and beating area, but also cTnT expression, and the area of these parameters was also nearly identical. These results indicate that field potential can be used as a suitable cardiac differentiation marker, and can be a promising parameter of EST.


Environmental drivers of forest structure and stem turnover across Venezuelan tropical forests.
Using data from 50 long-term permanent plots from across Venezuelan forests in northern South America, we explored large-scale patterns of stem turnover, aboveground biomass (AGB) and woody productivity (AGWP), and the relationships between them and with potential climatic drivers. We used principal component analysis coupled with generalized least squares models to analyze the relationship between climate, forest structure and stem dynamics. Two major axes associated with orthogonal temperature and moisture gradients effectively described more than 90% of the environmental variability in the dataset. Average turnover was 1.91 ± 0.10% year-1 with mortality and recruitment being almost identical, and close to average rates for other mature tropical forests. Turnover rates were significantly different among regions (p < 0.001), with the lowland forests in Western alluvial plains being the most dynamic, and Guiana Shield forests showing the lowest turnover rates. We found a weak positive relationship between AGB and AGWP, with Guiana Shield forests having the highest values for both variables (204.8 ± 14.3 Mg C ha-1 and 3.27 ± 0.27 Mg C ha-1 year-1 respectively), but AGB was much more strongly and negatively related to stem turnover. Our data suggest that moisture is a key driver of turnover, with longer dry seasons favoring greater rates of tree turnover and thus lower biomass, having important implications in the context of climate change, given the increases in drought frequency in many tropical forests. Regional variation in AGWP among Venezuelan forests strongly reflects the effects of climate, with greatest woody productivity where both precipitation and temperatures are high. Overall, forests in wet, low elevation sites and with slow turnover stored the greatest amounts of biomass. Although faster stand dynamics are closely associated with lower carbon storage, stem-level turnover rates and woody productivity did not show any correlation, indicating that stem dynamics and carbon dynamics are largely decoupled from one another.


Dicarboxylic amino acid influx across brush border of rabbit ileum. Effects of amino acid charge on the sodium-amino acid interaction.
Glutamate and aspartate influxes across the brush border of rabbit intestine are saturable processes that are subject to competitive inhibition and are markedly influenced by the Na concentration in the mucosal solution. Lowering the Na concentration increases the amino acid concentration needed to elicit a half-maximal influx but does not significantly affect the maximal influx. The interaction between Na and anionic amino acid influx can be described by the same kinetic model that has been applied to the influxes of neutral amino acids and lysine. Comparison of the kinetic parameters for anionic, neutral, and cationic amino acids suggests that amino acid charge influences (a) the stability of the binary (amino acid-site) complex and (b) the affinity of this binary complex for the subsequent binding of Na. A mechanistic interpretation of these interactions is proposed.


Successful Suicide Screening in the Pediatric Emergency Department: Youth, Parent, Researcher, and Clinician Perspectives.
The purpose of this study was to include youth, parents, researchers, and clinicians in the identification of feasible and acceptable strategies for teen suicide screening in the pediatric emergency department (ED). Concept mapping methodology was used to elicit stakeholder responses. Regarding the most important result of suicide screening for teens in the pediatric ED, suicide prevention and education for parents, friends, and community members was rated easiest to implement, while short- and long-term follow-up and treatment was rated most important. In terms of successful suicide screening for teens in the pediatric ED, provision of resources and information was rated most feasible, and a safe, friendly, private screening environment was rated most important. The concept maps can be used to align suicide risk screening with the priorities and recommendations of pediatric ED stakeholders.


Spurious observation of splenic cyp2b1 expression.
Phenobarbital (PB) induction of the CYP2B subfamily was studied in the livers and spleens of male and female rats. Animals were treated with either PB (10 mg/kg) or vehicle for 4 consecutive days. A reverse transcriptase-polymerase chain reaction (RT-PCR), quantitative Northern blotting, Western blotting, and a radioenzymatic assay were used to observe differential levels of CYP2B1 and CYP2B2 mRNAs, proteins, and catalytic activities. CYP2B2 expression was limited to the livers of PB-treated male and female rats and was not detected in spleen. Low constitutive levels of CYP2B1 mRNA were markedly induced approximately 7- to 17-fold in the livers of PB-treated male and female rats, respectively. However, using the same standard oligonucleotide probe for CYP2B1 mRNA, we observed considerably greater constitutive concentrations of the transcript in spleen than in liver. Putative splenic CYP2B1 mRNA was significantly elevated by the PB treatment, although not as profoundly as the hepatic response. In contrast, only the livers of the barbiturate-treated rats expressed CYP2B1 proteins or specific catalytic activity (androstenedione 16beta-hydroxylase). Protein and catalytic activities of the isoforms were undetectable in spleen of either male or female vehicle- and PB-treated rats. In agreement, RT-PCR was unable to demonstrate the expression of splenic CYP2B1 mRNAs. Investigating the possibility that the Northern probe for CYP2B1 was identifying a similar sequence isoform, we performed RT-PCR using primers for CYP2B12 and CYP2B15. Since neither of these isoforms was expressed in spleen, we conclude that the spurious results using the Northern probe for CYP2B1 mRNA were due to the presence of a cross-reacting, PB-responsive transcript not currently identifiable in existing databases.


Peripherally inserted central catheter-related bloodstream infection due to Tsukamurella pulmonis: a case report and literature review.
Tsukamurella pulmonis is an aerobic gram-positive and rod-shaped organism that causes central catheter-related bloodstream infections in immunocompromised hosts. However, peripherally inserted central catheter (PICC)-related bloodstream infections due to this organism have not been reported. We describe a case of a 48-year-old man with acquired immunodeficiency syndrome and diffuse large B cell lymphoma who received five courses of chemotherapy including rituximab , cyclophosphamide , doxorubicin hydrochloride , vincristine , and prednisone via a PICC. Five days after the last chemotherapy course, he presented with a high fever and shaking chills. His absolute neutrophil count was 4200/μL. Cultures obtained from blood and PICC culture revealed T. pulmonis. The colony count of T. pulmonis grown from PICC culture was 103 colony-forming units. Therefore, he was diagnosed with T. pulmonis bacteremia resulting from PICC-related bloodstream infection. The patient's condition improved and he became afebrile within 48 h after intravenous administration of cefozopran hydrochloride, which is a fourth generation cephalosporin. PICCs can be associated with T. pulmonis bacteremia, and fourth generation cephalosporins may be effective treatment.


New perspectives in pheochromocytoma.
Pheochromocytomas are tumors that arise from chromaffin cells. Although the majority occur in the adrenal gland, these tumors can be found anywhere from the neck to the base of the pelvis. This condition, which is the cause of hypertension in only a small percentage of patients, can be cured in approximately 90 per cent of cases but can be lethal if left untreated. Pheochromocytomas may be familial in 10 per cent of patients, and in such cases, they may be associated with a variety of other conditions. The authors review the biochemical and radiologic diagnosis of adrenal and extra-adrenal tumors and discuss the treatment, including the special problems of pheochromocytoma during pregnancy.


Georeferenced data in epidemiologic research.
This paper reviews some conceptual and practical issues regarding the application of georeferenced data in epidemiologic research. Starting with the disease mapping tradition of geographical medicine, topics such as types of georeferenced data, implications for data analysis, spatial autocorrelation and main analytical approaches are heuristically discussed, relying on examples from the epidemiologic literature, most of them concerning mapping disease distribution, detection of disease spatial clustering, evaluation of exposure in environmental health investigation and ecological correlation studies. As for concluding remarks, special topics that deserve further development, including the misuses of the concept of space in epidemiologic research, issues related to data quality and confidentiality, the role of epidemiologic designs for spatial research, sensitivity analysis and spatiotemporal modeling, are presented.


Chinese haze versus Western smog: lessons learned.
Air pollution in many Chinese cities has been so severe in recent years that a special terminology, the "Chinese haze", was created to describe China's air quality problem. Historically, the problem of Chinese haze has developed several decades after Western high-income countries have significantly improved their air quality from the smog-laden days in the early- and mid-20(th) century. Hence it is important to provide a global and historical perspective to help China combat the current air pollution problems. In this regard, this article addresses the followings specific questions: (I) What is the Chinese haze in comparison with the sulfurous (London-type) smog and the photochemical (Los Angeles-type) smog? (II) How does Chinese haze fit into the current trend of global air pollution transition? (III) What are the major mitigation measures that have improved air quality in Western countries? and (IV) What specific recommendations for China can be derived from lessons and experiences from Western countries?


[Psoriasis and leprosy in the light of common history. A contribution on epidemiology and differential diagnosis].
Over the centuries psoriasis was described as a variety of leprosy. It is only in the last century, however, that it was described as a separate entity. Many early medical descriptions of this woridwide affliction can be found, which attempt to distinguish between the various kinds of leprosy. These attempts led to cultural-religious consequences on one hand and to different theories about their contagiousness and curability on the other. Today the current conviction, that with the proof for the existence of a lepra bacillus the question of heritability of the disease can finally be discarded, is being reconsidered. For both psoriasis and leprosy the search for relationships between these diseases and genetically determined markers in the blood-, serum protein- and enzyme-group systems has led to applicable results which, insofar as they can be interpreted as indications of selective factors, will enable us to understand the geographical distribution of both diseases.


Construct validity of the Iowa Gambling Task.
The Iowa Gambling Task (IGT) was created to assess real-world decision making in a laboratory setting and has been applied to various clinical populations (i.e., substance abuse, schizophrenia, pathological gamblers) outside those with orbitofrontal cortex damage, for whom it was originally developed. The current review provides a critical examination of lesion, functional neuroimaging, developmental, and clinical studies in order to examine the construct validity of the IGT. The preponderance of evidence provides support for the use of the IGT to detect decision making deficits in clinical populations, in the context of a more comprehensive evaluation. The review includes a discussion of three critical issues affecting the validity of the IGT, as it has recently become available as a clinical instrument: the lack of a concise definition as to what aspect of decision making the IGT measures, the lack of data regarding reliability of the IGT, and the influence of personality and state mood on IGT performance.


A Comparison of Tissue Dissection Techniques for Diagnostic, Prognostic, and Theragnostic Analysis of Human Disease.
Histopathology has historically been the critical technique for the diagnosis and treatment of human disease. Today, genomics, transcriptomics, and proteomics from specific cells, rather than bulk tissue, have become key to understanding underlying disease mechanisms and rendering useful diagnostic information. Extraction of desired analytes, i.e., nucleic acids or proteins, from easily accessible formalin-fixed paraffin-embedded tissues allows for clinically relevant activities, such as sequencing biomarker mutations or typing amyloidogenic proteins. Genetic profiling has become routine for cancers as varied as non-small cell lung cancer and prostatic carcinoma. The five main tissue dissection techniques that have been developed thus far include: bulk scraping, manual macrodissection, manual microdissection, laser-capture microdissection, and expression microdissection. In this review, we discuss the importance of tissue dissection in clinical practice and research, the basic methods, applications, as well as some advantages and disadvantages for each modality.


Selectivity of dobutamine for adrenergic receptor subtypes: in vitro analysis by radioligand binding.
The cardiovascular responses elicited by dobutamine are distinctly different from those produced by other adrenergic or dopaminergic agonists. To test the hypothesis that dobutamine could have differential affinities for adrenergic receptor subtypes, and that such subtype selectivity could be related to its relatively unique pharmacologic properties, we assessed the ability of dobutamine to displace adrenergic radioligands from membrane receptors in a number of tissues of previously characterized adrenergic receptor subtype. For beta adrenergic receptors identified by (-) [(3)H]dihydroalprenolol (DHA), dobutamine had significantly greater affinity for the beta(1) subtype (K(D) = 2.5 muM in rat heart and 2.6 muM in turkey erythrocyte) than for the beta(2) subtype (K(D) = 14.8 muM in frog heart and 25.4 muM in rat lung) (P < 0.001). For alpha adrenergic receptors, dobutamine had markedly greater affinity for the alpha(1)-subtype identified by [(3)H]prazosin (K(D) = 0.09 muM in rat heart and 0.14 muM in rabbit uterus) than for the alpha(2)-subtype identified by [(3)H]dihydroergocryptine (DHE) (K(D) = 9.3 muM in human platelet) or by [(3)H]yohimbine (K(D) = 5.7 muM in rabbit uterus) (P < 0.001). Like other beta(1)-agonists, in the absence of guanine nucleotide, dobutamine competition curves for DHA binding in rat heart demonstrated two classes of binding sites, with one site of significantly higher affinity (K(D) = 0.5 muM, P = 0.008) than the single class of binding sites (K(D) = 5.2 muM) identified in the presence of guanine nucleotide. However, unlike beta(2)- or alpha(2)-agonists, dobutamine displacement of DHA binding in rat lung or of DHE binding in human platelets demonstrated only a single class of binding sites, and guanine nucleotide had only minimal effects. We conclude that dobutamine is selective for beta(1) as opposed to beta(2), and for alpha(1) as opposed to alpha(2) adrenergic receptors. Furthermore, guanine nucleotide effects on dobutamine binding, and biochemical response data in vitro suggest that dobutamine is a beta(1)-agonist, but has little intrinsic activity at beta(2) and alpha(2)-receptors. This selectivity for adrenergic receptor subtypes may be part of the basis for dobutamine's distinctive pharmacologic properties in vivo.


The management of varicella-zoster virus exposure and infection in pregnancy and the newborn period. Australasian Subgroup in Paediatric Infectious Diseases of the Australasian Society for Infectious Diseases.
Zoster immunoglobulin (ZIG) should be offered to pregnant, varicella-seronegative women with significant exposure to varicella-zoster virus (VZV) (chickenpox) infection. Oral aciclovir prophylaxis should be considered for susceptible pregnant women exposed to VZV who did not receive ZIG or have risk factors for severe disease. Intravenous aciclovir should be given to pregnant women who develop complicated varicella at any stage of pregnancy. Counselling on the risk of congenital varicella syndrome is recommended for pregnant women who develop chickenpox. ZIG should be given to a baby whose mother develops chickenpox up to 7 days before delivery or up to 28 days after delivery. Intravenous aciclovir should be given to babies presenting unwell with chickenpox, whether or not they received ZIG. Breastfeeding of babies infected with or exposed to VZV is encouraged. A mother with chickenpox or zoster does not need to be isolated from her own baby. If siblings at home have chickenpox, a newborn baby should be given ZIG if its mother is seronegative. The newborn baby does not need to be isolated from its siblings with chickenpox, whether or not the baby was given ZIG. After significant nursery exposure to VZV, ZIG should be given to seronegative babies and to all babies born before 28 weeks' gestation.


An electrophysiological investigation of the macular photostress test.
The speed of macular recovery from photostress was measured as a function of age by subjective and objective techniques. The subjective procedures involved measuring the speed of recovery of habitual visual acuities after dazzling of the macula with a bright light. Macular function was assessed objectively by monitoring the development of the transient pattern VER after photostress. Both techniques indicated that the ability to recover from photostress decreased with age. However, the recovery of visual acuity and the restoration of baseline VER parameters had very different temporal characteristics. The time for restoration of baseline VER amplitude was usually several times longer than the time for subjective recovery of prestress visual acuity levels. Furthermore, subjects less than 30 years of age differed from the older group in that their recovery VER amplitudes tended to quickly surpass the baseline VER values, more so in the youngest age group. Most subjects over 30 did not show such super normal VERs and failed to recover baseline VEP amplitudes within 60 sec after photostress. These observations suggest age-related functional differences in retrobulbar components of the visual system.


Time course and topographic distribution of ocular fundus pulsation measured by low-coherence tissue interferometry.
Low-coherence tissue interferometry is a technique for the depth-resolved measurement of ocular fundus pulsations. Whereas fundus pulsation amplitudes at preselected axial positions can readily be assessed by this method, coupling of the interferometer with a pulse oximeter additionally allows for the reconstruction of the time course of ocular fundus pulsation with respect to the cardiac cycle of the subject. For this purpose, the interferogram resulting from the superposition of waves reflected at the cornea and the ocular fundus is recorded synchronously with the plethysmogram. A new method for evaluating the time course of synthetic interferograms in combination with plethysmograms based on averaging several pulse periods has been developed. This technique allows for the analysis of amplitudes, time courses, and phase differences of fundus pulsations at preselected axial and transversal positions and for creating fundus pulsation movies. Measurements are performed in three healthy emmetropic subjects at angles from 0 deg to 18 deg to the axis of vision. Considerably different time courses, amplitudes, and phases with respect to the cardiac cycle are found at different angles. Data on ocular fundus pulsation obtained with this technique can--among other applications--be used to verify and to improve biomechanical models of the eye.


Interaction of JMJD6 with single-stranded RNA.
JMJD6 is a Jumonji C domain-containing hydroxylase. JMJD6 binds alpha-ketoglutarate and iron and has been characterized as either a histone arginine demethylase or U2AF65 lysyl hydroxylase. Here, we describe the structures of JMJD6 with and without alpha-ketoglutarate, which revealed a novel substrate binding groove and two positively charged surfaces. The structures also contain a stack of aromatic residues located near the active center. The side chain of one residue within this stack assumed different conformations in the two structures. Interestingly, JMJD6 bound efficiently to single-stranded RNA, but not to single-stranded DNA, double-stranded RNA, or double-stranded DNA. These structural features and truncation analysis of JMJD6 suggest that JMJD6 may bind and modify single-stand RNA rather than the previously reported peptide substrates.


Relative contribution of arms and legs in humans to propulsion in 25-m sprint front-crawl swimming.
Eight male subjects were asked to swim 25 m at maximal velocity while the use of the arm(s) and legs was alternately restricted. Four situations were examined using one arm (1A), two arms (2A), one arm and two legs (1A2L) and both arms and legs (2A2L, normal swim) for propulsion. A significant mean increase of 10% on maximal velocity was obtained in 1A2L and 2A2L compared to 1A and 2A. A non-significant 4% effect was obtained in 1A. This study focused on the actual contribution of leg kick in the 10% gain in maximal velocity. It was clear that the underwater trajectory of the wrist was modified by the action of the legs (most comparisons P < 0.001). Therefore it was thought that the legs enhanced the generated propulsive force by improving the propulsive action of the arm. The arm action was quantified by selecting typical phases from the filmed trajectory of the wrist, namely forward (F), downwards (D) and backwards (B). Although there was a tendency for individual changes in kinematic parameters (F, D and B) to occur with individual changes in velocity when 2A was compared to 2A2L, no relationship was found between the relative changes in F, D and B and relative changes in velocity. This was illustrated by describing the responses of three individuals who could represent three patterns of contribution by legs and arms to propulsion in high speed swimming.


Association of IFN-gamma and IFN regulatory factor 1 polymorphisms with childhood atopic asthma.
IFN-gamma and related molecules play important roles in the differentiation and function of TH2 cells. We sought to determine whether IFNG and related genes contribute to any susceptibility to atopic asthma, a representative TH2-dominant disorder. We investigated the association of IFNG (CA repeat polymorphism within the first intron), IRF1 (GT repeat polymorphism within the intron 7), IFNGR1 (Val 14 Met), and IFNGR2 (Gln 64 Arg) gene polymorphisms with atopic asthma in the Japanese child population. A significant association (P =.0018) was observed between IFNG gene polymorphism and atopic asthma. The tendency was more prominent in patients with age of onset of 3 years or younger (P =.0004) or patients with a family history of allergic diseases (P =.0038). Furthermore, there was a significant association between IRF1 gene whole-allele distribution and atopic asthma (P =.044). The tendency was more prominent in patients with onset at 3 years of age or less (P =.0058). On the other hand, IFNGR1 and IFNGR2 gene polymorphisms showed no association with atopic asthma. These results suggested that among IFNG and related genes, IFNG and IRF1 genes confer genetic susceptibility to atopic asthma in Japanese children.


Acute exogenous intoxications in childhood: factors related to hospitalization.
To describe the profile of children assisted in the Emergency Room of a Children reference Hospital in the South of Brazil, victims of acute poisoning between 2016 and 2021, to characterize the toxic agents and to present the factors related to hospitalization. Retrospective, descriptive and observational study with data collection from medical records at a children's hospital from July 2016 to June 2021 based on the compulsory notification forms. The characteristics of victims, of the incident, the type and class of the substance involved, the procedures demanded and the need for the Intensive Care Unit were evaluated. The outcome considered was hospitalization. Absolute and relative frequencies were calculated for the categorical variables and measures of central tendency and dispersion for the numerical ones. Binary logistic regression was performed to identify variables related to hospitalization. There were 411 consultations, with the mean age of 7.2±5.5 years, and predominance of females (59.9%). Most of the poisoning occurred at home (82.1%) and orally (93.7%). Chemicals or cleaning products were the main agents in children up to 1 year of age, whereas in the other age groups accidents occurred most frequently with medicines. Hospital admission occurred in 38.7% of the cases, with related variables being: number of agents, type of substance involved, medication that acts on the Central Nervous System, recurrence, motivation (accidental/intentional), and performance of simple exams. More preventive actions are needed, such as legislation, as well as greater guidance to parents on how to store products in the domestic environment, in order to reduce the number of exogenous intoxications in the pediatric population.


Therapeutic effect of human ghrelin and growth hormone: Attenuation of immunosuppression in septic aged rats.
Sepsis is a leading cause of mortality in intensive care units, and is more common in the geriatric population. The control of hyperinflammation has been suggested as a therapeutic approach in sepsis, but to date clinical trials utilizing this strategy have not lead to an effective treatment. In addition to hyperinflammation, patients with sepsis often experience a state of immunosuppression, which serves as an important determinant for increased morbidity and mortality. We previously used aged animals to demonstrate the effectiveness of combined treatment with human ghrelin (Ghr) and human growth hormone (GH) in improving organ injury and survival in septic animals. Here, we hypothesized that combined treatment with Ghr and GH could improve immune function in septic aged animals. Male 24-month-old rats were subjected to cecal ligation and puncture (CLP) for sepsis induction. Human Ghr (80nmol/kg BW) plus GH (50μg/kg BW) or vehicle (normal saline) was administrated subcutaneously at 5h after CLP. The ex vivo production of TNF-α, IL-6 and IL-10 to LPS-stimulation, as well as TNF-α, IL-6, IL-10 and IFN-γ production to anti-CD3/anti-CD28 antibody-stimulation, in splenocytes isolated 20h after CLP, was significantly decreased compared to production of these cytokines in splenocytes from sham animals. The production of cytokines from splenocytes isolated from septic animals that received the combined treatment, however, was significantly higher than from those isolated from vehicle-treated septic animals. Combined treatment prevented the loss of splenic CD4+ and CD8+ T cells in septic aged rats, and reduced lymphocyte apoptosis. Combined treatment also inhibited an increase in the regulatory T cell (Treg) population and expression of the immune co-inhibitory molecule PD-1 in the spleens of septic aged rats. In contrast, expression of HLA-DR was increased after combined treatment with Ghr and GH. Based on these findings, we conclude that co-administration of Ghr and GH is a promising therapeutic tool for reversing immunosuppression caused by sepsis in the geriatric population. This article is part of a Special Issue entitled: Immune and Metabolic Alterations in Trauma and Sepsis edited by Dr. Raghavan Raju.


Lichen planus and cicatricial conjunctivitis: disease course and response to therapy of 11 patients.
Lichen planus is an autoimmune, inflammatory dermatosis of unknown cause that affects the skin and mucous membranes. The aim of this study was to report the clinical features and response to therapy in a series of patients with ocular lichen planus. A retrospective chart review was performed to identify patients with ocular lichen planus. Information about clinical presentation, treatment, and therapeutic response was extracted from the medical records. Eleven patients with ocular lichen planus were identified. The diagnosis was confirmed histologically for 10 patients. Nine patients were women. The average time from onset of ocular symptoms to diagnosis was 4.1 years. Eight patients had mucous membrane involvement at other sites. Disease was well controlled in eight patients. Lichen planus should be considered in the differential diagnosis of cicatricial conjunctivitis, especially when severe lichen planus is noted at other sites.


Cost-Effective Live Cell Density Determination of Liquid Cultured Microorganisms.
Live monitoring of microorganisms growth in liquid medium is a desired parameter for many research fields. A wildly used approach for determining microbial liquid growth quantification is based on light scattering as the result of the physical interaction of light with microbial cells. These measurements are generally achieved using costly table-top instruments; however, a live, reliable, and straight forward instrument constructed using parts that are inexpensive may provide opportunities for many researchers. Here, such an instrument has been constructed and tested. It consists of modular test tube holding chambers, each with a low power monochromatic light-emitting diode, and a monolithic photodiode. A microcontroller connects to all modular chambers to control the diodes, and send the live data to either an LCD screen, or a computer. This work demonstrate that this modular instrument can determine precise cell concentrations for the bacteria Escherichia coli and Pseudomonas syringae pv. tomato DC3000, as well as Saccharomyces cerevisiae yeast.


[Postsynaptic reactions of neurons of the sensomotor cortex of the cat during evoked and self-sustained rhythmic activity of the "peak-wave" type].
Intracellular correlates of evoked rhythmic cortical "spike-and-wave" potentials produced in sensorimotor cortex during 3/s stimulation of the thalamic relay nucleus (VPL) and of self-sustained "spike-and-wave" afterdischarges following 8-14/s stimulation of the same nucleus were studied in acute experiments on cats immobilized by myorelaxants. Intracellular recordings of pyramidal tract neurons revealed that different components of evoked "spike-and-wave" potentials, i. e. the spike-like negative wave and the long lasting negative wave, are postsynaptic in origin: the first is due to EPSPs with spike discharges, and the latter--to IPSPs of cortical neurons. Components of "spike-and-wave" afterdischarge mostly reflect the paroxysmal depolarizing shifts of the membrane potential of cortical neurons. After cessation of sustained "spike-and-wave" activity the long-lasting hyperpolarization accompanied by inhibition of spike discharges and subsequent recovery was observed in cortical neurons. It is presumed that the negative wave of the evoked "spike-and-wave" potential as well as slow negative potentials of direct cortical and primary responses reflect IPSPs of deeper parts of pyramidal tract neurons, while the waves of the sustained "spike-and-wave" afterdischarges are due to paroxysmal depolarizing shifts in cortical neurons.


Empirical measurement of pressure in negative pressure wound therapy for infected wounds: How long can it really stay under pressure?
Surgical site infections represent one of the most common surgical complications. Negative pressure wound therapy is considered an effective wound management system, based on the principle that a negative pressure inside the wound can suction fluids and approximate wound edges. With the negative pressure wound therapy systems commercially available it is assumed that the pressure inside the wound is stable at the set values. We conducted a prospective experimental study to investigate this. The negative pressure level achieved inside the dressing was investigated at a standard distance from the pad of suction and at specific times in patients with surgical site infections. Pressure measurements were performed in 28 dressings in 14 patients admitted to the Emergency Surgery Department between April 2016 and June 2017. In general, the machine was set at a pressure of -100 mmHg. Negative pressure was measured with a portable pressure reader in the dressing sponge at a distance of 0, 1.5, and 3 cm from the suction pad at the time of dressing change at 24 and 48 hours. The data suggest that there is a significant decrease in negative pressure at both 1.5 and 3 cm from the suction pad at 48 hours from the dressing change (p = 0.0001 and p < 0.0001, respectively). This preliminary study shows with statistical significance that the pressure inside the sponge of a negative pressure wound therapy system decreases after 48 hours, confirming that the pressure inside the system is not the same as the pressure reported by the machine and, instead of being stable, changes overtime.


Immature neutrophils in the blood smears of young febrile children.
To determine whether the immature neutrophil (band) count in the peripheral blood smear helps to distinguish young febrile children with bacterial or respiratory viral infections. A prospective cohort study in 3 pediatric emergency departments. A convenience sample of 100 febrile children aged 2 years or younger with either laboratory-documented bacterial infections (n = 31; 24 with urinary tract infections, 7 with bacteremia) or laboratory-documented respiratory viral infections (n = 69). Each patient received a clinical appearance score using the Yale Observation Scale prior to laboratory evaluation. A complete blood cell count was obtained from all patients and manual differential count of the peripheral blood smear was performed by 1 senior technician masked to clinical information. Band counts, represented as a percentage of white blood cells in the peripheral blood smear, the absolute band count, and band-neutrophil ratio. Logistic regression analysis was performed to determine whether the band count helps to distinguish bacterial infections from viral infections after adjusting for age, temperature, Yale Observation Scale score, and absolute neutrophil count. Patients with bacterial infections had a higher mean absolute neutrophil count (11.3 vs 5.9 x 10(9)/L; P<.01) than patients with respiratory viral infections. There was no difference, however, in percentage band count (13.5% vs 13.3%; P = .90), absolute band count (2.2 vs 1.9 X 10(9)/L; P= .31), or band-neutrophil ratio (0.24 vs 0.33; P = .08, bacterial vs viral, respectively); the band count did not help to distinguish bacterial and viral infections after adjusting for age, temperature, Yale Observation Scale score, and absolute neutrophil count in the regression analysis. The band count in the peripheral blood smear does not routinely help to distinguish bacterial infections from respiratory viral infections in young febrile children.


Helcococcus ovis, an emerging pathogen in bovine valvular endocarditis.
The initial isolation of Helcococcus ovis from a valvular thrombus prompted us to investigate the prevalence of this bacterium in bovine valvular endocarditis. Specimens from 55 affected hearts were examined by culture using Columbia blood agar and cross streaking the inoculated plate with a Staphylococcus aureus strain. As confirmed by 16S rRNA gene sequencing, H. ovis was isolated with an unexpectedly high frequency of 33%, predominantly as heavy growth and pure culture. The majority of H. ovis isolates showed distinct satellitism around S. aureus and pyridoxal dependency, resembling "nutritionally variant streptococci" (now assigned to the genera Abiotrophia and Granulicatella). Using the API rapid ID 32 Strep, API ZYM, and Rosco Diatabs systems, incongruent results were obtained for alkaline phosphatase, beta-galactosidase, beta-glucuronidase, and leucine aminopeptidase activities. Based on the satellitism/pyridoxal dependency; hemolysis on blood agar; the API rapid ID 32 Strep results for arginine dihydrolase, alpha-galactosidase, beta-galactosidase, beta-glucuronidase, and pyroglutamic acid arylamidase activities; hippurate hydrolysis; and acidification of sucrose, a scheme for the identification of H. ovis and its differentiation from other members of the Helcococcus genus and the pyridoxal-dependent species Abiotrophia defectiva, Granulicatella adiacens, and Granulicatella elegans is proposed. By establishing specific fluorescence in situ hybridization, large H. ovis aggregates were specifically detected within the fibrinous exudate of the valvular thrombi. Our results demonstrate for the first time that H. ovis represents an emerging pathogen in bovine valvular endocarditis that is frequently isolated if appropriate culture conditions are used.


Factors associated with hospitalisations of patients with chronic heart failure approaching the end of life: A systematic review.
Heart failure has high mortality and is linked to substantial burden for patients, carers and health care systems. Patients with chronic heart failure frequently experience recurrent hospitalisations peaking at the end of life, but most prefer to avoid hospital. The drivers of hospitalisations are not well understood. We aimed to synthesise the evidence on factors associated with all-cause and heart failure hospitalisations of patients with advanced chronic heart failure. Systematic review of studies quantitatively evaluating factors associated with all-cause or heart failure hospitalisations in adult patients with advanced chronic heart failure. Five electronic databases were searched from inception to September 2020. Additionally, searches for grey literature, citation searching and hand-searching were performed. We assessed the quality of individual studies using the QualSyst tool. Strength of evidence was determined weighing number, quality and consistency of studies. Findings are reported narratively as pooling was not deemed feasible. In 54 articles, 68 individual, illness-level, service-level and environmental factors were identified. We found high/moderate strength evidence for specialist palliative or hospice care being associated with reduced risk of all-cause and heart failure hospitalisations, respectively. Based on high strength evidence, we further identified black/non-white ethnicity as a risk factor for all-cause hospitalisations. Efforts to integrate hospice and specialist palliative services into care may reduce avoidable hospitalisations in advanced heart failure. Inequalities in end-of-life care in terms of race/ethnicity should be addressed. Further research should investigate the causality of the relationships identified here.


Molecular imaging of activated platelets via antibody-targeted ultra-small iron oxide nanoparticles displaying unique dual MRI contrast.
Magnetic resonance imaging (MRI) is a powerful and indispensable tool in medical research, clinical diagnosis, and patient care due to its high spatial resolution and non-limited penetration depth. The simultaneous use of positive and negative MRI imaging that employs the same contrast agents will significantly improve detection accuracy. Here we report the development of functional multimodal iron oxide nanoparticles for targeted MRI of atherothrombosis using a combination of chemical and biological conjugation techniques. Monodisperse, water-soluble and biocompatible ultra-small magnetic dual contrast iron oxide nanoparticles (DCIONs) were generated using a high-temperature co-precipitation route and appeared to be efficient positive and negative dual contrast agents for magnetic resonance imaging. Using a unique chemo-enzymatic approach involving copper-free click chemistry and Staphylococcus aureus sortase A enzyme conjugation, DCIONs were functionalized with single-chain antibodies (scFv) directed against activated platelets for targeting purposes. The DCIONs were also labelled with fluorescent molecules to allow for optical imaging. The antigen binding activity of the scFv was retained and resulted in the successful targeting of contrast agents to thrombosis as demonstrated in a range of in vitro and in vivo experiments. T1- and T2-weighted MRI of thrombi was recorded and demonstrated the great potential of dual T1/T2 contrast iron oxide particles in imaging of cardiovascular disease.


Accelerated lung function decline in swine confinement workers.
We conducted a longitudinal study to determine the annual rate decline in pulmonary function measurements in male swine confinement workers. For comparison, a grain farming group and a nonfarming rural-dwelling control group were also chosen for the longitudinal study. Two hundred seventeen swine confinement workers, 218 grain farmers, and 179 nonfarming control subjects had valid pulmonary function measurements at the baseline observation conducted in 1990 to 1991 and at the second observation conducted in 1994 to 1995. The swine confinement workers were younger (mean age=38.3+/-11.7 [SD] years) than the nonfarming control subjects (42.6+/-10.4 years) and the grain farmers (44.5+/-11.9 years). When stratified by age, nonfarming control subjects had the lowest mean annual rate decline in FEV1 and FVC in all age categories. The swine confinement workers had the largest annual rate decline in FEV1 and FVC, and this was most obvious in the middle age categories. After controlling for age, height, smoking, and baseline pulmonary function, swine confinement workers had excess annual decline of 26.1 mL in FEV1 (p=0.0005), 33.5 mL in FVC (p=0.0002), and 42.0 mL/s in forced expiratory flow between 25% and 75% of FVC (FEF[25-75%]) (p=0.02) over nonfarming control subjects. Grain farmers had excess annual decline of 16.4 mL in FEV1 (p=0.03), 26.7 mL in FVC (p=0.002), and 11.2 mL/s in FEF(25-75%) (p=0.38) over control subjects. These findings suggest that workers engaged in the swine industry and grain farmers appear prone to accelerated yearly losses in lung function and may therefore be at risk for the future development of chronic airflow limitation.


A novel mechanism for vasoconstrictor action of 8-isoprostaglandin F2 alpha on retinal vessels.
Using a video-imaging technique, we characterized the effects of 8-isoprostaglandin F2 alpha (8-iso-PGF2 alpha) on retinal vasculature from piglets. 8-Iso-PGF2 alpha potently contracted (EC50 = 5.9 +/- 0.5 nM) retinal vessels. These effects were completely antagonized by the cyclooxygenase inhibitor indomethacin, the thromboxane synthase blocker CGS-12970, the thromboxane receptor antagonist L-670596, and the putative inhibitor of the non-voltage-dependent receptor-operated Ca2+ pathway SKF-96365; constrictor effects of 8-iso-PGF2 alpha were also partly attenuated by the ETA-receptor blocker BQ-123 and an inhibitor of endothelin-converting enzyme, phosphoramidon, but was negligibly affected by the L-type voltage-gated Ca2+ channel blocker nifedipine. Correspondingly, 8-iso-PGF2 alpha elicited endothelin release from retinal preparations, which was markedly reduced by SKF-96365. 8-Iso-PGF2 alpha also increased thromboxane production in the retina and cultured endothelial cells, but not on retinovascular smooth muscle cells; these effects of 8-iso-PGF2 alpha were blocked by indomethacin, CGS-12970, SKF-96365, and EGTA, but not by nifedipine. 8-Iso-PGF2 alpha also increased Ca2+ transients in retinal endothelial cells, which were inhibited by SKF-96365 and EGTA, but not by nifedipine, whereas in smooth muscle cells U-46619, but not 8-iso-PGF2 alpha, stimulated a rise in Ca2+ transients. Finally, H2O2 + FeCl2 (in vitro) and anoxia followed by reoxygenation (in vivo) stimulated formation of 8-iso-PGF2 alpha in the retina. In conclusion, 8-iso-PGF2 alpha-induced retinal vasoconstriction is mediated by cyclooxygenase-generated formation of thromboxane and, to a lesser extent, by endothelin after Ca2+ entry into cells, possibly through receptor-operated channels. Retinal vasoconstriction to 8-isoprostanes might play a role in the genesis of ischemic retinopathies.


Site-specific integration of H-ras in transformed rat embryo cells.
A karyotypic analysis was performed on seven independently derived clones of primary rat embryo cells transformed by the ras oncogene plus the cooperating oncogene myc. The transfected oncogenes were sometimes present in amplified copy number, with heterogeneity in the levels of amplification. Some chromosomal features, such as aberrantly banding regions and double-minute chromosomes, typical of cells carrying amplified genes, were also seen in three of the seven cell lines. Underlying this heterogeneity there was an unexpected finding. All seven lines showed a common integration site for ras on the q arm of rat chromosome 3 (3q12), though some lines also had other sites of integration. In four of the lines integration of ras was accompanied by deletion of the p arm of chromosome 3 or its possible translocation to chromosome 12.


Cellular mechanisms in the response of the arterial wall to injury and repair.
This synopsis emphasizes the inappropriateness of a "single stimulus-single response" approach in understanding the response of the arterial wall to injury and repair. The outcome of any injurious stimulus is a series of interactive cascades among the endogenous and exogenous cellular and non-cellular components of the arterial wall, and the cellular and non-cellular elements of the blood. Both genetic and hemodynamic factors can further influence this response. The more prominent of the cellular and non-cellular components have been discussed. These include: 1) the vascular endothelium, its dynamic interaction with macromolecules and formed elements of the blood, its role in the transport of plasma proteins, its influence on the function of arterial smooth muscle cells and the recruitment of blood-born monocytes; 2) the arterial smooth muscle cell, its role in the vasomotor function of the artery wall, arterial repair and reconstruction, metabolism of lipids and the secretion of cytokines regulating monocyte recruitment; 3) the mononuclear phagocyte, its role in arterial debridement, metabolism of modified LDLs, a precursor of the cholesteryl ester-rich foam cell, and the secretion of neutral hydrolases, bioactive lipids and cytokines; 4) lymphocytes, as mediators of the inflammatory response and possible autoimmune reactions; 5) platelets, their roles in hemostasis, thrombosis, atherogenesis, and the repair process and 6) plasma LDLs, their oxidative modification by cells of the vessel wall and their roles in the injury process. The interactive processes among arterial and circulating components in both injury and repair is emphasized.


Duration of preventive antibiotic administration for penetrating abdominal trauma.
Eighty-two patients with penetrating abdominal trauma and visceral injuries requiring laparotomy were prospectively randomized to receive either 12 hours or five days of penicillin G potassium and doxycycline hyclate beginning before operation. Distribution between groups was equivalent for all risk factors except shock, which was more prevalent in the 12-hour group. Antibiotics were first administered an average of 64 minutes following injury, and 90% of all patients had received antibiotics and were being operated on within 3 hours 15 minutes. Overall infection rates were 17% in patients with colon penetration, 14% in patients without colon penetration but with other intestinal penetration, and 0% in patients without intestinal penetration. Twelve-hour and five-day antibiotic regimens were comparable in the prevention of postoperative infectious complications following penetrating abdominal injuries. Intestinal penetration was the most important risk factor for developmental of infectious complications in this patient population.


The Impact of Suprascapular Nerve Interventions in Patients with Frozen Shoulder: A Systematic Review and Meta-Analysis.
Frozen shoulder is a common condition resulting in severe pain and restricted range of motion. An assessment of the effectiveness of interventions may provide an improved understanding of the development and management of frozen shoulder. A literature search was conducted using Embase, the Cumulative Index of Nursing and Allied Health (CINAHL), the Cochrane Central Register of Controlled Trials (CENTRAL), and National Center for Biotechnology Information PubMed using relevant terms. Studies were included if they assessed the outcomes of interventions on the suprascapular nerve that aim to improve the symptoms of frozen shoulder. A database search returned 196 articles. After review, 9 articles met the inclusion criteria and were included in the analysis. Suprascapular nerve interventions (nerve block, pulsed radiofrequency lesioning) are associated with improvement in pain, motion, and function. Meta-analysis showed that pain (Hedges g, -3.084 [95% confidence interval (CI), -4.273 to -1.894]; p < 0.001) and range of motion (Hedges g, 2.204 [95% CI, 0.992 to 3.415]; p < 0.001) improved significantly following suprascapular nerve block (SSNB). SSNB is associated with significant improvements in shoulder pain and range of motion in patients with frozen shoulder. Further randomized controlled trials comparing SSNB with intra-articular injection and other nonoperative treatments are required to fully define its role in the management of frozen shoulder. Therapeutic Level III. See Instructions for Authors for a complete description of levels of evidence.


Migratory divides and their consequences for dispersal, population size and parasite-host interactions.
Populations of migratory birds differ in their direction of migration with neighbouring populations often migrating in divergent directions separated by migratory divides. A total of 26% of 103 passerine bird species in Europe had migratory divides that were located disproportionately often along a longitudinal gradient in Central Europe, consistent with the assumption of a Quaternary glacial origin of such divides in the Iberian and Balkan peninsulas followed by recolonization. Given that studies have shown significant genetic differentiation and reduced gene flow across migratory divides, we hypothesized that an absence of migratory divides would result in elevated rates of gene flow and hence a reduced level of local adaptation. In a comparative study, species with migratory divides had larger population sizes and population densities and longer dispersal distances than species without migratory divides. Species with migratory divides tended to be habitat generalists. Bird species with migratory divides had higher richness of blood parasites and higher growth rates of Staphylococcus on their eggs during the incubation period. There was weaker cell-mediated immunity in adults and stronger cell lysis in species with migratory divides. These findings may suggest that migratory divides constitute barriers to dispersal with consequences for ecology and evolution of distributions, population sizes, habitats and parasite-host interactions. They also suggest that migratory divides may play a role in local adaptation in host-parasite interactions.


Biotic and abiotic interactions in aquatic microcosms determine fate and toxicity of Ag nanoparticles. Part 1. Aggregation and dissolution.
To better understand their fate and toxicity in aquatic environments, we compared the aggregation and dissolution behavior of gum arabic (GA) and polyvinylpyrrolidone (PVP) coated Ag nanoparticles (NPs) in aquatic microcosms. There were four microcosm types: surface water; water and sediment; water and aquatic plants; or water, sediment, and aquatic plants. Dissolution and aggregation behavior of AgNPs were examined using ultracentrifugation, ultrafiltration, and asymmetrical flow field flow fractionation coupled to ultraviolet-visible spectroscopy, dynamic and static laser light scattering, and inductively coupled plasma mass spectrometry. Plants released dissolved organic matter (DOM) into the water column either through active or passive processes in response to Ag exposure. This organic matter fraction readily bound Ag ions. The plant-derived DOM had the effect of stabilizing PVP-AgNPs as primary particles, but caused GA-AgNPs to be removed from the water column, likely by dissolution and binding of released Ag ions on sediment and plant surfaces. The destabilization of the GA-AgNPs also corresponded with X-ray absorption near edge spectroscopy results which suggest that 22-28% of the particulate Ag was associated with thiols and 5-14% was present as oxides. The results highlight the potential complexities of nanomaterial behavior in response to biotic and abiotic modifications in ecosystems, and may help to explain differences in toxicity of Ag observed in realistic exposure media compared to simplified laboratory exposures.


Structure--activity study of the antibacterial peptide fallaxin.
Fallaxin is a 25-mer antibacterial peptide amide, which was recently isolated from the West Indian mountain chicken frog Leptodactylus fallax. Fallaxin has been shown to inhibit the growth of several Gram-negative bacteria including Enterobacter cloacae, Escherichia coli, Klebsiella pneumoniae, and Pseudomonas aeruginosa. Here, we report a structure-activity study of fallaxin based on 65 analogs, including a complete alanine scan and a full set of N- and C-terminal truncated analogs. The fallaxin analogs were tested for hemolytic activity and antibacterial activity against methicillin-resistant Staphylococcus aureus (MRSA), vancomycin-intermediate resistant S. aureus, (VISA), methicillin-susceptible S. aureus (MSSA), E. coli, K. pneumoniae, and P. aeruginosa. We identified several analogs, which showed improved antibacterial activity compared to fallaxin. Our best candidate was FA12, which displayed MIC values of 3.12, 25, 25, and 50 muM against E. coli, K. pneumoniae, MSSA, and VISA, respectively. Furthermore, we correlated the antibacterial activity with various structural parameters such as charge, hydrophobicity H, mean hydrophobic moment mu(H), and alpha-helicity. We were able to group the active and inactive analogs according to mean hydrophobicity H and mean hydrophobic moment mu(H). Far-UV CD-spectroscopy experiments on fallaxin and several analogs in buffer, in TFE, and in membrane mimetic environments (small unilamellar vesicles) indicated that a coiled-coil conformation could be an important structural trait for antibacterial activity. This study provides data that support fallaxin analogs as promising lead structures in the development of new antibacterial agents.


The immunomodulatory effect of sevoflurane in endotoxin-injured alveolar epithelial cells.
Endotoxin-induced lung injury is a useful experimental system for the characterization of immunopathologic mechanisms in acute lung injury. Although alveolar epithelial cells (AEC) are directly exposed to volatile anesthetics, there is limited information about the effect of anesthetics on these cells. In this study we investigated the effect of pretreatment with the inhaled anesthetic sevoflurane on lipopolysaccharide (LPS)-injured AEC. AEC were incubated with 1.1 vol % sevoflurane for 0.5 h, followed by LPS stimulation for 5 h. Expression of monocyte chemoattractant protein-1 (MCP-1), macrophage inflammatory protein-1beta (MIP-1beta), macrophage inflammatory protein-2 (MIP-2), cytokine-induced neutrophil chemoattractant-1 (CINC-1), and intercellular adhesion molecule-1 (ICAM-1) was analyzed. In addition, functional tests were performed through chemotaxis and adherence assays to underline the biological relevance of the findings. Exposure of AEC to sevoflurane resulted in a 50% downregulation of MCP-1 protein in the sevoflurane-LPS group when compared with non-sevoflurane- LPS cells (P < 0.05). MIP-1beta concentration in LPS-stimulated cells decreased by 32% with sevoflurane (P < 0.05), MIP-2 by 29% (P < 0.05), and CINC-1 by 20% (P < 0.05). ICAM-1 protein expression was attenuated by 36% (P < 0.05). This inhibition caused substantial changes in the inflammatory response of neutrophils. 33% less chemotactic activity was seen in sevoflurane-treated LPS cells (P < 0.001) as well as 47% decreased adhesion of neutrophils to AEC (P < 0.001). This study shows that sevoflurane alters the LPS-induced inflammatory response, not only with respect to the expression pattern of inflammatory mediators, but also regarding the biological consequences with less accumulation of effector cells such as neutrophils.


[Corneal wound healing after hyperopic PRK and LASIK].
The purpose of this study was to investigate the influence of different postoperative treatments on the wound healing reaction in the anterior stroma after PRK and in the interface area after LASIK. Seventy-two corneal buttons of refractively treated rabbit eyes underwent different postoperative eyedrop regimens with antibiotics and/or steroids or additional UV-B irradiation. Morphological and immunohistological investigations were performed 6 months postoperatively by light and transmission electron microscopy. PRK eyes showed interdigitations between the epithelia and the anterior stroma. LASIK-treated eyes showed only minor changes between epithelia and stroma in the incisional region. Only a slight increase in deposits of fibrillar extracellular matrix components were detectable in the interface region. The clinically important problem of haze after PRK is caused by the interdigitations between epithelia and anterior stroma. The delicate wound healing reactions in the interface region in LASIK eyes corresponded to the clinically visible minor changes in these corneas.


THUNDER 2: THeragnostic Utilities for Neoplastic DisEases of the Rectum by MRI guided radiotherapy.
Neoadjuvant chemoradiation therapy (nCRT) is the standard treatment modality in locally advanced rectal cancer (LARC). Since response to radiotherapy (RT) is dose dependent in rectal cancer, dose escalation may lead to higher complete response rates. The possibility to predict patients who will achieve complete response (CR) is fundamental. Recently, an early tumour regression index (ERI) was introduced to predict pathological CR (pCR) after nCRT in LARC patients. The primary endpoints will be the increase of CR rate and the evaluation of feasibility of delta radiomics-based predictive MRI guided Radiotherapy (MRgRT) model. Patients affected by LARC cT2-3, N0-2 or cT4 for anal sphincter involvement N0-2a, M0 without high risk features will be enrolled in the trial. Neoadjuvant CRT will be administered using MRgRT. The initial RT treatment will consist in delivering 55 Gy in 25 fractions on Gross Tumor Volume (GTV) plus the corresponding mesorectum and 45 Gy in 25 fractions on the drainage nodes. Chemotherapy with 5-fluoracil (5-FU) or oral capecitabine will be administered continuously. A 0.35 Tesla MRI will be acquired at simulation and every day during MRgRT. At fraction 10, ERI will be calculated: if ERI will be inferior than 13.1, the patient will continue the original treatment; if ERI will be higher than 13.1 the treatment plan will be reoptimized, intensifying the dose to the residual tumor at the 11th fraction to reach 60.1 Gy. At the end of nCRT instrumental examinations are to be performed in order to restage patients. In case of stable disease or progression, the patient will undergo surgery. In case of major or complete clinical response, conservative approaches may be chosen. Patients will be followed up to evaluate toxicity and quality of life. The number of cases to be enrolled will be 63: all the patients will be treated at Fondazione Policlinico Universitario A. Gemelli IRCCS in Rome. This clinical trial investigates the impact of RT dose escalation in poor responder LARC patients identified using ERI, with the aim of increasing the probability of CR and consequently an organ preservation benefit in this group of patients. ClinicalTrials.gov Identifier: NCT04815694 (25/03/2021).


Toddler dysregulated fear predicts continued risk for social anxiety symptoms in early adolescence.
Identifying early risk factors for the development of social anxiety symptoms has important translational implications. Accurately identifying which children are at the highest risk is of critical importance, especially if we can identify risk early in development. We examined continued risk for social anxiety symptoms at the transition to adolescence in a community sample of children (n = 112) that had been observed for high fearfulness at age 2 and tracked for social anxiety symptoms from preschool through age 6. In our previous studies, we found that a pattern of dysregulated fear (DF), characterized by high fear in low threat contexts, predicted social anxiety symptoms at ages 3, 4, 5, and 6 years across two samples. In the current study, we re-evaluated these children at 11-13 years of age by using parent and child reports of social anxiety symptoms, parental monitoring, and peer relationship quality. The scores for DF uniquely predicted adolescents' social anxiety symptoms beyond the prediction that was made by more proximal measures of behavioral (e.g., kindergarten social withdrawal) and concurrent environmental risk factors (e.g., parental monitoring, peer relationships). Implications for early detection, prevention, and intervention are discussed.


Nonexpanded mesenchymal stem cells for regenerative medicine: yield in stromal vascular fraction from adipose tissues.
The adipose-derived stromal vascular fraction (SVF) represents a rich source of mesenchymal cells, potentially able to differentiate into adipocytes, chondrocytes, osteoblasts, myocytes, cardiomyocytes, hepatocytes, and neuronal, epithelial, and endothelial cells. These cells are ideal candidates for use in regenerative medicine, tissue engineering, including gene therapy, and cell replacement cancer therapies. In this work, we aimed to the optimization of the adipose SVF-based therapy, and the effect of the collection site, surgical procedure, and tissue processing techniques on SVF yield was evaluated in terms of cell recovery and live cells, taking into account the effect of gender, age, and body mass index. Adipose tissue samples were recovered from 125 informed subjects (37 males and 88 females; mean age: 51.31 years; range: 15-87 years), and digested in different condition with collagenase. A multivariate linear model put in evidence that in males the best collection site in terms of yield is located in the abdomen, whereas in females the biopsy region do not influence cell recovery; the collection technique, the age, and the body mass index of donor seem not to influence the cell yield. The tissue-processing procedures strongly modify the yield and the vitality of cells: a collagenase concentration of 0.2% and a digestion time of 1 h could be chosen as the best operating conditions.


A fractal model for nuclear organization: current evidence and biological implications.
Chromatin is a multiscale structure on which transcription, replication, recombination and repair of the genome occur. To fully understand any of these processes at the molecular level under physiological conditions, a clear picture of the polymorphic and dynamic organization of chromatin in the eukaryotic nucleus is required. Recent studies indicate that a fractal model of chromatin architecture is consistent with both the reaction-diffusion properties of chromatin interacting proteins and with structural data on chromatin interminglement. In this study, we provide a critical overview of the experimental evidence that support a fractal organization of chromatin. On this basis, we discuss the functional implications of a fractal chromatin model for biological processes and propose future experiments to probe chromatin organization further that should allow to strongly support or invalidate the fractal hypothesis.


Lack of evidence does not justify neglect: how can we address unmet medical needs in calciphylaxis?
Calcific uraemic arteriolopathy (CUA), or calciphylaxis, is a rare disease predominantly occurring in comorbidity with dialysis. Due to the very low frequency of CUA, prospective studies on its management are lacking and even anecdotal reports on treatment remain scarce. Therefore, calciphylaxis is still a challenging disease with dismal prognosis urgently requiring adequate strategies for diagnosis and treatment.In an attempt to fill some of the current gaps in evidence on various, highly debated and controversial aspects of dialysis-associated calciphylaxis, 13 international experts joined the 1st Consensus Conference on CUA, held in Leuven, Belgium on 21 September 2015. The conference was supported by the European Calciphylaxis Network (EuCalNet), which is a task force of the ERA-EDTA scientific working group on Chronic Kidney Disease-Mineral and Bone Disorders (CKD-MBD). After an intense discussion, a 9-point Likert scale questionnaire regarding 20 items on calciphylaxis was anonymously answered by each participant. These 20 items addressed unsolved issues in terms of diagnosis and management of calciphylaxis. On the one hand, the analysis of the expert opinions identified areas of general consensus, which might be a valuable aid for physicians treating such a disease with less experience in the field. On the other hand, some topics such as the pertinence of skin biopsy and administration of certain treatments revealed divergent opinions. The aim of the present summary report is to provide some guidance for clinicians who face patients with calciphylaxis in the current setting of absence of evidence-based medicine.


A STRP-ed definition of Structured Tandem Repeats in Proteins.
Tandem Repeat Proteins (TRPs) are a class of proteins with repetitive amino acid sequences that have been studied extensively for over two decades. Different features at the level of sequence, structure, function and evolution have been attributed to them by various authors. And yet many of its salient features appear only when looking at specific subclasses of protein tandem repeats. Here, we attempt to rationalize the existing knowledge on Tandem Repeat Proteins (TRPs) by pointing out several dichotomies. The emerging picture is more nuanced than generally assumed and allows us to draw some boundaries of what is not a "proper" TRP. We conclude with an operational definition of a specific subset, which we have denominated STRPs (Structural Tandem Repeat Proteins), which separates a subclass of tandem repeats with distinctive features from several other less well-defined types of repeats. We believe that this definition will help researchers in the field to better characterize the biological meaning of this large yet largely understudied group of proteins.