Direct interaction between endothelial nitric-oxide synthase and dynamin-2. Implications for nitric-oxide synthase function.
Endothelial nitric-oxide synthase (eNOS) is regulated in part through specific protein interactions. Dynamin-2 is a large GTPase residing within similar membrane compartments as eNOS. Here we show that dynamin-2 binds directly with eNOS thereby augmenting eNOS activity. Double label confocal immunofluorescence demonstrates colocalization of eNOS and dynamin in both Clone 9 cells cotransfected with green fluorescent protein-dynamin and eNOS, as well as in bovine aortic endothelial cells (BAEC) expressing both proteins endogenously, predominantly in a Golgi membrane distribution. Immunoprecipitation of eNOS from BAEC lysate coprecipitates dynamin and, conversely, immunoprecipitation of dynamin coprecipitates eNOS. Additionally, the calcium ionophore, a reagent that promotes nitric oxide release, enhances coprecipitation of dynamin with eNOS in BAEC, suggesting the interaction between the proteins can be regulated by intracellular signals. In vitro studies demonstrate that glutathione S-transferase (GST)-dynamin-2 quantitatively precipitates both purified recombinant eNOS protein as well as in vitro transcribed (35)S-labeled eNOS from solution indicating a direct interaction between the proteins in vitro. Scatchard analysis of binding studies demonstrates an equilibrium dissociation constant (K(d)) of 27.6 nm. Incubation of purified recombinant eNOS protein with GST-dynamin-2 significantly increases eNOS activity as does overexpression of dynamin-2 in ECV 304 cells stably transfected with eNOS-green fluorescent protein. These studies demonstrate a direct protein-protein interaction between eNOS and dynamin-2, thereby identifying a new NOS-associated protein and providing a novel function for dynamin. These events may have relevance for eNOS regulation and trafficking within vascular endothelium.


Microarray analysis of global changes in gene expression during cardiac myocyte differentiation.
Significant progress has been made in defining pathways that mediate the formation of the mammalian heart. Little is known, however, about the genetic program that directs the differentiation of cardiac myocytes from their precursor cells. A major hindrance to this kind of investigation has been the absence of an appropriate cell culture model of cardiac myocyte differentiation. Recently, a subline of P19 cells (P19CL6) was derived that, following dimethyl sulfoxide (DMSO) treatment, differentiate efficiently over 10 days into spontaneously beating cardiac myocytes. We demonstrate that these cells are indeed cardiac myocytes as they express cell type-specific markers and exhibit electrophysiological properties indicative of cardiac myocytes. The requirement for DMSO stimulation in this paradigm was shown to be limited to the first 4 days, suggesting that critical events in the differentiation process occur over this interval. To uncover relationships among known genes and identify novel genes that mediate cardiac myocyte differentiation, a detailed time course of changes in global gene expression was carried out using cDNA microarrays. In addition to the activation of genes encoding cardiac transcription factors and structural proteins, increases were noted in the expression of multiple known genes and expressed sequence tags (ESTs). Analysis of the former suggested the involvement of a variety of signaling pathways in cardiac myocyte differentiation. The 16 ESTs whose expression was increased during the early, stimulus-dependent phase of cardiac myocyte differentiation may be novel regulators of this process. Thus this first report of large-scale changes in gene expression during cardiac myocyte differentiation has delineated relationships among the expression patterns of known genes and identified a number of novel genes that merit further study.


Scrotal imaging.
Pathological lesions within the scrotum are relatively rare in imaging except for ultrasonography. The diseases presented in the paper are usually found in men at the age of 15-45, i.e. men of reproductive age, and therefore they are worth attention. Scrotal ultrasound in infertile individuals should be conducted on a routine basis owing to the fact that pathological scrotal lesions are frequently detected in this population. Malignant testicular cancers are the most common neoplasms in men at the age of 20-40. Ultrasound imaging is the method of choice characterized by the sensitivity of nearly 100% in the differentiation between intratesticular and extratesticular lesions. In the case of doubtful lesions that are not classified for intra-operative verification, nuclear magnetic resonance is applied. Computed tomography, however, is performed to monitor the progression of a neoplastic disease, in pelvic trauma with scrotal injury as well as in rare cases of scrotal hernias involving the ureters or a fragment of the urinary bladder.


Radiolabelling of glucose-Tyr3-octreotate with 125I and analysis of its metabolism in rats: comparison with radiolabelled DOTA-Tyr3-octreotate.
Somatostatin analogues labelled with radiometals or radiohalogens are useful for the imaging and treatment of somatostatin receptor-containing tumours. In this study, the procedures for the radioiodination of glucose-Tyr3-octreotate (gluc-Tyr3-tate) and radiolabelling of DOTA-Tyr3-octreotate (DOTA-Tyr3-tate) with 111In, 177Lu and 125I were compared and their metabolism in rats was analyzed. The usefulness of high performance liquid chromatography (HPLC) analysis and instant thin-layer chromatography on silica gel (ITLC-SG) for both radiochemical purity determination and analysis of metabolism in urine was investigated. For labelling with radiometals, the formation of a complex with the 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA) functionality of the peptide was employed. Radioiodination was performed by the chloramime-T method. The radiochemical purity of radiolabelled peptides and the analyses of rat urine were determined by HPLC and/or ITLC-SG methods. Male Wistar rats were used in the elimination studies. DOTA-Tyr3-tate was simply radiolabelled with radiometals with high yield and high radiochemical purity. Stopping of the reaction was a critical step for radioiodination, therefore labelling of gluc-Tyr3-tate and DOTA-Tyr3-tate with 125I was not so simple and the reaction product had to be purified by preparative HPLC analysis. Whereas 111In-DOTA-Tyr3-tate and 177Lu-DOTA-Tyr3-tate were eliminated in rat urine in a practically unchanged form, a significant proportion of metabolites was observed with radioiodinated peptides, particularly at longer time intervals. Labelling of DOTA-Tyr3-tate with radiometals is simple and the radiochemical purity of prepared compounds is very high, while iodination of the peptides demands purification of the product by preparative HPLC. The analysis of rat urine showed that excretion of radioiodinated peptides included a significant proportion of metabolites.


Dancing Salsa with Machines-Filling the Gap of Dancing Learning Solutions.
Dancing is an activity that positively enhances the mood of people that consists of feeling the music and expressing it in rhythmic movements with the body. Learning how to dance can be challenging because it requires proper coordination and understanding of rhythm and beat. In this paper, we present the first implementation of the Dancing Coach (DC), a generic system designed to support the practice of dancing steps, which in its current state supports the practice of basic salsa dancing steps. However, the DC has been designed to allow the addition of more dance styles. We also present the first user evaluation of the DC, which consists of user tests with 25 participants. Results from the user test show that participants stated they had learned the basic salsa dancing steps, to move to the beat and body coordination in a fun way. Results also point out some direction on how to improve the future versions of the DC.


College campuses and COVID-19 mitigation: clinical and economic value.
Decisions around US college and university operations will affect millions of students and faculty amidst the COVID-19 pandemic. We examined the clinical and economic value of different COVID-19 mitigation strategies on college campuses. We used the Clinical and Economic Analysis of COVID-19 interventions (CEACOV) model, a dynamic microsimulation that tracks infections accrued by students and faculty, accounting for community transmissions. Outcomes include infections, $/infection-prevented, and $/quality-adjusted-life-year ($/QALY). Strategies included extensive social distancing (ESD), masks, and routine laboratory tests (RLT). We report results per 5,000 students (1,000 faculty) over one semester (105 days). Mitigation strategies reduced COVID-19 cases among students (faculty) from 3,746 (164) with no mitigation to 493 (28) with ESD and masks, and further to 151 (25) adding RLTq3 among asymptomatic students and faculty. ESD with masks cost $168/infection-prevented ($49,200/QALY) compared to masks alone. Adding RLTq3 ($10/test) cost $8,300/infection-prevented ($2,804,600/QALY). If tests cost $1, RLTq3 led to a favorable cost of $275/infection-prevented ($52,200/QALY). No strategies without masks were cost-effective. Extensive social distancing with mandatory mask-wearing could prevent 87% of COVID-19 cases on college campuses and be very cost-effective. Routine laboratory testing would prevent 96% of infections and require low cost tests to be economically attractive.


Effects of diffusion and mixing pattern on microfluidic-assisted synthesis of chitosan/ATP nanoparticles.
Chitosan (CHI) nanoparticles present promising applications in pharmaceutical and biomedical fields, including drug and gene delivery. Among different approaches, microfluidics emerges as a resourceful tool for nanoparticle production in low-cost, reproducible processes with predictable fluid dynamics. However, microfluidic-assisted synthesis of CHI nanoparticles has not been widely explored in the literature. In this context, we systematically investigated different process parameters that influence the synthesis of CHI/ATP nanoparticles. We highlight the effects and limitations of diffusion and distinct mixing patterns developed through the microchannels on the final physicochemical characteristics of CHI/ATP nanoparticles produced. To address these hurdles, here we describe a simple, feasible, and reproducible method for the production of CHI/ATP nanoparticles. This strategy enables the development of a continuous and homogeneous production process for CHI nanoparticles to be applied in the most varied fields of research.


Components of the transforming growth factor-beta family and the pathogenesis of human Achilles tendon pathology--a genetic association study.
Achilles tendon pathology is a multifactorial condition for which various risk factors, including genetic factors, have been identified. Gene transfection of two members of the TGF-β family, TGF-β1 and growth/differentiation factor-5 (GDF-5), have been shown to enhance tendon repair and mechanical strength within animal Achilles tendon injury models. The objective of this study was to investigate whether two functional 5' untranslated region (UTR) single nucleotide polymorphisms (SNPs), the TGFB1 rs1800469 variant and the GDF5 rs143383 variant, were associated with ATP within an Australian ('AUS') and a South African ('SA') case-control cohort. One hundred and seventy-one subjects (58 AUS and 112 SA) with Achilles tendon pathology (ATP group) and 235 (142 AUS and 96 SA) asymptomatic control (CON group) subjects were genotyped for the selected SNPs using custom-designed Taqman assays. A χ(2)-analysis or Fisher's exact test was used to analyse any differences in the genotype and allele frequencies. Significance was accepted when P < 0.05. There were no significant TGFB1 rs1800469 genotype (P = 0.491) or allele (P = 0.400) frequency differences between the ATP and CON groups. The TT genotype of the GDF5 rs143383 variant was significantly over-represented in the ATP group of the AUS cohort [P = 0.011; odds ratio (OR) = 2.24; 95% CI 1.21, 4.16], and when the AUS and SA cohorts were combined (P = 0.004; OR = 1.82; 95% CI 1.23, 2.74). In conclusion, this study suggests that individuals with a TT genotype of the functional GDF5 rs143383 variant have twice the risk of developing ATP. This finding highlights a role of GDF-5 in the pathogenesis of Achilles tendon pathology.


Inhibition of intestinal polyp formation by pitavastatin, a HMG-CoA reductase inhibitor.
It has been suggested that hyperlipidemia is positively associated with colon carcinogenesis. Statins, 3-hydroxy-3-methyl-glutaryl-coenzyme A reductase inhibitors, reduce serum lipid levels. In this study, we clarified the effects of a novel chemically synthesized statin, pitavastatin, on intestinal polyp formation in Min mice, and further examined serum lipid and adipocytokine levels, and proinflammatory and adipocytokine gene levels in intestinal mucosa of Min mice. Treatment with pitavastatin at doses of 20 and 40 ppm decreased the total number of polyps dose-dependently to 85.2% and 65.8% (P < 0.05) of the untreated value, respectively. Serum levels of total cholesterol and triglyceride were slightly reduced and those of IL-6, leptin, and MCP-1 were decreased by 40-ppm pitavastatin treatment. mRNA expression levels of cyclooxygenase-2, IL-6, inducible nitric oxide (iNOS), MCP-1, and Pai-1 were significantly reduced in intestinal nonpolyp parts by pitavastatin treatment. Among them, iNOS mRNA levels were also reduced in the intestinal polyps. Moreover, oxidative stress represented by 8-nitroguanosine in the small intestinal epithelial cells was reduced by pitavastatin treatment. Related to these proinflammatory genes, PPARγ activity was activated in the intestinal nonpolyp parts and in the liver of Min mice with pitavastatin treatment. These results indicated that pitavastatin has potential benefit for the suppression of intestinal polyp development.


Alterations in cerebrospinal fluid proteins in a presymptomatic primary glioma model.
Understanding the early relationship between brain tumor cells and their environment could lead to more sensitive biomarkers and new therapeutic strategies. We have been using a rodent model of neurocarcinogenesis in which all animals develop brain tumors by six months of age to establish two early landmarks in glioma development: the appearance of a nestin(+) cell at thirty days of age and the appearance of cellular hyperplasia between 60 and 120 days of age. We now report an assessment of the CSF proteome to determine the changes in protein composition that occur during this period. Nestin(+) cell clusters and microtumors were assessed in 63 ethylnitrosourea-exposed rats on 30, 60, and 90 days of age. CSF was obtained from the cisterna magna from 101 exposed and control rats at 30, 60, and 90 days and then analyzed using mass spectrometry. Differentially expressed peaks were isolated and identified. Nestin(+) cells were noted in all ethylnitrosourea-exposed rats assessed pathologically. Small microtumors were noted in 0%, 18%, and 67% of 30-, 60-, and 90-day old rats, respectively (p<0.05, Chi square). False Discovery Rate analysis of peak intensities showed that the number of true discoveries with p<0.05 increased markedly with increasing age. Isolation and identification of highly differentially detected proteins at 90 days of age revealed increases in albumin and a fragment of α1 macroglobulin and alterations in glutathionylated transthyretin. The presence of increased albumin, fragments of cerebrospinal fluid proteins, and glutathione breakdown in temporal association with the development of cellular hyperplasia, suggests that, similar to many other systemic cancers, inflammation and oxidative stress is playing an important early role in the host's response to brain tumor development and may be involved in affecting the early growth of brain tumor.


Dietary indoles and isothiocyanates that are generated from cruciferous vegetables can both stimulate apoptosis and confer protection against DNA damage in human colon cell lines.
The natural indoles 3,3'-diindolylmethane (DIM), ascorbigen (ASG), indole-3-carbinol (I3C), and indolo[3,2-b]carbazole (ICZ), as well as the natural isothiocyanates sulforaphane (SUL), benzyl isothiocyanate (BITC) and phenethyl isothiocyanate (PEITC), all possess cancer chemopreventive properties. It is now shown that DIM, ICZ, SUL, and BITC can each stimulate apoptosis in human colon adenocarcinoma LS-174 and Caco-2 cells. Treatment of LS-174 cells with nontoxic doses of DIM, ASG, I3C, or ICZ affected an increase of up to 21-fold in cytochrome P450 1A1 (CYP1A1). None of these indoles caused an elevation in either aldo-keto reductase 1C1 (AKR1C1) or the gamma-glutamylcysteine synthetase heavy subunit (GCS(h)), but DIM, I3C, and ICZ produced a very modest increase in NAD(P)H:quinone oxidoreductase 1 (NQO1). By contrast, nontoxic doses of SUL, BITC, or PEITC failed to induce expression of CYP1A1 in LS-174 cells, but caused an increase of between 11- and 17-fold in the protein levels of AKR1C1, NQO1, and GCS(h). Treatment of the colon cell line with ICZ or SUL caused increases in the levels of mRNA for CYP1A1, AKR1C1, and NQO1 that were consistent with the enzyme data. Exposure of Caco-2 cells to media containing indoles or isothiocyanates gave similar results to those obtained using LS-174 cells. Evidence is presented that the ability of indoles and isothiocyanates to stimulate either xenobiotic response element- or antioxidant response element-driven gene expression accounts for the two groups of phytochemicals inducing different gene batteries. Pretreatment of LS-174 cells for 24 h with ICZ and SUL before exposure for 24 h to benzo(a)pyrene (BaP) reduced to <20% the number of single-strand DNA breaks produced by the carcinogen. Neither ICZ alone nor SUL alone were able to confer the same degree of protection against DNA damage produced by BaP as they achieved in combination. Similar results were obtained with H(2)O(2) as the genotoxic agent. Together, these phytochemicals may prevent colon tumorigenesis by both stimulating apoptosis and enhancing intracellular defenses against genotoxic agents.


Successful outcome of shell fractures in freshwater turtles treated with plate or screws and wire fixation techniques: 51 cases (2014-2019).
To evaluate the outcome of surgical fixation of shell fractures in rehabilitated wild freshwater turtles. 51 freshwater turtles with 86 shell fractures. The medical record database of a wildlife rehabilitation center in Wisconsin was searched from 2014 through 2019 for records of freshwater turtles with shell fractures repaired with a plate technique, screws and wire technique, or both. Signalment, fracture location, therapeutic approach (including the type of hardware used for repair), dry-docking duration, time to hardware removal, postremoval care, and outcome were evaluated. 36 of 51 (71%) turtles with shell fractures experienced successful fracture healing following surgical hardware fixation, and 33 (65%) were released. Shells of 38 (75%) turtles were repaired with plates only, 5 (10%) turtles with wire only, and 8 (16%) turtles with a combination of plates and screws and wires. Of the 51 turtles, 13 (25%) did not survive > 4 weeks following hardware repair, leaving 38 animals available to assess fracture healing. Median time to start staged removal was 42 days (range, 35 to 49 days) and to complete removal of the applied hardware was 56 days (range, 26 to 77 days). Complications associated with placement of the hardware occurred in 6 of 38 (16%) turtles. Complications included screw hole infection (4/38 [11%]), fracture necrosis (1/38 [3%]), and deep screw hole penetration (1/38 [3%]). Results indicated that shell fractures in freshwater turtles treated with surgical fixation techniques had a successful outcome. Most complications were minor, and fractures improved with time, resulting in acceptable fracture healing for release.


Human alpha 7 acetylcholine receptor: cloning of the alpha 7 subunit from the SH-SY5Y cell line and determination of pharmacological properties of native receptors and functional alpha 7 homomers expressed in Xenopus oocytes.
The alpha-bungarotoxin-binding acetylcholine receptors from the human neuroblastoma cell line SH-SY5Y were found to cross-react with some monoclonal antibodies to alpha 7 subunits of nicotinic acetylcholine receptors from chicken brain. The human alpha 7 subunit cDNA from SH-SY5Y was cloned, revealing 94% amino acid sequence identity to rat alpha 7 subunits and 92% identity to chicken alpha 7 subunits. Native human alpha 7 receptors showed affinities for some ligands similar to those previously observed with native chicken alpha 7 receptors, but for other ligands there were large species-specific differences in binding affinity. These results paralleled properties of alpha 7 homomers expressed in Xenopus oocytes. Human alpha 7 homomers exhibited rapidly desensitizing, inwardly rectifying, agonist-induced, cation currents that triggered Ca(2+)-sensitive Cl- channels in the oocytes. A change in efficacy from partial agonist for chicken alpha 7 homomers to full agonist for human alpha 7 homomers was exhibited by 1,1-dimethyl-4-phenylpiperazinium. This result reveals a large species-specific pharmacological difference, despite small differences in alpha 7 sequences. This is important for understanding the effects of these drugs in humans and for identifying amino acids that may contribute to the acetylcholine binding site, for analysis by in vitro mutagenesis. These results also characterize properties of native alpha 7 receptors and alpha 7 homomers that will provide criteria for functional properties expected of structural subunits, when these can be identified, cloned, and coexpressed with alpha 7 subunits.


A critical assessment of the WHO responsiveness tool: lessons from voluntary HIV testing and counselling services in Kenya.
Health, fair financing and responsiveness to the user's needs and expectations are seen as the essential objectives of health systems. Efforts have been made to conceptualise and measure responsiveness as a basis for evaluating the non-health aspects of health systems performance. This study assesses the applicability of the responsiveness tool developed by WHO when applied in the context of voluntary HIV counselling and testing services (VCT) at a district level in Kenya. A mixed method study was conducted employing a combination of quantitative and qualitative research methods concurrently. The questionnaire proposed by WHO was administered to 328 VCT users and 36 VCT counsellors (health providers). In addition to the questionnaire, qualitative interviews were carried out among a total of 300 participants. Observational field notes were also written. A majority of the health providers and users indicated that the responsiveness elements were very important, e.g. confidentiality and autonomy were regarded by most users and health providers as very important and were also reported as being highly observed in the VCT room. However, the qualitative findings revealed other important aspects related to confidentiality, autonomy and other responsiveness elements that were not captured by the WHO tool. Striking examples were inappropriate location of the VCT centre, limited information provided, language problems, and concern about the quality of counselling. The results indicate that the WHO developed responsiveness elements are relevant and important in measuring the performance of voluntary HIV counselling and testing. However, the tool needs substantial revision in order to capture other important dimensions or perspectives. The findings also confirm the importance of careful assessment and recognition of locally specific aspects when conducting comparative studies on responsiveness of HIV testing services.


Enhancement of Bacillus subtilis Lipopeptide Biosurfactants Production through Optimization of Medium Composition and Adequate Control of Aeration.
Interest in biosurfactants has increased considerably in recent years, as they are potentially used in many commercial applications in petroleum, pharmaceuticals, biomedical, and food processing industries. Since improvement of their production was of great importance to reduce the final coast, cultural conditions were analyzed to optimize biosurfactants production from Bacillus subtilis SPB1 strain. A high yield of biosurfactants was obtained from a culture of B. subtilis using carbohydrate substrate as a carbon source; among carbohydrates, glucose enhanced the best surfactin production. The optimum glucose concentration was 40 g/L. Higher amount of biosurfactants was obtained using 5 g/L of urea as organic nitrogen source and applying C/N ratio of 7 with ammonium chloride as inorganic nitrogen source. The highest amount of biosurfactants was recorded with the addition of 2% kerosene. Moreover, it was shown, using an automated full-controlled 2.6 L fermenter, that aeration of the medium, which affected strongly the growth regulated biosurfactants synthesis by the producing cell. So that, low or high aerations lead to a decrease of biosurfactants synthesis yields. It was found that when using dissolved oxygen saturation of the medium at 30%, biosurfactants production reached 4.92 g/L.


Continuous tuneable droplet ejection via pulsed surface acoustic wave jetting.
We report a miniaturised platform for continuous production of single or multiple liquid droplets with diameters between 60 and 500 μm by interfacing a capillary-driven self-replenishing liquid feed with pulsed excitation of focussed surface acoustic waves (SAWs). The orifice-free operation circumvents the disadvantages of conventional jetting systems, which are often prone to clogging that eventuates in rapid degradation of the operational performance. Additionally, we show the possibility for flexibly tuning the ejected droplet size through the pulse width duration, thus avoiding the need for a separate device for every different droplet size required, as is the case for systems in which the droplet size is set by nozzles and orifices, as well as preceding ultrasonic jetting platforms where the droplet size is controlled by the operating frequency. Further, we demonstrate that cells can be jetted and hence printed onto substrates with control over the cell density within the droplets down to single cells. Given that the jetting does not lead to significant loss to the cell's viability or ability to proliferate, we envisage that this versatile jetting method can potentially be exploited with further development for cell encapsulation, dispensing and 3D bioprinting applications.


There are two sides to every story: implications of asymmetry on breast support requirements for sports bra manufacturers.
This study aimed to investigate: (1) the prevalence and magnitude of breast movement asymmetry, (2) the interaction between static and dynamic breast asymmetry and (3) the influence of sports bras on breast asymmetry during running. Position data were collected from 167 females whilst treadmill running and then a sub-group of 12 participants in different bra conditions. Breast movement asymmetry existed in 89% of participants, with resultant static breast position asymmetry larger in participants displaying dynamic asymmetry. Asymmetry was most commonly caused (60% to 75%) by greater movement of the left breast. No significant relationships were found between asymmetry and bra size or breast pain. Sports bras reduced asymmetry prevalence from 75% to 33% of participants in the antero-posterior direction but only from 75% to 67% of participants in the infero-superior direction. The magnitude of range-of-motion asymmetry reduced from 67 mm with no bra to between 6 and 64 mm in-bra in the infero-superior direction, with the best performing bra incorporating encapsulating cups and adjustable straps and underband. It is recommended that sports bras allow underband and strap adjustment to facilitate individual breast support and that asymmetry is considered when designing and fitting bras, which could utilise resultant asymmetry measured statically.


Tolvaptan resistance is related with a short-term poor prognosis in patients with lung cancer and syndrome of inappropriate anti-diuresis.
Purpose: Tolvaptan (TVP), a vasopressin receptor antagonist, represents a therapeutic option in the syndrome of inappropriate anti-diuresis (SIAD). The aim of this study was to evaluate the effect of TVP to treat and solve hyponatremia in oncologic patients. Methods: 15 oncologic patients who developed SIAD have been enrolled. Patients receiving TVP belonged to group A, whereas group B was characterized by hyponatremic patients treated with hypertonic saline solutions and fluid restriction. Results: In group A, the correction of serum sodium was achieved after 3.7±2.8 days. In group B, the target levels were obtained more slowly, after 5.2±3.1 days (p: 0.01) than in group A. The hospital stay and incidence of re-hospitalization were higher in group B than in group A. In this latter, 37% of patients had hyponatremic relapses, notwithstanding the progressive increase of doses from 7.5 to 60 mg per day of TVP, revealing a complete lack of response to TVP. In these patients, a growth of tumor mass or new metastatic lesions has been revealed. Conclusion: TVP improved hyponatremia more efficiently and stably than hypertonic solutions and fluid restrictions. Positive consequences have been obtained about the rate of chemotherapeutical cycles concluded, hospital stay, rate of relapse of hyponatremia, and re-hospitalization. Our study also suggested potential prognostic information that could be deduced from TVP patients, in whom sudden and progressive hyponatremia occurred, despite TVP dosage increase. A re-staging of these patients to rule out tumor mass growth or new metastatic lesions is suggested.


Caveolin-1/3 double-knockout mice are viable, but lack both muscle and non-muscle caveolae, and develop a severe cardiomyopathic phenotype.
The caveolin gene family consists of caveolins 1, 2, and 3. Caveolins 1 and 2 are co-expressed in many cell types, such as endothelial cells, fibroblasts, smooth muscle cells and adipocytes, where they form a heteroligomeric complex. In contrast, the expression of caveolin-3 is muscle-specific. Thus, the expression of caveolin-1 is required for caveolae formation in non-muscle cells, while the expression of caveolin-3 drives caveolae formation in striated muscle cell types (cardiac and skeletal). To create a truly caveolae-deficient mouse, we interbred Cav-1 null mice and Cav-3 null mice to generate Cav-1/Cav-3 double-knockout (Cav-1/3 dKO) mice. Here, we report that Cav-1/3 dKO mice are viable and fertile, despite the fact that they lack morphologically identifiable caveolae in endothelia, adipocytes, smooth muscle cells, skeletal muscle fibers, and cardiac myocytes. We also show that these mice are deficient in all three caveolin gene products, as caveolin-2 is unstable in the absence of caveolin-1. Interestingly, Cav-1/3 dKO mice develop a severe cardiomyopathy. At 2 months of age, analysis of Cav-1/3 dKO hearts via gated magnetic resonance imaging reveals a dramatic increase in left ventricular wall thickness, as compared with Cav-1-KO, Cav-3 KO, and wild-type mice. Further functional analysis of Cav-1/3 dKO hearts via transthoracic echocardiography demonstrates hypertrophy and dilation of the left ventricle, with a significant decrease in fractional shortening. As predicted, Northern analysis of RNA derived from the left ventricle of Cav-1/3 dKO mice shows a dramatic up-regulation of the atrial natriuretic factor message, a well-established biochemical marker of cardiac hypertrophy. Finally, histological analysis of Cav-1/3 dKO hearts reveals hypertrophy, disorganization, and degeneration of the cardiac myocytes, as well as chronic interstitial fibrosis and inflammation. Thus, dual ablation of both Cav-1 and Cav-3 genes in mice leads to a pleiotropic defect in caveolae formation and severe cardiomyopathy.


Pharmacokinetics of pimobendan following oral administration to New Zealand White rabbits (Oryctolagus cuniculus).
To determine the pharmacokinetics and potential adverse effects of pimobendan after oral administration in New Zealand White rabbits (Ocytolagus cuniculi). 10 adult sexually intact (5 males and 5 females) rabbits. 2 pilot studies were performed with a pimobendan suspension or oral tablets. Eight rabbits received 7.5 mg of pimobendan (mean 2.08 mg/kg) suspended in a critical care feeding formula. Plasma concentrations of pimobendan and O-demethylpimobendan (ODMP) were measured, and pharmacokinetic parameters were calculated for pimobendan by noncompartmental analysis. Body weight, food and water consumption, mentation, urine, and fecal output were monitored. Mean ± SD maximum concentration following pimobendan administration was 15.7 ± 7.54 ng/mL and was detected at 2.79 ± 1.25 hours. The half-life was 3.54 ± 1.32 hours. Plasma concentrations of pimobendan were detectable for up to 24 hours. The active metabolite, ODMP, was detected in rabbits for 24 to 36 hours. An adverse event occurred following administration of pimobendan in tablet form in 1 pilot study, resulting in death secondary to aspiration. No other adverse events occurred. Plasma concentrations of pimobendan were lower than previously reported for dogs and cats, despite administration of higher doses, and had longer time to maximum concentration and half-life. Based on this study, 2 mg/kg of pimobendan in a critical care feeding formulation should maintain above a target plasma concentration for 12 to 24 hours. However, further studies evaluating multiple-dose administration as well as pharmacodynamic studies and clinical trials in rabbits with congestive heart failure are needed to determine accurate dose and frequency recommendations.


The role of the endocannabinoid system in Alzheimer's disease: facts and hypotheses.
Unlike other neuroinflammatory disorders, like Parkinson's disease, Huntington's disease and multiple sclerosis, little is still known of the role of the endocannabinoid system in Alzheimer's disease (AD). This is partly due to the poor availability of animal models that are really relevant to the human disease, and to the complexity of AD as compared to other neurological states. Nevertheless, the available data indicate that endocannabinoids are likely to play in this disorder a role similar to that suggested in other neurodegenerative diseases, that is, to represent an endogenous adaptive response aimed at counteracting both the neurochemical and inflammatory consequences of beta-amyloid-induced tau protein hyperactivity, possibly the most important underlying cause of AD. Furthermore, plant and synthetic cannabinoids, and particularly the non-psychotropic cannabidiol, might also exert other, non-cannabinoid receptor-mediated protective effects, including, but not limited to, anti-oxidant actions. There is evidence, from in vivo studies on beta-amyloid-induced neurotoxicity, also for a possible causative role of endocannabinoids in the impairment in memory retention, which is typical of AD. This might open the way to the use of cannabinoid receptor antagonists as therapeutic drugs for the treatment of cognitive deficits in the more advanced phases of this disorder. The scant, but nevertheless important literature on the regulation and role of the endocannabinoid system in AD, and on the potential treatment of this disorder with cannabinoids and endocannabinoid-based drugs, are discussed in this mini-review.


Pro-apoptotic Par-4 and dopamine D2 receptor in temporal cortex in schizophrenia, bipolar disorder and major depression.
Although the etiology of schizophrenia remains unknown, diverse neuropathological evidence suggests a disorder of synaptic connectivity. Apoptosis is a form of cell death that helps determine synaptic circuitry during neurodevelopment and altered regulation of apoptosis has been implicated in schizophrenia. Prostate apoptosis response-4 (Par-4) is an upstream regulator of apoptosis preferentially localized to synapses. Brain Par-4 levels are upregulated in response to pro-apoptotic stimuli in rodent models and in patients with classic neurodegenerative diseases. Recently, Par-4 was also found to form a complex with the dopamine D2 receptor (D2DR) in competition with the calcium-binding protein calmodulin, implicating Par-4 as an important regulatory component in normal dopamine signaling. Interestingly, mutant mice with disrupted Par-4/D2DR interaction demonstrated depressive-like behaviors, suggesting a potential role for Par-4 in both depression and schizophrenia. In this study, Par-4, D2DR and calmodulin protein levels were measured using semiquantitative Western blotting in postmortem temporal cortex in subjects with schizophrenia, major depression and bipolar disorder. Compared to normal controls, mean Par-4 levels appeared slightly lower in schizophrenia and bipolar disorder. However, in major depression, Par-4 was decreased by 67% compared to normal controls. No differences were found between any groups for calmodulin or for the D2DR 48 kDa band. The D2DR 98 kDa band was lower by 50% in the schizophrenia compared to control groups. Changes in the Par-4/D2DR signaling pathway represent a novel mechanism that may link apoptotic and dopamine signaling pathways in major depression and schizophrenia.


Human coagulation abnormalities during acute exposure to hypobaric hypoxia.
Multiple coagulation studies were carried out in eight healthy young men at sea level (SL) and after 1, 24, and 48 h at a simulated altitude of 4,400 m. Platelet aggregation, as induced by ADP, epinephrine, and collagen, was not significantly altered by high-altitude (HA) exposure. Mean 2,3-diphosphoglycerate, a physiological inhibitor of platelet aggregation, rose (P less than 0.001) after 24 h at HA and remained elevated while no changes in circulating catecholamines were observed. Platelet count, factor 3 availability, and membrane lipid peroxide formation were likewise unaltered at HA, as were prothrombin and thrombin times and protamine paracoagulation test. However, mean partial thromboplastin time was significantly shortened (P less than 0.01) after 1 and 24 h at HA, recovering to SL control by 48 h. Fibrinogen and factor VIII levels also fell (P less than 0.01 and P less than 0.02) after 1 h at HA but returned to the preexposure values by 24 h. Fibrin degradation products were transiently detectable in three subjects at HA. Thus, although normal platelet function did not appear to be modified by short-term exposure to simulated high altitude, evidence for a coagulopathy was obtained.


Reviewing dengue: still a neglected tropical disease?
Dengue is currently listed as a "neglected tropical disease" (NTD). But is dengue still an NTD or not? Classifying dengue as an NTD may carry advantages, but is it justified? This review considers the criteria for the definition of an NTD, the current diverse lists of NTDs by different stakeholders, and the commonalities and differences of dengue with other NTDs. We also review the current research gaps and research activities and the adequacy of funding for dengue research and development (R&D) (2003-2013). NTD definitions have been developed to a higher precision since the early 2000s, with the following main features: NTDs are characterised as a) poverty related, b) endemic to the tropics and subtropics, c) lacking public health attention, d) having poor research funding and shortcomings in R&D, e) usually associated with high morbidity but low mortality, and f) often having no specific treatment available. Dengue meets most of these criteria, but not all. Although dengue predominantly affects resource-limited countries, it does not necessarily only target the poor and marginalised in those countries. Dengue increasingly attracts public health attention, and in some affected countries it is now a high profile disease. Research funding for dengue has increased exponentially in the past two decades, in particular in the area of dengue vaccine development. However, despite advances in dengue research, dengue epidemics are increasing in frequency and magnitude, and dengue is expanding to new areas. Specific treatment and a highly effective vaccine remain elusive. Major research gaps exist in the area of integrated surveillance and vector control. Hence, although dengue differs from many of the NTDs, it still meets important criteria commonly used for NTDs. The current need for increased R&D spending, shared by dengue and other NTDs, is perhaps the key reason why dengue should continue to be considered an NTD.


Unprecedentedly enhanced solar photocatalytic activity of a layered titanate simply integrated with TiO2 nanoparticles.
We report a simple, low-cost methodology for unprecedentedly enhancing the photocatalytic activity of layered inorganic semiconductors. A layered titanate with a lepidocrocite-type structure scarcely showed photocatalytic activity for a test reaction, the oxidative decomposition of formic acid in water into CO2, under simulated solar light, but it showed highly enhanced photocatalytic activity upon mixing with a much smaller amount (approximately 10 wt%) of commercially available TiO2 nanoparticles (P25) in water. The photocatalytic activity of the mixture was approximately 5 times that of P25, a benchmark photocatalyst. From various analyses, the enhancement resulted from the transfer of photoexcited electrons from the layered titanate to P25 at their particle interfaces and retardation of charge recombination. When applied to a photocatalyst for H2 production from water containing methanol under simulated solar light, the layered titanate/P25 mixture showed considerably enhanced activity and the apparent quantum yield was 23% (at 320 nm). By replacing P25 with Pt co-catalyst-loaded P25, the apparent quantum yield of the mixture increased from 23 to 73%, although an extremely small amount (below 0.06%) of Pt was used in the system.


Genotype-phenotype analysis of the Crohn's disease susceptibility haplotype on chromosome 5q31.
Recent molecular data suggest that genetic factors may underlie the disease heterogeneity observed in both ulcerative colitis (UC) and Crohn's disease (CD). A locus on chromosome 5q has been implicated in susceptibility to CD, and recently refined by linkage disequilibrium mapping to a conserved 250 kb haplotype (5q31). No data regarding the contribution of this locus to clinical phenotype exist. In this case control study, we investigated the contribution of this haplotype to both susceptibility and phenotype of CD and UC. We studied 330 Caucasian CD and 457 UC patients recruited from a single UK centre. Association with disease susceptibility and phenotype was analysed with haplotypes reconstructed from three single nucleotide polymorphisms chosen to span this susceptibility region. Evidence for possible genetic epistasis between IBD5 and NOD2/CARD15 was sought. Linkage disequilibrium across this region was confirmed, with two haplotypes comprising 88% of all chromosomes. Susceptibility to CD, but not to UC, was associated with homozygosity for a common haplotype, H2 (p(c)=0.002; relative risk (RR) 2.0). Genotype-phenotype analyses demonstrated that this association was particularly strong in patients with perianal disease (p(c)=0.0005; RR 1.7), especially in individuals homozygous for this haplotype (p(c)=0.0005; RR 3.0). Importantly, no association with H2 was found in 186 patients without perianal disease. No evidence of epistasis between IBD5 and NOD2/CARD15 was demonstrated. The IBD5 risk haplotype is associated with CD only. Genotype-phenotype analysis reveals that the strongest association is observed in patients with perianal CD. While the precise gene involved is unclear, these data provide further molecular evidence for a genetic basis of the clinical heterogeneity of CD.


Designing for redundancy: nurses experiences with the wireless nurse call system.
Patients rely on the nurse call system when in need of nurses' assistance, while nurses rely on the nurse call system to coordinate work. In order to handle the highly mobile nature of nurses' work, a wireless nurse call system has been introduced at a Norwegian hospital. In light of findings from previous research that identify challenges associated with wireless nurse call systems, we have conducted qualitative and ethnographically inspired fieldwork, i.e. workshops including both ordinary and student nurses to describe nurses' present system use. We further identify challenges related to wireless nurse call system usage, in particular how nurses manage the interruptive nature of wireless nurse calls so as to ensure continuous patient support. Our analysis acknowledges that High Reliability Organization (HRO) principles are transferable to a hospital organization. Based on our data, we propose improvements for design that hopefully can help reduce disruptive interruptions, ensure quick responses to patients, and further improve quality of care and patient safety. Further, we suggest that technology should facilitate and strengthen the redundancy of function.


Structural insight into the mechanisms of transport across the Salmonella enterica Pdu microcompartment shell.
Bacterial microcompartments are a functionally diverse group of proteinaceous organelles that confine specific reaction pathways in the cell within a thin protein-based shell. The propanediol utilizing (Pdu) microcompartment contains the reactions for metabolizing 1,2-propanediol in certain enteric bacteria, including Salmonella. The Pdu shell is assembled from a few thousand protein subunits of several different types. Here we report the crystal structures of two key shell proteins, PduA and PduT. The crystal structures offer insights into the mechanisms of Pdu microcompartment assembly and molecular transport across the shell. PduA forms a symmetric homohexamer whose central pore appears tailored for facilitating transport of the 1,2-propanediol substrate. PduT is a novel, tandem domain shell protein that assembles as a pseudohexameric homotrimer. Its structure reveals an unexpected site for binding an [Fe-S] cluster at the center of the PduT pore. The location of a metal redox cofactor in the pore of a shell protein suggests a novel mechanism for either transferring redox equivalents across the shell or for regenerating luminal [Fe-S] clusters.


Upper-body power as measured by medicine-ball throw distance and its relationship to class level among 10- and 11-year-old female participants in club gymnastics.
The purpose of this study was to determine whether or not a relationship existed between upper-body power and class level among female club gymnasts. Sixty female gymnasts between the ages of 10 and 11 and between class levels 5 and 8 participated in the study. The distance of a medicine-ball throw was used to measure upper-body power. Three types of throws--overhead forward throw, overhead backward throw, and chest pass--were performed with a 6-lb rubber medicine ball. The mean distances of 2 trails were calculated and categorized into age group and class level. An analysis of variance design was used to determine the relationship between mean throw distances and throw type, age, and class level. No significant differences were found between mean throw distances and throw type, age, or class level. The results of this study show no relationship between upper-body power of female gymnasts and throw type, age, and class level.


Appropriateness of antibiotic usage for gastrointestinal disorders in a tertiary care hospital.
To assess antibiotic usage in gastrointestinal disorders with respect to appropriateness, pattern of resistance, and incidence of adverse drug reactions (ADRs). Antibiotic prescribing in the gastroenterology department of a tertiary care hospital was evaluated using the Gyssens criteria and also by assessing drug related problems (DRPs) using the Pharmaceutical Care Network Europe V.6.2. A total of 173 patients were studied prospectively by a team of clinical pharmacists. Antibiotic susceptibility was prospectively studied; in addition, retrospective data on culture and sensitivity reports of commonly isolated organisms from 1 October 2012 to 30 September 2014 were collected to determine the resistance pattern in previous years. ADRs were evaluated using the Naranjo scale. Antibiotic therapy was appropriate in 60% of patients and inappropriate in the remaining patients due to incorrect decision, choice, and use. A total of 184 DRPs and 30 ADRs of antibiotics were identified. In the study patients, the most commonly isolated organism was Escherichia coli (27.3%) followed by Klebsiella pneumoniae (16.7%). Both E coli and K pneumoniae exhibited 100% resistance towards cefotaxime. There was an increase in the resistance of E coli and K pneumoniae against various antibiotics tested in 2013-2014 as compared to the previous year. An empirical antibiotic policy was developed which was endorsed by the gastroenterology department. Although antibiotic therapy was appropriate in the majority of patients, irrational use occurred due to incorrect choice, improper dosage, and improper duration of therapy. E coli and K pneumoniae isolates showed an increase in resistance towards various antibiotics tested.


An analysis of current pulmonary embolism therapy.
The clinical diagnosis of venous thromboembolism, and especially pulmonary embolism, is inaccurate due to the nonspecific nature of presenting symptoms and signs which overlap with other frequently coexistent conditions. In recent years, the operating characteristics of more objective diagnostic strategies (ie, ventilation/perfusion lung scan, impedance plethysmography, venous compression ultrasound) have been more clearly defined. However, data regarding the natural history of pulmonary embolism primarily come from an era when objective diagnostic tests were unavailable which calls into question the validity of such data. Furthermore, despite a long experience there are few data regarding the clinical course of pulmonary embolism as currently treated with heparin/warfarin anticoagulation. Acute mortality correlates with the presence of systemic hypotension regardless of embolus size, suggesting that right heart functional reserve is the major determinant of acute survival. Long-term mortality has been attributed to underlying co-morbid disease. However, there are no studies of age- and sex-specific survival which control for co-morbid disease. Pulmonary embolism may be a marker for severe and imminently fatal co-morbid disease, or pulmonary embolism may add "excess mortality" over and above that co-morbid disease. Thrombolytic therapy clears pulmonary artery thrombus more rapidly than heparin and can rapidly restore normal right heart function. Current research aims to identify the ideal plasminogen activator and dosing regimen which maximizes thrombolysis and minimizes disturbance of hemostasis. In addition, more data regarding the natural history and clinical course of pulmonary embolism is needed to stratify patients into subsets with better or worse prognosis.(ABSTRACT TRUNCATED AT 250 WORDS)


Modularity of intrinsic disorder in the human proteome.
Predicting regions of disorder has become of increasing interest when determining protein structure and function. With approximately 33% of eukaryotic proteins having significant disordered regions, and an increasing occurrence of disorder in higher organisms, an analysis of the importance of disorder from an evolutionary perspective was clearly warranted. Focusing on the human proteome, we have studied how abundant disorder is and its relevance to protein function and structure. We have shown that disordered regions frequently appear to be independent functional units, and judged by complete association to certain protein domains, may be evolutionarily conserved. Our work also supports previous analyses on association between disorder and alternate splicing and provides support for the modularity of disorder by showing that with respect to splicing events, disordered regions frequently appear to be spliced as whole units.


Far-field optical nanoscopy.
In 1873, Ernst Abbe discovered what was to become a well-known paradigm: the inability of a lens-based optical microscope to discern details that are closer together than half of the wavelength of light. However, for its most popular imaging mode, fluorescence microscopy, the diffraction barrier is crumbling. Here, I discuss the physical concepts that have pushed fluorescence microscopy to the nanoscale, once the prerogative of electron and scanning probe microscopes. Initial applications indicate that emergent far-field optical nanoscopy will have a strong impact in the life sciences and in other areas benefiting from nanoscale visualization.


Employer contribution and premium growth in health insurance.
We study whether employer premium contribution schemes could impact the pricing behavior of health plans and contribute to rising premiums. Using 1991-2011 data before and after a 1999 premium subsidy policy change in the Federal Employees Health Benefits Program (FEHBP), we find that the employer premium contribution scheme has a differential impact on health plan pricing based on two market incentives: 1) consumers are less price sensitive when they only need to pay part of the premium increase, and 2) each health plan has an incentive to increase the employer's premium contribution to that plan. Both incentives are found to contribute to premium growth. Counterfactual simulation shows that average premium would have been 10% less than observed and the federal government would have saved 15% per year on its premium contribution had the subsidy policy change not occurred in the FEHBP. We discuss the potential of similar incentives in other government-subsidized insurance systems such as the Medicare Part D and the Health Insurance Marketplace under the Affordable Care Act.


Myotome adaptability confers developmental robustness to somitic myogenesis in response to fibre number alteration.
Balancing the number of stem cells and their progeny is crucial for tissue development and repair. Here we examine how cell numbers and overall muscle size are tightly regulated during zebrafish somitic muscle development. Muscle stem/precursor cell (MPCs) expressing Pax7 are initially located in the dermomyotome (DM) external cell layer, adopt a highly stereotypical distribution and thereafter a proportion of MPCs migrate into the myotome. Regional variations in the proliferation and terminal differentiation of MPCs contribute to growth of the myotome. To probe the robustness of muscle size control and spatiotemporal regulation of MPCs, we compared the behaviour of wild type (wt) MPCs with those in mutant zebrafish that lack the muscle regulatory factor Myod. Myodfh261 mutants form one third fewer multinucleate fast muscle fibres than wt and show a significant expansion of the Pax7+ MPC population in the DM. Subsequently, myodfh261 mutant fibres generate more cytoplasm per nucleus, leading to recovery of muscle bulk. In addition, relative to wt siblings, there is an increased number of MPCs in myodfh261 mutants and these migrate prematurely into the myotome, differentiate and contribute to the hypertrophy of existing fibres. Thus, homeostatic reduction of the excess MPCs returns their number to normal levels, but fibre numbers remain low. The GSK3 antagonist BIO prevents MPC migration into the deep myotome, suggesting that canonical Wnt pathway activation maintains the DM in zebrafish, as in amniotes. BIO does not, however, block recovery of the myodfh261 mutant myotome, indicating that homeostasis acts on fibre intrinsic growth to maintain muscle bulk. The findings suggest the existence of a critical window for early fast fibre formation followed by a period in which homeostatic mechanisms regulate myotome growth by controlling fibre size. The feedback controls we reveal in muscle help explain the extremely precise grading of myotome size along the body axis irrespective of fish size, nutrition and genetic variation and may form a paradigm for wider matching of organ size.


Regulation of CXCR4-mediated invasion by DARPP-32 in gastric cancer cells.
Although Dopamine and cAMP-regulated phosphoprotein, Mr 32000 (DARPP-32) is overexpressed in two-thirds of gastric cancers, its impact on molecular functions has not been fully characterized. In this study, we examined the role of DARPP-32 in gastric cancer cell invasion. Using matrigel-coated Boyden chamber invasion assay, DARPP-32-overexpressing AGS cells showed a three-fold increase in invasion relative to the vector control (P < 0.01). We also tested the transendothelial cell invasion as a measure of cell aggressiveness using the impedance-based human umbilical vein endothelial cells invasion assay and obtained similar results (P < 0.001). Western blot analysis indicated that overexpression of DARPP-32 mediated an increase in the membrane-type 1 matrix metalloproteinase (MT1-MMP) and CXCR4 protein levels. Consistent with the role of MT1-MMP in cleaving extracellular matrix proteins initiating the activation of soluble MMPs, we detected a robust increase in MMP-2 activity in DARPP-32-overexpressing cells. The knockdown of endogenous DARPP-32 in the MKN-45 cells reversed these signaling events and decreased cell invasive activity. We tested whether the invasive activity mediated by DARPP-32 might involve sustained signaling via CXCR4-dependent activation of the MT1-MMP/MMP-2 pathway. The small-molecule CXCR4 antagonist (AMD3100) and CXCR4-siRNA blocked DARPP-32-induced cell invasion. We further examined our hypothesis that DARPP-32 could interact with CXCR4 and stabilize its levels following stimulation with its ligand, CXCL12. Using reciprocal coimmunoprecipitation and immunofluorescence experiments, we found that DARPP-32 and CXCR4 coexist in the same protein complex. DARPP-32 prolonged the CXCR4 protein half-life and reduced ubiquitination of the CXCR4 protein, following treatment with its ligand, CXCL12. In conclusion, these findings show a novel mechanism by which DARPP-32 promotes cell invasion by regulating CXCR4-mediated activation of the MT1-MMP/MMP-2 pathway.


CBSX3-Trxo-2 regulates ROS generation of mitochondrial complex II (succinate dehydrogenase) in Arabidopsis.
Despite being toxic at a high concentrations, reactive oxygen species (ROS) play a pivotal role as signaling molecules in responses to stress and regulation of plant development. The mitochondrial electron transport chain (ETC) is the major source of ROS in cells. Although the regulation of ROS in mitochondria has been well elucidated, the protein-protein interaction-based regulation of ETC members has not been well elucidated. In this study, we identified a CBS domain-containing protein, CBSX3, and found that CBSX3 activates o-type thioredoxin (Trx-o2) in mitochondria. In addition, we found that Trx-o2 interacts with SDH1, a subunit of ETC complex II. Knockdown (KD) of CBSX3 revealed anther indehiscence due to deficient lignin deposition caused by insufficient ROS accumulation, and increased expression of genes related to cell cycle and accelerated plant growth. However, in the CBSX3-overexpression plants, ROS accumulation increased, and cell cycle-related gene expression decreased, and thereby plant growth was retarded and leaf size decreased. Moreover, KD of CBSX3 and Trx-o2 conferred resistance to mitochondria ETC inhibitors in terms of ROS release. Taken together, we suggest that CBSX3-Trx-o2 is a ROS generation regulator of mitochondria in plants and plays an important role in regulating plant development and the redox system.


Activation-induced death by apoptosis in CD4+ T cells from human immunodeficiency virus-infected asymptomatic individuals.
In immature thymocytes, T cell receptor for antigen (TCR) mobilization leads to an active T cell suicide process, apoptosis, which is involved in the selection of the T cell repertoire. We have proposed that inappropriate induction of such a cell death program in the mature CD4+ T cell population could account for both early qualitative and late quantitative CD4+ T lymphocyte defects of human immunodeficiency virus (HIV)-infected individuals (Ameisen, J.C., and A. Capron. 1991. Immunol. Today. 4:102). Here, we report that the selective failure of CD4+ T cells from 59 clinically asymptomatic HIV-infected individuals to proliferate in vitro to TCR mobilization by major histocompatibility complex class II-dependent superantigens and to pokeweed mitogen (PWM) is due to an active CD4+ T cell death process, with the biochemical and ultrastructural features of apoptosis. Activation-induced cell death occurred only in the CD4+ T cell population from HIV-infected asymptomatic individuals and was not observed in T cells from any of 58 HIV-seronegative controls, including nine patients with other acute or chronic infectious diseases. Activation-induced CD4+ T cell death was prevented by cycloheximide, cyclosporin A, and a CD28 monoclonal antibody (mAb). The CD28 mAb not only prevented apoptosis but also restored T cell proliferation to stimuli, including PWM, superantigens, and the tetanus and influenza recall antigens. These findings may have implications for the understanding of the pathogenesis of acquired immune deficiency syndrome and for the design of specific therapeutic strategies.


Developing health website quality assessment guidelines for the voluntary sector: outcomes from the Judge Project.
This paper describes the Judge project, the aim of which was to explore the need for health website quality assessment guidelines for the voluntary sector. Such guidelines would enable health consumers to assess the quality of health sites and assist support groups to produce their own good quality sites. The project was a partnership between the Information Management Research Institute, School of Informatics, Northumbria University and Contact a Family. It was supported by the Health Foundation. The views of health consumers and support group members and workers were obtained by focus groups (35 attendees) and questionnaires (55 responses). They were asked questions about quality issues and concerns about Internet health information and any help they needed with judging the quality of health websites. The results supported the need for guidelines. The guidelines were written to reflect the requirements of health consumers and support groups articulated from the focus group and questionnaire data. They were then disseminated via a website. There is a need to improve communication and information exchange between health consumers and professionals and official organizations; publicise support groups to patients and carers; set up local Webs of Trust, linking together voluntary and statutory organizations within a locality. Professionals and official organizations in the public sector should make information provision to patients and health consumers a priority. Information provision should be embedded in the clinical setting via guidelines, protocols and administrative structures. Professionals should be trained in the role of information and how to provide it in appropriate ways. Support groups should use the Judge guidelines to assist them in producing good quality websites. Support groups should promote the Judge guidelines to health consumers, to help them make their own quality judgement about health-information websites.


Amylin: localization, effects on cerebral arteries and on local cerebral blood flow in the cat.
Amylin and adrenomedullin are two peptides structurally related to calcitonin gene-related peptide (CGRP). We studied the occurrence of amylin in trigeminal ganglia and cerebral blood vessels of the cat with immunocytochemistry and evaluated the role of amylin and adrenomedullin in the cerebral circulation by in vitro and in vivo pharmacology. Immunocytochemistry revealed that numerous nerve cell bodies in the trigeminal ganglion contained CGRP immunoreactivity (-ir); some of these also expressed amylin-ir but none adrenomedullin-ir. There were numerous nerve fibres surrounding cerebral blood vessels that contained CGRP-ir. Occasional fibres contained amylin-ir while we observed no adrenomedullin-ir in the vessel walls. With RT-PCR and Real-Time-PCR we revealed the presence of mRNA for calcitonin receptor-like receptor (CLRL) and receptor-activity-modifying proteins (RAMPs) in cat cerebral arteries. In vitro studies revealed that amylin, adrenomedullin, and CGRP relaxed ring segments of the cat middle cerebral artery. CGRP and amylin caused concentration-dependent relaxations at low concentrations of PGF 2alpha-precontracted segment (with or without endothelium) whereas only at high concentration did adrenomedullin cause relaxation. CGRP8-37 blocked the CGRP and amylin induced relaxations in a parallel fashion. In vivo studies of amylin, adrenomedullin, and CGRP showed a brisk reproducible increase in local cerebral blood flow as examined using laser Doppler flowmetry applied to the cerebral cortex of the alpha-chloralose-anesthetized cat. The responses to amylin and CGRP were blocked by CGRP8-37. The studies suggest that there is a functional sub-set of amylin-containing trigeminal neurons which probably act via CGRP receptors.


Alternative translation of human fibroblast growth factor 2 mRNA occurs by internal entry of ribosomes.
Alternative initiations of translation of the human fibroblast growth factor 2 (FGF-2) mRNA, at three CUG start codons and one AUG start codon, result in the synthesis of four isoforms of FGF-2. This process has important consequences on the fate of FGF-2: the CUG-initiated products are nuclear and their constitutive expression is able to induce cell immortalization, whereas the AUG-initiated product, mostly cytoplasmic, can generate cell transformation. Thus, the different isoforms probably have distinct targets in the cell. We show here that translation initiation of the FGF-2 mRNA breaks the rule of the cap-dependent ribosome scanning mechanism. First, translation of the FGF-2 mRNA was shown to be cap independent in vitro. This cap-independent translation required a sequence located between nucleotides (nt) 192 and 256 from the 5' end of the 318-nt-long 5' untranslated region. Second, expression of bicistronic vectors in COS-7 cells indicated that the FGF-2 mRNA is translated through a process of internal ribosome entry mediated by the mRNA leader sequence. By introducing additional AUG codons into the RNA leader sequence, we localized an internal ribosome entry site to between nt 154 and 318 of the 5' untranslated region, just upstream of the first CUG. The presence of an internal ribosome entry site in the FGF-2 mRNA suggests that the process of internal translation initiation, by controlling the expression of a growth factor, could have a crucial role in the control of cell proliferation and differentiation.


Effects of the cyclin-dependent kinase inhibitor CYC202 (R-roscovitine) on the physiology of cultured human keratinocytes.
CYC202 (R-roscovitine) is a potent cyclin-dependent kinase inhibitor, investigated as a potential anti-cancer agent. The knowledge of the action of this pharmacological agent on normal human cells is still limited. In this study, we have explored the effects of the cyclin-dependent kinase inhibitor CYC202 on normal human epidermal keratinocytes. The loss of cell viability induced by this compound was strongly dependent on the rate of keratinocyte proliferation. At slightly cytotoxic doses, CYC202 inhibited the proliferation of subconfluent keratinocytes in a dose-dependent manner, and at higher concentrations induction of early apoptosis was observed, evidenced by caspase-3 activation. The signal transduction pathways in subconfluent keratinocytes were altered, as CYC202 increased the phosphorylation of p38 MAP kinase. The activation of this kinase was confirmed by the increased phosphorylation of p38 MAPK substrate, the small heat shock protein HSP27. Prolonged inhibition of highly proliferative cells with CYC202 for 48 and 72 h altered the expression of epidermal differentiation markers. The use of the selective p38 kinase inhibitor PD169316 demonstrated that involucrin mRNA was upregulated by CYC202 via p38 MAPK pathway. These effects were strongly dependent on cell density and were observed only in highly proliferative keratinocytes. We concluded that CYC202 although highly potent against cancer cells inhibits also the proliferation and induces early apoptotic events in autocrine culture of normal human keratinocytes, activates p38 MAP kinase pathway and alters the expression of the epidermal differentiation markers. These results suggest that despite this potency against tumour cells, CYC202 must be used attentively in the clinical practice.


Validation of sick leave measures: self-reported sick leave and sickness benefit data from a Danish national register compared to multiple workplace-registered sick leave spells in a Danish municipality.
Previous validation studies of sick leave measures have focused on self-reports. Register-based sick leave data are considered to be valid; however methodological problems may be associated with such data. A Danish national register on sickness benefit (DREAM) has been widely used in sick leave research. On the basis of sick leave records from 3,554 and 2,311 eldercare workers in 14 different workplaces, the aim of this study was to: 1) validate registered sickness benefit data from DREAM against workplace-registered sick leave spells of at least 15 days; 2) validate self-reported sick leave days during one year against workplace-registered sick leave. Agreement between workplace-registered sick leave and DREAM-registered sickness benefit was reported as sensitivities, specificities and positive predictive values. A receiver-operating characteristic curve and a Bland-Altman plot were used to study the concordance with sick leave duration of the first spell. By means of an analysis of agreement between self-reported and workplace-registered sick leave sensitivity and specificity was calculated. Ninety-five percent confidence intervals (95% CI) were used. The probability that registered DREAM data on sickness benefit agrees with workplace-registered sick leave of at least 15 days was 96.7% (95% CI: 95.6-97.6). Specificity was close to 100% (95% CI: 98.3-100). The registered DREAM data on sickness benefit overestimated the duration of sick leave spells by an average of 1.4 (SD: 3.9) weeks. Separate analysis on pregnancy-related sick leave revealed a maximum sensitivity of 20% (95% CI: 4.3-48.1).The sensitivity of self-reporting at least one or at least 56 sick leave day/s was 94.5 (95% CI: 93.4 - 95.5) % and 58.5 (95% CI: 51.1 - 65.6) % respectively. The corresponding specificities were 85.3 (95% CI: 81.4 - 88.6) % and 98.9 (95% CI: 98.3 - 99.3) %. The DREAM register offered valid measures of sick leave spells of at least 15 days among eldercare employees. Pregnancy-related sick leave should be excluded in studies planning to use DREAM data on sickness benefit. Self-reported sick leave became more imprecise when number of absence days increased, but the sensitivity and specificity were acceptable for lengths not exceeding one week.


Serum immunoglobulin G antibody subclass response to respiratory syncytial virus F and G glycoproteins after first, second, and third infections.
Serum samples from 31 children who experienced two or three infections with respiratory syncytial virus (RSV) in the first four years of life were tested in an enzyme-linked immunosorbent assay to examine the immunoglobulin G (IgG) subclass responses to the RSV F and G surface glycoproteins associated with primary infection and reinfection. We sought to determine whether the greater degree of glycosylation of the G glycoprotein was reflected in an IgG subclass immune response more like that to a polysaccharide antigen than to a protein antigen. We found that the IgG1/IgG2 ratio of postinfection antibody titers to F was fourfold higher than that to the G glycoprotein after RSV infections 1, 2, and 3. The IgG2 response to the heavily glycosylated G glycoprotein differed from that to a polysaccharide antigen in that the IgG1/IgG2 ratio remained constant with age, whereas the response to a polysaccharide antigen decreased as the IgG2 response increased with age. We also noted that antibody responses to both surface glycoproteins in the IgG1 and IgG2 subclasses reached their maximum levels after RSV infection 2.


A scalable method for O-antigen purification applied to various Salmonella serovars.
The surface lipopolysaccharide of gram-negative bacteria is both a virulence factor and a B cell antigen. Antibodies against O-antigen of lipopolysaccharide may confer protection against infection, and O-antigen conjugates have been designed against multiple pathogens. Here, we describe a simplified methodology for extraction and purification of the O-antigen core portion of Salmonella lipopolysaccharide, suitable for large-scale production. Lipopolysaccharide extraction and delipidation are performed by acetic acid hydrolysis of whole bacterial culture and can take place directly in a bioreactor, without previous isolation and inactivation of bacteria. Further O-antigen core purification consists of rapid filtration and precipitation steps, without using enzymes or hazardous chemicals. The process was successfully applied to various Salmonella enterica serovars (Paratyphi A, Typhimurium, and Enteritidis), obtaining good yields of high-quality material, suitable for conjugate vaccine preparations.


Rapid generation of markerless recombinant MVA vaccines by en passant recombineering of a self-excising bacterial artificial chromosome.
The non-replicating poxviral vector modified vaccinia virus Ankara (MVA) is currently a leading candidate for development of novel recombinant vaccines against globally important diseases. The 1980s technology for making recombinant MVA (and other poxviruses) is powerful and robust, but relies on rare recombination events in poxviral-infected cells. In the 21st century, it has become possible to apply bacterial artificial chromosome (BAC) technology to poxviruses, as first demonstrated by B. Moss' lab in 2002 for vaccinia virus. A similar BAC clone of MVA was subsequently derived, but while recombination-mediated genetic engineering for rapid production was used of deletion mutants, an alternative method was required for efficient insertion of transgenes. Furthermore "markerless" viruses, which carry no trace of the selectable marker used for their isolation, are increasingly required for clinical trials, and the viruses derived via the new method contained the BAC sequence in their genomic DNA. Two methods are adapted to MVA-BAC to provide more rapid generation of markerless recombinants in weeks rather than months. "En passant" recombineering is applied to the insertion of a transgene expression cassette and the removal of the selectable marker in bacteria; and a self-excising variant of MVA-BAC is constructed, in which the BAC cassette region is rapidly and efficiently lost from the viral genome following rescue of the BAC into infectious virus. These methods greatly facilitate and accelerate production of recombinant MVA, including markerless constructs.


Intravenous microinjections of zebrafish larvae to study acute kidney injury.
In this video article we describe a zebrafish model of AKI using gentamicin as the nephrotoxicant. The technique consists of intravenous microinjections on 2 dpf zebrafish. This technique represents an efficient and rapid method to deliver soluble substances into the bloodstream of zebrafish larvae, allowing for the injection of 15-20 fish per hour. In addition to AKI studies, this microinjection technique can also be used for other types of experimental studies such as angiography. We provide a detailed protocol of the technique from equipment required to visual measures of decreased kidney function. In addition, we also demonstrate the process of fixation, whole mount immunohistochemistry with a kidney tubule marker, plastic embedding and sectioning of the larval zebrafish. We demonstrate that zebrafish larvae injected with gentamicin show morphological features consistent with AKI: edema, loss of cell polarity in proximal tubular epithelial cells, and morphological disruption of the tubule.


Capsaicin cough receptor sensitivity test in children.
Capsaicin has been used as a tussive agent in studies in adults to determine cough receptor sensitivity. The aim of this study was to determine the tolerance, repeatability and influence of inspiratory flow on the capsaicin cough receptor sensitivity test in children. Thirty children (mean age 11 yrs; range 6-16 yrs) were tested on two different days, to determine the lowest concentration of capsaicin required to stimulate two or more coughs (cough threshold (Cth)), 2-4 coughs (C2), and five or more coughs (C5). Capsaicin was nebulized through a dosimeter, with an arrangement that allowed the subjects to visualize and regulate their inspiratory flow. Using a constant inspiratory flow of 20 L x min(-1), tests were reproducible for C2, C5 and Cth (doubling dose changes of 1.13, 1.03 and 1.08, respectively). An increase in the inspiratory flow from 20 to 60 L x min(-1) significantly increased C2 (19.5 to 46.8 microM; p=0.016) and C5 (46.8 to 128.8 microM; p=0.008). We conclude that in children, the capsaicin cough challenge test: 1) is well-tolerated; 2) is highly repeatable; and that 3) the inspiratory flow significantly influences cough receptor sensitivity and repeatability of the test and should, thus, be regulated.


The impact of price promotions on confectionery and snacks on the energy content of shopping baskets: A randomised controlled trial in an experimental online supermarket.
Overconsumption of foods high in fat, sugars, and salt (HFSS) poses a significant risk to health. The government in England has passed legislation that would limit some price promotions of HFSS foods within supermarkets, but evidence regarding likely impacts of these policies, especially in online settings, is limited. This study aimed to determine whether there were any differences in the energy and nutrient content of shopping baskets after removing promotions on HFSS foods in an online experimental supermarket. UK adults (n = 511) were asked to select food from four categories with a £10 budget in an online experimental supermarket: confectionery; biscuits and crackers; crisps, nuts and snacking fruit; cakes and tarts. They were randomly allocated to one of two trial arms: (1) promotions present (matched to promotion frequency seen in a major UK retailer) (n = 257), or (2) all promotions removed from all products within the target food categories (n = 254). The primary outcome analysis used linear regression to compare total energy (kcal) of items placed in shopping baskets when promotions were present vs. absent, while secondary analyses investigated differences in nutrients and energy purchased from individual food categories. Mean energy in food selected without promotions was 5156 kcal per basket (SD 1620), compared to 5536 kcal (SD 1819) with promotions, a difference of -552kcal (95%CIs: -866, -238), equivalent to 10%. There were no significant differences in energy purchased for any individual category between groups. No evidence was found of other changes in nutritional composition of baskets or of significant interactions between the impact of promotions and participant characteristics (gender, age, ethnicity) on energy purchased. Removing promotions on HFSS foods resulted in significantly less total energy selected in an online experimental supermarket study.


A Full Quantum Mechanical Approach Assessing the Chemical and Electromagnetic Effect in TERS.
Tip-enhanced Raman spectroscopy (TERS) is a valuable method for surface analysis with nanometer to angstrom-scale resolution; however, the accurate simulation of particular TERS signals remains a computational challenge. We approach this challenge by combining the two main contributors to plasmon-enhanced Raman spectroscopy and to the high resolution in TERS, in particular, the electromagnetic and the chemical effect, into one quantum mechanical simulation. The electromagnetic effect describes the sample's interaction with the strong, highly localized, and inhomogeneous electric fields associated with the plasmonic tip and is typically the thematic focus for most mechanistic studies. On the other hand, the chemical effect covers the different responses to the extremely close-range and highly position-sensitive chemical interaction between the apex tip atom(s) and the sample, and, as we could show in previous works, plays an often underestimated role. Starting from a (time-dependent) density functional theory description of the chemical model system, comprised of a tin(II) phthalocyanine sample molecule and a single silver atom as the tip, we introduce the electromagnetic effect through a series of static point charges that recreate the electric field in the vicinity of the plasmonic Ag nanoparticle. By scanning the tip over the molecule along a 3D grid, we can investigate the system's Raman response on each position for nonresonant and resonant illumination. Simulating both effects on their own already hints at the achievable signal enhancement and resolution, but the combination of both creates even stronger evidence that TERS is capable of resolving submolecular features.


Formaldehyde catabolism is essential in cells deficient for the Fanconi anemia DNA-repair pathway.
Metabolism is predicted to generate formaldehyde, a toxic, simple, reactive aldehyde that can damage DNA. Here we report a synthetic lethal interaction in avian cells between ADH5, encoding the main formaldehyde-detoxifying enzyme, and the Fanconi anemia (FA) DNA-repair pathway. These results define a fundamental role for the combined action of formaldehyde catabolism and DNA cross-link repair in vertebrate cell survival.


Excess early mortality in schizophrenia.
Schizophrenia is often referred to as one of the most severe mental disorders, primarily because of the very high mortality rates of those with the disorder. This article reviews the literature on excess early mortality in persons with schizophrenia and suggests reasons for the high mortality as well as possible ways to reduce it. Persons with schizophrenia have an exceptionally short life expectancy. High mortality is found in all age groups, resulting in a life expectancy of approximately 20 years below that of the general population. Evidence suggests that persons with schizophrenia may not have seen the same improvement in life expectancy as the general population during the past decades. Thus, the mortality gap not only persists but may actually have increased. The most urgent research agenda concerns primary candidates for modifiable risk factors contributing to this excess mortality, i.e., side effects of treatment and lifestyle factors, as well as sufficient prevention and treatment of physical comorbidity.


Withholding or termination of resuscitation in pediatric out-of-hospital traumatic cardiopulmonary arrest.
This multiorganizational literature review was undertaken to provide an evidence base for determining whether recommendations for out-of-hospital termination of resuscitation could be made for children who are victims of traumatic cardiopulmonary arrest. Although there is increasing acceptance of out-of-hospital termination of resuscitation for adult traumatic cardiopulmonary arrest when there is no expectation of a good outcome, children are routinely excluded from state termination-of-resuscitation protocols. The decision to withhold resuscitative efforts in a child under specific circumstances (decapitation or dependent lividity, rigor mortis, etc) is reasonable. If there is any doubt as to the circumstances or timing of the traumatic cardiopulmonary arrest, under the current status of limiting termination of resuscitation in the field to persons older than 18 years in most states, resuscitation should be initiated and continued until arrival to the appropriate facility. If the patient has arrested, resuscitation has already exceeded 30 minutes, and the nearest facility is more than 30 minutes away, involvement of parents and family of these children in the decision-making process with assistance and guidance from medical professionals should be considered as part of an emphasis on family-centered care because the evidence suggests that either death or a poor outcome is inevitable.


Computed Tomography and Shifts to Alternate Imaging Modalities in Hospitalized Children.
Many studies have demonstrated a rise in computed tomography (CT) utilization in abstract children's hospitals. However, CT utilization may be declining, perhaps due to awareness of potential hazards of pediatric ionizing radiation, such as increased risk of malignancy. Th e objective is to assess the trend in CT utilization in hospitalized children at freestanding children's hospitals from 2004 to 2012 and we hypothesize decreases are associated with shifts to alternate imaging modalities. Multicenter cross-sectional study of children admitted to 33 pediatric tertiary-care hospitals participating in the Pediatric Health Information System between January 1, 2004, and December 31, 2012. The rates of CT, ultrasound, and MRI for the top 10 All-Patient Refined Diagnosis Related Groups (APR-DRGs) for which CT was performed in 2004 were determined by billing data. Rates of each imaging modality for those top 10 APR-DRGs were followed through the study period. Odds ratios of imaging were adjusted for demographics and illness severity. For all included APR-DRGs except ventricular shunt procedures and nonbacterial gastroenteritis, the number of children imaged with any modality increased. CT utilization decreased for all APR-DRGs (P values , .001). For each of the APR-DRGs except seizure and infections of upper respiratory tract, the decrease in CT was associated with a significant rise in an alternative imaging modality (P values # .005). For the 10 most common APR-DRGs for which children received CT in 2004,a decrease in CT utilization was found in 2012. Alternative imaging modalities for 8 of the diagnoses were used.


Human diseases versus mouse models: insights into the regulation of genomic imprinting at the human 11p15/mouse distal chromosome 7 region.
The 11p15 region is organised into two independent imprinted domains controlled by imprinting control regions, which carry opposite germline imprints. Dysregulation of 11p15 genomic imprinting results in two human fetal growth disorders (Silver-Russell syndrome (SRS, MIM 180860) and Beckwith-Wiedemann syndrome (BWS, MIM 130650)) with opposite growth phenotypes. The mouse orthologous region on distal chromosome 7 (dist7) is well conserved in its organisation and its regulation. Targeted mutagenesis in mice has provided highly valuable clues in terms of the mechanisms involved in the regulation of genomic imprinting of the 11p15/dist7 imprinted region. On the other hand, the recent identification of unexpected genetic defects in BWS and SRS patients also brought new insights into the mechanisms of 11p15 imprinting regulation. However, some mouse models and human genetic defects show contradictions in term of growth phenotypes and parental transmission. In this review, we extensively analyse those various mouse and human models and more particularly models with mutations affecting the two imprinting centres, in order to improve our understanding of regulation of 11p15/dist7 genomic imprinting.


Adenosine receptor antagonists and retinal neovascularization in vivo.
The role of adenosine receptor (AdoR) antagonists in human retinal endothelial cell function in vitro has previously been determined. In this study, efficacy of AdoR antagonist administration in reducing retinal neovascularization was examined in a mouse pup model of oxygen-induced retinopathy. A previously described model of oxygen-induced retinal neovascularization in newborn mouse pups was used to examine the effect of various AdoR antagonists on neovascularization. The nonselective AdoR antagonist xanthine amine congener (XAC), the A(2A)-selective antagonist ZM241385, the A(2B)-selective antagonists 3-N-propylxanthine (enprofylline) and 3-isobutyl-8-pyrrolidinoxanthine (IPDX), and the A(1)-selective antagonist cyclopentyl-1,3-dipropylxanthine (CPX) were used. After the hyperoxia exposure the animals received daily intraperitoneal injections of pharmacologically relevant doses of AdoR antagonists for 5 days. Control animals received vehicle (0.1% dimethyl sulfoxide [DMSO]) alone. The animals were then killed and perfused with fluorescein-dextran. Wholemounts of retinas from one eye were prepared and examined, whereas the retinas of the contralateral eye were embedded, sectioned, and stained for counting neovascular nuclei extending beyond the internal limiting membrane into the vitreous. Angiography of wholemount retinas showed reduction of neovascular tufts in animals treated with selective A(2B) AdoR antagonists. Quantification of the extraretinal neovascular nuclei showed that only animals treated with XAC, enprofylline, or IPDX showed a significant reduction in retinal neovascularization. By contrast, neither CPX nor ZM241385 had an effect on neovascularization. The A(2B)-selective AdoR antagonists inhibited oxygen-induced retinal neovascularization in vivo and may provide a basis for developing pharmacologic therapies for the treatment of proliferative retinopathies.


A cord blood monocyte-derived cell therapy product accelerates brain remyelination.
Microglia and monocytes play important roles in regulating brain remyelination. We developed DUOC-01, a cell therapy product intended for treatment of demyelinating diseases, from banked human umbilical cord blood (CB) mononuclear cells. Immunodepletion and selection studies demonstrated that DUOC-01 cells are derived from CB CD14+ monocytes. We compared the ability of freshly isolated CB CD14+ monocytes and DUOC-01 cells to accelerate remyelination of the brains of NOD/SCID/IL2Rγnull mice following cuprizone feeding-mediated demyelination. The corpus callosum of mice intracranially injected with DUOC-01 showed enhanced myelination, a higher proportion of fully myelinated axons, decreased gliosis and cellular infiltration, and more proliferating oligodendrocyte lineage cells than those of mice receiving excipient. Uncultured CB CD14+ monocytes also accelerated remyelination, but to a significantly lesser extent than DUOC-01 cells. Microarray analysis, quantitative PCR studies, Western blotting, and flow cytometry demonstrated that expression of factors that promote remyelination including PDGF-AA, stem cell factor, IGF1, MMP9, MMP12, and triggering receptor expressed on myeloid cells 2 were upregulated in DUOC-01 compared to CB CD14+ monocytes. Collectively, our results show that DUOC-01 accelerates brain remyelination by multiple mechanisms and could be beneficial in treating demyelinating conditions.


Global estimate of the prevalence of post-traumatic stress disorder among adults living with HIV: a systematic review and meta-analysis.
Although people living with HIV (PLWH) have been disproportionately affected by post-traumatic stress disorder (PTSD), the global prevalence of PTSD among PLWH is unknown. This study aimed to systematically review the prevalence of PTSD among PLWH worldwide and explore variation in prevalence across sociodemographic and methodological factors. A meta-analysis using a random-effects model was conducted to pool the prevalence estimated from individual studies, and subgroup analyses were used to analyse heterogeneities. Observational studies providing PTSD prevalence data in an adult HIV population were searched from January 2000 to November 2019. Measurements were not restricted, although the definition of PTSD had to align with the Diagnostic and Statistical Manual of Mental Disorders or the International Classification of Diseases diagnostic criteria. A total of 38 articles were included among 2406 records identified initially. The estimated global prevalence of PTSD in PLWH was 28% (95% CI 24% to 33%). Significant heterogeneity was detected in the proportion of PLWH who reported PTSD across studies, which was partially explained by geographic area, population group, measurement and sampling method (p<0.05). PTSD among PLWH is common worldwide. This review highlights that PTSD should be routinely screened for and that more effective prevention strategies and treatment packages targeting PTSD are needed in PLWH.


The consequences of Bacillus axarquiensis Ruiz-García et al. 2005, Bacillus malacitensis Ruiz-García et al. 2005 and Brevibacterium halotolerans Delaporte and Sasson 1967 (Approved Lists 1980) being treated as heterotypic synonyms.
In a recent publication, data was presented supporting Bacillus axarquiensis Ruiz-García et al. 2005, Bacillus malacitensis Ruiz-García et al. 2005 and Brevibacterium halotolerans Delaporte and Sasson 1967 (Approved Lists 1980) being treated as heterotypic synonyms. The nomenclatural consequences proposed were that under these circumstances the correct name to be used is Bacillus axarquiensis Ruiz-García et al. 2005, but this is not consistent with the wording of the International Code of Nomenclature of Prokaryotes and it is, therefore, it is necessary to establish the correct name to be used.


Fluorescein dye as a novel cost-effective approach for staining raw specimens in ophthalmic pathology.
To study the usefulness of sodium fluorescein dye for staining raw specimens in ophthalmic pathology. Laboratory-based observational study. Eye specimens received in the ocular pathology department of a tertiary eye care center in northeast India were included in the study after obtaining the informed consent. The study period was from 2016 to 2019. Specimens received were a corneal button, lid, orbital tissues, enucleated eyeballs, eviscerated eye, explanted intraocular lens (IOLs), optic nerve and ocular parasites. Sections of the gross specimens were stained with sodium fluorescein (C20H12O5NA) dye. The average duration of tissue-stain contact time was 45 s. The sections were analyzed under the compound microscope. The intensity of illumination of the microscope was modulated to obtain high contrast digital photographs. 26 corneal buttons with or without limbal tissue specimens were analyzed with fluorescein staining procedure; limbus with its pigmented cells were seen in the enucleated eyeballs. 33 enucleated eyes (retinoblastoma [RB] (n = 24), phthisical eyes (n = 4), choroidal melanomas (n = 2), and others (n = 3) were included in the study cohort. In these 33 enucleated eyes, vitreous were also examined for the presence of hyalocytes and other pathological cells. Retinal pigment epithelial cells were also seen (n = 11). RB seeds were seen with fluorescein stain and documented in 14 specimens. The RB seeds were mostly in vitreous (n = 9) and subretinal space (n = 5). Fat cells (n = 8) from orbital tissues and sebaceous cells (n = 5) from frozen section specimens were also observed and documented. This study highlights a novel method of rapid staining of gross ophthalmic pathology specimens.


Ethnic differences in incidence of type 1 diabetes among second-generation immigrants and adoptees from abroad.
The incidence of type 1 diabetes shows a large variation worldwide, but whether the causes are environmental or genetic has not been settled. We examine here the incidence of type 1 diabetes among second-generation immigrants and adoptees from abroad to disentangle genetic/ethnic vs. environmental influence, assuming adoptees from abroad have similar environmental exposures compared with the native Swedes, with the only difference in their genetic background. Second-generation immigrants and adoptees from abroad were retrieved from the MIGMED2 database, and they were followed up until the diagnosis of type 1 diabetes, death, or the end of study. Standardized incidence ratios (SIRs) were calculated for type 1 diabetes among these immigrants compared with native Swedes. A total of 1,050,569 children were defined as second-generation immigrants and the overall SIR of type 1 diabetes was significantly decreased. A decreased risk was observed for all countries of origin, with an exception for children with parents from Finland. A total of 51,557 children born in foreign countries were adopted by Swedes. Adoptees from Eastern Europe, Soviet countries, India, Pakistan, Afghanistan, East and Southeast Asia, Chile, and other Central and South American countries had a significantly decreased SIR. The decreased incidence of type 1 diabetes observed in some second-generation immigrants and adoptees from abroad strongly suggests that ethnic genetic heterogeneity could play an important role on type 1 diabetes.


"Becoming myself": how participants in a longitudinal substance use disorder recovery study experienced receiving continuous feedback on their results.
Being a participant in longitudinal follow-up studies is not commonly a factor considered when investigating useful self-change aspects for individuals attempting recovery from substance use disorder (SUD). This study reports on how ongoing monitoring, and feedback on data results in a longitudinal follow-up study of SUD recovery were perceived by individuals who had achieved long-term abstinence and social recovery. Interviewers with first-hand experience with the topic conducted interviews with 30 participants and analysed the data using a thematic analytic approach within an interpretative-phenomenological framework. Analyses resulted in the following themes. 1) Ongoing short text messaging (SMS) monitoring: helped participants by offering recovery milestones and reminders of the past. 2) Feedback on data results helped participants track physical and cognitive recovery: "I am more like myself". 3) Using feedback in treatment: understanding the importance of a functional brain to participants may help with long-term retention in treatment. Self-changes that were challenging to detect on a day-to-day basis were available for reflection through longitudinal study participation, including ongoing monitoring and feedback on the results, allowing personal consolidation of change processes. Clinical services could benefit from continuing development and implementation of such technology for ongoing monitoring and feedback on assessments to motivate self-change in SUD recovery. The development of guidelines for providing the results of research assessments to individuals could help reduce attrition in research projects and support recovery and healthy choices for study participants.