SU-E-T-276: Treatment Planning Strategies for Lung Injury Studies in Rat Models in 6 MV Delivery.
To study planning strategies that can be used in small animal radiation-induced lung toxicity experiments using 6 MV accelerator with high density MLC. Three different types of plans were designed on CT images of a Sprague Dawley rat model to irradiate 50% of the total lung volume (lung divided into apex and base) with a prescription dose of 24 Gy to the partial lung. Two VMAT arc therapy plans were optimized to cover to the prescription dose, either the apex or base of the lung. Two AP- PA plans were designed to completely block either lung apex or base while irradiating the remaining 50% of the lung. Finally, two AP-PA plans were designed to cover, to the prescription dose, the apex or base of the lung. The plans were designed and optimized using the Eclipse AAA algorithm and recalculated using the MMCTP/EGS/Beam Monte Carlo system. When completely blocking the lung base, the apex will be underdosed by up to 30%; when completely covering the apex by the prescribed dose, the base will receive overdosing (V50%=73%). The VMAT plan leads to a more conformal dose distribution and spares unnecessary skin exposure when compared to AP-PA MV or kV delivery. Despite the small size of rat model, the 6 MV VMAT delivery is superior in terms of dose conformality and sparing of the heart and the non-irradiated 50% of the lung compared to the standard, simpler, AP-PA delivery. MC dosimetry in lung shows that the delivered dose is 10% higher than predicted by AAA because of the predominance of small fields in the delivery. Clinical state-of- the-art planning and delivery techniques can be scaled down accurately to rats. The use of these techniques is essential in small animal studies to render conclusions of radiation response investigations translatable to human studies.


Constitutive expression of a chimeric receptor that delivers IL-2/IL-15 signals allows antigen-independent proliferation of CD8+CD44high but not other T cells.
We have prepared transgenic mice whose T cells constitutively express a chimeric receptor combining extracellular human IL-4R and intracellular IL-2Rbeta segments. This receptor can transmit IL-2/IL-15-like signals in response to human, but not mouse, IL-4. We used these animals to explore to what extent functional IL-2R/IL-15R expression controls the capacity of T cells to proliferate in response to IL-2/IL-15-like signals. After activation with Con A, naive transgenic CD8+ and CD4+ T cells respond to human IL-4 as well as to IL-2. Without prior activation, they failed to proliferate in response to human IL-4, although human IL-4 did prolong their survival. Thus, IL-2-induced proliferation of activated T cells requires at least one other Ag-induced change apart from the induction of a functional IL-2R. However, a fraction of CD8+CD44high T cells proliferate in human IL-4 without antigenic stimulation or syngeneic feeder cells. In contrast, CD4+CD44high T cells are not constitutively responsive to human IL-4. We conclude that although all transgenic T cells express a functional chimeric receptor, only some CD8+CD44high T cells contain all molecules required for entry into the cell cycle in response to human IL-4 or IL-15.


The Oncolytic Herpes Simplex Virus Talimogene Laherparepvec Shows Promising Efficacy in Neuroendocrine Cancer Cell Lines.
Metastatic neuroendocrine cancer still constitutes a palliative situation, lacking promising treatment options. Oncolytic virotherapy, a novel type of virus-based immunotherapy, lyses tumor cells using genetically engineered viruses thereby activating the immune system to induce an optimized antitumor response which could bring down tumor masses to a stage of minimal residual tumor disease. The oncolytic vector talimogene laherparepvec (T-VEC, herpes simplex virus [HSV] type 1) has already shown excellent safety profiles in clinical studies and has become the first ever FDA/EMA-approved oncolytic virus (OV). This work presents a first preclinical assessment of this state-of-the-art OV, using a panel of human neuroendocrine tumor/neuroendocrine carcinoma (NET/NEC) cell lines. Cytotoxicity, transgene expression, and viral replication patterns were studied. Furthermore, the antiproliferative activity was compared to the one of mTOR inhibitor Everolimus and also interactions between the OV and Everolimus were evaluated. Moreover, virostatic effects of ganciclovir (GCV) on replication of T-VEC were assessed and electron microscopic pictures were taken to comprehend viral envelopment and details of the replication cycle of T-VEC in human neuroendocrine cancer. It could be shown that T-VEC infects, replicates in, and lyses human NET/NEC cells exhibiting high oncolytic efficiencies already at quite low virus concentrations. Interestingly, Everolimus was not found to have any relevant impact on rates of viral replication, but no additive effects could be proved using a combinatorial therapy regimen. On the other hand, GCV was shown to be able to limit replication of T-VEC, thus establishing an important safety feature for future treatments of NET/NEC patients. Taken together, T-VEC opens up a promising novel treatment option for NET/NEC patients, warranting its further preclinical and clinical development.


A novel automated discontinuous venous blood monitoring system for ex vivo glucose determination in humans.
Intensive insulin therapy reduces mortality and morbidity in critically ill patients but imposes great demands on medical staff who must take frequent blood samples for the determination of glucose levels. A solution to this resourcing problem would be provided by an automated blood monitoring system. The aim of the present clinical study was to evaluate such a system comprising an automatic blood sampling unit linked to a glucose biosensor. Our approach was to determine the correlation and system error of the sampling unit alone and of the combined system with respect to reference levels over 12h in humans. Two venous cannulae were inserted to connect the automatic and reference systems to the subjects. Blood samples were taken at 15 and 30 min intervals. The median Pearson coefficient of correlation between manually and automatically withdrawn blood samples was 0.982 for the sampling unit alone and 0.950 for the complete system. The biosensor had a linear range up to 20 mmoll(-1) and a 95% response time of <2 min. Clark Error Grid analysis showed that 96.93% of the data (228 data pairs) was in zone A and 3.07% in zone B. Insulin Titration Error Grid analysis suggested an acceptable treatment in 99.56% of cases. Implementation of a "Keep Vein Open" saline infusion into the automated blood sampling system reduced blood withdrawal failures through occluded catheters fourfold. In summary, automated blood sampling from a peripheral vein coupled with automatic glucose determination is a promising alternative to frequent manual blood sampling.


Atypical fat necrosis of the breast: the 'mycetoma' appearance.
A very unusual appearance of fat necrosis in the breast is described. The low density/hypo-echoic rim and high density/hyperechoic central part are shown on mammography and ultrasound, respectively. The mass had the appearance of a 'mycetoma' in a cavity in the lung. The radiological diagnosis of fat necrosis was confirmed by histopathology.


Factors associated with exclusive breastfeeding practices among mothers in Goba district, south east Ethiopia: a cross-sectional study.
Exclusive breastfeeding is defined as feeding infants only breast milk, be it directly from breast or expressed, with no addition of any liquid or solids apart from drops or syrups consisting of vitamins, mineral supplements or medicine, and nothing else. Several studies have shown that exclusive breastfeeding for the first six months plays a great role in preventing morbidity and mortality. However, in Ethiopia a large portion of infants are not exclusively breastfed according to the infant feeding recommendations. Understanding the factors that influence exclusive breastfeeding is crucial to promoting the practice. This study was carried out to identify factors predicting exclusive breastfeeding among mothers in Bale Goba district, south east Ethiopia. A community-based cross-sectional study was conducted from March to February 2010 involving both quantitative and qualitative data. A total of 608 mothers were selected randomly. A convenience sampling technique was used to generate the qualitative data. The qualitative data were analyzed using thematic frameworks. A multivariable logistic regression analysis was used to identify independent predictors of exclusive breastfeeding after controlling for background variables. The prevalence of exclusive breastfeeding in the last 24 hours preceding the survey was 71.3%. The median duration of exclusive breastfeeding was three months and mean frequency of breastfeeding was six times per day. Being unemployed [AOR: 10.4 (95% CI: 1.51, 71.50)] and age of infants of less than two months [AOR: 5.6 (95% CI: 2.28, 13.60)] were independently associated with exclusive breastfeeding. A large proportion of infants are not exclusively breastfed during the first 6 months, despite what is recommended in the national and global infant and young child feeding (IYCF) guidelines. Employed mothers were less likely to practice exclusive breastfeeding, implying the need for promoting workplace breastfeeding practices and creating an enabling environment for exclusive breastfeeding. Extensions of maternity leave up to the first six month of child's age to achieve optimal level of exclusive breastfeeding practices should also be looked into as an alternative solution.


Increasing incidence of testicular germ cell tumors among black men in the United States.
There has been marked disparity in the incidence of testicular germ cell tumors (TGCT) among white and black men for a number of decades in the United States. Since at least the beginning of the Surveillance, Epidemiology, and End Results (SEER) Program in 1973, incidence rates among white men have been five times higher than rates among black men. In addition, rates among white men have been increasing, whereas rates among black men have remained stable. However, a recent examination of ethnic-specific rates suggested that the incidence among black men may have begun to change in the 1990s. TGCT incidence data from nine registries of the SEER Program were analyzed for the years 1973 to 2001. Trends were examined separately for seminoma and nonseminoma. Analyses found that the incidence of TGCT began to increase among black men between the 1988 to 1992 and 1993 to 1997 periods. Before that time, incidence among black men had decreased by 14.8%. Between 1988 to 1992 and 1998 to 2001, however, the incidence increased by 100%, with the incidence of seminoma increasing twice as much (124.4%) as the incidence of nonseminoma (64.3%). Over the 29-year time period, there was no evidence of a change in the proportion of tumors diagnosed at earlier stages among black men. In contrast, the proportion of tumors diagnosed at localized stages significantly increased among white men. The incidence of TGCT among black men has increased since 1988 to 1992. Although the reasons for this increase are unclear, screening and earlier diagnosis of TGCT do not seem to be factors.


Fecal microbiota transplantation for recurrent Clostridioides difficile infection in patients with concurrent ulcerative colitis.
Treatment of recurrent Clostridioides difficile infection (rCDI) has emerged as an important management dilemma particularly in patients with underlying inflammatory bowel disease (IBD). Fecal microbiota transplantation (FMT) has been used as a safe and highly effective treatment option for rCDI refractory to standard antibiotic therapies. The aim of this study was to report the efficacy of FMT in Iranian rCDI patients with concurrent IBD. A total of seven consecutive patients with ulcerative colitis (UC) who had experienced 3 episodes of rCDI were enrolled in this study. All patients received at least a single FMT administered during colonoscopy by direct infusion of minimally processed donor stool. Patients were followed for a minimum of 6 months for assessment of treatment efficacy and adverse events (AEs) attributable to FMT. All 7 UC patients (100%) experienced a durable clinical response to a single FMT following 2 months after the procedure. One patient received a second FMT in which a successful resolution of rCDI was ultimately achieved. No serious AEs from FMT were noted. FMT through colonoscopy was a safe, simple and effective alternative treatment approach for rCDI in patients with underlying IBD. However, its use and efficacy should be pursued in long-term prospective controlled trials.


Placenta growth factor-1 exerts time-dependent stabilization of adherens junctions following VEGF-induced vascular permeability.
Increased vascular permeability is an early event characteristic of tissue ischemia and angiogenesis. Although VEGF family members are potent promoters of endothelial permeability the role of placental growth factor (PlGF) is hotly debated. Here we investigated PlGF isoforms 1 and 2 and present in vitro and in vivo evidence that PlGF-1, but not PlGF-2, can inhibit VEGF-induced permeability but only during a critical window post-VEGF exposure. PlGF-1 promotes VE-cadherin expression via the trans-activating Sp1 and Sp3 interaction with the VE-cadherin promoter and subsequently stabilizes transendothelial junctions, but only after activation of endothelial cells by VEGF. PlGF-1 regulates vascular permeability associated with the rapid localization of VE-cadherin to the plasma membrane and dephosphorylation of tyrosine residues that precedes changes observed in claudin 5 tyrosine phosphorylation and membrane localization. The critical window during which PlGF-1 exerts its effect on VEGF-induced permeability highlights the importance of the translational significance of this work in that PLGF-1 likely serves as an endogenous anti-permeability factor whose effectiveness is limited to a precise time point following vascular injury. Clinical approaches that would pattern nature's approach would thus limit treatments to precise intervals following injury and bring attention to use of agents only during therapeutic windows.


A phase III clinical trial of 0.5% levofloxacin ophthalmic solution versus 0.3% ofloxacin ophthalmic solution for the treatment of bacterial conjunctivitis.
To compare the efficacy and safety of 0.5% levofloxacin ophthalmic solution (QUIXIN) with 0.3% ofloxacin ophthalmic solution for the treatment of bacterial conjunctivitis. Prospective, randomized, active-controlled, double-masked, multicenter study. Four hundred twenty-three patients with a clinical diagnosis of bacterial conjunctivitis were enrolled. Patients were randomly assigned to receive either 0.5% levofloxacin (n = 211) or 0.3% ofloxacin (n = 212) for 5 days (every 2 hours on days 1 and 2 and every 4 hours on days 3-5). Conjunctival cultures were obtained, and ocular signs and symptoms were evaluated on day 1 (baseline), days 3 to 5 (interim), and days 6 to 10 (final). End point was defined as the last evaluable observation. Primary microbial and clinical outcomes were based on culture results and resolution of cardinal signs, respectively. Secondary efficacy assessments included evaluations of ocular signs and symptoms. Two hundred eight patients (levofloxacin, n = 109; ofloxacin, n = 99) were evaluated for efficacy. Microbial eradication rates were significantly greater in the 0.5% levofloxacin treatment group compared with the 0.3% ofloxacin group at both the final visit (89% vs. 80%, P = 0.034) and at end point (90% vs. 81%; P = 0.038). Treatment with 0.5% levofloxacin was significantly more effective in resolving photophobia than was 0.3% ofloxacin treatment (94% vs. 73%, P = 0.006). Both study medications were well tolerated, with a low incidence of adverse events. Although clinical cure rates in the 0.5% levofloxacin and 0.3% ofloxacin treatment groups were similar, a 5-day treatment regimen with 0.5% levofloxacin achieved microbial eradication rates that were statistically superior to those attained with 0.3% ofloxacin. Despite the higher concentration of active drug in 0.5% levofloxacin versus 0.3% ofloxacin, there was no difference between treatment groups in the incidence of treatment-related adverse events.


Changes in intracellular free calcium concentration during long exposures to simulated ischemia in isolated mammalian ventricular muscle.
Intracellular free calcium concentration ([Ca2+]i) was measured in isolated ferret ventricular papillary muscles during and after long exposures to ischemia. All experiments were performed at 37 degrees C, and the muscles were stimulated at 1 Hz. Ischemia was simulated by changing from superfusion with oxygenated Tyrode's solution to superfusion with water-saturated gas (95% N2-5% CO2), thus simultaneously stopping oxygenation and restricting the extracellular space. [Ca2+]i was measured with aequorin, which was microinjected into superficial cells of the preparation. Exposure to ischemia caused a complex series of changes in [Ca2+]i. In the first few minutes the changes in [Ca2+]i were variable; however, after approximately 5 minutes all preparations exhibited a progressive increase in amplitude and duration of the stimulated rise in [Ca2+]i (the calcium transient). The amplitude of the calcium transients peaked after approximately 18 minutes of ischemia, when they were 339% of the control value. After this peak, the calcium transients progressively failed to occur in response to stimulation and declined in amplitude; simultaneously, spontaneous oscillations of [Ca2+]i appeared and increased in size and frequency. The oscillations in turn then gradually became less frequent until a large, prolonged (5-10 minute) increase in [Ca2+]i occurred, after which [Ca2+]i returned to a low level. There were no further oscillations after this event, which was seen on average after 37 minutes of ischemia. A slowly progressive contracture often began to develop at about this time. A gradual rise in resting [Ca2+]i occurred during the remainder of the exposure to ischemia. When muscles were reperfused after long exposures to ischemia, there was a very large and prolonged increase in [Ca2+]i, which was usually associated with a contracture and failure of recovery of developed tension. The large increase in [Ca2+]i could be reduced by the inclusion of 3 mM nickel chloride in the reperfusing solution. Comparison between reperfusion with O2 gas versus reperfusion with anoxic Tyrode's solution indicated that reoxygenation was more beneficial to the muscle than resumption of bulk flow. These results reveal the complex spectrum of changes in [Ca2+]i that occur during ischemia and on reperfusion. These changes in [Ca2+]i are likely to play an important role in the generation of ischemic arrhythmias and muscle damage.


[Does ADHD medication stop working after 2-3 years? On the surprising, but little-known follow-up of the MTA study].
The large Multimodal Treatment Study of Children with ADHD (MTA) has become widely known especially because in the treatment phase intensive medication management appeared to be superior (on some outcome-measures) to other treatment modalities. The second followup, 22 months after the end of the treatment phase, shows several surprising and controversial results. For example, the advantage of having partaken in the intensive medication algorithm has disappeared and continued stimulant treatment is a marker not of beneficial outcome, but of deterioration (higher symptom scores). The balance of advantages and disadvantages of longterm stimulant treatment appears to be considerably less favourable than previously thought. A reconsideration of several widely held views on stimulant treatment would seem in order. However, in The Netherlands the results of the second follow-up do not seem to draw the amount of attention and publicity the previous findings did. A possible reason for this is that they contradict prevailing ideas on ADHD. The goal of this article is to draw attention to the most important findings and clinical implications of the second follow-up. At the same time the article illustrates the extent to which the dissemination of scientific knowledge is a social process.


Double transduction of a Cre/LoxP lentiviral vector: a simple method to generate kidney cell-specific knockdown mice.
In a lentivirus-based gene delivery system, the incorporated gene is continuously expressed for a long time. In this study, we devised a simple way to knock down a specific gene in a kidney cell-specific pattern in adult mice by lentivirus-assisted transfer of short hairpin RNA (shRNA). Kidney collecting duct (CD)-specific aquaporin-3 (AQP3)-knockdown mice were generated by consecutive injection of Hoxb7-Cre-expressing lentivirus (LV-Hoxb7 Cre) and loxP-AQP3 shRNA-expressing lentivirus (LV-loxP shAQP3) in adult C57BL6/J mice. LV-Hoxb7 Cre was designed to express mCherry, while LV-loxP shAQP3 was designed with a floxed enhanced green fluorescent protein (EGFP)-tagged stop sequence, and thus EGFP would be expressed only in the absence of Cre recombination. In mice treated with LV-Hoxb7 Cre alone, mCherry protein expression, which indicates the presence of Cre recombinase, occurred only in CD cells. However, LV-loxP shAQP3 injection alone resulted in an increase in EGFP expression in all kidney cells, indicating the transcription of the floxed region. When LV-Hoxb7 Cre and LV-loxP shAQP3 were sequentially transduced, EGFP expression was attenuated while mCherry expression was sustained in CD cells, demonstrating a CD cell-specific recombination of the floxed region. AQP3 expression in mice injected with LV-Hoxb7 Cre or LV-loxP shAQP3 alone did not differ, but consecutive injection of LV-Hoxb7 Cre and LV-loxP shAQP3 significantly reduced AQP3 expression in CD cells. However, the expression levels of AQP3 were not altered in other cell types. Double transduction of Cre- and loxP-based lentivirus can easily generate kidney cell-specific knockdown mice, and this method might be applicable to other species.


Metal-organic Kagome lattices M3(2,3,6,7,10,11-hexaiminotriphenylene)2 (M = Ni and Cu): from semiconducting to metallic by metal substitution.
Motivated by recent experimental synthesis of a semiconducting metal-organic graphene analogue (J. Am. Chem. Soc., 2014, 136, 8859), i.e., Ni3(2,3,6,7,10,11-hexaiminotriphenylene)2 [Ni3(HITP)2], a new Kagome lattice, Cu3(HITP)2, is designed by substituting the coordination of Ni by Cu. Such substitution results in interesting changes in electronic properties of the M3(HITP)2 bulk and two-dimensional (2D) sheets. In Ni3(HITP)2, each Ni atom adopts the dsp(2) hybridization, forming a perfect 2D conjugation, whereas in Cu3(HITP)2, each Cu atom adopts the sp(3) hybridization, resulting in a distorted 2D sheet. The M3(HITP)2 bulks, assembled from M3(HITP)2 sheets via both strong π-π interaction and weak metal-metal interaction, are metallic. However, the 2D Ni3(HITP)2 sheet is a semiconductor with a narrow band gap whereas the 2D Cu3(HITP)2 sheet is a metal. Remarkably, both the 2D M3(HITP)2 Kagome lattices possess Dirac bands in the vicinity of the Fermi level. Additional ab initio molecular dynamics simulations show that both sheets exhibit high thermal stability at elevated temperatures. Our theoretical study offers new insights into tunability of electronic properties for the 2D metal-organic frameworks (MOFs).


Characterization of the metabolic phenotype of rapamycin-treated CD8+ T cells with augmented ability to generate long-lasting memory cells.
Cellular metabolism plays a critical role in regulating T cell responses and the development of memory T cells with long-term protections. However, the metabolic phenotype of antigen-activated T cells that are responsible for the generation of long-lived memory cells has not been characterized. Using lymphocytic choriomeningitis virus (LCMV) peptide gp33-specific CD8(+) T cells derived from T cell receptor transgenic mice, we characterized the metabolic phenotype of proliferating T cells that were activated and expanded in vitro in the presence or absence of rapamycin, and determined the capability of these rapamycin-treated T cells to generate long-lived memory cells in vivo. Antigen-activated CD8(+) T cells treated with rapamycin gave rise to 5-fold more long-lived memory T cells in vivo than untreated control T cells. In contrast to that control T cells only increased glycolysis, rapamycin-treated T cells upregulated both glycolysis and oxidative phosphorylation (OXPHOS). These rapamycin-treated T cells had greater ability than control T cells to survive withdrawal of either glucose or growth factors. Inhibition of OXPHOS by oligomycin significantly reduced the ability of rapamycin-treated T cells to survive growth factor withdrawal. This effect of OXPHOS inhibition was accompanied with mitochondrial hyperpolarization and elevation of reactive oxygen species that are known to be toxic to cells. Our findings indicate that these rapamycin-treated T cells may represent a unique cell model for identifying nutrients and signals critical to regulating metabolism in both effector and memory T cells, and for the development of new methods to improve the efficacy of adoptive T cell cancer therapy.


Cancer/testis (CT) antigens, carcinogenesis and spermatogenesis.
During spermatogenesis, spermatogonial stem cells, undifferentiated and differentiated spermatogonia, spermatocytes, spermatids and spermatozoa all express specific antigens, yet the functions of many of these antigens remain unexplored. Studies in the past three decades have shown that many of these transiently expressed genes in developing germ cells are proto-oncogenes and oncogenes, which are expressed only in the testis and various types of cancers in humans and rodents. As such, these antigens are designated cancer/testis antigens (CT antigens). Since the early 1980s, about 70 families of CT antigens have been identified with over 140 members are known to date. Due to their restricted expression in the testis and in various tumors in humans, they have been used as the target of immunotherapy. Multiple clinical trials at different phases are now being conducted with some promising results. Interestingly, in a significant number of cancer patients, antibodies against some of these CT antigens were detected in their sera. However, antibodies against these CT antigens in humans under normal physiological conditions have yet to be reported even though many of these antigens are residing outside of the blood-testis barrier (BTB), such as in the basal compartment of the seminiferous epithelium and in the stem cell niche in the testis. In this review, we summarize latest findings in the field regarding several selected CT antigens which may be intimately related to spermatogenesis due to their unusual restricted expression during different discrete events of spermatogenesis, such as cell cycle progression, meiosis and spermiogenesis. This information should be helpful to investigators in the field to study the roles of these oncogenes in spermatogenesis.


Human immunodeficiency virus type 1 Vpr induces G2 checkpoint activation by interacting with the splicing factor SAP145.
Vpr, the viral protein R of human immunodeficiency virus type 1, induces G(2) cell cycle arrest and apoptosis in mammalian cells via ATR (for "ataxia-telangiectasia-mediated and Rad3-related") checkpoint activation. The expression of Vpr induces the formation of the gamma-histone 2A variant X (H2AX) and breast cancer susceptibility protein 1 (BRCA1) nuclear foci, and a C-terminal domain is required for Vpr-induced ATR activation and its nuclear localization. However, the cellular target of Vpr, as well as the mechanism of G(2) checkpoint activation, was unknown. Here we report that Vpr induces checkpoint activation and G(2) arrest by binding to the CUS1 domain of SAP145 and interfering with the functions of the SAP145 and SAP49 proteins, two subunits of the multimeric splicing factor 3b (SF3b). Vpr interacts with and colocalizes with SAP145 through its C-terminal domain in a speckled distribution. The depletion of either SAP145 or SAP49 leads to checkpoint-mediated G(2) cell cycle arrest through the induction of nuclear foci containing gamma-H2AX and BRCA1. In addition, the expression of Vpr excludes SAP49 from the nuclear speckles and inhibits the formation of the SAP145-SAP49 complex. To conclude, these results point out the unexpected roles of the SAP145-SAP49 splicing factors in cell cycle progression and suggest that cellular expression of Vpr induces checkpoint activation and G(2) arrest by interfering with the function of SAP145-SAP49 complex in host cells.


A simple dipstick assay for the detection of antibodies to phenolic glycolipid-I of Mycobacterium leprae.
Among the many reported applications of the detection of antibodies to phenolic glycolipid-I (PGL-I) of Mycobacterium leprae, in particular, the use of seroprevalence as an indicator of the magnitude of the leprosy problem may turn out to be very useful in leprosy control programs. An operational function of serology within the leprosy control services requires a simple test system. We have developed a simple dipstick assay for the detection of antibodies to PGL-I and compared its performance with that of an ELISA. A high degree of agreement (97.2%) was observed between the ELISA and the dipstick assay when tested on 435 sera; the agreement beyond chance (Kappa value) was 0.92. No significant difference was found between the dipstick assay and the ELISA when seropositivity rates obtained in groups of leprosy patients, household contacts, and controls were compared. The interpretation of the dipstick results as positive or negative was unequivocal, as illustrated by the high agreement between different persons reading the test (Kappa values > 0.88). Storage of the only reagents required, the dipsticks and the stabilized detection reagent, up to three weeks under tropical conditions of high temperatures, high humidity, and exposure to light, did not influence the results of the assay. The dipstick assay described here is an easy-to-perform method for the detection of IgM antibodies to PGL-I of M. leprae; it does not require any special equipment and the highly stable reagents make the test robust and suitable for use in tropical countries. An internal control validates the performance of the assay. This dipstick assay may be the method of choice for epidemiologic mapping of leprosy.


The impact of newer nucleos(t)ide analogues on patients with hepatitis B decompensated cirrhosis.
Patients with HBV-related decompensated cirrhosis (HBV-DeCi) should be treated with potent nucleos(t)ide analogues (NA)[entecavir (ETV) or tenofovir (TDF)]. The aim was the evaluation of safety and efficacy in terms of changes in liver disease course in HBV-DeCi patients treated with ETV or TDF. In 52 HBV-DeCi patients clinical and laboratory data, including glomerular filtration rates (GFR), were recorded. The changes in MELD (DMELD) and Child-Pugh (DCTP) scores between baseline and after 6 months of treatment were evaluated. The independent factors associated with survival were evaluated. 31 patients under TDF and 21 under ETV were evaluated. During a median follow-up of 22.5 months (range: 6-68), there were no differences between the two groups in GFR and serum phosphate levels. At the end of follow up, in the TDF group, 2 patients died and 3 received liver transplantations (LT), while in the ETV group, 1 patient died and 3 received LT. In multivariable Cox regression analysis, DMELD was independently associated with the outcome in the total cohort (HR: 1.78, 95%C.I.:1.12-2.79, P=0.013) as well as in the subgroup of naïve (n=37) patients (HR: 1.8, 95%C.I.:1.19-4.5, P=0.03). Finally, in the non-hepatocellular carcinoma patients, the DCTP score was independently associated with the outcome in the total cohort (HR: 2.64, 95%C.I.: 1.21-7.29, P=0.015). TDF and ETV appear to have similar renal safety profile in HBV-DeCi patients. DMELD score in the total cohort and DCTP score in non-HCC patients were independently associated with the outcome; these findings need confirmation in larger studies.


Plasticity in single neuron and circuit computations.
Plasticity in neural circuits can result from alterations in synaptic strength or connectivity, as well as from changes in the excitability of the neurons themselves. To better understand the role of plasticity in the brain, we need to establish how brain circuits work and the kinds of computations that different circuit structures achieve. By linking theoretical and experimental studies, we are beginning to reveal the consequences of plasticity mechanisms for network dynamics, in both simple invertebrate circuits and the complex circuits of mammalian cerebral cortex.


Surface-emitting red, green, and blue colloidal quantum dot distributed feedback lasers.
We demonstrate surface emitting distributed feedback (DFB) lasers across the red, green, and blue from densely packed colloidal quantum dot (CQD) films. The solid CQD films were deposited on periodic grating patterns to enable 2nd-order DFB lasing action at mere 120, 280, and 330 μJ/cm2 of optical pumping energy densities for red, green, and blue DFB lasers, respectively. The lasers operated in single mode operation with less than 1 nm of full-width-half-maximum. We measured far-field patterns showing high degree of spatial beam coherence. Specifically, by taking advantage of single exciton optical gain regime from our engineered CQDs, we can significantly suppress the Auger recombination to reduce lasing threshold and achieve quasi-steady state, optically pumped operation.


Fedratinib Improves Myelofibrosis-related Symptoms and Health-related Quality of Life in Patients with Myelofibrosis Previously Treated with Ruxolitinib: Patient-reported Outcomes from the Phase II JAKARTA2 Trial.
Myelofibrosis symptoms compromise health-related quality of life (HRQoL). Ruxolitinib can reduce myelofibrosis symptom severity, but many patients discontinue ruxolitinib due to loss of response or unacceptable toxicity. Fedratinib is an oral, selective JAK2 inhibitor approved in the United States for treatment of patients with intermediate-2 or high-risk myelofibrosis. The single-arm, phase II JAKARTA2 trial assessed fedratinib 400 mg/d (starting dose) in patients with myelofibrosis previously treated with ruxolitinib. Patient-reported changes in myelofibrosis symptom severity using the modified Myelofibrosis Symptom Assessment Form (MFSAF), and overall HRQoL and functional status using the EORTC QLQ-C30, were evaluated at each cycle. Clinically meaningful changes from baseline HRQoL scores were based on effect sizes. Ninety patients were MFSAF-evaluable. Myelofibrosis symptoms were mild-to-moderate at baseline. Patients showed statistically significant and clinically meaningful improvements in total symptom scores from baseline on the MFSAF at all post baseline visits through the end of cycle 6 (EOC6). Baseline global health status/QoL and functional domain scores on the EORTC QLQ-C30 were meaningfully worse than in the general population. At EOC6, 44% of patients reported clinically meaningful improvements in global health status/QoL, and 30%-53% of patients experienced clinically meaningful improvement in QLQ-C30 functional domains across post baseline timepoints. Over 80% of ongoing patients perceived fedratinib as beneficial on the Patient's Global Impression of Change questionnaire. Fedratinib effects were consistent among prognostically relevant patient subgroups. Patients with myelofibrosis previously treated with ruxolitinib experienced clinically meaningful improvements in myelofibrosis symptom burden, overall HRQoL, and functional status in the first 6 months of fedratinib treatment.


Group-constrained sparse fMRI connectivity modeling for mild cognitive impairment identification.
Emergence of advanced network analysis techniques utilizing resting-state functional magnetic resonance imaging (R-fMRI) has enabled a more comprehensive understanding of neurological disorders at a whole-brain level. However, inferring brain connectivity from R-fMRI is a challenging task, particularly when the ultimate goal is to achieve good control-patient classification performance, owing to perplexing noise effects, curse of dimensionality, and inter-subject variability. Incorporating sparsity into connectivity modeling may be a possible solution to partially remedy this problem since most biological networks are intrinsically sparse. Nevertheless, sparsity constraint, when applied at an individual level, will inevitably cause inter-subject variability and hence degrade classification performance. To this end, we formulate the R-fMRI time series of each region of interest (ROI) as a linear representation of time series of other ROIs to infer sparse connectivity networks that are topologically identical across individuals. This formulation allows simultaneous selection of a common set of ROIs across subjects so that their linear combination is best in estimating the time series of the considered ROI. Specifically, l 1-norm is imposed on each subject to filter out spurious or insignificant connections to produce sparse networks. A group-constraint is hence imposed via multi-task learning using a l 2-norm to encourage consistent non-zero connections across subjects. This group-constraint is crucial since the network topology is identical for all subjects while still preserving individual information via different connectivity values. We validated the proposed modeling in mild cognitive impairment identification and promising results achieved demonstrate its superiority in disease characterization, particularly greater sensitivity to early stage brain pathologies. The inferred group-constrained sparse network is found to be biologically plausible and is highly associated with the disease-associated anatomical anomalies. Furthermore, our proposed approach achieved similar classification performance when finer atlas was used to parcellate the brain space.


Transcription of HLA-G transgenes commences shortly after implantation during embryonic development in mice.
We studied the pattern of transcription of a human HLA-G transgene in mice using polymerase chain reaction (PCR) techniques. Transcription of the HLA-G transgene commenced in cells derived from embryos as soon as 48 hr after implantation of embryos in the uterine wall and continued for at least a further 48 hr during embryonic development. HLA-G transcripts were also present in RNA extracted from thymus, spleen and liver of adult HLA-G transgenic mice, although transcripts were not detected in RNA from any other tissues except testes of male transgenic mice. These results demonstrate that the restricted pattern of HLA-G transcription in embryo-derived trophoblast cells during the first trimester of human pregnancy is reproducible in mice. This suggests that transcription factors required for a highly regulated pattern of gene expression during embryonic development are present in murine trophoblast cells and provide a means to investigate the factors and study the consequences of HLA-G expression during development of the embryo.


Antifungal Activity of the Biphosphinic Cyclopalladate C7a against Candida albicans Yeast Forms In Vitro and In Vivo.
Vulvovaginal and invasive candidiasis are frequent conditions in immunosuppressed individuals caused by Candida albicans and non-albicans Candida spp. Fluconazole and Amphotericin B are the main drugs used to fight the infection. However, resistance to fluconazole and other azole antifungal drugs is an important clinical problem that encourages the search for new therapeutic alternatives. In this work, we evaluate the antifungal activity of the biphosphinic cyclopalladate C7a in the in vitro and in vivo model. Our results showed fungicidal activity, with low values of minimal inhibitory concentrations and minimum fungicidal concentrations, even for fluconazole and/or miconazole resistant Candida isolates. Fluorescence microscopy and transmission electron microscopy revealed that the compound was able to inhibit the formation of hyphae/pseudohyphae and, moreover, promoted morphological alterations in cellular organelles and structures, such as disruption of cell wall, apparent mitochondrial swelling, chromatin marginalization into the nuclei and increased numbers of electron-lucent vacuoles. C7a significantly decreased the biofilm formation and reduced the viability of yeast cells in mature biofilms when tested against a virulent C. albicans strain. In vivo assays demonstrated a significant decrease of fungal burden in local (vaginal canal) and disseminated (kidneys) infection. In addition, we observed a significant increase in the survival of the systemically infected animals treated with C7a. Our results suggest C7a as a novel therapeutic agent for vaginal and disseminated candidiasis, and an alternative for conventional drug-resistant Candida.


The conservation physiology toolbox: status and opportunities.
For over a century, physiological tools and techniques have been allowing researchers to characterize how organisms respond to changes in their natural environment and how they interact with human activities or infrastructure. Over time, many of these techniques have become part of the conservation physiology toolbox, which is used to monitor, predict, conserve, and restore plant and animal populations under threat. Here, we provide a summary of the tools that currently comprise the conservation physiology toolbox. By assessing patterns in articles that have been published in 'Conservation Physiology' over the past 5 years that focus on introducing, refining and validating tools, we provide an overview of where researchers are placing emphasis in terms of taxa and physiological sub-disciplines. Although there is certainly diversity across the toolbox, metrics of stress physiology (particularly glucocorticoids) and studies focusing on mammals have garnered the greatest attention, with both comprising the majority of publications (>45%). We also summarize the types of validations that are actively being completed, including those related to logistics (sample collection, storage and processing), interpretation of variation in physiological traits and relevance for conservation science. Finally, we provide recommendations for future tool refinement, with suggestions for: (i) improving our understanding of the applicability of glucocorticoid physiology; (ii) linking multiple physiological and non-physiological tools; (iii) establishing a framework for plant conservation physiology; (iv) assessing links between environmental disturbance, physiology and fitness; (v) appreciating opportunities for validations in under-represented taxa; and (vi) emphasizing tool validation as a core component of research programmes. Overall, we are confident that conservation physiology will continue to increase its applicability to more taxa, develop more non-invasive techniques, delineate where limitations exist, and identify the contexts necessary for interpretation in captivity and the wild.


Adipose concentrations of organochlorine compounds and breast cancer recurrence in Long Island, New York.
Several studies have measured the association between blood or adipose concentrations of organochlorinated compounds (OCs), such as pesticides and polychlorinated biphenyls (PCBs), and breast cancer. The estrogenic effects of OCs might adversely affect breast cancer recurrence. The participants were 224 women with nonmetastatic breast cancer enrolled in a New York-based case-control study. Supercritical fluid extraction followed by gas chromatography was conducted on adipose surgical specimens to determine OC concentrations. The mean follow-up time from surgery was 3.6 years. Thirty women (13.4%) were diagnosed with a recurrence. The concentration of pesticides and PCBs was correlated with baseline age and body mass index, but not with cancer stage. The highest tertile of total PCB concentration was associated with an increased risk of recurrence [relative risk (RR), 2.9; 95% confidence interval (CI), 1.02-8.2 versus the lowest tertile]. The risk for the highest tertile of the PCB congener Ballschmiter and Zell 118 was 4.0 (95% CI, 1.3-4.9). There was an increased risk for the middle level of the most abundant pesticide, 1,1-dichloro-2,2-di(4-chlorophenyl)ethylene (RR, 2.3; 95% CI, 0.9-5.7), and for beta-hexachlorocyclohexane, but not for their highest levels. Self-reported home termiticide exposure, alcohol consumption (> or = 1 drink/day), and race were not associated with prognosis. The RR for current cigarette smoking at diagnosis was 2.1 (95% CI, 0.9-5.1). In contrast to previous data showing no relationship between OC exposure and risk of breast cancer in these women, adipose PCB concentrations were associated with tumor recurrence. Pesticide levels were not related to recurrence.


A novel method for screening viral interferon-resistance genes.
Many viruses have evolved mechanisms to antagonize the interferon (IFN) system, targeting all the major components involved in receptor binding and signaling. Although a number of these vital proteins are homologous to cellular proteins involved in IFN downregulation (e.g., viral IFN regulatory factors [vIRFs]), many share little resemblance to known proteins. To determine the IFN-blocking properties of these proteins, functional assays are required. Here, we present a new and rapid functional screening method, based on the 2fTGH cell line, which is able to determine viral gene products that inhibit the IFN-alpha/Jak-Stat signaling pathway. Expression cloning of viral IFN-blocking genes into 2fTGH and consequent selection with IFN-alpha and 6-thioguanine result in the outgrowth of cells that are no longer responsive to IFN-alpha. We also demonstrate that selection occurs if members of the Jak-Stat signaling pathway are lost. To show the utility of our system, we have used a known suppressor of IFN signaling, the human papillomavirus (HPV) E7 gene. Expression of E7 causes the loss of ability of 2fTGH cells to respond to IFN-alpha treatment because of a functional disruption of the signaling pathway. This approach offers a new strategy for identifying novel viral genes or new functions of already described viral genes that have a role in IFN-alpha signaling inhibition.


Transcriptomics of morphological color change in polychromatic Midas cichlids.
Animal pigmentation has received much attention in evolutionary biology research due to its strong implications for adaptation and speciation. However, apart from a few cases the genetic changes associated with these evolutionary processes remain largely unknown. The Midas cichlid fish from Central America are an ideal model system for investigating pigmentation traits that may also play a role in speciation. Most Midas cichlids maintain their melanophores and exhibit a grayish (normal) color pattern throughout their lives. A minority of individuals, however, undergo color change and exhibit a distinctive gold or even white coloration in adulthood. The ontogenetic color change in the Midas cichlids may also shed light on the molecular mechanisms underlying pigmentation disorders in humans. Here we use next-generation sequencing (Illumina) RNAseq analyses to compare skin transcriptome-wide expression levels in three distinct stages of color transformation in Midas cichlids. cDNA libraries of scale tissue, for six biological replicates of each group, were generated and sequenced using Illumina technology. Using a combination of three differential expression (DE) analyses we identified 46 candidate genes that showed DE between the color morphs. We find evidence for two key DE patterns: a) genes involved in melanosomal pathways are up-regulated in normally pigmented fish; and b) immediate early and inflammatory response genes were up-regulated in transitional fish, a response that parallels some human skin disorders such as melanoma formation and psoriasis. One of the DE genes segregates with the gold phenotype in a genetic cross and might be associated with incipient speciation in this highly "species-rich" lineage of cichlids. Using transcriptomic analyses we successfully identified key expression differences between different color morphs of Midas cichlid fish. These differentially expressed genes have important implications for our understanding of the molecular mechanisms underlying speciation in this lineage of extremely young species since they mate strongly assortatively, and new species may arise by sexual selection due to this color polymorphism. Some of the human orthologues of the genes identified here may also be involved in pigmentation differences and diseases and therefore provide genetic markers for the detection of human pigmentation disorders.


A Machine Learning Approach to Assess Injury Risk in Elite Youth Football Players.
To assess injury risk in elite-level youth football (soccer) players based on anthropometric, motor coordination and physical performance measures with a machine learning model. A total of 734 players in the U10 to U15 age categories (mean age, 11.7 ± 1.7 yr) from seven Belgian youth academies were prospectively followed during one season. Football exposure and occurring injuries were monitored continuously by the academies' coaching and medical staff, respectively. Preseason anthropometric measurements (height, weight, and sitting height) were taken and test batteries to assess motor coordination and physical fitness (strength, flexibility, speed, agility, and endurance) were performed. Extreme gradient boosting algorithms (XGBoost) were used to predict injury based on the preseason test results. Subsequently, the same approach was used to classify injuries as either overuse or acute. During the season, half of the players (n = 368) sustained at least one injury. Of the first occurring injuries, 173 were identified as overuse and 195 as acute injuries. The machine learning algorithm was able to identify the injured players in the hold-out test sample with 85% precision, 85% recall (sensitivity) and 85% accuracy (f1 score). Furthermore, injuries could be classified as overuse or acute with 78% precision, 78% recall, and 78% accuracy. Our machine learning algorithm was able to predict injury and to distinguish overuse from acute injuries with reasonably high accuracy based on preseason measures. Hence, it is a promising approach to assess injury risk among elite-level youth football players. This new knowledge could be applied in the development and improvement of injury risk management strategies to identify youth players with the highest injury risk.


On the locus of the semantic satiation effect: evidence from event-related brain potentials.
The present study sought to determine whether semantic satiation is merely a by-product of adaptation or satiation of upstream, nonsemantic perceptual processes or whether the effect can have a locus in semantic memory. This was done by measuring event-related brain potentials (ERPs) in a semantic word-detection task involving multiple presentations of primes and critical related and unrelated words in three experiments involving visual (Experiment 1) and auditory (Experiments 2A and 2B) stimuli. Primes varied in their type case (Experiment 1) or pitch (Experiment 2B) in order to discourage sensory adaptation. Prime satiation and relatedness of the primes to the critical word had interacting effects on ERP amplitude to critical words, particularly within the time-window of the N400 component. Because numerous studies have indicated a role for the N400 in semantic processing, modulation of the N400 relatedness effect by prime satiation (with little or no contribution from perceptual adaptation) suggests that semantic memory can be directly satiated, rather than the cost to semantic processing necessarily resulting from impoverishment of perceptual inputs.


A comparison of dental treatment utilization and costs by HMO members living in fluoridated and nonfluoridated areas.
To compare dental treatment experiences and costs in members of a health maintenance organization (HMO) in areas with and without community water fluoridation. HMO members with continuous dental eligibility (January 1, 1990 to December 31, 1995) who resided in Oregon and Washington were identified using administrative databases. Fluoridation status was determined by geocoding subscriber address. Measures were utilization of dental procedures, fluoride dispensings, and associated costs. Costs were based on nonmember fees, adjusted to 1995 dollar values. Data were analyzed using analysis of covariance, controlling for age and interactions. About 85 percent of eligible members (n = 51,683) were classified as residing either in a fluoridated (n = 12,194) or nonfluoridated (n = 39,489) area. Mean age was 40.0 years; 52.3 percent were women. More than 92 percent of members had one or more dental visits. Community water fluoridation was associated with reduced total and restorative costs among members with one or more visits, but the magnitude and direction of the effect varied with locale and age and the effects were generally small. In two locales, the cost of restorations was higher in nonfluoridated areas in young people (<age 18) and older adults (>age 58). In younger adults, the opposite effect was observed. The impact of fluoridation may be attenuated by higher use of preventive procedures, in particular supplemental fluorides, in the nonfluoridated areas. These results are particularly relevant to insured populations with established access to dental care. Differences in treatment costs (savings) associated with water fluoridation should be estimated and included in future cost-effectiveness analyses of community water fluoridation.


Absolute coding of stimulus novelty in the human substantia nigra/VTA.
Novelty exploration can enhance hippocampal plasticity in animals through dopaminergic neuromodulation arising in the substantia nigra/ventral tegmental area (SN/VTA). This enhancement can outlast the exploration phase by several minutes. Currently, little is known about dopaminergic novelty processing and its relationship to hippocampal function in humans. In two functional magnetic resonance imaging (fMRI) studies, SN/VTA activations in humans were indeed driven by stimulus novelty rather than other forms of stimulus salience such as rareness, negative emotional valence, or targetness of familiar stimuli, whereas hippocampal responses were less selective. SN/VTA novelty responses were scaled according to absolute rather than relative novelty in a given context, unlike adaptive SN/VTA responses recently reported for reward outcome in animal studies. Finally, novelty enhanced learning and perirhinal/parahippocampal processing of familiar items presented in the same context. Thus, the human SN/VTA can code absolute stimulus novelty and might contribute to enhancing learning in the context of novelty.


Effect of Repeated Recruitment Manoeuvres on Patients with Severe Acute Respiratory Distress Syndrome.
The study aimed to evaluate the influence of repeated recruitment manoeuvres (RRMs) on lung injury in patients with acute respiratory distress syndrome (ARDS). Forty-one ventilated patients with severe ARDS were selected for this study. Recruitment manoeuvres (RMs) were conducted with continuous positive airway pressure (CPAP; 30 cm H2O for 40 seconds). Recruitment manoeuvres were repeated every two hours for all three groups. Changes in haemodynamics, pulmonary compliance, gas exchange and extravascular lung water index (EVLWI) were monitored before RM (pre-RM), 10 minutes after each RM, and four hours after RM3 (4 hours post-RRM). Pulmonary inflammatory factors (tumour necrosis factor-alpha [TNF-α] and interleukin [IL]-6 and -10) were also analysed. Compared with those in pre-RM, pulmonary compliance, oxygenation index (ratio of partial pressure of arterial oxygen to fraction of inspired oxygen [PaO2/FiO2]) and EVLWI remarkably improved in RM1, RM2, RM3 and 4 hours post-RRM (p < 0.05). The PaO2/FiO2 ratio increased significantly in RM1 and RM3 (p < 0.05). Extravascular lung water index decreased significantly in RM1 compared with that in RM3 and 4 hours post-RRM (p < 0.05). There was no significant difference in cytokines. Repeated recruitment manoeuvres during lung-protected ventilation can improve pulmonary compliance and oxygenation and significantly decrease extravascular lung water in ARDS patients. Lung injury was not worsened by RRMs in patients with severe ARDS.


Gallium nitrate inhibits calcium resorption from bone and is effective treatment for cancer-related hypercalcemia.
Approximately two-thirds of patients who receive the anticancer drug gallium nitrate develop mild hypocalcemia. To evaluate the mechanism of drug-induced hypocalcemia, we tested the effects of gallium nitrate upon in vitro release of 45Ca++ from explanted fetal rat bones. The drug significantly inhibited 45Ca++ release in response to stimulation with both parathyroid hormone and a lymphokine preparation with osteoclast activating factor activity. The inhibitory effects on bone resorption were both time- and dose-dependent. Later, in a pilot study, we treated 10 patients who had cancer-related hypercalcemia with gallium nitrate administered by continuous infusion. All patients responded by a reduction of total serum calcium to normal or subnormal concentrations (13.8 +/- 1.05 mg/dl, mean +/- SD pretreatment, to 8.03 +/- 1.03 mg/dl, mean posttreatment nadir). Our results indicate that gallium nitrate effectively treats cancer-related hypercalcemia and that it probably acts by inhibiting calcium release from bone.


Mechanisms of Action of Autophagy Modulators Dissected by Quantitative Systems Pharmacology Analysis.
Autophagy plays an essential role in cell survival/death and functioning. Modulation of autophagy has been recognized as a promising therapeutic strategy against diseases/disorders associated with uncontrolled growth or accumulation of biomolecular aggregates, organelles, or cells including those caused by cancer, aging, neurodegeneration, and liver diseases such as α1-antitrypsin deficiency. Numerous pharmacological agents that enhance or suppress autophagy have been discovered. However, their molecular mechanisms of action are far from clear. Here, we collected a set of 225 autophagy modulators and carried out a comprehensive quantitative systems pharmacology (QSP) analysis of their targets using both existing databases and predictions made by our machine learning algorithm. Autophagy modulators include several highly promiscuous drugs (e.g., artenimol and olanzapine acting as activators, fostamatinib as an inhibitor, or melatonin as a dual-modulator) as well as selected drugs that uniquely target specific proteins (~30% of modulators). They are mediated by three layers of regulation: (i) pathways involving core autophagy-related (ATG) proteins such as mTOR, AKT, and AMPK; (ii) upstream signaling events that regulate the activity of ATG pathways such as calcium-, cAMP-, and MAPK-signaling pathways; and (iii) transcription factors regulating the expression of ATG proteins such as TFEB, TFE3, HIF-1, FoxO, and NF-κB. Our results suggest that PKA serves as a linker, bridging various signal transduction events and autophagy. These new insights contribute to a better assessment of the mechanism of action of autophagy modulators as well as their side effects, development of novel polypharmacological strategies, and identification of drug repurposing opportunities.


Overcoming Obstacles To Enable Access To Medicines For Noncommunicable Diseases In Poor Countries.
The modern access-to-medicines movement grew largely out of the civil-society reaction to the HIV/AIDS pandemic three decades ago. While the movement was successful with regard to HIV/AIDS medications, the increasingly urgent challenge to address access to medicines for noncommunicable diseases has lagged behind-and, in some cases, has been forgotten. In this article we first ask what causes the access gap with respect to lifesaving essential noncommunicable disease medicines and then what can be done to close the gap. Using the example of the push for access to antiretrovirals for HIV/AIDS patients for comparison, we highlight the problems of inadequate global financing and procurement for noncommunicable disease medications, intellectual property barriers and concerns raised by the pharmaceutical industry, and challenges to building stronger civil-society organizations and a patient and humanitarian response from the bottom up to demand treatment. We provide targeted policy recommendations, specific to the public sector, the private sector, and civil society, with the goal of improving access to noncommunicable disease medications globally.


Erk1 positively regulates osteoclast differentiation and bone resorptive activity.
The extracellular signal-regulated kinases (ERK1 and 2) are widely-expressed and they modulate proliferation, survival, differentiation, and protein synthesis in multiple cell lineages. Altered ERK1/2 signaling is found in several genetic diseases with skeletal phenotypes, including Noonan syndrome, Neurofibromatosis type 1, and Cardio-facio-cutaneous syndrome, suggesting that MEK-ERK signals regulate human skeletal development. Here, we examine the consequence of Erk1 and Erk2 disruption in multiple functions of osteoclasts, specialized macrophage/monocyte lineage-derived cells that resorb bone. We demonstrate that Erk1 positively regulates osteoclast development and bone resorptive activity, as genetic disruption of Erk1 reduced osteoclast progenitor cell numbers, compromised pit formation, and diminished M-CSF-mediated adhesion and migration. Moreover, WT mice reconstituted long-term with Erk1(-/-) bone marrow mononuclear cells (BMMNCs) demonstrated increased bone mineral density as compared to recipients transplanted with WT and Erk2(-/-) BMMNCs, implicating marrow autonomous, Erk1-dependent osteoclast function. These data demonstrate Erk1 plays an important role in osteoclast functions while providing rationale for the development of Erk1-specific inhibitors for experimental investigation and/or therapeutic modulation of aberrant osteoclast function.


Household-level expenditure on protective measures against mosquitoes on the island of La Réunion, France.
For decades La Réunion has experienced a number of epidemics that have resulted in efforts to control the density of Aedes species on this Island. This study was conducted to assess household-level expenditure on protective measures against mosquito nuisance on the Island of La Réunion in 2012. Data was collected during a cross-sectional survey of 1024 households and used to determine the relationship between the use of chemically-based protective measures and subjective and objective indicators of the density of Aedes albopictus. The average household expenditure in July 2012 was USD 9.86 and the total household-level expenditure over a one-year period was extrapolated to USD 28.05million (range: USD 25.58 million to USD 30.76 million). Much of this money was spent on measures thought to be relatively ineffective against Aedes mosquitoes. Expenditure on protective measures was not influenced by the level of knowledge on mosquitoes or by the visual nuisance they generated at home, but rather by the perception of risk related to a future epidemic of chikungunya and socioeconomic factors. Most importantly, household spending on protective measures was found to be influenced by a measure of zone-level mosquito density (the Breteau index), but not by objective indicators of the presence of mosquitoes within or around the house. Household-level expenditure on chemically-based protective measures is high when compared to the investment made by public entities to achieve vector control, and it is differentially influenced by subjective and objective measures of mosquito density. The current situation could be improved, firstly by ensuring that the public is well-informed about mosquitoes and the effectiveness of various protective measures, and secondly by implementing interventions that could either complement current vector-control strategies and improve their effectiveness on a country-level, or that would steer the population toward the appropriate behaviours.


Single-fraction simulation of relative cell survival in response to uniform versus hypoxia-targeted dose escalation.
The purpose of this study was to investigate the increase in cell kill that can be achieved by tumor irradiation with heterogeneous dose distributions targeting hypoxic regions that can be visualized with non-invasive imaging. Starting with a heterogeneous distribution of microvessels, a microscopic two-dimensional model of tumor oxygenation was developed using planar simulation of oxygen diffusion. Non-invasive imaging of hypoxia was simulated taking partial volume effect into account. A dose-modulation scheme was implemented with the goal of delivering higher doses to the hypoxic pixels, as seen in simulated hypoxia images. To determine the relative cell kill in response to hypoxia-targeting irradiation, tumor cell survival fractions were compared to those resulting from treatments delivering the same average dose to the lesion in a spatially uniform fashion. It was shown that hypoxia-targeting dose modulation may be better suited for tumors with low α/β, low hypoxic fraction and spatially aggregated hypoxic features. Most importantly, it was determined that at low fraction doses there is no cell kill increase from targeting hypoxic regions alone versus escalating the total tumor dose. However, for higher doses per fraction (≥8 Gy/fraction), the effectiveness of hypoxia-targeting irradiation increases, resulting in the tumoricidal effect of up to 30% higher than that of uniform tumor irradiation delivering the same average tumor dose.


Histological validation of high-resolution DTI in human post mortem tissue.
Diffusion tensor imaging (DTI) is amongst the simplest mathematical models available for diffusion magnetic resonance imaging, yet still by far the most used one. Despite the success of DTI as an imaging tool for white matter fibers, its anatomical underpinnings on a microstructural basis remain unclear. In this study, we used 65 myelin-stained sections of human premotor cortex to validate modeled fiber orientations and oft used microstructure-sensitive scalar measures of DTI on the level of individual voxels. We performed this validation on high spatial resolution diffusion MRI acquisitions investigating both white and gray matter. We found a very good agreement between DTI and myelin orientations with the majority of voxels showing angular differences less than 10°. The agreement was strongest in white matter, particularly in unidirectional fiber pathways. In gray matter, the agreement was good in the deeper layers highlighting radial fiber directions even at lower fractional anisotropy (FA) compared to white matter. This result has potentially important implications for tractography algorithms applied to high resolution diffusion MRI data if the aim is to move across the gray/white matter boundary. We found strong relationships between myelin microstructure and DTI-based microstructure-sensitive measures. High FA values were linked to high myelin density and a sharply tuned histological orientation profile. Conversely, high values of mean diffusivity (MD) were linked to bimodal or diffuse orientation distributions and low myelin density. At high spatial resolution, DTI-based measures can be highly sensitive to white and gray matter microstructure despite being relatively unspecific to concrete microarchitectural aspects.


Photodissociation spectroscopy of protonated leucine enkephalin.
Protonated leucine enkephalin (YGGFL) was studied by ultraviolet photodissociation (UVPD) from 225 to 300 nm utilizing an optical parametric oscillator tunable wavelength laser system (OPO). Fragments were identified by absolute mass measurement in a 9.4 T Fourier transform ion cyclotron resonance mass spectrometer (FT-ICR MS). Bond cleavage was preferred in the vicinity of the two aromatic residues, resulting in high ion abundances for a4, a1, b3, y2 and y1 fragments. a, b and y ions dominated the mass spectrum, and full sequence coverage was achieved for those types. Photodissociation was most effective at the short wavelength end of the studied range, which is assigned to the onset of the La π-π* transition of the tyrosine chromophore, but worked well also at the Lb π-π* chromophore absorption maxima in the 35 000-39 000 cm-1 region. Several side-chain and internal fragments were observed. H atom loss is observed only above 41 000 cm-1, consistent with the requirement of a curve crossing to a repulsive 1πσ* state. It is suggested that the photochemically generated mobile H atom plays a role in further backbone cleavages, similar to the mechanism for electron capture dissociation. The b4 fragment is most intense at the Lb chromophore absorptions, undergoing additional fragmentation at higher photon energies. The high resolution of the FT-ICR MS revealed that out of all x and z-type fragments only x3 and x4 were formed, with low intensity. Other previously reported x- and z-fragments were re-assigned to internal fragments, based on exact mass measurement.


Identification of novel small molecule inhibitors of activated protein C.
Activated protein C (APC) is the central enzyme of the anticoagulant protein C pathway. Low concentrations of APC circulate in plasma and are believed to contribute to the maintenance of a normal haemostatic balance. We have used a structure-based virtual screening approach to discover small drug-like molecules that inhibit the interaction between APC and its substrate FVa through inhibition of a predominant APC exosite, known to be involved in FVa substrate binding. We have combined in silico selection with functional screening and direct binding analysis to identify novel molecules and to ascertain and characterize the inhibition of the interaction between APC and FVa. We have identified a number of novel molecules that bind to APC and protein C with Kd values in the range of 10(-3)- 10(-5)M. Inhibition by these molecules is incomplete, which most likely reflects the extended surface that is involved in the interaction between APC and its substrates. Direct binding of hit molecules to variant APC molecules that were mutated in the targeted binding site revealed that several of the molecules presented a 100-500 fold lower affinity for the variant molecule, suggesting that these molecules indeed bind the exosite of APC. The protein-protein interaction inhibitors discovered here, could function as starting molecules for further development of small molecules with anti-APC properties. Such molecules may be of clinical interest, in particular in individuals where thrombin formation is compromised and the haemostatic balance is tipped towards bleeding tendencies, such as in haemophilia A.


Irritable bowel syndrome and chronic fatigue 6 years after giardia infection: a controlled prospective cohort study.
Functional gastrointestinal disorders and fatigue may follow acute infections. This study aimed to estimate the persistence, prevalence, and risk of irritable bowel syndrome and chronic fatigue 6 years after Giardia infection. We performed a controlled prospective study of a cohort of 1252 individuals who had laboratory-confirmed Giardia infection during a waterborne outbreak in 2004. In total, 748 cohort cases (exposed) and 878 matched controls responded to a postal questionnaire 6 years later (in 2010). Responses were compared to data from the same cohort 3 years before (in 2007). The prevalences of irritable bowel syndrome (39.4%) by Rome III criteria and chronic fatigue (30.8%) in the exposed group 6 years after giardiasis were significantly elevated compared with controls, with adjusted relative risks (RRs) of 3.4 (95% confidence interval [CI], 2.9-3.9) and 2.9 (95% CI, 2.3-3.4), respectively. In the exposed group, the prevalence of irritable bowel syndrome decreased by 6.7% (RR, 0.85 [95% CI, .77-.93]), whereas the prevalence of chronic fatigue decreased by 15.3% from 3 to 6 years after Giardia infection (RR, 0.69 [95% CI, .62-.77]). Giardia exposure was a significant risk factor for persistence of both conditions, and increasing age was a risk factor for persisting chronic fatigue. Giardia infection in a nonendemic setting is associated with an increased risk for irritable bowel syndrome and chronic fatigue 6 years later. The prevalences of both conditions decrease over time, indicating that this intestinal protozoan parasite may elicit very long-term, but slowly self-limiting, complications.


Calcitonin gene-related peptide immunostained axons provide evidence for fine primary afferent fibers in the dorsal and dorsolateral funiculi of the rat spinal cord.
The hypothesis being tested in the present paper is that there are large numbers of fine primary afferent axons in the dorsal and dorsolateral funiculi of the lumbar spinal cord of the rat. The data show numerous calcitonin gene-related peptide labeled fine myelinated and unmyelinated axons in these funiculi. Approximately 95% of the labeled axons disappear after dorsal rhizotomy. Accordingly, the hypothesis is confirmed. Thus it is becoming apparent that fine primary afferent fibers are more widely distributed in spinal white matter than had been previously recognized. Implications are that it is not possible to find areas in the spinal white matter that contain only large myelinated sensory axons and that significant numbers of fine primary afferent fibers will be lost even if lesions are restricted to the dorsal funiculus. The sizable population of fine myelinated primary afferent axons in the dorsal funiculus is emphasized. An obvious question, suggested by significant differences in average diameters of the axons in the different pathways, is whether there are differences in the types of information carried by the fine afferent fibers in their different locations in the white matter of the lumbar cord.


Sequence analyses of Thogoto viral RNA segment 3: evidence for a distant relationship between an arbovirus and members of the Orthomyxoviridae.
The genome of Thogoto (THO) virus, an unclassified tick-borne virus, comprises six segments of single-stranded RNA. The complete sequence of the third largest RNA segment has been determined from overlapping cDNA clones and by primer extension studies. Segment 3 RNA consists of 1865 nucleotides (approx. 6.2 x 10(5) Mr). It has a large open reading frame (ORF1;597 amino acids, 68.6K) in its virus-complementary sequence, confirming that the RNA has a negative-sense coding strategy. A transcription termination (polyadenylation) site located after the end of ORF1 has been identified. A second ORF (ORF2;98 amino acids in length), overlapping ORF1, is also present in the virus-complementary sequence although whether it is translated is not known. The 3' and 5' sequences of the segment 3 RNA are complementary and similar to those of the tick-borne Dhori (DHO) and the mammalian and avian influenza viruses. Protein database searches have identified regions of homology between the sequence of the THO ORF1 gene product and regions of the PA protein of influenza virus strain A/NT/60/68 (approx. 20% aligned homology) and the corresponding protein of influenza B/Sing/222/79 virus (approx. 15% aligned homology). Although the THO protein sequence is not as closely related to those of the influenza viruses as they are to each other (40% aligned homology), the indicated sequence data provide further evidence of relationships between the tick-borne THO and DHO viruses and the vertebrate orthomyxoviruses.


An exported rhodanese-like protein is induced during growth of Acidithiobacillus ferrooxidans in metal sulfides and different sulfur compounds.
By proteomic analysis we found a 21-kDa protein (P21) from Acidithiobacillus ferrooxidans ATCC 19859 whose synthesis was greatly increased by growth of the bacteria in pyrite, thiosulfate, elemental sulfur, CuS, and ZnS and was almost completely repressed by growth in ferrous iron. After we determined the N-terminal amino acid sequence of P21, we used the available preliminary genomic sequence of A. ferrooxidans ATCC 23270 to isolate the DNA region containing the p21 gene. The nucleotide sequence of this DNA fragment contained a putative open reading frame (ORF) coding for a 23-kDa protein. This difference in size was due to the presence of a putative signal peptide in the ORF coding for P21. When p21 was cloned and overexpressed in Escherichia coli, the signal peptide was removed, resulting in a mature protein with a molecular mass of 21 kDa and a calculated isoelectric point of 9.18. P21 exhibited 27% identity and 42% similarity to the Deinococcus radiodurans thiosulfate-sulfur transferase (rhodanese; EC 184.108.40.206) and similar values in relation to other rhodaneses, conserving structural domains and an active site with a cysteine, both characteristic of this family of proteins. However, the purified recombinant P21 protein did not show rhodanese activity. Unlike cytoplasmic rhodaneses, P21 was located in the periphery of A. ferrooxidans cells, as determined by immunocytochemical analysis, and was regulated depending on the oxidizable substrate. The genomic context around gene p21 contained other ORFs corresponding to proteins such as thioredoxins and sulfate-thiosulfate binding proteins, clearly suggesting the involvement of P21 in inorganic sulfur metabolism in A. ferrooxidans.


Constipation is not associated with an increased rate of findings on colonoscopy: results from a national endoscopy consortium.
There are no definite guidelines regarding colonoscopic evaluation for the indication of constipation, a common gastrointestinal complaint. The aim of our study was to determine the risk of finding significant lesions in patients undergoing colonoscopy for the indication of constipation alone compared with constipation with another indication or average-risk screening. A retrospective review of the Clinical Outcomes Research Initiative database was carried out for colonoscopies undertaken between 1 January 2000 and 30 June 2003. A total of 41,775 index colonoscopies performed for the indications of average-risk screening, constipation only or constipation with another indication were identified. Logistic regression analyses were performed for constipation alone versus constipation with another indication, and for constipation alone versus average-risk screening. Constipation alone did not show any increased risk of significant findings on colonoscopy. Constipation and the presence of another indication, however, had a statistically significant increased risk of a significant finding on colonoscopy. The indication of constipation alone had a lower risk of significant findings on colonoscopy compared with average-risk screening. Variations in the definition of constipation used was a limitation of the study. Colonoscopy for constipation alone has a lower yield for significant findings compared with average-risk screening and constipation with another indication; hence, colonoscopy should not be done for constipation alone.


Critical observations on and suggested ways forward for healthcare communication during COVID-19: pEACH position paper.
Communication in healthcare has influenced and been influenced by the COVID-19 pandemic. In this position paper, we share observations based on the latest available evidence and experiential knowledge that have emerged during the pandemic, with a specific focus on policy and practice. This is a position paper that presents observations relating to policy and practice in communication in healthcare related to COVID-19. Through our critical observations as experts in the field of healthcare communication, we share our stance how healthcare communication has occured during the pandemic and suggest possible ways of improving policy and professional practice. We make recommendations for policy makers, healthcare providers, and communication experts while also highlighting areas that merit further investigation regarding healthcare communication in times of healthcare crises. We have witnessed an upheaval of healthcare practice and the development of policy on-the-run. To ensure that policy and practice are evidence-based, person-centred, more inclusive and equitable, we advocate for critical reflection on this symbiotic relationship between COVID-19 and the central role of communication in healthcare. This paper provides a summary of the key areas for development in communication in healthcare during COVID-19. It offers recommendations for improvement and a call to review policies and practice to build resilience and inclusive and equitable responsiveness in communication in healthcare.


Mapping the social network: tracking lice in a wild primate (Microcebus rufus) population to infer social contacts and vector potential.
Studies of host-parasite interactions have the potential to provide insights into the ecology of both organisms involved. We monitored the movement of sucking lice (Lemurpediculus verruculosus), parasites that require direct host-host contact to be transferred, in their host population of wild mouse lemurs (Microcebus rufus). These lemurs live in the rainforests of Madagascar, are small (40 g), arboreal, nocturnal, solitary foraging primates for which data on population-wide interactions are difficult to obtain. We developed a simple, cost effective method exploiting the intimate relationship between louse and lemur, whereby individual lice were marked, without removal from their host, with an individualized code, and tracked throughout the lemur population. We then tested the hypotheses that 1) the frequency of louse transfers, and thus interactions, would decrease with increasing distance between paired individual lemurs; 2) due to host polygynandry, social interactions and hence louse transfers would increase during the onset of the breeding season; and 3) individual mouse lemurs would vary in their contributions to the spread of lice. We show that louse transfers involved 43.75% of the studied lemur population, exclusively males. Louse transfers peaked during the breeding season, perhaps due to increased social interactions between lemurs. Although trap-based individual lemur ranging patterns are restricted, louse transfer rate does not correlate with the distance between lemur trapping locales, indicating wider host ranging behavior and a greater risk of rapid population-wide pathogen transmission than predicted by standard trapping data alone. Furthermore, relatively few lemur individuals contributed disproportionately to the rapid spread of lice throughout the population. Using a simple method, we were able to visualize exchanges of lice in a population of cryptic wild primates. This method not only provided insight into the previously unseen parasite movement between lemurs, but also allowed us to infer social interactions between them. As lice are known pathogen vectors, our method also allowed us to identify the lemurs most likely to facilitate louse-mediated epidemics. Our approach demonstrates the potential to uncover otherwise inaccessible parasite-host, and host social interaction data in any trappable species parasitized by sucking lice.


Pancreatic Cancer: An Emphasis on Current Perspectives in Immunotherapy.
Pancreatic cancer affects both male and female individuals with higher incidences and death rates among the male population. Detection of this malignancy is delayed due to the lack of symptoms in the early-stage cancer, which makes it extremely difficult to treat. Identifying effective strategies has been a challenge for improving the survival rates in pancreatic cancer patients. Resistance to chemotherapy is often developed in pancreatic cancer treatment. Although many strategies are under clinical trials to target certain markers associated with cancer, immunotherapeutic approaches are currently gaining importance. Immunotherapy for pancreatic cancer is in the limelight after preclinical research showed some promise. Immunotherapy approaches were tested along with other treatment options to enhance the treatment effect. Adoptive cell transfer and immune checkpoint inhibitors are currently in clinical trials. The Food and Drug Administration approved pembrolizumab in a fast-tracked review for advanced pancreatic cancer patients. Pembrolizumab blocks the checkpoint protein, programmed cell death protein 1 (PD-1), on T cells to boost the response of the immune system against cancer cells, thereby shrinking tumors. The recent developments in immunotherapy and the early success in other cancers are encouraging to further test immunotherapy in pancreatic cancer. The combination of pembrolizumab and pelareorep, an isolate of human reovirus, is in phase II clinical study in metastatic disease. Depending on the results of current clinical trials and testing, the strategies in the pipeline are expected to increase the use of immunotherapy in the clinical testing setting. Success in immunotherapy is urgently needed to address the side-effects, treating patients with advanced disease and reducing metastasis for increasing the survival rate in pancreatic cancer patients.


Plant growth strategies are remodeled by spaceflight.
Arabidopsis plants were grown on the International Space Station within specialized hardware that combined a plant growth habitat with a camera system that can capture images at regular intervals of growth. The Imaging hardware delivers telemetric data from the ISS, specifically images received in real-time from experiments on orbit, providing science without sample return. Comparable Ground Controls were grown in a sister unit that is maintained in the Orbital Environment Simulator at Kennedy Space Center. One of many types of biological data that can be analyzed in this fashion is root morphology. Arabidopsis seeds were geminated on orbit on nutrient gel Petri plates in a configuration that encouraged growth along the surface of the gel. Photos were taken every six hours for the 15 days of the experiment. In the absence of gravity, but the presence of directional light, spaceflight roots remained strongly negatively phototropic and grew in the opposite direction of the shoot growth; however, cultivars WS and Col-0 displayed two distinct, marked differences in their growth patterns. First, cultivar WS skewed strongly to the right on orbit, while cultivar Col-0 grew with little deviation away from the light source. Second, the Spaceflight environment also impacted the rate of growth in Arabidopsis. The size of the Flight plants (as measured by primary root and hypocotyl length) was uniformly smaller than comparably aged Ground Control plants in both cultivars. Skewing and waving, thought to be gravity dependent phenomena, occur in spaceflight plants. In the presence of an orienting light source, phenotypic trends in skewing are gravity independent, and the general patterns of directional root growth typified by a given genotype in unit gravity are recapitulated on orbit, although overall growth patterns on orbit are less uniform. Skewing appears independent of axial orientation on the ISS - suggesting that other tropisms (such as for oxygen and temperature) do not influence skewing. An aspect of the spaceflight environment also retards the rate of early Arabidopsis growth.


Children's Views on Long-Term Screening for Type 1 Diabetes.
There are an increasing number of medical research studies involving children, including many long-term birth cohort studies. Involving children raises many issues, and little is known about children's own views. This study explored children's views (N = 5,851) on participation in a long-term screening study for type 1 diabetes. The results show that children 10 to 13 years of age have in general a positive attitude to pediatric research and emphasized trust in researchers. The children stressed the importance to receive information and to be involved in decisions. The children also reported feeling concerned about blood sampling and disease risk. Researchers involved in long-term pediatric research need to address these issues to promote involvement and decrease worry.


Risk factor screening to identify women requiring oral glucose tolerance testing to diagnose gestational diabetes: A systematic review and meta-analysis and analysis of two pregnancy cohorts.
Easily identifiable risk factors including: obesity and ethnicity at high risk of diabetes are commonly used to indicate which women should be offered the oral glucose tolerance test (OGTT) to diagnose gestational diabetes (GDM). Evidence regarding these risk factors is limited however. We conducted a systematic review (SR) and meta-analysis and individual participant data (IPD) analysis to evaluate the performance of risk factors in identifying women with GDM. We searched MEDLINE, Medline in Process, Embase, Maternity and Infant Care and the Cochrane Central Register of Controlled Trials (CENTRAL) up to August 2016 and conducted additional reference checking. We included observational, cohort, case-control and cross-sectional studies reporting the performance characteristics of risk factors used to identify women at high risk of GDM. We had access to IPD from the Born in Bradford and Atlantic Diabetes in Pregnancy cohorts, all pregnant women in the two cohorts with data on risk factors and OGTT results were included. Twenty nine published studies with 211,698 women for the SR and a further 14,103 women from two birth cohorts (Born in Bradford and the Atlantic Diabetes in Pregnancy study) for the IPD analysis were included. Six studies assessed the screening performance of guidelines; six examined combinations of risk factors; eight evaluated the number of risk factors and nine examined prediction models or scores. Meta-analysis using data from published studies suggests that irrespective of the method used, risk factors do not identify women with GDM well. Using IPD and combining risk factors to produce the highest sensitivities, results in low specificities (and so higher false positives). Strategies that use the risk factors of age (>25 or >30) and BMI (>25 or 30) perform as well as other strategies with additional risk factors included. Risk factor screening methods are poor predictors of which pregnant women will be diagnosed with GDM. A simple approach of offering an OGTT to women 25 years or older and/or with a BMI of 25kg/m2 or more is as good as more complex risk prediction models. Research to identify more accurate (bio)markers is needed. Systematic Review Registration: PROSPERO CRD42013004608.


A matched set of rat/mouse chimeric antibodies. Identification and biological properties of rat H chain constant regions mu, gamma 1, gamma 2a, gamma 2b, gamma 2c, epsilon, and alpha.
Rat C regions mu, gamma 1, gamma 2a, gamma 2b, gamma 2c, epsilon, and alpha have been characterized by means of chimeric antibody technology. A set of rat/mouse Ag-specific (anti-4-hydroxy-3-nitrophenacetyl) antibodies was constructed that differ only in the H chain constant region but carry identical V region and L chain, both of which are of mouse origin. All rat constant regions could be expressed and m.w. were as expected from the protein sequence. A slight variation in mobility within the IgG subclasses allowed us to establish a hierarchy for the sizes of the four gamma H chains; gamma 2b greater than gamma 1 greater than gamma 2c greater than gamma 2a. Rat IgG2c and IgG2b could be purified on both protein A and protein G while rat IgG2a could only be purified on protein G. Rat IgM and IgG2b were the most potent in C-mediated hemolysis. This was not simply a consequence of the amount of C1q bound because IgG2c bound C1q efficiently but was relatively poor in cell lysis. In ADCC using human effector and target cells, IgG2b and IgG1 were the most effective.


A transcriptional analysis of carotenoid, chlorophyll and plastidial isoprenoid biosynthesis genes during development and osmotic stress responses in Arabidopsis thaliana.
The carotenoids are pure isoprenoids that are essential components of the photosynthetic apparatus and are coordinately synthesized with chlorophylls in chloroplasts. However, little is known about the mechanisms that regulate carotenoid biosynthesis or the mechanisms that coordinate this synthesis with that of chlorophylls and other plastidial synthesized isoprenoid-derived compounds, including quinones, gibberellic acid and abscisic acid. Here, a comprehensive transcriptional analysis of individual carotenoid and isoprenoid-related biosynthesis pathway genes was performed in order to elucidate the role of transcriptional regulation in the coordinated synthesis of these compounds and to identify regulatory components that may mediate this process in Arabidopsis thaliana. A global microarray expression correlation analysis revealed that the phytoene synthase gene, which encodes the first dedicated and rate-limiting enzyme of carotenogenesis, is highly co-expressed with many photosynthesis-related genes including many isoprenoid-related biosynthesis pathway genes. Chemical and mutant analysis revealed that induction of the co-expressed genes following germination was dependent on gibberellic acid and brassinosteroids (BR) but was inhibited by abscisic acid (ABA). Mutant analyses further revealed that expression of many of the genes is suppressed in dark grown plants by Phytochrome Interacting transcription Factors (PIFs) and activated by photoactivated phytochromes, which in turn degrade PIFs and mediate a coordinated induction of the genes. The promoters of PSY and the co-expressed genes were found to contain an enrichment in putative BR-auxin response elements and G-boxes, which bind PIFs, further supporting a role for BRs and PIFs in regulating expression of the genes. In osmotically stressed root tissue, transcription of Calvin cycle, methylerythritol 4-phosphate pathway and carotenoid biosynthesis genes is induced and uncoupled from that of chlorophyll biosynthesis genes in a manner that is consistent with the increased synthesis of carotenoid precursors for ABA biosynthesis. In all tissues examined, induction of β-carotene hydroxylase transcript levels are linked to an increased demand for ABA. This analysis provides compelling evidence to suggest that coordinated transcriptional regulation of isoprenoid-related biosynthesis pathway genes plays a major role in coordinating the synthesis of functionally related chloroplast localized isoprenoid-derived compounds.


Satisfaction with outpatient health care services in Manica Province, Mozambique.
The objective of the study was to describe ambulatory health care services, determine the level of client satisfaction, and identify obstacles to care in a rural area of Mozambique. Exit surveys at 34 health clinics in Manica Province were completed on a sample of 879 adults representing between 1% and 2% of the average monthly visit totals at each clinic. Eighty-three per cent of interviewees were women. Just over half of the visits were for paediatric patients. Men were more likely to be at the clinic for their own health care needs than women (81% vs. 40%, p < 0.001). Of patients seen for acute illness, 45% were examined, 22% received preventive education, and 23% received prognostic information. Overall, 55% of interviewees believed that the service they received was good or very good, 32% rated it as fair, and 13% as poor. Satisfaction was positively associated with increased training level of the provider (p < 0.005), and shorter waiting times (p < 0.001). The most common complaints about the clinic visits were lack of adequate transportation, long waiting times, lack of physical examinations, and failure to receive prescribed medications. These findings suggest that the majority of Mozambicans interviewed are moderately satisfied with the available outpatient services in Manica. Provider training, provider availability and distribution of medicines were areas identified by respondents as needing improvement.


Separation and characterization of metallosupramolecular libraries by ion mobility mass spectrometry.
The self-assembly of Zn(II) ions and bis(terpyridine) (tpy) ligands carrying 120° or 180° angles between their metal binding sites was utilized to prepare metallosupramolecular libraries with the <tpy-Zn(II)-tpy> connectivity. These combinatorial libraries were separated and characterized by ion mobility mass spectrometry (IM MS) and tandem mass spectrometry (MS(2)). The 180°-angle building blocks generate exclusively linear complexes, which were used as standards to determine the architectures of the assemblies resulting from the 120°-angle ligands. The latter ligand geometry promotes the formation of macrocyclic hexamers, but other n-mers with smaller (n = 5) or larger ring sizes (n = 7-9) were identified as minor products, indicating that the angles in the bis(terpyridine) ligand and within the coordinative tpy-Zn(II)-tpy bonds are not as rigid, as previously believed. Macrocyclic and linear isomers were detected in penta- and heptameric assemblies; in the larger octa- and nonameric assemblies, ring-opened conformers with compact and folded geometries were observed in addition to linear extended and cyclic architectures. IM MS(2) experiments provided strong evidence that the macrocycles present in the libraries were already formed in solution, during the self-assembly process, not by dissociation of larger complexes in the gas phase. The IM MS/MS(2) methods provide a means to analyze, based on size and shape (architecture), supramolecular libraries that are not amenable to liquid chromatography, LC-MS, NMR, and/or X-ray techniques.