The role of tolloid/mini fin in dorsoventral pattern formation of the zebrafish embryo.
A highly conserved TGF-&bgr; signaling pathway is involved in the establishment of the dorsoventral axis of the vertebrate embryo. Specifically, Bone Morphogenetic Proteins (Bmps) pattern ventral tissues of the embryo while inhibitors of Bmps, such as Chordin, Noggin and Follistatin, are implicated in dorsal mesodermal and neural development. We investigated the role of Tolloid, a metalloprotease that can cleave Chordin and increase Bmp activity, in patterning the dorsoventral axis of the zebrafish embryo. Injection of tolloid mRNA into six dorsalized mutants rescued only one of these mutants, mini fin. Through chromosomal mapping, linkage and cDNA sequence analysis of several mini fin alleles, we demonstrate that mini fin encodes the tolloid gene. Characterization of the mini fin mutant phenotype reveals that Mini fin/Tolloid activity is required for patterning ventral tissues of the tail: the ventral fin, and the ventroposterior somites and vasculature. Gene expression studies show that mfn mutants exhibit reduced expression of ventrally restricted markers at the end of gastrulation, suggesting that the loss of ventral tail tissues is caused by a dorsalization occurring at the end of gastrulation. Based on the mini fin mutant phenotype and the expression of tolloid, we propose that Mini fin/Tolloid modifes the Bmp activity gradient at the end of gastrulation, when the ventralmost marginal cells of the embryo are in close proximity to the dorsal Chordin-expressing cells. At this time, unimpeded Chordin may diffuse to the most ventral marginal regions and inhibit high Bmp activity levels. In the presence of Mini fin/Tolloid, however, Chordin activity would be negatively modulated through proteolytic cleavage, thereby increasing Bmp signaling activity. This extracellular mechanism is amplified by an autoregulatory loop for bmp gene expression.


Interactive national virtual water-energy nexus networks.
Across the globe, many regions import virtual resources to support their development. Although many researchers have studied transfers of a single virtual resource, interactions across two types of virtual resource transfer networks - energy and water, for example - have rarely been explored simultaneously. To address these knowledge gaps, we constructed and analyzed interprovincial virtual water and energy transfer networks, using China (the largest energy consumer and is undergoing severe water scarcity) as a demonstration. The results unexpectedly showed that more than 40% of provinces gained one kind of resource (either water or energy) through trade at the expense of losing the other kind of internal resource (energy or water), and 20% of provinces suffered a double loss of both water and energy. The remaining provinces gained both water and energy. Surprisingly, approximately 40% of transferred water/energy was from relatively water/energy-scarce provinces to water/energy-abundant provinces, further deepening resource inequality. Moreover, 33.3% and 26.7% of the provinces relied more on cross-border trade than on internal resources to support their water and energy consumption, respectively. Furthermore, in terms of total trade volume, 83.3% and 73.3% of provinces depended more on distant provinces via trade than adjacent ones to support their water and energy consumption, respectively. Overall, virtual water-energy networks tended to enhance each other. Trade largely shaped the nexus relationship between water and energy consumption in provinces. Our study suggests that there is an urgent need to assess multiple virtual resource networks simultaneously in other countries to uncover unintended consequences and to develop cross-sectoral and holistic policies to achieve global sustainability and human well-being.


Diabetic neuropathy and surface sway-referencing disrupt somatosensory information for postural stability in stance.
In order to determine the type of somatosensory information for postural control that is most affected by neuropathy, we compared the relative effects of three methods of sway-referencing the surface in a group of subjects with profound loss of somatosensory function associated with sensory polyneuropathy from diabetes with age-matched control subjects. Sway-referencing disrupted somatosensory feedback for postural control by servo-controlling the dorsi- and plantar-flexion rotation of the support surface in proportion to anterior-posterior excursion of (1) ankle angle, (2) center of body mass (CoM) angle or (3) filtered center of pressure (CoP). Postural sway in subjects with somatosensory loss was significantly larger than normal on a firm surface but not on the sway-referenced surfaces, suggesting that sway-referencing disrupts somatosensory information for postural control already disrupted by neuropathy. Control subjects standing on any sway-referenced surface swayed significantly more than neuropathy subjects who stood on a firm surface, suggesting that sway-referencing disrupts more somatosensory information than disrupted by severe neuropathy. CoP sway-referencing was less sensitive than ankle or CoM sway-referencing for distinguishing postural sway in subjects with somatosensory loss from age-matched control subjects. Given that filtered CoP sway-referencing disrupts the ability to utilize somatosensory information related to surface reactive force to a greater extent than the other two methods of sway-referencing, then these results support the hypothesis that subjects with diabetic peripheral neuropathy have lost more CoP information, than ankle or CoM angle information, for controlling postural sway in stance.


Spectral anomaly detection in deep shadows.
Although several hyperspectral anomaly detection algorithms have proven useful when illumination conditions provide for enough light, many of these same detection algorithms fail to perform well when shadows are also present. To date, no general approach to the problem has been demonstrated. In this paper, a novel hyperspectral anomaly detection algorithm that adapts the dimensionality of the spectral detection subspace to multiple illumination levels is described. The novel detection algorithm is applied to reflectance domain hyperspectral data that represents a variety of illumination conditions: well illuminated and poorly illuminated (i.e., shadowed). Detection results obtained for objects located in deep shadows and light-shadow transition areas suggest superiority of the novel algorithm over standard subspace RX detection.


[Comparison of NO contents and cutaneous electric conduction quantity at the acupoints and the non-acupoints].
To observe the effects of acupuncture at acupoints and non-acupoints on the subcutaneous nitric oxide (NO) content and skin electric conduction quantity. A total of 40 healthy volunteers (half males and half females at ages of 25-45 years) signed informed consent for detection of NO (n=20) contents and skin electric conduction quantity (electric current, EC, n=20) of the acupoint and non-acupoint area. The two series of 20 subjects were equally randomized into acupuncture group and non-acupuncture group, respectively. Jianshi (PC 5), Chengshan (BL 57) were punctured with filiform needles, with the needles twirled for 2 min and retained for 20 min. Dermal electric current were detected by using Dermatron (electrodermal screening device), and NO contents in the microdialysis fluid collected from the subcutaneous tissues of Ximen (PC 4), Chengjin (BL 56) and non-acupoints 1.0 approximately 1.5 cm lateral to PC 4 and BL 56 were detected by chemiluminescence technique. (1) NO contents of PC 4, BL 56 and two non-acupoint regions in acupuncture group were all significantly higher than those of two acupoints and two non-acupoints in non-acupuncture group (P<0.05). In acupuncture group, the NO contents of both PC 4 and BL 56 were strikingly higher than those of two non-acupoints (P<0.05). In non-acupuncture group there were no significant differences between PC 4 and non-acupoint and between BL 56 and non-acupoint (P>0.05). (2) The skin electric current values of PC 4 and BL 56 were significantly higher than those of two corresponding non-acupoints in both acupuncture and non-acupuncture groups (P<0.05). Following acupuncture stimulation, an increasing tendency was found in the electric current in comparison with non-acupuncture group but without significance (P>0.05). Acupuncture stimulation can upregulate dermal NO content and and the skin electric current of acupoint region was significantly higher than that of non-acupoint in healthy subjects, suggesting an increase of potential activity after acupuncture.


Endothelin-1 induces connective tissue growth factor expression in human lung fibroblasts by ETAR-dependent JNK/AP-1 pathway.
Endothelin-1 (ET-1) acts as a key mediator of vasoconstriction and tissue repair. Overproduction of connective tissue growth factor (CTGF) underlies the development of lung fibrosis. ET-1 induces expression of matrix-associated genes in lung fibroblasts, however, little is known about the signaling pathway of CTGF expression caused by ET-1. In this study, we found that ET-1 caused concentration- and time-dependently increases in CTGF expression in human embryonic lung fibroblast cell line (WI-38). ET-1-induced CTGF expression was inhibited by BQ123 (ETAR antagonist), but not BQ788 (ETBR antagonist). Moreover, ET-1-induced CTGF expression was significantly reduced by JNK inhibitor (SP600125), the dominant-negative mutants of JNK1/2 (JNK1/2 DN), and AP-1 inhibitor (curcumin). ET-1 induced phosphorylations of JNK and c-Jun in time-dependent manners. AP-1 luciferase activity was concentration-dependently increased by ET-1, and this effect was attenuated by SP600125. We also found that ET-1-induced CTGF expression was most controlled by the AP-1 binding region of CTGF promoter. ET-1-indiced CTGF luciferase activity was predominately controlled by the sequence -747 to -408 bp upstream of the transcription start site on the human CTGF promoter. Furthermore, ET-1 caused the formation of AP-1-specific DNA-protein complex and the recruitment of c-Jun to the CTGF promoter. Moreover, we found that ET-1 induced α-smooth muscle actin (α-SMA) expression, which was inhibited by BQ123, SP600125, curcumin, and anti-CTGF antibody. These results suggest that ET-1 stimulates expressions of CTGF and α-SMA through ETAR/JNK/AP-1 signaling pathway, and CTGF is required for ET-1-induced α-SMA expression in human lung fibroblasts.


A comparison of lesions in small intracerebral arteries among Japanese men in Hawaii and Japan.
This report examines the hypothesis that the higher risk of stroke among Japanese men in Japan compared with those in Hawaii is related to pathology in small intracerebral arteries by comparing the prevalence of such lesions in autopsied participants from two cohorts of Japanese men in Japan and Hawaii. Existing histological sections from the left basal ganglia from 232 men from Japan and 175 men of Japanese ancestry in Hawaii were examined for selected abnormalities in arteries between 100 and 300 microns in diameter by three pathologists. The presence of lacunar infarcts was also noted, and information about cerebral infarcts, cerebral hemorrhages, and atherosclerosis in the circle of Willis was available for the Hawaii group. Lacunar infarcts and all small intracerebral artery lesions except medial fibrosis were more common at every age in Japan than in Hawaii. By cause of death, all lesions were three or more times more prevalent among men who died of stroke than of noncardiovascular causes in both areas. In the Hawaii group, the small intracerebral artery lesions were significantly associated with autopsy evidence of cerebral and lacunar infarcts, and with atherosclerosis in the large arteries of the circle of Willis. Among a large number of risk factors measured at the baseline examination in Hawaii, only high blood pressure and reported usual Asian diet were significantly associated with one or more measures of small intracerebral artery lesions. An overview of the accumulated data indicated that small intracerebral artery pathology plays an important role in the high risk of stroke in Japanese men in Japan compared with those in Hawaii. These studies support the idea that hypertension is a necessary factor in the causal pathway, but also indicate that some other factors are involved. Some aspect of an Asian diet continues to be of importance for future research.


Noninvasive monitoring of internal carotid artery dissection.
Internal carotid artery dissection has increasingly been reported as a cause of transient ischemic attack or stroke. However, scarce data exist on the natural history of the arterial lesions and the temporal profile of recanalization. We followed 48 patients with 50 angiographically confirmed internal carotid artery dissections by sequential duplex Doppler studies in 2- to 4-day intervals during the first weeks after the onset of symptoms and after 4 weeks in 1- to 2-month intervals for up to 2 years. We assessed sonographic features as well as the frequency and time course of resolution. Initial Doppler findings were abnormal in all patients, most of whom (68%) presented with a characteristic bidirectional high-resistance Doppler signal in the internal carotid artery. Gradual recanalization was found in 68% of the dissections after an average interval of 51 days. Changes of Doppler flow patterns in follow-up studies and features of intra-arterial angiography correlated with the development of internal carotid artery dissection and mirrored the recanalization process. Our findings suggest that Doppler sonography provides early recognition of internal carotid artery dissection and monitoring of its resolution. Thus, ultrasound studies may guide clinical decisions according to the development of the dissection.


A case-control nutrigenomic study on the synergistic activity of folate and vitamin B12 in cervical cancer progression.
The present study was designed to identify the role of folate, B12, homocysteine, and polymorphisms of methylene tetrahydrofolatereductase (MTHFR) gene in cervical carcinogenesis among 322 women from Kerala, South India. Serum folate, vitamin B12 (chemiluminescence assay), and homocysteine (EIA) along with genetic polymorphisms of MTHFR gene (polymerase chain reaction/restriction fragment length polymorphism) were analyzed for 136 control subjects, 92 low-grade squamous intraepithelial lesions (LSIL) subjects, and 94 invasive cervical cancer cases (ICC). Statistically significant associations between MTHFR polymorphisms, serum homocysteine, and folate levels with cervical carcinogenesis were not evident, but we found that these parameters acted as effect modifiers of serum vitamin B12. The risk estimates observed for B12 became prominent only when there was a deficiency in serum folate levels [LSIL-odds ratio (OR): 14.9 (95% CI: 2.65 to 84.4); ICC-OR = 8.72 (95% CI = 1.55 to 48.8)] or when MTHFR A1298C polymorphic variant was present [LSIL-OR = 9.8 (95% CI = 2.61 to 36.7); ICC-OR = 10.0 (95%CI = 2.5 to 39.3)]. The statistical significance of this effect modification was further studied using an interaction model, where only folate was observed to have an influence on B12 levels as suggested by the odds ratio of 7.11 (95% CI = 0.45 to 111.9) obtained for ICC group, implicating a synergistic role of these 2 vitamins in invasive cervical cancer.


Adjuvant Dabrafenib plus Trametinib in Stage III BRAF-Mutated Melanoma.
Combination therapy with the BRAF inhibitor dabrafenib plus the MEK inhibitor trametinib improved survival in patients with advanced melanoma with BRAF V600 mutations. We sought to determine whether adjuvant dabrafenib plus trametinib would improve outcomes in patients with resected, stage III melanoma with BRAF V600 mutations. In this double-blind, placebo-controlled, phase 3 trial, we randomly assigned 870 patients with completely resected, stage III melanoma with BRAF V600E or V600K mutations to receive oral dabrafenib at a dose of 150 mg twice daily plus trametinib at a dose of 2 mg once daily (combination therapy, 438 patients) or two matched placebo tablets (432 patients) for 12 months. The primary end point was relapse-free survival. Secondary end points included overall survival, distant metastasis-free survival, freedom from relapse, and safety. At a median follow-up of 2.8 years, the estimated 3-year rate of relapse-free survival was 58% in the combination-therapy group and 39% in the placebo group (hazard ratio for relapse or death, 0.47; 95% confidence interval [CI], 0.39 to 0.58; P<0.001). The 3-year overall survival rate was 86% in the combination-therapy group and 77% in the placebo group (hazard ratio for death, 0.57; 95% CI, 0.42 to 0.79; P=0.0006), but this level of improvement did not cross the prespecified interim analysis boundary of P=0.000019. Rates of distant metastasis-free survival and freedom from relapse were also higher in the combination-therapy group than in the placebo group. The safety profile of dabrafenib plus trametinib was consistent with that observed with the combination in patients with metastatic melanoma. Adjuvant use of combination therapy with dabrafenib plus trametinib resulted in a significantly lower risk of recurrence in patients with stage III melanoma with BRAF V600E or V600K mutations than the adjuvant use of placebo and was not associated with new toxic effects. (Funded by GlaxoSmithKline and Novartis; COMBI-AD ClinicalTrials.gov, NCT01682083 ; EudraCT number, 2012-001266-15 .).


A new peptide-based fluorescent probe selective for zinc(ii) and copper(ii).
A novel metal ion-sensitive fluorescent peptidyl-probe has been designed based on the most common five-residue repeat in mammalian histidine rich glycoproteins (HRGs). A dansyl-amide moiety at the N-terminus and a tryptophan residue at the C-terminus of the peptide were added as they can act as a FRET (fluorescence resonance energy transfer) pair. The dansyl fluorophore was chosen also because it frequently shows strong CHEF (chelation enhanced fluorescence) and solvatochromic effects. The designed peptide, dansyl-HPHGHW-NH2 (dH3w), showed a selective fluorescence turn-on response to Zn2+ in aqueous solutions at pH 7.0 when excited at both 295 nm and 340 nm, thus indicating that both FRET and CHEF or solvatochromic effects are active in the metal/peptide complex. Steady-state fluorescence and isothermal titration calorimetry (ITC) measurements demonstrated that two peptide molecules bind to one zinc ion with an association constant Ka = 5.7 × 105 M-1 at 25 °C and pH 7.0. The fluorescence response to Zn2+ was not influenced by Pb2+, Cd2+, Mn2+, Fe2+, Fe3+, Mg2+, Ca2+, K+ and Na+ ions and only slightly influenced by Co2+ and Ni2+. Copper(ii), at concentrations as low as 5 μM, caused a strong quenching of both free and Zn2+ complexed dH3w. The determination of the binding parameters for Cu2+ has shown that one copper ion binds to one dH3w molecule with an association constant of 1.2 × 106 M-1 thus confirming the higher affinity of peptide for Cu2+ than for Zn2+. Finally, we demonstrated that dH3w can penetrate into HeLa cells and could thus be used for the determination of intracellular Zn2+ and Cu2+ concentrations.


NEIL1 excises 3' end proximal oxidative DNA lesions resistant to cleavage by NTH1 and OGG1.
Base excision repair is the major pathway for the repair of oxidative DNA damage in human cells that is initiated by a damage-specific DNA glycosylase. In human cells, the major DNA glycosylases for the excision of oxidative base damage are OGG1 and NTH1 that excise 8-oxoguanine and oxidative pyrimidines, respectively. We find that both enzymes have limited activity on DNA lesions located in the vicinity of the 3' end of a DNA single-strand break, suggesting that other enzymes are involved in the processing of such lesions. In this study, we identify and characterize NEIL1 as a major DNA glycosylase that excises oxidative base damage located in close proximity to the 3' end of a DNA single-strand break.


Self-harm and suicide in adolescents.
Self-harm and suicide are major public health problems in adolescents, with rates of self-harm being high in the teenage years and suicide being the second most common cause of death in young people worldwide. Important contributors to self-harm and suicide include genetic vulnerability and psychiatric, psychological, familial, social, and cultural factors. The effects of media and contagion are also important, with the internet having an important contemporary role. Prevention of self-harm and suicide needs both universal measures aimed at young people in general and targeted initiatives focused on high-risk groups. There is little evidence of effectiveness of either psychosocial or pharmacological treatment, with particular controversy surrounding the usefulness of antidepressants. Restriction of access to means for suicide is important. Major challenges include the development of greater understanding of the factors that contribute to self-harm and suicide in young people, especially mechanisms underlying contagion and the effect of new media. The identification of successful prevention initiatives aimed at young people and those at especially high risk, and the establishment of effective treatments for those who self-harm, are paramount needs.


Bronchial artery embolization in the treatment of massive hemoptysis.
To evaluate the efficacy of bronchial arteriography and bronchial artery embolization (BAE) in the management of massive hemoptysis in a developing Asian country. A retrospective review was carried out from March 2000 to March 2005 to evaluate the demographics, clinical presentation, radiographic studies, bronchoscopy results, and complications of bronchial arteriography and BAE at a tertiary care hospital in Pakistan. Fourteen patients (9 males, 5 females) with a mean age of 49 years underwent bronchial arteriography and BAE for massive hemoptysis. Hemoptysis was caused by bronchiectasis (10 patients), active pulmonary tuberculosis (3 patients), and lung malignancy (one patient). A CT scan of the chest was carried out in 11 patients, which revealed bronchiectasis (8 patients), cavity with infiltrates (3 patients), and mass lesion (one patient). Bronchoscopy was performed in all patients. Bleeding lobe or segment was identified in 12 patients. Bronchial arteriography revealed hypervascularity (13 patients), bronchial artery hypertrophy (5 patients), hypervascularity with shunting (one patient), dense soft tissue staining (7 patients), extravasation of contrast (one patient) and pseudoaneurysm (one patient). Bronchial artery embolization was carried out in all patients. Rebleeding occurred within 24 hours in 2 patients who underwent surgery, and within one week in another 2 patients who were managed with repeat BAE. The complication of embolization occurred in one patient (transverse myelitis). Thirteen patients improved and were discharged home. One patient with terminal lung carcinoma died due to cardiogenic shock secondary to acute myocardial infarction. Bronchial artery embolization is an effective method for management of massive hemoptysis in developing countries and has a low complication rate.


In situ catalytic pyrolysis of lignocellulose using alkali-modified amorphous silica alumina.
Canadian pinewood was pyrolyzed at 450 °C in an Infrared oven and the pyrolysis vapors were converted by passing through a catalyst bed at 450 °C. The catalysts studied were amorphous silica alumina (ASA) containing alkali metal or alkaline earth metal species including Na, K, Cs, Mg and Ca. The catalysts effectiveness to reduce the bio-oil oxygen content, to enhance the bio-oil energy density and to change the liquid and gas product distribution were evaluated using different techniques including gravimetric analysis, elemental analysis, Karl-Fischer titration, GC/MS and micro-GC analysis. According to the results K/ASA found to be the most effective catalysts for conversion of hollocellulose (hemicellulose and cellulose)-derived vapors of pinewood while Cs/ASA catalyst was the most effective catalyst for conversion of lignin-derived vapors and production of hydrocarbons.


Sustained depletion of FXIII-A by inducing acquired FXIII-B deficiency.
The activated form of coagulation factor XIII (FXIII-A2B2), FXIII-A*, is a hemostatic enzyme essential for inhibiting fibrinolysis by irreversibly crosslinking fibrin and antifibrinolytic proteins. Despite its importance, there are no modulatory therapeutics. Guided by the observation that humans deficient in FXIII-B have reduced FXIII-A without severe bleeding, we hypothesized that a suitable small interfering RNA (siRNA) targeting hepatic FXIII-B could safely decrease FXIII-A. Here we show that knockdown of FXIII-B with siRNA in mice and rabbits using lipid nanoparticles resulted in a sustained and controlled decrease in FXIII-A. The concentration of FXIII-A in plasma was reduced by 90% for weeks after a single injection and for more than 5 months with repeated injections, whereas the concentration of FXIII-A in platelets was unchanged. Ex vivo, crosslinking of α2-antiplasmin and fibrin was impaired and fibrinolysis was enhanced. In vivo, reperfusion of carotid artery thrombotic occlusion was also enhanced. Re-bleeding events were increased after challenge, but blood loss was not significantly increased. This approach, which mimics congenital FXIII-B deficiency, provides a potential pharmacologic and experimental tool to modulate FXIII-A2B2 activity.


Epidemiological Imaginaries of the Social: Epidemiologists and Pathologies of Modernization in Postcolonial Africa.
There is a growing anthropological literature analyzing the place that epidemiological surveillance occupies in contemporary global health. In this article, I build on this literature and take it into new directions by exploring what I call the epidemiological imaginaries of the social. Drawing on science and technology studies, I suggest that epidemiologists help make up the world, articulating complex and normatively loaded visions of social life that both enable and constrain action. More specifically, I argue that epidemiologists tell stories about the type of societies and people that compose the world and that these stories often shape global health policies and programs in powerful ways. To substantiate this argument, I examine epidemiologists' efforts to map smoking in postcolonial Africa, documenting how they have imagined smokers and smoking through the lense of modernization theory and showing how these imaginaries have shaped tobacco control policies in the region up to this day.


Modeling the inherent optical properties of the ocean based on the detailed composition of the planktonic community.
We describe an approach to modeling the ocean's inherent optical properties (IOPs) that permits extensive analyses of IOPs as the detailed composition of suspended particulate matter is varied in a controlled manner. Example simulations of the IOP model, which includes 18 planktonic components covering a size range from submicrometer viruses and heterotrophic bacteria to microplanktonic species of 30-mum cell diameter, are discussed. Input data to the model include the spectral optical cross sections on a per particle basis and the particle-number concentration for each individual component. This approach represents a significant departure from traditional IOP and bio-optical models in which the composition of seawater is described in terms of a few components only or chlorophyll concentration alone. The simulations illustrate how the separation and understanding of the effects of various types of particle present within a water body can be achieved. In an example simulation representing an oligotrophic water body with a chlorophyll a concentration of 0.18 mg m(-3), the planktonic microorganisms altogether are the dominant particulate component in the process of light absorption, but their relative contribution to light scattering is smaller than that of nonliving particles. A series of simulations of water bodies with the same chlorophyll a concentration but dominated by different phytoplankton species shows that composition of the planktonic community is an important source of optical variability in the ocean.


No added benefit from nebulized amiloride in patients with cystic fibrosis.
In cystic fibrosis (CF) airway epithelial sodium absorption is increased 2-3 fold. Since sodium absorption is inhibited by the sodium channel blocker amiloride, our aim was to assess its therapeutic benefit in cystic fibrosis. A randomized, double-blind, placebo-controlled, cross-over trial of nebulized amiloride was performed in 23 patients with cystic fibrosis. Amiloride or placebo was administered four times daily for two six month periods. Existing treatment was continued, and any infective exacerbations treated in the usual way. Fourteen patients completed the study. No significant changes occurred in forced expiratory volume in one second, forced vital capacity, oxygen saturation, body weight, sputum volume, culture and rheology, serum urea, and electrolytes, white cell count and erythrocyte sedimentation rate during either treatment period. The frequency of infective exacerbations was also not different in either treatment period. We were thus unable to confirm the benefit shown in the only other clinical trial of nebulized amiloride in cystic fibrosis and conclude that, in the presence of established treatment for cystic fibrosis lung disease, nebulized amiloride offers no additional clinical benefit.


An electrochemical biosensor for rapid detection of anti-dsDNA antibodies in absolute scale.
Autoimmune diseases are chronic inflammatory pathologies that are characterized by the presence of antibodies against the body's own epitopes in serum (autoantibodies). Systemic lupus erythematosus (SLE) is a common autoimmune pathology characterized by the presence of antinuclear antibodies (ANAs). These include anti-dsDNA (α-dsDNA) antibodies, which are widely used for diagnosis and disease monitoring. Their determination is carried out using traditional techniques such as Indirect Immunofluorescence (IFI) or Enzyme-Linked Immunosorbent Assay (ELISA), which are time consuming, require qualified technicians, and are not compatible with decentralized analysis outside a laboratory facility. Here, we show a sandwich-format electrochemical biosensor-based method for α-dsDNA determination in a rapid and simple manner. The total assay time is only 30 minutes, and the sensor is capable of detecting 16 ng (8 μg mL-1) of α-dsDNA antibodies. Using the current derived from the detection limit of the method as a cut-off, we could discriminate positive from negative serum samples with 90% sensitivity and 100% specificity. Using monoclonal antibodies for calibration curves, our results are presented in absolute scale (i.e., concentration instead of serum titer) which will help us to perform comparisons between methods and carry out further improvements of this protocol. In an effort to render the sensor compatible with automation, we minimized the manipulation steps without compromising the analytical performance, even in complex samples such as serum.


Diffusive shunting of gases and other molecules in the renal vasculature: physiological and evolutionary significance.
Countercurrent systems have evolved in a variety of biological systems that allow transfer of heat, gases, and solutes. For example, in the renal medulla, the countercurrent arrangement of vascular and tubular elements facilitates the trapping of urea and other solutes in the inner medulla, which in turn enables the formation of concentrated urine. Arteries and veins in the cortex are also arranged in a countercurrent fashion, as are descending and ascending vasa recta in the medulla. For countercurrent diffusion to occur, barriers to diffusion must be small. This appears to be characteristic of larger vessels in the renal cortex. There must also be gradients in the concentration of molecules between afferent and efferent vessels, with the transport of molecules possible in either direction. Such gradients exist for oxygen in both the cortex and medulla, but there is little evidence that large gradients exist for other molecules such as carbon dioxide, nitric oxide, superoxide, hydrogen sulfide, and ammonia. There is some experimental evidence for arterial-to-venous (AV) oxygen shunting. Mathematical models also provide evidence for oxygen shunting in both the cortex and medulla. However, the quantitative significance of AV oxygen shunting remains a matter of controversy. Thus, whereas the countercurrent arrangement of vasa recta in the medulla appears to have evolved as a consequence of the evolution of Henle's loop, the evolutionary significance of the intimate countercurrent arrangement of blood vessels in the renal cortex remains an enigma.


Tissue-specific transcriptional regulation of seven heavy metal stress-responsive miRNAs and their putative targets in nickel indicator castor bean (R. communis L.) plants.
R.communis L. has high capability to accumulate nickel which is a trace nutrient for higher plants and also an environmental contaminant causes toxicity related symptoms at higher concentrations. MicroRNAs (miRNAs) are known to be important modulators of responses against heavy metal stress for detoxification of the metal. In this study, we experimentally measured and validated the transcript levels of the seven heavy metal stress response-related miRNAs and the expression levels of target genes in both leaf and root tissues of R. communis L. subjected to three different concentrations of nickel stress via qRT-PCR quantification. The results demonstrated differential regulations of heavy metal stress-responsive miRNAs and their putative targets in both tissues in same stress treatments. This dynamic regulation suggest that regulatory processes differ between the tissues under nickel stress. Our data suggest that, miR838 was the most responsive to the Ni2+ stress. miR398 target gene Cu-Zn/SOD was found to be up-regulated in both root and leaf tissues. The relations between TCP and expression levels of miR159 and miR319 were also found statistically significant exclusive to leaf tissues. In leaf tissue, changes in miR395 level and its putative target genes, sulphate transporter and sulphate adenyltransferase gene were found in relation whereas, only expression level of sulphate transporter represented a statistically significant relation in root tissue. The sharp decrease in transcript levels of 2r3 myb gene at lower nickel dose suggest to investigate the role of r2r3 myb and the all MYB family members in primary and secondary metabolisms against nickel stress.


Coverage of HPV vaccination programme in Friuli Venezia Giulia Region (Northern Italy).
to assess the completeness and timeliness of Human papillomavirus (HPV) vaccination programme in Friuli Venezia Giulia (FVG) Region (Northern Italy), notably by monitoring 2-dose coverage among girls before they turn 15 years old (referred to as "at 15") in each year between 2009 and 2018 and making a preliminary evaluation of coverage among boys at 13 years in 2016-2018. retrospective study. for each vaccine recipient, demographic information and history of HPV vaccine uptake from the digital FVG Vaccination Registry updated as of 31.12.2018 were extracted. Numerator data comprised all doses allocated to FVG residents. Age-specific denominators were derived from the FVG census in each examined year. Coverage estimates for the year 2018 were also provided by number of doses. coverage for a full course of HPV vaccine, defined as 2 doses in girls and boys younger than age 15 years but 3 doses in less young women. In FVG 52,217 females had received >=1 dose since 2008 and 12,152 males since 2015. >=2-dose coverage in girls at 15 increased from 42% in 2009 to 76% in 2015 and slightly declined afterwards (69% in 2018). In 2008, 3-dose coverage was 65%, 74%, and 59% in females aged 16-17, 18-19, and 20-26 years, respectively. In the same year, 2-dose coverage in boys at 13 years was 54%, similar to the coverage in girls at 13 years (57%). this paper shows the achievements of routine and catch-up HPV vaccination in FVG. While coverage in girls at 15 years of age peaked in 2015 and slightly diminished in subsequent years, the coverage in boys at 13 in 2018 had already approached the coverage in same-age girls (57%). On account of the signs of weakening in girls' coverage, campaigns in support to HPV vaccination must be repeated, especially in favour of the most cost-effective group, i.e., girls before 15 years of age. The heavy burden posed by the COVID-19 emergency on other prevention-related activities makes a better targeted use of HPV vaccination even more necessary.


Adipose tissue and the immune system.
Adipocytes anatomically associated with lymph nodes (and omental milky spots) have many special properties including fatty acid composition and the control of lipolysis that equip them to interact locally with lymphoid cells. Lymph node lymphocytes and tissue dendritic cells acquire their fatty acids from the contiguous adipocytes. Lymph node-derived dendritic cells suppress lipolysis in perinodal adipocytes but those that permeate the adipose tissue stimulate lipolysis, especially after minor, local immune stimulation. Inflammation alters the composition of fatty acids incorporated into dendritic cells, and that of node-containing adipose tissue, counteracting the effects of dietary lipids. Thus these specialised adipocytes partially emancipate the immune system from fluctuations in the abundance and composition of dietary lipids. Prolonged, low-level immune stimulation induces the local formation of more adipocytes, especially adjacent to the inflamed lymph node. This mechanism may contribute to hypertrophy of the mesentery and omentum in chronic inflammatory diseases such as HIV-infection, and in smokers. Paracrine interactions between adipose and lymphoid tissues are enhanced by diets rich in n-6 fatty acids and attentuated by fish oils. The latter improve immune function and body conformation in animals and people. The partitioning of adipose tissue in many depots, some specialised for local, paracrine interactions with other tissues, is a fundamental feature of mammals.


Spirochetes in the spleen of a patient with chronic Lyme disease.
A 54-year-old man had intermittent evening fever, arthralgia, transient erythematous macular eruption on the skin, and splenomegaly of two year's duration. Immunofluorescence tests for Borrelia burgdorferi serum antibodies had positive results, but G-penicillin treatment was ineffective. Splenectomy with lymph node biopsy was performed to rule out lymphoproliferative disorders. Borrelia-like spirochetes were identified histologically in the spleen; this finding was consistent with persistence of B. burgdorferi organisms in inner organs in chronic Lyme disease.


CD36-specific antibodies block release of HIV-1 from infected primary macrophages and its transmission to T cells.
HIV-1-infected macrophages likely represent viral reservoirs, as they accumulate newly formed virions in internal virus-containing compartments (VCCs). However, the nature and biogenesis of VCCs remain poorly defined. We show that upon HIV-1 infection of primary human macrophages, Gag is recruited to preexisting compartments containing the scavenger receptor CD36, which then become VCCs. Silencing of CD36 in HIV-1-infected macrophages decreases the amount of virions released. Strikingly, soluble anti-CD36 antibodies, but not the natural ligands of CD36, inhibit release of virions from HIV-1-infected macrophages and the transmission of virus to CD4(+) T cells. The effect of the antibodies is potent, rapid, and induces the retention of virions within VCCs. Ectopic expression of CD36 in HeLa cells renders them susceptible to the inhibitory effect of the anti-CD36 mAb upon HIV-1 infection. We show that the anti-CD36 mAb inhibits HIV-1 release by clustering newly formed virions at their site of budding, and that signaling via CD36 is not required. Thus, HIV-1 reservoirs in macrophages may be tackled therapeutically using anti-CD36 antibodies to prevent viral dissemination.


Quantifying charge carrier concentration in ZnO thin films by Scanning Kelvin Probe Microscopy.
In the last years there has been a renewed interest for zinc oxide semiconductor, mainly triggered by its prospects in optoelectronic applications. In particular, zinc oxide thin films are being widely used for photovoltaic applications, in which the determination of the electrical conductivity is of great importance. Being an intrinsically doped material, the quantification of its doping concentration has always been challenging. Here we show how to probe the charge carrier density of zinc oxide thin films by Scanning Kelvin Probe Microscopy, a technique that allows measuring the contact potential difference between the tip and the sample surface with high spatial resolution. A simple electronic energy model is used for correlating the contact potential difference with the doping concentration in the material. Limitations of this technique are discussed in details and some experimental solutions are proposed. Two-dimensional doping concentration images acquired on radio frequency-sputtered intrinsic zinc oxide thin films with different thickness and deposited under different conditions are reported. We show that results inferred with this technique are in accordance with carrier concentration expected for zinc oxide thin films deposited under different conditions and obtained from resistivity and mobility measurements.


Virus-encoded cyclin.
Herpesvirus saimiri contains an open reading frame called eclf2 with homology to the cellular type D cyclins. We now show that the eclf2 gene product is a novel virus-encoded cyclin (v-cyclin). The protein encoded by the v-cyclin gene of this oncogenic herpesvirus was found to have an apparent molecular size of 29 kDa in transformed cells. v-Cyclin protein was found to be associated with cdk6, a cellular cyclin-dependent kinase known to interact with cellular type D cyclins. cdk6/v-cyclin complexes strongly phosphorylated Rb fusion protein and histone H1 as substrates in vitro. Mutational analyses showed that highly conserved amino acids in the cyclin box of v-cyclin were important for association with cdk6 and for activation of cdk6 kinase activity. Thus, v-cyclin resembles cellular type D cyclins in primary sequence, in its association with cdk6, by its ability to activate protein kinase activity, and by the presence of functional cyclin box sequences. v-Cyclin exhibited a selective preference for association with cdk6 over other cyclin-dependent kinases and a high level of kinase activation. The properties of v-cyclin suggest a likely role in oncogenic transformation by this T-lymphotropic herpesvirus.


Stress behaviour and physiology of developing Arctic barnacle goslings ( Branta leucopsis) is affected by legacy trace contaminants.
Natural populations are persistently exposed to environmental pollution, which may adversely impact animal physiology and behaviour and even compromise survival. Responding appropriately to any stressor ultimately might tip the scales for survival, as mistimed behaviour and inadequate physiological responses may be detrimental. Yet effects of legacy contamination on immediate physiological and behavioural stress coping abilities during acute stress are virtually unknown. Here, we assessed these effects in barnacle goslings ( Branta leucopsis) at a historical coal mine site in the Arctic. For three weeks we led human-imprinted goslings, collected from nests in unpolluted areas, to feed in an abandoned coal mining area, where they were exposed to trace metals. As control we led their siblings to feed on clean grounds. After submitting both groups to three well-established stress tests (group isolation, individual isolation, on-back restraint), control goslings behaved calmer and excreted lower levels of corticosterone metabolites. Thus, legacy contamination may decisively change stress physiology and behaviour in long-lived vertebrates exposed at a young age.


Identifying Measures Used for Assessing Quality of YouTube Videos with Patient Health Information: A Review of Current Literature.
Recent publications on YouTube have advocated its potential for patient education. However, a reliable description of what could be considered quality information for patient education on YouTube is missing. To identify topics associated with the concept of quality information for patient education on YouTube in the scientific literature. A literature review was performed in MEDLINE, ISI Web of Knowledge, Scopus, and PsychINFO. Abstract selection was first conducted by two independent reviewers; discrepancies were discussed in a second abstract review with two additional independent reviewers. Full text of selected papers were analyzed looking for concepts, definitions, and topics used by its authors that focused on the quality of information on YouTube for patient education. In total, 456 abstracts were extracted and 13 papers meeting eligibility criteria were analyzed. Concepts identified related to quality of information for patient education are categorized as expert-driven, popularity-driven, or heuristic-driven measures. These include (in descending order): (1) quality of content in 10/13 (77%), (2) view count in 9/13 (69%), (3) health professional opinion in 8/13 (62%), (4) adequate length or duration in 6/13 (46%), (5) public ratings in 5/13 (39%), (6) adequate title, tags, and description in 5/13 (39%), (7) good description or a comprehensive narrative in 4/13 (31%), (8) evidence-based practices included in video in 4/13 (31%), (9) suitability as a teaching tool in 4/13 (31%), (10) technical quality in 4/13 (31%), (11) credentials provided in video in 4/13 (31%), (12) enough amount of content to identify its objective in 3/13 (23%), and (13) viewership share in 2/13 (15%). Our review confirms that the current topics linked to quality of information for patient education on YouTube are unclear and not standardized. Although expert-driven, popularity-driven, or heuristic-driven measures are used as proxies to estimate the quality of video information, caution should be applied when using YouTube for health promotion and patient educational material.


Transforming growth factor beta as endogenous growth inhibitor of chronic lymphocytic leukemia B cells.
Chronic lymphocytic leukemia (CLL) B cells are hyporesponsive or refractory to mitogens and growth factors in vitro. This study examined whether transforming growth factor beta (TGF-beta), a potent inhibitor of lymphocyte proliferation may play a role in the growth regulation of CLL B cells. CLL B cells from all donors treated expressed detectable TGF-beta 1 mRNA. In vitro release of TGF-beta by unstimulated cultures, or cultures stimulated by antibody to cell surface immunoglobulin (anti-mu) plus phorbol 12-myristate 13-acetate (PMA) was higher in CLL than in normal B cells. High levels of TGF-beta activity were also detected in plasma samples of CLL patients. The role of TGF-beta in growth regulation of CLL B cells was tested in assays using different B cell activators. Purified neoplastic B cells from most CLL patients proliferated in response to anti-mu, or the combination of anti-mu plus PMA. Levels of CLL B cell proliferation were lower than observed in normal B cells. Some CLL were refractory to these stimuli. Antibody to CD40 induced proliferation of CLL B cells from all donors tested when presented on Fc gamma RII (CDw32)-expressing L cells. Neutralizing antibodies to TGF-beta increased CLL B cell proliferation in the absence or presence of additional stimuli. These effects were dose dependent and specific. Exogenous TGF-beta completely inhibited CLL B cell proliferation induced by anti-mu, PMA, and anti-TGF-beta. CLL B cell proliferation induced by anti-CD40 was reduced by exogenous TGF-beta. However, even at high doses, TGF-beta did not completely inhibit the anti-CD40 effect. In summary, TGF-beta is overexpressed in CLL. CLL B cells are sensitive to TGF-beta and this cytokine functions as an autocrine growth inhibitor accounting at least in part for reduced proliferative responses of these leukemic cells and for the slow progression of the malignant process in vivo.


Cerebellar volume of musicians.
There is evidence that the cerebellum is involved in motor learning and cognitive function in humans. Animal experiments have found structural changes in the cerebellum in response to long-term motor skill activity. We investigated whether professional keyboard players, who learn specialized motor skills early in life and practice them intensely throughout life, have larger cerebellar volumes than matched non-musicians by analyzing high-resolution T(1)-weighted MR images from a large prospectively acquired database (n = 120). Significantly greater absolute (P = 0.018) and relative (P = 0.006) cerebellar volume but not total brain volume was found in male musicians compared to male non-musicians. Lifelong intensity of practice correlated with relative cerebellar volume in the male musician group (r = 0.595, P = 0.001). In the female group, there was no significant difference noted in volume measurements between musicians and non-musicians. The significant main effect for gender on relative cerebellar volume (F = 10.41, P < 0.01), with females having a larger relative cerebellar volume, may mask the effect of musicianship in the female group. We propose that the significantly greater cerebellar volume in male musicians and the positive correlation between relative cerebellar volume and lifelong intensity of practice represents structural adaptation to long-term motor and cognitive functional demands in the human cerebellum.


Association of native Ca2+ channel beta subunits with the alpha 1 subunit interaction domain.
beta Subunits of voltage-dependent Ca2+ channels play an important role in regulating Ca2+ channel function. The sites of alpha 1-beta subunit interaction have been localized recently to cytoplasmic domains of both subunits. The alpha 1 subunit interaction domain (AID) is an 18-amino-acid conserved motif located between repeats I and II on all alpha 1 subunits which is essential for the binding of beta subunits. In order to further study the interaction of beta subunits with AID, we have expressed a 50-amino-acid glutathione S-transferase (GST) fusion protein from the alpha 1A subunit that contains the AID. Mutant GST fusion proteins that contain a single amino acid change (Y392S, Y392F, and Y392W) in the AIDA along with control GST were coupled to glutathione-Sepharose beads to form affinity beads. Binding assays using these affinity beads with in vitro synthesized 35S-labeled beta 2 and beta 3 subunits demonstrate that the hydroxyl group on tyrosine 392 of AIDA is critical for binding to beta subunits. The affinity bead assay was also used to identify and characterize native beta subunits from detergent extracts of different tissues. The AIDA affinity beads, but not the control or Y392S beads, specifically bind beta subunits from detergent extracts of skeletal muscle, cardiac muscle, and brain. Immunoblot analyses demonstrate the presence of beta 1a in skeletal muscle, beta 2 and beta 3 in cardiac muscle, and beta 1b, beta 3, and beta 4 in brain. The assays also demonstrate the AIDA beads bind to beta subunits from tissue homogenates extracted with low salt and no detergent suggesting the existence of a pool of beta subunits which is not always associated with alpha 1 subunits. Also, beta subunits from solubilized skeletal muscle triads can be affinity-purified using AIDA CNBr-Sepharose. Our data demonstrate that the AID binds to native beta subunits from detergent and non-detergent tissue extracts illustrating that this domain on the alpha 1 subunit is the major anchoring site for the beta subunit.


Psychomotor testing predicts rate of skill acquisition for proficiency-based laparoscopic skills training.
Laparoscopic simulator training translates into improved operative performance. Proficiency-based curricula maximize efficiency by tailoring training to meet the needs of each individual; however, because rates of skill acquisition vary widely, such curricula may be difficult to implement. We hypothesized that psychomotor testing would predict baseline performance and training duration in a proficiency-based laparoscopic simulator curriculum. Residents (R1, n = 20) were enrolled in an IRB-approved prospective study at the beginning of the academic year. All completed the following: a background information survey, a battery of 12 innate ability measures (5 motor, and 7 visual-spatial), and baseline testing on 3 validated simulators (5 videotrainer [VT] tasks, 12 virtual reality [minimally invasive surgical trainer-virtual reality, MIST-VR] tasks, and 2 laparoscopic camera navigation [LCN] tasks). Participants trained to proficiency, and training duration and number of repetitions were recorded. Baseline test scores were correlated to skill acquisition rate. Cutoff scores for each predictive test were calculated based on a receiver operator curve, and their sensitivity and specificity were determined in identifying slow learners. Only the Cards Rotation test correlated with baseline simulator ability on VT and LCN. Curriculum implementation required 347 man-hours (6-person team) and 795,000 dollars of capital equipment. With an attendance rate of 75%, 19 of 20 residents (95%) completed the curriculum by the end of the academic year. To complete training, a median of 12 hours (range, 5.5-21), and 325 repetitions (range, 171-782) were required. Simulator score improvement was 50%. Training duration and repetitions correlated with prior video game and billiard exposure, grooved pegboard, finger tap, map planning, Rey Figure Immediate Recall score, and baseline performance on VT and LCN. The map planning cutoff score proved most specific in identifying slow learners. Proficiency-based laparoscopic simulator training provides improvement in performance and can be effectively implemented as a routine part of resident education, but may require significant resources. Although psychomotor testing may be of limited value in the prediction of baseline laparoscopic performance, its importance may lie in the prediction of the rapidity of skill acquisition. These tests may be useful in optimizing curricular design by allowing the tailoring of training to individual needs.


Resurfacing arthroplasty of the hip in osteopetrosis.
A 34-year-old woman with a benign form of osteopetrosis developed osteoarthritis of the hip. In order to avoid the difficulties associated with inserting the femoral component of a conventional total hip arthroplasty, a hybrid metal-on-metal resurfacing was performed. There were several technical challenges associated with the procedure, including the sizing of the component, press-fit fixation of the acetabular component and femoral head preparation, as well as trying to avoid a fracture. No surgical complication occurred. After more than a year following surgery, the patient showed excellent clinical function and remained satisfied with the outcome. We conclude that the hybrid metal-on-metal resurfacing arthroplasty represents a valuable option for the treatment of patients with osteopetrosis and secondary hip osteoarthritis.


Recruiting trainees for a global health research workforce: the National Institutes of Health Fogarty International Clinical Research Scholars Program selection process.
Between 2004 and 2012, the National Institutes of Health Fogarty International Clinical Research Scholars (FICRS) Program provided 1-year mentored research training at low- and middle-income country sites for American and international health science doctoral students. We describe the centralized application process, US applicant characteristics, and predictors of selection/enrollment. FICRS received 1,084 applicants representing many health professions and biomedical disciplines at 132 US academic institutions; 219 students from 72 institutions were accepted and enrolled. Medical/osteopathic students comprised 88.9% of applicants and 85.8% of enrollees. Applicants from institutions with higher applicant numbers were two times as likely to be selected. In 2012, FICRS was decentralized among 20 institutions in five consortia (Global Health Fellows), with autonomous selection processes that emphasize post-doctoral trainees. If academia, government, or charitable foundations offer future opportunities to health professions students for international research, the FICRS experience predicts that they can attract substantial numbers of motivated trainees from diverse backgrounds.


Value of MR contrast media in image-guided body interventions.
In the past few years, there have been multiple advances in magnetic resonance (MR) instrumentation, in vivo devices, real-time imaging sequences and interventional procedures with new therapies. More recently, interventionists have started to use minimally invasive image-guided procedures and local therapies, which reduce the pain from conventional surgery and increase drug effectiveness, respectively. Local therapy also reduces the systemic dose and eliminates the toxic side effects of some drugs to other organs. The success of MR-guided procedures depends on visualization of the targets in 3D and precise deployment of ablation catheters, local therapies and devices. MR contrast media provide a wealth of tissue contrast and allows 3D and 4D image acquisitions. After the development of fast imaging sequences, the clinical applications of MR contrast media have been substantially expanded to include pre- during- and post-interventions. Prior to intervention, MR contrast media have the potential to localize and delineate pathologic tissues of vital organs, such as the brain, heart, breast, kidney, prostate, liver and uterus. They also offer other options such as labeling therapeutic agents or cells. During intervention, these agents have the capability to map blood vessels and enhance the contrast between the endovascular guidewire/catheters/devices, blood and tissues as well as direct therapies to the target. Furthermore, labeling therapeutic agents or cells aids in visualizing their delivery sites and tracking their tissue distribution. After intervention, MR contrast media have been used for assessing the efficacy of ablation and therapies. It should be noted that most image-guided procedures are under preclinical research and development. It can be concluded that MR contrast media have great value in preclinical and some clinical interventional procedures. Future applications of MR contrast media in image-guided procedures depend on their safety, tolerability, tissue specificity and effectiveness in demonstrating success of the interventions and therapies.


Biocontrol of Listeria monocytogenes on fresh-cut produce by treatment with lytic bacteriophages and a bacteriocin.
The fresh-cut produce industry has been the fastest-growing portion of the food retail market during the past 10 years, providing consumers with convenient and nutritious food. However, fresh-cut fruits and vegetables raise food safety concerns, because exposed tissue may be colonized more easily by pathogenic bacteria than intact produce. This is due to the higher availability of nutrients on cut surfaces and the greater potential for contamination because of the increased amount of handling. We found that applied Listeria monocytogenes populations survived and increased only slightly on fresh-cut Red Delicious apples stored at 10 degrees C but increased significantly on fresh-cut honeydew melons stored at 10 degrees C over 7 days. In addition, we examined the effect of lytic, L. monocytogenes-specific phages via two phage application methods, spraying and pipetting, on L. monocytogenes populations in artificially contaminated fresh-cut melons and apples. The phage mixture reduced L. monocytogenes populations by 2.0 to 4.6 log units over the control on honeydew melons. On apples, the reduction was below 0.4 log units. In combination with nisin (a bacteriocin), the phage mixture reduced L. monocytogenes populations by up to 5.7 log units on honeydew melon slices and by up to 2.3 log units on apple slices compared to the control. Nisin alone reduced L. monocytogenes populations by up to 3.2 log units on honeydew melon slices and by up to 2.0 log units on apple slices compared to the control. The phage titer was stable on melon slices, but declined rapidly on apple slices. The spray application of the phage and phage plus nisin reduced the bacterial numbers at least as much as the pipette application. The effectiveness of the phage treatment also depended on the initial concentration of L. monocytogenes.


Measurement of endothelial function and its clinical utility for cardiovascular risk.
Over the past two decades, the central role of the endothelium in the initiation, progression, and clinical sequelae of atherosclerosis has been increasingly recognized. Assessment of the pathobiology of the endothelium and its ability to act as a potential therapeutic target remains an area of active research interest. Whilst endothelial function has been shown to be a marker for risk of cardiovascular events in high-risk groups, there remains considerable debate about the most appropriate way to assess this. We discuss the different clinical methods to assess endothelial function, focusing on flow-mediated dilatation (FMD) of the brachial artery, highlighting the importance of using a standardized methodology, as well as discussing the clinical limitations of using FMD in individuals.


Cost-effectiveness of varenicline compared with nicotine patches for smoking cessation--results from four European countries.
The aim of this study was to evaluate and compare the cost-effectiveness of varenicline with nicotine replacement therapy (NRT) for smoking cessation in four European countries (Belgium, France, Sweden and the UK). Markov simulations, using the Benefits of Smoking Cessation on Outcomes (BENESCO) model, were performed. We simulated the incidence of four smoking-related morbidities: lung cancer, chronic obstructive pulmonary disease, coronary heart disease and stroke. The model computes quality-adjusted life-years gained and incremental cost-effectiveness ratios. Incremental cost-utility ratios were calculated, adopting a lifetime perspective. Efficacy data were obtained from a randomized open-label trial: Week 52 continuous abstinence rates were 26.1% for varenicline and 20.3% for NRT. The analyses imply that for countries analysed, smoking cessation using varenicline versus NRT was associated with reduced smoking-related morbidity and mortality. The number of morbidities avoided, per 1000 smokers attempting to quit, ranged from 9.7 in Belgium to 6.5 in the UK. The number of quality-adjusted life-years gained, per 1000 smokers, was 23 (Belgium); 19.5 (France); 29.9 (Sweden); and 23.7 (UK). In all base-case simulations (except France), varenicline dominated (more effective and cost saving) NRT regarding costs per quality-adjusted life-year gained; for France the incremental cost-effectiveness ratio was 2803. This cost-effectiveness analysis demonstrated that since varenicline treatment was more effective, the result was increased healthcare cost savings in Belgium, Sweden and the UK. Our results suggest that funding varenicline as a smoking cessation aid is justifiable from a healthcare resource allocation perspective.


Oncogenic K-Ras and loss of Smad4 mediate invasion by activating an EGFR/NF-κB Axis that induces expression of MMP9 and uPA in human pancreas progenitor cells.
Activating K-Ras mutations and inactivating mutations of Smad4 are two common genetic alterations that occur in the development and progression of pancreatic ductal adenocarcinomas (PDAC). To further study the individual and combinatorial roles of these two mutations in the pathogenesis of PDAC, immortalized human pancreas nestin postive cells (HPNE) were genetically modified by either expressing oncogenic K-Ras (HPNE/K-Ras), by shRNA knock down of Smad4 (HPNE/ShSmad4) or by creating both alterations in the same cell line (HPNE/K-Ras/ShSmad4). We previously found that expression of oncogenic K-Ras caused an increase in expression of EGFR and loss of Smad4 further enhanced the up regulation in expression of EGFR and that this increase in EGFR was sufficient to induce invasion. Here we further investigated the mechanism that links mutational alterations and EGFR expression with invasion. The increase in EGFR signaling was associated with up regulation of MMP9 and uPA protein and activity. Moreover, the increase in EGFR signaling promoted a nuclear translocation and binding of RelA (p65), a subunit of NF-κB, to the promoters of both MMP-9 and uPA. Treatment of HPNE/K-Ras/ShSmad4 cells with an inhibitor of EGFR reduced EGF-mediated NF-κB nuclear translocation and inhibitors of either EGFR or NF-κB reduced the increase in MMP-9 or uPA expression. In conclusion, this study provides the mechanism of how a combination of oncogenic K-Ras and loss of Smad4 causes invasion and provides the basis for new strategies to inhibit metastases.


Understanding decomposition and encapsulation energies of structure I and II clathrate hydrates.
When compressed with water or ice under high pressure and low temperature conditions, some gases form solid gas hydrate inclusion compounds which have higher melting points than ice under those pressures. In this work, we study the balance of the guest-water and water-water interaction energies that lead to the formation of the clathrate hydrate phases. In particular, molecular dynamics simulations with accurate water potentials are used to study the energetics of the formation of structure I (sI) and II (sII) clathrate hydrates of methane, ethane, and propane. The dissociation enthalpy of the clathrate hydrate phases, the encapsulation enthalpy of methane, ethane, and propane guests in the corresponding phases, and the average bonding enthalpy of water molecules are calculated and compared with accurate calorimetric measurements and previous classical and quantum mechanical calculations, when available. The encapsulation energies of methane, ethane, and propane guests stabilize the small and large sI and sII hydrate cages, with the larger molecules giving larger encapsulation energies. The average water-water interactions are weakened in the sI and sII phases compared to ice. The relative magnitudes of the van der Waals potential energy in ice and the hydrate phases are similar, but in the ice phase, the electrostatic interactions are stronger. The stabilizing guest-water "hydrophobic" interactions compensate for the weaker water-water interactions and stabilize the hydrate phases. A number of common assumptions regarding the guest-cage water interactions are used in the van der Waals-Platteeuw statistical mechanical theory to predict the clathrate hydrate phase stability under different pressure-temperature conditions. The present calculations show that some of these assumptions may not accurately reflect the physical nature of the interactions between guest molecules and the lattice waters.


Cost-effectiveness of two screening strategies for Chlamydia trachomatis and Neisseria gonorrhoeae as part of the PrEP programme in the Netherlands: a modelling study.
Pre-exposure prophylaxis (PrEP) users are routinely tested four times a year (3 monthly) for asymptomatic Chlamydia trachomatis (CT) and Neisseria gonorrhoeae (NG) infections on three anatomical locations. Given the high costs of this testing to the PrEP programme, we assessed the impact of 3 monthly screening(current practice), compared with 6 monthly on the disease burden. We quantified the difference in impact of these two testing frequencies on the prevalence of CT and NG among all men who have sex with men (MSM) who are at risk of an STI, and explored the cost-effectiveness of 3-monthly screening compared with a baseline scenario of 6-monthly screening. A dynamic infection model was developed to simulate the transmission of CT and NG among sexually active MSM (6500 MSM on PrEP and 29 531 MSM not on PrEP), and the impact of two different test frequencies over a 10-year period. The difference in number of averted infections was used to calculate incremental costs and quality-adjusted life-years (QALY) as well as an incremental cost-effectiveness ratio (ICER) from a societal perspective. Compared with 6-monthly screening, 3-monthly screening of PrEP users for CT and NG cost an additional €46.8 million over a period of 10 years. Both screening frequencies would significantly reduce the prevalence of CT and NG, but 3-monthly screening would avert and extra ~18 250 CT and NG infections compared with 6-monthly screening, resulting in a gain of ~81 QALYs. The corresponding ICER was ~€430 000 per QALY gained, which exceeded the cost-effectiveness threshold of €20 000 per QALY. Three-monthly screening for CT and NG among MSM on PrEP is not cost-effective compared with 6-monthly screening. The ICER becomes more favourable when a smaller fraction of all MSM at risk for an STI are screened. Reducing the screening frequency could be considered when the PrEP programme is established and the prevalence of CT and NG decline.


Marital Transitions and Depressive Symptoms among Older Adults: Examining Educational Differences.
Later decades of the life course have undergone rapid transformations due to demographic changes in ageing societies, such as more frequent occurrences of later-life marital transitions. Adaption to these transitions, even when welcomed, brings novel chances and challenges in negotiating new social roles in old age, which could reinforce preexisting disparities in the acquisition and mastery of resources, social ties, and coping strategies. Because the ability to weather later-life marital transitions may depend on the long arm of education acquired earlier in the life course, the present study aims to identify and track trends in the prevalence of marriage, divorce/separation, and widowhood among sociodemographic subgroups; link the occurrence of those transitions with mental health; and test the influence of educational attainment on these associations. We employ an intraindividual, within-person approach to quantify the occurrence of marital transitions and their impact using data from the Health and Retirement Study (n = 22,013; 1992-2010). Measures of transition occurrence, depressive symptoms, and educational attainment were available across up to 10 biennial assessments. Individuals with less than a high school diploma displayed the highest likelihood of losing their significant other through divorce/separation or death. Marital loss was associated with increasing, and marital gain with decreasing, depressive symptoms. Compared to those with less than a high school diploma, individuals with a high school or general equivalency diploma exhibited larger increases in depressive symptoms associated with widowhood, even though their average levels of depressive symptoms were lower in the absence of this transition. Our findings revealed a predictable educational gradient for the occurrence of marital transitions and later-life mental health. Yet higher, formalized education did not protect the participants from increased depression in the presence of a loss-related transition, which could suggest that the strains of spousal loss may to some degree function as a leveler of the preexisting social inequalities of stratified life courses. We conclude that the benefits conferred by education are not necessarily ubiquitous, and its impact on the adaptation to spousal loss may be more complex and nuanced depending on the range of prior experiences and available coping strategies.


G protein-dependent inhibition of L-type Ca2+ currents by acetylcholine in mouse pancreatic B-cells.
1. The effect of acetylcholine (ACh) on voltage-dependent Ca2+ currents in mouse pancreatic B-cells was studied using the whole-cell configuration of the patch-clamp technique. 2. ACh (0.25-250 microM) reversibly and dose-dependently inhibited the Ca2+ current elicited by depolarizations from -80 mV to +10 mV. Maximal inhibition was observed at concentrations > 25 microM where it amounted to approximately 35%. The effect was voltage independent and prevented by atropine (10 microM) suggesting that it was mediated by muscarinic receptors. 3. The inhibitory action of ACh on the Ca2+ current was abolished when the cytoplasmic solution contained GDP beta S (2 mM) and became irreversible when the non-hydrolysable GTP analogue GTP gamma S (10 microM) was included in the pipette. This indicates the participation of G proteins in the inhibitory effect of ACh but pretreatment of the cells with either pertussis or cholera toxin failed to prevent the effect of ACh on the Ca2+ current. 4. ACh remained equally effective as an inhibitor of the whole-cell Ca2+ current in the presence of the L-type Ca2+ channel agonist (-)-Bay K 8644 and after partial inhibition of the current by nifedipine. Addition of omega-agatoxin IVA, omega-conotoxin GVIA or omega-conotoxin MVIIC neither affected the peak Ca2+ current amplitude nor the extent of inhibition produced by ACh. These pharmacological properties indicate that ACh acts by inhibiting L-type Ca2+ channels. 5. The inhibitory action of ACh on the B-cell Ca2+ current was not secondary to elevation of [Ca2+]i and ACh remained equally effective as an inhibitor when Ba2+ was used as the charge carrier, when [Ca2+]i was buffered to low concentrations using EGTA and under experimental conditions preventing the mobilization of Ca2+ from intracellular stores. 6. These results suggest that ACh reduces the whole-cell Ca2+ current in the B-cell through a G protein-regulated, voltage- and Ca(2+)-independent inhibition of L-type Ca2+ channels.


Evidence that ursodeoxycholic acid prevents steroid-resistant rejection in adult liver transplantation.
Steroid-resistant rejection continues to be a serious problem in liver transplantation. Since ursodeoxycholic acid (UDCA) is beneficial in several cholestatic disorders and possesses in vitro immunomodulatory and immunosuppressive effects, we have tested in a pilot study the effects of adjuvant UDCA in the prevention of steroid-resistant rejection. Fifty consecutive liver transplant patients were treated with a standard cyclosporine immunosuppressive regimen. Treatment with UDCA (10 mg/kg/d) was initiated in each patient who developed biopsy-proven rejection or biochemical evidence of cholestasis. Clinical and laboratory features were monitored for evidence of rejection. Data were analyzed after at least a 10-month follow-up period was available in each patient. Seven patients died during the study period, all within 4 wk of surgery. No evidence of rejection was documented in these patients. Twenty three of the 43 survivors 53% (23/43) developed an episode of rejection, and UDCA was initiated in each of them. Only one patient had a second episode of rejection, which responded to intravenous methylprednisolone therapy; no patient required antilymphocyte therapy. There was no evidence of toxicity for UDCA. These data suggest that UDCA can be given safely following OLT and may contribute to prevention of steroid-resistant rejection in liver transplant recipients.


Scanning electron microscopic studies of the oral tissue responses to dental implants.
Scanning electron microscopy (SEM) and its associated technologies have proven invaluable in elucidating the interfacial oral tissue responses to dental implants. Since the dental implant must extend from the mandibular or maxillary jaw, through the oral mucosa, and into the oral cavity, these tissue responses include epithelium, connective tissue and bone. The continual occlusal forces acting upon these tissues reinforce the dynamic character of these tissue responses. Immediately upon implantation, a healing phase begins as a response to the implanted biomaterial. Following this immediate response a longer healing phase occurs, beginning approximately 1 week after implantation, resulting in the modeling of bone to the implant as well as the formation of epithelial attachment to the implant. This later, delayed healing continues throughout the lifetime of the implant since these tissues must die and be replaced by similar tissues. Current dental research employing scanning electron microscopy is now documenting these tissue responses. This paper reviews, in detail, SEM observations of these tissue responses.


Sustained Long-Term Outcomes With Closed-Loop Spinal Cord Stimulation: 12-Month Results of the Prospective, Multicenter, Open-Label Avalon Study.
Spinal cord stimulation (SCS) activates the dorsal column fibers using electrical stimuli. Current SCS systems function in fixed-output mode, delivering the same stimulus regardless of spinal cord (SC) activation. To present long-term outcomes of a novel closed-loop SCS system that aims to maintain the SC activation near a set target level and within a therapeutic window for each patient. SC activation is measured through the evoked compound action potential (ECAP) generated by each stimulus pulse. Fifty patients with lower back and/or leg pain who were successfully trialed received a permanent system (Evoke; Saluda Medical, Sydney, Australia). Ratings of pain (visual analog scale), quality of life, function, sleep, and medication use were collected at baseline and at each visit. SC activation levels were reported in summary statistics. The therapeutic window for each individual patient was defined as the range of ECAP amplitudes between sensation threshold and uncomfortably strong stimulation. At 12 mo, the proportion of patients with ≥50% relief was 76.9% (back), 79.3% (leg), and 81.4% (overall), and the proportion with ≥80% pain relief was 56.4% (back), 58.6% (leg), and 53.5% (overall). Patients spent a median of 84.9% of their time with stimulation in their therapeutic window, and 68.8% (22/32) eliminated or reduced their opioid intake. Statistically significant improvements in secondary outcomes were observed. The majority of patients experienced more than 80% pain relief with stable SC activation, as measured by ECAP amplitude at 12 mo, providing evidence for the long-term effectiveness of the Evoke closed-loop SCS system.


Changing how we evaluate research is difficult, but not impossible.
The San Francisco Declaration on Research Assessment (DORA) was published in 2013 and described how funding agencies, institutions, publishers, organizations that supply metrics, and individual researchers could better evaluate the outputs of scientific research. Since then DORA has evolved into an active initiative that gives practical advice to institutions on new ways to assess and evaluate research. This article outlines a framework for driving institutional change that was developed at a meeting convened by DORA and the Howard Hughes Medical Institute. The framework has four broad goals: understanding the obstacles to changes in the way research is assessed; experimenting with different approaches; creating a shared vision when revising existing policies and practices; and communicating that vision on campus and beyond.


Aldosterone blockade in heart failure.
Aldosterone plays a key role in the pathophysiology of heart failure. Angiotensin converting enzyme inhibitors and angiotensin II receptor blockers may not suppress aldosterone production in the long term. This allows aldosterone to exert its effects on myocardial fibrosis and cardiac remodelling, endothelial function, electrolytes and baroreceptor response. The Randomized Aldactone Evaluation Study (RALES) tested spironolactone against placebo in patients with severe heart failure. The study found a 30% reduction in the risk of death among patients treated with spironolactone and a 31% reduction in the risk of death from cardiac causes. Patients in the spironolactone group had significantly lower risks of death from progression of heart failure and sudden cardiac death. The Eplerenone Post-Acute Myocardial Infarction Heart Failure Efficacy and Survival Study (EPHESUS) investigated the effects of eplerenone against placebo in patients with myocardial infarction complicated by left ventricular dysfunction. Compared to placebo, the relative risk of death from any cause was 0.85 in eplerenone-treated patients, and the relative risk of death or hospitalisation for cardiovascular events was 0.87. The reduction in the risk of sudden death from cardiac causes was statistically significant. In conclusion, aldosterone blockade should form part of optimal therapy for patients with heart failure.


Tandem 18O stable isotope labeling for quantification of N-glycoproteome.
A new strategy using tandem (18)O stable isotope labeling (TOSIL) to quantify the N-glycosylation site occupancy is reported. Three heavy oxygen atoms are introduced into N-glycosylated peptides: two (18)O atoms are incorporated into the carboxyl terminal of all peptides during a tryptic digestion, and the third (18)O atom is incorporated into the N-glycosylation site of asparagines-linked sugar chains specifically via a N-glycosidase F (PNGase F)-mediated hydrolysis. Comparing samples treated in H(2)(18)O and samples treated in H(2)(16)O, a unique mass shift of 6 Da can be shown for N-glycosylated peptide with single glycosylation site, which could be easily distinguished from those nonglycosite peptide pairs with a mass difference of 4 Da only. The relative quantities of N-glycosylated and its parent protein-levels were obtained simultaneous by measuring the intensity ratios of (18)O/(16)O for glycosylated (6 Da) and for nonglycosylated (4 Da) peptides, respectively. Thus, a comparison of these two ratios can be utilized to evaluate the changes of occupancy of N-glycosylation at specific sites between healthy and diseased individuals. The TOSIL approach yielded good linearity in quantitative response within 10-fold dynamic range with the correlation coefficient r(2) > 0.99. The standard deviation (SD) ranged from 0.06 to 0.21, for four glycopeptides from two model glycoproteins. Furthermore, serums from a patient with ovarian cancer and healthy individual were used as test examples to validate the novel TOSIL method. A total of 86 N-glycosylation sites were quantified and N-glycosylation levels of 56 glycopeptides showed significant changes. Most changes in N-glycosylation at specific sites have the same trends as those of protein expression levels; however, the occupancies of three N-glycosylation sites were significantly changed with no change in proteins levels.


Temporomandibular joint disorders.
Temporomandibular joint disorders are common in adults; as many as one third of adults report having one or more symptoms, which include jaw or neck pain, headache, and clicking or grating within the joint. Most symptoms improve without treatment, but various noninvasive therapies may reduce pain for patients who have not experienced relief from self-care therapies. Physical therapy modalities (e.g., iontophoresis, phonophoresis), psychological therapies (e.g., cognitive behavior therapy), relaxation techniques, and complementary therapies (e.g., acupuncture, hypnosis) are all used for the treatment of temporomandibular joint disorders; however, no therapies have been shown to be uniformly superior for the treatment of pain or oral dysfunction. Noninvasive therapies should be attempted before pursuing invasive, permanent, or semi-permanent treatments that have the potential to cause irreparable harm. Dental occlusion therapy (e.g., oral splinting) is a common treatment for temporomandibular joint disorders, but a recent systematic review found insufficient evidence for or against its use. Some patients with intractable temporomandibular joint disorders develop chronic pain syndrome and may benefit from treatment, including antidepressants or cognitive behavior therapy.


The spectrum of beta-thalassaemia mutations on the Indian subcontinent: the basis for prenatal diagnosis.
The beta-thalassaemia mutations in 702 unrelated carriers originating from seven different regions of the Indian subcontinent have been characterized using allele specific priming of the polymerase chain reaction (PCR). It was possible to identify the mutations in 688 (98%) of the individuals studied. Eleven different mutations were identified, of which five common ones accounted for 93.6%; namely the ones at IVS-1 position 5 (G-C), codons 8/9 (+G), IVS-1 position 1 (G-T), codons 41/42 (-CTTT) and the 619 bp deletion at the 3' end of the gene. The mutations at IVS-2 position 1 (G-A) and codon 30 (G-C), previously undescribed in Asian Indians, were found in two and six individuals respectively. Some regional variation in the distribution of beta-thalassaemia alleles was noted. These findings should prove useful for the development of a first trimester prenatal diagnosis programme based on direct detection of mutations.


The orphan nuclear receptor REV-ERBalpha controls circadian transcription within the positive limb of the mammalian circadian oscillator.
Mammalian circadian rhythms are generated by a feedback loop in which BMAL1 and CLOCK, players of the positive limb, activate transcription of the cryptochrome and period genes, components of the negative limb. Bmal1 and Per transcription cycles display nearly opposite phases and are thus governed by different mechanisms. Here, we identify the orphan nuclear receptor REV-ERBalpha as the major regulator of cyclic Bmal1 transcription. Circadian Rev-erbalpha expression is controlled by components of the general feedback loop. Thus, REV-ERBalpha constitutes a molecular link through which components of the negative limb drive antiphasic expression of components of the positive limb. While REV-ERBalpha influences the period length and affects the phase-shifting properties of the clock, it is not required for circadian rhythm generation.


Suppressor mutations within the core binding factor (CBF/AML1) binding site of a T-cell lymphomagenic retrovirus.
The transcriptional enhancer of the lymphomagenic mouse retrovirus SL3 contains a binding site for the transcription factor core binding factor (CBF; also called AML1, PEBP2, and SEF1). The SL3 CBF binding site is called the core. It differs from the core of the weakly lymphomagenic mouse retrovirus Akv by one nucleotide (the sequences are TGTGGTTAA and TGTGGTCAA, respectively). A mutant virus called SAA that was identical to SL3 except that its core was mutated to the Akv sequence was only moderately attenuated for lymphomagenicity. In most SAA-infected mice, tumor proviruses contained either reversions of the original mutation or one of two novel core sequences. In 20% of the SAA-infected mice, tumor proviruses retained the original SAA/Akv core mutation but acquired one of two additional mutations (underlined), TGCGGTCAA or TGTGGTCTA, that generated core elements called So and T*, respectively. We tested whether the novel base changes in the So and T* cores were suppressor mutations. SL3 mutants that contained So or T* cores in place of the wild-type sequence were generated. These viruses induced T-cell lymphomas in mice more quickly than SAA. Therefore, the mutations in the So and T* cores are indeed second-site suppressor mutations. The suppressor mutations increased CBF binding in vitro and transcriptional activity of the viral long terminal repeats (LTRs) in T lymphocytes to levels comparable to those of SL3. Thus, CBF binding was increased by any of three different nucleotide changes within the sequence of the SAA core. Increased CBF binding resulted in increased LTR transcriptional activity in T cells and in increased viral lymphomagenicity.


Molecular characterization of the microbial communities in the subcaudal gland secretion of the European badger (Meles meles).
Many mammals possess specialized scent glands, which convey information about the marking individual. As the chemical profile of scent marks is likely to be affected by bacteria metabolizing the primary gland products, the variation in bacterial communities between different individuals has been proposed to underpin olfactory communication. However, few studies have investigated the dependency of microbiota residing in the scent organs on the host's individual-specific parameters. Here, we used terminal restriction fragment length polymorphism analysis of the PCR-amplified 16S rRNA gene and clone library construction to investigate the microbial communities in the subcaudal gland secretion of the European badger (Meles meles). As the secretion has been shown to encode individual-specific information, we investigated the correlation of the microbiota with different individual-specific parameters (age, sex, body condition, reproductive status, and season). We discovered a high number of bacterial species (56 operational taxonomic units from four phyla: Actinobacteria, Firmicutes, Proteobacteria, and Bacteroidetes), dominated by Actinobacteria (76.0%). The bacterial communities of cubs and adults differed significantly. Cubs possessed considerably more diverse communities dominated by Firmicutes, while in adults the communities were less diverse and dominated by Actinobacteria, suggesting that the acquisition of a 'mature bacterial community' is an ontogenetic process related to physiological changes during maturation.


Driver Visual Attention Before and After Take-Over Requests During Automated Driving on Public Roads.
This study aims to understand drivers' visual attention before and after take-over requests during automated driving (AD), when the vehicle is fully responsible for the driving task on public roads. Existing research on transitions of control from AD to manual driving has mainly focused on take-over times. Despite its relevance for vehicle safety, drivers' visual attention has received little consideration. Thirty participants took part in a Wizard of Oz study on public roads. Drivers' visual attention was analyzed before and after four take-over requests. Visual attention during manual driving was also recorded to serve as a baseline for comparison. During AD, the participants showed reduced visual attention to the forward road and increased duration of single off-road glances compared to manual driving. In response to take-over requests, the participants looked away from the forward road toward the instrument cluster. Levels of visual attention towards the forward road did not return to the levels observed during manual driving until after 15 s had passed. During AD, drivers may look toward non-driving related task items (e.g., mobile phone) instead of forward. Further, when a transition of control is required, drivers may take over control before they are aware of the driving environment or potential threat(s). Thus, it cannot be assumed that drivers are ready to respond to events shortly after the take-over request. It is important to consider the effect of the design of take-over requests on drivers' visual attention alongside take-over times.


Pervasive haplotypic variation in the spliceo-transcriptome of the human major histocompatibility complex.
The human major histocompatibility complex (MHC) on chromosome 6p21 is a paradigm for genomics, showing remarkable polymorphism and striking association with immune and non-immune diseases. The complex genomic landscape of the MHC, notably strong linkage disequilibrium, has made resolving causal variants very challenging. A promising approach is to investigate gene expression levels considered as tractable intermediate phenotypes in mapping complex diseases. However, how transcription varies across the MHC, notably relative to specific haplotypes, remains unknown. Here, using an original hybrid tiling and splice junction microarray that includes alternate allele probes, we draw the first high-resolution strand-specific transcription map for three common MHC haplotypes (HLA-A1-B8-Cw7-DR3, HLA-A3-B7-Cw7-DR15, and HLA-A26-B18-Cw5-DR3-DQ2) strongly associated with autoimmune diseases including type 1 diabetes, systemic lupus erythematosus, and multiple sclerosis. We find that haplotype-specific differences in gene expression are common across the MHC, affecting 96 genes (46.4%), most significantly the zing finger protein gene ZFP57. Differentially expressed probes are correlated with polymorphisms between haplotypes, consistent with cis effects that we directly demonstrate for ZFP57 in a cohort of healthy volunteers (P = 1.2 × 10(-14)). We establish that alternative splicing is significantly more frequent in the MHC than genome-wide (72.5% vs. 62.1% of genes, P ≤ 1 × 10(-4)) and shows marked haplotypic differences. We also unmask novel and abundant intergenic transcription involving 31% of transcribed blocks identified. Our study reveals that the renowned MHC polymorphism also manifests as transcript diversity, and our novel haplotype-based approach marks a new step toward identification of regulatory variants involved in the control of MHC-associated phenotypes and diseases.


Between-city variation in frequency of injection among Puerto Rican injection drug users: East Harlem, New York, and Bayamon, Puerto Rico.
Frequency of injection has been consistently found to be higher among Puerto Rican Injection drug users (IDUs) than among other groups of IDUs. Several explanations have been suggested, but an empirical explanation has yet to be presented. This study compares the frequency of injection of Puerto Rican IDUs in East Harlem, New York, with that of IDUs in Bayamon, Puerto Rico. Study subjects comprised 521 Puerto Rican IDUs from East Harlem and 303 IDUs from Bayamon. The mean frequency of injection among IDUs in East Harlem was 2.8, the corresponding mean in Bayamon was 5.4. Younger IDUs reported a higher number of daily injection episodes than older IDUs, and the IDU group in Bayamon was 5 years younger than the group in East Harlem. The drug use variables accounted for a greater portion of the between-city difference than the demographic and psychosocial variables. Use of noninjected drugs, as well as the use of prescribed methadone, were found to be associated with a lower number of daily injections. Conversely, injection of cocaine, injection of cocaine mixed with heroin ("speedball"), and injection of larger amounts of drug solution were found to be associated with a higher number of daily injections.


A miR172 target-deficient AP2-like gene correlates with the double flower phenotype in roses.
One of the well-known floral abnormalities in flowering plants is the double-flower phenotype, which corresponds to flowers that develop extra petals, sometimes even containing entire flowers within flowers. Because of their highly priced ornamental value, spontaneous double-flower variants have been found and selected for in a wide range of ornamental species. Previously, double flower formation in roses was associated with a restriction of AGAMOUS expression domain toward the centre of the meristem, leading to extra petals. Here, we characterized the genomic region containing the mutation associated with the switch from simple to double flowers in the rose. An APETALA2-like gene (RcAP2L), a member of the Target Of EAT-type (TOE-type) subfamily, lies within this interval. In the double flower rose, two alleles of RcAP2L are present, one of which harbours a transposable element inserted into intron 8. This insertion leads to the creation of a miR172 resistant RcAP2L variant. Analyses of the presence of this variant in a set of simple and double flower roses demonstrate a correlation between the presence of this allele and the double flower phenotype. These data suggest a role of this miR172 resistant RcAP2L variant in regulating RcAGAMOUS expression and double flower formation in Rosa sp.


Genome-wide analysis reveals a highly diverse CD8 T cell response to murine cytomegalovirus.
Human CMV establishes a lifelong latent infection in the majority of people worldwide. Although most infections are asymptomatic, immunocompetent hosts devote an extraordinary amount of immune resources to virus control. To increase our understanding of CMV immunobiology in an animal model, we used a genomic approach to comprehensively map the C57BL/6 CD8 T cell response to murine CMV (MCMV). Responses to 27 viral proteins were detectable directly ex vivo, the most diverse CD8 T cell response yet described within an individual animal. Twenty-four peptide epitopes were mapped from 18 Ags, which together account for most of the MCMV-specific response. Most Ags were from genes expressed at early times, after viral genes that interfere with Ag presentation are expressed, consistent with the hypothesis that the CD8 T cell response to MCMV is largely driven by cross-presented Ag. Titration of peptide epitopes in a direct ex vivo intracellular cytokine staining assay revealed a wide range of functional avidities, with no obvious correlation between functional avidity and the strength of the response. The immunodominance hierarchy varied only slightly between mice and between experiments. However, H-2(b)-expressing mice with different genetic backgrounds responded preferentially to different epitopes, indicating that non-MHC-encoded factors contribute to immunodominance in the CD8 T cell response to MCMV.