bluuebunny/crossref_metadata_2025 huggingface dataset:
Individual sympathetic postganglionic neurons coinnervate myenteric ganglia and smooth muscle layers in the gastrointestinal tract of the rat
Authors: Walter Gary C. , Phillips Robert J. , McAdams Jennifer L. , Powley Terry L.
Published: February 2016

A full description of the terminal architecture of sympathetic axons innervating the gastrointestinal (GI) tract has not been available. To label sympathetic fibers projecting to the gut muscle wall, dextran biotin was injected into the celiac and superior mesenteric ganglia (CSMG) of rats. Nine days postinjection, animals were euthanized and stomachs and small intestines were processed as whole mounts (submucosa and mucosa removed) to examine CSMG efferent terminals. Myenteric neurons were counterstained with Cuprolinic Blue; catecholaminergic axons were stained immunohistochemically for tyrosine hydroxylase. Essentially all dextran‐labeled axons (135 of 136 sampled) were tyrosine hydroxylase‐positive. Complete postganglionic arbors (n = 154) in the muscle wall were digitized and analyzed morphometrically. Individual sympathetic axons formed complex arbors of varicose neurites within myenteric ganglia/primary plexus and, concomitantly, long rectilinear arrays of neurites within circular muscle/secondary plexus or longitudinal muscle/tertiary plexus. Very few CSMG neurons projected exclusively (i.e., ∼100% of an arbor's varicose branches) to myenteric plexus (∼2%) or smooth muscle (∼14%). With less stringent inclusion criteria (i.e., ≥85% of an axon's varicose branches), larger minorities of neurons projected predominantly to either myenteric plexus (∼13%) or smooth muscle (∼27%). The majority (i.e., ∼60%) of all individual CSMG postganglionics formed mixed, heterotypic arbors that coinnervated extensively (>15% of their varicose branches per target) both myenteric ganglia and smooth muscle. The fact that ∼87% of all sympathetics projected either extensively or even predominantly to smooth muscle, while simultaneously contacting myenteric plexus, is consistent with the view that these neurons control GI muscle directly, if not exclusively. J. Comp. Neurol. 524:2577–2603, 2016. © 2016 Wiley Periodicals, Inc.
Tectothalamic inhibitory projection neurons in the avian torus semicircularis
Authors: Ito Tetsufumi , Atoji Yasuro
Published: February 2016

Inhibitory feedforward projection is one of key features of the organization of the central auditory system. In mammals, the inferior colliculus (IC) is the origin of a substantial inhibitory feedforward projection as well as an excitatory projection to the auditory thalamus. This inhibitory feedforward projection is provided by large γ‐aminobutyric acid (GABA)ergic (LG) neurons, which are characterized by their receipt of dense excitatory axosomatic terminals positive for vesicular glutamate transporter (VGLUT) 2. In the avian torus semicircularis (TS), which is the homolog of the IC, neither the homology of cell types nor the presence of inhibitory feedforward inhibition have been established. In this study, we tested the presence of LG neurons in pigeon and chicken by neuroanatomical techniques. The TS contained two types of GABAergic neurons of different soma size. Of these, larger GABA + cells were encircled by dense VGLUT2 + axosomatic terminals. Ultrastructural analyses revealed that more than 30% of the perimeter of a large GABA+, but not small GABA + or GABA−, soma was covered by presumptive excitatory axosomatic terminals, suggesting that large GABA + cells are the sole recipient of dense excitatory axosomatic synapses. After injection of a retrograde tracer into the auditory thalamus, many retrogradely labeled neurons were found bilaterally in the TS, a few of which were GABA+. Almost all tectothalamic GABA + neurons had large somata, and received dense VGLUT2 + axosomatic terminals. These results clearly demonstrated the presence of LG neurons in birds. The similar morphology of LG neurons implies that the function of tectothalamic inhibition is similar among amniotes. J. Comp. Neurol. 524:2604–2622, 2016. © 2016 Wiley Periodicals, Inc.
A quantitative comparison of the hemispheric, areal, and laminar origins of sensory and motor cortical projections to the superior colliculus of the cat
Authors: Butler Blake E. , Chabot Nicole , Lomber Stephen G.
Published: February 2016

The superior colliculus (SC) is a midbrain structure central to orienting behaviors. The organization of descending projections from sensory cortices to the SC has garnered much attention; however, rarely have projections from multiple modalities been quantified and contrasted, allowing for meaningful conclusions within a single species. Here, we examine corticotectal projections from visual, auditory, somatosensory, motor, and limbic cortices via retrograde pathway tracers injected throughout the superficial and deep layers of the cat SC. As anticipated, the majority of cortical inputs to the SC originate in the visual cortex. In fact, each field implicated in visual orienting behavior makes a substantial projection. Conversely, only one area of the auditory orienting system, the auditory field of the anterior ectosylvian sulcus (fAES), and no area involved in somatosensory orienting, shows significant corticotectal inputs. Although small relative to visual inputs, the projection from the fAES is of particular interest, as it represents the only bilateral cortical input to the SC. This detailed, quantitative study allows for comparison across modalities in an animal that serves as a useful model for both auditory and visual perception. Moreover, the differences in patterns of corticotectal projections between modalities inform the ways in which orienting systems are modulated by cortical feedback. J. Comp. Neurol. 524:2623–2642, 2016. © 2016 Wiley Periodicals, Inc.
Implementation of a pharmacogenomics consult service to support the INGENIOUS trial
Authors: Eadon MT , Desta Z , Levy KD , Decker BS , Pierson RC , Pratt VM , Callaghan JT , Rosenman MB , Carpenter JS , Holmes AM , McDonald CA , Benson EA , Patil AS , Vuppalanchi R , Gufford BT , Dave N , Robarge JD , Hyder MA , Haas DM , Kreutz RP , Dexter PR , Skaar TC , Flockhart DA
Published: February 2016

Hospital systems increasingly utilize pharmacogenomic testing to inform clinical prescribing. Successful implementation efforts have been modeled at many academic centers. In contrast, this report provides insights into the formation of a pharmacogenomics consultation service at a safety‐net hospital, which predominantly serves low‐income, uninsured, and vulnerable populations. The report describes the INdiana GENomics Implementation: an Opportunity for the UnderServed (INGENIOUS) trial and addresses concerns of adjudication, credentialing, and funding.
Intestinal Microbiota in Inflammatory Bowel Disease and Carcinogenesis: Implication for Therapeutics
Authors: Bruner SD , Jobin C
Published: February 2016

Trillions of bacteria inhabit our intestine, forming a community called the microbiota, whose contributions are essential to maintain host homeostasis. Disruption of this normal microbial‐host communication network has deleterious consequences for the host and is associated with intestinal pathologies such as inflammatory bowel diseases (IBD) and colorectal cancer (CRC). Here we present key concepts and mechanisms by which bacteria may participate in intestinal pathology, and discuss possible means to therapeutically target the microbiome.
Rapid analysis and quantification of major neutral lipid species and free fatty acids by HPLC‐ELSD from microalgae
Authors: Donot Florentin , Strub Caroline , Fontana Angélique , Jouy Nicolas , Delbes Christelle , Gunata Ziya , Schorr‐Galindo Sabine
Published: February 2016

A method based on normal‐phase high performance liquid chromatography combined with evaporative light‐scattering detector was developed to quantify major neutral lipids from microalgae. A binary gradient enabled a good resolution of hydrocarbons and major neutral lipids by classes in less than 13 min. A good linearity was observed from 0.9 to 30 μg. Limits of detection (LOD) and quantification (LOQ) was from 3 to 64 ng depending on the analyte. The intra‐ and inter‐run precision (% RSD) of analysis was found less than 10%. The method was successfully applied to microalgae lipid extracts from Botryococcus braunii, Chlorella vulgaris, Spirulina platensis, and Porphyridium cruentum. B. braunii was distinguished by the highest levels of lipids, triacylglycerol (TAGs), and diacylglycerol (DAGs) being the major compounds.

Practical applications: Neutral lipids produced by the microalgae are an interesting way for the development of biofuel from renewable carbon. A simple quantification of these lipids is important in the implementation of oleaginous microalgae screening as well as in the bioprocess optimization of algal lipid production. Besides, determining quantitatively the major lipid classes provides information in the understanding of the biosynthetic pathways and the accumulation of these lipids in relation with algal culture conditions.

Normal‐phase HPLC conditions were studied to establish a method allowing a rapid separation of TAGs, DAGs, MAGs, and FFA but also HC by classes in a single run. Their quantification through evaporative light‐scattering detector (ELSD) was performed. The developed method was applied on lipid extracts from the different algal cultures.
Comparative analysis of Cu (I)‐catalyzed alkyne‐azide cycloaddition (CuAAC) and strain‐promoted alkyne‐azide cycloaddition (SPAAC) in O‐GlcNAc proteomics
Authors: Li Shanshan , Zhu He , Wang Jiajia , Wang Xiaomin , Li Xu , Ma Cheng , Wen Liuqing , Yu Bingchen , Wang Yuehua , Li Jing , Wang Peng George
Published: February 2016

O‐linked β‐N‐acetylglucosamine (O‐GlcNAc) is emerging as an essential protein post‐translational modification in a range of organisms. It is involved in various cellular processes such as nutrient sensing, protein degradation, gene expression, and is associated with many human diseases. Despite its importance, identifying O‐GlcNAcylated proteins is a major challenge in proteomics. Here, using peracetylated N‐azidoacetylglucosamine (Ac4GlcNAz) as a bioorthogonal chemical handle, we described a gel‐based mass spectrometry method for the identification of proteins with O‐GlcNAc modification in A549 cells. In addition, we made a labeling efficiency comparison between two modes of azide‐alkyne bioorthogonal reactions in click chemistry: copper‐catalyzed azide‐alkyne cycloaddition (CuAAC) with Biotin‐Diazo‐Alkyne and stain‐promoted azide‐alkyne cycloaddition (SPAAC) with Biotin‐DIBO‐Alkyne. After conjugation with click chemistry in vitro and enrichment via streptavidin resin, proteins with O‐GlcNAc modification were separated by SDS‐PAGE and identified with mass spectrometry. Proteomics data analysis revealed that 229 putative O‐GlcNAc modified proteins were identified with Biotin‐Diazo‐Alkyne conjugated sample and 188 proteins with Biotin‐DIBO‐Alkyne conjugated sample, among which 114 proteins were overlapping. Interestingly, 74 proteins identified from Biotin‐Diazo‐Alkyne conjugates and 46 verified proteins from Biotin‐DIBO‐Alkyne conjugates could be found in the O‐GlcNAc modified proteins database dbOGAP (http://cbsb.lombardi.georgetown.edu/hulab/OGAP.html). These results suggested that CuAAC with Biotin‐Diazo‐Alkyne represented a more powerful method in proteomics with higher protein identification and better accuracy compared to SPAAC. The proteomics credibility was also confirmed by the molecular function and cell component gene ontology (GO). Together, the method we reported here combining metabolic labeling, click chemistry, affinity‐based enrichment, SDS‐PAGE separation, and mass spectrometry, would be adaptable for other post‐translationally modified proteins in proteomics.
Dysgenetic polycystic disease of the minor and submandibular salivary glands
Authors: Koudounarakis Eleftherios , Willems Stefan , Karakullukcu Baris
Published: February 2016

Background

Dysgenetic polycystic salivary gland disease is a rare pathology. No case of minor salivary gland involvement has been reported in the literature.

Methods

A female patient presented with bilateral tumors of the parotid glands, bilateral submandibular gland enlargement, and multiple cystic lesions of the oral mucosa. MRI of the neck was performed along with fine‐needle aspiration (FNA) of the parotid tumors and excisional biopsy of an oral lesion.

Results

Imaging and FNA findings of the parotid glands were suggestive of bilateral Warthin tumors. Excisional biopsy of the oral lesion revealed a minor salivary gland with histopathological features of dysgenetic polycystic salivary gland disease. Submandibular gland enlargement was also attributed to this rare entity based on the imaging characteristics.

Conclusion

This is the first report of dysgenetic polycystic salivary gland disease of the minor salivary glands, combined with involvement of the submandibular glands and bilateral Warthin tumors of the parotid gland. © 2016 Wiley Periodicals, Inc. Head Neck 38: E2437–E2439, 2016
Abstinence from cocaine‐self‐administration activates the nELAV/GAP ‐43 pathway in the hippocampus: A stress‐related effect?
Authors: Pascale Alessia , Osera Cecilia , Moro Federico , Di Clemente Angelo , Giannotti Giuseppe , Caffino Lucia , Govoni Stefano , Fumagalli Fabio , Cervo Luigi
Published: February 2016

We previously demonstrated that nELAV/GAP‐43 pathway is pivotal for learning and its hippocampal expression is up‐regulated by acute stress following repeated cocaine administration. We therefore hypothesized that abstinence‐induced stress may sustain nELAV/GAP‐43 pathway during early abstinence following 2 weeks of cocaine self‐administration. We found that contingent, but not non‐contingent, cocaine exposure selectively increases hippocampal nELAV, but not GAP‐43, expression immediately after the last self‐administration session, an effect that wanes after 24 h and that comes back 7 days later when nELAV activation becomes associated with increased expression of GAP‐43, an effect again observed only in animals self‐administering the psychostimulant. Such effect is specific for nELAV since the ubiquitous ELAV/HuR is unchanged. This nELAV profile suggests that its initial transient alteration is perhaps related to the daily administration of cocaine, while the increase in the nELAV/GAP‐43 pathway following a week of abstinence may reflect the activation of this cascade as a target of stressful conditions associated with drug‐related memories. © 2016 Wiley Periodicals, Inc.
Probability of combined high sea levels and large rains in Malmö, Sweden, southern Öresund
Authors: Bengtsson Lars
Published: February 2016

Observations of sea level and precipitation in Malmö, Sweden in the southeastern part of the sound Öresund have been used to estimate the probabilities of local compound events of high sea level and large daily and hourly rains. There are observations of sea level and daily rains extending back to 1930. The observations of short‐term rainfall are from 1980 and onwards. Most large rainfalls come in the summer, while the highest sea levels are in the autumn and in the winter. The highest observed sea level is about 130 cm above mean sea level, and the largest daily rain is close to 100 mm. However, the highest sea level observed during a day with rainfall corresponding to the 1‐year rain is less than 60 cm. The highest sea level observed during an hour with 1‐year hourly rainfall is 30 cm.

From the statistics of daily rains, hourly rains and sea level, extreme values for each of them have been computed. For events with frequency higher than one per four years the probabilities of combined events sea level – rainfall are determined directly from the observations. For more rare events, marginal distributions of sea level and rainfall are determined. Copulas and conditional probabilities are used. When the sea level exceeds 20 cm above mean sea level, daily rains exceeding 10 mm are almost independent of the sea level and so are hourly rains exceeding 5 mm. It is extremely rare that large rains occur when the sea level is very high. The combination of 1‐year rainfall and the 1‐year sea level has a return period of more than 200 years.
Home‐based HPV self‐sampling improves participation by never‐screened and under‐screened women: Results from a large randomized trial (iPap) in Australia
Authors: Sultana Farhana , English Dallas R. , Simpson Julie A. , Drennan Kelly T. , Mullins Robyn , Brotherton Julia M.L. , Wrede C. David , Heley Stella , Saville Marion , Gertig Dorota M.
Published: February 2016

We conducted a randomized controlled trial to determine whether HPV self‐sampling increases participation in cervical screening by never‐ and under‐screened (not screened in past 5 years) women when compared with a reminder letter for a Pap test. Never‐ or under‐screened Victorian women aged 30–69 years, not pregnant and with no prior hysterectomy were eligible. Within each stratum (never‐screened and under‐screened), we randomly allocated 7,140 women to self‐sampling and 1,020 to Pap test reminders. The self‐sampling kit comprised a nylon tipped flocked swab enclosed in a dry plastic tube. The primary outcome was participation, as indicated by returning a swab or undergoing a Pap test; the secondary outcome, for women in the self‐sampling arm with a positive HPV test, was undergoing appropriate clinical investigation. The Roche Cobas® 4800 test was used to measure presence of HPV DNA. Participation was higher for the self‐sampling arm: 20.3 versus 6.0% for never‐screened women (absolute difference 14.4%, 95% CI: 12.6–16.1%, p < 0.001) and 11.5 versus 6.4% for under‐screened women (difference 5.1%, 95% CI: 3.4–6.8%, p < 0.001). Of the 1,649 women who returned a swab, 45 (2.7%) were positive for HPV16/18 and 95 (5.8%) were positive for other high‐risk HPV types. Within 6 months, 28 (62.2%) women positive for HPV16/18 had colposcopy as recommended and nine (20%) had cytology only. Of women positive for other high‐risk HPV types, 78 (82.1%) had a Pap test as recommended. HPV self‐sampling improves participation in cervical screening for never‐ and under‐screened women and most women with HPV detected have appropriate clinical investigation.
Multidecadal climate variability over northern France during the past 500 years and its relation to large‐scale atmospheric circulation
Authors: Dieppois B. , Lawler D. M. , Slonosky V. , Massei N. , Bigot S. , Fournier M. , Durand A.
Published: February 2016

We examine secular changes and multidecadal climate variability on a seasonal scale in northern France over the last 500 years and examine the extent to which they are driven by large‐scale atmospheric variability. Multiscale trend analysis and segmentation procedures show statistically significant increases of winter and spring precipitation amounts in Paris since the end of the 19th century. This changes the seasonal precipitation distribution from one with a pronounced summer peak at the end of the Little Ice Age to an almost uniform distribution in the 20th century. This switch is linked to an early warming trend in winter temperature. Changes in spring precipitation are also correlated with winter precipitation for time scales greater than 50 years, which suggests a seasonal persistence. Hydrological modelling results show similar rising trends in river flow for the Seine at Paris. However, such secular trends in the seasonal climatic conditions over northern France are substantially modulated by irregular multidecadal (50–80 years) fluctuations. Furthermore, since the end of the 19th century, we find an increasing variance in multidecadal hydroclimatic winter and spring, and this coincides with an increase in the multidecadal North Atlantic Oscillation (NAO) variability, suggesting a significant influence of large‐scale atmospheric circulation patterns. However, multidecadal NAO variability has decreased in summer. Using Empirical Orthogonal Function analysis, we detect multidecadal North Atlantic sea‐level pressure anomalies, which are significantly linked to the NAO during the Modern period. In particular, a south‐eastward (south‐westward) shift of the Icelandic Low (Azores High) drives substantial multidecadal changes in spring. Wetter springs are likely to be driven by potential changes in moisture advection from the Atlantic, in response to northward shifts of North Atlantic storm tracks over European regions, linked to periods of positive NAO. Similar, but smaller, changes in rainfall are observed in winter.
Relevance of activated hepatic stellate cells in predicting the development of pediatric liver allograft fibrosis
Authors: Venturi Carla , Reding Raymond , Quinones Jorge Abarca , Sokal Etienne , Rahier Jacques , Bueno Javier , Sempoux Christine
Published: February 2016

Activated hepatic stellate cells (HSCs) are the main collagen‐producing cells in liver fibrogenesis. With the purpose of analyzing their presence and relevance in predicting liver allograft fibrosis development, 162 liver biopsies of 54 pediatric liver transplantation (LT) recipients were assessed at 6 months, 3 years, and 7 years after LT. The proportion of activated HSCs, identified by α‐smooth muscle actin (ASMA) immunostaining, and the amount of fibrosis, identified by picrosirius red (PSR%) staining, were determined by computer‐based morphometric analysis. Fibrosis was also staged by using the semiquantitative liver allograft fibrosis score (LAFSc), specifically designed to score fibrosis in the pediatric LT population. Liver allograft fibrosis displayed progression over time by PSR% (P < 0.001) and by LAFSc (P < 0.001). The ASMA expression decreased in the long term, with inverse evolution with respect to fibrosis (P < 0.01). Patients with ASMA‐positive HSCs area ≥ 8% at 6 months (n = 20) developed a higher fibrosis proportion compared to those with ASMA‐positive HSCs area ≤ 8% (n = 34) at the same period of time and in the long term (P = 0.03 and P < 0.01, respectively), but not at 3 years (P = 0.8). ASMA expression ≥ 8% at 6 months was found to be an independent risk factor for 7‐year fibrosis development by PSR% (r
2 = 0.5; P < 0.01) and by LAFSc (r
2 = 0.3; P = 0.03). Furthermore, ASMA expression ≥ 8% at 3 years showed an association with the development of fibrosis at 7 years (P = 0.02). In conclusion, there is a high proportion of activated HSCs in pediatric LT recipients. ASMA ≥ 8% at 6 months seems to be a risk factor for early and longterm fibrosis development. In addition, activated HSCs showed inverse evolution with respect to fibrosis in the long term. Liver Transplantation 22 822–829 2016 AASLD.
Effect of diabetes and acute rejection on liver transplant outcomes: An analysis of the organ procurement and transplantation network/united network for organ sharing database
Authors: Kuo Hung‐Tien , Lum Erik , Martin Paul , Bunnapradist Suphamai
Published: February 2016

The effects of diabetic status and acute rejection (AR) on liver transplant outcomes are largely unknown. We studied 13,736 liver recipients from the United Network for Organ Sharing/Organ Procurement Transplant Network database who underwent transplantation between 2004 and 2007 with a functioning graft for greater than 1 year. The association of pretransplant diabetes mellitus (PDM), new‐onset diabetes after transplant (NODAT), and AR rates on allograft failure, all‐cause mortality, and cardiovascular mortality were determined. To determine the differential and joint effects of diabetic status and AR on transplant outcomes, recipients were further stratified into 6 groups: neither (reference, n = 6600); NODAT alone (n = 2054); PDM alone (n = 2414); AR alone (n = 1448); NODAT and AR (n = 707); and PDM and AR (n = 513). An analysis with hepatitis C virus (HCV) serostatus was also performed (HCV recipients, n = 6384; and non‐HCV recipient, n = 5934). The median follow‐up was 2537 days. The prevalence of PDM was 21.3%. At 1 year after transplant, the rates of NODAT and AR were 25.5% and 19.4%, respectively. Overall, PDM, NODAT, and AR were associated with increased risks for graft failure (PDM, hazard ratio [HR] = 1.31, P < 0.01; NODAT, HR = 1.11, P = 0.02; AR, HR = 1.28, P < 0.01). A multivariate Cox regression analysis of the 6 recipient groups demonstrated that NODAT alone was not significantly associated with any study outcomes. The presence of PDM, AR, NODAT and AR, and PDM and AR were associated with higher overall graft failure risk and mortality risk. The presence of PDM was associated with higher cardiovascular mortality risk. The analyses in both HCV‐positive and HCV‐negative cohorts showed a similar trend as in the overall cohort. In conclusion, PDM and AR, but not NODAT, is associated with increased mortality and liver allograft failure. Liver Transplantation 22 796–804 2016 AASLD.
Regionally progressive accumulation of iron in Parkinson's disease as measured by quantitative susceptibility mapping
Authors: Guan Xiaojun , Xuan Min , Gu Quanquan , Huang Peiyu , Liu Chunlei , Wang Nian , Xu Xiaojun , Luo Wei , Zhang Minming
Published: February 2016

The progression of Parkinson's disease (PD) seems to vary according to the disease stage, which greatly influences the management of PD patients. However, the underlying mechanism of progression in PD remains unclear. This study was designed to explore the progressive pattern of iron accumulation at different stages in PD patients. Sixty right‐handed PD patients and 40 normal controls were recruited. According to the disease stage, 45 patients with Hoehn–Yahr stage ≤ 2.5 and 15 patients with Hoehn–Yahr stage ≥ 3 were grouped into early‐stage PD (EPD) and late‐stage PD (LPD) groups, respectively. The iron content in the cardinal subcortical nuclei covering the cerebrum, cerebellum and midbrain was measured using quantitative susceptibility mapping (QSM). The substantia nigra pars compacta (SNc) showed significantly increased QSM values in the EPD patients compared with the controls. In the LPD patients, while the SNc continued to show increased QSM values compared with the controls and EPD patients, the regions showing increased QSM values spread to include the substantia nigra pars reticulata (SNr), red nucleus (RN) and globus pallidus (GP). Our data also indicated that iron deposition was more significant in the GP internal segment (GPi) than in the GP external segment. No other regions showed significant changes in QSM values among the groups. Therefore, we were able to confirm a regionally progressive pattern of iron accumulation in the different stages of PD, indicating that iron deposition in the SNc is affected exclusively in the early stages of the disease, while the SNr, RN and GP, and particularly the GPi segment, become involved in advanced stages of the disease. This is a preliminary study providing objective evidence of the iron‐related progression in PD. Copyright © 2016 John Wiley & Sons, Ltd.
Y‐chromosome identification in circulating cell‐free fetal DNA using surface plasmon resonance
Authors: Breveglieri Giulia , Bassi Elisabetta , Carlassara Silvia , Cosenza Lucia Carmela , Pellegatti Patrizia , Guerra Giovanni , Finotti Alessia , Gambari Roberto , Borgatti Monica
Published: February 2016

Objective

Since the discovery of cell‐free fetal DNA (cffDNA) in maternal plasma, diagnostic non‐invasive prenatal methods have been developed or optimized for fetal sex determination and identification of genetic diseases. As far as fetal sex determination, this might be important for therapeutic intervention on sex‐associated pathologies such as Duchenne muscular dystrophy, hemophilia and congenital adrenal hyperplasia. Surface plasmon resonance (SPR)‐based biosensors might be useful for these studies, because they allow to monitor the molecular interactions in real‐time providing qualitative and quantitative information, through kinetics, affinity and concentration analyses.

Methods

The BiacoreTM X100 has been applied to identify Y‐chromosome sequence in cffDNA obtained from plasma samples of 26 pregnant women at different gestational ages. We have performed SPR‐based analysis of SRY PCR products using SRY‐specific probes immobilized on the sensor chip.

Results

We have demonstrated that there is a statistically significant difference between samples collected by pregnancies carrying male or female fetuses. Moreover, cffDNA obtained at early gestational ages and not detectable by conventional quantitative real‐time PCR can be discriminated with high accuracy and reliability using SPR‐based biosensors.

Conclusions

These data, in addition to their direct applicability in more extensive diagnostic trials, should be considered as the basis of future developments. © 2016 John Wiley & Sons, Ltd.
Model for the allosteric regulation of the Na+/Ca2+ exchanger NCX
Authors: Abiko Layara Akemi , Vitale Phelipe M. , Favaro Denize C. , Hauk Pricila , Li Da‐Wei , Yuan Jiaqi , Bruschweiler‐Li Lei , Salinas Roberto K. , Brüschweiler Rafael
Published: February 2016

The Na+/Ca2+ exchanger provides a major Ca2+ extrusion pathway in excitable cells and plays a key role in the control of intracellular Ca2+ concentrations. In Canis familiaris, Na+/Ca2+ exchanger (NCX) activity is regulated by the binding of Ca2+ to two cytosolic Ca2+‐binding domains, CBD1 and CBD2, such that Ca2+‐binding activates the exchanger. Despite its physiological importance, little is known about the exchanger's global structure, and the mechanism of allosteric Ca2+‐regulation remains unclear. It was found previously that for NCX in the absence of Ca2+ the two domains CBD1 and CBD2 of the cytosolic loop are flexibly linked, while after Ca2+‐binding they adopt a rigid arrangement that is slightly tilted. A realistic model for the mechanism of the exchanger's allosteric regulation should not only address this property, but also it should explain the distinctive behavior of Drosophila melanogaster's sodium/calcium exchanger, CALX, for which Ca2+‐binding to CBD1 inhibits Ca2+ exchange. Here, NMR spin relaxation and residual dipolar couplings were used to show that Ca2+ modulates CBD1 and CBD2 interdomain flexibility of CALX in an analogous way as for NCX. A mechanistic model for the allosteric Ca2+ regulation of the Na+/Ca2+ exchanger is proposed. In this model, the intracellular loop acts as an entropic spring whose strength is modulated by Ca2+‐binding to CBD1 controlling ion transport across the plasma membrane. Proteins 2016; 84:580–590. © 2016 Wiley Periodicals, Inc.
Anti‐inflammatory effects of the novel inhaled phosphodiesterase type 4 inhibitor CHF6001 on virus‐inducible cytokines
Authors: Edwards Michael R. , Facchinetti Fabrizio , Civelli Maurizio , Villetti Gino , Johnston Sebastian L.
Published: February 2016

Respiratory virus infections precipitate asthma and chronic obstructive pulmonary disease (COPD) exacerbations, with most exacerbations due to rhinovirus infection. Both asthma and COPD exacerbations are not well controlled by steroid therapies, and there is a much research interest in finding improved therapies or combinations of therapies for controlling exacerbations. CHF6001 is a new, inhaled highly potent and selective phosphodiesterase type 4 (PDE4) inhibitor. Using in vitro human bronchial epithelial cells (BEAS‐2B), we investigated the potential anti‐inflammatory effects of CHF6001 on rhinovirus (RV1B)‐induced cytokines. Cytokine mRNA was measured by real‐time PCR, while protein release was measured by ELISA. CHF6001 was used in a 7‐point dose–response curve (1000–0.001 nmol/L) as a 1.5‐h pretreatment prior to infection in comparison with roflumilast. Both roflumilast and CHF6001 reduced RV1B‐induced IL‐8, IL‐29, IP‐10, and RANTES mRNA and protein in a concentration‐dependent manner. Generally, CHF6001 was 13‐ to 16‐fold more potent (subnanomolar EC50 values) than roflumilast at reducing IL‐8, IL‐29, IP‐10, and RANTES mRNA and protein release, but had similar efficacies. In combination with the steroid fluticasone propionate (1 nmol/L), CHF6001 had additive effects, significantly reducing RV‐induced cytokines when compared with steroid or CHF6001 alone. Combined low‐dose steroid and low‐dose CHF6001 had a similar efficacy as high‐dose steroid or CHF6001 alone, indicating the combination had steroid and PDE4 inhibitor sparing effects. Overall results indicate that PDE4 inhibitors have anti‐inflammatory activity against virus‐induced inflammatory mediators and that CHF6001 is more potent than roflumilast.
Lack of cross‐resistance to FF‐10501, an inhibitor of inosine‐5′‐monophosphate dehydrogenase, in azacitidine‐resistant cell lines selected from SKM‐1 and MOLM‐13 leukemia cell lines
Authors: Murase Motohiko , Iwamura Hiroyuki , Komatsu Kensuke , Saito Motoki , Maekawa Toshihiko , Nakamura Takaaki , Yokokawa Takuya , Shimada Yasuhiro
Published: February 2016

Resistance to azacitidine is a major issue in the treatments of myelodysplastic syndrome and acute myeloid leukemia, and previous studies suggest that changes in drug metabolism are involved in the resistance. Therefore, drugs with mechanisms resistant or alternative to such metabolic changes have been desired for the treatment of resistant disease. We generated azacitidine‐resistant cells derived from SKM‐1 and MOLM‐13 leukemia cell lines in vitro, analyzed the mechanisms, and examined the impact on the efficacy of other antimetabolic drugs. It appeared that the cell growth‐inhibitory effect of azacitidine, expression levels of uridine–cytidine kinase 2, and the concentrations of azacitidine triphosphate were remarkably decreased in the resistant cells compared with those in parent cells. These results were consistent with previous observations that azacitidine resistance is derived from metabolic changes. Cross‐resistance of greater than 10‐fold (shift in IC50 value) was observed in azacitidine‐resistant cells for decitabine and for cytarabine, but not for gemcitabine or the inosine‐5′‐monophosphate dehydrogenase (IMPDH) inhibitors FF‐10501 and mycophenolate mofetil (cross‐resistance to 5‐fluorouracil was cell line dependent). The IMPDH inhibitors maintained their cell growth‐inhibitory activities in the azacitidine‐resistant cell lines, in which the levels of adenine phosphoribosyltransferase (which converts FF‐10501 to its active form, FF‐10501 ribosylmonophosphate [FF‐10501RMP]), FF‐10501RMP, and the target enzyme, IMPDH, were equivalent to those in the parent cell lines. These results suggest that an IMPDH inhibitor such as FF‐10501 could be an alternative therapeutic treatment for leukemia patients with acquired resistance to azacitidine.
Slow drug delivery decreased total body clearance and altered bioavailability of immediate‐ and controlled‐release oxycodone formulations
Authors: Li Yan , Sun Duxin , Palmisano Maria , Zhou Simon
Published: February 2016

Oxycodone is a commonly used analgesic with a large body of pharmacokinetic data from various immediate‐release or controlled‐release formulations, under different administration routes, and in diverse populations. Longer terminal half‐lives from extravascular administration as compared to IV administration have been attributed to flip‐flop pharmacokinetics with the rate constant of absorption slower than elimination. However, PK parameters from the extravascular studies showed faster absorption than elimination. Sustained release formulations guided by the flip‐flop concept produced mixed outcomes in formulation development and clinical studies. This research aims to develop a mechanistic knowledge of oxycodone ADME, and provide a consistent interpretation of diverging results and insight to guide further extended release development and optimize the clinical use of oxycodone. PK data of oxycodone in human studies were collected from literature and digitized. The PK data were analyzed using a new PK model with Weibull function to describe time‐varying drug releases/ oral absorption, and elimination dependent upon drug input to the portal vein. The new and traditional PK models were coded in NONMEM. Sensitivity analyses were conducted to address the relationship between rates of drug release/absorption and PK profiles plus terminal half‐lives. Traditional PK model could not be applied consistently to describe drug absorption and elimination of oxycodone. Errors were forced on absorption, elimination, or both parameters when IV and PO profiles were fitted separately. The new mechanistic PK model with Weibull function on absorption and slower total body clearance caused by slower absorption adequately describes the complex interplay between oxycodone absorption and elimination in vivo. Terminal phase of oxycodone PK profile was shown to reflect slower total body drug clearance due to slower drug release/absorption from oral formulations. Mechanistic PK models with Weibull absorption functions, and release rate‐dependent saturable total body clearance well described the diverging oxycodone absorption and elimination kinetics in the literature. It showed no actual drug absorption during the terminal phase, but slower drug clearance caused by slower release/absorption producing the appearance of flip‐flop and offered new insight for the development of modified release formulations and clinical use of oxycodone.
Role of endoplasmic reticulum stress in drug‐induced toxicity
Authors: Foufelle Fabienne , Fromenty Bernard
Published: February 2016

Drug‐induced toxicity is a key issue for public health because some side effects can be severe and life‐threatening. These adverse effects can also be a major concern for the pharmaceutical companies since significant toxicity can lead to the interruption of clinical trials, or the withdrawal of the incriminated drugs from the market. Recent studies suggested that endoplasmic reticulum (ER) stress could be an important event involved in drug liability, in addition to other key mechanisms such as mitochondrial dysfunction and oxidative stress. Indeed, drug‐induced ER stress could lead to several deleterious effects within cells and tissues including accumulation of lipids, cell death, cytolysis, and inflammation. After recalling important information regarding drug‐induced adverse reactions and ER stress in diverse pathophysiological situations, this review summarizes the main data pertaining to drug‐induced ER stress and its potential involvement in different adverse effects. Drugs presented in this review are for instance acetaminophen (APAP), arsenic trioxide and other anticancer drugs, diclofenac, and different antiretroviral compounds. We also included data on tunicamycin (an antibiotic not used in human medicine because of its toxicity) and thapsigargin (a toxic compound of the Mediterranean plant Thapsia garganica) since both molecules are commonly used as prototypical toxins to induce ER stress in cellular and animal models.
Random points in halfspheres
Authors: Bárány Imre , Hug Daniel , Reitzner Matthias , Schneider Rolf
Published: February 2016

A random spherical polytope Pn in a spherically convex set as considered here is the spherical convex hull of n independent, uniformly distributed random points in K. The behaviour of Pn for a spherically convex set K contained in an open halfsphere is quite similar to that of a similarly generated random convex polytope in a Euclidean space, but the case when K is a halfsphere is different. This is what we investigate here, establishing the asymptotic behaviour, as n tends to infinity, of the expectation of several characteristics of Pn, such as facet and vertex number, volume and surface area. For the Hausdorff distance from the halfsphere, we obtain also some almost sure asymptotic estimates. © 2016 Wiley Periodicals, Inc. Random Struct. Alg., 50, 3–22, 2017
Nonparametric survival analysis using Bayesian Additive Regression Trees (BART)
Authors: Sparapani Rodney A. , Logan Brent R. , McCulloch Robert E. , Laud Purushottam W.
Published: February 2016

Bayesian additive regression trees (BART) provide a framework for flexible nonparametric modeling of relationships of covariates to outcomes. Recently, BART models have been shown to provide excellent predictive performance, for both continuous and binary outcomes, and exceeding that of its competitors. Software is also readily available for such outcomes. In this article, we introduce modeling that extends the usefulness of BART in medical applications by addressing needs arising in survival analysis. Simulation studies of one‐sample and two‐sample scenarios, in comparison with long‐standing traditional methods, establish face validity of the new approach. We then demonstrate the model's ability to accommodate data from complex regression models with a simulation study of a nonproportional hazards scenario with crossing survival functions and survival function estimation in a scenario where hazards are multiplicatively modified by a highly nonlinear function of the covariates. Using data from a recently published study of patients undergoing hematopoietic stem cell transplantation, we illustrate the use and some advantages of the proposed method in medical investigations. Copyright © 2016 John Wiley & Sons, Ltd.
Großbrücken in Deutschland
Authors: Reitz Dieter
Published: February 2016

Die aktuelle Marktsituation steht im Bereich der Infrastruktur im Allgemeinen und vor allem im Brückenbau in einem besonderen Kontext. Waren die beiden großen Epochen des Stahlbrückenbaus, der Beginn der Massenmobilität in den 60er und Anfang der 70er Jahre des vergangenen Jahrhunderts sowie die Maßnahmen nach der Wiedervereinigung mit Ende der 90er und zu Beginn des 21. Jahrhunderts, geprägt durch Neubauten weitestgehend auf der grünen Wiese, so stehen wir heute womöglich am Anfang einer weiteren Phase mit wesentlich geänderten Rahmenbedingungen. Der Neubau von Fernstraßen wird sich auf einige Lückenschlüsse reduzieren. Dafür gilt es, das vorhandene Straßennetz fit für das nächste Jahrhundert zu machen. Nicht nur die in die Jahre gekommenen Bauwerke, auch die zu keiner Zeit erwartete Zunahme des Verkehrs und insbesondere des Schwerverkehrs führt zu Sanierungen und Erneuerungsmaßnahmen in nie gedachtem Umfang. Dabei stellt das Bauen unter der Aufrechterhaltung des Verkehrs eine besondere Herausforderung dar.

Major bridges in Germany – Yesterday – today – tomorrow. The current market situation in the field of infrastructure in general and especially the superstructure are placed in a special context. Whereas the history of steel bridges started with the beginning of mass mobility in the 60s and early 70s of the last century as well as the reunification in the late 90s, our century is now battled with completely different conditions. A new area has started and is challenging us with all its pros and cons. The construction of new highways will be reduced to the closing of gaps. Therefore it is really important to focus on the existing road network and its future. Not only are old constructions a huge problem, especially the increase of traffic and particularly heavy load traffic are leading to redevelopment and renewal in never existing extent. All this leads to a very special challenge during the execution of all road and construction works.
Knockdown of DNMT1 and DNMT3a Promotes the Angiogenesis of Human Mesenchymal Stem Cells Leading to Arterial Specific Differentiation
Authors: Zhang Rui , Wang Nan , Zhang Li-Nan , Huang Na , Song Tie-Feng , Li Zheng-Zheng , Li Man , Luo Xue-Gang , Zhou Hao , He Hong-Peng , Zhang Xiao-Yu , Ma Wenjian , Zhang Tong-Cun
Published: February 2016

Human mesenchymal stem cells (hMSCs) possess the potential to differentiate into endothelial cells (EC). DNA methylation plays an important role in cell differentiation during development. However, the role of the DNA methyltransferases Dnmt1 and Dnmt3a in specific arterial differentiation of hMSCs is not clear. Here, we show that the CpG islands in the promoter regions of the EC specification and arterial marker genes were highly methylated in hMSCs based on bisulfite genomic sequencing. Treatment with the DNMT inhibitor 5-aza-dc induced the reactivation of EC specification and arterial marker genes by promoting demethylation of these genes as well as stimulating tube-like structure formation. The hMSCs with stable knockdown of Dnmt1/Dnmt3a were highly angiogenic and expressed several arterial specific transcription factors and marker genes. A Matrigel plug assay confirmed that Dnmt1/Dnmt3a stable knockdown hMSCs enhanced blood vessel formation compared with WT MSCs. We also identified that the transcription factor E2F1 could upregulate the transcription of arterial marker genes by binding to the promoters of arterial genes, suggesting its critical role for arterial specification. Moreover, miRNA gain/loss-of-function analyses revealed that miR152 and miR30a were involved in endothelial differentiation of hMSCs by targeting Dnmt1 and Dnmt3a, respectively. Taken together, these data suggest that Dnmt1 and Dnmt3a are critical regulators for epigenetic silencing of EC marker genes and that E2F1 plays an important role in promoting arterial cell determination.
Concise Review: Heteroplasmic Mitochondrial DNA Mutations and Mitochondrial Diseases: Toward iPSC-Based Disease Modeling, Drug Discovery, and Regenerative Therapeutics
Authors: Hatakeyama Hideyuki , Goto Yu-ichi
Published: February 2016

Mitochondria contain multiple copies of their own genome (mitochondrial DNA; mtDNA). Once mitochondria are damaged by mutant mtDNA, mitochondrial dysfunction is strongly induced, followed by symptomatic appearance of mitochondrial diseases. Major genetic causes of mitochondrial diseases are defects in mtDNA, and the others are defects of mitochondria-associating genes that are encoded in nuclear DNA (nDNA). Numerous pathogenic mutations responsible for various types of mitochondrial diseases have been identified in mtDNA; however, it remains uncertain why mitochondrial diseases present a wide variety of clinical spectrum even among patients carrying the same mtDNA mutations (e.g., variations in age of onset, in affected tissues and organs, or in disease progression and phenotypic severity). Disease-relevant induced pluripotent stem cells (iPSCs) derived from mitochondrial disease patients have therefore opened new avenues for understanding the definitive genotype-phenotype relationship of affected tissues and organs in various types of mitochondrial diseases triggered by mtDNA mutations. In this concise review, we briefly summarize several recent approaches using patient-derived iPSCs and their derivatives carrying various mtDNA mutations for applications in human mitochondrial disease modeling, drug discovery, and future regenerative therapeutics.
Jun-Mediated Changes in Cell Adhesion Contribute to Mouse Embryonic Stem Cell Exit from Ground State Pluripotency
Authors: Veluscek Giulia , Li Yaoyong , Yang Shen-Hsi , Sharrocks Andrew D.
Published: February 2016

Embryonic stem cells (ESC) are able to give rise to any somatic cell type. A lot is known about how ESC pluripotency is maintained, but comparatively less is known about how differentiation is promoted. Cell fate decisions are regulated by interactions between signaling and transcriptional networks. Recent studies have shown that the overexpression or downregulation of the transcription factor Jun can affect the ESC fate. Here we have focussed on the role of the Jun in the exit of mouse ESCs from ground state pluripotency and the onset of early differentiation. Transcriptomic analysis of differentiating ESCs reveals that Jun is required to upregulate a programme of genes associated with cell adhesion as ESCs exit the pluripotent ground state. Several of these Jun-regulated genes are shown to be required for efficient adhesion. Importantly this adhesion is required for the timely regulated exit of ESCs from ground state pluripotency and the onset of early differentiation events.