Isolation of Radial Glia-Like Neural Stem Cells from Fetal and Adult Mouse Forebrain via Selective Adhesion to a Novel Adhesive Peptide-Conjugate

Preferential adhesion of neural stem cells to surfaces covered with a novel synthetic adhesive polypeptide (AK-cyclo[RGDfC]) provided a unique, rapid procedure for isolating radial glia-like cells from both fetal and adult rodent brain. Radial glia-like (RGl) neural stem/progenitor cells grew readily on the peptide-covered surfaces under serum-free culture conditions in the presence of EGF as the only growth factor supplement. Proliferating cells derived either from fetal (E 14.5) forebrain or from different regions of the adult brain maintained several radial glia-specific features including nestin, RC2 immunoreactivity and Pax6, Sox2, Blbp, Glast gene expression. Proliferating RGl cells were obtained also from non-neurogenic zones including the parenchyma of the adult cerebral cortex and dorsal midbrain. Continuous proliferation allowed isolating one-cell derived clones of radial glia-like cells. All clones generated neurons, astrocytes and oligodendrocytes under appropriate inducing conditions. Electrophysiological characterization indicated that passive conductance with large delayed rectifying potassium current might be a uniform feature of non-induced radial glia-like cells. Upon induction, all clones gave rise to GABAergic neurons. Significant differences were found, however, among the clones in the generation of glutamatergic an

Fasciola spp: Mapping of the MF6 epitope and antigenic analysis of the MF6p/HDM family of heme-binding proteins

MF6p/FhHDM-1 is a small cationic heme-binding protein which is recognized by the monoclonal antibody (mAb) MF6, and abundantly present in parenchymal cells and secreted antigens of Fasciola hepatica. Orthologs of this protein (MF6p/HDMs) also exist in other causal agents of important foodborne trematodiasis, such as Clonorchis sinensis, Opisthorchis viverrini and Paragonimus westermani. Considering that MF6p/FhHDM-1 is relevant for heme homeostasis in Fasciola and was reported to have immunomodulatory properties, this protein is expected to be a useful target for vaccination. Thus, in this study we mapped the epitope recognized by mAb MF6 and evaluated its antigenicity in sheep. The sequence of the MF6p/FhHDM-1 ortholog from F. gigantica (MF6p/FgHDM-1) was also reported. By means of ELISA inhibitions with overlapping synthetic peptides, we determined that the epitope recognized by mAb MF6 is located within the C-terminal moiety of MF6p/FhHDM-1, which is the most conserved region of MF6p/HDMs. By immunoblotting analysis of parasite extracts and ELISA inhibitions with synthetic peptides we also determined that mAb MF6 reacted with the same intensity with F. hepatica and F. gigantica, and in decreasing order of intensity with C. sinensis, O.viverrini and P. westermani orthologs. On the contrary, mAb MF6 showed no reactivity against Dicrocoelium dendriticum and Schistosoma mansoni. The study of the recognition of peptides covering different regions of MF6p/FhHDM-1 by sera from immunized sheep revealed that the C-terminal moiety is the most antigenic, thus being of potential interest for vaccination. We also demonstrated that the production of antibodies to MF6p/FhHDM-1 in sheep infected by F. hepatica occurs relatively early and follows the same pattern as those produced against L-cathepsins.This work was funded by the Ministerio de Ciencia e Innovación, Spain (Grants AGL2011-30563-C03-01, AGL2011-30563-C03-02 and AGL2011-30563-C03-03); Xunta de Galicia, Spain (Grants GPC 2014/058 and ED431B 2017/18); Network of Biomedical Research on Tropical Diseases (RICET), Spain (Grant RD12/0018/0013) and the European Fund for Regional Development (FEDER). VMS holds a predoctoral fellowship from the Spanish Ministerio de Educación, Cultura y Deporte (Programa de Formación del Profesorado Universitario). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.S

Development and Evaluation of a New Lateral Flow Immunoassay for Serodiagnosis of Human Fasciolosis

Fasciolosis is an important plant-borne trematode zoonosis. This disease is of both clinical and veterinary relevance and, according to the WHO, is considered a re-emerging disease that is spreading around the world. Fasciolosis has a serious impact on health because of the large size of the parasite and the effects of the parasite in down-regulating the host immune response. Human fasciolosis can be distinguished by an acute phase, in which the parasite migrates through different tissues, and a chronic phase in which it invades the bile ducts. Here we describe the development of a rapid, simple and inexpensive immunochromatographic diagnostic method, based on the use of a recombinant cathepsin L1 protein, which performs better than other more complex indirect methods, providing similar specificity and higher sensitivity. The simplicity of the method represents a great advantage for the intervention systems applied in different endemic areas by WHO, such as passive case finding (e.g. Vietnam) and selective treatment (e.g. Egypt). Because of its characteristics, the system can be applied to both phases of the disease, and in holo, meso and hyperendemic areas where point-of-care testing is required

Transmission of Calicophoron daubneyi and Fasciola hepatica in Galicia (Spain): Temporal follow-up in the intermediate and definitive hosts

Background Paramphistomosis caused by Calicophoron daubneyi and fasciolosis caused by Fasciola hepatica are common parasitic diseases of livestock animals. Transmission of the diseases depends on the presence of intermediate hosts, i.e. freshwater gastropods such as lymnaeids. We carried out a 2-year-long study of the dynamics of the snail population acting as the intermediate host for these parasites, considering the population structure in terms of size/age and infection status. In addition, we determined the kinetics of trematode egg excretion in grazing cows. Generalized Additive Models (GAMs) were used to analyze the associations between different response variables and snail size, sampling month and weather-related variables. Results Of the molluscan species examined, Galba truncatula, Radix peregra, Anisus (Anisus) leucostoma and Pisidium casertanum (n = 2802), only G. truncatula was infected with C. daubneyi or F. hepatica, at prevalence rates of 8.2% and 4.4% respectively. The probability of infection with C. daubneyi or F. hepatica was linearly related to snail size, although in different ways (negative for C. daubneyi and positive for F. hepatica). The total snail population increased in winter, when specimens of all size classes were found. Infected snails were more abundant during spring-autumn. Mature cercariae of both parasites were found in most seasons. In the statistical models, the sampling month accounted for a high percentage (71.9–78.2%) of the observed variability in snail abundance. The inclusion of climatic variables in the models moderately increased the percentage of deviance explained (77.7–91.9%). Excretion of C. daubneyi eggs in cow faeces was always higher than that of F. hepatica eggs. Conclusions Particular care should be taken to prevent pastures and the surrounding environment being contaminated with parasite eggs during winter-spring, when the number of snails susceptible to miracidial infections is maximal. This is therefore the optimal time for treating grazing animals. Nevertheless, control of trematodosis based only on chemotherapy is difficult in an area such as the study area, where environmental factors favour the regular appearance of snail populations harbouring mature cercariaeThe present study was financially supported by the Ministerio de Economía y Competitividad (AGL2011-30563-C03-03). The funders did not have any role in the study design, data collection and analyses, decision to publish or preparation of the manuscriptS

HD 181068: A Red Giant in a Triply-Eclipsing Compact Hierarchical Triple System

Hierarchical triple systems comprise a close binary and a more distant component. They are important for testing theories of star formation and of stellar evolution in the presence of nearby companions. We obtained 218 days of Kepler photometry of HD 181068 (magnitude of 7.1), supplemented by groundbased spectroscopy and interferometry, which show it to be a hierarchical triple with two types of mutual eclipses. The primary is a red giant that is in a 45-day orbit with a pair of red dwarfs in a close 0.9-day orbit. The red giant shows evidence for tidally-induced oscillations that are driven by the orbital motion of the close pair. HD 181068 is an ideal target for studies of dynamical evolution and testing tidal friction theories in hierarchical triple systems.Comment: 22 pages, including supporting on-line material. This is the author's version of the work. It is posted here by permission of the AAAS for personal use, not for redistribution. The definitive version was published in Science Vol. 332 no. 6026 pp. 216-218 (8 April 2011), doi:10.1126/science.1201762. http://www.sciencemag.org/content/332/6026/216.ful

In-plate recapturing of a dual-tagged recombinant Fasciola antigen (FhLAP) by a monoclonal antibody (US9) prevents non-specific binding in ELISA

Recombinant proteins expressed in E. coli are frequently purified by immobilized metal affinity chromatography (IMAC). By means of this technique, tagged proteins containing a polyhistidine sequence can be obtained up to 95% pure in a single step, but some host proteins also bind with great affinity to metal ions and contaminate the sample. A way to overcome this problem is to include a second tag that is recognized by a preexistent monoclonal antibody (mAb) in the gene encoding the target protein, allowing further purification. With this strategy, the recombinant protein can be directly used as target in capture ELISA using plates sensitized with the corresponding mAb. As a proof of concept, in this study we engineered a Trichinella-derived tag (MTFSVPIS, recognized by mAb US9) into a His-tagged recombinant Fasciola antigen (rFhLAP) to make a new chimeric recombinant protein (rUS9-FhLAP), and tested its specificity in capture and indirect ELISAs with sera from sheep and cattle. FhLAP was selected since it was previously reported to be immunogenic in ruminants and is expressed in soluble form in E. coli, which anticipates a higher contamination by host proteins than proteins expressed in inclusion bodies. Our results showed that a large number of sera from non-infected ruminants (mainly cattle) reacted in indirect ELISA with rUS9-FhLAP after single-step purification by IMAC, but that this reactivity disappeared testing the same antigen in capture ELISA with mAb US9. These results demonstrate that the 6XHis and US9 tags can be combined when double purification of recombinant proteins is required.This work was supported by: Ministerio de Economía y Competitividad (Spain) [grant number AGL2011-30563-C03 and AGL2014-57125R], Ministerio de Economía, Industria y Competitividad (INIA, Spain) [grants numbers RTA2017-00010-C02-01 and RTA2017-00010-C02-02] and the Consellería de Cultura, Educación e Ordenación Universitaria (Xunta de Galicia, Spain) [grant number ED431B 2017/18]. RAOM holds a predoctoral fellowship from the Spanish Ministerio de Economía y Competitividad (Programa de Formación de Personal Investigador). VMS is supported by a contract under the grant ED431B 2017/18. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.S

Somatostatin subtype-2 receptor-targeted metal-based anticancer complexes

Conjugates of a dicarba analogue of octreotide, a potent somatostatin agonist whose receptors are overexpressed on tumor cells, with [PtCl 2(dap)] (dap = 1-(carboxylic acid)-1,2-diaminoethane) (3), [(η 6-bip)Os(4-CO 2-pico)Cl] (bip = biphenyl, pico = picolinate) (4), [(η 6-p-cym)RuCl(dap)] + (p-cym = p-cymene) (5), and [(η 6-p-cym)RuCl(imidazole-CO 2H)(PPh 3)] + (6), were synthesized by using a solid-phase approach. Conjugates 3-5 readily underwent hydrolysis and DNA binding, whereas conjugate 6 was inert to ligand substitution. NMR spectroscopy and molecular dynamics calculations showed that conjugate formation does not perturb the overall peptide structure. Only 6 exhibited antiproliferative activity in human tumor cells (IC 50 = 63 ± 2 μ in MCF-7 cells and IC 50 = 26 ± 3 μ in DU-145 cells) with active participation of somatostatin receptors in cellular uptake. Similar cytotoxic activity was found in a normal cell line (IC 50 = 45 ± 2.6 μ in CHO cells), which can be attributed to a similar level of expression of somatostatin subtype-2 receptor. These studies provide new insights into the effect of receptor-binding peptide conjugation on the activity of metal-based anticancer drugs, and demonstrate the potential of such hybrid compounds to target tumor cells specifically. © 2012 American Chemical Society

Calicophoron daubneyi (Paramphistomidae) in slaughtered cattle in Castilla y León (Spain)

The prevalence and aetiology of natural paramphistomosis was investigated in cattle slaughtered in the Castilla y León region (Spain) over a 3 year-period. The overall prevalence of positive animals was 6.20%. The parasite burden per animal ranged from 8 to 8005 (median = 144) and the ruminal atrium had the highest parasite burden whereas the ruminal dorsal sac the lowest. The prevalence and parasite burden increased with age while these parameters were lower in cattle under intensive management. Calicophoron daubneyi was the only Paramphistomidae species identified using morphoanatomical, histological and molecular methods in the studied animals.This work was supported by grant LE023A10-2 fromJunta de Castilla y León (JCyL). A.M. Martínez-Ibeas was supported by the JCyL andthe European Social Funds (ESF) and J. Benavides by theJAE-Doc programme (CSIC-ESF).Peer Reviewe

An UXor among FUors: Extinction-related Brightness Variations of the Young Eruptive Star V582 Aur

V582 Aur is an FU Ori-type young eruptive star in outburst since similar to 1985. The eruption is currently in a relatively constant plateau phase, with photometric and spectroscopic variability superimposed. Here we will characterize the progenitor of the outbursting object, explore its environment, and analyze the temporal evolution of the eruption. We are particularly interested in the physical origin of the two deep photometric dips, one that occurred in 2012 and one that is ongoing since 2016. We collected archival photographic plates and carried out new optical, infrared, and millimeter-wave photometric and spectroscopic observations between 2010 and 2018, with a high sampling rate during the current minimum. Besides analyzing the color changes during fading, we compiled multiepoch spectral energy distributions and fitted them with a simple accretion disk model. Based on pre-outburst data and a millimeter continuum measurement, we suggest that the progenitor of the V582 Aur outburst is a low-mass T Tauri star with average properties. The mass of an unresolved circumstellar structure, probably a disk, is 0.04M(circle dot). The optical and near-infrared spectra demonstrate the presence of hydrogen and metallic lines, show the CO band head in absorption, and exhibit a variable Ha profile. The color variations strongly indicate that both the similar to 1 yr long brightness dip in 2012 and the current minimum since 2016 are caused by increased extinction along the line of sight. According to our accretion disk models, the reddening changed from A(V) = 4.5 to 12.5mag, while the accretion rate remained practically constant. Similarly to the models of the UXor phenomenon of intermediate- and low-mass young stars, orbiting disk structures could be responsible for the eclipses

A Multi-Site Campaign To Detect The Transit Of The Second Planet In Hat-P-13 (Research Note)

Aims. A possible transit of HAT-P-13c had been predicted to occur on 2010 April 28. Here we report on the results of our multi-site campaign organised to detect the event. Methods. CCD photometric observations were carried out at five observatories in five countries. We reached 30% time coverage in a 5-day interval centered on the suspected transit of HAT-P-13c. Two transits of HAT-P-13b were also observed. Results. No transit of HAT-P-13c was detected during the campaign. By a numerical experiment with 10(5) model systems, we conclude that HAT-P-13c is not a transiting exoplanet with a significance level from 65% to 72%, depending on the planet parameters and the prior assumptions. We present two times of transit of HAT-P-13b ocurring at BJD 2 455 141.5522 +/- 0.0010 and BJD 2 455 249.4508 +/- 0.0020. The TTV of HAT-P-13b is consistent with zero within 0.001 days. The refined orbital period of HAT-P-13b is 2.916293 +/- 0.000010 days.Hungarian OTKA Grants K76816, K68626,K81421, MB08C 81013Lendület Young Researchers’ Program of the Hungarian Academy of SciencesBolyai Research Fellowship of the Hungarian Academy of SciencesAstronom