Inhibition of Notch3 signalling induces rhabdomyosarcoma cell differentiation promoting p38 phosphorylation and p21(Cip1) expression and hampers tumour cell growth in vitro and in vivo

Rhabdomyosarcoma (RMS) is a paediatric soft-tissue sarcoma arising from skeletal muscle precursors coexpressing markers of proliferation and differentiation. Inducers of myogenic differentiation suppress RMS tumourigenic phenotype. The Notch target gene HES1 is upregulated in RMS and prevents tumour cell differentiation in a Notch-dependent manner. However, Notch receptors regulating this phenomenon are unknown. In agreement with data in RMS primary tumours, we show here that the Notch3 receptor is overexpressed in RMS cell lines versus normal myoblasts. Notch3-targeted downregulation in RMS cells induces hyper-phosphorylation of p38 and Akt essential for myogenesis, resulting in the differentiation of tumour cells into multinucleated myotubes expressing Myosin Heavy Chain. These phenomena are associated to a marked decrease in HES1 expression, an increase in p21(Cip1) level and the accumulation of RMS cells in the G1 phase. HES1-forced overexpression in RMS cells reverses, at least in part, the pro-differentiative effects of Notch3 downregulation. Notch3 depletion also reduces the tumourigenic potential of RMS cells both in vitro and in vivo. These results indicate that downregulation of Notch3 is sufficient to force RMS cells into completing a correct full myogenic program providing evidence that it contributes, partially through HES1 sustained expression, to their malignant phenotype. Moreover, they suggest Notch3 as a novel potential target in human RMS.     

MR-Based PET Motion Correction Procedure for Simultaneous MR-PET Neuroimaging of Human Brain

Positron Emission Tomography (PET) images are prone to motion artefacts due to the long acquisition time of PET measurements. Recently, simultaneous magnetic resonance imaging (MRI) and PET have become available in the first generation of Hybrid MR-PET scanners. In this work, the elimination of artefacts due to head motion in PET neuroimages is achieved by a new approach utilising MR-based motion tracking in combination with PET list mode data motion correction for simultaneous MR-PET acquisitions. The method comprises accurate MR-based motion measurements, an intra-frame motion minimising and reconstruction time reducing temporal framing algorithm, and a list mode based PET reconstruction which utilises the Ordinary Poisson Algorithm and avoids axial and transaxial compression. Compared to images uncorrected for motion, an increased image quality is shown in phantom as well as in vivo images. In vivo motion corrected images show an evident increase of contrast at the basal ganglia and a good visibility of uptake in tiny structures such as superior colliculi.     

Embryo quality and transcervical technique are not the limiting factors in donkey embryo transfer outcome

Embryo transfer (ET) in the donkey resulted in a very low recipient pregnancy rates. The aim of these studies was to investigate if nonsurgical transfer techniques or donkey embryo quality affect donkey recipient pregnancy failure. In Study 1, the impact of transfer technique was investigated by evaluating if cervical catheterization is associated with prostaglandin release and suppression of luteal function and if donkey recipients would become pregnant after nonsurgical transfer of horse embryos. Four jennies, from 5 to 8 d after ovulation, were submitted to a sham transcervical ET and to evaluation of PGFM and progesterone plasma concentrations. Five 8 d horse embryos were nonsurgically transferred into synchronized donkey recipients (HD). Cervical stimulation caused a transient PGF_2α release in two of four jennies in the absence of a significant decrease in progesterone plasma concentration. All transferred horse embryos resulted in pregnancies in the jenny recipients. In Study 2, donkey embryo viability was investigated by 1.2 meters, 6-diamidino-2-phenylindole (DAPI) staining of 10 embryos and by the transfer of 6 and 12 donkey embryos in synchronized mare (DH) and donkey (DD) recipients, respectively, of known fertility. The estimated proportion of dead cells in DAPI stained embryos was 0.9% (range 0-3.9%) and below what is considered normal (20%) for horse embryos. Three of six and six of 12 of the DH and DD ETs, respectively resulted in pregnancies at 14 and 25 d (50%), a higher pregnancy rate than previously reported after DD ET. The overall results of this study suggest that the transcervical technique for ET and donkey embryo viability are not the reasons for the low pregnancy rates that have previously been described in donkey recipients, and that nonsurgical ET in donkeys can result in acceptable results.     

Effects of Magnetic Fields of up to 9.4 T on Resolution and Contrast of PET Images as Measured with an MR-BrainPET

Simultaneous, hybrid MR-PET is expected to improve PET image resolution in the plane perpendicular to the static magnetic field of the scanner. Previous papers have reported this either by simulation or experiment with simple sources and detector arrangements. Here, we extend those studies using a realistic brain phantom in a recently installed MR-PET system comprising a 9.4 T MRI-scanner and an APD-based BrainPET insert in the magnet bore. Point and line sources and a 3D brain phantom were filled with ¹⁸F (low-energy positron emitter), ⁶⁸Ga (medium energy positron emitter) or ¹²⁰I, a non-standard positron emitter (high positron energies of up to 4.6 MeV). Using the BrainPET insert, emission scans of the phantoms were recorded at different positions inside and outside the magnet bore such that the magnetic field was 0 T, 3 T, 7 T or 9.4 T. Brain phantom images, with the ‘grey matter’ compartment filled with ¹⁸F, showed no obvious resolution improvement with increasing field. This is confirmed by practically unchanged transaxial FWHM and ‘grey/white matter’ ratio values between at 0T and 9.4T. Field-dependent improvements in the resolution and contrast of transaxial PET images were clearly evident when the brain phantom was filled with ⁶⁸Ga or ¹²⁰I. The grey/white matter ratio increased by 7.3% and 16.3%, respectively. The greater reduction of the FWTM compared to FWHM in ⁶⁸Ga or ¹²⁰I line-spread images was in agreement with the improved contrast of ⁶⁸Ga or ¹²⁰I images. Notwithstanding elongations seen in the z-direction of ⁶⁸Ga or ¹²⁰I point source images acquired in foam, brain phantom images show no comparable extension. Our experimental study confirms that integrated MR-PET delivers improved PET image resolution and contrast for medium- and high-energy positron emitters even though the positron range is reduced only in directions perpendicular to the magnetic field.     

Infection risk dictates immunological divergence among populations in a Mediterranean lizard

The ability of vertebrates to evolve different defence strategies in response to varying parasitism regimes remains poorly understood. Hosts may adopt two different strategies to defend themselves against parasites: tolerance (hosts alleviate the negative fitness consequences of parasite infection) and resistance (hosts strengthen their immune response as parasite burden increases). Both strategies are effective, but fitness has been reported to decline faster in less‐tolerant individuals. Here, we assessed the number of splenocytes and the cell‐mediated response (proxies for resistance) and body condition (a proxy for tolerance) in four populations of a Greek endemic lizard (Podarcis gaigeae), each exposed to different infection risks (defined as the cumulative effect of parasite burden and duration of exposure). We anticipated that populations with heavy parasite burden would enhance the efficacy of their immune response (resistance) compared to lizards deriving from parasite‐poor habitats. We also predicted that populations with longer exposure to parasites would be adopted and be more tolerant. Each factor (duration of exposure and parasite burden) had a distinct effect on the immune response, and thus, our results were rather complicated. Lizards with heavy parasite burden and aperiodic exposure demonstrated resistance, whereas lizards with heavy parasite burden and chronic exposure were more tolerant. Populations with low parasite burden and minimal exposure were more resistant. Our results suggest that the development of some immunological strategies may be differentiated under different infection risks, even within the same species.     

Retrospective study of factors affecting multiple ovulations,embryo recovery,quality, and diameter in a commercial equine embryo transfer program

In this study, 198 donor mares of different breeds, ages, and reproductive category were inseminated with fresh, cooled and frozen or frozen and cooled semen at the embryo transfer station or in private artificial insemination centers during 10 breeding seasons. The results of this activity were retrospectively analyzed by Pearson Chi-square test and logistic regression to evaluate factors affecting multiple ovulations, embryo recovery, embryo quality, and embryo diameter. Out of the 661 cycles, 937 ovulations were recorded (mean ovulations/cycle: 1.42 ± 0.58). Ovulation rate and incidence of multiple ovulations were significantly affected by age, breed, and reproductive category. Uterine flushings for embryo recovery were performed between 7 and 10 days after ovulation and resulted in the recovery of 338 embryos (51.1% embryos/cycle and 36.1% embryos/ovulation, respectively). At least one embryo was recovered in 298 flushings (45.1%). The factors affecting embryo recovery were age, breed, reproductive category, type of semen, number of ovulations, and location of artificial insemination. Flushing protocol and day of flushing had no effect on embryo recovery. Age, type of semen, number of ovulations, and day of flushing had a significant influence on embryo diameter (N = 215). None of the factors included in the model had an effect on embryo quality distribution.     

Data mining for simple sequence repeats in expressed sequence tags from barley,maize, rice,sorghum and wheat

Plant genomics projects involving model species and many agriculturally important crops are resulting in a rapidly increasing database of genomic and expressed DNA sequences. The publicly available collection of expressed sequence tags (ESTs) from several grass species can be used in the analysis of both structural and functional relationships in these genomes. We analyzed over 260000 EST sequences from five different cereals for their potential use in developing simple sequence repeat (SSR) markers. The frequency of SSR-containing ESTs (SSR-ESTs) in this collection varied from 1.5% for maize to 4.7% for rice. In addition, we identified several ESTs that are related to the SSR-ESTs by BLAST analysis. The SSR-ESTs and the related sequences were clustered within each species in order to reduce the redundancy and to produce a longer consensus sequence. The consensus and singleton sequences from each species were pooled and clustered to identify cross-species matches. Overall a reduction in the redundancy by 85% was observed when the resulting consensus and singleton sequences (3569) were compared to the total number of SSR-EST and related sequences analyzed (24606). This information can be useful for the development of SSR markers that can amplify across the grass genera for comparative mapping and genetics. Functional analysis may reveal their role in plant metabolism and gene evolution.