Sequence Homology at the Breakpoint and Clinical Phenotype of Mitochondrial DNA Deletion Syndromes

Mitochondrial DNA (mtDNA) deletions are a common cause of mitochondrial disorders. Large mtDNA deletions can lead to a broad spectrum of clinical features with different age of onset, ranging from mild mitochondrial myopathies (MM), progressive external ophthalmoplegia (PEO), and Kearns-Sayre syndrome (KSS), to severe Pearson syndrome. The aim of this study is to investigate the molecular signatures surrounding the deletion breakpoints and their association with the clinical phenotype and age at onset. MtDNA deletions in 67 patients were characterized using array comparative genomic hybridization (aCGH) followed by PCR-sequencing of the deletion junctions. Sequence homology including both perfect and imperfect short repeats flanking the deletion regions were analyzed and correlated with clinical features and patients' age group. In all age groups, there was a significant increase in sequence homology flanking the deletion compared to mtDNA background. The youngest patient group (<6 years old) showed a diffused pattern of deletion distribution in size and locations, with a significantly lower sequence homology flanking the deletion, and the highest percentage of deletion mutant heteroplasmy. The older age groups showed rather discrete pattern of deletions with 44% of all patients over 6 years old carrying the most common 5 kb mtDNA deletion, which was found mostly in muscle specimens (22/41). Only 15% (3/20) of the young patients (<6 years old) carry the 5 kb common deletion, which is usually present in blood rather than muscle. This group of patients predominantly (16 out of 17) exhibit multisystem disorder and/or Pearson syndrome, while older patients had predominantly neuromuscular manifestations including KSS, PEO, and MM. In conclusion, sequence homology at the deletion flanking regions is a consistent feature of mtDNA deletions. Decreased levels of sequence homology and increased levels of deletion mutant heteroplasmy appear to correlate with earlier onset and more severe disease with multisystem involvement

N-(4-iodophenyl)-N′-(2-chloroethyl)urea as a microtubule disrupter: in vitro and in vivo profiling of antitumoral activity on CT-26 murine colon carcinoma cell line cultured and grafted to mice

The antitumoral profile of the microtubule disrupter N-(4-iodophenyl)-N′-(2-chloroethyl)urea (ICEU) was characterised in vitro and in vivo using the CT-26 colon carcinoma cell line, on the basis of the drug uptake by the cells, the modifications of cell cycle, and β-tubulin and lipid membrane profiles. N-(4-iodophenyl)-N′-(2-chloroethyl)urea exhibited a rapid and dose-dependent uptake by CT-26 cells suggesting its passive diffusion through the membranes. Intraperitoneally injected ICEU biodistributed into the grafted CT-26 tumour, resulting thus in a significant tumour growth inhibition (TGI). N-(4-iodophenyl)-N′-(2-chloroethyl)urea was also observed to accumulate within colon tissue. Tumour growth inhibition was associated with a slight increase in the number of G2 tetraploid tumour cells in vivo, whereas G2 blockage was more obvious in vitro. The phenotype of β-tubulin alkylation that was clearly demonstrated in vitro was undetectable in vivo. Nuclear magnetic resonance analysis showed that cells blocked in G2 phase underwent apoptosis, as confirmed by an increase in the methylene group resonance of mobile lipids, parallel to sub-G1 accumulation of the cells. In vivo, a decrease of the signals of both the phospholipid precursors and the products of membrane degradation occurred concomitantly with TGI. This multi-analysis established, at least partly, the ICEU activity profile, in vitro and in vivo, providing additional data in favour of ICEU as a tubulin-interacting drug accumulating within the intestinal tract. This may provide a starting point for researches for future efficacious tubulin-interacting drugs for the treatment of colorectal cancers

Race, colonial history and national identity: Resident Evil 5 as a Japanese game

Resident Evil 5 is a zombie game made by Capcom featuring a White American protagonist and set in Africa. This paper argues that approaching this as a Japanese game reveals aspects of a Japanese racial and colonial social imaginary that are missed if this context of production is ignored. In terms of race, the game presents hybrid racial subjectivities that can be related to Japanese perspectives of Blackness and Whiteness where these terms are two poles of difference and identity through which an essentialised Japanese identity is constructed in what Iwabuchi calls “strategic hybridism” (Iwabuchi, 2002). In terms of colonialism, the game echoes structures of Japanese colonialism through which Japanese colonialism is obliquely memorialised and a “normal” Japanese global subjectivity can be performed

Simulation de la dose biologique produite par des protons de 65 MeV (faisceau clinique) et des ions carbone

International audienceIntroduction. Pour optimiser les traitements en hadronthérapie, il est primordial de prendre en compte la dose absorbée physique ainsi que l’impact biologique des rayonnements sur les structures irradiées. Pour cela est estimée l’efficacité biologique relative (EBR) qui permet d’évaluer la réponse des cellules à un rayonnement étudié. Ici nous nous intéressons à un faisceau clinique de protons de 65 MeV délivré par la ligne MediCyc du centre Antoine Lacassagne à Nice et un rayonnement d’ions carbones délivrés par le Heavy-Ion Medical Accelerator de Chiba (Japon). Pour prédire cette efficacité biologique relative, il existe différents modèles biophysiques intégrés par des codes Monte Carlo dont certains sont déjà utilisés dans les logiciels de planification de traitement [1]. Parmi ces modèles, le modèle microcinétique (MKM) [2] et le modèle NanOx [1] sont testés dans cette étude. Nous proposons d’utiliser la plateforme Monte Carlo GATE à laquelle sont combinés ces modèles pour prédire la dose biologique.References 1. Monini, Caterina, Étienne Testa, and Michael Beuve. NanOx Prediction of cell survival probabilities for three cell lines. Acta Physica Polonica B 48.10 (2017).2. Y. Kase et al., Microdosimetric calculation of relative biological effectiveness for design of therapeutic proton beams. Journal of Radiation Research, 54. 485–493 (2013)

CPOP: An open source C++ cell POPulation modeler for radiation biology applications

International audiencePurpose: Multicellular tumor spheroids are realistic in-vitro systems used in radiation biology research to studythe effect of anticancer drugs or to evaluate the resistance of cancer cells under specific conditions. Whencombining the modeling of spheroids together with the simulation of radiation using Monte Carlo methods, onecould estimate cell and DNA damage to be compared with experimental data. We developed a Cell Population(CPOP) modeler combined to Geant4 simulations in order to tackle how energy depositions are allocated to cells,especially when enhancing radiation outcomes using high-Z nanoparticles. CPOP manages to model large threedimensionalcell populations with independent deformable cells described with their nucleus, cytoplasm andmembranes together with force law systems to manage cell–cell interactions.Methods: CPOP is an opensource platform written in C++. It is divided into two main libraries: a “Modeler”library, for cell geometry modeling using meshes, and a Multi Agent System (MAS) library, simulating all agent(cell) interactions among the population. CPOP is fully interfaced with the Geant4 Monte Carlo toolkit and is ableto directly launch Geant4 simulations after compilation.We modeled a full and realistic 3D cell population from SK-MEL28 melanoma cell population cultured experimentally.The spheroid diameter of 550 ± 40 μm corresponds to a population of approximately 1000 cells havinga diameter of 17.2 ± 2.5 μm and a nucleus diameter of 11.2 ± 2.0 μm. We decided to reproduce cell irradiationsperformed with a X-RAD 320 Biological Irradiator (Precision XRay Inc., North Branford, CT).Results: We simulated the energy spectrum of secondary particles generated in the vicinity of the spheroid andplotted the different energy spectra recovered internally to the spheroid. We evaluated also the impact of AGuIX(Gadolinium) nanoparticles modeled into the spheroid with their corresponding secondary energy spectra.Conclusions: We succeeded into modeling cell populations and combined them with Geant4 simulations. The nextstep will be to integrate DNA geometrical models into cell nuclei and to use the Geant4-DNA physics andradiolysis modeling capabilities in order to evaluate early strand breaks induced on DNA

The presence of modified nucleotides is required for cloverleaf folding of a human mitochondrial tRNA.

Direct sequencing of human mitochondrial tRNALysshows the absence of editing and the occurrence of six modified nucleotides (m1A9, m2G10, Psi27, Psi28 and hypermodified nucleotides at positions U34 and A37). This tRNA folds into the expected cloverleaf, as confirmed by structural probing with nucleases. The solution structure of the corresponding in vitro transcript unexpectedly does not fold into a cloverleaf but into an extended bulged hairpin. This non-canonical fold, established according to the reactivity to a large set of chemical and enzymatic probes, includes a 10 bp aminoacyl acceptor stem (the canonical 7 bp and 3 new pairs between residues 8-10 and 65-63), a 13 nt large loop and an anticodon-like domain. It is concluded that modified nucleotides have a predominant role in canonical folding of human mitochondrial tRNALys. Phylogenetic comparisons as well as structural probing of selected in vitro transcribed variants argue in favor of a major contribution of m1A9 in this process

Radiomarquage du NTP 15-5 ciblant les protéoglycanes pour l’imagerie TEP des pathologies du cartilage : comparaison d’un traceur TEP à un traceur TEMP

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Impact of 2D Vs 3D Cell Survivals On the RBE for Proton Therapy Using GATE

International audiencePurpose: Proton therapy treatments are based on a constant RBE (relative biological effectiveness) of 1.1. However, it has been shown that proton RBE varies with linear energy transfer (LET), physiological and biological factors, and clinical endpoint. After many years of improvement, the micro-kinetic model (MKM) allows now to obtain a fine description of the impact of radiation on tissues by using LET distributions and experimental cell survival curves. But, these measurements are generally performed using a 2D cell configuration, which is far to be realistic, compared to the natural arrangement of cells in tissues.To address this issue, we propose the prediction of RBE using the GATE Monte Carlo platform (www.opengatecollaboration.org) for 2D and 3D melanoma cell models irradiated with a therapeutic proton beam.Methods: Cell lines in 2D and 3D configurations are irradiated using a reference X-ray beam of 250 kVp and a therapeutic proton beam of 65 MeV. For the proton beam, cell survival curves are measured at several positions of a SOBP (spread-out Bragg peak) profile. The measured survival curves are then used to set the biological MKM parameters for both configurations.The MKM method combined with the LET simulation is implemented in the GATE platform. The LET distribution is obtained using a tissue-equivalent proportional counter geometry emulating a sphere of tissue equivalent to a cell nucleus (1 µm in diameter).Finally,Results: RBE calculated along proton SOBP depth profiles using 2D and the 3D cell configurations were compared. Survival curves obtained with the reference X-ray beam for 2D and 3D configurations revealed a higher sensitivity to radiation for 3D cell configuration.Conclusion: The setting of the MKM parameters (α₀, β, rd) in GATE for melanoma cells has been performed; the same methodology will be applied for other cell lines during 2017.Funding Support, Disclosures, and Conflict of Interest: This work was supported by grants from LabEx PRIMES (Physique, Radiobiologie, Imagerie Medicale et Simulation