This fMRI method shows potential for evaluating language dominance in patients with a variety of brain lesions.

Gene electrotransfer is a safe and efficient nonviral technique for the transfer of nucleic acids of all sizes. Using a small reporter plasmid (3.5 kbp), electrotransfer of more than 90% of the cells, with ~70% viability, can be routinely achieved even in primary cells like mesenchymal stem cells. However, under the same experimental conditions, electrotransfer of larger plasmids (from 6 to 16 kbp) results in very low viability and transfection efficacy. Here, we show that these strong decreases are directly linked to the physical size of the plasmid molecule. Moreover, large plasmids are toxic only when the cells are exposed to electrotransfer pulses. This specific toxicity of large plasmids during electrotransfer is not due to transgene expression and occurs within less than 45 minutes. Indeed, postpulses recovery times of up to 45 minutes are able to entirely abolish the specific toxicity of large plasmid electrotransfer, resulting in a survival and transfection efficacy identical to that of small plasmids. Finally, electrotransfer of small and large plasmids can reach 90–99% of transfection with 60–90% survival considering the findings here reported.

Dysfunction or death of retinal pigment epithelial (RPE) cells is involved in some forms of Retinitis Pigmentosa and in age-related macular degeneration (AMD). Since there is no cure for most patients affected by these diseases, the transplantation of RPE cells derived from human pluripotent stem cells (hPSCs) represents an attractive therapeutic alternative. First attempts to transplant hPSC-RPE cells in AMD and Stargardt patients demonstrated the safety and suggested the potential efficacy of this strategy. However, it also highlighted the need to upscale the production of the cells to be grafted in order to treat the millions of potential patients. Automated cell culture systems are necessary to change the scale of cell production. In the present study, we developed a protocol amenable for automation that combines in a sequential manner Nicotinamide, Activin A and CHIR99021 to direct the differentiation of hPSCs into RPE cells. This novel differentiation protocol associated with the use of cell culture robots open new possibilities for the production of large batches of hPSC-RPE cells while maintaining a high cell purity and functionality. Such methodology of cell culture automation could therefore be applied to various differentiation processes in order to generate the material suitable for cell therapy.

Rostro-caudal patterning of vertebrates depends on the temporally progressive activation of HOX genes within axial stem cells that fuel axial embryo elongation. Whether the pace of sequential activation of HOX genes, the 'HOX clock', is controlled by intrinsic chromatin-based timing mechanisms or by temporal changes in extrinsic cues remains unclear. Here, we studied HOX clock pacing in human pluripotent stem cell-derived axial progenitors differentiating into diverse spinal cord motor neuron subtypes. We show that the progressive activation of caudal HOX genes is controlled by a dynamic increase in FGF signaling. Blocking the FGF pathway stalled induction of HOX genes, while a precocious increase of FGF, alone or with GDF11 ligand, accelerated the HOX clock. Cells differentiated under accelerated HOX induction generated appropriate posterior motor neuron subtypes found along the human embryonic spinal cord. The pacing of the HOX clock is thus dynamically regulated by exposure to secreted cues. Its manipulation by extrinsic factors provides synchronized access to multiple human neuronal subtypes of distinct rostro-caudal identities for basic and translational applications.This article has an associated ‘The people behind the papers’ interview.

Mesenchymal stem cells (MSCs) are multipotent nonhematopoietic cells with the ability to differentiate into various specific cell types, thus holding great promise for regenerative medicine. Early clinical trials have proven that MSC-based therapy is safe, with possible efficacy in various diseased states. Moreover, genetic modification of MSCs to improve their function can be safely achieved using electrogene transfer. We previously achieved transfection efficiencies of up to 32% with preserved viability in rat MSCs. In this study, we further improved the transfection efficiency and transgene expression in human MSCs (hMSCs), while preserving the cells viability and ability to differentiate into osteoblasts and adipocytes by increasing the plasmid concentration and altering the osmotic pressure of the electrotransfer buffer. Using a square-wave electric pulse generator, we achieved a transfection efficiency of more than 80%, with around 70% viability and a detectable transgene expression of up to 30 days. Moreover, we demonstrated that this transfection efficiency can be reproduced reliably on two different sources of hMSCs: the bone marrow and adipose tissue. We also showed that there was no significant donor variability in terms of their transfection efficiency and viability. The cell confluency before electrotransfer had no significant effect on the transfection efficiency and viability. Cryopreservation of transfected cells maintained their transgene expression and viability upon thawing. In summary, we are reporting a robust, safe, and efficient protocol of electrotransfer for hMSCs with several practical suggestions for an optimal use of genetically engineered hMSCs for clinical application.

Nanoparticles of a mesoporous iron(iii) trimesate MIL-100 nanocarrier encapsulating high amounts of the challenging antineoplastic busulfan were administered to rats and compared with the commercial Busilvex®.

Background: Localizing critical brain functions such as language in children is difficult and generally requires invasive techniques. Recently sensory, motor and language functions in adults have been mapped to specific brain locations using functional imaging techniques. Of these techniques, functional MRI (fMRI) is the least invasive and has the highest spatial and temporal resolution. Its use in adults is well documented but application to children has not been as well described. In the present study lateralization and localization of language was evaluated with fMRI prior to epilepsy surgery in a nine-year-old male with complex partial seizures, attentional difficulty and decreased verbal proficiency. Methods: Two language paradigms well studied in adults (read, verb generation) and two additional language paradigms (antonym generation, letter fluency) were studied using whole brain fMRI after stimulus items and timing were adjusted to achieve the desired performance level during imaging. The patient was also conditioned to the magnet environment prior to imaging. Results: Word reading and letter fluency tasks produced lateralized and localized activation similar to that seen in adults. The patient had no language deficits following an anterior 2/3 dominant temporal lobe resection. Conclusions: With modifications of protocols such as those detailed in this report, this non-invasive method for localizing language function is feasible for the presurgical evaluation of children as well being applicable for a variety of developmental language issues. RESUME: Localisation fonctionnelle du langage par RMN chez un enfant de 9 ans. Introduction: II est difficile de localiser des fonctions critiques du cerveau comme le langage chez les enfants et une telle Ulche requiert g6n6ralement des techniques invasives. Dernierement, on a localise 1 a certaines r6gions spdcifiques du cerveau des fonctions sensorielles, motrices et linguistiques chez des adultes au moyen de techniques d'imagerie fonctionnelle. Parmi ces techniques, la RMN fonctionnelle (RMNf) est la moins invasive et poss&de la meilleure resolution spatiale et temporale. Son utilisation chez les adultes est bien document6e, mais son application chez les enfants n'a pas 6t6 bien d6crite. Dans cette etude, la latSralisation et la localisation du langage ont et6 6valu6es par RMNf avant une chirurgie pour epilepsie chez un enfant de neuf ans qui avait des crises partielles complexes, des difficultfes d'attention et une habiletfi verbale limit6e. Mithodes: Deux paradigmes du langage qui ont bit bien 6tudi6s chez les adultes (la lecture, la generation des verbes) et deux paradigmes additionnels du langage (la g6n6ration des antonymes, la maitrise des lettres) ont 6t6 Studies au moyen de la RMNf du cerveau entier aprds avoir ajuste les items servant au stimulus et le chronomdtrage pour obtenir le niveau de performance d6sir6 pendant l'imagerie. Le patient avait aussi 6t6 condition^ a l'environnement avant l'imagerie. Rhuuats: Les taches impliquant lecture de ...

Four experiments investigated the representation and integration in memory of spatial and nonspatial relations. Subjects learned two-dimensional spatial arrays in which critical pairs of object names were semantically related (Experiment 1), semantically and episodically related (Experiment 2), or just episodically related (Experiments 3a and 3b). Episodic relatedness was established in a paired-associate learning task that preceded array learning. After learning an array, subjects participated in two tasks: item recognition, in which the measure of interest was priming; and distance estimation. Priming in item recognition was sensitive to the Euclidean distance between object names and, for neighbouring locations, to nonspatial relations. Errors in distance estimations varied as a function of distance but were unaffected by nonspatial relations. These and other results indicated that nonspatial relations influenced the probability of encoding spatial relations between locations but did not lead to distorted spatial memories.