Implication de la voie de dégradation ubiquitine-dépendante dans la pathologie des maladies de surchage lysosomale

Mémoire numérisé par la Direction des bibliothèques de l'Université de Montréal

An Evaluation Of Advertising Models In Emergent Countries – The Case Of Albania

The market today is confronted with cornucopia of challenges and opportunities. The unfolding scenario is being closely watched by marketers across the globe and they are competing with each other to grasp the market share. In order to win over the consumers, marketers are constantly evolving strategies. The western firms based and operating from market-based economies are progressively being confronted with the growing challenge of creating brand awareness amongst the perspective consumers. As a result, advertising has become an essential marketing tool for these foreign firms in establishing their relatively less or unknown brands and products. The study aims to report the results of a survey about changing attitudes towards advertising in Albania. It examines the questionnaire - based response of a sample population of consumers from the city of Vlora, in their general attitudes towards advertising. Findings will contribute to the understanding of theoretical explanations for advertising in emerging markets, and of western firms using advertising marketing tool to penetrate these markets

Efficient and Robust NK-Cell Transduction With Baboon Envelope Pseudotyped Lentivector

NK-cell resistance to transduction is a major technical hurdle for developing NK-cell immunotherapy. By using Baboon envelope pseudotyped lentiviral vectors (BaEV-LVs) encoding eGFP, we obtained a transduction rate of 23.0 ± 6.6% (mean ± SD) in freshly-isolated human NK-cells (FI-NK) and 83.4 ± 10.1% (mean ± SD) in NK-cells obtained from the NK-cell Activation and Expansion System (NKAES), with a sustained transgene expression for at least 21 days. BaEV-LVs outperformed Vesicular Stomatitis Virus type-G (VSV-G)-, RD114- and Measles Virus (MV)- pseudotyped LVs (p < 0.0001). mRNA expression of both BaEV receptors, ASCT1 and ASCT2, was detected in FI-NK and NKAES, with higher expression in NKAES. Transduction with BaEV-LVs encoding for CAR-CD22 resulted in robust CAR-expression on 38.3 ± 23.8% (mean ± SD) of NKAES cells, leading to specific killing of NK-resistant pre-B-ALL-RS4;11 cell line. Using a larger vector encoding a dual CD19/CD22-CAR, we were able to transduce and re-expand dual-CAR-expressing NKAES, even with lower viral titer. These dual-CAR-NK efficiently killed both CD19KO- and CD22KO-RS4;11 cells. Our results suggest that BaEV-LVs may efficiently enable NK-cell biological studies and translation of NK-cell-based immunotherapy to the clinic

Human mesenchymal stromal cell-secreted lactate induces M2-macrophage differentiation by metabolic reprogramming

Human mesenchymal stromal cells (MSC) have been shown to dampen immune response and promote tissue repair, but the underlying mechanisms are still under investigation. Herein, we demonstrate that umbilical cord-derived MSC (UC-MSC) alter the phenotype and function of monocyte-derived dendritic cells (DC) through lactate-mediated metabolic reprogramming. UC-MSC can secrete large quantities of lactate and, when present during monocyte-to-DC differentiation, induce instead the acquisition of M2-macrophage features in terms of morphology, surface markers, migratory properties and antigen presentation capacity. Microarray expression profiling indicates that UC-MSC modify the expression of metabolic-related genes and induce a M2-macrophage expression signature. Importantly, monocyte-derived DC obtained in presence of UC-MSC, polarize naïve allogeneic CD4+ T-cells into Th2 cells. Treatment of UC-MSC with an inhibitor of lactate dehydrogenase strongly decreases lactate concentration in culture supernatant and abrogates the effect on monocyte-to- DC differentiation. Metabolic analysis further revealed that UC-MSC decrease oxidative phosphorylation in differentiating monocytes while strongly increasing the spare respiratory capacity proportional to the amount of secreted lactate. Because both MSC and monocytes are recruited in vivo at the site of tissue damage and inflammation, we propose the local increase of lactate concentration induced by UC-MSC and the consequent enrichment in M2-macrophage generation as a mechanism to achieve immunomodulation

Implication de la voie de dégradation ubiquitine-dépendante dans la pathologie des maladies de surchage lysosomale

Mémoire numérisé par la Direction des bibliothèques de l'Université de Montréal

Molecular signatures and properties of Pitx3-dependent and Pitx3-independent subsets of midbrain dopaminergic neurons

Midbrain dopaminergic (mDA) neurons have been implicated in neurological disorders and Parkinson’s disease (PD), the latter being characterized by progressive degeneration of particular subsets of mDA neurons. For example, mDA neurons localized in ventral substantia nigra compacta (vSNc) display an increased propensity to degenerate in typical PD subjects and animal models, while other subsets in dorsal SNc (dSNc) and VTA are more resistant. The molecular basis for this selective vulnerability remains largely unknown. Using Pitx3-null mice as a PD model for selective mDA cell loss, we performed a genome-wide transcriptome comparison of FACS-purified mDA subpopulations in order to gain insight into their representative gene expression profiles that may explain their respective biology and propensity to degenerate. This strategy first led us to discover an array of novel markers enriched in mDA neurons of vSNc, dSNc and VTA. Secondly, among these mDA subset-enriched markers, we identified those that are likely to be dependent on Pitx3. Third, we provide evidence for a putative subset-specific role of Pitx3 in regulation of primary cilium signaling in Pitx3-dependent mDA neurons of the VTA. Fourth, we identified conserved genomic regulatory regions where Pitx3 binds to mediate its gene regulatory actions. Fifth, we propose survival factors and pathways that are preferentially enriched in mDA neurons of vSNc, and that could participate in degenerative processes. Lastly, we study one such potential vSNc-enriched survival factor, that is Regulator of G-protein Signaling 6 (Rgs6) and demonstrate its implication in adult maintenance of vSNc mDA neurons. Indeed, inactivation of the Rgs6 gene in mice results in late-onset degeneration of vSNc mDA neurons associated with decreased expression of Pitx3 and its target genes. The ensemble of this work opened novel perspectives on the molecular understanding of mDA neuron biology in normal and disease states.Les neurones dopaminergiques du mésencéphale (DAm) ont été impliqués dans des désordres neurologiques et la maladie de Parkinson (MP), cette dernière étant caractérisée par la dégénérescence progressive de sous-populations particulières de neurones DAm. Par exemple, les neurones DAm de la partie ventrale de la substance noire (vSNc) et une faible proportion de ceux de l’aire tegmentée (VTA) montrent une tendance élevée pour la dégénérescence dans les patients typiques de la MP et certains modèles animaux, tandis que les sous-populations de neurones DAm dans la partie dorsale de la SNc (dSNc) et celles du VTA sont généralement plus résistantes. La base moléculaire pour cette vulnérabilité sélective reste largement inconnue. En ayant recours à des souris nulles pour Pitx3 en tant que modèles de la perte neuronale sélective, nous avons entrepris des comparaisons du transcriptome de diverses sous-populations de neurones DAm triés par FACS. Ceci a été fait afin de révéler leur profil d’expression génique qui, selon nous, expliquerait leurs propriétés biologiques ainsi que leur vulnérabilité. Cette stratégie a premièrement permis la découverte de marqueurs moléculaires propres aux neurones DAm de la vSNc, dSNc et du VTA. Deuxièmement, parmi ces marqueurs spécifiques, nous avons identifié ceux dont l’expression dépendrait de Pitx3. Troisièmement, nous avons montré une expression prédominante de la signalisation reliée au cil primaire uniquement dans la sous-population de neurones DAm Pitx3-dépendants du VTA. Quatrièmement, nous avons identifié les profils de liaison du facteur de transcription Pitx3 sur l’ensemble du génome. Cinquièmement, nous avons identifié des facteurs et voies de survie qui sont préférentiellement actives dans les neurones DAm de la vSNc et qui participeraient aux processus de dégénérescence. Enfin, nous avons étudié un de ces facteurs de survie enrichi dans la vSNc, le Regulator of G-protein Signaling 6 (Rgs6), et nous démontrons son implication dans la survie des neurones DAm de la vSNc chez la souris adulte. En effet, l’inactivation du gène Rgs6 chez les souris produit une dégénérescence tardive de ces derniers, en association avec une baisse d’expression de Pitx3 et de ses gènes cibles. L’ensemble de ce travail ouvre de nouvelles perspectives pour la compréhension moléculaire de la biologie des neurones DAm dans l’état normal et la maladie

Subset-specific expression signatures of mDA neurons.

<p>(A) Differentially expressed probesets were clustered in an unbiased manner into three-dimensional Cartesian-type coordinates defined according to the relative enrichment attributes for each probeset in the comparisons between SNc WT/VTA WT, SNc KO/SNc WT and VTA KO/VTA WT. Each cluster was named according to the observed preferential expression pattern (label and shown in red in diagrams). The heatmaps show relative enrichments for 20 genes that are highly enriched and/or that have documented functions (the complete gene lists for each subset is provided in <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1004863#pgen.1004863.s005" target="_blank">Table S1</a>). The total number of genes (not probesets) in each cluster is indicated together with the number of those that are either activated or repressed by Pitx3 based on the VTA KO/WT comparison. (B) qRT-PCR quantification of mRNAs with preferential expression in either SN or VTA assessed in FACS-sorted cells from <i>WT</i> or <i>Pitx3</i>^−/− (KO) tissues. mRNA levels are normalized relative to Gapdh mRNA. Data are shown as means ± S.D.</p