Chapter 2 Genetics at the Cell Level

Image-based and sequencing-based spatial technologies can currently be used to locate cells anatomically within a tissue; the Human Vell Atlas allows its users to navigate the human body at various levels of resolution to identify patterns and interactions among its fundamental elements, zooming in and out depending on the research goals. Integrating data types or simultaneously measuring multiple modalities is invaluable when defining cell identities

El delito de genocidio, con especial estudio del caso Núremberg

It was during the course of World War II that the most cruel and heinous crimes have occurred. That brought with it a change to international law, being the reason why the typification of acts that until now were not regulated, nor did a conviction be established for them. It was, therefore, the need for a legal instrument to punish these and the will to prevent crimes like those committed from being committed again that motivated an action of legal creation, whose main objective was to ensure international peace and security, and the collaboration of all nations was needed to achieve it. This was how the typification of genocide occurred, whose elements of the type are, not only the realization of typical conduct, but also realizing this with a special intention and awareness of destroying a particular group. In this way, we will proceed to the study of the Nuremberg Statute and the legacy that led to the condemnation of the acts committed during World War II, making a special deal of the crime of genocide.Ha sido durante el transcurso de la Segunda Guerra Mundial cuando se han producido los crímenes más crueles y atroces jamás vistos. Ello trajo consigo un cambio para el derecho internacional, al ser la razón por la cual se procedió a la tipificación de actos que hasta el momento no eran regulados, ni se establecía una condena por ellos. Fue, por tanto, la necesidad de un instrumento jurídico que castigara estos y la voluntad de evitar que crímenes como los cometidos se volvieran a cometer que motivaron una acción de creación jurídica, cuyo principal objetivo era garantizar la paz y seguridad internacional, siendo necesaria la colaboración de todas las naciones para conseguirlo. Así fue como se produjo la tipificación del delito de genocidio, cuyos elementos del tipo son, no sólo la realización de la conducta típica, sino también la realización de esta con una especial intención y consciencia de destruir a un grupo determinado. De esta manera, procederemos al estudio del Estatuto de Núremberg y el legado que conllevó la condena de los actos cometidos durante la II Guerra Mundial, haciendo un especial detenimiento en el delito de genocidio

Negative regulation of urokinase receptor activity by a GPI-specific phospholipase C in breast cancer cells.

The urokinase receptor (uPAR) is a glycosylphosphatidylinositol (GPI)-anchored protein that promotes tissue remodeling, tumor cell adhesion, migration and invasion. uPAR mediates degradation of the extracellular matrix through protease recruitment and enhances cell adhesion, migration and signaling through vitronectin binding and interactions with integrins. Full-length uPAR is released from the cell surface, but the mechanism and significance of uPAR shedding remain obscure. Here we identify transmembrane glycerophosphodiesterase GDE3 as a GPI-specific phospholipase C that cleaves and releases uPAR with consequent loss of function, whereas its homologue GDE2 fails to attack uPAR. GDE3 overexpression depletes uPAR from distinct basolateral membrane domains in breast cancer cells, resulting in a less transformed phenotype, it slows tumor growth in a xenograft model and correlates with prolonged survival in patients. Our results establish GDE3 as a negative regulator of the uPAR signaling network and, furthermore, highlight GPI-anchor hydrolysis as a cell-intrinsic mechanism to alter cell behavior

Single-cell imaging of α and β cell metabolic response to glucose in living human Langerhans islets

Here we use a combination of two-photon Fluorescence Lifetime Imaging Microscopy (FLIM) of NAD(P)H free/bound ratio in living HIs with post-fixation, immunofluorescence-based, cell-type identification. FLIM allowed to measure variations in the NAD(P)H free/bound ratio induced by glucose; immunofluorescence data allowed to identify single α and β cells; finally, matching of the two datasets allowed to assign metabolic shifts to cell identity. 312 α and 654 β cells from a cohort of 4 healthy donors, 15 total islets, were measured. Both α and β cells display a wide spectrum of responses, towards either an increase or a decrease in NAD(P)H free/bound ratio. Yet, if single-cell data are averaged according to the respective donor and correlated to donor insulin secretion power, a non-random distribution of metabolic shifts emerges: robust average responses of both α and β cells towards an increase of enzyme-bound NAD(P)H belong to the donor with the lowest insulin-secretion power; by contrast, discordant responses, with α cells shifting towards an increase of free NAD(P)H and β cells towards an increase of enzyme-bound NAD(P)H, correspond to the donor with the highest insulin-secretion power. Overall, data reveal neat anti-correlation of tissue metabolic responses with respect to tissue insulin secretion power

Detection of circulating immunosuppressive cytokines in malignant pleural mesothelioma patients for prognostic stratification.

Abstract Background No data on circulating biomarkers for the prognostic stratification of Malignant Pleural Mesothelioma (MPM) patients are available. We prospectively explored the prognostic role of circulating monocyte and cytokine levels and their dynamic change during chemotherapy. Patients and Methods MPM patients receiving a first line treatment based on a platinum compound plus pemetrexed were eligible. Blood samples were collected at the baseline and at the end of induction chemotherapy. CCL-2, IL-10 and TGF-β levels in plasma were quantified by Enzyme-Linked Immunosorbent Assay (ELISA); white blood cells, monocytes and platelets were evaluated by blood count test. Results Thirty-one patients were included in the study. Median overall survival (OS) was 12.13 months versus 9.6 months in patients with lower and higher monocytes count, respectively (p value = 0.02). We further stratified patients according to a combined score based on the association of IL-10, TGF-β levels and monocytes count. High combined score was associated with shorter OS and PFS in univariate and multivariate analysis. Chemotherapy induced an increase in monocytes, IL-10, but not TGF-β levels. Conclusion The prognostic value of circulating levels of multiple immunosuppressive cytokines and inflammatory cells should be confirmed in a wider validation set of MPM patients

Investigating the mechanism of action of DNA-loaded PEGylated lipid nanoparticles

PEGylated lipid nanoparticles (LNPs) are commonly used to deliver bioactive molecules, but the role of PEGylation in DNA-loaded LNP interactions at the cellular and subcellular levels remains poorly understood. In this study, we investigated the mechanism of action of DNA-loaded PEGylated LNPs using gene reporter technologies, dynamic light scattering (DLS), synchrotron small angle X-ray scattering (SAXS), and fluorescence confocal microscopy (FCS). We found that PEG has no significant impact on the size or nanostructure of DNA LNPs but reduces their zeta potential and interaction with anionic cell membranes. PEGylation increases the structural stability of LNPs and results in lower DNA unloading. FCS experiments revealed that PEGylated LNPs are internalized intact inside cells and largely shuttled to lysosomes, while unPEGylated LNPs undergo massive destabilization on the plasma membrane. These findings can inform the design, optimization, and validation of DNA-loaded LNPs for gene delivery and vaccine development

Combining Multi-Task Learning and Multi-Channel Variational Auto-Encoders to Exploit Datasets with Missing Observations -Application to Multi-Modal Neuroimaging Studies in Dementia

The joint modeling of neuroimaging data across multiple datasets requires to consistently analyze high-dimensional and heterogeneous information in presence of often non-overlapping sets of views across data samples (e.g. imaging data, clinical scores, biological measurements). This analysis is associated with the problem of missing information across datasets, which can happen in two forms: missing at random (MAR), when the absence of a view is unpredictable and does not depend on the dataset (e.g. due to data corruption); missing not at random (MNAR), when a specific view is absent by design for a specific dataset. In order to take advantage of the increased variability and sample size when pooling together observations from many cohorts and at the same time cope with the ubiquitous problem of missing information, we propose here a multi-task generative latent-variable model where the common variability across datasets stems from the estimation of a shared latent representation across views. Our formulation allows to retrieve a consistent latent representation common to all views and datasets, even in the presence of missing information. Simulations on synthetic data show that our method is able to identify a common latent representation of multi-view datasets, even when the compatibility across datasets is minimal. When jointly analyzing multi-modal neuroimaging and clinical data from real independent dementia studies, our model is able to mitigate the absence of modalities without having to discard any available information. Moreover, the common latent representation inferred with our model can be used to define robust classifiers gathering the combined information across different datasets. To conclude, both on synthetic and real data experiments, our model compared favorably to state of the art benchmark methods, providing a more powerful exploitation of multi-modal observations with missing views

SimulAD: A dynamical model for personalized simulation and disease staging in Alzheimer's disease

International audienceSimulAD is a computational disease progression model (DPM) originally developed on the ADNI database to simulate the evolution of clinical and imaging markers characteristic of AD, and to quantify the disease severity (DS) of a subject. In this work, we assessed the validity of this estimated DS, as well as the generalization of the DPM, by applying SimulAD on a new cohort from the Geneva Memory Center (GMC). The differences between the estimated DS of healthy, mild cognitive impairment and AD dementia groups were statistically significant (p-values < 0.05; d ≥ 0.8). DS correlated with MMSE (ρ =-0.55), hippocampal atrophy (ρ =-0.62), glucose hypometabolism (ρ =-0.67), amyloid burden (ρ = 0.31) and tau deposition (ρ = 0.62) (p-values < 0.01). Based on the dynamics estimated on the ADNI cohort, we simulated a DPM for the subjects of the GMC cohort. The difference between the temporal evolution of similar biomarkers simulated on the ADNI and GMC cohorts remained below 10%. This study illustrates the robustness and good generalization of SimulAD, highlighting its potential for clinical and pharmaceutical studies

Integration of bovine herpesvirus 4 genome into cultured persistently infected host cell genome

Persistent infection of macrophages with bovine herpesvirus 4 (BoHV-4) has been proposed to play a secondary causal role, along with bacterial infection, in bovine post-partum metritis. Mechanisms of maintenance of BoHV-4 persistent infection are not understood. We previously generated in vitro models of BoHV-4 persistent infection in human rhadomyosarcoma and bovine macrophage cell lines by drug selection of cells infected with BoHV-4 carrying a drug-resistance marker, and demonstrated circular episomal BoHV-4 genomes. In the present study, we used fluorescent in situ hybridization (FISH) to demonstrate BoHV-4 genomes also integrated into the genomes of these persistently infected cells

DMRT1 regulates human germline commitment

Germline commitment following primordial germ cell (PGC) specification during early human development establishes an epigenetic programme and competence for gametogenesis. Here we follow the progression of nascent PGC-like cells derived from human embryonic stem cells in vitro. We show that switching from BMP signalling for PGC specification to Activin A and retinoic acid resulted in DMRT1 and CDH5 expression, the indicators of migratory PGCs in vivo. Moreover, the induction of DMRT1 and SOX17 in PGC-like cells promoted epigenetic resetting with striking global enrichment of 5-hydroxymethylcytosine and locus-specific loss of 5-methylcytosine at DMRT1 binding sites and the expression of DAZL representing DNA methylation-sensitive genes, a hallmark of the germline commitment programme. We provide insight into the unique role of DMRT1 in germline development for advances in human germ cell biology and in vitro gametogenesis