The impact of phosphoinositide metabolic enzymes in glioblastoma: a tale of phosphoinositide-specific phospholipase C PLCβ1 and 5-phosphatase SKIP

Background: Glioblastoma multiforme (GBM) is one of the deadliest and most aggressive form of primary brain tumor. Unfortunately, current GBM treatment therapies are not effective in treating GBM patients. They usually experience very poor prognosis with a median survival of approximately 12 months. Only 3-5% survive up to 3 years or more. A large-scale gene profile study revealed that several genes involved in essential cellular processes are altered in GBM, thus, explaining why existing therapies are not effective. The survival of GBM patients depends on understanding the molecular and key signaling events associated with these altered physiological processes in GBM. Phosphoinositides (PI) form just a tiny fraction of the total lipid content in humans, however they are implicated in almost all essential biological processes, such as acting as second messengers in spatio-temporal regulation of cell signaling, cytoskeletal reorganization, cell adhesion, migration, apoptosis, vesicular trafficking, differentiation, cell cycle and post-translational modifications. Interestingly, these essential processes are altered in GBM. More importantly, incoming reports have associated PI metabolism, which is mediated by several PI phosphatases such as SKIP, lipases such as PLCβ1, and other kinases, to regulate GBM associated cellular processes. Even as PLCβ1 and SKIP are involved in regulating aberrant cellular processes in several other cancers, very few studies, of which majority are in-silico-based, have focused on the impact of PLCβ1 and SKIP in GBM. Hence, it is important to employ clinical, in vitro, and in vivo GBM models to define the actual impact of PLCβ1 and SKIP in GBM. AIM: Since studies of PLCβ1 and SKIP in GBM are limited, this study aimed at determining the pathological impact of PI metabolic enzymes, PLCB1 and SKIP, in GBM patient samples, GBM cell line models, and xenograft models for SKIP. Results: For the first time, this study confirmed through qPCR that PLCβ1 gene expression is lower in human GBM patient samples. Moreover, PLCβ1 gene expression inversely correlates with pathological grades of glioma; it decreases as glioma grades increases or worsens. Silencing PLCβ1 in U87MG GBM cells produces a dual impact in GBM by participating in both pro-tumoral and anti-tumoral roles. PLCβ1 knockdown cells were observed to have more migratory abilities, increased cell to extracellular matrix (ECM) adhesion, transition from epithelial phenotype to mesenchymal phenotype through the upregulation of EMT transcription factors Twist1 and Slug, and mesenchymal marker, vimentin. On the other hand, p-Akt and p-mTOR protein expression were downregulated in PLCβ1 knockdown cells. Thus, the oncogenic pathway PI3K/Akt/mTOR pathway is inhibited during PLCβ1 knockdown. Consistently, cell viability in PLCβ1 knockdown cells were significantly decreased compared to controls. As for SKIP, this study demonstrated that about 48% of SKIP colocalizes with nuclear PtdIns(4,5)P2 to nuclear speckles and that SKIP knockdown alters nuclear PtdIns(4,5)P2 in a cell-type dependent manner. In addition, SKIP silencing increased tumor volume and weight in xenografts than controls by reducing apoptosis and increasing viability. All in all, these data confirm that PLCβ1 and SKIP are involved in GBM pathology and a complete understanding of their roles in GBM may be beneficial.

A methodological approach for the performance optimization of transient seepage models through inverse analysis

The topic of the Ph.D project focuses on the modelling of the soil-water dynamics inside an instrumented embankment section along Secchia River (Cavezzo (MO)) in the period from 2017 to 2018 and the quantification of the performance of the direct and indirect simulations . The commercial code Hydrus2D by Pc-Progress has been chosen to run the direct simulations. Different soil-hydraulic models have been adopted and compared. The parameters of the different hydraulic models are calibrated using a local optimization method based on the Levenberg - Marquardt algorithm implemented in the Hydrus package. The calibration program is carried out using different types of dataset of observation points, different weighting distributions, different combinations of optimized parameters and different initial sets of parameters. The final goal is an in-depth study of the potentialities and limits of the inverse analysis when applied to a complex geotechnical problem as the case study. The second part of the research focuses on the effects of plant roots and soil-vegetation-atmosphere interaction on the spatial and temporal distribution of pore water pressure in soil. The investigated soil belongs to the West Charlestown Bypass embankment, Newcastle, Australia, that showed in the past years shallow instabilities and the use of long stem planting is intended to stabilize the slope. The chosen plant species is the Malaleuca Styphelioides, native of eastern Australia. The research activity included the design and realization of a specific large scale apparatus for laboratory experiments. Local suction measurements at certain intervals of depth and radial distances from the root bulb are recorded within the vegetated soil mass under controlled boundary conditions. The experiments are then reproduced numerically using the commercial code Hydrus 2D. Laboratory data are used to calibrate the RWU parameters and the parameters of the hydraulic model.

GÉOARCHÉOLOGIE DU VILLAGE: Variabilité des modes d’occupation de la fin du Néolithique à l’âge du Bronze en Italie et en France méridionale. Implications culturelles

Entre la fin du Néolithique et l’âge du Bronze, la présence d’habitats groupés de type village est un phénomène diffus, tant en Italie qu’en France méridionale. Néanmoins, la prise en compte de la variabilité des formes de la stratification des sites interroge. En quoi l’enregistrement sédimentaire des sols d’habitat permet-il d’appréhender la question de l’organisation villageoise et de sa variabilité entre la fin du Néolithique et l’âge du Bronze ? Quelle image cet enregistrement sédimentaire donne-t-il de l’organisation sociale et économique du village ? Afin d’aborder ces questions, nous avons choisi de mener une étude géoarchéologique sur des sites de formes différentes, issus de contextes chrono-culturels et environnementaux variés. La démarche, fondée sur l’emploi de la micromorphologie des sols en tant qu’outil analytique, vise à caractériser l’organisation spatio-temporelle des sols d’occupation à l’échelle du site, selon une approche spatiale des processus de formation de la stratification archéologique. L’élaboration d’un modèle, qui repose sur une classification des micro-faciès sédimentaires selon le système d’activité, et son application à des sites-laboratoires permettent de qualifier les techniques de construction en terre, l’usage du sol et les dynamiques d’occupation propres à chaque site, dans le but de déterminer les comportements socio-économiques et les spécificités du mode de vie villageois enregistrées par les sols. Cette approche permet d’évaluer les constantes et les variables qui qualifient les différents types d’occupation. Le sol, conçu comme matérialité de l’espace villageois, devient ainsi un témoignage direct de la variabilité culturelle et des différentes formes d’organisation des communautés de la fin du Néolithique et de l’âge du Bronze.

Preharvest factors affecting quality and antioxidant levels on Abate Fétel pears: study of superficial scald with a multivariate statistical approach and neural networks

There are only a few insights concerning the influence that agronomic and management variability may have on superficial scald (SS) in pears. Abate Fétel pears were picked during three seasons (2018, 2019 and 2020) from thirty commercial orchards in the Emilia Romagna region, Italy. Using a multivariate statistical approach, high heterogeneity between farms for SS development after cold storage with regular atmosphere was demonstrated. Indeed, some factors seem to affect SS in all growing seasons: high yields, soil texture, improper irrigation and Nitrogen management, use of plant growth regulators, late harvest, precipitations, Calcium and cow manure, presence of nets, orchard age, training system and rootstock. Afterwards, we explored the spatio/temporal variability of fruit attributes in two pear orchards. Environmental and physiological spatial variables were recorded by a portable RTK GPS. High spatial variability of the SS index was observed. Through a geostatistical approach, some characteristics, including soil electrical conductivity and fruit size, have been shown to be negatively correlated with SS. Moreover, regression tree analyses were applied suggesting the presence of threshold values of antioxidant capacity, total phenolic content, and acidity against SS. High pulp firmness and IAD values before storage, denoting a more immature fruit, appeared to be correlated with low SS. Finally, a convolution neural networks (CNN) was tested to detect SS and the starch pattern index (SPI) in pears for portable device applications. Preliminary statistics showed that the model for SS had low accuracy but good precision, and the CNN for SPI denoted good performances compared to the Ctifl and Laimburg scales. The major conclusion is that Abate Fétel pears can potentially be stored in different cold rooms, according to their origin and quality features, ensuring the best fruit quality for the final consumers. These results might lead to a substantial improvement in the Italian pear industry.

Mathematical methods to analyze and interpret calcium signals of astrocytes

Astrocytes are the most numerous glial cell type in the mammalian brain and permeate the entire CNS interacting with neurons, vasculature, and other glial cells. Astrocytes display intracellular calcium signals that encode information about local synaptic function, distributed network activity, and high-level cognitive functions. Several studies have investigated the calcium dynamics of astrocytes in sensory areas and have shown that these cells can encode sensory stimuli. Nevertheless, only recently the neuro-scientific community has focused its attention on the role and functions of astrocytes in associative areas such as the hippocampus. In our first study, we used the information theory formalism to show that astrocytes in the CA1 area of the hippocampus recorded with 2-photon fluorescence microscopy during spatial navigation encode spatial information that is complementary and synergistic to information encoded by nearby "place cell" neurons. In our second study, we investigated various computational aspects of applying the information theory formalism to astrocytic calcium data. For this reason, we generated realistic simulations of calcium signals in astrocytes to determine optimal hyperparameters and procedures of information measures and applied them to real astrocytic calcium imaging data. Calcium signals of astrocytes are characterized by complex spatiotemporal dynamics occurring in subcellular parcels of the astrocytic domain which makes studying these cells in 2-photon calcium imaging recordings difficult. However, current analytical tools which identify the astrocytic subcellular regions are time consuming and extensively rely on user-defined parameters. Here, we present Rapid Astrocytic calcium Spatio-Temporal Analysis (RASTA), a novel machine learning algorithm for spatiotemporal semantic segmentation of 2-photon calcium imaging recordings of astrocytes which operates without human intervention. We found that RASTA provided fast and accurate identification of astrocytic cell somata, processes, and cellular domains, extracting calcium signals from identified regions of interest across individual cells and populations of hundreds of astrocytes recorded in awake mice.

Moving toward stock units identification based on spatial population structure of marine species in the Mediterranean Sea and adjacent waters through multidisciplinary and holistic approaches for the sustainability of fisheries resources

Investigating stock identity of marine species in a multidisciplinary holistic approach can reveal patterns of complex spatial population structure and signatures of potential local adaptation. The population structure of common sole (Solea solea) in the Mediterranean Sea was delineated using genomic and otolith data, including single nucleotide polymorphisms (SNPs) markers and otolith data. SNPs were correlated with environmental and spatial variables to evaluate the impact of these features on the actual genetic population structure. Integrated holistic approach was applied to combine the tracers with different spatio-temporal scales. SNPs data was also used to illustrate the population structure of European hake (Merluccius merluccius) within the Alboran Sea, extending into the neighboring Mediterranean Sea and Atlantic Ocean. The aim was to identify patterns of neutral and potential adaptive genetic variation by applying seascape genomic framework. Results from both genetic and otolith data suggested significant divergence among putative populations of common sole, confirming a clear separation between Western, Adriatic Sea and Eastern Mediterranean Sea. Evidence of fine-scale population structure in the Western Mediterranean Sea was observed at outlier loci level and in the Adriatic. Our study not only indicates that separation among Mediterranean sole population is led primarily by neutral processes, but it also suggests the presence of local adaptation influenced by environmental and spatial factors. The holistic approach by considering the spatio-temporal scales of variation confirmed that the same pattern of separation between these geographical sites is currently occurring and has occurred for many generations. Results showed the occurrence of population structure in Merluccius merluccius by detecting westward–eastward differentiation among populations and distinct subgroups at a fine geographical scale using outlier SNPs. These results enhance the knowledge of the population structure of commercially relevant species to support the application of spatial stock assessment models, including a redefinition of fishery management units.

Idee, Natur, Geist. Schellings allgemeine Ontologie des Wirklichen nach der "Darstellung des Naturprocesses" von 1843/44

Ziel dieser Dissertation ist es, die grundlegenden philosophisch-theoretischen Implikationen von Schellings letzter systematischer Darlegung seiner Naturphilosophie nach dem Berliner Textfragment von 1843/44, der “Darstellung des Naturprocesses”, zu untersuchen. Angesichts der sich zwischen den 1830 und den 1860 Jahren in Berlin abzeichnenden neuen intellektuellen Tendenzen und der Entwicklungen in den Naturwissenschaften legt Schelling hier die Grundlagen für eine allgemeine Ontologie des Wirklichen in kritischer Auseinandersetzung mit Kants transzendentalem Idealismus. Innerhalb des systematischen Horizonts der "apriorischen Vernunftwissenschaft" oder "negativen Philosophie" stellt er im ersten Teil seines Werkes die Prinzipien fest, die die „Idee des Existierenden“ ausmachen, und beschreibt die rationale Operation, die durchgeführt werden muss, um zum Gedanken einer „Welt außer der Idee“ zu gelangen. Die philosophisch-systematischen Annahmen, die mit dem Übergang von der bloßen Idee des Existierenden zum Gedanken der außeridealen Welt verbunden sind, werden im ersten Kapitel dieser Dissertation untersucht. Im zweiten Teil seines Werkes definiert Schelling durch eine detaillierte Analyse von Kants Transzendentalen Ästhetik den Raum als diejenige Form, in der uns die Existenzen als voneinander getrennt vorstellen lassen. Obwohl der Zeitbegriff von Schelling nur am Rande behandelt wird, trägt er zusammen mit dem Raum dazu bei, die erste ontologische Grundstruktur der außeridealen Welt zu definieren. Die Analyse von Schellings Konzeption der raumzeitlichen Grundstruktur der außeridealen Welt stellt das Thema des zweiten Kapitels dieser Dissertation dar. Schließlich bestimmt Schelling im dritten Teil seines Werkes die Finalität als diejenige Kausalitätsform, die es ermöglicht, die außerideale Welt als einen werdenden Kontext zu verstehen, dessen Entwicklungsstufen die siderische Welt, die unorganische Welt und die organische Welt sind. Die Schelling‘sche Definition der Teleologie der Natur als zweite ontologische Grundstruktur der außeridealen Welt ist das Thema des dritten Kapitels dieser Dissertation.