Letteratura scientifica selezionata sul tema "In cellulo and in vivo models"
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Articoli di riviste sul tema "In cellulo and in vivo models":
1
Morant, Laura, Maria-Luise Petrovic-Erfurth e Albena Jordanova. "An Adapted GeneSwitch Toolkit for Comparable Cellular and Animal Models: A Proof of Concept in Modeling Charcot-Marie-Tooth Neuropathy". International Journal of Molecular Sciences 24, n. 22 (9 novembre 2023): 16138. http://dx.doi.org/10.3390/ijms242216138.
Abstract (sommario):
Investigating the impact of disease-causing mutations, their affected pathways, and/or potential therapeutic strategies using disease modeling often requires the generation of different in vivo and in cellulo models. To date, several approaches have been established to induce transgene expression in a controlled manner in different model systems. Several rounds of subcloning are, however, required, depending on the model organism used, thus bringing labor-intensive experiments into the technical approach and analysis comparison. The GeneSwitch™ technology is an adapted version of the classical UAS-GAL4 inducible system, allowing the spatial and temporal modulation of transgene expression. It consists of three components: a plasmid encoding for the chimeric regulatory pSwitch protein, Mifepristone as an inducer, and an inducible plasmid. While the pSwitch-containing first plasmid can be used both in vivo and in cellulo, the inducible second plasmid can only be used in cellulo. This requires a specific subcloning strategy of the inducible plasmid tailored to the model organism used. To avoid this step and unify gene expression in the transgenic models generated, we replaced the backbone vector with standard pUAS-attB plasmid for both plasmids containing either the chimeric GeneSwitch™ cDNA sequence or the transgene cDNA sequence. We optimized this adapted system to regulate transgene expression in several mammalian cell lines. Moreover, we took advantage of this new system to generate unified cellular and fruit fly models for YARS1-induced Charco–Marie–Tooth neuropathy (CMT). These new models displayed the expected CMT-like phenotypes. In the N2a neuroblastoma cells expressing YARS1 transgenes, we observed the typical “teardrop” distribution of the synthetase that was perturbed when expressing the YARS1CMT mutation. In flies, the ubiquitous expression of YARS1CMT induced dose-dependent developmental lethality and pan-neuronal expression caused locomotor deficit, while expression of the wild-type allele was harmless. Our proof-of-concept disease modeling studies support the efficacy of the adapted transgenesis system as a powerful tool allowing the design of studies with optimal data comparability.
2
Dasnur Nanjappa, Madhusudan, Anup Pandith, Svetlana Sankaran, Dorothy Priyanka Dorairaj, Anusha Anjaneya Reddy e Hari Prasad Badubanahalli Ramesh. "Recent Advancements in Developments of Novel Fluorescent Probes: In Cellulo Recognitions of Alkaline Phosphatases". Symmetry 14, n. 8 (9 agosto 2022): 1634. http://dx.doi.org/10.3390/sym14081634.
Abstract (sommario):
Alkaline phosphatase (ALP) is one of the vital phospho-ester bond cleaving biocatalysts that has inevitable significance in cellular systems, viz., early-stage osteoblast differentiation, cell integrity in tissues, bone mineralization, cancer biomarker, liver dysfunction, cellular osmotic pressure, protein folding and many more. Variation from optimal levels of ALP in intra and extracellular fluids can cause severe diseases, including death. Due to these reasons, ALP is considered as a vital biomarker for various preclinical and medical diagnosis. Fluorescence image-based diagnosis is the most widely used method, owing to its simplicity, robustness, non-invasive properties and excellent spatio-temporal resolution (up to the nM/pM level), as compared to conventional analytical techniques, such as the electroanalytical method, nuclear magnetic resonance (NMR) and high-performance liquid chromatography (HPLC). Most of the reviews reported for ALP’s recognition in the literature scarcely explain the structurally related, photophysical and biophysical parameters; and the sub-cellular localizations. Considering these facts, in order to enhance the opto-analytical parameters of fluorescence-based diagnostic materials at the cellular level, herein we have systematically documented recent developments in the opto-analytical capabilities of quencher-free probes for ALP, used in in vitro (biological buffers) to in cellulo conditions, along with in vivo models.
3
Sailer, Alexander, Franziska Ermer, Yvonne Kraus, Rebekkah Bingham, Ferdinand H. Lutter, Julia Ahlfeld e Oliver Thorn-Seshold. "Potent hemithioindigo-based antimitotics photocontrol the microtubule cytoskeleton in cellulo". Beilstein Journal of Organic Chemistry 16 (27 gennaio 2020): 125–34. http://dx.doi.org/10.3762/bjoc.16.14.
Abstract (sommario):
Background: Hemithioindigo is a promising molecular photoswitch that has only recently been applied as a photoswitchable pharmacophore for control over bioactivity in cellulo. Uniquely, in contrast to other photoswitches that have been applied to biology, the pseudosymmetric hemithioindigo scaffold has allowed the creation of both dark-active and lit-active photopharmaceuticals for the same binding site by a priori design. However, the potency of previous hemithioindigo photopharmaceuticals has not been optimal for their translation to other biological models. Results: Inspired by the structure of tubulin-inhibiting indanones, we created hemithioindigo-based indanone-like tubulin inhibitors (HITubs) and optimised their cellular potency as antimitotic photopharmaceuticals. These HITubs feature reliable and robust visible-light photoswitching and high fatigue resistance. The use of the hemithioindigo scaffold also permitted us to employ a para-hydroxyhemistilbene motif, a structural feature which is denied to most azobenzenes due to the negligibly short lifetimes of their metastable Z-isomers, which proved crucial to enhancing the potency and photoswitchability. The HITubs were ten times more potent than previously reported hemithioindigo photopharmaceutical antimitotics in a series of cell-free and cellular assays, and allowed robust photocontrol over tubulin polymerisation, microtubule (MT) network structure, cell cycle, and cell survival. Conclusions: HITubs represent a powerful addition to the growing toolbox of photopharmaceutical reagents for MT cytoskeleton research. Additionally, as the hemithioindigo scaffold allows photoswitchable bioactivity for substituent patterns inaccessible to the majority of current photopharmaceuticals, wider adoption of the hemithioindigo scaffold may significantly expand the scope of cellular and in vivo targets addressable by photopharmacology.
4
Thompson, John W., Omar Elwardany, David J. McCarthy, Dallas L. Sheinberg, Carlos M. Alvarez, Ahmed Nada, Brian M. Snelling, Stephanie H. Chen, Samir Sur e Robert M. Starke. "In vivo cerebral aneurysm models". Neurosurgical Focus 47, n. 1 (luglio 2019): E20. http://dx.doi.org/10.3171/2019.4.focus19219.
Abstract (sommario):
Cerebral aneurysm rupture is a devastating event resulting in subarachnoid hemorrhage and is associated with significant morbidity and death. Up to 50% of individuals do not survive aneurysm rupture, with the majority of survivors suffering some degree of neurological deficit. Therefore, prior to aneurysm rupture, a large number of diagnosed patients are treated either microsurgically via clipping or endovascularly to prevent aneurysm filling. With the advancement of endovascular surgical techniques and devices, endovascular treatment of cerebral aneurysms is becoming the first-line therapy at many hospitals. Despite this fact, a large number of endovascularly treated patients will have aneurysm recanalization and progression and will require retreatment. The lack of approved pharmacological interventions for cerebral aneurysms and the need for retreatment have led to a growing interest in understanding the molecular, cellular, and physiological determinants of cerebral aneurysm pathogenesis, maturation, and rupture. To this end, the use of animal cerebral aneurysm models has contributed significantly to our current understanding of cerebral aneurysm biology and to the development of and training in endovascular devices. This review summarizes the small and large animal models of cerebral aneurysm that are being used to explore the pathophysiology of cerebral aneurysms, as well as the development of novel endovascular devices for aneurysm treatment.
5
Lepore Signorile, Martina, Valentina Grossi, Candida Fasano, Giovanna Forte, Vittoria Disciglio, Paola Sanese, Katia De Marco et al. "c-MYC Protein Stability Is Sustained by MAPKs in Colorectal Cancer". Cancers 14, n. 19 (4 ottobre 2022): 4840. http://dx.doi.org/10.3390/cancers14194840.
Abstract (sommario):
c-MYC is one of the most important factors involved in colorectal cancer (CRC) initiation and progression; indeed, it is found to be upregulated in up to 80% of sporadic cases. During colorectal carcinogenesis, c-MYC is maintained upregulated through β-catenin-mediated transcriptional activation and ERK-mediated post-translational stabilization. Our data demonstrate that p38α, a kinase involved in CRC metabolism and survival, contributes to c-Myc protein stability. Moreover, we show that p38α, like ERK, stabilizes c-MYC protein levels by preventing its ubiquitination. Of note, we found that p38α phosphorylates c-MYC and interacts with it both in vitro and in cellulo. Extensive molecular analyses in the cellular and in vivo models revealed that the p38α kinase inhibitors, SB202190 and ralimetinib, affect c-MYC protein levels. Ralimetinib also exhibited a synthetic lethality effect when used in combination with the MEK1 inhibitor trametinib. Overall, our findings identify p38α as a promising therapeutic target, acting directly on c-MYC, with potential implications for countering c-MYC-mediated CRC proliferation, metastatic dissemination, and chemoresistance.
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Mode, Selma, Maren Ketterer, Maxime Québatte e Christoph Dehio. "Antibiotic persistence of intracellular Brucella abortus". PLOS Neglected Tropical Diseases 16, n. 7 (26 luglio 2022): e0010635. http://dx.doi.org/10.1371/journal.pntd.0010635.
Abstract (sommario):
Background Human Brucellosis caused by the facultative intracellular pathogen Brucella spp. is an endemic bacterial zoonosis manifesting as acute or chronic infections with high morbidity. Treatment typically involves a combination therapy of two antibiotics for several weeks to month, but despite this harsh treatment relapses occur at a rate of 5–15%. Although poor compliance and reinfection may account for a fraction of the observed relapse cases, it is apparent that the properties of the infectious agent itself may play a decisive role in this phenomenon. Methodology/Principal findings We used B. abortus carrying a dual reporter in a macrophage infection model to gain a better understanding of the efficacy of recommended therapies in cellulo. For this we used automated fluorescent microscopy as a prime read-out and developed specific CellProfiler pipelines to score infected macrophages at the population and the single cell level. Combining microscopy of constitutive and induced reporters with classical CFU determination, we quantified the protective nature of the Brucella intracellular lifestyle to various antibiotics and the ability of B. abortus to persist in cellulo despite harsh antibiotic treatments. Conclusion/Significance We demonstrate that treatment of infected macrophages with antibiotics at recommended concentrations fails to fully prevent growth and persistence of B. abortus in cellulo, which may be explained by a protective nature of the intracellular niche(s). Moreover, we show the presence of bona fide intracellular persisters upon antibiotic treatment, which are metabolically active and retain the full infectious potential, therefore constituting a plausible reservoir for reinfection and relapse. In conclusion, our results highlight the need to extend the spectrum of models to test new antimicrobial therapies for Brucellosis to better reflect the in vivo infection environment, and to develop therapeutic approaches targeting the persister subpopulation.
7
Qiu, Xudong, Seth T. Gammon, James R. Johnson, Federica Pisaneschi, Steven W. Millward, Edward M. Barnett e David Piwnica-Worms. "Apoptosis Detection in Retinal Ganglion Cells Using Quantitative Changes in Multichannel Fluorescence Colocalization". Biosensors 12, n. 9 (28 agosto 2022): 693. http://dx.doi.org/10.3390/bios12090693.
Abstract (sommario):
KcapTR488 is a dual-fluorophore peptide sensor for the real-time reporting of programmed cell death by fluorescence imaging. KcapTR488 contains a nuclear localization sequence (NLS) conjugated with Texas Red, a caspase-cleavable sequence (DEVD), and a C-terminus conjugated to Alexa Fluor 488 (AF488). The synthesis and preliminary evaluation in cellulo of KcapTR488 for monitoring cell death by fluorescence imaging has been previously reported, but its utility in vivo has yet to be tested or validated. Herein, in vitro solution experiments verified the intramolecular fluorescence resonance energy transfer (FRET) between the two fluorophores and enabled a quantitative analysis of enzyme rates and selectivity. The sensor delivery kinetics in live rat models were quantified by ex vivo fluorescence microscopy. Studies in healthy control retinas demonstrated that KcapTR488 concentrated in the nucleus of retinal ganglion cells (RGC), with a strong colocalization of red and green fluorescence signals producing robust FRET signals, indicating an intact reporter. By contrast, using an acute but mild NMDA-induced retinal injury model, dual-color confocal ex vivo microscopy of cleaved KcapTR488 identified sensor activation as early as 2 h after injection. Quantitative changes in fluorescence colocalization were superior to changes in FRET for monitoring injury progression. Longitudinal monitoring revealed that the NLS-Texas Red fragment of the cleaved sensor moved out of the cell body, down the axon, and exited the retina, consistent with anterograde axonal transport. Thus, KcapTR488 may be a powerful tool to study RGC death pathways in live preclinical models of glaucoma.
8
Reinhardt, Christoph, e Heiko Rühl. "Animal and Cellular Models in Thrombosis and Hemostasis". Hämostaseologie 43, n. 05 (ottobre 2023): 319–20. http://dx.doi.org/10.1055/a-2031-7975.
Abstract (sommario):
Abstract Standardized In Vitro and In Vivo Model Systems to Simplify Complexity—That's How We Learn The discovery of new target molecules and translational progress in the development and refinement of antithrombotic therapies as well as the improved treatment of bleeding disorders strongly relies on standardized ex vivo and in vivo models that closely resemble the respective human pathologies. The standardization of these models requires sound training in specialized hemostasis and thrombosis research laboratories as well as a consistent daily routine. In this theme issue of Hämostaseologie—Progress in Haemostasis, four review articles cover key models that have proven instrumental to gain mechanistic insights on thrombogenesis and hemostatic processes. In recent decades, these models have moved our field forward and enabled translation across scales, from cell-based research to isolated flow chamber systems, to mouse thrombosis models reflecting the pathologic situations as observed in patients, to large animal models.
9
Irrera, Pietro, Miriam Roberto, Lorena Consolino, Annasofia Anemone, Daisy Villano, Victor Navarro-Tableros, Antonella Carella, Walter Dastrù, Silvio Aime e Dario Livio Longo. "Effect of Esomeprazole Treatment on Extracellular Tumor pH in a Preclinical Model of Prostate Cancer by MRI-CEST Tumor pH Imaging". Metabolites 13, n. 1 (28 dicembre 2022): 48. http://dx.doi.org/10.3390/metabo13010048.
Abstract (sommario):
Novel anticancer treatments target the pH regulating system that plays a major role in tumor progression by creating an acidic microenvironment, although few studies have addressed their effect on tumor acidosis. In this study, we investigated in vivo several proton pump inhibitors (PPIs) targeting NHE-1 (Amiloride and Cariporide) and V-ATPase (Esomeprazole and Lansoprazole) proton transporters in the DU145 androgen-insensitive human prostate cancer model. In cellulo results showed that DU145 are sensitive, with decreasing efficacy, to Amiloride, Esomeprazole and Lansoprazole, with marked cell toxicity both in normoxia and in hypoxia, with almost any change in pH. In vivo studies were performed upon administration of Esomeprazole to assess both the acute and chronic effects, and Iopamidol-based tumor pH imaging was performed to evaluate tumor acidosis. Although statistically significant tumor pH changes were observed a few hours after Esomeprazole administration in both the acute study and up to one week of treatment in the chronic study, longer treatment resulted in a lack of changes in tumor acidosis, which was associated to similar tumor growth curves between treated and control groups in both the subcutaneous and orthotopic models. Overall, this study highlights MRI-CEST tumor pH imaging as a valid approach to monitoring treatment response to PPIs.
10
Kuhn, Deborah J., Qing Chen, Peter M. Voorhees, John S. Strader, Kevin D. Shenk, Congcong M. Sun, Susan D. Demo, Mark K. Bennett, Fred W. van Leeuwen e Robert Z. Orlowski. "The Novel, Irreversible Proteasome Inhibitor PR-171 Demonstrates Potent Anti-Tumor Activity in Pre-Clinical Models of Multiple Myeloma, and Overcomes Bortezomib Resistance." Blood 108, n. 11 (16 novembre 2006): 3461. http://dx.doi.org/10.1182/blood.v108.11.3461.3461.
Abstract (sommario):
Abstract Introduction: The ubiquitin-proteasome pathway has been validated as a therapeutic target with the approval of the small molecule proteasome inhibitor, bortezomib (VELCADE®), in multiple myeloma and non-Hodgkin lymphoma. However, the overall response rate of patients with multiple myeloma in phase III clinical trials was 43%, underscoring the need for a next generation of inhibitors with the potential for greater efficacy. Methods: PR-171 is a novel, tetrapeptide epoxomicin-related inhibitor that binds the proteasome irreversibly, and our objectives were to evaluate its activity and mechanism of action in pre-clinical models of multiple myeloma. Results: PR-171 potently bound and inhibited the chymotrypsin-like subunit of the proteasome in vitro, in cellulo, and in vivo at low concentrations. At higher concentrations, however, unlike bortezomib, which targeted the chymotrypsin-like and peptidyl-glutamyl peptide hydrolyzing activities in vivo, PR-171 also displayed significant inhibition of the trypsin-like and the peptidyl-glutamyl peptide hydrolyzing activities. PR-171-induced proteasome inhibition was associated with accumulation of polyubiquitinated substrates and pro-apoptotic Bax. Brief pulse PR-171 exposure, which simulates the in vivo pharmacokinetics of bortezomib, led to PR-171-mediated inhibition of cellular proliferation linked to induction of caspase-3-dependent apoptosis through both intrinsic (caspase-9) and extrinsic (caspase-8-dependent) pathways. Pretreatment with caspase-3, -8, and -9 inhibitors rescued the anti-proliferative effect of PR-171. Furthermore, pulse PR-171 treatment activated c-Jun-N-terminal kinase, a key-signaling molecule in proteasome inhibitor-induced apoptosis, and cleavage of poly-ADP-ribose polymerase, while abrogation of c-Jun-N-terminal kinase signaling with a dominant-negative c-Jun inhibited PR-171-induced effects. PR-171 displayed enhanced anti-proliferative activity compared to bortezomib in multiple myeloma cell lines and freshly isolated patient-derived CD138+ plasma cells, associated with enhanced phosphorylation of c-Jun-N-terminal kinase and capase-3, -8, and -9 activation. Lastly, PR-171 was a potent inhibitor of proliferation in a multiple myeloma cell line model resistant to bortezomib and in isolates from two patients, one with primary and the other with acquired bortezomib-resistance. Conclusions: These data indicate that PR-171 has enhanced activity against preclinical models of multiple myeloma, perhaps owing to its irreversible binding and subunit specificity, and provide a rationale for its translation into the clinic.
Tesi sul tema "In cellulo and in vivo models":
1
Fischer, Jared Michael. "Mouse Models of Mutation in vivo". University of Cincinnati / OhioLINK, 2008. http://rave.ohiolink.edu/etdc/view?acc_num=ucin1227214862.
2
Passeri, Elodie. "Use of nanoliposomes rich in omega-3 for the prevention of neurodegenerative diseases : bioavailability in vivo and in cortical neurons". Electronic Thesis or Diss., Université de Lorraine, 2021. https://docnum.univ-lorraine.fr/ulprive/DDOC_T_2021_0337_PASSERI.pdf.
Abstract (sommario):
La maladie d’Alzheimer (MA) est la cause d’affection neurodégénérative la plus fréquente et représente un enjeu majeur de santé publique au niveau mondial. De nombreuses stratégies thérapeutiques sont explorées depuis plusieurs décennies, néanmoins, il n'existe toujours aucun traitement curatif et la priorité reste la prévention. Dans ce travail de thèse, nous nous sommes focalisés sur l’approche préventive basée sur les lipides, en particulier les acides gras polyinsaturés oméga-3 (AGPI n-3). Les AGPI n-3 jouent un rôle important dans le développement, le maintien et le fonctionnement du cerveau et ils ont fait l'objet d'un intérêt particulier dans la prévention des déficits cognitifs liés aux maladies neurodégénératives. L’objectif de ce travail est d’étudier la fonctionnalité et la biodisponibilité des nanoliposomes (NL) riches en AGPI n-3, afin d’évaluer leur potentiel neuroprotecteur pour développer de nouvelles stratégies préventives dans les maladies liées au vieillissement comme la MA. Pour ce faire, une technique d'extraction verte a été utilisée pour préparer des NL, provenant de ressources naturelles à partir de lécithine de saumon riche en AGPI n-3. Cette thèse repose sur trois parties. La première partie est consacrée à une revue bibliographique des nouvelles techniques pour faciliter l’accès des molécules au cerveau. Les NL montrent un rôle prometteur, ils sont capables d’améliorer le transport de molécules à travers la barrière hémato-encéphalique et atteindre les régions cérébrales ciblées. Dans la deuxième partie de ce travail, la biodisponibilité des NL riches en AGPI n-3 a été étudiée dans une culture primaire de cellules neuronales corticales d’embryons de rat et dans un modèle de souris. Cette étude montre pour la première fois la biodisponibilité cérébrale des NL riches en AGPI n-3 dans des modèles in vitro et in vivo. Et enfin, dans la troisième partie de cette thèse, les propriétés physico-chimiques et des mécanismes de transfert des NL ont été étudiés dans une culture primaire de neurones corticaux. Ces résultats apportent de nouvelles informations sur l'interaction entre les NL et les neurones et sont prometteurs quant à l'utilisation des NL riches en AGPI n-3, ouvrant de nouvelles possibilités dans le développement de stratégies thérapeutiques préventives et neuroprotectrices pour les maladies neurodégénératives telles que la MAAlzheimer's disease (AD) is the most common cause of neurodegenerative disease and represents a major public health issue worldwide. Many therapeutic strategies have been explored for several decades, however, there is still no curative treatment and the priority remains prevention. In this thesis work, we focused on the preventive approach based on lipids, in particular omega-3 polyunsaturated fatty acids (n-3 PUFAs). N-3 PUFAs play an important role in the development, maintenance and function of the brain, and they have been the subject of particular interest in the prevention of cognitive deficits associated with neurodegenerative diseases. The objective of this work is to study the functionality and bioavailability of nanoliposomes (NL) rich in n-3 PUFAs, in order to assess their neuroprotective potential to develop new preventive strategies in aging-related diseases such as AD. To do this, a green extraction technique was used to prepare NL, from natural resources from salmon lecithin rich in n-3 PUFA. This thesis is based on three parts. The first part is devoted to a bibliographical review of new techniques to facilitate the access of molecules to the brain. NL shows a promising role, being able to improve the transport of molecules across the blood-brain barrier and reach relevant brain regions. In the second part of this work, the bioavailability of NL rich in n-3 PUFA was studied in a primary culture of cortical neuronal cells from rat embryos and in a mouse model. This study shows for the first time the brain bioavailability of NL rich in n-3 PUFA in in vitro and in vivo models. Finally, in the third part of this thesis, the physicochemical properties and transfer mechanisms of NL were studied in a primary culture of cortical neurons. These results provide new information on the interaction between NL and neurons and are promising with regards to the use of NL rich in n-3 PUFA, opening up new possibilities in the development of preventive and neuroprotective therapeutic strategies for neurodegenerative diseases such as AD
3
Rose, Nicolas. "New ex vivo models to study the mechanical interplay between muscle cells and their microenvironment". Electronic Thesis or Diss., Sorbonne université, 2021. http://www.theses.fr/2021SORUS440.
Abstract (sommario):
Un intérêt croissant est porté aux systèmes ex vivo pour des raisons bioéthiques, légales et économiques. Afin d'étudier la fonction et la régénération musculaire, la recherche médicale nécessite la mise au point de plateformes d’analyse fonctionnelle de myotubes matures et d’outils de culture de cellules souches musculaires (MuSCs) non conventionnelle intégrant les contraintes mécaniques. Dans le cadre de mes travaux, une puce de culture de myotubes en 3D avec capacité de mesure de la contraction a été développée. La combinaison d'une technologie de photo-patterning et de micro-substrats a permis d'obtenir des rendements de culture élevés dans des conditions physico-chimiques contrôlées. Des contractions de myotubes 3D dérivés de myoblastes humains primaires ont été observées, la force générée a été mesurée et le schéma de contraction caractérisé. L'impact de la culture 3D sur la morphologie nucléaire a été analysé, confirmant la similitude d'organisation entre les myotubes 3D obtenus et les myofibres in vivo. De plus, le système a permis de modéliser la dystrophie musculaire congénitale due à une mutation de LMNA avec la culture de myotubes 3D mutants présentant un phénotype de laminopathie typique. Enfin, des MuSCs ont été cultivées sur des hydrogels, démontrant l’effet des variations d'élasticité de la niche sur l'activation des MuSCs au cours de la régénération musculaireEx vivo systems are under increasing interest for bioethical, legal and financial stakes. To study muscle function and regeneration, medical research requires functional analysis platforms for mature myotubes and unconventional muscle stem cells (MuSCs) culture tools integrating mechanical consraints. During my doctoral works, a 3D myotube culture chip with contraction monitoring capacity has been developed. Combining photopatterning technology and microsubstrate design allowed to obtain high culture yield in controlled physical and chemical microenvironment. Spontaneous contractions of human primary myoblast-derived 3D myotubes has been observed, their generated forces measured and their contraction pattern characterised. The impact of 3D culture on nuclear morphology has been analysed, confirming organizational similarities betaween obtained 3 myotubes and in vivo myofibers. Moreover, LMNA-related Congenital Muscular Dystrophy has been modelled in 3D mutants myotubes displaying typical laminopathy phenotype. Finally, MuSCs has ben cultured on hydrogels, demonstrating the effect of niche elasticity variations on MuSCs activation during muscle regeneration
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BAZZINI, CHIARA. "STUDY OF MOLECULAR MECHANISMS AND NEW STRATEGIES AGAINST A CYTOTOXICITY AND NEUROINFLAMMATION IN EX VIVO CELLULAR MODELS FROM ALZHEIMER’S DISEASE PATIENTS". Doctoral thesis, Università degli Studi di Milano-Bicocca, 2021. http://hdl.handle.net/10281/306480.
Abstract (sommario):
La malattia di Alzheimer (AD) rappresenta una delle principali problematiche per la salute pubblica ed è stata identificata come una priorità per la ricerca. Le due caratteristiche patologiche fondamentali della malattia sono le placche amiloidi e i grovigli neuro fibrillari che sono alla base della neuroinfiammazione e del deterioramento cognitivo.Le forme solubili degli oligomeri sono la specie più tossica della β-amiloide (Aβ) e interagiscono con diverse chinasi proteiche coinvolte nella trasduzione del segnale intracellulare come Ras/MAPK e PI3K/AKT che regolano molti processi cellulari e funzioni cognitive, e alcuni meccanismi molecolari coinvolti nella degenerazione neuronale, come l'iperfosforilazione di tau e l'eccitotossicità del glutammato. Negli ultimi anni molta attenzione è stata focalizzata sull'utilizzo di composti naturali come agenti neuroprotettivi. Il luppolo (Humulus Lupulus) contiene flavonoidi, molecole aromatiche che hanno proprietà antiossidanti e antinfiammatorie. È stato dimostrato che l'estratto di luppolo ha effetti antiaggreganti sull’Aβ e sembra impedire la sua produzione nelle cellule in coltura. L'accumulo di Aβ induce anche l'attivazione della proteina 3 del recettore Nod-like receptor 3 (NLRP3) dell’inflammosoma e il conseguente rilascio di citochine proinfiammatorie, il quale svolge un ruolo fondamentale nella neuroinfiammazione associata all'AD. NLRP3 attivato induce la produzione e il rilascio di mediatori infiammatori, tra cui i complessi proteici ASC (ASC specks), IL-1β e IL-18, che facilitano la deposizione di Aβ in un ciclo che si auto alimenta. Impedire l’assemblaggio e l'attivazione del complesso dell’inflammosoma potrebbe essere una possibile strategia per la terapia dell'AD.
L'obiettivo generale di questo studio è quello di indagare i meccanismi molecolari coinvolti nelle malattie neurodegenerative e nella neuroinfiammazione utilizzando modelli cellulari periferici ex vivo di AD.Al fine di caratterizzare le interazioni Aβ e vie di trasduzione del segnale MAPK e AKT, abbiamo utilizzato fibroblasti di pazienti AD sporadici con diversa gravità della malattia. Per valutare i meccanismi molecolari che potrebbero prevenire o modulare la tossicità indotta da Aβ, sono stati studiati anche i potenziali effetti citoprotettivi dell'estratto di luppolo e il relativo signaling intracellulare. Inoltre, è stato dato particolare interesse alla via di attivazione del NLRP3-infiammasoma. Abbiamo studiato il coinvolgimento dell'attivazione di NLRP3 sulle vie MAPK e AKT e sui loro bersagli a valle, utilizzando una combinazione di studi in vitro e di campioni ottenuti dai pazienti. In particolare, abbiamo utilizzato monociti umani THP-1 di derivazione macrofagica e monociti derivati da cellule mononucleate del sangue periferico (PBMC) di soggetti sani (HC) e pazienti affetti da AD, per analizzare la modulazione autofagica e gli effetti della Stavudina (D4T), un inibitore nucleosidico della trascrittasi inversa, che riduce l'attivazione dell'inflammosoma bloccando il recettore purinergico P2X7R. Inoltre, abbiamo analizzato il pathway di attivazione dell'inflammosoma NLRP3 e il ruolo di CRID3 un inibitore selettivo, per confrontare gli effetti dell’inibizione dell’inflammosoma attraverso due pathway differenti. I monociti derivati da HC e AD sono stati differenziati in cellule microglia-like (MDMIs) e caratterizzati per l'espressione di proteine intracellulari e di superficie tipiche delle cellule mieloidi. Funzioni tipiche della microglia come il rilascio di citochine infiammatorie, la fagocitosi e la degradazione sono state valutate anche in seguito all'esposizione di attivatori dell'inflammosoma con o senza CRID3. MDMIs riflettono molte caratteristiche della microglia e sono un modello cellulare utile per comprendere la patogenesi dell'AD, identificare i target terapeutici e consentire lo screening farmacologico su larga scala dei nuovi composti per uso terapeutico.Alzheimer's disease (AD) is a major public health concern and has been identified as a priority for research in Life Science. The two core pathological hallmarks of AD are extracellular amyloid plaques and intracellular neurofibrillary tangles which underlie microglial and neuronal damage, neuroinflammation and cognitive impairment. Soluble oligomers are the most toxic species of β-amyloid (Aβ) and interact with several protein kinases such as Ras/MAPK and PI3K/AKT pathways, which regulate many cellular processes and cognitive functions. These pathways mediate Aβ toxicity, regulating some molecular mechanisms involved in neuronal degeneration such as cytoskeletal impairment, glutamate excitotoxicity and neuroinflammation. In the last years much attention has been focused on the potential role of natural compounds as neuroprotective agents. Hop (Humulus Lupulus) contains flavonoids, aromatic molecules which have antioxidant, anti-inflammatory and anti-atherogenic properties. In fact, hop extract has anti-aggregating effects on Aβ, and it seems to prevent its production in cultured cells. Aβ induces also the activation of the pattern recognition receptor Nod-like receptor protein 3 (NLRP3) inflammasome complex in microglia and the consequent release of proinflammatory cytokines, playing a pivotal role in AD-associated neuroinflammation. NLRP3 activation results in the release of inflammatory mediators, including ASC protein complexes (ASC specks), IL-1β and IL-18, that facilitate Aβ deposition and neuroinflammation in a self-feeding pathogenic loop. Since specific therapeutical strategies are still lacking, the dampening of the inflammasome assembly and activation could be a new strategy for AD. The overall focus of this study is to investigate molecular mechanisms involved in neurodegenerative diseases and in neuroinflammation, using peripheral ex vivo cellular models from AD, to check new potential therapeutical targets. In order to characterize the complex interactions among Aβ, MAPK and AKT signaling, we used fibroblasts from sporadic AD patients with different disease severity. To evaluate any molecular mechanisms that could prevent or modulate Aβ-induced toxicity, the potential cytoprotective effects of Hop extract and related intracellular signaling were also investigated. Fibroblasts provide a useful cellular model for studying AD, since they could be differentiated into patient-specific neural cell lines, using iPSC technologies. Moreover, particular interest was given to NLRP3-inflammasome activation pathway. We investigated the involvement of NLRP3 inflammasome activation on intracellular pathways and their downstream targets, using a combination of in vitro studies and patient-derived samples. In particular, we used macrophage-derived THP-1 human monocytes and peripheral blood mononuclear cells (PBMC)-derived monocytes from healthy control (HC) subjects and AD patients, to analyse phagocytosis, autophagy and apoptosis modulation and the effects of the nucleoside reverse transcriptase inhibitor Stavudine (D4T), that reduces NLRP3 inflammasome activation blocking the purinergic receptor P2X7R. Furthermore, we analyzed the NLRP3 inflammasome pathway and the role of the selective NLRP3 inhibitor CRID3, to compare the effects of inflammasome inhibition through two different mechanisms. At this purpose, HC and AD-derived monocytes were differentiated into microglia-like cells (MDMIs) and characterized for myeloid surface and intracellular proteins expression. Key microglia functions such as inflammatory cytokines release, Aβ phagocytosis and degradation were evaluated upon exposure to NLRP3 inflammasome activators with or without CRID3. MDMIs reflected many features of microglia and, as fibroblasts-derived iPSCs, they are attractive cellular models helpful to understand AD pathogenesis, identify therapeutic targets and allow large-scale drug screening of the novel therapeutic candidates.
5
Zangrossi, Manuela. "Study of the extra-telomeric functions of telomerase in in vitro and in vivo models". Doctoral thesis, Università degli studi di Padova, 2018. http://hdl.handle.net/11577/3426233.
Abstract (sommario):
Maintenance of telomere length, required for the unlimited cell proliferation displayed by cancer cells, is provided by telomerase, a ribonucleoprotein complex containing a specialized reverse transcriptase, encoded by TERT gene, that uses an internal RNA template to maintain telomeres length, thus playing a critical role in tumor formation and progression. TERT is usually repressed in normal somatic cells, but is detectable in the vast majority of tumors.
Recent studies have suggested that TERT, besides maintaining telomere, is involved in other cellular functions, and it may contribute to carcinogenesis also via telomere length-independent mechanisms; therefore its inhibition could represent a promising strategy to improve cancer treatment, regardless of telomere length. The possible therapeutic effects of BIBR1532 (BIBR), a specific TERT inhibitor, have been evaluated in different cellular backgrounds, but no data are currently available regarding Epstein-Barr virus (EBV)-driven and virus-unrelated B-cell malignancies.
The aim of this study was to characterize the biological effects of short-term TERT inhibition by BIBR on EBV-immortalized lymphoblastoid cell lines (LCLs) and fully transformed Burkitt’s lymphoma (BL) cell lines; in addition, we investigated the effects of short-term BIBR treatment in vivo in wild type zebrafish embryos.
We found that short-term inhibition of TERT by BIBR, in in vitro models of B-cell malignancies, led to decreased cell proliferation, accumulation of cells in the S-phase and ultimately increased apoptosis. The cell cycle arrest and apoptosis, consequent upon short-term TERT inhibition, were associated with and likely dependent on the activation of the DNA damage response (DDR), highlighted by the increased levels of γH2AX and activation of ATM and ATR pathways. Analyses of the mean and range of telomere lengths and telomere dysfunction-induced foci indicated that DDR after short-term TERT inhibition was not related to telomere dysfunction, thus suggesting that TERT, besides stabilizing telomere, may protect DNA via telomere-independent mechanisms. Notably, TERT-positive LCLs treated with BIBR in combination with fludarabine or cyclophosphamide showed a significant increase in the number of apoptotic cells with respect to those treated with chemotherapeutic agents alone.
In agreement with in vitro results, short-term inhibition of Tert by BIBR in wild type zebrafish embryos reduced cell proliferation, induced an accumulation of cells in S-phase, increased apoptosis, and triggered the activation of DDR. These effects were telomere length-unrelated, since the range of telomere length was not affected by the short-term BIBR treatment and the DNA damage foci were distributed randomly, rather than specifically located at telomeres. All these effects were specifically related to Tert inhibition since BIBR treatment showed no effect in Tert-negative zebrafish embryos.
Taken together these data demonstrate that TERT inhibition impairs cell proliferation and induces pro-apoptotic effects unrelated to telomere dysfunction, enforcing the concept that TERT per se exerts telomere length-independent tumor-promoting effects, and thus supporting the introduction of TERT inhibitors to complement current anticancer treatment modalities.Il mantenimento dei telomeri, necessario per la proliferazione illimitata delle cellule tumorali, è esercitato dalla telomerasi, un complesso ribonucleoproteico contenente una trascrittasi inversa specializzata, codificata dal gene TERT, che utilizza un templato ad RNA per sintetizzare nuove sequenze telomeriche, svolgendo quindi un ruolo critico nella formazione e nella progressione dei tumori. TERT viene infatti solitamente represso in normali cellule somatiche, mentre è rilevabile nella maggior parte dei tumori. Studi recenti hanno suggerito che TERT è coinvolto in altre funzioni cellulari e può contribuire alla carcinogenesi anche attraverso meccanismi indipendenti dal mantenimento dei telomeri, quindi la sua inibizione potrebbe rappresentare una strategia promettente per migliorare il trattamento antitumorale, al di là dell’effetto sui telomeri. I possibili effetti terapeutici di BIBR1532 (BIBR), un inibitore specifico del TERT, sono stati valutati in diversi contesti cellulari, ma non sono attualmente disponibili dati ottenuti su modelli di neoplasie delle cellule B sia associate al virus di Epstein-Barr (EBV) che virus-indipendenti. Lo scopo di questo studio era di caratterizzare gli effetti biologici dell'inibizione di TERT a breve termine da parte del BIBR su linee cellulari linfoblastoidi immortalizzate da EBV (LCL) e su modelli in vitro di linfoma di Burkitt (BL); inoltre, sono stati studiati gli effetti del trattamento con BIBR a breve termine in vivo negli embrioni di zebrafish. I risultati ottenuti hanno dimostrato che l'inibizione a breve termine di TERT da parte di BIBR, in modelli in vitro di tumori delle cellule B, ha portato a una diminuzione della proliferazione cellulare, all'accumulo di cellule nella fase S e infine all'aumento dell'apoptosi. L'arresto del ciclo cellulare e l'apoptosi, conseguenti all'inibizione di TERT a breve termine, erano associati e probabilmente dipendenti dall'attivazione della risposta al danno del DNA, come evidenziato dall’aumento dei livelli di γH2AX e dall'attivazione dei pathway di ATM e ATR. L’analisi della media e del range di lunghezza dei telomeri e dei foci di danno al DNA ha indicato che la risposta al danno attivata in seguito all’inibizione TERT a breve termine non era legata a disfunzioni telomeriche, suggerendo quindi che TERT, oltre a stabilizzare il telomero, può proteggere il DNA tramite meccanismi telomero-indipendenti. In particolare, LCL-TERT positive trattate con BIBR in combinazione con fludarabina o ciclofosfamide hanno mostrato un aumento significativo del numero di cellule apoptotiche rispetto a quelle trattate con agenti chemioterapici da soli. In accordo con i risultati in vitro, l'inibizione a breve termine di Tert da parte del BIBR in embrioni di zebrafish ha ridotto la proliferazione cellulare, indotto un accumulo di cellule nella fase S, aumentato il tasso di apoptosi e innescato l'attivazione della risposta al danno al DNA. Questi effetti non erano legati a disfunzioni telomeriche, poiché il range di lunghezza dei telomeri non era influenzato dal trattamento a breve termine con BIBR e i foci di danno al DNA erano distribuiti casualmente, piuttosto che localizzati in modo specifico sui telomeri. Tutti questi effetti erano specificamente associati all'inibizione di Tert poiché il trattamento con BIBR non mostrava alcun effetto negli embrioni di zebrafish Tert-negativi. Nel complesso questi dati dimostrano che l'inibizione del TERT compromette la proliferazione cellulare e induce effetti pro-apoptotici non associati a disfunzioni telomeriche, rafforzando il concetto che TERT esercita di per sé funzioni pro-tumorali indipendenti dalla lunghezza del telomero e quindi supportando l'introduzione di inibitori di TERT per integrare le attuali modalità di trattamento antitumorale.
6
Ronayne, Rachel E. "Human Ependymin-1 Neurotrophic Factor Mimetics Reduce Tau Phosphorylation and Cellular Apoptosis in Vitro and in Vivo in Alzheimer’s Disease Models". Digital WPI, 2008. https://digitalcommons.wpi.edu/etd-theses/1018.
Abstract (sommario):
"Alzheimer’s disease (AD) is the most widespread neurodegenerative disorder, affecting approximately 20 million people worldwide. AD pathology is primarily characterized by the formation of extracellular amyloid plaques resulting from the aggregation of insoluble amyloid-beta 1-42 (A-beta), and neurofibrillary tangles (NFT’s) resulting from intracellular aggregation of hyperphosphorylated tau protein. The current FDA-approved AD treatments do not stop or reverse neurodegeneration, but only treat the symptoms by increasing acetylcholine neurotransmitter. Our laboratory is attempting to provide an additional therapeutic approach by using neurotrophic factors to block apoptosis or to restore neurons. We previously demonstrated that, in an in vitro model for AD, hEPN-1 neurotrophic factor mimetics can block synthetic A-beta-induced neuronal cell death when added to cultures, presumably by blocking caspase activation. In this thesis, we extended these findings to study the effect of A-beta and hEPN-1 on tau hyperphosphorylation (as measured by immunoblots with phospho-specific antibodies) and nuclear DNA fragmentation (as measured by TUNEL staining), both in vitro and in vivo in AD transgenic mice. We found that A-beta induces the hyperphosphorylation of tau in both mouse N2a and human SHSY neuronal cells, and that hEPN-1 may lower this phosphorylation in N2a cells. Furthermore, we discovered that hEPN-1 can reduce nuclear DNA fragmentation when added both simultaneously to A-beta and 3 and 6 hours post A-beta addition. Finally, in vivo hEPN-1 may lower both tau hyperphosphorylation and caspase-7 related protein (C7RP) in AD transgenic (Tg) mice. The overall results validate our in vitro AD model, show the efficacy of hEPN-1 at blocking A-beta-induced DNA fragmentation even when added post-insult, and show that hEPN-1 may work in an AD mouse model. However, more studies must be conducted to confirm these findings. "
7
Lu, Yen-Zhen. "The effects of 670nm light on retinal Müller cell gliosis following retinal stress or injury: exploring the underlying cellular mechanisms using in vivo and in vitro models". Phd thesis, Canberra, ACT : The Australian National University, 2018. http://hdl.handle.net/1885/154729.
Abstract (sommario):
Photobiomodulation (PBM) describes a process whereby light
wavelengths of 600-
1000nm are used to initiate biological responses. PBM has been
shown to attenuate
inflammation and accelerate wound healing in skin and mucosal
tissues. In the nervous
system, it promotes recovery of injured spinal cord and optic
nerve. Our laboratory has
found that irradiation with 670nm light, applied prior to retinal
insult, reduced
photoreceptor death in retinal degeneration in vivo. However,
very little attention was
paid to the non-neuronal component of the retina, the macroglia
of the retina.
Müller cells (MCs), the principal macroglia of the retina, are
involved in supporting
retinal structure and maintaining its homeostasis. MCs react to
retinal stress or injuries,
described as gliosis, aiming to protect neurons. However when it
enters into a
progressive state, it becomes detrimental to the retina.
Activated MCs, if uncontrolled,
release a large amount of proinflammatory cytokines and
chemokines, recruiting
microglias (MGs) and monocytes, which lead to further retinal
inflammation. While
the retinal damage is extensive, MCs undergo mitosis and
thickening of their processes,
which reach the subretinal space to form glial scars that inhibit
nutrient delivery,
leading to further neuronal death. Thus, the aim of this thesis
is to investigate the effects
of 670nm irradiation on activated MCs using in vivo and in vitro
stress models,
exploring a new avenue that may prevent irreversible retinal
degeneration.
In Chapter 3, the effects of 670nm light on activated MCs using
in vitro and in vivo
stress models of retinal injury were investigated. Our results
demonstrated that 670nm
modified MC activation, both its proinflammatory and
proliferative processes. This
chapter additionally draws attention to the importance of
appropriate timing of
treatment, as there is a finite therapeutic window to effectively
mitigate gliotic changes.
In Chapter 4 investigated the effects of 670nm light on
interaction between MCs and
photoreceptors, and on subsequent MC-derived MG activation in
vitro. Results
confirmed that 670nm light mitigated MC gliosis induced by
photo-oxidative damage
(PD), subsequently reducing MG activation. This protective
mechanism of action of
670nm light in MCs was associated with increased mitochondrial
activity. Chapter 5
explored the cell communication between MCs and MGs in vitro,
focusing on the role
of exosomes. Exosomes have been discovered to carry microRNAs
(miRNAs), which
allows the transfer of genetic information between cells. In this
chapter, IL-1β
stimulation of MCs led to the release of exosomes, which
stimulated MGs to upregulate
expression of proinflammatory cytokines. Several miRNAs
implicated in regulating
inflammatory processes were identified in MC-derived exosomes in
stressed MCs.
Treatment with 670nm significantly reduced the expression of some
of the proinflammatory
genes.
The results from this thesis collectively indicate that following
MC activation in retinal
damage, 670nm treatment post-damage can mitigate MC gliosis and
subsequently
ameliorate retinal inflammation. Furthermore, this effect may be
achieved through
down-regulation of proinflammatory cytokine production in the
retina and modification
of exosome contents. Therefore, targeting activated MCs using
670nm light may be a
potential therapeutic strategy in mitigating inflammation
associated with the
irreversible retinal degeneration.
8
Toutain, Hervé. "Développement et caractérisation de modèles expérimentaux pour l'étude ex-vivo et in-vitro de la cellule tubulaire proximale de rein de lapin". Rouen, 1989. http://www.theses.fr/1989ROUES036.
Abstract (sommario):
Une suspension de cellules tubulaires proximales de rein de lapin est isolée par une nouvelle méthode qui ne fait pas intervenir d'enzymes protéolytiques. Ces cellules, après une caractérisation biochimique, morphologique, et métabolique, sont séparées en deux populations hautement purifiées par une technique d'électrophorèse ou flux libre en veine liquide. Présentation d'un modèle de culture primaire de cellules tubulaires proximales de rein de lapin
9
Sibeko, Sengeziwe. "The impact of macrophage inflammatory protein-3 alpha and other innate immune markers on susceptibility/resistance to HIV infection in the female genital tract mucosa using cellular and ex vivo tissue models". Thesis, University of Oxford, 2016. https://ora.ox.ac.uk/objects/uuid:a7ecf529-b94e-492f-8374-675cb495ef05.
Abstract (sommario):
The distinctive feature of the Human Immunodeficiency Virus (HIV) epidemic in the 21st century is the burden it places on women. Scientists believe that the best opportunities for successful interventions to prevent sexual HIV transmission lie in the initial stages of infection at the portal of entry, the genital tract (GT), which offers the greatest host advantages and viral vulnerabilities. However, understanding of the correlates of protection/vulnerability and innate immunity at the portal of entry is poor. First and foremost, there is no agreement about which GT sub-compartment is the primary site of HIV/SIV infection. Second, the epithelium, previously studied solely for its function as a barrier, has hardly been investigated for its role in innate immunity in the context of SIV/HIV infection. MIP-3α, a chemokine secreted by epithelial cells, was previously proposed to have a role in amplifying the early Simian Immunodeficiency Virus (SIV) infection events in the GT of female macaques. Specifically, MIP-3α was shown to be secreted by epithelial cells of the endocervix, accumulating subepithelially within the first 24 hours post exposure, following deposition of an intravaginal inoculum of SIV. Similar studies in humans have not been reported. We hence undertook to study MIP-3α for its role in early HIV infection events in the endocervix of humans. In order to achieve this, we first characterised MIP-3α constitutive secretion patterns in different sub-compartments of the GT before proceeding to determine its induced secretion patterns, stimulating with HIV-1 and various Toll-like receptor ligands. For completeness we determined constitutive and induced secretion patterns of multiple soluble proteins (SPs) and antimicrobial peptides (AMPs) in the endocervices of humans and macaques. The GT being an immunohormonal system, we further studied the influence of endogenous hormonal changes on the stability of MIP-3α and that of other innate immune markers. We quantified MIP-3α with a sandwich Elisa, and SPs and AMPs with the Luminex multiplex bead assay. Our results showed that the GT is a rich source of MIP-3α with its levels being among those of the highest SPs in the GT. Constitutive levels were highest in the endocervical sub-compartment of all the sub-compartments studied. Further, the GT is an inflammatory environment, which would explain the high levels of MIP-3α. The primary driver of MIP-3α levels appears to be inflammation rather than hormonal levels. MIP-3α levels are significantly higher in the GT of humans than in macaques. There was no evidence that MIP-3α levels are elevated on exposure to HIV and SIV in humans and macaques, respectively. We therefore concluded that since the endocervix is unlikely to respond to HIV/SIV by secreting MIP-3α in vivo, contrary to the previous reports, MIP-3α is hence not a key player in amplifying early events in infection. And as such, it should not be a prime target for preventive therapy. Further, the human GT having a pre-existing inflammatory profile may explain the high rates of HIV sexual transmission. Lastly, we concluded that the infection mechanisms described in the macaque model (i.e. the 'outside-in' signaling) are likely not required for human infection.
10
Body, Simon. "Physiopathologie du lymphome à cellules du manteau : de la mécanistique aux modèles précliniques". Thesis, Normandie, 2017. http://www.theses.fr/2017NORMC419/document.
Abstract (sommario):
Le lymphome à cellules du manteau (LCM) est une hémopathie maligne B mature, appartenant à la famille des lymphomes non hodgkiniens. Le LCM est caractérisé par la translocation t(11;14)(q13;q32) qui provoque une expression aberrante de cycline D1. C’est une pathologie rare mais à haut risque de rechute, et qui reste le plus souvent incurable suite à l’apparition de clones chimiorésistants. L’acquisition de résistance est intimement liée aux interactions entre les cellules tumorales et leur microenvironnement. Afin de mimer de la manière la plus pertinente possible ces interactions, nous avons mis en place un modèle murin de xénogreffe en utilisant les lignées cellulaires de LCM JeKo1, REC1, Z138 et Granta-519 que nous avons modifiées afin qu’elles expriment un fluorophore (GFP ou m-cherry) et/ou le gène codant pour la luciférase. Après injection aux souris du substrat de la luciférase, la luciférine, nous sommes en mesure de suivre au cours du temps la progression tumorale. Nous pouvons également évaluer le degré d’infiltration tumorale dans la moelle osseuse, la rate, le cerveau et le sang après euthanasie des animaux, par des techniques de cytométrie en flux et d’immunocytochimie. Ce modèle nous a permis de montrer l’intérêt thérapeutique d’un inhibiteur de l’exportine 1 (XPO1) : le KPT 330 (ou selinexor) qui est capable de contenir cycline D1 uniquement au niveau nucléaire. Nous avons montré que la localisation subcellulaire de cycline D1, est retrouvée majoritairement cytoplasmique dans certaines lignées cellulaires de LCM (2/7) et chez un certain nombre de patients (6/42, 14%), et est associée à un fort potentiel d’invasion, de migration et à un phénotype agressif. Par ailleurs, grâce à ce modèle, nous avons pu objectiver le manque d’efficacité in vivo d’agonistes aux récepteurs aux œstrogènes de type β (ER β). Ces récepteurs, présents sur les lymphocytes B étaient supposés inhiber la prolifération cellulaire et provoquer la mort des cellules par apoptose. L’utilisation de deux agonistes des ER β, le diarylpropionitrile (DPN) et l’ERB-041 a montré une absence d’effet de ces molécules, lorsque les cellules tumorales sont au contact de leur microenvironnement. D’autre part, afin de mieux comprendre les mécanismes de résistance aux chimiothérapies, nous avons étudié la résistance de la lignée cellulaire REC-1 traitée par des agents génotoxiques. Nous avons montré que cette lignée présentait une anomalie de dégradation de cycline D1 associée à une activité diminuée du protéasome 26S. Enfin, nous avons montré dans des travaux préliminaires que la protéine fused in sarcoma (FUS) pourrait, lorsqu’elle est associée à cycline D1, être capable de réguler les voies de réparation des dommages à l’ADN. Des anomalies de ces voies induisent une grande instabilité génétique responsable de l’échappement des tumeurs aux traitements, le ciblage de FUS pourrait par conséquent présenter un intérêt thérapeutique.Pris dans leur ensemble, ces résultats permettent de renforcer ou d’infirmer l’intérêt de certaines cibles thérapeutiques dans l’espoir de pouvoir continuer à améliorer la prise en charge des patients. Ils fournissent également un outil pour l’évaluation de nouvelles molécules dans un modèle murin prenant en compte les interactions entre la cellule tumorale et son microenvironnementMantle cell lymphoma (MCL) is a mature malignant hemopathy, belonging to the non-Hodgkin's lymphoma family. The MCL is characterized by the translocation t(11;14)(q13;q32) which causes an aberrant expression of cyclin D1. It is a rare disease but at high risk of relapse, and it is most often incurable due to the appearance of chemoresistant clones. The acquisition of resistance is intimately linked to the interactions between the tumor cells and their microenvironment. In order to mimic, in the most relevant way, these interactions, we have implemented a mouse xenograft model using the MCL cell lines JeKo1, REC1, Z138 and Granta-519 which we have modified so that they express a fluorophore (GFP or m-cherry) and / or the gene encoding the luciferase. After injection to the mice of the luciferase substrate, luciferin, we are able to follow over time the tumor progression. We can also assess the degree of tumor infiltration in bone marrow, spleen, brain and blood after euthanasia of animals, by flow cytometry and immunocytochemistry. This model allowed us to show the therapeutic interest of an inhibitor of exportin 1 (XPO1): the KPT 330 (or selinexor) which is able to contain cyclin D1 only on the nuclear level. We have shown that the subcellular localization of cyclin D1 is mainly cytoplasmic in some LCM (2/7) cell lines and in a number of patients (6/42, 14%), and is associated with a high potential Invasion, migration and an aggressive phenotype. Moreover, thanks to this model, we have been able to objectify the in vivo lack of efficacy of agonists to β-type estrogen receptors (ER β). These receptors, present on B lymphocytes, were thought to inhibit cell proliferation and cause cell death by apoptosis. The use of two ER β agonists, diarylpropionitrile (DPN) and ERB-041 showed an absence of effect of these molecules, when the tumor cells are in contact with their microenvironment. On the other hand, in order to better understand the mechanisms of resistance to chemotherapies, we studied the resistance of the REC-1 cell line treated with genotoxic agents. We have shown that this line has an abnormality of cyclin D1 degradation associated with decreased activity of the 26S proteasome. Finally, we have shown in preliminary work that the fused in sarcoma protein (FUS) could, when associated with cyclin D1, be able to regulate the repair pathways of DNA damage. Abnormalities of these pathways induce a great genetic instability responsible for the escape of tumors to treatments, the targeting of FUS could therefore be of therapeutic interest.Taken as a whole, these results reinforce or invalidate the interest of certain therapeutic targets in the hope of continuing to improve the management of patients. They also provide a tool for evaluating new molecules in a murine model that takes into account the interactions between the tumor cell and its microenvironment
Libri sul tema "In cellulo and in vivo models":
1
Ferrick, David A. Transgenic mice as a in vivo model for self reactivity. Austin: R.G. Landes Co., 1994.
2
Morgan, Douglas W., e Lisa A. Marshall, a cura di. In Vivo Models of Inflammation. Basel: Birkhäuser Basel, 1999. http://dx.doi.org/10.1007/978-3-0348-7775-6.
3
Stevenson, Christopher S., Lisa A. Marshall e Douglas W. Morgan, a cura di. In Vivo Models of Inflammation. Basel: Birkhäuser Basel, 2006. http://dx.doi.org/10.1007/978-3-7643-7520-1.
4
Stevenson, Christopher S., Lisa A. Marshall e Douglas W. Morgan, a cura di. In Vivo Models of Inflammation. Basel: Birkhäuser Basel, 2006. http://dx.doi.org/10.1007/978-3-7643-7760-1.
5
1974-, Stevenson Christopher S., Morgan Douglas W. 1951- e Marshall Lisa A. 1954-, a cura di. In vivo models of inflammation. 2a ed. Basel: Birkhäuser, 2006.
6
1951-, Morgan Douglas W., e Marshall Lisa A. 1954-, a cura di. In vivo models of inflammation. Basel: Birkhauser Verlag, 1999.
7
Ntziachristos, Vasilis. Textbook of in vivo imaging in vertebrates. Chichester, West Sussex: J. Wiley, 2007.
8
Friedman, Herman, Steven Specter e Mauro Bendinelli, a cura di. In vivo Models of HIV Disease and Control. Boston, MA: Springer US, 2006. http://dx.doi.org/10.1007/b135975.
9
J, Ellis Kenneth, Eastman Jerry D e International Symposium on In Vivo Body Composition Studies (1992 : Houston, Tex.), a cura di. Human body composition: In vivo methods, models, and assessment. New York: Plenum Press, 1993.
10
C, Sahu Saura, a cura di. Hepatotoxicity: From genomics to in vitro and in vivo models. Chichester, England: John Wiley & Sons, 2007.
Capitoli di libri sul tema "In cellulo and in vivo models":
1
Andes, David R. "In Vivo Candida Device Biofilm Models". In Candida albicans: Cellular and Molecular Biology, 93–113. Cham: Springer International Publishing, 2017. http://dx.doi.org/10.1007/978-3-319-50409-4_7.
2
Cwykiel, Joanna, e Maria Z. Siemionow. "Cellular Therapy Models: Ex Vivo Chimera Model by Cell Fusion". In Plastic and Reconstructive Surgery, 593–603. London: Springer London, 2014. http://dx.doi.org/10.1007/978-1-4471-6335-0_72.
3
Flaherty, Lynne, John M. Harlan e Robert K. Winn. "Blockade of Leukocyte Adhesion in in Vivo Models of Inflammation". In Cellular Adhesion, 153–66. Boston, MA: Springer US, 1994. http://dx.doi.org/10.1007/978-1-4615-2466-3_9.
4
Chernokal, Brea, Cailin R. Gonyea e Jason P. Gleghorn. "Lung Development in a Dish: Models to Interrogate the Cellular Niche and the Role of Mechanical Forces in Development". In Advances in Experimental Medicine and Biology, 29–48. Cham: Springer International Publishing, 2023. http://dx.doi.org/10.1007/978-3-031-26625-6_3.
Abstract (sommario):
AbstractOver the past decade, emphasis has been placed on recapitulating in vitro the architecture and multicellular interactions found in organs in vivo [1, 2]. Whereas traditional reductionist approaches to in vitro models enable teasing apart the precise signaling pathways, cellular interactions, and response to biochemical and biophysical cues, model systems that incorporate higher complexity are needed to ask questions about physiology and morphogenesis at the tissue scale. Significant advancements have been made in establishing in vitro models of lung development to understand cell-fate specification, gene regulatory networks, sexual dimorphism, three-dimensional organization, and how mechanical forces interact to drive lung organogenesis [3–5]. In this chapter, we highlight recent advances in the rapid development of various lung organoids, organ-on-a-chip models, and whole lung ex vivo explant models currently used to dissect the roles of these cellular signals and mechanical cues in lung development and potential avenues for future investigation (Fig. 3.1).
5
Welsch, Clifford W. "Rodent Models To ExamineIn Vivo Hormonal Regulation of Mammary Gland Tumorigenesis". In Cellular and Molecular Biology of Mammary Cancer, 163–79. Boston, MA: Springer US, 1987. http://dx.doi.org/10.1007/978-1-4613-0943-7_10.
6
Mazière, J. C., S. Salmon, C. Candide, C. Mazière, R. Santus, J. P. Reyftmann, P. Morlière e L. Dubertret. "Lipid Peroxidation and Cellular Functions: in Vitro Models and Relation to in Vivo Observations". In Free Radicals, Lipoproteins, and Membrane Lipids, 327–42. New York, NY: Springer US, 1990. http://dx.doi.org/10.1007/978-1-4684-7427-5_31.
7
Wiegand, C., J. Tittelbach, U. C. Hipler e P. Elsner. "Water-Filtered Infrared A Irradiation: From Observations in Clinical Studies to Complex In Vitro Models". In Water-filtered Infrared A (wIRA) Irradiation, 203–12. Cham: Springer International Publishing, 2022. http://dx.doi.org/10.1007/978-3-030-92880-3_17.
Abstract (sommario):
AbstractSuccessful treatment of recalcitrant common hand and foot warts in a prospective randomized controlled blind trial using wIRA and PDT has been reported. In addition, in wound healing wIRA is mostly investigated in vitro based on the resolution of mechanical damage to confluent cell layers using the “scratch wound assay.” The latter enables the direct measurement of cell migration and regeneration of the cell layer. Preliminary studies for wIRA effects on wound closure in vitro have shown beneficial effects of single 10 min treatments. Although cellular processes induced and mediators involved still need to be elucidated, it is apparent that the observed clinical benefits of wIRA on wound healing can be investigated in vitro using adequate models and experimental settings. The next step is to employ 3D skin models for morphological investigations closely simulating in vivo conditions.
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Schaefer, H., e W. Schalla. "Human Cutaneous Pharmacokinetics In Vivo". In Skin Models, 94–102. Berlin, Heidelberg: Springer Berlin Heidelberg, 1986. http://dx.doi.org/10.1007/978-3-642-70387-4_12.
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Stephens, L. C., L. Milas, K. K. Ang, K. A. Mason e R. E. Meyn. "Apoptosis In Vivo". In Tumor Models in Cancer Research, 625–40. Totowa, NJ: Humana Press, 2010. http://dx.doi.org/10.1007/978-1-60761-968-0_25.
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Garcia, Kristen, Marcus Hock, Vikrant Jaltare, Can Uysalel, Kimberly J. McCabe, Abigail Teitgen e Daniela Valdez-Jasso. "Investigating the Multiscale Impact of Deoxyadenosine Triphosphate (dATP) on Pulmonary Arterial Hypertension (PAH) Induced Heart Failure". In Computational Physiology, 77–90. Cham: Springer International Publishing, 2022. http://dx.doi.org/10.1007/978-3-031-05164-7_7.
Abstract (sommario):
Abstract2-deoxy-ATP (dATP) is a myosin activator known to improve cardiac contractile force [1]. In vitro studies have shown that dATP alters the calcium transient profile in addition to the kinetics of the cross-bridge cycle [2]. Furthermore, in vivo studies of transgenic mice with increased production of dATP show elevated left ventricular systolic function [3]. Pulmonary arterial hypertension (PAH) is a rare disease of the pulmonary vasculature in which pressure overload in the right ventricle results in reduced contractile function and right heart failure [4]. We hypothesize that dATP may have a therapeutic effect on PAH-induced heart failure, by improving contractile function and restoring cardiac output and ejection fraction. However, because the effects of dATP cannot easily be assessed experimentally, we propose using a computational multiscale modeling approach to predict cardiac function. By altering parameters in an existing multiscale biventricular cardiac model [5], we were able to reproduce end-systolic and end-diastolic pressures and volumes that reflect the PAH condition, as well as healthy hearts. dATP was simulated by adjusting parameters in the model at the molecular and cellular levels based on experimental data [1], allowing us to predict the effects of dATP on PAH at the organ level. Our results show that the molecular effects of dATP can increase cardiac output and restore ejection fraction in PAH conditions, though at the cost of elevated mean arterial pressure, and may provide a new approach to treating this disease. Our multiscale modeling approach paves the way for further studies mapping out cardiovascular mechanics. As novel therapeutics continue to be discovered, their application and mechanism can be further explored through these computational models.
Atti di convegni sul tema "In cellulo and in vivo models":
1
Boppart, Stephen A., Wolfgang Drexler, Uwe Morgner, Franz X. Kärtner e James G. Fujimoto. "Ultrahigh Resolution and Spectroscopic OCT Imaging of Cellular Morphology and Function". In In Vivo optical Imaging at the NIH. Washington, D.C.: Optica Publishing Group, 1999. http://dx.doi.org/10.1364/ivoi.1999.msi56.
Abstract (sommario):
Optical coherence tomography (OCT) is a promising optical microscopy technique which enables ultrahigh-resolution, spectroscopic, in vivo imaging in transparent and non-transparent biological specimens. This is achieved by exploiting the short temporal coherence of ultrabroad bandwidth light sources to image morphological features at subcellular resolution at depths beyond that of conventional bright-field and confocal microscopes. Extraction of spatially resolved spectroscopic information is feasible to improve image contrast and to obtain functional or biochemical properties of the investigated tissue. The potential for using OCT to image the morphological expression of genes involved in normal and abnormal development has been shown in common developmental biology animal models. In vivo imaging of single cell morphology, mitosis, and migration, as well as preliminary spectroscopic imaging, is demonstrated.
2
Yasubumi Sakakibara, Hirotaka Nakagawa, Yusaku Nakashima e Katsuyuki Yugi. "Implementing in vivo cellular automata using toggle switch and inter-bacteria communication mechanism". In 2007 2nd Bio-Inspired Models of Network, Information and Computing Systems (BIONETICS). IEEE, 2007. http://dx.doi.org/10.1109/bimnics.2007.4610137.
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Sakakibara, Yasubumi, Hirotaka Nakagawa, Yusaku Nakashima e Katsuyuki Yugi. "Implementing in vivo Cellular Automata using Toggle Switch and Inter-Bacteria Communication Mechanism". In 2nd International ICST Conference on Bio-Inspired Models of Network, Information, and Computing Systems. IEEE, 2007. http://dx.doi.org/10.4108/icst.bionetics2007.2410.
4
Williams, Amber, e Mark Niedre. "Detection of Circulating Tumor Cells and Clusters Using a 2-Fluorophore Diffuse in Vivo Flow Cytometer". In Optical Molecular Probes, Imaging and Drug Delivery. Washington, D.C.: Optica Publishing Group, 2023. http://dx.doi.org/10.1364/omp.2023.om4e.4.
Abstract (sommario):
We discuss development and validation of a new two-fluorophore diffuse in vivo flow cytometer instrument designed to detect two populations of circulating tumor cells and multi-cellular clusters in small animal metastasis models.
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Ehlert, Jan E., Bettina Mutschler, Melanie Müller, Andreas Lingnau, Steffen Hoffmann e Michael HG Kubbutat. "Abstract 3225: Cellular and in vivo models for the analyses of B-Raf and c-Src inhibitors". In Proceedings: AACR 101st Annual Meeting 2010‐‐ Apr 17‐21, 2010; Washington, DC. American Association for Cancer Research, 2010. http://dx.doi.org/10.1158/1538-7445.am10-3225.
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Marshall, Lauren, Isabel Löwstedt, Paul Gatenholm e Joel Berry. "Prevascularized, Co-Culture Model for Breast Cancer Drug Development". In ASME 2012 Summer Bioengineering Conference. American Society of Mechanical Engineers, 2012. http://dx.doi.org/10.1115/sbc2012-80409.
Abstract (sommario):
The objective of this study was to create 3D engineered tissue models to accelerate identification of safe and efficacious breast cancer drug therapies. It is expected that this platform will dramatically reduce the time and costs associated with development and regulatory approval of anti-cancer therapies, currently a multi-billion dollar endeavor [1]. Existing two-dimensional (2D) in vitro and in vivo animal studies required for identification of effective cancer therapies account for much of the high costs of anti-cancer medications and health insurance premiums borne by patients, many of whom cannot afford it. An emerging paradigm in pharmaceutical drug development is the use of three-dimensional (3D) cell/biomaterial models that will accurately screen novel therapeutic compounds, repurpose existing compounds and terminate ineffective ones. In particular, identification of effective chemotherapies for breast cancer are anticipated to occur more quickly in 3D in vitro models than 2D in vitro environments and in vivo animal models, neither of which accurately mimic natural human tumor environments [2]. Moreover, these 3D models can be multi-cellular and designed with extracellular matrix (ECM) function and mechanical properties similar to that of natural in vivo cancer environments [3].
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Boppart, Stephen A., Gary J. Tearney, Brett E. Bouma, James G. Fujimoto e Mark E. Brezinski. "Optical Coherence Tomography of Embryonic Morphology During Cellular Differentiation". In Advances in Optical Imaging and Photon Migration. Washington, D.C.: Optica Publishing Group, 1996. http://dx.doi.org/10.1364/aoipm.1996.cit231.
Abstract (sommario):
Improved imaging of morphological changes has the potential of offering new insight into the complex process of embryonic development. Optical coherence tomography (OCT), is a new imaging technique for performing in vivo cross-sectional imaging of architectural morphology by measuring backscattered infrared light. This study investigates the application of OCT for imaging developing structure in Xenopus laevis (African frog) and Brachydanio rerio (zebra fish), two developmental biology animal models. Images are compared to corresponding histological preparations. Cross sectional imaging can be performed and structural morphology identified at greater imaging depths than possible with confocal and light microscopy. Repeated OCT imaging may be performed in vivo in order to track structural changes throughout development. Imaging in vivo microscopic embryonic morphology with OCT is a fundamental biological research application for the study of genetic disease.
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Baker, Jonathan, Genki Kimura, Yuki Nishimoto, Shyreen Hassibi, Yasuo Kizawa e Kazuhiro Ito. "The senolytic effect of Dasatinib and Quercetin on cellular senescence in COPD in vitro and in vivo models". In ERS International Congress 2023 abstracts. European Respiratory Society, 2023. http://dx.doi.org/10.1183/13993003.congress-2023.pa4046.
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Uddin, Sardar M. Zia, e Yi-Xian Qin. "Anabolic Effects of Ultrasound as Countermeasures of Simulated Microgravity in In-Vitro and In-Vivo Functional Disuse Models". In ASME 2011 Summer Bioengineering Conference. American Society of Mechanical Engineers, 2011. http://dx.doi.org/10.1115/sbc2011-53796.
Abstract (sommario):
Microgravity (MG) during space flight has been known to cause adverse effect on bone quality. Data collected from studies done on spaceflights show loss of 1–1.6% bone mineral density (BMD) per space-flight-month[1]. Most BMD has been recorded in load-bearing bones [2]. Some studies has considered using drugs and different growth factors to maintain bone mass in microgravity conditions but it can be too expensive to maintain over longer periods of time besides the systematic effects of such treatments [3]. Considering the effects of microgravity are partially attributed to lack of mechanical force on bone tissue, which alters gene expression, reduction in transcription factors and growth factors. Furthermore, lack of gravity effects cell growth, proliferation, differentiation, cytoskeleton polymerization and cellular morphology [4, 5]. Thus to reverse these adverse effects on bone physiology, it is important to provide cells with mechanical stimulus which can provide essential mechanical signal for cells to counter the effects of microgravity. Ultrasound acoustic vibrations can be readily applied in, in vivo and human studies and has shown anabolic effects on osteopenic bone tissue [6]. Furthermore, ultrasound is a non-invasive and more target specific treatment relative to cyclic strain and vibration. The objective of this study is to see effects of low intensity pulsed ultrasound (LIPUS) on disused bone model and osteogenic activity of osteoblast cells cultures in simulated microgravity. This will help us understand that effects of ultrasound on microgravity and mechanotransduction pathway responsible for anabolic effect on bone cells.
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Zwick, H., B. E. Stuck, W. R. Elliott, D. J. Lund, S. T. Schuschereba e G. Li. "An Animal Model for In-Vivo Characterization of Laser Induced Retinal cellular Pathology and Recovery Processes". In In Vivo optical Imaging at the NIH. Washington, D.C.: Optica Publishing Group, 1999. http://dx.doi.org/10.1364/ivoi.1999.msi31.
Abstract (sommario):
In the high numerical aperture eye of the snake, the photoreceptor matrix can be imaged in vivo under anesthetized conditions, providing a unique capability to image acute photic damage effects on photoreceptor and anterior retinal blood cell dynamics in response to retinal laser injury. New insights into photic damage and neural repair mechanisms become available from such in vivo cellular observations.
Rapporti di organizzazioni sul tema "In cellulo and in vivo models":
1
Eldar, Avigdor, e Donald L. Evans. Streptococcus iniae Infections in Trout and Tilapia: Host-Pathogen Interactions, the Immune Response Toward the Pathogen and Vaccine Formulation. United States Department of Agriculture, dicembre 2000. http://dx.doi.org/10.32747/2000.7575286.bard.
Abstract (sommario):
In Israel and in the U.S., Streptococcus iniae is responsible for considerable losses in various fish species. Poor understanding of its virulence factors and limited know-how-to of vaccine formulation and administration are the main reasons for the limited efficacy of vaccines. Our strategy was that in order to Improve control measures, both aspects should be equally addressed. Our proposal included the following objectives: (i) construction of host-pathogen interaction models; (ii) characterization of virulence factors and immunodominant antigens, with assessment of their relative importance in terms of protection and (iii) genetic identification of virulence factors and genes, with evaluation of the protective effect of recombinant proteins. We have shown that two different serotypes are involved. Their capsular polysaccharides (CPS) were characterized, and proved to play an important role in immune evasion and in other consequences of the infection. This is an innovative finding in fish bacteriology and resembles what, in other fields, has become apparent in the recent years: S. iniae alters surface antigens. By so doing, the pathogen escapes immune destruction. Immunological assays (agar-gel immunodiffusion and antibody titers) confirmed that only limited cross recognition between the two types occurs and that capsular polysaccharides are immunodominant. Vaccination with purified CPS (as an acellular vaccine) results in protection. In vitro and ex-vivo models have allowed us to unravel additional insights of the host-pathogen interactions. S. iniae 173 (type II) produced DNA fragmentation of TMB-8 cells characteristic of cellular necrosis; the same isolate also prevented the development of apoptosis in NCC. This was determined by finding reduced expression of phosphotidylserine (PS) on the outer membrane leaflet of NCC. NCC treated with this isolate had very high levels of cellular necrosis compared to all other isolates. This cellular pathology was confirmed by observing reduced DNA laddering in these same treated cells. Transmission EM also showed characteristic necrotic cellular changes in treated cells. To determine if the (in vitro) PCD/apoptosis protective effects of #173 correlated with any in vivo activity, tilapia were injected IV with #173 and #164 (an Israeli type I strain). Following injection, purified NCC were tested (in vitro) for cytotoxicity against HL-60 target cells. Four significant observations were made : (i) fish injected with #173 had 100-400% increased cytotoxicity compared to #164 (ii) in vivo activation occurred within 5 minutes of injection; (iii) activation occurred only within the peripheral blood compartment; and (iv) the isolate that protected NCC from apoptosis in vitro caused in vivo activation of cytotoxicity. The levels of in vivo cytotoxicity responses are associated with certain pathogens (pathogen associated molecular patterns/PAMP) and with the tissue of origin of NCC. NCC from different tissue (i.e. PBL, anterior kidney, spleen) exist in different states of differentiation. Random amplified polymorphic DNA (RAPD) analysis revealed the "adaptation" of the bacterium to the vaccinated environment, suggesting a "Darwinian-like" evolution of any bacterium. Due to the selective pressure which has occurred in the vaccinated environment, type II strains, able to evade the protective response elicited by the vaccine, have evolved from type I strains. The increased virulence through the appropriation of a novel antigenic composition conforms with pathogenic mechanisms described for other streptococci. Vaccine efficacy was improved: water-in-oil formulations were found effective in inducing protection that lasted for a period of (at least) 6 months. Protection was evaluated by functional tests - the protective effect, and immunological parameters - elicitation of T- and B-cells proliferation. Vaccinated fish were found to be resistant to the disease for (at least) six months; protection was accompanied by activation of the cellular and the humoral branches.
2
Parada, Luis F. In Vivo Models of NF-1: The Nervous System and Tumorigenesis. Fort Belvoir, VA: Defense Technical Information Center, settembre 2000. http://dx.doi.org/10.21236/ada386287.
3
Parada, Luis. In Vivo Models of NF-1: The Nervous System and Tumorigenesis. Fort Belvoir, VA: Defense Technical Information Center, settembre 1999. http://dx.doi.org/10.21236/ada391279.
4
Geisbert, Thomas W. Pathogenesis of Ebola Hemorrhagic Fever in Primate Models In Vivo and In Vitro. Fort Belvoir, VA: Defense Technical Information Center, gennaio 2003. http://dx.doi.org/10.21236/ad1012627.
5
Young, Pamela R. Advanced Imaging Approaches to Characterize Stromal and Metabolic Changes in In Vivo Mammary Tumor Models. Fort Belvoir, VA: Defense Technical Information Center, marzo 2013. http://dx.doi.org/10.21236/ada580941.
6
Spiers, Donald, Arieh Gertler, Harold Johnson e James Spain. An In Vitro and In Vivo Investigation of the Diverse Biological Activities of Bovine Placental Lactogen. United States Department of Agriculture, agosto 1993. http://dx.doi.org/10.32747/1993.7568087.bard.
Abstract (sommario):
In order to understand the structure-function relationship of bovine placental lactogen (bPL) and initiate production of material for in vivo testing, 28 different bPL analogues were prepared by either truncation or site-directed mutagenesis. The effect of these mutations was determined by measuring binding capacity, ability to homodimerize extracellular domains (ECDs) of several lactogenic and somatogenic receptors, and by in vitro bioassays. Two analogues were prepared in large amounts for in vivo studies. These studies (a) identified the residues responsible for the somatogenic activity of bPL (K73, G133, T188) and for both lactogenic and somatogenic activity (N-terminus, K185, Y190); (b) allowed preparation of bPL analogues with selectively abolished or reduced somatogenic activity; and (c) provided a tool to understand the kinetic difference between lactogenic and somatogenic receptors. In vivo studies using rodent and dairy models showed that bovine growth hormone (bGH) is superior to bPL in stimulating growth and lactation. Likewise, bGH has greater somatogenic activity in different age groups and thermal environments. Initial studies of bPL analog T188 suggest that its lactogenic potential is superior to bGH. Effective experimental models have now been developed and tested for analysis of new bPL analogs.
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Kopeć, Aneta, Ewa Piątkowska e Joanna Skoczylas. Sardines and sprats as the potential source of nutrients required for supporting proper function of immune system in in vitro and in vivo models. Publishing House of the University of Agriculture in Krakow, 2024. http://dx.doi.org/10.15576/repourk/2024.1.01.
8
Lioy, P. J., M. Gallo, P. Georgopoulos, R. Tate e B. Buckley. Comparison of the bioavailability of elemental waste laden soils using in vivo and in vitro analytical methodology and refinement of exposure/dose models. 1998 annual progress report. Office of Scientific and Technical Information (OSTI), giugno 1998. http://dx.doi.org/10.2172/13580.
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Boisclair, Yves R., e Arieh Gertler. Development and Use of Leptin Receptor Antagonists to Increase Appetite and Adaptive Metabolism in Ruminants. United States Department of Agriculture, gennaio 2012. http://dx.doi.org/10.32747/2012.7697120.bard.
Abstract (sommario):
Objectives The original project had 2 major objectives: (1) To determine the effects of centrally administered leptin antagonist on appetite and adaptive metabolism in the sheep; (2) To develop and prepare second-generation leptin antagonists combining high binding affinity and prolonged in vivo half-life. Background Periods of suboptimal nutrition or exaggerated metabolic activity demands lead to a state of chronic energy insufficiency. Ruminants remain productive for a surprisingly long period of time under these circumstances by evoking adaptations sparing available energy and nutrients. The mechanism driving these adaptations in ruminant remains unknown, but could involve a reduction in plasma leptin, a hormone acting predominantly in the brain. In laboratory animals, reduced leptin signaling promotes survival during nutritional insufficiency by triggering energy sparing adaptations such as reduced thyroid hormone production and insulin resistance. Our overall hypothesis is that similar adaptations are triggered by reduced leptin signaling in the brain of ruminants. Testing of this hypothesis in ruminants has not been possible due to inability to block the actions of endogenous leptin and access to ruminant models where leptin antagonistic therapy is feasible and effective. Major achievements and conclusions The Israeli team had previously mutated 3 residues in ovine leptin, with no effect on receptor binding. This mutant was renamed ovine leptin antagonist (OLA) because it cannot activate signaling and therefore antagonizes the ability of wild type leptin to activate its receptor. To transform OLA into an effective in vivo antagonist, the Israeli made 2 important technical advances. First, it incorporated an additional mutation into OLA, increasing its binding affinity and thus transforming it into a super ovine leptin antagonist (SOLA). Second, the Israeli team developed a method whereby polyethylene glycol is covalently attached to SOLA (PEG-SOLA) with the goal of extending its half-life in vivo. The US team used OLA and PEG-SOLA in 2 separate animal models. First, OLA was chronically administered directly into the brain of mature sheep via a cannula implanted into the 3rdcerebroventricule. Unexpectedly, OLA had no effect of voluntary feed intake or various indicators of peripheral insulin action but reduced the plasma concentration of thyroid hormones. Second, the US team tested the effect of peripheral PEG-SOLA administration in an energy sensitive, rapidly growing lamb model. PEG-SOLA was administered for 14 consecutive days after birth or for 5 consecutive days before sacrifice on day 40 of life. Plasma PEG-SOLA had a half-life of over 16 h and circulated in 225- to 288-fold excess over endogenous leptin. PEG-SOLA administration reduced plasma thyroid hormones and resulted in a higher fat content in the carcass at slaughter, but had no effects on feed intake, body weight, plasma glucose or insulin. These results show that the team succeeded in developing a leptin antagonist with a long in vivo half-life. Moreover, in vivo results show that reduced leptin signaling promotes energy sparing in ruminants by repressing thyroid hormone production. Scientific and agricultural implications The physiological role of leptin in ruminants has been difficult to resolve because peripheral administration of wild type leptin causes little effects. Our work with leptin antagonists show for the first time in ruminants that reduced leptin signaling induces energy sparing mechanisms involving thyroid hormone production with little effect on peripheral insulin action. Additional work is needed to develop even more potent leptin antagonists, to establish optimal administration protocols and to narrow down phases of the ruminant life cycle when their use will improve productivity.
10
Cao, Siyang, Yihao Wei, Tiantian Qi, Peng Liu, Yingqi Chen, Fei Yu, Hui Zeng e Jian Weng. Stem cell therapy for peripheral nerve injury: An up-to-date meta-analysis of 55 preclinical researches. INPLASY - International Platform of Registered Systematic Review and Meta-analysis Protocols, ottobre 2022. http://dx.doi.org/10.37766/inplasy2022.10.0083.
Abstract (sommario):
Review question / Objective: It has been the gold standard for decades to reconstruct a large peripheral nerve injury with a nerve autograft, and this remains true today as well. In addition to nerve autografts, biological conduits and vessels can also be applied. A fair amount of studies have examined the benefits of adding stem cells to the lumen of a nerve conduit. The aim of this meta-analysis was to summarize animal experiments related to the utilization of stem cells as a luminal additive when rebuilding a peripheral nerve injury using nerve grafts. Eligibility criteria: The inclusion criteria were as following: 1.Reconstruction of peripheral nerve injury; 2.Complete nerve transection with gap defect created; 3.Animal in-vivo models; 4.Experimental comparisons between nerve conduits containing and not containing one type of stem cell; 5.Functional testing and electrophysiology evaluations are performed. The exclusion criteria were as following: 1.Repair of central nervous system; 2.Nerve repair is accomplished by end-to-end anastomosis; 3.Animal models of entrapment injuries, frostbite, traction injuries and electric injuries; 4.Nerve conduits made from autologous epineurium; 5.Clinical trials, reviews, letters, conference papers, meta-analyses or commentaries; 6.Same studies have been published in different journals under the same or a different title.