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  • About
  • The Global ETD Search service is a free service for researchers to find electronic theses and dissertations. This service is provided by the Networked Digital Library of Theses and Dissertations.
    Our metadata is collected from universities around the world. If you manage a university/consortium/country archive and want to be added, details can be found on the NDLTD website.
1

Maternal exposure to volatile anesthetics induces IL-6 in fetal brains and affects neuronal development / 母体への揮発性麻酔薬投与は胎児脳においてIL-6を誘導し神経発達に影響を及ぼす

Hirotsu, Akiko 23 March 2020 (has links)
京都大学 / 0048 / 新制・課程博士 / 博士(医学) / 甲第22310号 / 医博第4551号 / 新制||医||1040(附属図書館) / 京都大学大学院医学研究科医学専攻 / (主査)教授 渡邉 大, 教授 万代 昌紀, 教授 影山 龍一郎 / 学位規則第4条第1項該当 / Doctor of Medical Science / Kyoto University / DFAM
2

Assessing Epidermal Growth Factor Expression in the Rodent Hippocampus Following Traumatic Brain Injury

Daus, Janice Mabutas 01 January 2006 (has links)
Hippocampal neurons are vulnerable to injury, as indicated by the prevalence of learning and memory deficits following traumatic brain injury. Research indicates that proliferation of neural precursor cells increases following brain injury, which implies that there is an endogenous response in the hippocampus to replenish neurons and restore cognitive function. Studies show that mitogenic growth factors may drive this proliferative response; one of which is epidermal growth factor. Because adults and the elderly manifest the most enduring deficits following TBI, it is critical to investigate how EGF expression following injury may relate to injury-induced cell proliferation and the degree of cognitive recovery observed with aging. In the current study, we assessed the temporal and spatial expression of EGF in the injured hippocampus with age. Our results suggest that EGF expression increases following TBI, and this increase is more significant in the younger brain. Additionally, we investigated the phenotype and localization of cells that express EGF following injury.
3

An in vitro model of the brain tissue reaction to chronically implanted recording electrodes reveals essential roles for serum and bFGF in glial scarring

Polikov, Vadim Steven January 2009 (has links)
<p>Chronically implanted recording electrode arrays linked to prosthetics have the potential to make positive impacts on patients suffering from full or partial paralysis [1;2]. Such arrays are implanted into the patient's cortical tissue and record extracellular potentials from nearby neurons, allowing the information encoded by the neuronal discharges to control external devices. While such systems perform well during acute recordings, they often fail to function reliably in clinically relevant chronic settings [3]. Available evidence suggests that a major failure mode of electrode arrays is the brain tissue reaction against these implants (termed the glial scar), making the biocompatibility of implanted electrodes a primary concern in device design. Previous studies have focused on modifying the form factor of recording arrays, implanting such arrays in experimental animals, and, upon explantation, evaluating the glial scarring in response to the implant after several weeks in vivo. Because of a lack of information regarding the mechanisms involved in the tissue reaction to implanted biomaterials in the brain, it is not surprising that these in vivo studies have met with limited success. This dissertation describes the development of a simple, controlled in vitro model of glial scarring and the utilization of that model to probe the cellular and molecular mechanisms behind glial scarring.</p><p>A novel in vitro model of glial scarring was developed by adapting a primary cell-based system previously used for studying neuroinflammatory processes in neurodegenerative disease [4]. Midbrains from embryonic day 14 Fischer 344 rats were mechanically dissociated and grown on poly-D-lysine coated 24 well plates to a confluent layer of neurons, astrocytes, and microglia. The culture was injured with either a mechanical scrape or foreign-body placement (segments of 50 mm diameter stainless steel microwire), fixed at time points from 6 h to 10 days, and assessed by immunocytochemistry. Microglia invaded the scraped wound area at early time points and hypertrophied activated astrocytes repopulated the wound after 7 days. The chronic presence of microwire resulted in a glial scar forming at 10 days, with microglia forming an inner layer of cells coating the microwire, while astrocytes surrounded the microglial core with a network of cellular processes containing upregulated GFAP. Neurons within the culture did not repopulate the scrape wound and did not respond to the microwire, although they were determined to be electrically active through patch clamp recording. </p><p>This initial model recreated many of the hallmarks of glial scarring around electrodes used for recording in the brain; however, the model lacked the reproducibility necessary to establish a useful characterization tool. After the protocol was amended to resemble protocols typically used to culture neural stem/precursor cells, an intense scarring reaction was consistently seen [5]. To further optimize and characterize the reaction, six independent cell culture variables (growth media, seeding density, bFGF addition day, serum concentration in treatment media, treatment day, and duration of culture) were varied systematically and the resulting scars were quantified. The following conditions were found to give the highest level of scarring: Neurobasal medium supplemented with B27, 10% fetal bovine serum at treatment, 10 ng/ml b-FGF addition at seeding and at treatment, treatment at least 6 days after seeding and scar growth of at least 5 days. Seeding density did not affect scarring as long as at least 500,000 cells were seeded per well, but appropriate media, bFGF, and serum were essential for significant scar formation. </p><p>The optimized in vitro model was then used to help uncover the underlying molecular and cellular mechanisms behind glial scarring. A microwire coating that mimics the basal lamina present within glial scars was developed that allows cells responding to the coated microwire to be isolated and evaluated (i.e. through cell counting or cell staining). A panel of soluble factors known to be involved in glial scar formation was added to the media and the cellular response was recorded. The extent of cell accumulation on the coated microwires was significantly increased by titration of the culture with serum, the pleotropic growth factor bFGF, the inflammatory cytokines IL-1&alpha; and IL-1&beta;, and the growth factors PDGF and BMP-2. The other fourteen soluble factors tested had little to no effect on the number of cells that attached to the coated microwires, although a specific blocker of the bFGF receptor was able to abrogate the effect of bFGF. This study proposes essential roles in glial scarring of serum, which infiltrates brain tissue upon disruption of the blood-brain barrier, and bFGF, which is a necessary growth and survival factor for the neural precursor cells that respond to injury. These insights suggest repeated rounds of implant micromotion-induced cellular damage, with the resultant neuronal death, serum release, and bFGF deposition may thicken the glial scar and lead to recording signal loss.</p> / Dissertation
4

Defining the Mechanisms By Which Transplanted Neural Precursor Cells Mediate Functional Recovery Following Spinal Cord Injury

Hawryluk, Gregory 15 August 2013 (has links)
Spinal cord injury (SCI) is uniquely devastating. Cellular transplantation strategies for SCI are showing promise. Little, however, is known about how transplanted neural precursor cells (NPCs) enhance functional recovery or the mechanisms by which they interact with the host spinal cord. Better understanding of these critical issues may lead to improved strategies to enhance recovery after SCI. Given this background, I hypothesized that NPCs mediate functional recovery by a number of mechanisms including trophin production, neuroprotection, modulation of the host inflammatory response or glial scarring, and/or remyelination. I thus endeavored to characterize trophin production by NPCs in vitro and in vivo in rats with clip compression SCI of the thoracic spinal cord, to determine if preservation of host cells and tissue contribute to functional recovery and to determine how NPC transplantation influences the host inflammatory response and glial scarring. Here I present unique and novel insights into NPC-host interactions following SCI. We show that NPCs are poised to provide trophic support to the injured spinal cord. We also show that the combination of NPCs, pharmacotherapy and trophin infusion is associated with sparing of grey and white matter, enhanced numbers of oligodendrocytes but not axons as well as an increased inflammatory response. To assess the potential impact of myelination as a mechanism underlying NPC-mediated functional recovery after SCI, experiments were undertaken using NPCs derived from shiverer mutant mice unable to produce central myelin. These experiments showed that while NPCs from wild-type mice generate myelin and mediate functional recovery after SCI; transplanted shiverer NPCs impede neurobehavioural recovery. In summary, my work provides unique insights into the functional effects of NPC transplantation after SCI. Of importance, this thesis provides novel evidence that remyelination is a key mechanism of action by which NPCs mediate recovery after SCI. Hence, this work has important implications for patients with SCI.
5

Defining the Mechanisms By Which Transplanted Neural Precursor Cells Mediate Functional Recovery Following Spinal Cord Injury

Hawryluk, Gregory 15 August 2013 (has links)
Spinal cord injury (SCI) is uniquely devastating. Cellular transplantation strategies for SCI are showing promise. Little, however, is known about how transplanted neural precursor cells (NPCs) enhance functional recovery or the mechanisms by which they interact with the host spinal cord. Better understanding of these critical issues may lead to improved strategies to enhance recovery after SCI. Given this background, I hypothesized that NPCs mediate functional recovery by a number of mechanisms including trophin production, neuroprotection, modulation of the host inflammatory response or glial scarring, and/or remyelination. I thus endeavored to characterize trophin production by NPCs in vitro and in vivo in rats with clip compression SCI of the thoracic spinal cord, to determine if preservation of host cells and tissue contribute to functional recovery and to determine how NPC transplantation influences the host inflammatory response and glial scarring. Here I present unique and novel insights into NPC-host interactions following SCI. We show that NPCs are poised to provide trophic support to the injured spinal cord. We also show that the combination of NPCs, pharmacotherapy and trophin infusion is associated with sparing of grey and white matter, enhanced numbers of oligodendrocytes but not axons as well as an increased inflammatory response. To assess the potential impact of myelination as a mechanism underlying NPC-mediated functional recovery after SCI, experiments were undertaken using NPCs derived from shiverer mutant mice unable to produce central myelin. These experiments showed that while NPCs from wild-type mice generate myelin and mediate functional recovery after SCI; transplanted shiverer NPCs impede neurobehavioural recovery. In summary, my work provides unique insights into the functional effects of NPC transplantation after SCI. Of importance, this thesis provides novel evidence that remyelination is a key mechanism of action by which NPCs mediate recovery after SCI. Hence, this work has important implications for patients with SCI.
6

Veränderungen der adulten Neurogenese im Hippocampus von Drogenabhängigen

Bayer, Ronny 07 April 2015 (has links) (PDF)
Die Neubildung von Neuronen persistiert lebenslang in der Subgranularzellschicht des Hippocampus und der Subventrikularzone des Großhirns und wird als adulte Neuroge-nese bezeichnet. Es wird vermutet, dass diese beim erwachsenen Menschen einen rele-vanten Einfluss auf degenerative Veränderungen, verschiedene neurologische Krank-heitsbilder und auf die (Dys-)Funktion des Gedächtnisses hat. Im Tiermodell wurde eine Verringerung der Neurogenese nach chronischer Morphingabe nachgewiesen. Vorarbeiten zeigten einen Zusammenhang zwischen chronischem Heroinmissbrauch und reaktiver Astrogliose, Mikrogliose und einer vermehrten Expression des polysialylated neural cell adhesion molecule im humanen Hippocampus. Daraus leitet sich die Hypothese ab, dass chronischer Heroinmissbrauch, als Modell für eine Abhängigkeitserkrankung, einen Einfluss auf die adulte humane Neurogenese hat. Es wurden in Formalin fixierte Gewebeproben aus dem Hippocampus von Verstorbenen mit einer letalen Heroinintoxikation und mit bekanntem Heroinmissbrauch (n = 20) un-tersucht und mit einer nach Alter und Geschlecht angepassten Kontrollgruppe (n = 28) verglichen. Hierbei wurden spezifische Neurogenesemarker mittels immunhistochemi-scher Methoden angewendet und ausgewertet. Es bestand eine generell sehr geringe zelluläre Proliferationsrate und eine signifikante Reduktion Musashi-1 positiver neuro-naler Vorläuferzellen bei gleichzeitig unveränderter Anzahl Nestin positiver reifender und Calretinin positiver migrierender postmitotischer Neurone. Zudem wurde ein ver-ändertes Calretinin-Expressionsmuster als Hinweis auf eventuelle funktionelle neuronale Defizite bei Drogenabhängigen festgestellt. Der potentielle Einfluss von chronischem Heroinmissbrauch auf die adulte humane Neurogenese wird erstmals gezeigt. Die Ergebnisse weisen auf eine negative Beeinflus-sung im Stadium neuronaler Vorläuferzellen und der Zellfunktion migrierender Neurone in der Fallgruppe im Vergleich zu einer gesunden Kontrollgruppe hin. Diese Hemmung der Neurogenese könnte eine Erklärungsmöglichkeit für kognitive Defizite und Funktionsstörungen des Gedächtnisses infolge chronischen Drogenkonsums bieten und zugleich eine Bedeutung bei der Entstehung von Abhängigkeitserkrankungen haben. Insofern könnte sich hier ein Ansatzpunkt für zukünftige Therapiestrategien derartiger Erkrankungen oder ihrer Folgen bieten.
7

Veränderungen der adulten Neurogenese im Hippocampus von Drogenabhängigen: Immunhistochemische Untersuchungen mit ausgewählten Neurogenesemarkern

Bayer, Ronny 02 March 2015 (has links)
Die Neubildung von Neuronen persistiert lebenslang in der Subgranularzellschicht des Hippocampus und der Subventrikularzone des Großhirns und wird als adulte Neuroge-nese bezeichnet. Es wird vermutet, dass diese beim erwachsenen Menschen einen rele-vanten Einfluss auf degenerative Veränderungen, verschiedene neurologische Krank-heitsbilder und auf die (Dys-)Funktion des Gedächtnisses hat. Im Tiermodell wurde eine Verringerung der Neurogenese nach chronischer Morphingabe nachgewiesen. Vorarbeiten zeigten einen Zusammenhang zwischen chronischem Heroinmissbrauch und reaktiver Astrogliose, Mikrogliose und einer vermehrten Expression des polysialylated neural cell adhesion molecule im humanen Hippocampus. Daraus leitet sich die Hypothese ab, dass chronischer Heroinmissbrauch, als Modell für eine Abhängigkeitserkrankung, einen Einfluss auf die adulte humane Neurogenese hat. Es wurden in Formalin fixierte Gewebeproben aus dem Hippocampus von Verstorbenen mit einer letalen Heroinintoxikation und mit bekanntem Heroinmissbrauch (n = 20) un-tersucht und mit einer nach Alter und Geschlecht angepassten Kontrollgruppe (n = 28) verglichen. Hierbei wurden spezifische Neurogenesemarker mittels immunhistochemi-scher Methoden angewendet und ausgewertet. Es bestand eine generell sehr geringe zelluläre Proliferationsrate und eine signifikante Reduktion Musashi-1 positiver neuro-naler Vorläuferzellen bei gleichzeitig unveränderter Anzahl Nestin positiver reifender und Calretinin positiver migrierender postmitotischer Neurone. Zudem wurde ein ver-ändertes Calretinin-Expressionsmuster als Hinweis auf eventuelle funktionelle neuronale Defizite bei Drogenabhängigen festgestellt. Der potentielle Einfluss von chronischem Heroinmissbrauch auf die adulte humane Neurogenese wird erstmals gezeigt. Die Ergebnisse weisen auf eine negative Beeinflus-sung im Stadium neuronaler Vorläuferzellen und der Zellfunktion migrierender Neurone in der Fallgruppe im Vergleich zu einer gesunden Kontrollgruppe hin. Diese Hemmung der Neurogenese könnte eine Erklärungsmöglichkeit für kognitive Defizite und Funktionsstörungen des Gedächtnisses infolge chronischen Drogenkonsums bieten und zugleich eine Bedeutung bei der Entstehung von Abhängigkeitserkrankungen haben. Insofern könnte sich hier ein Ansatzpunkt für zukünftige Therapiestrategien derartiger Erkrankungen oder ihrer Folgen bieten.:I. Inhaltsverzeichnis 1 II. Bibliografische Zusammenfassung 2 III. Abkürzungsverzeichnis 3 1. Einführung 4 1.1. Drogenabhängigkeit und Epidemiologie 4 1.2. Heroin 6 1.3. Hippocampus 9 1.4. Adulte Neurogenese 11 1.5. Aufgabenstellung und Ziel der Arbeit 14 2. Materialen und Methoden 18 2.1. Fall- und Kontrollgruppe 18 2.2. Toxikologisch-chemische Untersuchungen 20 2.3. Immunhistochemie 21 2.4. Immunfluoreszenz und konfokale Mikroskopie 25 2.5. Quantifizierung, Datenanalyse und Statistik 26 3. Ergebnisse 28 3.1. Deskriptive Datenanalyse 28 3.2. Musashi-1 30 3.3. Nestin 31 3.4. Calretinin 32 3.5. Ki-67 34 3.6. Doublecortin 35 3.7. Doppelimmunfluoreszenz 36 4. Diskussion 37 4.1. Neurogenese – Proliferation (Ki-67) 38 4.2. Neurogenese – Differenzierung (MSI-1, Nestin) 39 4.3. Neurogenese – Reifung (Calretinin) 42 4.4. Methodische Grenzen und Fehlerbetrachtung 43 4.5. Fazit und Ausblick 46 5. Zusammenfassung der Arbeit 48 6. Literaturverzeichnis 52 7. Anlagen 1-8 65 IV. Selbständigkeitserklärung 73 V. Curriculum vitae 74 VI. Publikationen 75 VII. Danksagung 76
8

Signaling Cascade Involved in Rapid Stimulation of Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) by Dexamethasone

Bossmann, Miriam, Ackermann, Benjamin W., Thome, Ulrich H., Laube, Mandy 15 January 2024 (has links)
Impairment of mucociliary clearance with reduced airway fluid secretion leads to chronically inflamed airways. Cystic fibrosis transmembrane conductance regulator (CFTR) is crucially involved in airway fluid secretion and dexamethasone (dexa) has previously been shown to elevate CFTR activity in airway epithelial cells. However, the pathway by which dexa increases CFTR activity is largely unknown. We aimed to determine whether the increase of CFTR activity by dexa is achieved by non-genomic signaling and hypothesized that the phosphoinositide 3-kinase (PI3K) pathway is involved in CFTR stimulation. Primary rat airway epithelial cells and human bronchial submucosal gland-derived Calu-3 cells were analyzed in Ussing chambers and kinase activation was determined byWestern blots. Results demonstrated a critical involvement of PI3K and protein kinase B (AKT) signaling in the dexa-induced increase of CFTR activity, while serum and glucocorticoid dependent kinase 1 (SGK1) activity was not essential. We further demonstrated a reduced neural precursor cell expressed, developmentally downregulated 4-like (NEDD4L) ubiquitin E3 ligase activity induced by dexa, possibly responsible for the elevated CFTR activity. Finally, increases of CFTR activity by dexa were demonstrated within 30 min accompanied by rapid activation of AKT. In conclusion, dexa induces a rapid stimulation of CFTR activity which depends on PI3K/AKT signaling in airway epithelial cells. Glucocorticoids might thus represent, in addition to their immunomodulatory actions, a therapeutic strategy to rapidly increase airway fluid secretion.
9

Electronic Devices for the Combination of Electrically Controlled Drug Release, Electrostimulation, and Optogenetic Stimulation for Nerve Tissue Regeneration

Monreal Trigo, Javier 02 June 2023 (has links)
[ES] La capacidad de las células madre para proliferar formando distintas células especializadas les otorga la potencialidad de servir de base para terapias efectivas para patologías cuyo tratamiento era inimaginable hasta hace apenas dos décadas. Sin embargo, esta capacidad se encuentra mediada por estímulos fisiológicos, químicos, y eléctricos, específicos y complejos, que dificultan su traslación a la rutina clínica. Por ello, las células madre representan un campo de estudio en el que se invierten amplios esfuerzos por parte de la comunidad científica. En el ámbito de la regeneración nerviosa, para modular su desarrollo y diferenciación el tratamiento farmacológico, la electroestimulación, y la estimulación optogenética son técnicas que están consiguiendo prometedores resultados. Es por ello por lo que en la presente tesis se ha desarrollado un conjunto de sistemas electrónicos para permitir la aplicación combinada de estas técnicas in vitro, con perspectiva a su aplicación in vivo. Hemos diseñado una novedosa tecnología para la liberación eléctricamente controlada de fármacos. Esta tecnología está basada en nanopartículas de sílice mesoporosa y puertas moleculares de bipiridina-heparina. Las puertas moleculares son electroquímicamente reactivas, y encierran los fármacos en el interior de las nanopartículas, liberándolos ante un estímulo eléctrico. Hemos caracterizado esta tecnología, y la hemos validado mediante la liberación controlada de rodamina en cultivos celulares de HeLa. Para la combinación de liberación controlada de fármacos y electroestimulación hemos desarrollado dispositivos que permiten aplicar los estímulos eléctricos de forma configurable desde una interfaz gráfica de usuario. Además, hemos diseñado un módulo de expansión que permite multiplexar las señales eléctricas a diferentes cultivos celulares. Además, hemos diseñado un dispositivo de estimulación optogenética. Este tipo de estimulación consiste en la modificación genética de las células para que sean sensibles a la radiación lumínica de determinada longitud de onda. En el ámbito de la regeneración de tejido mediante células precursoras neurales, es de interés poder inducir ondas de calcio, favoreciendo su diferenciación en neuronas y la formación de circuitos sinápticos. El dispositivo diseñado permite obtener imágenes en tiempo real mediante microscopía confocal de las respuestas transitorias de las células al ser irradiadas. El dispositivo se ha validado irradiando neuronas modificadas con luz pulsada de 100 ms. También hemos diseñado un dispositivo electrónico complementario de medida de irradiancia con el doble fin de permitir la calibración del equipo de irradiancia y medir la irradiancia en tiempo real durante los experimentos in vitro. Los resultados del uso de los bioactuadores en procesos complejos y dinámicos, como la regeneración de tejido nervioso, son limitados en lazo abierto. Uno de los principales aspectos analizados es el desarrollo de biosensores que permitiesen la cuantización de ciertas biomoléculas para ajustar la estimulación suministrada en tiempo real. Por ejemplo, la segregación de serotonina es una respuesta identificada en la elongación de células precursoras neurales, pero hay otras biomoléculas de interés para la implementación de un control en lazo cerrado. Entre las tecnologías en el estado del arte, los biosensores basados en transistores de efecto de campo (FET) funcionalizados con aptámeros son realmente prometedores para esta aplicación. Sin embargo, esta tecnología no permitía la medición simultánea de más de una biomolécula objetivo en un volumen reducido debido a las interferencias entre los distintos FETs, cuyos terminales se encuentran inmersos en la solución. Por ello, hemos desarrollado instrumentación electrónica capaz de medir simultáneamente varios de estos biosensores, y la hemos validado mediante la medición simultánea de pH y la detección preliminar de serotonina y glutamato. / [CA] La capacitat de les cèl·lules mare per a proliferar formant diferents cèl·lules especialitzades els atorga la potencialitat de servir de base per a teràpies efectives per a patologies el tractament de les quals era inimaginable fins fa a penes dues dècades. No obstant això, aquesta capacitat es troba mediada per estímuls fisiològics, químics, i elèctrics, específics i complexos, que dificulten la seua translació a la rutina clínica. Per això, les cèl·lules mare representen un camp d'estudi en el qual s'inverteixen amplis esforços per part de la comunitat científica. En l'àmbit de la regeneració nerviosa, per a modular el seu desenvolupament i diferenciació el tractament farmacològic, l'electroestimulació, i l'estimulació optogenética són tècniques que estan aconseguint prometedors resultats. És per això que en la present tesi s'ha desenvolupat un conjunt de sistemes electrònics per a permetre l'aplicació combinada d'aquestes tècniques in vitro, amb perspectiva a la seua aplicació in vivo. Hem dissenyat una nova tecnologia per a l'alliberament elèctricament controlat de fàrmacs. Aquesta tecnologia està basada en nanopartícules de sílice mesoporosa i portes moleculars de bipiridina-heparina. Les portes moleculars són electroquímicament reactives, i tanquen els fàrmacs a l'interior de les nanopartícules, alliberant-los davant un estímul elèctric. Hem caracteritzat aquesta tecnologia, i l'hem validada mitjançant l'alliberament controlat de rodamina en cultius cel·lulars de HeLa. Per a la combinació d'alliberament controlat de fàrmacs i electroestimulació hem desenvolupat dispositius que permeten aplicar els estímuls elèctrics de manera configurable des d'una interfície gràfica d'usuari. A més, hem dissenyat un mòdul d'expansió que permet multiplexar els senyals elèctrics a diferents cultius cel·lulars. A més, hem dissenyat un dispositiu d'estimulació optogenètica. Aquest tipus d'estimulació consisteix en la modificació genètica de les cèl·lules perquè siguen sensibles a la radiació lumínica de determinada longitud d'ona. En l'àmbit de la regeneració de teixit mitjançant cèl·lules precursores neurals, és d'interés poder induir ones de calci, afavorint la seua diferenciació en neurones i la formació de circuits sinàptics. El dispositiu dissenyat permet obtindré imatges en temps real mitjançant microscòpia confocal de les respostes transitòries de les cèl·lules en ser irradiades. El dispositiu s'ha validat irradiant neurones modificades amb llum polsada de 100 ms. També hem dissenyat un dispositiu electrònic complementari de mesura d'irradiància amb el doble fi de permetre el calibratge de l'equip d'irradiància i mesurar la irradiància en temps real durant els experiments in vitro. Els resultats de l'ús dels bioactuadors en processos complexos i dinàmics, com la regeneració de teixit nerviós, són limitats en llaç obert. Un dels principals aspectes analitzats és el desenvolupament de biosensors que permeteren la quantització de certes biomolècules per a ajustar l'estimulació subministrada en temps real. Per exemple, la segregació de serotonina és una resposta identificada amb l'elongació de les cèl·lules precursores neurals, però hi ha altres biomolècules d'interés per a la implementació d'un control en llaç tancat. Entre les tecnologies en l'estat de l'art, els biosensors basats en transistors d'efecte de camp (FET) funcionalitzats amb aptàmers són realment prometedors per a aquesta aplicació. No obstant això, aquesta tecnologia no permetia el mesurament simultani de més d'una biomolècula objectiu en un volum reduït a causa de les interferències entre els diferents FETs, els terminals dels quals es troben immersos en la solució. Per això, hem desenvolupat instrumentació electrònica capaç de mesurar simultàniament diversos d'aquests biosensors i els hem validat amb mesurament simultani del pH i la detecció preliminar de serotonina i glutamat. / [EN] The stem cells' ability to proliferate to form different specialized cells gives them the potential to serve as the basis for effective therapies for pathologies whose treatment was unimaginable until just two decades ago. However, this capacity is mediated by specific and complex physiological, chemical, and electrical stimuli that complicate their translation to clinical routine. For this reason, stem cells represent a field of study in which the scientific community is investing a great deal of effort. In the field of nerve regeneration, to modulate their development and differentiation, pharmacological treatment, electrostimulation, and optogenetic stimulation are techniques that are achieving promising results. For this reason, we have developed a set of electronic systems to allow the combined application of these techniques in vitro, with a view to their application in vivo. We have designed a novel technology for the electrically controlled release of drugs. This technology is based on mesoporous silica nanoparticles and bipyridine-heparin molecular gates. The molecular gates are electrochemically reactive and entrap the drugs inside the nanoparticles, releasing them upon electrical stimulus. We have characterized this technology and validated it by controlled release of rhodamine in HeLa cell cultures. For combining electrostimulation and controlled drug release we have developed devices that allow applying the different electrical stimuli in a configurable way from a graphical user interface. In addition, we have designed an expansion module that allows multiplexing electrical signals to different cell cultures. In addition, we have designed an optogenetic stimulation device. This type of stimulation consists of genetically modifying cells to make them sensitive to light radiation of a specific wavelength. In tissue regeneration using neural precursor cells, it is interesting to be able to induce calcium waves, favoring the cell differentiation into neurons and the formation of synaptic circuits. The designed device enable the obtention of real-time images through confocal microscopy of the transient responses of cells upon irradiation. The device has been validated by irradiating modified neurons with 100 ms pulsed light stimulation. We have also designed a complementary electronic irradiance measurement device to allow calibration of the irradiator equipment and measuring irradiance in real time during in vitro experiments. The results of using bioactuators in complex and dynamic processes, such as nerve tissue regeneration, are limited in an open loop. One of the main aspects analyzed is the development of biosensors that would allow quantifying of specific biomolecules to adjust the stimulation provided in real time. For instance, serotonin secretion is an identified response of neural precursor cells elongation, among other biomolecules of interest for the implementation of a closed-loop control. Among the state-of-the-art technologies, biosensors based on field effect transistors (FETs) functionalized with aptamers are promising for this application. However, this technology did not allow the simultaneous measurement of more than one target biomolecule in a small volume due to interferences between the different FETs, whose terminals are immersed in the solution. This is why we have developed electronic instrumentation capable of simultaneously measuring several of these biosensors, and we have validated it with the simultaneous pH measurement and the preliminary detection of serotonin and glutamate. / Monreal Trigo, J. (2023). Electronic Devices for the Combination of Electrically Controlled Drug Release, Electrostimulation, and Optogenetic Stimulation for Nerve Tissue Regeneration [Tesis doctoral]. Universitat Politècnica de València. https://doi.org/10.4995/Thesis/10251/193841

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