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Mitochondria-Endoplasmic Reticulum Contacts in Neuronal Cells: From Physiology to Therapeutics.

紀錄類型:	書目-電子資源 : Monograph/item
正題名/作者:	Mitochondria-Endoplasmic Reticulum Contacts in Neuronal Cells: From Physiology to Therapeutics./
作者:	Dentoni, Giacomo.
出版者:	Ann Arbor : ProQuest Dissertations & Theses, : 2021,
面頁冊數:	109 p.
附註:	Source: Dissertations Abstracts International, Volume: 83-02, Section: B.
Contained By:	Dissertations Abstracts International83-02B.
標題:	Apoptosis. -
電子資源:	http://pqdd.sinica.edu.tw/twdaoapp/servlet/advanced?query=28621539
ISBN:	9798522950521

Mitochondria-Endoplasmic Reticulum Contacts in Neuronal Cells: From Physiology to Therapeutics.
Dentoni, Giacomo.

Mitochondria-Endoplasmic Reticulum Contacts in Neuronal Cells: From Physiology to Therapeutics. - Ann Arbor : ProQuest Dissertations & Theses, 2021 - 109 p.

Source: Dissertations Abstracts International, Volume: 83-02, Section: B.

Thesis (Ph.D.)--Karolinska Institutet (Sweden), 2021.

This item must not be sold to any third party vendors.

Mitochondria and the endoplasmic reticulum (ER) are intracellular organelles that play vital physiological functions. Mitochondria are key players in energy production through adenosinetriphosphate (ATP) production and calcium (Ca2+) buffering, while the ER is involved inprotein and lipid synthesis along with Ca2+ signalling in the cell. In the last 10 years scientistshave realised the importance of intracellular organelle communication as a pivotal process for physiological functions. Among these interactions, mitochondria and ER functionally and structurally interact with each other forming mitochondria-ER contact sites (MERCS).Importantly, these structures oversee a variety of pathways including intracellular Ca2+signalling. Indeed, ER to mitochondria Ca2+ shuttling has been shown to impact onmitochondrial respiration and bioenergetics. On the other hand, sustained increase in Ca2+signalling between these two organelles can cause activation of apoptosis mediators leading to cell death. In Alzheimer´s disease (AD), cerebral hypometabolism, mitochondrial dysfunction,and functional and structural upregulation of ER to mitochondria apposition appear as early events in disease pathogenesis. Despite over 30 years of studies, the causes of AD are essentially unknown and only two symptomatic drugs have been approved for treatment, which means that AD leads to decline of quality of life and ultimately death.In this thesis, using human brain biopsies from idiopathic normal pressure hydrocephalus(iNPH) patients, mouse models of AD and cellular models, we investigated the role of mitochondria and MERCS in synapses and exocytotic mechanism and their role in the development of pathology in AD. Additionally, we have set up a high throughput screen (HTS)to find potential modulators of mitochondrial function with the overarching aim to find drugs to target neurodegenerationIn PAPER I, for the first time we have shown the presence of several organelle contact sites in human brain material and we have confirmed the presence of MERCS in human synapses.In this study we have also shown that patients suffering from dementia have more MERCS compared to non-demented patients. Furthermore, we have shown correlation of soluble Aβ levels, thought to be one of the initiators of AD, and MERCS number in iNPH patients.In PAPER II, through knockdown of Mitofusin 2 (Mfn2) in SH-SY5Y cells, a negativeregulator of MERCS, we have detected substantial increased juxtaposition between ER and mitochondria. Upon Mfn2 knockdown, we have observed decreased levels of cytoplasmicvesicle and increased vesicle release upon cellular depolarization. Furthermore, we have shown that this mechanism was dependent on IP3Rs activity, an important channel for Ca2+ transfer from ER to mitochondria.In PAPER III we have characterised in vitro a novel knock-in model of AD, the AppNL-F model, which overcomes the problem of over expressing amyloid precursor protein (APP). We have shown that embryonic cells derived from AppNL-F mice are capable of secreting levels of Aβ similar to adult brains, causing bioenergetics impairments, movement abnormalities along neurites and increased MERCS functions. Furthermore, these cells seem to be more susceptible to cell death upon inhibition of mitochondrial respiration compared to WT cells.In PAPER IV, we have assessed whether the other pathological protein in AD, tau, impacts on mitochondrial function and MERCS using the pure tauopathy model P301s.

ISBN: 9798522950521Subjects--Topical Terms:

600650
Apoptosis.

Mitochondria-Endoplasmic Reticulum Contacts in Neuronal Cells: From Physiology to Therapeutics.
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506 $a This item must not be sold to any third party vendors.
520 $a Mitochondria and the endoplasmic reticulum (ER) are intracellular organelles that play vital physiological functions. Mitochondria are key players in energy production through adenosinetriphosphate (ATP) production and calcium (Ca2+) buffering, while the ER is involved inprotein and lipid synthesis along with Ca2+ signalling in the cell. In the last 10 years scientistshave realised the importance of intracellular organelle communication as a pivotal process for physiological functions. Among these interactions, mitochondria and ER functionally and structurally interact with each other forming mitochondria-ER contact sites (MERCS).Importantly, these structures oversee a variety of pathways including intracellular Ca2+signalling. Indeed, ER to mitochondria Ca2+ shuttling has been shown to impact onmitochondrial respiration and bioenergetics. On the other hand, sustained increase in Ca2+signalling between these two organelles can cause activation of apoptosis mediators leading to cell death. In Alzheimer´s disease (AD), cerebral hypometabolism, mitochondrial dysfunction,and functional and structural upregulation of ER to mitochondria apposition appear as early events in disease pathogenesis. Despite over 30 years of studies, the causes of AD are essentially unknown and only two symptomatic drugs have been approved for treatment, which means that AD leads to decline of quality of life and ultimately death.In this thesis, using human brain biopsies from idiopathic normal pressure hydrocephalus(iNPH) patients, mouse models of AD and cellular models, we investigated the role of mitochondria and MERCS in synapses and exocytotic mechanism and their role in the development of pathology in AD. Additionally, we have set up a high throughput screen (HTS)to find potential modulators of mitochondrial function with the overarching aim to find drugs to target neurodegenerationIn PAPER I, for the first time we have shown the presence of several organelle contact sites in human brain material and we have confirmed the presence of MERCS in human synapses.In this study we have also shown that patients suffering from dementia have more MERCS compared to non-demented patients. Furthermore, we have shown correlation of soluble Aβ levels, thought to be one of the initiators of AD, and MERCS number in iNPH patients.In PAPER II, through knockdown of Mitofusin 2 (Mfn2) in SH-SY5Y cells, a negativeregulator of MERCS, we have detected substantial increased juxtaposition between ER and mitochondria. Upon Mfn2 knockdown, we have observed decreased levels of cytoplasmicvesicle and increased vesicle release upon cellular depolarization. Furthermore, we have shown that this mechanism was dependent on IP3Rs activity, an important channel for Ca2+ transfer from ER to mitochondria.In PAPER III we have characterised in vitro a novel knock-in model of AD, the AppNL-F model, which overcomes the problem of over expressing amyloid precursor protein (APP). We have shown that embryonic cells derived from AppNL-F mice are capable of secreting levels of Aβ similar to adult brains, causing bioenergetics impairments, movement abnormalities along neurites and increased MERCS functions. Furthermore, these cells seem to be more susceptible to cell death upon inhibition of mitochondrial respiration compared to WT cells.In PAPER IV, we have assessed whether the other pathological protein in AD, tau, impacts on mitochondrial function and MERCS using the pure tauopathy model P301s.
520 $a I mitocondri e il reticolo endoplasmatico (ER) sono organelli intracellulari che hanno importanti funzioni per la salute della cellula. Il primo organello media la produzione di ATP e il buffering del calcio mentre il secondo la sintesi delle proteine, dei lipidi e il buffering del calcio. Negli ultimi dieci anni si e iniziato ad apprezzare come questi organelli non agiscono indipendentemente nella cellula, ma interagiscono tra di loro strutturalmente e funzionalmente in siti di contatto tra i mitochondri e l'ER, chiamati MERCS. Di particolar importanza e il ruolo di queste strutture subcellulari nelle vie di segnalazione intracellulari dipendenti dal calcio (Ca2+). Infatti, e stato dimostrato che il flusso di Ca2+ tra ER e mitochondri e in grado di attivare la respirazione mitocondriale. D'altra parte, se questo flusso di Ca2+ rimanendo costante puo attivare meccanismi apoptotici nella cellula, causando la morte cellulare. Nella malattia di Alzheimer (AD), una forma di demenza senile, ipometabolismo cerebrale, disfunzione mitocondriale e incrementato strutturale e funzionale di MERCS sembrano essere tratti distintivi del decorso della malattia. Tutt'oggi i meccanismi che determinano la malattia sono ancora sconosciuti. Solo due farmaci sono stati approvati per mediare i sintomi, ma nessuna cura per bloccare la malattia e stata trovata nonostate gli ultimi 20 anni di studi.In questa tesi, usando biopsie del cervello di pazienti affetti da idrocefalo normoteso (iNPH), modelli murini di Alzheimer e modelli cellulari, abbiamo cercato di capire come i mitochondri e particolarmente MERCS, siano importanti nella fisiologia sinaptica, esocitosi e nello sviluppo della patologia neuronale nelle malattie neurodegenerative come l'Alzheimer. Inoltre, abbiamo creato una piattaforma cellulare per lo studio di farmaci che potrebbero potenziare l´attivita mitocondriale.Nello specifico, nel PAPER I abbiamo osservato per la prima volta siti di contatto tra i mitocondri e altri organelli in boipsie del cervello e abbiamo appurato la presenza dei MERCS nelle sinapsi umane. Abbiamo inoltre visto che pazienti affetti da varie demenze dimostrano un numero elevato di MERCS rispetto a pazienti non affetti da demenza e notato come il numero di MERCS si correla con i livelli di β-amiloide, una delle possibili cause scatenanti dell'Alzheimer, in pazienti con iNPH.Nel PAPER II, abbiamo artificialmente incrementato i MERCS attraverso il knockdown della proteina Mfn2, un regolatore negativo dei contatti. In questo studio abbiamo visto che aumentando il numero di questi contatti, si osserva un decremento nei livelli di vescicole citoplasmatiche e incremento del rilascio di queste vescicole. Inoltre, abbiamo visto che questo meccanismo e dipendente dall'attivita del recettore dell'inositolo trifosfato (IP3R), importante per il trasferimento di Ca2+ tra ER e mitocondri.Nel PAPER III, abbiamo caratterizzato in vitro un nuovo modello knock-in di AD, chiamato AppNL-F, caratterizzato dall´ assenza di sovraespressione della Proteina precorritrice della betaamiloide (APP), che appunto genera β-amiloide. Abbiamo osservato che cellule embrionali in vitro sono in grado di secernere β-amiloide come nei cervelli adulti, causando un'alterazione dei mitocondri, alterazioni bioenergetiche e di trasporto lungo dendriti e assoni, e dei MERCS. Inoltre, queste cellule sono piu suscettibili a danni quando inibitori della respirazione mitochondriale sono usati, rispetto alle cellule sane.Nel PAPER IV, abbiamo considerato come l'altra proteina resposabile dell´Alzheimer, tau, danneggi i mitochondri e MERCS usando il modello murino di taupatia pura P301s. Abbiamo visto che prima che la patologia dovuta a tau si sviluppasse, sin da 22 giorni le cavie dimostrino gia prolemi nella respirazione mitocondriale e incremento nel numero di MERCS. Questa patologia e sostenuta durante l'invecchiamento delle cavie fino a 10 mesi.Nel PAPER V, creando una piattaforma cellulare per lo studio di farmaci mirati ai mitocondri, abbiamo scoperto che la luteolina, un composto naturale della famiglia dei flavonoidi, e in grado di aumentare fino al venti percento l´ATP cellulare e attivita mitocondriale in vitro in cellule neuroblastoma, cellule corticali primarie ed ex vivo in mitocondri isolati e sinaptosomi. Questo incremento in ATP e dovuto all'aumento in numero di MERCS e trasfer di Ca2+ tra i due organelli. Abbiamo testato la luteolina in cellue derivanti dal modello murino e c.elegans della malattia di Huntington, riportando miglioramenti nelle respirazione mitochondriale in vitro e miglioramento nel movimento in vivo.In sintesi, questa tesi ha contribuito a sviluppare la conoscienza del ruolo dei mitochondri e MERCS nelle sinapsi e nei meccanimsi esocitotici, nella disfunzione precoce nei modelli tau e beta amilodide di questi organelli. Abbiamo inoltre fornito una piattaforma per lo studio di farmaci in cellule neuronali con la scoperta della luteolina come promettente mediatore di potenziamento funzione mitocondriale.
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