Inhibitors of p53 and HIF-prolyl-4-hydroxylases provide mitochondrial protection in a model of oxytosis

Mitochondrial dysfunction and demise are hallmarks of many neurological disorders and neurodegenerative diseases. Since mitochondria are the key organelles providing energy, they are regarded as the power house of the cell. Under conditions of stress, mitochondria regulate the ‘point of no return’ in intrinsic cell death pathways, which marks the decision point between life and death. Once the mitochondria are irretrievably damaged, they drive the cell into death. Mitochondrial protection displays a promising strategy to protect neurons from death. Previously, the two proteins p53 and PHD1 have been associated with neurodegeneration. Hence, the aim of this study was to investigate the role of those two proteins in neuronal cell death and to evaluate the neuroprotective potential of their particular inhibitors PFTα and DFO, DHB, CPO and oxyquinoline, respectively. Additionally, underlying mechanisms should be elucidated by which these inhibitors mediate their neuroprotective effects. In this study, immortalised mouse hippocampal HT-22 cells were used since they represent an established model of caspase-independent cell death induced by glutamate, termed oxytosis. Moreover, erastin was applied in the same cell line to induce a mode of cell death called ferroptosis. The first part of this study revealed that siRNA-mediated knockdown of p53 delayed glutamate-induced cell death in HT-22 cells for about 2 h, but failed to prevent lipid peroxidation or mitochondrial damage depicted as enhanced mitochondrial fragmentation, depolarisation of the mitochondrial membrane, increased mitochondrial ROS formation and a loss of ATP levels. Both p53 and phospho-p53 did not translocate to the mitochondria upon glutamate challenge indicating that oxytosis was not attributed to a direct action of p53 at the level of mitochondria. The inhibition of p53 transcriptional activity by knockdown of p53, which was determined by a reporter assay established in this work, could serve as a possible explanation for the observed delay of cell death. In contrast, the pharmacological p53-inhibitor PFTα prevented glutamate-induced cell death of HT-22 cells more efficiently and was still able to rescue these cells when applied up to 4 h after the onset of glutamate treatment. Furthermore, PFTα abolished lipid peroxidation and subsequently preserved mitochondrial integrity which was indicated by reduced mitochondrial fission, attenuated formation of mitochondrial ROS and restored mitochondrial membrane potential and ATP levels. Notably, PFTα rescued HT-22 cells depleted of p53 from glutamate-induced cell death. These results exposed a pronounced neuroprotective potential of PFTα which occurred at the level of mitochondria and independently of p53. The second part of this thesis demonstrated the neuroprotective potential of PHD inhibition by the use of structural diverse PHD-inhibitors and siRNAs selectively targeting PHD1. Both concepts of PHD inhibition reduced generation of lipid peroxides and preserved mitochondrial morphology and function indicated by restored mitochondrial respiration and membrane potential and abolished mitochondrial ROS formation, revealing that PHD inhibition acts upstream of mitochondrial demise. Remarkably, the effects by siRNA-mediated PHD1 silencing were less pronounced than those achieved by pharmacological inhibitors. These differences in efficacy were likely attributed to the insufficient knockdown by the siRNA approach. Nevertheless, these findings exposed the selective inhibition of PHD1 and the broad pharmacological inhibition of the PHD enzyme family as promising strategies to achieve mitochondrial rescue and subsequent neuroprotection. Previously, PHDs have been shown to interact with the transcription factor ATF4. The present study revealed that oxyquinoline was able to prevent the glutamate-induced down regulation of ATF4. However, oxyquinoline was still able to prevent oxytosis in cells depleted of ATF4. Therefore, the observed regulation of ATF4 protein levels after oxyquinoline application emerged as dispensable for oxyquinoline mediated protection in oxytosis, although previous studies in vivo suggested a modification of ATF4 transcriptional activity as mode of action for oxyquinoline. Overall, the exact mechanism by which PHD inhibition and, in particular oxyquinoline induced neuroprotection in the paradigm of oxytosis remains elusive so far. Finally, PHD inhibition was also shown to protect HT-22 cells against erastin-induced ferroptosis further supporting the pivotal role of PHDs in neuronal demise and the potential of PHD inhibition as a promising therapeutic strategy in the treatment of neurodegenerative diseases, where oxidative stress contributes to progressive mitochondrial dysfunction and neuronal death

Nicotin-analoge Strukturen : biologische Bedeutung, Synthese und Anwendung in der organischen Chemie

[no abstract

Can neural quantum states learn volume-law ground states?

We study whether neural quantum states based on multi-layer feed-forward networks can find ground states which exhibit volume-law entanglement entropy. As a testbed, we employ the paradigmatic Sachdev-Ye-Kitaev model. We find that both shallow and deep feed-forward networks require an exponential number of parameters in order to represent the ground state of this model. This demonstrates that sufficiently complicated quantum states, although being physical solutions to relevant models and not pathological cases, can still be difficult to learn to the point of intractability at larger system sizes. This highlights the importance of further investigations into the physical properties of quantum states amenable to an efficient neural representation

Mitochondrial damage by α-synuclein causes cell death in human dopaminergic neurons

Evolving concepts on Parkinson's disease (PD) pathology suggest that α-synuclein (aSYN) promote dopaminergic neuron dysfunction and death through accumulating in the mitochondria. However, the consequence of mitochondrial aSYN localisation on mitochondrial structure and bioenergetic functions in neuronal cells are poorly understood. Therefore, we investigated deleterious effects of mitochondria-targeted aSYN in differentiated human dopaminergic neurons in comparison with wild-type (WT) aSYN overexpression and corresponding EGFP (enhanced green fluorescent protein)-expressing controls. Mitochondria-targeted aSYN enhanced mitochondrial reactive oxygen species (ROS) formation, reduced ATP levels and showed severely disrupted structure and function of the dendritic neural network, preceding neuronal death. Transmission electron microscopy illustrated distorted cristae and many fragmented mitochondria in response to WT-aSYN overexpression, and a complete loss of cristae structure and massively swollen mitochondria in neurons expressing mitochondria-targeted aSYN. Further, the analysis of mitochondrial bioenergetics in differentiated dopaminergic neurons, expressing WT or mitochondria-targeted aSYN, elicited a pronounced impairment of mitochondrial respiration. In a pharmacological compound screening, we found that the pan-caspase inhibitors QVD and zVAD-FMK, and a specific caspase-1 inhibitor significantly prevented aSYN-induced cell death. In addition, the caspase inhibitor QVD preserved mitochondrial function and neuronal network activity in the human dopaminergic neurons overexpressing aSYN. Overall, our findings indicated therapeutic effects by caspase-1 inhibition despite aSYN-mediated alterations in mitochondrial morphology and function

Performance Increase in Sales

Die Arbeit beschäftigt sich mit der Leistung von Versicherungsvermittlern in der Ausschließlichkeitsorganisation. Dabei wird untersucht, welche Faktoren die Verkaufsleistung von Versicherungsvermittlern beeinflussen. Zunächst werden die Faktoren untersucht, die den Vermittler selbst betreffen. Anschließend wird geprüft welchen Einfluß die Führungskräfte und die Struktur der Ausschließlichkeitsorganisation auf die Leistung der Vermittler haben. Aus diesen Erkenntnissen werden Schlussfolgerungen gezogen, welche Rahmenbedingungen ein Versicherungsunternehmen schaffen muss, um eine Leistungssteigerung im Vertrieb zu erreichen

Inhibitors of p53 and HIF-prolyl-4-hydroxylases provide mitochondrial protection in a model of oxytosis

Mitochondrial dysfunction and demise are hallmarks of many neurological disorders and neurodegenerative diseases. Since mitochondria are the key organelles providing energy, they are regarded as the power house of the cell. Under conditions of stress, mitochondria regulate the ‘point of no return’ in intrinsic cell death pathways, which marks the decision point between life and death. Once the mitochondria are irretrievably damaged, they drive the cell into death. Mitochondrial protection displays a promising strategy to protect neurons from death. Previously, the two proteins p53 and PHD1 have been associated with neurodegeneration. Hence, the aim of this study was to investigate the role of those two proteins in neuronal cell death and to evaluate the neuroprotective potential of their particular inhibitors PFTα and DFO, DHB, CPO and oxyquinoline, respectively. Additionally, underlying mechanisms should be elucidated by which these inhibitors mediate their neuroprotective effects. In this study, immortalised mouse hippocampal HT-22 cells were used since they represent an established model of caspase-independent cell death induced by glutamate, termed oxytosis. Moreover, erastin was applied in the same cell line to induce a mode of cell death called ferroptosis. The first part of this study revealed that siRNA-mediated knockdown of p53 delayed glutamate-induced cell death in HT-22 cells for about 2 h, but failed to prevent lipid peroxidation or mitochondrial damage depicted as enhanced mitochondrial fragmentation, depolarisation of the mitochondrial membrane, increased mitochondrial ROS formation and a loss of ATP levels. Both p53 and phospho-p53 did not translocate to the mitochondria upon glutamate challenge indicating that oxytosis was not attributed to a direct action of p53 at the level of mitochondria. The inhibition of p53 transcriptional activity by knockdown of p53, which was determined by a reporter assay established in this work, could serve as a possible explanation for the observed delay of cell death. In contrast, the pharmacological p53-inhibitor PFTα prevented glutamate-induced cell death of HT-22 cells more efficiently and was still able to rescue these cells when applied up to 4 h after the onset of glutamate treatment. Furthermore, PFTα abolished lipid peroxidation and subsequently preserved mitochondrial integrity which was indicated by reduced mitochondrial fission, attenuated formation of mitochondrial ROS and restored mitochondrial membrane potential and ATP levels. Notably, PFTα rescued HT-22 cells depleted of p53 from glutamate-induced cell death. These results exposed a pronounced neuroprotective potential of PFTα which occurred at the level of mitochondria and independently of p53. The second part of this thesis demonstrated the neuroprotective potential of PHD inhibition by the use of structural diverse PHD-inhibitors and siRNAs selectively targeting PHD1. Both concepts of PHD inhibition reduced generation of lipid peroxides and preserved mitochondrial morphology and function indicated by restored mitochondrial respiration and membrane potential and abolished mitochondrial ROS formation, revealing that PHD inhibition acts upstream of mitochondrial demise. Remarkably, the effects by siRNA-mediated PHD1 silencing were less pronounced than those achieved by pharmacological inhibitors. These differences in efficacy were likely attributed to the insufficient knockdown by the siRNA approach. Nevertheless, these findings exposed the selective inhibition of PHD1 and the broad pharmacological inhibition of the PHD enzyme family as promising strategies to achieve mitochondrial rescue and subsequent neuroprotection. Previously, PHDs have been shown to interact with the transcription factor ATF4. The present study revealed that oxyquinoline was able to prevent the glutamate-induced down regulation of ATF4. However, oxyquinoline was still able to prevent oxytosis in cells depleted of ATF4. Therefore, the observed regulation of ATF4 protein levels after oxyquinoline application emerged as dispensable for oxyquinoline mediated protection in oxytosis, although previous studies in vivo suggested a modification of ATF4 transcriptional activity as mode of action for oxyquinoline. Overall, the exact mechanism by which PHD inhibition and, in particular oxyquinoline induced neuroprotection in the paradigm of oxytosis remains elusive so far. Finally, PHD inhibition was also shown to protect HT-22 cells against erastin-induced ferroptosis further supporting the pivotal role of PHDs in neuronal demise and the potential of PHD inhibition as a promising therapeutic strategy in the treatment of neurodegenerative diseases, where oxidative stress contributes to progressive mitochondrial dysfunction and neuronal death