Luana Naia1, Catarina M Pinho1, Giacomo Dentoni1, Jianping Liu2, Nuno Santos Leal1, Duarte M S Ferreira3, Bernadette Schreiner1, Riccardo Filadi4,5, Lígia Fão6, Niamh M C Connolly7, Pontus Forsell8, Gunnar Nordvall8, Makoto Shimozawa1, Elisa Greotti4,5, Emy Basso4,5, Pierre Theurey4, Anna Gioran9, Alvin Joselin10, Marie Arsenian-Henriksson11, Per Nilsson1, A Cristina Rego6,12, Jorge L Ruas3, David Park10, Daniele Bano9, Paola Pizzo4,5, Jochen H M Prehn7, Maria Ankarcrona13. 1. Center for Alzheimer Research, Division of Neurogeriatrics, Department of Neurobiology Care Sciences and Society, Karolinska Institutet, Stockholm, Sweden. 2. Department of Medicine-Huddinge, Karolinska Institutet, Stockholm, Sweden. 3. Department of Physiology and Pharmacology, Karolinska Institutet, Stockholm, Sweden. 4. Department of Biomedical Sciences, University of Padua, Padua, Italy. 5. Neuroscience Institute, National Research Council (CNR), 35131, Padua, Italy. 6. CNC-Center for Neuroscience and Cell Biology, University of Coimbra, Coimbra, Portugal. 7. Royal College of Surgeons in Ireland, Department of Physiology & Medical Physics Department, Dublin, Ireland. 8. AlzeCure Pharma AB, Huddinge, Sweden. 9. German Center for Neurodegenerative Diseases (DZNE), Bonn, Germany. 10. Department of Clinical Neurosciences, Hotchkiss Brain Institute, Cumming School of Medicine, University of Calgary, Calgary, Canada. 11. Department of Microbiology, Tumor and Cell Biology, Karolinska Institutet, Stockholm, Sweden. 12. Faculty of Medicine, Institute of Biochemistry, University of Coimbra, Coimbra, Portugal. 13. Center for Alzheimer Research, Division of Neurogeriatrics, Department of Neurobiology Care Sciences and Society, Karolinska Institutet, Stockholm, Sweden. maria.ankarcrona@ki.se.
Abstract
BACKGROUND: Mitochondrial dysfunction is a common feature of aging, neurodegeneration, and metabolic diseases. Hence, mitotherapeutics may be valuable disease modifiers for a large number of conditions. In this study, we have set up a large-scale screening platform for mitochondrial-based modulators with promising therapeutic potential. RESULTS: Using differentiated human neuroblastoma cells, we screened 1200 FDA-approved compounds and identified 61 molecules that significantly increased cellular ATP without any cytotoxic effect. Following dose response curve-dependent selection, we identified the flavonoid luteolin as a primary hit. Further validation in neuronal models indicated that luteolin increased mitochondrial respiration in primary neurons, despite not affecting mitochondrial mass, structure, or mitochondria-derived reactive oxygen species. However, we found that luteolin increased contacts between mitochondria and endoplasmic reticulum (ER), contributing to increased mitochondrial calcium (Ca2+) and Ca2+-dependent pyruvate dehydrogenase activity. This signaling pathway likely contributed to the observed effect of luteolin on enhanced mitochondrial complexes I and II activities. Importantly, we observed that increased mitochondrial functions were dependent on the activity of ER Ca2+-releasing channels inositol 1,4,5-trisphosphate receptors (IP3Rs) both in neurons and in isolated synaptosomes. Additionally, luteolin treatment improved mitochondrial and locomotory activities in primary neurons and Caenorhabditis elegans expressing an expanded polyglutamine tract of the huntingtin protein. CONCLUSION: We provide a new screening platform for drug discovery validated in vitro and ex vivo. In addition, we describe a novel mechanism through which luteolin modulates mitochondrial activity in neuronal models with potential therapeutic validity for treatment of a variety of human diseases.
BACKGROUND:Mitochondrial dysfunction is a common feature of aging, neurodegeneration, and metabolic diseases. Hence, mitotherapeutics may be valuable disease modifiers for a large number of conditions. In this study, we have set up a large-scale screening platform for mitochondrial-based modulators with promising therapeutic potential. RESULTS: Using differentiated humanneuroblastoma cells, we screened 1200 FDA-approved compounds and identified 61 molecules that significantly increased cellular ATP without any cytotoxic effect. Following dose response curve-dependent selection, we identified the flavonoidluteolin as a primary hit. Further validation in neuronal models indicated that luteolin increased mitochondrial respiration in primary neurons, despite not affecting mitochondrial mass, structure, or mitochondria-derived reactive oxygen species. However, we found that luteolin increased contacts between mitochondria and endoplasmic reticulum (ER), contributing to increased mitochondrial calcium (Ca2+) and Ca2+-dependent pyruvate dehydrogenase activity. This signaling pathway likely contributed to the observed effect of luteolin on enhanced mitochondrial complexes I and II activities. Importantly, we observed that increased mitochondrial functions were dependent on the activity of ERCa2+-releasing channels inositol 1,4,5-trisphosphate receptors (IP3Rs) both in neurons and in isolated synaptosomes. Additionally, luteolin treatment improved mitochondrial and locomotory activities in primary neurons and Caenorhabditis elegans expressing an expanded polyglutamine tract of the huntingtin protein. CONCLUSION: We provide a new screening platform for drug discovery validated in vitro and ex vivo. In addition, we describe a novel mechanism through which luteolin modulates mitochondrial activity in neuronal models with potential therapeutic validity for treatment of a variety of human diseases.
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