Proteomic Characterization of Human Neural Stem Cells and Their Secretome During in vitro Differentiation
Cell therapies represent a promising approach to slow down the progression of currently untreatable neurodegenerative diseases (e.g., Alzheimer's and Parkinson's disease or amyotrophic lateral sclerosis), as well as to support the reconstruction of functional neural circuits after spinal c...
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Published in | Frontiers in cellular neuroscience Vol. 14; p. 612560 |
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Main Authors | , , , , , , , , , , |
Format | Journal Article |
Language | English |
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28.01.2021
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Abstract | Cell therapies represent a promising approach to slow down the progression of currently untreatable neurodegenerative diseases (e.g., Alzheimer's and Parkinson's disease or amyotrophic lateral sclerosis), as well as to support the reconstruction of functional neural circuits after spinal cord injuries. In such therapies, the grafted cells could either functionally integrate into the damaged tissue, partially replacing dead or damaged cells, modulate inflammatory reaction, reduce tissue damage, or support neuronal survival by secretion of cytokines, growth, and trophic factors. Comprehensive characterization of cells and their proliferative potential, differentiation status, and population purity before transplantation is crucial to preventing safety risks, e.g., a tumorous growth due to the proliferation of undifferentiated stem cells. We characterized changes in the proteome and secretome of human neural stem cells (NSCs) during their spontaneous (EGF/FGF2 withdrawal) differentiation and differentiation with trophic support by BDNF/GDNF supplementation. We used LC-MS/MS in SWATH-MS mode for global cellular proteome profiling and quantified almost three thousand cellular proteins. Our analysis identified substantial protein differences in the early stages of NSC differentiation with more than a third of all the proteins regulated (including known neuronal and NSC multipotency markers) and revealed that the BDNF/GDNF support affected more the later stages of the NSC differentiation. Among the pathways identified as activated during both spontaneous and BDNF/GDNF differentiation were the HIF-1 signaling pathway, Wnt signaling pathway, and VEGF signaling pathway. Our follow-up secretome analysis using Luminex multiplex immunoassay revealed significant changes in the secretion of VEGF and IL-6 during NSC differentiation. Our results further demonstrated an increased expression of neuropilin-1 as well as catenin β-1, both known to participate in the regulation of VEGF signaling, and showed that VEGF-A isoform 121 (VEGF121), in particular, induces proliferation and supports survival of differentiating cells. |
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AbstractList | Cell therapies represent a promising approach to slow down the progression of currently untreatable neurodegenerative diseases (e.g., Alzheimer's and Parkinson's disease or amyotrophic lateral sclerosis), as well as to support the reconstruction of functional neural circuits after spinal cord injuries. In such therapies, the grafted cells could either functionally integrate into the damaged tissue, partially replacing dead or damaged cells, modulate inflammatory reaction, reduce tissue damage, or support neuronal survival by secretion of cytokines, growth, and trophic factors. Comprehensive characterization of cells and their proliferative potential, differentiation status, and population purity before transplantation is crucial to preventing safety risks, e.g., a tumorous growth due to the proliferation of undifferentiated stem cells. We characterized changes in the proteome and secretome of human neural stem cells (NSCs) during their spontaneous (EGF/FGF2 withdrawal) differentiation and differentiation with trophic support by BDNF/GDNF supplementation. We used LC-MS/MS in SWATH-MS mode for global cellular proteome profiling and quantified almost three thousand cellular proteins. Our analysis identified substantial protein differences in the early stages of NSC differentiation with more than a third of all the proteins regulated (including known neuronal and NSC multipotency markers) and revealed that the BDNF/GDNF support affected more the later stages of the NSC differentiation. Among the pathways identified as activated during both spontaneous and BDNF/GDNF differentiation were the HIF-1 signaling pathway, Wnt signaling pathway, and VEGF signaling pathway. Our follow-up secretome analysis using Luminex multiplex immunoassay revealed significant changes in the secretion of VEGF and IL-6 during NSC differentiation. Our results further demonstrated an increased expression of neuropilin-1 as well as catenin β-1, both known to participate in the regulation of VEGF signaling, and showed that VEGF-A isoform 121 (VEGF121), in particular, induces proliferation and supports survival of differentiating cells. Cell therapies represent a promising approach to slow down the progression of currently untreatable neurodegenerative diseases (e.g., Alzheimer's and Parkinson's disease or amyotrophic lateral sclerosis), as well as to support the reconstruction of functional neural circuits after spinal cord injuries. In such therapies, the grafted cells could either functionally integrate into the damaged tissue, partially replacing dead or damaged cells, modulate inflammatory reaction, reduce tissue damage and support neuronal survival by secretion of cytokines, growth, and trophic factors. Comprehensive characterization of cells and their proliferative potential, differentiation status and population purity before transplantation is crucial to prevent safety risks, e.g., a tumorous growth due to the proliferation of undifferentiated stem cells. We characterized changes in proteome and secretome of human neural stem cells (NSCs) during their spontaneous (EGF/FGF2 withdrawal) differentiation and differentiation with trophic support by BDNF/GDNF supplementation. We used LC-MS/MS in SWATH-MS mode for global cellular proteome profiling and quantified almost three thousand cellular proteins. Our analysis identified substantial proteins differences in the early stages of NSC differentiation with more than a third of all the proteins regulated (including known neuronal and NSC multipotency markers) and revealed that the BDNF/GDNF support affected more the later stages of the NSC differentiation. Among the pathways identified as activated during both spontaneous and BDNF/GDNF differentiation were HIF-1 signaling pathway, Wnt signaling pathway and VEGF signaling pathway. Our follow-up secretome analysis using Luminex multiplex immunoassay revealed significant changes in the secretion of VEGF and IL-6 during NSC differentiation. Our results further demonstrated an increased expression of Neuropilin-1 as well as Catenin β-1, both known to participate in regulation of VEGF signaling, and showed that VEGF-A isoform 121 (VEGF121), in particular, induces proliferation and supports survival of differentiating cells. |
Author | Kupcová Skalníková, Helena Kolář, Michal Červenka, Jakub Vaškovičová, Michaela Pánková, Tereza Valeková, Ivona Tylečková, Jiřina Vodičková Kepková, Kateřina Pfeiferová, Lucie Vodička, Petr Poliakh, Ievgeniia |
AuthorAffiliation | 3 Laboratory of Cell Regeneration and Plasticity, Research Center PIGMOD, Institute of Animal Physiology and Genetics of the Czech Academy of Sciences , Liběchov , Czechia 2 Department of Cell Biology, Faculty of Science, Charles University , Prague , Czechia 5 Department of Informatics and Chemistry, Faculty of Chemical Technology, University of Chemistry and Technology , Prague , Czechia 1 Laboratory of Applied Proteome Analyses, Research Center PIGMOD, Institute of Animal Physiology and Genetics of the Czech Academy of Sciences , Liběchov , Czechia 4 Laboratory of Genomics and Bioinformatics, Institute of Molecular Genetics of the Czech Academy of Sciences , Prague , Czechia 6 Laboratory of DNA Integrity, Research Center PIGMOD, Institute of Animal Physiology and Genetics of the Czech Academy of Sciences , Liběchov , Czechia |
AuthorAffiliation_xml | – name: 6 Laboratory of DNA Integrity, Research Center PIGMOD, Institute of Animal Physiology and Genetics of the Czech Academy of Sciences , Liběchov , Czechia – name: 4 Laboratory of Genomics and Bioinformatics, Institute of Molecular Genetics of the Czech Academy of Sciences , Prague , Czechia – name: 2 Department of Cell Biology, Faculty of Science, Charles University , Prague , Czechia – name: 1 Laboratory of Applied Proteome Analyses, Research Center PIGMOD, Institute of Animal Physiology and Genetics of the Czech Academy of Sciences , Liběchov , Czechia – name: 3 Laboratory of Cell Regeneration and Plasticity, Research Center PIGMOD, Institute of Animal Physiology and Genetics of the Czech Academy of Sciences , Liběchov , Czechia – name: 5 Department of Informatics and Chemistry, Faculty of Chemical Technology, University of Chemistry and Technology , Prague , Czechia |
Author_xml | – sequence: 1 givenname: Jakub surname: Červenka fullname: Červenka, Jakub organization: Department of Cell Biology, Faculty of Science, Charles University, Prague, Czechia – sequence: 2 givenname: Jiřina surname: Tylečková fullname: Tylečková, Jiřina organization: Laboratory of Applied Proteome Analyses, Research Center PIGMOD, Institute of Animal Physiology and Genetics of the Czech Academy of Sciences, Liběchov, Czechia – sequence: 3 givenname: Helena surname: Kupcová Skalníková fullname: Kupcová Skalníková, Helena organization: Laboratory of Applied Proteome Analyses, Research Center PIGMOD, Institute of Animal Physiology and Genetics of the Czech Academy of Sciences, Liběchov, Czechia – sequence: 4 givenname: Kateřina surname: Vodičková Kepková fullname: Vodičková Kepková, Kateřina organization: Laboratory of Applied Proteome Analyses, Research Center PIGMOD, Institute of Animal Physiology and Genetics of the Czech Academy of Sciences, Liběchov, Czechia – sequence: 5 givenname: Ievgeniia surname: Poliakh fullname: Poliakh, Ievgeniia organization: Department of Cell Biology, Faculty of Science, Charles University, Prague, Czechia – sequence: 6 givenname: Ivona surname: Valeková fullname: Valeková, Ivona organization: Laboratory of Cell Regeneration and Plasticity, Research Center PIGMOD, Institute of Animal Physiology and Genetics of the Czech Academy of Sciences, Liběchov, Czechia – sequence: 7 givenname: Lucie surname: Pfeiferová fullname: Pfeiferová, Lucie organization: Department of Informatics and Chemistry, Faculty of Chemical Technology, University of Chemistry and Technology, Prague, Czechia – sequence: 8 givenname: Michal surname: Kolář fullname: Kolář, Michal organization: Laboratory of Genomics and Bioinformatics, Institute of Molecular Genetics of the Czech Academy of Sciences, Prague, Czechia – sequence: 9 givenname: Michaela surname: Vaškovičová fullname: Vaškovičová, Michaela organization: Laboratory of DNA Integrity, Research Center PIGMOD, Institute of Animal Physiology and Genetics of the Czech Academy of Sciences, Liběchov, Czechia – sequence: 10 givenname: Tereza surname: Pánková fullname: Pánková, Tereza organization: Department of Cell Biology, Faculty of Science, Charles University, Prague, Czechia – sequence: 11 givenname: Petr surname: Vodička fullname: Vodička, Petr organization: Laboratory of Applied Proteome Analyses, Research Center PIGMOD, Institute of Animal Physiology and Genetics of the Czech Academy of Sciences, Liběchov, Czechia |
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Copyright | Copyright © 2021 Červenka, Tylečková, Kupcová Skalníková, Vodičková Kepková, Poliakh, Valeková, Pfeiferová, Kolář, Vaškovičová, Pánková and Vodička. 2021. This work is licensed under http://creativecommons.org/licenses/by/4.0/ (the “License”). Notwithstanding the ProQuest Terms and Conditions, you may use this content in accordance with the terms of the License. Copyright © 2021 Červenka, Tylečková, Kupcová Skalníková, Vodičková Kepková, Poliakh, Valeková, Pfeiferová, Kolář, Vaškovičová, Pánková and Vodička. 2021 Červenka, Tylečková, Kupcová Skalníková, Vodičková Kepková, Poliakh, Valeková, Pfeiferová, Kolář, Vaškovičová, Pánková and Vodička |
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Keywords | neural stem cell proliferation VEGF SWATH-MS neural differentiation secretome proteome |
Language | English |
License | Copyright © 2021 Červenka, Tylečková, Kupcová Skalníková, Vodičková Kepková, Poliakh, Valeková, Pfeiferová, Kolář, Vaškovičová, Pánková and Vodička. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms. |
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Notes | ObjectType-Article-1 SourceType-Scholarly Journals-1 ObjectType-Feature-2 content type line 23 Reviewed by: Nunzio Vicario, University of Catania, Italy; Hugo Guerrero-Cazares, Mayo Clinic, United States This article was submitted to Cellular Neuropathology, a section of the journal Frontiers in Cellular Neuroscience Edited by: Antonio Salgado, University of Minho, Portugal |
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SubjectTerms | Alzheimer's disease Amyotrophic lateral sclerosis Brain-derived neurotrophic factor Cell culture Cell differentiation Cell growth Cell proliferation Cellular Neuroscience Epidermal growth factor Fibroblast growth factor 2 Functional morphology Glial cell line-derived neurotrophic factor Inflammation Injuries Interleukin 6 Movement disorders Nervous system neural differentiation Neural networks neural stem cell Neural stem cells Neurodegenerative diseases Neuropilin Parkinson's disease proliferation Proteins proteome Proteomes Secretome Signal transduction Stem cells Systems development Transplantation Trophic factors Vascular endothelial growth factor VEGF |
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Title | Proteomic Characterization of Human Neural Stem Cells and Their Secretome During in vitro Differentiation |
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