Neural stem cells-derived neurons obtained from human pluripotent stem cells as an in vitro model for studying CDK5/p25 mediated neurodegenerative processes

Autores
Mucci, Sofia; Rodríguez Varela, Maria Soledad; Isaja, Luciana; Sevlever, Gustavo Emilio; Scassa, Maria Elida; Romorini, Leonardo
Año de publicación
2020
Idioma
inglés
Tipo de recurso
documento de conferencia
Estado
versión publicada
Descripción
Human embryonic and induced pluripotent stem cells (hESCs and IPSCs)can differentiate into a wide range of specialized cells,including neural stem cells (NSC). Moreover, NSC-derived neurons are proposed as a model for studying neurodegeneration. CDK5/ p35 complex is involved in neuronal homeostasis and development. However, its functionis deregulated in neurodegeneration by p35 cleavage in p25, which allows the aberrant phosphorylation of targets through the constitution of a more stable complex CDK5/p25. In this work we aimed to set up an in vitroCDK5/p25 neurodegenerative model using NSC-derived neurons obtained from hESCs and hIPSCs subjected to different cellular stressors. For this purpose, we first derived NSC from hESC (H9 line) and hIPSC (FN 2.1 line), which were further differentiated into neuronsusin a neural induction kit. NSC and neuron-like phenotype were validated by immunofluoresce and RT-qPCR of cell specific markers (sox-1, Sox-2, pax-6 and nestin for NSC; MAP5, MAP2 and Tuj-1 for neurons). Then, CDK5 and p35 mRNA and protein expression levels were analyzed in hESCs, hIPSCs, NSC and neurons by RT-qPCR and western blot. Interestingly, we observed that althought CDK5 was ubiquitous, p35 mRNA and protein were mainly expressed in neurons. We next evaluated how different stress stimuli (rotenone, glutamate and calcium ionophore A23187) affected NSC andneurons viability. We determined the percentage of cell viability after 24 hours treatment with increasing concentrations of rotenone and glutamate and 2 hours treatment with A23187 using an XTT vital dye assay. Cell viability felt downsignificantly in the case of rotenone and A23187 treatments in a concetration dependent manner. In conclusion, NSC-derived neurons obtained from hESCs andhiPSCs expressed high levels of p35 and responded to rotenone stressor, making them a suitable CDK5/p25 neurodegenerative invitro model.
Fil: Mucci, Sofia. Fundación para la Lucha contra las Enfermedades Neurológicas de la Infancia; Argentina
Fil: Rodríguez Varela, Maria Soledad. Fundación para la Lucha contra las Enfermedades Neurológicas de la Infancia. Instituto de Neurociencias - Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Neurociencias; Argentina
Fil: Isaja, Luciana. Fundación para la Lucha contra las Enfermedades Neurológicas de la Infancia. Instituto de Neurociencias - Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Neurociencias; Argentina
Fil: Sevlever, Gustavo Emilio. Fundación para la Lucha contra las Enfermedades Neurológicas de la Infancia; Argentina
Fil: Scassa, Maria Elida. Fundación para la Lucha contra las Enfermedades Neurológicas de la Infancia; Argentina
Fil: Romorini, Leonardo. Fundación para la Lucha contra las Enfermedades Neurológicas de la Infancia. Instituto de Neurociencias - Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Neurociencias; Argentina
LXIII Reunión Anual de la de la Sociedad Argentina de Investigación Clínica; LXVI Reunión Anual de la Sociedad; Reunión Anual de la Sociedad Argentina de Fisiología
Mar del Plata
Argentina
Sociedad Argentina de Investigación Clínica
Sociedad Argentina de Inmunología
Sociedad Argentina de Fisiología
Sociedad Argentina de Virología
Asociación Argentina de Naomedicinas
Materia
celulas madre neurales
neuronas
celulas madres pluripotentes inducidas
neurodegeneración
Nivel de accesibilidad
acceso abierto
Condiciones de uso
https://creativecommons.org/licenses/by-nc-sa/2.5/ar/
Repositorio
CONICET Digital (CONICET)
Institución
Consejo Nacional de Investigaciones Científicas y Técnicas
OAI Identificador
oai:ri.conicet.gov.ar:11336/263805
Acceder
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oai_identifier_str oai:ri.conicet.gov.ar:11336/263805
network_acronym_str CONICETDig
repository_id_str 3498
network_name_str CONICET Digital (CONICET)
spelling Neural stem cells-derived neurons obtained from human pluripotent stem cells as an in vitro model for studying CDK5/p25 mediated neurodegenerative processesMucci, SofiaRodríguez Varela, Maria SoledadIsaja, LucianaSevlever, Gustavo EmilioScassa, Maria ElidaRomorini, Leonardocelulas madre neuralesneuronascelulas madres pluripotentes inducidasneurodegeneraciónhttps://purl.org/becyt/ford/1.6https://purl.org/becyt/ford/1Human embryonic and induced pluripotent stem cells (hESCs and IPSCs)can differentiate into a wide range of specialized cells,including neural stem cells (NSC). Moreover, NSC-derived neurons are proposed as a model for studying neurodegeneration. CDK5/ p35 complex is involved in neuronal homeostasis and development. However, its functionis deregulated in neurodegeneration by p35 cleavage in p25, which allows the aberrant phosphorylation of targets through the constitution of a more stable complex CDK5/p25. In this work we aimed to set up an in vitroCDK5/p25 neurodegenerative model using NSC-derived neurons obtained from hESCs and hIPSCs subjected to different cellular stressors. For this purpose, we first derived NSC from hESC (H9 line) and hIPSC (FN 2.1 line), which were further differentiated into neuronsusin a neural induction kit. NSC and neuron-like phenotype were validated by immunofluoresce and RT-qPCR of cell specific markers (sox-1, Sox-2, pax-6 and nestin for NSC; MAP5, MAP2 and Tuj-1 for neurons). Then, CDK5 and p35 mRNA and protein expression levels were analyzed in hESCs, hIPSCs, NSC and neurons by RT-qPCR and western blot. Interestingly, we observed that althought CDK5 was ubiquitous, p35 mRNA and protein were mainly expressed in neurons. We next evaluated how different stress stimuli (rotenone, glutamate and calcium ionophore A23187) affected NSC andneurons viability. We determined the percentage of cell viability after 24 hours treatment with increasing concentrations of rotenone and glutamate and 2 hours treatment with A23187 using an XTT vital dye assay. Cell viability felt downsignificantly in the case of rotenone and A23187 treatments in a concetration dependent manner. In conclusion, NSC-derived neurons obtained from hESCs andhiPSCs expressed high levels of p35 and responded to rotenone stressor, making them a suitable CDK5/p25 neurodegenerative invitro model.Fil: Mucci, Sofia. Fundación para la Lucha contra las Enfermedades Neurológicas de la Infancia; ArgentinaFil: Rodríguez Varela, Maria Soledad. Fundación para la Lucha contra las Enfermedades Neurológicas de la Infancia. Instituto de Neurociencias - Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Neurociencias; ArgentinaFil: Isaja, Luciana. Fundación para la Lucha contra las Enfermedades Neurológicas de la Infancia. Instituto de Neurociencias - Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Neurociencias; ArgentinaFil: Sevlever, Gustavo Emilio. Fundación para la Lucha contra las Enfermedades Neurológicas de la Infancia; ArgentinaFil: Scassa, Maria Elida. Fundación para la Lucha contra las Enfermedades Neurológicas de la Infancia; ArgentinaFil: Romorini, Leonardo. Fundación para la Lucha contra las Enfermedades Neurológicas de la Infancia. Instituto de Neurociencias - Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Neurociencias; ArgentinaLXIII Reunión Anual de la de la Sociedad Argentina de Investigación Clínica; LXVI Reunión Anual de la Sociedad; Reunión Anual de la Sociedad Argentina de FisiologíaMar del PlataArgentinaSociedad Argentina de Investigación ClínicaSociedad Argentina de InmunologíaSociedad Argentina de FisiologíaSociedad Argentina de VirologíaAsociación Argentina de NaomedicinasFundación Revista Medicina2020info:eu-repo/semantics/publishedVersioninfo:eu-repo/semantics/conferenceObjectReuniónJournalhttp://purl.org/coar/resource_type/c_5794info:ar-repo/semantics/documentoDeConferenciaapplication/pdfapplication/pdfapplication/pdfhttp://hdl.handle.net/11336/263805Neural stem cells-derived neurons obtained from human pluripotent stem cells as an in vitro model for studying CDK5/p25 mediated neurodegenerative processes; LXIII Reunión Anual de la de la Sociedad Argentina de Investigación Clínica; LXVI Reunión Anual de la Sociedad; Reunión Anual de la Sociedad Argentina de Fisiología; Mar del Plata; Argentina; 2018; 158-1581669-9106CONICET DigitalCONICETenginfo:eu-repo/semantics/altIdentifier/url/https://www.saic.org.ar/revista-medicinaNacionalinfo:eu-repo/semantics/openAccesshttps://creativecommons.org/licenses/by-nc-sa/2.5/ar/reponame:CONICET Digital (CONICET)instname:Consejo Nacional de Investigaciones Científicas y Técnicas2025-09-03T09:48:14Zoai:ri.conicet.gov.ar:11336/263805instacron:CONICETInstitucionalhttp://ri.conicet.gov.ar/Organismo científico-tecnológicoNo correspondehttp://ri.conicet.gov.ar/oai/requestdasensio@conicet.gov.ar; lcarlino@conicet.gov.arArgentinaNo correspondeNo correspondeNo correspondeopendoar:34982025-09-03 09:48:14.254CONICET Digital (CONICET) - Consejo Nacional de Investigaciones Científicas y Técnicasfalse
dc.title.none.fl_str_mv Neural stem cells-derived neurons obtained from human pluripotent stem cells as an in vitro model for studying CDK5/p25 mediated neurodegenerative processes
title Neural stem cells-derived neurons obtained from human pluripotent stem cells as an in vitro model for studying CDK5/p25 mediated neurodegenerative processes
spellingShingle Neural stem cells-derived neurons obtained from human pluripotent stem cells as an in vitro model for studying CDK5/p25 mediated neurodegenerative processes
Mucci, Sofia
celulas madre neurales
neuronas
celulas madres pluripotentes inducidas
neurodegeneración
title_short Neural stem cells-derived neurons obtained from human pluripotent stem cells as an in vitro model for studying CDK5/p25 mediated neurodegenerative processes
title_full Neural stem cells-derived neurons obtained from human pluripotent stem cells as an in vitro model for studying CDK5/p25 mediated neurodegenerative processes
title_fullStr Neural stem cells-derived neurons obtained from human pluripotent stem cells as an in vitro model for studying CDK5/p25 mediated neurodegenerative processes
title_full_unstemmed Neural stem cells-derived neurons obtained from human pluripotent stem cells as an in vitro model for studying CDK5/p25 mediated neurodegenerative processes
title_sort Neural stem cells-derived neurons obtained from human pluripotent stem cells as an in vitro model for studying CDK5/p25 mediated neurodegenerative processes
dc.creator.none.fl_str_mv Mucci, Sofia
Rodríguez Varela, Maria Soledad
Isaja, Luciana
Sevlever, Gustavo Emilio
Scassa, Maria Elida
Romorini, Leonardo
author Mucci, Sofia
author_facet Mucci, Sofia
Rodríguez Varela, Maria Soledad
Isaja, Luciana
Sevlever, Gustavo Emilio
Scassa, Maria Elida
Romorini, Leonardo
author_role author
author2 Rodríguez Varela, Maria Soledad
Isaja, Luciana
Sevlever, Gustavo Emilio
Scassa, Maria Elida
Romorini, Leonardo
author2_role author
author
author
author
author
dc.subject.none.fl_str_mv celulas madre neurales
neuronas
celulas madres pluripotentes inducidas
neurodegeneración
topic celulas madre neurales
neuronas
celulas madres pluripotentes inducidas
neurodegeneración
purl_subject.fl_str_mv https://purl.org/becyt/ford/1.6
https://purl.org/becyt/ford/1
dc.description.none.fl_txt_mv Human embryonic and induced pluripotent stem cells (hESCs and IPSCs)can differentiate into a wide range of specialized cells,including neural stem cells (NSC). Moreover, NSC-derived neurons are proposed as a model for studying neurodegeneration. CDK5/ p35 complex is involved in neuronal homeostasis and development. However, its functionis deregulated in neurodegeneration by p35 cleavage in p25, which allows the aberrant phosphorylation of targets through the constitution of a more stable complex CDK5/p25. In this work we aimed to set up an in vitroCDK5/p25 neurodegenerative model using NSC-derived neurons obtained from hESCs and hIPSCs subjected to different cellular stressors. For this purpose, we first derived NSC from hESC (H9 line) and hIPSC (FN 2.1 line), which were further differentiated into neuronsusin a neural induction kit. NSC and neuron-like phenotype were validated by immunofluoresce and RT-qPCR of cell specific markers (sox-1, Sox-2, pax-6 and nestin for NSC; MAP5, MAP2 and Tuj-1 for neurons). Then, CDK5 and p35 mRNA and protein expression levels were analyzed in hESCs, hIPSCs, NSC and neurons by RT-qPCR and western blot. Interestingly, we observed that althought CDK5 was ubiquitous, p35 mRNA and protein were mainly expressed in neurons. We next evaluated how different stress stimuli (rotenone, glutamate and calcium ionophore A23187) affected NSC andneurons viability. We determined the percentage of cell viability after 24 hours treatment with increasing concentrations of rotenone and glutamate and 2 hours treatment with A23187 using an XTT vital dye assay. Cell viability felt downsignificantly in the case of rotenone and A23187 treatments in a concetration dependent manner. In conclusion, NSC-derived neurons obtained from hESCs andhiPSCs expressed high levels of p35 and responded to rotenone stressor, making them a suitable CDK5/p25 neurodegenerative invitro model.
Fil: Mucci, Sofia. Fundación para la Lucha contra las Enfermedades Neurológicas de la Infancia; Argentina
Fil: Rodríguez Varela, Maria Soledad. Fundación para la Lucha contra las Enfermedades Neurológicas de la Infancia. Instituto de Neurociencias - Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Neurociencias; Argentina
Fil: Isaja, Luciana. Fundación para la Lucha contra las Enfermedades Neurológicas de la Infancia. Instituto de Neurociencias - Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Neurociencias; Argentina
Fil: Sevlever, Gustavo Emilio. Fundación para la Lucha contra las Enfermedades Neurológicas de la Infancia; Argentina
Fil: Scassa, Maria Elida. Fundación para la Lucha contra las Enfermedades Neurológicas de la Infancia; Argentina
Fil: Romorini, Leonardo. Fundación para la Lucha contra las Enfermedades Neurológicas de la Infancia. Instituto de Neurociencias - Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Neurociencias; Argentina
LXIII Reunión Anual de la de la Sociedad Argentina de Investigación Clínica; LXVI Reunión Anual de la Sociedad; Reunión Anual de la Sociedad Argentina de Fisiología
Mar del Plata
Argentina
Sociedad Argentina de Investigación Clínica
Sociedad Argentina de Inmunología
Sociedad Argentina de Fisiología
Sociedad Argentina de Virología
Asociación Argentina de Naomedicinas
description Human embryonic and induced pluripotent stem cells (hESCs and IPSCs)can differentiate into a wide range of specialized cells,including neural stem cells (NSC). Moreover, NSC-derived neurons are proposed as a model for studying neurodegeneration. CDK5/ p35 complex is involved in neuronal homeostasis and development. However, its functionis deregulated in neurodegeneration by p35 cleavage in p25, which allows the aberrant phosphorylation of targets through the constitution of a more stable complex CDK5/p25. In this work we aimed to set up an in vitroCDK5/p25 neurodegenerative model using NSC-derived neurons obtained from hESCs and hIPSCs subjected to different cellular stressors. For this purpose, we first derived NSC from hESC (H9 line) and hIPSC (FN 2.1 line), which were further differentiated into neuronsusin a neural induction kit. NSC and neuron-like phenotype were validated by immunofluoresce and RT-qPCR of cell specific markers (sox-1, Sox-2, pax-6 and nestin for NSC; MAP5, MAP2 and Tuj-1 for neurons). Then, CDK5 and p35 mRNA and protein expression levels were analyzed in hESCs, hIPSCs, NSC and neurons by RT-qPCR and western blot. Interestingly, we observed that althought CDK5 was ubiquitous, p35 mRNA and protein were mainly expressed in neurons. We next evaluated how different stress stimuli (rotenone, glutamate and calcium ionophore A23187) affected NSC andneurons viability. We determined the percentage of cell viability after 24 hours treatment with increasing concentrations of rotenone and glutamate and 2 hours treatment with A23187 using an XTT vital dye assay. Cell viability felt downsignificantly in the case of rotenone and A23187 treatments in a concetration dependent manner. In conclusion, NSC-derived neurons obtained from hESCs andhiPSCs expressed high levels of p35 and responded to rotenone stressor, making them a suitable CDK5/p25 neurodegenerative invitro model.
publishDate 2020
dc.date.none.fl_str_mv 2020
dc.type.none.fl_str_mv info:eu-repo/semantics/publishedVersion
info:eu-repo/semantics/conferenceObject
Reunión
Journal
http://purl.org/coar/resource_type/c_5794
info:ar-repo/semantics/documentoDeConferencia
status_str publishedVersion
format conferenceObject
dc.identifier.none.fl_str_mv http://hdl.handle.net/11336/263805
Neural stem cells-derived neurons obtained from human pluripotent stem cells as an in vitro model for studying CDK5/p25 mediated neurodegenerative processes; LXIII Reunión Anual de la de la Sociedad Argentina de Investigación Clínica; LXVI Reunión Anual de la Sociedad; Reunión Anual de la Sociedad Argentina de Fisiología; Mar del Plata; Argentina; 2018; 158-158
1669-9106
CONICET Digital
CONICET
url http://hdl.handle.net/11336/263805
identifier_str_mv Neural stem cells-derived neurons obtained from human pluripotent stem cells as an in vitro model for studying CDK5/p25 mediated neurodegenerative processes; LXIII Reunión Anual de la de la Sociedad Argentina de Investigación Clínica; LXVI Reunión Anual de la Sociedad; Reunión Anual de la Sociedad Argentina de Fisiología; Mar del Plata; Argentina; 2018; 158-158
1669-9106
CONICET Digital
CONICET
dc.language.none.fl_str_mv eng
language eng
dc.relation.none.fl_str_mv info:eu-repo/semantics/altIdentifier/url/https://www.saic.org.ar/revista-medicina
dc.rights.none.fl_str_mv info:eu-repo/semantics/openAccess
https://creativecommons.org/licenses/by-nc-sa/2.5/ar/
eu_rights_str_mv openAccess
rights_invalid_str_mv https://creativecommons.org/licenses/by-nc-sa/2.5/ar/
dc.format.none.fl_str_mv application/pdf
application/pdf
application/pdf
dc.coverage.none.fl_str_mv Nacional
dc.publisher.none.fl_str_mv Fundación Revista Medicina
publisher.none.fl_str_mv Fundación Revista Medicina
dc.source.none.fl_str_mv reponame:CONICET Digital (CONICET)
instname:Consejo Nacional de Investigaciones Científicas y Técnicas
reponame_str CONICET Digital (CONICET)
collection CONICET Digital (CONICET)
instname_str Consejo Nacional de Investigaciones Científicas y Técnicas
repository.name.fl_str_mv CONICET Digital (CONICET) - Consejo Nacional de Investigaciones Científicas y Técnicas
repository.mail.fl_str_mv dasensio@conicet.gov.ar; lcarlino@conicet.gov.ar
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