Human Schwann cell senescence is not prevented by ectopic expression of human telomerase reverse transcriptase

Autores
Andersen, Natalia Denise; Kuo, Blanche; Piñero, Gonzalo Miguel; Ravelo, Kristine; Rai, Priyamvada; Monje, Paula
Año de publicación
2016
Idioma
inglés
Tipo de recurso
documento de conferencia
Estado
versión publicada
Descripción
Isolated human Schwann cells (hSCs) typically become senescent and unresponsive to mitogenic factors with continued in vitro expansion. Data from our lab has shown that adult nerve-derived hSCs, in contrast to rodent (rat) SCs, inevitably stop proliferating and acquire a senescent phenotype characterized by high levels of senescence-associated (SA)-β galactosidase activity and morphological changes that include cell enlargement and appearance of multinucleated cells. As opposed to rodent SCs, hSC cultures consist of mixed populations of proliferating cells, senescent cells and cells at different stages of differentiation regardless of the nerve of origin and other donor-specific factors. RNA-seq analysis of representative cultures of hSCs did not reveal the presence of telomerase reverse transcriptase (TERT) mRNA while other TERT-related genes (e.g. TERF1, telomeric repeat binding factor, and TEP1, telomeraseassociated protein) were well-represented in the hSC transcriptome. In an attempt to overcome senescence, we used retroviral vectors and antibiotic selection to generate hSC lines ectopically expressing human (h)-TERT. For these experiments, highly proliferative, non-senescent, early passage hSC cultures were stably transduced with the retroviruses h-TERT-hygro or h-TERTpuro, each encoding the h-TERT gene along with hygromycin or puromycin resistance genes, respectively. Transduced hSC cultures from three different donors were selected and subjected to three rounds of expansion in medium containing chemical mitogens. Subsequently, the cultures were analyzed for their rate of proliferation by means of EdU incorporation assays and the acquisition of senescence by means of SA-β galactosidase activity assays in each round. We found that whereas ectopic h-TERT expression extended the lifespan of cultured hSCs when compared to non-infected or GFP-expressing cells, it was not sufficient to confer immortalization and overcome senescence. In sum, our results suggest that progression of the hSCs to a senescent state likely is stress-induced rather than dependent on replication-associated telomere shortening
Fil: Andersen, Natalia Denise. Miami University; Estados Unidos. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina
Fil: Kuo, Blanche. Miami University; Estados Unidos
Fil: Piñero, Gonzalo Miguel. Miami University; Estados Unidos. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina
Fil: Ravelo, Kristine. Miami University; Estados Unidos
Fil: Rai, Priyamvada. Miami University. School Of Medicine; Estados Unidos
Fil: Monje, Paula. Miami University. School Of Medicine; Estados Unidos
Society for Neuroscience 46th Annual Meeting
San Diego
Estados Unidos
Society for Neuroscience
Materia
SCHWANN CELLS
SENESCENCE
PROLIFERATION
RETROVIRAL VECTORS
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:11336/235392
Acceder
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oai_identifier_str oai:ri.conicet.gov.ar:11336/235392
network_acronym_str CONICETDig
repository_id_str 3498
network_name_str CONICET Digital (CONICET)
spelling Human Schwann cell senescence is not prevented by ectopic expression of human telomerase reverse transcriptaseAndersen, Natalia DeniseKuo, BlanchePiñero, Gonzalo MiguelRavelo, KristineRai, PriyamvadaMonje, PaulaSCHWANN CELLSSENESCENCEPROLIFERATIONRETROVIRAL VECTORShttps://purl.org/becyt/ford/1.6https://purl.org/becyt/ford/1Isolated human Schwann cells (hSCs) typically become senescent and unresponsive to mitogenic factors with continued in vitro expansion. Data from our lab has shown that adult nerve-derived hSCs, in contrast to rodent (rat) SCs, inevitably stop proliferating and acquire a senescent phenotype characterized by high levels of senescence-associated (SA)-β galactosidase activity and morphological changes that include cell enlargement and appearance of multinucleated cells. As opposed to rodent SCs, hSC cultures consist of mixed populations of proliferating cells, senescent cells and cells at different stages of differentiation regardless of the nerve of origin and other donor-specific factors. RNA-seq analysis of representative cultures of hSCs did not reveal the presence of telomerase reverse transcriptase (TERT) mRNA while other TERT-related genes (e.g. TERF1, telomeric repeat binding factor, and TEP1, telomeraseassociated protein) were well-represented in the hSC transcriptome. In an attempt to overcome senescence, we used retroviral vectors and antibiotic selection to generate hSC lines ectopically expressing human (h)-TERT. For these experiments, highly proliferative, non-senescent, early passage hSC cultures were stably transduced with the retroviruses h-TERT-hygro or h-TERTpuro, each encoding the h-TERT gene along with hygromycin or puromycin resistance genes, respectively. Transduced hSC cultures from three different donors were selected and subjected to three rounds of expansion in medium containing chemical mitogens. Subsequently, the cultures were analyzed for their rate of proliferation by means of EdU incorporation assays and the acquisition of senescence by means of SA-β galactosidase activity assays in each round. We found that whereas ectopic h-TERT expression extended the lifespan of cultured hSCs when compared to non-infected or GFP-expressing cells, it was not sufficient to confer immortalization and overcome senescence. In sum, our results suggest that progression of the hSCs to a senescent state likely is stress-induced rather than dependent on replication-associated telomere shorteningFil: Andersen, Natalia Denise. Miami University; Estados Unidos. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Kuo, Blanche. Miami University; Estados UnidosFil: Piñero, Gonzalo Miguel. Miami University; Estados Unidos. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Ravelo, Kristine. Miami University; Estados UnidosFil: Rai, Priyamvada. Miami University. School Of Medicine; Estados UnidosFil: Monje, Paula. Miami University. School Of Medicine; Estados UnidosSociety for Neuroscience 46th Annual MeetingSan DiegoEstados UnidosSociety for NeuroscienceSociety for Neuroscience2016info:eu-repo/semantics/publishedVersioninfo:eu-repo/semantics/conferenceObjectCongresoBookhttp://purl.org/coar/resource_type/c_5794info:ar-repo/semantics/documentoDeConferenciaapplication/pdfapplication/pdfapplication/pdfhttp://hdl.handle.net/11336/235392Human Schwann cell senescence is not prevented by ectopic expression of human telomerase reverse transcriptase; Society for Neuroscience 46th Annual Meeting; San Diego; Estados Unidos; 2016; 1-1CONICET DigitalCONICETenginfo:eu-repo/semantics/altIdentifier/url/https://www.sfn.org/Annual-Meeting/Neuroscience-2016/Sessions-and-Events/Program/~/media/SfN/Documents/Annual%20Meeting/FinalProgram/NS2016/Full%20Abstract%20PDFs%202016/SfN16_Abstract%20PDFs%20%20Posters_3_Mon_PM.ashxinfo:eu-repo/semantics/altIdentifier/url/https://www.sfn.org/meetings/past-and-future-annual-meetingsInternacionalinfo:eu-repo/semantics/openAccesshttps://creativecommons.org/licenses/by-nc-sa/2.5/ar/reponame:CONICET Digital (CONICET)instname:Consejo Nacional de Investigaciones Científicas y Técnicas2025-10-22T12:03:30Zoai:ri.conicet.gov.ar:11336/235392instacron: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:34982025-10-22 12:03:30.919CONICET Digital (CONICET) - Consejo Nacional de Investigaciones Científicas y Técnicasfalse
dc.title.none.fl_str_mv Human Schwann cell senescence is not prevented by ectopic expression of human telomerase reverse transcriptase
title Human Schwann cell senescence is not prevented by ectopic expression of human telomerase reverse transcriptase
spellingShingle Human Schwann cell senescence is not prevented by ectopic expression of human telomerase reverse transcriptase
Andersen, Natalia Denise
SCHWANN CELLS
SENESCENCE
PROLIFERATION
RETROVIRAL VECTORS
title_short Human Schwann cell senescence is not prevented by ectopic expression of human telomerase reverse transcriptase
title_full Human Schwann cell senescence is not prevented by ectopic expression of human telomerase reverse transcriptase
title_fullStr Human Schwann cell senescence is not prevented by ectopic expression of human telomerase reverse transcriptase
title_full_unstemmed Human Schwann cell senescence is not prevented by ectopic expression of human telomerase reverse transcriptase
title_sort Human Schwann cell senescence is not prevented by ectopic expression of human telomerase reverse transcriptase
dc.creator.none.fl_str_mv Andersen, Natalia Denise
Kuo, Blanche
Piñero, Gonzalo Miguel
Ravelo, Kristine
Rai, Priyamvada
Monje, Paula
author Andersen, Natalia Denise
author_facet Andersen, Natalia Denise
Kuo, Blanche
Piñero, Gonzalo Miguel
Ravelo, Kristine
Rai, Priyamvada
Monje, Paula
author_role author
author2 Kuo, Blanche
Piñero, Gonzalo Miguel
Ravelo, Kristine
Rai, Priyamvada
Monje, Paula
author2_role author
author
author
author
author
dc.subject.none.fl_str_mv SCHWANN CELLS
SENESCENCE
PROLIFERATION
RETROVIRAL VECTORS
topic SCHWANN CELLS
SENESCENCE
PROLIFERATION
RETROVIRAL VECTORS
purl_subject.fl_str_mv https://purl.org/becyt/ford/1.6
https://purl.org/becyt/ford/1
dc.description.none.fl_txt_mv Isolated human Schwann cells (hSCs) typically become senescent and unresponsive to mitogenic factors with continued in vitro expansion. Data from our lab has shown that adult nerve-derived hSCs, in contrast to rodent (rat) SCs, inevitably stop proliferating and acquire a senescent phenotype characterized by high levels of senescence-associated (SA)-β galactosidase activity and morphological changes that include cell enlargement and appearance of multinucleated cells. As opposed to rodent SCs, hSC cultures consist of mixed populations of proliferating cells, senescent cells and cells at different stages of differentiation regardless of the nerve of origin and other donor-specific factors. RNA-seq analysis of representative cultures of hSCs did not reveal the presence of telomerase reverse transcriptase (TERT) mRNA while other TERT-related genes (e.g. TERF1, telomeric repeat binding factor, and TEP1, telomeraseassociated protein) were well-represented in the hSC transcriptome. In an attempt to overcome senescence, we used retroviral vectors and antibiotic selection to generate hSC lines ectopically expressing human (h)-TERT. For these experiments, highly proliferative, non-senescent, early passage hSC cultures were stably transduced with the retroviruses h-TERT-hygro or h-TERTpuro, each encoding the h-TERT gene along with hygromycin or puromycin resistance genes, respectively. Transduced hSC cultures from three different donors were selected and subjected to three rounds of expansion in medium containing chemical mitogens. Subsequently, the cultures were analyzed for their rate of proliferation by means of EdU incorporation assays and the acquisition of senescence by means of SA-β galactosidase activity assays in each round. We found that whereas ectopic h-TERT expression extended the lifespan of cultured hSCs when compared to non-infected or GFP-expressing cells, it was not sufficient to confer immortalization and overcome senescence. In sum, our results suggest that progression of the hSCs to a senescent state likely is stress-induced rather than dependent on replication-associated telomere shortening
Fil: Andersen, Natalia Denise. Miami University; Estados Unidos. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina
Fil: Kuo, Blanche. Miami University; Estados Unidos
Fil: Piñero, Gonzalo Miguel. Miami University; Estados Unidos. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina
Fil: Ravelo, Kristine. Miami University; Estados Unidos
Fil: Rai, Priyamvada. Miami University. School Of Medicine; Estados Unidos
Fil: Monje, Paula. Miami University. School Of Medicine; Estados Unidos
Society for Neuroscience 46th Annual Meeting
San Diego
Estados Unidos
Society for Neuroscience
description Isolated human Schwann cells (hSCs) typically become senescent and unresponsive to mitogenic factors with continued in vitro expansion. Data from our lab has shown that adult nerve-derived hSCs, in contrast to rodent (rat) SCs, inevitably stop proliferating and acquire a senescent phenotype characterized by high levels of senescence-associated (SA)-β galactosidase activity and morphological changes that include cell enlargement and appearance of multinucleated cells. As opposed to rodent SCs, hSC cultures consist of mixed populations of proliferating cells, senescent cells and cells at different stages of differentiation regardless of the nerve of origin and other donor-specific factors. RNA-seq analysis of representative cultures of hSCs did not reveal the presence of telomerase reverse transcriptase (TERT) mRNA while other TERT-related genes (e.g. TERF1, telomeric repeat binding factor, and TEP1, telomeraseassociated protein) were well-represented in the hSC transcriptome. In an attempt to overcome senescence, we used retroviral vectors and antibiotic selection to generate hSC lines ectopically expressing human (h)-TERT. For these experiments, highly proliferative, non-senescent, early passage hSC cultures were stably transduced with the retroviruses h-TERT-hygro or h-TERTpuro, each encoding the h-TERT gene along with hygromycin or puromycin resistance genes, respectively. Transduced hSC cultures from three different donors were selected and subjected to three rounds of expansion in medium containing chemical mitogens. Subsequently, the cultures were analyzed for their rate of proliferation by means of EdU incorporation assays and the acquisition of senescence by means of SA-β galactosidase activity assays in each round. We found that whereas ectopic h-TERT expression extended the lifespan of cultured hSCs when compared to non-infected or GFP-expressing cells, it was not sufficient to confer immortalization and overcome senescence. In sum, our results suggest that progression of the hSCs to a senescent state likely is stress-induced rather than dependent on replication-associated telomere shortening
publishDate 2016
dc.date.none.fl_str_mv 2016
dc.type.none.fl_str_mv info:eu-repo/semantics/publishedVersion
info:eu-repo/semantics/conferenceObject
Congreso
Book
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/11336/235392
Human Schwann cell senescence is not prevented by ectopic expression of human telomerase reverse transcriptase; Society for Neuroscience 46th Annual Meeting; San Diego; Estados Unidos; 2016; 1-1
CONICET Digital
CONICET
url http://hdl.handle.net/11336/235392
identifier_str_mv Human Schwann cell senescence is not prevented by ectopic expression of human telomerase reverse transcriptase; Society for Neuroscience 46th Annual Meeting; San Diego; Estados Unidos; 2016; 1-1
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.sfn.org/Annual-Meeting/Neuroscience-2016/Sessions-and-Events/Program/~/media/SfN/Documents/Annual%20Meeting/FinalProgram/NS2016/Full%20Abstract%20PDFs%202016/SfN16_Abstract%20PDFs%20%20Posters_3_Mon_PM.ashx
info:eu-repo/semantics/altIdentifier/url/https://www.sfn.org/meetings/past-and-future-annual-meetings
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 Internacional
dc.publisher.none.fl_str_mv Society for Neuroscience
publisher.none.fl_str_mv Society for Neuroscience
dc.source.none.fl_str_mv reponame:CONICET Digital (CONICET)
instname:Consejo Nacional de Investigaciones Científicas y Técnicas
reponame_str CONICET Digital (CONICET)
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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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