Direct reprogramming of dermal fibroblasts derived from Northern elephant seals into muscle cells
Ex vivo tissue culture allows the study of complex cellular mechanisms that are relevant to physiological responses while overcoming the challenges presented by studying animals that are not tractable. In a primary cell culture system, certain proliferating cells can be functionally reprogrammed int...
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Online Access: | http://dx.doi.org/10.1096/fasebj.2022.36.s1.r6331 |
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crwiley:10.1096/fasebj.2022.36.s1.r6331 2024-06-02T08:06:01+00:00 Direct reprogramming of dermal fibroblasts derived from Northern elephant seals into muscle cells Lam, Emily Torres‐Velarde, Julia M. Allen, Kaitlin Arango, B. Gabriela Crocker, Daniel Vázquez‐Medina, José P. 2022 http://dx.doi.org/10.1096/fasebj.2022.36.s1.r6331 en eng Wiley http://onlinelibrary.wiley.com/termsAndConditions#vor The FASEB Journal volume 36, issue S1 ISSN 0892-6638 1530-6860 journal-article 2022 crwiley https://doi.org/10.1096/fasebj.2022.36.s1.r6331 2024-05-03T12:04:09Z Ex vivo tissue culture allows the study of complex cellular mechanisms that are relevant to physiological responses while overcoming the challenges presented by studying animals that are not tractable. In a primary cell culture system, certain proliferating cells can be functionally reprogrammed into other cell types via overexpression of key genes. Dermal fibroblasts can be reprogrammed into muscle cells, which are often challenging to obtain but offer a unique system to study metabolic responses, by overexpression of the myogenic transcription factor myod. We isolated cells from Northern elephant seal (NES) skin samples and propagated them in primary culture. NES cells respire, stain positive for fibroblast markers (vimentin and PDGFR), and are amenable to electroporation. We overexpressed GFP‐myod in NES fibroblasts and conducted antibiotic selection and purification by FACS. As expected, expression of myod was higher in transfected cells than in controls according to qPCR analysis (t‐test p< 0.05). Treatment with small molecules (CHIR99021, Forskolin and Repsox) enhanced myod expression. Furthermore, fibroblasts overexpressing myod expressed downstream markers of myogenesis (myogenin, myosin heavy chain 1 and myosin heavy chain 8) and the effect was enhanced when myod‐overexpressing cells were supplemented with small molecules. We are currently evaluating the capacity of myod‐overexpressing cells to differentiate into myotubes and comparing metabolic profiles with primary NES myoblasts. Establishing differentiated muscle fibers from other mature cell types could provide a unique platform to conduct mechanistic studies in species where muscle tissue samples cannot be obtained from live animals. Article in Journal/Newspaper Elephant Seal Elephant Seals Wiley Online Library Nes ENVELOPE(7.634,7.634,62.795,62.795) Nes’ ENVELOPE(44.681,44.681,66.600,66.600) The FASEB Journal 36 S1 |
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Open Polar |
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Wiley Online Library |
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crwiley |
language |
English |
description |
Ex vivo tissue culture allows the study of complex cellular mechanisms that are relevant to physiological responses while overcoming the challenges presented by studying animals that are not tractable. In a primary cell culture system, certain proliferating cells can be functionally reprogrammed into other cell types via overexpression of key genes. Dermal fibroblasts can be reprogrammed into muscle cells, which are often challenging to obtain but offer a unique system to study metabolic responses, by overexpression of the myogenic transcription factor myod. We isolated cells from Northern elephant seal (NES) skin samples and propagated them in primary culture. NES cells respire, stain positive for fibroblast markers (vimentin and PDGFR), and are amenable to electroporation. We overexpressed GFP‐myod in NES fibroblasts and conducted antibiotic selection and purification by FACS. As expected, expression of myod was higher in transfected cells than in controls according to qPCR analysis (t‐test p< 0.05). Treatment with small molecules (CHIR99021, Forskolin and Repsox) enhanced myod expression. Furthermore, fibroblasts overexpressing myod expressed downstream markers of myogenesis (myogenin, myosin heavy chain 1 and myosin heavy chain 8) and the effect was enhanced when myod‐overexpressing cells were supplemented with small molecules. We are currently evaluating the capacity of myod‐overexpressing cells to differentiate into myotubes and comparing metabolic profiles with primary NES myoblasts. Establishing differentiated muscle fibers from other mature cell types could provide a unique platform to conduct mechanistic studies in species where muscle tissue samples cannot be obtained from live animals. |
format |
Article in Journal/Newspaper |
author |
Lam, Emily Torres‐Velarde, Julia M. Allen, Kaitlin Arango, B. Gabriela Crocker, Daniel Vázquez‐Medina, José P. |
spellingShingle |
Lam, Emily Torres‐Velarde, Julia M. Allen, Kaitlin Arango, B. Gabriela Crocker, Daniel Vázquez‐Medina, José P. Direct reprogramming of dermal fibroblasts derived from Northern elephant seals into muscle cells |
author_facet |
Lam, Emily Torres‐Velarde, Julia M. Allen, Kaitlin Arango, B. Gabriela Crocker, Daniel Vázquez‐Medina, José P. |
author_sort |
Lam, Emily |
title |
Direct reprogramming of dermal fibroblasts derived from Northern elephant seals into muscle cells |
title_short |
Direct reprogramming of dermal fibroblasts derived from Northern elephant seals into muscle cells |
title_full |
Direct reprogramming of dermal fibroblasts derived from Northern elephant seals into muscle cells |
title_fullStr |
Direct reprogramming of dermal fibroblasts derived from Northern elephant seals into muscle cells |
title_full_unstemmed |
Direct reprogramming of dermal fibroblasts derived from Northern elephant seals into muscle cells |
title_sort |
direct reprogramming of dermal fibroblasts derived from northern elephant seals into muscle cells |
publisher |
Wiley |
publishDate |
2022 |
url |
http://dx.doi.org/10.1096/fasebj.2022.36.s1.r6331 |
long_lat |
ENVELOPE(7.634,7.634,62.795,62.795) ENVELOPE(44.681,44.681,66.600,66.600) |
geographic |
Nes Nes’ |
geographic_facet |
Nes Nes’ |
genre |
Elephant Seal Elephant Seals |
genre_facet |
Elephant Seal Elephant Seals |
op_source |
The FASEB Journal volume 36, issue S1 ISSN 0892-6638 1530-6860 |
op_rights |
http://onlinelibrary.wiley.com/termsAndConditions#vor |
op_doi |
https://doi.org/10.1096/fasebj.2022.36.s1.r6331 |
container_title |
The FASEB Journal |
container_volume |
36 |
container_issue |
S1 |
_version_ |
1800750895591325696 |