Immortalized human hair follicle-derived mesenchymal-like stromal cells for the long-term production of scalable immunomodulatory and regenerative secretome

Background: Mesenchymal stromal cells (MSCs) exert therapeutic effects primarily through their secretome, rich in bioactive factors with immunomodulatory and regenerative properties. However, clinical application of MSC-derived secretome is hindered by donor variability, limited expansion, and repli...

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Autores: Rosselló Gelabert, María, Las Heras Zapata, Kevin, González Pujana, Ainhoa, Igartua Olaechea, Manuela, Santos Vizcaíno, Edorta, Hernández Martín, Rosa María
Tipo de recurso: artículo
Fecha de publicación:2025
País:España
Institución:Universidad del País Vasco
Repositorio:Addi. Archivo Digital para la Docencia y la Investigación
OAI Identifier:oai:addi.ehu.eus:10810/75740
Acceso en línea:http://hdl.handle.net/10810/75740
Access Level:acceso abierto
Palabra clave:cell-free therapy
haie follicle
immortalization
immunomodulation
mesenchymal stromal cell (MSC)
regenerative medicine
secretome
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spelling Immortalized human hair follicle-derived mesenchymal-like stromal cells for the long-term production of scalable immunomodulatory and regenerative secretomeRosselló Gelabert, MaríaLas Heras Zapata, KevinGonzález Pujana, AinhoaIgartua Olaechea, ManuelaSantos Vizcaíno, EdortaHernández Martín, Rosa Maríacell-free therapyhaie follicleimmortalizationimmunomodulationmesenchymal stromal cell (MSC)regenerative medicinesecretomeBackground: Mesenchymal stromal cells (MSCs) exert therapeutic effects primarily through their secretome, rich in bioactive factors with immunomodulatory and regenerative properties. However, clinical application of MSC-derived secretome is hindered by donor variability, limited expansion, and replicative senescence. To address these issues, we developed immortalized human hair follicle-derived mesenchymal-like stromal cells (iHF-MSCs) as a consistent and scalable source of therapeutic secretome. Methods: HF-MSCs were isolated using a minimally invasive procedure and immortalized with lentiviral SV40 large T antigen. From 576 single-cell clones, C18 and C26 were selected based on proliferation capacity, absence of senescence, and retention of a mesenchymal-like phenotype. Characterization included RT-PCR, flow cytometry for surface markers, trilineage differentiation, and CD56 expression analysis. For immunomodulatory evaluation, cells were licensed with IFN-γ and TNF-α, and the resulting conditioned media (licensed CM) were analyzed by ELISA and cytokine arrays. PBMC proliferation and Treg induction were assessed using licensed CM, while unlicensed CM was tested for its regenerative effects on dermal and epidermal cell functions, including wound repair, tube formation assay as an indicator of angiogenic potential, and oxidative stress response. Results: The selected clones (C18 and C26) exhibited long-term stability, rapid expansion, and preserved mesenchymal-like phenotype and multipotency. Secretome profiling revealed an enriched composition of immunoregulatory (e.g., Gal-9, IL-1Ra, TSG-6) and pro-regenerative (e.g., VEGF, PDGF-AA, EGF) factors, with enhanced responsiveness to inflammatory licensing. Notably, iHF-MSCs maintained low immunogenicity and demonstrated superior functional performance: inhibition of PBMC proliferation, Treg induction, and promotion of skin cell proliferation, migration, tube formation, and oxidative stress protection. Conclusion: These results position iHF-MSCs as a robust and scalable platform for consistent production of therapeutic secretome, paving the way for innovative, cell-free immunomodulatory and regenerative therapies.This work was supported by Grant PID2021-122577OB-I00 funded by MCIN/AEI/ https://doi.org/10.13039/501100011033 and by “ERDF A way of making Europe”. Additionally, Grant IT1448-22 funded by Basque Government. M. Rossello-Gelabert thanks the grant PRE2022-102058, funded by MCIN/AEI/https://doi.org/10.13039/501100011033 and by the FSE+. A. Gonzalez-Pujana thanks the University of the Basque Country (UPV/EHU) for the postdoctoral grant (ESPDOC20/119).BMC202520252025info:eu-repo/semantics/articleapplication/pdfhttp://hdl.handle.net/10810/75740reponame:Addi. Archivo Digital para la Docencia y la Investigacióninstname:Universidad del País VascoInglésinfo:eu-repo/grantAgreement/MICINN/PID2021-122577OB-I00/https://stemcellres.biomedcentral.com/articles/10.1186/s13287-025-04775-8info:eu-repo/semantics/openAccesshttp://creativecommons.org/licenses/by-nc-nd/4.0/© The Author(s) 2025. This article is licensed under a Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International Licenseoai:addi.ehu.eus:10810/757402026-06-18T09:23:17Z
dc.title.none.fl_str_mv Immortalized human hair follicle-derived mesenchymal-like stromal cells for the long-term production of scalable immunomodulatory and regenerative secretome
title Immortalized human hair follicle-derived mesenchymal-like stromal cells for the long-term production of scalable immunomodulatory and regenerative secretome
spellingShingle Immortalized human hair follicle-derived mesenchymal-like stromal cells for the long-term production of scalable immunomodulatory and regenerative secretome
Rosselló Gelabert, María
cell-free therapy
haie follicle
immortalization
immunomodulation
mesenchymal stromal cell (MSC)
regenerative medicine
secretome
title_short Immortalized human hair follicle-derived mesenchymal-like stromal cells for the long-term production of scalable immunomodulatory and regenerative secretome
title_full Immortalized human hair follicle-derived mesenchymal-like stromal cells for the long-term production of scalable immunomodulatory and regenerative secretome
title_fullStr Immortalized human hair follicle-derived mesenchymal-like stromal cells for the long-term production of scalable immunomodulatory and regenerative secretome
title_full_unstemmed Immortalized human hair follicle-derived mesenchymal-like stromal cells for the long-term production of scalable immunomodulatory and regenerative secretome
title_sort Immortalized human hair follicle-derived mesenchymal-like stromal cells for the long-term production of scalable immunomodulatory and regenerative secretome
dc.creator.none.fl_str_mv Rosselló Gelabert, María
Las Heras Zapata, Kevin
González Pujana, Ainhoa
Igartua Olaechea, Manuela
Santos Vizcaíno, Edorta
Hernández Martín, Rosa María
author Rosselló Gelabert, María
author_facet Rosselló Gelabert, María
Las Heras Zapata, Kevin
González Pujana, Ainhoa
Igartua Olaechea, Manuela
Santos Vizcaíno, Edorta
Hernández Martín, Rosa María
author_role author
author2 Las Heras Zapata, Kevin
González Pujana, Ainhoa
Igartua Olaechea, Manuela
Santos Vizcaíno, Edorta
Hernández Martín, Rosa María
author2_role author
author
author
author
author
dc.subject.none.fl_str_mv cell-free therapy
haie follicle
immortalization
immunomodulation
mesenchymal stromal cell (MSC)
regenerative medicine
secretome
topic cell-free therapy
haie follicle
immortalization
immunomodulation
mesenchymal stromal cell (MSC)
regenerative medicine
secretome
description Background: Mesenchymal stromal cells (MSCs) exert therapeutic effects primarily through their secretome, rich in bioactive factors with immunomodulatory and regenerative properties. However, clinical application of MSC-derived secretome is hindered by donor variability, limited expansion, and replicative senescence. To address these issues, we developed immortalized human hair follicle-derived mesenchymal-like stromal cells (iHF-MSCs) as a consistent and scalable source of therapeutic secretome. Methods: HF-MSCs were isolated using a minimally invasive procedure and immortalized with lentiviral SV40 large T antigen. From 576 single-cell clones, C18 and C26 were selected based on proliferation capacity, absence of senescence, and retention of a mesenchymal-like phenotype. Characterization included RT-PCR, flow cytometry for surface markers, trilineage differentiation, and CD56 expression analysis. For immunomodulatory evaluation, cells were licensed with IFN-γ and TNF-α, and the resulting conditioned media (licensed CM) were analyzed by ELISA and cytokine arrays. PBMC proliferation and Treg induction were assessed using licensed CM, while unlicensed CM was tested for its regenerative effects on dermal and epidermal cell functions, including wound repair, tube formation assay as an indicator of angiogenic potential, and oxidative stress response. Results: The selected clones (C18 and C26) exhibited long-term stability, rapid expansion, and preserved mesenchymal-like phenotype and multipotency. Secretome profiling revealed an enriched composition of immunoregulatory (e.g., Gal-9, IL-1Ra, TSG-6) and pro-regenerative (e.g., VEGF, PDGF-AA, EGF) factors, with enhanced responsiveness to inflammatory licensing. Notably, iHF-MSCs maintained low immunogenicity and demonstrated superior functional performance: inhibition of PBMC proliferation, Treg induction, and promotion of skin cell proliferation, migration, tube formation, and oxidative stress protection. Conclusion: These results position iHF-MSCs as a robust and scalable platform for consistent production of therapeutic secretome, paving the way for innovative, cell-free immunomodulatory and regenerative therapies.
publishDate 2025
dc.date.none.fl_str_mv 2025
2025
2025
dc.type.none.fl_str_mv info:eu-repo/semantics/article
format article
dc.identifier.none.fl_str_mv http://hdl.handle.net/10810/75740
url http://hdl.handle.net/10810/75740
dc.language.none.fl_str_mv Inglés
language_invalid_str_mv Inglés
dc.relation.none.fl_str_mv info:eu-repo/grantAgreement/MICINN/PID2021-122577OB-I00/
https://stemcellres.biomedcentral.com/articles/10.1186/s13287-025-04775-8
dc.rights.none.fl_str_mv info:eu-repo/semantics/openAccess
http://creativecommons.org/licenses/by-nc-nd/4.0/
eu_rights_str_mv openAccess
rights_invalid_str_mv http://creativecommons.org/licenses/by-nc-nd/4.0/
dc.format.none.fl_str_mv application/pdf
dc.publisher.none.fl_str_mv BMC
publisher.none.fl_str_mv BMC
dc.source.none.fl_str_mv reponame:Addi. Archivo Digital para la Docencia y la Investigación
instname:Universidad del País Vasco
instname_str Universidad del País Vasco
reponame_str Addi. Archivo Digital para la Docencia y la Investigación
collection Addi. Archivo Digital para la Docencia y la Investigación
repository.name.fl_str_mv
repository.mail.fl_str_mv
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