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Human peripheral blood mononuclear cells enriched in endothelial progenitor cells via quality and quantity controlled culture accelerate vascularization and wound healing in a porcine wound model

The transplantation of endothelial progenitor cells (EPCs) is used to promote wound angiogenesis. In patients with chronic wounds and accompanying morbidities, EPCs are often compromised in number and function. To overcome these limitations, we previously developed a quality and quantity controlled...

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Autores principales: Kado, Makiko, Tanaka, Rica, Arita, Kayo, Okada, Kayoko, Ito-Hirano, Rie, Fujimura, Satoshi, Mizuno, Hiroshi
Formato: Online Artículo Texto
Lenguaje:English
Publicado: SAGE Publications 2018
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6158547/
https://www.ncbi.nlm.nih.gov/pubmed/29974793
http://dx.doi.org/10.1177/0963689718780307
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author Kado, Makiko
Tanaka, Rica
Arita, Kayo
Okada, Kayoko
Ito-Hirano, Rie
Fujimura, Satoshi
Mizuno, Hiroshi
author_facet Kado, Makiko
Tanaka, Rica
Arita, Kayo
Okada, Kayoko
Ito-Hirano, Rie
Fujimura, Satoshi
Mizuno, Hiroshi
author_sort Kado, Makiko
collection PubMed
description The transplantation of endothelial progenitor cells (EPCs) is used to promote wound angiogenesis. In patients with chronic wounds and accompanying morbidities, EPCs are often compromised in number and function. To overcome these limitations, we previously developed a quality and quantity controlled (QQ) culture system to enrich peripheral blood mononuclear cells (PBMNCs) in EPCs. To evaluate the wound healing efficacy of mononuclear cells (MNCs) harvested after QQ culture (QQMNCs), preclinical studies were performed on large animals. MNCs harvested from the blood of healthy human subjects were cultured in the presence of angiogenic cytokines and growth factors in a serum-free medium for 7 days. A total of 5 × 10(6) QQMNCs per full-thickness skin defect or control saline was injected into wounds induced in cyclosporine-immunosuppressed pigs. EPC colony-forming assays revealed a significantly higher number of definitive (partially differentiated) EPC colony-forming units in QQMNCs. Flow cytometry evaluation of QQMNC surface markers showed enrichment of CD34(+) and CD133(+) stem cell populations, significant reduction in CCR2(+) cell percentages, and a greater than 10-fold increase in the percentage of anti-inflammatory M2-type macrophages (CD206(+) cells) compared with PBMNCs. Wounds treated with QQMNCs had a significantly higher closure rate. Wounds were harvested, frozen, and sectioned at day 21 postoperatively. Hematoxylin and eosin staining revealed that the epithelization of QQMNC-treated wounds was more advanced than in controls. Treated wounds developed granulation tissue with more mature collagen and larger capillary networks. CD31 and human mitochondrial co-staining confirmed the presence of differentiated human cells within newly formed vessels. Real-time polymerase chain reaction (PCR) showed upregulation of interleukin 6 (IL-6), IL-10, and IL-4 in the wound bed, suggesting paracrine activity of the transplanted QQMNCs. Our data demonstrate for the first time that QQ culture of MNCs obtained from a small amount of peripheral blood yields vasculogenic and therapeutic cells effective in wound healing.
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spelling pubmed-61585472018-10-01 Human peripheral blood mononuclear cells enriched in endothelial progenitor cells via quality and quantity controlled culture accelerate vascularization and wound healing in a porcine wound model Kado, Makiko Tanaka, Rica Arita, Kayo Okada, Kayoko Ito-Hirano, Rie Fujimura, Satoshi Mizuno, Hiroshi Cell Transplant Original Articles The transplantation of endothelial progenitor cells (EPCs) is used to promote wound angiogenesis. In patients with chronic wounds and accompanying morbidities, EPCs are often compromised in number and function. To overcome these limitations, we previously developed a quality and quantity controlled (QQ) culture system to enrich peripheral blood mononuclear cells (PBMNCs) in EPCs. To evaluate the wound healing efficacy of mononuclear cells (MNCs) harvested after QQ culture (QQMNCs), preclinical studies were performed on large animals. MNCs harvested from the blood of healthy human subjects were cultured in the presence of angiogenic cytokines and growth factors in a serum-free medium for 7 days. A total of 5 × 10(6) QQMNCs per full-thickness skin defect or control saline was injected into wounds induced in cyclosporine-immunosuppressed pigs. EPC colony-forming assays revealed a significantly higher number of definitive (partially differentiated) EPC colony-forming units in QQMNCs. Flow cytometry evaluation of QQMNC surface markers showed enrichment of CD34(+) and CD133(+) stem cell populations, significant reduction in CCR2(+) cell percentages, and a greater than 10-fold increase in the percentage of anti-inflammatory M2-type macrophages (CD206(+) cells) compared with PBMNCs. Wounds treated with QQMNCs had a significantly higher closure rate. Wounds were harvested, frozen, and sectioned at day 21 postoperatively. Hematoxylin and eosin staining revealed that the epithelization of QQMNC-treated wounds was more advanced than in controls. Treated wounds developed granulation tissue with more mature collagen and larger capillary networks. CD31 and human mitochondrial co-staining confirmed the presence of differentiated human cells within newly formed vessels. Real-time polymerase chain reaction (PCR) showed upregulation of interleukin 6 (IL-6), IL-10, and IL-4 in the wound bed, suggesting paracrine activity of the transplanted QQMNCs. Our data demonstrate for the first time that QQ culture of MNCs obtained from a small amount of peripheral blood yields vasculogenic and therapeutic cells effective in wound healing. SAGE Publications 2018-07-05 2018-07 /pmc/articles/PMC6158547/ /pubmed/29974793 http://dx.doi.org/10.1177/0963689718780307 Text en © The Author(s) 2018 http://creativecommons.org/licenses/by/4.0/ This article is distributed under the terms of the Creative Commons Attribution 4.0 License (http://www.creativecommons.org/licenses/by/4.0/) which permits any use, reproduction and distribution of the work without further permission provided the original work is attributed as specified on the SAGE and Open Access pages (https://us.sagepub.com/en-us/nam/open-access-at-sage).
spellingShingle Original Articles
Kado, Makiko
Tanaka, Rica
Arita, Kayo
Okada, Kayoko
Ito-Hirano, Rie
Fujimura, Satoshi
Mizuno, Hiroshi
Human peripheral blood mononuclear cells enriched in endothelial progenitor cells via quality and quantity controlled culture accelerate vascularization and wound healing in a porcine wound model
title Human peripheral blood mononuclear cells enriched in endothelial progenitor cells via quality and quantity controlled culture accelerate vascularization and wound healing in a porcine wound model
title_full Human peripheral blood mononuclear cells enriched in endothelial progenitor cells via quality and quantity controlled culture accelerate vascularization and wound healing in a porcine wound model
title_fullStr Human peripheral blood mononuclear cells enriched in endothelial progenitor cells via quality and quantity controlled culture accelerate vascularization and wound healing in a porcine wound model
title_full_unstemmed Human peripheral blood mononuclear cells enriched in endothelial progenitor cells via quality and quantity controlled culture accelerate vascularization and wound healing in a porcine wound model
title_short Human peripheral blood mononuclear cells enriched in endothelial progenitor cells via quality and quantity controlled culture accelerate vascularization and wound healing in a porcine wound model
title_sort human peripheral blood mononuclear cells enriched in endothelial progenitor cells via quality and quantity controlled culture accelerate vascularization and wound healing in a porcine wound model
topic Original Articles
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6158547/
https://www.ncbi.nlm.nih.gov/pubmed/29974793
http://dx.doi.org/10.1177/0963689718780307
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