Cargando…
Generation of hematopoietic stem cells from human embryonic stem cells using a defined, stepwise, serum-free, and serum replacement-free monolayer culture method
BACKGROUND: Embryonic stem cells (ESCs) can be expanded infinitely in vitro and have the potential to differentiate into hematopoietic stem cells (HSCs); thus, they are considered a useful source of cells for HSC production. Although several technical in vitro methods for engineering HSCs from pluri...
Autores principales: | , , , , , |
---|---|
Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Korean Society of Hematology; Korean Society of Blood and Marrow Transplantation; Korean Society of Pediatric Hematology-Oncology; Korean Society on Thrombosis and Hemostasis
2017
|
Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5383586/ https://www.ncbi.nlm.nih.gov/pubmed/28401100 http://dx.doi.org/10.5045/br.2017.52.1.37 |
_version_ | 1782520302408826880 |
---|---|
author | Kim, So-Jung Jung, Ji-Won Ha, Hye-Yeong Koo, Soo Kyung Kim, Eung-Gook Kim, Jung-Hyun |
author_facet | Kim, So-Jung Jung, Ji-Won Ha, Hye-Yeong Koo, Soo Kyung Kim, Eung-Gook Kim, Jung-Hyun |
author_sort | Kim, So-Jung |
collection | PubMed |
description | BACKGROUND: Embryonic stem cells (ESCs) can be expanded infinitely in vitro and have the potential to differentiate into hematopoietic stem cells (HSCs); thus, they are considered a useful source of cells for HSC production. Although several technical in vitro methods for engineering HSCs from pluripotent stem cells have been developed, clinical application of HSCs engineered from pluripotent stem cells is restricted because of the possibility of xenogeneic contamination resulting from the use of murine materials. METHODS: Human ESCs (CHA-hES15) were cultured on growth factor-reduced Matrigel-coated dishes in the mTeSR1 serum-free medium. When the cells were 70% confluent, we initiated HSC differentiation by three methods involving (1) knockout serum replacement (KSR), cytokines, TGFb1, EPO, and FLT3L; (2) KSR, cytokines, and bFGF; or (3) cytokines and bFGF. RESULTS: Among the three differentiation methods, the minimal number of cytokines without KSR resulted in the greatest production of HSCs. The optimized method resulted in a higher proportion of CD34(+)CD43(+) hematopoietic progenitor cells (HPCs) and CD34(+)CD45(+) HPCs compared to the other methods. In addition, the HSCs showed the potential to differentiate into multiple lineages of hematopoietic cells in vitro. CONCLUSION: In this study, we optimized a two-step, serum-free, animal protein-free, KSR-free, feeder-free, chemically defined monolayer culture method for generation of HSCs and hematopoietic stem and progenitor cells (HSPCs) from human ESCs. |
format | Online Article Text |
id | pubmed-5383586 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2017 |
publisher | Korean Society of Hematology; Korean Society of Blood and Marrow Transplantation; Korean Society of Pediatric Hematology-Oncology; Korean Society on Thrombosis and Hemostasis |
record_format | MEDLINE/PubMed |
spelling | pubmed-53835862017-04-11 Generation of hematopoietic stem cells from human embryonic stem cells using a defined, stepwise, serum-free, and serum replacement-free monolayer culture method Kim, So-Jung Jung, Ji-Won Ha, Hye-Yeong Koo, Soo Kyung Kim, Eung-Gook Kim, Jung-Hyun Blood Res Original Article BACKGROUND: Embryonic stem cells (ESCs) can be expanded infinitely in vitro and have the potential to differentiate into hematopoietic stem cells (HSCs); thus, they are considered a useful source of cells for HSC production. Although several technical in vitro methods for engineering HSCs from pluripotent stem cells have been developed, clinical application of HSCs engineered from pluripotent stem cells is restricted because of the possibility of xenogeneic contamination resulting from the use of murine materials. METHODS: Human ESCs (CHA-hES15) were cultured on growth factor-reduced Matrigel-coated dishes in the mTeSR1 serum-free medium. When the cells were 70% confluent, we initiated HSC differentiation by three methods involving (1) knockout serum replacement (KSR), cytokines, TGFb1, EPO, and FLT3L; (2) KSR, cytokines, and bFGF; or (3) cytokines and bFGF. RESULTS: Among the three differentiation methods, the minimal number of cytokines without KSR resulted in the greatest production of HSCs. The optimized method resulted in a higher proportion of CD34(+)CD43(+) hematopoietic progenitor cells (HPCs) and CD34(+)CD45(+) HPCs compared to the other methods. In addition, the HSCs showed the potential to differentiate into multiple lineages of hematopoietic cells in vitro. CONCLUSION: In this study, we optimized a two-step, serum-free, animal protein-free, KSR-free, feeder-free, chemically defined monolayer culture method for generation of HSCs and hematopoietic stem and progenitor cells (HSPCs) from human ESCs. Korean Society of Hematology; Korean Society of Blood and Marrow Transplantation; Korean Society of Pediatric Hematology-Oncology; Korean Society on Thrombosis and Hemostasis 2017-03 2017-03-27 /pmc/articles/PMC5383586/ /pubmed/28401100 http://dx.doi.org/10.5045/br.2017.52.1.37 Text en © 2017 Korean Society of Hematology http://creativecommons.org/licenses/by-nc/4.0 This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/4.0) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited. |
spellingShingle | Original Article Kim, So-Jung Jung, Ji-Won Ha, Hye-Yeong Koo, Soo Kyung Kim, Eung-Gook Kim, Jung-Hyun Generation of hematopoietic stem cells from human embryonic stem cells using a defined, stepwise, serum-free, and serum replacement-free monolayer culture method |
title | Generation of hematopoietic stem cells from human embryonic stem cells using a defined, stepwise, serum-free, and serum replacement-free monolayer culture method |
title_full | Generation of hematopoietic stem cells from human embryonic stem cells using a defined, stepwise, serum-free, and serum replacement-free monolayer culture method |
title_fullStr | Generation of hematopoietic stem cells from human embryonic stem cells using a defined, stepwise, serum-free, and serum replacement-free monolayer culture method |
title_full_unstemmed | Generation of hematopoietic stem cells from human embryonic stem cells using a defined, stepwise, serum-free, and serum replacement-free monolayer culture method |
title_short | Generation of hematopoietic stem cells from human embryonic stem cells using a defined, stepwise, serum-free, and serum replacement-free monolayer culture method |
title_sort | generation of hematopoietic stem cells from human embryonic stem cells using a defined, stepwise, serum-free, and serum replacement-free monolayer culture method |
topic | Original Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5383586/ https://www.ncbi.nlm.nih.gov/pubmed/28401100 http://dx.doi.org/10.5045/br.2017.52.1.37 |
work_keys_str_mv | AT kimsojung generationofhematopoieticstemcellsfromhumanembryonicstemcellsusingadefinedstepwiseserumfreeandserumreplacementfreemonolayerculturemethod AT jungjiwon generationofhematopoieticstemcellsfromhumanembryonicstemcellsusingadefinedstepwiseserumfreeandserumreplacementfreemonolayerculturemethod AT hahyeyeong generationofhematopoieticstemcellsfromhumanembryonicstemcellsusingadefinedstepwiseserumfreeandserumreplacementfreemonolayerculturemethod AT koosookyung generationofhematopoieticstemcellsfromhumanembryonicstemcellsusingadefinedstepwiseserumfreeandserumreplacementfreemonolayerculturemethod AT kimeunggook generationofhematopoieticstemcellsfromhumanembryonicstemcellsusingadefinedstepwiseserumfreeandserumreplacementfreemonolayerculturemethod AT kimjunghyun generationofhematopoieticstemcellsfromhumanembryonicstemcellsusingadefinedstepwiseserumfreeandserumreplacementfreemonolayerculturemethod |