A scalable solution for isolating human multipotent clinical-grade neural stem cells from ES precursors
Abstract Background A well-characterized method has not yet been established to reproducibly, efficiently, and safely isolate large numbers of clinical-grade multipotent human neural stem cells (hNSCs) from embryonic stem cells (hESCs). Consequently, the transplantation of neurogenic/gliogenic precu...
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2019-03-01
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Series: | Stem Cell Research & Therapy |
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Online Access: | http://link.springer.com/article/10.1186/s13287-019-1163-7 |
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Article |
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DOAJ |
language |
English |
format |
Article |
sources |
DOAJ |
author |
Dasa Bohaciakova Marian Hruska-Plochan Rachel Tsunemoto Wesley D. Gifford Shawn P. Driscoll Thomas D. Glenn Stephanie Wu Silvia Marsala Michael Navarro Takahiro Tadokoro Stefan Juhas Jana Juhasova Oleksandr Platoshyn David Piper Vickie Sheckler Dara Ditsworth Samuel L. Pfaff Martin Marsala |
spellingShingle |
Dasa Bohaciakova Marian Hruska-Plochan Rachel Tsunemoto Wesley D. Gifford Shawn P. Driscoll Thomas D. Glenn Stephanie Wu Silvia Marsala Michael Navarro Takahiro Tadokoro Stefan Juhas Jana Juhasova Oleksandr Platoshyn David Piper Vickie Sheckler Dara Ditsworth Samuel L. Pfaff Martin Marsala A scalable solution for isolating human multipotent clinical-grade neural stem cells from ES precursors Stem Cell Research & Therapy Human embryonic stem cell (hESC) Neural stem cell (NSC) Spinal cord Amyotrophic lateral sclerosis (ALS) Spinal traumatic injury Bioinformatic tools to study xenografts |
author_facet |
Dasa Bohaciakova Marian Hruska-Plochan Rachel Tsunemoto Wesley D. Gifford Shawn P. Driscoll Thomas D. Glenn Stephanie Wu Silvia Marsala Michael Navarro Takahiro Tadokoro Stefan Juhas Jana Juhasova Oleksandr Platoshyn David Piper Vickie Sheckler Dara Ditsworth Samuel L. Pfaff Martin Marsala |
author_sort |
Dasa Bohaciakova |
title |
A scalable solution for isolating human multipotent clinical-grade neural stem cells from ES precursors |
title_short |
A scalable solution for isolating human multipotent clinical-grade neural stem cells from ES precursors |
title_full |
A scalable solution for isolating human multipotent clinical-grade neural stem cells from ES precursors |
title_fullStr |
A scalable solution for isolating human multipotent clinical-grade neural stem cells from ES precursors |
title_full_unstemmed |
A scalable solution for isolating human multipotent clinical-grade neural stem cells from ES precursors |
title_sort |
scalable solution for isolating human multipotent clinical-grade neural stem cells from es precursors |
publisher |
BMC |
series |
Stem Cell Research & Therapy |
issn |
1757-6512 |
publishDate |
2019-03-01 |
description |
Abstract Background A well-characterized method has not yet been established to reproducibly, efficiently, and safely isolate large numbers of clinical-grade multipotent human neural stem cells (hNSCs) from embryonic stem cells (hESCs). Consequently, the transplantation of neurogenic/gliogenic precursors into the CNS for the purpose of cell replacement or neuroprotection in humans with injury or disease has not achieved widespread testing and implementation. Methods Here, we establish an approach for the in vitro isolation of a highly expandable population of hNSCs using the manual selection of neural precursors based on their colony morphology (CoMo-NSC). The purity and NSC properties of established and extensively expanded CoMo-NSC were validated by expression of NSC markers (flow cytometry, mRNA sequencing), lack of pluripotent markers and by their tumorigenic/differentiation profile after in vivo spinal grafting in three different animal models, including (i) immunodeficient rats, (ii) immunosuppressed ALS rats (SOD1G93A), or (iii) spinally injured immunosuppressed minipigs. Results In vitro analysis of established CoMo-NSCs showed a consistent expression of NSC markers (Sox1, Sox2, Nestin, CD24) with lack of pluripotent markers (Nanog) and stable karyotype for more than 15 passages. Gene profiling and histology revealed that spinally grafted CoMo-NSCs differentiate into neurons, astrocytes, and oligodendrocytes over a 2–6-month period in vivo without forming neoplastic derivatives or abnormal structures. Moreover, transplanted CoMo-NSCs formed neurons with synaptic contacts and glia in a variety of host environments including immunodeficient rats, immunosuppressed ALS rats (SOD1G93A), or spinally injured minipigs, indicating these cells have favorable safety and differentiation characteristics. Conclusions These data demonstrate that manually selected CoMo-NSCs represent a safe and expandable NSC population which can effectively be used in prospective human clinical cell replacement trials for the treatment of a variety of neurodegenerative disorders, including ALS, stroke, spinal traumatic, or spinal ischemic injury. |
topic |
Human embryonic stem cell (hESC) Neural stem cell (NSC) Spinal cord Amyotrophic lateral sclerosis (ALS) Spinal traumatic injury Bioinformatic tools to study xenografts |
url |
http://link.springer.com/article/10.1186/s13287-019-1163-7 |
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doaj-f87a5d79962b40ba83ca9d1341131d902020-11-25T00:42:02ZengBMCStem Cell Research & Therapy1757-65122019-03-0110111910.1186/s13287-019-1163-7A scalable solution for isolating human multipotent clinical-grade neural stem cells from ES precursorsDasa Bohaciakova0Marian Hruska-Plochan1Rachel Tsunemoto2Wesley D. Gifford3Shawn P. Driscoll4Thomas D. Glenn5Stephanie Wu6Silvia Marsala7Michael Navarro8Takahiro Tadokoro9Stefan Juhas10Jana Juhasova11Oleksandr Platoshyn12David Piper13Vickie Sheckler14Dara Ditsworth15Samuel L. Pfaff16Martin Marsala17Department of Anesthesiology, University of California San Diego School of MedicineDepartment of Anesthesiology, University of California San Diego School of MedicineGene Expression Laboratory, Howard Hughes Medical Institute and Salk Institute for Biological StudiesGene Expression Laboratory, Howard Hughes Medical Institute and Salk Institute for Biological StudiesGene Expression Laboratory, Howard Hughes Medical Institute and Salk Institute for Biological StudiesGene Expression Laboratory, Howard Hughes Medical Institute and Salk Institute for Biological StudiesDepartment of Anesthesiology, University of California San Diego School of MedicineDepartment of Anesthesiology, University of California San Diego School of MedicineDepartment of Anesthesiology, University of California San Diego School of MedicineDepartment of Anesthesiology, University of California San Diego School of MedicineInstitute of Animal Physiology and Genetics, v.v.i., AS CRInstitute of Animal Physiology and Genetics, v.v.i., AS CRDepartment of Anesthesiology, University of California San Diego School of MedicinePrimary and Stem Cell Systems, Life Technologies (Thermo Fisher Scientific)Sanford Stem Cell Clinical Center, University of California San DiegoDepartment of Cellular and Molecular Medicine, University of California San DiegoGene Expression Laboratory, Howard Hughes Medical Institute and Salk Institute for Biological StudiesDepartment of Anesthesiology, University of California San Diego School of MedicineAbstract Background A well-characterized method has not yet been established to reproducibly, efficiently, and safely isolate large numbers of clinical-grade multipotent human neural stem cells (hNSCs) from embryonic stem cells (hESCs). Consequently, the transplantation of neurogenic/gliogenic precursors into the CNS for the purpose of cell replacement or neuroprotection in humans with injury or disease has not achieved widespread testing and implementation. Methods Here, we establish an approach for the in vitro isolation of a highly expandable population of hNSCs using the manual selection of neural precursors based on their colony morphology (CoMo-NSC). The purity and NSC properties of established and extensively expanded CoMo-NSC were validated by expression of NSC markers (flow cytometry, mRNA sequencing), lack of pluripotent markers and by their tumorigenic/differentiation profile after in vivo spinal grafting in three different animal models, including (i) immunodeficient rats, (ii) immunosuppressed ALS rats (SOD1G93A), or (iii) spinally injured immunosuppressed minipigs. Results In vitro analysis of established CoMo-NSCs showed a consistent expression of NSC markers (Sox1, Sox2, Nestin, CD24) with lack of pluripotent markers (Nanog) and stable karyotype for more than 15 passages. Gene profiling and histology revealed that spinally grafted CoMo-NSCs differentiate into neurons, astrocytes, and oligodendrocytes over a 2–6-month period in vivo without forming neoplastic derivatives or abnormal structures. Moreover, transplanted CoMo-NSCs formed neurons with synaptic contacts and glia in a variety of host environments including immunodeficient rats, immunosuppressed ALS rats (SOD1G93A), or spinally injured minipigs, indicating these cells have favorable safety and differentiation characteristics. Conclusions These data demonstrate that manually selected CoMo-NSCs represent a safe and expandable NSC population which can effectively be used in prospective human clinical cell replacement trials for the treatment of a variety of neurodegenerative disorders, including ALS, stroke, spinal traumatic, or spinal ischemic injury.http://link.springer.com/article/10.1186/s13287-019-1163-7Human embryonic stem cell (hESC)Neural stem cell (NSC)Spinal cordAmyotrophic lateral sclerosis (ALS)Spinal traumatic injuryBioinformatic tools to study xenografts |