The Bioactive Peptide SL-13R Expands Human Umbilical Cord Blood Hematopoietic Stem and Progenitor Cells In Vitro

Hematopoietic stem and progenitor cell (HSPC) transplantation is a curative treatment of hematological disorders that has been utilized for several decades. Although umbilical cord blood (UCB) is a promising source of HSPCs, the low dose of HSPCs in these preparations limits their use, prompting need for ex vivo HSPC expansion. To establish a more efficient method to expand UCB HSPCs, we developed the bioactive peptide named SL-13R and cultured UCB HSPCs (CD34+ cells) with SL-13R in animal component-free medium containing a cytokine cocktail. Following 9 days of culture with SL-13R, the numbers of total cells, CD34+, CD38− cells, and hematopoietic stem cell (HSC)-enriched cells were significantly increased relative to control. Transplantation of cells cultured with SL-13R into immunodeficient NOD/Shi-scid/IL-2Rγ knockout mice confirmed that they possess long-term reconstitution and self-renewal ability. AHNAK, ANXA2, and PLEC all interact with SL-13R. Knockdown of these genes in UCB CD34+ cells resulted in reduced numbers of hematopoietic colonies relative to SL-13R-treated and non-knockdown controls. In summary, we have identified a novel bioactive peptide SL-13R promoting expansion of UCB CD34+ cells with long-term reconstitution and self-renewal ability, suggesting its clinical use in the future.

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Hoechst dye efflux reveals a novel CD7+CD34− lymphoid progenitor in human umbilical cord blood

Blood ◽

10.1182/blood.v96.6.2125 ◽

2000 ◽

Vol 96 (6) ◽

pp. 2125-2133 ◽

Cited By ~ 101

Author(s):

Robert W. Storms ◽

Margaret A. Goodell ◽

Alan Fisher ◽

Richard C. Mulligan ◽

Clay Smith

Keyword(s):

Stem Cells ◽

Cord Blood ◽

Umbilical Cord ◽

Umbilical Cord Blood ◽

Large Population ◽

Lymphoid Cells ◽

Human Umbilical Cord Blood ◽

Human Umbilical Cord ◽

Hematopoietic Stem ◽

Cd34 Cells

Abstract A novel Hoechst 33342 dye efflux assay was recently developed that identifies a population of hematopoietic cells termed side population (SP) cells. In the bone marrow of multiple species, including mice and primates, the SP is composed primarily of CD34−cells, yet has many of the functional properties of hematopoietic stem cells (HSCs). This report characterizes SP cells from human umbilical cord blood (UCB). The SP in unfractionated UCB was enriched for CD34+ cells but also contained a large population of CD34− cells, many of which were mature lymphocytes. SP cells isolated from UCB that had been depleted of lineage-committed cells (Lin− UCB) contained CD34+ and CD34− cells in approximately equivalent proportions. Similar to previous descriptions of human HSCs, the CD34+Lin− SP cells were CD38dimHLA-DRdimThy-1dimCD45RA−CD71−and were enriched for myelo-erythroid precursors. In contrast, the CD34−Lin− SP cells were CD38−HLA-DR−Thy-1−CD71−and failed to generate myelo-erythroid progeny in vitro. The majority of these cells were CD7+CD11b+CD45RA+, as might be expected of early lymphoid cells, but did not express other lymphoid markers. The CD7+CD34−Lin− UCB SP cells did not proliferate in simple suspension cultures but did differentiate into natural killer cells when cultured on stroma with various cytokines. In conclusion, the human Lin− UCB SP contains both CD34+ multipotential stem cells and a novel CD7+CD34−Lin− lymphoid progenitor. This observation adds to the growing body of evidence that CD34− progenitors exist in humans.

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High Integrity and Fidelity of Long-Term Cryopreserved Umbilical Cord Blood for Transplantation

Journal of Clinical Medicine ◽

10.3390/jcm10020293 ◽

2021 ◽

Vol 10 (2) ◽

pp. 293

Author(s):

Gee-Hye Kim ◽

Jihye Kwak ◽

Sung Hee Kim ◽

Hee Jung Kim ◽

Hye Kyung Hong ◽

...

Keyword(s):

Stem Cell ◽

Cord Blood ◽

Umbilical Cord ◽

Umbilical Cord Blood ◽

Clinical Applications ◽

Hematopoietic Stem ◽

Cd34 Cells ◽

Hsc Transplantation

Umbilical cord blood (UCB) is used as a source of donor cells for hematopoietic stem cell (HSC) transplantation. The success of transplantation is dependent on the quality of cord blood (CB) units for maximizing the chance of engraftment. Improved outcomes following transplantation are associated with certain factors of cryopreserved CB units: total volume and total nucleated cell (TNC) count, mononuclear cell (MNC) count, and CD34+ cell count. The role of the storage period of CB units in determining the viability and counts of cells is less clear and is related to the quality of cryopreserved CB units. Herein, we demonstrate the recovery of viable TNCs and CD34+ cells, as well as the MNC viability in 20-year-old cryopreserved CB units in a CB bank (MEDIPOST Co., Ltd., Seongnam-si, Gyeonggi-do, Korea). In addition, cell populations in CB units were evaluated for future clinical applications. The stable recovery rate of the viability of cryopreserved CB that had been stored for up to 20 years suggested the possibility of uses of the long-term cryopreservation of CB units. Similar relationships were observed in the recovery of TNCs and CD34+ cells in units of cryopreserved and fresh CB. The high-viability recovery of long-term cryopreserved CB suggests that successful hematopoietic stem cell (HSC) transplantation and other clinical applications, which are suitable for treating incurable diseases, may be performed regardless of long-term storage.

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Transplantation Induces Profound Changes in the Transcriptional Asset of Hematopoietic Stem Cells: Identification of Specific Signatures Using Machine Learning Techniques

Journal of Clinical Medicine ◽

10.3390/jcm9061670 ◽

2020 ◽

Vol 9 (6) ◽

pp. 1670

Author(s):

Daniela Cilloni ◽

Jessica Petiti ◽

Valentina Campia ◽

Marina Podestà ◽

Margherita Squillario ◽

...

Keyword(s):

Machine Learning ◽

Bone Marrow ◽

Cord Blood ◽

Umbilical Cord ◽

Umbilical Cord Blood ◽

Machine Learning Techniques ◽

Specific Gene ◽

Self Renewal ◽

Hematopoietic Stem ◽

Cd34 Cells

During the phase of proliferation needed for hematopoietic reconstitution following transplantation, hematopoietic stem/progenitor cells (HSPC) must express genes involved in stem cell self-renewal. We investigated the expression of genes relevant for self-renewal and expansion of HSPC (operationally defined as CD34+ cells) in steady state and after transplantation. Specifically, we evaluated the expression of ninety-one genes that were analyzed by real-time PCR in CD34+ cells isolated from (i) 12 samples from umbilical cord blood (UCB); (ii) 15 samples from bone marrow healthy donors; (iii) 13 samples from bone marrow after umbilical cord blood transplant (UCBT); and (iv) 29 samples from patients after transplantation with adult hematopoietic cells. The results show that transplanted CD34+ cells from adult cells acquire an asset very different from transplanted CD34+ cells from cord blood. Multivariate machine learning analysis (MMLA) showed that four specific gene signatures can be obtained by comparing the four types of CD34+ cells. In several, but not all cases, transplanted HSPC from UCB overexpress reprogramming genes. However, these remarkable changes do not alter the commitment to hematopoietic lineage. Overall, these results reveal undisclosed aspects of transplantation biology.

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Enhanced self-renewal of human long-term hematopoietic stem cells by a sulfamoyl benzoate derivative targeting p18INK4C

Blood Advances ◽

10.1182/bloodadvances.2020004054 ◽

2021 ◽

Vol 5 (17) ◽

pp. 3362-3372

Author(s):

Yinghui Li ◽

Wenshan Zhang ◽

Yu Zhang ◽

Yahui Ding ◽

Ming Yang ◽

...

Keyword(s):

Cord Blood ◽

Progenitor Cells ◽

Umbilical Cord ◽

Umbilical Cord Blood ◽

Kinase Inhibitor ◽

Ex Vivo ◽

The Self ◽

Self Renewal ◽

Hematopoietic Stem

Abstract The use of umbilical cord blood transplant has been substantially limited by the finite number of hematopoietic stem and progenitor cells in a single umbilical cord blood unit. Small molecules that not only quantitatively but also qualitatively stimulate enhancement of hematopoietic stem cell (HSC) self-renewal ex vivo should facilitate the clinical use of HSC transplantation and gene therapy. Recent evidence has suggested that the cyclin-dependent kinase inhibitor, p18INK4C (p18), is a critical regulator of mice HSC self-renewal. The role of p18 in human HSCs and the effect of p18 inhibitor on human HSC expansion ex vivo need further studies. Here we report that knockdown of p18 allowed for an increase in long-term colony-forming cells in vitro. We then identified an optimized small molecule inhibitor of p18, 005A, to induce ex vivo expansion of HSCs that was capable of reconstituting human hematopoiesis for at least 4 months in immunocompromised mice, and hence, similarly reconstituted secondary recipients for at least 4 more months, indicating that cells exposed to 005A were still competent in secondary recipients. Mechanistic studies showed that 005A might delay cell division and activate both the Notch signaling pathway and expression of transcription factor HoxB4, leading to enhancement of the self-renewal of long-term engrafting HSCs and the pool of progenitor cells. Taken together, these observations support a role for p18 in human HSC maintenance and that the p18 inhibitor 005A can enhance the self-renewal of long-term HSCs.

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Hoechst dye efflux reveals a novel CD7+CD34− lymphoid progenitor in human umbilical cord blood

Blood ◽

10.1182/blood.v96.6.2125.h8002125_2125_2133 ◽

2000 ◽

Vol 96 (6) ◽

pp. 2125-2133 ◽

Cited By ~ 1

Author(s):

Robert W. Storms ◽

Margaret A. Goodell ◽

Alan Fisher ◽

Richard C. Mulligan ◽

Clay Smith

Keyword(s):

Stem Cells ◽

Cord Blood ◽

Umbilical Cord ◽

Umbilical Cord Blood ◽

Large Population ◽

Lymphoid Cells ◽

Human Umbilical Cord Blood ◽

Human Umbilical Cord ◽

Hematopoietic Stem ◽

Cd34 Cells

A novel Hoechst 33342 dye efflux assay was recently developed that identifies a population of hematopoietic cells termed side population (SP) cells. In the bone marrow of multiple species, including mice and primates, the SP is composed primarily of CD34−cells, yet has many of the functional properties of hematopoietic stem cells (HSCs). This report characterizes SP cells from human umbilical cord blood (UCB). The SP in unfractionated UCB was enriched for CD34+ cells but also contained a large population of CD34− cells, many of which were mature lymphocytes. SP cells isolated from UCB that had been depleted of lineage-committed cells (Lin− UCB) contained CD34+ and CD34− cells in approximately equivalent proportions. Similar to previous descriptions of human HSCs, the CD34+Lin− SP cells were CD38dimHLA-DRdimThy-1dimCD45RA−CD71−and were enriched for myelo-erythroid precursors. In contrast, the CD34−Lin− SP cells were CD38−HLA-DR−Thy-1−CD71−and failed to generate myelo-erythroid progeny in vitro. The majority of these cells were CD7+CD11b+CD45RA+, as might be expected of early lymphoid cells, but did not express other lymphoid markers. The CD7+CD34−Lin− UCB SP cells did not proliferate in simple suspension cultures but did differentiate into natural killer cells when cultured on stroma with various cytokines. In conclusion, the human Lin− UCB SP contains both CD34+ multipotential stem cells and a novel CD7+CD34−Lin− lymphoid progenitor. This observation adds to the growing body of evidence that CD34− progenitors exist in humans.

Download Full-text