scholarly journals Short- and Long-Term Multilineage Repopulating Hematopoietic Stem Cells in Late Fetal and Newborn Mice: Models for Human Umbilical Cord Blood

Blood ◽  
1997 ◽  
Vol 90 (1) ◽  
pp. 174-181 ◽  
Author(s):  
David E. Harrison ◽  
Clinton M. Astle
Keyword(s):  
Stem Cells ◽  
Cord Blood ◽  
Umbilical Cord Blood ◽  
Functional Abilities ◽  
Newborn Mice ◽  
Hematopoietic Stem ◽  
Short And Long Term ◽  
Marrow Cells

Abstract Blood from late fetal and newborn mice is similar to umbilical cord blood obtained at birth in human beings, an important source of stem cells for clinical transplantation. The mouse model is useful because long-term functions can be readily assayed in vivo. To evaluate the functions of hematopoietic precursors in the blood and other tissues of late fetal and newborn mice, short- and long-term multilineage repopulating abilities were measured in vivo by competitive repopulation. Manipulations that might affect cell function, such as enrichment, tissue culture, or retroviral marking, were avoided. Hematopoietic stem cell functions of late fetal or newborn blood, liver, and spleen, were assayed as myeloid and lymphoid repopulating abilities relative to standard adult marrow cells. Donor cells from these tissues as well as adult control donor marrow cells were all of the same genotype. Cells from each donor tissue were mixed with portions from a pool of standard adult “competitor” marrow distinguished from the donors by genetic differences in hemoglobin and glucosephosphate isomerase. After 21 to 413 days, percentages of donor type myeloid and lymphoid cells in recipient blood were measured to assay the functional abilities of donor precursors relative to the standard. These relative measures are expressed as repopulating units, where each unit is equivalent to the repopulating ability found in 100,000 standard adult marrow cells. Thus, measures of repopulating units do not compare single cells but overall repopulating abilities of donor cell populations. Relative functional abilities in 1 million nucleated cells from late fetal or newborn blood were several times less than those found in adult marrow, but far more than in normal adult blood, and appeared to include long-term functional primitive hematopoietic stem cells (PHSC) similar to those in marrow. To estimate functional abilities of individual PHSC, variances among large groups of identical recipients were analyzed using both the binomial model and competitive dilution, a new model based on the Poisson distribution. The data best fit the hypothesis that individual PHSC from adult marrow, late fetal blood, or newborn blood each produce similar fractions of the total lymphoid and erythroid cells found in the recipient for many months.

scholarly journals Short- and Long-Term Multilineage Repopulating Hematopoietic Stem Cells in Late Fetal and Newborn Mice: Models for Human Umbilical Cord Blood

Blood ◽  
1997 ◽  
Vol 90 (1) ◽  
pp. 174-181 ◽  
Author(s):  
David E. Harrison ◽  
Clinton M. Astle
Keyword(s):  
Stem Cells ◽  
Cord Blood ◽  
Umbilical Cord Blood ◽  
Functional Abilities ◽  
Newborn Mice ◽  
Hematopoietic Stem ◽  
Short And Long Term ◽  
Marrow Cells

Blood from late fetal and newborn mice is similar to umbilical cord blood obtained at birth in human beings, an important source of stem cells for clinical transplantation. The mouse model is useful because long-term functions can be readily assayed in vivo. To evaluate the functions of hematopoietic precursors in the blood and other tissues of late fetal and newborn mice, short- and long-term multilineage repopulating abilities were measured in vivo by competitive repopulation. Manipulations that might affect cell function, such as enrichment, tissue culture, or retroviral marking, were avoided. Hematopoietic stem cell functions of late fetal or newborn blood, liver, and spleen, were assayed as myeloid and lymphoid repopulating abilities relative to standard adult marrow cells. Donor cells from these tissues as well as adult control donor marrow cells were all of the same genotype. Cells from each donor tissue were mixed with portions from a pool of standard adult “competitor” marrow distinguished from the donors by genetic differences in hemoglobin and glucosephosphate isomerase. After 21 to 413 days, percentages of donor type myeloid and lymphoid cells in recipient blood were measured to assay the functional abilities of donor precursors relative to the standard. These relative measures are expressed as repopulating units, where each unit is equivalent to the repopulating ability found in 100,000 standard adult marrow cells. Thus, measures of repopulating units do not compare single cells but overall repopulating abilities of donor cell populations. Relative functional abilities in 1 million nucleated cells from late fetal or newborn blood were several times less than those found in adult marrow, but far more than in normal adult blood, and appeared to include long-term functional primitive hematopoietic stem cells (PHSC) similar to those in marrow. To estimate functional abilities of individual PHSC, variances among large groups of identical recipients were analyzed using both the binomial model and competitive dilution, a new model based on the Poisson distribution. The data best fit the hypothesis that individual PHSC from adult marrow, late fetal blood, or newborn blood each produce similar fractions of the total lymphoid and erythroid cells found in the recipient for many months.

CD45−/ALDHlow/SSEA-4+/Oct-4+/CD133+/CXCR4+/Lin− Very Small Embryonic-Like (VSEL) Stem Cells Isolated from Umbilical Cord Blood as Potential Long Term Repopulating Hematopoietic Stem Cells

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 2444-2444
Author(s):  
Ewa K Zuba-Surma ◽  
Magdalena Kucia ◽  
Rui Liu ◽  
Mariusz Z Ratajczak ◽  
Janina Ratajczak
Keyword(s):  
Stem Cells ◽  
Cord Blood ◽  
Umbilical Cord ◽  
Umbilical Cord Blood ◽  
Mononuclear Cells ◽  
Hematopoietic Stem ◽  
Hematopoietic Activity

Abstract Recently, we identified a population of very small embryonic-like (VSEL) stem cells in umbilical cord blood (CB) (Leukemia2007;21:297–303) These VSELs are: smaller than erythrocytes; SSEA-4+/Oct-4+/CD133+/CXCR4+/Lin−/CD45−; responsive to SDF-1 gradient; and iv) possessing large nuclei that contain unorganized chromatin (euchromatin). Data obtained in a murine model indicate that a similar cell population isolated from bone marrow (BM) does not reveal hematopoietic activity after isolation. However, in appropriate models (i.e., in vitro co-culture over OP-9 cells or in vivo after intra bone injection), these cells contribute to hematopoiesis and thus possesses potential of long term repopulating hematopoietic stem cells (LT-HSCs). To investigate the hematopoietic activity of CB-derived, CD45 negative VSELs, we employed staining with Aldefluor detecting aldehyde dehydrogenase (ALDH), the enzyme expressed in primitive hematopoietic cells. We sorted CD133+/CD45−/ALDHhigh and CD133+/CD45−/ALDHlow sub-fractions of VSELs from CB samples and established that both freshly sorted CB-derived populations did not grow hematopoietic colonies in vitro. However, when activated/expanded over OP-9 stroma cells, they exhibit hematopoietic potential and initiate hematopoietic colonies composed of CD45+ cells when replated into methylcellulose cultures. Furthermore, while CD133+/CD45−/ALDHhigh VSELs gave raise to hematopoietic colonies after the first replating, the formation of colonies by CD133+/CD45−/ALDHlow VSELs was delayed. The data indicate that both populations of CD45− cells may acquire hematopoietic potential; however hematopoietic specification is delayed for CD133+/CD45−/ALDHlow cells (Fig. 1A). In parallel, real time PCR analysis revealed that freshly isolated CD133+/CD45−/ALDHhigh VSELs express more hematopoietic transcripts (e.g., c-myb, 80.2±27.4 fold difference) while CD133+/CD45−/ALDHlow exhibit higher levels of pluripotent stem cell markers (e.g., Oct-4, 119.5±15.5 fold difference) as compared to total CB mononuclear cells (Fig. B). Furthermore and somewhat unexpectedly, we found that because of their unusually small size, these important cells may be partially depleted (in 42.5±12.6%) during standard preparation strategies of CB units for storage that employ volume reduction. In conclusion, our data suggest very small CB mononuclear cells expressing VSEL markers that are CD133+/CD45−/ALDHlow are highly enriched for the most primitive population of LT-HSCs. These cells may be responsible for long term CB engraftment and be a population of cells from which HSCs should be expanded. We are currently testing this in an in vivo model by performing heterotransplants of CD45− ALDHlow VSELs into immunodeficient mice. It is important to stress that currently employed, routine CB processing strategies may lead up to ~50% loss of these small cells that are endowed with such remarkable hematopoietic activity. Figure Figure

scholarly journals Hematopoietic stem/progenitor cells, generation of induced pluripotent stem cells, and isolation of endothelial progenitors from 21- to 23.5-year cryopreserved cord blood

Blood ◽  
2011 ◽  
Vol 117 (18) ◽  
pp. 4773-4777 ◽  
Author(s):  
Hal E. Broxmeyer ◽  
Man-Ryul Lee ◽  
Giao Hangoc ◽  
Scott Cooper ◽  
Nutan Prasain ◽  
...  
Keyword(s):  
Stem Cells ◽  
Cord Blood ◽  
Progenitor Cells ◽  
Proliferative Potential ◽  
Hematopoietic Stem ◽  
Immunodeficient Mice ◽  
Induced Pluripotent

Abstract Cryopreservation of hematopoietic stem cells (HSCs) and hematopoietic progenitor cells (HPCs) is crucial for cord blood (CB) banking and transplantation. We evaluated recovery of functional HPC cryopreserved as mononuclear or unseparated cells for up to 23.5 years compared with prefreeze values of the same CB units. Highly efficient recovery (80%-100%) was apparent for granulocyte-macrophage and multipotential hematopoietic progenitors, although some collections had reproducible low recovery. Proliferative potential, response to multiple cytokines, and replating of HPC colonies was extensive. CD34+ cells isolated from CB cryopreserved for up to 21 years had long-term (≥ 6 month) engrafting capability in primary and secondary immunodeficient mice reflecting recovery of long-term repopulating, self-renewing HSCs. We recovered functionally responsive CD4+ and CD8+ T lymphocytes, generated induced pluripotent stem (iPS) cells with differentiation representing all 3 germ cell lineages in vitro and in vivo, and detected high proliferative endothelial colony forming cells, results of relevance to CB biology and banking.

Hematopoietic stem cells express Tie-2 receptor in the murine fetal liver

Blood ◽  
2000 ◽  
Vol 96 (12) ◽  
pp. 3757-3762 ◽  
Author(s):  
Hsiang-Chun Hsu ◽  
Hideo Ema ◽  
Mitsujiro Osawa ◽  
Yukio Nakamura ◽  
Toshio Suda ◽  
...  
Keyword(s):  
Stem Cells ◽  
Hematopoietic Stem Cells ◽  
Receptor Tyrosine Kinase ◽  
Fetal Liver ◽  
Newborn Mice ◽  
Hematopoietic Stem ◽  
Fetal Liver Cells

Tie-2 receptor tyrosine kinase expressed in endothelial and hematopoietic cells is believed to play a role in both angiogenesis and hematopoiesis during development of the mouse embryo. This article addressed whether Tie-2 is expressed on fetal liver hematopoietic stem cells (HSCs) at day 14 of gestation. With the use of anti–Tie-2 monoclonal antibody, its expression was detected in approximately 7% of an HSC population of Kit-positive, Sca-1–positive, lineage-negative or -low, and AA4.1-positive (KSLA) cells. These Tie-2–positive KSLA (T+ KSLA) cells represent 0.01% to 0.02% of fetal liver cells. In vitro colony and in vivo competitive repopulation assays were performed for T+ KSLA cells and Tie-2–negative KSLA (T− KSLA) cells. In the presence of stem cell factor, interleukin-3, and erythropoietin, 80% of T+ KSLA cells formed colonies in vitro, compared with 40% of T− KSLA cells. Long-term multilineage repopulating cells were detected in T+ KSLA cells, but not in T− KSLA cells. An in vivo limiting dilution analysis revealed that at least 1 of 8 T+ KSLA cells were such repopulating cells. The successful secondary transplantation initiated with a limited number of T+ KSLA cells suggests that these cells have self-renewal potential. In addition, engraftment of T+ KSLA cells in conditioned newborn mice indicates that these HSCs can be adapted equally by the adult and newborn hematopoietic environments. The data suggest that T+ KSLA cells represent HSCs in the murine fetal liver.

Hematopoietic stem cells express Tie-2 receptor in the murine fetal liver

Blood ◽  
2000 ◽  
Vol 96 (12) ◽  
pp. 3757-3762 ◽  
Author(s):  
Hsiang-Chun Hsu ◽  
Hideo Ema ◽  
Mitsujiro Osawa ◽  
Yukio Nakamura ◽  
Toshio Suda ◽  
...  
Keyword(s):  
Stem Cells ◽  
Hematopoietic Stem Cells ◽  
Receptor Tyrosine Kinase ◽  
Fetal Liver ◽  
Newborn Mice ◽  
Hematopoietic Stem ◽  
Fetal Liver Cells

Abstract Tie-2 receptor tyrosine kinase expressed in endothelial and hematopoietic cells is believed to play a role in both angiogenesis and hematopoiesis during development of the mouse embryo. This article addressed whether Tie-2 is expressed on fetal liver hematopoietic stem cells (HSCs) at day 14 of gestation. With the use of anti–Tie-2 monoclonal antibody, its expression was detected in approximately 7% of an HSC population of Kit-positive, Sca-1–positive, lineage-negative or -low, and AA4.1-positive (KSLA) cells. These Tie-2–positive KSLA (T+ KSLA) cells represent 0.01% to 0.02% of fetal liver cells. In vitro colony and in vivo competitive repopulation assays were performed for T+ KSLA cells and Tie-2–negative KSLA (T− KSLA) cells. In the presence of stem cell factor, interleukin-3, and erythropoietin, 80% of T+ KSLA cells formed colonies in vitro, compared with 40% of T− KSLA cells. Long-term multilineage repopulating cells were detected in T+ KSLA cells, but not in T− KSLA cells. An in vivo limiting dilution analysis revealed that at least 1 of 8 T+ KSLA cells were such repopulating cells. The successful secondary transplantation initiated with a limited number of T+ KSLA cells suggests that these cells have self-renewal potential. In addition, engraftment of T+ KSLA cells in conditioned newborn mice indicates that these HSCs can be adapted equally by the adult and newborn hematopoietic environments. The data suggest that T+ KSLA cells represent HSCs in the murine fetal liver.

Identification of a Hierarchy of Multipotent Hematopoietic Progenitors in Human Cord Blood.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 2237-2237
Author(s):  
Ravindra Majeti ◽  
Christopher Y. Park ◽  
Irving L. Weissman
Keyword(s):  
Bone Marrow ◽  
Cord Blood ◽  
Human Cord Blood ◽  
Self Renewal ◽  
Newborn Mice ◽  
Hematopoietic Stem ◽  
Multipotent Progenitors

Abstract Mouse hematopoiesis is initiated by long-term hematopoietic stem cells (HSC) that differentiate into a series of multipotent progenitors that exhibit progressively diminished self-renewal ability. In human hematopoiesis, populations enriched for HSC have been identified, as have downstream lineage-committed progenitors, but not multipotent progenitors. Previous reports indicate that human HSC are enriched in Lin-CD34+CD38- cord blood and bone marrow, and express CD90. We demonstrate that the Lin-CD34+CD38- fraction of cord blood and bone marrow can be subdivided into three subpopulations: CD90+CD45RA-, CD90-CD45RA-, and CD90-CD45RA+. While, the function of the CD90- subpopulations is unknown, the CD90+CD45RA- subpopulation presumably contains HSC. We report here in vitro and in vivo functional studies of these three subpopulations from normal human cord blood. In vitro, CD90+CD45RA- cells formed all types of myeloid colonies in methylcellulose and were able to replate with 70% efficiency. CD90-CD45RA- cells also formed all types of myeloid colonies, but replated with only 33% efficiency. CD90-CD45RA+ cells failed to form myeloid colonies in methylcellulose. In liquid culture, CD90+CD45RA- cells gave rise to all three subpopulations; CD90-CD45RA- cells gave rise to both CD90- subpopulations, but not CD90+ cells; CD90-CD45RA+ cells gave rise to themselves only. These data establish an in vitro differentiation hierarchy from CD90+CD45RA- to CD90-CD45RA- to CD90-CD45RA+ cells among Lin-CD34+CD38- cord blood. In vivo, xenotransplantation of CD90+CD45RA- cells into NOD/SCID/IL-2R?-null newborn mice resulted in long-term multilineage engraftment with transplantation of as few as 10 purified cells. Secondary transplants from primary engrafted mice also resulted in long-term multilineage engraftment, indicating the presence of self-renewing HSC. Transplantation of CD90-CD45RA- cells also resulted in long-term multilineage engraftment; however, secondary transplants did not reliably result in long-term engraftment, indicating a reduced capacity for self-renewal. Transplantation of CD90-CD45RA+ cells did not result in any detectable human hematopoietic cells, indicating that the function of these cells is undetermined. Finally, transplantation of limiting numbers of CD90-CD45RA- cells (less than 100) resulted in multilineage human engraftment at 4 weeks, that was no longer detectable by 12 weeks. Thus, the CD90-CD45RA- subpopulation is capable of multilineage differentiation while exhibiting limited self-renewal ability. We believe this study represents the first prospective identification of a population of human multipotent progenitors, Lin-CD34+CD38-CD90-CD45RA- cord blood.

In Vitro and In Vivo Evidence That Umbilical Cord Blood (UCB)-Derived CD45-/SSEA-4+/OCT-4+/CD133+/CXCR4+/Lin- Very Small Embryonic/Epiblast Like Stem Cells (VSELs) Do Not Contain Clonogenic Hematopoietic Progenitors but Are Highly Enriched in More Primitive Stem Cells - Novel View On Hierarchy of UCB Stem Cell Compartment.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 35-35 ◽  
Author(s):  
Ewa K. Zuba-Surma ◽  
Izabela Klich ◽  
Marcin Wysoczynski ◽  
Nicholas J Greco ◽  
Mary J. Laughlin ◽  
...  
Keyword(s):  
Stem Cells ◽  
Stem Cell ◽  
Hematopoietic Stem Cells ◽  
Cord Blood ◽  
Umbilical Cord ◽  
Umbilical Cord Blood ◽  
Hematopoietic Stem ◽  
Hematopoietic Activity

Abstract Abstract 35 Recently, we identified in umbilical cord blood (UCB) a population of very small embryonic/epiblast-like (VSEL) stem cells (Leukemia 2007;21:297–303) that are i) smaller than erythrocytes, ii) SSEA-4+/Oct-4+/CD133+/CXCR4+/Lin−/CD45−, iii) respond to SDF-1 gradient and iv) possess large nuclei containing primitive euchromatin. We have demonstrated in vitro that UCB-derived VSELs did not reveal hematopoietic activity freshly after isolation, but grow hematopoietic colonies following co-culture/activation over OP-9 cells. To investigate the hierarchy of UCB-derived, CD45 negative VSELs, we employed staining with Aldefluor - detecting aldehyde dehydrogenase (ALDH), the enzyme expressed in primitive hematopoietic cells. Subsequently, we sorted CD45−/CD133+/ALDHhigh and CD45−/CD133+/ALDHlow sub-fractions of VSELs from UCB samples and established that freshly sorted from UCB VSELs in contrast to sorted CD45+/ CD133+/ALDHhigh and CD45+/CD133+/ALDHlow hematopoietic stem cells (HSC) did not grow colonies in vitro. However, when CD45− VSELs were activated/expanded over OP-9 stroma cells, they exhibit hematopoietic potential and grew in routine methylcellulose cultures hematopoietic colonies composed of CD45+ cells. Interestingly, while CD45−/CD133+/ALDHhigh VSELs gave raise to hematopoietic colonies after the first replating, the formation of colonies by CD45−/CD133+/ALDHlow VSELs was somehow delayed, what suggest that they needed more time to acquire hematopoietic commitment. Thus our in vitro data indicate that both populations of CD45− cells may acquire hematopoietic potential; however hematopoietic specification is delayed for CD45−/CD133+/ALDHlow cells, suggesting their more primitive nature. In parallel, real time PCR analysis confirmed that while freshly isolated CD45−/CD133+/ALDHhigh VSELs express more hematopoietic transcripts (e.g., c-myb, 80.2±27.4 fold difference), CD45−/CD133+/ALDHlow exhibit higher levels of pluripotent stem cell markers (e.g., Oct-4, 119.5±15.5 fold difference as compared to total UCB mononuclear cells) (Figure 1 panel A). Next hematopoietic potential of UCB-derived VSELs was tested in vivo after transplantation into NOD/SCID mice (Figure 1 panel B and C). We noticed that both CD45−/CD133+/ALDHhigh and CD45−/CD133+/ALDHlow VSELs, give rise to human lympho-hematopoietic chimerism in lethally irradiated NOD/SCID mice as assayed 4–6 weeks after transplantation. The level of human hematopoietic CD45+ cells in murine peripheral blood (PB), bone marrow (BM) and spleen (SP) were comparable for both transplanted UCB-VSELs fractions - 7.1±2.9% (PB), 23.2±0.2% (SP) and 25.2±1.0% (BM). In conclusion, our data suggest that freshly isolated very small CD45 negative UCB-VSELs are depleted from clonogeneic progenitors, however they are highly enriched for primitive HSC. Based on our in vitro and in vivo data we postulate following hierarchy of hematopoietic stem cells in UCB (from most primitive to more differentiated) i) CD45−/CD133+/ALDHlow, ii) CD45−/CD133+/ALDHhigh , iii) CD45+/CD133+/ALDHlow and iv) CD45−/CD133+/ALDHhigh. We also postulate that as we have already shown for murine BM-derived VSELs, human UCB-derived CD45 negative VSELs correspond to a population of most primitive long term repopulating HSC (LT-HSC). Of note, we also found that currently employed, routine UCB processing strategies may lead up to ∼50% unwanted loss of these small cells that are endowed with such remarkable hematopoietic activity! Disclosures: No relevant conflicts of interest to declare.

Expansion of Umbilical Cord Blood Hematopoietic Stem Cells for Clinical Use.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 4049-4049
Author(s):  
Rowayda Peters ◽  
Gregor Hutter ◽  
Adriano Aguzzi ◽  
Mathias Heikenwaelder ◽  
Christoph Renner ◽  
...  
Keyword(s):  
Stem Cells ◽  
Hematopoietic Stem Cells ◽  
Cord Blood ◽  
Cell Count ◽  
Umbilical Cord ◽  
Umbilical Cord Blood ◽  
Induction Medium ◽  
Adherent Cells ◽  
Hematopoietic Stem

Abstract Umbilical cord blood (UCB) is an alternative donor source for allogeneic hematopoietic stem cell transplantation. However, transplantation in adults is frequently limited by the small number of cells available in a unit. We have previously developed the technology to expand hematopoietic stem cells (HSC) and stromal/mesenchymal stem cells (SMSC) from all UCB frozen samples (13, 3). The incubation of thawed UCB mononuclear cells (MNC) in the presence of SCF (25ng/ml), FLt-3 (25ng/ml), MGDF (10ng/ml) & IL-6 (20ng/ml) and 10% human serum in stroma-free liquid culture not only generated long term expansion of transplantable UCB HSC (non-adherent). Also, long-term expansion of SMSC (adherent cells) was successful. In order to upscale the expansion to use for clinical applications, we analyzed 3 frozen UCB comparing the expansion from 24-well plates (previous) versus expansion in bags (VueLife™) after 10 and 14 days of culture. Results show substantial expansion in total cell count (TCC, 4.8, 10.9: 2.4, 3.8 fold) at d10 and d14 from wells & bags respectively. TCC increased further in the presence of SMSC (38% & 33% in CD34+ cell count cultured in wells). CD133+CD34+CD38- HSC multiplied (11, 25 fold, d10/d14 & colony forming cells (CFC) 19 fold at d14 bags respectively). Heterogeneous cell populations were detected after d 14 in bags: T and B -lymphoid (%CD3/%CD19; 65/4:50w/3), megakaryocytic (%CD61; 7:4) and myeloid (%CD33; 31:43) at d0/d14 respectively. Further, expanded cells (250,000) containing a small number of CD133+CD34+CD38- (15,000–30,000) were injected into the liver of sub-lethally irradiated newborn Balb/C Rag2-Cγ−/− mice. Our preliminary data show no engraftment from cells expanded in wells and bags after 6 weeks of transplantation from d10 cultures (human CD45 + <1%). However, positive engraftment in mouse PB was detected from cells expanded for 14 d (wells, 1.16–2.5% & bags, 1.21–3.9%) as compared to control mice (CD45; 30% PB & 70% BM) receiving selected CD34 + (300,000 CD34+ at d0). Primitive repopulating cells (PRC), and multilineage human CFC were detected after transplantation. On d14 of HSC expansion, UCB MNC derived adherent cells (SMSC) were enriched by trypsinization. SMSC were established in serum free and serum plus culture as well. The immunophenotype of harvested SMSC was CD29+, CD44+ and CD45−, CD34− and CD133− at percentages + >90%. Following repeated trypsinization, SMSC count increased 41–96 folds. CFU-Fibroblast colonies (92–173) were generated from 104 SMSC after 2 weeks in MesenCult medium. We have previously differentiated SMSC into hepatocytes. Now we also generated adipocytes in an induction medium containing, Insulin, dexamethasone & indomethacin. SMSC formed Oil-droplet vacuoles in the cytoplasm in 3 weeks. The culture conditions we defined to maintain UCB PRC should be developed clinically. SMSC described herein exhibit in vitro properties of multipotent stem cells.

scholarly journals Ex Vivo Expansion and Transplantation of Hematopoietic Stem/Progenitor Cells Supported by Mesenchymal Stem Cells from Human Umbilical Cord Blood

2007 ◽  
Vol 16 (6) ◽  
pp. 579-585 ◽  
Author(s):  
Guo-Ping Huang ◽  
Zhi-Jun Pan ◽  
Bing-Bing Jia ◽  
Qiang Zheng ◽  
Chun-Gang Xie ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Mesenchymal Stem Cells ◽  
Cord Blood ◽  
Progenitor Cells ◽  
Umbilical Cord Blood ◽  
Ex Vivo ◽  
Hematopoietic Stem ◽  
Cd34 Cells

Human mesenchymal stem cells (MSCs) are multipotential and are detected in bone marrow (BM), adipose tissue, placenta, and umbilical cord blood (UCB). In this study, we examined the ability of UCB-derived MSCs (UCB-MSCs) to support ex vivo expansion of hematopoietic stem/progenitor cells (HSPCs) from UCB and the engraftment of expanded HSPCs in NOD/SCID mice. The result showed that UCB-MSCs supported the proliferation and differentiation of CD34+ cells in vitro. The number of expanded total nucleated cells (TNCs) in MSC-based culture was twofold higher than cultures without MSC (control cultures). UCB-MSCs increased the expansion capabilities of CD34+ cells, long-term culture-initiating cells (LTC-ICs), granulocyte-macrophage colony-forming cells (GM-CFCs), and high proliferative potential colony-forming cells (HPP-CFCs) compared to control cultures. The expanded HSPCs were transplanted into lethally irradiated NOD/SCID mice to assess the effects of expanded cells on hematopoietic recovery. The number of white blood cells (WBCs) in the peripheral blood of mice transplanted with expanded cells from both the MSC-based and control cultures returned to pretreatment levels at day 25 posttransplant and then decreased. The WBC levels returned to pretreatment levels again at days 45–55 posttransplant. The level of human CD45+ cell engraftment in primary recipients transplanted with expanded cells from the MSC-based cultures was significantly higher than recipients transplanted with cells from the control cultures. Serial transplantation demonstrated that the expanded cells could establish long-term engraftment of hematopoietic cells. UCB-MSCs similar to those derived from adult bone marrow may provide novel targets for cellular and gene therapy.

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