scholarly journals Stem cells with short term and long term repopulating ability in the mouse

1996 ◽  
Vol 7 ◽  
pp. 15-18
Author(s):  
W.E. Fibbe ◽  
J. Mark ◽  
J.M. Zijlmans ◽  
R. Willemze
Keyword(s):  
Stem Cells ◽  
Short Term

Stability of Self-Renewal in Hematopoietic Stem Cells

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. SCI-42-SCI-42
Author(s):  
Norman N. Iscove
Keyword(s):  
Stem Cells ◽  
Hematopoietic Stem Cells ◽  
Conflicts Of Interest ◽  
Expression Profiles ◽  
Single Cells ◽  
Gene Expression Profiles ◽  
Short Term ◽  
Blood Leukocytes ◽  
Hematopoietic Stem

Abstract Abstract SCI-42 For many years a distinction was drawn between prospectively separable murine HSC populations with long-term, essentially permanent reconstituting potential (LT-HSC), versus HSC populations yielding short-term engraftment lasting only 4 – 6 weeks after transplantation (ST-HSC). Recent work based on transplantation of single cells shows that highly purified populations of LT-HSC prepared by standard sorting parameters consist in fact predominantly of a distinct, newly recognized class of intermediate- term reconstituting cells (IT-HSC) whose grafts endure longer than short-term HSC but also eventually fail (1). IT-HSC are separable from long-term reconstituting cells on the basis of expression of more alpha2 integrin and less SLAM150. Crucial to recognition of the distinction between LT- and IT-HSC are the endpoints used to evaluate reconstitution. If blood erythroid or myeloid reconstitution is measured, IT reconstitution is readily distinguished by the disappearance of these elements by 16 wk post-transplant. If instead reconstitution is measured simply by presence of blood leukocytes of donor origin, which in the mouse are almost entirely lymphocytes, the distinction is not made because lymphoid elements persist even in fading IT clones to 24 wk or beyond. The observations imply a need for reinterpretation of most of the published descriptions of the biology and gene expression profiles previously attributed to LT-HSC but in fact derived from analysis of populations that consisted mainly of IT-HSC. The capacity now to separate LT- from IT-HSC creates new opportunities for probing the mechanisms that specify and sustain long term function in the former but not the latter. 1. Benveniste P, Frelin C, Janmohamed S, Barbara M, Herrington R, Hyam D, Iscove NN. Intermediate-term hematopoietic stem cells with extended but time-limited reconstitution potential. Cell Stem Cell. 2010;6:48–58 Disclosures: No relevant conflicts of interest to declare.

scholarly journals Nerve growth factor is involved in the supportive effect by bone marrow- -derived stromal cells of the factor-dependent human cell line UT-7 [published erratum appears in Blood 1996 Oct 1;88(7):2818]

Blood ◽  
1996 ◽  
Vol 88 (5) ◽  
pp. 1608-1618 ◽  
Author(s):  
I Auffray ◽  
S Chevalier ◽  
J Froger ◽  
B Izac ◽  
W Vainchenker ◽  
...  
Keyword(s):  
Stem Cells ◽  
Stem Cell ◽  
Nerve Growth Factor ◽  
Growth Factor ◽  
Northern Blot ◽  
Stromal Cells ◽  
Synergistic Action ◽  
Short Term ◽  
Colony Assays

Abstract We previously demonstrated that murine MS-5 and SI/SI4 cell lines induce the proliferation of human factor-dependent UT-7 cells in the absence of normally required human cytokines and also stimulate the differentiation of CD34+/CD38-LTC-ICs. We report in this study that the effect of MS-5 cells on UT-7 cells can be completely explained by the synergistic action of nerve growth factor (NGF) and stem cell factor (SCF) produced by these murine stromal cells. Purified murine NGF was able to support short-term clone formation and long-term growth of UT-7 cells in suspension cultures as efficiently as rhu-granulocyte- macrophage colony-stimulating factor. NGF action was mediated through the TrkA receptor, in which messenger RNA (mRNA) was easily detected in UT-7 cells by Northern blot. MS-5 cells strongly expressed NGF mRNA in Northern blot, and direct implication of MS-5-derived NGF in the induction of UT-7 cells proliferation was demonstrated in inhibition assays with an anti-NGF monoclonal antibody (MoAb) that neutralized by 84% +/- 4.1% (n = 5) UT-7 clone formation. However, NGF did not act alone, and several arguments demonstrated the synergistic action of MS- 5-derived SCF: (1) an anti-c-kit partially inhibited UT-7 cells clone formation in coculture assays, (2) SCF and NGF synergized in an H3-TdR incorporation assay, and (3) the stimulatory effect of 10x-concentrated MS-5 supernatant was completely inhibited by an anti-c-kit but not by an anti-NGF, and levels of soluble NGF (1.2 ng/mL) detected by enzyme- linked immunosorbent assay in 10x supernatant of MS-5 cells cultures were below the biologically active concentrations. In contrast, although MS-5 cells also promoted the differentiation of very primitive CD34+/CD38- human stem cells both in colony assays and long-term cultures, we could not incriminate MS-5-derived NGF in the observed effect: an anti-NGF MoAb did not inhibit the synergistic effect of MS-5 cells in colony assays or long-term cultures nor did soluble muNGF duplicate MS-5 effect and survival of CD34+/CD38- clonogenic progenitor cells promoted by MS-5 was unaffected by an anti-NGF and was not induced by soluble NGF alone or combined with SCF. In contrast, NGF in synergy with SCF supported the short-term maintenance of high numbers of CD34+/CD38+ mature erythroid progenitors probably through an indirect mechanism implying macrophages. These results suggest that NGF, in which the primary target cells are outside the hematopoietic system, is present in the marrow environment and might act at some steps of hematopoietic stem cell development. These results also underline that the response of cell lines and normal stem cells to stromal cells is mediated by different pathways.

scholarly journals Hydroxyurea Can Be Used to Increase Mouse c-kit+Thy-1.1loLin−/loSca-1+ Hematopoietic Cell Number and Frequency in Cell Cycle In Vivo

Blood ◽  
1997 ◽  
Vol 90 (11) ◽  
pp. 4354-4362 ◽  
Author(s):  
Nobuko Uchida ◽  
Annabelle M. Friera ◽  
Dongping He ◽  
Michael J. Reitsma ◽  
Ann S. Tsukamoto ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Stem Cells ◽  
Bone Marrow ◽  
Cell Number ◽  
Short Term ◽  
Synthesis Inhibitor ◽  
Hematopoietic Stem ◽  
Cell Cycle Status

Abstract The DNA synthesis inhibitor hydroxyurea (HU) was administered to determine whether it induces changes in the cell-cycle status of primitive hematopoietic stem cells (HSCs)/progenitors. Administration of HU to mice leads to bone marrow accumulation of c-kit+Thy-1.1loLin−/loSca-1+ (KTLS) cells in S/G2/M phases of the cell cycle. HU is a relatively nontoxic, reversible cell-cycle agent that can lead to approximately a threefold expansion of KTLS cells in vivo and approximately an eightfold increase in the number of KTLS cells in S/G2/M. HSCs in HU-treated mice have undiminished multilineage long-term and short-term clonal reconstitution activity.

scholarly journals Pharmacologic Activation of Aldehyde Metabolism to Protect Hematopoietic Stem Cells (HSC) in Murine Models of Fanconi Anemia (FA)

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 105-105
Author(s):  
Jennifer Tsai ◽  
Kelsey R. Logas ◽  
Lauren D. Van Wassenhove ◽  
Beruh Dejene ◽  
Che-Hong Chen ◽  
...  
Keyword(s):  
Gene Expression ◽  
Stem Cells ◽  
Hematopoietic Stem Cells ◽  
Aldehyde Dehydrogenase ◽  
Principal Component ◽  
Patent Office ◽  
Aldehyde Dehydrogenase 2 ◽  
Short Term ◽  
Hematopoietic Stem

HSC loss in FA is due to failure to resolve DNA inter-strand crosslinks (ICL), which can be induced by reactive aldehydes, radiation, or other clastogenic agents. Aldehyde exposure may occur through environmental sources, e.g. ingestion, absorption, and inhalation, or endogenously as a byproduct of cellular metabolism. The ALDH2*2 genotype, a dominant-negative point mutation in the aldehyde dehydrogenase 2 (ALDH2) gene, causes the "Asian flushing syndrome" when ethanol (EtOH) is ingested, due to decreased metabolism of small aldehydes, particularly acetaldehyde. ALDH2*2 is a disease modifier in FA, causing more rapid bone marrow failure and earlier leukemia onset in doubly affected children. Similarly, mice experimentally doubly knocked out for FANCD2 and ALDH2 demonstrate increased HSC loss, which is accelerated by EtOH exposure. To reduce aldehyde exposure, we developed a small molecule ALDH activator, Alda-1, which increases the enzymatic activity of both wild type (WT) and mutant ALDH2. We hypothesized that DNA damage and HSC loss in FA would be prevented by reduction of the aldehyde load. To test the effects of Alda-1 mediated ALDH2 activation, we generated a novel murine FA model with FANCD2 KO and knock-in of the ALDH2*2 mutation into the murine locus. The FANCD2-/- ALDH2*1/*2 genetic model and parental controls were then tested after exogenous aldehydic challenge and/or therapeutic intervention with Alda-1. Increased aldehydic load was experimentally induced by EtOH administration 10 mg/kg/day IP, while Alda-1 10 ug/kg/day was continuously administered via osmotic pump. For each of these conditions, marrow was analyzed for HSC and progenitor cell (HSPC) number, HSC gene expression, and function. The importance of the altered aldehyde metabolism due to ALDH2*2 genotype was demonstrated by progressive loss of HSPC in ALDH2*2/*2 and FANCD2-/- ALDH2*1/*2 mice, e.g., 5-fold and 2-fold decline in long-term HSC (LT-HSC), respectively, by 36 weeks. Experimental EtOH challenge to increase the aldehyde load precipitously decreased HSC numbers of all genotypes. After 5 weeks of EtOH challenge, LT-HSC decreased 35-fold in FANCD2-/- ALDH2*1/*2, 12.5-fold in FANCD2-/-ALDH2*1/*1, and 10.5-fold in WT mice. Long-term Alda-1 treatment to decrease aldehydic load rescued FA mice from HSC loss. After 7 months of Alda-1 treatment, LT-HSC numbers in FANCD2-/-ALDH2*1/*2 and FANCD2-/-ALDH2*1/*1 were approximately 10-fold higher than untreated controls. There were no clinically observed adverse effects. Aldehyde exposure and Alda-1 treatment altered gene expression of HSC. Principal component analysis and clustering of HSC gene expression showed that the first principal component representing 40% of the variation in gene expression could be attributed to increased aldehydic load, either genetically (ALDH2*2 genotype) or environmentally (EtOH administration) induced, while Alda-1 treatment obviated these effects. HSC from Alda-1 treated mice clustered with those from control WT mice. To test whether Alda-1 improved HSC function as well as phenotypic number, engraftment potential was assessed with competitive repopulation assays of sorted HSC from congenic untreated donors vs short-term (3 weeks) Alda-1 treated donors. HSC from Alda-1 treated mice had 2-4 fold greater granulocyte repopulating capacity than those from the untreated donors. Our results demonstrate that Alda-1 treatment rescues HSC from aldehyde induced loss, whether from genetic variation (FANCD2- and/or ALDH2*2) or experimental challenge (EtOH administration). Furthermore, Alda-1 treatment prevents the abnormal HSC gene expression induced by increased aldehydic load. HSC function is improved by Alda-1 with greater repopulating capacity observed even after short-term treatment. These preclinical experiments provide compelling proof-of-concept that sustained activation of ALDH2 can prevent HSC loss in FA. Potential applications include long-term administration to prevent the development of marrow failure or leukemia, and HSC expansion to increase the number of cells available for gene therapy with autologous HSC. Our results suggest that a clinical trial of ALDH2 activation in FA patients to prevent marrow failure is warranted. Disclosures Van Wassenhove: U.S. Patent Office: Patents & Royalties: patent pending - submitted for ALDH2 activators to expand hematopoietic stem cells. Chen:Foresee Pharmaceuticals: Patents & Royalties: patents licensed to Foresee related to compounds that activate aldehyde dehydrogenase 2 (ALDH2) and correct the dysfunction in ALDH2*2; U.S. Patent Office: Patents & Royalties: patent pending - submitted for aldehyde dehydrogenase 2 (ALDH2) activators to expand hematopoietic stem cells. Mochly-Rosen:Foresee Pharmaceuticals: Membership on an entity's Board of Directors or advisory committees, Patents & Royalties: patents licensed to Foresee related to compounds that activate aldehyde dehydrogenase 2 (ALDH2) and correct the dysfunction in ALDH2*2; U.S. Patent Office: Patents & Royalties: patent pending - submitted for aldehyde dehydrogenase 2 (ALDH2) activators to expand hematopoietic stem cells. Weinberg:U.S. Patent Office: Patents & Royalties: patent pending - submitted for aldehyde dehydrogenase 2 (ALDH2) activators to expand hematopoietic stem cells.

Lymphoid-Restricted Development From Multipotent Candidate Murine Stem Cells: Distinct and Complimentary Functions of the c-kit and flt3-Ligands

Blood ◽  
1999 ◽  
Vol 94 (11) ◽  
pp. 3781-3790 ◽  
Author(s):  
Ole Johan Borge ◽  
Jörgen Adolfsson ◽  
Annica Mårtensson, Inga-Lill Mårtensson, and Sten E.W. Jacobsen
Keyword(s):  
Stem Cells ◽  
Cell Formation ◽  
Stem Cell Population ◽  
Tyrosine Kinase Receptors ◽  
Short Term ◽  
Interleukin 7 ◽  
Potential Interactions

Abstract The two tyrosine kinase receptors, c-kit and flt3, and their respective ligands KL and FL, have been demonstrated to play key and nonredundant roles in regulating the earliest events in hematopoiesis. However, their precise roles and potential interactions in promoting early lymphoid commitment and development remain unclear. Here we show that most if not all murine Lin−/loSca1+c-kit+ bone marrow (BM) cells generating B220+CD19+proB-cells in response to FL and interleukin-7 (IL-7) also have a myeloid potential. In contrast to FL + IL-7, KL + IL-7 could not promote proB-cell formation from Lin−/loSca1+c-kit+ cells. However, KL potently enhanced FL + IL-7–stimulated proB-cell formation, in part through enhanced recruitment of FL + IL-7–unresponsive Lin−/loSca1+c-kit+progenitors, and in part by enhancing the growth of proB-cells. The enhanced recruitment (4-fold) in response to KL occurred exclusively from the Lin−/loSca1+c-kit+flt3−long-term repopulating stem cell population, whereas KL had no effect on FL + IL-7–stimulated recruitment of Lin−/loSca1+c-kit+flt3+short-term repopulating cells. The progeny of FL + IL-7–stimulated Lin−/loSca1+c-kit+ cells lacked in vitro and in vivo myeloid potential, but efficiently reconstituted both B and T lymphopoiesis. In agreement with this FL, but not KL, efficiently induced expression of B220 and IL-7 receptor- on Lin−/loSca1+c-kit+flt3+cells. Thus, whereas KL appears crucial for recruitment of FL + IL-7–unresponsive candidate (c-kit+flt3−) murine stem cells, FL is essential and sufficient for development toward lymphoid restricted progenitors from a population of (c-kit+flt3+) multipotent short-term reconstituting progenitors.

scholarly journals Partially differentiated ex vivo expanded cells accelerate hematologic recovery in myeloablated mice transplanted with highly enriched long- term repopulating stem cells

Blood ◽  
1996 ◽  
Vol 88 (9) ◽  
pp. 3642-3653 ◽  
Author(s):  
SJ Szilvassy ◽  
KP Weller ◽  
B Chen ◽  
CA Juttner ◽  
A Tsukamoto ◽  
...  
Keyword(s):  
Stem Cells ◽  
Blood Cells ◽  
Ex Vivo ◽  
Granulocyte Colony Stimulating Factor ◽  
Short Term ◽  
Hematopoietic Stem ◽  
Colony Forming Units ◽  
Sustained Recovery ◽  
Stimulating Factor

The ability of an infusion of ex vivo expanded hematopoietic cells to ameliorate cytopenia following transplantation of hematopoietic stem cells (HSCs) is controversial. To address this issue, we measured the recovery of circulating leukocytes, erythrocytes, and platelets in lethally irradiated mice transplanted with 10(3) enriched HSCs, with or without their expanded equivalent (EE) generated after 7 days of culture in interleukin-3 (IL-3), IL-6, granulocyte colony-stimulating factor and Steel Factor. Two HSC populations differing in their content of short-term repopulating progenitors were evaluated. Thy-1loLIN-Sca- 1+ (TLS) bone marrow (BM) is enriched in colony-forming cells (CFCs), day 8 and day 12 spleen colony-forming units (CFU-S) (435 +/- 19, 170 +/- 30, and 740 +/- 70 per 10(3) cells, respectively), and stem cells with competitive long-term repopulating potential (> or = 1 per 43 cells). Thy-1loSca-1+H-2Khl cells (TSHFU) isolated from BM 1 day after treatment of donor mice with 5-fluorouracil (5-FU) are also highly enriched in competitive repopulating units (CRU, > or = 1 per 55 cells), but are depleted of CFCs, day 8 and day 12 CFU-S (171 +/- 8, 0 and 15 +/- 4 per 10(3) cells, respectively). Recipients of 10(3) TLS cells transiently recovered leukocytes to > or = 2,000/microL in 12 days, but sustained engraftment required 25 days. Platelets recovered to > or = 200,000/microL in 15 days, and erythrocytes never decreased below 50% of normal. Mice transplanted with 10(3) TSHFU cells recovered leukocytes in 15 days, and platelets and erythrocytes in 18 days. Recipients of unseparated normal or 5-FU-treated BM cells (containing 10(3) TLS or TSHFU cells) recovered safe levels of blood cells in 9 to 12 days, suggesting that unseparated marrow contains early engrafting cells that were depleted by sorting. Upon ex vivo expansion, total cells, CFCs and day 12 CFU-S were amplified 2,062-,83- and 13-fold, respectively, from TLS cells; and 1,279-, 259- and 708-fold, respectively, from TSHFU cells. Expanded cells could regenerate the majority of lymphocytes and granulocytes in primary (17 weeks) and secondary (26 weeks) hosts and were only moderately impaired compared to fresh HSCs. The EE of TSHFU cells was more potent than that of TLS cells, suggesting that more highly enriched HSCs are more desirable starting populations for this application. When mice were transplanted with 10(3) TSHFU cells and their EE, the duration of thrombocytopenia was shortened from 18 to 12 days, and anemia was abolished. Leukocytes were also elevated on days 9 to 12, although sustained recovery was not accelerated. Anemia was also abrogated in recipients of 10(3) TLS cells and their EE. Early platelet counts were slightly higher than with TLS cells alone, but leukocyte recovery was not improved. These data confirm that TLS cells contribute to early and sustained hematopoiesis, and demonstrate a benefit of ex vivo expanded cells in accelerating engraftment of more primitive TSHFU stem cells depleted of progenitors.

Short-Term Hematopoietic Stem Cells (ST-HSC) Have Full Long-Term Capacity with Sustained but Reduced Potential Compared with LT-HSC.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 2550-2550
Author(s):  
Gerald A. Colvin ◽  
David Berz ◽  
Peter J. Quesenberry ◽  
Elaine Papa ◽  
Liansheng Liu
Keyword(s):  
Stem Cells ◽  
Stem Cell ◽  
Bone Marrow Cells ◽  
Conflicts Of Interest ◽  
Cell Number ◽  
Large Animal ◽  
Short Term ◽  
Hematopoietic Stem ◽  
One Year

Abstract Abstract 2550 Poster Board II-527 We evaluated the hypothesis that there was a homing defect between long-term (LT) hematopoietic stem cell (HSC) (KLS-Flk2-) and ST-HSC (KLS-Flk2+) that explained differences in engraftment potential and duration. Short-term HSC by definition have limited self-renewal capacity, generally described as giving rise to lymphohematopoiesis for 4–12 weeks before senescence. We performed three large animal engraftment studies into lethally ablated mice (950cGy split dose) looking at engraftment of both ST and LT-HSC cells delivered via intravenous, intraperitoneal and intra-femoral route. Two-hundred or 500 cells derived from B6/SJL mice were administered to each animal along with 300,000 recipient (C57/BLK) whole bone marrow cells for radioprotection following prior published studied [PNAS:98;14541, Stem Cells 24:1087] with optimization of flourochromes for better discrimination with our Cytopeia sorter. The animals were serially transplanted after eight months or one year to secondary recipients. In our hands, the ST-HSC engrafted animals did not lose chimerism over time. Review of the literature revealed that there were not confirmatory studies from those published from the initial one publication describing the ST-HSC. We found the ST-HSC were not short-term and persisted for one year in primary recipients and at least 3 months in secondary recipients. Engraftment kinetics favored LT-HSC over ST-HSC with engraftment examples at one year of 62% compared with 30% respectively when administered intravenously, 10% verses 4% given intra-femoral and 0.5% verse 0.3% given intraperitoneal. Chimerism was on average 50% better for the LT-HSC when compared with the ST-HSC and was irrespective of route proving that the differences seen are not due to homing deficiency but rather intrinsic differences in the two stem cell pools. Prior studies gave a maximum of 100 cells. Cell number was purposely increased for better differentiate of subtle differences in engraftment kinetics for statistical reasons. To avoid contamination of Flk2+ cells in the Flk2- cohort and vise-versa, discrimination of the gates were enhanced from that which was published prior. Double sorting of the cells confirmed that there was no appreciable cross contamination but obviously we cannot totally rule that out as a potentially confounding factor. In conclusion we found that ST-HSC as described have long-term capacity with intrinsic differences in engraftment potential that is not driven by a homing defect. Disclosures: No relevant conflicts of interest to declare.

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