Effect of donor age on long-term culture of bone marrow in vitro

1984 ◽  
Vol 24 (1) ◽  
pp. 119-127 ◽  
Author(s):  
D.A. Lipschitz ◽  
K.B. Udupa
Keyword(s):  
Bone Marrow ◽  
Donor Age ◽  
Long Term Culture

scholarly journals Differential Maintenance of Primitive Human SCID-Repopulating Cells, Clonogenic Progenitors, and Long-Term Culture-Initiating Cells After Incubation on Human Bone Marrow Stromal Cells

Blood ◽  
1997 ◽  
Vol 90 (2) ◽  
pp. 641-650 ◽  
Author(s):  
Olga I. Gan ◽  
Barbara Murdoch ◽  
Andre Larochelle ◽  
John E. Dick
Keyword(s):  
Bone Marrow ◽  
Bone Marrow Stromal Cells ◽  
Cultured Cells ◽  
Ex Vivo ◽  
Hematopoietic Cells ◽  
Long Term Culture ◽  
Cell Engraftment ◽  
Ex Vivo Culture

Abstract Many experimental and clinical protocols are being developed that involve ex vivo culture of human hematopoietic cells on stroma or in the presence of cytokines. However, the effect of these manipulations on primitive hematopoietic cells is not known. Our severe combined immune-deficient mouse (SCID)-repopulating cell (SRC) assay detects primitive human hematopoietic cells based on their ability to repopulate the bone marrow (BM) of immune-deficient non-obese diabetic/SCID (NOD/SCID) mice. We have examined here the maintenance of SRC, colony-forming cells (CFC), and long-term culture-initiating cells (LTC-IC) during coculture of adult human BM or umbilical cord blood (CB) cells with allogeneic human stroma. Transplantation of cultured cells in equivalent doses as fresh cells resulted in lower levels of human cell engraftment after 1 and 2 weeks of culture for BM and CB, respectively. Similar results were obtained using CD34+-enriched CB cells. By limiting dilution analysis, the frequency of SRC in BM declined sixfold after 1 week of culture. In contrast to the loss of SRC as measured by reduced repopulating capacity, the transplanted inocula of cultured cells frequently contained equal or higher numbers of CFC and LTC-IC compared with the inocula of fresh cells. The differential maintenance of CFC/LTC-IC and SRC suggests that SRC are biologically distinct from the majority of these in vitro progenitors. This report demonstrates the importance of the SRC assay in the development of ex vivo conditions that will allow maintenance of primitive human hematopoietic cells with repopulating capacity.

scholarly journals Isolation and Expansion of Mesenchymal Stem/Stromal Cells, Functional Assays and Long-Term Culture Associated Alterations of Cellular Properties

2021 ◽  
Author(s):  
Chenghai Li
Keyword(s):  
Bone Marrow ◽  
Stem Cell ◽  
Mhc Class Ii ◽  
Stromal Cells ◽  
Replicative Senescence ◽  
Cell Types ◽  
Long Term Culture ◽  
Medicine Research

Mesenchymal stem cell/stromal cells (MSCs) can differentiate into a variety of cell types, including osteocytes, adipocytes and chondrocytes. MSCs are present in the multiple types of adult tissue, such as bone marrow, adipose tissue, and various neonatal birth-associated tissues. Given their self-renewal and differentiation potential, immunomodulatory and paracrine properties, and lacking major histocompatibility complex (MHC) class II molecules, MSCs have attracted much attention for stem cell-based translational medicine research. Due to a very low frequency in different types of tissue, MSCs can be isolated and expanded in vitro to derive sufficient cell numbers prior to the clinical applications. In this chapter, the methodology to obtain primary bone marrow-derived MSCs as well as their in vitro culture expansion will be described. To assess the functional properties, differentiation assays, including osteogenesis, chondrogenesis and adipogenesis, 3-D culture of MSCs and co-culture of MSCs and tumor cells are also provided. Finally, the long-term culture associated alterations of MSCs, such as replicative senescence and spontaneous transformation, will be discussed for better understanding of the use of MSCs at the early stages for safe and effective cell-based therapy.

scholarly journals Different stimulative effects of human bone marrow and fetal liver stromal cells on erythropoiesis in long-term culture

Blood ◽  
1987 ◽  
Vol 69 (1) ◽  
pp. 135-139 ◽  
Author(s):  
I Slaper-Cortenbach ◽  
R Ploemacher ◽  
B Lowenberg
Keyword(s):  
Bone Marrow ◽  
Stromal Cells ◽  
Fetal Liver ◽  
The Other ◽  
Long Term Culture ◽  
Human Fetal Liver ◽  
Relative Paucity ◽  
Fetal Erythropoiesis

The factors determining the predominantly erythroid direction of human fetal liver hematopoiesis are unknown. We compared the capacities of human fetal liver and bone marrow stromas to sustain fetal and adult hematopoiesis in long-term cultures. In various marrow-fetal liver combinations of stroma and recharge, the maintenance of erythroid (BFU- e) and myeloid (CFU-GM) precursors in the nonadherent phase was determined. The morphology of the fetal liver nucleated cells during culture was also examined. This study shows that fetal liver stromas efficiently support fetal BFU-e for 6 to 7 weeks in vitro. Bone marrow stromas were not able to maintain fetal BFU-e beyond 4 weeks. Significant numbers of marrow BFU-e were not sustained in vitro on either source of stroma. On the other hand, the stroma layers of fetal liver and marrow origin were equally effective in maintaining fetal CFU- GM and adult CFU-GM in long-term culture. These findings show that the human embryonic liver stroma is a preferential site for stimulating fetal erythropoiesis. They do not demonstrate differences in stroma function to explain the relative paucity of myelopoiesis in the fetal liver.

Distinct Requirements for Optimal Growth and In Vitro Expansion of Human CD34+CD38− Bone Marrow Long-Term Culture-Initiating Cells (LTC-IC), Extended LTC-IC, and Murine In Vivo Long-Term Reconstituting Stem Cells

Blood ◽  
1999 ◽  
Vol 94 (12) ◽  
pp. 4093-4102 ◽  
Author(s):  
Veslemøy Ramsfjell ◽  
David Bryder ◽  
Helga Björgvinsdóttir ◽  
Sten Kornfält ◽  
Lars Nilsson ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Stem Cell ◽  
Cell Activity ◽  
Optimal Growth ◽  
Cell Manipulation ◽  
Long Term Culture ◽  
Human Cd34

Abstract Recently, primitive human bone marrow (BM) progenitors supporting hematopoiesis in extended (>60 days) long-term BM cultures were identified. Such extended long-term culture-initiating cells (ELTC-IC) are of the CD34+CD38− phenotype, are quiescent, and are difficult to recruit into proliferation, implicating ELTC-IC as the most primitive human progenitor cells detectable in vitro. However, it remains to be established whether ELTC-IC can proliferate and potentially expand in response to early acting cytokines. Here, CD34+CD38− BM ELTC-IC (12-week) were efficiently recruited into proliferation and expanded in vitro in response to early acting cytokines, but conditions for expansion of ELTC-IC activity were distinct from those of traditional (5-week) LTC-IC and murine long-term repopulating cells. Whereas c-kit ligand (KL), interleukin-3 (IL-3), and IL-6 promoted proliferation and maintenance or expansion of murine long-term reconstituting activity and human LTC-IC, they dramatically depleted ELTC-IC activity. In contrast, KL, flt3 ligand (FL), and megakaryocyte growth and development factor (MGDF) (and KL + FL + IL-3) expanded murine long-term reconstituting activity as well as human LTC-IC and ELTC-IC. Expansion of LTC-IC was most optimal after 7 days of culture, whereas optimal expansion of ELTC-IC activity required 12 days, most likely reflecting the delayed recruitment of quiescent CD34+CD38− progenitors. The need for high concentrations of KL, FL, and MGDF (250 ng/mL each) and serum-free conditions was more critical for expansion of ELTC-IC than of LTC-IC. The distinct requirements for expansion of ELTC-IC activity when compared with traditional LTC-IC suggest that the ELTC-IC could prove more reliable as a predictor for true human stem cell activity after in vitro stem cell manipulation.

Distinct Requirements for Optimal Growth and In Vitro Expansion of Human CD34+CD38− Bone Marrow Long-Term Culture-Initiating Cells (LTC-IC), Extended LTC-IC, and Murine In Vivo Long-Term Reconstituting Stem Cells

Blood ◽  
1999 ◽  
Vol 94 (12) ◽  
pp. 4093-4102 ◽  
Author(s):  
Veslemøy Ramsfjell ◽  
David Bryder ◽  
Helga Björgvinsdóttir ◽  
Sten Kornfält ◽  
Lars Nilsson ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Stem Cell ◽  
Cell Activity ◽  
Optimal Growth ◽  
Cell Manipulation ◽  
Long Term Culture ◽  
Human Cd34

Recently, primitive human bone marrow (BM) progenitors supporting hematopoiesis in extended (>60 days) long-term BM cultures were identified. Such extended long-term culture-initiating cells (ELTC-IC) are of the CD34+CD38− phenotype, are quiescent, and are difficult to recruit into proliferation, implicating ELTC-IC as the most primitive human progenitor cells detectable in vitro. However, it remains to be established whether ELTC-IC can proliferate and potentially expand in response to early acting cytokines. Here, CD34+CD38− BM ELTC-IC (12-week) were efficiently recruited into proliferation and expanded in vitro in response to early acting cytokines, but conditions for expansion of ELTC-IC activity were distinct from those of traditional (5-week) LTC-IC and murine long-term repopulating cells. Whereas c-kit ligand (KL), interleukin-3 (IL-3), and IL-6 promoted proliferation and maintenance or expansion of murine long-term reconstituting activity and human LTC-IC, they dramatically depleted ELTC-IC activity. In contrast, KL, flt3 ligand (FL), and megakaryocyte growth and development factor (MGDF) (and KL + FL + IL-3) expanded murine long-term reconstituting activity as well as human LTC-IC and ELTC-IC. Expansion of LTC-IC was most optimal after 7 days of culture, whereas optimal expansion of ELTC-IC activity required 12 days, most likely reflecting the delayed recruitment of quiescent CD34+CD38− progenitors. The need for high concentrations of KL, FL, and MGDF (250 ng/mL each) and serum-free conditions was more critical for expansion of ELTC-IC than of LTC-IC. The distinct requirements for expansion of ELTC-IC activity when compared with traditional LTC-IC suggest that the ELTC-IC could prove more reliable as a predictor for true human stem cell activity after in vitro stem cell manipulation.

scholarly journals Differential Maintenance of Primitive Human SCID-Repopulating Cells, Clonogenic Progenitors, and Long-Term Culture-Initiating Cells After Incubation on Human Bone Marrow Stromal Cells

Blood ◽  
1997 ◽  
Vol 90 (2) ◽  
pp. 641-650 ◽  
Author(s):  
Olga I. Gan ◽  
Barbara Murdoch ◽  
Andre Larochelle ◽  
John E. Dick
Keyword(s):  
Bone Marrow ◽  
Bone Marrow Stromal Cells ◽  
Cultured Cells ◽  
Ex Vivo ◽  
Hematopoietic Cells ◽  
Long Term Culture ◽  
Cell Engraftment ◽  
Ex Vivo Culture

Many experimental and clinical protocols are being developed that involve ex vivo culture of human hematopoietic cells on stroma or in the presence of cytokines. However, the effect of these manipulations on primitive hematopoietic cells is not known. Our severe combined immune-deficient mouse (SCID)-repopulating cell (SRC) assay detects primitive human hematopoietic cells based on their ability to repopulate the bone marrow (BM) of immune-deficient non-obese diabetic/SCID (NOD/SCID) mice. We have examined here the maintenance of SRC, colony-forming cells (CFC), and long-term culture-initiating cells (LTC-IC) during coculture of adult human BM or umbilical cord blood (CB) cells with allogeneic human stroma. Transplantation of cultured cells in equivalent doses as fresh cells resulted in lower levels of human cell engraftment after 1 and 2 weeks of culture for BM and CB, respectively. Similar results were obtained using CD34+-enriched CB cells. By limiting dilution analysis, the frequency of SRC in BM declined sixfold after 1 week of culture. In contrast to the loss of SRC as measured by reduced repopulating capacity, the transplanted inocula of cultured cells frequently contained equal or higher numbers of CFC and LTC-IC compared with the inocula of fresh cells. The differential maintenance of CFC/LTC-IC and SRC suggests that SRC are biologically distinct from the majority of these in vitro progenitors. This report demonstrates the importance of the SRC assay in the development of ex vivo conditions that will allow maintenance of primitive human hematopoietic cells with repopulating capacity.

scholarly journals Different stimulative effects of human bone marrow and fetal liver stromal cells on erythropoiesis in long-term culture

Blood ◽  
1987 ◽  
Vol 69 (1) ◽  
pp. 135-139 ◽  
Author(s):  
I Slaper-Cortenbach ◽  
R Ploemacher ◽  
B Lowenberg
Keyword(s):  
Bone Marrow ◽  
Stromal Cells ◽  
Fetal Liver ◽  
The Other ◽  
Long Term Culture ◽  
Human Fetal Liver ◽  
Relative Paucity ◽  
Fetal Erythropoiesis

Abstract The factors determining the predominantly erythroid direction of human fetal liver hematopoiesis are unknown. We compared the capacities of human fetal liver and bone marrow stromas to sustain fetal and adult hematopoiesis in long-term cultures. In various marrow-fetal liver combinations of stroma and recharge, the maintenance of erythroid (BFU- e) and myeloid (CFU-GM) precursors in the nonadherent phase was determined. The morphology of the fetal liver nucleated cells during culture was also examined. This study shows that fetal liver stromas efficiently support fetal BFU-e for 6 to 7 weeks in vitro. Bone marrow stromas were not able to maintain fetal BFU-e beyond 4 weeks. Significant numbers of marrow BFU-e were not sustained in vitro on either source of stroma. On the other hand, the stroma layers of fetal liver and marrow origin were equally effective in maintaining fetal CFU- GM and adult CFU-GM in long-term culture. These findings show that the human embryonic liver stroma is a preferential site for stimulating fetal erythropoiesis. They do not demonstrate differences in stroma function to explain the relative paucity of myelopoiesis in the fetal liver.

Sign in / Sign up

Export Citation Format

Share Document