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.

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.

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 MRI Tracking of SPIO- and Fth1-Labeled Bone Marrow Mesenchymal Stromal Cell Transplantation for Treatment of Stroke

2019 ◽  
Vol 2019 ◽  
pp. 1-10 ◽  
Author(s):  
Xiaolei Huang ◽  
Yang Xue ◽  
Jinliang Wu ◽  
Qing Zhan ◽  
Jiangmin Zhao
Keyword(s):  
Bone Marrow ◽  
Prussian Blue ◽  
Mesenchymal Stromal Cells ◽  
Stromal Cells ◽  
Signal Intensity ◽  
Immunofluorescent Staining ◽  
Blue Staining

We aimed to identify a suitable method for long-term monitoring of the migration and proliferation of mesenchymal stromal cells in stroke models of rats using ferritin transgene expression by magnetic resonance imaging (MRI). Bone marrow mesenchymal stromal cells (BMSCs) were transduced with a lentivirus containing a shuttle plasmid (pCDH-CMV-MCS-EF1-copGFP) carrying the ferritin heavy chain 1 (Fth1) gene. Ferritin expression in stromal cells was evaluated with western blotting and immunofluorescent staining. The iron uptake of Fth1-BMSCs was measured with Prussian blue staining. Following surgical introduction of middle cerebral artery occlusion, Fth1-BMSCs and superparamagnetic iron oxide- (SPIO-) labeled BMSCs were injected through the internal jugular vein. The imaging and signal intensities were monitored by diffusion-weighted imaging (DWI), T2-weighted imaging (T2WI), and susceptibility-weighted imaging (SWI) in vitro and in vivo. Pathology was performed for comparison. We observed that the MRI signal intensity of SPIO-BMSCs gradually reduced over time. Fth1-BMSCs showed the same signal intensity between 10 and 60 days. SWI showed hypointense lesions in the SPIO-BMSC (traceable for 30 d) and Fth1-BMSC groups. T2WI was not sensitive enough to trace Fth1-BMSCs. After transplantation, Prussian blue-stained cells were observed around the infarction area and in the infarction center in both transplantation models. Fth1-BMSCs transplanted for treating focal cerebral infarction were safe, reliable, and traceable by MRI. Fth1 labeling was more stable and suitable than SPIO labeling for long-term tracking. SWI was more sensitive than T2W1 and suitable as the optimal MRI-tracking sequence.

scholarly journals Stromal cells in myeloid and lymphoid long-term bone marrow cultures can support multiple hemopoietic lineages and modulate their production of hemopoietic growth factors

Blood ◽  
1986 ◽  
Vol 68 (6) ◽  
pp. 1348-1354 ◽  
Author(s):  
A Johnson ◽  
K Dorshkind
Keyword(s):  
Bone Marrow ◽  
Stromal Cells ◽  
Stromal Cell ◽  
Culture Conditions ◽  
B Lymphopoiesis ◽  
Bone Marrow Cultures ◽  
Factor Secretion ◽  
Marrow Cultures

Abstract Hemopoiesis in long-term bone marrow cultures (LTBMC) is dependent on adherent stromal cells that form an in vitro hemopoietic microenvironment. Myeloid bone marrow cultures (MBMC) are optimal for myelopoiesis, while lymphoid bone marrow cultures (LBMC) only support B lymphopoiesis. The experiments reported here have made a comparative analysis of the two cultures to determine whether the stromal cells that establish in vitro are restricted to the support of myelopoiesis or lymphopoiesis, respectively, and to examine how the different culture conditions affect stromal cell physiology. In order to facilitate this analysis, purified populations of MBMC and LBMC stroma were prepared by treating the LTBMC with the antibiotic mycophenolic acid; this results in the elimination of hemopoietic cells while retaining purified populations of functional stroma. Stromal cell cultures prepared and maintained under MBMC conditions secreted myeloid growth factors that stimulated the growth of granulocyte-macrophage colonies, while no such activity was detected from purified LBMC stromal cultures. However, this was not due to the inability of LBMC stroma to mediate this function. Transfer of LBMC stromal cultures to MBMC conditions resulted in an induction of myeloid growth factor secretion. When seeded under these conditions with stromal cell- depleted populations of hemopoietic cells, obtained by passing marrow through nylon wool columns, the LBMC stromal cells could support long- term myelopoiesis. Conversely, transfer of MBMC stroma to LBMC conditions resulted in a cessation of myeloid growth factor secretion; on seeding these cultures with nylon wool-passed marrow, B lymphopoiesis, but not myelopoiesis, initiated. These findings indicate that the stroma in the different LTBMC are not restricted in their hemopoietic support capacity but are sensitive to culture conditions in a manner that may affect the type of microenvironment formed.

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.

Perivascular/Mesenchymal Stem Cells from Multiple Human Tissues Support Hematopoiesis in Vitro.

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 1361-1361
Author(s):  
Elisa Montelatici ◽  
Gabriella Andriolo ◽  
Mihaela Crisan ◽  
Rosaria Giordano ◽  
Paolo Rebulla ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Stem Cells ◽  
Bone Marrow ◽  
Mesenchymal Stem Cells ◽  
Stromal Cells ◽  
Marrow Stromal Cells ◽  
Hematopoietic Cell ◽  
Human Tissues

Abstract Mesenchymal stem cells (MSC) can be derived selectively in culture from multiple organs, an omnipresence we have recently suggested to be explained by the perivascular location of native MSC ancestors within intact tissues (Crisan et al. 2008, in press). We have now analyzed the ability of MSC extracted pro- or retrospectively from different human tissues to support hematopoiesis. MSC were either classically derived in primary cultures of umbilical cord blood (UCB) lineage-depleted mononuclear cells (n=3) or enzymatically dissociated adult adipose tissue (n=3), or grown as CD146+ NG2+ CD34-CD56- CD45- pericytes (n=2) purified by flow cytometry from fetal skeletal muscle and cultured over the long term. In both settings, identical MSC were obtained that maintained a stable CD146+ CD90+ CD73+ CD105+ CD34- CD45- surface phenotype and could differentiate into skeletal muscle, fat, bone and cartilage. CD34+ hematopoietic progenitors (n=3) immunoselected from term UCB were seeded (5×10e3cells/cm2 in triplicate) onto confluent irradiated layers of MSC derived from UCB, adipose tissue or fetal muscle pericytes (MSCu, MSCa and MSCmp, respectively) or, as a control, MS5 bone marrow stromal cells that allow the proliferation of very primitive human progenitor cells. All studies were approved by the relevant institutional regulatory board. The cells were cocultured for 5 weeks in a classical long-term culture-initiating cell assay in a complete medium (MyeloCult H5100, Stem Cell Technologies) containing hydrocortisone but no added cytokine. Wells were scored daily for the presence of cobblestone areas (CA) and half of the medium was replaced every week. Eventually, trypsinized cells from each well were characterized by flow cytometry for the expression of hematopoietic cell markers and assayed for CFC potential. After 14 days of incubation, colonies grown in semi-solid medium were scored as derived from colony forming units (CFU)-granulocyte, erythroid, macrophage, megakaryocyte (GEMM) and as high-proliferative-potential colony precursors (HPPC), the most primitive hematopoietic cell so far identified in a clonogenic assay in vitro. Within the CD45+ gate, all trypsinized cultures contained comparable percentages of CD34+lin- cells (MSCu: 51±9%; MSCa: 58±14%; MSCmp: 61±19%; MS5: 59±18%), the most immature hematopoietic cell compartment maintained during the long-term coculture. MSCu and MSCmp supported a similar cell proliferation during the whole culture while on MSCa, CA formed very rapidly and consistently but eventually decreased over the long-term culture. Interestingly, MSCu and MSCmp supported the development of the highest numbers of HPPC and of CFU giving rise to the largest GEMM colonies, as compared to MSCa that gave the same results as the control MS5 cell line. In summary, all MSCs tested were able to support hematopoiesis and CA formation, albeit with differences in growth kinetics and morphology of the colonies. Herein we show for the first time that purified human perivascular cells exhibit robust hematopoiesis support in vitro, in addition to multilineage mesodermal developmental potential. In conclusion, we demonstrate that MSC from novel sources distinct from the bone marrow are able to support hematopoiesis. These results further sustain the identity, beyond acronyms, between marrow stromal cells, long known for their support of hematopoiesis, and mesenchymal stem cells that gained more recent credit in the field of regenerative medicine because of their multilineage differentiation potential.

Long-term culture of infant leukemia cells: Dependence upon stromal cells from the bone marrow and bilineage differentiation

1986 ◽  
Vol 10 (8) ◽  
pp. 1007-1013 ◽  
Author(s):  
Tehila Umiel ◽  
Sari Friedman ◽  
Rina Zaizov ◽  
Ian J. Cohen ◽  
Yehoshua Gozes ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Stromal Cells ◽  
Leukemia Cells ◽  
Long Term Culture ◽  
Infant Leukemia

scholarly journals Stromal cells in myeloid and lymphoid long-term bone marrow cultures can support multiple hemopoietic lineages and modulate their production of hemopoietic growth factors

Blood ◽  
1986 ◽  
Vol 68 (6) ◽  
pp. 1348-1354 ◽  
Author(s):  
A Johnson ◽  
K Dorshkind
Keyword(s):  
Bone Marrow ◽  
Stromal Cells ◽  
Stromal Cell ◽  
Culture Conditions ◽  
B Lymphopoiesis ◽  
Bone Marrow Cultures ◽  
Factor Secretion ◽  
Marrow Cultures

Hemopoiesis in long-term bone marrow cultures (LTBMC) is dependent on adherent stromal cells that form an in vitro hemopoietic microenvironment. Myeloid bone marrow cultures (MBMC) are optimal for myelopoiesis, while lymphoid bone marrow cultures (LBMC) only support B lymphopoiesis. The experiments reported here have made a comparative analysis of the two cultures to determine whether the stromal cells that establish in vitro are restricted to the support of myelopoiesis or lymphopoiesis, respectively, and to examine how the different culture conditions affect stromal cell physiology. In order to facilitate this analysis, purified populations of MBMC and LBMC stroma were prepared by treating the LTBMC with the antibiotic mycophenolic acid; this results in the elimination of hemopoietic cells while retaining purified populations of functional stroma. Stromal cell cultures prepared and maintained under MBMC conditions secreted myeloid growth factors that stimulated the growth of granulocyte-macrophage colonies, while no such activity was detected from purified LBMC stromal cultures. However, this was not due to the inability of LBMC stroma to mediate this function. Transfer of LBMC stromal cultures to MBMC conditions resulted in an induction of myeloid growth factor secretion. When seeded under these conditions with stromal cell- depleted populations of hemopoietic cells, obtained by passing marrow through nylon wool columns, the LBMC stromal cells could support long- term myelopoiesis. Conversely, transfer of MBMC stroma to LBMC conditions resulted in a cessation of myeloid growth factor secretion; on seeding these cultures with nylon wool-passed marrow, B lymphopoiesis, but not myelopoiesis, initiated. These findings indicate that the stroma in the different LTBMC are not restricted in their hemopoietic support capacity but are sensitive to culture conditions in a manner that may affect the type of microenvironment formed.

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