Formation of Bone and Foci of Ectopic Hemopoiesis at Joint Application of Calcium Scaffolds with Bone Marrow Cells or Cultivated Mesenchymal Stromal Cells

Potential application of calcium scaffolds in combination with either bone marrow or adherent cell layers from long-term cultures containing mesenchymal stem cell for the induction of bone tissue growth was studied in mice. Two scaffolds, i.e. Osteoset® and Prodens® were tested on the model of ectopic grafting under the skin and renal capsule of mice. It was demonstrated that Prodens® and less effectively Osteoset® could be used for the induction of bone growth in combination with bone marrow cells but even more effectively in combination with cultivated mesenchymal stromal cells. Both the site of transplantation and preliminary induction of bone differentiation of stromal cells exerted a great influence upon the process of bone formation.

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Bisecting GlcNAc structure is implicated in suppression of stroma-dependent haemopoiesis in transgenic mice expressing N-acetylglucosaminyltransferase III

Biochemical Journal ◽

10.1042/bj3310733 ◽

1998 ◽

Vol 331 (3) ◽

pp. 733-742 ◽

Cited By ~ 8

Author(s):

Masafumi YOSHIMURA ◽

Yoshito IHARA ◽

Tetsuo NISHIURA ◽

Yu OKAJIMA ◽

Megumu OGAWA ◽

...

Keyword(s):

Bone Marrow ◽

Transgenic Mice ◽

Stromal Cells ◽

Bone Marrow Cells ◽

Adherent Cell ◽

Wild Type ◽

Spleen Cells ◽

Bisecting Glcnac ◽

Marrow Cells

Several sugar structures have been reported to be necessary for haemopoiesis. We analysed the haematological phenotypes of transgenic mice expressing β-1,4 N-acetylglucosaminyltransferase III (GnT-III), which forms bisecting N-acetylglucosamine on asparagine-linked oligosaccharides. In the transgenic mice, the GnT-III activity was elevated in bone marrow, spleen and peripheral blood and in isolated mononuclear cells from these tissues, whereas no activity was found in these tissues of wild-type mice. Stromal cells after long-term cultures of transgenic-derived bone marrow and spleen cells also showed elevated GnT-III activity, compared with an undetectable activity in wild-type stromal cells. As judged by HPLC analysis, lectin blotting and lectin cytotoxicity assay, bisecting GlcNAc residues were increased on both blood cells and stromal cells from bone marrow and spleen in transgenic mice. The transgenic mice displayed spleen atrophy, hypocellular bone marrow and pancytopenia. Bone marrow cells and spleen cells from transgenic mice produced fewer haemopoietic colonies. After lethal irradiation followed by bone marrow transplantation, transgenic recipient mice showed pancytopenia compared with wild-type recipient mice. Bone marrow cells from transgenic donors gave haematological reconstitution at the same level as wild-type donor cells. In addition, non-adherent cell production was decreased in long-term bone marrow cell cultures of transgenic mice. Collectively these results indicate that the stroma-supported haemopoiesis is compromised in transgenic mice expressing GnT-III, providing the first demonstration that the N-glycans have some significant roles in stroma-dependent haemopoiesis.

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Mesenchymal Stromal Cells Enhance G-CSF-Induced Mobilization Of Hematopoietic Stem- and Progenitor Cells

Blood ◽

10.1182/blood.v122.21.2460.2460 ◽

2013 ◽

Vol 122 (21) ◽

pp. 2460-2460

Author(s):

Evert-Jan F. M. de Kruijf ◽

Ingmar van Hengel ◽

Jorge M Perez-Galarza ◽

Willem E. Fibbe ◽

Melissa van Pel

Keyword(s):

Bone Marrow ◽

B Lymphocytes ◽

Mesenchymal Stromal Cells ◽

Stromal Cells ◽

Peripheral Blood ◽

Bone Marrow Cells ◽

Calf Serum ◽

Intraperitoneal Administration ◽

Hematopoietic Stem ◽

Marrow Cells

Abstract Hematopoietic stem- and progenitor cell (HSPC) mobilization is a property of most hematopoietic growth factors, such as Granulocyte Colony Stimulating Factor (G-CSF). Not all donors mobilize equally well and therefore the number of HSPC that are obtained following mobilization may be limited. Mesenchymal stromal cells (MSC) have the capacity to differentiate into cells of the mesodermal lineage and have immunomodulatory properties in vivo and in vitro. Here, we have investigated the effect of MSC co-administration on G-CSF-induced HSPC mobilization. MSC were obtained from bone marrow cells (bone marrow-derived) or bone fragments (bone-derived) and were expanded in alpha-MEM containing 10% fetal calf serum until sufficient cell numbers were obtained. Bone marrow or bone-derived MSC were administered intravenously for three days at a dose of 200 x103 cells per day to male C57BL/6 recipients that were simultaneously mobilized with G-CSF (10 μg per day intraperitoneally for 3 days) or PBS as a control. Co-injection of G-CSF and MSC lead to a 2-fold increase in HSPC mobilization compared to G-CSF alone (8,563 ± 3,309 vs. 4,268 ± 1,314 CFU-C per ml peripheral blood respectively; n=13, p<0.01). Administration of MSC alone did not induce HSPC mobilization (273 ± 229 CFU-C/ml blood; n=13). Furthermore, co-injection of splenocytes and G-CSF did not enhance HSPC mobilization, showing that the administration of exogeneous cells as such is not sufficient for enhancement of HSPC mobilization. It has been reported that G-CSF-induced HSPC mobilization is associated with a decrease in the number of osteal macrophages, B lymphocytes and erythroid progenitors. Administration of MSC alone induced a significant decrease in the frequency of osteal macrophages (7.9 ± 1.2 vs 6.2 ± 1.4% bone marrow cells for PBS vs. MSC respectively; n=8, p<0.05), but did not affect osteoblast numbers. Furthermore, the frequency of B lymphocytes was significantly decreased following MSC administration (29.9 ± 4.0 vs. 16.5 ± 4.9% bone marrow cells for PBS vs. MSC respectively; n=13, p<0.0001). No differences were observed in erythroid numbers following MSC administration. To investigate the mechanisms underlying these observations, the migratory capacity of luciferase transduced MSC was studied through bioluminescence imaging. Following intravenous injection, MSC were detected in the lungs, but not in other organs. In addition, no difference in MSC migration was observed between G-CSF and PBS treated mice. Moreover, intraperitoneal administration of G-CSF and MSC resulted in increased HSPC mobilization compared to G-CSF alone (10,178 ±3,039 vs. 5,158 ± 2,436 CFU-C per ml peripheral blood; n=5-12). Together, these data point to an endocrine effect of MSC on G-CSF-induced HSPC mobilization. No differences in IL-6, CXCL-12 or M-CSF levels in bone marrow extracellular fluid were observed. In conclusion, G-CSF-induced HSPC mobilization is enhanced by injection of MSC. We hypothesize that the MSC-induced partial depletion of B lymphocytes and osteal macrophages in the bone marrow are crucial factors involved in the enhancement of G-CSF-induced HSPC mobilization. Disclosures: No relevant conflicts of interest to declare.

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No increased trapping of multipotent mesenchymal stromal cells in bone marrow filters compared with other bone marrow cells

Cytotherapy ◽

10.1080/14653240801965164 ◽

2008 ◽

Vol 10 (3) ◽

pp. 238-242 ◽

Cited By ~ 6

Author(s):

M. Sundin ◽

M. Remberger ◽

H. Lönnies ◽

B. Sundberg ◽

O. Ringdén ◽

...

Keyword(s):

Bone Marrow ◽

Mesenchymal Stromal Cells ◽

Stromal Cells ◽

Bone Marrow Cells ◽

Multipotent Mesenchymal Stromal Cells ◽

Marrow Cells

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Assessment of DNA Damage in Human Bone Marrow Cells and Multipotent Mesenchymal Stromal Cells

Bulletin of Experimental Biology and Medicine ◽

10.1007/s10517-011-1379-8 ◽

2011 ◽

Vol 151 (4) ◽

pp. 550-552 ◽

Cited By ~ 4

Author(s):

V. A. Nikitina ◽

A. I. Chausheva ◽

A. K. Zhanataev ◽

E. Yu. Osipova ◽

A. D. Durnev ◽

...

Keyword(s):

Bone Marrow ◽

Dna Damage ◽

Mesenchymal Stromal Cells ◽

Stromal Cells ◽

Bone Marrow Cells ◽

Multipotent Mesenchymal Stromal Cells ◽

Human Bone ◽

Human Bone Marrow ◽

Marrow Cells

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Colony formation by primitive haemopoietic progenitors in cocultures of bone marrow cells and stromal cells

British Journal of Haematology ◽

10.1111/j.1365-2141.1985.tb07393.x ◽

1985 ◽

Vol 60 (1) ◽

pp. 129-136 ◽

Cited By ~ 47

Author(s):

M. Y. Gordon ◽

J. A. Hibbin ◽

L. U. Kearney ◽

E. C. Gordon-Smith ◽

J. M. Goldman

Keyword(s):

Bone Marrow ◽

Stromal Cells ◽

Colony Formation ◽

Bone Marrow Cells ◽

Marrow Cells

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Thrombopoietin Is Synthesized by Bone Marrow Stromal Cells

Blood ◽

10.1182/blood.v90.9.3444 ◽

1997 ◽

Vol 90 (9) ◽

pp. 3444-3455 ◽

Cited By ~ 72

Author(s):

Anastasia Guerriero ◽

Lydia Worford ◽

H. Kent Holland ◽

Gui-Rong Guo ◽

Kevin Sheehan ◽

...

Keyword(s):

Bone Marrow ◽

Stem Cell ◽

Progenitor Cells ◽

Stromal Cells ◽

Cell Sorting ◽

Bone Marrow Cells ◽

Adult Bone Marrow ◽

Hla Dr ◽

Adult Bone ◽

Marrow Cells

Abstract We have previously characterized stromal progenitor cells contained in fetal bone marrow by fluorescence-activated cell sorting (FACS) using the differential expression of CD34, CD38, and HLA-DR, and found that a small number were contained within the CD34+ cell fraction. In the present study, the frequency of stromal progenitors in both the CD34+ and CD34− subpopulations from samples of fetal and adult bone marrow was approximately one in 5,000 of the mononuclear cell fraction. Using multiparameter single-cell sorting, one in 20 fetal bone marrow cells with the CD34+, CD38−, HLA-DR−, CDw90+ phenotype were clonogenic stromal progenitors, whereas greater than one in five single cells with the CD34−, CD38−, HLA-DR−, CDw90+ phenotype formed stromal cultures. We found that cultures initiated by hematopoietic and stromal progenitors contained within the CD34+ fraction of bone marrow cells formed mixed hematopoietic/stromal cell cultures that maintained the viability of the hematopoietic progenitor cells for 3 weeks in the absence of added hematopoietic cytokines. We characterized some of the hematopoietic cytokines synthesized by stromal cultures derived from either CD34+ or CD34− bone marrow cells using reverse transcriptase–polymerase chain reaction (RT-PCR) amplification of interleukin-3 (IL-3), stem cell factor (SCF), CD34, Flt3/Flk2 ligand (FL), and thrombopoietin (TPO) mRNA sequences. We found ubiquitous expression of TPO mRNA in greater than 90% of stromal cultures initiated by either CD34+ or CD34− cells, and variable expression of SCF, FL, and CD34 mRNA. In particular, SCF and CD34 mRNA were detected only in stromal cultures initiated by CD34+ bone marrow cells, although the differences between CD34+ and CD34− stromal cells were not statistically significant. IL-3 mRNA was not found in any stromal cultures. An enzyme-linked immunosorbent assay (ELISA) of soluble SCF and TPO present in culture supernatants demonstrated that biologically significant amounts of protein were secreted by some cultured stromal cells: eight of 16 samples of conditioned media from stromal cultures initiated by fetal and adult bone marrow contained more than 32 pg/mL SCF (in the linear range of the ELISA), with a median value of 32 pg/mL (range, 9 to 230), while 13 of 24 samples of conditioned media had more than 16 pg/mL TPO (in the linear range of the ELISA), with a median of 37 pg/mL (range, 16 to 106). Our data indicate that stromal cultures initiated by single bone marrow cells can make FL, SCF, and TPO. Local production of early-acting cytokines and TPO by stromal cells may be relevant to the regulation of hematopoietic stem cell self-renewal and megakaryocytopoiesis in the bone marrow microenvironment.

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Direct stimulation of osteoclastogenesis by MIP-1alpha: evidence obtained from studies using RAW264 cell clone highly responsive to RANKL

Journal of Endocrinology ◽

10.1677/joe.0.1800193 ◽

2004 ◽

Vol 180 (1) ◽

pp. 193-201 ◽

Cited By ~ 93

Author(s):

T Watanabe ◽

T Kukita ◽

A Kukita ◽

N Wada ◽

K Toh ◽

...

Keyword(s):

Bone Marrow ◽

Stromal Cells ◽

Bone Marrow Cells ◽

Tnf Alpha ◽

Adhesion Properties ◽

Multinucleated Cells ◽

Raw264 Cell ◽

Receptor Activator ◽

Rat Bone ◽

Marrow Cells

Macrophage inflammatory protein-1alpha (MIP-1alpha) is a member of the CC chemokines. We have previously reported the use of a whole bone marrow culture system to show that MIP-1alpha stimulates the formation of osteoclast-like multinucleated cells. Here we use rat bone marrow cells deprived of stromal cells, and clones obtained from murine macrophage-like cell line RAW264 to show that MIP-1alpha acts directly on cells in osteoclast lineage. We obtained several types of RAW264 cell clones, one of these clones, designated as RAW264 cell D clone (D clone), showed an extremely high response to receptor activator of NFkappaB ligand (RANKL) and tumor necrosis factor-alpha (TNF-alpha), while the other clone, RAW264 cell N clone (N clone), demonstrated no response to RANKL or TNF-alpha. Although both clones expressed receptor activator NFkappaB (RANK) before being stimulated for differentiation, only the D clone expressed cathepsin K when cells were stimulated to differentiate to osteoclasts. MIP-1alpha stimulated the formation of mononuclear preosteoclast-like cells from rat bone marrow cells deprived of stromal cells. MIP-1alpha also stimulated formation of osteoclast-like multinucleated cells from the D clone, when these cells were stimulated with RANKL and TNF-alpha. These findings provide strong evidence to show that MIP-1alpha acts directly on cells in the osteoclast lineage to stimulate osteoclastogenesis. Furthermore, pretreatment of RAW264 cell D clone with MIP-1alpha significantly induced adhesion properties of these cells to primary osteoblasts, suggesting a crucial role for MIP-1alpha in the regulation of the interaction between osteoclast precursors and osteoblasts in osteoclastogenesis.

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Erythropoietin Gene Expression in the Stromal Cell Increased by Silica to Induce Erythrocyte Differentiation

Advanced Materials Research ◽

10.4028/www.scientific.net/amr.647.494 ◽

2013 ◽

Vol 647 ◽

pp. 494-498

Author(s):

Wei Chung Liu ◽

Chang Shu Tsai ◽

Ya Yun Chen ◽

Nien Tzu Keng

Keyword(s):

Gene Expression ◽

Bone Marrow ◽

Stromal Cells ◽

Bone Marrow Cells ◽

Biological Effects ◽

Interaction Mechanism ◽

Silica Particles ◽

Control Group ◽

Stimulate Bone Marrow ◽

Marrow Cells

Silica containing materials are often applied in bone tissue engineering, which may contact with bone marrow cells. However, the biological effects have not always been observed in studies of bone marrow cells exposed to silica. In this experiment, the relevant biological effects were evaluated. Bone marrow cells and stromal cells treated with silica particles (0.5-10 μm) were applied to investigate the possible interaction mechanism. HEL-92 cells were culture with the condition medium of stromal cells treated with or without silica particles. The erythrogenesis of bone marrow cells treated with silica particles was increased significantly. The expression level of glycophorin A the erythroid marker in HEL-92 cells treated by condition medium was higher than control group. The silica particles could also up-regulate the erythropoietin gene expression of stromal cells. The results indicate that bone marrow cells can be stimulated by silica particles to differentiate into erythrocytes. Our results suggest that silica particles can stimulate bone marrow cells to differentiate erythrocytes possibly via enhancing gene expression of erythropoietin.

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