The Role of Human Leukocyte Antigen G in Inducing Immune Tolerance After Allogeneic Hematopoietic Stem Cell Transplantation.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 4478-4478
Author(s):  
Xuan Du ◽  
Xiuli Wu ◽  
Rui Li ◽  
Zhiping Fan ◽  
Yu Zhang ◽  
...  
Keyword(s):  
Bone Marrow ◽  
T Lymphocytes ◽  
Peripheral Blood ◽  
Mononuclear Cells ◽  
Culture Supernatant ◽  
Hematopoietic Stem ◽  
Ifn Γ ◽  
High Level

Abstract Abstract 4478 Background and Objective Allogeneic hematopoietic stem cell transplantation (allo-HSCT) is now applied widely for the treatment of hematological or non-hematological malignancies, aplastic anemia and hereditary diseases. Recently, a protocol for haploidentical allo-HSCT that combines granulocyte-colony stimulating factor (G-CSF) primed bone marrow (G-BM) and peripheral blood stem cells (G-PBSC) without in vitro T-cell depletion received great success. But the mechanism of G-CSF inducing immunotolerance in haploidentical-HSCT has not yet been clarified. Human leucocyte antigen G (HLA-G) is a nonclassical HLA class I molecule, the tolerogenic role of HLA-G is highly supported in pregnancy immunization, tumor immune escape and organ transplant. Because HLA-G closely related to immunotolerance, we investigate the role of HLA-G in inducing immune tolerance after allo-HSCT and the effects of G-CSF on the expression and secretion level of HLA-G. Methods Flow cytometry was used to detect the expression of membrane-bound HLA-G (mHLA-G) on donor peripheral blood cells (PBC) or bone marrow (BM) cells. The levels of soluble HLA-G (sHLA-G) in the plasma and bone marrow fluid were determined by enzyme-linked immunosorbent assay (ELISA). In vitro, the expression and secretion level of HLA-G in bone marrow mononuclear cells (BMMCs) and peripheral blood mononuclear cells (PBMCs) after G-CSF stimulated were detected by flow cytometry and ELISA, respectively; Separated T lymphocytes which expressed high level of HLA-G were cultured with allogeneic T lymphocytes and relative response index (RPI) was measured with MTT assay; Furthermore, separated T lymphocytes were cultured with allogeneic BMMCs and the levels of IFN-γ and IL-10 in culture supernatant were determined by ELISA. Results The mean level of mHLA-G after G-CSF mobilization in the PBC or BM cells was significantly higher than that before G-CSF mobilization (P<0.05). The level of mHLA-G or sHLA-G in BM cells was higher than that in PBC after G-CSF mobilization (P<0.05). The level of mHLA-G or sHLA-G in BMMCs or PBSCs which were stimulated by G-CSF was higher than that of the controls (P<0.05), and the level of HLA-G in BMMCs was higher than that in PBSCs. HLA-G predominant expressed in CD3+ T cells; The results of allogeneic mixed lymphocyte culture revealed that immunological function of the separated T lymphocytes which expressed high level of HLA-G was inhibited (RPI: 54.3%). The separated T lymphocytes co-cultured with allogeneic BMMCs, the levels of IFN-γ and IL-10 in culture supernatant were significantly higher than the controls (P<0.05). Conclusions HLA-G is rich in G-BM that might be interpret G-BM could induce better immunotolerance than G-PBSC. The G-CSF could regulate HLA-G expression and secretion directly. The mechanism of G-CSF inducing immunotolerance might be related to the inhibition of allogeneic T cell reactivity and the increase of IFN-γ and IL-10 secretion through HLA-G. Disclosures: No relevant conflicts of interest to declare.

Differential Role for VLA-5 in Mobilization of Heamotopoieic Stem Cells Toward Peripheral Blood and Homing to Bone Marrow and Spleen.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 2190-2190 ◽  
Author(s):  
Pieter K. Wierenga ◽  
Ellen Weersing ◽  
Bert Dontje ◽  
Gerald de Haan ◽  
Ronald P. van Os
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Stem Cell ◽  
Progenitor Cells ◽  
Peripheral Blood ◽  
Hematopoietic Stem ◽  
Late Antigen ◽  
Cell Mobilization

Abstract Adhesion molecules have been implicated in the interactions of hematopoietic stem and progenitor cells with the bone marrow extracellular matrix and stromal cells. In this study we examined the role of very late antigen-5 (VLA-5) in the process of stem cell mobilization and homing after stem cell transplantation. In normal bone marrow (BM) from CBA/H mice 79±3 % of the cells in the lineage negative fraction express VLA-5. After mobilization with cyclophosphamide/G-CSF, the number of VLA-5 expressing cells in mobilized peripheral blood cells (MPB) decreases to 36±4%. The lineage negative fraction of MPB cells migrating in vitro towards SDF-1α (M-MPB) demonstrated a further decrease to 3±1% of VLA-5 expressing cells. These data are suggestive for a downregulation of VLA-5 on hematopoietic cells during mobilization. Next, MPB cells were labelled with PKH67-GL and transplanted in lethally irradiated recipients. Three hours after transplantation an increase in VLA-5 expressing cells was observed which remained stable until 24 hours post-transplant. When MPB cells were used the percentage PKH-67GL+ Lin− VLA-5+ cells increased from 36% to 88±4%. In the case of M-MPB cells the number increased from 3% to 33±5%. Although the increase might implicate an upregulation of VLA-5, we could not exclude selective homing of VLA-5+ cells as a possible explanation. Moreover, we determined the percentage of VLA-5 expressing cells immediately after transplantation in the peripheral blood of the recipients and were not able to observe any increase in VLA-5+ cells in the first three hours post-tranpslant. Finally, we separated the MPB cells in VLA-5+ and VLA-5− cells and plated these cells out in clonogenic assays for progenitor (CFU-GM) and stem cells (CAFC-day35). It could be demonstared that 98.8±0.5% of the progenitor cells and 99.4±0.7% of the stem cells were present in the VLA-5+ fraction. Hence, VLA-5 is not downregulated during the process of mobilization and the observed increase in VLA-5 expressing cells after transplantation is indeed caused by selective homing of VLA-5+ cells. To shed more light on the role of VLA-5 in the process of homing, BM and MPB cells were treated with an antibody to VLA-5. After VLA-5 blocking of MPB cells an inhibition of 59±7% in the homing of progenitor cells in bone marrow could be found, whereas homing of these subsets in the spleen of the recipients was only inhibited by 11±4%. For BM cells an inhibition of 60±12% in the bone marrow was observed. Homing of BM cells in the spleen was not affected at all after VLA-5 blocking. Based on these data we conclude that mobilization of hematopoietic progenitor/stem cells does not coincide with a downregulation of VLA-5. The observed increase in VLA-5 expressing cells after transplantation is caused by preferential homing of VLA-5+ cells. Homing of progenitor/stem cells to the bone marrow after transplantation apparantly requires adhesion interactions that can be inhibited by blocking VLA-5 expression. Homing to the spleen seems to be independent of VLA-5 expression. These data are indicative for different adhesive pathways in the process of homing to bone marrow or spleen.

Novel Double Labeled Mixed Lymphocyte in Vitro Assay to Predict the Dominant Graft in 2 Unit UCB Transplantation

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 4699-4699
Author(s):  
Shicheng Yang ◽  
Xiao Huang ◽  
Hongyan Lu ◽  
Amandeep Salhotra ◽  
Alexander Wendling ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
T Cells ◽  
Peripheral Blood ◽  
In Vitro Assay ◽  
Proliferation Index ◽  
Vitro Assay ◽  
Hematopoietic Stem ◽  
Ifn Γ

Abstract Abstract 4699 Introduction: Umbilical cord blood cells (UCB) from allogeneic donors have been established as an alternative source for HSC transplantation in patients who lack suitably HLA matched bone marrow or peripheral blood stem cells from adult donors. Transplantation using 2 unit UCB has been shown to compensate the low engraftment and slow hematopoietic recovery resulting from 1 unit UCB transplantation in full stature adult patients. At present, there are no unit specific factors that reliably predicts for the “winning unit” in 2 unit UCB transplantation, e.g. cell viability, number of infused total nucleated cells, CD34+ or CD3+ cells, sex mismatch, ABO blood group, and degree of HLA mismatch. In vivo mouse models suggest that CD34 negative subsets play an important role. Among CD34 negative subsets, CD8 T subset accounts for approximately 34.0+/−23.3% of T lymphocytes from UCB. In bone marrow transplantation CD8 T cells have been found to facilitate donor hematopoietic cell engraftment. Moreover, it has been reported that 1 dominant unit coincides with a specific CD8 T cell response against the non-engrafted unit which was not observed from CD4 or NK cells. Methods: In this study, we used volunteer donated UCB research units (kindly provided by P. Rubinstein, MD, New York Blood Center). Mononuclear cells (MNC) were purified by Ficoll gradient centrifugation, and CD3 T cells were isolated with CD3 MicroBeads (Miltenyi Biotec; autoMACS). The purified CD3 (confirmed by FACS >95% purity) cells were labeled with CFSE and DDAO-SE. After labeling, the cells from two different donors were mixed in 96-well U-bottom plates for continued culture in 37 °C 5% CO2. The expansion from each labeled donor cells was evaluated using flow cytometry; the dead cells were gated out using propidium iodide, and the data was analyzed using FlowJo software. For proper T cells activation, we also compared different activation conditions using i.) anti-CD3/CD28 Beads, ii.) anti-CD3 antibody plus anti-CD28 antibody, and iii.) cytokine IL-2. The schematic illustration of methods is shown in Figure 1. Results and discussion: We noted that T cells from UCB are primarily at naïve stage as determined by CD45RA (93.8 +/− 7.11%) and CCR7 (84.9 +/− 12.0%) expression. We also determined the optimal activation condition using a modified mixed lymphocyte reaction from 2 UCB units. Four days after incubation, the proliferation from 2 units labeled with CFSE and DDAO-SE could be reproducibly distinguished using FL1 channel for CFSE and FL4 channel for DDAO-SE (Figure 1). The optimal concentration for labeling using CFSE (1 mM) and DDAO (1 μM or 3 mM) was determined by titration. To avoid cell toxicity resulting from CFSE and DDAO-SE labeling, as well as self-crossing from each donor using two dyes, we examined additional mixed lymphocyte analyses in which each donor was labeled with CFSE or DDAO-SE respectively and vice versa. As shown in Figure 1, we found consistently that the predicated dominant unit accounted for the majority of culture (73.2% stained with DDAO; 63.5% stained with CFSE) after 4 days co-culture. The dominance was not correlated with cell proliferation indicated by the proliferation index (1.12 for dominant and 1.48 for another unit). After confirmation of this in vitro assay, further studies were conducted to evaluate the IFN-γ release of 2 UCB units in this optimized mixed lymphocyte assay in the condition using cytokine IL-2. Interestingly, we could only detect IFN-γ by intracellular staining in one unit when co-culture was set-up using CD3 T cells from each unit; the expression of IFN-γ was not detected when we used CD3 T cells from 1 unit. The correlation between dominance and the expression of IFN-γ is currently under investigation. Conclusion: UCB Transplantation is an important alternative for patients lacking bone marrow or peripheral blood stem cell donors. With the establishment of this novel modified mixed lymphocyte in vitro assay for prediction of the “winning” immune dominant unit, routine analyses can be performed to guide unit selection. Further interventions can be exploited to preferentially treat the expected dominant unit with glycosylation, cytokines, prostaglandins, or C3a compliments to further enhance hematopoietic stem cells trafficking and engraftment to the marrow. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Polyfunctional T Lymphocytes Are in the Peripheral Blood of Donors Naturally Immune to Coccidioidomycosis and Are Not Induced by Dendritic Cells

2009 ◽  
Vol 78 (1) ◽  
pp. 309-315 ◽  
Author(s):  
Lance Nesbit ◽  
Suzanne M. Johnson ◽  
Demosthenes Pappagianis ◽  
Neil M. Ampel
Keyword(s):  
Dendritic Cells ◽  
T Lymphocytes ◽  
Peripheral Blood ◽  
Mononuclear Cells ◽  
Interleukin 2 ◽  
High Expression ◽  
Peripheral Blood Mononuclear ◽  
Necrosis Factor Alpha ◽  
Ifn Γ

ABSTRACT Coccidioidomycosis is a fungal infection endemic in the southwestern United States that is increasing in incidence. While cellular immunity correlates with protection from clinical illness, the precise elements of that response are undefined. Using the coccidioidal antigen preparation T27K and multiparametric flow cytometry, the in vitro frequency of polyfunctional T lymphocytes in the peripheral blood of naturally immune healthy donors and those who were nonimmune was determined. Polyfunctional CD4 lymphocytes, defined as producing intracellular interleukin 2 (IL-2), gamma interferon (IFN-γ), and tumor necrosis factor alpha simultaneously, had a frequency of 137 per 400,000 events among peripheral blood mononuclear cells (PBMC) of immune donors compared to 11 per 400,000 PBMC from nonimmune donors (P = 0.03). When monocyte-derived mature dendritic cells pulsed with T27K (mDCT27K) were used for antigen presentation, the frequency of polyfunctional CD4 T lymphocytes did not significantly increase for either group, although mDCT27K did significantly increase the concentrations of IL-2 and IFN-γ released by PBMC from nonimmune donors (P = 0.02). After in vitro stimulation with T27K, polyfunctional CD4 and CD8 lymphocytes of PBMC from immune donors had a mixture of low- and high-expression CCR7 cells, suggesting both effector and central memory, compared with predominantly high-expression CCR7 cells when PBMC were incubated with the mitogen phytohemagglutinin (P = 0.03). These data demonstrate the presence of polyfunctional T lymphocytes in the peripheral blood of individuals with coccidioidal immunity and suggest a model for the in vitro testing of vaccine candidates for coccidioidomycosis.

scholarly journals Characterization of the potentiation effect of activin on human erythroid colony formation in vitro

Blood ◽  
1989 ◽  
Vol 73 (4) ◽  
pp. 952-960 ◽  
Author(s):  
J Yu ◽  
L Shao ◽  
J Vaughan ◽  
W Vale ◽  
AL Yu
Keyword(s):  
Bone Marrow ◽  
T Lymphocytes ◽  
Dna Synthesis ◽  
Peripheral Blood ◽  
Colony Formation ◽  
Mononuclear Cells ◽  
Erythroid Progenitors ◽  
Potentiation Effect ◽  
Proliferation And Differentiation

Activin, also named FSH-releasing protein, was previously shown to induce hemoglobin accumulation in K562 cells and potentiate the proliferation and differentiation of CFU-E in human bone marrow cultures. Present studies indicate that the potentiation effect of activin is lineage specific. In addition to CFU-E, activin caused an increase in the colony formation of BFU-E from either bone marrow or peripheral blood. It had little effect on the colony formation of CFU- GM and the mixed colonies from CFU-GEMM. In serum-depleted culture, the effect of activin was shown to be dose-dependent with doses effective at picomolar concentrations. The potentiation effect of activin was exerted indirectly through mediation of both monocytes and T lymphocytes. Activin was also found to increase specifically the proportion of DNA-synthesizing erythroid progenitors from both bone marrow and peripheral blood. It had little effect on DNA synthesis in CFU-GM and in mitogen-stimulated lymphocytes. Addition of the monocytes or T lymphocytes to their respective depleted subpopulations of mononuclear cells reconstituted the enhancing effect of activin on the colony formation and DNA synthesis of erythroid progenitors. These results strongly suggest a specific role of activin in potentiating the proliferation and differentiation of erythroid progenitors in vitro.

Homing and Engraftment of Mobilized Hematopoietic Stem Cells: Involvement of VLA-5.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 4943-4943
Author(s):  
Pieter K. Wierenga ◽  
Gerald de Haan ◽  
Bert Dontje ◽  
Ellen Weersing ◽  
Ronald van Os
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Stem Cell ◽  
Progenitor Cells ◽  
Peripheral Blood ◽  
Hematopoietic Stem ◽  
Post Transplant ◽  
Hematopoietic Reconstitution

Abstract VLA-5 has been implicated in the adhesive interactions of stem and progenitor cells with the bone marrow extracellular matrix and stromal cells and is therefore considered to play an important role in the hematopoietic reconstitution after stem cell transplantation. In normal bone marrow (BM) from CBA/H mice 79±3% of the cells in the lineage negative fraction express VLA-5. After mobilization with cyclophosphamide/G-GSF, the number of VLA-5 expressing cells in mobilized peripheral blood cells (MPB) decreases to 38±3%. Despite this low frequency of VLA-5+ cells, however, even when equal numbers of progenitor cells are transplanted MPB cells provide a much faster hematopoietic recovery compared to BM cells. To shed more light on the role of VLA-5 in the process of homing and engraftment, we investigated whether differences in homing potential of the stem cell subsets might be responsible for this enhanced reconstitution. At 3 hours post-transplant, however, no differences in homing efficiency of progenitor and stem cells from MPB and BM grafts in both bone marrow and spleen could be detected. It should be realized that MPB and BM grafts demonstrate different ratios of stem/progenitor cells which might be another explanation for the observed differences in repopulation potential. Furthermore, MPB cells migrating in vitro towards SDF-1α showed potent reconstitution while VLA-5 expression was reduced on these cells. In fact, in vitro treatment with SDF-1α showed further decrease in VLA-5 expressing cells (from 38% to 4%) in the lin- fraction. When equal numbers of MPB were transplanted with and without SDF-1α pretreatment, no difference in hematopoietic reconstitution was observed suggesting a minor role of VLA-5 in homing and engraftment. On the other hand, after VLA-5 blocking an inhibition of 59±7% in the homing of MPB progenitor cells in the bone marrow could be found, whereas homing in the spleen of the the recipients is only inhibited by 11±4%. To elucidate whether the observed enhanced reconstitution could be explained by a selective homing of VLA-5+ cells or a rapid upregulation of VLA-5 expression, cells were labelled with PKH67-GL and transplanted in lethally irradiated recipients. It could be demonstrated that at 3 hours post-transplant cells from MPB grafts showed a rapid increase from 38±3% up to 66±9% of VLA-5+ cells in the bone marrow of the recipient. In the spleen no significant increase in VLA-5+ cells was observed. When MPB cells were transplanted after pretreatment with SDF-1α an increase from 2±1% up to 33±5% of VLA-5+ cells in the bone marrow was detected. When calculating the number of cells recovered from bone marrow, a selective homing of VLA-5+ cells cannot be excluded. Therefore, we also assessed the number of VLA-5+ cells in the PKH+ fraction in peripheral blood from the recipient immediately (½-1 hour) after transplantation but found no increase during that time period. So far it can be concluded that MPB cells show low number of VLA-5+ cells but these cells possess an enhanced hematopoietic reconstitution potential. Homing of progenitor cells to the spleen seems to be less dependent on VLA-5 expression than homing to the bone marrow. A rapid upregulation of VLA-5 expression on engrafting MPB cells early after transplantation does not occur and hence our data are suggestive for the preferential homing of VLA-5+ cells in the bone marrow after transplantation.

scholarly journals The role of chorionic gonadotropin in the formation of immunological tolerance during pregnancy

2011 ◽  
Vol 57 (5) ◽  
pp. 52-56 ◽  
Author(s):  
S V Shirshev ◽  
S A Zamorina
Keyword(s):  
T Lymphocytes ◽  
Chorionic Gonadotropin ◽  
Mononuclear Cells ◽  
Active Oxygen Species ◽  
Relative Number ◽  
Immunological Tolerance ◽  
Interleukin 4 ◽  
Ifn Γ

We have studied the role of pregnancy hormone (chorionic gonadotropin, CG) in the control of expression of Foxp3 (a marker of T-regulatory lymphocytes, Treg), synthesis of interleukin-4 (IL-4) and interferon-gamma (IFH-γ) in lymphocytes, and secretion of indolamine-2,3-dioxygenase (IDO) by monocytes. Moreover, we evaluated the phagocytic and oxidative activities of these cells. It was shown that incubation of CG at a dose of 100 IU/ml with mononuclear cells from peripheral blood (MPC) resulted in a rise in the relative number of IL-4+CD3+CD4+T-lymphocytes whereas the number of IFN-γ+CD3+CD4+T-cells remained unaltered; the intracellular production of IFN-γ and IL-4 in the subpopulation of CD3+CD8+T-lymphocytes did not change either. In vitro, CG (10 and 100 IU/ml) significantly increased percentage of CD4+CD25 brightFoxp3+ cells and enhanced activity of indolamine-2,3-doxygenase in lipopolysaccharide-stimulated MPC. CG suppressed phagocitosis of E. coli lux+ by monocytes and simultaneously inhibited the production of active oxygen species in the reaction of spontaneous luminol-dependent chemiluminescence. Taken together, these properties of CG account for its role in the promotion of the formation of immunological tolerance during pregnancy.

Mobilization of Human Hemopoietic Progenitor Stem Cell Via Modulating CXCR4/SDF-1 Using Galactofucan Sulfate.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 1971-1971
Author(s):  
Mohammad R. Irhimeh ◽  
Ray M. Lowenthal ◽  
J. Helen Fitton
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Peripheral Blood ◽  
Mononuclear Cells ◽  
Autologous Transplantation ◽  
White Blood Cells ◽  
Healthy Human ◽  
Cytokine Analysis ◽  
Ifn Γ

Abstract Mobilization of peripheral blood progenitor cells for hematopoietic rescue following autologous transplantation is usually achieved with the chemokine G-CSF. Engraftment potential is increased when higher levels of the receptor CXCR4 are noted on the hematopoietic progenitor stem cells (HPCs). It was previously reported that intravenous treatment with fucoidan or dextran sulfate, which are sulfated high molecular weight compounds, increased numbers of circulating mature white blood cells and HPCs in mice and nonhuman primates. This treatment also led to an increase in the pro-inflammatory cytokines IFN-γ and IL-12 levels in mice. In vitro treatment of bone marrow mononuclear cells with IFN-γ can up-regulate the expression of CXCR4 on granulocyte precursors and monocytes. We obtained ethics approval and informed consent to study the mobilization effect of orally ingested GFSTM (Galactofucan Sulfate), a seaweed-derived fucoidan, in healthy human volunteers in a single blinded placebo controlled phase I/II clinical study. Flow cytometry was used to monitor CXCR4 receptor on CD34+ stem cells. When moderate quantities (3 g/day) of seaweed containing 10% GFSTM were ingested, a slight increase in the total number of HPCs (CD34+) in the peripheral blood (PB) was observed, from 1.38 to 1.69 cells/μL (p=0.22, n=6). In addition, there was a small increase in the percentage of HPCs that expressed CXCR4 surface receptor, from 0.59 to 1.47 cells/μL, which is equivalent to 43% to 63% (p=0.19, n=6). Moreover, when 3 g/day of 75% GFSTM was ingested, a greater increase in the total number of HPCs (CD34+) in PB was observed, from 1.65 to 1.84 cells/μL (p=0.04, n=23). Furthermore, the percentage of HPCs that expressed CXCR4 increased from 0.746–1.652 cells/μL, which is equivalent to 45% to 90% (p=0.0002, n=23). Cytokine analysis, which was performed using ELISA to test for SDF-1 and IFN-γ, showed a significant increase in the plasma level of these cytokines. SDF-1 level was elevated from 1979 to 2068 pg/mL (p=0.051, n=10) and the level of IFN-γ from 9.04 to 9.90 pg/mL (p=0.007, n=10). These results suggest that GFSTM may modulate CXCR4 or disturb the SDF-1 gradient between bone marrow and PB. IFN-γ might play a role in the up-regulation of the expression of CXCR4 on CD34+ cells. To the authors’ knowledge, this is the first report of mobilization of HPCs by disruption of CXCR4/SDF-1 interaction using oral fucoidan.

scholarly journals Characterization of the potentiation effect of activin on human erythroid colony formation in vitro

Blood ◽  
1989 ◽  
Vol 73 (4) ◽  
pp. 952-960 ◽  
Author(s):  
J Yu ◽  
L Shao ◽  
J Vaughan ◽  
W Vale ◽  
AL Yu
Keyword(s):  
Bone Marrow ◽  
T Lymphocytes ◽  
Dna Synthesis ◽  
Peripheral Blood ◽  
Colony Formation ◽  
Mononuclear Cells ◽  
Erythroid Progenitors ◽  
Potentiation Effect ◽  
Proliferation And Differentiation

Abstract Activin, also named FSH-releasing protein, was previously shown to induce hemoglobin accumulation in K562 cells and potentiate the proliferation and differentiation of CFU-E in human bone marrow cultures. Present studies indicate that the potentiation effect of activin is lineage specific. In addition to CFU-E, activin caused an increase in the colony formation of BFU-E from either bone marrow or peripheral blood. It had little effect on the colony formation of CFU- GM and the mixed colonies from CFU-GEMM. In serum-depleted culture, the effect of activin was shown to be dose-dependent with doses effective at picomolar concentrations. The potentiation effect of activin was exerted indirectly through mediation of both monocytes and T lymphocytes. Activin was also found to increase specifically the proportion of DNA-synthesizing erythroid progenitors from both bone marrow and peripheral blood. It had little effect on DNA synthesis in CFU-GM and in mitogen-stimulated lymphocytes. Addition of the monocytes or T lymphocytes to their respective depleted subpopulations of mononuclear cells reconstituted the enhancing effect of activin on the colony formation and DNA synthesis of erythroid progenitors. These results strongly suggest a specific role of activin in potentiating the proliferation and differentiation of erythroid progenitors in vitro.

scholarly journals Primitive Human Hematopoietic Cells Are Enriched in Cord Blood Compared With Adult Bone Marrow or Mobilized Peripheral Blood as Measured by the Quantitative In Vivo SCID-Repopulating Cell Assay

Blood ◽  
1997 ◽  
Vol 89 (11) ◽  
pp. 3919-3924 ◽  
Author(s):  
Jean C.Y. Wang ◽  
Monica Doedens ◽  
John E. Dick
Keyword(s):  
Bone Marrow ◽  
Cord Blood ◽  
Peripheral Blood ◽  
Hematopoietic Cells ◽  
Allogeneic Transplantation ◽  
Functional Assay ◽  
Hematopoietic Stem ◽  
Mobilized Peripheral Blood

Abstract We have previously reported the development of in vivo functional assays for primitive human hematopoietic cells based on their ability to repopulate the bone marrow (BM) of severe combined immunodeficient (SCID) and nonobese diabetic/SCID (NOD/SCID) mice following intravenous transplantation. Accumulated data from gene marking and cell purification experiments indicate that the engrafting cells (defined as SCID-repopulating cells or SRC) are biologically distinct from and more primitive than most cells that can be assayed in vitro. Here we demonstrate through limiting dilution analysis that the NOD/SCID xenotransplant model provides a quantitative assay for SRC. Using this assay, the frequency of SRC in cord blood (CB) was found to be 1 in 9.3 × 105 cells. This was significantly higher than the frequency of 1 SRC in 3.0 × 106 adult BM cells or 1 in 6.0 × 106 mobilized peripheral blood (PB) cells from normal donors. Mice transplanted with limiting numbers of SRC were engrafted with both lymphoid and multilineage myeloid human cells. This functional assay is currently the only available method for quantitative analysis of human hematopoietic cells with repopulating capacity. Both CB and mobilized PB are increasingly being used as alternative sources of hematopoietic stem cells in allogeneic transplantation. Thus, the findings reported here will have important clinical as well as biologic implications.

scholarly journals Transcription factor Gfi-1 induced by G-CSF is a negative regulator of CXCR4 in myeloid cells

Blood ◽  
2007 ◽  
Vol 110 (7) ◽  
pp. 2276-2285 ◽  
Author(s):  
Maria De La Luz Sierra ◽  
Paola Gasperini ◽  
Peter J. McCormick ◽  
Jinfang Zhu ◽  
Giovanna Tosato
Keyword(s):  
Bone Marrow ◽  
Dna Sequences ◽  
Peripheral Blood ◽  
Cell Function ◽  
Myeloid Cell ◽  
Cxcr4 Expression ◽  
Negative Regulator ◽  
Hematopoietic Stem

The mechanisms underlying granulocyte-colony stimulating factor (G-CSF)–induced mobilization of granulocytic lineage cells from the bone marrow to the peripheral blood remain elusive. We provide evidence that the transcriptional repressor growth factor independence-1 (Gfi-1) is involved in G-CSF–induced mobilization of granulocytic lineage cells from the bone marrow to the peripheral blood. We show that in vitro and in vivo G-CSF promotes expression of Gfi-1 and down-regulates expression of CXCR4, a chemokine receptor essential for the retention of hematopoietic stem cells and granulocytic cells in the bone marrow. Gfi-1 binds to DNA sequences upstream of the CXCR4 gene and represses CXCR4 expression in myeloid lineage cells. As a consequence, myeloid cell responses to the CXCR4 unique ligand SDF-1 are reduced. Thus, Gfi-1 not only regulates hematopoietic stem cell function and myeloid cell development but also probably promotes the release of granulocytic lineage cells from the bone marrow to the peripheral blood by reducing CXCR4 expression and function.

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