Elevation of Plasma Leukotriene B4 on G-CSF-Induced Hematopoietic Stem Cells Mobilization In Normal Individuals

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 4437-4437
Author(s):  
Yeung-Chul Mun ◽  
Jee Young Ahn ◽  
Solip Lee ◽  
Eun-Sun Yoo ◽  
Jung Yeon Cho ◽  
...  
Keyword(s):  
Stem Cells ◽  
Murine Model ◽  
Peripheral Blood ◽  
Receptor Activation ◽  
Enzyme Linked Immunosorbent Assay ◽  
Base Line ◽  
Receptor Antagonists ◽  
Hematopoietic Stem ◽  
Healthy Donors ◽  
Ltb4 Receptor

Abstract Abstract 4437 Background: Previously, we had reported that mobilization kinetics of CD34+ cells association of CD44 and CD31 expression during continuous intravenous administration of G-CSF in normal donors(Stem Cells 18:281-286, 2000, BMT 36:1027-1032, 2005). Meanwhile, a number of studies have reported the mechanisms of G-CSF-induced HSCs mobilization, but the underlying mechanisms are not clear yet. Some chemokines, macrophage inflammatory protein-1α (MIP-1α/CCL3), stem cell-derived factor 1α (SDF-1α/CCL12), interleukin 8 (IL-8/CXCL8), and GROβ/CXCL2, can mobilize HSCs, and we also reported that LTB4, which has considerable functional overlap with the chemokine family of chemoattractant peptides, had mobilized HSCs in murine model. In this study, we have investigated whether G-CSF may affect plasma LTB4 level during HSCs mobilization and G-CSF-induced HSCs mobilization may be modulated by LTB4. Method: To investigate that G-CSF may modulate plasma LTB4 level, G-CSF (Filgrastim, Kirin Brewery Co. Tokyo, Japan; 10 microgram/kg/day) was administered subcutaneously into 4 healthy donors for 5 days and then, apheresis were performed on day 4 and 5, and were collected for their serum plasma at base line, 24hours, 48hours and 72hours after 1st dose of G-CSF. LTB4 concentration was measured by enzyme-linked immunosorbent assay (Parameter™ LTB4 Assay; R&D Systems, Mineapolis, MN, USA). Meanwhile, to evaluate the effect of LTB4 inhibition on G-CSF-induced HSCs mobilization, LTB4APA and U75302, which are LTB4 receptor antagonists were given to C57BL/6 mice followed by G-CSF 5μ g or LTB4 1μ g administration intravenously 2 hours later. 24 hours after the G-CSF injection or 4 hours after LTB4 injection, peripheral blood samples were obtained and analyzed for HSCs mobilization by flow cytometry using Sca-1, CD45R(B220), CD116, Gr-1 and TER119. Results: Plasma LTB4 levels of healthy donors demonstrated increases in 24hrs after G-CSF administration; 415.3±112.1 pg/ml before treatment of G-CSF and 706.4±154.7 pg/ml 24hours after 1st dose of G-CSF (p=0.005), and then plasma LTB4 levels were decreased continuously after peak level on 24hours. Meanwhile, when LTB4 receptor antagonists were given, the number of HSCs were decreased in the G-CSF mobilized mice; 4.69×103 cells/ml blood before treatment of G-CSF, 80.08×103 cells/ml blood after treatment G-CSF without LTB4 receptor antagonist, 9.24×103 cells/ml blood after treatment of G-CSF with LTB4APA 1μ g respectively(p<0.05, compared with the data of G-CSF alone), 22.86×103 cells/ml blood after treatment of G-CSF with U75302 1μ g (p<0.05, compared with the data of G-CSF alone). The blocking effects on mobilization of HSCs by LTB4 receptor antagonists were also demonstrated in the LTB4 mobilized mice (data not shown). Conclusions: We observed that G-CSF increases plasma LTB4 levels during HSCs mobilization in healthy donors, and that LTB4 inhibition by LTB4 receptor antagonists in murine model downregulate the G-CSF-induced HSCs mobilization. These indicate that LTB4 may be involved in the downstream pathway of G-CSF-induced HSCs mobilization. Through the our results, we hypothesize that G-CSF increase LTB4 level in plasma during HSCs mobilization, and LTB4 receptor activation by increased LTB4 level in plasma may contribute HSCs mobilization in vivo. Currently, we are investigating the cellular and molecular mechanism(s) of potential role of LTB4 during G-CSF induced peripheral blood progenitor cell mobilization. Disclosures: No relevant conflicts of interest to declare.

Successful Hematopoietic STEM CELL Mobilization with GRANULOCYTE Colony Stimulating FACTOR PLUS Plerixafor (MOZOBIL, AMD3100) IN POOR Mobilizers: A Single CENTER STUDY

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 4444-4444
Author(s):  
Despina Mallouri ◽  
Ioanna Sakellari ◽  
Chrisa Apostolou ◽  
Panayotis Baliakas ◽  
Apostolia Papalexandri ◽  
...  
Keyword(s):  
Stem Cells ◽  
Peripheral Blood ◽  
Median Number ◽  
Granulocyte Colony Stimulating Factor ◽  
Colony Stimulating Factor ◽  
Hematopoietic Stem ◽  
Cd 34 ◽  
Healthy Donors ◽  
Stimulating Factor ◽  
Poor Mobilizers

Abstract Abstract 4444 Background: According to published data mobilization of sufficient number of CD 34+ cells with cytokines alone or with chemo-mobilization fails in 5–30% of patients. Plerixafor is a novel chemokine receptor 4 antagonist (CXCR4) that reversibly inhibits the interaction with its ligand SDF-1 (Stromal Derived Factor 1). Phase III studies have demonstrated that plerixafor combined with granulocyte-colony stimulating factor (GCSF) improves CD 34+ cell collection in patients with Multiple Myeloma (MM) or Non Hodgkin Lymphoma (NHL). It has been shown to be efficacious in combination with GCSF to mobilize adequate number of CD 34+ cells in patients proven to be poor mobilizers yielding a success rate of 60–100% in several reports. Plerixafor is currently approved for administration in combination with GCSF to enhance mobilization of hematopoietic stem cells in patients with lymphoma and MM whose cells mobilize poorly. Patients/methods: We administered plerixafor in combination with GCSF in 14 patients (in 4/14 as part of compassionate use protocol) and 2 sibling donors after an informed consent was obtained. Individuals were defined as poor mobilizers either due to unsuccessful collection of CD34+ >2×106/kg or due to peripheral blood CD34+ peak <20/μ l in spite of adequate mobilization treatment. Data of the individuals were collected retrospectively. Eight patients suffered from MM, 3 of NHL, 3 of H°dgkin Lymphoma (HL). Marrow involvement was present in 1 patient suffering from MM. The median number of previous chemotherapy regimens was 4(1-8). Two patients had a history of previous autologous hematopoietic cell transplantation (autoHCT) and 4 patients had received multiple radiotherapy courses. Patients had a median of 2 (1-3) previous unsuccessful attempts of mobilization before plerixafor plus GCSF administration. Nine patients had received GCSF alone and 5 patients chemotherapy plus GCSF. Patients received GCSF for 4 consecutive days and plerixafor was administered at the evening of the forth day, 10–11 hours before the scheduled aphaeresis procedure. In case of not sufficient or suboptimal number of CD34+ cells collection the procedure was repeated for maximum of 3 days plerixafor administration (7 days of GCSF). Results: Mobilization with plerixafor plus GCSF and collection of adequate number of CD 34+ cells was successful in 12/14 patients. The median number of CD 34+ cells collected was 2.5×106/kg in a median of 2 (1-4) apheresis days. Two of 14 patients proceeded to a second mobilization with plerixafor plus GCSF, eventually succeeding a sufficient cell dose graft collection. In 2 sibling female donors, aged 47 and 54 years, after administration of GCSF 10μ g/kg/day for 5 consecutive days mobilization was poor and collection of a graft with an acceptable CD 34+ cell dose was not possible. Administration of plerixafor improved mobilization and eventually grafts consisting of 2.55 ×106 and 5.34×106 CD34+/kg were collected by apheresis. Patients reported a grade ≤ II according to WHO scale toxicity following plerixafor administration. Most common side effects were hyperhidrosis, facial numbness and abdominal pain. None of the two healthy donors reported any adverse side effect. Engraftment was uneventfully in predictive time according to our historical data. Discussion: In our experience mobilization with either cytokines alone or cytokines following chemotherapy fails in a number of otherwise eligible for transplantation patients, mostly heavily pretreated or with advanced disease. In addition a small minority of healthy donors with no identifiable risk factors for poor mobilization, also fail mobilization with GCSF. The combination of plerixafor and GCSF seems to augment peripheral blood stem cells mobilization in poor mobilizers and offers a new treatment to collect sufficient CD 34+ cells and benefit from the transplantation procedure Disclosures: Off Label Use: Plerixafor was used for the mobilization of two healthy sibling donors after a signed concept was optained, due to poor mobilization with GCSF and failure of addequate graft collection.

Allograft Cell Content and GVHD Differ in Murine Recipients of AMD3100 Versus G-CSF Mobilized Peripheral Blood Stem Cells.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 3108-3108
Author(s):  
A. Lundqvist ◽  
N. Hamoui ◽  
J. Andersson ◽  
M. Elshal ◽  
Y. Takahashi ◽  
...  
Keyword(s):  
Stem Cells ◽  
T Cells ◽  
T Cell ◽  
Murine Model ◽  
Peripheral Blood ◽  
Lower Percentage ◽  
Peripheral Blood Stem Cells ◽  
Hematopoietic Stem ◽  
Recipient Group ◽  
Blood Stem Cells

Abstract G-CSF is currently the preferred agent to mobilize peripheral blood stem cells (PBSC) for allogeneic hematopoietic cell transplantation. AMD3100, a selective antagonist of SDF-1, has recently proven to rapidly mobilize hematopoietic stem cells in both humans and mice. It is currently unknown whether GVHD will differ in recipients of T-cell replete allografts mobilized with AMD3100 versus G-CSF. Therefore, we investigated the effects of AMD3100 on GVHD in a murine model of PBSC transplantation in which Balb/c recipients received 15x106 splenocytes from allogeneic MHC matched B10.d2 mice following 950cGy of irradiation. Splenocytes from donor mice were harvested six hours after a single subcutaneous injection of AMD3100 (100μg). Controls consisted of Balb/c recipients of splenocytes harvested after five daily subcutaneous injections of G-CSF (10μg) or saline. In addition, one Balb/c cohort received splenocytes mobilized with the combination of G-CSF and AMD3100. Significantly higher numbers of stem cells (KLS cells; cKit+, lineage-, sca-1+) were mobilized after AMD3100 compared to saline controls (mean=32,300±3,900 vs. 14,700±4,900; p=0.02). The absolute number of KLS cells was significantly lower after AMD3100 mobilization compared to G-CSF (mean=52,400±8,600; p=0.03). No difference in number of KLS cells was observed in mice mobilized with AMD3100 and G-CSF compared to G-CSF alone. Splenocytes from G-CSF mobilized B10.d2 donors contained a significantly (p&lt;0.01) lower percentage of T cells (15.9±3.1%) than AMD3100 mobilized donors (20.3±3.5%). The incidence of skin GVHD was higher in Balb/c recipients of AMD3100 mobilized splenocytes (19/20) compared to G-CSF mobilized splenocytes (5/18; p&lt;0.01), while recipients of donors mobilized with the combination of G-CSF and AMD3100 had a slightly higher incidence of skin GVHD (9/20, p&lt;0.01) compared to G-CSF alone. Using a cumulative clinical GVHD scoring system (maximum 9 points), the severity of GVHD was higher in mice receiving AMD3100 compared to mice receiving G-CSF (day +45 mean score=1.8 vs. 0.4 respectively; p&lt;0.01) or G-CSF + AMD3100 (mean score=0.8; p=0.03) mobilized splenocytes. When the T cell dose was adjusted to equal numbers in all transplant groups, the difference in GVHD between cohorts was less pronounced; 8/8 (100%) in the AMD3100 recipient group compared to 4/9 (44%; p=0.03) in the G-CSF recipient group developed skin GVHD. Th1/Th2 serum cytokine profiles following mobilization were similar in all donor groups. Compared to G-CSF, AMD3100, given alone or in combination with G-CSF, mobilized comparable numbers of CD4/CD25+ regulatory T cells with similar MLR suppressive effects. In contrast, the percentage of memory T cells (CD62L-) was significantly increased in mice mobilized with G-CSF (89.1±3.0%) compared to AMD3100 (43.9±1.3%), potentially accounting for the lower incidence of GVHD in recipients of G-CSF mobilized PBSC. This murine model provides the first insight into differences in GVHD that may occur when allogeneic transplantation is performed using T-cell replete PBSC allografts mobilized with AMD3100. Whether the higher incidence of GVHD observed in recipients of AMD3100 mobilized PBSC will enhance graft vs. tumor effects in tumor bearing Balb/c recipients is currently under investigation.

scholarly journals PIG-A mutations in normal hematopoiesis

Blood ◽  
2005 ◽  
Vol 105 (10) ◽  
pp. 3848-3854 ◽  
Author(s):  
Rong Hu ◽  
Galina L. Mukhina ◽  
Steven Piantadosi ◽  
Jamie P. Barber ◽  
Richard J. Jones ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
T Cells ◽  
Peripheral Blood ◽  
Paroxysmal Nocturnal Hemoglobinuria ◽  
Healthy Individuals ◽  
Hematopoietic Stem ◽  
Class A ◽  
Healthy Donors ◽  
Mobilized Peripheral Blood

AbstractParoxysmal nocturnal hemoglobinuria (PNH) is caused by phosphatidylinositol glycan–class A (PIG-A) mutations in hematopoietic stem cells (HSCs). PIG-A mutations have been found in granulocytes from most healthy individuals, suggesting that these spontaneous PIG-A mutations are important in the pathogenesis of PNH. It remains unclear if these PIG-A mutations have relevance to those found in PNH. We isolated CD34+ progenitors from 4 patients with PNH and 27 controls. The frequency of PIG-A mutant progenitors was determined by assaying for colony-forming cells (CFCs) in methylcellulose containing toxic doses of aerolysin (1 × 10-9 M). Glycosylphosphatidylinositol (GPI)–anchored proteins serve as receptors for aerolysin; thus, PNH cells are resistant to aerolysin. The frequency of aerolysin resistant CFC was 14.7 ± 4.0 × 10-6 in the bone marrow of healthy donors and was 57.0 ± 6.7 × 10-6 from mobilized peripheral blood. DNA was extracted from individual day-14 aerolysin-resistant CFCs and the PIG-A gene was sequenced to determine clonality. Aerolysin-resistant CFCs from patients with PNH exhibited clonal PIG-A mutations. In contrast, PIG-A mutations in the CFCs from controls were polyclonal, and did not involve T cells. Our data confirm the finding that PIG-A mutations are relatively common in normal hematopoiesis; however, the finding suggests that these mutations occur in differentiated progenitors rather than HSCs.

PIG-A Mutations in Normal Hematopoiesis.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 2825-2825
Author(s):  
Rong Hu ◽  
Galina Mukhina ◽  
Steven Piantadosi ◽  
Richard J. Jones ◽  
Robert A. Brodsky
Keyword(s):  
Stem Cells ◽  
Hematopoietic Stem Cells ◽  
Peripheral Blood ◽  
Genetic Fidelity ◽  
Large Mammals ◽  
Hematopoietic Stem ◽  
Differentiated Cells ◽  
Mutational Frequency ◽  
Healthy Donors ◽  
Normal Individuals

Abstract Introduction: Paroxysmal nocturnal hemoglobinuria (PNH) is caused by somatic mutations of the X-linked gene, PIG-A, in hematopoietic stem cells (HSCs). The product of this gene is necessary for the assembly of glycosylphosphatidylinositol (GPI) anchors. Consequently, PNH cells lack the expression of GPI-anchored proteins on their cell surface. PIG-A mutations have been found in granulocytes and T lymphocytes from most normal individuals. Although the significance of these mutations is unclear, it suggests that they are important in the pathogenesis of PNH. Methods: We isolated CD34+ progenitors from 4 PNH patients, 18 healthy donors, and 9 non-PNH patients undergoing peripheral blood stem cell tranplantation. The frequency of PIG-A mutant progenitors was determined by assaying for colony forming cells (CFC) in methylcellulose containing toxic doses of aerolysin. Aerolysin is a pore-forming toxin that uses the GPI anchor as it receptor; hence, PNH cells are unique in their resistance to aerolysin. DNA was extracted from individual day 14 aerolysin resistant CFC and the PIG-A gene was sequenced to determine clonality. We performed a Poisson distribution of the mutational frequency to determine the probability that the PIG-A mutation in controls arose from HSC. Results: In PNH patients, 67% of the CFC were aerolysin resistant. The frequency of aerolysin resistant CFC was 14.7 ± 4.0 x 10−6 in the bone marrow of healthy donors and was 57.0 ± 6.7 x 10−6 from mobilized peripheral blood. Aerolysin resistant CFC from PNH patients exhibited clonalPIG-A mutations, and thus, arose from HSC. In contrast, PIG-A mutations in the CFC from controls were polyclonal (up to 15 different mutations from one individual). Recent evidence suggests that humans and other large mammals possess only 10,000 primitive HSC, and that only 1000 of these cells are thought to contribute to hematopoiesis at any one time. Poisson statistics show that only 5% of normals would be expected to harbor a PIG-A mutation in 1000 HSC, and &lt; 1 x 10−9 persons would harbor 10 or more different PIG-A mutations even if all 10,000 hematopoietic stem cells were contributing to hematopoiesis. Thus, the high frequency of PIG-A mutations in controls, coupled with their polyclonality, suggests that they do not arise at the level of HSC; rather, PIG-A mutations in normals appear to arise as a consequence of hematopoietic differentiation, between the level of an HSC and a CFC. Conclusion: Our data confirm the findings that PIG-A mutations are relatively common in normal hematopoiesis. Although we cannot rule out that a rare PIG-A mutant blood cell in normals does in fact arise from a mutant HSC, our data suggest that most of the mutations occur with differentiation. Genetic fidelity can be lost with differentiation without consequence, as mutations in differentiated cells would not be propagated. These data also call into question the relevance of PIG-A mutations in normals to the pathogenesis of PNH.

High-activity samarium-153-EDTMP therapy followed by autologous peripheral blood stem cell support in unresectable osteosarcoma

2001 ◽  
Vol 40 (06) ◽  
pp. 215-220 ◽  
Author(s):  
S. Bielack ◽  
S. Flege ◽  
J. Eckardt ◽  
J. Sciuk ◽  
H. Jürgens ◽  
...  
Keyword(s):  
Stem Cells ◽  
Stem Cell ◽  
High Activity ◽  
Peripheral Blood ◽  
Peripheral Blood Stem Cell ◽  
Blood Stem Cell ◽  
High Dose ◽  
External Radiotherapy ◽  
Primary Tumor Site ◽  
Hematopoietic Stem

Summary Purpose: Despite highly efficacious chemotherapy, patients with osteosarcomas still have a poor prognosis if adequate surgical control cannot be obtained. These patients may benefit from therapy with radiolabeled phosphonates. Patients and Methods: Six patients (three male, three female; seven to 41 years) with unresectable primary osteosarcoma (n = 3) or unresectable recurrent sites of osteosarcomas (n = 3) were treated with high-activity of Sm-153-EDTMP (150 MBq/kg BW). In all patients autologous peripheral blood stem cells had been collected before Sm-153-EDTMP therapy. Results: No immediate adverse reactions were observed in the patients. In one patient bone pain increased during the first 48 hrs after therapy. Three patients received pain relief. Autologous peripheral blood stem cell reinfusion was performed on day +12 to +27 in all patients to overcome potentially irreversible damage to the hematopoietic stem cells. In three patient external radiotherapy of the primary tumor site was performed after Sm-153-EDTMP therapy and in two of them polychemotherapy was continued. Thirty-six months later one of these patients is still free of progression. Two further patients are still alive. However, they have developed new metastases. The three patients who had no accompanying external radiotherapy, all died of disease progression five to 20 months after therapy. Conclusion: These preliminary results show that high-dose Sm-153-EDTMP therapy is feasible and warrants further evaluation of efficacy. The combination with external radiation and polychemotherapy seems to be most promising. Although osteosarcoma is believed to be relatively radioresistant, the total focal dose achieved may delay local progression or even achieve permanent local tumor control in patients with surgically inaccessible primary or relapsing tumors.

Efficient lentiviral gene transfer to canine repopulating cells using an overnight transduction protocol

Blood ◽  
2004 ◽  
Vol 103 (10) ◽  
pp. 3710-3716 ◽  
Author(s):  
Peter A. Horn ◽  
Kirsten A. Keyser ◽  
Laura J. Peterson ◽  
Tobias Neff ◽  
Bobbie M. Thomasson ◽  
...  
Keyword(s):  
Stem Cells ◽  
Stem Cell ◽  
Gene Transfer ◽  
Peripheral Blood ◽  
Blood Cells ◽  
Lentiviral Vectors ◽  
Large Animal ◽  
Hematopoietic Stem ◽  
Lentiviral Transduction ◽  
Mobilized Peripheral Blood

Abstract The use of lentiviral vectors for the transduction of hematopoietic stem cells has evoked much interest owing to their ability to stably integrate into the genome of nondividing cells. However, published large animal studies have reported highly variable gene transfer rates of typically less than 1%. Here we report the use of lentiviral vectors for the transduction of canine CD34+ hematopoietic repopulating cells using a very short, 18-hour transduction protocol. We compared lentiviral transduction of hematopoietic repopulating cells from either stem cell factor (SCF)– and granulocyte-colony stimulating factor (G-CSF)–primed marrow or mobilized peripheral blood in a competitive repopulation assay in 3 dogs. All dogs engrafted rapidly within 9 days. Transgene expression was detected in all lineages (B cells, T cells, granulocytes, and red blood cells as well as platelets) indicating multilineage engraftment of transduced cells, with overall long-term marking levels of up to 12%. Gene transfer levels in mobilized peripheral blood cells were slightly higher than in primed marrow cells. In conclusion, we show efficient lentiviral transduction of canine repopulating cells using an overnight transduction protocol. These results have important implications for the design of stem cell gene therapy protocols, especially for those diseases in which the maintenance of stem cells in culture is a major limitation.

scholarly journals Long-term repopulating hematopoietic stem cells and “side population” in human steady state peripheral blood

2013 ◽  
Vol 11 (1) ◽  
pp. 625-633 ◽  
Author(s):  
Philippe Brunet de la Grange ◽  
Marija Vlaski ◽  
Pascale Duchez ◽  
Jean Chevaleyre ◽  
Veronique Lapostolle ◽  
...  
Keyword(s):  
Stem Cells ◽  
Steady State ◽  
Hematopoietic Stem Cells ◽  
Peripheral Blood ◽  
Side Population ◽  
Hematopoietic Stem
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