The aim of this research was to understand the influence of rhG-CSF on the sphingosine kinase (SphK) activity of monocytes. The peripheral blood monocytes were collected from 6 peripheral blood progenitor cell donors on the fifth day of mobilization with rhG-CSF and from 5 blood donors' buffy coats. The mRNA expressions of monocyte G-CSF receptor and SphK were tested with RT-PCR. The changes of SphK activity of monocytes were assayed after being treated with rhG-CSF. The results showed that the two kinds monocytes collected from both blood donors and peripheral blood progenitor cell donors mobilized with rhG-CSF expressed mRNA of G-CSF receptor and SphK. The SphK activity of monocytes collected from blood donors was not changed significantly after being treated with rhG-CSF (P > 0.05). The SphK activity of monocytes collected from peripheral blood progenitor cell donors transiently increased by (39.6 - 87.2)% after being treated by means of rhG-CSF (P < 0.05) without obviously dose-dependent effect. It is concluded that the SphK activity of monocytes collected from peripheral blood progenitor cell donors can be activated by rhG-CSF.
Granulocyte Colony-Stimulating Factor ; pharmacology ; Hematopoietic Stem Cell Mobilization ; Humans ; Monocytes ; cytology ; enzymology ; Phosphotransferases (Alcohol Group Acceptor) ; drug effects ; metabolism ; Receptors, Granulocyte Colony-Stimulating Factor ; biosynthesis ; genetics ; Recombinant Proteins

To investigate the effects of rhG-CSF on apoptosis of HL-60 cell induced by cytarabine (Ara-C), HL-60 cell were cultured with Ara-C, with or without rhG-CSF, and the changes of HL-60 cells were detected by morphology, including Wright's and acridine orange (AO) and ethidium bromide (EB) staining, and MTT assay. The results showed that comparing with Ara-C group, the apoptosis level of rhG-CSF + Ara-C group was enhanced (P < 0.01), and the OD value of MTT assay declined (P < 0.01). It is concluded that rhG-CSF can induce the apoptosis of HL-60 cells and enhance the apoptosis induced by Ara-C.
Antimetabolites, Antineoplastic ; pharmacology ; Apoptosis ; drug effects ; Cell Survival ; drug effects ; Cytarabine ; pharmacology ; Drug Synergism ; Granulocyte Colony-Stimulating Factor ; pharmacology ; HL-60 Cells ; Humans ; Recombinant Proteins

OBJECTIVETo investigate the effect of in vivo administration of rhG-CSF and/or rhIL-11 on mice immune system function.

METHODST cell subgroups, suppressor T cells (CD8+ CD28-, CD4+ CD25+, CD3+ CD4- CD8- T cells), expression of CD28 on T cells, and spleen T cells intracellular IL4/IFN-gamma secretion were determined by multicolor flow cytometry. MTT was used to determine the T cell proliferation capacity and mixed lymphocyte reactions.

RESULTSIn vivo administration of cytokines decreased the percentage of lymphocytes (P < 0.05), rhIL-11 and rhG-CSF in combination significantly decreased the CD4+/CD8+ ratio and increased the percentage of CD8+ CD28- suppressor T cells compared to either cytokine alone (P < 0.01). There was no difference in the percentage of CD3+ CD4- CD8- and CD4+ CD25+ suppressor T cells between either of the cytokines. Furthermore, cytokines treatments significantly decreased the capacities of splenic T cells proliferation and the response to alloantigens compared with the PBS treatment (P < 0.05), the combination group being more significantly decreased (P < 0.01). And cytokines treatment significantly decreased the production of IFN-gamma and increased the production of IL-4 compared with the PBS treatment(P < 0.05). The ratio of IFN-gamma/IL-4 were significantly decreased after the combination compared with either of them alone.

CONCLUSIONThe combination of rhIL-11 and rhG-CSF is potentially synergistic in the induction of immune tolerance by their effects on the proliferation capacity and function of T lymphocytes.

Animals ; CD28 Antigens ; metabolism ; Granulocyte Colony-Stimulating Factor ; pharmacology ; Immune Tolerance ; Interleukin-11 ; pharmacology ; Mice ; Mice, Inbred C57BL ; Recombinant Proteins ; T-Lymphocytes ; drug effects ; immunology ; metabolism

This study was purposed to evaluate the effects of recombinant human granulocyte colony-stimulating factor (rhG-CSF) on hematopoietic reconstruction and survival in beagles exposed to mixed fission neutron and γ-ray. 13 beagles were unilaterally exposed to single dose of 2.3 Gy 90% neutrons. The experiments were divided into 3 groups: irradiation control group (no any treatment, n = 4), supportive care group (n = 5) and rhG-CSF plus supportive care group (n = 4, abbreviated as rhG-CSF group) in which the beagles were subcutaneously injected with 200 µg/kg of rhG-CSF early at half an hour and 24 hours post-irradiation respectively. The results showed that 2.3 Gy 90% neutron irradiation induced a severe acute radiation sickness of bone marrow type. The administration of rhG-CSF increased the survival rate from 60% in supportive care group to 100%. Twice injection of rhG-CSF in the first 24 hours reduced duration of neutropenia, enhanced neutrophil nadir and promoted neutrophil recovery when compared with control cohort administered clinical support. The number of colony-forming cells (CFU-GM, CFU-E, and BFU-E) in peripheral blood of rhG-CSF treated canines increased 2-to 5-fold relative to those of the supportive care group on day 3. All canines treated with rhG-CSF achieved hematopoietic reconstruction as evidenced by the pathological section of sternum while severe shortage of hemopoietic cells remained in the cohorts given supportive care alone. It is concluded that the combination of supportive care and high-dose rhG-CSF can accelerate hematopoietic recovery and enhance survival of dogs exposed to 2.3 Gy mixed neutron and gamma ray.
Animals ; Dogs ; Gamma Rays ; adverse effects ; Granulocyte Colony-Stimulating Factor ; administration & dosage ; pharmacology ; Hematopoietic System ; drug effects ; radiation effects ; Neutron Diffraction ; Recombinant Proteins ; administration & dosage ; pharmacology ; Survival Rate

BACKGROUNDEndothelial progenitor cells (EPCs) derived from bone marrow may differentiate into endothelial cells and participate in endothelial repair. These cells can be mobilized into peripheral blood by cytokines, including granulocyte colony-stimulating factor (G-CSF). In the present study, we investigated the effects of G-CSF on neointimal formation and restenosis in a canine model of arterial balloon injury.

METHODSSixteen male beagle dogs were injected subcutaneously with 20 microg x kg(-1) x d(-1) recombinant human G-CSF (n = 8) or normal saline (n = 8) for 1 week. On the fifth day of treatment, the dogs underwent renal arterial angioplasty. At 8 weeks after arterial balloon injury, angiographic observations were made and injured arteries were processed for morphometric analysis of neointimal formation.

RESULTSPeripheral white blood cell counts were increased by 3.34-fold compared to baseline on the fifth day of administration of G-CSF. Angiographies revealed that one stenosis had occurred among the eight injured renal arteries from dogs treated with G-CSF, whereas all injured renal arteries from dogs treated with normal saline remained patent. The mean extent of stenosis among injured arteries was 18.3% +/- 17.9% in the G-CSF treated group compared to 12.5% +/- 7.6% in the saline treated control group (P = 0.10). G-CSF treatment slightly increased neointimal thickness (0.42 +/- 0.15 mm vs 0.25 +/- 0.06 mm, P = 0.08) with an intima to media ratio of 0.83 +/- 0.49 vs 0.54 +/- 0.18 (P = 0.11).

CONCLUSIONSG-CSF treatment does not attenuate neointimal hyperplasia and restenosis formation in a canine model of renal arterial injury, suggesting that the therapeutic strategy for preventing restenosis by stem cell mobilization should be investigated further.

Animals ; Dogs ; Granulocyte Colony-Stimulating Factor ; pharmacology ; Hematopoietic Stem Cell Mobilization ; Hyperplasia ; Male ; Recombinant Proteins ; Renal Artery ; injuries ; pathology ; Tunica Intima ; pathology

CD4+ T cells mainly interact with Sphingosine 1-phosphate (S1P) to regulate immune function through Sphingosine 1-phosphate receptor 1 (S1P1). This study was aimed to investigate the effects of recombinant human granulocyte-colony-stimulating factor (rhG-CSF) mobilization on S1P1 expression in CD4+ T cells of donor's peripheral blood. The CD4+T cells of peripheral blood were isolated by magnetic beads from 17 allo-hematopoietic stem cell transplantation (allo-HSCT) donors before and at fourth day of mobilization with rhG-CSF. The S1P1 expression was detected by real time quantitative PCR in the RNA extracted from CD4+ T cells collected before and after rhG-CSF mobilization. The results showed that the expression of S1P1 was found in CD4+ cells before and after rhG-CSF mobilization, but the expression level of SIP1 in CD4+ cells after rhG-CSF mobilization was significantly lower than that before rhG-CSF mobilization (p<0.01). It is concluded that the mobilization with rhG-CSF obviously down-regulates the expression of S1P1 in CD4+ T cells of donor's peripheral blood.
CD4-Positive T-Lymphocytes ; drug effects ; metabolism ; Granulocyte Colony-Stimulating Factor ; pharmacology ; Hematopoietic Stem Cell Mobilization ; methods ; Hematopoietic Stem Cell Transplantation ; Humans ; Receptors, Lysosphingolipid ; metabolism ; Recombinant Proteins

OBJECTIVETo investigate the effect and mechanism of high dose Vitamin B3 on granulopoiesis in normal rat.

METHODSTwenty one healthy SD rats were randomly divided into three groups: the Vitamin B3 group (Vit B3 500 mg·kg⁻¹·d⁻¹, × 7 d), the rhG-CSF group (rhG-CSF 25 μg·kg⁻¹·d⁻¹, × 7 d) and the normal saline group (2 ml/d, × 7 d). The peripheral blood cell counts were analyzed by automatic blood cell counter before (day 0) treatment, the third day (day 3) and the seventh day (day 7) after administration of drugs, respectively. The concentration of serum nicotinamide adenine dinucleotide (NAD⁺) level was measured by enzymatic cycling assay before and after drugs treatment. The expressions of G-CSF, G-CSFR, SIRT1, C/EBPα, C/EBPβ, C/EBPε and NAMPT mRNA were detected by reverse transcription real-time fluorescent quantitative PCR.

RESULTSThe neutrophil counts increased significantly after 7 days of Vitamin B3 and rhG-CSF treatment compared with that of control group [(1.64 ± 0.19) × 10⁹/L, (1.88 ± 0.37)× 10⁹/L vs (0.86 ± 0.18) × 10⁹/L, P<0.01]; the level of serum NAD⁺ increased significantly [(0.96 ± 0.08) nmol/L, (0.65 ± 0.12) nmol/L vs (0.36 ± 0.15) nmol/L, P<0.01]; the expression of G-CSF, G-CSFR, SIRT1, C/EBPα, C/EBPε and NAMPT mRNA in bone marrow mononuclear cells were increased significantly compared with that of control group (P<0.01).

CONCLUSIONHigh dose of Vitamin B3 may play an important role in increasing absolute neutrophil count in healthy rat under steady state, and the mechanism may be dependent on NAMPT-NAD⁺-SIRT1 signaling pathways.

Animals ; Bone Marrow Cells ; Granulocyte Colony-Stimulating Factor ; Leukocyte Count ; Neutrophils ; drug effects ; Niacinamide ; pharmacology ; Rats ; Rats, Sprague-Dawley ; Recombinant Proteins

To evaluate the effects of rhG-CSF on mobilization of mesenchymal stem cells (MSCs) of mouse bone marrow at different time point, thirty mice were randomly divided into rhG-CSF treatment group and control group. The mice were subcutaneously injected with rhG-CSF in a dose of 80 microg/kg or saline for 5 days. The bone marrow and peripheral blood were obtained at time points of 6, 12, 168 hours after final injection of rhG-CSF or saline. Bone marrow mononuclear cells (BMMNCs) were seeded at density of 1 x 10(6) MNCs onto 12-well plate for culture expansion in DMEM supplemented with 10% FBS, and the number of colony forming unit - fibroblast (CFU-F) was counted after 14 days. The cells were collected by trypsinization and the surface antigens CD34, CD133, CD90 and CD105 were analyzed by flow cytometry. The multi-differentiation of MSCs were done in the culture condition of induced-adipocyte and osteocyte. Peripheral blood MNCs examination was same as the bone marrow. The results indicated that the number of CFU-F of bone marrow in rhG-CSF group was more than that in control group (p < 0.01), the number of CFU-F in rhG-CSF group at 6 hours was more than that at 12 hours and 168 hours, respectively (p < 0.01). There was no obvious difference between CFU-F at 12 hours and at 168 hours (p > 0.05). MSCs were positive for CD90, CD105 and negative for CD34 and CD133. MSCs were found to differentiate into adipocyte and osteocyte in vitro. The CFU-F of PBMNCs obtained and cultured in vitro in the same culture conditions could be observed after the rhG-CSF injection at 6 hours, but cloning efficiency was (0.50 +/- 0.11) x 10(-6) MNCs and showed statistical difference as compared with control. It is concluded that rhG-CSF to mobilize hemopoietic stem cells can be used to induce mouse MSCs in vivo expansion, which showed the peak value within 6 hours after final injection of rhG-CSF. rhG-CSF have the mini-mobilization effect on murine MSCs derived from bone marrow.
Animals ; Cells, Cultured ; Female ; Granulocyte Colony-Stimulating Factor ; pharmacology ; Hematopoietic Stem Cell Mobilization ; Mesenchymal Stromal Cells ; cytology ; Mice ; Random Allocation ; Recombinant Proteins

OBJECTIVETo evaluate the time course of granulocyte-colony-stimulating-factor (G-CSF), estrogen and various doses of atorvastatin on endothelial progenitor cells (EPCs) mobilization.

METHODA total of 48 male New Zealand White rabbits were treated with placebo, estrogen (0.25 mg.k(-1).d(-1)), Atorvastatin (2.5, 5, or 10 mg) and G-CSF (50 microg/rabbit/d), respectively. Peripheral EPCs number was surveyed weekly for 4 weeks by FACS analysis (double-positive for PE-CD34/FITC-CD133) and under fluorescent microscope (double-positive for FITC-UEA-1/Dil-acLDL). Serum nitric oxide (NO) and lipids were also measured at the third week.

RESULTSPeripheral EPCs was significantly increased in G-CSF treated animals and remained constant for 4 weeks compared to placebo treated animals. Atorvastatin increased peripheral EPCs dose-dependently from 2.5 to 5 mg and peaked at the third week while peripheral EPCs number was not affected by 10 mg.k(-1).d(-1) atorvastatin during the first 3 weeks and was significantly higher only in the fourth week compared to placebo group. Estrogen also significantly increased peripheral EPCs at the third and fourth week compared to placebo group. At the third week, serum NO was similar in G-CSF group, significantly higher in atorvastatin 5 mg.k(-1).d(-1) and estrogen groups while significantly lower in atorvastatin 10 mg.k(-1).d(-1) group compared to placebo group. Serum lipids were similar among various groups.

CONCLUSIONAtorvastatin, estrogen and G-CSF could mobilize EPCs. The mobilization efficacy is as follows: G-CSF > atorvastatin 5 mg.k(-1).d(-1) > estrogen > atorvastatin 2.5 mg.k(-1).d(-1) > atorvastatin 10 mg.k(-1).d(-1). NO might partly contribute to the mobilizing effect of estrogen and atorvastatin.

Animals ; Atorvastatin Calcium ; Endothelial Cells ; cytology ; drug effects ; Estrogens ; pharmacology ; Granulocyte Colony-Stimulating Factor ; pharmacology ; Heptanoic Acids ; pharmacology ; Hypolipidemic Agents ; pharmacology ; Lipids ; blood ; Male ; Nitric Oxide ; blood ; Pyrroles ; pharmacology ; Rabbits ; Recombinant Proteins ; Stem Cells ; drug effects

To evaluate the effects of rhG-CSF and rhSCF on mobilization of the peripheral blood stem cells, 15 monkeys were divided into control, rhG-CSF 10 micro g/(kg x day) and rhG-CSF 10 micro g/(kg x day) + rhSCF 50 micro g/(kg x day) treated groups. Monkeys were administered with vehicle, rhG-CSF and rhG-CSF + rhSCF subcutaneously once daily for 14 days, respectively. The results showed that the highest counts of leukocyte of rhG-CSF treated group were 411% of baseline value on day 7 after administration, compared with that of rhG-CSF + rhSCF treated group which were 538% on day 9. The highest counts of leukocytes lasted for 3 days in combined treated group. CFU-GM from peripheral blood in the two groups were 8.37 and 11.75 times higher at 5 and 9 days respectively after the mobilization. It is concluded that rhG-CSF significantly increases the number of peripheral blood leukocytes and CFU-GM, and a better effect can be obtained by rhSCF + rhG-CSF combined administration.
Animals ; Drug Therapy, Combination ; Female ; Granulocyte Colony-Stimulating Factor ; administration & dosage ; pharmacology ; Hematopoietic Stem Cells ; drug effects ; Leukocyte Count ; Leukocytes ; drug effects ; Macaca mulatta ; Male ; Recombinant Proteins ; pharmacology ; Stem Cell Factor ; administration & dosage ; pharmacology