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Sample records for human granulocyte colony-stimulating

  1. Recombinant human granulocyte colony-stimulating factor reverts vascular dysfunction.

    PubMed

    Squadrito, F; Altavilla, D; Squadrito, G; Campo, G M; Ioculano, M; Serranò, M; Minutoli, L; Arlotta, M; Musolino, C; Saitta, A; Caputi, A P

    1997-01-01

    The aim of our study was to investigate the vascular effects of recombinant human granulocyte colony-stimulating factor (rh G-CSF) in a rat model of irreversible vascular failure. Male anesthetized rats were subjected to the clamping of the splanchnic arteries for 45 min. This surgical procedure resulted in an irreversible state of shock (splanchnic artery occlusion shock) characterized by high mortality rate (0% survival, 120 min following the release of clamps), a profound hypotension and vascular dysfunction consisting of a marked hyporeactivity to phenylephrine (PE 1 nM-10 microM) of aortic rings. Administration of recombinant human granulocyte colony-stimulating factor (20 micrograms/kg i.v. 5 min after the release of occlusion) increased survival rate (90% 4 h after the release of occlusion), blunted the profound hypotension and reverted the marked vascular dysfunction. Finally, rh G-CSF inhibited the activity of inducible nitric oxide synthase in peritoneal macrophages activated with endotoxin. Our data suggest that rh G-CSF may influence vascular function when low-flow states occur.

  2. Recovery from severe hematopoietic suppression using recombinant human granulocyte-macrophage colony-stimulating factor

    SciTech Connect

    Monroy, R.L.; Skelly, R.R.; Taylor, P.; Dubois, A.; Donahue, R.E.; MacVittie, T.J.

    1988-06-01

    The ability of recombinant human granulocyte-macrophage colony-stimulating factor (rhGM-CSF) to enhance recovery of a radiation-suppressed hematopoietic system was evaluated in a nonuniform radiation exposure model using the rhesus monkey. Recombinant human GM-CSF treatment for 7 days after a lethal, nonuniform radiation exposure of 800 cGy was sufficient to enhance hematopoietic reconstitution, leading to an earlier recovery. Monkeys were treated with 72,000 U/kg/day of rhGM-CSF delivered continuously through an Alzet miniosmotic pump implanted subcutaneously on day 3. Treated monkeys demonstrated effective granulocyte and platelet levels in the peripheral blood, 4 and 7 days earlier, respectively, than control monkeys. Granulocyte-macrophage colony-forming unit (CFU-GM) activity in the bone marrow was monitored to evaluate the effect of rhGM-CSF on marrow recovery. Treatment with rhGM-CSF led to an early recovery of CFU-GM activity suggesting that rhGM-CSF acted on an earlier stem cell population to generate CFU-GM. Thus, the effect of rhGM-CSF on hematopoietic regeneration, granulocyte recovery, and platelet recovery are evaluated in this paper.

  3. Recovery from severe hematopoietic suppression using recombinant human granulocyte-macrophage colony-stimulating factor

    SciTech Connect

    Monroy, R.L.; Skelly, R.R.; Taylor, P.; Dubois, A.; Donahue, R.E.

    1988-01-01

    The ability of recombinant human granulocytemacrophage colony-stimulating factor (rhGM-CSF) to enhance recovery of a radiation-suppressed hematopoietic system was evaluated in a nonuniform radiation-exposure model using the rhesus monkey. Recombinant human GM-CSF treatment for 7 days after a lethal, nonuniform radiation exposure of 800 cGy was sufficient to enhance hematopoietic reconstitution, leading to an earlier recovery. Monkeys were treated with 72,000 U/kg/day of rhGm-CSF delivered continuously through an Alzet mini-osmotic pump implanted subcutaneously on day 3. Treated monkeys demonstrated effective granulocyte and platelet levels in the peripheral blood, 4 and 7 days earlier, respectively, than control monkeys. Granulocyte-macrophage colony-forming unit (CFU-GM) activity in the bone marrow was monitored to evaluate the effect of rhGM-CSF on marrow recovery. Treatment with rhGM-CSF led to an early recovery of CFU-GM activity suggesting that rhGM-CSF acted on an earlier stem cell population to generate CFU-GM. Thus, the effect of rhGM-CSF on hematopoietic regeneration, granulocyte recovery, and platelet recovery are evaluated.

  4. Neutrophil kinetics of recombinant human granulocyte colony-stimulating factor-induced neutropenia in rats

    SciTech Connect

    Okada, Yuji; Kawagishi, Mayumi; Kusaka, Masaru )

    1990-01-01

    Single injection of recombinant human granulocyte colony-stimulating factor (rhG-CSF) immediately induced a decrease in the number of circulating neutrophils in rats. This neutropenia occurred 10 minutes after the injection but disappeared 40 minutes after injection. This transient neutropenia was dose-dependently induced by rhG-CSF and also induced by repeated injections. We studied the kinetics of circulating neutrophils in transient neutropenia. rhG-CSF markedly decreased the number of {sup 3}H-diisopropylfluorophosphate ({sup 3}H-DFP) labeled neutrophils in the circulation 10 minutes after injection but the labeled neutrophils recovered to near the control level 40 minutes after the injection. These results indicate that the neutrophil margination accounts for the neutrophenia and the marginated neutrophils return to the circulation.

  5. Action of granulocyte-macrophage colony-stimulating factors: studies using a human leukemia cell line.

    PubMed Central

    Lusis, A J; Koeffler, H P

    1980-01-01

    Granulocyte-macrophage colony-stimulating factors (CSFs) have previously been shown to stimulate colony formation in soft agar culture by a human myelogenous leukemia cell line known as KG-1. We have used KG-1 cells as a model system to investigate the interaction of CSF with myeloid cells. We now report that exposure of KG-1 cells to human CSFs in liquid culture results in a rapid (within 3 hr) burst of RNA synthesis and, after a lag of about 10 hr, a stimulation of DNA and protein synthesis. RNA and protein synthesis were maximally stimulated about 2-fold and DNA synthesis was stimulated about 2.5-fold. The stimulation was specific; various growth factors, hormones, and mouse CSFs had no effect on KG-1 macromolecular synthesis. Treatment with CSF did not discernibly alter the morphological appearance of the KG-1 cells (primarily myeloblasts) nor did it qualitatively affect the pattern of newly synthesized proteins separable by one- and two-dimensional electrophoresis. Several myeloid leukemia cell lines that were not responsive to CSF in agar culture, including a dedifferentiated variant of KG-1, showed little or no stimulation of macromolecular synthesis upon exposure to CSF. We have used the CSF-dependent stimulation of macromolecular synthesis of KG-1 to develop a rapid, sensitive microassay for human CSFs. The assay, involving thymidine incorporation by the cells, should be useful for characterization and purification of human CSFs. Images PMID:6159645

  6. Endotoxin down-modulates granulocyte colony-stimulating factor receptor (CD114) on human neutrophils.

    PubMed

    Hollenstein, U; Homoncik, M; Stohlawetz, P J; Marsik, C; Sieder, A; Eichler, H G; Jilma, B

    2000-07-01

    During infection, the development of nonresponsiveness to granulocyte colony-stimulating factor (G-CSF) may be influenced by the down-modulation of G-CSF receptor (G-CSFR) by cytokines. This down-modulation was studied during experimental human endotoxemia. Healthy volunteers received either 2 ng/kg endotoxin (lipopolysaccharide [LPS], n=20) or placebo (n=10) in a randomized, controlled trial. Endotoxin infusion increased the mean fluorescence intensity of the neutrophil activation marker CD11b >300% after 1 h (P<.001 vs. placebo). LPS infusion down-modulated G-CSFR expression in as early as 60 min (-17%; P=.001 vs. placebo). Down-modulation was almost maximal at 90 min and persisted for 6 h (-50% from baseline; P<.0001 vs. placebo). Plasma levels of G-CSF started to increase only after G-CSFR down-modulation had occurred and peaked 37-fold above baseline at 4 h (P<.0001 vs. placebo). In conclusion, LPS down-modulates G-CSFR expression in humans, which may render neutrophils less responsive to the effects of G-CSF and, thereby, compromise host defense mechanisms.

  7. Effect of recombinant human granulocyte colony-stimulating factor on granulocytopenia in mice induced by irradiation

    SciTech Connect

    Fushiki, M.; Ono, K.; Sasai, K.; Shibamoto, Y.; Tsutsui, K.; Nishidai, T.; Takahashi, M.; Abe, M. )

    1990-02-01

    We report the effect of human granulocyte colony-stimulating factor (hG-CSF) on the recovery from granulocytopenia induced by irradiation. Female 9-week old C3H/He mice were used. The irradiation schedule was as follows: Group 1 and 2 received whole-body irradiation of 1 Gy and 5 Gy, respectively, on day 0; Group 3 and 4 received whole-body irradiation of 0.5 and 1.0 Gy, respectively, for 5 consecutive days; Group 5 received upper hemibody irradiation of 3 Gy for 5 consecutive days. Daily subcutaneous injections of G-CSF (3 x 10(5) Unit/mouse) or 0.3 ml of saline to each group were started from the day after the first irradiation and continued for 18 days. Mice were sampled randomly from each group, and the total number of leukocytes, erythrocytes of peripheral blood, nucleated cells in femur, and spleen weight were counted and measured, respectively, on day 0, 3, 5, 7, 9, 12, and 18. The leukocyte counts decreased with an increase in radiation doses. In Group 1 and 2 mice, G-CSF enhanced the leukocyte count more than saline. In Group 3 mice, the recovery of leukocytopenia was facilitated by G-CSF, but in Group 4 mice, G-CSF had no effect on the leukocyte count decrease or on leukocytopenia recovery. In Group 5 mice, G-CSF greatly affected leukocytopenia recovery. Increase in spleen weight paralleled the peripheral leukocyte count. Daily administration of recombinant hG-CSF accelerated the granulocytopenia recovery which was induced by irradiation, and it may be a useful therapeutic agent for treating myelosuppressive cases.

  8. Clinical safety of tbo-filgrastim, a short-acting human granulocyte colony-stimulating factor.

    PubMed

    Pettengell, Ruth; Bias, Peter; Mueller, Udo; Lang, Nicole

    2016-06-01

    The recombinant human granulocyte colony-stimulating factor (G-CSF) known as filgrastim (Tevagrastim(®), Ratiograstim(®), Biograstim(®)) in Europe (approved in 2008) and tbo-filgrastim (Granix(®)) in the USA (approved in 2012; Teva Pharmaceutical Industries Ltd., Petach Tikva, Israel) is indicated to reduce the duration of severe neutropenia in patients with non-myeloid malignancies receiving myelosuppressive anti-cancer drugs associated with a clinically significant incidence of febrile neutropenia. This article presents pooled clinical data for tbo-filgrastim compared with Neupogen(®) (Amgen, Thousand Oaks, CA, USA) as well as tbo-filgrastim post-marketing safety data. The safety and efficacy of tbo-filgrastim were evaluated in three phase III studies in 677 patients receiving myelosuppressive chemotherapy and study drug (348 patients with breast cancer, 237 with lung cancer, 92 with non-Hodgkin lymphoma). In each study, the efficacy of tbo-filgrastim was similar to that of Neupogen. Overall, 633 (93.5 %) patients receiving the study drug experienced 6093 treatment-emergent adverse events (AEs), most of which were related to chemotherapy. Adverse events related to the study drug (tbo-filgrastim or Neupogen) were experienced by 185 (27.3 %) patients; 19 (2.8 %) had severe drug-related AEs, 5 (0.7 %) had drug-related serious AEs, and 6 (0.9 %) discontinued the study due to drug-related AEs. Overall, the most common drug-related AEs were bone pain (7.1 %), myalgia (4.0 %), and asthenia (4.4 %). The post-marketing safety profile of tbo-filgrastim was consistent with that observed during the clinical studies. The availability of tbo-filgrastim, a G-CSF with safety and efficacy comparable to those of Neupogen, provides physicians with an alternative treatment option for supportive care of patients with non-myeloid malignancies receiving myelosuppressive chemotherapy. PMID:26780505

  9. Enhancement of in vitro and in vivo anti-tumor activity of anti-GD2 monoclonal antibody 220-51 against human neuroblastoma by granulocyte-macrophage colony-stimulating factor and granulocyte colony-stimulating factor.

    PubMed

    Fukuda, M; Horibe, K; Furukawa, K

    1998-10-01

    We have evaluated the anti-tumor effect of anti-GD2 mouse monoclonal antibody (mAb) 220-51 against human neuroblastoma cell line TGW in vitro and in vivo. The mAb 220-51 was able to mediate complement-dependent cytotoxicity (CDC) and antibody-dependent cellular cytotoxicity (ADCC) using human effector cells. In the presence of recombinant human granulocyte-macrophage colony-stimulating factor (GM-CSF), granulocyte ADCC was significantly augmented in vitro. When mAb 220-51 was administered to tumor-bearing nude mice, tumor growth was significantly inhibited as compared with untreated controls. Administration of recombinant murine GM-CSF in combination with mAb 220-51 significantly enhanced the anti-tumor effect of mAb in vivo. Recombinant human granulocyte colony-stimulating factor (G-CSF) combined with mAb 220-51 was also able to enhance it, although granulocyte ADCC was not affected by the presence of recombinant human G-CSF in vitro. Moreover, GM-CSF and G-CSF work additively to enhance the anti-tumor effect of mAb 220-51 in vivo. The GM-CSF and G-CSF may have a clinical potency in immunotherapy with anti-GD2 mAb for the treatment of neuroblastoma.

  10. Growth of human hemopoietic colonies in response to recombinant gibbon interleukin 3: comparison with human recombinant granulocyte and granulocyte-macrophage colony-stimulating factor

    SciTech Connect

    Messner, H.A.; Yamasaki, K.; Jamal, N.; Minden, M.M.; Yang, Y.C.; Wong, G.G.; Clark, S.C.

    1987-10-01

    Supernatants of COS-1 cells transfected with gibbon cDNA encoding interleukin 3 (IL-3) with homology to sequences for human IL-3 were tested for ability to promote growth of various human hemopoietic progenitors. The effect of these supernatants as a source of recombinant IL-3 was compared to that of recombinant human granulocyte-macrophage colony-stimulating factor (GM-CSF) and granulocyte colony-stimulating factor (G-CSF) as well as to that of medium conditioned by phytohemagglutinin-stimulated leukocytes. The frequency of multilineage colonies, erythroid bursts, and megakaryocyte colonies in cultures containing the COS-1 cell supernatant was equivalent to the frequency observed in the controls and significantly higher than found in cultures plated with recombinant GM-CSF. G-CSF did not support the formation of multilineage colonies, erythroid bursts, and megakaryocyte colonies. In contrast, growth of granulocyte-macrophage colonies was best supported with GM-CSF, while recombinant IL-3 yielded colonies at lower or at best equivalent frequency. The simultaneous addition of higher concentrations of GM-CSF to cultures containing IL-3 in optimal amounts did not enhance the formation of multilineage colonies, erythroid bursts, and megakaryocyte colonies. However, the frequency of such colonies and bursts increased with GM-CSF when cultures were plated with suboptimal concentrations of IL-3. Growth of colonies within the granulocyte-macrophage lineage is optimally supported by GM-CSF and does not increase with further addition of IL-3.

  11. In vivo effect of human granulocyte-macrophage colony-stimulating factor on megakaryocytopoiesis

    SciTech Connect

    Aglietta, M.; Monzeglio, C.; Sanavio, F.; Apra, F.; Morelli, S.; Stacchini, A.; Piacibello, W.; Bussolino, F.; Bagnara, G.; Zauli, G. )

    1991-03-15

    The effect of granulocyte-macrophage colony-stimulating factor (GM-CSF) on megakaryocytopoiesis and platelet production was investigated in patients with normal hematopoiesis. Three findings indicated that GM-CSF plays a role in megakaryocytopoiesis. During treatment with GM-CSF (recombinant mammalian, glycosylated; Sandoz/Schering-Plough, 5.5 micrograms protein/kg/d, subcutaneously for 3 days) the percentage of megakaryocyte progenitors (megakaryocyte colony forming unit (CFU-Mk)) in S phase (evaluated by the suicide technique with high 3H-Tdr doses) increased from 31% +/- 16% to 88% +/- 11%; and the maturation profile of megakaryocytes was modified, with a relative increase in more immature stage I-III forms. Moreover, by autoradiography (after incubation of marrow cells with 125I-labeled GM-CSF) specific GM-CSF receptors were detectable on megakaryocytes. Nevertheless, the proliferative stimulus induced on the progenitors was not accompanied by enhanced platelet production (by contrast with the marked granulomonocytosis). It may be suggested that other cytokines are involved in the regulation of the intermediate and terminal stages of megakaryocytopoiesis in vivo and that their intervention is an essential prerequisite to turn the GM-CSF-induced proliferative stimulus into enhanced platelet production.

  12. Construction of recombinant Escherichia coli strains for secretory expression of artificial genes for human granulocyte-macrophage colony stimulating factor

    SciTech Connect

    Petrovskaya, L.E.; Ruzin, A.V.; Shingarova, L.N.; Korobko, V.G.

    1995-11-01

    A number of recombinant plasmids for expression of artificial genes encoding human granulocyte-macrophage colony stimulating factor (GM-CSF) were constructed. A hybrid gene was obtained that contains a sequence encoding the leader peptide and a tandem of two IgG-binding domains of protein A from Staphylococcus aureus coupled, through an enteropepdidase linker, to a synthetic gmcsf gene. The construction enables Escherichia coli to carry out biosynthesis of the hybrid protein and its subsequent transport into the periplasmic space of bacteria. Another hybrid gene, combining sequences for the signal peptide of the E. coli outer membrane protein OmpA and GM-CSF, was obtained using polymerase chain reaction. The localization of the mature protein produced by the hybrid gene was found to depend on the strength of the promoter used. 39 refs., 6 figs.

  13. Weekly CODE chemotherapy with recombinant human granulocyte colony-stimulating factor for relapsed or refractory small cell lung cancer.

    PubMed

    Sato, K; Tsuchiya, S; Minato, K; Sunaga, N; Ishihara, S I; Makimoto, T; Naruse, I; Hoshino, H; Watanabe, S; Saitoh, R; Mori, M

    2000-01-01

    We used cisplatin, vincristine, doxorubicin, and etoposide (CODE) plus recombinant human granulocyte colony-stimulating factor (rhG-CSF) weekly for salvage chemotherapy in relapsed or refractory small cell lung cancer (SCLC). We reviewed the medical charts of patients between January 1993 and December 1996 at the National Nishi-Gunma Hospital. Twenty patients were treated with salvage chemotherapy. The overall response rate was 55.0%. The median survival time of extensive disease patients from the start of CODE therapy was 23 weeks and the 1-year survival rate was 21.0%. Toxicities were severe, especially in myelosuppression. CODE could be selected as a salvage therapy for chemotherapy- relapsed SCLC cases.

  14. Augmentation of Human Macrophage Candidacidal Capacity by Recombinant Human Myeloperoxidase and GranulocyteMacrophage Colony-Stimulating Factor

    PubMed Central

    Maródi, László; Tournay, Christophe; Káposzta, Rita; Johnston, Richard B.; Moguilevsky, Nicole

    1998-01-01

    Phagocyte myeloperoxidase (MPO) is believed to be particularly important in defense against candida infection. We reported earlier that monocytes, rich in MPO, killed Candida albicans at a significantly higher rate and extent than did monocyte-derived macrophages, known to lack MPO, and that C. albicans is less resistant to MPO-dependent oxidants than less pathogenic Candida species. We hypothesized, therefore, that the capacity of macrophages to kill C. albicans might be improved in the presence of MPO. In this study, we evaluated the ability of recombinant human MPO (rhMPO) to augment the killing of C. albicans by resident macrophages and macrophages activated by recombinant human granulocyte-macrophage colony-stimulating factor. Addition of rhMPO (concentration range, 0.8 to 6.4 U/ml) to suspensions of resident and activated macrophages and opsonized C. albicans resulted in concentration-dependent and significant increases in candida killing. This enhancement was particularly pronounced with activated macrophages, whether C. albicans was opsonized or unopsonized and ingested through the macrophage mannose receptor. rhMPO did not affect the killing of C. albicans by monocytes, nor did it affect phagocytosis of opsonized or unopsonized C. albicans. These results indicate that exogenous rhMPO can augment the candidacidal capacity of both resident and activated macrophages, with a more profound effect on activated cells. We suggest that rhMPO may be effective in the treatment of invasive candidiasis. PMID:9596743

  15. Enhanced expression of DNA topoisomerase II by recombinant human granulocyte colony-stimulating factor in human leukemia cells.

    PubMed

    Towatari, M; Ito, Y; Morishita, Y; Tanimoto, M; Kawashima, K; Morishima, Y; Andoh, T; Saito, H

    1990-11-15

    The effect of recombinant human granulocyte colony-stimulating factor (G-CSF) on DNA topoisomerase II (topo II) expression was studied in two human acute myelogenous leukemia cell lines, NKM-1 and NOMO-1, which express G-CSF receptor and proliferate in response to exogenous G-CSF. Northern blot analysis revealed that the level of topo II mRNA in 16-h stimulated cells in serum-free medium with G-CSF (10 ng/ml) was approximately 2-fold higher than that in cells without G-CSF. Enhanced topo II mRNA expression was detectable within 3 h after the addition of G-CSF. Topo II activity in crude nuclear extracts from 16-h G-CSF-stimulated cells was also found to be approximately 2-fold greater than that from unstimulated cells. According to in vitro cytotoxic assay, the sensitivity of G-CSF-stimulated cells to intercalating (daunorubicin) and nonintercalating (etoposide) topo II-targeting drugs increased significantly, whereas no enhancement of sensitivity was observed with an alkylating agent (4-hydroperoxycyclophosphamide). The augmented drug sensitivity observed was not due to the increased level of drug transport, as suggested by the similar extent of [3H]etoposide uptake between G-CSF-stimulated and unstimulated cells. By measuring the topo II mRNA and the cytotoxicity of the above mentioned drugs, we obtained essentially the same results in G-CSF-responsive leukemia cells isolated from three acute myeloblastic leukemia patients, as observed in the cultured cell lines. These findings strongly suggest that the sensitivity to "topo II-targeting drugs" could be augmented by exogenous G-CSF through elevated topo II activity in G-CSF-responsive leukemia cells. PMID:1699657

  16. Structure of the chromosomal gene for granulocyte-macrophage colony stimulating factor: comparison of the mouse and human genes.

    PubMed Central

    Miyatake, S; Otsuka, T; Yokota, T; Lee, F; Arai, K

    1985-01-01

    A cDNA clone that expresses granulocyte-macrophage colony stimulating factor (GM-CSF) activity in COS-7 cells has been isolated from a pcD library prepared from mRNA derived from concanavalin A-activated mouse helper T cell clones. Based on homology with the mouse GM-CSF cDNA sequence, the mouse GM-CSF gene was isolated. The human GM-CSF gene was also isolated based on homology with the human GM-CSF cDNA sequence. The nucleotide sequences determined for the genes and their flanking regions revealed that both the mouse and human GM-CSF genes are composed of three introns and four exons. The organization of the mouse and human GM-CSF genes are highly homologous and strong sequence homology between the two genes is found both in the coding and non-coding regions. A 'TATA'-like sequence was found 20-25 bp upstream from the transcription initiation site. In the 5'-flanking region, there is a highly homologous region extending 330 bp upstream of the putative TATA box. This sequence may play a role in regulation of expression of the GM-CSF gene. These structures are compared with those of different lymphokine genes and their regulatory regions. Images Fig. 2. Fig. 6. PMID:3876930

  17. Affinity purification of human granulocyte macrophage colony-stimulating factor receptor alpha-chain. Demonstration of binding by photoaffinity labeling

    SciTech Connect

    Chiba, S.; Shibuya, K.; Miyazono, K.; Tojo, A.; Oka, Y.; Miyagawa, K.; Takaku, F. )

    1990-11-15

    The human granulocyte macrophage colony-stimulating factor (GM-CSF) receptor alpha-chain, a low affinity component of the receptor, was solubilized and affinity-purified from human placenta using biotinylated GM-CSF. Scatchard analysis of {sup 125}I-GM-CSF binding to the placental membrane extract disclosed that the GM-CSF receptor had a dissociation constant (Kd) of 0.5-0.8 nM, corresponding to the Kd value of the GM-CSF receptor alpha-chain on the intact placental membrane. Affinity labeling of the solubilized protein using a photoreactive cross-linking agent, N-hydroxysuccinimidyl-4-azidobenzoate (HSAB), demonstrated a single specific band of 70-95 kDa representing a ligand-receptor complex. Approximately 2 g of the placental membrane extract was subjected to a biotinylated GM-CSF-fixed streptavidin-agarose column, resulting in a single major band at 70 kDa on a silver-stained sodium dodecyl sulfate gel. The radioiodination for the purified material disclosed that the purified protein had an approximate molecular mass of 70 kDa and a pI of 6.6. Binding activity of the purified material was demonstrated by photoaffinity labeling using HSAB-{sup 125}I-GM-CSF, producing a similar specific band at 70-95 kDa as was demonstrated for the crude protein.

  18. Phenotypic features of first-generation transgenic goats for human granulocyte-colony stimulation factor production in milk.

    PubMed

    Batista, Ribrio I T P; Melo, Carlos H S; Souza-Fabjan, Joanna M G; Teixeira, Dárcio I A; Melo, Luciana M; Freitas, Vicente J F

    2014-11-01

    Human granulocyte-colony stimulating factor (hG-CSF) is a hematopoietic growth factor used in neutropenic patients. It is produced in transgenic bacteria or cultured mammalian cells. As an alternative, we now show that hG-CSF can be expressed in the mammary gland of first-generation (F1) transgenic goats during induced lactation. Despite lower milk production, transgenic females presented a similar milk composition (fat, protein and lactose) when compared to non-transgenic (p > 0.05) ones. The mean concentration (±SD) of recombinant hG-CSF in milk during lactation was 360 ± 178 µg ml(-1). All clinical parameters, as well as kidney and liver function, indicated that F1 transgenic goats were healthy. Additionally, no ectopic hG-CSF expression was detected in studied tissues of F1 transgenic males. Thus, F1 hG-CSF-transgenic goats can express the recombinant protein in milk at quantities compatible with their use as bioreactors in a commercial-scale protein-production program.

  19. Treatment of methimazole-induced severe aplastic anemia with recombinant human granulocyte-monocyte colony-stimulating factor and glucocorticosteroids.

    PubMed

    López-Karpovitch, X; Ulloa-Aguirre, A; von Eiff, C; Hurtado-Monroy, R; Alanis, A

    1992-01-01

    The in vivo response to recombinant human granulocyte-monocyte colony-stimulating factor (rHu GM-CSF) in facilitating the reconstitution of granulomonopoiesis was evaluated in a patient with Graves' disease who developed severe aplastic anemia during methimazole therapy. After 10 days of treatment with rHu GM-CSF, the neutrophil and monocyte counts rose to 1.65 x 10(9)/l and 0.41 x 10(9)/l, respectively. However, the patient was still dependent on erythrocyte and platelet transfusions. Two days after rHu GM-CSF withdrawal, the neutrophil count dropped off to 0.41 x 10(9)/l.rHu GM-CSF was reinitiated for 2 days along with glucocorticosteroids. With this combined therapeutic approach, the neutrophil count returned to normal and remained stable, and both Hb and platelet values began to improve. It is concluded that the combination of rHu GM-CSF and glucocorticosteroids can be used as a therapeutic option that may lead to beneficial results in drug-induced aplastic anemia.

  20. Modulation of neutrophil and monocyte function by recombinant human granulocyte macrophage colony-stimulating factor in patients with lymphoma.

    PubMed

    Kharazmi, A; Nielsen, H; Hovgaard, D; Borregaard, N; Nissen, N I

    1991-04-01

    Granulocyte macrophage colony-stimulating factor (GM-CSF) has been shown to inhibit the chemotaxis and enhance the oxidative burst response of human neutrophils in vitro. The present study describes the effect of recombinant GM-CSF on the neutrophil and monocyte function in patients with lymphoma undergoing GM-CSF treatment. Patients with either Hodgkin's or non-Hodgkin's lymphoma were treated with various dosages (2-16 micrograms kg-1 body weight per day for 5 days) of rhGM-CSF by intravenous or subcutaneous route. Prior to and on day 5 of rhGM-CSF treatment, neutrophil and monocyte chemotaxis and chemiluminescence responses to f-Met-Leu-Phe, zymosan activated serum (ZAS) and opsonized zymosan (OZ) were determined. It was observed that chemotactic response of neutrophils to f-Met-Leu-Phe and ZAS was reduced, whereas the chemiluminescence response of both cell types to f-Met-Leu-Phe and zymosan was enhanced by up to 43-fold. rhGM-CSF treatment did not affect degranulation of the neutrophils as measured by release of vitamin B12 binding protein. Degree of modulation of neutrophil and monocyte function by rhGM-CSF was independent of rhGM-CSF dosages administered. These data suggest that phagocytic defence system may be enhanced by GM-CSF treatment and that this cytokine may be a useful therapeutic adjunct in compromised patients.

  1. Vaccination with Irradiated Autologous Melanoma Cells Engineered to Secrete Human Granulocyte--Macrophage Colony-Stimulating Factor Generates Potent Antitumor Immunity in Patients with Metastatic Melanoma

    NASA Astrophysics Data System (ADS)

    Soiffer, Robert; Lynch, Thomas; Mihm, Martin; Jung, Ken; Rhuda, Catherine; Schmollinger, Jan C.; Hodi, F. Stephen; Liebster, Laura; Lam, Prudence; Mentzer, Steven; Singer, Samuel; Tanabe, Kenneth K.; Benedict Cosimi, A.; Duda, Rosemary; Sober, Arthur; Bhan, Atul; Daley, John; Neuberg, Donna; Parry, Gordon; Rokovich, Joseph; Richards, Laurie; Drayer, Jan; Berns, Anton; Clift, Shirley; Cohen, Lawrence K.; Mulligan, Richard C.; Dranoff, Glenn

    1998-10-01

    We conducted a Phase I clinical trial investigating the biologic activity of vaccination with irradiated autologous melanoma cells engineered to secrete human granulocyte--macrophage colony-stimulating factor in patients with metastatic melanoma. Immunization sites were intensely infiltrated with T lymphocytes, dendritic cells, macrophages, and eosinophils in all 21 evaluable patients. Although metastatic lesions resected before vaccination were minimally infiltrated with cells of the immune system in all patients, metastatic lesions resected after vaccination were densely infiltrated with T lymphocytes and plasma cells and showed extensive tumor destruction (at least 80%), fibrosis, and edema in 11 of 16 patients examined. Antimelanoma cytotoxic T cell and antibody responses were associated with tumor destruction. These results demonstrate that vaccination with irradiated autologous melanoma cells engineered to secrete granulocyte--macrophage colony-stimulating factor stimulates potent antitumor immunity in humans with metastatic melanoma.

  2. Efficient Process Development of Recombinant Human Granulocyte Colony-Stimulating Factor (rh-GCSF) Production in Escherichia coli

    PubMed Central

    Babaeipour, Valiollah; Khanchezar, Sirwan; Mofid, Mohammad Reza; Pesaran Hagi Abbas, Mahdi

    2015-01-01

    Background: The protein hormone granulocyte colony-stimulating factor (GCSF) stimulates the production of white blood cells and plays an important role in medical treatment of cancer patients. Methods: An efficient process was developed for heterologous expression of the human GCSF in E. coli BL21 (DE3). The feeding rate was adjusted to achieve the maximum attainable specific growth rate under critical value. In this method, specific growth rate was maintained at the maximum value of 0.55 h-1 at the beginning of feeding to 0.4 h-1 at the induction time. Recombinant human GCSF (rh-GCSF) was produced as inclusion body. At first, inclusion bodies were released by cell disruption and then washed, solubilized and refolded. Finally, the rh-GCSF was purified by cation exchange chromatography. Results: Obviouly, higher specific growth rate decreases process time and consequently increases productivity. The final concentration of biomass and GCSF was achieved 126 g DCW.l-1 and 32.1 g.l-1. Also, the final specific yield (YP/X) and total productivity of rh-GCSF were obtained 254 mg.g-1 DCW and 1.83 g.l-1.h-1, respectively. According to the available data, this is one of the highest YP/X and productivity that has been reported for any human protein which is expressed in E. coli. Recovery yield of purification process was %40 and purity of recombinant protein was over than 99%. The circular dichroism spectra of purified rh-GCSF, Neupogen® and PD-Grastim showed that all proteins have a similar secondary structure. Conclusion: Modified exponential feeding strategy for fed-batch cultivation of recombinant E. coli, results in minimum fed-batch duration and maximum productivity. PMID:25864815

  3. Improved Production and Characterization of Recombinant Human Granulocyte Colony Stimulating Factor from E. coli under Optimized Downstream Processes.

    PubMed

    Vemula, Sandeep; Thunuguntla, Rahul; Dedaniya, Akshay; Kokkiligadda, Sujana; Palle, Chaitanya; Ronda, Srinivasa Reddy

    2015-04-01

    This work reports the upstream and downstream process of recombinant human granulocyte colony stimulating factor (rhG-CSF) expressed in Escherichia coli BL21 (DE3)pLysS. The fed batch mode was selected for the maximum output of biomass (6.4g/L) and purified rhG-CSF (136mg/L) under suitable physicochemical environment. The downstream processing steps viz., recovery, solubilization, refolding and concentration were optimized in this study. The maximum rhG-CSF inclusion bodies recovery yield (97%) was accomplished with frequent homogenization and sonication procedure. An efficient solubilization (96%) of rhG-CSF inclusion bodies were observed with 8M urea at pH 9.5. Refolding efficiency studies showed maximum refolding ⩾86% and ⩾84% at 20°C and pH 9 respectively. The renatured protein solution was concentrated, clarified and partially purified (⩾95%) by the cross flow filtration technique. The concentrated protein was further purified by a single step size exclusion chromatography with ⩾98% purity. The characterization of purified rhG-CSF molecular mass as evidenced by SDS-PAGE, western blot and LC/MS analysis was shown to be 18.8kDa. The secondary structure of rhG-CSF was evaluated by the CD spectroscopic technique based on the helical structural components. The biological activity of the purified rhG-CSF showed a similar activity of cell proliferation with the standard rhG-CSF. Overall, the results demonstrate an optimized downstream process for obtaining high yields of biologically active rhG-CSF.

  4. Effect of recombinant human granulocyte colony-stimulating factor on efficacy of radiation therapy in tumor-bearing rats

    SciTech Connect

    Koji Kabaya; Masahiko Watanabe; Masaru Kusaka; Hiromichi Akahori; Masatoshi Seki; Masato Fushiki

    1994-07-01

    The effect of recombinant human granulocyte colony-stimulating factor on radiation-induced neutropenia and on growth of transplanted tumors treated by irradiation was investigated using tumor-bearing rats as a model for radiation therapy. In a preliminary study using normal rats, neutropenia induced by upper hemi-body irradiation at 3 Gy/day 5 times a week for 3 weeks was prevented by consecutive subcutaneous injections of rhG-CSF at 100 {mu}g/kg/day. Rats bearing Walker-256, a mammary tumor, were scheduled to receive upper hemibody irradiation at 3 Gy/day for 15 times in 3 weeks if white blood cell (WBC) counts were maintained above 3,000/{mu}l. In control tumor-bearing rats not receiving rhG-CSF, irradiation was often withheld because of the decrease in WBC counts below 3,000/{mu}l. In contrast, a decrease in WBC counts below 3,000/{mu}l was rarely found in tumor-bearing rats injected daily with rhG-CSF. The average number of radiation treatments in control rats and rats treated with rhG-CSF was about 8 and 14, respectively, out of the scheduled 15 treatments in 3 weeks. Treatment with rgG-CSF made it possible to complete the radiation therapy regimen and thus inhibit the growth of the transplanted tumor more effectively. These results suggest that rgG-CSF may be useful to ensure radiation therapy on schedule in cancer patients. 20 refs., 4 figs., 1 tab.

  5. Up-regulation of transferrin receptor gene expression by granulocyte colony-stimulating factor in human myeloid leukemia cells.

    PubMed

    Morishita, Y; Kataoka, T; Towatari, M; Ito, T; Inoue, H; Ogura, M; Morishima, Y; Saito, H

    1990-12-15

    Granulocyte colony-stimulating factor (G-CSF) enhanced surface transferrin receptor (TfR) expression in two human myeloid leukemia cell lines, NKM-1 and NOMO-1, which possess G-CSF receptors. Radioligand-binding assay revealed that 10 ng/ml G-CSF significantly increased TfR to 186 +/- 20 and 276 +/- 38% of control for NKM-1 cells and NOMO-1 cells, respectively, in a 24-h culture. Scatchard analysis showed the increase of transferrin (Tf)-binding sites but no change in the receptor affinity. The enhanced TfR expression was not mediated either by the kinetic change of receptor cycling or by cellular iron content. Immunoprecipitation with anti-TfR antibody was used, and the increased biosynthesis of the receptor was demonstrated in G-CSF-stimulated cells. Northern blot analysis showed a 2- to 3-fold increase of TfR mRNA of NKM-1 cells cultured in medium containing Tf and G-CSF, whereas the mRNA declined without G-CSF. The effect of G-CSF on the TfR mRNA was observed within 2 h, which preceded the increase of surface TfR and the transition to the S phase of the cell cycle. G-CSF also potentiated TfR expression in freshly obtained myeloid leukemia cells. The present study shows up-regulation of TfR expression by G-CSF in myeloid leukemia cells and provides evidence that the regulation is mediated by controlling the steady-state level of the mRNA. PMID:1701357

  6. Human granulocyte colony stimulating factor (G-CSF) produced in the filamentous fungus Aspergillus niger.

    PubMed

    Kraševec, Nada; Milunović, Tatjana; Lasnik, Marija Anžur; Lukančič, Irena; Komel, Radovan; Porekar, Vladka Gaberc

    2014-01-01

    For the first time, a fungal production system is described for expression and secretion of the medically important human protein G-CSF, in Aspergillus niger. A reliable strategy was chosen with in-frame fusion of G-CSF behind a KEX2 cleavage site downstream of the coding region of the highly secreted homologous glucoamylase. This provided secretion levels of 5-10 mg/l culture medium of correctly processed G-CSF, although the majority of the protein (>90%) was biologically inactive. Following denaturation/ concentration and chromatographic separation/ renaturation, the G-CSF proliferation activity increased considerably, and analytical immobilised metal affinity chromatography confirmed the monomeric and correctly folded protein. These data suggest that this human secretory protein secreted into the medium of A. niger was not correctly folded, and that it escaped the endoplasmic reticulum folding control systems. This is compared to the folding of G-CSF produced in bacteria and yeast. PMID:25551710

  7. Kinetics of human hemopoietic cells after in vivo administration of granulocyte-macrophage colony-stimulating factor.

    PubMed Central

    Aglietta, M; Piacibello, W; Sanavio, F; Stacchini, A; Aprá, F; Schena, M; Mossetti, C; Carnino, F; Caligaris-Cappio, F; Gavosto, F

    1989-01-01

    The kinetic changes induced by granulocyte-macrophage colony-stimulating factor (GM-CSF) on hemopoietic cells were assessed in physiological conditions by administering GM-CSF (8 micrograms/kg per d) for 3 d to nine patients with solid tumors and normal bone marrow (BM), before chemotherapy. GM-CSF increased the number of circulating granulocytes and monocytes; platelets, erythrocytes, lymphocyte number, and subsets were unmodified. GM-CSF increased the percentage of BM S phase BFU-E (from 32 +/- 7 to 79 +/- 16%), day 14 colony-forming unit granulocyte-macrophage (CFU-GM) (from 43 +/- 20 to 82 +/- 11%) and day 7 CFU-GM (from 41 +/- 14 to 56 +/- 20%). The percentage of BM myeloblasts, promyelocytes, and myelocytes in S phase increased from 26 +/- 14 to 41 +/- 6%, and that of erythroblasts increased from 25 +/- 12 to 30 +/- 12%. This suggests that GM-CSF activates both erythroid and granulomonopoietic progenitors but that, among the morphologically recognizable BM precursors, only the granulomonopoietic lineage is a direct target of the molecule. GM-CSF increased the birth rate of cycling cells from 1.3 to 3.4 cells %/h and decreased the duration of the S phase from 14.3 to 9.1 h and the cell cycle time from 86 to 26 h. After treatment discontinuation, the number of circulating granulocytes and monocytes rapidly fell. The proportion of S phase BM cells dropped to values lower than pretreatment levels, suggesting a period of relative refractoriness to cell cycle-active antineoplastic agents. PMID:2643633

  8. Granulocyte colony stimulating factor in neutropenic patients with infective endocarditis

    PubMed Central

    Borgbjerg, B. M.; Hovgaard, D.; Laursen, J. B.; Aldershvile, J.

    1998-01-01

    A well known complication in the treatment of infectious endocarditis is development of neutropenia caused by treatment with antibiotics in high concentrations over long periods. Neutropenia often necessitates discontinuation of antibiotic treatment. Three patients with infectious endocarditis who developed neutropenia are reported. The patients were treated with granulocyte colony stimulating factor (G-CSF), a haematopoietic growth factor that stimulates neutrophils. G-CSF induced an immediate increase in white blood cell count, primarily neutrophils. G-CSF may be effective in ameliorating neutropenia in patients who receive antibiotics for treatment of infectious endocarditis.

 Keywords: granulocyte colony stimulating factor;  neutropenia;  endocarditis PMID:9505928

  9. Administration of recombinant human granulocyte colony-stimulating factor (rhG-CSF) for the intracranial hemorrhage in two dogs: a case report.

    PubMed

    Kang, M H; Park, H M

    2016-01-01

    Two dogs with generalized seizures were evaluated. The dogs were diagnosed with traumatic intracranial hemorrhages based on the history, neurological examinations, and magnetic resonance imaging (MRI) of the brain. Treatment was started with oxygen, prednisolone and anticonvulsant agents. No further seizure activity was observed after treatment in both dogs, however cushing reflex was detected in case 1 and a left-sided hemi-paresis was detected in case 2. Further supportive treatment with recombinant human granulocyte colony-stimulating factor (rhG-CSF) was attempted. No abnormal signs were noted in either of the dogs and no recurrence was noted 16 and 14 months later, in case 1 and 2, respectively. These cases indicate that a combination of rhG-CSF treatment with previous therapy could be used in dogs with traumatic brain injury. PMID:27656233

  10. Administration of recombinant human granulocyte colony-stimulating factor (rhG-CSF) for the intracranial hemorrhage in two dogs: a case report

    PubMed Central

    Kang, M. H.; Park, H. M.

    2016-01-01

    Two dogs with generalized seizures were evaluated. The dogs were diagnosed with traumatic intracranial hemorrhages based on the history, neurological examinations, and magnetic resonance imaging (MRI) of the brain. Treatment was started with oxygen, prednisolone and anticonvulsant agents. No further seizure activity was observed after treatment in both dogs, however cushing reflex was detected in case 1 and a left-sided hemi-paresis was detected in case 2. Further supportive treatment with recombinant human granulocyte colony-stimulating factor (rhG-CSF) was attempted. No abnormal signs were noted in either of the dogs and no recurrence was noted 16 and 14 months later, in case 1 and 2, respectively. These cases indicate that a combination of rhG-CSF treatment with previous therapy could be used in dogs with traumatic brain injury.

  11. Administration of recombinant human granulocyte colony-stimulating factor (rhG-CSF) for the intracranial hemorrhage in two dogs: a case report

    PubMed Central

    Kang, M. H.; Park, H. M.

    2016-01-01

    Two dogs with generalized seizures were evaluated. The dogs were diagnosed with traumatic intracranial hemorrhages based on the history, neurological examinations, and magnetic resonance imaging (MRI) of the brain. Treatment was started with oxygen, prednisolone and anticonvulsant agents. No further seizure activity was observed after treatment in both dogs, however cushing reflex was detected in case 1 and a left-sided hemi-paresis was detected in case 2. Further supportive treatment with recombinant human granulocyte colony-stimulating factor (rhG-CSF) was attempted. No abnormal signs were noted in either of the dogs and no recurrence was noted 16 and 14 months later, in case 1 and 2, respectively. These cases indicate that a combination of rhG-CSF treatment with previous therapy could be used in dogs with traumatic brain injury. PMID:27656233

  12. Infection of human synovial cells by human T cell lymphotropic virus type I. Proliferation and granulocyte/macrophage colony-stimulating factor production by synovial cells.

    PubMed Central

    Sakai, M; Eguchi, K; Terada, K; Nakashima, M; Yamashita, I; Ida, H; Kawabe, Y; Aoyagi, T; Takino, H; Nakamura, T

    1993-01-01

    The present study was performed to clarify the relationship between human T cell lymphotropic virus type I (HTLV-I) infection and chronic inflammatory arthropathy. To determine the ability of HTLV-I to infect synovial cells and the effect on synovial cell proliferation, synovial cells were cocultured with the HTLV-I-producing T cell lines (MT-2 or HCT-1). After coculture with HTLV-I-infected T cells, the synovial cells expressed HTLV-I-specific core antigens, and HTLV-I proviral DNA was detected from the synovial cells by polymerase chain reaction. These cocultured synovial cells with HTLV-I-infected T cells proliferated more actively than the synovial cells cocultured with uninfected T cells. This stimulatory effect of HTLV-I-infected T cells on synovial cell proliferation seems necessary to contact each other. After being cocultured with MT-2 cells, synovial cells proliferated more actively than control cells even after several passages. Furthermore, HTLV-I-infected synovial cells produced significant amounts of granulocyte/macrophage colony-stimulating factor. These results suggest that HTLV-I can infect synovial cells, resulting their active proliferation and may be involved in the pathogenesis of proliferative synovitis similar to that found in rheumatoid arthritis. Images PMID:8408648

  13. Granulocyte colony-stimulating factor (G-CSF): A saturated fatty acid-induced myokine with insulin-desensitizing properties in humans

    PubMed Central

    Ordelheide, Anna-Maria; Gommer, Nadja; Böhm, Anja; Hermann, Carina; Thielker, Inga; Machicao, Fausto; Fritsche, Andreas; Stefan, Norbert; Häring, Hans-Ulrich; Staiger, Harald

    2016-01-01

    Objective Circulating long-chain free fatty acids (FFAs) are important metabolic signals that acutely enhance fatty acid oxidation, thermogenesis, energy expenditure, and insulin secretion. However, if chronically elevated, they provoke inflammation, insulin resistance, and β-cell failure. Moreover, FFAs act via multiple signaling pathways as very potent regulators of gene expression. In human skeletal muscle cells differentiated in vitro (myotubes), we have shown in previous studies that the expression of CSF3, the gene encoding granulocyte colony-stimulating factor (G-CSF), is markedly induced upon FFA treatment and exercise. Methods and results We now report that CSF3 is induced in human myotubes by saturated, but not unsaturated, FFAs via Toll-like receptor 4-dependent and -independent pathways including activation of Rel-A, AP-1, C/EBPα, Src, and stress kinases. Furthermore, we show that human adipocytes and myotubes treated with G-CSF become insulin-resistant. In line with this, a functional polymorphism in the CSF3 gene affects adipose tissue- and whole-body insulin sensitivity and glucose tolerance in human subjects with elevated plasma FFA concentrations. Conclusion G-CSF emerges as a new player in FFA-induced insulin resistance and thus may be of interest as a target for prevention and treatment of type 2 diabetes. PMID:27069870

  14. Recombinant human granulocyte-macrophage colony-stimulating factor (rH GM-CSF) regulates f Met-Leu-Phe receptors on human neutrophils.

    PubMed Central

    Atkinson, Y H; Lopez, A F; Marasco, W A; Lucas, C M; Wong, G G; Burns, G F; Vadas, M A

    1988-01-01

    The regulation of mature human neutrophil function by recombinant human granulocyte-macrophage colony-stimulating factor (rH GM-CSF) was studied. Preincubation of neutrophils with this CSF did not stimulate superoxide anion directly but enhanced the subsequent release of superoxide anion in response to stimulation with the bacterial product formylmethionylleucyl-phenylalanine (f Met-Leu-Phe). Enhanced superoxide anion production was evident by 5 min and reached a plateau at 30 min. In contrast, neutrophils preincubated with rH GM-CSF exhibited reduced chemotaxis under agarose in response to a gradient of f Met-Leu-Phe. The inhibition of neutrophil migration was dependent on the dose of rH GM-CSF and exhibited a time-course similar to the effect on superoxide production. Binding studies of f Met-Leu-[3H]Phe to purified human neutrophils revealed heterogeneous binding to unstimulated cells. Two affinity components were identified. The high-affinity component consisted of approximately 2000 sites/cell and had an average Kd of 4 +/- 2 nM (n = 6). The low-affinity component consisted of approximately 40,000 sites/cell and had an average Kd of 220 +/- 130 nM (n = 6). rH GM-CSF caused conversion to a linear Scatchard plot showing no significant change in total binding sites but a single Kd of 30 +/- 10 nM. These data indicate that rH GM-CSF may influence neutrophil responses to f Met-Leu-Phe by regulating the affinity of f Met-Leu-Phe receptors. PMID:2842255

  15. Granulocyte colony-stimulating factor and reproductive medicine: A review

    PubMed Central

    Cavalcante, Marcelo Borges; Costa, Fabrício DA Silva; Barini, Ricardo; Araujo Júnior, Edward

    2015-01-01

    Background: Recently, the use of granulocyte colony-stimulating factor (G-CSF) has been proposed to improve pregnancy outcomes in reproductive medicine. Objective: A systematic review of the current use of G-CSF in patients who have difficulty conceiving and maintaining pregnancy was performed. Materials and Methods: Two electronic databases (PubMed/ Medline and Scopus) were searched. Study selection, data extraction and quality assessment were performed in duplicate. The subject codes used were granulocyte colony-stimulating factor, G-CSF, recurrent miscarriage, IVF failure, and endometrium. Results: The search of electronic databases resulted in 215 citations (PubMed/ Medline: 139 and Scopus: 76), of which 38 were present in both databases. Of the remaining 177 publications, seven studies were included in the present review. Conclusion: Treatment with G-CSF is a novel proposal for immune therapy in patients with recurrent miscarriage and implantation failure following cycles of IVF. However, a larger number of well-designed studies are required for this treatment to be established. PMID:26131007

  16. High pH solubilization and chromatography-based renaturation and purification of recombinant human granulocyte colony-stimulating factor from inclusion bodies.

    PubMed

    Li, Ming; Fan, Hua; Liu, Jiahua; Wang, Minhong; Wang, Lili; Wang, Chaozhan

    2012-03-01

    Recombinant human granulocyte colony-stimulating factor (rhG-CSF) is a very efficient therapeutic protein drug which has been widely used in human clinics to treat cancer patients suffering from chemotherapy-induced neutropenia. In this study, rhG-CSF was solubilized from inclusion bodies by using a high-pH solution containing low concentration of urea. It was found that solubilization of the rhG-CSF inclusion bodies greatly depended on the buffer pH employed; alkalic pH significantly favored the solubilization. In addition, when small amount of urea was added to the solution at high pH, the solubilization was further enhanced. After solubilization, the rhG-CSF was renatured with simultaneous purification by using weak anion exchange, strong anion exchange, and hydrophobic interaction chromatography, separately. The results indicated that the rhG-CSF solubilized by the high-pH solution containing low concentration of urea had much higher mass recovery than the one solubilized by 8 M urea when using anyone of the three refolding methods employed in this work. In the case of weak anion exchange chromatography, the high pH solubilized rhG-CSF could get a mass recovery of 73%. The strategy of combining solubilization of inclusion bodies at high pH with refolding of protein using liquid chromatography may become a routine method for protein production from inclusion bodies.

  17. Effect of recombinant human granulocyte colony-stimulating factor on hemodynamic and cytokine response in a porcine model of Pseudomonas sepsis.

    PubMed

    Haberstroh, J; Breuer, H; Lücke, I; Massarrat, K; Früh, R; Mand, U; Hagedorn, P; Brunnberg, L; von Specht, B U

    1995-09-01

    To investigate the effect of recombinant human granulocyte colony-stimulating factor (rhG-CSF) on sepsis, chronically catheterized conscious pigs were challenged with Pseudomonas aeruginosa (8 x 10(7) colony-forming units kg-1 h-1) for 84 h (Group A, n = 8). Group B (n = 7) also received rhG-CSF at 5 micrograms kg-1 d-1, the first dose being given 30 min before starting bacterial infusion. Two of the animals in Group A died from pulmonary failure, whereas all those treated with rh-GCSF survived. Fever, severe pulmonary hypertension and systemic hypotension--the latter accompanied at first by a transient hypodynamic, and later a hyperdynamic response--were observed in all of the animals. In Group B, however, the rise in temperature, mean pulmonary arterial pressure (at a later stage of the observation), plasma levels of tumor necrosis factor, and endotoxin were significantly less than in Group A. In the rhG-CSF-treated pigs, an initial leukopenia completely recovered within 24 h (p < .05 vs. Group A). These data suggest that rhG-CSF might be beneficial in the treatment of sepsis.

  18. A Randomized Controlled Phase III Trial of Recombinant Human Granulocyte Colony-Stimulating Factor (Filgrastim) for Treatment of Severe Chronic Neutropenia

    PubMed Central

    Dale, David C.; Bonilla, Mary Ann; Davis, Mark W.; Nakanishi, Arline M.; Hammond, William P.; Kurtzberg, Joanne; Wang, Winfred; Jakubowski, Ann; Winton, Elliott; Lalezari, Parviz; Robinson, William; Glaspy, John A.; Emerson, Steve; Gabrilove, Janice; Vincent, Martha; Boxer, Laurence A.

    2014-01-01

    Patients with idiopathic, cyclic, and congenital neutropenia have recurrent severe bacterial infections. One hundred twenty-three patients with recurrent infections and severe chronic neutropenia (absolute neutrophil count < 0.5 × 109/L) due to these diseases were enrolled in this multi-center phase III trial. They were randomized to either immediately beginning recombinant human granulocyte colony-stimulating factor (filgrastim) (3.45 to 11.50 μg/kg/d, subcutaneously) or entering a 4-month observation period followed by filgrastim administration. Blood neutrophil counts, bone marrow (BM) cell histology, and incidence and duration of infection-related events were monitored. Of the 123 patients enrolled, 120 received filgrastim. On therapy, 108 patients had a median absolute neutrophil count of ≥ 1.5 × 109/L. Examination of BM aspirates showed increased proportions of maturing neutrophils. Infection-related events were significantly decreased (P < .05) with approximately 50% reduction in the incidence and duration of infection-related events and almost 70% reduction in duration of antibiotic use. Asymptomatic splenic enlargement occurred frequently: adverse events frequently reported were bone pain, headache, and rash, which were generally mild and easily manageable. These data indicate that treatment of patients with severe chronic neutropenia with filgrastim results in a stimulation of BM production and maturation of neutrophils, an increase in circulating neutrophils, and a reduction in infection-related events. PMID:8490166

  19. Comparative pharmacokinetics of single-dose administration of mammalian and bacterially-derived recombinant human granulocyte-macrophage colony-stimulating factor.

    PubMed

    Hovgaard, D; Mortensen, B T; Schifter, S; Nissen, N I

    1993-01-01

    Pharmacokinetics of recombinant human non-glycosylated bacterially-synthesized (E. coli) granulocyte-macrophage colony-stimulating factor (GM-CSF) were studied following single intravenous (i.v.) and subcutaneous (s.c.) bolus injection, and compared to equivalent doses of glycosylated mammalian-derived CHO-GM-CSF. Each route of administration gave a different GM-CSF concentration-time profile. The highest peak serum concentrations (Cmax) were observed following i.v. bolus injection. After i.v. administration, a two-phase decline in concentration was noted for both types of GM-CSF with a significantly shorter t1/2 alpha of 7.8 minutes for the E. coli GM-CSF versus 20.0 min for the CHO-GM-CSF, while no significant difference was observed for the terminal phase. Following s.c. administration of equivalent doses, a higher peak serum concentration was observed in the E. coli-treated patients and, again, a faster elimination where pretreatment serum levels were reached after 16-20 h, versus more than 48 h after administration of CHO-GM-CSF. Although the non-glycosylated E. coli GM-CSF thus seems to undergo a faster elimination that the glycosylated CHO-GM-CSF no significant difference could be demonstrated in the in vivo effect of corresponding doses of the two compounds with respect to stimulation of granulopoiesis--with reservation for small patient numbers and a large individual variations in response.

  20. Granulocyte-Macrophage Colony-Stimulating Factor- and Tumor Necrosis Factor Alpha-Mediated Matrix Metalloproteinase Production by Human Osteoblasts and Monocytes after Infection with Brucella abortus ▿

    PubMed Central

    Scian, Romina; Barrionuevo, Paula; Giambartolomei, Guillermo H.; Fossati, Carlos A.; Baldi, Pablo C.; Delpino, M. Victoria

    2011-01-01

    Osteoarticular complications are common in human brucellosis, but the pathogenic mechanisms involved are largely unknown. Since matrix metalloproteinases (MMPs) are involved in joint and bone damage in inflammatory and infectious diseases, we investigated the production of MMPs by human osteoblasts and monocytes, either upon Brucella abortus infection or upon reciprocal stimulation with factors produced by each infected cell type. B. abortus infection of the normal human osteoblastic cell line hFOB 1.19 triggered a significant release of MMP-2, which was mediated in part by granulocyte-macrophage colony-stimulating factor (GM-CSF) acting on these same cells. Supernatants from infected osteoblasts exhibited increased levels of monocyte chemoattractant protein 1 and induced the migration of human monocytes (THP-1 cell line). Infection with B. abortus induced a high MMP-9 secretion in monocytes, which was also induced by heat-killed B. abortus and by the Omp19 lipoprotein from B. abortus. These effects were mediated by Toll-like receptor 2 and by the action of tumor necrosis factor alpha (TNF-α) produced by these same cells. Supernatants from B. abortus-infected monocytes induced MMP-2 secretion in uninfected osteoblasts, and this effect was mediated by TNF-α. Similarly, supernatants from infected osteoblasts induced MMP-9 secretion in uninfected monocytes. This effect was mediated by GM-CSF, which induced TNF-α production by monocytes, which in turn induced MMP-9 in these cells. These results suggest that MMPs could be potentially involved in the tissue damage observed in osteoarticular brucellosis. PMID:20956574

  1. Antigen activation of THP-1 human monocytic cells after stimulation with lipopolysaccharide from oral microorganisms and granulocyte-macrophage colony-stimulating factor.

    PubMed

    Baqui, A A; Meiller, T F; Kelley, J I; Turng, B F; Falkler, W A

    1999-05-01

    A human THP-1 monocyte cell line culture system has been utilized to evaluate the morphological changes in THP-1 cells and to measure expression of activation antigens (CD-11b, CD-11c, CD-14, CD-35, CD-68, CD-71 and HLA-DR) as evidence of maturation of THP-1 cells in response to stimulation by lipopolysaccharide (LPS) from the oral microorganisms, Fusobacterium nucleatum and Porphyromonas gingivalis, and granulocyte-macrophage colony-stimulating factor. THP-1 cells were stimulated with LPS (1 microgram/ml) of P. gingivalis or F. nucleatum for different time periods (1, 2, 4 and 7 d). Detection of different activation antigens on THP-1 cells was performed by indirect immunohistochemical staining followed by light microscopy. Confirmational studies were performed in parallel using indirect immunofluorescence and immunogold electron microscopy for detection of the corresponding activation antigens. Expression of different activation antigens by resting THP-1 cells revealed HLA-DR to be on 3% of the cells; CD-11b, 9%; CD-11c, 8%; CD-14, 22%; CD-35, 9% and CD-68, 7%. The CD-71 activation antigen was not expressed in untreated THP-1 cells. LPS stimulation increased expression of all activation antigens. A significant (p < 0.05) increase in expression of CD-11b, CD-11c, CD-14, CD-35, CD-68 and CD-71 was observed when GM-CSF (50 IU/ml) was supplemented during the treatment of THP-1 cells with LPS of F. nucleatum or P. gingivalis. Activation and differentiation of THP-1 cells by LPS from oral microorganisms in the presence of GM-CSF supports a role for human macrophages in acute and chronic periodontal diseases and may explain the clinically observable periodontal exacerbations in some patients after GM-CSF therapy.

  2. Osteoporosis in severe congenital neutropenia treated with granulocyte colony-stimulating factor.

    PubMed

    Bishop, N J; Williams, D M; Compston, J C; Stirling, D M; Prentice, A

    1995-04-01

    Recombinant human granulocyte colony-stimulating factor (G-CSF) has substantially improved life expectancy for children with severe congenital neutropenia (SCN). Severe osteoporosis, reported in this population, may relate to the disease process, or be a therapeutic side-effect. This report details bone loss, quantitated absorptiometrically and histomorphometrically, in a child with SCN and vertebral collapse, and the positive response to anabolic steroid and bisphosphonate therapy.

  3. [Successful treatment of agranulocytosis caused by carbimazole using recombinant granulocyte-macrophage colony stimulating factor].

    PubMed

    Kreze, A; Kuviková, A; Laca, L; Kompis, S; Dobáková, M; Babusík, P; Matecek, L; Pekárová, E

    1995-07-01

    The authors describe a patient with hyperthyroidism who developed after three weeks' treatment with carbimazole agranulocytosis and the condition was complicated by septic shock. The authors used for treatment in addition to antibiotics and corticoids, for the first time in their practice, human recombinant granulocyte macrophage colonies stimulating factor (rHuGM-CSF) which produced a very favourable effect. In the meantime the patient was prepared for goitrectomy, operated and discharged in a satisfactory condition to have ambulatory treatment.

  4. Granulocyte/macrophage colony-stimulating factor stimulates the expression of the 5-lipoxygenase-activating protein (FLAP) in human neutrophils

    PubMed Central

    1994-01-01

    The synthesis of leukotrienes in human blood neutrophils chiefly relies on the activity of two enzymes, phospholipase A2 and 5-lipoxygenase (5- LO). In turn, the activation of the 5-LO requires the participation of a recently characterized membrane-bound protein, the 5-LO-activating protein (FLAP). In this study, we have investigated conditions under which FLAP expression in neutrophils may be modulated. Of several cytokines tested, only granulocyte/macrophage colony-stimulating factor (GM-CSF) (and to a lesser extent tumor necrosis factor alpha) significantly increased expression of FLAP. GM-CSF increased FLAP mRNA steady-state levels in a time- and dose-dependent manner. The stimulatory effect of GM-CSF on FLAP mRNA was inhibited by prior treatment of the cells with the transcription inhibitor, actinomycin D, and pretreatment of the cells with the protein synthesis inhibitor, cycloheximide, failed to prevent the increase in FLAP mRNA induced by GM-CSF. The accumulation of newly synthesized FLAP, as determined by immunoprecipitation after incorporation of 35S-labeled amino acids, was also increased after incubation of neutrophils with GM-CSF. In addition, the total level of FLAP protein was increased in GM-CSF- treated neutrophils, as determined by two-dimensional gel electrophoresis, followed by Western blot. GM-CSF did not alter the stability of the FLAP protein, indicating that the effect of GM-CSF on FLAP accumulation was the consequence of increased de novo synthesis as opposed to decreased degradation of FLAP. Finally, incubation of neutrophils with the synthetic glucocorticoid dexamethasone directly stimulated the upregulation of FLAP mRNA and protein, and enhanced the effect of GM-CSF. Taken together, these data demonstrate that FLAP expression may be upmodulated after appropriate stimulation of neutrophils. The increase in FLAP expression induced by GM-CSF in inflammatory conditions could confer upon neutrophils a prolonged capacity to synthesize

  5. A trial of recombinant human granulocyte colony stimulating factor for the treatment of very low birthweight infants with presumed sepsis and neutropenia

    PubMed Central

    Russell, A; Emmerson, A; Wilkinson, N; Chant, T; Sweet, D; Halliday, H; Holland, B; Davies, E

    2001-01-01

    OBJECTIVES—The primary objective was to investigate the safety of recombinant human granulocyte colony stimulating factor (rhG-CSF) for the treatment of very low birthweight infants (VLBW) with sepsis and relative neutropenia, specifically with regard to worsening of respiratory distress and thrombocytopenia and all cause mortality. Secondary objectives were to evaluate duration of ventilation, intensive care, and antibiotic use as markers of efficacy.
DESIGN—Neonates (⩽ 28 days) in intensive care, with birth weights of 500-1500 g, absolute neutrophil count (ANC) of ⩽ 5 × 109/l, and clinical evidence of sepsis, were randomly assigned to receive either rhG-CSF (10 µg/kg/day) administered intravenously (n = 13), or placebo (n = 15) for a maximum of 14 days, in addition to standard treatment and antibiotics. All adverse events, oxygenation index, incidence of thrombocytopenia, all cause mortality, duration of ventilation, intensive care and antibiotic treatment, and ANC recovery were compared between the two groups.
RESULTS—Adverse events and oxygenation index were not increased by, and thrombocytopenia was not attributable to, treatment with rhG-CSF. At 6 and 12 months postmenstrual age, there were significantly fewer deaths in the group receiving rhG-CSF (1/13 v 7/15; p ⩽ 0.038). There was a non-significant trend towards a reduction in duration of ventilation, intensive care, and antibiotic use in the rhG-CSF group. There was a significantly more rapid increase in ANC in the rhG-CSF treated babies (p < 0.001).
CONCLUSIONS—In a small randomised placebo controlled trial in a highly selected group of neonates, adjuvant treatment with rhG-CSF increased ANC rapidly, and no treatment related adverse events were identified. Mortality at 6 and 12 months postmenstrual age was significantly lower in the treatment group. A large trial investigating efficacy in a similar group of neonates is warranted.
 PMID:11320043

  6. Differential augmentation of in vivo natural killer cytotoxicity in normal primates with recombinant human interleukin-1 and granulocyte-macrophage colony-stimulating factor.

    PubMed Central

    Davis, T A; Monroy, R L; Skelly, R R; Donahue, R E; MacVittie, T J

    1990-01-01

    The effect of recombinant human interleukin-1 (IL-1) alpha, granulocyte-macrophage colony-stimulating factor (GM-CSF), and combined factor therapy (CFT) on Rhesus monkey peripheral blood natural killer (NK) activity in vivo was compared. During a 14-day treatment period, IL-1-treated animals demonstrated a 170% increase in NK activity against K562 target cells by day 4, reaching maximal levels (300%) at day 16, and returning to baseline by day 30. NK activity of GM-CSF-treated monkeys increased slightly (60-100%) during days 4-12, as did saline-treated monkeys, but returned to baseline values by day 16. A delayed increase in NK activity resulted after GM-CSF treatment, reaching a peak (260%) on day 23 and remaining elevated through day 39. CFT resulted in a bimodal response pattern, with two peaks of NK activity: one at day 16 and a second at day 39. The first peak of activity (223%) was significantly less than the activity attained with IL-1 alone; the second peak (300%) was of greater duration and occurred later than the peak observed in GM-CSF-treated monkeys. Unlike IL-1, GM-CSF treatment did not lead to a immediate stimulation of NK activity; augmentation was delayed by more than 7 days post treatment. CFT results suggest that GM-CSF reduced the direct NK response to IL-1; while IL-1 led to an enhanced delayed NK response. Therefore, IL-1 and GM-CSF augment NK activity through different but interrelated pathways. PMID:2180599

  7. Antibiotics and production of granulocyte-macrophage colony-stimulating factor by human bronchial epithelial cells in vitro. A comparison of cefodizime and ceftriaxone.

    PubMed

    Pacheco, Y; Hosni, R; Dagrosa, E E; Gormand, F; Guibert, B; Chabannes, B; Lagarde, M; Perrin-Fayolle, M

    1994-04-01

    Cultured human bronchial epithelial cells (HBEC) produce both granulocyte-macrophage colony-stimulating factor (GM-CSF) and interleukin 8 (IL-8). The influence of cefodizime (CAS 69739-16-8), a new broad spectrum cephalosporin with immunostimulatory effects, and ceftriaxone on the production of GM-CSF and IL-8 in HBEC primary cultures was investigated. HBEC were isolated from biopsy specimens obtained during fibreoptic bronchoscopy in 12 patients (most frequent diagnosis: chronic bronchitis). Confluent monolayers of HBEC cultured on collagen were incubated for 24 h in a medium without study drugs (spontaneous production) or containing cefodizime or ceftriaxone at the clinically relevant concentrations of 1, 10 and 100 mg/l, with or without tumor necrosis factor alpha (TNF alpha, 100 U/ml). GM-CSF and IL-8 were measured in supernatant by ELISA technique. TNF alpha alone led to a significant (p < 0.005) increase in both GM-CSF and IL-8 production. Cefodizime induced a significant (p < 0.05), dose-dependent increase in GM-CSF release. No additive effect of cefodizime with TNF alpha was observed. Cefodizime did not affect IL-8 production and ceftriaxone had no influence on cytokine production. This is the first report of a stimulatory effect of a beta-lactam antibiotic on cytokine production by epithelial cells. GM-CSF production by epithelial cells is an important immunological step for neutrophil and monocyte recruitment and cell priming during lung defence. Previous studies with cefodizime in immunodepressed subjects have shown activation of phagocytosis and phagocytosis-related functions in non-lung phagocytes. An indirect mechanism of action, similar to that indicated by our results, may have been responsible for these stimulatory effects.(ABSTRACT TRUNCATED AT 250 WORDS)

  8. Granulocyte/macrophage colony-stimulating factor is an intrinsic keratinocyte-derived growth factor for human melanocytes in UVA-induced melanosis.

    PubMed

    Imokawa, G; Yada, Y; Kimura, M; Morisaki, N

    1996-01-15

    Recently we demonstrated that endothelins secreted from human keratinocytes act as intrinsic mitogens and melanogens for human melanocytes in UVB-induced melanosis. We show here that UVA-induced melanosis is associated with other keratinocyte-derived growth factors, secretion of which is specifically stimulated after exposure of human keratinocytes to UVA. Medium conditioned by UVA-exposed human keratinocytes elicited a significant increase in DNA synthesis by cultured human melanocytes in a UVA dose-dependent manner. Analysis of endothelin-1 and interleukin (IL)-1 alpha in the conditioned medium by ELISA, both of which are major keratinocyte-derived cytokines involved in UVB-associated melanocyte activation, revealed that UVA exposure did not cause human keratinocytes to stimulate the secretion of the two cytokines. In contrast, the levels of several other cytokines such as IL-6, IL-8 and granulocyte/macrophage colony-stimulating factor (GM-CSF) were significantly increased in the conditioned medium of human keratinocytes after exposure to UVA at a dose of 1.0 J/cm2. The gel chromatographic profile of UVA-exposed keratinocyte-conditioned medium demonstrated that there were two factors (P-1 and P-2) with molecular masses of approx. 20 and 1 kDa respectively that stimulate DNA synthesis in human melanocytes, and the larger species (P-1) also increased melanization as assessed by [14C]thiouracil incorporation. Quantitative analysis of cytokines in chromatographic fractions by ELISA revealed the P-1 fraction to be consistent with the molecular mass profile of GM-CSF. Furthermore the stimulatory effect of the P-1 fraction on DNA synthesis in human melanocytes was neutralized by antibodies to GM-CSF, but not to basic fibroblast growth factor or stem cell factor. Binding and proliferation assays with recombinant GM-CSF demonstrated that human melanocytes possess specific binding sites for GM-CSF(Kd 2.11 nM; binding sites, 2.5-3.5 x 10(4) per cell), and recombinant GM

  9. Modulation of colony stimulating factor release and apoptosis in human colon cancer cells by anticancer drugs

    PubMed Central

    Calatayud, S; Warner, T D; Mitchell, J A

    2002-01-01

    Modulation of the immune response against tumour cells is emerging as a valuable approach for cancer treatment. Some experimental studies have shown that secretion of colony stimulating factors by cancer cells reduces their tumorigenicity and increases their immunogenicity probably by promoting the cytolitic and antigen presenting activities of leukocytes. We have observed that human colon cancer cells (HT-29) are able to secrete granulocyte-macrophage-colony stimulating factor, granulocyte-colony stimulating factor and macrophage-colony stimulating factor when stimulated with cytokines (IL-1β and TNF-α). In this study we assessed, for the first time, the effects of several anticancer drugs on colony stimulating factor release or apoptosis in HT-29 cells. Cytokine-induced release of granulocyte-macrophage-colony stimulating factor, granulocyte-colony stimulating factor and macrophage-colony stimulating factor was significantly increased by cisplatin and 6-mercaptopurine. Taxol only increased macrophage-colony stimulating factor release while reduced that of granulocyte-colony stimulating factor. No changes in colony stimulating factor secretion were observed after treatment with methotrexate. Only cisplatin and taxol induced apoptosis in these cells. Secretion of colony stimulating factors by colon cancer cells may contribute to the immune host response against them. Anticancer drugs such as cisplatin and 6-mercaptopurine increase colony stimulating factor secretion by cytokine stimulated cancer cells probably through mechanisms different to those leading to cell apoptosis, an effect that may contribute to their anti-neoplasic action. British Journal of Cancer (2002) 86, 1316–1321. DOI: 10.1038/sj/bjc/6600240 www.bjcancer.com © 2002 Cancer Research UK PMID:11953891

  10. Recombinant granulocyte colony-stimulating factor (G-CSF) in infectious diseases: still a debate.

    PubMed

    Wiedermann, F J; Mittermayr, M; Hoffmann, G; Schobersberger, W

    2001-02-15

    Granulocyte colony-stimulating factor (G-CSF), a central mediator of the endogenous response to infection and inflammation, is approved for use in the prevention of infection-related complications in patients with nonmyeloid malignancies during antineoplastic therapy associated with high risk of severe neutropenia. Administration of granulocyte colony-stimulating factor results in improvement of host defence paired with anti-inflammatory effects. There is evidence from animal and clinical studies that administration of granulocyte colony-stimulating factor may also be beneficial in non-neutropenic infections. This review focuses mainly on the results of different animal and clinical studies of granulocyte colony stimulating factor used in the treatment of severe infections and sepsis.

  11. Neuroprotective effects of recombinant human granulocyte colony-stimulating factor (G-CSF) in a rat model of anterior ischemic optic neuropathy (rAION).

    PubMed

    Chang, Chung-Hsing; Huang, Tzu-Lun; Huang, Shun-Ping; Tsai, Rong-Kung

    2014-01-01

    The purpose of this study was to investigate the neuroprotective effects of recombinant human granulocyte colony stimulating factor (G-CSF), as administered in a rat model of anterior ischemic optic neuropathy (rAION). Using laser-induced photoactivation of intravenously administered Rose Bengal in the optic nerve head of 60 adult male Wistar rats, an anterior ischemic optic neuropathy (rAION) was inducted. Rats either immediately received G-CSF (subcutaneous injections) or phosphate buffered saline (PBS) for 5 consecutive days. Rats were euthanized at 4 weeks post infarct. Density of retinal ganglion cells (RGCs) was counted using retrograde labeling of Fluoro-gold. Visual function was assessed by flash visual-evoked potentials (FVEP) at 4 weeks. TUNEL assay in the retinal sections and immunohistochemical staining of ED1 (marker of macrophage/microglia) were investigated in the optic nerve (ON) specimens. The RGC densities in the central and mid-peripheral retinas in the G-CSF treated rats were significantly higher than those of the PBS-treated rats (survival rate was 71.4% vs. 33.2% in the central retina; 61.8% vs. 22.7% in the mid-peripheral retina, respectively; both p < 0.05). FVEP measurements showed a significantly better preserved latency and amplitude of the p1 wave in the G-CSF-treated rats than that of the PBS-treated rats (latency120 ± 11 ms vs. 142 ± 12 ms, p = 0.03; amplitude 50 ± 11 μv vs. 31 ± 13 μv, p = 0.04). TUNEL assays showed fewer apoptotic cells in the retinal ganglion cell layers of G-CSF treated rats [2.1 ± 1.0 cells/high power field (HPF) vs. 8.0 ± 1.5/HPF; p = 0.0001]. In addition, the number of ED1 positive cells was attenuated at the optic nerve sections of G-CSF-treated rats (16 ± 6/HPF vs. 35 ± 10/HPF; p = 0.016). In conclusion, administration of G-CSF is neuroprotective in the rat model of anterior ischemic optic neuropathy, as demonstrated both structurally by RGC density and functionally by

  12. Clinical observation of the therapeutic effects of pegylated recombinant human granulocyte colony-stimulating factor in patients with concurrent chemoradiotherapy-induced grade IV neutropenia

    PubMed Central

    WU, FENG-PENG; WANG, JUN; WANG, HUI; LI, NA; GUO, YIN; CHENG, YUN-JIE; LIU, QING; YANG, XIANG-RAN

    2015-01-01

    The aim of the present study was to investigate the efficacy and side-effects of preventive treatment with pegylated recombinant human granulocyte colony-stimulating factor (PEG-rhG-CSF) on concurrent chemoradiotherapy-induced grade IV neutropenia and to provide a rational basis for its clinical application. A total of 114 patients with concurrent chemoradiotherapy-induced grade IV neutropenia were enrolled. A randomized approach was used to divide the patients into an experimental group and a control group. The experimental group included three subgroups, namely a P-50 group, P-100 group and P + R group. The P-50 group had 42 cases, which were given a single 50-μg/kg subcutaneous injection of PEG-rhG-CSF. The P-100 group had 30 cases, which received a single 100-μg/kg subcutaneous injection of PEG-rhG-CSF. The P + R group comprised 22 cases, which were given a single 50-μg/kg subcutaneous injection of PEG-rhG-CSF and rhG-CSF 5 μg/kg/day; when the absolute neutrophil count (ANC) was ≥2.0×109/l, the administration of rhG-CSF was stopped. The control group (RC group) comprised 20 patients, who received rhG-CSF 5 μg/kg/day by subcutaneous injection until the ANC was ≥2.0×109/l. Changes in the neutrophil proliferation rate and ANC values over time, the neutropenic symptom remission time and incidence of adverse drug reactions were analyzed statistically in each group of patients. In the experimental group, the neutrophil proliferation rate and ANC values were significantly higher than those in the control group; the clinical effects began 12–24 h after treatment in the experimental group, and indicated that the treatment improved neutropenia in ~48 h after treatment. There was no significant difference in the neutrophil proliferation rate and ANC values between the P-50 and P+R groups. In the experimental group, the remission time of neutropenia-induced fever and muscle pain after administration was significantly shorter than that in the control group

  13. Prevention of corticosteroid-induced suppression of human polymorphonuclear leukocyte-induced damage of Aspergillus fumigatus hyphae by granulocyte colony-stimulating factor and gamma interferon.

    PubMed Central

    Roilides, E; Uhlig, K; Venzon, D; Pizzo, P A; Walsh, T J

    1993-01-01

    Neutrophils (PMNs) are a critical line of defense against Aspergillus fumigatus infection. Increased frequency of invasive aspergillosis has been observed in patients receiving corticosteroids, suggesting a deleterious effect of these compounds on PMN antifungal function. To investigate this hypothesis and to determine the potential preventive utility of granulocyte colony-stimulating factor (G-CSF) and gamma interferon (IFN-gamma), the effects of hydrocortisone (HCS) and dexamethasone (DXS) on PMN-induced damage of Aspergillus fumigatus hyphae were studied with or without pretreatment of PMNs with G-CSF and IFN-gamma. PMNs treated with HCS (> or = 3,000 microM) or DXS (> or = 10 microM) during a 2-h colorimetric tetrazolium metabolic assay (using methylthiotetrazolium) showed suppressed percentage of hyphal damage (P < 0.02). In addition, both HCS (> or = 30 microM) and DXS (> or = 1 microM) significantly suppressed oxidative burst measured as superoxide anion release by PMNs in response to opsonized and nonopsonized hyphae as well as to N-formylmethionyl leucyl phenylalanine. Pretreatment of PMNs with G-CSF (4,000 U/ml) and/or IFN-gamma (100 and 1,000 U/ml) for 90 min prevented the suppression of hyphal damage that occurred in the presence of HCS (3,000 microM; P < 0.01) or DXS (10 microM; P < or = 0.001). G-CSF (4,000 U/ml) and IFN-gamma (100 U/ml) combined had an additive effect on increasing the antifungal activity of HCS-treated but not of DXS-treated PMNs compared with IFN-gamma alone (P = 0.015). Thus, these findings reveal that corticosteroids impair PMN function in response to A. fumigatus and that G-CSF and IFN-gamma prevent this impairment. PMID:7691757

  14. [Sequential treatment with granulocyte-colony stimulating factor (GCSF) and human recombinant erythropoietin (rH-EPO) in anemia of a patient with myelodysplastic syndrome and high blood transfusion requirements)].

    PubMed

    Borbolla-Escoboza, J R; González-Avante, C M; López-Hernández, M A; Flores-Chapa, J D; Collados-Larumbe, M T

    1997-01-01

    We describe the case of a patient with myelodysplastic syndrome (MDS) classified as Refractory Anemia with our Excess blasts, who suffered from high transfusional requirements and who did not respond to the administrations of B12 vitamin, folates, danazol, low dose cytarabine or recombinant human erythropoietin (rHuEPO). The patient was administered two cytokines: granulocyte colony stimulating factor (G-CSF) followed by rHuEPO. The patient remained transfusion free for more than 4 months until his death from causes not related to MDS or the therapy he received. It is the opinion of the authors that the initial G-CSF administration stimulated the early erythroid precursors, making them capable of finishing their maturation when rHuEPO was administered. We believe that this could be a useful therapeutic measure in the treatment of patients with MDS and high transfusional requirements.

  15. Isolation and characterization of a resistant core peptide of recombinant human granulocyte-macrophage colony-stimulating factor (GM-CSF); confirmation of the GM-CSF amino acid sequence by mass spectrometry.

    PubMed Central

    Tsarbopoulos, A.; Pramanik, B. N.; Labdon, J. E.; Reichert, P.; Gitlin, G.; Patel, S.; Sardana, V.; Nagabhushan, T. L.; Trotta, P. P.

    1993-01-01

    A trypsin-resistant core peptide of recombinant human granulocyte-macrophage colony-stimulating factor (rhGM-CSF) was isolated and analyzed by high-energy Cs+ liquid secondary-ion (LSI) mass spectrometric analysis. This analysis provided successful detection of the high-mass disulfide-linked core peptide as well as information confirming the existence of disulfide pairing. Similarly, LSI mass spectrometric analysis of the peptide fragments isolated chromatographically from a Staphylococcus aureus V8 protease digest of rhGM-CSF provided rapid confirmation of the cDNA-derived sequence and determination of the existing disulfide bonds between cysteine residues 54-96 and 88-121. Electrospray ionization mass spectrometry was employed to measure the molecular weight of the intact protein and to determine the number of the disulfide bonds in the protein molecule by comparative analysis of the protein before and after reduction with beta-mercaptoethanol. PMID:8268804

  16. Immunostimulation using granulocyte- and granulocyte-macrophage colony stimulating factor in patients with severe sepsis and septic shock.

    PubMed

    Schefold, Joerg C

    2011-01-01

    Sepsis is associated with failure of multiple organs, including failure of the immune system. The resulting 'sepsis-associated immunosuppression' resembles a state of immunological anergy that is characterized by repeated 'infectious hits', prolonged multiple-organ failure, and death. As a consequence, adjunctive treatment approaches using measures of immunostimulation with colony-stimulating factors (CSFs) were tested in animal experiments and clinical trials. Herein, data from randomized clinical trials will be discussed in the context of a recently published meta-analysis investigating the effects of granulocyte- and granulocyte-macrophage colony-stimulating factor therapy in patients with severe sepsis and septic shock.

  17. Protective effects of granulocyte colony-stimulating factor on endotoxin shock in mice with retrovirus-induced immunodeficiency syndrome.

    PubMed

    Toki, S; Hiromatsu, K; Aoki, Y; Makino, M; Yoshikai, Y

    1997-10-01

    Mice with retrovirus-induced murine acquired immunodeficiency syndrome (MAIDS) were hypersensitive to lipopolysaccharide (LPS)-induced lethal shock accompanied by marked elevations of systematic interleukin 1beta (IL-beta) and interferon gamma (IFN-gamma) after LPS challenge. Pretreatment with 10 microg of recombinant human granulocyte colony-stimulating factor (rhG-CSF) protected MAIDS mice from hypersensitivity to LPS-induced lethal shock and this protection was concomitant with suppression of IFN-gamma production.

  18. Establishment of a retinoic acid-resistant human acute promyelocytic leukaemia (APL) model in human granulocyte-macrophage colony-stimulating factor (hGM-CSF) transgenic severe combined immunodeficiency (SCID) mice.

    PubMed Central

    Fukuchi, Y.; Kizaki, M.; Kinjo, K.; Awaya, N.; Muto, A.; Ito, M.; Kawai, Y.; Umezawa, A.; Hata, J.; Ueyama, Y.; Ikeda, Y.

    1998-01-01

    To understand the mechanisms and identify novel approaches to overcoming retinoic acid (RA) resistance in acute promyelocytic leukaemia (APL), we established the first human RA-resistant APL model in severe combined immunodeficiency (SCID) mice. UF-1 cells, an RA-resistant APL cell line established in our laboratory, were transplanted into human granulocyte-macrophage colony-stimulating factor (GM-CSF)-producing SCID (hGMTg SCID) mice and inoculated cells formed subcutaneous tumours in all hGMTg SCID mice, but not in the non-transgenic control SCID mice. Single-cell suspensions (UF-1/GMTg SCID cells) were similar in morphological, immunological, cytogenetic and molecular genetic features to parental UF-1 cells. All-trans RA did not change the morphological features of cells or their expression of CD11b. RA did not alter the growth curve of cells as determined by MTT assay, suggesting that UF-1/GMTg SCID cells are resistant to RA. These results demonstrate that this is the first RA-resistant APL animal model that may be useful for investigating the biology of this myeloid leukaemia in vivo, as well as for evaluating novel therapeutic approaches including patients with RA-resistant APL. Images Figure 1 Figure 2 Figure 3 Figure 4 PMID:9764578

  19. The Granulocyte-colony stimulating factor has a dual role in neuronal and vascular plasticity

    PubMed Central

    Wallner, Stephanie; Peters, Sebastian; Pitzer, Claudia; Resch, Herbert; Bogdahn, Ulrich; Schneider, Armin

    2015-01-01

    Granulocyte-colony stimulating factor (G-CSF) is a growth factor that has originally been identified several decades ago as a hematopoietic factor required mainly for the generation of neutrophilic granulocytes, and is in clinical use for that. More recently, it has been discovered that G-CSF also plays a role in the brain as a growth factor for neurons and neural stem cells, and as a factor involved in the plasticity of the vasculature. We review and discuss these dual properties in view of the neuroregenerative potential of this growth factor. PMID:26301221

  20. Cytokine refacing effect reduces granulocyte macrophage colony-stimulating factor susceptibility to antibody neutralization.

    PubMed

    Heinzelman, Pete; Carlson, Sharon J; Cox, George N

    2015-10-01

    Crohn's Disease (CD) afflicts over half a million Americans with an annual economic impact exceeding $10 billion. Granulocyte macrophage colony-stimulating factor (GM-CSF) can increase patient immune responses against intestinal microbes that promote CD and has been effective for some patients in clinical trials. We have made important progress toward developing GM-CSF variants that could be more effective CD therapeutics by virtue of being less prone to neutralization by the endogenous GM-CSF autoantibodies that are highly expressed in CD patients. Yeast display engineering revealed mutations that increase GM-CSF variant binding affinity by up to ∼3-fold toward both GM-CSF receptor alpha and beta subunits in surface plasmon resonance experiments. Increased binding affinity did not reduce GM-CSF half-maximum effective concentration (EC50) values in conventional in vitro human leukocyte proliferation assays. Affinity-enhancing mutations did, however, promote a 'refacing effect' that imparted all five evaluated GM-CSF variants with increased in vitro bioactivity in the presence of GM-CSF-neutralizing polyclonal antisera. The most improved variant, H15L/R23L, was 6-fold more active than wild-type GM-CSF. Incorporation of additional known affinity-increasing mutations could augment the refacing effect and concomitant bioactivity improvements described here. PMID:25855658

  1. RUNX1 haploinsufficiency results in granulocyte colony-stimulating factor hypersensitivity.

    PubMed

    Chin, D W L; Sakurai, M; Nah, G S S; Du, L; Jacob, B; Yokomizo, T; Matsumura, T; Suda, T; Huang, G; Fu, X-Y; Ito, Y; Nakajima, H; Osato, M

    2016-01-01

    RUNX1/AML1 is among the most commonly mutated genes in human leukemia. Haploinsufficiency of RUNX1 causes familial platelet disorder with predisposition to myeloid malignancies (FPD/MM). However, the molecular mechanism of FPD/MM remains unknown. Here we show that murine Runx1(+/-) hematopoietic cells are hypersensitive to granulocyte colony-stimulating factor (G-CSF), leading to enhanced expansion and mobilization of stem/progenitor cells and myeloid differentiation block. Upon G-CSF stimulation, Runx1(+/-) cells exhibited a more pronounced phosphorylation of STAT3 as compared with Runx1(+/+) cells, which may be due to reduced expression of Pias3, a key negative regulator of STAT3 signaling, and reduced physical sequestration of STAT3 by RUNX1. Most importantly, blood cells from a FPD patient with RUNX1 mutation exhibited similar G-CSF hypersensitivity. Taken together, Runx1 haploinsufficiency appears to predispose FPD patients to MM by expanding the pool of stem/progenitor cells and blocking myeloid differentiation in response to G-CSF. PMID:26745853

  2. Hematologic improvement in dogs with parvovirus infection treated with recombinant canine granulocyte-colony stimulating factor.

    PubMed

    Duffy, A; Dow, S; Ogilvie, G; Rao, S; Hackett, T

    2010-08-01

    Previously, dogs with canine parvovirus-induced neutropenia have not responded to treatment with recombinant human granulocyte-colony stimulating factor (rhG-CSF). However, recombinant canine G-CSF (rcG-CSF) has not been previously evaluated for treatment of parvovirus-induced neutropenia in dogs. We assessed the effectiveness of rcG-CSF in dogs with parvovirus-induced neutropenia with a prospective, open-label, nonrandomized clinical trial. Endpoints of our study were time to recovery of WBC and neutrophil counts, and duration of hospitalization. 28 dogs with parvovirus and neutropenia were treated with rcG-CSF and outcomes were compared to those of 34 dogs with parvovirus and neutropenia not treated with rcG-CSF. We found that mean WBC and neutrophil counts were significantly higher (P < 0.05) in the 28 dogs treated with rcG-CSF compared to disease-matched dogs not treated with rcG-CSF. In addition, the mean duration of hospitalization was reduced (P = 0.01) in rcG-CSF treated dogs compared to untreated dogs. However, survival times were decreased in dogs treated with rcG-CSF compared to untreated dogs. These results suggest that treatment with rcG-CSF was effective in stimulating neutrophil recovery and shortening the duration of hospitalization in dogs with parvovirus infection, but indicate the need for additional studies to evaluate overall safety of the treatment.

  3. In vivo kinetics of 111indium-labelled autologous granulocytes following i.v. administration of granulocyte-macrophage colony-stimulating factor (GM-CSF).

    PubMed

    Hovgaard, D; Schifter, S; Rabøl, A; Mortensen, B T; Nissen, N I

    1992-04-01

    Administration of both glycosylated and non-glycosylated recombinant human granulocyte-macrophage colony-stimulating factor (rhGM-CSF) induces an immediate transient granulocytopenia of 1-3 hours' duration. In order to explore this phenomenon, granulocytes were labelled with 111Indium and the effect on the kinetics of granulocytes after administration of rhGM-CSF was studied in 10 previously untreated patients with malignant lymphoma. For both types and doses of rhGM-CSF, a significant and dramatic accumulation of the 111Indium-labelled granulocytes was observed in the lung within a few minutes after i.v. injection of rhGM-CSF. The accumulation of radioactivity coincided with the pronounced and transient granulocytopenia in peripheral blood. The 111Indium-labelled granulocytes later reappeared in the peripheral blood, indicating reversible pulmonary vascular margination of the granulocytes. Half-life of labelled granulocytes after reappearance was comparable to half-life values under normal conditions. The transient accumulation of granulocytes in the pulmonary vessels seems not to be of clinical importance in the management of patients, but it may to some degree explain previously described side-effects, such as transient hypoxemia ("first-dose" reaction) following administration of rhGM-CSF.

  4. Delivery of granulocyte-macrophage colony-stimulating factor in bioadhesive hydrogel stimulates migration of dendritic cells in models of human papillomavirus-associated (pre)neoplastic epithelial lesions.

    PubMed

    Hubert, Pascale; Evrard, Brigitte; Maillard, Catherine; Franzen-Detrooz, Elizabeth; Delattre, Luc; Foidart, Jean-Michel; Noël, Agnes; Boniver, Jacques; Delvenne, Philippe

    2004-11-01

    Because of the central role of dendritic cells and/or Langerhans cells(DC/LC) in the induction of cellular immune responses, pharmacological agents that modulate the recruitment of these cells might have a clinical interest. The present study was designed to evaluate the capacity of several pharmaceutical formulations to topically deliver granulocyte-macrophage colony-stimulating factor (GM-CSF) on human papillomavirus (HPV)-associated genital (pre)neoplastic lesions. The formulations were evaluated for their bioactivity and for their potential to recruit DC in organotypic cultures of HPV-transformed keratinocytes. We found that a bioadhesive polycarbophil gel (Noveon) at pH 5.5 is able to maintain the bioactivity of GM-CSF at 4 or 37 degrees C for at least 7 days, whereas a decreased activity of GM-CSF was observed when the molecule is included in other polymer gels. GM-CSF incorporated in the polycarbophil gel was also a potent factor in enhancing the colonization of DC into organotypic cultures of HPV-transformed keratinocytes since the infiltration of DC in the in vitro-formed (pre)neoplastic epithelium was very low under basal conditions and dramatically increased in the presence of GM-CSF gel. We next demonstrated that GM-CSF incorporated in polycarbophil gel induces the recruitment of human DC in a human (pre)neoplastic epithelium grafted into NOD/SCID mice. The efficacy of GM-CSF in this formulation was equivalent to that observed with liquid GM-CSF. These results suggest that GM-CSF incorporated in polycarbophil gel could play an important role in the recruitment of DC/LC in mucosal surfaces and be useful as a new immunotherapeutic approach for genital HPV-associated (pre)neoplastic lesions.

  5. High-level expression of cancer/testis antigen NY-ESO-1 and human granulocyte-macrophage colony-stimulating factor in dendritic cells with a bicistronic retroviral vector.

    PubMed

    Batchu, R B; Moreno, A M; Szmania, S; Gupta, S K; Zhan, F; Rosen, N; Kozlowski, M; Spencer, T; Spagnoli, G C; Shaughnessy, J; Barlogie, B; Tricot, G; van Rhee, F

    2003-09-20

    Tumor-specific genes delivered to dendritic cells (DCs) have been used for the generation of cytotoxic T cells (CTLs), but their application has been limited on the one hand by low viral titers resulting in low transduction efficiency and poor protein production, and on the other hand by immunogenicity of the selectable marker and poor viability of the DCs. We addressed these limitations by creating a multipurpose master vector (pMV) and cloning the tumor gene NY-ESO-1, which is highly expressed in more than 50% of advanced myeloma patients. pMV was constructed from a Moloney murine leukemia virus (Mo-MuLV)-based retroviral backbone with the following features: (1) an extended packaging signal to achieve high viral titers, (2) a splice acceptor region to facilitate protein production, (3) a nonimmunogenic selectable marker, dihydrofolate reductase-L22Y (DHFR(L22Y)), to exclude the generation of CTLs against the selectable marker, (4) an internal ribosomal entry site between the tumor-specific gene (NY-ESO-1) and the selectable marker DHFR(L22Y) for coexpression of two heterologous gene products from a single bicistronic mRNA, minimizing the possibility of differential expression of these two genes, and (5) human granulocyte-macrophage colony-stimulating factor (hGM-CSF) cDNA driven by the human T-lymphotropic virus promoter to enhance DC function and viability. Recombinant virus of pMV-NY-ESO-1 was generated with vesicular stomatitis virus G envelope protein (VSV-G) in the GP2-293 cell line for efficient transduction. We present evidence that the DC phenotype is unaltered after transduction and that more than 85% of DCs express NY-ESO-1, which secrete approximately 40 ng of GM-CSF per 10(6) DCs. PMID:14503968

  6. Characterization of the receptor binding determinants of granulocyte colony stimulating factor.

    PubMed Central

    Young, D. C.; Zhan, H.; Cheng, Q. L.; Hou, J.; Matthews, D. J.

    1997-01-01

    We performed a series of experiments using alanine-scanning mutagenesis to locate side chains within human granulocyte colony-stimulating factor (G-CSF) that are involved in human G-CSF receptor binding. We constructed a panel of 28 alanine mutants that examined all surface exposed residues on helices A and D, as well as all charged residues on the surface of G-CSF. The G-CSF mutants were expressed in a transiently transfected mammalian cell line and quantitated by a sensitive biosensor method. We measured the activity of mutant proteins using an in vitro proliferation assay and an ELISA binding competition assay. These studies show that there is a region of five charged residues on helices A and C employed by G-CSF in binding its receptor, with the most important residue in this binding patch being Glu 19. Both wild-type G-CSF and the E19A mutant were expressed in E. coli. The re-folded proteins were found to have proliferative activities similar to the analogous proteins from mammalian cells: furthermore, biophysical analysis indicated that the E19A mutation does not cause gross structural perturbations in G-CSF. Although G-CSF is likely to signal through receptor homo-dimerization, we found no compelling evidence for a second receptor binding region. We also found no evidence of self-antagonism at high G-CSF concentrations, suggesting that, in contrast to human growth hormone (hGH) and erythropoietin (EPO), G-CSF probably does not signal via a pure 2:1 receptor ligand complex. Thus, G-CSF, while having a similar tertiary structure to hGH and EPO, uses different areas of the four helix bundle for high-affinity interaction with its receptor. PMID:9194183

  7. Expression of the human granulocyte-macrophage colony stimulating factor (hGM-CSF) gene under control of the 5'-regulatory sequence of the goat alpha-S1-casein gene with and without a MAR element in transgenic mice.

    PubMed

    Burkov, I A; Serova, I A; Battulin, N R; Smirnov, A V; Babkin, I V; Andreeva, L E; Dvoryanchikov, G A; Serov, O L

    2013-10-01

    Expression of the human granulocyte-macrophage colony-stimulating factor (hGM-CSF) gene under the control of the 5'-regulatory sequence of the goat alpha-S1-casein gene with and without a matrix attachment region (MAR) element from the Drosophila histone 1 gene was studied in four and eight transgenic mouse lines, respectively. Of the four transgenic lines carrying the transgene without MAR, three had correct tissues-specific expression of the hGM-CSF gene in the mammary gland only and no signs of cell mosaicism. The concentration of hGM-CSF in the milk of transgenic females varied from 1.9 to 14 μg/ml. One line presented hGM-CSF in the blood serum, indicating ectopic expression. The values of secretion of hGM-CSF in milk of 6 transgenic lines carrying the transgene with MAR varied from 0.05 to 0.7 μg/ml, and two of these did not express hGM-CSF. Three of the four examined animals from lines of this group showed ectopic expression of the hGM-CSF gene, as determined by RT-PCR and immunofluorescence analyses, as well as the presence of hGM-CSF in the blood serum. Mosaic expression of the hGM-CSF gene in mammary epithelial cells was specific to all examined transgenic mice carrying the transgene with MAR but was never observed in the transgenic mice without MAR. The mosaic expression was not dependent on transgene copy number. Thus, the expected "protective or enhancer effect" from the MAR element on the hGM-CSF gene expression was not observed.

  8. PyNTTTTGT and CpG immunostimulatory oligonucleotides: effect on granulocyte/monocyte colony-stimulating factor (GM-CSF) secretion by human CD56+ (NK and NKT) cells.

    PubMed

    Rodriguez, Juan M; Marchicio, José; López, Mariela; Ziblat, Andrea; Elias, Fernanda; Fló, Juan; López, Ricardo A; Horn, David; Zorzopulos, Jorge; Montaner, Alejandro D

    2015-01-01

    CD56+ cells have been recognized as being involved in bridging the innate and acquired immune systems. Herein, we assessed the effect of two major classes of immunostimulatory oligonucleotides (ODNs), PyNTTTTGT and CpG, on CD56+ cells. Incubation of human peripheral blood mononuclear cells (hPBMC) with some of these ODNs led to secretion of significant amounts of interferon gamma (IFN-γ), tumor necrosis factor alpha (TNF-α) and granulocyte/monocyte colony-stimulating factor (GM-CSF), but only if interleukin 2 (IL2) was present. IMT504, the prototype of the PyNTTTTGT ODN class, was the most active. GM-CSF secretion was very efficient when non-CpG ODNs with high T content and PyNTTTTGT motifs lacking CpGs were used. On the other hand, CpG ODNs and IFNα inhibited this GM-CSF secretion. Selective cell type removal from hPBMC indicated that CD56+ cells were responsible for GM-CSF secretion and that plasmacytoid dendritic cells (PDCs) regulate this process. In addition, PyNTTTTGT ODNs inhibited the IFNα secretion induced by CpG ODNs in PDCs by interference with the TLR9 signaling pathway. Since IFNα is essential for CD56+ stimulation by CpG ODNs, there is a reciprocal interference of CpG and PyNTTTTGT ODNs when acting on this cell population. This suggests that these synthetic ODNs mimic different natural alarm signals for activation of the immune system. PMID:25706946

  9. PyNTTTTGT and CpG Immunostimulatory Oligonucleotides: Effect on Granulocyte/Monocyte Colony-Stimulating Factor (GM-CSF) Secretion by Human CD56+ (NK and NKT) Cells

    PubMed Central

    Rodriguez, Juan M.; Marchicio, José; López, Mariela; Ziblat, Andrea; Elias, Fernanda; Fló, Juan; López, Ricardo A.; Horn, David; Zorzopulos, Jorge; Montaner, Alejandro D.

    2015-01-01

    CD56+ cells have been recognized as being involved in bridging the innate and acquired immune systems. Herein, we assessed the effect of two major classes of immunostimulatory oligonucleotides (ODNs), PyNTTTTGT and CpG, on CD56+ cells. Incubation of human peripheral blood mononuclear cells (hPBMC) with some of these ODNs led to secretion of significant amounts of interferon gamma (IFN-γ), tumor necrosis factor alpha (TNF-α) and granulocyte/monocyte colony-stimulating factor (GM-CSF), but only if interleukin 2 (IL2) was present. IMT504, the prototype of the PyNTTTTGT ODN class, was the most active. GM-CSF secretion was very efficient when non-CpG ODNs with high T content and PyNTTTTGT motifs lacking CpGs were used. On the other hand, CpG ODNs and IFNα inhibited this GM-CSF secretion. Selective cell type removal from hPBMC indicated that CD56+ cells were responsible for GM-CSF secretion and that plasmacytoid dendritic cells (PDCs) regulate this process. In addition, PyNTTTTGT ODNs inhibited the IFNα secretion induced by CpG ODNs in PDCs by interference with the TLR9 signaling pathway. Since IFNα is essential for CD56+ stimulation by CpG ODNs, there is a reciprocal interference of CpG and PyNTTTTGT ODNs when acting on this cell population. This suggests that these synthetic ODNs mimic different natural alarm signals for activation of the immune system. PMID:25706946

  10. PyNTTTTGT and CpG immunostimulatory oligonucleotides: effect on granulocyte/monocyte colony-stimulating factor (GM-CSF) secretion by human CD56+ (NK and NKT) cells.

    PubMed

    Rodriguez, Juan M; Marchicio, José; López, Mariela; Ziblat, Andrea; Elias, Fernanda; Fló, Juan; López, Ricardo A; Horn, David; Zorzopulos, Jorge; Montaner, Alejandro D

    2015-01-01

    CD56+ cells have been recognized as being involved in bridging the innate and acquired immune systems. Herein, we assessed the effect of two major classes of immunostimulatory oligonucleotides (ODNs), PyNTTTTGT and CpG, on CD56+ cells. Incubation of human peripheral blood mononuclear cells (hPBMC) with some of these ODNs led to secretion of significant amounts of interferon gamma (IFN-γ), tumor necrosis factor alpha (TNF-α) and granulocyte/monocyte colony-stimulating factor (GM-CSF), but only if interleukin 2 (IL2) was present. IMT504, the prototype of the PyNTTTTGT ODN class, was the most active. GM-CSF secretion was very efficient when non-CpG ODNs with high T content and PyNTTTTGT motifs lacking CpGs were used. On the other hand, CpG ODNs and IFNα inhibited this GM-CSF secretion. Selective cell type removal from hPBMC indicated that CD56+ cells were responsible for GM-CSF secretion and that plasmacytoid dendritic cells (PDCs) regulate this process. In addition, PyNTTTTGT ODNs inhibited the IFNα secretion induced by CpG ODNs in PDCs by interference with the TLR9 signaling pathway. Since IFNα is essential for CD56+ stimulation by CpG ODNs, there is a reciprocal interference of CpG and PyNTTTTGT ODNs when acting on this cell population. This suggests that these synthetic ODNs mimic different natural alarm signals for activation of the immune system.

  11. Culture perfusion schedules influence the metabolic activity and granulocyte-macrophage colony-stimulating factor production rates of human bone marrow stromal cells.

    PubMed

    Caldwell, J; Palsson, B O; Locey, B; Emerson, S G

    1991-05-01

    The metabolic function and GM-CSF production rates of adherent human bone marrow stromal cells were investigated as functions of medium and serum feeding rates. A range of medium exchange schedules was studied, ranging from a typical Dexter culture protocol of one weekly medium exchange to a full media exchange daily, which more closely approximates what bone marrow cells experience in situ. Glucose consumption was found to be significantly higher at full daily exchange rate than at any other exchange schedule examined. However, the lactate yield on glucose was a constant, at 1.8 mol/mol, under all conditions considered. Differential serum vs. medium exchange experiment showed that both serum supply and medium nutrients were responsible for the altered behavior at high exchange rates. Glutamine consumption was found to be insignificant under all culture conditions examined. A change in exchange schedule from 50% daily medium exchange to full daily medium exchange after 14 days of culture was found to result in a transient production of GM-CSF and a change in metabolic behavior to resemble that of cultures which had full daily exchange from day one. These results suggest that both stromal cell metabolism and GM-CSF production are sensitive to medium exchange schedules. Taken together, the data presented indicate that attempts to model the function of human bone marrow in vitro may be well served by beginning with medium exchange schedules that more closely mimic the in vivo physiologic state of bone marrow. PMID:2040665

  12. The effects of granulocyte colony-stimulating factor in preclinical models of infection and acute inflammation.

    PubMed

    Marshall, John C

    2005-12-01

    The cytokine granulocyte colony-stimulating factor (G-CSF) is a potent endogenous trigger for the release of neutrophils from bone marrow stores and for their activation for enhanced antimicrobial activity. G-CSF has been widely evaluated in preclinical models of acute illness, with generally promising though divergent results. A recombinant G-CSF molecule has recently undergone clinical trials to assess its efficacy as an adjuvant therapy in community-acquired and nosocomial pneumonia, however, these studies failed to provide convincing evidence of benefit. We undertook a systematic review of the published literature reporting the effects of modulation of G-CSF in preclinical in vivo models to determine whether evidence of differential efficacy might explain the disappointing results of human studies and point to disease states that might be more likely to benefit from G-CSF therapy. G-CSF has been evaluated in 86 such studies involving a variety of different models. The strongest evidence of benefit was seen in studies involving intraperitoneal challenge with live organisms; benefit was evident whether the agent was given before or after challenge. G-CSF demonstrates anti-inflammatory activity in models of systemic challenge with viable organisms or endotoxin, but only when the agent is given before challenge; evidence of benefit after challenge was minimal. Preclinical models of intrapulmonary challenge only show efficacy when the cytokine is administered before the infectious challenge, and suggested harm in gram-negative pneumonia resulting from challenge with Escherichia coli or Klebsiella. There is little evidence for therapeutic efficacy in noninfectious models of acute illness. We conclude that the most promising populations for evaluation of G-CSF are neutropenic patients with invasive infection and patients with intra-abdominal infection, particularly those with the syndrome of tertiary, or recurrent, peritonitis. Significant variability in the design

  13. Increased platelet aggregation and in vivo platelet activation after granulocyte colony-stimulating factor administration. A randomised controlled trial.

    PubMed

    Spiel, Alexander O; Bartko, Johann; Schwameis, Michael; Firbas, Christa; Siller-Matula, Jolanta; Schuetz, Matthias; Weigl, Manuela; Jilma, Bernd

    2011-04-01

    Granulocyte colony-stimulating factor (G-CSF) stimulates the bone marrow to produce granulocytes and stem cells and is widely used to accelerate neutrophil recovery after chemotherapy. Interestingly, specific G-CSF receptors have been demonstrated not only on myeloid cells, but also on platelets. Data on the effects of G-CSF on platelet function are limited and partly conflicting. The objective of this study was to determine the effect of G-CSF on platelet aggregation and in vivo platelet activation. Seventy-eight, healthy volunteers were enrolled into this randomised, placebo-controlled trial. Subjects received 5 μg/kg methionyl human granulocyte colony-stimulating factor (r-metHuG-CSF, filgrastim) or placebo subcutaneously for four days. We determined platelet aggregation with a whole blood impedance aggregometer with various, clinically relevant platelet agonists (adenosine diphosphate [ADP], collagen, arachidonic acid [AA], ristocetin and thrombin receptor activating peptide 6 [TRAP]). Filgrastim injection significantly enhanced ADP (+40%), collagen (+60%) and AA (+75%)-induced platelet aggregation (all p<0.01 as compared to placebo and p<0.001 as compared to baseline). In addition, G-CSF enhanced ristocetin-induced platelet aggregation (+18%) whereas TRAP-induced platelet aggregation decreased slightly (-14%) in response to filgrastim. While baseline aggregation with all agonists was only slightly but insignificantly higher in women than in men, this sex difference was enhanced by G-CSF treatment, and became most pronounced for ADP after five days (p<0.001). Enhanced platelet aggregation translated into a 75% increase in platelet activation as measured by circulating soluble P-selectin. G-CSF enhances platelet aggregation and activation in humans. This may put patients suffering from cardiovascular disease and cancer at risk for thrombotic events. PMID:21301783

  14. Autopsy of anaplastic carcinoma of the pancreas producing granulocyte colony-stimulating factor.

    PubMed

    Hayashi, Haruna; Eguchi, Noriaki; Sumimoto, Kyoku; Matsumoto, Kenta; Azakami, Takahiro; Sumida, Tomonori; Tamura, Tadamasa; Sumii, Masaharu; Uraoka, Naohiro; Shimamoto, Fumio

    2016-08-01

    A 50-year-old man presented to a nearby hospital with high fever and anorexia. An abdominal tumor was detected, and he was referred to our hospital. A pancreatic tumor was detected by computed tomography and abdominal ultrasonography. He had high fever, leukocytosis, and high serum granulocyte colony-stimulating factor (G-CSF). We performed a tumor biopsy and histological examination revealed anaplastic carcinoma of the pancreas. Based on the diagnosis, we initiated chemotherapy using gemcitabine plus S-1. However, the tumor rapidly progressed and he deteriorated and died 123 days after admission. As immunohistochemical study showed positive staining for G-CSF in the tumor cell, we diagnosed the tumor producing G-CSF during autopsy. Anaplastic carcinoma of the pancreas producing G-CSF is very rare, with 10 cases, including ours, reported in the literature. PMID:27498938

  15. Human macrophage colony-stimulating factor induces macrophage colonies after L-phenylalanine methylester treatment of human marrow.

    PubMed

    Rosenfeld, C S; Evans, C; Shadduck, R K

    1990-11-01

    Macrophage-colony stimulating factor (M-CSF) has well-known effects on murine bone marrow, but its colony stimulating activity for human bone marrow is controversial. After treatment of human bone marrow with L-phenylalanine methylester (PME), macrophage-colonies (CFU-M) were induced by M-CSF in a dose-dependent fashion. The optimal concentration of recombinant human-macrophage colony stimulating factor (rhM-CSF) was 1,000 U/mL. Purified human urine M-CSF had colony stimulating activity similar to rhM-CSF. Further studies were performed to determine the factors responsible for the enhanced CFU-M formation from PME treated marrow. Compared with nylon wool and carbonyl iron monocyte depletion methods, PME eliminated significantly more monocytes and myeloid cells. This observation suggested that these cells may release hematopoietic inhibitory factors for CFU-M. Low concentrations (1%) but not normal (10%) concentrations of blood monocytes were inhibitory (mean inhibition, 48%) to CFU-M. High concentrations of monocytes (50%) augmented CFU-M colonies. HL-60 conditioned media was used to simulate secretory products of early myeloid cells. HL-60 conditioned media (1%) inhibited CFU-M formation but not granulocyte macrophage or granulocyte colonies. We conclude that M-CSF has colony stimulating activity for human marrow that can be recognized after removal of inhibitory cells by PME treatment. PMID:2224128

  16. Endothelin receptor B protects granulocyte macrophage colony-stimulating factor mRNA from degradation.

    PubMed

    Jungck, David; Knobloch, Jürgen; Körber, Sandra; Lin, Yingfeng; Konradi, Jürgen; Yanik, Sarah; Stoelben, Erich; Koch, Andrea

    2015-06-01

    Evidence is lacking on the differential effects of the two therapeutic concepts of endothelin receptor antagonists (ERAs): the blockade of only the endothelin receptor A (ETAR; selective antagonism) versus both ETAR and endothelin receptor B (ETBR; dual blockade). Ambrisentan, a selective ERA, and bosentan, a dual blocker, are both available for therapy. We hypothesized that there are differences in the potential of ERAs to ameliorate inflammatory processes in human airway smooth muscle cells (HASMCs) and aimed to unravel underlying mechanisms. We used HASMC culture, enzyme-linked immunosorbent assay, and quantitative reverse-transcription polymerase chain reaction. Tumor necrosis factor α (TNFα) induced transcription and expression of chemokine (C-X-C motif) ligand 2 (CXCL2), chemokine (C-X-C motif) ligand 3 (CXCL3), granulocyte macrophage colony-stimulating factor (GM-CSF), and matrix metalloproteinase 12 (MMP12) in HASMCs. In concentration-response experiments, bosentan led to a significantly greater reduction of GM-CSF and MMP12 protein release than ambrisentan, whereas there was no significant difference in their effect on GM-CSF and MMP12 mRNA. Both ERAs reduced CXCL3 protein and mRNA equally but had no effect on CXCL2. Blocking mitogen-activated protein kinases revealed that both ETAR and ETBR signal through p38 mitogen-activated protein kinase, but ETBR also signals through extracellular signal-regulated kinase (ERK) 1/2 to induce GM-CSF expression. In the presence of the transcription inhibitor actinomycin D, bosentan, but not ambrisentan, reduced GM-CSF but not MMP12 or CXCL3 mRNA. In conclusion, blockade of each endothelin receptor subtype reduces GM-CSF transcription, but blocking ETBR additionally protects GM-CSF mRNA from degradation via ERK-1/2. Accordingly, blocking both ETAR and ETBR leads to a stronger reduction of TNFα-induced GM-CSF protein expression. This mechanism might be specific to GM-CSF. Our data stress the anti-inflammatory potential

  17. Regulation of Wound Healing by Granulocyte-Macrophage Colony-Stimulating Factor after Vocal Fold Injury

    PubMed Central

    Lim, Jae-Yol; Choi, Byung Hyune; Lee, Songyi; Jang, Yun Ho; Choi, Jeong-Seok; Kim, Young-Mo

    2013-01-01

    Objectives Vocal fold (VF) scarring remains a therapeutic challenge. Granulocyte-macrophage colony-stimulating factor (GM-CSF) facilitates epithelial wound healing, and recently, growth factor therapy has been applied to promote tissue repair. This study was undertaken to investigate the effect of GM-CSF on VF wound healing in vivo and in vitro. Methods VF scarring was induced in New Zealand white rabbits by direct injury. Immediately thereafter, either GM-CSF or PBS was injected into the VFs of rabbits. Endoscopic, histopathological, immunohistochemical, and biomechanical evaluations of VFs were performed at 3 months post-injury. Human vocal fold fibroblasts (hVFFs) were cultured with GM-CSF. Production of type I and III collagen was examined immunocytochemically, and the synthesis of elastin and hyaluronic acids was evaluated by ELISA. The mRNA levels of genes related to ECM components and ECM production-related growth factors, such as HGF and TGF-ß1, were examined by real time RT-PCR. Results The GM-CSF-treated VFs showed reduced collagen deposition in comparison to the PBS-injected controls (P<0.05). Immunohistochemical staining revealed lower amounts of type I collagen and fibronectin in the GM-CSF-treated VFs (P<0.05 and P<0.01, respectively). Viscous and elastic shear moduli of VF samples were significantly lower in the GM-CSF group than in the PBS-injected group (P<0.001 and P<0.01, respectively). Mucosal waves in the GM-CSF group showed significant improvement when compared to the PBS group (P = 0.0446). GM-CSF inhibited TGF-β1-induced collagen synthesis by hVFFs (P<0.05) and the production of hyaluronic acids increased at 72 hours post-treatment (P<0.05). The expressions of HAS-2, tropoelastin, MMP-1, HGF, and c-Met mRNA were significantly increased by GM-CSF, although at different time points (P<0.05). Conclusion The present study shows that GM-CSF offers therapeutic potential for the remodeling of VF wounds and the promotion of VF regeneration

  18. Differential regulation of early response genes and cell proliferation through the human granulocyte macrophage colony-stimulating factor receptor: selective activation of the c-fos promoter by genistein.

    PubMed Central

    Watanabe, S; Muto, A; Yokota, T; Miyajima, A; Arai, K

    1993-01-01

    Granulocyte macrophage colony-stimulating factor (GM-CSF) binds to the high-affinity GM-CSF receptor (GMR) consisting of alpha and beta subunits and induces rapid tyrosine phosphorylation, activation of early response genes, and proliferation of hematopoietic cells. The alpha subunit is the primary cytokine binding component and the beta subunit is required for high-affinity binding as well as for signal transduction. Using tyrosine kinase inhibitors and cytoplasmic deletion mutants of the beta subunit, we obtained evidence that there are at least two distinct pathways downstream of the GMR in BA/F3 cell, one which is essential for proliferation, leads to the c-myc gene activation, and is sensitive to herbimycin and genistein. Activation of this pathway depends on the cytoplasmic region between amino acid positions 455 and 517 of the beta subunit. The second pathway, which leads to activation of c-fos and c-jun genes, is only partially sensitive to herbimycin, is resistant to genistein and depends on the region between amino acid positions 626 and 763 of the beta subunit. Unexpectedly, the c-fos mRNA induction was augmented by genistein. The enhanced expression of c-fos mRNA by genistein also occurred with stimulation with cAMP, PMA, or EGF in NIH3T3 cells. It thus seems likely that genistein affects a common pathway downstream of these signals. Images PMID:8298195

  19. Regulatory elements responsible for inducible expression of the granulocyte colony-stimulating factor gene in macrophages.

    PubMed Central

    Nishizawa, M; Nagata, S

    1990-01-01

    Granulocyte colony-stimulating factor (G-CSF) plays an essential role in granulopoiesis during bacterial infection. Macrophages produce G-CSF in response to bacterial endotoxins such as lipopolysaccharide (LPS). To elucidate the mechanism of the induction of G-CSF gene in macrophages or macrophage-monocytes, we have examined regulatory cis elements in the promoter of mouse G-CSF gene. Analyses of linker-scanning and internal deletion mutants of the G-CSF promoter by the chloramphenicol acetyltransferase assay have indicated that at least three regulatory elements are indispensable for the LPS-induced expression of the G-CSF gene in macrophages. When one of the three elements was reiterated and placed upstream of the TATA box of the G-CSF promoter, it mediated inducibility as a tissue-specific and orientation-independent enhancer. Although this element contains a conserved NF-kappa B-like binding site, the gel retardation assay and DNA footprint analysis with nuclear extracts from macrophage cell lines demonstrated that nuclear proteins bind to the DNA sequence downstream of the NF-kappa B-like element, but not to the conserved element itself. The DNA sequence of the binding site was found to have some similarities to the LPS-responsive element which was recently identified in the promoter of the mouse class II major histocompatibility gene. Images PMID:1691438

  20. Granulocyte colony-stimulating factor-producing hepatocellular carcinoma with abrupt changes

    PubMed Central

    Nagata, Hiroaki; Komatsu, Shuhei; Takaki, Wataru; Okayama, Tokunari; Sawabe, Yasunori; Ishii, Michiaki; Kishimoto, Mitsuo; Otsuji, Eigo; Konosu, Hiroshi

    2016-01-01

    Granulocyte colony-stimulating factor (G-CSF)-producing tumor is one of the rare types of cancer clinically characterized by an elevated fever and white blood cell (WBC) increment. Although G-CSF producing tumors have been reported in several types of cancer including those of the lungs, cervix and bladder, G-CSF producing hepatocellular carcinoma is extremely rare. Here, we report the case of a rapidly growing and poorly differentiated hepatocellular carcinoma producing G-CSF. The patient showed symptoms of continuous high fever, stomach pain and cough, and high serum WBC counts, C-reactive protein (CRP) and G-CSF levels were found in laboratory tests. After a radical hepatectomy, the patient completely recovered from the above symptoms and inflammatory state. The serum levels of G-CSF were reduced to normal levels after radical surgery. An immunohistochemical analysis revealed the overexpression of G-CSF in the cytoplasm of certain hepatocellular carcinoma (HCC) cell. The patient’s serum WBC, CRP and G-CSF levels remained within normal levels in the six months after surgery without recurrence. This is the 9th case report of G-CSF producing hepatocellular carcinoma in English literature. We review the clinical characteristics of the G-CSF producing HCC and discuss a possible treatment strategy. PMID:27777880

  1. Granulocyte/macrophage colony-stimulating factor attenuates endothelial hyperpermeability after thermal injury.

    PubMed

    Zhao, Jingling; Chen, Lei; Shu, Bin; Tang, Jinming; Zhang, Lijun; Xie, Julin; Liu, Xusheng; Xu, Yingbin; Qi, Shaohai

    2015-01-01

    Microvascular hyperpermeability followed by burn injury is the main cause of shock, and cardiovascular collapse can result if the condition is treated improperly. Our previous studies demonstrated that granulocyte/macrophage colony-stimulating factor (GM-CSF) clearly reduces microvascular permeability and protects microvessels against burn injury. However, the mechanism underlying the protective function of GM-CSF on burn-injured microvessels remains unknown. This study aimed to investigate the effect and mechanism of GM-CSF on endothelial cells after exposure to burn serum. We demonstrated that GM-CSF reduced post-burn endothelial "capillary leak" by inhibiting the activity of RhoA and maintaining the membrane localization of VE-cadherin. Membranous VE-cadherin enhances adherens junctions between endothelial cells and co-localizes with and activates VEGFR2, which protect cells from burn serum-induced apoptosis. Our findings suggest that the protective mechanism of GM-CSF on burn serum-injured endothelial monolayer hyperpermeability is achieved by strengthening cell adherens junctions and improving cell viability.

  2. Granulocyte-colony stimulating factor therapy to induce neovascularization in ischemic heart disease.

    PubMed

    Ripa, Rasmus Sejersten

    2012-03-01

    Cell based therapy for ischemic heart disease has the potential to reduce post infarct heart failure and chronic ischemia. Treatment with granulocyte-colony stimulating factor (G-CSF) mobilizes cells from the bone marrow to the peripheral blood. Some of these cells are putative stem or progenitor cells. G-CSF is injected subcutaneously. This therapy is intuitively attractive compared to other cell based techniques since repeated catheterizations and ex vivo cell purification and expansion are avoided. Previous preclinical and early clinical trials have indicated that treatment with G-CSF leads to improved myocardial perfusion and function in acute or chronic ischemic heart disease. The hypothesis of this thesis is that patient with ischemic heart disease will benefit from G-CSF therapy. We examined this hypothesis in two clinical trials with G-CSF treatment to patients with either acute myocardial infarction or severe chronic ischemic heart disease. In addition, we assed a number of factors that could potentially affect the effect of cell based therapy. Finally, we intended to develop a method for in vivo cell tracking in the heart. Our research showed that subcutaneous G-CSF along with gene therapy do not improve myocardial function in patients with chronic ischemia despite a large increase in circulation bone marrow-derived cells. Also, neither angina pectoris nor exercise capacity was improved compared to placebo treatment. We could not identify differences in angiogenic factors or bone marrow-derived cells in the blood that could explain the neutral effect of G-CSF. Next, we examined G-CSF as adjunctive therapy following ST segment elevation myocardial infarction. We did not find any effect of G-CSF neither on the primary endpoint--regional myocardial function--nor on left ventricular ejection fraction (secondary endpoint) compared to placebo treatment. In subsequent analyses, we found significant differences in the types of cells mobilized from the bone marrow

  3. Effect of in vivo infusion of granulocyte colony-stimulating factor on immune function.

    PubMed

    Valente, John F; Alexander, J Wesley; Li, Bing-Guo; Noel, J Gregory; Custer, David A; Ogle, James D; Ogle, Cora K

    2002-01-01

    As the applications of hematopoietic growth factors increase, their complex impact on host defense and immune responses continues to unfold. The effect of the administration of granulocyte colony-stimulating factor (G-CSF) on bacterial defense, proliferation of lymphocytes, and cytokine production by lymphocytes and peripheral blood mononuclear cells (PBMC) was studied. The effect of G-CSF administration on the phenotype of the cells in the major hematopoietic organs was studied as well. ACI rats were given 10 mg/kg/day G-CSF or vehicle daily for 4 days. Isolated bone marrow neutrophils and enterocytes from treated animals showed a greater bactericidal activity than controls. Proliferation of mitogen-stimulated lymphocytes and PBMC was reduced in G-CSF-treated animals. The production of proinflammatory cytokines, tumor necrosis factor (TNF), and interleukin 6 (IL-6) by lymphocytes and PBMC was reduced by G-CSF pretreatment. G-CSF administration caused an increase in IL-4 (Th2 cytokine) release and a decrease in interferon-gamma (IFNgamma, Th1 cytokine) release by mitogen-stimulated lymphocytes. Cytometric analysis of cells in the progenitor cell region indicated a large increase in immature cells in the bone marrow of G-CSF-treated animals compared with sham along with an increase in B cells and a decrease in polymorphonuclear leukocytes (PMNs). In addition, cytometric analysis showed a large increase in PMNs in blood and splenocytes of the treated animals compared with sham. This study confirms and extends previous observations that G-CSF administration has a number of effects that might simultaneously enhance host defense while reducing the risk of developing uncontrolled systemic inflammation. This may also be efficacious in prolonging graft survival and reducing graft vs. host disease.

  4. Granulocyte colony-stimulating factor in repeated IVF failure, a randomized trial.

    PubMed

    Aleyasin, Ashraf; Abediasl, Zhila; Nazari, Atefeh; Sheikh, Mahdi

    2016-06-01

    Recent studies have revealed key roles for granulocyte colony-stimulating factor (GCSF) in embryo implantation process and maintenance of pregnancy, and some studies showed promising results by using local intrauterine infusion of GCSF in patients undergoing in vitro fertilization (IVF). This multicenter, randomized, controlled trial included 112 infertile women with repeated IVF failure to evaluate the efficacy of systemic single-dose subcutaneous GCSF administration on IVF success in these women. In this study, the Long Protocol of ovarian stimulation was used for all participants. Sealed, numbered envelopes assigned 56 patients to receive subcutaneous 300 µg GCSF before implantation and 56 in the control group. The implantation (number of gestational sacs on the total number of transferred embryos), chemical pregnancy (positive serum β-HCG), and clinical pregnancy (gestational sac and fetal heart) rates were compared between the two groups. This trial is registered at www.irct.ir (IRCT201503119568N11). The successful implantation (18% vs 7.2%, P=0.007), chemical pregnancy (44.6% vs 19.6%, P=0.005), and clinical pregnancy (37.5% vs 14.3%, P=0.005) rates were significantly higher in the intervention group than in the control group. After adjustment for participants' age, endometrial thickness, good-quality oocyte counts, number of transferred embryos, and anti-Mullerian hormone levels, GCSF treatment remained significantly associated with successful implantation (OR=2.63, 95% CI=1.09-6.96), having chemical pregnancy (OR= 2.74, 95% CI=1.11-7.38) and clinical pregnancy (OR=2.94, 95% CI=1.23-8.33). In conclusion, administration of single-dose systemic subcutaneous GCSF before implantation significantly increases the IVF success, implantation, and pregnancy rates in infertile women with repeated IVF failure. PMID:26980809

  5. Use of granulocyte colony-stimulating factor: a survey among Italian medical oncologists.

    PubMed

    Danova, Marco; Rosti, Giovanni; De Placido, Sabino; Bencardino, Katia; Venturini, Marco

    2005-12-01

    In October 2003, the Italian Association of Medical Oncology (AIOM) published its own guidelines on the use of granulocyte colony-stimulating factor (G-CSF). The present survey was conducted during the same period with the aim of collecting data on the current use of G-CSF to provide a starting point for future evaluations of the implementation of AIOM guidelines. From October 2003 to January 2004, 1591 AIOM members were asked to complete a questionnaire based on specific clinical scenarios, regarding the use of G-CSF for primary and secondary prophylaxis and treatment of neutropenia. The rate of response was 22%. For primary prophylaxis, the majority of physicians avoid using G-CSF, with no difference in cases of adjuvant, curative or palliative chemotherapy (CT). In fact, 67.2% to 74.9% would 'rarely or never' use G-CSF in the proposed clinical scenarios. In chemosensitive tumors, rather than reducing CT doses, 55.7% would use G-CSF as a secondary prophylaxis after afebrile neutropenia (AN), and 68.8% after febrile neutropenia (FN). In elderly patients experiencing FN, 35.7% would reduce the adjuvant CT doses and 23.1% would change the regimen. Most oncologists would use G-CSF to treat neutropenia, and the median duration of G-CSF treatment is less than 1 week and would depend on neutrophil count. Our survey shows that Italian oncologists are particularly oriented towards the use of G-CSF in clinical practice to maintain the CT dose intensity, and are sensitive to the prevention and treatment of not only FN, but also AN. Finally, Italian medical oncologists appear to be very cautious in introducing G-CSF when treating elderly patients. PMID:16273232

  6. Granulocyte-macrophage colony-stimulating factor primes interleukin-13 production by macrophages via protease-activated receptor-2.

    PubMed

    Aoki, Manabu; Yamaguchi, Rui; Yamamoto, Takatoshi; Ishimaru, Yasuji; Ono, Tomomichi; Sakamoto, Arisa; Narahara, Shinji; Sugiuchi, Hiroyuki; Hirose, Eiji; Yamaguchi, Yasuo

    2015-04-01

    Chronic inflammation is often linked to the presence of type 2-polarized macrophages, which are induced by the T helper type 2 cytokines interleukin-4 and interleukin-13 (IL-13). IL-13 is a key mediator of tissue fibrosis caused by T helper type 2-based inflammation. Human neutrophil elastase (HNE) plays a pivotal role in the pathogenesis of pulmonary fibrosis. This study investigated the priming effect of granulocyte-macrophage colony-stimulating factor (GM-CSF) on IL-13 expression by macrophages stimulated with HNE. Adherent macrophages were obtained from primary cultures of human mononuclear cells. Expression of IL-13 mRNA and protein by GM-CSF-dependent macrophages was investigated after stimulation with HNE, using the polymerase chain reaction and enzyme-linked immunosorbent assay. GM-CSF had a priming effect on IL-13 mRNA and protein expression by macrophages stimulated with HNE, while this effect was not observed for various other cytokines. GM-CSF-dependent macrophages showed a significant increase in the expression of protease activated receptor-2 (PAR-2) mRNA and protein. The response of IL-13 mRNA to HNE was significantly decreased by pretreatment with alpha1-antitrypsin, a PAR-2 antibody (SAM11), or a PAR-2 antagonist (ENMD-1068). These findings suggest that stimulation with HNE can induce IL-13 production by macrophages, especially GM-CSF-dependent macrophages. Accordingly, neutrophil elastase may have a key role in fibrosis associated with chronic inflammation.

  7. A randomized, placebo-controlled trial of granulocyte-macrophage colony-stimulating factor and nucleoside analogue therapy in AIDS.

    PubMed

    Brites, C; Gilbert, M J; Pedral-Sampaio, D; Bahia, F; Pedroso, C; Alcantara, A P; Sasaki, M D; Matos, J; Renjifo, B; Essex, M; Whitmore, J B; Agosti, J M; Badaro, R

    2000-11-01

    Preliminary preclinical and clinical data suggest that granulocyte-macrophage colony-stimulating factor (GM-CSF) may decrease viral replication. Therefore, 105 individuals with AIDS who were receiving nucleoside analogue therapy were enrolled in a placebo-controlled, double-blind study and were randomized to receive either 125 microgram/m(2) of yeast-derived, GM-CSF (sargramostim) or placebo subcutaneously twice weekly for 6 months. Subjects were evaluated for toxicity and disease progression. A significant decrease in mean virus load (VL) was observed for the GM-CSF treatment group at 6 months (-0.07 log(10) vs. -0.60 log(10); P=.02). More subjects achieved human immunodeficiency virus (HIV)-RNA levels <500 copies/mL at >/=2 evaluations (2% on placebo vs. 11% on GM-CSF; P=.04). Genotypic analysis of 46 subjects demonstrated a lower frequency of zidovudine-resistant mutations among those receiving GM-CSF (80% vs. 50%; P=.04). No difference was observed in the incidence of opportunistic infections (OIs) through 6 months or survival, despite a higher risk for OI among GM-CSF recipients. GM-CSF reduced VL and limited the evolution of zidovudine-resistant genotypes, potentially providing adjunctive therapy in HIV disease.

  8. Granulocyte Macrophage-Colony Stimulating Factor-induced Zn Sequestration Enhances Macrophage Superoxide and Limits Intracellular Pathogen Survival

    PubMed Central

    Vignesh, Kavitha Subramanian; Landero Figueroa, Julio A.; Porollo, Aleksey; Caruso, Joseph A.; Deepe, George S.

    2013-01-01

    SUMMARY Macrophages possess numerous mechanisms to combat microbial invasion, including sequestration of essential nutrients, like Zn. The pleiotropic cytokine granulocyte macrophage-colony stimulating factor (GM-CSF) enhances antimicrobial defenses against intracellular pathogens such as Histoplasma capsulatum, but its mode of action remains elusive. We have found that GM-CSF activated infected macrophages sequestered labile Zn by inducing binding to metallothioneins (MTs) in a STAT3 and STAT5 transcription factor-dependent manner. GM-CSF upregulated expression of Zn exporters, Slc30a4 and Slc30a7 and the metal was shuttled away from phagosomes and into the Golgi apparatus. This distinctive Zn sequestration strategy elevated phagosomal H+ channel function and triggered reactive oxygen species (ROS) generation by NADPH oxidase. Consequently, H. capsulatum was selectively deprived of Zn, thereby halting replication and fostering fungal clearance. GM-CSF mediated Zn sequestration via MTs in vitro and in vivo in mice and in human macrophages. These findings illuminate a GM-CSF-induced Zn-sequestration network that drives phagocyte antimicrobial effector function. PMID:24138881

  9. Recombinant hybrid protein, Shiga toxin and granulocyte macrophage colony stimulating factor effectively induce apoptosis of colon cancer cells

    PubMed Central

    Roudkenar, Mehryar Habibi; Bouzari, Saeid; Kuwahara, Yoshikazu; Roushandeh, Amaneh Mohammadi; Oloomi, Mana; Fukumoto, Manabu

    2006-01-01

    AIM: To investigate the selective cytotoxic effect of constructed hybrid protein on cells expressing granulocyte macrophage colony stimulating factor (GM-CSF) receptor. METHODS: HepG2 (human hepatoma) and LS174T (colon carcinoma) were used in this study. The fused gene was induced with 0.02 % of arabinose for 4 h and the expressed protein was detected by Western blotting. The chimeric protein expressed in E.coli was checked for its cytotoxic activity on these cells and apoptosis was measured by comet assay and nuclear staining. RESULTS: The chimeric protein was found to be cytotoxic to the colon cancer cell line expressing GM-CSFRs, but not to HepG2 lacking these receptors. Maximum activity was observed at the concentration of 40 ng/mL after 24 h incubation. The IC50 was 20  ±  3.5 ng/mL. CONCLUSION: Selective cytotoxic effect of the hybrid protein on the colon cancer cell line expressing GM-CSF receptors (GM-CSFRs) receptor and apoptosis can be observed in this cell line. The hybrid protein can be considered as a therapeutic agent. PMID:16688822

  10. Granulocyte-Colony Stimulating Factor (G-CSF) Improves Motor Recovery in the Rat Impactor Model for Spinal Cord Injury

    PubMed Central

    Dittgen, Tanjew; Pitzer, Claudia; Plaas, Christian; Kirsch, Friederike; Vogt, Gerhard; Laage, Rico; Schneider, Armin

    2012-01-01

    Granulocyte-colony stimulating factor (G-CSF) improves outcome after experimental SCI by counteracting apoptosis, and enhancing connectivity in the injured spinal cord. Previously we have employed the mouse hemisection SCI model and studied motor function after subcutaneous or transgenic delivery of the protein. To further broaden confidence in animal efficacy data we sought to determine efficacy in a different model and a different species. Here we investigated the effects of G-CSF in Wistar rats using the New York University Impactor. In this model, corroborating our previous data, rats treated subcutaneously with G-CSF over 2 weeks show significant improvement of motor function. PMID:22253813

  11. Molecular cloning, sequencing and structural studies of granulocyte-macrophage colony-stimulating factor (GM-CSF) from Indian water buffalo (Bubalus bubalis).

    PubMed

    Sugumar, Thennarasu; Pugalenthi, Ganesan; Harishankar, Murugesan; Dhinakar Raj, G

    2014-02-01

    Granulocyte-macrophage colony-stimulating factor (GM-CSF) is a cytokine that is essential for growth and development of progenitors of granulocytes and monocytes/macrophages. In this study, we report molecular cloning, sequencing and characterization of GM-CSF from Indian water buffalo, Bubalus bubalis. In addition, we performed sequence and structural analysis for buffalo GM-CSF. Buffalo GM-CSF has been compared with 17 mammalian GM-CSFs using multiple sequence alignment and phylogenetic tree. Three-dimensional model for buffalo GM-CSF and human receptor complex was built using homology modelling to study cross-reactivity between two species. Detailed analysis was performed to study GM-CSF interface and various interactions at the interface.

  12. The efficiency of granulocyte colony-stimulating factor in hemorrhagic mucositis and febrile neutropenia resulted from methotrexate toxicity.

    PubMed

    Ozkol, Hatice Uce; Toptas, Tayfur; Calka, Omer; Akdeniz, Necmettin

    2015-01-01

    Methotrexate (MTX) remains one of the most frequently used anti-metabolite agents in dermatology. MTX is an analog of folate that competitively and irreversibly inhibits dihydrofolate reductase. Oral mucositis is a common side effect of chemotherapy drugs and is characterized by erythema, pain, poor oral intake, pseudomembranous destruction, open ulceration and hemorrhage of the oral mucosa. In this paper, we report a 32-year-old female with a case of mucositis due to MTX intoxication that resulted from an overdose for rheumatoid arthritis. The patient had abdominal pain, vomiting, and nausea. During follow-up, the patient's white blood cell count was found to be 0.9 × 10(9)/L (4-10 × 10(9)/L). The patient developed fever exceeding 40 °C. The patient was consulted to the hematology service. They suggested using granulocyte colony-stimulating factor for febrile neutropenia. On the fifth day of treatment, the white blood cell count reached 5.3 × 10(9)/L and the patient's fever and mucositis started to resolve. Here, we presented a case of hemorrhagic mucositis and febrile neutropenia resulted from high-dose MTX that responded very well to granulocyte colony-stimulating factor treatment and we reviewed the literature.

  13. Bone pain from granulocyte colony stimulating factor: does clinical trial sponsorship by a pharmaceutical company influence its reporting?

    PubMed

    Aldairy, Y; Nguyen, P L; Jatoi, A

    2011-01-01

    It is alleged that pharmaceutical companies sometimes unfairly present clinical trial results. To our knowledge, studies have not explored whether such alleged unfair reporting also occurs in the testing of palliative care agents in cancer patients, a particularly vulnerable group. Therefore, a systematic search was conducted to retrieve all published, prospective clinical trials that used granulocyte colony stimulating factor starting in 2003. Because granulocyte colony stimulating factor can cause severe bone pain - a concerning but historically under-reported symptom in cancer patients - this symptom was assessed to determine whether differences in reporting occurred based on pharmaceutical company-sponsorship. A total of 239 published clinical trials met the present study's eligibility criteria and were retrievable. Within this entire group of studies, 65 (27%) were pharmaceutical company-sponsored, and only 31 (13%) reported on bone pain. However, pharmaceutical company-sponsored trials reported on bone pain at a higher rate compared with other studies: 23% versus 9% (P= 0.005), and this conclusion did not change after adjusting for dose, use of the slow release formulation and year of publication. The reporting of adverse events from cancer symptom control and palliative care interventions should be improved - especially in trials not sponsored by pharmaceutical companies.

  14. A bioassay system for two types of colony stimulating factor in human serum.

    PubMed

    Francis, G E

    1980-07-01

    The biological assay of factors with granulocyte-macrophage colony stimulating activity in human serum poses special problems. Assays based on colony counts suggest, often erroneously, that serum lacks colony stimulating factors (CSF) but interacts with other materials to potentiate or produce factors. Although the scoring of total clone numbers reveals the presence of CSF in serum, this is still no sufficient for the interpretation of the effects of mixing materials containing CSF, because increments in clone numbers are not directly proportional to increments in CSF. This is particularly important in serum assays because two types of activity are present, one which stimulates progenitor cells directly, and another which results from the interation of serum and bone marrow adherent CSF-producing) cells or peripheral blood leucocytes, indicating the presence of both direct acting and "adherent cell dependent" CSF. A biological assay is described which uses analysis of dose-response curves of clone formation in agar culture, and allows simultaneous assay of both types of activity. The criteria for the selection of suitable target progenitor cell populations are discussed.

  15. Differential induction of bone marrow macrophage proliferation by mycoplasmas involves granulocyte-macrophage colony-stimulating factor.

    PubMed Central

    Stuart, P M; Cassell, G H; Woodward, J G

    1990-01-01

    We have studied the ability of three different Mycoplasma species to induce proliferation of bone marrow-derived macrophages (BMM). We observed a significant mitogenic effect when BMM cells from BALB/c, DBA/2J, SJL, and C57BL/6 mice were incubated with membranes derived from Mycoplasma arginini or M. arthritidis but not when they were incubated with an equivalent amount of M. pulmonis membrane. We also determined that pretreatment of mycoplasma membrane preparations with papain eliminated the ability of these preparations to induce BMM proliferation. To determine whether these membrane fractions acted indirectly by stimulating the production of soluble factors known to stimulate proliferation of BMM cells, we performed blocking studies with antibodies directed against colony-stimulating factor 1 (CSF-1), interleukin-3 (IL-3), and granulocyte-macrophage colony-stimulating factor. Our results indicate that antibodies directed against either CSF-1 or IL-3 failed to block mycoplasma-initiated proliferation of BMM cells. However, when anti-GM-CSF was added to proliferative cultures at the time of initiation, we saw a dose-dependent reduction of mycoplasma-initiated proliferation. We conclude that the ability of mycoplasma membranes to initiate the proliferation of BMM is not shared by all species of mycoplasma and that it involves the production of GM-CSF by an as yet undetermined cell. PMID:2228227

  16. Giant Cell Arteritis which Developed after the Administration of Granulocyte-colony Stimulating Factor for Cyclic Neutropenia.

    PubMed

    Umeda, Masataka; Ikenaga, Jin; Koga, Tomohiro; Michitsuji, Toru; Shimizu, Toshimasa; Fukui, Shoichi; Nishino, Ayako; Nakasima, Yoshikazu; Kawashiri, Sin-Ya; Iwamoto, Naoki; Ichinose, Kunihiro; Hirai, Yasuko; Tamai, Mami; Nakamura, Hideki; Origuchi, Tomoki; Kawakami, Atsushi

    2016-01-01

    A 78-year-old woman diagnosed with cyclic neutropenia 5 years previously had been treated with recombinant granulocyte-colony stimulating factor (G-CSF). She developed fever, tenderness and distension of temporal arteries after the treatment with G-CSF. Magnetic resonance imaging and ultrasonography revealed wall thickening of the temporal arteries. She was therefore diagnosed with giant cell arteritis (GCA). Small vessel vasculitis has been reported as a complication of G-CSF. However, the development of large vessel vasculitis after G-CSF treatment is quite rare. To our knowledge, the present case is the first report of GCA suspected to be associated with coexisting cyclic neutropenia and G-CSF treatment. PMID:27523011

  17. A 3,387 bp 5'-flanking sequence of the goat alpha-S1-casein gene provides correct tissue-specific expression of human granulocyte colony-stimulating factor (hG-CSF) in the mammary gland of transgenic mice.

    PubMed

    Serova, Irina A; Dvoryanchikov, Gennady A; Andreeva, Ludmila E; Burkov, Ivan A; Dias, Luciene P B; Battulin, Nariman R; Smirnov, Alexander V; Serov, Oleg L

    2012-06-01

    A new expression vector containing the 1,944 bp 5'-flanking regulatory region together with exon 1 and intron 1 of the goat alpha-S1-casein gene (CSN1S1), the full-sized human granulocyte colony-stimulating factor gene (hGCSF) and the 3'-flanking sequence of the bovine CSN1S1, was created. The vector DNA was used for generation of four mouse transgenic lines. The transgene was integrated into chromosomes 8 and 12 of two founders as 2 and 5 copies, respectively. Tissue-specific secretion of hG-CSF into the milk of transgenic mice was in the range of 19-40 μg/ml. RT-PCR analysis of various tissues of the transgenic mice demonstrated that expression of hGCSF was detected in only the mammary gland in the progeny of all founders. Moreover, cells were shown to be positive for hG-CSF by immunofluorescent analysis in the mammary glands but not in any other tissues. There were no signs of mosaic expression in the mammary gland. Trace amounts of hG-CSF were detected in the serum of females of two transgenic lines during lactation only. However, no transgenic mice showed any changes in hematopoiesis based on the number of granulocytes in blood. Immunoblotting of hG-CSF in the milk of transgenic mice revealed two forms, presumably the glycosylated and non-glycosylated forms. The hematopoietic activity of hG-CSF in the milk of transgenic females is comparable to that of recombinant G-CSF. In general, the data obtained in this study show that the new expression vector is able to provide correct tissue-specific expression of hG-CSF with high biological activity in transgenic mice.

  18. Amifostine plus granulocyte colony-stimulating factor therapy enhances recovery from supralethal radiation exposures: preclinical experience in animals models.

    PubMed

    Patchen, M L

    1995-01-01

    A murine model was used to explore whether the cytoprotective agent amifostine (WR-2721) can be used to protect a critical fraction of haemopoietic stem cells against radiation, and whether granulocyte colony-stimulating factor (G-CSF) can then be used to stimulate the protected cells to proliferate and reconstitute the haematopoietic system. Groups of C3H/HeN mice treated with 200 mg/kg amifostine i.p. 30 min before 60Co irradiation and/or 125 micrograms/kg G-CSF subcutaneously from days 1-16 post irradiation were compared. The dose reduction factor (DRF) of the combination of amifostine and G-CSF from LD50/30 values was greater than the sum of the DRFs for amifostine and G-CSF individually. Acceleration of recovery bone marrow and splenic multipotent stem cells (CFU-s) and granulocyte-macrophage progenitor cells (GM-CFC), as well as of peripheral blood red and white cells and platelets, was greatest in mice treated with amifostine plus G-CSF. These studies suggest that amifostine and recombinant haematopoietic growth factors can be used in combination to reduce myelosuppression and lethality associated with radiation or radiomimetic drugs

  19. Carcinoembryonic antigen-related cell adhesion molecule-1 regulates granulopoiesis by inhibition of granulocyte colony-stimulating factor receptor.

    PubMed

    Pan, Hao; Shively, John E

    2010-10-29

    Although carcinoembryonic antigen-related cell adhesion molecule-1 (CEACAM1) is an activation marker for neutrophils and delays neutrophil apoptosis, the role of CEACAM1 in granulopoiesis and neutrophil-dependent host immune responses has not been investigated. CEACAM1 expression correlated with granulocytic differentiation, and Ceacam1(-/-) mice developed neutrophilia because of loss of the Src-homology-phosphatase-1 (SHP-1)-dependent inhibition of granulocyte colony-stimulating factor receptor (G-CSFR) signal transducer and activator of transcription (Stat3) pathway provided by CEACAM1. Moreover, Ceacam1(-/-) mice were hypersensitive to Listeria Monocytogenes (LM) infection with an accelerated mortality. Reintroduction of CEACAM1 into Ceacam1(-/-) bone marrow restored normal granulopoiesis and host sensitivity to LM infection, while mutation of its immunoreceptor tyrosine-based inhibitory motifs (ITIMs) abrogated this restoration. shRNA-mediated reduction of Stat3 amounts rescued normal granulopoiesis, attenuating host sensitivity to LM infection in Ceacam1(-/-) mice. Thus, CEACAM1 acted as a coinhibitory receptor for G-CSFR regulating granulopoiesis and host innate immune response to bacterial infections.

  20. Granulocyte colony-stimulating factor (G-CSF) production in hemorrhagic shock requires both the ischemic and resuscitation phase.

    PubMed

    Hierholzer, C; Kelly, E; Billiar, T R; Tweardy, D J

    1997-01-01

    Granulocyte colony-stimulating factor (G-CSF) is the cytokine that is critical for polymorphonuclear neutrophilic granulocyte (PMN) production as well as being a potent agonist of PMN activation. We have recently reported that in the lung and the liver of rats resuscitated after hemorrhagic shock (HS) G-CSF mRNA expression is induced. It is not known if both phases of HS, the ischemic and the reperfusion phase, are required for G-CSF mRNA induction. The present study was designed to test the hypothesis that the upregulation of G-CSF mRNA expression is the consequence of HS followed by resuscitation and that ischemia alone is insufficient to induce G-CSF mRNA expression in the affected organs. Male Sprague-Dawley rats were subjected to resuscitated and unresuscitated shock protocols of varying severity. Control animals were subjected to anesthesia and all surgical preparations except for hemorrhage. Lungs and livers were isolated and their RNA extracted. Using semiquantitative reverse transcriptase-polymerase chain reaction (RT-PCR), we demonstrated that G-CSF mRNA was induced in the lung and liver of shock animals above the level observed in control animals. Upregulation of G-CSF mRNA relative to controls occurred only in animals undergoing resuscitated HS and not in ones subjected to unresuscitated HS. These results indicate that G-CSF production specific for the hemorrhage component of shock is dependent on resuscitation. As a consequence, the production of this cytokine may be decreased through modifications in the resuscitation protocols.

  1. Replication of human immunodeficiency virus in monocytes. Granulocyte/macrophage colony-stimulating factor (GM-CSF) potentiates viral production yet enhances the antiviral effect mediated by 3'-azido- 2'3'-dideoxythymidine (AZT) and other dideoxynucleoside congeners of thymidine

    PubMed Central

    1989-01-01

    We have investigated the influence of granulocyte-macrophage CSF (GM- CSF) on the replication of HIV-1 in cells of monocyte/macrophage (M/M) lineage, and its effect on the anti-HIV activity of several 2'3'- dideoxynucleoside congeners of thymidine in these cells in vitro. We found that replication of both HTLV-IIIBa-L (a monocytotropic strain of HIV-1) and HTLV-IIIB (a lymphocytotropic strain) is markedly enhanced in M/M, but not in lymphocytes exposed to GM-CSF in culture. Moreover, GM-CSF reduced the dose of HIV required to obtain productive infection in M/M. Even in the face of this increased infection, GM-CSF also enhanced the net anti-HIV activity of 3'-azido-2'3'-dideoxythymidine (AZT) and several related congeners: 2'3'-dideoxythymidine (ddT), 2'3'- dideoxy-2'3'-didehydrothymidine (D4T), and 3'-azido-2'3'-dideoxyuridine (AZddU). Inhibition of viral replication in GM-CSF-exposed M/M was achieved with concentrations of AZT and related drugs, which were 10- 100 times lower than those inhibitory for HIV-1 in monocytes in the absence of GM-CSF. Other dideoxynucleosides not related to AZT showed unchanged or decreased anti-HIV activity in GM-CSF-exposed M/M. To investigate the possible biochemical basis for these effects, we evaluated the metabolism of several drugs in M/M exposed to GM-CSF. We observed in these cells markedly increased levels of both parent and mono-, di-, and triphosphate anabolites of AZT and D4T compared with M/M not exposed to GM-CSF. By contrast, only limited increases of endogenous competing 2'-deoxynucleoside-5'-triphosphate pools were observed after GM-CSF exposure. Thus, the ratio of AZT-5'- triphosphate/2'-deoxythymidine-5'-triphosphate and 2'3'-dideoxy-2'3'- didehydrothymidine-5'-triphosphate/2'-deoxythymi dine- 5'-triphosphate is several-fold higher in GM-CSF-exposed M/M, and this may account for the enhanced activity of such drugs in these cells. Taken together, these findings suggest that GM-CSF increases HIV-1 replication in M

  2. Randomized Trial of Two Dosages of Prophylactic Granulocyte Colony-Stimulating Factor after Induction Chemotherapy in Pediatric Acute Myeloid Leukemia

    PubMed Central

    Inaba, Hiroto; Cao, Xueyuan; Pounds, Stanley; Pui, Ching-Hon; Rubnit, Jeffrey E.; Ribeiro, Raul C.; Razzouk, Bassem I.

    2010-01-01

    Background Granulocyte colony-stimulating factor (G-CSF) is effective in accelerating neutrophil recovery after intensive chemotherapy for acute myeloid leukemia (AML). However, the optimal G-CSF dosage for patients with AML has not been determined. To our knowledge, G-CSF dosages have not been compared in a randomized AML study. Methods Patients enrolled on the St. Jude AML97 protocol who remained on study after window therapy were eligible to participate. The effect of the dosage of G-CSF given after induction chemotherapy courses 1 and 2 was analyzed in 46 patients randomly assigned in a double-blinded manner to receive 5 or 10 μg/kg/day of G-CSF. The number of days of G-CSF treatment, neutropenia (absolute neutrophil count < 0.5 × 109/L), and hospitalization; the number of episodes of febrile neutropenia, grade 2-4 infection, and antimicrobial therapy; transfusion requirements; the cost of supportive care; and survival were compared between the two study arms. Results We found no statistically significant difference between the two arms in any of the endpoints measured. Conclusions The higher G-CSF dosage (10 μg/kg/day) offers no greater benefit than the lower dosage (5 μg/kg/day) in patients undergoing intensive chemotherapy for AML. PMID:21381017

  3. Granulocyte colony-stimulating factor administration to healthy volunteers: analysis of the immediate activating effects on circulating neutrophils.

    PubMed

    de Haas, M; Kerst, J M; van der Schoot, C E; Calafat, J; Hack, C E; Nuijens, J H; Roos, D; van Oers, R H; von dem Borne, A E

    1994-12-01

    In four healthy volunteers, we analyzed in detail the immediate in vivo effects on circulating neutrophils of subcutaneous administration of 300 micrograms of granulocyte colony-stimulating factor (G-CSF). Neutrophil activation was assessed by measurement of degranulation. Mobilization of secretory vesicles was shown by a decrease in leukocyte alkaline phosphatase content of the circulating neutrophils. Furthermore, shortly postinjection, Fc gamma RIII was found to be upregulated from an intracellular pool that we identified by immunoelectron microscopy as secretory vesicles. Intravascular release of specific granules was shown by increased plasma levels of lactoferrin and by upregulation of the expression of CD66b and CD11b on circulating neutrophils. Moreover, measurement of fourfold elevated plasma levels of elastase, bound to its physiologic inhibitor alpha 1-antitrypsin, indicated mobilization of azurophil granules. However, no expression of CD63, a marker of azurophil granules, was observed on circulating neutrophils. G-CSF--induced mobilization of secretory vesicles and specific granules could be mimicked in whole blood cultures in vitro, in contrast to release of azurophil granules. Therefore, we postulate that the most activated neutrophils leave the circulation, as observed shortly postinjection, and undergo subsequent stimulation in the endothelial microenvironment, resulting in mobilization of azurophil granules. Our data demonstrate that G-CSF should be regarded as a potent immediate activator of neutrophils in vivo.

  4. The granulocyte-macrophage colony-stimulating factor receptor: linking its structure to cell signaling and its role in disease.

    PubMed

    Hercus, Timothy R; Thomas, Daniel; Guthridge, Mark A; Ekert, Paul G; King-Scott, Jack; Parker, Michael W; Lopez, Angel F

    2009-08-13

    Already 20 years have passed since the cloning of the granulocyte-macrophage colony-stimulating factor (GM-CSF) receptor alpha-chain, the first member of the GM-CSF/interleukin (IL)-3/IL-5 family of hemopoietic cytokine receptors to be molecularly characterized. The intervening 2 decades have uncovered a plethora of biologic functions transduced by the GM-CSF receptor (pleiotropy) and revealed distinct signaling networks that couple the receptor to biologic outcomes. Unlike other hemopoietin receptors, the GM-CSF receptor has a significant nonredundant role in myeloid hematologic malignancies, macrophage-mediated acute and chronic inflammation, pulmonary homeostasis, and allergic disease. The molecular mechanisms underlying GM-CSF receptor activation have recently been revealed by the crystal structure of the GM-CSF receptor complexed to GM-CSF, which shows an unexpected higher order assembly. Emerging evidence also suggests the existence of intracellular signosomes that are recruited in a concentration-dependent fashion to selectively control cell survival, proliferation, and differentiation by GM-CSF. These findings begin to unravel the mystery of cytokine receptor pleiotropy and are likely to also apply to the related IL-3 and IL-5 receptors as well as other heterodimeric cytokine receptors. The new insights in GM-CSF receptor activation have clinical significance as the structural and signaling nuances can be harnessed for the development of new treatments for malignant and inflammatory diseases.

  5. Granulocyte-Macrophage Colony-Stimulating Factor Auto-Antibodies: A Marker of Aggressive Crohn’s Disease

    PubMed Central

    Gathungu, Grace; Kim, Mi-Ok; Ferguson, John P.; Sharma, Yashoda; Zhang, Wei; Ng, Sok Meng E.; Bonkowski, Erin; Ning, Kaida; Simms, Lisa A.; Croft, Anthony R.; Stempak, Joanne M.; Walker, Nicole; Huang, Ning; Xiao, Yang; Silverberg, Mark S.; Trapnell, Bruce C.; Cho, Judy H.; Radford-Smith, Graham L.; Denson, Lee A.

    2013-01-01

    Background & Aims Neutralizing auto-antibodies (Ab) against granulocyte-macrophage colony-stimulating factor (GM-CSF Ab) have been associated with stricturing ileal Crohn’s disease (CD) in a largely pediatric patient cohort (total 394, adult CD 57). The aim of this study was to examine this association in two independent predominantly adult inflammatory bowel disease patient cohorts. Methods Serum samples from 745 subjects from the NIDDK IBD Genetics Consortium and 737 patients from Australia were analyzed for GM-CSF Ab and genetic markers. We conducted multiple regression analysis with backwards elimination to assess the contribution of GM-CSF Ab levels, established CD risk alleles and smoking on ileal disease location in the 477 combined CD subjects from both cohorts. We also determined associations of GM-CSF Ab levels with complications requiring surgical intervention in combined CD subjects in both cohorts. Results Serum samples from CD patients expressed significantly higher concentrations of GM-CSF Ab when compared to Ulcerative Colitis or controls in each cohort. Non-smokers with ileal CD expressed significantly higher GM-CSF Ab concentrations in the Australian cohort (p= 0.002). Elevated GM-CSF Ab, ileal disease location and disease duration greater than 3 years were independently associated with stricturing/penetrating behavior and intestinal resection for CD. Conclusions The expression of high GM-CSF Ab is a risk marker for aggressive CD behavior and complications including surgery. Modifying factors include environmental exposure to smoking and genetic risk markers. PMID:23749272

  6. Granulocyte-macrophage colony-stimulating factor dependent monocyte-mediated cytotoxicity post-autologous bone marrow transplantation.

    PubMed

    Nagler, A; Shur, I; Barak, V; Fabian, I

    1996-08-01

    We investigated the in vitro antitumor activity of monocytes derived from autologous bone marrow transplanted (ABMT) patients treated in vivo with granulocyte-macrophage colony-stimulating factor (GM-CSF). Thirty-four patients (17 female, 17 male), median age 42 (range 3-57) years, were enrolled in the study. Fourteen patients were diagnosed with non-Hodgkin's lymphoma (NHL), eight with Hodgkin's disease (HD), nine with breast cancer and three with neuroblastoma. Six patients who did not receive GM-CSF post-ABMT served as controls. We assessed cytotoxicity, antibody-dependent cellular cytotoxicity (ADCC), expression of the activation antigen CD16, and cytokine production by an enriched population of monocytes (> 90% CD+14) pre-, during and post-GM-CSF administration. Within the group of patients receiving treatment, ADCC was significantly higher during in vivo GM-CSF administration than post-therapy (P < 0.05) and in 50% of these patients, ADCC increased during in vivo GM-CSF administration over pretreatment values. In addition, in vivo GM-CSF administration caused the monocytes to secrete elevated levels of tumor necrosis factor-alpha (TNF-alpha) and GM-CSF (P < 0.05). We conclude that GM-CSF augments monocyte-mediated cytotoxicity post-ABMT, and therefore may have a role in controlling minimal residual disease post-transplant.

  7. GRANULOCYTE COLONY-STIMULATING FACTOR: MOLECULAR MECHANISMS OF ACTION DURING STEADY STATE AND ‘EMERGENCY’ HEMATOPOIESIS

    PubMed Central

    Panopoulos, Athanasia D.; Watowich, Stephanie S.

    2008-01-01

    Neutrophils are phagocytes whose principal function is to maintain anti-bacterial immunity. Neutrophils ingest and kill invading bacteria, releasing cytotoxic, chemotactic and inflammatory mediators at sites of infection. This serves to control the immediate host immune response and attract other cells, such as macrophages and dendritic cells, which are important for establishing long-term adaptive immunity. Neutrophils thus contribute to both the initiation and the maintenance of inflammation at sites of infection. Aberrant neutrophil activity is deleterious; suppressed responses can cause extreme susceptibility to infection while overactivation can lead to excessive inflammation and tissue damage. This review will focus on neutrophil regulation by granulocyte colony-stimulating factor (G-CSF), the principal cytokine controlling neutrophil development and function. The review will emphasize the molecular aspects of G-CSF-driven granulopoiesis in steady state (healthy) conditions and during demand-driven or ‘emergency’ conditions elicited by infection or clinical administration of G-CSF. Understanding the molecular control of granulopoiesis will aid in the development of new approaches designed to treat disorders of neutrophil production and function. PMID:18400509

  8. Effect of granulocyte colony stimulating factor (G-CSF) on IVF outcomes in infertile women: An RCT

    PubMed Central

    Eftekhar, Maryam; Hosseinisadat, Robabe; Baradaran, Ramesh; Naghshineh, Elham

    2016-01-01

    Background: Despite major advances in assisted reproductive techniques, the implantation rates remain relatively low. Some studies have demonstrated that intrauterine infusion of granulocyte colony stimulating factor (G-CSF) improves implantation in infertile women. Objective: To assess the G-CSF effects on IVF outcomes in women with normal endometrial thickness. Materials and methods: In this randomized controlled clinical trial, 100 infertile women with normal endometrial thickness who were candidate for IVF were evaluated in two groups. Exclusion criteria were positive history of repeated implantation failure (RIF), endocrine disorders, severe endometriosis, congenital or acquired uterine anomaly and contraindication for G-CSF (renal disease, sickle cell disease, or malignancy). In G-CSF group (n=50), 300 µg trans cervical intrauterine of G-CSF was administered at the oocyte retrieval day. Controls (n=50) were treated with standard protocol. Chemical, clinical and ongoing pregnancy rates, implantation rate, and miscarriage rate were compared between groups. Results: Number of total and mature oocytes (MII), two pronuclei (2PN), total embryos, transferred embryos, quality of transferred embryos, and fertilization rate did not differ significantly between two groups. So there were no significant differences between groups in chemical, clinical and ongoing pregnancy rate, implantation rate, and miscarriage rate Conclusion: our result showed in normal IVF patients with normal endometrial thickness, the intrauterine infusion of G-CSF did not improve pregnancy outcomes. PMID:27326420

  9. Granulocyte-colony stimulating factor decreases the extent of ovarian damage caused by cisplatin in an experimental rat model

    PubMed Central

    Akdemir, Ali; Akman, Levent; Ergenoglu, Ahment Mete; Yeniel, Ahmet Ozgur; Erbas, Oytun; Yavasoglu, Altug; Terek, Mustafa Cosan; Taskiran, Dilek

    2014-01-01

    Objective To investigate whether granulocyte-colony stimulating factor (G-CSF) can decrease the extent of ovarian follicle loss caused by cisplatin treatment. Methods Twenty-one adult female Sprague-Dawley rats were used. Fourteen rats were administered 2 mg/kg/day cisplatin by intraperitoneal injection twice per week for five weeks (total of 20 mg/kg). Half of the rats (n=7) were treated with 1 mL/kg/day physiological saline, and the other half (n=7) were treated with 100 µg/kg/day G-CSF. The remaining rats (n=7, control group) received no therapy. The animals were then euthanized, and both ovaries were obtained from all animals, fixed in 10% formalin, and stored at 4℃ for paraffin sectioning. Blood samples were collected by cardiac puncture and stored at -30℃ for hormone assays. Results All follicle counts (primordial, primary, secondary, and tertiary) and serum anti-Müllerian hormone levels were significantly increased in the cisplatin+G-CSF group compared to the cisplatin+physiological saline group. Conclusion G-CSF was beneficial in decreasing the severity of follicle loss in an experimental rat model of cisplatin chemotherapy. PMID:25142624

  10. Intranasal Delivery of Granulocyte Colony-Stimulating Factor Enhances Its Neuroprotective Effects Against Ischemic Brain Injury in Rats.

    PubMed

    Sun, Bao-liang; He, Mei-qing; Han, Xiang-yu; Sun, Jing-yi; Yang, Ming-feng; Yuan, Hui; Fan, Cun-dong; Zhang, Shuai; Mao, Lei-lei; Li, Da-wei; Zhang, Zong-yong; Zheng, Cheng-bi; Yang, Xiao-yi; Li, Yang V; Stetler, R Anne; Chen, Jun; Zhang, Feng

    2016-01-01

    Granulocyte colony-stimulating factor (G-CSF) is a hematopoietic growth factor with strong neuroprotective properties. However, it has limited capacity to cross the blood-brain barrier and thus potentially limiting its protective capacity. Recent studies demonstrated that intranasal drug administration is a promising way in delivering neuroprotective agents to the central nervous system. The current study therefore aimed at determining whether intranasal administration of G-CSF increases its delivery to the brain and its neuroprotective effect against ischemic brain injury. Transient focal cerebral ischemia in rat was induced with middle cerebral artery occlusion. Our resulted showed that intranasal administration is 8-12 times more effective than subcutaneous injection in delivering G-CSF to cerebrospinal fluid and brain parenchyma. Intranasal delivery enhanced the protective effects of G-CSF against ischemic injury in rats, indicated by decreased infarct volume and increased recovery of neurological function. The neuroprotective mechanisms of G-CSF involved enhanced upregulation of HO-1 and reduced calcium overload following ischemia. Intranasal G-CSF application also promoted angiogenesis and neurogenesis following brain ischemia. Taken together, G-CSF is a legitimate neuroprotective agent and intranasal administration of G-CSF is more effective in delivery and neuroprotection and could be a practical approach in clinic.

  11. Characterization of Stress-Exposed Granulocyte Colony Stimulating Factor Using ELISA and Hydrogen/Deuterium Exchange Mass Spectrometry

    NASA Astrophysics Data System (ADS)

    Tsuchida, Daisuke; Yamazaki, Katsuyoshi; Akashi, Satoko

    2014-10-01

    Information on the higher-order structure is important in the development of biopharmaceutical drugs. Recently, hydrogen/deuterium exchange coupled with mass spectrometry (HDX-MS) has been widely used as a tool to evaluate protein conformation, and unique automated systems for HDX-MS are now commercially available. To investigate the potential of this technique for the prediction of the activity of biopharmaceuticals, granulocyte colony stimulating factor (G-CSF), which had been subjected to three different stress types, was analyzed using HDX-MS and through comparison with receptor-binding activity. It was found that HDX-MS, in combination with ion mobility separation, was able to identify conformational changes in G-CSF induced by stress, and a good correlation with the receptor-binding activity was demonstrated, which cannot be completely determined by conventional peptide mapping alone. The direct evaluation of biological activity using bioassay is absolutely imperative in biopharmaceutical development, but HDX-MS can provide the alternative information in a short time on the extent and location of the structural damage caused by stresses. Furthermore, the present study suggests the possibility of this system being a versatile evaluation method for the preservation stability of biopharmaceuticals.

  12. Using hydrogen/deuterium exchange mass spectrometry to study conformational changes in granulocyte colony stimulating factor upon PEGylation.

    PubMed

    Wei, Hui; Ahn, Joomi; Yu, Ying Qing; Tymiak, Adrienne; Engen, John R; Chen, Guodong

    2012-03-01

    PEGylation is the covalent attachment of polyethylene glycol to proteins, and it can be used to alter immunogenicity, circulating half life and other properties of therapeutic proteins. To determine the impact of PEGylation on protein conformation, we applied hydrogen/deuterium exchange mass spectrometry (HDX MS) to analyze granulocyte colony stimulating factor (G-CSF) upon PEGylation as a model system. The combined use of HDX automation technology and data analysis software allowed reproducible and robust measurements of the deuterium incorporation levels for peptic peptides of both PEGylated and non-PEGylated G-CSF. The results indicated that significant differences in deuterium incorporation were induced by PEGylation of G-CSF, although the overall changes observed were quite small. PEGylation did not result in gross conformational rearrangement of G-CSF. The data complexity often encountered in HDX MS measurements was greatly reduced through a data processing and presentation format designed to facilitate the comparison process. This study demonstrates the practical utility of HDX MS for comparability studies, process monitoring, and protein therapeutic characterization in the biopharmaceutical industry.

  13. Effects of a granulocyte colony stimulating factor, Neulasta, in mini pigs exposed to total body proton irradiation

    NASA Astrophysics Data System (ADS)

    Sanzari, Jenine K.; Krigsfeld, Gabriel S.; Shuman, Anne L.; Diener, Antonia K.; Lin, Liyong; Mai, Wilfried; Kennedy, Ann R.

    2015-04-01

    Astronauts could be exposed to solar particle event (SPE) radiation, which is comprised mostly of proton radiation. Proton radiation is also a treatment option for certain cancers. Both astronauts and clinical patients exposed to ionizing radiation are at risk for loss of white blood cells (WBCs), which are the body's main defense against infection. In this report, the effect of Neulasta treatment, a granulocyte colony stimulating factor, after proton radiation exposure is discussed. Mini pigs exposed to total body proton irradiation at a dose of 2 Gy received 4 treatments of either Neulasta or saline injections. Peripheral blood cell counts and thromboelastography parameters were recorded up to 30 days post-irradiation. Neulasta significantly improved WBC loss, specifically neutrophils, in irradiated animals by approximately 60% three days after the first injection, compared to the saline treated, irradiated animals. Blood cell counts quickly decreased after the last Neulasta injection, suggesting a transient effect on WBC stimulation. Statistically significant changes in hemostasis parameters were observed after proton radiation exposure in both the saline and Neulasta treated irradiated groups, as well as internal organ complications such as pulmonary changes. In conclusion, Neulasta treatment temporarily alleviates proton radiation-induced WBC loss, but has no effect on altered hemostatic responses.

  14. Effects of a granulocyte colony stimulating factor, Neulasta, in mini pigs exposed to total body proton irradiation

    PubMed Central

    Sanzari, Jenine K.; Krigsfeld, Gabriel S.; Shuman, Anne L.; Diener, Antonia K.; Lin, Liyong; Mai, Wilfried; Kennedy, Ann R.

    2015-01-01

    Astronauts could be exposed to solar particle event (SPE) radiation, which is comprised mostly of proton radiation. Proton radiation is also a treatment option for certain cancers. Both astronauts and clinical patients exposed to ionizing radiation are at risk for white blood cell (WBC) loss, which are the body’s main defense against infection. In this report, the effect of Neulasta treatment, a granulocyte colony stimulating factor, after proton radiation exposure is discussed. Mini pigs exposed to total body proton irradiation at a dose of 2 Gy received 4 treatments of either Neulasta or saline injections. Peripheral blood cell counts and thromboelastography parameters were recorded up to 30 days post-irradiation. Neulasta significantly improved white blood cell (WBC), specifically neutrophil, loss in irradiated animals by approximately 60% three days after the first injection, compared to the saline treated irradiated animals. Blood cell counts quickly decreased after the last Neulasta injection, suggesting a transient effect on WBC stimulation. Statistically significant changes in hemostasis parameters were observed after proton radiation exposure in both the saline and Neulasta treated irradiated groups, as well internal organ complications such as pulmonary changes. In conclusion, Neulasta treatment temporarily alleviates proton radiation-induced WBC loss, but has no effect on altered hemostatic responses. PMID:25909052

  15. Protective effect of recombinant murine granulocyte-macrophage colony-stimulating factor against Pseudomonas aeruginosa infection in leukocytopenic mice.

    PubMed Central

    Tanaka, T.; Okamura, S.; Okada, K.; Suga, A.; Shimono, N.; Ohhara, N.; Hirota, Y.; Sawae, Y.; Niho, Y.

    1989-01-01

    The effects of recombinant murine granulocyte-macrophage colony-stimulating factor (rmGM-CSF) against Pseudomonas aeruginosa infection in ICR mice were investigated. Mice were treated with cyclophosphamide (CPA) and were then injected intraperitoneally with rmGM-CSF three times daily, beginning on the day after CPA treatment, for 7 days. The number of peripheral blood leukocytes in both CPA- and rmGM-CSF-treated mice and control CPA-treated mice reached a nadir on day 4, when P. aeruginosa was injected intraperitoneally. The administration of rmGM-CSF significantly increased the proportion of survivors among mice infected with a lethal dose of P. aeruginosa. This effect was further analyzed by monitoring sequential changes in leukocyte count and bacterial growth in various organs. The number of bacteria in the peritoneal cavities, peripheral blood samples, and livers of GM-CSF-treated mice decreased to an undetectable level after a transient increase, and the number was significantly lower than that in control mice. In GM-CSF-treated mice, the neutrophil levels in peripheral blood started to increase 5 days after CPA administration and were consistently higher than those in controls. Furthermore, the neutrophils in GM-CSF-treated mice were more mature morphologically. Thus, the prophylactic effect of rmGM-CSF against P. aeruginosa infection may result from a rapid recovery of myelopoiesis and a partial enhancement of mature neutrophil function. PMID:2656523

  16. The role of stem cell factor and granulocyte-colony stimulating factor in brain repair during chronic stroke.

    PubMed

    Piao, Chun-Shu; Gonzalez-Toledo, Maria E; Xue, Yue-Qiang; Duan, Wei-Ming; Terao, Satoshi; Granger, D Neil; Kelley, Roger E; Zhao, Li-Ru

    2009-04-01

    Chronic stroke is a highly important but under-investigated scientific problem in neurologic research. We have reported earlier that stem cell factor (SCF) in combination with granulocyte-colony stimulating factor (G-CSF) treatment during chronic stroke improves functional outcomes. Here we have determined the contribution of bone marrow-derived cells in angiogenesis and neurogenesis, which are enhanced by SCF+G-CSF treatment during chronic stroke. Using bone marrow tracking, flow cytometry, 2-photon live brain imaging, and immunohistochemistry, we observed that the levels of circulating bone marrow stem cells (BMSCs) (CD34+/c-kit+) were significantly increased by SCF+G-CSF treatment. In addition, live brain imaging revealed that numerous bone marrow-derived cells migrate into the brain parenchyma in the treated mice. We also found that bone marrow-derived cells, bone marrow-derived endothelial cells, vascular density, and bone marrow-derived neurons were significantly augmented by SCF+G-CSF. It is interesting that, in addition to the increase in bone marrow-derived endothelial cells, the number of bone marrow-derived pericytes was reduced after SCF+G-CSF treatment during chronic stroke. These data suggest that SCF+G-CSF treatment can enhance repair of brain damage during chronic stroke by mobilizing BMSCs, and promoting the contribution of bone marrow-derived cells to angiogenesis and neurogenesis. PMID:19209180

  17. Isolated abdominal aortitis following administration of granulocyte colony stimulating factor (G-CSF).

    PubMed

    Miller, Edward B; Grosu, Roy; Landau, Zvi

    2016-06-01

    G-CSF is a myeloid growth factor produced by monocytes, macrophages, fibroblasts, and endothelial cells. Clinical uses of G-CSF includes mobilization of peripheral blood progenitor cells from healthy donors before hematopoietic stem cell transplantation, acceleration of neutrophil recovery following chemotherapy, and in the management of neutropenia due to other causes including AIDS and genetic disorders of granulocyte production. G-CSF is well tolerated and reports to be safe in healthy donors, although follow-up studies are limited in duration (D'Souza et al. in Transfus Med Rev 22(4):280-290, 2008).Isolated abdominal aortitis (IAA) is a rare disorder most commonly caused by the large-vessel vasculitides giant cell arteritis (GCA) and Takayasu arteritis, although it may also be associated with several other rheumatologic diseases and infections (Gornik and Creager in Circulation 117:3039-3051, 2008). To our knowledge, there only two cases have been published of IAA occurring in patients who had received G-CSF therapy (Dariea et al. in Rev Med Interne 25(3):225-229, 2004; Adiga et al. in Clin Drug Investig 29:821-825, 2009).We describe a case of a 55-year-old male, with peripheral vascular disease who after receiving Neupogen (G-CSF) developed a latent case of IAA. After further investigation and exclusion of other possible causative factors, we conclude that the most probable etiology is induction by G-CSF.

  18. Isolated abdominal aortitis following administration of granulocyte colony stimulating factor (G-CSF).

    PubMed

    Miller, Edward B; Grosu, Roy; Landau, Zvi

    2016-06-01

    G-CSF is a myeloid growth factor produced by monocytes, macrophages, fibroblasts, and endothelial cells. Clinical uses of G-CSF includes mobilization of peripheral blood progenitor cells from healthy donors before hematopoietic stem cell transplantation, acceleration of neutrophil recovery following chemotherapy, and in the management of neutropenia due to other causes including AIDS and genetic disorders of granulocyte production. G-CSF is well tolerated and reports to be safe in healthy donors, although follow-up studies are limited in duration (D'Souza et al. in Transfus Med Rev 22(4):280-290, 2008).Isolated abdominal aortitis (IAA) is a rare disorder most commonly caused by the large-vessel vasculitides giant cell arteritis (GCA) and Takayasu arteritis, although it may also be associated with several other rheumatologic diseases and infections (Gornik and Creager in Circulation 117:3039-3051, 2008). To our knowledge, there only two cases have been published of IAA occurring in patients who had received G-CSF therapy (Dariea et al. in Rev Med Interne 25(3):225-229, 2004; Adiga et al. in Clin Drug Investig 29:821-825, 2009).We describe a case of a 55-year-old male, with peripheral vascular disease who after receiving Neupogen (G-CSF) developed a latent case of IAA. After further investigation and exclusion of other possible causative factors, we conclude that the most probable etiology is induction by G-CSF. PMID:27094941

  19. Interleukin-10 inhibits burst-forming unit-erythroid growth by suppression of endogenous granulocyte-macrophage colony-stimulating factor production from T cells.

    PubMed

    Oehler, L; Kollars, M; Bohle, B; Berer, A; Reiter, E; Lechner, K; Geissler, K

    1999-02-01

    Numerous cytokines released from accessory cells have been shown to exert either stimulatory or inhibitory growth signals on burst-forming unit-erythroid (BFU-E) growth. Because of its cytokine synthesis-inhibiting effects on T cells and monocytes, interleukin-10 (IL-10) may be a potential candidate for indirectly affecting erythropoiesis. We investigated the effects of IL-10 on BFU-E growth from normal human peripheral blood mononuclear cells (PBMC) using a clonogenic progenitor cell assay. The addition of recombinant human IL-10 to cultures containing recombinant human erythropoietin suppressed BFU-E growth in a dose-dependent manner (by 55.2%, range 47.3-63.3%, p < 0.01, at 10 ng/mL). In contrast, no inhibitory effect of IL-10 was seen when cultivating highly enriched CD34+ cells. BFU-E growth from PBMC also was markedly suppressed in the presence of a neutralizing anti-granulocyte-macrophage colony-stimulating factor (GM-CSF) antibody (by 48.7%, range 32.9-61.2% inhibition,p < 0.01), but not by neutralizing antibodies against granulocyte colony-stimulating factor and interleukin-3. This suggests a stimulatory role of endogenously released GM-CSF on BFU-E formation. Also, the addition of exogenous GM-CSF completely restored IL-10-induced suppression of BFU-E growth. To determine the cellular source of GM-CSF production, we analyzed GM-CSF levels in suspension cultures containing PBMC that were either depleted of monocytes or T cells. Monocyte-depleted PBMC showed spontaneous production of increasing amounts of GM-CSF on days 3, 5, and 7, respectively, which could be suppressed by IL-10, whereas GM-CSF levels did not increase in cultures containing T-cell-depleted PBMC. Our data indicate that IL-10 inhibits the growth of erythroid progenitor cells in vitro, most likely by suppression of endogenous GM-CSF production from T cells.

  20. Characterization of recombinant human granulocyte colony-stimulating factor expression by FT-IR spectroscopy: Studies on thermal induction and media formulation on the stability of the protein secondary structure.

    PubMed

    Vemula, Sandeep; Vemula, Sushma; Dedaniya, Akshay; Kante, Rajesh Kumar; Ronda, Srinivasa Reddy

    2016-08-17

    The Fourier-transform infrared (FT-IR) spectroscopic approach has been employed to understand the recombinant human G-CSF (rhG-CSF) protein accumulation, secondary structure, and thermal stability in Escherichia coli grown under a temperature shift strategy (37 and 28°C) in various media formulations. The choline + sodium pyruvate (37°C) and sodium pyruvate (28°C) formulations have shown the highest inclusion body (IB) accumulation of 0.41 and 0.46 mg/mL, respectively. Furthermore, insights on the structure of the rhG-CSF within IBs and intact cells have been investigated through secondary structure analysis. Thermal stability experiments were also carried out to explain the pattern of the second derivative structure of rhG-CSF. The studies showed that choline + sodium pyruvate formulation has preserved the protein secondary structure even at 82°C. Overall, the FT-IR spectroscopic technique can also be adopted to accelerate the characterization of other recombinant therapeutic proteins of E. coli origin.

  1. Pharmacokinetic studies of intravenous glycosylated recombinant human granulocyte colony-stimulating factor in various hematological disorders: inverse correlation between the half-life and bone marrow myeloid cell pool.

    PubMed

    Watari, K; Ozawa, K; Takahashi, S; Tojo, A; Tani, K; Kamachi, S; Asano, S

    1997-07-01

    The pharmacokinetics of an intravenous bolus dose of glycosylated recombinant human G-CSF (rhG-CSF) was examined in 15 patients with various hematological disorders and 3 normal volunteers. The elimination half-life of rhG-CSF varied with the disorder. The half-life of an initial dose of rhG-CSF (2 micrograms/weight kg) was significantly prolonged in patients with aplastic anemia (2.7 +/- 0.3 h, n = 3) and myelodysplastic syndrome-refractory anemia (2.0 +/- 0.3 h, n = 3) when compared with those in normal controls (0.9 +/- 0.5 h, n = 3). In contrast, in patients with acute myelogenous leukemia which was overt leukemia from myelodysplastic syndrome-refractory anemia with excess of blasts in transformation, the half-life was shortened after chemotherapy (0.2 +/- 0.1 h, n = 3). The half-life of rhG-CSF in 2 patients with acute lymphoblastic leukemia in complete remission was prolonged (2.0 and 2.7 h) at the time of marrow-suppression after chemotherapy and then shortened (0.5, 1.0 h, respectively) in the recovery phase. The half-life of rhG-CSF was very weakly, inversely correlated with absolute neutrophil count in blood (n = 24, r2 = 0.32, P < 0.01), and was inversely correlated with the absolute count of bone-marrow myeloid cells (nucleated cell count in bone-marrow aspirates x the percentage of myeloid cells/100) of patients with aplastic anemia and myelodysplastic syndrome-refractory anemia (n = 12, r2 = 0.63, P = 0.002). These results suggest that the half-life of intravenously administered rhG-CSF (2 micrograms/kg) reflects the size of the myeloid cell compartment in vivo, and support the hypothesis that receptor-mediated consumption mainly accounts for the clearance of exogenous G-CSF.

  2. Biologic Activity of Autologous, Granulocyte-Macrophage Colony Stimulating Factor Secreting Alveolar Soft Parts Sarcoma and Clear Cell Sarcoma Vaccines

    PubMed Central

    Goldberg, John; Fisher, David E.; Demetri, George D.; Neuberg, Donna; Allsop, Stephen A.; Fonseca, Catia; Nakazaki, Yukoh; Nemer, David; Raut, Chandrajit P.; George, Suzanne; Morgan, Jeffrey A.; Wagner, Andrew J.; Freeman, Gordon J.; Ritz, Jerome; Lezcano, Cecilia; Mihm, Martin; Canning, Christine; Hodi, F. Stephen; Dranoff, Glenn

    2015-01-01

    Purpose Alveolar soft parts sarcoma (ASPS) and clear cell sarcoma (CCS) are rare mesenchymal malignancies driven by chromosomal translocations that activate members of the microphthalmia transcription factor (MITF) family. However, in contrast to malignant melanoma, little is known about their immunogenicity. To learn more about the host response to ASPS and CCS, we conducted a phase I clinical trial of vaccination with irradiated, autologous sarcoma cells engineered by adenoviral mediated gene transfer to secrete granulocyte-macrophage colony stimulating factor (GM-CSF). Experimental Design Metastatic tumors from ASPS and CCS patients were resected, processed to single cell suspensions, transduced with a replication defective adenoviral vector encoding GM-CSF, and irradiated. Immunizations were administered subcutaneously and intradermally weekly times three and then every other week. Results Vaccines were successfully manufactured for 11 of the 12 enrolled patients. Eleven subjects received from 3 to 13 immunizations. Toxicities were restricted to grade 1–2 skin reactions at inoculation sites. Vaccination elicited local dendritic cell infiltrates and stimulated T cell mediated delayed type-hypersensitivity reactions to irradiated, autologous tumor cells. Antibody responses to tissue-type plasminogen activator (tTPA) and angiopoietins-1/2 were detected. Tumor biopsies showed programmed death-1 (PD-1) positive CD8+ T cells in association with PD ligand-1 (PD-L1) expressing sarcoma cells. No tumor regressions were observed. Conclusions Vaccination with irradiated, GM-CSF secreting autologous sarcoma cell vaccines is feasible, safe, and biologically active. Concurrent targeting of angiogenic cytokines and antagonism of the PD-1 negative regulatory pathway might intensify immune-mediated tumor destruction. PMID:25805798

  3. A novel combinatorial therapy with pulp stem cells and granulocyte colony-stimulating factor for total pulp regeneration.

    PubMed

    Iohara, Koichiro; Murakami, Masashi; Takeuchi, Norio; Osako, Yohei; Ito, Masataka; Ishizaka, Ryo; Utunomiya, Shinji; Nakamura, Hiroshi; Matsushita, Kenji; Nakashima, Misako

    2013-07-01

    Treatment of deep caries with pulpitis is a major challenge in dentistry. Stem cell therapy represents a potential strategy to regenerate the dentin-pulp complex, enabling conservation and restoration of teeth. The objective of this study was to assess the efficacy and safety of pulp stem cell transplantation as a prelude for the impending clinical trials. Clinical-grade pulp stem cells were isolated and expanded according to good manufacturing practice conditions. The absence of contamination, abnormalities/aberrations in karyotype, and tumor formation after transplantation in an immunodeficient mouse ensured excellent quality control. After autologous transplantation of pulp stem cells with granulocyte-colony stimulating factor (G-CSF) in a dog pulpectomized tooth, regenerated pulp tissue including vasculature and innervation completely filled in the root canal, and regenerated dentin was formed in the coronal part and prevented microleakage up to day 180. Transplantation of pulp stem cells with G-CSF yielded a significantly larger amount of regenerated dentin-pulp complex compared with transplantation of G-CSF or stem cells alone. Also noteworthy was the reduction in the number of inflammatory cells and apoptotic cells and the significant increase in neurite outgrowth compared with results without G-CSF. The transplanted stem cells expressed angiogenic/neurotrophic factors. It is significant that G-CSF together with conditioned medium of pulp stem cells stimulated cell migration and neurite outgrowth, prevented cell death, and promoted immunosuppression in vitro. Furthermore, there was no evidence of toxicity or adverse events. In conclusion, the combinatorial trophic effects of pulp stem cells and G-CSF are of immediate utility for pulp/dentin regeneration, demonstrating the prerequisites of safety and efficacy critical for clinical applications.

  4. Granulocyte colony-stimulating factor impairs CD8(+) T cell functionality by interfering with central activation elements.

    PubMed

    Bunse, C E; Tischer, S; Lahrberg, J; Oelke, M; Figueiredo, C; Blasczyk, R; Eiz-Vesper, B

    2016-07-01

    Besides mobilizing stem cells into the periphery, granulocyte colony-stimulating factor (G-CSF) has been shown to influence various types of innate and adaptive immune cells. For example, it impairs the effector function of cytotoxic T lymphocytes (CTLs). It is assumed that this effect is mediated indirectly by monocytes, regulatory T cells and immunomodulatory cytokines influenced by G-CSF. In this study, isolated G-CSF-treated CD8(+) T cells were stimulated antigen-dependently with peptide-major histocompatibility complex (pMHC)-coupled artificial antigen-presenting cells (aAPCs) or stimulated antigen-independently with anti-CD3/CD28 stimulator beads. By measuring the changes in interferon (IFN)-γ and granzyme B expression at the mRNA and protein level, we showed for the first time that G-CSF has a direct effect on CD8(+) CTLs, which was confirmed based on the reduced production of IFN-γ and granzyme B by the cytotoxic T cell line TALL-104 after G-CSF treatment. By investigating further elements affected by G-CSF in CTLs from stem cell donors and untreated controls, we found a decreased phosphorylation of extracellular-regulated kinase (ERK)1/2, lymphocyte-specific protein tyrosine kinase (Lck) and CD3ζ after G-CSF treatment. Additionally, miRNA-155 and activation marker expression levels were reduced. In summary, our results show that G-CSF directly influences the effector function of cytotoxic CD8(+) T cells and affects various elements of T cell activation. PMID:26990855

  5. Clinical practice in febrile neutropenia risk assessment and granulocyte colony-stimulating factor primary prophylaxis of febrile neutropenia in Poland

    PubMed Central

    Chmielowska, Ewa; Filipczyk-Cisarż, Emilia; Krzemieniecki, Krzysztof; Leśniewski-Kmak, Krzysztof; Litwiniuk, Maria M.; Wieruszewska-Kowalczyk, Karolina; Kosno-Kruszewska, Elżbieta

    2014-01-01

    Aim of the study The first aim was to investigate the knowledge and awareness of oncologists concerning febrile neutropenia (FN) risk assessment and indications for granulocyte colony-stimulating factor (G-CSF) primary prophylaxis (PP), based on current therapeutic guidelines (PTOK and EORTC). The second aim was to educate the oncologists on best practices for risk assessment and neutropenia management. Material and methods The project participants included 169 oncologists from 7 regions working in large specialist oncological centres, university hospitals, regional and city hospitals, specialist outpatient clinics, and oncological wards in small local hospitals. The participants completed a questionnaire based on seven prepared clinical cases of patients with different tumour types and patient characteristics, receiving chemotherapy (CT), and with different levels of FN risk. Participants answered questions related to FN risk assessment and G-CSF use. After completing the questionnaire, the participants proceeded to an educational module in which they were provided with an analysis of correct diagnostic and therapeutic procedures according to the PTOK and EORTC guidelines. Results and Conclusions Febrile neutropenia risk assessment was found to be a routine procedure performed for over 90% of the clinical cases by the participant oncologists. However, the FN risk assessment of clinical cases was correct and consistent with therapeutic guidelines in only 65% of responses. Indications for G-CSF PP were properly identified in 76% of responses and it appeared that indications for G-CSF PP were more likely to be correctly identified in patients receiving high-risk or low-risk regimens than in those receiving intermediate-risk regimens, where the decision to give G-CSF PP is based on additional assessment of patient risk factors. The vast majority of participants who correctly identified the need for PP administered G-CSF in accordance with the dose and schedule

  6. Granulocyte-colony stimulating factor for acute-on-chronic liver failure: systematic review and meta-analysis.

    PubMed

    Chavez-Tapia, Norberto C; Mendiola-Pastrana, Indira; Ornelas-Arroyo, Victoria J; Noreña-Herrera, Camilo; Vidaña-Perez, Desiree; Delgado-Sanchez, Guadalupe; Uribe, Misael; Barrientos-Gutierrez, Tonatiuh

    2015-01-01

    Acute-on-chronic liver failure (ACLF) is associated with increased short and long-term mortality. Animal models of liver failure have demonstrated that granulocyte-colony stimulating factor (G-CSF) accelerates the liver regeneration process and improves survival. However, clinical evidence regarding the use of G-CSF in ACLF remains scarce. The aim of this study was to assess the benefits and harms of G-CSF in patients with acute-on-chronic liver failure. An electronic search was made in The Cochrane Central Register of Controlled Trials (CENTRAL), MEDLINE, EMBASE and LILACS up to November 2013. Randomized clinical trials comparing the use of any regimen of G-CSF against placebo or no intervention in patients with ACLF were included. Primary outcomes included overal mortality, mortality due multi-organ failure, and adverse events. Relative risk (RR) and mean difference (MD) were used. Two trials involving 102 patients were included. A significant reduction in short-term overall mortality was observed in patients receiving G-CSF compared to controls (RR 0.56; 95%CI 0.39,0.80). G-CSF failed to reduce mortality secondary to gastrointestinal bleeding (RR 1.45; 95%CI 0.50, 4.27). Adverse effects reported included: fever, rash, herpes zoster, headache and nausea. In conclusion, the use of G-CSF for the treatment of patients with ACLF significantly reduced short-term mortality. While the evidence is still limited, the apparent benefit observed on short-term mortality, mild adverse effects and lack of an alternative therapy make the use of G-CSF in ACLF patients a reasonable alternative when liver transplantation is contraindicated or unavailable.

  7. Identification of a novel Stat3 recruitment and activation motif within the granulocyte colony-stimulating factor receptor.

    PubMed

    Chakraborty, A; Dyer, K F; Cascio, M; Mietzner, T A; Tweardy, D J

    1999-01-01

    Stat3 is essential for early embryonic development and for myeloid differentiation induced by the cytokines granulocyte colony-stimulating factor (G-CSF) and interleukin-6 (IL-6). Two isoforms of Stat3 have been identified, (p92) and beta (p83), which have distinct transcriptional and biological functions. Activation of both Stat3 and Stat3beta requires the distal cytoplasmic domain of the G-CSFR, which contains four Tyr at positions 704, 729, 744, and 764. The studies reported here were undertaken to determine which, if any, of these tyrosine residues participated in Stat3/beta recruitment and activation. We showed that Stat3 and Stat3beta were affinity purified using phosphopeptides containing Y704 and Y744 but not by nonphosphorylated peptide analogues or by phosphopeptides containing Y729 and Y764. Complementary results were obtained in studies examining the ability of these peptides to destabilize and inhibit DNA binding of activated Stat3. Both Y704 and Y744 contributed to optimal activation of Stat3/beta in M1 murine myeloid leukemia cells containing wild-type and Y-to-F mutant G-CSFR constructs. Carboxy-terminal to Y704 at the +3 position is Gln; YXXQ represents a consensus Stat3 recruitment and activation motif. Y744 is followed at the +3 position by Cys (C); YXXC, represents a novel motif implicated in the recruitment and activation of Stat3. Modeling of the SH2 domain of Stat3 based on homologous SH2 domains of known structure revealed polar residues whose side chains contact the +3 position. This substitution may confer specificity for the Y704- and Y744-based ligands by allowing H-bond formation between the binding surface and the Gln or Cys found at the respective +3 position.

  8. Granulocyte-macrophage colony-stimulating factor (GM-CSF) induces antiapoptotic and proapoptotic signals in acute myeloid leukemia.

    PubMed

    Faderl, Stefan; Harris, David; Van, Quin; Kantarjian, Hagop M; Talpaz, Moshe; Estrov, Zeev

    2003-07-15

    High levels of cytokines are associated with a poor prognosis in acute myeloid leukemia (AML). However, cytokines may induce, on one hand, survival factor expression and cell proliferation and, on the other hand, expression of inhibitory signals such as up-regulation of suppressors of cytokine signaling (SOCS) and induce apoptotic cell death. Because blasts from patients with AML express high procaspase protein levels, we asked whether granulocyte-macrophage colony-stimulating factor (GM-CSF) enhances procaspase protein production in AML cells. In the GM-CSF-responsive OCIM2 AML cell line, GM-CSF induced signal transducer and activator of transcription 5 (Stat 5) phosphorylation, up-regulated cyclin D2, and stimulated cell cycle progression. Concurrently, GM-CSF stimulated expression of SOCS-2 and -3 and of procaspases 2 and 3 and induced caspase 3 activation, poly(ADP[adenosine 5'-diphosphate]-ribose) polymerase (PARP) cleavage, and apoptotic cell death. The Janus kinase (Jak)-Stat inhibitor AG490 abrogated GM-CSF-induced expression of procaspase 3 and activation of caspase 3. Under the same conditions GM-CSF up-regulated production of BAX as well as Bcl-2, Bcl-XL, survivin, and XIAP. GM-CSF also increased procaspase 3 protein levels in OCI/AML3 and Mo7e cells, suggesting that this phenomenon is not restricted to a single leukemia cell line. Our data suggest that GM-CSF exerts a dual effect: it stimulates cell division but contemporaneously up-regulates Jak-Stat-dependent proapoptotic proteins. Up-regulation of procaspase levels in AML is thus a beacon for an ongoing growth-stimulatory signal.

  9. Granulocyte colony-stimulating factor administration to HIV-infected subjects augments reduced leukotriene synthesis and anticryptococcal activity in neutrophils.

    PubMed Central

    Coffey, M J; Phare, S M; George, S; Peters-Golden, M; Kazanjian, P H

    1998-01-01

    Neutrophil (PMN) dysfunction occurs in HIV infection. Leukotrienes (LT) are mediators derived from the 5-lipoxygenase (5-LO) pathway that play a role in host defense and are synthesized by PMN. We investigated the synthesis of LT by PMN from HIV-infected subjects. There was a reduction (4.0+/-1.3% of control) in LT synthesis in PMN from HIV-infected compared with normal subjects. This was associated with reduced expression of 5-LO-activating protein (31.2+/-9.6% of normal), but not of 5-LO itself. Since HIV does not directly infect PMN, we considered that these effects were due to reduced release of cytokines, such as granulocyte colony-stimulating factor (G-CSF). We examined the effect of G-CSF treatment (300 microgram daily for 5 d) on eight HIV-infected subjects. PMN were studied in vitro before therapy (day 1) and on days 4 and 7. LTB4 synthesis was increased on day 4 of G-CSF treatment, and returned toward day 1 levels on day 7. 5-LO and 5-LO-activating protein expression were increased in parallel. As a functional correlate to this increase in PMN LT synthesis by G-CSF, we examined the effects on killing of Cryptococcus neoformans. Anticryptococcal activity of PMN from HIV-infected subjects was less than that of PMN from normal subjects. G-CSF treatment improved fungistatic activity of PMN. This increase in antifungal activity was attenuated by in vitro treatment with the LT synthesis inhibitor, MK-886. In conclusion, PMN from HIV-infected subjects demonstrate reduced 5-LO metabolism and antifungal activity in vitro, which was reversed by in vivo G-CSF therapy. PMID:9710433

  10. Granulocyte-colony stimulating factor improves Parkinson's disease associated with co-morbid depression: An experimental exploratory study

    PubMed Central

    Prakash, Ajay; Chopra, Kanwaljit; Medhi, Bikash

    2013-01-01

    Introduction: The present study was designed to evaluate the effect of granulocyte-colony stimulating factor (G-CSF) in the treatment of Parkinson's disease (PD), the second most common neurodegenerative disease characterized by muscle and movement disorder, often associated with depression. PD is very difficult to treat. Hence, the present study was aimed to evaluate the effect of G-CSF in PD associated with depression. Materials and Methods: Adult Wistar male rats weighing about 180-250 g were selected and divided into five groups in parallel designed method namely; control group (n = 5); sham operated group (n = 5); Vehicle group (n = 5); G-CSF group (70 μg/kg, s.c.) (n = 5) and L-DOPA group (n = 5). The rats were treated with 6-hydroxydopamine (6-OHDA) on day 0 and then treatment was continued for 14 day of L-DOPA/carbidopa, whereas G-CSF (70 μg/kg, s.c.) was given from day 1 to 6. Thereafter, adhesive removal and forced swim tests were conducted to evaluate the behavioral outcome of G-CSF treatment. The finding was correlated and analyzed with Nissl staining findings for the final conclusion. Results: The behavioral parameters were assessed and found to be ameliorate the symptoms of Parkinson's and reduced the depression like behavior in PD. The histological findings were supported the behavioral findings and showed pathological improvement. Conclusion: As a preliminary work, the present study first time suggested that G-CSF have a potential role in PD and associated depression. PMID:24347771

  11. Synergy of interleukin 1 and granulocyte colony-stimulating factor: in vivo stimulation of stem-cell recovery and hematopoietic regeneration following 5-fluorouracil treatment of mice

    SciTech Connect

    Moore, M.A.S.; Warren, D.J.

    1987-10-01

    The human bladder carcinoma cell line 5637 produces hematopoietic growth factors (granulocyte and granulocyte/macrophage colony-stimulating factors (G-CSF and GM-CSF)) and hemopoietin 1, which synergizes with CSFs to stimulate colony formation by primitive hematopoietic stem cells in 5-fluorouracil-treated mouse bone marrow. Molecular and functional properties of hemopoietin 1 identified it as identical to interleukin 1..cap alpha.. (IL-1..cap alpha..). When bone marrow cells from 5-fluorouracil-treated mice were cultured in suspension for 7 days with recombinant human IL-1..cap alpha.. and/or G-CSF, it was found that the two factors synergized to enhance recovery of myelopoietic cells and colony-forming cells of both high and low proliferative potential. G-CSF alone did not sustain these populations, but the combination had greater-than-additive stimulating capacity. In vivo, 5-fluorouracil (150 mg/kg) produced profound myelosuppression and delayed neutrophil regeneration for up to 2 weeks in C3H/HeJ mice. Daily administration of recombinant human G-CSF or human IL-1..cap alpha.. accelerated recovery of stem cells, progenitor cells, and blood neutrophils by up to 4 days in 5-fluorouracil-treated C3H/HeJ and B6D2F/sub 1/ mice. The combination of IL-1..cap alpha.. and G-CSF acted synergistically, reducing neutropenia and accelerating recovery of normal neutrophil numbers by up to 7 days. These results indicate the possible therapeutic potential of combination therapy with IL-1 and hematopoietic growth factors such as G-CSF in the treatment of chemotherapy- or radiation-induced myelosuppression.

  12. Thiotepa cyclophosphamide followed by granulocyte colony-stimulating factor mobilized allogeneic peripheral blood cells in adults with advanced leukemia.

    PubMed

    Bacigalupo, A; Van Lint, M T; Valbonesi, M; Lercari, G; Carlier, P; Lamparelli, T; Gualandi, F; Occhini, D; Bregante, S; Valeriani, A; Piaggio, G; Pitto, A; Benvenuto, F; Figari, O; De Stefano, G; Caimo, A; Sessarego, M

    1996-07-01

    Thirty-one patients (median age, 44 years) with advanced hematologic malignancies were given thiotepa 15 mg/kg, and cyclophosphamide 120 (n = 14) or 150 (n = 17) mg/kg followed by unfractionated peripheral blood stem cell transplants (PBSCT) from genotypically identical siblings (n = 28) or one antigen mismatched family donor (n = 3). Donors were mobilized with granulocyte colony-stimulating factor 5 to 10 microgram/kg/d for 6 days and underwent two to three leukapheresis on days +5, +6, +7. The median cell yield per donor expressed/kg of recipients body weight was as follows: nucleated cells 13 x 10(8)/kg; CD34+ cells 6 x 10(6)/kg; colony-forming unit-granulocyte macrophage 38 x 10(4)/kg, and CD3+ cells 449 x 10(6)/kg. The diagnoses were chronic myeloid leukemia (n = 4), acute myeloid (n = 9) or lymphoid leukemia (n = 2), acute myelofibrosis (n = 2), multiple myeloma (n = 1), lymphoma (n = 6), chronic lymphocytic leukemia (n = 1) myelodysplasia (n = 6). Twenty-eight patients had advanced disease, 29 patients were first grafts, and 2 were second transplants 3 and 9 years after the first. Neutrophil counts of 0.5 x 10(9)/L and platelet counts of 30 x 10(9)/L platelets were both achieved on day +14 (median). Engraftment could be proven by sex markers or DNA polymorphism in 29 of 31 patients: one had early leukemia relapse and one patient was unevaluable because of early death. Acute graft-versus-host disease (GVHD) was scored as minimal or absent (grade 0 to 1) in 14 patients, moderate (grade II) in 13, and severe (grade III to IV) in four. Causes of death were leukemia (n = 4), acute GVHD (n = 4, with associated cytomegalovirus infections in three), sepsis (n = 1), liver failure (n = 1), multiorgan failure (n = 1), and hemorrhage (n = 1). The actuarial transplant mortality is 29%, the actuarial relapse rate 22%. Nineteen patients survive with a median follow up of 288 days (100-690). The actuarial 2-year survival is 57%. Three patients received PBSCT from family

  13. Effects of Granulocyte-Macrophage Colony-Stimulating (GM-CSF) Factor on Corneal Epithelial Cells in Corneal Wound Healing Model.

    PubMed

    Rho, Chang Rae; Park, Mi-young; Kang, Seungbum

    2015-01-01

    Granulocyte-macrophage colony-stimulating factor (GM-CSF) is a pleiotropic cytokine that activates granulocyte and macrophage cell lineages. It is also known to have an important function in wound healing. This study investigated the effect of GM-CSF in wound healing of human corneal epithelial cells (HCECs). We used human GM-CSF derived from rice cells (rice cell-derived recombinant human GM-CSF; rhGM-CSF). An in vitro migration assay was performed to investigate the migration rate of HCECs treated with various concentrations of rhGM-CSF (0.1, 1.0, and 10.0 μg/ml). MTT assay and flow cytometric analysis were used to evaluate the proliferative effect of rhGM-CSF. The protein level of p38MAPK was analyzed by western blotting. For in vivo analysis, 100 golden Syrian hamsters were divided into four groups, and their corneas were de-epithelialized with alcohol and a blade. The experimental groups were treated with 10, 20, or 50 μg/ml rhGM-CSF four times daily, and the control group was treated with phosphate-buffered saline. The corneal wound-healing rate was evaluated by fluorescein staining at the initial wounding and 12, 24, 36, and 48 hours after epithelial debridement. rhGM-CSF accelerated corneal epithelial wound healing both in vitro and in vivo. MTT assay and flow cytometric analysis revealed that rhGM-CSF treatment had no effects on HCEC proliferation. Western blot analysis demonstrated that the expression level of phosphorylated p38MAPK increased with rhGM-CSF treatment. These findings indicate that rhGM-CSF enhances corneal wound healing by accelerating cell migration.

  14. Co-expression of HIV-1 virus-like particles and granulocyte-macrophage colony stimulating factor by GEO-D03 DNA vaccine.

    PubMed

    Hellerstein, Michael; Xu, Yongxian; Marino, Tracie; Lu, Shan; Yi, Hong; Wright, Elizabeth R; Robinson, Harriet L

    2012-11-01

    Here, we report on GEO-D03, a DNA vaccine that co-expresses non-infectious HIV-1 virus-like particles (VLPs) and the human cytokine, granulocyte-macrophage colony-stimulating factor (GM-CSF). The virus-like particles display the native gp160 form of the HIV-1 Envelope glycoprotein (Env) and are designed to elicit antibody against the natural form of Env on virus and virus-infected cells. The DNA-expressed HIV Gag, Pol and Env proteins also have the potential to elicit virus-specific CD4 and CD8 T cells. The purpose of the co-expressed GM-CSF is to target a cytokine that recruits, expands and differentiates macrophages and dendritic cells to the site of VLP expression. The GEO-D03 DNA vaccine is currently entered into human trials as a prime for a recombinant modified vaccinia Ankara (MVA) boost. In preclinical studies in macaques using an SIV prototype vaccine, this vaccination regimen elicited both anti-viral T cells and antibody, and provided 70% protection against acquisition during 12 weekly rectal exposures with a heterologous SIV. Higher avidity of the Env-specific Ab for the native form of the Env in the challenge virus correlated with lower likelihood of SIV infection.

  15. Granulocyte colony-stimulating factor promotes tumor angiogenesis via increasing circulating endothelial progenitor cells and Gr1+CD11b+ cells in cancer animal models.

    PubMed

    Okazaki, Tatsuma; Ebihara, Satoru; Asada, Masanori; Kanda, Akio; Sasaki, Hidetada; Yamaya, Mutsuo

    2006-01-01

    Recombinant granulocyte colony-stimulating factor (G-CSF) is used for cancer patients with myelosuppression induced by chemotherapy. G-CSF has been reported to progress tumor growth and angiogenesis, but the precise mechanism of tumor angiogenesis activated by G-CSF has not been fully clarified. N-terminal-mutated recombinant human G-CSF administration increased WBCs and neutrophils in peripheral blood and reduced bone marrow stromal cell-derived factor-1 in mice, indicating its biological relevance. Mice were inoculated with Lewis lung carcinoma cells (LLCs) or KLN205 cells and treated with G-CSF. G-CSF accelerated tumor growth and intratumoral vessel density, while it did not accelerate proliferation of LLCs, KLN205 cells or human umbilical vein endothelial cells in vitro. In the absence of tumors, G-CSF did not increase circulating cells that displayed phenotypic characteristics of endothelial progenitor cells (EPCs). In the presence of tumors, G-CSF increased circulating EPCs. In addition, G-CSF treatment increased immune suppressor and endothelial cell-differentiating Gr1+CD11b+ cells in tumor-bearing mice. We conclude that G-CSF promotes tumor growth by activating tumor angiogenesis via increasing circulating EPCs and Gr1+CD11b+ cells in cancer animal models.

  16. Purified murine granulocyte/macrophage progenitor cells express a high-affinity receptor for recombinant murine granulocyte/macrophage colony-stimulating factor

    SciTech Connect

    Williams, D.E.; Bicknell, D.C.; Park, L.S.; Straneva, J.E.; Cooper, S.; Broxmeyer, H.E.

    1988-01-01

    Purified recombinant murine granulocyte/macrophage colony-stimulating factor (GM-CSF) was labeled with /sup 125/I and used to examine the GM-CSF receptor on unfractionated normal murine bone marrow cells, casein-induced peritoneal exudate cells, and highly purified murine granulocyte/macrophage progenitor cells (CFU-GM). CFU-GM were isolated from cyclophosphamide-treated mice by Ficoll-Hypaque density centrifugation followed by counterflow centrifugal elutriation. The resulting population had a cloning efficiency of 62-99% in cultures containing conditioned medium from pokeweed mitogen-stimulated spleen cells and 55-86% in the presence of a plateau concentration of purified recombinant murine GM-CSF. Equilibrium binding studies with /sup 125/I-labeled GM-CSF showed that normal bone marrow cells, casein-induced peritoneal exudate cells, and purified CFU-GM had a single class of high-affinity receptor. Affinity crosslinking studies demonstrated that /sup 125/I-labeled GM-CSF bound specifically to two species of M/sub r/ 180,000 and 70,000 on CFU-GM, normal bone marrow cells, and peritoneal exudate cells. The M/sub r/ 70,000 species is thought to be a proteolytic fragment of the intact M/sub r/ 180,000 receptor. The present studies indicate that the GM-CSF receptor expressed on CFU-GM and mature myeloid cells are structurally similar. In addition, the number of GM-CSF receptors on CFU-GM is twice the average number of receptors on casein-induced mature myeloid cells, suggesting that receptor number may decrease as CFU-GM mature.

  17. Alteration of the functional effects of granulocyte-macrophage colony-stimulating factor on polymorphonuclear leukocytes by membrane-fluidizing agents.

    PubMed Central

    Buescher, E S; McIlheran, S M; Banks, S M; Vadhan-Raj, S

    1990-01-01

    Locomotion and oxidative metabolism of polymorphonuclear leukocytes from 15 patients receiving recombinant human granulocyte-macrophage colony-stimulating factor (GM-CSF) were examined in vitro. At the end of each GM-CSF treatment course, polymorphonuclear leukocyte (PMN) chemotactic responses were suppressed and no enhancement of formyl-peptide-stimulated superoxide production was observed. The priming of PMN superoxide production normally seen after in vitro GM-CSF exposure was also blunted in these cells. By using control donor PMN, two membrane-fluidizing agents, pentoxifylline and butanol, were shown to normalize suppressed PMN chemotaxis caused by in vitro GM-CSF (1 nM) exposure. Pentoxifylline, but not butanol, also reversed the effects of in vitro GM-CSF on PMN superoxide production. When PMN obtained from six patients at the end of GM-CSF therapy were exposed to pentoxifylline in vitro, the chemotactic suppression typically observed was significantly improved. The data suggest that GM-CSF may affect PMN function via mechanisms involving membrane fluidity or cell deformability or both. PMID:2167293

  18. The Role of Granulocyte Colony-Stimulating Factor in the Neutrophilia Observed in the Fetal Inflammatory Response Syndrome

    PubMed Central

    Chaiworapongsa, Tinnakorn; Romero, Roberto; Berry, Stanley M.; Hassan, Sonia S.; Yoon, Bo Hyun; Edwin, Samuel; Mazor, Moshe

    2012-01-01

    OBJECTIVE Fetal neutrophilia is present in two-thirds of cases with the fetal inflammatory response syndrome (FIRS). The mechanisms responsible for this finding have not been elucidated. Granulocyte Colony-Stimulating Factor (G-CSF) is the primary physiologic regulator of neutrophil production and plays a key role in the rapid generation and release of neutrophils in stressful conditions (i.e., infection). The objective of this study was to determine: 1) whether FIRS was associated with changes in fetal plasma G-CSF concentrations; and 2) if fetal plasma G-CSF concentrations correlated with fetal neutrophil counts, chorioamnionitis, neonatal morbidity/mortality and cordocentesis-to-delivery interval. STUDY DESIGN Percutaneous umbilical cord blood sampling was performed in a population of patients with preterm labor (n=107). A fetal plasma interleukin-6 (IL-6) concentration >11 pg/mL was used to define FIRS. Cord blood G-CSF was measured by a sensitive and specific immunoassay. An absolute neutrophil count was determined and corrected for gestational age. Receiver operating characteristic (ROC) curve, survival analysis and Cox proportional hazard model were employed. RESULTS 1) G-CSF was detected in all fetal blood samples; 2) fetuses with FIRS had a higher median fetal plasma G-CSF concentration than those without FIRS (p<0.001); 3) a fetal plasma G-CSF concentration ≥134 pg/mL (derived from an ROC curve) was associated with a shorter cordocentesis-to-delivery interval, a higher frequency of chorioamnionitis (clinical and histological), intra-amniotic infection, and composite neonatal morbidity/mortality than a fetal plasma concentration below this cut-off; and 4) a fetal plasma G-CSF concentration ≥134 pg/mL was associated with a shorter cordocentesis-to-delivery interval (hazard ratio 3.2; 95% confidence interval 1.8-5.8) after adjusting for confounders. CONCLUSIONS 1) G-CSF concentrations are higher in the peripheral blood of fetuses with FIRS than in

  19. Myelodysplastic syndrome and acute myeloid leukemia following adjuvant chemotherapy with and without granulocyte colony-stimulating factors for breast cancer.

    PubMed

    Calip, Gregory S; Malmgren, Judith A; Lee, Wan-Ju; Schwartz, Stephen M; Kaplan, Henry G

    2015-11-01

    Risk of myelodysplastic syndrome (MDS) and acute myeloid leukemia (AML) post-breast cancer treatment with adjuvant chemotherapy and granulocyte colony-stimulating factors (G-CSF) is not fully characterized. Our objective was to estimate MDS/AML risk associated with specific breast cancer treatments. We conducted a retrospective cohort study of women aged ≥66 years with stage I-III breast cancer between 2001 and 2009 using the Surveillance, Epidemiology, and End Results-Medicare database. Women were classified as receiving treatment with radiation, chemotherapy, and/or G-CSF. We used multivariable Cox proportional hazards models to estimate adjusted hazard ratios (HR) and 95 % confidence intervals (CI) for MDS/AML risk. Among 56,251 breast cancer cases, 1.2 % developed MDS/AML during median follow-up of 3.2 years. 47.1 % of women received radiation and 14.3 % received chemotherapy. Compared to breast cancer cases treated with surgery alone, those treated with chemotherapy (HR = 1.38, 95 %-CI 0.98-1.93) and chemotherapy/radiation (HR = 1.77, 95 %-CI 1.25-2.51) had increased risk of MDS/AML, but not radiation alone (HR = 1.08, 95 % CI 0.86-1.36). Among chemotherapy regimens and G-CSF, MDS/AML risk was differentially associated with anthracycline/cyclophosphamide-containing regimens (HR = 1.86, 95 %-CI 1.33-2.61) and filgrastim (HR = 1.47, 95 %-CI 1.05-2.06), but not pegfilgrastim (HR = 1.10, 95 %-CI 0.73-1.66). We observed increased MDS/AML risk among older breast cancer survivors treated with anthracycline/cyclophosphamide chemotherapy that was enhanced by G-CSF. Although small, this risk warrants consideration when determining adjuvant chemotherapy and neutropenia prophylaxis for breast cancer patients.

  20. The effects of granulocyte-macrophage colony-stimulating factor on tumour-infiltrating lymphocytes from renal cell carcinoma.

    PubMed Central

    Steger, G. G.; Kaboo, R.; deKernion, J. B.; Figlin, R.; Belldegrun, A.

    1995-01-01

    It has been shown that granulocyte-macrophage colony-stimulating factor (GM-CSF) can induce specific and non-specific anti-tumour cytotoxicity and also stimulates the proliferation and function of peripheral lymphocytes and thymocytes. GM-CSF and interleukin 2 (IL-2) act synergistically on peripheral lymphocytes for the induction of a highly effective cytotoxic cell population. Thus, the goal of our investigation was to study the effects of GM-CSF upon expansion, proliferation and in vitro killing activity of tumour-infiltrating lymphocytes (TILs) from renal cell carcinoma (RCC). TILs from seven consecutive tumours were cultured with GM-CSF (500 or 1000 nmol ml-1) without IL-2 supplementation, with suboptimal doses of IL-2 (8 and 40 U ml-1) plus GM-CSF (1000 nmol ml-1), and with a dose of IL-2 (400 U ml-1) which sufficed alone to induce TIL development plus GM-CSF (500 or 1000 nmol ml-1). GM-CSF alone or together with suboptimal doses of IL-2 was not able to induce or facilitate TIL development in these cultures. When GM-CSF at both concentrations studied was added to optimal doses of IL-2 the resulting TIL populations proliferated significantly better and faster (+66%), resulting in a higher cell yield (+24%) at the time of maximal expansion of the TIL cultures. The length of the culture periods of TILs was not affected by GM-CSF when compared with the control cultures supplemented with IL-2 alone. In vitro killing activity of TIL populations stimulated with IL-2 and GM-CSF remained unspecific, but lysis of the autologous tumour targets as well as the allogeneic renal tumour targets was significantly enhanced (+138%) as compared with the corresponding control TILs stimulated with IL-2 alone. Lysis of the natural killer (NK)-sensitive control cell line K562 and the NK-resistant Daudi cell line remained unchanged even though FACS analysis of TILs cultured with IL-2 and 1000 nmol of GM-CSF demonstrated a significantly higher proportion of cells expressing the CD56

  1. Functional granulocyte/macrophage colony stimulating factor receptor is constitutively expressed on neoplastic plasma cells and mediates tumour cell longevity.

    PubMed

    Villunger, A; Egle, A; Kos, M; Egle, D; Tinhofer, I; Henn, T; Uberall, F; Maly, K; Greil, R

    1998-09-01

    It has been shown that granulocyte/macrophage colony stimulating factor (GM-CSF) is able to support myeloma cell propagation in cooperation with interleukin (IL)-6, the major growth factor for malignant plasma cells, although the biological mechanisms involved remain unknown. Therefore we investigated (i) the expression levels of the GM-CSF receptor (GM-CSFR) constituents in three malignant plasma cell lines and in native malignant plasma cells, (ii) the ability of the receptor to mediate common signalling pathways regulating proliferation and cell survival in malignant plasma cell lines, and (iii) the effects of GM-CSF on tumour cell biology. The GM-CSFRalpha subunit was detected in the malignant plasma cell lines RPMI-8226, MC/CAR, IM-9 as well as 6/6 native myeloma cell samples derived from the bone marrow of patients with overt disease. Furthermore, GM-CSFR expression was also detected in the CD19+ fraction from 2/3 bone marrow samples and 5/8 peripheral blood samples derived from patients with malignant plasma cell disorders, but not in the CD19+ fraction of peripheral blood from healthy donors. The expressed cytokine receptor alpha-subunit was able to constitute a functional signalling complex with the ubiquitously expressed GM-CSFRbeta subunit, as demonstrated by the fact that GM-CSF induced the p21-ras/mitogen-activated protein kinase (MAPK) signalling cascade in malignant plasma cell lines. Since this signalling cascade plays an essential role in the mediation of both proliferation and cell survival, we investigated the impact of GM-CSF on these two events. Application of GM-CSF led to an increase of DNA-synthesis in MC/CAR, IM-9 and RPMI-8226 cells. Furthermore, it increased longevity of these malignant plasma cell lines by reducing the rates of spontaneous apoptosis. We conclude that (i) the functional GM-CSFR is commonly expressed on malignant plasma cells and that (ii) GM-CSF promotes the clonal expansion of myeloma cells by inhibiting spontaneous

  2. Stromal cell-derived factor-1 enhances pro-angiogenic effect of granulocyte-colony stimulating factor

    PubMed Central

    Tan, Yaohong; Shao, Hongwei; Eton, Darwin; Yang, Zhe; Alonso-Diaz, Luis; Zhang, Hongkun; Schulick, Andrew; Livingstone, Alan S.; Yu, Hong

    2008-01-01

    Objective Granulocyte colony-stimulating factor (G-CSF) mobilizes bone marrow mononuclear cells into the peripheral circulation. Stromal cell-derived factor-1 (SDF-1) enhances the homing of progenitor cells mobilized from the bone marrow and augments neovascularization in ischemic tissue. We hypothesize that SDF-1 will boost the pro-angiogenic effect of G-CSF. Methods and results NIH 3T3 cells retrovirally transduced with SDF-1α gene (NIH 3T3/SDF-1) were used to deliver SDF-1 in vitro and in vivo. Endothelial progenitor cells (EPCs) co-cultured with NIH 3T3/SDF-1 cells using cell culture inserts migrated faster and were less apoptotic compared to those not exposed to SDF-1. NIH 3T3/SDF-1 (106 cells) were injected into the ischemic muscles immediately after resection of the left femoral artery and vein of C57BL/6J mice. G-CSF (25 μg/kg/day) was injected intraperitioneally daily for 3 days after surgery. Blood perfusion was examined using a laser Doppler perfusion imaging system. The perfusion ratio of ischemic/non-ischemic limb increased to 0.57±0.03 and 0.50±0.06 with the treatment of either SDF-1 or G-CSF only, respectively, 3 weeks after surgery, which was significantly higher than the saline-injected control group (0.41±0.01, P<0.05). Combined treatment with both SDF-1 and G-CSF resulted in an even better perfusion ratio of 0.69±0.08 (P<0.05 versus the single treatment groups). Mice were sacrificed 21 days after surgery. Immunostaining and Western blot assay of the tissue lysates showed that the injected NIH 3T3/SDF-1 survived and expressed SDF-1. CD34+ cells were detected with immunostaining, capillary density was assessed with alkaline phosphatase staining, and the apoptosis of muscle cells was viewed using an in situ cell death detection kit. More CD34+ cells, increased capillary density, and less apoptotic muscle cells were found in both G-CSF and SDF-1 treated group (P<0.05 versus other groups). Conclusion Combination of G-CSF-mediated progenitor cell

  3. Effect of immobilized granulocyte colony-stimulating factor on hemopoietic precursors of various classes during cytostatic-induced myelosuppression.

    PubMed

    Dygai, A M; Skurikhin, E G; Andreeva, T V; Madonov, P G; Vereshagin, E I; Kinsht, D N; Pershina, O V; Khmelevskaya, E S

    2010-09-01

    Experiments were performed on the model of cytostatic myelosuppression induced by cyclophosphamide. We compared the effect of immobilized granulocyte CSF (the preparation was created in Russia) and reference standard preparation of granulocyte CSF on the development of neutrophilic leukopenia and hemopoietic precursors of various classes. It was found that preparations of granulocyte CSF decreased the duration and degree of peripheral blood neutropenia. The granulocytopoiesis-stimulating effect was related to stimulation of multipotent hemopoietic precursors, granulocyte-erythroid-macrophage-megakaryocyte precursors, and granulocyte precursors. Induction of division and maturation of multipotent hemopoietic precursors, granulocyte-erythroid-macrophage-megakaryocyte precursors, and granulocyte precursors and recovery of cellularity of the granulocytic hemopoietic stem after administration of immobilized granulocyte CSF were observed at later terms compared to treatment with the reference preparation of granulocyte CSF.

  4. Engineering superactive granulocyte macrophage colony-stimulating factor transferrin fusion proteins as orally-delivered candidate agents for treating neurodegenerative disease.

    PubMed

    Heinzelman, Pete; Priebe, Molly C

    2015-01-01

    Intravenously injected granulocyte macrophage colony-stimulating factor (GM-CSF) has shown efficacy in Alzheimer's Disease (AD) and Parkinson's Disease (PD) animal studies and is undergoing clinical evaluation. The likely need for dosing of GM-CSF to patients over months or years motivates pursuit of avenues for delivering GM-CSF to circulation via oral administration. Flow cytometric screening of 37 yeast-displayed GM-CSF saturation mutant libraries revealed residues P12, H15, R23, R24, and K72 as key determinants of GM-CSF's CD116 and CD131 GM-CSF receptor (GM-CSFR) subunit binding affinity. Screening combinatorial GM-CSF libraries mutated at positions P12, H15, and R23 yielded variants with increased affinities toward both CD116 and CD131. Genetic fusion of GM-CSF to human transferrin (Trf), a strategy that enables oral delivery of other biopharmaceuticals in animals, yielded bioactive wild type and variant cytokines upon secretion from cultured Human Embryonic Kidney cells. Surface plasmon resonance (SPR) measurements showed that all evaluated variants possess decreases in CD116 and CD131 binding KD values of up to 2.5-fold relative to wild type. Improved affinity led to increased in vitro bioactivity; the most bioactive variant, P12D/H15L/R23L, had a leukocyte proliferation assay EC50 value 3.5-fold lower than the wild type GM-CSF/Trf fusion. These outcomes are important first steps toward our goal of developing GM-CSF/Trf fusions as orally available AD and PD therapeutics. PMID:25737095

  5. Health, economic, and quality-of-life effects of erythropoietin and granulocyte colony-stimulating factor for the treatment of myelodysplastic syndromes: a randomized, controlled trial.

    PubMed

    Casadevall, Nicole; Durieux, Pierre; Dubois, Stéphanie; Hemery, François; Lepage, Eric; Quarré, Marie-Catherine; Damaj, Gandhi; Giraudier, Stéphane; Guerci, Agnès; Laurent, Guy; Dombret, Hervé; Chomienne, Christine; Ribrag, Vincent; Stamatoullas, Aspasia; Marie, Jean-Pierre; Vekhoff, Anne; Maloisel, Frédéric; Navarro, Robert; Dreyfus, François; Fenaux, Pierre

    2004-07-15

    In myelodysplastic syndromes (MDS), anemia responds to recombinant human erythropoietin (rHuEPO) alone and in combination with recombinant human granulocyte-colony-stimulating factor (rHuGCSF) in 10% to 20% and in 35% to 40% of patients, respectively. We randomly divided 60 patients with low-grade anemic MDS and serum EPO levels lower than 500 IU/L (500 mU/mL) into 2 groups: rHuEPO + rHuG-CSF (arm A) and supportive care (arm B). After 12 weeks, those who had erythroid responses were given rHuEPO alone for 40 additional weeks. They were also given rHuG-CSF if they had relapses. A response was considered major if the hemoglobin (Hb) level was 115 g/L (11.5 g/dL) or higher and minor Hb increase was 15 g/L (1.5 g/dL) or more or if it remained stable without transfusion. Ten of 24 patients responded in arm A, and 0 of 26 responded in arm B (P =.01). Eight patients in arm A continued rHuEPO therapy alone, and 6 had relapses. Responses were always restored when rHuG-CSF was reintroduced. Mean direct costs per patient were 26,723 euros (arm A) and 8,746 euros (arm B). Quality of life was assessed with a Functional Assessment of Cancer Therapy-Anemia (FACT-An) scale. Similar percentages of patients from both arms showed significant clinical improvement. rHuEPO plus rHuG-CSF led to responses in 41.7% of MDS patients. This treatment was expensive. No effect on quality of life was demonstrated.

  6. Granulocyte-colony stimulating factor for hematopoietic stem cell donation from healthy female donors during pregnancy and lactation: what do we know?

    PubMed

    Pessach, Ilias; Shimoni, Avichai; Nagler, Arnon

    2013-01-01

    BACKGROUND Hematopoietic growth factors (HGFs) are mostly used as supportive measures to reduce infectious complications associated with neutropenia. Over the past decade, the use of HGFs became a common method for mobilizing human CD34+ stem cells, either for autologous or allogeneic transplantation. However, since their introduction the long-term safety of the procedure has become a major focus of discussion and research. Most information refers to healthy normal donors and data concerning pregnant and lactating women are scarce. The clinical question, which is the core of this review, is whether stem cell donation, preceded by administration of granulocyte-colony stimulating factor (G-CSF) for mobilization, is a safe procedure for pregnant donors. METHODS Literature searches were performed in Pubmed for English language articles published before the end of May 2012, focusing on G-CSF administration during pregnancy, lactation and hematopoietic stem cell donation. Searches included animal and human studies. RESULTS Data from animals (n = 15 studies) and women (n = 46 studies) indicate that G-CSF crosses the placenta, stimulates fetal granulopoiesis, improves neonatal survival mostly for very immature infants, promotes trophoblast growth and placental metabolism and has an anti-abortive role. Granulocyte macrophage-CSF is a key cytokine in the maternal immune tolerance towards the implanted embryo and exerts protective long-term programming effects to preimplantation embryos. The available data suggest that probably CSFs should not be administered during the time of most active organogenesis (first trimester), except perhaps for the first week during which implantation takes place. Provided CSF is administered during the second and third trimesters, it appears to be safe, and pregnant women receiving the CSF treatment can become hematopoietic stem cell donors. There are also risks related to the anesthesia, which is required for the bone marrow aspiration. During

  7. Mobilization and collection of CD34+ cells for autologous transplantation of peripheral blood hematopoietic progenitor cells in children: analysis of two different granulocyte-colony stimulating factor doses

    PubMed Central

    Eid, Kátia Aparecida de Brito; Miranda, Eliana Cristina Martins; Aguiar, Simone dos Santos

    2015-01-01

    Introduction The use of peripheral hematopoietic progenitor cells (HPCs) is the cell choice in autologous transplantation. The classic dose of granulocyte-colony stimulating factor (G-CSF) for mobilization is a single daily dose of 10 μg/kg of patient body weight. There is a theory that higher doses of granulocyte-colony stimulating factor applied twice daily could increase the number of CD34+ cells collected in fewer leukapheresis procedures. Objective The aim of this study was to compare a fractionated dose of 15 μg G-CSF/kg of body weight and the conventional dose of granulocyte-colony stimulating factor in respect to the number of leukapheresis procedures required to achieve a minimum collection of 3 × 106 CD34+ cells/kg body weight. Methods Patients were divided into two groups: Group 10 – patients who received a single daily dose of 10 μg G-CSF/kg body weight and Group 15 – patients who received a fractioned dose of 15 μg G-CSF/kg body weight daily. The leukapheresis procedure was carried out in an automated cell separator. The autologous transplantation was carried out when a minimum number of 3 × 106 CD34+ cells/kg body weight was achieved. Results Group 10 comprised 39 patients and Group 15 comprised 26 patients. A total of 146 apheresis procedures were performed: 110 (75.3%) for Group 10 and 36 (24.7%) for Group 15. For Group 10, a median of three (range: 1–7) leukapheresis procedures and a mean of 8.89 × 106 CD34+ cells/kg body weight (±9.59) were collected whereas for Group 15 the corresponding values were one (range: 1–3) and 5.29 × 106 cells/kg body weight (±4.95). A statistically significant difference was found in relation to the number of apheresis procedures (p-value <0.0001). Conclusions To collect a minimum target of 3 × 106 CD34+ cells/kg body weight, the administration of a fractionated dose of 15 μg G-CSF/kg body weight significantly decreased the number of leukapheresis procedures performed. PMID:26041417

  8. Drugs elevating extracellular adenosine promote regeneration of haematopoietic progenitor cells in severely myelosuppressed mice: their comparison and joint effects with the granulocyte colony-stimulating factor.

    PubMed

    Hofer, Michal; Pospísil, Milan; Znojil, Vladimír; Vacek, Antonín; Weiterova, Lenka; Holá, Jirina; Vácha, Jirí

    2002-01-01

    We tested capabilities of drugs elevating extracellular adenosine and of granulocyte colony-stimulating factor (G-CSF) given alone or in combination to modulate regeneration from severe myelosuppression resulting from combined exposure of mice to ionizing radiation and carboplatin. Elevation of extracellular adenosine was induced by joint administration of dipyridamole (DP), a drug inhibiting the cellular uptake of adenosine, and adenosine monophosphate (AMP), serving as an adenosine prodrug. DP+AMP, G-CSF or all these drugs in combination were administered in a 4-d treatment regimen starting on day 3 after induction of myelosuppression. Comparable enhancements of haematopoietic regeneration due to elevation of extracellular adenosine or to action of G-CSF were demonstrated as shown by elevated numbers of haematopoietic progenitor cells for granulocytes/macrophages (GM-CFC) and erythrocytes (BFU-E) in the bone marrow and spleen in early time intervals after termination of the drug treatment, i.e. on days 7 and 10 after induction of myelosuppression. Coadministration of all the drugs further potentiated the restoration of progenitor cell pools in the haematopoietic organs. The effects of the drug treatments on progenitor cells were reflected in the peripheral blood in later time intervals of days 15 and 20 after induction of myelosuppression, especially as significantly elevated numbers of granulocytes and less pronounced elevation of lymphocytes and erythrocytes. The results substantiate the potential of drugs elevating extracellular adenosine for clinical utilization in myelosuppressive states, e.g. those accompanying oncological radio- and chemotherapy.

  9. Cell acidification in apoptosis: granulocyte colony-stimulating factor delays programmed cell death in neutrophils by up-regulating the vacuolar H(+)-ATPase.

    PubMed Central

    Gottlieb, R A; Giesing, H A; Zhu, J Y; Engler, R L; Babior, B M

    1995-01-01

    Neutrophils in tissue culture spontaneously undergo programmed cell death (apoptosis), a process characterized by well-defined morphological alterations affecting the cell nucleus. We found that these morphological changes were preceded by intracellular acidification and that acidification and the apoptotic changes in nuclear morphology were both delayed by granulocyte colony-stimulating factor (G-CSF). Among the agents that defend neutrophils against intracellular acidification is a vacuolar H(+)-ATPase that pumps protons out of the cytosol. When this proton pump was inhibited by bafilomycin A1, G-CSF no longer protected the neutrophils against apoptosis. We conclude that G-CSF delays apoptosis in neutrophils by up-regulating the cells' vacuolar H(+)-ATPase and that intracellular acidification is an early event in the apoptosis program. Images Fig. 1 Fig. 2 PMID:7541139

  10. Granulocyte colony stimulating factor (G-CSF) can allow treatment with clozapine in a patient with severe benign ethnic neutropaenia (BEN): a case report.

    PubMed

    Spencer, Benjamin W J; Williams, Hugh R J; Gee, Siobhan H; Whiskey, Eromona; Rodrigues, Joseph P; Mijovic, Aleksandar; MacCabe, James H

    2012-09-01

    Clozapine is the treatment of choice for treatment-resistant schizophrenia, but it is associated with a risk of neutropaenia and agranulocytosis. Clozapine use is regulated by mandatory blood monitoring in the UK, requiring cessation of treatment should the absolute neutrophil count (ANC) drop below specified values. Benign reductions in the ANC in non-white populations are common, and this can preclude a patient from receiving treatment with clozapine. A diagnosis of benign ethnic neutropaenia can reduce these treatment restrictions (UK specific), but the degree of neutropaenia can be significant enough to still prevent treatment. In this report, we show that response to granulocyte colony stimulating factor (G-CSF) may be quite variable and difficult to predict, but with careful monitoring it can be used to increase the ANC count and allow continued treatment with clozapine.

  11. Granulocyte colony stimulating factor (G-CSF) can allow treatment with clozapine in a patient with severe benign ethnic neutropaenia (BEN): a case report.

    PubMed

    Spencer, Benjamin W J; Williams, Hugh R J; Gee, Siobhan H; Whiskey, Eromona; Rodrigues, Joseph P; Mijovic, Aleksandar; MacCabe, James H

    2012-09-01

    Clozapine is the treatment of choice for treatment-resistant schizophrenia, but it is associated with a risk of neutropaenia and agranulocytosis. Clozapine use is regulated by mandatory blood monitoring in the UK, requiring cessation of treatment should the absolute neutrophil count (ANC) drop below specified values. Benign reductions in the ANC in non-white populations are common, and this can preclude a patient from receiving treatment with clozapine. A diagnosis of benign ethnic neutropaenia can reduce these treatment restrictions (UK specific), but the degree of neutropaenia can be significant enough to still prevent treatment. In this report, we show that response to granulocyte colony stimulating factor (G-CSF) may be quite variable and difficult to predict, but with careful monitoring it can be used to increase the ANC count and allow continued treatment with clozapine. PMID:22719015

  12. Regulation of Granulocyte Colony-Stimulating Factor and Its Receptor in Skeletal Muscle is Dependent Upon the Type of Inflammatory Stimulus.

    PubMed

    Wright, Craig Robert; Brown, Erin Louise; Della Gatta, Paul A; Fatouros, Ioannis G; Karagounis, Leonidas G; Terzis, Gerasimos; Mastorakos, Georgios; Michailidis, Yannis; Mandalidis, Dimitris; Spengos, Kontantinos; Chatzinikolaou, Athanasios; Methenitis, Spiros; Draganidis, Dimitrios; Jamurtas, Athanasios Z; Russell, Aaron Paul

    2015-09-01

    The cytokine granulocyte colony-stimulating factor (G-CSF) binds to its receptor (G-CSFR) to stimulate hematopoietic stem cell mobilization, myelopoiesis, and the production and activation of neutrophils. In response to exercise-induced muscle damage, G-CSF is increased in circulation and G-CSFR has recently been identified in skeletal muscle cells. While G-CSF/G-CSFR activation mediates pro- and anti-inflammatory responses, our understanding of the role and regulation in the muscle is limited. The aim of this study was to investigate, in vitro and in vivo, the role and regulation of G-CSF and G-CSFR in skeletal muscle under conditions of muscle inflammation and damage. First, C2C12 myotubes were treated with lipopolysaccharide (LPS) with and without G-CSF to determine if G-CSF modulates the inflammatory response. Second, the regulation of G-CSF and its receptor was measured following eccentric exercise-induced muscle damage and the expression levels we investigated for redox sensitivity by administering the antioxidant N-acetylcysteine (NAC). LPS stimulation of C2C12 myotubes resulted in increases in G-CSF, interleukin (IL)-6, monocyte chemoattractant protein-1 (MCP-1), and tumor necrosis factor-α (TNFα) messenger RNA (mRNA) and an increase in G-CSF, IL-6, and MCP-1 release from C2C12 myotubes. The addition of G-CSF following LPS stimulation of C2C12 myotubes increased IL-6 mRNA and cytokine release into the media, however it did not affect MCP-1 or TNFα. Following eccentric exercise-induced muscle damage in humans, G-CSF levels were either marginally increased in circulation or remain unaltered in skeletal muscle. Similarly, G-CSFR levels remained unchanged in response to damaging exercise and G-CSF/G-CSFR did not change in response to NAC. Collectively, these findings suggest that G-CSF may cooperate with IL-6 and potentially promote muscle regeneration in vitro, whereas in vivo aseptic inflammation induced by exercise did not change G-CSF and G

  13. Granulocyte-Macrophage Colony-Stimulating Factor Expressed by Recombinant Respiratory Syncytial Virus Attenuates Viral Replication and Increases the Level of Pulmonary Antigen-Presenting Cells

    PubMed Central

    Bukreyev, Alexander; Belyakov, Igor M.; Berzofsky, Jay A.; Murphy, Brian R.; Collins, Peter L.

    2001-01-01

    An obstacle to developing a vaccine against human respiratory syncytial virus (RSV) is that natural infection typically does not confer solid immunity to reinfection. To investigate methods to augment the immune response, recombinant RSV (rRSV) was constructed that expresses murine granulocyte-macrophage colony-stimulating factor (mGM-CSF) from a transcription cassette inserted into the G-F intergenic region. Replication of rRSV/mGM-CSF in the upper and lower respiratory tracts of BALB/c mice was reduced 23- to 74- and 5- to 588-fold, respectively, compared to that of the parental rRSV. Despite this strong attenuation of replication, the level of RSV-specific serum antibodies induced by rRSV/mGM-CSF was comparable to, or marginally higher than, that of the parental rRSV. The induction of RSV-specific CD8+ cytotoxic T cells was moderately reduced during the initial infection, which might be a consequence of reduced antigen expression. Mice infected with rRSV/mGM-CSF had elevated levels of pulmonary mRNA for gamma interferon (IFN-γ) and interleukin 12 (IL-12) p40 compared to animals infected by wild-type rRSV. Elevated synthesis of IFN-γ could account for the restriction of RSV replication, as was observed previously with an IFN-γ-expressing rRSV. The accumulation of total pulmonary mononuclear cells and total CD4+ T lymphocytes was accelerated in animals infected with rRSV/mGM-CSF compared to that in animals infected with the control virus, and the level of IFN-γ-positive or IL-4-positive pulmonary CD4+ cells was elevated approximately twofold. The number of pulmonary lymphoid and myeloid dendritic cells and macrophages was increased up to fourfold in mice infected with rRSV/mGM-CSF compared to those infected with the parental rRSV, and the mean expression of major histocompatibility complex class II molecules, a marker of activation, was significantly increased in the two subsets of dendritic cells. Enhanced antigen presentation likely accounts for the

  14. Plerixafor and granulocyte colony-stimulating factor for first-line steady-state autologous peripheral blood stem cell mobilization in lymphoma and multiple myeloma: results of the prospective PREDICT trial

    PubMed Central

    Russell, Nigel; Douglas, Kenny; Ho, Anthony D.; Mohty, Mohamad; Carlson, Kristina; Ossenkoppele, G.J.; Milone, Giuseppe; Pareja, Macarena Ortiz; Shaheen, Daniel; Willemsen, Arnold; Whitaker, Nicky; Chabannon, Christian

    2013-01-01

    In Europe, the combination of plerixafor + granulocyte colony-stimulating factor is approved for the mobilization of hematopoietic stem cells for autologous transplantation in patients with lymphoma and myeloma whose cells mobilize poorly. The purpose of this study was to further assess the safety and efficacy of plerixafor + granulocyte colony-stimulating factor for front-line mobilization in European patients with lymphoma or myeloma. In this multicenter, open label, single-arm study, patients received granulocyte colony-stimulating factor (10 μg/kg/day) subcutaneously for 4 days; on the evening of day 4 they were given plerixafor (0.24 mg/kg) subcutaneously. Patients underwent apheresis on day 5 after a morning dose of granulocyte colony-stimulating factor. The primary study objective was to confirm the safety of mobilization with plerixafor. Secondary objectives included assessment of efficacy (apheresis yield, time to engraftment). The combination of plerixafor + granulocyte colony-stimulating factor was used to mobilize hematopoietic stem cells in 118 patients (90 with myeloma, 25 with non-Hodgkin's lymphoma, 3 with Hodgkin's disease). Treatment-emergent plerixafor-related adverse events were reported in 24 patients. Most adverse events occurred within 1 hour after injection, were grade 1 or 2 in severity and included gastrointestinal disorders or injection-site reactions. The minimum cell yield (≥2×106 CD34+ cells/kg) was harvested in 98% of patients with myeloma and in 80% of those with non-Hodgkin's lymphoma in a median of one apheresis. The optimum cell dose (≥5×106 CD34+ cells/kg for non-Hodgkin's lymphoma or ≥6×106 CD34+ cells/kg for myeloma) was harvested in 89% of myeloma patients and 48% of non-Hodgkin's lymphoma patients. In this prospective, multicenter European study, mobilization with plerixafor + granulocyte colony-stimulating factor allowed the majority of patients with myeloma or non-Hodgkin's lymphoma to undergo transplantation with

  15. An early granulocyte colony-stimulating factor treatment attenuates neuropathic pain through activation of mu opioid receptors on the injured nerve

    PubMed Central

    Liao, Ming-Feng; Yeh, Shin-Rung; Lo, Ai-Lun; Chao, Po-Kuan; Lee, Yun-Lin; Hung, Yu-Hui; Lu, Kwok-Tung; Ro, Long-Sun

    2016-01-01

    Several studies have shown that the mu opioid receptor (MOR) located in the peripheral nerves can be activated after nerve injury and that it attenuates peripheral nociceptive signals to the spinal dorsal horn. Various cytokines and phosphorylated-p38 (p-p38) activation in the dorsal horn also play an important role in neuropathic pain development. Granulocyte-colony stimulating factor (GCSF) is a growth factor that can stimulate granulocyte formation and has been shown to exert an analgesic effect on neuropathic pain through recruiting opioid-containing leukocytes to the injured nerve. However, the underlying mechanisms are not well understood. Herein, the results of behavior tests in addition to MOR levels in the injured sciatic nerve and the levels of p-p38 and various cytokines in the spinal dorsal horn were studied in vehicle-treated or GCSF-treated chronic constriction injured (CCI) rats at different time points (i.e., 1, 3, and 7 days, respectively) after nerve injury. The results showed that a single early systemic GCSF treatment after nerve injury can up-regulate MORs in the injured nerve, which can decrease peripheral nociceptive signals. Thereafter, those changes suppress the pro-inflammatory cytokine IL-6 but enhance the anti-inflammatory cytokine IL-4, followed by decreases in p-p38 in the dorsal horn, and thus further attenuate neuropathic pain. PMID:27180600

  16. Synergistic action of the benzene metabolite hydroquinone on myelopoietic stimulating activity of granulocyte/macrophage colony-stimulating factor in vitro.

    PubMed Central

    Irons, R D; Stillman, W S; Colagiovanni, D B; Henry, V A

    1992-01-01

    The effects of in vitro pretreatment with benzene metabolites on colony-forming response of murine bone marrow cells stimulated with recombinant granulocyte/macrophage colony-stimulating factor (rGM-CSF) were examined. Pretreatment with hydroquinone (HQ) at concentrations ranging from picomolar to micromolar for 30 min resulted in a 1.5- to 4.6-fold enhancement in colonies formed in response to rGM-CSF that was due to an increase in granulocyte/macrophage colonies. The synergism equaled or exceeded that reported for the effects of interleukin 1, interleukin 3, or interleukin 6 with GM-CSF. Optimal enhancement was obtained with 1 microM HQ and was largely independent of the concentration of rGM-CSF. Pretreatment with other authentic benzene metabolites, phenol and catechol, and the putative metabolite trans, trans-muconaldehyde did not enhance growth factor response. Coadministration of phenol and HQ did not enhance the maximal rGM-CSF response obtained with HQ alone but shifted the optimal concentration to 100 pM. Synergism between HQ and rGM-CSF was observed with nonadherent bone marrow cells and lineage-depleted bone marrow cells, suggesting an intrinsic effect on recruitment of myeloid progenitor cells not normally responsive to rGM-CSF. Alterations in differentiation in a myeloid progenitor cell population may be of relevance in the pathogenesis of acute myelogenous leukemia secondary to drug or chemical exposure. PMID:1570288

  17. Synergistic action of the benzene metabolite hydroquinone on myelopoietic stimulating activity of granulocyte/macrophage colony-stimulating factor in vitro

    NASA Technical Reports Server (NTRS)

    Irons, R. D.; Stillman, W. S.; Colagiovanni, D. B.; Henry, V. A.; Clarkson, T. W. (Principal Investigator)

    1992-01-01

    The effects of in vitro pretreatment with benzene metabolites on colony-forming response of murine bone marrow cells stimulated with recombinant granulocyte/macrophage colony-stimulating factor (rGM-CSF) were examined. Pretreatment with hydroquinone (HQ) at concentrations ranging from picomolar to micromolar for 30 min resulted in a 1.5- to 4.6-fold enhancement in colonies formed in response to rGM-CSF that was due to an increase in granulocyte/macrophage colonies. The synergism equaled or exceeded that reported for the effects of interleukin 1, interleukin 3, or interleukin 6 with GM-CSF. Optimal enhancement was obtained with 1 microM HQ and was largely independent of the concentration of rGM-CSF. Pretreatment with other authentic benzene metabolites, phenol and catechol, and the putative metabolite trans, trans-muconaldehyde did not enhance growth factor response. Coadministration of phenol and HQ did not enhance the maximal rGM-CSF response obtained with HQ alone but shifted the optimal concentration to 100 pM. Synergism between HQ and rGM-CSF was observed with nonadherent bone marrow cells and lineage-depleted bone marrow cells, suggesting an intrinsic effect on recruitment of myeloid progenitor cells not normally responsive to rGM-CSF. Alterations in differentiation in a myeloid progenitor cell population may be of relevance in the pathogenesis of acute myelogenous leukemia secondary to drug or chemical exposure.

  18. Crystallization of a 2:2 complex of granulocyte-colony stimulating factor (GCSF) with the ligand-binding region of the GCSF receptor

    SciTech Connect

    Honjo, Eijiro; Tamada, Taro; Maeda, Yoshitake; Koshiba, Takumi; Matsukura, Yasuko; Okamoto, Tomoyuki; Ishibashi, Matsujiro; Tokunaga, Masao; Kuroki, Ryota

    2005-08-01

    A 2:2 complex of highly purified GCSF receptor (Ig-CRH) with GCSF was crystallized. The crystal diffracted to 2.8 Å resolution with sufficient quality for further structure determination. The granulocyte-colony stimulating factor (GCSF) receptor receives signals for regulating the maturation, proliferation and differentiation of the precursor cells of neutrophilic granulocytes. The signalling complex composed of two GCSFs (GCSF, 19 kDa) and two GCSF receptors (GCSFR, 34 kDa) consisting of an Ig-like domain and a cytokine-receptor homologous (CRH) domain was crystallized. A crystal of the complex was grown in 1.0 M sodium formate and 0.1 M sodium acetate pH 4.6 and belongs to space group P4{sub 1}2{sub 1}2 (or its enantiomorph P4{sub 3}2{sub 1}2), with unit-cell parameters a = b = 110.1, c = 331.8 Å. Unfortunately, this crystal form did not diffract beyond 5 Å resolution. Since the heterogeneity of GCSF receptor appeared to prevent the growth of good-quality crystals, the GCSF receptor was fractionated by anion-exchange chromatography. Crystals of the GCSF–fractionated GCSF receptor complex were grown as a new crystal form in 0.2 M ammonium phosphate. This new crystal form diffracted to beyond 3.0 Å resolution and belonged to space group P3{sub 1}21 (or its enantiomorph P3{sub 2}21), with unit-cell parameters a = b = 134.8, c = 105.7 Å.

  19. Modulation of JAK2, STAT3 and Akt1 proteins by granulocyte colony stimulating factor following carbon monoxide poisoning in male rat.

    PubMed

    Hashemzaei, Mahmoud; Imen Shahidi, Mohsen; Moallem, Seyyed Adel; Abnous, Khalil; Ghorbani, Maryam; Mohamadpour, Amir Hooshang

    2016-10-01

    Carbon monoxide (CO) is an odorless, colorless, tasteless and non-irritating by-product of inefficient combustion of hydrocarbon fuels such as motor vehicle exhausted gases. It is the leading cause of mortality in the USA among all unintentional toxicants. Male rats exposed to CO poisoning in the heart has many cardiovascular effects such as, cardiomyopathy, tachycardia, arrhythmias, and ischemia and in severe cases, myocardial infarction (MI) and cardiac arrest. Cardiomyocyte apoptosis is one of the most frequent consequences in the heart. Granulocyte colony stimulating factor (G-CSF) is a cytokine that mobilizes and differentiates granulocytes from stem cells. It can stimulate many anti-apoptotic pathways such as JAK2-STAT3 and PI3-Akt kinases following cardiac ischemia. G-CSF exerts its anti-apoptotic effects through binding to its specific cell surface receptor. The purpose of this study was to elucidate the mechanism of anti-apoptotic effect of G-CSF following CO poisoning. Rats were exposed to CO 1500 or 3000 ppm for 60 min. Animals received G-CSF 100 μg/kg subcutaneously for five consecutive days after CO intoxication. Western blot analysis was used to evaluate the expression of six proteins namely JAK2, p-JAK2, STAT3, p-STAT3, Akt1 and p-Akt1 following G-CSF 100 μg/kg consecutive dose administration after CO poisoning. There was a significant difference between phosphorylated proteins including p-JAK2, p-STAT3 and p-Akt1 in the G-CSF groups and those in control groups and there were not any significant differences in total protein among the groups. PMID:26810905

  20. Case Report: Combination Therapy with Mesenchymal Stem Cells and Granulocyte-Colony Stimulating Factor in a Case of Spinal Cord Injury

    PubMed Central

    Derakhshanrad, Nazi; Saberi, Hooshang; Tayebi Meybodi, Keyvan; Taghvaei, Mohammad; Arjmand, Babak; Aghayan, Hamid Reza; Kohan, Amir Hassan; Haghpanahi, Mohammad; Rahmani, Shahrokh

    2015-01-01

    Introduction: Various neuroregenerative procedures have been recently employed along with neurorehabilitation programs to promote neurological function after Spinal Cord Injury (SCI), and recently most of them have focused on the acute stage of spinal cord injury. In this report, we present a case of acute SCI treated with neuroprotective treatments in conjunction with conventional rehabilitation program. Methods: A case of acute penetrative SCI (gunshot wound), 40 years old, was treated with intrathecal bone marrow derived stem cells and parenteral Granulocyte-Colony Stimulating Factor (G-CSF) along with rehabilitation program. The neurological outcomes as well as safety issues have been reported. Results: Assessment with American Spinal Injury Association (ASIA), showed neurological improvement, meanwhile he reported neuropathic pain, which was amenable to oral medication. Discussion: In the acute setting, combination therapy of G-CSF and intrathecal Mesenchymal Stem Cells (MSCs) was safe in our case as an adjunct to conventional rehabilitation programs. Further controlled studies are needed to find possible side effects, and establish net efficacy. PMID:26649168

  1. Repeated courses of granulocyte colony-stimulating factor in amyotrophic lateral sclerosis: clinical and biological results from a prospective multicenter study.

    PubMed

    Chiò, Adriano; Mora, Gabriele; La Bella, Vincenzo; Caponnetto, Claudia; Mancardi, Gianluigi; Sabatelli, Mario; Siciliano, Gabriele; Silani, Vincenzo; Corbo, Massimo; Moglia, Cristina; Calvo, Andrea; Mutani, Roberto; Rutella, Sergio; Gualandi, Francesca; Melazzini, Mario; Scimè, Rosanna; Petrini, Mario; Bondesan, Paola; Garbelli, Silvia; Mantovani, Stefania; Bendotti, Caterina; Tarella, Corrado

    2011-02-01

    Granulocyte colony-stimulating factor (G-CSF) induces a transient mobilization of hematopoietic progenitor cells from bone marrow to peripheral blood. Our aim was to evaluate safety of repeated courses of G-CSF in patients with amyotrophic lateral sclerosis (ALS), assessing disease progression and changes in chemokine and cytokine levels in serum and cerebrospinal fluid (CSF). Twenty-four ALS patients entered an open-label, multicenter trial in which four courses of G-CSF and mannitol were administered at 3-month intervals. Levels of G-CSF were increased after treatment in the serum and CSF. Few and transitory adverse events were observed. No significant reduction of the mean monthly decrease in ALSFRS-R score and forced vital capacity was observed. A significant reduction in CSF levels of monocyte chemoattractant protein-1 (MCP-1) and interleukin-17 (IL-17) was observed. G-CSF treatment was safe and feasible in a multicenter series of ALS patients. A decrease in the CSF levels of proinflammatory cytokines MCP-1 and IL-17 was found, indicating a G-CSF-induced central anti-inflammatory response. PMID:21254083

  2. The alpha subunit of the granulocyte-macrophage colony-stimulating factor receptor interacts with c-Kit and inhibits c-Kit signaling.

    PubMed

    Chen, Jian; Cárcamo, Juan M; Golde, David W

    2006-08-01

    The cytokine granulocyte-macrophage colony-stimulating factor (GM-CSF) regulates hematopoiesis and the function of mature host defense cells through the GM-CSF receptor (GMR), which is composed of alpha (alphaGMR) and beta (betaGMR) subunits. Stem cell factor is another important hematopoietic cytokine that signals through c-Kit, a receptor tyrosine kinase, and regulates hematopoietic stem cell maintenance and erythroid development. Like other cytokine receptors, GMR and c-Kit are generally deemed as independent adaptor molecules capable of transducing cytokine-specific signals. We found that the alphaGMR directly interacts with c-Kit and that the interaction is mediated by the cytoplasmic domains. Furthermore, alphaGMR inhibited c-Kit auto-phosphorylation induced by the ligand stem cell factor. Consistent with the inhibitory effect, the expression of alphaGMR was suppressed in cells whose viability was dependent on c-Kit signaling. In contrast, the alternatively spliced alpha2 isoform of the alphaGMR could not inhibit c-Kit signaling, providing a rationale for the existence of the alpha2 isoform. Our results suggest that in addition to having the commonly appreciated roles in cytokine signal transduction, the receptors alphaGMR and c-Kit could interact to coordinate their signal initiation.

  3. Vaccination with Irradiated Tumor Cells Engineered to Secrete Murine Granulocyte-Macrophage Colony-Stimulating Factor Stimulates Potent, Specific, and Long-Lasting Anti-Tumor Immunity

    NASA Astrophysics Data System (ADS)

    Dranoff, Glenn; Jaffee, Elizabeth; Lazenby, Audrey; Golumbek, Paul; Levitsky, Hyam; Brose, Katja; Jackson, Valerie; Hamada, Hirofumi; Pardoll, Drew; Mulligan, Richard C.

    1993-04-01

    To compare the ability of different cytokines and other molecules to enhance the immunogenicity of tumor cells, we generated 10 retroviruses encoding potential immunomodulators and studied the vaccination properties of murine tumor cells transduced by the viruses. Using a B16 melanoma model, in which irradiated tumor cells alone do not stimulate significant anti-tumor immunity, we found that irradiated tumor cells expressing murine granulocyte-macrophage colony-stimulating factor (GM-CSF) stimulated potent, long-lasting, and specific anti-tumor immunity, requiring both CD4^+ and CD8^+ cells. Irradiated cells expressing interleukins 4 and 6 also stimulated detectable, but weaker, activity. In contrast to the B16 system, we found that in a number of other tumor models, the levels of anti-tumor immunity reported previously in cytokine gene transfer studies involving live, transduced cells could be achieved through the use of irradiated cells alone. Nevertheless, manipulation of the vaccine or challenge doses made it possible to demonstrate the activity of murine GM-CSF in those systems as well. Overall, our results have important implications for the clinical use of genetically modified tumor cells as therapeutic cancer vaccines.

  4. Granulocyte colony-stimulating factor does not enhance recruitment of bone marrow-derived cells in rats with acute myocardial infarction.

    PubMed

    Sato, Daisuke; Otani, Hajime; Fujita, Masanori; Shimazu, Takayuki; Yoshioka, Kei; Enoki, Chiharu; Minato, Naoki; Iwasaka, Toshiji

    2012-09-01

    Despite the potential benefit of granulocyte colony-stimulating factor (G-CSF) therapy in patients with acute myocardial infarction (MI), the efficacy of G-CSF in regenerating the heart after MI remains controversial. The authors hypothesize that the limited efficacy of G-CSF is related to its inhibitory effect on recruitment of bone marrow-derived cells (BMCs) to the infarcted tissue. MI was induced in rats with intrabone marrow-bone marrow transplantation from syngenic rats expressing green fluorescence protein to track BMCs. G-CSF was administered for five days after the onset of MI. G-CSF increased the number of CD45(+) cells in the peripheral circulation but did not increase their recruitment to the heart. G-CSF had no effect on myocardial stromal-derived factor-1 alpha and chemokine (C-X-C motif) receptor 4 (CXCR4) expression in mononuclear cells in the peripheral blood and CXCR4(+) cells in the heart. G-CSF had no effect on angiogenesis, myocardial fibrosis or left ventricular function four weeks after MI. These results suggest that G-CSF mobilizes BMCs to the peripheral circulation but does not increase recruitment to the infarcted myocardium despite preservation of the stromal-derived factor-1 alpha/CXCR4 axis. PMID:23620693

  5. Role of Granulocyte-Macrophage Colony-Stimulating Factor Signaling in Regulating Neutrophil Antifungal Activity and the Oxidative Burst During Respiratory Fungal Challenge.

    PubMed

    Kasahara, Shinji; Jhingran, Anupam; Dhingra, Sourabh; Salem, Anand; Cramer, Robert A; Hohl, Tobias M

    2016-04-15

    Granulocyte-macrophage colony-stimulating factor (GM-CSF) is a pleiotropic cytokine that plays a critical role in regulating myeloid cell host defense. In this study, we demonstrated that GM-CSF signaling plays an essential role in antifungal defense against Aspergillus fumigatus. Mice that lack the GM-CSF receptor β chain (GM-CSFRβ) developed invasive hyphal growth and exhibited impaired survival after pulmonary challenge with A. fumigatus conidia. GM-CSFRβ signaling regulated the recruitment of inflammatory monocytes to infected lungs, but not the recruitment of effector neutrophils. Cell-intrinsic GM-CSFRβ signaling mediated neutrophil and inflammatory monocyte antifungal activity, because lung GM-CSFRβ(-/-) leukocytes exhibited impaired conidial killing compared with GM-CSFRβ(+/+) counterparts in mixed bone marrow chimeric mice. GM-CSFRβ(-/-) neutrophils exhibited reduced (hydrogenated) nicotinamide adenine dinucleotide phosphate (NADPH) oxidase activity in vivo. Conversely, administration of recombinant GM-CSF enhanced neutrophil NADPH oxidase function, conidiacidal activity, and lung fungal clearance in A. fumigatus-challenged mice. Thus, our study illustrates the functional role of GM-CSFRβ signaling on lung myeloid cell responses against inhaled A. fumigatus conidia and demonstrates a benefit for systemic GM-CSF administration. PMID:26908736

  6. Unrelated donor granulocyte colony-stimulating factor-mobilized peripheral blood mononuclear cell transplantation after nonmyeloablative conditioning: the effect of postgrafting mycophenolate mofetil dosing.

    PubMed

    Maris, Michael B; Sandmaier, Brenda M; Storer, Barry E; Maloney, David G; Shizuru, Judith A; Agura, Edward; Kliem, Constanze; Pulsipher, Michael; Maziarz, Richard T; McSweeney, Peter A; Wade, James; Langston, Amelia A; Chauncey, Thomas R; Bruno, Benedetto; Blume, Karl G; Storb, Rainer

    2006-04-01

    We previously reported results in 71 patients with advanced hematologic malignancies given HLA-matched unrelated granulocyte colony-stimulating factor-mobilized peripheral blood mononuclear cell (G-PBMC) grafts after fludarabine 90 mg/m(2), 2 Gy of total body irradiation, and postgrafting mycophenolate mofetil (MMF) 15 mg/kg twice daily and cyclosporine 6.25 mg/kg twice daily orally. Graft rejection was 15%; the cumulative probability of acute graft-versus-host disease (GVHD) was 52%. According to MMF pharmacokinetic studies, which showed a short half-life of its active metabolite, mycophenolic acid, we increased MMF dosing from 15 mg/kg twice daily to 15 mg/kg 3 times daily to increase immunosuppression and reduce the incidence of both graft rejection and acute GVHD. Among 103 patients so treated, graft rejection occurred in 5%, whereas acute GVHD remained at 53%. Outcomes were compared with results of previous G-PBMC recipients given MMF twice daily. Infection rates were slightly higher with MMF 3 times daily than with MMF twice daily. Nevertheless, 2-year nonrelapse mortality and overall and progression-free survivals were similar for MMF 3-times-daily and twice-daily patients (19%, 58%, and 49% versus 20%, 48%, and 37%, respectively). Nonmyeloablative conditioning with postgrafting cyclosporine and MMF given 3 times daily allowed 95% durable engraftment of unrelated donor G-PBMC grafts.

  7. Effect of granulocyte colony-stimulating factor priming combined with low-dose cytarabine and homoharringtonine in higher risk myelodysplastic syndrome patients.

    PubMed

    Wang, Fang-Xia; Zhang, Wang-Gang; He, Ai-Li; Cao, Xin-Mei; Chen, Yin-Xia; Zhao, Wan-Hong; Yang, Yun; Wang, Jian-Li; Zhang, Peng-Yu; Gu, Liu-Fang

    2016-09-01

    As sensitization of leukemia cells with granulocyte colony-stimulating factor (G-CSF) can enhance the cytotoxicity of chemotherapy in myeloid malignancies, a pilot study was conducted in order to evaluate the effect of G-CSF priming combined with low-dose chemotherapy in patients with higher risk myelodysplastic syndrome (MDS). The regimen, G-HA, consisted of cytarabine (Ara-C) 7.5mg/m(2)/12h by subcutaneous injection, days 1-14, homoharringtonine (HHT) 1.5mg/m(2)/day by intravenous continuous infusion, days 1-14, and G-CSF 150mg/m(2)/day by subcutaneous injection, days 0-14. 56 patients were enrolled, 34 patients (61%, 95% confidence interval: 51.44-70.56%) achieved complete remission (CR). Median duration of neutropenia was 7days (ranging from 2 to 16days). Grade 1-2 nonhematologic toxicities were documented, including nausea and vomiting (5%), liver function abnormality (5%), and heart function abnormality (2%). No central nervous system toxicity was found. Mortality within the first 4 weeks was 4%. The G-HA regimen is effective in remission induction for higher risk MDS patients and well tolerated due to the acceptable toxicity in maintenance therapy in the patients who cannot undergo Hematopoietic cell transplantation (HCT). PMID:27497340

  8. Tachyplesin III and granulocyte-colony stimulating factor enhance the efficacy of tazobactam/piperacillin in a neutropenic mouse model of polymicrobial peritonitis.

    PubMed

    Cirioni, Oscar; Ghiselli, Roberto; Kamysz, Wojciech; Orlando, Fiorenza; Silvestri, Carmela; Mocchegiani, Federico; Di Matteo, Fabio; Kamysz, Elzbieta; Riva, Alessandra; Rocchi, Marco; Saba, Vittorio; Scalise, Giorgio; Giacometti, Andrea

    2008-01-01

    We investigated the efficacy of tazobactam/piperacillin (TZP), tachyplesin III and granulocyte-colony stimulating factor (G-CSF) in an experimental murine neutropenic intraabdominal infection. BALB/c male mice were rendered neutropenic by intraperitoneal administration of cyclophosphamide on days -4 and -2 pre-infection. Septic shock was induced by cecal ligation and puncture. Animals received intravenously isotonic sodium chloride solution (control group C1), 1mg/kg of tachyplesin III, 120 mg/kg of TZP, 0.1mg/kg of G-CSF, tachyplesin III plus TZP, G-CSF plus TZP and finally tachyplesin III plus G-CSF plus TZP, respectively. Lethality, bacterial growth in blood, peritoneum, spleen, liver, and mesenteric lymph nodes, endotoxin, IL-6 and TNF-alpha concentrations in plasma were evaluated. All compounds reduced the lethality when compared to controls. Endotoxin and cytokine plasma levels were significantly higher in TZP-treated animals compared to tachyplesin III-treated animals. Finally, all drug combinations showed to be the most effective treatment in reducing all variables measured. Interestingly, the strongest results concerning the bacterial growth inhibition, lethality and endotoxemia were obtained when the three compounds were contemporaneously administered. The presence of their positive interaction makes tachyplesin III and G-CSF potentially valuable as an adjuvant for antimicrobial chemotherapy of sepsis.

  9. Immunomodulation Induced by Stem Cell Mobilization and Harvesting in Healthy Donors: Increased Systemic Osteopontin Levels after Treatment with Granulocyte Colony-Stimulating Factor

    PubMed Central

    Melve, Guro Kristin; Ersvaer, Elisabeth; Akkök, Çiğdem Akalın; Ahmed, Aymen Bushra; Kristoffersen, Einar K.; Hervig, Tor; Bruserud, Øystein

    2016-01-01

    Peripheral blood stem cells from healthy donors mobilized by granulocyte colony-stimulating factor (G-CSF) and harvested by leukapheresis are commonly used for allogeneic stem cell transplantation. The frequency of severe graft versus host disease is similar for patients receiving peripheral blood and bone marrow allografts, even though the blood grafts contain more T cells, indicating mobilization-related immunoregulatory effects. The regulatory phosphoprotein osteopontin was quantified in plasma samples from healthy donors before G-CSF treatment, after four days of treatment immediately before and after leukapheresis, and 18–24 h after apheresis. Myeloma patients received chemotherapy, combined with G-CSF, for stem cell mobilization and plasma samples were prepared immediately before, immediately after, and 18–24 h after leukapheresis. G-CSF treatment of healthy stem cell donors increased plasma osteopontin levels, and a further increase was seen immediately after leukapheresis. The pre-apheresis levels were also increased in myeloma patients compared to healthy individuals. Finally, in vivo G-CSF exposure did not alter T cell expression of osteopontin ligand CD44, and in vitro osteopontin exposure induced only small increases in anti-CD3- and anti-CD28-stimulated T cell proliferation. G-CSF treatment, followed by leukapheresis, can increase systemic osteopontin levels, and this effect may contribute to the immunomodulatory effects of G-CSF treatment. PMID:27447610

  10. Granulocyte Colony-Stimulating Factor for Amyotrophic Lateral Sclerosis: A Randomized, Double-Blind, Placebo-Controlled Study of Iranian Patients

    PubMed Central

    Amirzagar, Nasibeh; Nafissi, Shahriar; Tafakhori, Abbas; Modabbernia, Amirhossein; Amirzargar, Aliakbar; Ghaffarpour, Majid; Siroos, Bahaddin

    2015-01-01

    Background and Purpose The aim of this study was to determine the efficacy and tolerability of granulocyte colony-stimulating factor (G-CSF) in subjects with amyotrophic lateral sclerosis (ALS). Methods Forty subjects with ALS were randomly assigned to two groups, which received either subcutaneous G-CSF (5 µg/kg/q12h) or placebo for 5 days. The subjects were then followed up for 3 months using the ALS Functional Rating Scale-Revised (ALSFRS-R), manual muscle testing, ALS Assessment Questionnaire-40, and nerve conduction studies. CD34+/CD133+ cell count and monocyte chemoattractant protein-1 (MCP-1) levels were evaluated at baseline. Results The rate of disease progression did not differ significantly between the two groups. The reduction in ALSFRS-R scores was greater in female subjects in the G-CSF group than in their counterparts in the placebo group. There was a trend toward a positive correlation between baseline CSF MCP-1 levels and the change in ALSFRS-R scores in both groups (Spearman's ρ=0.370, p=0.070). Conclusions With the protocol implemented in this study, G-CSF is not a promising option for the treatment of ALS. Furthermore, it may accelerate disease progression in females. PMID:25851895

  11. Vaccination with irradiated tumor cells engineered to secrete murine granulocyte-macrophage colony-stimulating factor stimulates potent, specific, and long-lasting anti-tumor immunity.

    PubMed Central

    Dranoff, G; Jaffee, E; Lazenby, A; Golumbek, P; Levitsky, H; Brose, K; Jackson, V; Hamada, H; Pardoll, D; Mulligan, R C

    1993-01-01

    To compare the ability of different cytokines and other molecules to enhance the immunogenicity of tumor cells, we generated 10 retroviruses encoding potential immunomodulators and studied the vaccination properties of murine tumor cells transduced by the viruses. Using a B16 melanoma model, in which irradiated tumor cells alone do not stimulate significant anti-tumor immunity, we found that irradiated tumor cells expressing murine granulocyte-macrophage colony-stimulating factor (GM-CSF) stimulated potent, long-lasting, and specific anti-tumor immunity, requiring both CD4+ and CD8+ cells. Irradiated cells expressing interleukins 4 and 6 also stimulated detectable, but weaker, activity. In contrast to the B16 system, we found that in a number of other tumor models, the levels of anti-tumor immunity reported previously in cytokine gene transfer studies involving live, transduced cells could be achieved through the use of irradiated cells alone. Nevertheless, manipulation of the vaccine or challenge doses made it possible to demonstrate the activity of murine GM-CSF in those systems as well. Overall, our results have important implications for the clinical use of genetically modified tumor cells as therapeutic cancer vaccines. PMID:8097319

  12. Evaluating the effects of buffer conditions and extremolytes on thermostability of granulocyte colony-stimulating factor using high-throughput screening combined with design of experiments.

    PubMed

    Ablinger, Elisabeth; Hellweger, Monika; Leitgeb, Stefan; Zimmer, Andreas

    2012-10-15

    In this study, we combined a high-throughput screening method, differential scanning fluorimetry (DSF), with design of experiments (DoE) methodology to evaluate the effects of several formulation components on the thermostability of granulocyte colony stimulating factor (G-CSF). First we performed a primary buffer screening where we tested thermal stability of G-CSF in different buffers, pH values and buffer concentrations. The significance of each factor and the two-way interactions between them were studied by multivariable regression analysis. pH was identified as most critical factor regarding thermal stability. The most stabilizing buffer, sodium glutamate, and sodium acetate were determined for further investigations. Second we tested the effect of 6 naturally occurring extremolytes (trehalose, sucrose, ectoine, hydroxyectoine, sorbitol, mannitol) on the thermal stability of G-CSF, using a central composite circumscribed design. At low pH (3.8) and low buffer concentration (5 mM) all extremolytes led to a significant increase in thermal stability except the addition of ectoine which resulted in a strong destabilization of G-CSF. Increasing pH and buffer concentration led to an increase in thermal stability with all investigated extremolytes. The described systematic approach allowed to create a ranking of stabilizing extremolytes at different buffer conditions.

  13. Effect of Periodic Granulocyte Colony-Stimulating Factor Administration on Endothelial Progenitor Cells and Different Monocyte Subsets in Pediatric Patients with Muscular Dystrophies

    PubMed Central

    Sienkiewicz, Dorota; Grubczak, Kamil; Okurowska-Zawada, Bożena; Paszko-Patej, Grażyna; Miklasz, Paula; Singh, Paulina; Radzikowska, Urszula; Kulak, Wojciech

    2016-01-01

    Muscular dystrophies (MD) are heterogeneous group of diseases characterized by progressive muscle dysfunction. There is a large body of evidence indicating that angiogenesis is impaired in muscles of MD patients. Therefore, induction of dystrophic muscle revascularization should become a novel approach aimed at diminishing the extent of myocyte damage. Recently, we and others demonstrated that administration of granulocyte colony-stimulating factor (G-CSF) resulted in clinical improvement of patients with neuromuscular disorders. To date, however, the exact mechanisms underlying these beneficial effects of G-CSF have not been fully understood. Here we used flow cytometry to quantitate numbers of CD34+ cells, endothelial progenitor cells, and different monocyte subsets in peripheral blood of pediatric MD patients treated with repetitive courses of G-CSF administration. We showed that repetitive cycles of G-CSF administration induced efficient mobilization of above-mentioned cells including cells with proangiogenic potential. These findings contribute to better understanding the beneficial clinical effects of G-CSF in pediatric MD patients. PMID:26770204

  14. Pretransplant Mobilization with Granulocyte Colony-Stimulating Factor Improves B-Cell Reconstitution by Lentiviral Vector Gene Therapy in SCID-X1 Mice

    PubMed Central

    Huston, Marshall W.; Riegman, Adriaan R.A.; Yadak, Rana; van Helsdingen, Yvette; de Boer, Helen; van Til, Niek P.

    2014-01-01

    Abstract Hematopoietic stem cell (HSC) gene therapy is a demonstrated effective treatment for X-linked severe combined immunodeficiency (SCID-X1), but B-cell reconstitution and function has been deficient in many of the gene therapy treated patients. Cytoreductive preconditioning is known to improve HSC engraftment, but in general it is not considered for SCID-X1 since the poor health of most of these patients at diagnosis and the risk of toxicity preclude the conditioning used in standard bone marrow stem cell transplantation. We hypothesized that mobilization of HSC by granulocyte colony-stimulating factor (G-CSF) should create temporary space in bone marrow niches to improve engraftment and thereby B-cell reconstitution. In the present pilot study supplementing our earlier preclinical evaluation (Huston et al., 2011), Il2rg−/− mice pretreated with G-CSF were transplanted with wild-type lineage negative (Lin−) cells or Il2rg−/− Lin− cells transduced with therapeutic IL2RG lentiviral vectors. Mice were monitored for reconstitution of lymphocyte populations, level of donor cell chimerism, and antibody responses as compared to 2 Gy total body irradiation (TBI), previously found effective in promoting B-cell reconstitution. The results demonstrate that G-CSF promotes B-cell reconstitution similar to low-dose TBI and provides proof of principle for an alternative approach to improve efficacy of gene therapy in SCID patients without adverse effects associated with cytoreductive conditioning. PMID:25222508

  15. Granulocyte colony-stimulating factor receptor signalling via Janus kinase 2/signal transducer and activator of transcription 3 in ovarian cancer

    PubMed Central

    Kumar, J; Fraser, F W; Riley, C; Ahmed, N; McCulloch, D R; Ward, A C

    2014-01-01

    Background: Ovarian cancer remains a major cause of cancer mortality in women, with only limited understanding of disease aetiology at the molecular level. Granulocyte colony-stimulating factor (G-CSF) is a key regulator of both normal and emergency haematopoiesis, and is used clinically to aid haematopoietic recovery following ablative therapies for a variety of solid tumours including ovarian cancer. Methods: The expression of G-CSF and its receptor, G-CSFR, was examined in primary ovarian cancer samples and a panel of ovarian cancer cell lines, and the effects of G-CSF treatment on proliferation, migration and survival were determined. Results: G-CSFR was predominantly expressed in high-grade serous ovarian epithelial tumour samples and a subset of ovarian cancer cell lines. Stimulation of G-CSFR-expressing ovarian epithelial cancer cells with G-CSF led to increased migration and survival, including against chemotherapy-induced apoptosis. The effects of G-CSF were mediated by signalling via the downstream JAK2/STAT3 pathway. Conclusion: This study suggests that G-CSF has the potential to impact on ovarian cancer pathogenesis, and that G-CSFR expression status should be considered in determining appropriate therapy. PMID:24220695

  16. Granulocyte colony-stimulating factor inhibits Fas-triggered apoptosis in bone marrow cells isolated from patients with refractory anemia with ringed sideroblasts.

    PubMed

    Schmidt-Mende, J; Tehranchi, R; Forsblom, A M; Joseph, B; Christensson, B; Fadeel, B; Zhivotovsky, B; Hellström-Lindberg, E

    2001-05-01

    Treatment with granulocyte colony-stimulating factor (G-CSF) plus erythropoietin may synergistically improve hemoglobin levels and reduce bone marrow apoptosis in patients with refractory anemia with ringed sideroblasts (RARS). Fas-induced caspase activity is increased in RARS bone marrow cells. We showed that G-CSF significantly reduced Fas-mediated caspase-8 and caspase-3-like activity and the degree of nuclear apoptotic changes in bone marrow from nine RARS patients. A decrease in mitochondrial membrane potential and an increase in intracellular reactive oxygen species occurred in Fas-treated cells, but became significant only 24 h after changes in caspase activity and decrease in proliferation. G-CSF also reduced the magnitude of these late apoptotic changes. In CD34-selected normal cells, G-CSF induced myeloid colony growth, and an overall small decrease in the number of erythroid colonies. By contrast, G-CSF induced a 33-263% increase of erythroid colony formation in CD34+ cells from four of five RARS patients with severely reduced erythroid growth, while the normal or slightly reduced erythroid growth of three other patients was not influenced by G-CSF. This study suggests that G-CSF may reduce the pathologically increased caspase activity and concomitant apoptotic changes, and promote erythroid growth and differentiation of stem cells from RARS patients. Our data support the clinical benefit of G-CSF in this subgroup of myelodysplastic syndromes.

  17. Granulocyte colony stimulating factor permits dose intensification by interval compression in the treatment of Ewing's sarcomas and soft tissue sarcomas in children.

    PubMed

    Womer, R B; Daller, R T; Fenton, J G; Miser, J S

    2000-01-01

    71 children with sarcomas were treated in a prospective pilot study to determine whether granulocyte colony stimulating factor (G-CSF) permits compression of the interval between chemotherapy cycles. Patients had Ewing's sarcoma/primitive neuroectodermal tumour (PNET), rhabdomyosarcoma, non-rhabdo soft tissue sarcomas or other advanced soft tissue tumours. The chemotherapy alternated vincristine-doxorubicin-cyclophosphamide and ifosfamide-etoposide, with G-CSF between courses. Therapy had two phases: induction (six cycles) and continuation (six cycles), which included primary tumour treatment with surgery and/or radiation. Chemotherapy cycles began every 14 days, or upon absolute neutrophil count (ANC) and platelet count recovery. The median chemotherapy cycle interval was 16 (11-48) days in the induction phase, with a median average relative dose intensification (ARDI) of 1.27 compared with every-21-day therapy. In the continuation phase, the median cycle interval was 21 days, with a median ARDI of 1.10. Radiation therapy prolonged chemotherapy intervals, whilst erythropoietin shortened them. Toxicity was modest for such chemotherapy. Event-free survival is comparable with or superior to that in recent large studies. G-CSF permits intensification of this regimen through interval compression. The impact of this approach on efficacy remains to be determined in a randomised trial. PMID:10741300

  18. [Two cases of malignant lymphoma with high fever and C-reactive protein (CRP) elevation after treatment with granulocyte colony-stimulating factor (G-CSF)].

    PubMed

    Kanbayashi, Yuko; Mukoyama, Naoki; Nishida, Katsuji; Shimizu, Daisuke; Matsumoto, Yosuke; Nomura, Kenichi; Horiike, Shigeo; Taniwaki, Masafumi

    2006-01-01

    We present two cases of malignant lymphoma that developed a high fever that eventually reached an extremely high 38.9 degrees C. The C-reactive protein( CRP) elevation also climbed to a very high 12.2 mg/dl after treatment with granulocyte colony-stimulating factor (G-CSF). In the one case, after stem cells were mobilized with CHASER therapy (cyclophosphamide, cytarabine, etoposide, dexamethasone and rituximab) followed by G-CSF (filgrastim 600 microg/day) subcutaneous daily, the serum CRP level rose to a maximum of 5.6 mg/dl, with a maximum fever elevation of 38.9 degrees C. In the other case, after the subject was given CHASE therapy followed by subcutaneous treatment with G-CSF (filgrastim 75 microg/day) daily, the maximum serum CRP level was 12.2 mg/dl along with a maximum fever of 38.9 degrees C. Although no infection was found in either case, multiple antibacterial agents were ineffective;after discontinuation of G-CSF, fever dissipated and the serum CRP level became negative. G-CSF induces the proliferation and differentiation of neutrophils and also causes the mobilization of mature neutrophils from hematopoietic tissues. With the increasing propensity to G-CSF, we must keep in mind the possibility of such adverse reactions so as to serve the overall best interests of the patient.

  19. Induction of Specific Cellular and Humoral Responses against Renal Cell Carcinoma after Combination Therapy with Cryoablation and Granulocyte-Macrophage Colony Stimulating Factor: A Pilot Study

    PubMed Central

    Thakur, Archana; Littrup, Peter; Paul, Elyse N.; Adam, Barbara; Heilbrun, Lance K.; Lum, Lawrence G.

    2013-01-01

    Cryotherapy offers a minimally invasive treatment option for the management of both irresectable and localized prostate, liver, pulmonary and renal tumors. The anti-neoplastic effects of cryotherapy are mediated by direct tumor lysis and by indirect effects such as intracellular dehydration, pH changes, and microvascular damage resulting in ischemic necrosis. In this study, we investigated whether percutaneous cryoablation of lung metastasis from renal cell carcinoma (RCC) in combination with aerosolized granulocyte-macrophage colony stimulating factor (GM-CSF) can induce systemic cellular and humoral immune responses in 6 RCC patients. Peripheral blood mononuclear cells (PBMC) were sequentially studied up to 63 days post cryoimmunotherapy (CI). PBMC from pre and post CI were phenotyped for lymphocyte subsets and tested for cytotoxicity and IFNγ Elispots directed at RCC cells. Humoral responses were measured by in vitro antibody synthesis assay directed at RCC cells. The immune monitoring data showed that CI induced tumor specific CTL, specific in vitro anti-tumor antibody responses, and enhanced Th1 cytokine production in 4 out of 6 patients. More importantly, the magnitude of cellular and humoral anti-tumor response appears to be associated with clinical responses. These pilot data show that CI can induce robust and brisk cellular and humoral immune responses in metastatic RCC patients, but requires further evaluation in optimized protocols. PMID:21577139

  20. Therapeutic efficacy of T cells derived from lymph nodes draining a poorly immunogenic tumor transduced to secrete granulocyte-macrophage colony-stimulating factor.

    PubMed

    Arca, M J; Krauss, J C; Aruga, A; Cameron, M J; Shu, S; Chang, A E

    1996-01-01

    We examined the host immune response to the poorly immunogenic B16-BL6 melanoma, which was transduced to secrete granulocyte-macrophage colony-stimulating factor (GM-CSF) (450 ng/10(6)/24 h). Tumor growth after subcutaneous inoculation was not significantly altered, although an influx of neutrophils and monocytes/macrophages was evident within tumors and draining lymph nodes (LNs). Immunization with irradiated transduced cells did not induce systemic immunity to the parental tumor. However, vaccination with transduced tumors significantly augmented in vivo sensitization of draining LN cells. These tumor-draining LN (TDLN) cells, when secondarily stimulated in vitro with anti-CD3 monoclonal antibodies and expanded in interleukin-2 (10 U/ml), exhibited greater release of GM-CST and interferon-gamma against tumor compared with TDLN cells from animals with parental tumor. In adoptive immunotherapy, activated LN cells draining transduced tumors mediated significant reductions of the numbers of established pulmonary metastases compared with LN cells draining parental tumor, which were ineffective. In addition, the therapeutic efficacy of LN cells draining transduced tumors was significantly better than LN cells primed in vivo with tumor cells admixed with Corynebacterium parvum, which we have previously described as an approach to generate immune cells. Thus, GM-CSF appears to be an important adjuvant in the induction of tumor immunity. PMID:8785710

  1. Immune responses of two recombinant adenoviruses expressing VP1 antigens of FMDV fused with porcine granulocyte macrophage colony-stimulating factor.

    PubMed

    Du, Yijun; Jiang, Ping; Li, Yufeng; He, Hairong; Jiang, Wenming; Wang, Xinglong; Hong, Weibin

    2007-11-28

    Foot-and-mouth disease (FMD) is a highly contagious and economically devastating vesicular disease of cloven-hoofed animals. In the present report, we constructed and characterized the immune responses conferred by two recombinant adenoviruses expressing VP1 epitopes (three amino acid residues 21-60, 141-160 and 200-213 in VP1, designated VPe) or VP1 protein of FMDV fused with porcine granulocyte macrophage colony-stimulating factor (named rAd-GMCSF-VPe and rAd-GMCSF-VP1). Seven groups of female BALB/c mice each containing 18 mice were inoculated subcutaneously twice at 2-week intervals with the recombinant adenoviruses. Then the protective efficacy of the two adenoviruses was detected in guinea pigs and swine. The results showed that the highest levels of FMDV-specific T cell proliferation, IFN-gamma and IL-4 could be induced by rAd-GMCSF-VPe expressing fusion GMCSF-VPe, and the highest level of FMDV-specific humoral immune responses could be induced by rAd-GMCSF-VP1 expressing fusion GMCSF-VP1 in mice. All guinea pigs and swine co-administrated with rAd-GMCSF-VPe and rAd-GMCSF-VP1 were protected from viral challenge, even though the neutralizing antibody titers were significantly lower than those in the group inoculated with inactivated FMD vaccine. It demonstrated that co-administration of rAd-GMCSF-VPe and rAd-GMCSF-VP1 might be attractive candidate vaccines for preventing FMDV infection. PMID:17980939

  2. Effect of a long-term treatment with a low-dose granulocyte colony-stimulating factor on post-infarction process in the heart

    PubMed Central

    Okada, Hideshi; Takemura, Genzou; Li, Yiwen; Ohno, Takamasa; Li, Longhu; Maruyama, Rumi; Esaki, Masayasu; Miyata, Shusaku; Kanamori, Hiromitsu; Ogino, Atsushi; Nakagawa, Munehiro; Minatoguchi, Shinya; Fujiwara, Takako; Fujiwara, Hisayoshi

    2008-01-01

    Although beneficial effects of granulocyte colony-stimulating factor (G-CSF) have been demonstrated on post-myocardia infarction (MI) process, the mechanisms and feasibility are not fully agreed yet. We investigated effects of a long-term treatment with a low-dose G-CSF started 1 day after the onset of MI, on post-infarction process. One day after being made MI by left coronary ligation, mice were given G-CSF (10 μg/kg/day) for 4 weeks. The G-CSF treatment resulted in a significant mitigation of cardiac remodelling and dysfunction. In the G-CSF-treated hearts, the infarcted scar was smaller with less fibrosis and abundant vessels while in the non-infarcted area, hypertrophic cardiomyocytes with attenuated degenerative changes and reduced fibrosis were apparent. These effects were accompanied by activation of signal transducer and activator of transcription 3 (STAT3) and Akt and also by up-regulation of GATA-4, myosin heavy chain and matrix metalloproteinases-2 and -9. Apoptosis of cardiomyocytes appeared insignificant at any stages. Parthenolide, a STAT3 inhibitor, completely abolished the beneficial effects of G-CSF on cardiac function and remodelling with loss of effect on both anti-cardiomyocyte degeneration and anti-fibrosis. In contrast, wortmannin, an Akt inhibitor, did not affect G-CSF-induced benefis despite cancelling vessel increase. In conclusion, treatment with G-CSF at a small dose but for a long duration beneficially affects the post-infarction process possibly through STAT3-mediated anti-cardiomyocyte degeneration and anti-fibrosis, but not through anti-cardiomyocyte apoptosis or Akt-mediated angio-genesis. The findings may also imply a more feasible way of G-CSF administration in the clinical settings. PMID:18298650

  3. Plerixafor on-demand combined with chemotherapy and granulocyte colony-stimulating factor: significant improvement in peripheral blood stem cells mobilization and harvest with no increase in costs.

    PubMed

    Milone, Giuseppe; Martino, Massimo; Spadaro, Andrea; Leotta, Salvatore; Di Marco, Annalia; Scalzulli, Potito; Cupri, Alessandra; Di Martina, Valentina; Schinocca, Elena; Spina, Eleonora; Tripepi, Giovanni

    2014-01-01

    To date, no prospective study on Plerixafor 'on-demand' in combination with chemotherapy and granulocyte colony-stimulating factor (G-CSF) has been reported. We present an interim analysis of the first prospective study in which Plerixafor was administered on-demand in patients affected by multiple myeloma and lymphoma who received high dose cyclophosphamide or DHAP (dexamethasone, cytarabine, cisplatin) plus G-CSF to mobilize peripheral blood stem cells (PBSC). One hundred and two patients were evaluable for response. A cohort of 240 patients receiving the same mobilizing chemotherapy was retrospectively studied. Failure to mobilize CD34(+) cells in peripheral blood was reduced by 'on-demand' strategy compared to conventional mobilization; from 13·0 to 3·0% (P = 0·004). Failure to harvest CD34(+) cells 2 × 10(6) /kg decreased from 20·9 to 4·0% (P = 0·0001). The on-demand Plerixafor strategy also resulted in a lower rate of mobilization failure (P = 0·03) and harvest failure (P = 0·0008) when compared to a 'bias-adjusted set of controls'. Evaluation of economic costs of the two strategies showed that the overall cost of the two treatments were comparable when salvage mobilizations were taken into account. When in combination with cyclophosphamide or DHAP plus G-CSF, the 'on-demand' use of Plerixafor showed, in comparison to conventionally treated patients, a significant improvement in mobilization of PBSC with no increase in overall cost. PMID:24138497

  4. Increased Prevalence of Luminal Narrowing and Stricturing Identified by Enterography in Pediatric Crohn Disease Patients with Elevated Granulocyte-Macrophage Colony Stimulating Factor Auto-antibodies

    PubMed Central

    Dykes, Dana M.H.; Towbin, Alexander J.; Bonkowski, Erin; Chalk, Claudia; Bezold, Ramona; Lake, Kathleen; Kim, Mi-Ok; Heubi, James E.; Trapnell, Bruce C.; Podberesky, Daniel J.; Denson, Lee A.

    2013-01-01

    Background Crohn disease (CD) patients with elevated Granulocyte-Macrophage Colony-Stimulating Factor auto-antibodies (GM-CSF Ab) are more likely to develop stricturing behavior requiring surgery. Computed Tomography or Magnetic Resonance Enterography (CTE or MRE) may detect luminal narrowing (LN) prior to stricture development. Objective To determine whether CD patients with elevated GM-CSF Ab (≥ 1.6 mcg/mL) have a higher prevalence of LN and stricturing on CTE or MRE. Methods A single center, cross-sectional study of 153 pediatric CD patients and controls undergoing CTE or MRE. A novel scoring system evaluated for disease activity, presence of LN, stricture, intra-abdominal abscess, or fistulae Ouutcomes were compared with respect to antibody status using Fisher's exact test, logistic regression, and the unpaired t-test. Results GM-CSF Ab were elevated in CD patients (n=114) with a median (IQR) GM-CSF Ab level of 2.3 mcg/mL (0.5, 6.6) compared with healthy and disease controls, p=0.001. Ileal disease location was more common in CD patients with high GM-CSF Ab, p<0.001. Luminal narrowing increased from 39% in CD patients with low GM-CSF Ab to 71% in those with high levels (p=0.004). High GM-CSF Ab remained significantly associated with LN in a multivariate logistic model. Stricturing increased from 4% in CD patients with low GM-CSF Ab to 19% in those with high GM-CSF Ab (p=0.03). Conclusions Pediatric CD patients with high GM-CSF Ab levels have a higher prevalence of LN on CTE or MRE. Further study will be needed to determine whether medical therapy will reduce progression to stricturing behavior in these patients. PMID:23893081

  5. Effectiveness of daily versus non-daily granulocyte colony-stimulating factors in patients with solid tumours undergoing chemotherapy: a multivariate analysis of data from current practice

    PubMed Central

    Almenar Cubells, D; Bosch Roig, C; Jiménez Orozco, E; Álvarez, R; Cuervo, JM; Díaz Fernández, N; Sánchez Heras, AB; Galán Brotons, A; Giner Marco, V; Codes M De Villena, M

    2013-01-01

    We conducted a multicentre, retrospective, observational study including patients with solid tumours (excluding breast cancer) that received granulocyte colony-stimulating factors (G-CSF) and chemotherapy. We investigated the effectiveness of daily vs. non-daily G-CSFs (pegfilgrastim) adjusting by potential confounders. The study included 391 patients (211 daily G-CSF; 180 pegfilgrastim), from whom 47.3% received primary prophylaxis (PP) (57.8% pegfilgrastim), 26.3% secondary prophylaxis (SP: initiation after cycle 1 and no reactive treatment in any cycle) (51.5% pegfilgrastim) and 26.3% reactive treatment (19.4% pegfilgrastim). Only 42.2% of patients with daily G-CSF and 46.2% with pegfilgrastim initiated prophylaxis within 72 h after chemotherapy, and only 10.5% of patients with daily G-CSF received it for ≥7 days. In the multivariate models, daily G-CSF was associated with higher risk of grade 3-4 neutropenia (G3-4N) vs. pegfilgrastim [odds ratio (OR): 1.73, 95% confidence interval (CI): 1.004–2.97]. Relative to SP, PP protected against G3-4N (OR for SP vs. PP: 6.0, 95%CI: 3.2–11.4) and febrile neutropenia (OR: 3.1, 95%CI: 1.1–8.8), and was associated to less chemotherapy dose delays and reductions (OR for relative dose intensity <85% for SP vs. PP: 3.1, 95%CI: 1.7–5.4) and higher response rate (OR: 2.1, 95%CI: 1.2–3.7). Data suggest that pegfilgrastim, compared with a daily G-CSF, and PP, compared with SP, could be more effective in preventing neutropenia and its related events in the clinical practice. PMID:23331323

  6. pH responsive granulocyte colony-stimulating factor variants with implications for treating Alzheimer's disease and other central nervous system disorders.

    PubMed

    Heinzelman, Pete; Schoborg, Jennifer A; Jewett, Michael C

    2015-10-01

    Systemic injection of granulocyte colony-stimulating factor (G-CSF) has yielded encouraging results in treating Alzheimer's Disease (AD) and other central nervous system (CNS) disorders. Making G-CSF a viable AD therapeutic will, however, require increasing G-CSF's ability to stimulate neurons within the brain. This objective could be realized by increasing transcytosis of G-CSF across the blood brain barrier (BBB). An established correlation between G-CSF receptor (G-CSFR) binding pH responsiveness and increased recycling of G-CSF to the cell exterior after endocytosis motivated development of G-CSF variants with highly pH responsive G-CSFR binding affinities. These variants will be used in future validation of our hypothesis that increased BBB transcytosis can enhance G-CSF therapeutic efficacy. Flow cytometric screening of a yeast-displayed library in which G-CSF/G-CSFR interface residues were mutated to histidine yielded a G-CSF triple His mutant (L109H/D110H/Q120H) with highly pH responsive binding affinity. This variant's KD, measured by surface plasmon resonance (SPR), increases ∼20-fold as pH decreases from 7.4 to below histidine's pKa of ∼6.0; an increase 2-fold greater than for previously reported G-CSF His mutants. Cell-free protein synthesis (CFPS) enabled expression and purification of soluble, bioactive G-CSF triple His variant protein, an outcome inaccessible via Escherichia coli inclusion body refolding. This purification and bioactivity validation will enable future identification of correlations between pH responsiveness and transcytosis in BBB cell culture model and animal experiments. Furthermore, the library screening and CFPS methods employed here could be applied to developing other pH responsive hematopoietic or neurotrophic factors for treating CNS disorders. PMID:25877663

  7. Lipopolysaccharide-binding protein (LBP) blockade augments the protective effect of granulocyte colony-stimulating factor (G-CSF) in a rat sepsis model.

    PubMed

    Liu, Anding; Weiss, Stefanie; Fang, Haoshu; Claus, Ralf A; Rödel, Jürgen; Dirsch, Olaf; Dahmen, Uta

    2015-05-01

    The effect of granulocyte colony-stimulating factor (G-CSF) on sepsis is discussed controversially in clinical studies. We previously demonstrated that G-CSF treatment induced lipopolysaccharide (LPS) sensitization via up-regulation of LPS-binding protein (LBP). We hypothesized that the futile effect of G-CSF-treatment in sepsis might be due to its ability to up-regulate LBP. Therefore, blockade of LBP may attenuate the G-CSF-induced LPS sensitization and protect animals from polymicrobial sepsis. Endogenous LBP levels were up-regulated by pretreatment with G-CSF, and the LBP protein was blocked by administration of a specific blocking peptide-LBPK95A. Polymicrobial sepsis was induced by intraperitoneal injection of feces slurry. Rats were monitored every 3 up to 72 h to observe the survival rate. Tissue injury, bacterial infiltration, local inflammatory response, and neutrophil infiltration at 0, 2, and 12 h after the septic insult were analyzed. The survival benefit of G-CSF pretreatment was improved when combined with LBPK95A treatment (control vs. G-CSF vs. combi: 36% vs. 56% vs. 93%; P < 0.05). Combined treatment of G-CSF and LBPK95A was associated with the minimal tissue damage. Treatment with LBPK95A significantly inhibited the neutrophil infiltration without interfering with the bacterial clearance. The G-CSF-induced inflammatory sensitization effect was inhibited by LBPK95A, indicated by the decrease of cytokines expression, and the activation of nuclear factor kappa B and signal transducer and activator of transcription 3 signaling pathway. In conclusion, these results suggested that the effect of prophylactic augmentation of the host's response via G-CSF pretreatment was further enhanced by inhibition of the up-regulation of LBP.

  8. Increased susceptibility to liver injury after hemorrhagic shock in rats chronically fed ethanol: role of nuclear factor-kappa B, interleukin-6, and granulocyte colony-stimulating factor.

    PubMed

    Ono, Masafumi; Yu, Bi; Hardison, Edith G; Mastrangelo, Mary-Ann A; Tweardy, David J

    2004-06-01

    Chronic ethanol use preceding severe trauma and hemorrhagic shock (HS) is associated with an increased incidence of multiorgan failure (MOF) and death; however, the molecular basis for this increased susceptibility is unknown. We previously demonstrated that production of interleukin-6 (IL-6) and granulocyte colony-stimulating factor (G-CSF), mediated by nuclear factor-kappa B (NF-kappa B), each make essential contributions to organ injury and inflammation in a rodent model of controlled HS, and we proposed in this study to examine the hypothesis that the increased susceptibility to MOF after shock/trauma in the setting of chronic ethanol use is due to an exaggerated activation of NF-kappa B and production of these proinflammatory cytokines. We observed increased HS-induced liver injury 4 h after resuscitation in rats fed the ethanol-containing Lieber-DeCarli liquid diet for 8 weeks compared with rats fed the control liquid diet (3-fold increase in serum alanine aminotransferase [ALT], P = 0.008, and 2-fold increase in focal liver necrosis, P = 0.005). The increased liver injury in the ethanol-fed HS rats was accompanied by a 70% increase in liver NF-kappa B activation (P < 0.05), a 3- to 5-fold increase in hepatocyte and Kupffer cell production of IL-6 and G-CSF (P < 0.05 for each), and a 2-fold increase in neutrophil infiltration (P < 0.005) compared with the control diet-fed HS rats. Thus, increased susceptibility to HS-induced liver injury in the setting of chronic ethanol use may be mediated, at least in part, by increased NF-kappa B activation resulting in increased local production of IL-6 and G-CSF and increased infiltration of neutrophils, which can damage liver cells directly and contribute to impaired sinusoidal blood flow.

  9. Carcinoembryonic antigen-related cell adhesion molecule 1 negatively regulates granulocyte colony-stimulating factor production by breast tumor-associated macrophages that mediate tumor angiogenesis.

    PubMed

    Samineni, Sridhar; Zhang, Zhifang; Shively, John E

    2013-07-15

    Carcinoembryonic antigen-related cell adhesion molecule 1 (CEACAM1), a cell adhesion molecule expressed on epithelial cells and activated immune cells, is downregulated in many cancers and plays a role in inhibition of inflammation in part by inhibition of granulocyte colony-stimulating factor (G-CSF) production by myeloid cells. As macrophages are associated with a poor prognosis in breast cancer, but play important roles in normal breast, we hypothesized that CEACAM1 downregulation would lead to tumor promotion under inflammatory conditions. Cocultures of proinflammatory M1 macrophages with CEACAM1 negative MCF7 breast cells produced high levels of G-CSF (10 ng/mL) compared to CEACAM1-transfected MCF7/4S cells (1 ng/mL) or anti-inflammatory M2 macrophage cocultures (0.5 or 0.1 ng/mL, MCF7 or MCF7/4S, respectively). The expression of CEACAM1 on M1s was much greater than for M2s and was observed only in cocultures with either MCF7 or MCF7/4S cells. When M1 macrophages were mixed with MCF7 cells and implanted in murine mammary fat pads of nonobese diabetic/severe combined immunodeficient mice, tumor size and blood vessel density were significantly greater than MCF7 or MCF7/4S only tumors which were hardly detected after 8 weeks of growth. In contrast, M1 cells had a much reduced effect on MCF7/4S tumor growth and blood vessel density, indicating that the tumor inhibitory effect of CEACAM1 is most likely related to its anti-inflammatory action on inflammatory macrophages. These results support our previous finding that CEACAM1 inhibits both G-CSF production by myeloid cells and G-CSF-stimulated tumor angiogenesis.

  10. Effectiveness of daily versus non-daily granulocyte colony-stimulating factors in patients with solid tumours undergoing chemotherapy: a multivariate analysis of data from current practice.

    PubMed

    Almenar Cubells, D; Bosch Roig, C; Jiménez Orozco, E; Álvarez, R; Cuervo, J M; Díaz Fernández, N; Sánchez Heras, A B; Galán Brotons, A; Giner Marco, V; Codes M De Villena, M

    2013-05-01

    We conducted a multicentre, retrospective, observational study including patients with solid tumours (excluding breast cancer) that received granulocyte colony-stimulating factors (G-CSF) and chemotherapy. We investigated the effectiveness of daily vs. non-daily G-CSFs (pegfilgrastim) adjusting by potential confounders. The study included 391 patients (211 daily G-CSF; 180 pegfilgrastim), from whom 47.3% received primary prophylaxis (PP) (57.8% pegfilgrastim), 26.3% secondary prophylaxis (SP: initiation after cycle 1 and no reactive treatment in any cycle) (51.5% pegfilgrastim) and 26.3% reactive treatment (19.4% pegfilgrastim). Only 42.2% of patients with daily G-CSF and 46.2% with pegfilgrastim initiated prophylaxis within 72 h after chemotherapy, and only 10.5% of patients with daily G-CSF received it for ≥ 7 days. In the multivariate models, daily G-CSF was associated with higher risk of grade 3-4 neutropenia (G3-4N) vs. pegfilgrastim [odds ratio (OR): 1.73, 95% confidence interval (CI): 1.004-2.97]. Relative to SP, PP protected against G3-4N (OR for SP vs. PP: 6.0, 95%CI: 3.2-11.4) and febrile neutropenia (OR: 3.1, 95%CI: 1.1-8.8), and was associated to less chemotherapy dose delays and reductions (OR for relative dose intensity <85% for SP vs. PP: 3.1, 95%CI: 1.7-5.4) and higher response rate (OR: 2.1, 95%CI: 1.2-3.7). Data suggest that pegfilgrastim, compared with a daily G-CSF, and PP, compared with SP, could be more effective in preventing neutropenia and its related events in the clinical practice.

  11. pH responsive granulocyte colony-stimulating factor variants with implications for treating Alzheimer's disease and other central nervous system disorders.

    PubMed

    Heinzelman, Pete; Schoborg, Jennifer A; Jewett, Michael C

    2015-10-01

    Systemic injection of granulocyte colony-stimulating factor (G-CSF) has yielded encouraging results in treating Alzheimer's Disease (AD) and other central nervous system (CNS) disorders. Making G-CSF a viable AD therapeutic will, however, require increasing G-CSF's ability to stimulate neurons within the brain. This objective could be realized by increasing transcytosis of G-CSF across the blood brain barrier (BBB). An established correlation between G-CSF receptor (G-CSFR) binding pH responsiveness and increased recycling of G-CSF to the cell exterior after endocytosis motivated development of G-CSF variants with highly pH responsive G-CSFR binding affinities. These variants will be used in future validation of our hypothesis that increased BBB transcytosis can enhance G-CSF therapeutic efficacy. Flow cytometric screening of a yeast-displayed library in which G-CSF/G-CSFR interface residues were mutated to histidine yielded a G-CSF triple His mutant (L109H/D110H/Q120H) with highly pH responsive binding affinity. This variant's KD, measured by surface plasmon resonance (SPR), increases ∼20-fold as pH decreases from 7.4 to below histidine's pKa of ∼6.0; an increase 2-fold greater than for previously reported G-CSF His mutants. Cell-free protein synthesis (CFPS) enabled expression and purification of soluble, bioactive G-CSF triple His variant protein, an outcome inaccessible via Escherichia coli inclusion body refolding. This purification and bioactivity validation will enable future identification of correlations between pH responsiveness and transcytosis in BBB cell culture model and animal experiments. Furthermore, the library screening and CFPS methods employed here could be applied to developing other pH responsive hematopoietic or neurotrophic factors for treating CNS disorders.

  12. Stem cell mobilisation by granulocyte-colony stimulating factor in patients with acute myocardial infarction. Long-term results of the REVIVAL-2 trial.

    PubMed

    Steppich, Birgit; Hadamitzky, Martin; Ibrahim, Tareq; Groha, Philip; Schunkert, Heribert; Laugwitz, Karl-Ludwig; Kastrati, Adnan; Ott, Ilka

    2016-04-01

    Treatment with granulocyte-colony stimulating factor (G-CSF) mobilises cells from the bone marrow to the peripheral blood. Previous preclinical and early clinical trials may suggest that treatment with G-CSF leads to improved myocardial perfusion and function in acute or chronic ischaemic heart disease. In the REVIVAL-2 study we found that stem cell mobilisation by G-CSF does not influence infarct size, left ventricular function and coronary restenosis in patients with acute myocardial infarction (MI) that underwent successful percutaneous coronary intervention. The objective of the present analysis was to assess the impact of G-CSF treatment on seven-year clinical outcomes from the REVIVAL-2 trial. In the randomized, double-blind, placebo-controlled REVIVAL-2 study, 114 patients with the diagnosis of acute myocardial infarction were enrolled five days after successful reperfusion by percutaneous coronary intervention. Patients were assigned to receive 10 µg/kg G-CSF (n=56) or placebo (n=58) for five days. The primary endpoint for this long-term outcome analysis was the composite of death, myocardial infarction or stroke seven years after randomisation. The endpoint occurred in 14.3 % of patients in the G-CSF group versus 17.2 % assigned to placebo (p=0.67). The combined incidence of death or myocardial infarction occurred in 14.3 % of the patients assigned to G-CSF and 15.5 % of the patients assigned to placebo (p=0.85). In conclusion, these long-term follow-up data show that G-CSF does not improve clinical outcomes of patients with acute myocardial infarction.

  13. CHOP chemotherapy with preemptive granulocyte colony-stimulating factor in elderly patients with aggressive non-Hodgkin's lymphoma: a dose-intensity analysis.

    PubMed

    Jacobson, J O; Grossbard, M; Shulman, L N; Neuberg, D

    2000-12-01

    This prospective trial was designed to determine the safety and efficacy of full-dose, on-time chemotherapy in elderly patients with aggressive non-Hodgkin's lymphoma. Twenty patients (median age, 71 years; range, 66 to 80 years) were enrolled in a phase II, multicenter trial to receive cyclophosphamide/doxorubicin/vincristine/prednisone (CHOP) supported by granulocyte colony-stimulating factor (G-CSF). CHOP was given in standard doses. Six cycles were planned every 21 days, with G-CSF starting on day 3 and continuing until the absolute neutrophil count was greater than 10,000/microL. Consolidation radiation therapy was permitted. Restaging was performed following cycles 4 and 6. By the age-adjusted International Prognostic Index, four patients were low, 10 were low-intermediate, four were high-intermediate, and two were high risk. Eighteen cases completed all 6 cycles. The average cycle length for all 112 cycles was 21.7 days. The dose intensities (corrected for delay) for each agent were cyclophosphamide 97.3%, doxorubicin 97.3%, vincristine 91.5%, and prednisone 97.3%. Treatment-related complications included grade 4 leukopenia and grade 4 thrombocytopenia in 11.6% and 3.6% of cycles, respectively. Hospitalization for neutropenia and fever was needed for 7.1% of cycles. There was no grade 3/4 cardiac toxicity. No treatment-related mortality occurred. All toxicities were reversible. There were 12 (60%) complete responses, four (20%) gallium-negative partial responses, and four patients (20%) with progressive disease. With a median follow-up of 2.29 years, progression-free and overall survival rates at 2 years are 42% (90% confidence interval: 23%-61%) and 66% (90% confidence interval: 47%-85%), respectively. Using preemptive G-CSF, full-dose CHOP can be administered safely to elderly patients.

  14. Overexpression of granulocyte-macrophage colony-stimulating factor induces pulmonary granulation tissue formation and fibrosis by induction of transforming growth factor-beta 1 and myofibroblast accumulation.

    PubMed Central

    Xing, Z.; Tremblay, G. M.; Sime, P. J.; Gauldie, J.

    1997-01-01

    We have previously reported that transfer to rat lung of the granulocyte-macrophage colony-stimulating factor (GM-CSF) gene leads to high expression of GM-CSF between days 1 and 4 and granulation tissue formation followed by an irreversible fibrotic response starting from day 12 onward. In the current study, we investigated the underlying mechanisms. We found that GM-CSF overexpression did not enhance production of tumor necrosis factor-alpha in a significant manner at any time after GM-CSF gene transfer. However, the content of transforming growth factor-beta 1 in bronchoalveolar lavage fluid was markedly induced at day 4 and appeared to be maximal around day 7 and remained high at day 12. Macrophages purified from bronchoalveolar lavage fluid 7 days after GM-CSF gene transfer spontaneously released significant quantities of transforming growth factor-beta 1 protein in vitro. After peak transforming growth factor-beta 1 production was the emergence of alpha-smooth muscle actin-rich myofibroblasts. Accumulation of these cells was most prominent at day 12 within the granulation tissues and they were still present in fibrotic areas between days 12 and 24 and diminished markedly afterward. Thus, we provide the first in vivo evidence that tumor necrosis factor-alpha may be dissociated from participation in a fibrotic process in the lung and GM-CSF may play a more direct role in pulmonary fibrogenesis at least in part through its capability to induce transforming growth factor-beta 1 in macrophages and the subsequent emergence of myofibroblast phenotypes. This GM-CSF transgene lung model is useful for a stepwise dissection of both cellular and molecular events involved in pulmonary fibrosis. Images Figure 2 Figure 5 Figure 6 PMID:9006322

  15. [Recombinant granulocyte-colony stimulating factor (filgrastim): optimization of conditions of isolation and purification from inclusion body].

    PubMed

    Kononova, N V; Iakovlev, A V; Zhuravko, A M; Pankeev, N N; Minaev, S V; Bobruskin, A I; Mart'ianov, V A

    2014-01-01

    We developed a unified process platform for two recombinant human GCSF medicines--one with the non-prolonged and the other with prolonged action. This unified technology led to a simpler and cheaper production while introduction of the additional pegylation stage to the technological line eased obtaining of the medicines with different action and allowed to standardize technological process documenting according to GMP requirements.

  16. Enhancement of innate immunity with granulocyte colony-stimulating factor did not mitigate disease in pigs infected with a highly pathogenic Chinese PRRSV strain.

    PubMed

    Schlink, Sarah N; Lager, Kelly M; Brockmeier, Susan L; Loving, Crystal L; Miller, Laura C; Vorwald, Ann C; Yang, Han-Chun; Kehrli, Marcus E; Faaberg, Kay S

    2016-10-15

    Porcine reproductive and respiratory syndrome virus (PRRSV) is responsible for one of the most economically important diseases in swine worldwide. It causes reproductive failure in sows and pneumonia in pigs that predisposes them to secondary bacterial infections. Methods to control PRRSV and/or limit secondary bacterial infections are desired to reduce the impact of this virus on animal health. Neutrophils play a major role in combatting infection; they can act as phagocytes as well as produce and release lytic enzymes that have potent antimicrobial effects leading to the destruction and clearance of bacterial pathogens. Granulocyte-colony stimulating factor (G-CSF) is a cytokine that controls the production, differentiation and function of granulocytes (including neutrophils) from the bone marrow. Recent work from our laboratory has shown that encoding porcine G-CSF in a replication-defective adenovirus (Ad5-G-CSF) and delivering a single dose to pigs induced a neutrophilia lasting more than two weeks. As secondary bacterial infection is a common occurrence following PRRSV infection, particularly following challenge with highly pathogenic (HP)-PRRSV, the aim of the current study was to evaluate the effectiveness of a single prophylactic dose of adenovirus-encoded G-CSF to mitigate secondary bacterial disease associated with HP-PRRSV infection. Administration of Ad5-G-CSF induced a significant neutrophilia as expected. However, between 1 and 2days following HP-PRRSV challenge the number of circulating neutrophils decreased dramatically in the HP-PRRSV infected group, but not the non-infected Ad5-G-CSF group. Ad5-G-CSF administration induced monocytosis as well, which was also reduced by HP-PRRSV challenge. There was no difference in the progression of disease between the Ad5-G-CSF and Ad5-empty groups following HP-PRRSV challenge, with pneumonia and systemic bacterial infection occurring in both treatment groups. Given the impact of HP-PRRSV infection on the

  17. Treatment of adjuvant arthritis with granulocyte-colony stimulating factor and peptide derived from heat shock protein 65.

    PubMed

    Brendolan, Andrea; Higuchi, Masanori; Sibley, Richard; Strober, Samuel

    2003-01-01

    Adjuvant arthritis in Lewis rats is induced by the subcutaneous injection of Mycobacterium tuberculosis in mineral oil, and the predominant T cell immune reactivity is against the heat shock protein 65 derived peptide 176-190. We treated Lewis rats with human recombinant G-CSF followed by (i.v) administration of peptide 176-190 after induction of adjuvant arthritis (AA), and observed decreased disease severity, joint destruction, new bone formation and joint ankylosis. Treatment with G-CSF alone was also effective, but to a lesser extent. In addition, we found that splenocytes from rats treated with G-CSF had reduced antigen presenting capacity compared with splenocytes from vehicle treated rats. Primed lymph node cells from G-CSF plus peptide treated rats showed a marked reduction in proliferation and secretion of IFN-gamma after stimulation with the heat shock protein peptide in vitro as compared to controls.

  18. Role of Serum Amyloid A, Granulocyte-Macrophage Colony-Stimulating Factor, and Bone Marrow Granulocyte-Monocyte Precursor Expansion in Segmented Filamentous Bacterium-Mediated Protection from Entamoeba histolytica.

    PubMed

    Burgess, Stacey L; Saleh, Mahmoud; Cowardin, Carrie A; Buonomo, Erica; Noor, Zannatun; Watanabe, Koji; Abhyankar, Mayuresh; Lajoie, Stephane; Wills-Karp, Marsha; Petri, William A

    2016-10-01

    Intestinal segmented filamentous bacteria (SFB) protect from ameba infection, and protection is transferable with bone marrow dendritic cells (BMDCs). SFB cause an increase in serum amyloid A (SAA), suggesting that SAA might mediate SFB's effects on BMDCs. Here we further explored the role of bone marrow in SFB-mediated protection. Transient gut colonization with SFB or SAA administration alone transiently increased the H3K27 histone demethylase Jmjd3, persistently increased bone marrow Csf2ra expression and granulocyte monocyte precursors (GMPs), and protected from ameba infection. Pharmacologic inhibition of Jmjd3 H3K27 demethylase activity during SAA treatment or blockade of granulocyte-macrophage colony-stimulating factor (GM-CSF) signaling in SFB-colonized mice prevented GMP expansion, decreased gut neutrophils, and blocked protection from ameba infection. These results indicate that alteration of the microbiota and systemic exposure to SAA can influence myelopoiesis and susceptibility to amebiasis via epigenetic mechanisms. Gut microbiota-marrow communication is a previously unrecognized mechanism of innate protection from infection. PMID:27456830

  19. Combined suicide and granulocyte-macrophage colony-stimulating factor gene therapy induces complete tumor regression and generates antitumor immunity.

    PubMed

    Jones, R K; Pope, I M; Kinsella, A R; Watson, A J; Christmas, S E

    2000-12-01

    The use of prodrug-activated ("suicide") gene therapy has been shown to be effective in inducing tumor regression when only a small proportion of tumor cells contains the suicide gene. These experiments were designed to test whether additional therapeutic benefit may be obtained by stimulating the immune response. Murine MC26 colon carcinoma cells, either untransduced or transduced with genes for herpes simplex virus-1 thymidine kinase (HSV1-TK) or human GM-CSF, were injected subcutaneously into syngeneic BALB/c mice in various combinations. Inoculation of equal numbers of untransduced and HSV1-TK-containing cells followed by ganciclovir (GCV) treatment resulted in almost complete tumor regression, but by 7 weeks, tumors had recurred in all mice. A similar initial regression was obtained using equal numbers of cells containing HSV1-TK and GM-CSF genes, but >80% of these mice remained tumor-free after 3 months. Groups of tumor-free mice that had received GM-CSF-containing cells were left for different periods of time and rechallenged with unmodified MC26 cells on the opposite flank. Of the mice rechallenged 14, 28, and 108 days later, 100%, 88%, and 57%, respectively, showed complete resistance to unmodified tumor cells. In mice that showed tumor regrowth, tumor volume was much less than in control mice. Adoptive transfer of spleen cells from resistant mice to naïve syngeneic mice resulted in partial resistance to challenge with unmodified tumor cells. Specific cytotoxicity against MC26 cells was only demonstrable in mice receiving GM-CSF- and HSV1-TK-containing tumor cells. These experiments show that the presence of cells secreting GM-CSF in HSV1-TK-containing, regressing tumor is able to induce complete or partial resistance to tumor rechallenge. This indicates the potential usefulness of GM-CSF in enhancing other antitumor therapies.

  20. Effect of folic acid plus glycine supplement on uterine prostaglandin and endometrial granulocyte-macrophage colony-stimulating factor expression during early pregnancy in pigs.

    PubMed

    Guay, Frédéric; Matte, J Jacques; Girard, Christiane L; Palin, Marie-France; Giguère, Alain; Laforest, Jean-Paul

    2004-01-15

    The objective was to determine the effects of folic acid+glycine supplement on uterine metabolism of prostaglandin and mRNA expression of endometrial granulocyte-macrophage colony-stimulating factor (GM-CSF) in nulliparous (NYL) and multiparous Yorkshire-Landrace (YL) sows, and in multiparous Meishan-Landrace sows (ML). In each of these three groups, sows were randomly assigned to two treatments: 15 ppm folic acid+0.6% glycine or no supplement. The dietary supplement was given from the estrus before mating to slaughter on Day 25 of pregnancy. At slaughter, endometrial tissue was collected to determine endometrial expression levels of GM-CSF mRNA, cyclooxygenase-1 (COX1) and -2 (COX2) and to evaluate in vitro endometrial secretion of prostaglandin E2 (PGE2) secretion. Allantoic fluid samples were also collected to determine the concentration of PGE2, prostaglandin F2alpha (PGF2alpha), estradiol-17beta (E2), progesterone (P4), and transforming-growth factor-beta2 (TGF-beta2). The allantoic contents of PGF2alpha, E2 and P4, and endometrial in vitro secretion of PGE2 were not significantly influenced by the folic acid+glycine supplement. The folic acid+glycine supplement tended (P<0.07) to increase allantoic content of PGE2 and TGF-beta2 in all sows and increased (P<0.05) endometrial expression of COX2, especially in NYL sows. The endometrial expression of COX1 was decreased (P<0.05) by folic acid+glycine supplement, especially in multiparous YL sows. The allantoic contents of PGE2 and PGF2alpha were not significantly affected by sow type. However, NYL sows had higher (P<0.05) endometrial in vitro secretion of PGE2 and allantoic content of P4 than multiparous YL and ML sows. The allantoic content of E2 was also higher (P<0.05) in NYL sows than in multiparous ML sows only. The allantoic content of TGF-beta2 was lower (P<0.05) in multiparous ML than in multiparous YL only sows. Finally, in YL and NYL sows, folic acid+glycine supplement decreased (P<0.05) the endometrial

  1. CpG oligonucleotides enhance the tumor antigen-specific immune response of a granulocyte macrophage colony-stimulating factor-based vaccine strategy in neuroblastoma.

    PubMed

    Sandler, Anthony D; Chihara, Hiroshi; Kobayashi, Gen; Zhu, Xiaoyan; Miller, Michal A; Scott, David L; Krieg, Arthur M

    2003-01-15

    Granulocyte macrophage colony-stimulating factor (GM-CSF)-transduced autologous tumor cells form the basis of many immunotherapeutic strategies. We tested whether combining this approach with T-helper 1 (Th-1)-like immunostimulatory CpG oligodeoxynucleotides (CpG ODNs) would improve therapeutic efficacy in an established model of murine neuroblastoma. The weakly immunogenic Neuro-2a cell line was used in syngeneic A/J mice. CpG 1826 was tested for its antitumor effect alone and as an adjuvant to Neuro-2a cells retrovirally transduced to express murine GM-CSF (GM/Neuro-2a). Three days after wild-type (WT) tumor cell inoculation, mice in different groups were s.c. vaccinated in the opposite leg with combinations of WT neuro2a, irradiated (15 Gy) WT or GM/Neuro-2a transfectants with or without CpG 1826 (200 micro g). To test for the induction of memory responses, mice that rejected their tumor were rechallenged with WT Neuro-2a (1 x 10(6)) 7 weeks after vaccination. All of the mice in the control (unvaccinated) group died within 3 weeks after Neuro-2a inoculation. Most of the vaccinated groups had only minimal-to-modest antitumor responses, and the mice succumbed to tumor. Tumor growth was remarkably inhibited in the group of mice that received irradiated GM/Neuro-2a plus CpG and four (50%) of eight mice in this group survived tumor free. Tumor-free mice were resistant to further WT tumor cell challenge, indicating a memory response. Mechanistic studies showed that CpG alone induced a favorable Th-1-like cytokine immune response and vaccine-induced tumor cell killing was dependent on both CD4 and CD8 T cells that killed tumor cell targets by apoptosis. These results demonstrate that CpG ODNs enhanced the antitumor effect of irradiated GM-CSF secreting Neuro-2a cells. This vaccine strategy elicits a potent tumor antigen-specific immune response against established murine neuroblastoma and generates systemic neuroblastoma-specific immunity.

  2. Granulocyte-macrophage colony-stimulating factor (GM-CSF) secreted by cDNA-transfected tumor cells induces a more potent antitumor response than exogenous GM-CSF.

    PubMed

    Shi, F S; Weber, S; Gan, J; Rakhmilevich, A L; Mahvi, D M

    1999-01-01

    Clinical cancer gene therapy trials have generally focused on the transfer of cytokine cDNA to tumor cells ex vivo and with the subsequent vaccination of the patient with these genetically altered tumor cells. This approach results in high local cytokine concentrations that may account for the efficacy of this technique in animal models. We hypothesized that the expression of certain cytokines by tumor cells would be a superior immune stimulant when compared with local delivery of exogenous cytokines. Granulocyte-macrophage colony-stimulating factor (GM-CSF) cDNA in a nonviral expression vector was inserted into MDA-MB-231 (human breast cancer), M21 (human melanoma), B16 (murine melanoma), and P815 (mastocytoma) cells by particle-mediated gene transfer. The ability of transfected tumor cells to generate a tumor-specific immune response was evaluated in an in vitro mixed lymphocyte-tumor cell assay and in an in vivo murine tumor protection model. Peripheral blood lymphocytes cocultured with human GM-CSF-transfected tumor cells were 3- to 5-fold more effective at lysis of the parental tumor cells than were peripheral blood lymphocytes incubated with irradiated tumor cells and exogenous human GM-CSF. Mice immunized with murine GM-CSF-transfected irradiated B16 murine melanoma cells or P815 mastocytoma cells were protected from subsequent tumor challenge, whereas mice immunized with the nontransfected tumors and cutaneous transfection of murine GM-CSF cDNA at the vaccination site developed tumors more frequently. The results indicate that GM-CSF protein expressed in human and murine tumor cells is a superior antitumor immune stimulant compared with exogenous GM-CSF in the tumor microenvironment. PMID:10078967

  3. The pros and cons of split-dose granulocyte colony-stimulating factor alone rather than a single high dose for hematopoietic progenitor cell mobilization in small children (< 15 kg) with solid tumors.

    PubMed

    Merlin, Etienne; Piguet, Christophe; Auvrignon, Anne; Rubie, Hervé; Deméocq, François; Kanold, Justyna

    2006-07-01

    Hematopoietic progenitor cells were mobilized in 34 children with solid tumors weighing < or = 15 kg using granulocyte colony-stimulating factor alone at the doses of 10, 20 or 2 x 12 microg/kg/day. The mobilization with 2 x 12 microg/kg/day was more efficient than that with 10 mg/kg/day. Although the superiority of the split-dose compared to the single, high daily dose (20 microg/kg/day) was not statistically significant, our results suggest that the 2 x 12 microg/kg/day regimen is interesting.

  4. The intramuscular administration of granulocyte colony-stimulating factor as an adjunct to chemotherapy in pretreated ovarian cancer patients: an Italian Trials in Medical Oncology (ITMO) Group pilot study.

    PubMed Central

    Di Leo, A.; Bajetta, E.; Nolè, F.; Biganzoli, L.; Ferrari, L.; Oriana, S.; Riboldi, G.; Bohm, S.; Spatti, G.; Raspagliesi, F.

    1994-01-01

    No published data are available concerning the activity and tolerability of intramuscularly administered granulocyte colony-stimulating factor (G-CSF) in humans. To fill this gap, 19 patients with advanced ovarian cancer previously treated with at least one first-line chemotherapy cycle received the following myelosuppressive regimen: mitoxantrone (DHAD) 12 mg m-2 i.v. on day 1; ifosfamide (IFO) 4 g m-2 i.v. on days 1 and 2; mesna 800 mg m-2 i.v. t.i.d. on days 1 and 2. G-CSF (Filgrastim) was given at a dose of 5 micrograms/kg/day i.m. from day 6 to day 19, its pharmacokinetics being assessed in five patients. The neutrophil nadir was observed after a mean period of 8 days, and the neutrophil count was < 1.0 x 10(3) mm-3 for a mean of 6 days during the cycle of chemotherapy. The neutrophil count fell after the withdrawal of G-CSF on the 19th day of treatment. The difference in absolute neutrophil count between day 19 and day 21 was statistically significant (P = 0.0001); nevertheless, at day 21 no WHO grade 3-4 neutropenia was reported. DHAD and IFO were respectively given at 95% and 93% of the planned dose. The pharmacokinetics of G-CSF i.m. seems to be similar to that of the drug given subcutaneously. No evidence of cumulative myelosuppression was observed. G-CSF was well tolerated and no complications were observed at the injection sites. In conclusion, if the results obtained in this pilot study regarding the activity of i.m. G-CSF are confirmed by a randomised trial, the intramuscular administration of G-CSF could become a valid alternative for patients who dislike the subcutaneous route and who are being treated with chemotherapy that does not induce profound thrombocytopenia. PMID:7514030

  5. Establishment and characterization of a human thyroid carcinoma cell line (HOTHC) producing colony stimulating factor.

    PubMed

    Ishiwata, Isamu; Ono, Isao; Kiguchi, Kazushige; Ishiwata, Chieko; Soma, Masayuki; Ishikawa, Hiroshi

    2005-09-01

    A cell line designated HOTHC was established from an anaplastic carcinoma (giant cell type) of the thyroid gland of 80-year-old woman. The HOTHC line grew rapidly in multilayer without contact inhibition, and more than 120 serial passages were made within 27 months. The cells were spindle or polygonal in shape and revealed neoplastic and pleomorphic features. These cells were characterized as containing coloid droplets and poorly developed rough-endoplasmic reticulum in the cytoplasm. Doubling time was about 24 hours and plating efficiency was about 70%. The karyotype exhibits hyperploidy and marker chromosomes, and the modal chromosome number ranged between 77-90. The HOTHC cells were transplanted into the subcutis of BALB/c nude mice and produced anaplatic carcinomas (giant cell type) resembling the original tumor. The HOTHC cells produced colony stimulating factor (CSF) and caused granulocytosis in the mice. PMID:17022149

  6. Human Granulocytic Anaplasmosis

    PubMed Central

    Bakken, Johan S.

    2015-01-01

    Synopsis Human granulocytic anaplasmosis (HGA), a deer tick transmitted rickettsial infection caused by Anaplasma phagocytophilum, is a common cause of undifferentiated fever in the Northeast and Upper Midwest U.S. Patients are often initially diagnosed with a mild viral infection, and illness readily resolves in most cases. However, as many as 3% may develop life threatening complications and nearly 1% die from the infection. A history of tick bite and a high degree of clinical suspicion thus warrant consideration for doxycycline treatment in both adults and children, even in the absence of known tick bite, a negative blood film examination, or pending results of specific A. phagocytophilum diagnostic tests such as paired serology or PCR on acute phase blood. Antibody tests and titers should not be used to monitor active infection as detectable antibodies can remain present for years. Moreover, persistent infection has never been reported. While co-infections with Borrelia burgdorferi and Babesia microti occur, there is little evidence to suggest synergism of disease or a role for A. phagocytophilum in chronic illness. Preventive measures include avoiding tick-infested areas, use of tick repellents, and careful searches of skin to remove attached ticks; no vaccine is available. PMID:25999228

  7. Mouse neutrophilic granulocytes express mRNA encoding the macrophage colony-stimulating factor receptor (CSF-1R) as well as many other macrophage-specific transcripts and can transdifferentiate into macrophages in vitro in response to CSF-1.

    PubMed

    Sasmono, R Tedjo; Ehrnsperger, Achim; Cronau, Stephen L; Ravasi, Timothy; Kandane, Rangi; Hickey, Michael J; Cook, Andrew D; Himes, S Roy; Hamilton, John A; Hume, David A

    2007-07-01

    The differentiation of macrophages from their progenitors is controlled by macrophage colony-stimulating factor (CSF-1), which binds to a receptor (CSF-1R) encoded by the c-fms proto-oncogene. We have previously used the promoter region of the CSF-1R gene to direct expression of an enhanced green fluorescent protein (EGFP) reporter gene to resident macrophage populations in transgenic mice. In this paper, we show that the EGFP reporter is also expressed in all granulocytes detected with the Gr-1 antibody, which binds to Ly-6C and Ly-6G or with a Ly-6G-specific antibody. Transgene expression reflects the presence of CSF-1R mRNA but not CSF-1R protein. The same pattern is observed with the macrophage-specific F4/80 marker. Based on these findings, we performed a comparative array profiling of highly purified granulocytes and macrophages. The patterns of mRNA expression differed predominantly through granulocyte-specific expression of a small subset of transcription factors (Egr1, HoxB7, STAT3), known abundant granulocyte proteins (e.g., S100A8, S100A9, neutrophil elastase), and specific receptors (fMLP, G-CSF). These findings suggested that appropriate stimuli might mediate rapid interconversion of the major myeloid cell types, for example, in inflammation. In keeping with this hypothesis, we showed that purified Ly-6G-positive granulocytes express CSF-1R after overnight culture and can subsequently differentiate to form F4/80-positive macrophages in response to CSF-1.

  8. Granulocyte-macrophage colony stimulating factor promotes prolonged survival and the support of virulent infection by African swine fever virus of macrophages generated from porcine bone marrow and blood.

    PubMed

    Denham, S; Brookes, S M; Hutchings, G H; Parkhouse, R M

    1996-10-01

    Long-surviving cultures of non-adherent cells of the monocyte-macrophage lineage were established from the bone marrow and blood of weanling pigs by culturing cells from these tissues in the presence of recombinant porcine granulocyte-macrophage colony stimulating factor (GM-CSF). The cells increased in number, principally during the first 4 weeks of culture, bound monoclonal antibodies recognizing porcine macrophage antigens and avidly phagocytosed latex particles. The GM-CSF generated mononuclear phagocytes were highly infectible [correction of infectable] with a virulent Malawi isolate of African swine fever virus (ASFV) and able to generate levels of virus progeny similar to those produced by freshly isolated pig macrophages. The cultured cells retained their susceptibility to ASFV infection for as long as the cultures survived i.e. for up to 3 months.

  9. Key role for myeloid cells: phase II results of anti-G(D2) antibody 3F8 plus granulocyte-macrophage colony-stimulating factor for chemoresistant osteomedullary neuroblastoma.

    PubMed

    Cheung, Nai-Kong V; Cheung, Irene Y; Kramer, Kim; Modak, Shakeel; Kuk, Deborah; Pandit-Taskar, Neeta; Chamberlain, Elizabeth; Ostrovnaya, Irina; Kushner, Brian H

    2014-11-01

    Anti-G(D2) murine antibody 3F8 plus subcutaneously (sc) administered granulocyte-macrophage colony-stimulating factor (GM-CSF) was used against primary refractory neuroblastoma in metastatic osteomedullary sites. Large study size and long follow-up allowed assessment of prognostic factors in a multivariate analysis not reported with other anti-G(D2) antibodies. In a phase II trial, 79 patients without prior progressive disease were treated for persistent osteomedullary neuroblastoma documented by histology and/or metaiodobenzyl-guanidine (MIBG) scan. In the absence of human antimouse antibody, 3F8 + scGM-CSF cycles were repeated up to 24 months. Minimal residual disease (MRD) in bone marrow was measured by quantitative reverse transcription-polymerase chain reaction pre-enrollment and post-cycle #2, before initiation of 13-cis-retinoic acid. Study endpoints were: (i) progression-free survival (PFS) compared with the predecessor trial of 3F8 plus intravenously administered (iv) GM-CSF (26 patients) and (ii) impact of MRD on PFS. Using all 105 patients from the two consecutive 3F8 + GM-CSF trials, prognostic factors were analyzed by multivariate Cox regression model. Complete response rates to 3F8 + scGM-CSF were 87% by histology and 38% by MIBG. Five-year PFS was 24 ± 6%, which was significantly superior to 11 ± 7% with 3F8 + ivGM-CSF (p = 0.002). In the multivariate analysis, significantly better PFS was associated with R/R or H/R FCGR2A polymorphism, sc route of GM-CSF and early MRD response. MYCN amplification was not prognostic. Complement consumption was similar with either route of GM-CSF. Toxicities were manageable, allowing outpatient treatment. 3F8 + scGM-CSF is highly active against chemoresistant osteomedullary neuroblastoma. MRD response may be an indicator of tumor sensitivity to anti-G(D2) immunotherapy. Correlative studies highlight the antineoplastic potency of myeloid effectors.

  10. Murine Anti-GD2 Monoclonal Antibody 3F8 Combined With Granulocyte-Macrophage Colony-Stimulating Factor and 13-Cis-Retinoic Acid in High-Risk Patients With Stage 4 Neuroblastoma in First Remission

    PubMed Central

    Cheung, Nai-Kong V.; Cheung, Irene Y.; Kushner, Brian H.; Ostrovnaya, Irina; Chamberlain, Elizabeth; Kramer, Kim; Modak, Shakeel

    2012-01-01

    Purpose Anti-GD2 monoclonal antibody (MoAb) combined with granulocyte-macrophage colony-stimulating factor (GM-CSF) has shown efficacy against neuroblastoma (NB). Prognostic variables that could influence clinical outcome were explored. Patients and Methods One hundred sixty-nine children diagnosed with stage 4 NB (1988 to 2008) were enrolled onto consecutive anti-GD2 murine MoAb 3F8 ± GM-CSF ± 13-cis-retinoic acid (CRA) protocols after achieving first remission (complete remission/very good partial remission). Patients enrolled in regimen A (n = 43 high-risk [HR] patients) received 3F8 alone; regimen B (n = 41 HR patients), 3F8 + intravenous GM-CSF + CRA, after stem-cell transplantation (SCT); and regimen C (n = 85), 3F8 + subcutaneous GM-CSF + CRA, 46 of 85 after SCT, whereas 28 of 85 required additional induction therapy and were deemed ultra high risk (UHR). Marrow minimal residual disease (MRD) was measured by quantitative reverse transcription polymerase chain reaction. Survival probability was calculated by the Kaplan-Meier method, and prognostic variables were analyzed by multivariate Cox regression model. Results At 5 years from the start of immunotherapy, progression-free survival (PFS) improved from 44% for HR patients receiving regimen A to 56% and 62% for those receiving regimens B and C, respectively. Overall survival (OS) was 49%, 61%, and 81%, respectively. PFS and OS of UHR patients were 36% and 75%, respectively. Relapse was mostly at isolated sites. Independent adverse prognostic factors included UHR (PFS) and post–cycle two MRD (PFS and OS), whereas the prognostic factors for improved outcome were missing killer immunoglobulin-like receptor ligand (PFS and OS), human antimouse antibody response (OS), and regimen C (OS). Conclusion Retrospective analysis of consecutive trials from a single center demonstrated that MoAb 3F8 + GM-CSF + CRA is effective against chemotherapy-resistant marrow MRD. Its positive impact on long-term survival can only

  11. Recombinant human granulocyte colony stimulating factor pre-screening and screening of stabilizing carbohydrates and polyols.

    PubMed

    Pavisić, Renata; Hock, Karlo; Mijić, Ivana; Horvatić, Anita; Gecan, Martina; Sedić, Mirela; Krajacić, Mirjana Bukvić; Cindrić, Mario

    2010-03-15

    Protein stabilization by solvent additives is frequently used concept in formulation development, although new technologies implemented over the past decade can improve protein biophysical as well as clinical properties by protein structural design (e.g. PEGylation, acylation, hesylation). The scope of this work was to evaluate the effect of chosen carbohydrate or polyol stabilizer in the formulation; firstly on linear peptide sequences on instable model of rHuG-CSF cleaved macromolecule by novel method named protein and peptide stabilizer pre-screening PPSP (formulated tryptic digest mixture stability evaluation in 54 h) and on overall stability of rHuG-CSF macromolecule by quantifying all relevant degradation parameters. Comprehensive protein stabilizing screening study included conformational analysis of formulated rHuG-CSF protein to obtain information on its secondary structure conformational stability. Protein aggregation induced by modulating conditions in solution (e.g. thermal stress and agitation) was monitored over discrete time periods. Oxidation and deamidation, as well as truncation or hydrolysis were accurately quantified. Together with pre-screening data, obtained by fast and resourceful amino acid sequence degradation analysis by LC-MS, statistical data evaluation of stabilizing contribution of substances selected from group of carbohydrates and polyols was performed. According to the statistical interpretation of obtained results the stabilizers were ranked in the following order: turanose, D-trehalose, lactitol, acetate buffer (non-stabilized sample), xylitol, cellobiitol, sorbitol, D-lyxose, leucrose, sorbitol without polysorbate, cellobiose.

  12. New Role for Granulocyte Colony-Stimulating Factor-Induced Extracellular Signal-Regulated Kinase 1/2 in Histone Modification and Retinoic Acid Receptor α Recruitment to Gene Promoters: Relevance to Acute Promyelocytic Leukemia Cell Differentiation ▿

    PubMed Central

    Cassinat, B.; Zassadowski, F.; Ferry, C.; Llopis, L.; Bruck, N.; Lainey, E.; Duong, V.; Cras, A.; Despouy, G.; Chourbagi, O.; Beinse, G.; Fenaux, P.; Rochette Egly, C.; Chomienne, C.

    2011-01-01

    The induction of the granulocytic differentiation of leukemic cells by all-trans retinoic acid (RA) has been a major breakthrough in terms of survival for acute promyelocytic leukemia (APL) patients. Here we highlight the synergism and the underlying novel mechanism between RA and the granulocyte colony-stimulating factor (G-CSF) to restore differentiation of RA-refractory APL blasts. First, we show that in RA-refractory APL cells (UF-1 cell line), PML-RA receptor alpha (RARα) is not released from target promoters in response to RA, resulting in the maintenance of chromatin repression. Consequently, RARα cannot be recruited, and the RA target genes are not activated. We then deciphered how the combination of G-CSF and RA successfully restored the activation of RA target genes to levels achieved in RA-sensitive APL cells. We demonstrate that G-CSF restores RARα recruitment to target gene promoters through the activation of the extracellular signal-regulated kinase (ERK)/mitogen-activated protein kinase (MAPK) pathway and the subsequent derepression of chromatin. Thus, combinatorial activation of cytokines and RARs potentiates transcriptional activity through epigenetic modifications induced by specific signaling pathways. PMID:21262770

  13. Efficacy of delayed administration of post-chemotherapy granulocyte colony-stimulating factor: evidence from murine studies of bone marrow cell kinetics

    PubMed Central

    Yankelevich, Maxim; Goodell, Margaret A.; Kaplan, Joseph

    2008-01-01

    The optimal schedule of post-chemotherapy G-CSF administration has not been determined. G-CSF is customarily started 24 hours after chemotherapy; however, clinical data demonstrated that delaying the G-CSF until 5 days after completion of chemotherapy has not resulted in a longer duration of neutropenia. Here, we examined the optimal timing of post-chemotherapy G-CSF administration in a mouse model, to show that delayed administration does not postpone the appearance of mature granulocytes in the peripheral blood. We also investigated the mechanism of decreased efficacy of the early G-CSF application after chemotherapy by characterizing the changes in bone marrow cellular composition. To our knowledge, we demonstrate for the first time, that early after chemotherapy, the bone marrow is predominantly composed of mature residual granulocytes and very few progenitors and precursors, on which G-CSF would act to generate granulocytes. The point when immature progenitors reappear does not occur in murine bone marrow until 48 hours after a single dose of cyclophosphamide. Our results indicate that the bone marrow cellular composition early after discontinuation of chemotherapy is not optimal for G-CSF action on acceleration of myeloid recovery. Given the high cost of G-CSF prophylaxis, its delayed administration may potentially result in substantial economic benefits. PMID:17949891

  14. Cytokines in chronic inflammatory arthritis. V. Mutual antagonism between interferon-gamma and tumor necrosis factor-alpha on HLA-DR expression, proliferation, collagenase production, and granulocyte macrophage colony-stimulating factor production by rheumatoid arthritis synoviocytes.

    PubMed Central

    Alvaro-Gracia, J M; Zvaifler, N J; Firestein, G S

    1990-01-01

    The effects of a broad array of cytokines, individually and in combination, were determined on separate functions (proliferation, collagenase production, and granulocyte macrophage colony-stimulating factor [GM-CSF] production) and phenotype (expression of class II MHC antigens) of cultured fibroblast-like RA synoviocytes. The following recombinant cytokines were used: IL-1 beta, IL-2, IL-3, IL-4, IFN-gamma, tumor necrosis factor (TNF)-alpha, GM-CSF, and macrophage colony-stimulating factor (M-CSF). Only IFN-gamma induced HLA-DR (but not HLA-DQ) expression. TNF-alpha inhibited IFN-gamma-mediated HLA-DR expression (46.7 +/- 4.1% inhibition) and HLA-DR mRNA accumulation. This inhibitory effect was also observed in osteoarthritis synoviocytes. Only TNF-alpha and IL-1 increased synoviocyte proliferation (stimulation index 3.60 +/- 1.03 and 2.31 +/- 0.46, respectively). IFN-gamma (but none of the other cytokines) inhibited TNF-alpha-induced proliferation (70 +/- 14% inhibition) without affecting the activity of IL-1. Only IL-1 beta and TNF-alpha induced collagenase production (from less than 0.10 U/ml to 1.10 +/- 0.15 and 0.72 +/- 0.24, respectively). IFN-gamma decreased TNF-alpha-mediated collagenase production (69 +/- 19% inhibition) and GM-CSF production but had no effect on the action of IL-1. These data demonstrate mutual antagonism between IFN-gamma and TNF-alpha on fibroblast-like synoviocytes and suggest a novel homeostatic control mechanism that might be defective in RA where very little IFN-gamma is produced. Images PMID:2174906

  15. Accelerated phagocytosis of amyloid-beta by mouse and human microglia overexpressing the macrophage colony-stimulating factor receptor.

    PubMed

    Mitrasinovic, Olivera M; Murphy, Greer M

    2002-08-16

    Microglia surrounding A beta plaques in Alzheimer's disease and in the APPV717F transgenic mouse model of Alzheimer's disease have enhanced immunoreactivity for the macrophage colony-stimulating factor receptor (M-CSFR), encoded by the proto-oncogene c-fms. Increased expression of M-CSFR on cultured microglia results in proliferation and release of pro-inflammatory cytokines and expression of inducible nitric-oxide synthase. We transfected mouse BV-2 and human SV-A3 microglia to overexpress M-CSFR and examined microglial phagocytosis of fluorescein-conjugated A beta. Flow cytometry and laser confocal microscopy showed accelerated phagocytosis of A beta in mouse and human microglia because of M-CSFR overexpression that was time- and concentration-dependent. In contrast, microglial uptake of 1-microm diameter polystyrene microspheres was not enhanced by M-CSFR overexpression. Microglial uptake of A beta was blocked by cytochalasin D, which inhibits phagocytosis. M-CSFR overexpression increased the mRNA for macrophage scavenger receptor A, and fucoidan blocking of macrophage scavenger receptors inhibited uptake of A beta. M-CSFR antibody blocking experiments demonstrated that increased A beta uptake depended on the interaction of the M-CSFR with its ligand. These results suggest that overexpression of M-CSFR in APPV717F mice may prime microglia for phagocytosis of A beta after immunization.

  16. Early measurement of CD34+ cells in peripheral blood after cyclophosphamide and granulocyte colony-stimulating factor treatment predicts later CD34+ mobilisation failure and is a possible criterion for guiding “on demand” use of plerixafor

    PubMed Central

    Milone, Giuseppe; Tripepi, Giovanni; Martino, Massimo; Ancora, Flavia; Bartolozzi, Benedetta; Spadaro, Andrea; Nozzoli, Chiara; La Fauci, Alessia; Amico, Irene; Leotta, Salvatore; Poidomani, Massimo; Irrera, Giuseppe; Iacopino, Pasquale; Saccardi, Riccardo; Guidi, Stefano; Bosi, Alberto

    2013-01-01

    Background Early identification of predictive factors of failure to mobilise CD34+ cells could enable rational use of plerixafor during first mobilisation, avoiding the need for a second mobilisation course. However, “on demand” administration of plerixafor needs to be driven by established parameters to avoid inappropriate use. Materials and methods To address this issue, we studied the value of the peripheral blood CD34+ count, measured early (on days +10, +11, +12 and +13), in predicting the mobilisation outcome in the ensuing days. We retrospectively collected data from three Italian centres on 233 patients affected by multiple myeloma or lymphoma who underwent a first or second attempt at mobilisation with cyclophosphamide 4 g/m2 and granulocyte colony-stimulating factor. To assess the diagnostic value of peripheral blood white blood cell and CD34+ cell counts with respect to “mobilisation failure”, we considered failed mobilisation as “disease” and the CD34+ cell count in peripheral blood, on a specific day, as a “diagnostic test”. For various thresholds, we measured sensitivity, false positive rate, specificity and positive predictive value (PPV) as well as the area under the receiver-operating characteristic curves (AUC). Results A CD34+ cell count <10×106/L on day 13 had high sensitivity (1.00) and high specificity (1.00) for predicting subsequent mobilisation failure, with an AUC of 1.0. However, good prediction was also obtained using a lower threshold (CD34+ cell count: <6×106/L) at an earlier time (day 12). The PPV of the day 13 threshold was 1.00 while that of the day 12 one was 0.87. Discussion We propose that patients with <6×106/L CD34+ cells in peripheral blood on day 12 and <10×106/L on day 13 following mobilisation with cyclophosphamide 4 g/m2 and granulocyte colony-stimulating factor are candidates for “on demand” use of plerixafor, making the administration of this expensive agent more efficient and avoiding its

  17. A novel function of colony-stimulating factor 1 receptor in hTERT immortalization of human epithelial cells.

    PubMed

    Li, N F; Kocher, H M; Salako, M A; Obermueller, E; Sandle, J; Balkwill, F

    2009-02-01

    The receptor for macrophage colony-stimulating factor 1 receptor (CSF1R) is a product of the proto-oncogene c-fms and a member of the class III transmembrane tyrosine kinase receptor family. Earlier, we described increased mRNA expression of CSF1R in human telomerase reverse transcriptase (hTERT) immortalized human ovarian surface epithelial (IOSE) cell lines derived from a single donor. Here, we further describe that CSF1R is upregulated at both the mRNA and protein level in hTERT immortalized human normal OSE cells from two different donors and in hTERT immortalized human pancreatic ductal epithelial cells. CSF1R was not upregulated in hTERT immortalized epithelial clones that subsequently underwent senescence or in immortalized fibroblasts. Upon stimulation by the CSF1R ligand CSF1, the immortalized epithelial cell lines showed rapid internalization of CSF1R with concomitant down-modulation and colocalization of phosphorylated NFkappaBp65 with hTERT protein, hTERT translocation into the nucleus and the binding of c-Myc to the hTERT promoter region. Reducing the expression of CSF1R using short hairpin interfering RNA abolished these effects and also decreased cell survival and the number of population doublings under suboptimal culture conditions. The telomerase inhibitor GRN163L confirmed a role for telomerase in the cleavage of the intracellular domain of CSF1R. On the basis of these findings, we suggest that CSF1R may be a critical factor facilitating hTERT immortalization of epithelial cells.

  18. Nicotine can skew the characterization of the macrophage type-1 (M{Phi}1) phenotype differentiated with granulocyte-macrophage colony-stimulating factor to the M{Phi}2 phenotype

    SciTech Connect

    Yanagita, Manabu; Kobayashi, Ryohei; Murakami, Shinya

    2009-10-09

    Macrophages (M{Phi}s) exhibit functional heterogeneity and plasticity in the local microenvironment. Recently, it was reported that M{Phi}s can be divided into proinflammatory M{Phi}s (M{Phi}1) and anti-inflammatory M{Phi}s (M{Phi}2) based on their polarized functional properties. Here, we report that nicotine, the major ingredient of cigarette smoke, can modulate the characteristics of M{Phi}1. Granulocyte-macrophage colony-stimulating factor-driven M{Phi}1 with nicotine (Ni-M{Phi}1) showed the phenotypic characteristics of M{Phi}2. Like M{Phi}2, Ni-M{Phi}1 exhibited antigen-uptake activities. Ni-M{Phi}1 suppressed IL-12, but maintained IL-10 and produced high amounts of MCP-1 upon lipopolysaccharide stimulation compared with M{Phi}1. Moreover, we observed strong proliferative responses of T cells to lipopolysaccharide-stimulated M{Phi}1, whereas Ni-M{Phi}1 reduced T cell proliferation and inhibited IFN-{gamma} production by T cells. These results suggest that nicotine can change the functional characteristics of M{Phi} and skew the M{Phi}1 phenotype to M{Phi}2. We propose that nicotine is a potent regulator that modulates immune responses in microenvironments.

  19. Randomized study of granulocyte colony stimulating factor for childhood B-cell non-Hodgkin lymphoma: a report from the Japanese pediatric leukemia/lymphoma study group B-NHL03 study.

    PubMed

    Tsurusawa, Masahito; Watanabe, Tomoyuki; Gosho, Masahiko; Mori, Tetsuya; Mitsui, Tetsuo; Sunami, Shosuke; Kobayashi, Ryoji; Fukano, Reiji; Tanaka, Fumiko; Fujita, Naoto; Inada, Hiroko; Sekimizu, Masahiro; Koh, Katsuyoshi; Kosaka, Yoshiyuki; Komada, Yoshihiro; Saito, Akiko M; Nakazawa, Atsuko; Horibe, Keizo

    2016-07-01

    The objective of this study was to assess the impact of the primary prophylaxis of granulocyte colony-stimulating factor (G-CSF) in the management of childhood B-cell non-Hodgkin lymphoma (B-NHL). Patients with advanced-stage mature B-NHL were randomized to receive prophylactic G-CSF (G-CSF+) or not receive G-CSF (G-CSF-) based on protocols of the B-NHL03 study. The G-CSF group received 5 μg/kg/d Lenograstim from day 2 after each course of six chemotherapy courses. Fifty-eight patients were assessable, 29 G-CSF + and 29 G-CSF-. G-CSF + patients showed a positive impact on the meantime to neutrophil recovery and hospital stay. On the other hand, they had no impact in the incidences of febrile neutropenia, serious infections, stomatitis and total cost. Our study showed that administration of prophylactic G-CSF through all six chemotherapy courses for childhood B-NHL showed no clinical and economic benefits for the management of childhood B-NHL treatment.

  20. Sustained receptor activation and hyperproliferation in response to granulocyte colony-stimulating factor (G-CSF) in mice with a severe congenital neutropenia/acute myeloid leukemia-derived mutation in the G-CSF receptor gene.

    PubMed

    Hermans, M H; Antonissen, C; Ward, A C; Mayen, A E; Ploemacher, R E; Touw, I P

    1999-02-15

    In approximately 20% of cases of severe congenital neutropenia (SCN), mutations are found in the gene encoding the granulocyte colony-stimulating factor receptor (G-CSF-R). These mutations introduce premature stop codons, which result in truncation of 82-98 COOH-terminal amino acids of the receptor. SCN patients who develop secondary myelodysplastic syndrome and acute myeloid leukemia almost invariably acquired a GCSFR mutation, suggesting that this genetic alteration represents a key step in leukemogenesis. Here we show that an equivalent mutation targeted in mice (gcsfr-Delta715) results in the selective expansion of the G-CSF- responsive progenitor (G-CFC) compartment in the bone marrow. In addition, in vivo treatment of gcsfr-Delta715 mice with G-CSF results in increased production of neutrophils leading to a sustained neutrophilia. This hyperproliferative response to G-CSF is accompanied by prolonged activation of signal transducer and activator of transcription (STAT) complexes and extended cell surface expression of mutant receptors due to defective internalization. In view of the continuous G-CSF treatment of SCN patients, these data provide insight into why progenitor cells expressing truncated receptors clonally expand in vivo, and why these cells may be targets for additional genetic events leading to leukemia. PMID:9989983

  1. Sustained Receptor Activation and Hyperproliferation in Response to Granulocyte Colony-stimulating Factor (G-CSF) in Mice with a Severe Congenital Neutropenia/Acute Myeloid Leukemia–derived Mutation in the G-CSF Receptor Gene

    PubMed Central

    Hermans, Mirjam H.A.; Antonissen, Claudia; Ward, Alister C.; Mayen, Angelique E.M.; Ploemacher, Rob E.; Touw, Ivo P.

    1999-01-01

    In approximately 20% of cases of severe congenital neutropenia (SCN), mutations are found in the gene encoding the granulocyte colony-stimulating factor receptor (G-CSF–R). These mutations introduce premature stop codons, which result in truncation of 82–98 COOH-terminal amino acids of the receptor. SCN patients who develop secondary myelodysplastic syndrome and acute myeloid leukemia almost invariably acquired a GCSFR mutation, suggesting that this genetic alteration represents a key step in leukemogenesis. Here we show that an equivalent mutation targeted in mice (gcsfr-Δ715) results in the selective expansion of the G-CSF– responsive progenitor (G-CFC) compartment in the bone marrow. In addition, in vivo treatment of gcsfr-Δ715 mice with G-CSF results in increased production of neutrophils leading to a sustained neutrophilia. This hyperproliferative response to G-CSF is accompanied by prolonged activation of signal transducer and activator of transcription (STAT) complexes and extended cell surface expression of mutant receptors due to defective internalization. In view of the continuous G-CSF treatment of SCN patients, these data provide insight into why progenitor cells expressing truncated receptors clonally expand in vivo, and why these cells may be targets for additional genetic events leading to leukemia. PMID:9989983

  2. Sustained receptor activation and hyperproliferation in response to granulocyte colony-stimulating factor (G-CSF) in mice with a severe congenital neutropenia/acute myeloid leukemia-derived mutation in the G-CSF receptor gene.

    PubMed

    Hermans, M H; Antonissen, C; Ward, A C; Mayen, A E; Ploemacher, R E; Touw, I P

    1999-02-15

    In approximately 20% of cases of severe congenital neutropenia (SCN), mutations are found in the gene encoding the granulocyte colony-stimulating factor receptor (G-CSF-R). These mutations introduce premature stop codons, which result in truncation of 82-98 COOH-terminal amino acids of the receptor. SCN patients who develop secondary myelodysplastic syndrome and acute myeloid leukemia almost invariably acquired a GCSFR mutation, suggesting that this genetic alteration represents a key step in leukemogenesis. Here we show that an equivalent mutation targeted in mice (gcsfr-Delta715) results in the selective expansion of the G-CSF- responsive progenitor (G-CFC) compartment in the bone marrow. In addition, in vivo treatment of gcsfr-Delta715 mice with G-CSF results in increased production of neutrophils leading to a sustained neutrophilia. This hyperproliferative response to G-CSF is accompanied by prolonged activation of signal transducer and activator of transcription (STAT) complexes and extended cell surface expression of mutant receptors due to defective internalization. In view of the continuous G-CSF treatment of SCN patients, these data provide insight into why progenitor cells expressing truncated receptors clonally expand in vivo, and why these cells may be targets for additional genetic events leading to leukemia.

  3. Local administration of granulocyte macrophage colony-stimulating factor induces local accumulation of dendritic cells and antigen-specific CD8+ T cells and enhances dendritic cell cross-presentation

    PubMed Central

    Lee, Sung-Jong; Song, Liwen; Yang, Ming-Cheh; Mao, Chih-Ping; Yang, Benjamin; Yang, Andrew; Jeang, Jessica; Peng, Shiwen; Wu, T.-C.; Hung, Chien-Fu

    2015-01-01

    Immunotherapy has emerged as a promising treatment strategy for the control of HPV-associated malignancies. Various therapeutic HPV vaccines have elicited potent antigen-specific CD8+ T cell mediated antitumor immune responses in preclinical models and are currently being tested in several clinical trials. Recent evidence indicates the importance of local immune activation, and higher number of immune cells in the site of lesion correlates with positive prognosis. Granulocyte macrophage colony-stimulating factor (GMCSF) has been reported to posses the ability to induce migration of antigen presentation cells and CD8+ T cells. Therefore, in the current study, we employ a combination of systemic therapeutic HPV DNA vaccination with local GMCSF application in the TC-1 tumor model. We show that intramuscular vaccination with CRT/E7 DNA followed by GMCSF intravaginal administration effectively controls cervicovaginal TC-1 tumors in mice. Furthermore, we observe an increase in the accumulation of E7-specific CD8+ T cells and dendritic cells in vaginal tumors following the combination treatment. In addition, we show that GMCSF induces activation and maturation in dendritic cells and promotes antigen cross-presentation. Our results support the clinical translation of the combination treatment of systemic therapeutic vaccination followed by local GMCSF administration as an effective strategy for tumor treatment. PMID:25701675

  4. Prophylactic Administration of Vector-Encoded Porcine Granulocyte-Colony Stimulating Factor Reduces Salmonella Shedding, Tonsil Colonization, and Microbiota Alterations of the Gastrointestinal Tract in Salmonella-Challenged Swine

    PubMed Central

    Bearson, Shawn M. D.; Bearson, Bradley L.; Loving, Crystal L.; Allen, Heather K.; Lee, InSoo; Madson, Darin; Kehrli, Marcus E.

    2016-01-01

    Salmonella colonization of food animals is a concern for animal health and public health as a food safety risk. Various obstacles impede the effort to reduce asymptomatic Salmonella carriage in food animals, including the existence of numerous serovars and the ubiquitous nature of Salmonella. To develop an intervention strategy that is non-specific yet effective against diverse Salmonella serovars, we explored the prophylactic use of a cytokine to decrease Salmonella in swine by boosting the host’s innate immune system. Granulocyte-colony stimulating factor (G-CSF) is the major cytokine regulating the production, differentiation, function, and survival of neutrophils. Neutrophils play a critical role in the response to Salmonella; therefore, we evaluated the vectored-delivery of porcine G-CSF as a prophylactic to reduce Salmonella in pigs. Crossbred pigs, 5 weeks of age, were intramuscularly injected with a replication-defective human adenovirus (Ad5) engineered to express porcine G-CSF (Ad5-G-CSF, n = 9). Control pigs received the same Ad5 vector lacking the gene encoding G-CSF (Ad5-empty, n = 7). Four days later, all pigs (n = 16) were intranasally inoculated with 1 × 107 colony forming unit (CFU) of Salmonella enterica serovar Typhimurium UK1. At 2 and 3 days post-challenge with Salmonella, Ad5-G-CSF-treated pigs shed significantly less Salmonella (~103 CFU/g) in their feces than Ad5-empty-treated pigs (~104–105 CFU/g; P < 0.05). A significant 4-log reduction in tonsil colonization was also observed in the Ad5-G-CSF-treated pigs at 7 days post-challenge (P < 0.05). In the gastrointestinal tract, the Peyer’s patch region of the ileum exhibited a significant 0.5-log reduction in colonization in the Ad5-G-CSF-treated pigs (P < 0.05). The microbiota of all challenged pigs was assessed by sequencing and analyzing the V1–V3 region of the 16S rRNA gene from fecal DNA samples. The microbial community structure of

  5. Prophylactic Administration of Vector-Encoded Porcine Granulocyte-Colony Stimulating Factor Reduces Salmonella Shedding, Tonsil Colonization, and Microbiota Alterations of the Gastrointestinal Tract in Salmonella-Challenged Swine

    PubMed Central

    Bearson, Shawn M. D.; Bearson, Bradley L.; Loving, Crystal L.; Allen, Heather K.; Lee, InSoo; Madson, Darin; Kehrli, Marcus E.

    2016-01-01

    Salmonella colonization of food animals is a concern for animal health and public health as a food safety risk. Various obstacles impede the effort to reduce asymptomatic Salmonella carriage in food animals, including the existence of numerous serovars and the ubiquitous nature of Salmonella. To develop an intervention strategy that is non-specific yet effective against diverse Salmonella serovars, we explored the prophylactic use of a cytokine to decrease Salmonella in swine by boosting the host’s innate immune system. Granulocyte-colony stimulating factor (G-CSF) is the major cytokine regulating the production, differentiation, function, and survival of neutrophils. Neutrophils play a critical role in the response to Salmonella; therefore, we evaluated the vectored-delivery of porcine G-CSF as a prophylactic to reduce Salmonella in pigs. Crossbred pigs, 5 weeks of age, were intramuscularly injected with a replication-defective human adenovirus (Ad5) engineered to express porcine G-CSF (Ad5-G-CSF, n = 9). Control pigs received the same Ad5 vector lacking the gene encoding G-CSF (Ad5-empty, n = 7). Four days later, all pigs (n = 16) were intranasally inoculated with 1 × 107 colony forming unit (CFU) of Salmonella enterica serovar Typhimurium UK1. At 2 and 3 days post-challenge with Salmonella, Ad5-G-CSF-treated pigs shed significantly less Salmonella (~103 CFU/g) in their feces than Ad5-empty-treated pigs (~104–105 CFU/g; P < 0.05). A significant 4-log reduction in tonsil colonization was also observed in the Ad5-G-CSF-treated pigs at 7 days post-challenge (P < 0.05). In the gastrointestinal tract, the Peyer’s patch region of the ileum exhibited a significant 0.5-log reduction in colonization in the Ad5-G-CSF-treated pigs (P < 0.05). The microbiota of all challenged pigs was assessed by sequencing and analyzing the V1–V3 region of the 16S rRNA gene from fecal DNA samples. The microbial community structure of

  6. Prophylactic Administration of Vector-Encoded Porcine Granulocyte-Colony Stimulating Factor Reduces Salmonella Shedding, Tonsil Colonization, and Microbiota Alterations of the Gastrointestinal Tract in Salmonella-Challenged Swine.

    PubMed

    Bearson, Shawn M D; Bearson, Bradley L; Loving, Crystal L; Allen, Heather K; Lee, InSoo; Madson, Darin; Kehrli, Marcus E

    2016-01-01

    Salmonella colonization of food animals is a concern for animal health and public health as a food safety risk. Various obstacles impede the effort to reduce asymptomatic Salmonella carriage in food animals, including the existence of numerous serovars and the ubiquitous nature of Salmonella. To develop an intervention strategy that is non-specific yet effective against diverse Salmonella serovars, we explored the prophylactic use of a cytokine to decrease Salmonella in swine by boosting the host's innate immune system. Granulocyte-colony stimulating factor (G-CSF) is the major cytokine regulating the production, differentiation, function, and survival of neutrophils. Neutrophils play a critical role in the response to Salmonella; therefore, we evaluated the vectored-delivery of porcine G-CSF as a prophylactic to reduce Salmonella in pigs. Crossbred pigs, 5 weeks of age, were intramuscularly injected with a replication-defective human adenovirus (Ad5) engineered to express porcine G-CSF (Ad5-G-CSF, n = 9). Control pigs received the same Ad5 vector lacking the gene encoding G-CSF (Ad5-empty, n = 7). Four days later, all pigs (n = 16) were intranasally inoculated with 1 × 10(7) colony forming unit (CFU) of Salmonella enterica serovar Typhimurium UK1. At 2 and 3 days post-challenge with Salmonella, Ad5-G-CSF-treated pigs shed significantly less Salmonella (~10(3) CFU/g) in their feces than Ad5-empty-treated pigs (~10(4)-10(5) CFU/g; P < 0.05). A significant 4-log reduction in tonsil colonization was also observed in the Ad5-G-CSF-treated pigs at 7 days post-challenge (P < 0.05). In the gastrointestinal tract, the Peyer's patch region of the ileum exhibited a significant 0.5-log reduction in colonization in the Ad5-G-CSF-treated pigs (P < 0.05). The microbiota of all challenged pigs was assessed by sequencing and analyzing the V1-V3 region of the 16S rRNA gene from fecal DNA samples. The microbial community structure of

  7. Propofol pretreatment attenuates LPS-induced granulocyte-macrophage colony-stimulating factor production in cultured hepatocytes by suppressing MAPK/ERK activity and NF-{kappa}B translocation

    SciTech Connect

    Jawan, Bruno; Kao, Y.-H.; Goto, Shigeru; Pan, M.-C.; Lin, Y.-C.; Hsu, L.-W.; Nakano, Toshiaki; Lai, C.-Y.; Sun, C.-K.; Cheng, Y.-F.; Tai, M.-H.

    2008-06-15

    Propofol (PPF), a widely used intravenous anesthetic for induction and maintenance of anesthesia during surgeries, was found to possess suppressive effect on host immunity. This study aimed at investigating whether PPF plays a modulatory role in the lipopolysaccharide (LPS)-induced inflammatory cytokine expression in a cell line of rat hepatocytes. Morphological observation and viability assay showed that PPF exhibits no cytotoxicity at concentrations up to 300 {mu}M after 48 h incubation. Pretreatment with 100 {mu}M PPF for 24 h prior to LPS stimulation was performed to investigate the modulatory effect on LPS-induced inflammatory gene production. The results of semi-quantitative RT-PCR demonstrated that PPF pretreatment significantly suppressed the LPS-induced toll-like receptor (TLR)-4, CD14, tumor necrosis factor (TNF)-{alpha}, and granulocyte-macrophage colony-stimulating factor (GM-CSF) gene expression. Western blotting analysis showed that PPF pretreatment potentiated the LPS-induced TLR-4 downregulation. Flow cytometrical analysis revealed that PPF pretreatment showed no modulatory effect on the LPS-upregulated CD14 expression on hepatocytes. In addition, PPF pretreatment attenuated the phosphorylation of mitogen-activated protein kinase/extracellular signal-regulated kinase (MAPK/ERK) and I{kappa}B{alpha}, as well as the nuclear translocation of NF-{kappa}B primed by LPS. Moreover, addition of PD98059, a MAPK kinase inhibitor, significantly suppressed the LPS-induced NF-{kappa}B nuclear translocation and GM-CSF production, suggesting that the PPF-attenuated GM-CSF production in hepatocytes may be attributed to its suppressive effect on MAPK/ERK signaling pathway. In conclusion, PPF as an anesthetic may clinically benefit those patients who are vulnerable to sepsis by alleviating sepsis-related inflammatory response in livers.

  8. Low-dose cytarabine and aclarubicin combined with granulocyte colony-stimulating factor for the treatment of relapsed or primary refractory acute lymphocytic leukemia: a retrospective study of 25 Chinese patients.

    PubMed

    Xue, Sheng-Li; Cui, Hong-Xia; Zou, Jing-Ying; Xue, Meng-Xing; Tang, Xiao-Wen; Zhang, Yan-Ming; Wu, De-Pei

    2013-12-01

    Despite improvements in treatment, the prognosis of relapsed or primary refractory acute lymphocytic leukemia (ALL) remains poor, and outcomes are worse in older adults with the short first complete remission (CR). Attainment of the second CR by salvage therapy would improve the survival of these patients and may enable them to undergo curative treatment with allogeneic hematopoietic stem cell transplantation. The fact that there are diverse salvage protocols for these adult patients but without a striking CR-induction efficacy indicates that efforts are still needed to indentify new effective reinduction regimens. In this study, the CAG regimen (cytarabine, 10 mg/m(2) subcutaneously every 12 h on days 1-14; aclarubicin, 5-7 mg/m(2) intravenously daily on days 1-8; and concurrent granulocyte colony-stimulating factor, 200 µg/m(2) /day subcutaneously) was administered to 25 patients with relapsed or refractory ALL, including 11 T-cell ALL (T-ALL) and 14 B-cell (B-ALL) patients (age range, 11-61 years; median age, 26 years), to assess its efficacy as a salvage therapy. One course of the CAG regimen resulted in an overall response [CR or partial remission (PR)] rate of 64%, a CR rate of 56% and generally mild adverse effects. An overall response was observed in all 11 T-ALL patients (10 CR and 1 PR) and 35.7% of B-ALL patients (p = 0.0009). The significant treatment potential of CAG regimen for relapsed or primary refractory ALL, especially for T-ALL patients, described in this report would prepare them for a second CR to pursue longer survival.

  9. Influence of Granulocyte-Macrophage Colony-Stimulating Factor or Influenza Vaccination on HLA-DR, Infection and Delirium Days in Immunosuppressed Surgical Patients: Double Blind, Randomised Controlled Trial

    PubMed Central

    Lachmann, Gunnar; Renius, Markus; von Haefen, Clarissa; Wernecke, Klaus-Dieter; Bahra, Marcus; Schiemann, Alexander; Paupers, Marco; Meisel, Christian

    2015-01-01

    Purpose Surgical patients are at high risk for developing infectious complications and postoperative delirium. Prolonged infections and delirium result in worse outcome. Granulocyte-macrophage colony-stimulating factor (GM-CSF) and influenza vaccination are known to increase HLA-DR on monocytes and improve immune reactivity. This study aimed to investigate whether GM-CSF or vaccination reverses monocyte deactivation. Secondary aims were whether it decreases infection and delirium days after esophageal or pancreatic resection over time. Methods In this prospective, randomized, placebo-controlled, double-blind, double dummy trial setting on an interdisciplinary ICU of a university hospital 61 patients with immunosuppression (monocytic HLA-DR [mHLA-DR] <10,000 monoclonal antibodies [mAb] per cell) on the first day after esophageal or pancreatic resection were treated with either GM-CSF (250 μg/m2/d), influenza vaccination (Mutagrip 0.5 ml/d) or placebo for a maximum of 3 consecutive days if mHLA-DR remained below 10,000 mAb per cell. HLA-DR on monocytes was measured daily until day 5 after surgery. Infections and delirium were followed up for 9 days after surgery. Primary outcome was HLA-DR on monocytes, and secondary outcomes were duration of infection and delirium. Results mHLA-DR was significantly increased compared to placebo (p < 0.001) and influenza vaccination (p < 0.001) on the second postoperative day. Compared with placebo, GM-CSF-treated patients revealed shorter duration of infection (p < 0.001); the duration of delirium was increased after vaccination (p = 0.003). Conclusion Treatment with GM-CSF in patients with postoperative immune suppression was safe and effective in restoring monocytic immune competence. Furthermore, therapy with GM-CSF reduced duration of infection in immune compromised patients. However, influenza vaccination increased duration of delirium after major surgery. Trial Registration www.controlled-trials.com ISRCTN27114642 PMID

  10. Granulocyte colony-stimulating factor potentiates differentiation induction by all-trans retinoic acid and arsenic trioxide and enhances arsenic uptake in the acute promyelocytic leukemia cell line HT93A.

    PubMed

    Iriyama, Noriyoshi; Yuan, Bo; Hatta, Yoshihiro; Horikoshi, Akira; Yoshino, Yuta; Toyoda, Hiroo; Aizawa, Shin; Takeuchi, Jin

    2012-11-01

    The effects of arsenic trioxide (ATO), all-trans retinoic acid (ATRA) and granulocyte colony-stimulating factor (G-CSF), alone or in combination, were investigated by focusing on differentiation, growth inhibition and arsenic uptake in the acute promyelocytic leukemia (APL) cell line HT93A. ATO induced differentiation at low concentrations (0.125 µM) and apoptosis at high concentrations (1-2 µM). Furthermore, ATRA induced greater differentiation than ATO. No synergistic effect of ATRA and ATO was found on differentiation. G-CSF promoted differentiation-inducing activities of both ATO and ATRA. The combination of ATRA and G-CSF showed maximum differentiation and ATO addition was not beneficial. Addition of 1 µM ATRA and/or 50 ng/ml G-CSF to ATO did not affect apoptosis compared to ATO treatment alone. ATRA induced expression of aquaporin-9 (AQP9), a transmembrane transporter recognized as a major pathway of arsenic uptake, in a time- and dose-dependent manner. However, treatment with 1 µM ATRA decreased arsenic uptake by 43.7% compared to control subject. Although G-CSF addition did not enhance AQP9 expression in the cells, the reduced arsenic uptake was recovered to the same level as that in controls. ATRA decreased cell viability and addition of 50 ng/ml G-CSF to ATRA significantly increased the number of viable cells compared with that in ATRA alone treated cells. G-CSF not only promotes differentiation-inducing activities of both ATRA and ATO, but also makes APL cells vulnerable to increased arsenic uptake. These observations provide new insights into combination therapy using these three agents for the treatment of APL.

  11. Graft monocytic myeloid-derived suppressor cell content predicts the risk of acute graft-versus-host disease after allogeneic transplantation of granulocyte colony-stimulating factor-mobilized peripheral blood stem cells.

    PubMed

    Vendramin, Antonio; Gimondi, Silvia; Bermema, Anisa; Longoni, Paolo; Rizzitano, Sara; Corradini, Paolo; Carniti, Cristiana

    2014-12-01

    Myeloid-derived suppressor cells (MDSCs) are powerful immunomodulatory cells that in mice play a role in infectious and inflammatory disorders, including acute graft-versus-host disease (GVHD) after allogeneic hematopoietic stem cell transplantation. Their relevance in clinical acute GVHD is poorly known. We analyzed whether granulocyte colony-stimulating factor (G-CSF) administration, used to mobilize hematopoietic stem cells, affected the frequency of MDSCs in the peripheral blood stem cell grafts of 60 unrelated donors. In addition, we evaluated whether the MDSC content in the peripheral blood stem cell grafts affected the occurrence of acute GVHD in patients undergoing unrelated donor allogeneic stem cell transplantation. Systemic treatment with G-CSF induces an expansion of myeloid cells displaying the phenotype of monocytic MDSCs (Lin(low/neg)HLA-DR(-)CD11b(+)CD33(+)CD14(+)) with the ability to suppress alloreactive T cells in vitro, therefore meeting the definition of MDSCs. Monocytic MDSC dose was the only graft parameter to predict acute GVHD. The cumulative incidence of acute GVHD at 180 days after transplantation for recipients receiving monocytic MDSC doses below and above the median was 63% and 22%, respectively (P = .02). The number of monocytic MDSCs infused did not impact the relapse rate or the transplant-related mortality rate (P > .05). Although further prospective studies involving larger sample size are needed to validate the exact monocytic MDSC graft dose that protects from acute GVHD, our results strongly suggest the modulation of G-CSF might be used to affect monocytic MDSCs graft cell doses for prevention of acute GVHD.

  12. The impact of concurrent granulocyte macrophage-colony stimulating factor on radiation-induced mucositis in head and neck cancer patients: A double-blind placebo-controlled prospective Phase III study by Radiation Therapy Oncology Group 9901

    SciTech Connect

    Ryu, Janice K. . E-mail: janice.ryu@ucdmc.ucdavis.edu; Swann, Suzanne; LeVeque, Francis; Johnson, Darlene J.; Chen, Allan; Fortin, Andre; Kim, Harold; Ang, Kian K.

    2007-03-01

    Purpose: Based on early clinical evidence of potential mucosal protection by granulocyte-macrophage colony stimulating factor (GM-CSF), the Radiation Therapy Oncology Group conducted a double-blind, placebo-controlled, randomized study to test the efficacy and safety of GM-CSF in reducing the severity and duration of mucosal injury and pain (mucositis) associated with curative radiotherapy (RT) in head-and-neck cancer patients. Methods and Materials: Eligible patients included those with head-and-neck cancer with radiation ports encompassing >50% of oral cavity and/or oropharynx. Standard RT ports were used to cover the primary tumor and regional lymphatics at risk in standard fractionation to 60-70 Gy. Concurrent cisplatin chemotherapy was allowed. Patients were randomized to receive subcutaneous injection of GM-CSF 250 {mu}g/m{sup 2} or placebo 3 times a week. Mucosal reaction was assessed during the course of RT using the National Cancer Institute Common Toxicity Criteria and the protocol-specific scoring system. Results: Between October 2000 and September 2002, 130 patients from 36 institutions were accrued. Nine patients (7%) were excluded from the analysis, 3 as a result of drug unavailability. More than 80% of the patients participated in the quality-of-life endpoint of this study. The GM-CSF did not cause any increase in toxicity compared with placebo. There was no statistically significant difference in the average mean mucositis score in the GM-CSF and placebo arms by a t test (p = 0.4006). Conclusion: This placebo-controlled, randomized study demonstrated no significant effect of GM-CSF given concurrently compared with placebo in reducing the severity or duration of RT-induced mucositis in patients undergoing definitive RT for head-and-neck cancer.

  13. Improved Outcome of Refractory/Relapsed Acute Myeloid Leukemia after Post-Transplantation Cyclophosphamide-Based Haploidentical Transplantation with Myeloablative Conditioning and Early Prophylactic Granulocyte Colony-Stimulating Factor-Mobilized Donor Lymphocyte Infusions.

    PubMed

    Jaiswal, Sarita Rani; Zaman, Shamsuz; Chakrabarti, Aditi; Sen, Subrata; Mukherjee, Shashwata; Bhargava, Sneh; Ray, Kunal; O'Donnell, Paul V; Chakrabarti, Suparno

    2016-10-01

    We carried out post-transplantation cyclophosphamide (PTCy)-based haploidentical peripheral blood stem cell transplantation in 51 patients with refractory/relapsed acute myeloid leukemia not in remission. The first 10 patients received nonmyeloablative conditioning followed by planned granulocyte colony-stimulating factor (G-CSF)-mobilized donor lymphocyte infusions (DLIs) on days 35, 60, and 90. No patient developed graft-versus-host disease (GVHD), but 90% had disease progression between 3 and 6 months. A subsequent 41 patients received myeloablative conditioning (MAC); the first 20 patients did not receive DLIs (MAC group) and the next 21 patients received G-CSF-mobilized DLIs (G-DLI) on days 21, 35, and 60 (MAC-DLI group). The incidence of disease progression and progression-free survival at 18 months were 66% and 25% in the MAC group compared with 21.4% and 61.9% in the MAC-DLI group (P = .01). Chronic GVHD but not acute GVHD was increased in the MAC-DLI group (41.2% versus 11%, P = .05). Natural killer cell alloreactive donor was associated with lower incidence of disease progression in the MAC but not in MAC-DLI group. The only factor favorably influencing disease progression and progression-free survival was administration of G-DLI after myeloablative conditioning. Our study shows that early administration of G-DLI is feasible after PTCy-based haploidentical hematopoietic stem cell transplantation for refractory/relapsed acute myeloid leukemia and might be associated with improved survival after MAC. PMID:27470289

  14. Effect of Granulocyte-Macrophage Colony-Stimulating Factor on Chemotherapy-Related Neutropenia in Patients with Non-Hodgkin's Lymphomas-A Phase I/II Study of Dose and Mode of Administration.

    PubMed

    Hovgaard, D J; Nissen, N I

    1991-01-01

    The effect of mammalian glycosylated recombinant granulocyte-macrophage colony-stimulating factor was investigated in 24 patients with newly diagnosed non-Hodgkin's lymphoma in a phase I/II study. All patients received standard chemotherapy with CHOP. RhGM-CSF was administered after the first cycle for 5 days, and at one of four dose levels (2, 4, 8 and 16 μg/kg). Patients were randomized to receive the drug either by continuous intravenous infusion or twice daily as subcutaneous injection. No significant difference in results was observed between subcutaneous administration of rhGM-CSF and continuous i.v. infusion and these patient groups could therefore be combined in the analysis. Administration of rhGM-CSF resulted in a significant dose-dependent increase of total WBC, mainly neutrophils, eosinophils and monocytes. The increase was observed in 18/24 patients, reaching a peak 24-72 (median 24) hours after the start of rhGM-CSF. The CHOP chemotherapy-induced leucocyte nadir occurred on day 12 (mean) compared to day 14 for the 127 historical controls. The WBC nadir values were higher (2.4 ± 1.4) than for historical controls (1.8 ± 1.1) and the leucopenic/neutropenic period was of shorter duration. Following the chemotherapy nadir a more rapid recovery of WBC was seen than in controls. GM-CSF was well tolerated, the side effects were mild and transient, and included myalgias, low grade fever, headache, chest/bone discomfort, nausea, erythema at injection site and superficial phlebitis. The encouraging results of this phase I/II study indicate the need for a prospective controlled study of GM-CSF in chemotherapy of malignant lymphoma.

  15. Comparison of neurological and functional outcomes after administration of granulocyte-colony-stimulating factor in motor-complete versus motor-incomplete postrehabilitated, chronic spinal cord injuries: a phase I/II study.

    PubMed

    Saberi, Hooshang; Derakhshanrad, Nazi; Yekaninejad, Mir Saeed

    2014-01-01

    Granulocyte-colony-stimulating factor (G-CSF) is a major growth factor in the activation and differentiation of granulocytes. This cytokine has been widely and safely employed in different disease conditions over many years. The administration of the growth factors in spinal cord injury (SCI) has been reported elsewhere; here we have tried to see the effect of SCI severity on the neurological outcomes after neuroprotective treatment for SCI with G-CSF. Seventy-four consecutive patients with SCI of at least 6 months' duration, with stable neurological status in the last 3 months, having informed consent for the treatment were included in the study. All the patients had undergone at least 3 months of standard rehabilitation. Patients were assessed by the American Spinal Injury Association (ASIA) scale, Spinal Cord Independence Measure (SCIM) III, and International Association of Neurorestoratology-Spinal Cord Injury Functional Rating Scale (IANR-SCIFRS) just before intervention and periodically until 6 months after subcutaneous administration of 5 µg/kg per day of G-CSF for 7 consecutive days. Multiple linear regression models were performed for statistical evaluation of lesion completeness and level of injury on changes in ASIA motor, light touch, pinprick, IANR-SCIFRS, and SCIM III scores, as a phase I/II comparative study. The study consisted of 52 motor-complete and 22 motor-incomplete SCI patients. There was no significant difference regarding age and sex, chronicity, and level of SCI between the two groups. Motor-incomplete patients had significantly more improvement in ASIA motor score compared to the motor-complete patients (7.68 scores, p < 0.001); also they had significant improvement in light touch (6.42 scores, p = 0.003) and pinprick sensory scores (4.89 scores, p = 0.011). Therefore, G-CSF administration in motor-incomplete SCIs is associated with significantly higher motor improvement, and also the higher the initial ASIA Impairment Scale

  16. Late Onset Agranulocytosis with Clozapine Associated with HLA DR4 Responding to Treatment with Granulocyte Colony-stimulating Factor: A Case Report and Review of Literature

    PubMed Central

    Singh, Aakanksha; Grover, Sandeep; Malhotra, Pankaj; Varma, Subhash C.

    2016-01-01

    Agranulocytosis as a side effect of clozapine has been reported to be associated with initial phases of treatment, i.e., first six months. Agranulocytosis with clozapine during the initial phases of treatment has been linked to genetic vulnerability in the form of variations in the human leukocyte-antigen haplotypes. However, there is limited literature on late onset agranulocytosis with clozapine and this has very rarely been linked to human leukocyte-antigen haplotypes vulnerability. In this report we review the existing data on late onset agranulocytosis with clozapine and describe the case of a young man, who developed agranulocytosis with clozapine after 35 months of treatment and was found to have genetic vulnerability in form of being positive for HLA DR4. This case highlights underlying autoimmune immune mechanism in clozapine-induced agranulocytosis and the need for frequent blood count monitoring on clozapine even after the initial 6 months of starting treatment especially in patients with genetic vulnerability to develop this condition. PMID:27121434

  17. Establishment of the first international standard for PEGylated granulocyte colony stimulating factor (PEG-G-CSF): Report of an international collaborative study

    PubMed Central

    Wadhwa, Meenu; Bird, Chris; Dougall, Thomas; Rigsby, Peter; Bristow, Adrian; Thorpe, Robin

    2015-01-01

    We assessed the feasibility of developing a suitable international reference standard for determination of in vitro biological activity of human sequence recombinant PEG-G-CSF products with a 20 kD linear PEG linked to the N-terminal methionyl residue of G-CSF (INN Filgrastim), produced using a conjugation process and coupling chemistry similar to that employed for the lead PEGfilgrastim product. Based on initial data which showed that the current WHO 2nd international standard, IS for G-CSF (09/136) or alternatively, a PEG-G-CSF standard with a unitage traceable to the G-CSF IS may potentially serve as the IS for PEG-G-CSF products, two candidate preparations of PEG-G-CSF were formulated and lyophilized at NIBSC. These preparations were tested by 23 laboratories using in vitro bioassays in a multi-centre collaborative study. Results indicated that on the basis of parallelism, the current WHO 2nd IS for G-CSF or any of the PEG-G-CSF samples could be used as the international standard for PEG-G-CSF preparations. However, because of the variability in potency estimates seen when PEG-G-CSF preparations were compared with the current WHO 2nd IS for G-CSF, a candidate PEG-G-CSF was suitable as the WHO IS. The preparation 12/188 was judged suitable to serve as the WHO IS based on in vitro biological activity data. Therefore, the preparation coded 12/188 was established by the WHO Expert Committee on Biological Standardization (ECBS) in 2013 as the WHO 1st IS for human PEGylated G-CSF with an assigned in vitro bioactivity of 10,000 IU per ampoule. PMID:25450254

  18. Cyclic AMP-elevating agents down-regulate the oxidative burst induced by granulocyte-macrophage colony-stimulating factor (GM-CSF) in adherent neutrophils.

    PubMed Central

    Ottonello, L; Morone, M P; Dapino, P; Dallegri, F

    1995-01-01

    Human neutrophils, plated on fibronectin-precoated wells, were found to release large quantities of superoxide anion (O2-) in response to GM-CSF. O2- production was reduced by prostaglandin E2 (PGE2) and the phosphodiesterase type IV (PDE IV) inhibitor RO 20-1724. Both agents are known to increase intracellular cyclic AMP (cAMP) levels by inducing its production (PGE2) or blocking its catabolism (RO 20-1724). When added in combination, PGE2 and RO 20-1724 had a marked synergistic inhibitory effect, which was reproduced by replacing PGE2 with a direct activator of adenylate cyclase, i.e. forskolin (FK). Moreover, the neutrophil response to GM-CSF was inhibited by a membrane-permeable analogue of cAMP in a dose-dependent manner. As GM-CSF and PGE2 are known to be generated at tissue sites of inflammation, the results suggest the existence of a PGE2-dependent regulatory pathway potentially capable of controlling the neutrophil response to GM-CSF, in turn limiting the risk of local oxidative tissue injury. Moreover, owing to its susceptibility to amplification by RO 20-1724, the PGE2-dependent pathway and in particular PDE-IV may represent a pharmacological target to reduce the generation of histotoxic oxidants by GM-CSF-responding neutrophils. PMID:7664497

  19. cAMP suppresses p21ras and Raf-1 responses but not the Erk-1 response to granulocyte-colony-stimulating factor: possible Raf-1-independent activation of Erk-1.

    PubMed Central

    Csar, X F; Ward, A C; Hoffmann, B W; Guy, G G; Hamilton, J A

    1997-01-01

    The cAMP analogue 8-bromo-cAMP (8BrcAMP) inhibits granulocyte-colony-stimulating factor (G-CSF)-stimulated DNA synthesis in myeloid NFS-60 cells. We examined the effect of 8BrcAMP addition on the G-CSF-stimulated extracellular signal-related protein kinase 1 (Erk-1), p21ras and Raf-1 activation. The Erk-1 activity was not down-regulated by the increase in intracellular cAMP levels, whereas p21ras and Raf-1 activities were, suggesting that Erk-1 activity might not be dependent on upstream p21ras and/or Raf-1 activity in this system. To explore this possibility further, we sought to determine whether there were downstream substrates of Raf-1 that were distinguishable from those of Erk-1 by using two-dimensional SDS/PAGE analysis of the protein phosphorylation patterns of NFS-60 cell cytosolic extracts treated with exogenous Raf-1 or Erk-1 in the presence of [gamma-32P]ATP. The two phosphorylation patterns were found to have many differences. To gain further insights into the possible relevance of these phosphorylation patterns and as an approach to exploring in more detail the inhibitory effect of 8BrcAMP, two-dimensional SDS/PAGE analysis was performed on the cytosolic extracts of 32P-labelled NFS-60 cells treated with G-CSF, in the absence or presence of 8BrcAMP. It was found that the phosphorylated proteins whose appearance was specific to the action of exogenous Raf-1 were sensitive to the action of 8BrcAMP in vivo, whereas those whose appearance was specific to the action of exogenous Erk-1 alone, or common to the actions of Raf-1 and Erk-1, were 8BrcAMP-insensitive. The results are consistent with a Raf-1-independent pathway for Erk-1 activation in G-CSF treated myeloid cells, and a number of potential downstream substrates of these kinases have been identified for further characterization. PMID:9078246

  20. Induction of cytokine granulocyte-macrophage colony-stimulating factor and chemokine macrophage inflammatory protein 2 mRNAs in macrophages by Legionella pneumophila or Salmonella typhimurium attachment requires different ligand-receptor systems.

    PubMed Central

    Yamamoto, Y; Klein, T W; Friedman, H

    1996-01-01

    The attachment of bacteria to macrophages is mediated by different ligands and receptors and induces various intracellular molecular responses. In the present study, induction of cytokines and chemokines, especially granulocyte-macrophage colony-stimulating factor (GM-CSF) and macrophage inflammatory protein 2 (MIP-2), was examined, following bacterial attachment, with regard to the ligand-receptor systems involved. Attachment of Legionella pneumophila or Salmonella typhimurium to cultured mouse peritoneal macrophages increased the steady-state levels of cellular mRNAs for the cytokines interleukin 1beta (IL-1beta), IL-6, and GM-CSF as well as the chemokines MIP-1beta, MIP-2, and KC. However, when macrophages were treated with alpha-methyl-D-mannoside (alphaMM), a competitor of glycopeptide ligands, induction of cytokine mRNAs was inhibited, but the levels of chemokine mRNAs were not. Pretreatment of the bacteria with fresh mouse serum enhanced the level of GM-CSF mRNA but not the level of MIP-2 mRNA. In addition, serum treatment reduced the inhibitory effect of alphaMM on GM-CSF mRNA. These results indicate that bacterial attachment increases the steady-state levels of the cytokine and chemokine mRNAs tested by at least two distinct receptor-ligand systems, namely, one linked to cytokine induction and involving mannose or other sugar residues and the other linked to chemokine induction and relatively alphaMM insensitive. Furthermore, opsonization with serum engages other pathways in the cytokine response which are relatively independent of the alphaMM-sensitive system. Regarding bacterial surface ligands involved in cytokine mRNA induction, evidence is presented that the flagellum may be important in stimulating cytokine GM-CSF message but not chemokine MIP-2 message. Analysis of cytokine GM-CSF and chemokine MIP-2 signaling pathways with protein kinase inhibitors revealed the involvement of calmodulin and myosin light-chain kinase in GM-CSF but not MIP-2 m

  1. Combination treatment of biomechanical support and targeted intra-arterial infusion of peripheral blood stem cells mobilized by granulocyte-colony stimulating factor for the osteonecrosis of the femoral head: a randomized controlled clinical trial.

    PubMed

    Mao, Qiang; Wang, Weidong; Xu, Taotao; Zhang, Shanxing; Xiao, Luwei; Chen, Di; Jin, Hongting; Tong, Peijian

    2015-04-01

    The objective of this study was to determine the benefits of combination treatment with mechanical support and targeted intra-arterial infusion of peripheral blood stem cells (PBSCs) mobilized by granulocyte-colony stimulating factor (G-CSF) via the medial circumflex femoral artery on the progression of osteonecrosis of the femoral head (ONFH). Fifty-five patients (89 hips) with early and intermediate stage ONFH were recruited and randomly assigned to combination treatment or mechanical support treatment (control group). All hips received mechanical support treatment (porous tantalum rod implantation). Then, hips in the combination treatment group were performed targeted intra-arterial infusion of PBSCs. At each follow-up, Harris hip score (HHS) and Association Research Circulation Osseous (ARCO) classification were used to evaluate the symptoms and progression of osteonecrosis. Total hip arthroplasty (THA) was assessed as an endpoint at each follow-up. At 36 months, 9 of the 41 hips (21.95%) in the control group progressed to clinical failure and underwent THA whereas only 3 of the 48 hips (6.25%) in the combination treatment group required THA (p = 0.031). Kaplan-Meier survival analysis showed a significant difference in the survival time between the two groups (log-rank test; p = 0.025). Compared to the control group, combination treatment significantly improved the HHS at 36 months (p = 0.003). At the final follow-up examination, radiological progression was noted in 13 of 41 hips (31.71%) for the control group, but in only 4 of 48 hips (8.33%) for the combination treatment group (p = 0.005). The overall collapse rates were 15.15% (5/33 hips) and 8.11% (3/37 hips) in the control and combination treatment groups, respectively. Targeted intra-arterial infusion of PBSCs is capable of enhancing the efficacy of biomechanical support in the treatment of ONFH. This clinical trial confirmed that the combination treatment might be a safe and feasible

  2. Properties of bcr-abl-transformed mouse 12B1 cells secreting interleukin-2 and granulocyte-macrophage colony stimulating factor (GM-CSF): II. Adverse effects of GM-CSF.

    PubMed

    Petráčková, Martina; Staněk, Libor; Mandys, Václav; Dundr, Pavel; Vonka, Vladimír

    2012-06-01

    Granulocyte-macrophage colony stimulating factor (GM-CSF) is considered to be the most effective immunostimulating factor for the construction of gene-engineered anti-cancer vaccines. In some tumour cells, this type of genetic modification has resulted in the loss of the oncogenic potential. This was not the case with bcr-abl-transformed mouse 12B1 cells. A cell line, designated 12B1/GM-CSF/cl-5 producing more than 100 ng/106 cells/24 h, displayed higher pathogenicity than the parental, non-transduced cells. Although the tumours induced by the parental 12B1 cells and 12B1/GM-CSF/cl-5 cells appeared nearly at the same time and then grew at an approximately equal rate, the latter mice were in a much poorer clinical condition. In these animals the growth of the tumours was associated with gradually increasing blood levels of GM-CSF. In both groups of animals splenomegaly was observed; it was much more pronounced in the case of 12B1/GM-CSF/cl-5-inoculated animals. While in the case of animals inoculated with the parental cells the splenomegaly was probably mainly due to infiltration with tumour cells, in the animals inoculated with the GM-CSF-secreting cells splenomegaly and derangement of parenchymal organs, such as lungs, liver and kidneys, were more complex, including congestion and infiltration with hemopoietic cells, predominantly immature cells of myeloid lineage. The most conspicuous of these changes was the hyperaemia of the lungs. No such alterations were seen in animals inoculated with the parental cells. On the other hand, the contents of T regulatory cells were comparable in both groups and they increased in parallel at the end of the observation period. When GM-CSF neutralizing antibody was administered to animals inoculated with the 12B1/GM-CSF/cl-5 cells, the pathological changes observed within the organs were suppressed, this proving that the overproduced GM-CSF and not any other substance, played the key role in their induction.

  3. Identification of sequences within the murine granulocyte-macrophage colony-stimulating factor mRNA 3'-untranslated region that mediate mRNA stabilization induced by mitogen treatment of EL-4 thymoma cells.

    PubMed

    Iwai, Y; Bickel, M; Pluznik, D H; Cohen, R B

    1991-09-25

    Phorbol esters (TPA) and concanavalin A (ConA) are known to induce granulocyte-macrophage colony-stimulating factor (GM-CSF) production in murine thymoma EL-4 cells by mRNA stabilization. The role of the 3'-untranslated region (3'-UTR) in GM-CSF mRNA stabilization induced by TPA and ConA in EL-4 cells was examined by transfection studies using chloramphenicol acetyltransferase (CAT) constructions. The GM-CSF 3'-UTR contains a 63-nucleotide region at its 3' end with repeating ATTTA motifs which is responsible for mRNA degradation in a variety of cell types (Shaw, G., and Kamen, R. (1986) Cell 46, 659-666). We produced constructs containing most of the GM-CSF 3'-UTR (303 nucleotides, pRSV-CATgm) or the 3'-terminal AT-rich region (116 nucleotides, pRSV-CATau) and measured CAT enzyme activity and CAT mRNA after transient transfection into EL-4 and NIH 3T3 cells. Low levels of CAT activity were seen in both cells with either plasmid compared with levels of CAT activity obtained with pRSV-CAT. TPA treatment caused an approximately 10-fold increase in CAT activity and mRNA in EL-4 cells transfected with pRSV-CATgm. No increases were seen in EL-4 cells transfected with pRSV-CATau or pRSV-CAT. No response to TPA was detected in transfected NIH 3T3 cells, indicating that the response to TPA is relatively cell-specific. There was no increase in CAT activity after ConA treatment in EL-4 or NIH 3T3 cells transfected with any of the constructs suggesting that the GM-CSF 3'-UTR lacks elements that can respond alone to ConA. Nuclear run-on and actinomycin D chase experiments in EL-4 cells showed that TPA induces CAT activity via mRNA stabilization. By linker-substitution mutagenesis we show that TPA inducibility depends on a 60-nucleotide region of the 3'-UTR whose 5' end is located 160 nucleotides upstream of the 5' end of the AU-rich region. PMID:1917935

  4. cAMP suppresses p21ras and Raf-1 responses but not the Erk-1 response to granulocyte-colony-stimulating factor: possible Raf-1-independent activation of Erk-1.

    PubMed

    Csar, X F; Ward, A C; Hoffmann, B W; Guy, G G; Hamilton, J A

    1997-02-15

    The cAMP analogue 8-bromo-cAMP (8BrcAMP) inhibits granulocyte-colony-stimulating factor (G-CSF)-stimulated DNA synthesis in myeloid NFS-60 cells. We examined the effect of 8BrcAMP addition on the G-CSF-stimulated extracellular signal-related protein kinase 1 (Erk-1), p21ras and Raf-1 activation. The Erk-1 activity was not down-regulated by the increase in intracellular cAMP levels, whereas p21ras and Raf-1 activities were, suggesting that Erk-1 activity might not be dependent on upstream p21ras and/or Raf-1 activity in this system. To explore this possibility further, we sought to determine whether there were downstream substrates of Raf-1 that were distinguishable from those of Erk-1 by using two-dimensional SDS/PAGE analysis of the protein phosphorylation patterns of NFS-60 cell cytosolic extracts treated with exogenous Raf-1 or Erk-1 in the presence of [gamma-32P]ATP. The two phosphorylation patterns were found to have many differences. To gain further insights into the possible relevance of these phosphorylation patterns and as an approach to exploring in more detail the inhibitory effect of 8BrcAMP, two-dimensional SDS/PAGE analysis was performed on the cytosolic extracts of 32P-labelled NFS-60 cells treated with G-CSF, in the absence or presence of 8BrcAMP. It was found that the phosphorylated proteins whose appearance was specific to the action of exogenous Raf-1 were sensitive to the action of 8BrcAMP in vivo, whereas those whose appearance was specific to the action of exogenous Erk-1 alone, or common to the actions of Raf-1 and Erk-1, were 8BrcAMP-insensitive. The results are consistent with a Raf-1-independent pathway for Erk-1 activation in G-CSF treated myeloid cells, and a number of potential downstream substrates of these kinases have been identified for further characterization.

  5. Early applications of granulocyte colony-stimulating factor (G-CSF) can stabilize the blood–optic-nerve barrier and ameliorate inflammation in a rat model of anterior ischemic optic neuropathy (rAION)

    PubMed Central

    Wen, Yao-Tseng; Huang, Tzu-Lun; Huang, Sung-Ping; Chang, Chung-Hsing

    2016-01-01

    ABSTRACT Granulocyte colony-stimulating factor (G-CSF) was reported to have a neuroprotective effect in a rat model of anterior ischemic optic neuropathy (rAION model). However, the therapeutic window and anti-inflammatory effects of G-CSF in a rAION model have yet to be elucidated. Thus, this study aimed to determine the therapeutic window of G-CSF and investigate the mechanisms of G-CSF via regulation of optic nerve (ON) inflammation in a rAION model. Rats were treated with G-CSF on day 0, 1, 2 or 7 post-rAION induction for 5 consecutive days, and a control group were treated with phosphate-buffered saline (PBS). Visual function was assessed by flash visual evoked potentials at 4 weeks post-rAION induction. The survival rate and apoptosis of retinal ganglion cells were determined by FluoroGold labeling and TUNEL assay, respectively. ON inflammation was evaluated by staining of ED1 and Iba1, and ON vascular permeability was determined by Evans Blue extravasation. The type of macrophage polarization was evaluated using quantitative real-time PCR (qRT-PCR). The protein levels of TNF-α and IL-1β were analyzed by western blotting. A therapeutic window during which G-CSF could rescue visual function and retinal ganglion cell survival was demonstrated at day 0 and day 1 post-infarct. Macrophage infiltration was reduced by 3.1- and 1.6-fold by G-CSF treatment starting on day 0 and 1 post-rAION induction, respectively, compared with the PBS-treated group (P<0.05). This was compatible with 3.3- and 1.7-fold reductions in ON vascular permeability after G-CSF treatment compared with PBS treatment (P<0.05). Microglial activation was increased by 3.8- and 3.2-fold in the early (beginning treatment at day 0 or 1) G-CSF-treated group compared with the PBS-treated group (P<0.05). Immediate (within 30 mins of infarct) treatment with G-CSF also induced M2 microglia/macrophage activation. The cytokine levels were lower in the group that received immediate G-CSF treatment

  6. Granulocyte Colony Stimulating Factor and Physiotherapy after Stroke: Results of a Feasibility Randomised Controlled Trial: Stem Cell Trial of Recovery EnhanceMent after Stroke-3 (STEMS-3 ISRCTN16714730)

    PubMed Central

    Sprigg, Nikola; O’Connor, Rebecca; Woodhouse, Lisa; Krishnan, Kailash; England, Timothy J.; Connell, Louise A.; Walker, Marion F.; Bath, Philip M.

    2016-01-01

    Background Granulocyte-colony stimulating factor (G-CSF) mobilises endogenous haematopoietic stem cells and enhances recovery in experimental stroke. Recovery may also be dependent on an enriched environment and physical activity. G-CSF may have the potential to enhance recovery when used in combination with physiotherapy, in patients with disability late after stroke. Methods A pilot 2 x 2 factorial randomised (1:1) placebo-controlled trial of G-CSF (double-blind), and/or a 6 week course of physiotherapy, in 60 participants with disability (mRS >1), at least 3 months after stroke. Primary outcome was feasibility, acceptability and tolerability. Secondary outcomes included death, dependency, motor function and quality of life measured 90 and 365 days after enrolment. Results Recruitment to the trial was feasible and acceptable; of 118 screened patients, 92 were eligible and 32 declined to participate. 60 patients were recruited between November 2011 and July 2013. All participants received some allocated treatment. Although 29 out of 30 participants received all 5 G-CSF/placebo injections, only 7 of 30 participants received all 18 therapy sessions. G-CSF was well tolerated but associated with a tendency to more adverse events than placebo (16 vs 10 patients, p = 0.12) and serious adverse events (SAE) (9 vs 3, p = 0.10). On average, patients received 14 (out of 18 planned) therapy sessions, interquartile range [12, 17]. Only a minority (23%) of participants completed all physiotherapy sessions, a large proportion of sessions (114 of 540, 21%) were cancelled due to patient (94, 17%) and therapist factors (20, 4%). No significant differences in functional outcomes were detected in either the G-CSF or physiotherapy group at day 90 or 365. Conclusions Delivery of G-CSF is feasible in chronic stroke. However, the study failed to demonstrate feasibility for delivering additional physiotherapy sessions late after stroke therefore a definitive study using this trial design

  7. Immunizing and curative potential of replicating and nonreplicating murine mammary adenocarcinoma cells engineered with interleukin (IL)-2, IL-4, IL-6, IL-7, IL-10, tumor necrosis factor alpha, granulocyte-macrophage colony-stimulating factor, and gamma-interferon gene or admixed with conventional adjuvants.

    PubMed

    Allione, A; Consalvo, M; Nanni, P; Lollini, P L; Cavallo, F; Giovarelli, M; Forni, M; Gulino, A; Colombo, M P; Dellabona, P

    1994-12-01

    To evaluate the efficacy of vaccinations with cytokine-gene-transduced tumor cells, BALB/c mice were challenged with 1 x 10(5) parental cells of a syngeneic adenocarcinoma cell line (TSA-pc). No protection was observed in mice immunized 30 days earlier with 1 x 10(5) nonreplicating mitomycin-C-treated TSA-pc alone, or with Corynebacterium parvum or Complete Freund Adjuvant (CFA). Ten to 30% of mice immunized with nonreplicating cells engineered to produce interleukin (IL)-2, IL-4, IL-6, IL-7, IL-10, tumor necrosis factor alpha, granulocyte-macrophage colony-stimulating factor, and gamma-interferon gene were protected. Fifty % of mice immunized with replicating TSA-pc admixed with C. parvum and 80-100% of mice immunized with replicating tumor cells transduced with IL-2, IL-4, IL-7, IL-10, or gamma-interferon gene were protected. No cure was afforded by TSA cells admixed with C. parvum or CFA, nor by TSA cells engineered with IL-6, granulocyte-macrophage colony-stimulating factor, and tumor necrosis factor alpha gene injected starting 1 day after TSA-pc challenge. Complete tumor regression, however, was obtained in 10-20% of mice treated with TSA cells transduced with IL-2, IL-4, IL-7, or IL-10 and in 30% of those treated with TSA cells transduced with gamma-interferon gene. PMID:7954438

  8. Stimulation of human monocytes with macrophage colony-stimulating factor induces a Grb2-mediated association of the focal adhesion kinase pp125FAK and dynamin.

    PubMed Central

    Kharbanda, S; Saleem, A; Yuan, Z; Emoto, Y; Prasad, K V; Kufe, D

    1995-01-01

    Macrophage colony-stimulating factor (M-CSF) is required for the growth and differentiation of mononuclear phagocytes. In the present studies using human monocytes, we show that M-CSF induces interaction of the Grb2 adaptor protein with the focal adhesion kinase pp125FAK. The results demonstrate that tyrosine-phosphorylated pp125FAK directly interacts with the SH2 domain of Grb2. The findings indicate that a pYENV site at Tyr-925 in pp125FAK is responsible for this interaction. We also demonstrate that the Grb2-FAK complex associates with the GTPase dynamin. Dynamin interacts with the SH3 domains of Grb2 and exhibits M-CSF-dependent tyrosine phosphorylation in association with pp125FAK. These findings suggest that M-CSF-induced signaling involves independent Grb2-mediated pathways, one leading to Ras activation and another involving pp125FAK and a GTPase implicated in receptor internalization. Images Fig. 1 Fig. 2 Fig. 3 Fig. 4 Fig. 5 PMID:7597091

  9. (TH) diazepam binding to human granulocytes

    SciTech Connect

    Bond, P.A.; Cundall, R.L.; Rolfe, B.

    1985-07-08

    (TH)-diazepam binds to sites on human granulocyte membranes, with little or no binding to platelets or lymphocytes. These (TH)-diazepam binding sites are of the peripheral type, being strongly inhibited by R05-4864 (Ki=6.23nM) but only weakly by clonazepam (Ki=14 M). Binding of (TH) diazepam at 0 is saturable, specific and stereoselective. Scatchard analysis indicates a single class of sites with Bmax of 109 +/- 17f moles per mg of protein and K/sub D/ of 3.07 +/- 0.53nM. Hill plots of saturation experiments gave straight lines with a mean Hill coefficient of 1.03 +/- 0.014. Binding is time dependent and reversible and it varies linearly with granulocyte protein concentration over the range 0.025-0.300 mg of protein. 11 references, 3 figures, 1 table.

  10. In vitro and in vivo activation of endothelial cells by colony-stimulating factors.

    PubMed Central

    Bussolino, F; Ziche, M; Wang, J M; Alessi, D; Morbidelli, L; Cremona, O; Bosia, A; Marchisio, P C; Mantovani, A

    1991-01-01

    This study was designed to identify the set of functions activated in cultured endothelial cells by the hematopoietic growth factors, granulocyte colony-stimulating factor (G-CSF) and granulocyte macrophage-colony-stimulating factor (GM-CSF), and to compare them with those elicited by prototypic cytokines active on these cells. Moreover, indications as to the in vivo relevance of in vitro effects were obtained. G-CSF and GM-CSF induced endothelial cells to proliferate and migrate. In contrast, unlike appropriate reference cytokines (IL-1 and tumor necrosis factor, IFN-gamma), G-CSF and GM-CSF did not modulate endothelial cell functions related to hemostasis-thrombosis (production of procoagulant activity and of platelet activating factor), inflammation (expression of leukocyte adhesion molecule-1 and production of platelet activating factor), and accessory function (expression of class II antigens of MHC). Other colony-stimulating factors (IL-3 and macrophage-colony-stimulating factor) were inactive on all functions tested. In comparison to basic fibroblast growth factor (bFGF), G-CSF and GM-CSF induced lower maximal proliferation of endothelial cells, whereas migration was of the same order of magnitude. G-CSF and GM-CSF stimulated repair of mechanically wounded endothelial monolayers. Exposure to both cytokines induced shape changes and cytoskeletal reorganization consistent with a migratory phenotype. To explore the in vivo relevance of the in vitro effects of these cytokines on endothelium, we studied the angiogenic activity of human G-CSF in the rabbit cornea. G-CSF, but not the heat-inactivated molecule, had definite angiogenic activity, without any sign of inflammatory reactions. G-CSF was less active than bFGF. However, the combination of a nonangiogenic dose of bFGF with G-CSF resulted in an angiogenic response higher than that elicited by either individual cytokines. Thus, G-CSF and GM-CSF induce endothelial cells to express an activation

  11. Animal models of human granulocyte diseases.

    PubMed

    Schäffer, Alejandro A; Klein, Christoph

    2013-02-01

    In vivo animal models have proven very useful to the understanding of basic biologic pathways of the immune system, a prerequisite for the development of innovate therapies. This article addresses currently available models for defined human monogenetic defects of neutrophil granulocytes, including murine, zebrafish, and larger mammalian species. Strengths and weaknesses of each system are summarized, and clinical investigators may thus be inspired to develop further lines of research to improve diagnosis and therapy by use of the appropriate animal model system. PMID:23351993

  12. Comparative effect of recombinant IL-1, -2, -3, -4, and -6, IFN-gamma, granulocyte-macrophage-colony-stimulating factor, tumor necrosis factor-alpha, and histamine-releasing factors on the secretion of histamine from basophils

    SciTech Connect

    Alam, R.; Welter, J.B.; Forsythe, P.A.; Lett-Brown, M.A.; Grant, J.A. )

    1989-05-15

    Most cytokines possess multiple biologic activities. This study was undertaken to investigate the effect of rIL-1 beta, -2, -3, -4 and -6, IFN-gamma, TNF-alpha, and granulocyte-macrophage (GM)-CSF on basophils from 16 donors and the amount of histamine released was compared with that by partially purified mononuclear cell-derived histamine-releasing factor (HRF) and anti-IgE. We found that only IL-3 and GM-CSF at relatively high doses (50 to 500 ng/ml) released small amounts of histamine (3 to 14%) from two allergic donors. In contrast, both HRF and anti-IgE released significant amounts of histamine from all donors. Other cytokines did not release any measurable quantity of histamine. Simultaneous addition of several cytokines to the basophils also failed to release histamine. IL-3, GM-CSF, and IL-1 can also release histamine at lower concentrations (less than 5 ng/ml) when incubated with basophils in the presence of D{sub 2}O. Basophils from 6 out of 13 allergic donors released histamine in response to IL-3, whereas three donors responded to IL-1 beta and two responded to GM-CSF. The results of this study demonstrated that although IL-3 and GM-CSF release small amounts of histamine only from a select group of allergic patients, mononuclear cell-derived HRF is more potent in their action and release histamine from normals as well as allergic patients.

  13. Salvage chemotherapy followed by granulocyte colony-stimulating factor-primed donor leukocyte infusion with graft-vs.-host disease control for minimal residual disease in acute leukemia/myelodysplastic syndrome after allogeneic hematopoietic stem cell transplantation: prognostic factors and clinical outcomes.

    PubMed

    Mo, Xiao-Dong; Zhang, Xiao-Hui; Xu, Lan-Ping; Wang, Yu; Yan, Chen-Hua; Chen, Huan; Chen, Yu-Hong; Han, Wei; Wang, Feng-Rong; Wang, Jing-Zhi; Liu, Kai-Yan; Huang, Xiao-Jun

    2016-03-01

    This study investigated the prognostic factors and clinical outcomes of preemptive chemotherapy followed by granulocyte colony-stimulating factor-primed donor leukocyte infusion (Chemo-DLI) according to minimal residual disease (MRD) status in patients with acute leukemia and myelodysplastic syndromes who received allogeneic hematopoietic stem cell transplantation (HSCT) (n = 101). Patients received immunosuppressive drugs to prevent graft-vs.-host disease (GVHD) after Chemo-DLI. The 3-yr cumulative incidences of relapse, non-relapse mortality, and disease-free survival (DFS) after HSCT were 39.5%, 9.6%, and 51.7%, respectively. The cumulative incidences of relapse and DFS were significantly poorer in patients who exhibited early-onset MRD. Forty-four patients turned MRD negative 1 month after Chemo-DLI; their cumulative incidences of relapse and DFS were significantly better than those with persistent MRD 1 month after preemptive Chemo-DLI (relapse: 19.8% vs. 46.8%, P = 0.001; DFS: 69.6% vs. 46.4%, P = 0.004). The cumulative incidences of relapse and DFS after HSCT were significantly better in patients with chronic GVHD (cGVHD) than those without cGVHD (relapse: 19.6% vs. 63.7%, P < 0.001; DFS: 74.4% vs. 23.8%, P < 0.001). Early-onset MRD, persistent MRD after Chemo-DLI, and non-cGVHD after Chemo-DLI, which were associated with increased relapse and impaired DFS, suggest unsatisfactory response to preemptive Chemo-DLI.

  14. Interactions of dimethyl sulfoxide and granulocyte-macrophage colony-stimulating factor on the cell cycle kinetics and phosphoproteins of G1-enriched HL-60 cells: evidence of early effects on lamin B phosphorylation.

    PubMed

    Brennan, J K; Lee, K S; Frazel, M A; Keng, P C; Young, D A

    1991-03-01

    We have found that GM-CSF and DMSO have antagonistic effects on the proliferation but not maturation of asynchronously growing HL-60 cells such that growth in the presence of both more closely resembles normal hematopoiesis (Brennan et al., J. Cell Physiol. 132:246, 1987). Studies were undertaken to determine whether or not the agents affected the same mitogenic pathway and locus in the cell cycle. HL-60 populations containing at least 90% G1 cells were obtained by centrifugal elutriation, exposed to 100 u/ml recombinant human GM-CSF and/or 0-1.25% DMSO, and phosphoprotein changes quantified on autoradiograms of [32P]-orthophosphate-labeled cell proteins separated by giant 2-D gel electrophoresis. Results were correlated with 1) intracellular pH, determined by measurement of BCECF fluorescence; 2) [32P]-orthophosphate uptake; 3) cell cycle progression, determined by flow quantitation of DNA content in mithramycin or propidium iodide-stained cells; and 4) growth, determined by cell volume and concentration. GM-CSF stimulated and DMSO inhibited the GM-CSF-stimulated phosphorylation of 1 protein (approximately 65 kDa, p.i. 5.6) within 2 min of exposure. These effects were sustained through G1, not associated with changes in intracellular pH, and preceded similar antagonistic effects on phosphate uptake (15-30 minutes), cell volume change (16-24 hr), and cell concentration increase (28-32 hr). GM-CSF accelerated and DMSO inhibited G1 to S transit with the most marked antagonism observed in the second cycle following synchronization (28 to 40 hrs). Cell maturation (morphology, NBT reduction) was dominated by DMSO and not antagonized by GM-CSF. We have identified p65 as the nuclear intermediate filament protein, lamin B, on the basis of its locus on gels and its binding of a monoclonal antibody to intermediate filaments and antiserum to human lamin B on immunoblots. These studies suggest that at least part of the GM-CSF-DMSO antagonism is exerted through the same

  15. Colony-Stimulating Factors for Febrile Neutropenia during Cancer Therapy

    PubMed Central

    Bennett, Charles L.; Djulbegovic, Benjamin; Norris, LeAnn B.; Armitage, James O.

    2014-01-01

    A 55-year-old, previously healthy woman received a diagnosis of diffuse large-B-cell lymphoma after the evaluation of an enlarged left axillary lymph node obtained on biopsy. She had been asymptomatic except for the presence of enlarged axillary lymph nodes, which she had found while bathing. She was referred to an oncologist, who performed a staging evaluation. A complete blood count and test results for liver and renal function and serum lactate dehydrogenase were normal. Positron-emission tomography and computed tomography (PET–CT) identified enlarged lymph nodes with abnormal uptake in the left axilla, mediastinum, and retroperitoneum. Results on bone marrow biopsy were normal. The patient’s oncologist recommends treatment with six cycles of cyclophosphamide, doxorubicin, vincristine, and prednisone with rituximab (CHOP-R) at 21-day intervals. Is the administration of prophylactic granulocyte colony-stimulating factor (G-CSF) with the first cycle of chemotherapy indicated? PMID:23514290

  16. Yersinia enterocolitica Invasin Protein Triggers Differential Production of Interleukin-1, Interleukin-8, Monocyte Chemoattractant Protein 1, Granulocyte-Macrophage Colony-Stimulating Factor, and Tumor Necrosis Factor Alpha in Epithelial Cells: Implications for Understanding the Early Cytokine Network in Yersinia Infections

    PubMed Central

    Kampik, Daniel; Schulte, Ralf; Autenrieth, Ingo B.

    2000-01-01

    Yersinia enterocolitica infection of epithelial cells results in interleukin-8 (IL-8) mRNA expression. Herein we demonstrate that besides IL-8, increased mRNA levels of five other cytokines, IL-1α, IL-1β, monocyte chemoattractant protein 1 (MCP-1), granulocyte-macrophage colony-stimulating factor (GM-CSF), and tumor necrosis factor alpha (TNF-α), can be detected upon infection of HeLa cells with Yersinia. Yersinia-triggered cytokine production was not affected by blocking phosphatidylinositol-3-phosphate kinase with wortmannin, which inhibited bacterial invasion. Comparable cytokine mRNA responses were triggered by Escherichia coli expressing Yersinia inv, while no response was triggered by an inv-deficient Yersinia mutant. Moreover, cytokine responses were independent from metabolic activity of the bacteria, as killed bacterial cells were sufficient for triggering cytokine responses in HeLa cells. Semiquantitative reverse transcription-PCR analysis was used to assess the kinetics of cytokine mRNA expression in infected HeLa cells. IL-8, IL-1α, IL-1β, MCP-1, GM-CSF, and TNF-α mRNA expression increased within 1 h postinfection, reached a maximum after 3 to 4 h, and then declined to preinfection levels within 3 h. IL-8, MCP-1, and GM-CSF were secreted by HeLa cells, whereas IL-1α and IL-1β were not secreted and thus were found exclusively intracellularly. TNF-α protein could not be detected in cell lysates or supernatants. Stimulation of HeLa cells with IL-1α was followed by increased IL-8 mRNA expression, whereas stimulation with IL-8 did not induce cytokine production. Likewise, MCP-1 and GM-CSF did not induce significant cytokine responses in HeLa cells. Our results implicate that the initial host response to Yersinia infection might be sustained by IL-8, MCP-1, and GM-CSF produced by epithelial cells. PMID:10768935

  17. Large-Scale Hematopoietic Differentiation of Human Induced Pluripotent Stem Cells Provides Granulocytes or Macrophages for Cell Replacement Therapies

    PubMed Central

    Lachmann, Nico; Ackermann, Mania; Frenzel, Eileen; Liebhaber, Steffi; Brennig, Sebastian; Happle, Christine; Hoffmann, Dirk; Klimenkova, Olga; Lüttge, Doreen; Buchegger, Theresa; Kühnel, Mark Philipp; Schambach, Axel; Janciauskiene, Sabina; Figueiredo, Constanca; Hansen, Gesine; Skokowa, Julia; Moritz, Thomas

    2015-01-01

    Summary Interleukin-3 (IL-3) is capable of supporting the proliferation of a broad range of hematopoietic cell types, whereas granulocyte colony-stimulating factor (G-CSF) and macrophage CSF (M-CSF) represent critical cytokines in myeloid differentiation. When this was investigated in a pluripotent-stem-cell-based hematopoietic differentiation model, IL-3/G-CSF or IL-3/M-CSF exposure resulted in the continuous generation of myeloid cells from an intermediate myeloid-cell-forming complex containing CD34+ clonogenic progenitor cells for more than 2 months. Whereas IL-3/G-CSF directed differentiation toward CD45+CD11b+CD15+CD16+CD66b+ granulocytic cells of various differentiation stages up to a segmented morphology displaying the capacity of cytokine-directed migration, respiratory burst response, and neutrophil-extracellular-trap formation, exposure to IL-3/M-CSF resulted in CD45+CD11b+CD14+CD163+CD68+ monocyte/macrophage-type cells capable of phagocytosis and cytokine secretion. Hence, we show here that myeloid specification of human pluripotent stem cells by IL-3/G-CSF or IL-3/M-CSF allows for prolonged and large-scale production of myeloid cells, and thus is suited for cell-fate and disease-modeling studies as well as gene- and cell-therapy applications. PMID:25680479

  18. Antibody-dependent antitumor cytotoxicity by human monocytes cultured with recombinant macrophage colony-stimulating factor. Induction of efficient antibody-mediated antitumor cytotoxicity not detected by isotope release assays.

    PubMed

    Munn, D H; Cheung, N K

    1989-08-01

    Macrophage colony-stimulating factor (M-CSF) is known to stimulate proliferation of monocyte/macrophage progenitors and enhance in vitro antitumor cytotoxicity by murine macrophages. In this paper we have shown that recombinant human M-CSF causes human peripheral blood monocytes to differentiate in culture into metabolically active macrophage-like cells. These cells mediate very efficient antibody-dependent cellular cytotoxicity (ADCC) against human melanoma and neuroblastoma cell lines in the presence of two murine IgG3 mAbs (3F8 and R24). They also mediate antibody-independent cytotoxicity (or cytostasis) to a lesser extent. Human serum had an inconsistent effect on ADCC, but often induced similar high levels of ADCC. Cytotoxicity was measured using a novel ELISA to detect surviving tumor cells after ADCC. Two conventional isotope-release assays (51Cr and [3H]TdR) underestimated or entirely failed to detect ADCC by M-CSF-activated monocytes. Optimal activation occurred with 100-300 U/ml of M-CSF, and required 9-11 d for completion. Most of the M-CSF cultured monocytes expressed the low-affinity Fc receptor (CD16). ADCC by cells of the monocyte/macrophage lineage using murine IgG3 mAbs may have significance for the immunotherapy of human malignancies.

  19. Influence of receptor activator of nuclear factor (NF)-kappaB ligand (RANKL), macrophage-colony stimulating factor (M-CSF) and fetal calf serum on human osteoclast formation and activity.

    PubMed

    Kreja, Ludwika; Liedert, Astrid; Schmidt, Carla; Claes, Lutz; Ignatius, Anita

    2007-10-01

    Human osteoclast (OC) formation and activity was studied in cultures of peripheral blood mononuclear cells (PBMNC) from six healthy donors after stimulation with fetal calf serum (FCS), under the influence of the receptor activator of nuclear factor (NF)-kappaB ligand (RANKL) and the macrophage-colony stimulating factor (M-CSF). The results showed that selected FCS could stimulate OC formation without any medium supplementation with osteoclastogenic factors. The OC formation, investigated by quantification of multinucleated tartrate-resistant acid phosphatase-positive cells (TRAP+ cells), and the sensitivity of OC progenitors to RANKL and M-CSF, varied widely between individual donors. The OC resorption activity, measured in the "pit-assay" on dentine, was strictly dependent on the presence of RANKL and M-CSF in the medium and was also donor dependent. The considerable donor variability should be considered in culture studies investigating, e.g. the interactions of OC with biomaterials or the influence of cytokines, growth factors and drugs on osteoclastogenesis. PMID:18161075

  20. Monoclonal antibodies specific for human monocytes, granulocytes and endothelium.

    PubMed Central

    Hogg, N; MacDonald, S; Slusarenko, M; Beverley, P C

    1984-01-01

    Four monoclonal antibodies against antigens of human myeloid cells have been produced and thoroughly characterized in terms of their reactions with peripheral blood cells, cell lines, nine lymphoid and non-lymphoid tissues and the polypeptides with which they react. UCHM1 and SmO identify antigens present on the majority of blood monocytes and a variable, but lower, proportion of tissue macrophages. From their morphology and location in tissues, these cells appear to be recirculating monocytes. SMO antigen is also present on platelets. In addition, both antibodies stained endothelial cells, SMO in all tissues examined and UCHM1 variably. Biochemical investigation indicated that the UCHM1 antigen is a protein of 52,000 MW while the SMO antigen could not be indentified. The antibodies TG1 and 28 identify antigens mainly present on granulocytes. While mAb 28 reacted with neutrophils, TG1 also stained eosinophils and stained strongly a proportion of monocytes. TG1 also reacted variably with some non-haemopoietic cell lines. Both antibodies reacted predominantly with granulocytes in tissue sections. MAb TG1 precipitated a single polypeptide of 156,000 MW from monocytes and granulocytes, while mAb 28 precipitated non-convalently associated polypeptides of 83,000 and 155,000 MW from granulocytes but only a single molecule from monocytes, corresponding to the lower MW chain of 83,000. The epitope with which mAb 28 reacts appears not to be exposed on the surface of intact monocytes. This suggests that a similar or identical 83,000 MW molecule is made by both neutrophils and monocytes, but that its expression differs according to cell type. Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 PMID:6389324

  1. Accuracy of Diagnosis of Human Granulocytic Anaplasmosis in China.

    PubMed

    Wormser, Gary P

    2016-10-01

    In 2008, human granulocytic anaplasmosis (HGA) was reported from China. However, the clinical and laboratory findings, including reports of nosocomial transmission, were inconsistent with those reported for HGA in the United States. In 2012, it was demonstrated that the patients described in the 2008 report had all been infected with a newly discovered bunyavirus, severe fever with thrombocytopenia syndrome virus, which causes an illness with the same clinical features described for the patients in the 2008 report. This finding raises the question of HGA misdiagnosis in China and establishes the need for further studies to determine whether HGA occurs there. PMID:27648639

  2. Accuracy of Diagnosis of Human Granulocytic Anaplasmosis in China

    PubMed Central

    2016-01-01

    In 2008, human granulocytic anaplasmosis (HGA) was reported from China. However, the clinical and laboratory findings, including reports of nosocomial transmission, were inconsistent with those reported for HGA in the United States. In 2012, it was demonstrated that the patients described in the 2008 report had all been infected with a newly discovered bunyavirus, severe fever with thrombocytopenia syndrome virus, which causes an illness with the same clinical features described for the patients in the 2008 report. This finding raises the question of HGA misdiagnosis in China and establishes the need for further studies to determine whether HGA occurs there. PMID:27648639

  3. Colony-stimulating factors for the treatment of the hematopoietic component of the acute radiation syndrome (H-ARS): a review.

    PubMed

    Singh, Vijay K; Newman, Victoria L; Seed, Thomas M

    2015-01-01

    One of the greatest national security threats to the United States is the detonation of an improvised nuclear device or a radiological dispersal device in a heavily populated area. As such, this type of security threat is considered to be of relatively low risk, but one that would have an extraordinary high impact on health and well-being of the US citizenry. Psychological counseling and medical assessments would be necessary for all those significantly impacted by the nuclear/radiological event. Direct medical interventions would be necessary for all those individuals who had received substantial radiation exposures (e.g., >1 Gy). Although no drugs or products have yet been specifically approved by the United States Food and Drug Administration (US FDA) to treat the effects of acute radiation syndrome (ARS), granulocyte colony-stimulating factor (G-CSF), granulocyte macrophage colony-stimulating factor (GM-CSF), and pegylated G-CSF have been used off label for treating radiation accident victims. Recent threats of terrorist attacks using nuclear or radiologic devices makes it imperative that the medical community have up-to-date information and a clear understanding of treatment protocols using therapeutically effective recombinant growth factors and cytokines such as G-CSF and GM-CSF for patients exposed to injurious doses of ionizing radiation. Based on limited human studies with underlying biology, we see that the recombinants, G-CSF and GM-CSF appear to have modest, but significant medicinal value in treating radiation accident victims. In the near future, the US FDA may approve G-CSF and GM-CSF as ‘Emergency Use Authorization’ (EUA) for managing radiation-induced aplasia, an ARS-related pathology. In this article, we review the status of growth factors for the treatment of radiological/nuclear accident victims. PMID:25215458

  4. Induction of megakaryocytic colony-stimulating activity in mouse skin by inflammatory agents and tumor promoters

    SciTech Connect

    Clark, D.A.; Dessypris, E.N.; Koury, M.J.

    1987-03-01

    The production of megakaryocytic colony-stimulating activity (MEG-CSA) was assayed in acetic acid extracts of skin from mice topically treated with inflammatory and tumor-promoting agents. A rapid induction of MEG-CSA was found in skin treated both with phorbol 12-myristate 13-acetate (PMA), a strong tumor promoter, and with mezerein, a weak tumor promoter, but no induction was found in untreated skin. The time course of induction of MEG-CSA following treatment of skin with PMA or mezerein was very similar to that previously demonstrated for the induction of granulocyte-macrophage colony-stimulating activity in mouse skin by these agents. The induced MEG-CSA was found in both the epidermis and the dermis. Pretreatment of the skin with US -methasone abrogated the MEG-CSA induction. The cell number response curve suggests that the MEG-CSA acts directly on the progenitor cells of the megakaryocyte colonies. That topical administration of diterpene esters results in the rapid, local induction of MEG-CSA which can be blocked by US -methasone pretreatment suggests a mechanism for the thrombocytosis associated with some inflammatory states. The indirect action in which diterpene esters induce in certain cells the production or release of growth regulatory factors for other cell types may also aid in understanding their carcinogenic properties.

  5. Modulation of Decidual Macrophage Polarization by Macrophage Colony-Stimulating Factor Derived from First-Trimester Decidual Cells: Implication in Preeclampsia.

    PubMed

    Li, Min; Piao, Longzhu; Chen, Chie-Pein; Wu, Xianqing; Yeh, Chang-Ching; Masch, Rachel; Chang, Chi-Chang; Huang, S Joseph

    2016-05-01

    During human pregnancy, immune tolerance of the fetal semiallograft occurs in the presence of abundant maternal leukocytes. At the implantation site, macrophages comprise approximately 20% of the leukocyte population and act as primary mediators of tissue remodeling. Decidual macrophages display a balance between anti-inflammatory and proinflammatory phenotypes. However, a shift to an M1 subtype is reported in preeclampsia. Granulocyte-macrophage colony-stimulating-factor (GM-CSF) and macrophage colony-stimulating factor (M-CSF) are major differentiating factors that mediate M1 and M2 polarization, respectively. Previously, we observed the following: i) the preeclamptic decidua contains an excess of both macrophages and GM-CSF, ii) the preeclampsia-associated proinflammatory cytokines, IL-1β and tumor necrosis factor-α, markedly enhance GM-CSF and M-CSF expression in cultured leukocyte-free first-trimester decidual cells (FTDCs), iii) FTDC-secreted GM-CSF polarizes macrophages toward an M1 subtype. The microenvironment is a key determinant of macrophage phenotype. Thus, we examined proinflammatory stimulation of FTDC-secreted M-CSF and its role in macrophage development. Immunofluorescence staining demonstrated elevated M-CSF-positive decidual cell numbers in preeclamptic decidua. In FTDCs, IL-1β and tumor necrosis factor-α signal through the NF-κB pathway to induce M-CSF production, which does the following: i) enhances differentiation of and elevates CD163 expression in macrophages, ii) increases macrophage phagocytic capacity, and iii) inhibits signal-regulatory protein α expression by macrophages. These findings suggest that FTDC-secreted M-CSF modulates the decidual immune balance by inducing M2 macrophage polarization and phagocytic capacity in response to proinflammatory stimuli.

  6. Colony-stimulating factor 1 potentiates lung cancer bone metastasis.

    PubMed

    Hung, Jaclyn Y; Horn, Diane; Woodruff, Kathleen; Prihoda, Thomas; LeSaux, Claude; Peters, Jay; Tio, Fermin; Abboud-Werner, Sherry L

    2014-04-01

    Colony-stimulating factor 1 (CSF1) is essential for osteoclastogenesis that mediates osteolysis in metastatic tumors. Patients with lung cancer have increased CSF1 in serum and high levels are associated with poor survival. Adenocarcinomas metastasize rapidly and many patients suffer from bone metastasis. Lung cancer stem-like cells sustain tumor growth and potentiate metastasis. The purpose of this study was to determine the role of CSF1 in lung cancer bone metastasis and whether inhibition of CSF1 ameliorates the disease. Human lung adenocarcinoma A549 cells were examined in vitro for CSF1/CSF1R. A549-luc cells were injected intracardiac in NOD/SCID mice and metastasis was assessed. To determine the effect of CSF1 knockdown (KD) in A549 cells on bone metastasis, cells were stably transfected with a retroviral vector containing short-hairpin CSF1 (KD) or empty vector (CT). Results showed that A549 cells express CSF1/CSF1R; CSF1 increased their proliferation and invasion, whereas soluble CSF1R inhibited invasion. Mice injected with A549-luc cells showed osteolytic bone lesions 3.5 weeks after injection and lesions increased over 5 weeks. Tumors recapitulated adenocarcinoma morphology and showed osteoclasts along the tumor/bone interface, trabecular, and cortical bone loss. Analyses of KD cells showed decreased CSF1 protein levels, reduced colony formation in soft agar assay, and decreased fraction of stem-like cells. In CSF1KD mice, the incidence of tumor metastasis was similar to controls, although fewer CSF1KD mice had metastasis in both hind limbs. KD tumors showed reduced CSF1 expression, Ki-67+ cells, and osteoclasts. Importantly, there was a low incidence of large tumors >0.1 mm(2) in CSF1KD mice compared with control mice (10% vs 62.5%). This study established a lung osteolytic bone metastasis model that resembles human disease and suggests that CSF1 is a key determinant of cancer stem cell survival and tumor growth. Results may lead to novel strategies to

  7. Flow cytometry evidence of human granulocytes interaction with polyhedral oligomeric silsesquioxanes: effect of nanoparticle charge

    NASA Astrophysics Data System (ADS)

    Renò, Filippo; Carniato, Fabio; Rizzi, Manuela; Olivero, Francesco; Pittarella, Pamela; Marchese, Leonardo

    2013-05-01

    Nanoparticles (NPs) entering the human body are immediately confronted with the innate part of human immune system. In particular, monocyte and neutrophil granulocytes readily clear particles by phagocytosis, even if in the case of NPs the uptake mechanism may be classified as macropinocytosis. Among engineered nanoparticles, in the last years, siliceous materials have emerged as promising materials for several applications ranging from catalysis to biomedical. The polyhedral oligomeric silsesquioxanes (POSS) are nanodimensional, easily synthesizable molecular compounds and POSS-based systems are promising carriers for biological molecules. In this work, the ability of human granulocytes to uptake positively and negatively charged POSS was measured using a simple flow cytometry analysis based on cell size modifications. The data obtained showed that after a 30 min exposure only positive NPs were uptaken by human granulocyte using both macropinocytosis and clathrin-mediated mechanisms as demonstrated by uptake inhibition mediated by amiloride and chlorpromazine.

  8. H2O2 Release from Human Granulocytes during Phagocytosis

    PubMed Central

    Root, Richard K.; Metcalf, Julia A.

    1977-01-01

    Normal and cytochalasin B-treated human granulocytes have been studied to determine some of the interrelationships between phagocytosis-induced respiration and superoxide and hydrogen peroxide formation and release into the extracellular medium by intact cells. By using the scopoletin fluorescent assay to continuously monitor extracellular hydrogen peroxide concentrations during contact of cells with opsonized staphylococci, it was demonstrated that the superoxide scavengers ferricytochrome c and nitroblue tetrazolium significantly reduced the amount of H2O2 released with time from normal cells but did not abolish it. This inhibitory effect was reversed by the simultaneous addition of superoxide dismutase (SOD), whereas the addition of SOD alone increased the amount of detectable H2O2 in the medium. The addition of sodium azide markedly inhibited myeloperoxidase-H2O2-dependent protein iodination and more than doubled H2O2 release, including the residual amount remaining after exposure of the cells to ferricytochrome c, suggesting its origin from an intracellular pool shared by several pathways for H2O2 catabolism. When cells were pretreated with cytochalasin B and opsonized bacteria added, reduced oxygen consumption was observed, but this was in parallel to a reduction in specific binding of organisms to the cells when compared to normal. Under the influence of inhibited phagosome formation by cytochalasin B, the cells released an increased amount of superoxide and peroxide into the extracellular medium relative to oxygen consumption, and all detectable peroxide release could be inhibited by the addition of ferricytochrome c. Decreased H2O2 production in the presence of this compound could not be ascribed to diminished bacterial binding, decreased oxidase activity, or increased H2O2 catabolism and was reversed by the simultaneous addition of SOD. Furthermore, SOD and ferricytochrome c had similar effects on another H2O2-dependent reaction, protein iodination, in

  9. Group-specific human granulocyte antigens on a chronic myelogenous leukemia cell line with a Philadelphia chromosome marker.

    PubMed

    Drew, S I; Terasaki, P I; Billing, R J; Bergh, O J; Minowada, J; Klein, E

    1977-05-01

    Group-specific human granulocyte antigens are serologically detectable with granulocytotoxic-positive human alloantisera on a cell line, K562, of chronic myelogenous leukemia origin which bears a Philadelphia chromosomal marker. The same cell line lacks serologically detectable HLA, B2 microglobulin, and B-lymphocyte antigens. Granulocyte antigens are important cell markers for cell lines of suspected myeloid lineage.

  10. Membrane permeability of the human granulocyte to water, dimethyl sulfoxide, glycerol, propylene glycol and ethylene glycol

    PubMed Central

    Vian, Alex M.; Higgins, Adam Z.

    2015-01-01

    Granulocytes are currently transfused as soon as possible after collection because they rapidly deteriorate after being removed from the body. This short shelf life complicates the logistics of granulocyte collection, banking and safety testing. Cryopreservation has the potential to significantly increase shelf life; however, cryopreservation of granulocytes has proven to be difficult. In this study, we investigate the membrane permeability properties of human granulocytes, with the ultimate goal of using membrane transport modeling to facilitate development of improved cryopreservation methods. We first measured the equilibrium volume of human granulocytes in a range of hypo- and hypertonic solutions and fit the resulting data using a Boyle-van't Hoff model. This yielded an isotonic cell volume of 378 μm3 and an osmotically inactive volume of 165 μm3. To determine the permeability of the granulocyte membrane to water and cryoprotectant (CPA), cells were injected into well-mixed CPA solution while collecting volume measurements using a Coulter Counter. These experiments were performed at temperatures ranging from 4 to 37 °C for exposure to dimethyl sulfoxide, glycerol, ethylene glycol and propylene glycol. The best-fit water permeability was similar in the presence of all of the CPAs, with an average value at 21 °C of 0.18 μm atm−1 min−1. The activation energy for water transport ranged from 41 to 61 kJ/mol. The CPA permeability at 21 °C was 6.4, 1.0, 8.4 and 4.0 μm/min for dimethyl sulfoxide, glycerol, ethylene glycol and propylene glycol, respectively, and the activation energy for CPA transport ranged between 59 and 68 kJ/mol. PMID:24269528

  11. Membrane permeability of the human granulocyte to water, dimethyl sulfoxide, glycerol, propylene glycol and ethylene glycol.

    PubMed

    Vian, Alex M; Higgins, Adam Z

    2014-02-01

    Granulocytes are currently transfused as soon as possible after collection because they rapidly deteriorate after being removed from the body. This short shelf life complicates the logistics of granulocyte collection, banking, and safety testing. Cryopreservation has the potential to significantly increase shelf life; however, cryopreservation of granulocytes has proven to be difficult. In this study, we investigate the membrane permeability properties of human granulocytes, with the ultimate goal of using membrane transport modeling to facilitate development of improved cryopreservation methods. We first measured the equilibrium volume of human granulocytes in a range of hypo- and hypertonic solutions and fit the resulting data using a Boyle-van't Hoff model. This yielded an isotonic cell volume of 378 μm(3) and an osmotically inactive volume of 165 μm(3). To determine the permeability of the granulocyte membrane to water and cryoprotectant (CPA), cells were injected into well-mixed CPA solution while collecting volume measurements using a Coulter Counter. These experiments were performed at temperatures ranging from 4 to 37°C for exposure to dimethyl sulfoxide, glycerol, ethylene glycol, and propylene glycol. The best-fit water permeability was similar in the presence of all of the CPAs, with an average value at 21°C of 0.18 μmatm(-1)min(-1). The activation energy for water transport ranged from 41 to 61 kJ/mol. The CPA permeability at 21°C was 6.4, 1.0, 8.4, and 4.0 μm/min for dimethyl sulfoxide, glycerol, ethylene glycol, and propylene glycol, respectively, and the activation energy for CPA transport ranged between 59 and 68 kJ/mol. PMID:24269528

  12. Generation of slow-reacting substance (leukotrienes) by endotoxin and lipid A from human polymorphonuclear granulocytes.

    PubMed Central

    Bremm, K D; König, W; Spur, B; Crea, A; Galanos, C

    1984-01-01

    Leukotrienes were released from human polymorphonuclear granulocytes on incubation with endotoxins and lipid A. The analysis was performed by their smooth muscle contracting properties, reversed phase high-pressure liquid chromatography and radioimmunoassay for leukotrienes C4 and D4. The active component of the lipopolysaccharides seems to be the lipid A portion. PMID:6490085

  13. Current management of human granulocytic anaplasmosis, human monocytic ehrlichiosis and Ehrlichia ewingii ehrlichiosis

    PubMed Central

    Thomas, Rachael J; Stephen Dumler, J; Carlyon, Jason A

    2009-01-01

    Anaplasma phagocytophilum, Ehrlichia chaffeensis and Ehrlichia ewingii are emerging tick-borne pathogens and are the causative agents of human granulocytic anaplasmosis, human monocytic ehrlichiosis and E. ewingii ehrlichiosis, respectively. Collectively, these are referred to as human ehrlichioses. These obligate intracellular bacterial pathogens of the family Anaplasmataceae are transmitted by Ixodes spp. or Amblyomma americanum ticks and infect peripherally circulating leukocytes to cause infections that range in clinical spectra from asymptomatic seroconversion to mild, severe or, in rare instances, fatal disease. This review describes: the ecology of each pathogen; the epidemiology, clinical signs and symptoms of the human diseases that each causes; the choice methods for diagnosing and treating human ehrlichioses; recommendations for patient management; and is concluded with suggestions for potential future research. PMID:19681699

  14. Colony-stimulating factors for the management of neutropenia in cancer patients.

    PubMed

    Dale, David C

    2002-01-01

    Neutropenia and its subsequent infectious complications represent the most common dose-limiting toxicity of cancer chemotherapy. Febrile neutropenia (FN) occurs with common chemotherapy regimens in 25 to 40% of treatment-naive patients, and its severity depends on the dose intensity of the chemotherapy regimen, the patient's prior history of either radiation therapy or use of cytotoxic treatment, and comorbidities. The occurrence of FN often causes subsequent chemotherapy delays or dose reductions. It may also lengthen hospital stay, increase monitoring, diagnostic and treatment costs, and reduce patient quality of life. A decade after their introduction, colony-stimulating factors (CSFs) such as granulocyte colony-stimulating factor (G-CSF) and granulocyte-macrophage colony-stimulating factor (GM-CSF) are now an integral part of the prevention of potentially life-threatening FN; however, only G-CSF has US Food and Drug Administration approval for use in chemotherapy-induced neutropenia. These adjunctive agents accelerate formation of neutrophils from committed progenitors, thereby reducing the duration and severity of neutropenia. Important uses of CSFs in oncology are prevention of FN after chemotherapy, treatment of febrile neutropenic episodes and support following bone marrow transplantation, and collection of CSF-mobilised peripheral blood progenitor cells. G-CSF is used more frequently than GM-CSF for all of these indications because of fewer associated adverse effects. Clinical trials to date have not demonstrated a significant effect on overall survival or disease-free survival, which is most likely to be due to small sample size and lack of power to prove effect. However, they have demonstrated clinical utility in allowing the delivery of planned chemotherapy dose on schedule, an important clinical goal especially in curative tumour settings. The high cost of these agents limits their widespread use. Current American Society of Clinical Oncology

  15. The hematopoietic cytokine granulocyte-macrophage colony stimulating factor is important for cognitive functions

    PubMed Central

    Krieger, Markus; Both, Martin; Kranig, Simon A.; Pitzer, Claudia; Klugmann, Matthias; Vogt, Gerhard; Draguhn, Andreas; Schneider, Armin

    2012-01-01

    We recently reported expression of hematopoietic growth factor GM-CSF and its receptor (GM-CSFR) in CNS neurons. Here we evaluated this system in learning and memory formation using GM-CSF deficient mice. In complementation, GM-CSF signalling was manipulated specifically in adult murine hippocampus by adeno-associated virus (AAV)-mediated GM-CSFR alpha overexpression or knock-down. GM-CSF ablation caused various hippocampus and amygdala-dependent deficits in spatial and fear memory while rendering intact basic parameters like motor function, inherent anxiety, and pain threshold levels. Corroborating these data, spatial memory of AAV-injected mice was positively correlated with GM-CSFRα expression levels. Hippocampal neurons of knock-out mice showed markedly pruned dendritic trees, reduced spine densities, and lower percentages of mature spines. Despite such morphological alterations, long-term potentiation (LTP) was unimpaired in the knock-out hippocampus. Collectively, these results suggest that GM-CSF signalling plays a major role in structural plasticity relevant to learning and memory. PMID:23019518

  16. Interleukin 18 inhibits osteoclast formation via T cell production of granulocyte macrophage colony-stimulating factor.

    PubMed Central

    Horwood, N J; Udagawa, N; Elliott, J; Grail, D; Okamura, H; Kurimoto, M; Dunn, A R; Martin, T; Gillespie, M T

    1998-01-01

    IL-18 inhibits osteoclast (OCL) formation in vitro independent of IFN-gamma production, and this was abolished by the addition of neutralizing antibodies to GM-CSF. We now establish that IL-18 was unable to inhibit OCL formation in cocultures using GM-CSF-deficient mice (GM-CSF -/-). Reciprocal cocultures using either wild-type osteoblasts with GM-CSF -/- spleen cells or GM-CSF -/- osteoblasts with wild-type spleen cells were examined. Wild-type spleen cells were required to elicit a response to IL-18 indicating that cells of splenic origin were the IL-18 target. As T cells comprise a large proportion of the spleen cell population, the role of T cells in osteoclastogenesis was examined. Total T cells were removed and repleted in various combinations. Addition of wild-type T cells to a GM-CSF -/- coculture restored IL-18 inhibition of osteoclastogenesis. Major subsets of T cells, CD4+ and CD8+, were also individually depleted. Addition of either CD4+ or CD8+ wild-type T cells restored IL-18 action in a GM-CSF -/- background, while IL-18 was ineffective when either CD4+ or CD8+ GM-CSF -/- T cells were added to a wild-type coculture. These results highlight the involvement of T cells in IL-18-induced OCL inhibition and provide evidence for a new OCL inhibitory pathway whereby IL-18 inhibits OCL formation due to action upon T cells promoting the release of GM-CSF, which in turn acts upon OCL precursors. PMID:9449693

  17. Local transgenic expression of granulocyte macrophage-colony stimulating factor initiates autoimmunity.

    PubMed

    Biondo, M; Nasa, Z; Marshall, A; Toh, B H; Alderuccio, F

    2001-02-01

    Mechanisms leading to breakdown of immunological tolerance and initiation of autoimmunity are poorly understood. Experimental autoimmune gastritis is a paradigm of organ-specific autoimmunity arising from a pathogenic autoimmune response to gastric H/K ATPase. The gastritis is accompanied by autoantibodies to the gastric H/K ATPase. The best characterized model of experimental autoimmune gastritis requires neonatal thymectomy. This procedure disrupts the immune repertoire, limiting its usefulness in understanding how autoimmunity arises in animals with intact immune systems. Here we tested whether local production of GM-CSF, a pro-inflammatory cytokine, is sufficient to break tolerance and initiate autoimmunity. We generated transgenic mice expressing GM-CSF in the stomach. These transgenic mice spontaneously developed gastritis with an incidence of about 80% after six backcrosses to gastritis-susceptible BALBc/CrSlc mice. The gastritis is accompanied by mucosal hypertrophy, enlargement of draining lymph nodes and autoantibodies to gastric H/K ATPase. An infiltrate of dendritic cells and macrophages preceded CD4 T cells into the gastric mucosa. T cells from draining lymph nodes specifically proliferated to the gastric H/K ATPase. CD4 but not CD8 T cells transferred gastritis to nude mouse recipients. CD4(+) CD25(+) T cells from the spleen retained anergic suppressive properties that were reversed by IL-2. We conclude that local expression of GM-CSF is sufficient to break tolerance and initiate autoimmunity mediated by CD4 T cells. This new mouse model should be useful for studies of organ-specific autoimmunity.

  18. Reduced salmonella fecal shedding in swine administered porcine granulocyte-colony stimulating factor (G-CSF)

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Salmonella colonization of food animals is a concern for animal health, food safety and public health. Key objectives of pre-harvest food safety programs are to detect asymptomatic Salmonella carriage in food animals, reduce colonization, and prevent transmission of Salmonella to other animals and ...

  19. The Gottingen Minipig Is a Model of the Hematopoietic Acute Radiation Syndrome: G-Colony Stimulating Factor Stimulates Hematopoiesis and Enhances Survival From Lethal Total-Body γ-Irradiation

    SciTech Connect

    Moroni, Maria; Ngudiankama, Barbara F.; Christensen, Christine; Olsen, Cara H.; Owens, Rossitsa; Lombardini, Eric D.; Holt, Rebecca K.; Whitnall, Mark H.

    2013-08-01

    Purpose: We are characterizing the Gottingen minipig as an additional large animal model for advanced drug testing for the acute radiation syndrome (ARS) to enhance the discovery and development of novel radiation countermeasures. Among the advantages provided by this model, the similarities to human hematologic parameters and dynamics of cell loss/recovery after irradiation provide a convenient means to compare the efficacy of drugs known to affect bone marrow cellularity and hematopoiesis. Methods and Materials: Male Gottingen minipigs, 4 to 5 months old and weighing 9 to 11 kg, were used for this study. We tested the standard off-label treatment for ARS, rhG-CSF (Neupogen, 10 μg/kg/day for 17 days), at the estimated LD70/30 total-body γ-irradiation (TBI) radiation dose for the hematopoietic syndrome, starting 24 hours after irradiation. Results: The results indicated that granulocyte colony stimulating factor (G-CSF) enhanced survival, stimulated recovery from neutropenia, and induced mobilization of hematopoietic progenitor cells. In addition, the administration of G-CSF resulted in maturation of monocytes/macrophages. Conclusions: These results support continuing efforts toward validation of the minipig as a large animal model for advanced testing of radiation countermeasures and characterization of the pathophysiology of ARS, and they suggest that the efficacy of G-CSF in improving survival after total body irradiation may involve mechanisms other than increasing the numbers of circulating granulocytes.

  20. Human granulocyte/pollen-binding protein. Recognition and identification as transferrin.

    PubMed Central

    Sass-Kuhn, S P; Moqbel, R; Mackay, J A; Cromwell, O; Kay, A B

    1984-01-01

    Normal human serum was found to contain a heat-stable protein which promoted the binding of granulocytes to timothy grass pollen (granulocyte/pollen-binding protein [GPBP]). GPBP was purified by gel filtration, anion exchange, and affinity chromatography. Virtually all of the granulocyte/pollen-binding activity was associated with a beta-1-protein having a molecular mass of approximately 77,000 D and an isoelectric point of between 5.5 and 6.1. By immunoelectrophoresis and sodium dodecyl sulfate-polyacrylamide gel electrophoresis, the protein was identified as transferrin. Monospecific antisera raised against either GPBP or transferrin removed biological activity from GPBP preparations, and GPBP and transferrin gave lines of identity with these two antisera. The apparent heterogeneity in the molecular size and charge of GPBP observed during progressive purification was minimal when GPBP was saturated with ferric ions before the separation procedures. These experiments indicate that granulocyte/pollen binding is a hitherto unrecognized property of transferrin which appears to be unrelated to iron transport and raises the possibility that transferrin might have a physiological role in the removal of certain organic matter. Images PMID:6690479

  1. [EPIDEMIOLOGIC ASPECTS OF HUMAN GRANULOCYTIC ANAPLASMOSIS IN THE WESTERN REGION OF UKRAINE].

    PubMed

    Ben, I I; Biletska, H V

    2015-01-01

    This article presents data of complex study of human granulocytic anaplasmosis the human granulocytic anaplasmosis (HGA) in the western region of Ukraine. Natural HGA foci were identified, where the prevalence of A. phagocytophilum in the main vector (I. ricinus) amount to (12.0 +/- 0.7) %, and seroprevalence of HGA among the healthy population--(28.6 +/- 1.6) %. It's shown that A. phagocytophilum is the etiologic agent of (33.7 +/- 4.9) % of undiagnosed cases of seasonal febrile diseases. Principal characteristics of HGA epidemiology: spring-summer scasonality, the prevalence of disease in the age structure of people of active age (45.30 +/- 1.95 years), a significant level combination with (60.4 +/- 2.2) % Lyme-horreliosis and other tick-borne infections (mixed infections), the different level of activity of HGA epidemic process in forest and steppe geographical landscape zones--were revealed. PMID:27491171

  2. 1,25-dihydroxycholecalciferol-induced differentiation of myelomonocytic leukemic cells unresponsive to colony stimulating factors and phorbol esters

    SciTech Connect

    Bettens, F.; Schlick, E.; Farrar, W.; Ruscetti, F.

    1986-12-01

    The murine myelomonocytic leukemia cell line WEHI-3B D/sup +/, which differentiates in response to granulocyte colony stimulating factor (G-CSF), can also be induced to differentiate into monocyte-macrophages by phorbol myristate acetate (PMA) treatment, whereas the WEHI-3B D/sup -/ subline, which is unresponsive to G-CSF and PMA, can be induced to differentiate to granulocytes as well as monocytes by 1,25-dihydroxycholecalciferol (1,25-(OH)/sub 2/ D3), the biologically active metabolite of vitamin D3. A newly developed variant of the WEHI-3B D/sup +/ line, named WEHI-3B D/sup +/G, which was responsive to G-CSF but not to PMA, was also differentiated to granulocytes by 1,25-(OH)/sub 2/ D3. Although vitamin D3 has been reported to induce macrophage differentiation in responsive tumor cells, this is the first demonstration that 1,25-(OH)/sub 2/ D3 can induce granulocyte differentiation. In both differentiation pathways, cessation of cellular proliferation accompanies changes in morphologic and cytochemical properties of the cells. This suggests that leukemic cell lines unresponsive to differentiation agents acting at the cell surface retain their ability to differentiate in response to agents that do not act via the plasma membrane such as 1,25-(OH)/sub 2/ D3, which has cytosolic/nuclear receptors. These results suggest that low doses of 1,25-(OH)/sub 2/ D3 may be useful in combination with hemopoietic growth factors (CSFs) as therapeutic agent to induce leukemic cell differentiation in vivo.

  3. Role of macrophages in serum colony-stimulating factor induction by Lactobacillus casei in mice.

    PubMed Central

    Nanno, M; Shimizu, T; Mike, A; Ohwaki, M; Mutai, M

    1988-01-01

    Heat-killed Lactobacillus casei YIT9018 (LC9018), when injected intravenously into mice at a dose of 4 to 40 mg/kg, induced the production of serum colony-stimulating factor (CSF). Since this induction was observed in both C3H/HeJ and C3H/HeN mice, LC9018 was considered to act differently from lipopolysaccharide. The amount of serum CSF induced by LC9018 in nude mice and whole-body-X-ray-irradiated mice was similar to that in control mice, but the induction of serum CSF was suppressed by the previous administration of carrageenan, indicating that macrophages, but not T cells, were responsible for serum CSF induction by LC9018. To determine whether macrophages themselves produce CSF or help other cells produce CSF in response to LC9018, we prepared adherent cells from the peritoneal cavity of normal mice and examined CSF activity in their conditioned media. Peritoneal adherent cells did not produce CSF without LC9018, but when cultivated with 1 mg of LC9018 per ml, they produced CSF at the same time that serum CSF was induced after the intravenous administration of LC9018. Additionally, in vitro-induced CSF formed macrophage, granulocyte, and mixed colonies, as serum CSF did. CSF production by peritoneal adherent cells was completely inhibited by cycloheximide (50 micrograms/ml), and neither the elimination of T cells from the peritoneal adherent cells by treating them with anti-Thy-1.2 antibody plus complement nor the addition of T cells affected CSF production. These results suggest that heat-killed LC9018 induces serum CSF in mice via direct stimulation of macrophages to produce CSF de novo. PMID:3123388

  4. Biological role of granulocyte macrophage colony-stimulating factor (GM-CSF) and macrophage colony-stimulating factor (M-CSF) on cells of the myeloid lineage.

    PubMed

    Ushach, Irina; Zlotnik, Albert

    2016-09-01

    M-CSF and GM-CSF are 2 important cytokines that regulate macrophage numbers and function. Here, we review their known effects on cells of the macrophage-monocyte lineage. Important clues to their function come from their expression patterns. M-CSF exhibits a mostly homeostatic expression pattern, whereas GM-CSF is a product of cells activated during inflammatory or pathologic conditions. Accordingly, M-CSF regulates the numbers of various tissue macrophage and monocyte populations without altering their "activation" status. Conversely, GM-CSF induces activation of monocytes/macrophages and also mediates differentiation to other states that participate in immune responses [i.e., dendritic cells (DCs)]. Further insights into their function have come from analyses of mice deficient in either cytokine. M-CSF signals through its receptor (CSF-1R). Interestingly, mice deficient in CSF-1R expression exhibit a more significant phenotype than mice deficient in M-CSF. This observation was explained by the discovery of a novel cytokine (IL-34) that represents a second ligand of CSF-1R. Information about the function of these ligands/receptor system is still developing, but its complexity is intriguing and strongly suggests that more interesting biology remains to be elucidated. Based on our current knowledge, several therapeutic molecules targeting either the M-CSF or the GM-CSF pathways have been developed and are currently being tested in clinical trials targeting either autoimmune diseases or cancer. It is intriguing to consider how evolution has directed these pathways to develop; their complexity likely mirrors the multiple functions in which cells of the monocyte/macrophage system are involved. PMID:27354413

  5. Macrophage-Colony Stimulating Factor Derived from Injured Primary Afferent Induces Proliferation of Spinal Microglia and Neuropathic Pain in Rats.

    PubMed

    Okubo, Masamichi; Yamanaka, Hiroki; Kobayashi, Kimiko; Dai, Yi; Kanda, Hirosato; Yagi, Hideshi; Noguchi, Koichi

    2016-01-01

    Peripheral nerve injury induces proliferation of microglia in the spinal cord, which can contribute to neuropathic pain conditions. However, candidate molecules for proliferation of spinal microglia after injury in rats remain unclear. We focused on the colony-stimulating factors (CSFs) and interleukin-34 (IL-34) that are involved in the proliferation of the mononuclear phagocyte lineage. We examined the expression of mRNAs for macrophage-CSF (M-CSF), granulocyte macrophage-CSF (GM-CSF), granulocyte-CSF (G-CSF) and IL-34 in the dorsal root ganglion (DRG) and spinal cord after spared nerve injury (SNI) in rats. RT-PCR and in situ hybridization revealed that M-CSF and IL-34, but not GM- or G-CSF, mRNAs were constitutively expressed in the DRG, and M-CSF robustly increased in injured-DRG neurons. M-CSF receptor mRNA was expressed in naive rats and increased in spinal microglia following SNI. Intrathecal injection of M-CSF receptor inhibitor partially but significantly reversed the proliferation of spinal microglia and in early phase of neuropathic pain induced by SNI. Furthermore, intrathecal injection of recombinant M-CSF induced microglial proliferation and mechanical allodynia. Here, we demonstrate that M-CSF is a candidate molecule derived from primary afferents that induces proliferation of microglia in the spinal cord and leads to induction of neuropathic pain after peripheral nerve injury in rats.

  6. Characterization of macrophages from the bony fish gilthead seabream using an antibody against the macrophage colony-stimulating factor receptor.

    PubMed

    Mulero, Iván; Pilar Sepulcre, M; Roca, Francisco J; Meseguer, José; García-Ayala, Alfonsa; Mulero, Victoriano

    2008-01-01

    Two major professional phagocyte populations have been described in fish, namely granulocytes and monocytes/macrophages. Although the distribution and localization of macrophages have been documented in several teleost species using mainly light and/or electron microscopy, the lack of appropriate markers for these cells has hampered our in-depth knowledge of their biology. We report here the generation of a monospecific rabbit polyclonal antibody against the gilthead seabream macrophage colony-stimulating factor receptor (Mcsfr), which is an excellent marker of macrophages in mammals and the zebrafish. The anti-Mcsfr has been found to be very useful in immunohistochemistry (IHC) to specifically immunostain the purified macrophages (adherent cells) obtained from the head-kidney as well as different cell populations in paraffin-embedded organs, including the head-kidney, spleen, thymus, gills and liver. Unexpectedly, however, no Mcsfr immunoreactive (Mcsfr(+)) cells were observed in the brain and intestine of the gilthead seabream. We also show that the distribution of Mcsfr(+) cells in the head-kidney and the spleen is unaltered following infection with the fish pathogenic bacterium Vibrio anguillarum and that the Il1b-producing cells in these two organs after infection are exclusively acidophilic granulocytes. Finally, as the epitope recognized by the anti-Mcsfr is well conserved, we illustrate the potential usefulness of this antibody in other teleost species, such as the European seabass.

  7. Macrophage-Colony Stimulating Factor Derived from Injured Primary Afferent Induces Proliferation of Spinal Microglia and Neuropathic Pain in Rats.

    PubMed

    Okubo, Masamichi; Yamanaka, Hiroki; Kobayashi, Kimiko; Dai, Yi; Kanda, Hirosato; Yagi, Hideshi; Noguchi, Koichi

    2016-01-01

    Peripheral nerve injury induces proliferation of microglia in the spinal cord, which can contribute to neuropathic pain conditions. However, candidate molecules for proliferation of spinal microglia after injury in rats remain unclear. We focused on the colony-stimulating factors (CSFs) and interleukin-34 (IL-34) that are involved in the proliferation of the mononuclear phagocyte lineage. We examined the expression of mRNAs for macrophage-CSF (M-CSF), granulocyte macrophage-CSF (GM-CSF), granulocyte-CSF (G-CSF) and IL-34 in the dorsal root ganglion (DRG) and spinal cord after spared nerve injury (SNI) in rats. RT-PCR and in situ hybridization revealed that M-CSF and IL-34, but not GM- or G-CSF, mRNAs were constitutively expressed in the DRG, and M-CSF robustly increased in injured-DRG neurons. M-CSF receptor mRNA was expressed in naive rats and increased in spinal microglia following SNI. Intrathecal injection of M-CSF receptor inhibitor partially but significantly reversed the proliferation of spinal microglia and in early phase of neuropathic pain induced by SNI. Furthermore, intrathecal injection of recombinant M-CSF induced microglial proliferation and mechanical allodynia. Here, we demonstrate that M-CSF is a candidate molecule derived from primary afferents that induces proliferation of microglia in the spinal cord and leads to induction of neuropathic pain after peripheral nerve injury in rats. PMID:27071004

  8. Macrophage-Colony Stimulating Factor Derived from Injured Primary Afferent Induces Proliferation of Spinal Microglia and Neuropathic Pain in Rats

    PubMed Central

    Okubo, Masamichi; Yamanaka, Hiroki; Kobayashi, Kimiko; Dai, Yi; Kanda, Hirosato; Yagi, Hideshi; Noguchi, Koichi

    2016-01-01

    Peripheral nerve injury induces proliferation of microglia in the spinal cord, which can contribute to neuropathic pain conditions. However, candidate molecules for proliferation of spinal microglia after injury in rats remain unclear. We focused on the colony-stimulating factors (CSFs) and interleukin-34 (IL-34) that are involved in the proliferation of the mononuclear phagocyte lineage. We examined the expression of mRNAs for macrophage-CSF (M-CSF), granulocyte macrophage-CSF (GM-CSF), granulocyte-CSF (G-CSF) and IL-34 in the dorsal root ganglion (DRG) and spinal cord after spared nerve injury (SNI) in rats. RT-PCR and in situ hybridization revealed that M-CSF and IL-34, but not GM- or G-CSF, mRNAs were constitutively expressed in the DRG, and M-CSF robustly increased in injured-DRG neurons. M-CSF receptor mRNA was expressed in naive rats and increased in spinal microglia following SNI. Intrathecal injection of M-CSF receptor inhibitor partially but significantly reversed the proliferation of spinal microglia and in early phase of neuropathic pain induced by SNI. Furthermore, intrathecal injection of recombinant M-CSF induced microglial proliferation and mechanical allodynia. Here, we demonstrate that M-CSF is a candidate molecule derived from primary afferents that induces proliferation of microglia in the spinal cord and leads to induction of neuropathic pain after peripheral nerve injury in rats. PMID:27071004

  9. American Society of Clinical Oncology guidelines for the use of hematopoietic colony-stimulating factors.

    PubMed

    Ozer, H

    1996-01-01

    The hematopoietic colony-stimulating factors have been introduced into clinical practice as additional supportive measures that can reduce the likelihood of neutropenic complications due to chemotherapy. Clinical benefit has been shown, but the high cost of colony-stimulating factors has led to concern about their appropriate use. The American Society of Clinical Oncology has established evidence-based, clinical practice guidelines for the use of colony-stimulating factors in patients who are not enrolled in clinical trials. An expert multidisciplinary panel reviewed the clinical data documenting the activity of colony-stimulating factors. For each common clinical situation, the panel formulated a guideline to encourage reasonable use of colony-stimulating factors to preserve effectiveness but discourage excess use when little marginal benefit is anticipated. Outcomes considered in evaluating colony stimulating factor benefit included duration of neutropenia, incidence of febrile neutropenia, incidence and duration of antibiotic use, frequency and duration of hospitalization, infectious mortality, chemotherapy dose intensity, chemotherapy efficacy, quality of life, colony-stimulating factor toxicity, and economic impact. To the extent that these data were available, the panel placed greatest value on survival benefit, reduction in rates of febrile neutropenia, decreased hospitalization, and reduced costs. Lesser value was placed on alterations in absolute neutrophil counts.

  10. Effect of oxygenated sterol compounds on human bone marrow granulocytic progenitor cells

    SciTech Connect

    Hoffman, P.C.; Richman, C.M.; Hsu, R.C.; Chung, J.; Scanu, A.M.; Yachnin, S.

    1981-01-01

    Oxygenated sterol compounds are potent inhibitors of sterol and DNA synthesis in mammalian cells. We studied the effects of oxygenated sterols on human marrow granulocytic progenitor cells in vitro (CFU-C)*. 25-Hydroxycholesterol, as well as 6-ketocholestanol, 7-ketocholesterol, and 20 ..cap alpha..-hydroxycholesterol, demonstrated marked inhibition of CFU-C proliferation. The latter effect, which was not a result of direct cytoxicity of the compounds, was reversible by cholesterol, but not by mevalonic acid. We conclude that inhibition of sterol synthesis by oxygenated sterol compounds may be insufficient to explain their suppression of CFU-C proliferation.

  11. Reaction of the human granulocyte system in successfully treated bacterial bronchopneumonia.

    PubMed Central

    Kruszewski, J; Szczylik, C; Wiktor-Jedrzejczak, W

    1982-01-01

    The behaviour of peripheral blood granulocyte pools, nitroblue tetrazolium reduction by granulocytes, migration of granulocytes out of skin windows, serum muramidase activity and bone marrow reserve of granulocytes during uncomplicated bronchopneumonia were studied. The time course of changes in these parameters during infection was established, and suggests maximal changes on day 2 of infection with gradual regression until 30 days after the disease onset, when they disappear. PMID:7037862

  12. Adenylate pool and energy charge in human lymphocytes and granulocytes irradiated at 632 nm (HeNe laser)

    NASA Astrophysics Data System (ADS)

    Bolognani, Lorenzo; Venturelli, T.; Volpi, N.; Zirilli, O.

    1995-05-01

    Aim of this report was to investigate the adenylate pool and the energy charge in human white blood cells exposed to increasing time (15, 30 and 60 min) of HeNe laser treatment. EDTA treated human blood diluted 1:1 with 0.88% KCl was added (1:5) with NaCl-dextran solution to allow sedimentation of red blood cells. 6 ml of the white cells floating in the supernatant were layered on 3 ml of Lymphoprep in plastic tubes and each tube was centrifuged (from 50 to 5000 X g for 5 min). Granulocytes were concentrated in the lower phase, whilst lymphocytes were in the intermediated phase. After further purification cytological homogeneity was tested by a cell counter. Granulocytes and lymphocytes were irradiated at +22°C with HeNe (Space, Valfivre equipment). On these population ATP was tested by luminometric procedure, the adenylate pool was separated by HPLC (Jasco) on neutralyzed perchloric extracts. ATP concentration increased in lymphocytes (+63.9%, p < 0.01) and in granulocytes (+25.0%, p < 0.05) after 60 min irradiation. The adenylate pool (tested by HPLC) does not change significatively in lymphocytes or granulocytes after 30 min irradiation, whilst in 60 min irradiated lymphocytes and granulocytes a significative increment was observed in nucleotide concentration. No changes were observed in energy charge according to Atkinson.

  13. Multipronged attenuation of macrophage-colony stimulating factor signaling by Epstein-Barr virus BARF1

    SciTech Connect

    Shim, Ann Hye-Ryong; Chang, Rhoda Ahn; Chen, Xiaoyan; Longnecker, Richard; He, Xiaolin

    2014-10-02

    The ubiquitous EBV causes infectious mononucleosis and is associated with several types of cancers. The EBV genome encodes an early gene product, BARF1, which contributes to pathogenesis, potentially through growth-altering and immune-modulating activities, but the mechanisms for such activities are poorly understood. We have determined the crystal structure of BARF1 in complex with human macrophage-colony stimulating factor (M-CSF), a hematopoietic cytokine with pleiotropic functions in development and immune response. BARF1 and M-CSF form a high-affinity, stable, ring-like complex in both solution and the crystal, with a BARF1 hexameric ring surrounded by three M-CSF dimers in triangular array. The binding of BARF1 to M-CSF dramatically reduces but does not completely abolish M-CSF binding and signaling through its cognate receptor FMS. A three-pronged down-regulation mechanism is proposed to explain the biological effect of BARF1 on M-CSF:FMS signaling. These prongs entail control of the circulating and effective local M-CSF concentration, perturbation of the receptor-binding surface of M-CSF, and imposition of an unfavorable global orientation of the M-CSF dimer. Each prong may reduce M-CSF:FMS signaling to a limited extent but in combination may alter M-CSF:FMS signaling dramatically. The downregulating mechanism of BARF1 underlines a viral modulation strategy, and provides a basis for understanding EBV pathogenesis.

  14. Mechanisms of leukocyte accumulation and activation in chorioamnionitis: interleukin 1 beta and tumor necrosis factor alpha enhance colony stimulating factor 2 expression in term decidua.

    PubMed

    Arcuri, Felice; Toti, Paolo; Buchwalder, Lynn; Casciaro, Alessandra; Cintorino, Marcella; Schatz, Frederick; Rybalov, Basya; Lockwood, Charles J

    2009-05-01

    Chorioamnionitis is a major cause of prematurity as well as perinatal morbidity and mortality. The present study observed a marked increase in immunohistochemical staining for Colony Stimulating Factor 2 (CSF2; also known as granulocyte macrophage-colony stimulating factor), a potent neutrophil and macrophage chemoattractant and activator, in the decidua of patients with CAM compared with controls (n = 8; P = .001). To examine the regulation of this CSF2, cultured decidual cells primed with estradiol (E2) or E2 plus medroxyprogesterone acetate, were exposed to tumor necrosis factor-alpha or interleukin-1beta and secreted CSF2 measured by ELISA. Levels of CSF2 in E2 plus MPA-treated cultures increased 18- and 245-fold following treatment with TNF or IL1B (n = 7, P < .05). Quantitative RT-PCR demonstrated parallel changes in mRNA levels. This study reveals that CSF2 is strongly expressed in decidua from patients with CAM and indicates TNF or IL1B as important regulators of CAM-related decidual leukocyte infiltration and activation.

  15. Stimulation of proliferation, differentiation, and function of human cells by primate interleukin 3

    SciTech Connect

    Lopez, A.F.; To, L.B.; Yang, Y.C.; Gamble, J.R.; Shannon, M.F.; Burns, G.F.; Dyson, P.G.; Juttner, C.A.; Clark, S.; Vadas, M.A.

    1987-05-01

    Cloned gibbon interleukin 3 (gIL-3) was found to stimulate the proliferation and differentiation of human bone marrow cells to produce day-14 granulocyte, macrophage, granulocyte-macrophage, and eosinophil colonies in semisolid agar. In the presence of normal human plasma, gIL-3 stimulated megakaryocytes. In methylcellulose cultures, it stimulated erythroid colonies in the presence, but not in the absence, of erythropoietin. When mature human leukocytes were used, gIL-3 stimulated the function of purified mature eosinophils as measured by the capacity to kill /sup 51/Cr-labeled antibody-coated target cells, to produce superoxide anions, and to phagocytize opsonized yeast particles in a manner similar to recombinant human granulocyte-macrophage colony-stimulating factor. In contrast, gIL-3 did not significantly stimulate any of the neutrophil functions tested, whereas human recombinant granulocyte-macrophage colony-stimulating factor was active in these assay. Among cytokines that are active on human hematopoietic cells, gIL-3 thus has a distinct set of functions and may predict the range of actions of the human molecule.

  16. Isolation and characterization of elasnin, a new human granulocyte elastase inhibitor produced by a strain of Streptomyces.

    PubMed

    Ohno, H; Saheki, T; Awaya, J; Nakagawa, A; Omura, S

    1978-11-01

    Elasnin, a new human granulocyte elastase inhibitor, produced by the strain of KM-2753 designated as Streptomyces noboritoensis KM--2753 has been isolated from the fermentation broth by column chromatography on silica gel and neutral alumina. Elasnin is a neutral, colorless, and viscous oil (ND17 = 1.4983, [alpha]18D -0.9 degrees, lambdaEtOHmax 291 nm (epsilon, 7,760) having a molecular formula of C24H40O4 (MW 392) as shown by its elemental analysis and mass spectrum. Elasnin markedly inhibits human granulocyte elastase, but it is almost inactive against pancreatic elastase, chymotrypsin, and trypsin. At 1.3 microgram/ml (3.3 X 10(-6) M), elasnin is 50% inhibitory to human elastase, but it causes 50% inhibition of pancreatic elastase at 30.1 microgram/ml (76.8 X 10(-6) M). PMID:721707

  17. Natural infection of small mammal species in Minnesota with the agent of human granulocytic ehrlichiosis.

    PubMed

    Walls, J J; Greig, B; Neitzel, D F; Dumler, J S

    1997-04-01

    The natural reservoirs for the agent of human granulocytic ehrlichiosis (HGE) are suspected to be the small mammals that host immature stages of Ixodes scapularis ticks. To determine if such small mammals are naturally infected, we collected blood and serum samples from small mammal species in rural and suburban areas of Minneapolis and St. Paul, Minn. Samples were collected from white-footed mice (Peromyscus leucopus), eastern chipmunks (Tamias striatus), southern red-backed voles (Clethrionomys gapperi), and insectivorous shrews (Blarina brevicauda and Sorex cinereus). Blood samples were tested by PCR for active infection with the HGE agent, and sera from P. leucopus mice were tested for serologic evidence of infection by indirect immunofluorescence. PCR analyses revealed the presence of HGE agent DNA in 20 of the 190 samples (10.5%) tested. Of the 119 P. leucopus mouse serum samples that were analyzed, 12 (10.1%) contained Ehrlichia equi antibodies. In 3 of 119 (2.5%) P. leucopus mice from which both blood and serum were collected. HGE agent DNA and antibodies against E. equi were present. Animals with evidence of infection with the HGE agent are widely distributed around the Minneapolis-St. Paul area in regions with known I. scapularis tick activity. Small mammals that are frequent hosts for larval I. scapularis ticks and that are found in areas where HGE occurs are likely to be a major reservoir from which infected ticks that bite humans are derived. PMID:9157141

  18. Binding inhibition of type 1 fimbriae to human granulocytes: a flow cytometric inhibition assay using trivalent cluster mannosides.

    PubMed

    Horst, A K; Kötter, S; Lindhorst, T K; Ludwig, A; Brandt, E; Wagener, C

    2001-12-01

    The binding of type 1 fimbriae from Escherichia coli to vital human neutrophilic granulocytes was inhibited by synthetic trivalent cluster mannosides. Binding of type 1 fimbriae was measured in a flow cytometric assay. Based on the molarity of mannosyl residues, the clusters exceed the inhibitory potency of methyl alpha-D-mannoside by a factor of more than 1,000 and the inhibitory potency of p-nitrophenyl alpha-D-mannoside by a factor of more than 10. The inhibition studies indicate that the trivalent cluster mannosides are very potent inhibitors of the binding of type 1 fimbriae to human neutrophilic granulocytes. Based on their defined structure, cluster mannosides are well suited for characterizing the molecular interactions of mannose-sensitive fimbriae with their cell membrane receptors.

  19. Role of macrophage colony-stimulating factor in the development of neointimal thickening following arterial injury.

    PubMed

    Mishra, Vivek; Sinha, Satyesh K; Rajavashisth, Tripathi B

    2016-01-01

    Evidence suggests that macrophage colony-stimulating factor (M-CSF) participates critically in atherosclerosis; little is known about the role of M-CSF in the development of neointimal hyperplasia following mechanical vascular injury. We examined the expression of M-CSF and its receptor, c-fms, in rodent and rabbit models of arterial injury. Injured rat carotid arteries expressed 3- to 10-fold higher levels of M-CSF and c-fms mRNA and protein following balloon injury as compared to uninjured arteries. In the rabbit, M-CSF protein expression was greatest in neointimal smooth muscle cells (SMCs) postinjury, with some expression in medial SMCs. M-CSF-positive SMCs exhibited markers of proliferation. At 30days postinjury, neointimal SMCs in the adjacent healed area near the border between injured and uninjured zone lost both proliferative activity and overexpression of M-CSF. The presence of induced M-CSF and c-fms expression correlated with the initiation of SMCs proliferation. M-CSF stimulated incorporation of [(3)H] thymidine in human aortic smooth muscle cells in a concentration-dependent manner. Serum-free conditioned medium from aortic SMCs also promoted DNA synthesis, and this effect was blocked by M-CSF specific antibody. To test further the role of M-CSF in vivo, we induced arterial injury by placing a periadventitial collar around the carotid arteries in compound mutant mice lacking apolipoprotein apoE (apoE(-/-)) and M-CSF. Loss of M-CSF abolished the neointimal hyperplastic response to arterial injury in apoE(-/-) mice. Local delivery of M-CSF to the injured artery restored neointimal proliferation, suggesting a critical role of M-CSF for the development of neointimal thickening following arterial injury. PMID:27135205

  20. Positive interaction of thyme (red) essential oil with human polymorphonuclear granulocytes in eradicating intracellular Candida albicans.

    PubMed

    Tullio, Vivian; Mandras, Narcisa; Allizond, Valeria; Nostro, Antonia; Roana, Janira; Merlino, Chiara; Banche, Giuliana; Scalas, Daniela; Cuffini, Anna Maria

    2012-10-01

    The essential oils have started to be recognized for their potential antimicrobial role only in recent years. Clinical experience showed that the efficacy of antimicrobial agents depends not only on their direct effect on a given microorganism but also on the functional activity of the host immune system. Since data on the effects of essential oils on the innate immune system are scanty and fragmentary, the aim of this study was to evaluate the influence of thyme (red) essential oil (EO), at subinhibitory/inhibitory concentrations, on intracellular killing activity by human polymorphonuclear granulocytes (PMNs) against Candida albicans. In order to provide a frame of reference for the activity of this EO, its in vitro killing activity in the absence of PMNs was also evaluated.Results showed that EO at subminimal inhibitory (subMIC)/minimal inhibitory (MIC) concentrations significantly enhanced intracellular killing of C. albicans in comparison with EO-free controls and was comparable to the positive control (fluconazole). In in vitro killing assays without PMNs, we observed progressive growth of the yeast cells in the presence of EO subMIC/MIC concentrations. A positive antifungal interaction with phagocytes could explain why this EO, which appeared to be only fungistatic in time-kill assays, had efficacy in killing yeast cells once incubated with PMNs. PMID:22872591

  1. Serologic Testing for Human Granulocytic Ehrlichiosis at a National Referral Center

    PubMed Central

    Comer, James A.; Nicholson, William L.; Olson, James G.; Childs, James E.

    1999-01-01

    An indirect immunofluorescence assay (IFA) was used to identify patients with antibodies reactive to the human granulocytic ehrlichiosis (HGE) agent. Serum samples collected from clinically ill individuals were submitted to the Centers for Disease Control and Prevention by physicians via state health departments from throughout the United States and tested against a panel of ehrlichial and rickettsial pathogens. Antibodies reactive to the HGE agent were detected in 142 (8.9%) of 1,602 individuals tested. There were 19 confirmed and 59 probable (n = 78) cases of HGE as defined by seroconversion or a fourfold or higher titer to the HGE agent than to the Ehrlichia chaffeensis antigens. The average age of patients with HGE was 57 years, and males accounted for 53 (68%) of the patients. Cases of HGE occurred in 21 states; 47 (60%) of the cases occurred in Connecticut (n = 14), New York (n = 18), and Wisconsin (n = 15). Onset of HGE was identified from April through December, with cases peaking in June and July. The earliest confirmed cases of HGE occurred in 1987 in Wisconsin and 1988 in Florida. No fatalities were reported among the 78 patients with confirmed or probable HGE. Reactivity to the HGE agent and to either Coxiella burnetii, Rickettsia rickettsii, or Rickettsia typhi was infrequent; however, 74 (52%) of the 142 individuals who were positive for HGE had at least one serum sample that also reacted to the E. chaffeensis antigen. Thirty-four persons with confirmed or probable human monocytic ehrlichiosis due to E. chaffeensis also had antibodies to the HGE agent in at least one serum sample. The specific etiologic agent for 30 patients was not ascribed because of similarity of titers to both ehrlichial antigens. The use of both antigens may be required to correctly diagnose most cases of human ehrlichiosis, especially in geographic regions where both the HGE agent and E. chaffeensis occur. PMID:9986812

  2. Characterization of an immunoreactive protein from the agent of human granulocytic ehrlichiosis.

    PubMed Central

    Kolbert, C P; Bruinsma, E S; Abdulkarim, A S; Hofmeister, E K; Tompkins, R B; Telford, S R; Mitchell, P D; Adams-Stich, J; Persing, D H

    1997-01-01

    A gene that is homologous to the Ehrlichia chaffeensis groEL operon was recovered and characterized by broad-range PCR amplification of whole blood from patients with human granulocytic ehrlichiosis (HGE) and from infected HL60 cell cultures. Sequence analysis of an 820-bp DNA fragment recovered directly from human blood showed 76.5 and 76.3% identity with cognate sequences from E. chaffeensis and Cowdria ruminantium, respectively. Analysis of a 1.6-kb DNA fragment derived from an HGE agent-infected HL60 cell culture indicated a near-complete open reading frame that contained 75.6 and 75.2% sequence identity with the E. chaffeensis and C. ruminantium groEL sequences, respectively. Phylogenetic analysis of this fragment showed that the HGE agent-derived sequence was related to, but distinct from, the sequences of E. chaffeensis and C. ruminantium. Polyvalent antibody responses to a recombinant fusion protein based on the HGE agent groEL homolog were detected in three of three BALB/c mice that were infected by syringe inoculation with a Wisconsin strain of the HGE agent (WI-1) and nine of nine mice infected by Ixodes scapularis (Ixodes dammini) tick inoculation of an isolate from Nantucket Island, Mass. (NCH-1). No response was detected in mice infected with Borrelia burgdorferi or in control BALB/c mice. Further characterization of the sensitivity and specificity of immune responses to this protein will be facilitated by the use of recombinant fusion proteins or peptides based on the HGE agent-specific groEL homolog. PMID:9114402

  3. Mobilization of hematopoietic progenitor cells with granulocyte colony stimulating factors for autologous transplant in hematologic malignancies: a single center experience

    PubMed Central

    Gabús, Raul; Borelli, Gabriel; Ferrando, Martín; Bódega, Enrique; Citrín, Estela; Jiménez, Constanza Olivera; Álvarez, Ramón

    2011-01-01

    Background In 2006 the Hematology Service of Hospital Maciel published its experience with peripheral blood progenitor cell harvesting for autologous stem cell transplantation using Filgen JP (Clausen Filgrastim). After mobilization with a mean filgrastim dose of 78 mcg/Kg, 4.7 x 106 CD34+ cells/Kg were obtained by apheresis. Age above 50, multiple myeloma as underlying disease and a malignancy that was not in remission were identified as frequent characteristics among patients showing complex mobilization. Objective The aim of this study was to compare stem cell mobilization using different brands of filgrastim. Methods One hundred and fifty-seven mobilizations performed between 1997 and 2006 were analyzed. This retrospective analysis comparative two groups of patients: those mobilized with different brands of filgrastim (Group A) and those who received Filgen JP (Clausen Filgrastim) as mobilizing agent (Group B). A cluster analysis technique was used to identify four clusters of individuals with different behaviors differentiated by age, total dose of filgrastim required, number of apheresis and harvested CD34+ cells. Results The mean total dose of filgrastim administered was 105 mcg/Kg, the median number of apheresis was 2 procedures and the mean number of harvested stem cells was 4.98 x 106 CD34+ cells/Kg. No significant differences were observed between Groups A and B regarding the number of apheresis, harvested CD34+ cells and number of mobilization failures, however the total dose of filgrastim was significantly lower in Group B. Conclusions Among other factors, the origin of the cytokine used as mobilizing agent is an element to be considered when evaluating CD34+ cell mobilization results. PMID:23049356

  4. Interference between the agents of Lyme disease and human granulocytic ehrlichiosis in a natural reservoir host.

    PubMed

    Levin, M L; Fish, D

    2001-01-01

    Agents of Lyme disease (Borrelia burgdorferi) and human granulocytic ehrlichiosis (Ehrlichia phagocytophila) are perpetuated in a natural cycle involving the black-legged tick (Ixodes scapularis) and its vertebrate hosts. Using I. scapularis nymphs as the mode of infectious challenge, we studied how infection with one pathogen in white-footed mice (Peromyscus leucopus) affects their ability to acquire the other agent and subsequently to infect larvae, which these agents would do in nature. Two groups of mice were infected with either B. burgdorferi or E. phagocytophila. One week later, B. burgdorferi-infected mice were challenged with E. phagocytophila, and E. phagocytophila-infected mice were challenged with B. burgdorferi. Simultaneously, two control groups of uninfected mice were infected with each agent from the same tick cohorts used on the first groups of mice. Uninfected I. scapularis larvae were fed on all mice for xenodiagnosis at weekly intervals lasting 2 months. For the B. burgdorferi challenge, all control and E. phagocytophila-infected mice acquired B. burgdorferi. However, fewer xenodiagnostic larvae acquired B. burgdorferi from mice with mixed infections compared with mice infected with B. burgdorferi only. For the E. phagocytophila challenge, all five control mice acquired E. phagocytophila, but only two of five mice infected with B. burgdorferi subsequently acquired E. phagocytophila. Consequently, mice with both infections produced fewer xenodiagnostic ticks infected with E. phagocytophila than mice infected with E. phagocytophila only. Thus, a primary infection with either B. burgdorferi or E. phagocytophila in mice inhibited transmission of a second agent, suggesting interference between these two agents.

  5. Recombinant human interleukin-3: pharmacokinetics after intravenous and subcutaneous bolus injection and effects on granulocyte kinetics.

    PubMed

    Hovgaard, D J; Folke, M; Mortensen, B T; Nissen, N I

    1994-08-01

    The pharmacokinetics of E. coli derived recombinant human interleukin-3 (rhIL-3) was studied following intravenous (i.v.) and subcutaneous (s.c.) bolus injection of rhIL-3. After i.v. bolus injection in eight patients, serum peak levels of 34.5-135.0 ng/ml were reached, followed by a rapid decline with a t1/2 alpha of 17 +/- 2 min and a t1/2 beta of 59 +/- 7 min. After s.c. bolus injection in five patients, the absorption was more prolonged with peak serum levels reached at 2.8 +/- 0.4 h. Elimination was also more protracted, and serum base-line levels were reached at 14-24 h. The immediate effect of rhIL-3 on peripheral white blood cells was less pronounced and more variable than previously found for G- or GM-CSF. Following i.v. administration, neutrophils showed a moderate drop to median 64% of initial values (range 42-85%) at median 30 min after injection (range 15-60 min) followed by an increase at 24 h to 69-288% of initial values. Eosinophils dropped to a median nadir of 34% and then gradually increased to maximum values in the range 135-720% at 18-24 h. The effect of rhIL-3 was further examined following i.v. injection of autologous 111Indium-labelled granulocytes in six patients. In steady state, i.v. injection of rhIL-3 caused a moderate drop in 111Indium activity of peripheral blood within 20 min without tendency to subsequent recovery. No change occurred in the activity recorded over the lungs and liver. The activity over the spleen decreased moderately in two patients. These results are strikingly different from those previously obtained after i.v. injection of rhGM-CSF.

  6. Thalidomide inhibits granulocyte responses in healthy humans after ex vivo stimulation with bacterial antigens.

    PubMed

    Juffermans, N P; Verbon, A; Schultz, M J; Hack, C E; van Deventer, S J; Speelman, P; van der Poll, T

    2001-05-01

    Ingestion of thalidomide was associated with a reduction in the upregulation of the granulocyte activation marker CD11b and a reduced capacity to release elastase and lactoferrin after stimulation with lipopolysaccharide or lipoteichoic acid. A single oral dose of thalidomide attenuates neutrophil activation upon ex vivo stimulation with bacterial antigens.

  7. Regulation of urokinase-type plasminogen activator gene transcription by macrophage colony-stimulating factor.

    PubMed Central

    Stacey, K J; Fowles, L F; Colman, M S; Ostrowski, M C; Hume, D A

    1995-01-01

    The mouse urokinase-type plasminogen activator (uPA) gene was used as a model macrophage colony-stimulating factor 1 (CSF-1)-inducible gene to investigate CSF-1 signalling pathways. Nuclear run-on analysis showed that induction of uPA mRNA by CSF-1 and phorbol myristate acetate (PMA) was at the transcriptional level in bone marrow-derived macrophages. CSF-1 and PMA synergized strongly in the induction of uPA mRNA, showing that at least some components of CSF-1 action are mediated independently of protein kinase C. Promoter targets of CSF-1 signalling were investigated with NIH 3T3 cells expressing the human CSF-1 receptor (c-fms). uPA mRNA was induced in these cells by treatment with CSF-1, and a PEA3/AP-1 element at -2.4 kb in the uPA promoter was involved in this response. Ets transcription factors can act through PEA3 sequences, and the involvement of Ets factors in the induction of uPA was confirmed by use of a dominant negative Ets-2 factor. Expression of the DNA binding domain of Ets-2 fused to the lacZ gene product prevented CSF-1-mediated induction of uPA mRNA in NIH 3T3 cells expressing the CSF-1 receptor. Examination of ets-2 mRNA expression in macrophages showed that it was also induced synergistically by CSF-1 and PMA. In the macrophage cell line RAW264, the uPA PEA3/AP-1 element mediated a response to both PMA and cotransfected Ets-2. uPA promoter constructs were induced 60- to 130-fold by Ets-2 expression, and the recombinant Ets-2 DNA binding domain was able to bind to the uPA PEA3/AP-1 element. This work is consistent with a proposed pathway for CSF-1 signalling involving sequential activation of fms, ras, and Ets factors. PMID:7760840

  8. Cloning and expression of porcine Colony Stimulating Factor-1 (CSF-1) and Colony Stimulating Factor-1 Receptor (CSF-1R) and analysis of the species specificity of stimulation by CSF-1 and Interleukin 34.

    PubMed

    Gow, Deborah J; Garceau, Valerie; Kapetanovic, Ronan; Sester, David P; Fici, Greg J; Shelly, John A; Wilson, Thomas L; Hume, David A

    2012-12-01

    Macrophage Colony Stimulating Factor (CSF-1) controls the survival, differentiation and proliferation of cells of the mononuclear phagocyte system. A second ligand for the CSF-1R, Interleukin 34 (IL-34), has been described, but its physiological role is not yet known. The domestic pig provides an alternative to traditional rodent models for evaluating potential therapeutic applications of CSF-1R agonists and antagonists. To enable such studies, we cloned and expressed active pig CSF-1. To provide a bioassay, pig CSF-1R was expressed in the factor-dependent Ba/F3 cell line. On this transfected cell line, recombinant porcine CSF-1 and human CSF-1 had identical activity. Mouse CSF-1 does not interact with the human CSF-1 receptor but was active on pig. By contrast, porcine CSF-1 was active on mouse, human, cat and dog cells. IL-34 was previously shown to be species-specific, with mouse and human proteins demonstrating limited cross-species activity. The pig CSF-1R was equally responsive to both mouse and human IL-34. Based upon the published crystal structures of CSF-1/CSF-1R and IL34/CSF-1R complexes, we discuss the molecular basis for the species specificity.

  9. Isolation of the etiologic agent of human granulocytic ehrlichiosis from the white-footed mouse (Peromyscus leucopus).

    PubMed

    Ravyn, M D; Kodner, C B; Carter, S E; Jarnefeld, J L; Johnson, R C

    2001-01-01

    We examined white-footed mice (Peromyscus leucopus) from Minnesota for infection with the etiologic agent of human granulocytic ehrlichiosis (HGE). From April to September 1997, we collected P. leucopus from Washington County, Minnesota, an area enzootic for HGE. Blood was cultivated in HL60 cells for isolation of the HGE agent. Of 59 mice examined, only a single mouse was culture positive for the HGE agent. The 16S ribosomal DNA sequence of the isolate was determined to be identical to that of the HGE agent. The isolate was reactive with monoclonal antibodies to the 44-kDa antigen of the HGE agent and was infectious for laboratory mice.

  10. Measuring Granulocyte and Monocyte Phagocytosis and Oxidative Burst Activity in Human Blood.

    PubMed

    Meaney, Mary Pat; Nieman, David C; Henson, Dru A; Jiang, Qi; Wang, Fu-Zhang

    2016-01-01

    The granulocyte and monocyte phagocytosis and oxidative burst (OB) activity assay can be used to study the innate immune system. This manuscript provides the necessary methodology to add this assay to an exercise immunology arsenal. The first step in this assay is to prepare two aliquots ("H" and "F") of whole blood (heparin). Then, dihydroethidium is added to the H aliquot, and both aliquots are incubated in a warm water bath followed by a cold water bath. Next, Staphylococcus aureus (S. aureus) is added to the H aliquot and fluorescein isothiocyanate-labeled S. aureus is added to the F aliquot (bacteria:phagocyte = 8:1), and both aliquots are incubated in a warm water bath followed by a cold water bath. Then, trypan blue is added to each aliquot to quench extracellular fluorescence, and the cells are washed with phosphate-buffered saline. Next, the red blood cells are lysed, and the white blood cells are fixed. Finally, a flow cytometer and appropriate analysis software are used to measure granulocyte and monocyte phagocytosis and OB activity. This assay has been used for over 20 years. After heavy and prolonged exertion, athletes experience a significant but transient increase in phagocytosis and an extended decrease in OB activity. The post-exercise increase in phagocytosis is correlated with inflammation. In contrast to normal weight individuals, granulocyte and monocyte phagocytosis is chronically elevated in overweight and obese participants, and is modestly correlated with C-reactive protein. In summary, this flow cytometry-based assay measures the phagocytosis and OB activity of phagocytes and can be used as an additional measure of exercise- and obesity-induced inflammation. PMID:27684595

  11. Colony stimulating factor 1 is an extrinsic stimulator of mouse spermatogonial stem cell self-renewal

    PubMed Central

    Oatley, Jon M.; Oatley, Melissa J.; Avarbock, Mary R.; Tobias, John W.; Brinster, Ralph L.

    2009-01-01

    Summary Self-renewal and differentiation of spermatogonial stem cells (SSCs) provide the foundation for testis homeostasis, yet mechanisms that control their functions in mammals are poorly defined. We used microarray transcript profiling to identify specific genes whose expressions are augmented in the SSC-enriched Thy1+ germ cell fraction of mouse pup testes. Comparisons of gene expression in the Thy1+ germ cell fraction with the Thy1-depleted testis cell population identified 202 genes that are expressed 10-fold or higher in Thy1+ cells. This database provided a mining tool to investigate specific characteristics of SSCs and identify novel mechanisms that potentially influence their functions. These analyses revealed that colony stimulating factor 1 receptor (Csf1r) gene expression is enriched in Thy1+ germ cells. Addition of recombinant colony stimulating factor 1 (Csf1), the specific ligand for Csf1r, to culture media significantly enhanced the self-renewal of SSCs in heterogeneous Thy1+ spermatogonial cultures over a 63-day period without affecting total germ cell expansion. In vivo, expression of Csf1 in both pre-pubertal and adult testes was localized to clusters of Leydig cells and select peritubular myoid cells. Collectively, these results identify Csf1 as an extrinsic stimulator of SSC self-renewal and implicate Leydig and myoid cells as contributors of the testicular stem cell niche in mammals. PMID:19270176

  12. Differential modulation of IL-1-induced endothelial adhesion molecules and transendothelial migration of granulocytes by G-CSF.

    PubMed

    Eissner, G; Lindner, H; Reisbach, G; Klauke, I; Holler, E

    1997-06-01

    Granulocyte colony stimulating factor (G-CSF) is widely used for mobilization of haemopoietic stem cells into the peripheral blood. However, little is known about the mechanisms involved in mobilization and the immune modulatory effects of this growth factor. In this report we show that G-CSF down-regulated intercellular adhesion molecule 1 (ICAM-1) induced by Interleukin-1 (IL-1) on human endothelial cells. Interestingly, the G-CSF-mediated down-modulation of IL-1-induced ICAM-1 appeared to be biphasic. In pharmacological concentrations (> 300 ng/ml), and in dose ranges of plasma G-CSF levels above that of nonfebrile healthy individuals (30 pg/ml), a significant decrease in surface ICAM-1 could be observed. This could be explained, at least in part, by an increased autocrine G-CSF production by endothelial cells in response to IL-1 and exogenous G-CSF. In contrast to ICAM-1, IL-1-triggered VCAM-1 expression was superinduced by G-CSF with the optimal concentration of 30 pg/ml. To evaluate the functional significance of these findings, 51Cr adhesion assays with peripheral blood mononuclear cells (PBMC) or granulocytes known to lack the VCAM-1 counter-receptor very late antigen 4 (VLA-4) and IL-1-stimulated endothelial cells, in the presence or absence of G-CSF, were performed. G-CSF could not inhibit the IL-1-induced adhesion of PBMC to endothelial cells, which may be due to the differential adhesion molecule modulation. In contrast, granulocyte adhesion induced by IL-1 could effectively be blocked by co-incubation with G-CSF. Finally, G-CSF also inhibited transendothelial migration of granulocytes through IL-1-activated endothelial cells in a concentration-dependent manner.

  13. Prediction of the three-dimensional structure of human interleukin-7 by homology modeling.

    PubMed

    Kroemer, R T; Doughty, S W; Robinson, A J; Richards, W G

    1996-06-01

    The three-dimensional structure of human interleukin (IL)-7 has been predicted based on homology to human IL-2, IL-4, granulocyte-macrophage colony stimulating factor and growth hormone. The model has a topology common to other cytokines and displays a unique disulfide pattern. Knowledge of the tertiary structure of IL-7 has implications for analysis of key binding regions, suggestions for mutagenesis experiments and design of (ant)agonists. In this context, the model is discussed and compared with other cytokine structures. PMID:8862549

  14. Colony-stimulating factor 1 receptor (CSF1R) signaling in injured neurons facilitates protection and survival.

    PubMed

    Luo, Jian; Elwood, Fiona; Britschgi, Markus; Villeda, Saul; Zhang, Hui; Ding, Zhaoqing; Zhu, Liyin; Alabsi, Haitham; Getachew, Ruth; Narasimhan, Ramya; Wabl, Rafael; Fainberg, Nina; James, Michelle L; Wong, Gordon; Relton, Jane; Gambhir, Sanjiv S; Pollard, Jeffrey W; Wyss-Coray, Tony

    2013-01-14

    Colony-stimulating factor 1 (CSF1) and interleukin-34 (IL-34) are functional ligands of the CSF1 receptor (CSF1R) and thus are key regulators of the monocyte/macrophage lineage. We discovered that systemic administration of human recombinant CSF1 ameliorates memory deficits in a transgenic mouse model of Alzheimer's disease. CSF1 and IL-34 strongly reduced excitotoxin-induced neuronal cell loss and gliosis in wild-type mice when administered systemically before or up to 6 h after injury. These effects were accompanied by maintenance of cAMP responsive element-binding protein (CREB) signaling in neurons rather than in microglia. Using lineage-tracing experiments, we discovered that a small number of neurons in the hippocampus and cortex express CSF1R under physiological conditions and that kainic acid-induced excitotoxic injury results in a profound increase in neuronal receptor expression. Selective deletion of CSF1R in forebrain neurons in mice exacerbated excitotoxin-induced death and neurodegeneration. We conclude that CSF1 and IL-34 provide powerful neuroprotective and survival signals in brain injury and neurodegeneration involving CSF1R expression on neurons.

  15. Increased macrophage colony-stimulating factor in neonatal and adult autoimmune MRL-lpr mice.

    PubMed Central

    Yui, M. A.; Brissette, W. H.; Brennan, D. C.; Wuthrich, R. P.; Rubin-Kelley, V. E.

    1991-01-01

    Abnormal macrophages in MRL-lpr mice are implicated in the pathogenesis of autoimmune disease. These mice die of lupus nephritis by 5 to 6 months of age. This study reports that MRL-lpr mice have an increased level of circulating macrophage colony-stimulating factor (M-CSF) detectable as early as 1 week of age. Macrophage colony-stimulating factor decreased between 2 and 4 months and then steadily increased beginning at 4 months of age. In contrast, M-CSF was not detected in sera from congenic MRL-++ mice, normal C3H/FeJ mice, two other mouse strains with the lpr gene (B6-lpr and C3H-lpr), or another lupus model, the NZB/W mouse. These observations indicate that the lpr gene alone is not responsible for inducing this growth factor, and elevated M-CSF is not required for all forms of murine lupus. The entire source of serum M-CSF is not clear. The unique T cells regulated by the lpr gene are not responsible for the increased serum M-CSF levels, as no M-CSFs could be detected in supernatants from cultured lymph nodes from MRL-lpr mice, and the steady-state levels of M-CSF mRNA in lymph nodes and spleens in MRL-lpr, C3H-lpr mice and in their respective congenic strains were similar. The steady-state M-CSF mRNA transcripts in liver, lung, and bone marrow in MRL-lpr, MRL-++, and C3H/FeJ mice were also similar. Macrophage colony-stimulating factor transcripts were clearly elevated in the kidneys of MRL-lpr mice, suggesting a renal source of circulating M-CSF. The increase of M-CSF might be responsible for the increased numbers and enhanced functions of macrophages, which in turn cause tissue destruction in MRL-lpr mice. Images Figure 1 Figure 2 Figure 3 Figure 4 PMID:1867317

  16. Human granulocytic anaplasmosis in Austria: epidemiological, clinical, and laboratory findings in five consecutive patients from Tyrol, Austria.

    PubMed

    Walder, Gernot; Fuchs, Dietmar; Sarcletti, Mario; Berek, Klaus; Falkensammer, Barbara; Huber, Klaus; Petrovec, Miro; Dierich, Manfred P; Würzner, Reinhard

    2006-05-01

    We report five consecutive cases of Anaplasma (A.) phagocytophilum infection (the causative agent of human granulocytic anaplasmosis (HGA)) from western Austria. All infections were acquired between June and August in 2003 and 2004 in the Inn valley (Tyrol, Austria). Four patients required hospitalisation, one patient was treated as an outpatient. During the acute stage of illness, laboratory findings included thrombocytopenia (5/5), elevated C-reactive protein (5/5), elevated neopterin (5/5), elevated lactate dehydrogenase (4/5), and elevation of liver enzymes (4/5). Leukopenia (3/5) and elevated procalcitonin (2/5) were less frequently observed. All patients were treated with tetracyclines, which led to prompt improvement of the clinical conditions. Anti-platelet antibodies were observed in one of four patients, but remained unchanged after complete covalescence.

  17. Possible mechanisms and function of nuclear trafficking of the colony-stimulating factor-1 receptor.

    PubMed

    Rovida, Elisabetta; Dello Sbarba, Persio

    2014-10-01

    Receptor tyrosine kinases (RTK) have long being studied with respect to the "canonical" signaling. This includes ligand-induced activation of a receptor tyrosine kinase at the cell surface that leads to receptor dimerization, followed by its phosphorylation in the intracellular domain and activation. The activated receptor then recruits cytoplasmic signaling molecules including other kinases. Activation of the downstream signaling cascade frequently leads to changes in gene expression following nuclear translocation of downstream targets. However, RTK themselves may localize within the nucleus, as either full-length molecules or cleaved fragments, with or without their ligands. Significant differences in this mechanism have been reported depending on the individual RTK, cellular context or disease. Accumulating evidences indicate that the colony-stimulating factor-1 receptor (CSF-1R) may localize within the nucleus. To date, however, little is known about the mechanism of CSF-1R nuclear shuttling, as well as the functional role of nuclear CSF-1R.

  18. Granulocyte defects and opioid receptors in chronic exposure to heroin or methadone in humans.

    PubMed

    Mazzone, A; Mazzucchelli, I; Fossati, G; Gritti, D; Fea, M; Ricevuti, G

    1994-11-01

    In order to elucidate better the immunological effect of opioid abuse in the absence of HIV infection as a confounding factor, granulocyte function was investigated in three groups of HIV-negative subjects, including 20 active parenteral heroin abusers (H), 20 long-term methadone-maintained former opiate abusers (M) and 20 healthy controls (C). Chemotaxis to N-formyl methionyl-leucyl-phenylalanine (fMLP), casein and activated plasma were markedly and similarly reduced (approx. 50%) in both H and M groups, as was true for superoxide production after fMLP and PMA stimulation, 47% decrease of C values. Polymorphonuclear (PMN) of H and M subjects also exhibited a very marked and similar reduction in the expression of CD11b/CD18 integrin receptors after fMLP treatment, with values that were less than 10% of those in controls, as observed by flow cytometry. In parallel, PMN of H and M individuals presented an approximately four-fold increase in opioid receptors numbers compared to controls, a significant inverse correlation existing between the increase in opiate receptors and defective chemotaxis. The possible mechanism underlying the observed changes in PMN of H and M individuals is discussed.

  19. Entry of Sanfetrinem into Human Polymorphonuclear Granulocytes and Its Cell-Associated Activity against Intracellular, Penicillin-Resistant Streptococcus pneumoniae

    PubMed Central

    Cuffini, Anna Maria; Tullio, Vivian; Bonino, Alessandro; Allocco, Alessandra; Palarchio, Angela Ianni; Carlone, Nicola A.

    1998-01-01

    The entry of antibiotics into phagocytes is necessary for activity against intracellular pathogens. The ability of sanfetrinem, the first member of a new class of antibiotics, to penetrate human polymorphonuclear granulocytes and its consequences upon subsequent phagocytosis and killing of ingested penicillin-resistant Streptococcus pneumoniae have been evaluated. Sanfetrinem penetrated into human polymorphonuclear leukocytes (PMNs) at all concentrations tested, with cellular concentration/extracellular concentration ratios of 6.6 to 5.03 and 4.21 when sanfetrinem was used at 0.25 to 0.5 and 1 μg/ml, respectively, within 30 min of incubation. The uptake was complete within 5 min and was not energy dependent, since it was not affected by cell viability, environmental temperature, or the addition of a metabolic inhibitor. At a concentration of one-half the MIC, sanfetrinem significantly enhanced human PMN phagocytosis and increased intracellular bactericidal activity against penicillin-resistant S. pneumoniae. Following preexposure of PMNs to a concentration of one-half the MIC of sanfetrinem, there was a significant increase in both phagocytosis and killing compared with that for the controls, indicating the ability of sanfetrinem to interact with biological membranes and remain active within PMNs. Preexposure of streptococci to sanfetrinem made penicillin-resistant S. pneumoniae more susceptible to the bactericidal mechanisms of human PMNs than untreated organisms. PMID:9661015

  20. Colony-stimulating factor 1 regulates CTP: phosphocholine cytidylyltransferase mRNA levels.

    PubMed

    Tessner, T G; Rock, C O; Kalmar, G B; Cornell, R B; Jackowski, S

    1991-09-01

    Growth factor regulation of phosphatidylcholine (PtdCho) metabolism during the G1 stage of the cell cycle was investigated in the colony-stimulating factor 1 (CSF-1)-dependent murine macrophage cell-line BAC1.2F5. The transient removal of CSF-1 arrested the cells in G1. Incorporation of [3H]choline into PtdCho was stimulated significantly 1 h after growth factor addition to quiescent cells. Metabolic labeling experiments pointed to CTP:phosphocholine cytidylyltransferase (CT) as the rate-controlling enzyme for PtdCho biosynthesis in BAC1.2F5 cells. The amount of CT mRNA increased 4-fold within 15 min of CSF-1 addition and remained elevated for 2 h. The rise in CT mRNA levels was accompanied by a 50% increase in total CT specific activity in cell extracts within 4 h after the addition of CSF-1. CSF-1-dependent elevation of CT mRNA content was neither attenuated nor superinduced by the inhibition of protein synthesis with cycloheximide. The rate of CT mRNA turnover decreased in the presence of CSF-1 indicating that message stabilization was a key factor in determining the levels of CT mRNA. These data point to increased CT mRNA abundance as a component in growth factor-stimulated PtdCho synthesis.

  1. The clinical assessment, treatment, and prevention of lyme disease, human granulocytic anaplasmosis, and babesiosis: clinical practice guidelines by the Infectious Diseases Society of America.

    PubMed

    Wormser, Gary P; Dattwyler, Raymond J; Shapiro, Eugene D; Halperin, John J; Steere, Allen C; Klempner, Mark S; Krause, Peter J; Bakken, Johan S; Strle, Franc; Stanek, Gerold; Bockenstedt, Linda; Fish, Durland; Dumler, J Stephen; Nadelman, Robert B

    2006-11-01

    Evidence-based guidelines for the management of patients with Lyme disease, human granulocytic anaplasmosis (formerly known as human granulocytic ehrlichiosis), and babesiosis were prepared by an expert panel of the Infectious Diseases Society of America. These updated guidelines replace the previous treatment guidelines published in 2000 (Clin Infect Dis 2000; 31[Suppl 1]:1-14). The guidelines are intended for use by health care providers who care for patients who either have these infections or may be at risk for them. For each of these Ixodes tickborne infections, information is provided about prevention, epidemiology, clinical manifestations, diagnosis, and treatment. Tables list the doses and durations of antimicrobial therapy recommended for treatment and prevention of Lyme disease and provide a partial list of therapies to be avoided. A definition of post-Lyme disease syndrome is proposed.

  2. Human Granulocytic Anaplasmosis in the United States from 2008 to 2012: A Summary of National Surveillance Data

    PubMed Central

    Dahlgren, F. Scott; Heitman, Kristen Nichols; Drexler, Naomi A.; Massung, Robert F.; Behravesh, Casey Barton

    2015-01-01

    Human granulocytic anaplasmosis is an acute, febrile illness transmitted by the ticks Ixodes scapularis and Ixodes pacificus in the United States. We present a summary of passive surveillance data for cases of anaplasmosis with onset during 2008–2012. The overall reported incidence rate (IR) was 6.3 cases per million person-years. Cases were reported from 38 states and from New York City, with the highest incidence in Minnesota (IR = 97), Wisconsin (IR = 79), and Rhode Island (IR = 51). Thirty-seven percent of cases were classified as confirmed, almost exclusively by polymerase chain reaction. The reported case fatality rate was 0.3% and the reported hospitalization rate was 31%. IRs, hospitalization rates, life-threatening complications, and case fatality rates increased with age group. The IR increased from 2008 to 2012 and the geographic range of reported cases of anaplasmosis appears to have increased since 2000–2007. Our findings are consistent with previous case series and recent reports of the expanding range of the tick vector I. scapularis. PMID:25870428

  3. Granulocytic Myeloid-Derived Suppressor Cells Accumulate in Human Placenta and Polarize toward a Th2 Phenotype.

    PubMed

    Köstlin, Natascha; Hofstädter, Kathrin; Ostermeir, Anna-Lena; Spring, Bärbel; Leiber, Anja; Haen, Susanne; Abele, Harald; Bauer, Peter; Pollheimer, Jürgen; Hartl, Dominik; Poets, Christian F; Gille, Christian

    2016-02-01

    Tolerance induction toward the semiallogeneic fetus is crucial to enable a successful pregnancy; its failure is associated with abortion or preterm delivery. Skewing T cell differentiation toward a Th2-dominated phenotype seems to be pivotal in maternal immune adaption, yet underlying mechanisms are incompletely understood. Myeloid-derived suppressor cells (MDSCs) are innate immune cells that mediate T cell suppression and are increased in cord blood of healthy newborns and in peripheral blood of pregnant women. In this study, we demonstrate that granulocytic MDSCs (GR-MDSCs) accumulate in human placenta of healthy pregnancies but are diminished in patients with spontaneous abortions. Placental GR-MDSCs effectively suppressed T cell responses by expression of arginase I and production of reactive oxygen species and were activated at the maternal-fetal interface through interaction with trophoblast cells. Furthermore, GR-MDSCs isolated from placenta polarized CD4(+) T cells toward a Th2 cytokine response. These results highlight a potential role of GR-MDSCs in inducing and maintaining maternal-fetal tolerance and suggest them as a promising target for therapeutic manipulation of pregnancy complications. PMID:26712947

  4. The effects of vitamin D binding protein-macrophage activating factor and colony-stimulating factor-1 on hematopoietic cells in normal and osteopetrotic rats.

    PubMed

    Benis, K A; Schneider, G B

    1996-10-15

    Osteopetrosis is a heterogeneous group of bone disorders characterized by the failure of osteoclasts to resorb bone and by several immunological defects including macrophage dysfunction. Two compounds, colony-stimulating factor-1 (CSF-1) and vitamin D-binding protein-macrophage activating factor (DBP-MAF) were used in the present study to evaluate their effects on the peritoneal population of cells and on cells within the bone marrow microenvironment in normal and incisors absent (ia) osteopetrotic rats. Previous studies in this laboratory have demonstrated that administration of DBP-MAF to newborn ia animals results in a substantial increase in bone marrow cavity size due to upregulated osteoclast function. To study the effects of these compounds on the macrophage/osteoclast precursors, DBP-MAF, CSF-1, and the combination of these compounds were given to newborn ia and normal littermate animals. Both the normal and mutant phenotypes responded similarly when treated with these compounds. Rats exhibited a profound shift toward the macrophage lineage from the neutrophil lineage when compared with vehicle-treated control animals after treatment with these compounds. In the in vivo peritoneal lavage study, animals received injections of CSF-1, DBP-MAF or DBP-MAF/CSF-1 over a 4-week period. The various types of cells in the peritoneal cavity were then enumerated. The in vitro study consisted of cells isolated from the bone marrow microenvironment and cultured on feeder layers of CSF-1, DBP-MAF, or DBP-MAF/CSF-1 for colony enumeration. The increase in macrophage numbers at the expense of neutrophil numbers could be seen in both the in vivo and in vitro experiments. The macrophage/osteoclast and neutrophil lineages have a common precursor, the granulocyte/macrophage colony-forming cell (GM-CFC). With the addition of CSF-1, the GM-CFC precursor may be induced into the macrophage/osteoclast lineage rather than the granulocyte lineage. This increased pool of cells in the

  5. Origin and Role of a Subset of Tumor-Associated Neutrophils with Antigen-Presenting Cell Features in Early-Stage Human Lung Cancer.

    PubMed

    Singhal, Sunil; Bhojnagarwala, Pratik S; O'Brien, Shaun; Moon, Edmund K; Garfall, Alfred L; Rao, Abhishek S; Quatromoni, Jon G; Stephen, Tom Li; Litzky, Leslie; Deshpande, Charuhas; Feldman, Michael D; Hancock, Wayne W; Conejo-Garcia, Jose R; Albelda, Steven M; Eruslanov, Evgeniy B

    2016-07-11

    Based on studies in mouse tumor models, granulocytes appear to play a tumor-promoting role. However, there are limited data about the phenotype and function of tumor-associated neutrophils (TANs) in humans. Here, we identify a subset of TANs that exhibited characteristics of both neutrophils and antigen-presenting cells (APCs) in early-stage human lung cancer. These APC-like "hybrid neutrophils," which originate from CD11b(+)CD15(hi)CD10(-)CD16(low) immature progenitors, are able to cross-present antigens, as well as trigger and augment anti-tumor T cell responses. Interferon-γ and granulocyte-macrophage colony-stimulating factor are requisite factors in the tumor that, working through the Ikaros transcription factor, synergistically exert their APC-promoting effects on the progenitors. Overall, these data demonstrate the existence of a specialized TAN subset with anti-tumor capabilities in human cancer.

  6. Macrophage colony-stimulating factor receptor c-fms is a novel target of imatinib.

    PubMed

    Dewar, Andrea L; Cambareri, Antony C; Zannettino, Andrew C W; Miller, Bernadette L; Doherty, Kathleen V; Hughes, Timothy P; Lyons, A Bruce

    2005-04-15

    Imatinib is a tyrosine kinase inhibitor that suppresses the growth of bcr-abl-expressing chronic myeloid leukemia (CML) progenitor cells by blockade of the adenosine triphosphate (ATP)-binding site of the kinase domain of bcr-abl. Imatinib also inhibits the c-abl, platelet-derived growth factor (PDGF) receptor, abl-related gene (ARG) and stem-cell factor (SCF) receptor tyrosine kinases, and has been used clinically to inhibit the growth of malignant cells in patients with CML and gastrointestinal stromal tumors (GISTs). Although initially considered to have minimal effects of normal hematopoiesis, recent studies show that imatinib also inhibits the growth of some nonmalignant hematopoietic cells, including monocyte/macrophages. This inhibition could not be attributed to the known activity profile of imatinib. Here, we demonstrate for the first time that imatinib targets the macrophage colony-stimulating factor (M-CSF) receptor c-fms. Phosphorylation of c-fms was inhibited by therapeutic concentrations of imatinib, and this was not due to down-regulation in c-fms expression. Imatinib was also found to inhibit M-CSF-induced proliferation of a cytokine-dependent cell line, further supporting the hypothesis that imatinib affects the growth and development of monocyte and/or macrophages through inhibition of c-fms signaling. Importantly, these results identify an additional biologic target to those already defined for imatinib. Imatinib should now be assessed for activity in diseases where c-fms activation is implicated, including breast and ovarian cancer and inflammatory conditions.

  7. Biolistic expression of the macrophage colony stimulating factor receptor in organotypic cultures induces an inflammatory response.

    PubMed

    Mitrasinovic, Olivera M; Robinson, Christopher C; Tenen, Daniel G; Lee, Yuen Ling; Poon, Clara; Murphy, Greer M

    2004-08-01

    The receptor for macrophage colony-stimulating factor (M-CSFR; c-fms) is expressed at increased levels by microglia in Alzheimer's disease (AD) and in mouse models for AD. Increased expression of M-CSFR on cultured microglia results in a strong proinflammatory response, but the relevance of this cell culture finding to intact brain is unknown. To determine the effects of increased microglial expression of M-CSFR in a complex organotypic environment, we developed a system for biolistic transfection of microglia in hippocampal slice cultures. The promoter for the Mac-1 integrin alpha subunit CD11b is active in cells of myeloid origin. In the brain, CD11b expression is restricted to microglia. Constructs consisting of the promoter for CD11b and a c-fms cDNA or an enhanced green fluorescent protein (EGFP) cDNA were introduced into monotypic cultures of microglia, neurons, and astrocytes. Strong CD11b promoter activity was observed in microglia, whereas little activity was observed in other cell types. Biolistic transfection of organotypic hippocampal cultures with the CD11b/c-fms construct resulted in expression of the c-fms mRNA and protein that was localized to microglia. Furthermore, biolistic overexpression of M-CSFR on microglia resulted in significantly increased production by the hippocampal cultures of the proinflammatory cytokines interleukin (IL)-1alpha macrophage inflammatory protein (MIP-1alpha), and trends toward increased production of IL-6 and M-CSF. These findings demonstrate that microglial overexpression of M-CSFR in an organotypic environment induces an inflammatory response, and suggest that increased microglial expression of M-CSFR could contribute to the inflammatory response observed in AD brain.

  8. Mechanisms of amphibian macrophage development: characterization of the Xenopus laevis colony-stimulating factor-1 receptor.

    PubMed

    Grayfer, Leon; Edholm, Eva-Stina; Robert, Jacques

    2014-01-01

    Macrophage-lineage cells are indispensable to vertebrate homeostasis and immunity. In turn, macrophage development is largely regulated through colony-stimulating factor-1 (CSF1) binding to its cognate receptor (CSF1R). To study amphibian monopoiesis, we identified and characterized the X. laevis CSF1R cDNA transcript. Quantitative analysis revealed that CSF1R tissue gene expression increased with X. laevis development, with greatest transcript levels detected in the adult lung, spleen and liver tissues. Notably, considerable levels of CSF1R mRNA were also detected in the regressing tails of metamorphosing animals, suggesting macrophage involvement in this process, and in the adult bone marrow; corroborating the roles for this organ in Xenopus monopoiesis. Following animal infections with the ranavirus Frog Virus 3 (FV3), both tadpole and adult X. laevis exhibited increased kidney CSF1R gene expression. Conversely, while FV3-infected tadpoles increased their spleen and liver CSF1R mRNA levels, the FV3-challenged adults did not. Notably, FV3 induced elevated bone marrow CSF1R expression, and while stimulation of tadpoles with heat-killed E. coli had no transcriptional effects, bacterial stimulation of adult frogs resulted in significantly increased spleen, liver and bone marrow CSF1R expression. We produced the X. laevis CSF1R in recombinant form (rXlCSF1R) and determined, via in vitro cross-linking studies, that two molecules of rXlCSF1R bound the dimeric rXlCSF1. Finally, administration of rXlCSF1R abrogated the rXlCSF1-induced tadpole macrophage recruitment and differentiation as well as bacterial and FV3-elicited peritoneal leukocyte accumulation. This work marks a step towards garnering greater understanding of the unique mechanisms governing amphibian macrophage biology.

  9. Macrophage colony-stimulating factor induces prolactin expression in rat pituitary gland.

    PubMed

    Hoshino, Satoya; Kurotani, Reiko; Miyano, Yuki; Sakahara, Satoshi; Koike, Kanako; Maruyama, Minoru; Ishikawa, Fumio; Sakatai, Ichiro; Abe, Hiroyuki; Sakai, Takafumi

    2014-06-01

    We investigated the role of macrophage colony-stimulating factor (M-CSF) in the pituitary gland to understand the effect of M-CSF on pituitary hormones and the relationship between the endocrine and immune systems. When we attempted to establish pituitary cell lines from a thyrotropic pituitary tumor (TtT), a macrophage cell line, TtT/M-87, was established. We evaluated M-CSF-like activity in conditioned media (CM) from seven pituitary cell lines using TtT/M-87 cells. TtT/M-87 proliferation significantly increased in the presence of CM from TtT/GF cells, a pituitary folliculostellate (FS) cell line. M-CSF mRNA was detected in TtT/GF and MtT/E cells by reverse transcriptase-polymerase chain reaction (RT-PCR), and its expression in TtT/GF cells was increased in a lipopolysaccharide (LPS) dose-dependent manner. M-CSF mRNA expression was also increased in rat anterior pituitary glands by LPS. M-CSF receptor (M-CSFR) mRNA was only detected in TtT/ M-87 cells and increased in the LPS-stimulated rat pituitary glands. In rat pituitary glands, M-CSF and M-CSFR were found to be localized in FS cells and prolactin (PRL)-secreting cells, respectively, by immunohistochemistry. The PRL concentration in rat sera was significantly increased at 24 h after M-CSF administration, and mRNA levels significantly increased in primary culture cells of rat anterior pituitary glands. In addition, TNF-α mRNA was increased in the primary culture cells by M-CSF. These results revealed that M-CSF was secreted from FS cells and M-CSF regulated PRL expression in rat pituitary glands.

  10. Interferon-gamma enhances megakaryocyte colony-stimulating activity in murine bone marrow cells.

    PubMed

    Tsuji-Takayama, K; Tahata, H; Harashima, A; Nishida, Y; Izumi, N; Fukuda, S; Ohta, T; Kurimoto, M

    1996-09-01

    We have demonstrated previously that interferon-gamma (IFN-gamma) accelerates platelet recovery in mice with 5-FU induced-marrow aplasia in vivo. However, the mechanism for the regulation of megakaryocyte development induced by IFN-gamma in bone marrow cells in vivo remains unknown. To further study the effects of IFN-gamma on megakaryocyte development, various steps during IFN-gamma-mediated accelerated differentiation of the megakaryocytes were investigated in serum-free cultures of murine bone marrow cells in vitro. IFN-gamma markedly induced acetylcholine esterase (AChE) activity, a marker of murine megakaryocytic cells, accompanied by increased colony formation of the megakaryocyte lineage. A prominent increase in megakaryocyte number was observed after IFN-gamma treatment. All of these effects were dependent on the presence of IL-3, and, therefore, these results suggest that IFN-gamma acts as a megakaryocyte potentiator (Meg-POT). However, IFN-gamma did not enhance megakaryocyte maturation with respect to increase in cell size. The effects of IFN-gamma on megakaryocyte maturation were similar to those observed after treatment with higher doses of IL-3 alone. Meg-POT is defined as a factor that induces megakaryocyte maturation. Since IFN-gamma enhanced IL-3-dependent megakaryocyte colony formation and proliferation rather than megakaryocyte maturation, the effects on megakaryocyte development, which were induced by IFN-gamma treatment, seem to be different from the effects of a Meg-POT. We, therefore, propose a new function for IFN-gamma as an enhancer of megakaryocyte colony-stimulating factor activity. The effect of IFN-gamma in vitro appears to correlate well with the acceleration of platelet recovery in vivo.

  11. Colony-stimulating Factor-1 Receptor Utilizes Multiple Signaling Pathways to Induce Cyclin D2 Expression

    PubMed Central

    Dey, Arunangsu; She, Hongyun; Kim, Leopold; Boruch, Allan; Guris, Deborah L.; Carlberg, Kristen; Sebti, Saïd M.; Woodley, David T.; Imamoto, Akira; Li, Wei

    2000-01-01

    Colony-stimulating factor-1 (CSF-1) induces expression of immediate early gene, such as c-myc and c-fos and delayed early genes such as D-type cyclins (D1 and D2), whose products play essential roles in the G1 to S phase transition of the cell cycle. Little is known, however, about the cytoplasmic signal transduction pathways that connect the surface CSF-1 receptor to these genes in the nucleus. We have investigated the signaling mechanism of CSF-1-induced D2 expression. Analyses of CSF-1 receptor autophosphorylation mutants show that, although certain individual mutation has a partial inhibitory effect, only multiple combined mutations completely block induction of D2 in response to CSF-1. We report that at least three parallel pathways, the Src pathway, the MAPK/ERK kinase (MEK)/extracellular signal-regulated kinase (ERK) pathway, and the c-myc pathway, are involved. Induction of D2 is partially inhibited in Src−/− bone marrow-derived macrophages and by Src inhibitor PP1 and is enhanced in v-Src-overexpressing cells. Activation of myc's transactivating activity selectively induces D2 but not D1. Blockade of c-myc expression partially blocks CSF-1-induced D2 expression. Complete inhibition of the MEK/ERK pathway causes 50% decrease of D2 expression. Finally, simultaneous inhibition of Src, MEK activation, and c-myc expression additively blocks CSF-1-induced D2 expression. This study indicates that multiple signaling pathways are involved in full induction of a single gene, and this finding may also apply broadly to other growth factor-inducible genes. PMID:11071910

  12. Regulation of cytosolic phospholipase A2 phosphorylation and eicosanoid production by colony-stimulating factor 1.

    PubMed

    Xu, X X; Rock, C O; Qiu, Z H; Leslie, C C; Jackowski, S

    1994-12-16

    A colony-stimulating factor 1 (CSF-1)-dependent murine macrophage cell line (BAC1.2F5) and peritoneal macrophages were used to investigate the relationship between growth factor-dependent phosphorylation/activation of the 85-kDa cytosolic phospholipase A2 (cPLA2) and arachidonic acid metabolism. The addition of CSF-1 to quiescent BAC1.2F5 cells was followed by the rapid phosphorylation, electrophoretic gel retardation, and stable increase in the specific activity of cPLA2 that correlated with the activation of ERK kinases. cPLA2 phosphorylation depended on the presence of growth factor and persisted throughout the cell cycle. CSF-1 inhibited prostaglandin E2 production and did not enhance arachidonic acid release or increase the levels of lysophosphatidylcholine or glycerophosphocholine. Treatment of BAC1.2F5 cells with the calcium ionophore A23187 plus CSF-1 did not stimulate eicosanoid release. Instead, CSF-1 enhanced the rate of exogenous arachidonic acid incorporation into phosphatidylcholine and its subsequent transfer to phosphatidylethanolamine suggesting that higher rates of arachidonic acid acylation may contribute to the suppression of prostaglandin production. In peritoneal macrophages, ERK kinase activity was stimulated and cPLA2 was phosphorylated and activated in response to CSF-1. However, CSF-1 did not trigger eicosanoid release but did augment arachidonic acid mobilization and prostaglandin E2 production elicited by zymosan and A23187. Thus, cPLA2 phosphorylation/activation and calcium mobilization are not the only determinants for eicosanoid release, and additional components in differentiated tissue macrophages are also required.

  13. Enhancement of human granulopoiesis in vitro by biosynthetic insulin-like growth factor I/somatomedin C and human growth hormone.

    PubMed Central

    Merchav, S; Tatarsky, I; Hochberg, Z

    1988-01-01

    The effect of biosynthetic recombinant insulin-like growth factor I/somatomedin C (IGF-I/Sm-C) and human growth hormone (hGH) on the in vitro growth and maturation of human marrow myeloid progenitors was investigated. Myeloid colony formation was maximally enhanced by 60 ng/ml IGF-I/Sm-C and by 250 ng/ml hGH, resulting in an increase in colony numbers of 41 +/- 7 and 38 +/- 4%, respectively (P less than 0.001). Both peptides induced a 1.5-2.5-fold increase in the frequency of colonies composed of granulocytes alone, but did not alter the numbers of monocyte/macrophage or mixed granulocyte/macrophage colonies. IGF-I/Sm-C and hGH were also found to enhance myeloid maturation towards mature granulocytes in suspension cultures of human marrow cells. The effect of both peptides on human marrow granulopoiesis was similarly demonstrable in serum-free cultures stimulated with human recombinant granulocyte/macrophage colony-stimulating factor. Enhancement of human marrow granulopoiesis in vitro by hGH required the presence of marrow adherent cells and was abrogated by specific monoclonal antibodies directed against IGF-I/Sm-C receptors. The effect of hGH on marrow myeloid progenitors thus appears to be mediated by paracrine IGF-I/Sm-C. PMID:2963830

  14. Infection with agents of human granulocytic ehrlichiosis, lyme disease, and babesiosis in wild white-footed mice (Peromyscus leucopus) in Connecticut.

    PubMed

    Stafford, K C; Massung, R F; Magnarelli, L A; Ijdo, J W; Anderson, J F

    1999-09-01

    White-footed mice, Peromyscus leucopus, were captured in southern Connecticut during 1997 and 1998 to determine the prevalence of infections caused by granulocytic Ehrlichia sp., Borrelia burgdorferi, and Babesia microti. Of the 50 mice captured and recaptured, 25 of 47 (53.2%) and 23 of 48 (47.9%) contained antibodies to the BDS or NCH-1 Ehrlichia strains, respectively, as determined by indirect fluorescent antibody (IFA) staining methods. The majority (83.3%) of 48 mice also contained antibodies to B. burgdorferi, as determined by enzyme-linked immunosorbent assay. Moreover, 20 of 26 (76.9%) contained antibodies to B. microti by IFA staining methods. In nested PCR tests using the 16S rRNA gene, the DNA of the human granulocytic ehrlichiosis (HGE) agent was detected in 17 of 47 mice (36.2%), but only 4 (23.5%) of these 17 mice were PCR positive at each capture. Antibody-positive reactions to granulocytic Ehrlichia sp. organisms were detected in 17 of 23 (73. 9%) of the PCR-positive mice. The sequences from PCR products from nine positive blood samples were identical to the HGE agent. Ehrlichia spp. were cultured from three of five mice captured in April 1998, including one that was PCR positive in April 1997. In addition, 2 of 14 larval Ixodes scapularis pools, which were attached to two PCR-positive mice, contained DNA of the HGE agent. A high percentage of white-footed mice are infected or have been infected naturally by the HGE agent with low-level persistent infection or frequent reinfection in some individual mice. However, the changes noted in the presence of DNA and antibodies in repeated blood and serum samples from individual mice over several months of field collection suggests that infection with granulocytic Ehrlichia is transient in most wild P. leucopus.

  15. Association between colony-stimulating factor 1 receptor gene polymorphisms and asthma risk.

    PubMed

    Shin, Eun Kyong; Lee, Shin-Hwa; Cho, Sung-Hwan; Jung, Seok; Yoon, Sang Hyuk; Park, Sung Woo; Park, Jong Sook; Uh, Soo Taek; Kim, Yang Ki; Kim, Yong Hoon; Choi, Jae-Sung; Park, Byung-Lae; Shin, Hyoung Doo; Park, Choon-Sik

    2010-09-01

    Colony-stimulating factor 1 receptor (CSF1R) is expressed in monocytes/macrophages and dendritic cells. These cells play important roles in the innate immune response, which is regarded as an important aspect of asthma development. Genetic alterations in the CSF1R gene may contribute to the development of asthma. We investigated whether CSF1R gene polymorphisms were associated with the risk of asthma. Through direct DNA sequencing of the CSF1R gene, we identified 28 single nucleotide polymorphisms (SNPs) and genotyped them in 303 normal controls and 498 asthmatic patients. Expression of CSF1R protein and mRNA were measured on CD14-positive monocytes and neutrophils in peripheral blood of asthmatic patients using flow cytometry and real-time PCR. Among the 28 polymorphisms, two intronic polymorphism (+20511C>T and +22693T>C) were associated with the risk of asthma by logistic regression analysis. The frequencies of the minor allele at CSF1R +20511C>T and +22693T>C were higher in asthmatic subjects than in normal controls (4.6 vs. 7.7%, p = 0.001 in co-dominant and dominant models; 16.4 vs. 25.8%, p = 0.0006 in a recessive model). CSF1R mRNA levels in neutrophils of the asthmatic patients having the +22693CC allele were higher than in those having the +22693TT allele (p = 0.026). Asthmatic patients with the +22693CC allele also showed significantly higher CSF1R expression on CD14-positive monocytes and neutrophils than did those with the +22693TT allele (p = 0.045 and p = 0.044). The +20511C>T SNP had no association with CSF1R mRNA or protein expression. In conclusion, the minor allele at CSF1R +22693T>C may have a susceptibility effect in the development of asthma, via increased CSF1R protein and mRNA expression in inflammatory cells.

  16. Anaerobiosis increases resistance of Neisseria gonorrhoeae to O2-independent antimicrobial proteins from human polymorphonuclear granulocytes.

    PubMed Central

    Casey, S G; Shafer, W M; Spitznagel, J K

    1985-01-01

    We investigated the in vitro resistance of Neisseria gonorrhoeae FA19 to the O2-independent antimicrobial systems of human polymorphonuclear leukocytes. Acid extracts of polymorphonuclear leukocyte granules (crude granule extracts) and a purified granule protein (57 kilodaltons) were, at low concentrations, bactericidal for gonococci under aerobic conditions that permitted growth. However, they were less effective under anaerobic conditions that imposed bacteriostasis. We found that adding sodium nitrite to reduced growth media permitted the growth of strain FA19 in an anaerobic environment. Under these conditions with nitrite, anaerobic cultures of strain FA19 were no more resistant to the crude granule extract and the 57-kilodalton protein than aerobic cultures. In contrast, Salmonella typhimurium SL-1004, a facultative anaerobe, was readily killed by both the crude granule extract and the 57-kilodalton antimicrobial protein regardless of the presence or absence of free molecular oxygen. This is the first demonstration that an isolated antimicrobial protein from polymorphonuclear leukocyte granules is active against bacteria under anaerobic conditions. Our results also indicated that the efficacy of human polymorphonuclear leukocyte O2-independent killing of N. gonorrhoeae may, in part, be inhibited by bacteriostatic conditions imposed by hypoxia. Images PMID:3917976

  17. Functionally and morphologically distinct populations of extracellular vesicles produced by human neutrophilic granulocytes.

    PubMed

    Lőrincz, Ákos M; Schütte, Maria; Timár, Csaba I; Veres, Daniel S; Kittel, Ágnes; McLeish, Kenneth R; Merchant, Michael L; Ligeti, Erzsébet

    2015-10-01

    EVs in the microvesicle size range released during spontaneous death of human neutrophils were characterized and their properties compared with previously described EVs with antibacterial effect (aEVs, generated on specific activation) or produced spontaneously (sEVs). The 3 vesicle populations overlapped in size and in part of the constituent proteins were stained with annexin V and were impermeable to PI. However, none of them produced superoxide. In contrast, remarkable differences were observed in the morphology, abundance of proteins, and antibacterial function. EVs formed spontaneously in 30 min (sEVs) were more similar to EVs released during spontaneous death in 1-3 d than to EVs formed in 30 min on stimulation of opsonin receptors (aEVs). Spontaneously generated EVs had no antibacterial effect despite their large number and protein content. We hypothesized 2 parallel mechanisms: one that proceeds spontaneously and produces EVs without antibacterial effect and another process that is triggered by opsonin receptors and results in differential sorting of proteins into EVs with antibacterial capacity. Our results call attention to the functional and morphologic heterogeneity within the microvesicle/ectosome fraction of EVs.

  18. Effects of gadolinium oxide nanoparticles on the oxidative burst from human neutrophil granulocytes.

    PubMed

    Abrikossova, Natalia; Skoglund, Caroline; Ahrén, Maria; Bengtsson, Torbjörn; Uvdal, Kajsa

    2012-07-11

    We have previously shown that gadolinium oxide (Gd(2)O(3)) nanoparticles are promising candidates to be used as contrast agents in magnetic resonance (MR) imaging applications. In this study, these nanoparticles were investigated in a cellular system, as possible probes for visualization and targeting intended for bioimaging applications. We evaluated the impact of the presence of Gd(2)O(3) nanoparticles on the production of reactive oxygen species (ROS) from human neutrophils, by means of luminol-dependent chemiluminescence. Three sets of Gd(2)O(3) nanoparticles were studied, i.e. as synthesized, dialyzed and both PEG-functionalized and dialyzed Gd(2)O(3) nanoparticles. In addition, neutrophil morphology was evaluated by fluorescent staining of the actin cytoskeleton and fluorescence microscopy. We show that surface modification of these nanoparticles with polyethylene glycol (PEG) is essential in order to increase their biocompatibility. We observed that the as synthesized nanoparticles markedly decreased the ROS production from neutrophils challenged with prey (opsonized yeast particles) compared to controls without nanoparticles. After functionalization and dialysis, more moderate inhibitory effects were observed at a corresponding concentration of gadolinium. At lower gadolinium concentration the response was similar to that of the control cells. We suggest that the diethylene glycol (DEG) present in the as synthesized nanoparticle preparation is responsible for the inhibitory effects on the neutrophil oxidative burst. Indeed, in the present study we also show that even a low concentration of DEG, 0.3%, severely inhibits neutrophil function. In summary, the low cellular response upon PEG-functionalized Gd(2)O(3) nanoparticle exposure indicates that these nanoparticles are promising candidates for MR-imaging purposes.

  19. Effects of gadolinium oxide nanoparticles on the oxidative burst from human neutrophil granulocytes

    NASA Astrophysics Data System (ADS)

    Abrikossova, Natalia; Skoglund, Caroline; Ahrén, Maria; Bengtsson, Torbjörn; Uvdal, Kajsa

    2012-07-01

    We have previously shown that gadolinium oxide (Gd2O3) nanoparticles are promising candidates to be used as contrast agents in magnetic resonance (MR) imaging applications. In this study, these nanoparticles were investigated in a cellular system, as possible probes for visualization and targeting intended for bioimaging applications. We evaluated the impact of the presence of Gd2O3 nanoparticles on the production of reactive oxygen species (ROS) from human neutrophils, by means of luminol-dependent chemiluminescence. Three sets of Gd2O3 nanoparticles were studied, i.e. as synthesized, dialyzed and both PEG-functionalized and dialyzed Gd2O3 nanoparticles. In addition, neutrophil morphology was evaluated by fluorescent staining of the actin cytoskeleton and fluorescence microscopy. We show that surface modification of these nanoparticles with polyethylene glycol (PEG) is essential in order to increase their biocompatibility. We observed that the as synthesized nanoparticles markedly decreased the ROS production from neutrophils challenged with prey (opsonized yeast particles) compared to controls without nanoparticles. After functionalization and dialysis, more moderate inhibitory effects were observed at a corresponding concentration of gadolinium. At lower gadolinium concentration the response was similar to that of the control cells. We suggest that the diethylene glycol (DEG) present in the as synthesized nanoparticle preparation is responsible for the inhibitory effects on the neutrophil oxidative burst. Indeed, in the present study we also show that even a low concentration of DEG, 0.3%, severely inhibits neutrophil function. In summary, the low cellular response upon PEG-functionalized Gd2O3 nanoparticle exposure indicates that these nanoparticles are promising candidates for MR-imaging purposes.

  20. Granulocyte CEACAM3 is a phagocytic receptor of the innate immune system that mediates recognition and elimination of human-specific pathogens.

    PubMed

    Schmitter, Tim; Agerer, Franziska; Peterson, Lisa; Munzner, Petra; Hauck, Christof R

    2004-01-01

    Carcinoembryonic antigen-related cell adhesion molecules (CEACAMs) are used by several human pathogens to anchor themselves to or invade host cells. Interestingly, human granulocytes express a specific isoform, CEACAM3, that participates together with CEACAM1 and CEACAM6 in the recognition of CEACAM-binding microorganisms. Here we show that CEACAM3 can direct efficient, opsonin-independent phagocytosis of CEACAM-binding Neisseria, Moraxella, and Haemophilus species. CEACAM3- but not CEACAM6-mediated uptake is blocked by dominant-negative versions of the small GTPase Rac. Moreover, CEACAM3 engagement triggers membrane recruitment and increased GTP loading of Rac that are not observed upon bacterial binding to CEACAM6. Internalization and Rac stimulation are also inhibited by compromising the integrity of an immunoreceptor tyrosine-based activation motif (ITAM)-like sequence in the cytoplasmic tail of CEACAM3 or by interference with Src family protein tyrosine kinases that phosphorylate CEACAM3. In contrast to interfering with CEACAM6, blockage of CEACAM3-mediated events reduces the ability of primary human granulocytes to internalize and eliminate CEACAM-binding bacteria, indicating an important role of CEACAM3 in the control of human-specific pathogens by the innate immune system.

  1. Cloning, bacterial expression and biological characterization of recombinant human granulocyte chemotactic protein-2 and differential expression of granulocyte chemotactic protein-2 and epithelial cell-derived neutrophil activating peptide-78 mRNAs.

    PubMed

    Froyen, G; Proost, P; Ronsse, I; Mitera, T; Haelens, A; Wuyts, A; Opdenakker, G; Van Damme, J; Billiau, A

    1997-02-01

    Human osteosarcoma cells secrete a novel C-X-C chemokine called granulocyte chemotactic protein-2 (GCP-2), which was previously identified by amino acid sequencing of the purified natural protein. In order to understand the role of this new protein in inflammatory reactions, we cloned GCP-2 DNA sequences to generate recombinant protein and specific DNA probes and primers. By means of PCR on cloned cDNA of osteosarcoma cells induced by interleukin-1 beta and fibroblasts induced by lipopolysaccharide plus dsRNA, the complete coding domain of GCP-2 was isolated. This sequence was cloned into the bacterial expression vector pHEN1 and, after induction, GCP-2 was secreted into the periplasm of Escherichia coli. Recombinant GCP-2 (rGCP-2) was purified and characterized by SDS/PAGE as a monomeric 6.5-kDa protein and by amino-terminal sequencing. The chemoattractive potency of GCP-2 for neutrophilic granulocytes was about 10-times less than that of interleukin-8 and the minimal effective dose was 10 ng/ml. However, at optimal dose (100 ng/ml) the maximal chemotactic response was comparable with that of interleukin-8. Both characteristics correspond with those of natural GCP-2. In addition, intracellular calcium release in neutrophils by recombinant GCP-2 was achieved with as little as 10 ng/ml. Quantitation studies using reverse transcriptase and the polymerase chain reaction revealed higher GCP-2 mRNA production in normal fibroblasts than in tumor cells. When compared with epithelial-cell-derived neutrophil-activating peptide-78 (ENA-78) mRNA, the GCP-2 mRNA levels were higher in all cell lines tested. In addition, GCP-2 and ENA-78 expression seem to be differentially regulated in that phorbol ester and lipopolysaccharide have opposing effects on their mRNA induction in diploid fibroblasts and epithelial cells, respectively. Interleukin-1 was demonstrated to be a general inducer for both chemokines, while interferon-gamma down-regulates their mRNA expression. The

  2. HCMV infection of humanized mice after transplantation of G-CSF-mobilized peripheral blood stem cells from HCMV-seropositive donors.

    PubMed

    Hakki, Morgan; Goldman, Devorah C; Streblow, Daniel N; Hamlin, Kimberly L; Krekylwich, Craig N; Fleming, William H; Nelson, Jay A

    2014-01-01

    Human cytomegalovirus (HCMV) infection, including primary infection resulting from transmission from a seropositive donor to a seronegative recipient (D(+)/R(-)), remains a significant problem in the setting of peripheral blood stem cell transplantation (PBSCT). The lack of a suitable animal model for studying HCMV transmission after PBSCT is a major barrier to understanding this process and, consequently, developing novel interventions to prevent HCMV infection. Our previous work demonstrated that human CD34(+) progenitor cell-engrafted NOD-scid IL2Rγc(null) (NSG) mice support latent HCMV infection after direct inoculation and reactivation after treatment with granulocyte colony-stimulating factor. To more accurately recapitulate HCMV infection in the D(+)/R(-) PBSCT setting, granulocyte colony-stimulating factor-mobilized peripheral blood stem cells from seropositive donors were used to engraft NSG mice. All recipient mice demonstrated evidence of HCMV infection in liver, spleen, and bone marrow. These findings validate the NSG mouse model for studying HCMV transmission during PBSCT.

  3. An Epstein-Barr Virus Encoded Inhibitor of Colony Stimulating Factor-1 Signaling Is an Important Determinant for Acute and Persistent EBV Infection

    PubMed Central

    Ohashi, Makoto; Fogg, Mark H.; Orlova, Nina; Quink, Carol; Wang, Fred

    2012-01-01

    Acute Epstein-Barr virus (EBV) infection is the most common cause of Infectious Mononucleosis. Nearly all adult humans harbor life-long, persistent EBV infection which can lead to development of cancers including Hodgkin Lymphoma, Burkitt Lymphoma, nasopharyngeal carcinoma, gastric carcinoma, and lymphomas in immunosuppressed patients. BARF1 is an EBV replication-associated, secreted protein that blocks Colony Stimulating Factor 1 (CSF-1) signaling, an innate immunity pathway not targeted by any other virus species. To evaluate effects of BARF1 in acute and persistent infection, we mutated the BARF1 homologue in the EBV-related herpesvirus, or lymphocryptovirus (LCV), naturally infecting rhesus macaques to create a recombinant rhLCV incapable of blocking CSF-1 (ΔrhBARF1). Rhesus macaques orally challenged with ΔrhBARF1 had decreased viral load indicating that CSF-1 is important for acute virus infection. Surprisingly, ΔrhBARF1 was also associated with dramatically lower virus setpoints during persistent infection. Normal acute viral load and normal viral setpoints during persistent rhLCV infection could be restored by Simian/Human Immunodeficiency Virus-induced immunosuppression prior to oral inoculation with ΔrhBARF1 or infection of immunocompetent animals with a recombinant rhLCV where the rhBARF1 was repaired. These results indicate that BARF1 blockade of CSF-1 signaling is an important immune evasion strategy for efficient acute EBV infection and a significant determinant for virus setpoint during persistent EBV infection. PMID:23300447

  4. High-dose paclitaxel with granulocyte colony-stimulating factor in patients with advanced breast cancer refractory to anthracycline therapy: a European Cancer Center trial.

    PubMed

    Vermorken, J B; ten Bokkel Huinink, W W; Mandjes, I A; Postma, T J; Huizing, M T; Heimans, J J; Beijnen, J H; Bierhorst, F; Winograd, B; Pinedo, H M

    1995-08-01

    Paclitaxel (Taxol; Bristol-Myers Squibb Company, Princeton, NJ) is a novel cytostatic agent that has shown interesting antitumor activity in patients with advanced breast cancer. Depending on variable patient characteristics and amount and type of prior therapy, as well as the applied dose and schedule of paclitaxel, response rates have varied from 13% to 62%. However, optimal dose and schedule are still unknown. We studied a high-dose (250 to 300 mg/m2) 3-hour paclitaxel infusion schedule in a poor prognostic group of breast cancer patients who progressed or relapsed while taking anthracyclines. This regimen was given every 3 weeks. Twenty-one of the 36 patients studied had increased liver enzymes and 18 had documented liver metastases. The objective response rate was only 6%, but response rate by disease site indicated that soft tissue lesions responded in 30% of cases. For a better comparison with other reported data a uniform definition of "anthracycline refractory" is needed. Neuropathy, which was found to be dose limiting, and arthralgia/myalgia syndrome were the most frequently occurring toxicities. Both severe myelosuppression (and infections) and severe diarrhea and mucositis were reported more frequently in patients with liver dysfunction. As higher peak levels, increased areas under the concentration time curves, and longer times during which plasma concentrations were above the threshold level of 0.1 mumol/L were found in patients with elevated liver enzymes, a correlation with the observed toxicities is assumed. Further pharmacodynamic studies in such patients receiving a 3-hour infusion seem warranted.

  5. Recovery of pulmonary structure and exercise capacity by treatment with granulocyte-colony stimulating factor (G-CSF) in a mouse model of emphysema.

    PubMed

    Fortunato, Gustavo; Vidal, Daniel T A; Klein, Wilfried; Neto, Alberto; Angrizani, André; Vasconcelos, Juliana F; Kaneto, Carla; Souza, Bruno Solano de Freitas; Ribeiro-dos-Santos, Ricardo; Soares, Milena B P; Macambira, Simone G

    2014-04-01

    Emphysema is a chronic obstructive pulmonary disease characterized abnormal dilatation of alveolar spaces, which impairs alveolar gas exchange, compromising the physical capacity of a patient due to airflow limitations. Here we tested the effects of G-CSF administration in pulmonary tissue and exercise capacity in emphysematous mice. C57Bl/6 female mice were treated with elastase intratracheally to induce emphysema. Their exercise capacities were evaluated in a treadmill. Lung histological sections were prepared to evaluate mean linear intercept measurement. Emphysematous mice were treated with G-CSF (3 cycles of 200 μg/kg/day for 5 consecutive days, with 7-day intervals) or saline and submitted to a third evaluation 8 weeks after treatment. Values of run distance and linear intercept measurement were expressed as mean ± SD and compared applying a paired t-test. Effects of treatment on these parameters were analyzed applying a Repeated Measures ANOVA, followed by Tukey's post hoc analysis. p < 0.05 was considered statistically significant. Twenty eight days later, animals ran significantly less in a treadmill compared to normal mice (549.7 ± 181.2 m and 821.7 ± 131.3 m, respectively; p < 0.01). Treatment with G-CSF significantly increased the exercise capacity of emphysematous mice (719.6 ± 200.5 m), whereas saline treatment had no effect on distance run (595.8 ± 178.5 m). The PCR cytokines genes analysis did not detect difference between experimental groups. Morphometric analyses in the lung showed that saline-treated mice had a mean linear intercept significantly higher (p < 0.01) when compared to mice treated with G-CSF, which did not significantly differ from that of normal mice. Treatment with G-CSF promoted the recovery of exercise capacity and regeneration of alveolar structural alterations in emphysematous mice.

  6. Cross-talk between ICAM-1 and granulocyte-macrophage colony-stimulating factor receptor signaling modulates eosinophil survival and activation.

    PubMed

    Pazdrak, Konrad; Young, Travis W; Stafford, Susan; Olszewska-Pazdrak, Barbara; Straub, Christof; Starosta, Vitaliy; Brasier, Allan; Kurosky, Alexander

    2008-03-15

    Reversal of eosinophilic inflammation has been an elusive therapeutic goal in the management of asthma pathogenesis. In this regard, GM-CSF is a primary candidate cytokine regulating eosinophil activation and survival in the lung; however, its molecular mechanism of propagation and maintenance of stimulated eosinophil activation is not well understood. In this study, we elucidate those late interactions occurring between the GM-CSF receptor and activated eosinophil signaling molecules. Using coimmunoprecipitation with GM-CSF-stimulated eosinophils, we have identified that the GM-CSF receptor beta-chain (GMRbeta) interacted with ICAM-1 and Shp2 phosphatase, as well as Slp76 and ADAP adaptor proteins. Separate experiments using affinity binding with a tyrosine-phosphorylated peptide containing an ITIM (ICAM-1 residues 480-488) showed binding to Shp2 phosphatase and GMRbeta. However, the interaction of GMRbeta with the phosphorylated ICAM-1-derived peptide was observed only with stimulated eosinophil lysates, suggesting that the interaction of GMRbeta with ICAM-1 required phosphorylated Shp2 and/or phosphorylated GMRbeta. Importantly, we found that inhibition of ICAM-1 in activated eosinophils blocked GM-CSF-induced expression of c-fos, c-myc, IL-8, and TNF-alpha. Moreover, inhibition of ICAM-1 expression with either antisense oligonucleotide or an ICAM-1-blocking Ab effectively inhibited ERK activation and eosinophil survival. We concluded that the interaction between ICAM-1 and the GM-CSF receptor was essential for GM-CSF-induced eosinophil activation and survival. Taken together, these results provide novel mechanistic insights defining the interaction between ICAM-1 and the GM-CSF receptor and highlight the importance of targeting ICAM-1 and GM-CSF/IL-5/IL-3 receptor systems as a therapeutic strategy to counter eosinophilia in asthma.

  7. Chronic ethanol ingestion in rats decreases granulocyte-macrophage colony-stimulating factor receptor expression and downstream signaling in the alveolar macrophage.

    PubMed

    Joshi, Pratibha C; Applewhite, Lisa; Ritzenthaler, Jeffrey D; Roman, Jesse; Fernandez, Alberto L; Eaton, Douglas C; Brown, Lou Ann S; Guidot, David M

    2005-11-15

    Although it is well recognized that alcohol abuse impairs alveolar macrophage immune function and renders patients susceptible to pneumonia, the mechanisms are incompletely understood. Alveolar macrophage maturation and function requires priming by GM-CSF, which is produced and secreted into the alveolar space by the alveolar epithelium. In this study, we determined that although chronic ethanol ingestion (6 wk) in rats had no effect on GM-CSF expression within the alveolar space, it significantly decreased membrane expression of the GM-CSF receptor in alveolar macrophages. In parallel, ethanol ingestion decreased cellular expression and nuclear binding of PU.1, the master transcription factor that activates GM-CSF-dependent macrophage functions. Furthermore, treatment of ethanol-fed rats in vivo with rGM-CSF via the upper airway restored GM-CSF receptor membrane expression as well as PU.1 protein expression and nuclear binding in alveolar macrophages. Importantly, GM-CSF treatment also restored alveolar macrophage function in ethanol-fed rats, as reflected by endotoxin-stimulated release of TNF-alpha and bacterial phagocytosis. We conclude that ethanol ingestion dampens alveolar macrophage immune function by decreasing GM-CSF receptor expression and downstream PU.1 nuclear binding and that these chronic defects can be reversed relatively quickly with rGM-CSF treatment in vivo.

  8. Influence of preharvest tumor cell contamination in bone marrow or blood does not predict resultant tumor cell contamination of granulocyte colony-stimulating factor mobilized stem cells.

    PubMed

    Krüger, W; Kröger, N; Tögel, F; Badbaran, A; Renges, H; Gieseking, F; Gutensohn, K; Jänicke, F; Zander, A R

    2001-04-01

    Tumor cell contamination of stem cell collections harvested from breast cancer patients is a common phenomenon described by several investigators but with findings that vary among reports. Although so-called co-mobilization of these cells has been hypothesized, the origin of tumor cell contamination in stem cells is still unknown. A total of 47 G-CSF mobilized stem cell grafts from patients with nodal-positive (n = 30), chemosensitive metastatic (n = 11), and 5 women with inflammatory breast cancer were evaluated for cancer cells by immunocytochemistry. Additionally, 40 bone marrow aspirations and 23 peripheral blood samples collected prior to apheresis and after one to two cycles of conventional chemotherapy were available for examination. Tumor cell contamination of leukapheresis correlated best with preharvest blood state. This was valid when the nominal (positive/negative) presence of tumor cells in blood was compared to the nominal presence of tumor cells in apheresis samples and when the it was correlated to the tumor cell load of apheresis samples (TCL = tumor cells per 10(6) nucleated cells investigated). The correlation between blood and stem cells was better (nominal and quantitative) than that between marrow and stem cells, despite the larger sample size of marrow aspirations. The presence or absence of cancer cells in apheresis samples could not be safely predicted by the presence or absence of tumor cells in marrow or blood alone. Diagnostic specificity seems to improve from a combination of results from marrow and blood analysis. No correlation was found in quantitative analysis of tumor cell contamination between marrow and blood. In conclusion, the results suggest that blood and bone marrow represent different compartments for epithelial cancer cells and that contaminating tumor cells in stem cell harvests may be derived from the blood and/or marrow compartment. The tumor cell contamination of a stem cell harvest cannot be safely predicted by a preceding blood or marrow analysis.

  9. A role for granulocyte-macrophage colony-stimulating factor in the regulation of CD8{sup +} T cell responses to rabies virus

    SciTech Connect

    Wanjalla, Celestine N.; Goldstein, Elizabeth F.; Wirblich, Christoph; Schnell, Matthias J.

    2012-05-10

    Inflammatory cytokines have a significant role in altering the innate and adaptive arms of immune responses. Here, we analyzed the effect of GM-CSF on a RABV-vaccine vector co-expressing HIV-1 Gag. To this end, we immunized mice with RABV expressing HIV-1 Gag and GM-CSF and analyzed the primary and recall CD8{sup +} T cell responses. We observed a statistically significant increase in antigen presenting cells (APCs) in the spleen and draining lymph nodes in response to GM-CSF. Despite the increase in APCs, the primary and memory anti HIV-1 CD8{sup +} T cell response was significantly lower. This was partly likely due to lower levels of proliferation in the spleen. Animals treated with GM-CSF neutralizing antibodies restored the CD8{sup +} T cell response. These data define a role of GM-CSF expression, in the regulation of the CD8{sup +} T cell immune responses against RABV and has implications in the use of GM-CSF as a molecular adjuvant in vaccine development.

  10. Interferon-γ and granulocyte/monocyte colony-stimulating factor production by natural killer cells involves different signaling pathways and the adaptor stimulator of interferon genes (STING).

    PubMed

    Souza-Fonseca-Guimaraes, Fernando; Parlato, Marianna; de Oliveira, Rosane B; Golenbock, Douglas; Fitzgerald, Katherine; Shalova, Irina N; Biswas, Subhra K; Cavaillon, Jean-Marc; Adib-Conquy, Minou

    2013-04-12

    Natural killer (NK) cells are important for innate immunity in particular through the production of IFN-γ and GM-CSF. Both cytokines are important in restoration of immune function of tolerized leukocytes under inflammatory events. The expression of TLRs in NK cells has been widely studied by analyzing the mRNA of these receptors, rarely seeking their protein expression. We previously showed that murine spleen NK cells express TLR9 intracellularly and respond to CpG oligodeoxynucleotide (CpG-ODN) by producing IFN-γ and GM-CSF. However, to get such production the presence of accessory cytokines (such as IL-15 and IL-18) was required, whereas CpG-ODN or accessory cytokines alone did not induce IFN-γ or GM-CSF. We show here that TLR9 overlaps with the Golgi apparatus in NK cells. Furthermore, CpG-ODN stimulation in the presence of accessory cytokines induces the phosphorylation of c-Jun, STAT3, and IκBα. IFN-γ and GM-CSF production requires NF-κB and STAT3 activation as well as Erk-dependent mechanisms for IFN-γ and p38 signaling for GM-CSF. Using knock-out-mice, we show that UNC93b1 and IL-12 (produced by NK cells themselves) are also necessary for IFN-γ and GM-CSF production. IFN-γ production was found to be MyD88- and TLR9-dependent, whereas GM-CSF was TLR9-independent but dependent on STING (stimulator of interferon genes), a cytosolic adaptor recently described for DNA sensing. Our study thereby allows us to gain insight into the mechanisms of synergy between accessory cytokines and CpG-ODN in NK cells. It also identifies a new and alternative signaling pathway for CpG-ODN in murine NK cells.

  11. Granulocyte Colony Stimulating Factor (GCSF) Alters the Phenotype of Neuroblastoma Cells: Implications for disease free survival of high-risk patients

    PubMed Central

    Gay, Andre N.; Chang, Shirong; Rutland, Lindsey; Yu, Ling; Byeseda, Sarah; Naik-Mathuria, Bindi; Cass, Darrell L.; Russell, Heidi; Olutoye, Oluyinka O.

    2008-01-01

    Introduction GCSF is commonly employed for the treatment of chemotherapy-induced neutropenia. Despite high-dose intensive chemotherapy for advanced stage neuroblastoma, the survival rate remains poor. GCSF therapy is quite common in these children, thus we questioned its effect on neuroblastoma cells. We hypothesized that exogenous GCSF stimulates the proliferation and invasive character of neuroblastoma cells. Methods Expression of a GCSF receptor in five different neuroblastoma cell lines was determined by PCR. In addition, we determined the effect of increasing doses of GCSF (0, 1 ng/ml, 10 ng/ml, 1 µg/ml, 10 µg/ml) on DNA synthesis (BrdU assay), invasiveness (Matrigel invasion chambers) and cell proliferation. Results We tested five neuroblastoma cell lines; all expressed the GCSF receptor. GCSF treatment resulted in significantly increased proliferation of SK-N-SH, SK-N-AS and SHSY-5Y cells. Likewise, increased invasiveness of SK-N-SH cells was observed with GCSF treatment. Conclusions Our results indicate that neuroblastoma cell lines express the GCSF receptor and respond to exogenous GCSF by increased proliferation and invasiveness. These findings suggest that GCSF may stimulate the growth of neuroblastoma cells in patients undergoing high-dose chemotherapy with GCSF rescue and could have a significant impact on the ability to eradicate these tumors. PMID:18485949

  12. History of chronic comorbidity and risk of chemotherapy-induced febrile neutropenia in patients with non-Hodgkin lymphoma not receiving granulocyte colony-stimulating factor prophylaxis.

    PubMed

    Chao, Chun; Rodriguez, Roberto; Page, John H; Yang, Su-Jau; Huynh, Julie; Chia, Victoria M

    2015-01-01

    We conducted a cohort study to examine the association between a wide variety of chronic comorbidities and risk of febrile neutropenia (FN) in patients with non-Hodgkin lymphoma (NHL) from 2000 to 2009 treated with chemotherapy at Kaiser Permanente Southern California. History of comorbidities and FN events were identified using electronic medical records. Cox model adjusting for propensity score was used to determine the association between a comorbid condition and FN. Models that additionally adjusted for cancer stage, baseline absolute neutrophil count, chemotherapy regimen and dose reduction were also evaluated. A total of 2480 patients with NHL were included, and 60% received CHOP/R-CHOP (cyclophosphamide, doxorubicin, vincristine, prednisone, with or without rituximab). In total, 236 (9.5%) patients developed FN in the first chemotherapy cycle. Anemia (adjusted hazard ratio [HR] = 1.6, 95% confidence interval [1.2-2.2]), HIV infection (HR = 3.8 [2.0-6.7]) and rheumatoid diseases (HR = 2.4 [1.3-4.0]) were associated with significantly increased risk of FN. These results provide evidence that chronic comorbidity increases the risk of FN. PMID:24684228

  13. Viral vectored granulocyte-macrophage colony stimulating factor inhibits vaccine protection in an SIV challenge model: protection correlates with neutralizing antibody

    PubMed Central

    Schell, John B.; Bahl, Kapil; Rose, Nina F.; Buonocore, Linda; Hunter, Meredith; Marx, Preston A.; LaBranche, Celia C.; Montefiori, David C.; Rose, John K.

    2012-01-01

    In a previous vaccine study, we reported significant and apparently sterilizing immunity to high-dose, mucosal, simian immunodeficiency virus (SIV) quasispecies challenge (27). The vaccine consisted of vectors based on vesicular stomatitis virus (VSV) expressing simian immunodeficiency virus (SIV) gag and env genes, a boost with propagating replicon particles expressing the same SIV genes, and a second boost with VSV-based vectors. Concurrent with that published study we had a parallel group of macaques given the same doses of vaccine vectors, but in addition, we included a third VSV vector expressing rhesus macaque GM-CSF in the priming immunization only. We report here that addition of the vector expressing GM-CSF did not enhance CD8 T cell or antibody responses to SIV antigens, and almost completely abolished the vaccine protection against high-dose mucosal challenge with SIV. Expression of GM-CSF may have limited vector replication excessively in the macaque model. Our results suggest caution in the use of GM-CSF as a vaccine adjuvant, especially when expressed by a viral vector. Combining vaccine group animals from this study and the previous study we found that there was a marginal but significant positive correlation between the neutralizing antibody to a neutralization resistant SIV Env and protection from infection. PMID:22537983

  14. Case Report. Prevention of Clozapine-Induced Granulocytopenia/Agranulocytosis with Granulocyte-Colony Stimulating Factor (G-CSF) in an Intellectually Disabled Patient with Schizophrenia

    ERIC Educational Resources Information Center

    Rajagopal, G.; Graham, J. G.; Haut, F. F. A.

    2007-01-01

    Background: While clozapine is an effective treatment for refractory schizophrenia, its use is limited by haematological side effects. Treatment options that allow continued prescription of clozapine by tackling these side effects will greatly aid patients for whom this medication is all too often their only hope of recovery. Method: In this case…

  15. Enhancement of innate immunity with granulocyte colony-stimulating factor did not mitigate disease in pigs infected with a highly pathogenic Chinese PRRSV strain

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Porcine reproductive and respiratory syndrome virus (PRRSV) is responsible for one of the most economically important diseases in swine worldwide. It causes reproductive failure in sows and pneumonia in pigs that predisposes them to secondary bacterial infections. Methods to control PRRSV and/or lim...

  16. The role of biosimilar granulocyte colony stimulating factor (GCSF) Zarzio for progenitor cell mobilization and the treatment of therapy-induced neutropenia in adult hematopoietic stem cell transplantation.

    PubMed

    Severson, Cherie C

    2015-01-01

    Originator GCSF (Neupogen) has been used to mobilize progenitor stem cells and treat therapy-induced neutropenia in Canadian stem cell transplant settings for years. Although its benefit is not in question, viable alternatives are available. Biosimilar GCSF (Zarzio) is widely in use in Europe since 2009 and was recently approved in the U.S.for the same five indications as Neupogen. Zarzio is reported as safe, equally efficacious, more accessible and cost effective without negatively impacting patient outcomes. This paper summarizes the supporting evidence. PMID:26897866

  17. Correlations between Lymphocytes, Mid-Cell Fractions and Granulocytes with Human Blood Characteristics Using Low Power He-Ne Laser Radiation

    NASA Astrophysics Data System (ADS)

    Houssein, Hend A. A.; Jaafar, M. S.; Ramli, R. M.; Ismail, N. E.; Ahmad, A. L.; Bermakai, M. Y.

    2010-07-01

    In this study, the subpopulations of human blood parameters including lymphocytes, the mid-cell fractions (eosinophils, basophils, and monocytes), and granulocytes were determined by electronic sizing in the Health Centre of Universiti Sains Malaysia. These parameters have been correlated with human blood characteristics such as age, gender, ethnicity, and blood types; before and after irradiation with 0.95 mW He-Ne laser (λ = 632.8 nm). The correlations were obtained by finding patterns, paired non-parametric tests, and an independent non-parametric tests using the SPSS version 11.5, centroid and peak positions, and flux variations. The findings show that the centroid and peak positions, flux peak and total flux, were very much correlated and can become a significant indicator for blood analyses. Furthermore, the encircled flux analysis demonstrated a good future prospect in blood research, thus leading the way as a vibrant diagnosis tool to clarify diseases associated with blood.

  18. The transcriptional regulation of the Colony-Stimulating Factor 1 Receptor (csf1r) gene during hematopoiesis.

    PubMed

    Bonifer, Constanze; Hume, David A

    2008-01-01

    The colony-stimulating-factor 1 receptor (CSF-1 R) is a tyrosine kinase receptor that is absolutely required for macrophage differentiation and thus occupies a central role in hematopoiesis. Mice deficient for the csf1r gene show multiple defects in macrophage development, reproduction and tissue remodeling. Moreover, deregulation of this gene is a hallmark of many tumors. This includes repression of expression in acute myeloid leukemia and aberrant activation in certain solid tumors, such as breast cancer. Expression of this gene therefore needs to be tightly controlled. This review summarizes experiments providing a detailed picture of how transcription of csf1r gene expression is regulated. Aside from the direct relevance to hematopoiesis, studies of csf1r transcriptional regulation provide a model for understanding the molecular mechanisms that control mammalian cell fate.

  19. The macrophage-colony stimulating factor gene is a growth factor-inducible immediate early gene in fibroblasts.

    PubMed

    Ryseck, R P; Macdonald-Bravo, H; Bravo, R

    1991-02-01

    Polypeptide growth factors rapidly induce the expression of a group of genes during the onset of cell proliferation. We report that one of these genes, which is induced by several mitogens in NIH 3T3 cells, is identical to the gene for macrophage-colony stimulating factor (M-CSF). In contrast to other immediate early genes, the expression of the M-CSF gene lasted for several hours. Run-on assays demonstrated that the increased level of M-CSF mRNA following stimulation was mainly due to transcriptional activation. Our results support the notion that the products of the immediate early genes are not all mediators of fibroblasts growth but that some play an important role in other physiological responses such as wound repair. PMID:1712227

  20. Microglia overexpressing the macrophage colony-stimulating factor receptor are neuroprotective in a microglial-hippocampal organotypic coculture system.

    PubMed

    Mitrasinovic, Olivera M; Grattan, Alicia; Robinson, Christopher C; Lapustea, Nicolae B; Poon, Clara; Ryan, Heather; Phong, Connie; Murphy, Greer M

    2005-04-27

    Microglia with increased expression of the macrophage colony-stimulating factor receptor (M-CSFR; c-fms) are found surrounding plaques in Alzheimer's disease (AD) and in mouse models for AD and after ischemic or traumatic brain injury. Increased expression of M-CSFR causes microglia to adopt an activated state that results in proliferation, release of cytokines, and enhanced phagocytosis. To determine whether M-CSFR-induced microglial activation affects neuronal survival, we assembled a coculture system consisting of BV-2 microglia transfected to overexpress the M-CSFR and hippocampal organotypic slices treated with NMDA. Twenty-four hours after assembly of the coculture, microglia overexpressing M-CSFR proliferated at a higher rate than nontransfected control cells and exhibited enhanced migration toward NMDA-injured hippocampal cultures. Surprisingly, coculture with c-fms-transfected microglia resulted in a dramatic reduction in NMDA-induced neurotoxicity. Similar results were observed when cocultures were treated with the teratogen cyclophosphamide. Biolistic overexpression of M-CSFR on microglia endogenous to the organotypic culture also rescued neurons from excitotoxicity. Furthermore, c-fms-transfected microglia increased neuronal expression of macrophage colony-stimulating factor (M-CSF), the M-CSFR, and neurotrophin receptors in the NMDA-treated slices, as determined with laser capture microdissection. In the coculture system, direct contact between the exogenous microglia and the slice was necessary for neuroprotection. Finally, blocking expression of the M-CSF ligand by exogenous c-fms-transfected microglia with a hammerhead ribozyme compromised their neuroprotective properties. These results demonstrate a protective role for microglia overexpressing M-CSFR in our coculture system and suggest under certain circumstances, activated microglia can help rather than harm neurons subjected to excitotoxic and teratogen-induced injury.

  1. The profiles of gamma-H2AX along with ATM/DNA-PKcs activation in the lymphocytes and granulocytes of rat and human blood exposed to gamma rays.

    PubMed

    Wang, Jing; Yin, Lina; Zhang, Junxiang; Zhang, Yaping; Zhang, Xuxia; Ding, Defang; Gao, Yun; Li, Qiang; Chen, Honghong

    2016-08-01

    Establishing a rat model suitable for γ-H2AX biodosimeter studies has important implications for dose assessment of internal radionuclide contamination in humans. In this study, γ-H2AX, p-ATM and p-DNA-PKcs foci were enumerated using immunocytofluorescence method, and their protein levels were measured by Western blot in rat blood lymphocytes and granulocytes exposed to γ-rays compared with human blood lymphocytes and granulocytes. It was found that DNA double-strand break repair kinetics and linear dose responses in rat lymphocytes were similar to those observed in the human counterparts. Moreover, radiation induced clear p-ATM and p-DNA-PKcs foci formation and an increase in ratio of co-localization of p-ATM or p-DNA-PKcs with γ-H2AX foci in rat lymphocytes similar to those of human lymphocytes. The level of γ-H2AX protein in irradiated rat and human lymphocytes was significantly reduced by inhibitors of ATM and DNA-PKcs. Surprisingly, unlike human granulocytes, rat granulocytes with DNA-PKcs deficiency displayed a rapid accumulation, but delayed disappearance of γ-H2AX foci with essentially no change from 10 h to 48 h post-irradiation. Furthermore, inhibition of ATM activity in rat granulocytes also decreased radiation-induced γ-H2AX foci formation. In comparison, human granulocytes showed no response to irradiation regarding γ-H2AX, p-ATM or p-DNA-PKcs foci. Importantly, incidence of γ-H2AX foci in lymphocytes after total-body radiation of rats was consistent with that of in vitro irradiation of rat lymphocytes. These findings show that rats are a useful in vivo model for validation of γ-H2AX biodosimetry for dose assessment in humans. ATM and DNA-PKcs participate together in DSB repair in rat lymphocytes similar to that of human lymphocytes. Further, rat granulocytes, which have the characteristic of delayed disappearance of γ-H2AX foci in response to radiation, may be a useful experimental system for biodosimetry studies.

  2. The profiles of gamma-H2AX along with ATM/DNA-PKcs activation in the lymphocytes and granulocytes of rat and human blood exposed to gamma rays.

    PubMed

    Wang, Jing; Yin, Lina; Zhang, Junxiang; Zhang, Yaping; Zhang, Xuxia; Ding, Defang; Gao, Yun; Li, Qiang; Chen, Honghong

    2016-08-01

    Establishing a rat model suitable for γ-H2AX biodosimeter studies has important implications for dose assessment of internal radionuclide contamination in humans. In this study, γ-H2AX, p-ATM and p-DNA-PKcs foci were enumerated using immunocytofluorescence method, and their protein levels were measured by Western blot in rat blood lymphocytes and granulocytes exposed to γ-rays compared with human blood lymphocytes and granulocytes. It was found that DNA double-strand break repair kinetics and linear dose responses in rat lymphocytes were similar to those observed in the human counterparts. Moreover, radiation induced clear p-ATM and p-DNA-PKcs foci formation and an increase in ratio of co-localization of p-ATM or p-DNA-PKcs with γ-H2AX foci in rat lymphocytes similar to those of human lymphocytes. The level of γ-H2AX protein in irradiated rat and human lymphocytes was significantly reduced by inhibitors of ATM and DNA-PKcs. Surprisingly, unlike human granulocytes, rat granulocytes with DNA-PKcs deficiency displayed a rapid accumulation, but delayed disappearance of γ-H2AX foci with essentially no change from 10 h to 48 h post-irradiation. Furthermore, inhibition of ATM activity in rat granulocytes also decreased radiation-induced γ-H2AX foci formation. In comparison, human granulocytes showed no response to irradiation regarding γ-H2AX, p-ATM or p-DNA-PKcs foci. Importantly, incidence of γ-H2AX foci in lymphocytes after total-body radiation of rats was consistent with that of in vitro irradiation of rat lymphocytes. These findings show that rats are a useful in vivo model for validation of γ-H2AX biodosimetry for dose assessment in humans. ATM and DNA-PKcs participate together in DSB repair in rat lymphocytes similar to that of human lymphocytes. Further, rat granulocytes, which have the characteristic of delayed disappearance of γ-H2AX foci in response to radiation, may be a useful experimental system for biodosimetry studies. PMID:27260225

  3. Hemocytic rickettsia-like organisms in ticks: serologic reactivity with antisera to Ehrlichiae and detection of DNA of agent of human granulocytic ehrlichiosis by PCR.

    PubMed Central

    Magnarelli, L A; Stafford, K C; Mather, T N; Yeh, M T; Horn, K D; Dumler, J S

    1995-01-01

    Ixodid ticks were collected from Connecticut, Massachusetts, Missouri, Pennsylvania, Rhode Island, and British Columbia (Canada) during 1991 to 1994 to determine the prevalence of infection with hemocytic (blood cell), rickettsia-like organisms. Hemolymph obtained from these ticks was analyzed by direct and indirect fluorescent antibody (FA) staining methods with dog, horse, or human sera containing antibodies to Ehrlichia canis, Ehrlichia equi, or Rickettsia rickettsii. Of the 693 nymphal and adult Amblyomma americanum, Dermacentor variabilis, Ixodes scapularis, and Ixodes pacificus ticks tested with dog anti-E. canis antiserum, 209 (32.5%) contained hemocytic bacteria. The prevalence of infected ticks varied greatly with species and locale. In parallel tests of duplicate hemolymph preparations from adult I. scapularis ticks, the hemocytic organisms reacted positively with E. canis and/or E. equi antisera, including sera from persons who had granulocytic ehrlichiosis. In separate PCR analyses, DNA of the agent of human granulocytic ehrlichiosis was detected in 59 (50.0%) of 118 adult and in 1 of 2 nymphal I. scapularis ticks tested from Connecticut. There was no evidence of Ehrlichia chaffeensis DNA in these ticks. In indirect FA tests of hemolymph for spotted fever group rickettsiae, the overall prevalence of infection was less than 4%. Specificity tests of antigens and antisera used in these studies revealed no cross-reactivity between E. canis and E. equi or between any of the ehrlichial reagents and those of R. rickettsii. The geographic distribution of hemocytic microorganisms with shared antigens to Ehrlichia species or spotted fever group rickettsiae is widespread. PMID:8567911

  4. Sustained activation of proton channels and NADPH oxidase in human eosinophils and murine granulocytes requires PKC but not cPLA2α activity

    PubMed Central

    Morgan, Deri; Cherny, Vladimir V; Finnegan, Alison; Bollinger, James; Gelb, Michael H; DeCoursey, Thomas E

    2007-01-01

    The prevailing hypothesis that a signalling pathway involving cPLA2α is required to enhance the gating of the voltage-gated proton channel associated with NADPH oxidase was tested in human eosinophils and murine granulocytes. This hypothesis invokes arachidonic acid (AA) liberated by cPLA2α as a final activator of proton channels. In human eosinophils studied in the perforated-patch configuration, phorbol myristate acetate (PMA) stimulation elicited NADPH oxidase-generated electron current (Ie) and enhanced proton channel gating identically in the presence or absence of three specific cPLA2α inhibitors, Wyeth-1, pyrrolidine-2 and AACOCF3 (arachidonyl trifluoromethyl ketone). In contrast, PKC inhibitors GFX (GF109203X) or staurosporine prevented the activation of either proton channels or NADPH oxidase. PKC inhibition during the respiratory burst reversed the activation of both molecules, suggesting that ongoing phosphorylation is required. This effect of GFX was inhibited by okadaic acid, implicating phosphatases in proton channel deactivation. Proton channel activation by AA was partially reversed by GFX or staurosporine, indicating that AA effects are due in part to activation of PKC. In granulocytes from mice with the cPLA2α gene disrupted (knockout mice), PMA or fMetLeuPhe activated NADPH oxidase and proton channels in a manner indistinguishable from the responses of control cells. Thus, cPLA2α is not essential to activate the proton conductance or for a normal respiratory burst. Instead, phosphorylation of the proton channel or an activating molecule converts the channel to its activated gating mode. The existing paradigm for regulation of the concerted activity of proton channels and NADPH oxidase must be revised. PMID:17185330

  5. Activation of adenosine A(3) receptors potentiates stimulatory effects of IL-3, SCF, and GM-CSF on mouse granulocyte-macrophage hematopoietic progenitor cells.

    PubMed

    Hofer, M; Vacek, A; Pospísil, M; Holá, J; Streitová, D; Znojil, V

    2009-01-01

    Adenosine A(3) receptor agonist N(6)-(3-iodobenzyl)adenosine-5'-N-methyluronamide (IB-MECA) has been tested from the point of view of potentiating the effects of hematopoietic growth factors interleukin-3 (IL-3), stem cell factor (SCF), granulocyte-macrophage colony-stimulating factor (GM-CSF), and granulocyte colony-stimulating factor (G-CSF) on the growth of hematopoietic progenitor cells for granulocytes and macrophages (GM-CFC) in suspension of normal mouse bone marrow cells in vitro. IB-MECA alone induced no GM-CFC growth. Significant elevation of numbers of GM-CFC evoked by the combinations of IB-MECA with IL-3, SCF, or GM-CSF as compared with these growth factors alone has been noted. Combination of IB-MECA with G-CSF did not induce significantly higher numbers of GM-CFC in comparison with G-CSF alone. Joint action of three drugs, namely of IB-MECA + IL-3 + GM-CSF, produced significantly higher numbers of GM-CFC in comparison with the combinations of IB-MECA + IL-3, IB-MECA + GM-CSF, or IL-3 + GM-CSF. These results give evidence of a significant role of selective activation of adenosine A(3) receptors in stimulation of the growth of granulocyte/ macrophage hematopoietic progenitor cells.

  6. Combinatorial and Computational Approaches to Identify Interactions of Macrophage Colony-stimulating Factor (M-CSF) and Its Receptor c-FMS.

    PubMed

    Rosenfeld, Lior; Shirian, Jason; Zur, Yuval; Levaot, Noam; Shifman, Julia M; Papo, Niv

    2015-10-23

    The molecular interactions between macrophage colony-stimulating factor (M-CSF) and the tyrosine kinase receptor c-FMS play a key role in the immune response, bone metabolism, and the development of some cancers. Because no x-ray structure is available for the human M-CSF · c-FMS complex, the binding epitope for this complex is largely unknown. Our goal was to identify the residues that are essential for binding of the human M-CSF to c-FMS. For this purpose, we used a yeast surface display (YSD) approach. We expressed a combinatorial library of monomeric M-CSF (M-CSFM) single mutants and screened this library to isolate variants with reduced affinity for c-FMS using FACS. Sequencing yielded a number of single M-CSFM variants with mutations both in the direct binding interface and distant from the binding site. In addition, we used computational modeling to map the identified mutations onto the M-CSFM structure and to classify the mutations into three groups as follows: those that significantly decrease protein stability; those that destroy favorable intermolecular interactions; and those that decrease affinity through allosteric effects. To validate the YSD and computational data, M-CSFM and three variants were produced as soluble proteins; their affinity and structure were analyzed; and very good correlations with both YSD data and computational predictions were obtained. By identifying the M-CSFM residues critical for M-CSF · c-FMS interactions, we have laid down the basis for a deeper understanding of the M-CSF · c-FMS signaling mechanism and for the development of target-specific therapeutic agents with the ability to sterically occlude the M-CSF·c-FMS binding interface.

  7. Combinatorial and Computational Approaches to Identify Interactions of Macrophage Colony-stimulating Factor (M-CSF) and Its Receptor c-FMS.

    PubMed

    Rosenfeld, Lior; Shirian, Jason; Zur, Yuval; Levaot, Noam; Shifman, Julia M; Papo, Niv

    2015-10-23

    The molecular interactions between macrophage colony-stimulating factor (M-CSF) and the tyrosine kinase receptor c-FMS play a key role in the immune response, bone metabolism, and the development of some cancers. Because no x-ray structure is available for the human M-CSF · c-FMS complex, the binding epitope for this complex is largely unknown. Our goal was to identify the residues that are essential for binding of the human M-CSF to c-FMS. For this purpose, we used a yeast surface display (YSD) approach. We expressed a combinatorial library of monomeric M-CSF (M-CSFM) single mutants and screened this library to isolate variants with reduced affinity for c-FMS using FACS. Sequencing yielded a number of single M-CSFM variants with mutations both in the direct binding interface and distant from the binding site. In addition, we used computational modeling to map the identified mutations onto the M-CSFM structure and to classify the mutations into three groups as follows: those that significantly decrease protein stability; those that destroy favorable intermolecular interactions; and those that decrease affinity through allosteric effects. To validate the YSD and computational data, M-CSFM and three variants were produced as soluble proteins; their affinity and structure were analyzed; and very good correlations with both YSD data and computational predictions were obtained. By identifying the M-CSFM residues critical for M-CSF · c-FMS interactions, we have laid down the basis for a deeper understanding of the M-CSF · c-FMS signaling mechanism and for the development of target-specific therapeutic agents with the ability to sterically occlude the M-CSF·c-FMS binding interface. PMID:26359491

  8. Combinatorial and Computational Approaches to Identify Interactions of Macrophage Colony-stimulating Factor (M-CSF) and Its Receptor c-FMS*

    PubMed Central

    Rosenfeld, Lior; Shirian, Jason; Zur, Yuval; Levaot, Noam; Shifman, Julia M.; Papo, Niv

    2015-01-01

    The molecular interactions between macrophage colony-stimulating factor (M-CSF) and the tyrosine kinase receptor c-FMS play a key role in the immune response, bone metabolism, and the development of some cancers. Because no x-ray structure is available for the human M-CSF·c-FMS complex, the binding epitope for this complex is largely unknown. Our goal was to identify the residues that are essential for binding of the human M-CSF to c-FMS. For this purpose, we used a yeast surface display (YSD) approach. We expressed a combinatorial library of monomeric M-CSF (M-CSFM) single mutants and screened this library to isolate variants with reduced affinity for c-FMS using FACS. Sequencing yielded a number of single M-CSFM variants with mutations both in the direct binding interface and distant from the binding site. In addition, we used computational modeling to map the identified mutations onto the M-CSFM structure and to classify the mutations into three groups as follows: those that significantly decrease protein stability; those that destroy favorable intermolecular interactions; and those that decrease affinity through allosteric effects. To validate the YSD and computational data, M-CSFM and three variants were produced as soluble proteins; their affinity and structure were analyzed; and very good correlations with both YSD data and computational predictions were obtained. By identifying the M-CSFM residues critical for M-CSF·c-FMS interactions, we have laid down the basis for a deeper understanding of the M-CSF·c-FMS signaling mechanism and for the development of target-specific therapeutic agents with the ability to sterically occlude the M-CSF·c-FMS binding interface. PMID:26359491

  9. The role of colony-stimulating factor 1 and its receptor in the etiopathogenesis of endometrial adenocarcinoma.

    PubMed

    Smith, H O; Anderson, P S; Kuo, D Y; Goldberg, G L; DeVictoria, C L; Boocock, C A; Jones, J G; Runowicz, C D; Stanley, E R; Pollard, J W

    1995-03-01

    Colony-stimulating factor 1 (CSF-1) is a homodimeric growth factor that humorally regulates the growth and differentiation of mononuclear phagocytes, and locally regulates maternal-fetal interactions during pregnancy. It exerts these actions through a transmembrane tyrosine kinase receptor, colony-stimulating factor 1 receptor (CSF-1R), the product of the c-fms proto-oncogene. Recent studies have demonstrated overexpression of CSF-1 and its receptor in breast, ovarian, and endometrial adenocarcinomas. To further investigate the possible role of CSF-1 and its receptor in the pathogenesis of endometrial adenocarcinoma, a prospective study was undertaken to study CSF-1 expression in benign and neoplastic endometrial epithelium and to compare serum CSF-1 levels in endometrial adenocarcinoma patients with healthy perimenopausal women. The mean serum levels of CSF-1 in 71 patients with endometrial cancer (4.9 +/- 1.8 microgram/liter) were significantly elevated compared with levels found in the 32 controls (3.5 +/- 1.1 microgram/liter). Within the endometrial adenocarcinoma group, circulating CSF-1 levels were significantly elevated in patients with large tumor volume, high grade, myometrial invasion, residual disease, and circulating CA-125 levels. High serum levels of serum CSF-1 were associated with elevated serum CA19-9 and CA-125 levels. Immunohistochemistry results revealed in tumor epithelium intense staining for CSF-1R (27 of 54 cases, 50%) and elevated staining for CSF-1 (41 of 54 cases, 75.9%), with intense staining of CSF-1 in 16 of 54 cases (29.6%). Staining was significantly greater in intensity and number of cells involved in malignant compared with benign epithelium for CSF-1R and CSF-1 (P = 0.05 and <0.0001, respectively). A positive correlation between amount and intensity of CSF-1 and CSF-1R staining in endometrial adenocarcinoma tissue was also demonstrated (P = 0.007). CSF-1 and CSF-1R mRNA was also detected in the tumor samples, confirming the

  10. Uptake and Economic Impact of First-Cycle Colony-Stimulating Factor Use During Adjuvant Treatment of Breast Cancer

    PubMed Central

    Hershman, Dawn L.; Wilde, Elizabeth T.; Wright, Jason D.; Buono, Donna L.; Kalinsky, Kevin; Malin, Jennifer L.; Neugut, Alfred I.

    2012-01-01

    Purpose In 2002, pegfilgrastim was approved by the US Food and Drug Administration and the benefits of dose-dense breast cancer chemotherapy, especially for hormone receptor (HR) –negative tumors, were reported. We examined first-cycle colony-stimulating factor use (FC-CSF) before and after 2002 and estimated US expenditures for dose-dense chemotherapy. Methods We identified patients in Surveillance, Epidemiology, and End Results–Medicare greater than 65 years old with stages I to III breast cancer who had greater than one chemotherapy claim within 6 months of diagnosis(1998 to 2005) and classified patients with an average cycle length less than 21 days as having received dose-dense chemotherapy. The associations of patient, tumor, and physician-related factors with the receipt of any colony-stimulating factor (CSF) and FC-CSF use were analyzed by using generalized estimating equations. CSF costs were estimated for patients who were undergoing dose-dense chemotherapy. Results Among the 10,773 patients identified, 5,266 patients (48.9%) had a CSF claim. CSF use was stable between 1998 and 2002 and increased from 36.8% to 73.7% between 2002 and 2005, FC-CSF use increased from 13.2% to 67.9%, and pegfilgrastim use increased from 4.1% to 83.6%. In a multivariable analysis, CSF use was associated with age and chemotherapy type and negatively associated with black/Hispanic race, rural residence, and shorter chemotherapy duration. FC-CSF use was associated with high socioeconomic status but not with age or race/ethnicity. The US annual CSF expenditure for women with HR-positive tumors treated with dose-dense chemotherapy is estimated to be $38.8 million. Conclusion A rapid increase in FC-CSF use occurred over a short period of time, which was likely a result of the reported benefits of dose-dense chemotherapy and the ease of pegfilgrastim administration. Because of the increasing evidence that elderly HR-positive patients do not benefit from dose-dense chemotherapy

  11. Glioblastoma-derived Macrophage Colony-stimulating Factor (MCSF) Induces Microglial Release of Insulin-like Growth Factor-binding Protein 1 (IGFBP1) to Promote Angiogenesis.

    PubMed

    Nijaguna, Mamatha Bangalore; Patil, Vikas; Urbach, Serge; Shwetha, Shivayogi D; Sravani, Kotha; Hegde, Alangar S; Chandramouli, Bangalore A; Arivazhagan, Arimappamagan; Marin, Philippe; Santosh, Vani; Somasundaram, Kumaravel

    2015-09-18

    Glioblastoma (grade IV glioma/GBM) is the most common primary adult malignant brain tumor with poor prognosis. To characterize molecular determinants of tumor-stroma interaction in GBM, we profiled 48 serum cytokines and identified macrophage colony-stimulating factor (MCSF) as one of the elevated cytokines in sera from GBM patients. Both MCSF transcript and protein were up-regulated in GBM tissue samples through a spleen tyrosine kinase (SYK)-dependent activation of the PI3K-NFκB pathway. Ectopic overexpression and silencing experiments revealed that glioma-secreted MCSF has no role in autocrine functions and M2 polarization of macrophages. In contrast, silencing expression of MCSF in glioma cells prevented tube formation of human umbilical vein endothelial cells elicited by the supernatant from monocytes/microglial cells treated with conditioned medium from glioma cells. Quantitative proteomics based on stable isotope labeling by amino acids in cell culture showed that glioma-derived MCSF induces changes in microglial secretome and identified insulin-like growth factor-binding protein 1 (IGFBP1) as one of the MCSF-regulated proteins secreted by microglia. Silencing IGFBP1 expression in microglial cells or its neutralization by an antibody reduced the ability of supernatants derived from microglial cells treated with glioma cell-conditioned medium to induce angiogenesis. In conclusion, this study shows up-regulation of MCSF in GBM via a SYK-PI3K-NFκB-dependent mechanism and identifies IGFBP1 released by microglial cells as a novel mediator of MCSF-induced angiogenesis, of potential interest for developing targeted therapy to prevent GBM progression.

  12. Alteration of mineral crystallinity and collagen cross-linking of bones in osteopetrotic toothless (tl/tl) rats and their improvement after treatment with colony stimulating factor-1

    NASA Technical Reports Server (NTRS)

    Wojtowicz, A.; Dziedzic-Goclawska, A.; Kaminski, A.; Stachowicz, W.; Wojtowicz, K.; Marks, S. C. Jr; Yamauchi, M.

    1997-01-01

    A common feature of various types of mammalian osteopetroses is a marked increase in bone mass accompanied by spontaneous bone fractures. The toothless (tl/tl) rat osteopetrotic mutation is characterized by drastically reduced bone resorption due to a profound deficiency of osteoclasts and their precursors. An altered bone morphology has also been observed. The mutants cannot be cured by bone marrow transplantation, but skeletal defects are greatly reduced after treatment with colony stimulating factor 1 (CSF-1). The objectives of this study were to characterize mineral and collagen matrices in cancellous and compact bone isolated from long bones of 6-week-old normal littermates, tl/tl osteopetrotic mutants and mutants (tl/tl) treated with CSF-1. There were no differences in bone mineral content, but a significant decrease in the crystallinity of mineral evaluated by the method based on electron paramagnetic resonance spectrometry was observed in all bones of tl/tl mutants as compared to that of controls. Within the collagen matrix, slight decreases in the labile cross-links, but significant increases in the content of the stable cross-links, pyridinoline, and deoxypyridinoline, were observed in both cancellous and compact bone of osteopetrotic mutants. In tl/tl mutants treated with human recombinant CSF-1, the normalization of the crystallinity of bone mineral as well as collagen cross-links was found. Our results indicate that remodeling of bone matrix in tl/tl mutants is highly suppressed, but that after treatment with CSF-1, this activity recovers significantly. Taken together, these data provide further support for the hypothesis that CSF-1 is an essential factor for normal osteoclast differentiation and bone remodelling.

  13. Nerve growth factor promotes human hemopoietic colony growth and differentiation

    SciTech Connect

    Matsuda, H.; Coughlin, M.D.; Bienenstock, J.; Denburg, J.A. )

    1988-09-01

    Nerve growth factor (NGF) is a neurotropic polypeptide necessary for the survival and growth of some central neurons, as well as sensory afferent and sympathetic neurons. Much is now known of the structural and functional characteristics of NGF, whose gene has recently been clones. Since it is synthesized in largest amounts by the male mouse submandibular gland, its role exclusively in nerve growth is questionable. These experiments indicate that NGF causes a significant stimulation of granulocyte colonies grown from human peripheral blood in standard hemopoietic methylcellulose assays. Further, NGF appears to act in a relatively selective fashion to induce the differentiation of eosinophils and basophils/mast cells. Depletion experiments show that the NGF effect may be T-cell dependent and that NGF augments the colony-stimulating effect of supernatants from the leukemic T-cell (Mo) line. The hemopoietic activity of NGF is blocked by {sup 125}I-polyclonal and monoclonal antibodies to NGF. The authors conclude that NGF may indirectly act as a local growth factor in tissues other than those of the nervous system by causing T cells to synthesize or secrete molecules with colony-stimulating activity. In view of the synthesis of NGF in tissue injury, the involvement of basophils/mast cells and eosinophils in allergic and other inflammatory processes, and the association of mast cells with fibrosis and tissue repair, they postulate that NGF plays an important biological role in a variety of repair processes.

  14. Complement factor H modulates the activation of human neutrophil granulocytes and the generation of neutrophil extracellular traps.

    PubMed

    Schneider, Andrea E; Sándor, Noémi; Kárpáti, Éva; Józsi, Mihály

    2016-04-01

    Factor H (FH) is a major inhibitor of the alternative pathway of complement activation in plasma and on certain host surfaces. In addition to being a complement regulator, FH can bind to various cells via specific receptors, including binding to neutrophil granulocytes through complement receptor type 3 (CR3; CD11b/CD18), and modulate their function. The cellular roles of FH are, however, poorly understood. Because neutrophils are important innate immune cells in inflammatory processes and the host defense against pathogens, we aimed at studying the effects of FH on various neutrophil functions, including the generation of extracellular traps. FH co-localized with CD11b on the surface of neutrophils isolated from peripheral blood of healthy individuals, and cell-bound FH retained its cofactor activity and enhanced C3b degradation. Soluble FH supported neutrophil migration and immobilized FH induced cell spreading. In addition, immobilized but not soluble FH enhanced IL-8 release from neutrophils. FH alone did not trigger the cells to produce neutrophil extracellular traps (NETs), but NET formation induced by PMA and by fibronectin plus fungal β-glucan were inhibited by immobilized, but not by soluble, FH. Moreover, in parallel with NET formation, immobilized FH also inhibited the production of reactive oxygen species induced by PMA and by fibronectin plus β-glucan. Altogether, these data indicate that FH has multiple regulatory roles on neutrophil functions. While it can support the recruitment of neutrophils, FH may also exert anti-inflammatory effects and influence local inflammatory and antimicrobial reactions, and reduce tissue damage by modulating NET formation. PMID:26938503

  15. Thrombin binds to murine bone marrow-derived macrophages and enhances colony-stimulating factor-1-driven mitogenesis

    SciTech Connect

    Clohisy, D.R.; Erdmann, J.M.; Wilner, G.D. )

    1990-05-15

    The binding and mitogenic properties of thrombin have been established in various transformed cell lines. In such systems, thrombin induces cell division in the absence of exogenous growth factors, and the enzyme is considered to act directly as a mitogen. This study explores thrombin's interaction with nontransformed, growth factor-dependent cells. Binding of 125I-alpha-thrombin to colony-stimulating factor (CSF)-1-dependent bone marrow-derived macrophages is saturable, time-dependent, and displaceable by both unlabeled alpha-thrombin, and esterolytically inactive thrombin. Both dissociation studies of pre-bound radio-labeled thrombin and Scatchard analysis assisted by the program Ligand suggest adherence of thrombin-binding data to a multi-site model. There are an estimated 2 x 10(4) high affinity sites (Kd = 7 x 10(-9)M) and 2 x 10(6) low affinity sites (Kd = 9 x 10(-7)M) per cell. Quiescent bone marrow-derived macrophages were cultured with either 10(-8)M thrombin, 1000 units of CSF-1/ml, or both and (3H)thymidine incorporation was determined. Thrombin alone did not induce mitogenesis. CSF-1 induced mitogenesis with peak (3H) thymidine incorporation occurring 24 h after addition of the mitogen. This CSF-1-dependent mitogenic influence was enhanced greater than 2-fold by treatment with thrombin.

  16. Expression of colony-stimulating factor 1 is associated with occurrence of osteochondral change in pigmented villonodular synovitis.

    PubMed

    Ota, Takehiro; Urakawa, Hiroshi; Kozawa, Eiji; Ikuta, Kunihiro; Hamada, Shunsuke; Tsukushi, Satoshi; Shimoyama, Yoshie; Ishiguro, Naoki; Nishida, Yoshihiro

    2015-07-01

    Pigmented villonodular synovitis (PVNS) is a benign, translocation-derived neoplasm. Because of its high local recurrence rate after surgery and occurrence of osteochondral destruction, a novel therapeutic target is required. The present study aimed to evaluate the significance of protein expression possibly associated with the pathogenesis during the clinical course of PVNS. In 40 cases of PVNS, positivity of colony-stimulated factor 1 (CSF1), its receptor (CSF1R), and receptor activator of nuclear factor kappa-B ligand (RANKL) were immunohistochemically determined. The relationship between the positivity and clinical outcomes was investigated. High positivity of CSF1 staining intensity was associated with an increased incidence of osteochondral lesions (bone erosion and osteoarthritis) (p = 0.009), but not with the rate of local recurrence. Positivity of CSF1R and RANKL staining was not associated with any clinical variables. The number of giant cells was not correlated with positivity of any of the three proteins, or with the clinical outcome. Focusing on knee cases, CSF1 positivity was also associated with the incidence of osteochondal change (p = 0.02). CSF1R positivity was high in cases which had local recurrence, but not significantly so (p = 0.129). Determination of CSF1 and CSF1R expression may be useful as a prognosticator of the clinical course and/or outcomes of PVNS.

  17. Mutations in the colony stimulating factor 1 receptor (CSF1R) gene cause hereditary diffuse leukoencephalopathy with spheroids.

    PubMed

    Rademakers, Rosa; Baker, Matt; Nicholson, Alexandra M; Rutherford, Nicola J; Finch, NiCole; Soto-Ortolaza, Alexandra; Lash, Jennifer; Wider, Christian; Wojtas, Aleksandra; DeJesus-Hernandez, Mariely; Adamson, Jennifer; Kouri, Naomi; Sundal, Christina; Shuster, Elizabeth A; Aasly, Jan; MacKenzie, James; Roeber, Sigrun; Kretzschmar, Hans A; Boeve, Bradley F; Knopman, David S; Petersen, Ronald C; Cairns, Nigel J; Ghetti, Bernardino; Spina, Salvatore; Garbern, James; Tselis, Alexandros C; Uitti, Ryan; Das, Pritam; Van Gerpen, Jay A; Meschia, James F; Levy, Shawn; Broderick, Daniel F; Graff-Radford, Neill; Ross, Owen A; Miller, Bradley B; Swerdlow, Russell H; Dickson, Dennis W; Wszolek, Zbigniew K

    2011-12-25

    Hereditary diffuse leukoencephalopathy with spheroids (HDLS) is an autosomal-dominant central nervous system white-matter disease with variable clinical presentations, including personality and behavioral changes, dementia, depression, parkinsonism, seizures and other phenotypes. We combined genome-wide linkage analysis with exome sequencing and identified 14 different mutations affecting the tyrosine kinase domain of the colony stimulating factor 1 receptor (encoded by CSF1R) in 14 families with HDLS. In one kindred, we confirmed the de novo occurrence of the mutation. Follow-up sequencing identified an additional CSF1R mutation in an individual diagnosed with corticobasal syndrome. In vitro, CSF-1 stimulation resulted in rapid autophosphorylation of selected tyrosine residues in the kinase domain of wild-type but not mutant CSF1R, suggesting that HDLS may result from partial loss of CSF1R function. As CSF1R is a crucial mediator of microglial proliferation and differentiation in the brain, our findings suggest an important role for microglial dysfunction in HDLS pathogenesis.

  18. Identification of an isoform of colony-stimulating factor 1 receptor mRNA in the rat testis.

    PubMed

    Su, Hong; Wang, Yibiao; Söder, Olle; Hou, Mi

    2014-06-01

    Because alternative RNA splicing regulation in the testis is prevalent, we explored testes of Sprague-Dawley rats for existence of alternatively spliced colony-stimulating factor 1 receptor (CSF-1R) mRNA. Using RT-PCR and sequencing, we identified a variant of CSF-1R mRNA that was 284 bp shorter than the full-length CSF-1R transcript. This variant was present in the testis (late fetal stage to adult) and in other organs of rats (7 and 60 days old). The deletion of 284 bp disrupted the open reading frame, resulting in a noncoding mRNA product. When testicular macrophages were stimulated with CSF-1R ligand and lipopolysaccharide, proportionally increased expression of both short isoform and full-length CSF-1R mRNA was observed. Thus, the identified isoform of CSF-1R mRNA may interfere with the expression of full-length CSF-1R mRNA, thereby affecting the biological activity of the ligand/receptor signaling axis in Sprague-Dawley rats.

  19. Gene expression profiling of human erythroid progenitors by micro-serial analysis of gene expression.

    PubMed

    Fujishima, Naohito; Hirokawa, Makoto; Aiba, Namiko; Ichikawa, Yoshikazu; Fujishima, Masumi; Komatsuda, Atsushi; Suzuki, Yoshiko; Kawabata, Yoshinari; Miura, Ikuo; Sawada, Ken-ichi

    2004-10-01

    We compared the expression profiles of highly purified human CD34+ cells and erythroid progenitor cells by micro-serial analysis of gene expression (microSAGE). Human CD34+ cells were purified from granulocyte colony-stimulating factor-mobilized blood stem cells, and erythroid progenitors were obtained by cultivating these cells in the presence of stem cell factor, interleukin 3, and erythropoietin. Our 10,202 SAGE tags allowed us to identify 1354 different transcripts appearing more than once. Erythroid progenitor cells showed increased expression of LRBA, EEF1A1, HSPCA, PILRB, RANBP1, NACA, and SMURF. Overexpression of HSPCA was confirmed by real-time polymerase chain reaction analysis. MicroSAGE revealed an unexpected preferential expression of several genes in erythroid progenitor cells in addition to the known functional genes, including hemoglobins. Our results provide reference data for future studies of gene expression in various hematopoietic disorders, including myelodysplastic syndrome and leukemia.

  20. Prostaglandin E2 transactivates the colony-stimulating factor-1 receptor and synergizes with colony-stimulating factor-1 in the induction of macrophage migration via the mitogen-activated protein kinase ERK1/2.

    PubMed

    Digiacomo, Graziana; Ziche, Marina; Dello Sbarba, Persio; Donnini, Sandra; Rovida, Elisabetta

    2015-06-01

    Prostaglandin E2 (PGE2), a key mediator of immunity, inflammation, and cancer, acts through 4 G-protein-coupled E-prostanoid receptors (EPs 1-4). Crosstalk between EPs and receptor tyrosine kinases also occurs. Colony-stimulating factor-1 receptor (CSF-1R) is an RTK that sustains the survival, proliferation, and motility of monocytes/macrophages, which are an essential component of innate immunity and cancer development. The aim of this study was to investigate on a possible crosstalk between EP and CSF-1R. In BAC1.2F5 and RAW264.7 murine macrophages, CSF-1 (EC₅₀ = 18.1 and 10.2 ng/ml, respectively) and PGE2 (EC₅₀ = 1.5 and 5.5 nM, respectively) promoted migration. PGE2 induced rapid CSF-1R phosphorylation that was dependent on Src family kinases (SFKs). CSF-1R inhibition reduced PGE2-elicited ERK1/2 phosphorylation and macrophage migration, indicating that CSF-1R plays a role in PGE2-mediated immunoregulation. EP4 appeared responsible for functional PGE2/CSF-1R crosstalk. Furthermore, PGE2 synergized with CSF-1 in inducing ERK1/2 phosphorylation and macrophage migration. ERK1/2 inhibition completely blocked migration induced by the combination CSF-1/PGE2. CSF-1/PGE2 functional interaction with respect to migration also occurred in bone marrow-derived murine macrophages (EC₅₀ CSF-1, 6.7 ng/ml; EC₅₀ PGE2, 16.7 nM). These results indicated that PGE2 transactivates CSF-1R and synergizes with its signaling at ERK1/2 level in promoting macrophage migration.

  1. Clinically effective monoclonal antibody 3F8 mediates nonoxidative lysis of human neuroectodermal tumor cells by polymorphonuclear leukocytes.

    PubMed

    Kushner, B H; Cheung, N K

    1991-09-15

    Most studies of antibody-dependent cellular cytotoxicity (ADCC) by polymorphonuclear leukocytes (PMN) have supported oxidative lytic processes. This may be because the studies used nonhuman or nonneoplastic cells that were highly sensitive to reactive oxygen species or were small enough to be phagocytosed by PMN. We therefore investigated whether oxygen radicals participate in PMN cytotoxicity toward human neuroectodermal solid tumor cells sensitized by 3F8, which is an anti-ganglioside GD2 murine IgG3 monoclonal antibody with documented anticancer activity in humans. A 4-h 51Cr release assay was used to assess tumor cell lysis by hydrogen peroxide, superoxide, and hypochlorite. Nine of 11 GD2(+) human melanoma and neuroblastoma cell lines had equal or greater resistance to these oxidants as compared to a GD2(-) human carcinoma line (SKBr1-III) found by others (and confirmed by us) to be significantly more resistant to oxidative lysis than a murine cell line (P388D1) representative of those commonly used in cytotoxicity assays. To facilitate detection of oxidant-mediated lysis, subsequent studies of 3F8-mediated ADCC used GD2(+) targets that were relatively sensitive and others that were relatively resistant to oxygen radicals. Normal PMN and PMN obtained from children with chronic granulomatous disease, which do not generate reactive oxygen species, were equally effective in ADCC. Granulocyte-macrophage colony-stimulating factor, which primes oxidative responses of normal but not of chronic granulomatous disease PMN, enhanced ADCC by both kinds of PMN. During ADCC of 3F8-sensitized targets, with or without granulocyte-macrophage colony-stimulating factor, GD2(-) "innocent bystander" tumor cells (including P388D1) were not lysed, a finding consistent with unimportant extracellular release of cytotoxic mediators. Finally, antioxidant and antimyeloperoxidase moieties did not block ADCC. We conclude that oxidants are not key factors in 3F8-mediated lysis by PMN of

  2. Synthesis of granulocyte–macrophage colony-stimulating factor as homogeneous glycoforms and early comparisons with yeast cell-derived material

    PubMed Central

    Zhang, Qiang; Johnston, Eric V.; Shieh, Jae-Hung; Moore, Malcolm A. S.; Danishefsky, Samuel J.

    2014-01-01

    Granulocyte–macrophage colony-stimulating factor (GM-CSF) is a medicinally important glycoprotein, used as an immunostimulant following bone-marrow transplant. On the basis of reports of its potential utility as an anticancer vaccine adjuvant, we undertook to develop a synthetic route toward single-glycoform GM-CSF. We describe herein a convergent total synthesis of GM-CSF aglycone and two homogeneous glycoforms. Analytical and biological studies confirm the structure and activity of these synthetic congeners. PMID:24516138

  3. Macrophage colony-stimulating factor expressed in non-cancer tissues provides predictive powers for recurrence in hepatocellular carcinoma

    PubMed Central

    Kono, Hiroshi; Fujii, Hideki; Furuya, Shinji; Hara, Michio; Hirayama, Kazuyoshi; Akazawa, Yoshihiro; Nakata, Yuuki; Tsuchiya, Masato; Hosomura, Naohiro; Sun, Chao

    2016-01-01

    AIM To investigate the role of macrophage colony-stimulating factor (M-CSF) in patients with hepatocellular carcinoma (HCC) after surgery. METHODS Expression of M-CSF, distribution of M2 macrophages (MΦs), and angiogenesis were assessed in the liver, including tumors and peritumoral liver tissues. The prognostic power of these factors was assessed. Mouse isolated hepatic MΦs or monocytes were cultured with media containing M-CSF. The concentration of vascular endothelial growth factor (VEGF) in media was assessed. Furthermore, the role of the M-CSF-matured hepatic MΦs on proliferation of the vascular endothelial cell (VEC) was investigated. RESULTS A strong correlation between the expressions of M-CSF and CD163 was observed in the peritumoral area. Also, groups with high density of M-CSF, CD163 or CD31 showed a significantly shorter time to recurrence (TTR) than low density groups. Multivariate analysis revealed the expression of M-CSF or hepatic M2MΦs in the peritumoral area as the most crucial factor responsible for shorter TTR. Moreover, the expression of M-CSF and hepatic M2MΦs in the peritumoral area had better predictable power of overall survival. Values of VEGF in culture media were significantly greater in the hepatic MΦs compared with the monocytes. Proliferation of the VEC was greatest in the cells co-cultured with hepatic MΦs when M-CSF was present in media. CONCLUSION M-CSF increases hepatocarcinogenesis, most likely by enhancing an angiogenic factor derived from hepatic MΦ and could be a useful target for therapy against HCC.

  4. Colony stimulating factor-1 receptor signaling networks inhibit mouse macrophage inflammatory responses by induction of microRNA-21.

    PubMed

    Caescu, Cristina I; Guo, Xingyi; Tesfa, Lydia; Bhagat, Tushar D; Verma, Amit; Zheng, Deyou; Stanley, E Richard

    2015-02-19

    Macrophage polarization between the M2 (repair, protumorigenic) and M1 (inflammatory) phenotypes is seen as a continuum of states. The detailed transcriptional events and signals downstream of colony-stimulating factor 1 receptor (CSF-1R) that contributes to amplification of the M2 phenotype and suppression of the M1 phenotype are largely unknown. Macrophage CSF-1R pTyr-721 signaling promotes cell motility and enhancement of tumor cell invasion in vitro. Combining analysis of cellular systems for CSF-1R gain of function and loss of function with bioinformatic analysis of the macrophage CSF-1R pTyr-721-regulated transcriptome, we uncovered microRNA-21 (miR-21) as a downstream molecular switch controlling macrophage activation and identified extracellular signal-regulated kinase1/2 and nuclear factor-κB as CSF-1R pTyr-721-regulated signaling nodes. We show that CSF-1R pTyr-721 signaling suppresses the inflammatory phenotype, predominantly by induction of miR-21. Profiling of the miR-21-regulated messenger RNAs revealed that 80% of the CSF-1-regulated canonical miR-21 targets are proinflammatory molecules. Additionally, miR-21 positively regulates M2 marker expression. Moreover, miR-21 feeds back to positively regulate its own expression and to limit CSF-1R-mediated activation of extracellular signal-regulated kinase1/2 and nuclear factor-κB. Consistent with an anti-inflammatory role of miRNA-21, intraperitoneal injection of mice with a miRNA-21 inhibitor increases the recruitment of inflammatory monocytes and enhances the peritoneal monocyte/macrophage response to lipopolysaccharide. These results identify the CSF-1R-regulated miR-21 network that modulates macrophage polarization.

  5. A novel glycobiomarker, Wisteria floribunda agglutinin macrophage colony-stimulating factor receptor, for predicting carcinogenesis of liver cirrhosis.

    PubMed

    Iio, Etsuko; Ocho, Makoto; Togayachi, Akira; Nojima, Masanori; Kuno, Atsushi; Ikehara, Yuzuru; Hasegawa, Izumi; Yatsuhashi, Hiroshi; Yamasaki, Kazumi; Shimada, Noritomo; Ide, Tatsuya; Shinkai, Noboru; Nojiri, Shunske; Fujiwara, Kei; Joh, Takashi; Mizokami, Masashi; Narimatsu, Hisashi; Tanaka, Yasuhito

    2016-03-15

    Recently, we identified a novel liver fibrosis glycobiomarker, Wisteria floribunda agglutinin (WFA)-reactive colony stimulating factor 1 receptor (WFA(+) -CSF1R), using a glycoproteomics-based strategy. The aim of this study was to assess the value of measuring WFA(+) -CSF1R levels for the prognosis of carcinogenesis and outcome in liver cirrhosis (LC) patients with hepatitis C virus (HCV). WFA(+) -CSF1R and Total-CSF1R levels were measured in serum samples from 214 consecutive HCV-infected patients to evaluate their impact on carcinogenesis and the survival of LC patients. Serum WFA(+) -CSF1R levels were significantly higher in LC patients than chronic hepatitis (CH) patients (p < 0.001). The AUC of WFA(+) -CSF1R for predicting overall survival, calculated by time-dependent ROC analysis, was 0.691 and the HR (per 1-SD increase) was 1.80 (95% CI, 1.23-2.62, p < 0.001). Furthermore, the survival rate of LC patients with high WFA(+) -CSF1R levels (≥ 310 ng/ml) was significantly worse than those with lower levels (p < 0.01). The AUC of WFA(+) /total-CSF1R percentage (WFA(+) -CSF1R%) for predicting the cumulative carcinogenesis rate was 0.760, with an HR of 1.66 (95% CI 1.26-2.20, p < 0.001). In fact, the carcinogenesis rate was significantly higher in LC patients with a high WFA(+) -CSF1R% (≥ 35%, p = 0.006). Assessing serum levels of WFA(+) -CSF1R has diagnostic value for predicting carcinogenesis and the survival of LC patients.

  6. Macrophage colony-stimulating factor receptor marks and regulates a fetal myeloid-primed B-cell progenitor in mice.

    PubMed

    Zriwil, Alya; Böiers, Charlotta; Wittmann, Lilian; Green, Joanna C A; Woll, Petter S; Jacobsen, Sten Eirik W; Sitnicka, Ewa

    2016-07-14

    Although it is well established that unique B-cell lineages develop through distinct regulatory mechanisms during embryonic development, much less is understood about the differences between embryonic and adult B-cell progenitor cells, likely to underpin the genetics and biology of infant and childhood PreB acute lymphoblastic leukemia (PreB-ALL), initiated by distinct leukemia-initiating translocations during embryonic development. Herein, we establish that a distinct subset of the earliest CD19(+) B-cell progenitors emerging in the E13.5 mouse fetal liver express the colony-stimulating factor-1 receptor (CSF1R), previously thought to be expressed, and play a lineage-restricted role in development of myeloid lineages, and macrophages in particular. These early embryonic CSF1R(+)CD19(+) ProB cells also express multiple other myeloid genes and, in line with this, possess residual myeloid as well as B-cell, but not T-cell lineage potential. Notably, these CSF1R(+) myeloid-primed ProB cells are uniquely present in a narrow window of embryonic fetal liver hematopoiesis and do not persist in adult bone marrow. Moreover, analysis of CSF1R-deficient mice establishes a distinct role of CSF1R in fetal B-lymphopoiesis. CSF1R(+) myeloid-primed embryonic ProB cells are relevant for infant and childhood PreB-ALLs, which frequently have a bi-phenotypic B-myeloid phenotype, and in which CSF1R-rearrangements have recently been reported.

  7. Macrophage colony-stimulating factor receptor marks and regulates a fetal myeloid-primed B-cell progenitor in mice

    PubMed Central

    Zriwil, Alya; Böiers, Charlotta; Wittmann, Lilian; Green, Joanna C. A.; Woll, Petter S.; Jacobsen, Sten Eirik W.

    2016-01-01

    Although it is well established that unique B-cell lineages develop through distinct regulatory mechanisms during embryonic development, much less is understood about the differences between embryonic and adult B-cell progenitor cells, likely to underpin the genetics and biology of infant and childhood PreB acute lymphoblastic leukemia (PreB-ALL), initiated by distinct leukemia-initiating translocations during embryonic development. Herein, we establish that a distinct subset of the earliest CD19+ B-cell progenitors emerging in the E13.5 mouse fetal liver express the colony-stimulating factor-1 receptor (CSF1R), previously thought to be expressed, and play a lineage-restricted role in development of myeloid lineages, and macrophages in particular. These early embryonic CSF1R+CD19+ ProB cells also express multiple other myeloid genes and, in line with this, possess residual myeloid as well as B-cell, but not T-cell lineage potential. Notably, these CSF1R+ myeloid-primed ProB cells are uniquely present in a narrow window of embryonic fetal liver hematopoiesis and do not persist in adult bone marrow. Moreover, analysis of CSF1R-deficient mice establishes a distinct role of CSF1R in fetal B-lymphopoiesis. CSF1R+ myeloid-primed embryonic ProB cells are relevant for infant and childhood PreB-ALLs, which frequently have a bi-phenotypic B-myeloid phenotype, and in which CSF1R-rearrangements have recently been reported. PMID:27207794

  8. Colony-stimulating factor-1 plays a major role in the development of reproductive function in male mice.

    PubMed

    Cohen, P E; Hardy, M P; Pollard, J W

    1997-10-01

    Colony-stimulating factor-1 (CSF-1) is the principal regulator of cells of the mononuclear phagocytic lineage that includes monocytes, tissue macrophages, microglia, and osteoclasts. Macrophages are found throughout the reproductive tract of both males and females and have been proposed to act as regulators of fertility at several levels. Mice homozygous for the osteopetrosis mutation (csfm[op]) lack CSF-1 and, consequently, have depleted macrophage numbers. Further analysis has revealed that male csfm(op)/csfm(op) mice have reduced mating ability, low sperm numbers, and 90% lower serum testosterone levels. The present studies show that this low serum testosterone is due to reduced testicular Leydig cell steroidogenesis associated with severe ultrastructural abnormalities characterized by disrupted intracellular membrane structures. In addition, the Leydig cells from csfm(op)/ csfm(op) males have diminished amounts of the steroidogenic enzyme proteins P450 side chain cleavage, 3beta-hydroxysteroid dehydrogenase, and P450 17alpha-hydroxylase-lyase, with associated reductions in the activity of all these steroidogenic enzymes, as well as in 17beta-hydroxysteroid dehydrogenase. The CSF-1-deficient males also have reduced serum LH and disruption of the normal testosterone negative feedback response of the hypothalamus, as demonstrated by the failure to increase LH secretion in castrated males and their lack of response to exogenous testosterone. However, these males are responsive to GnRH and LH treatment. These studies have identified a novel role for CSF-1 in the development and/or regulation of the male hypothalamic-pituitary-gonadal axis.

  9. Colony stimulating factor-1 receptor signaling networks inhibit mouse macrophage inflammatory responses by induction of microRNA-21

    PubMed Central

    Caescu, Cristina I.; Guo, Xingyi; Tesfa, Lydia; Bhagat, Tushar D.; Verma, Amit; Zheng, Deyou

    2015-01-01

    Macrophage polarization between the M2 (repair, protumorigenic) and M1 (inflammatory) phenotypes is seen as a continuum of states. The detailed transcriptional events and signals downstream of colony-stimulating factor 1 receptor (CSF-1R) that contributes to amplification of the M2 phenotype and suppression of the M1 phenotype are largely unknown. Macrophage CSF-1R pTyr-721 signaling promotes cell motility and enhancement of tumor cell invasion in vitro. Combining analysis of cellular systems for CSF-1R gain of function and loss of function with bioinformatic analysis of the macrophage CSF-1R pTyr-721–regulated transcriptome, we uncovered microRNA-21 (miR-21) as a downstream molecular switch controlling macrophage activation and identified extracellular signal-regulated kinase1/2 and nuclear factor-κB as CSF-1R pTyr-721–regulated signaling nodes. We show that CSF-1R pTyr-721 signaling suppresses the inflammatory phenotype, predominantly by induction of miR-21. Profiling of the miR-21–regulated messenger RNAs revealed that 80% of the CSF-1–regulated canonical miR-21 targets are proinflammatory molecules. Additionally, miR-21 positively regulates M2 marker expression. Moreover, miR-21 feeds back to positively regulate its own expression and to limit CSF-1R–mediated activation of extracellular signal-regulated kinase1/2 and nuclear factor-κB. Consistent with an anti-inflammatory role of miRNA-21, intraperitoneal injection of mice with a miRNA-21 inhibitor increases the recruitment of inflammatory monocytes and enhances the peritoneal monocyte/macrophage response to lipopolysaccharide. These results identify the CSF-1R–regulated miR-21 network that modulates macrophage polarization. PMID:25573988

  10. Macrophage colony-stimulating factor receptor marks and regulates a fetal myeloid-primed B-cell progenitor in mice.

    PubMed

    Zriwil, Alya; Böiers, Charlotta; Wittmann, Lilian; Green, Joanna C A; Woll, Petter S; Jacobsen, Sten Eirik W; Sitnicka, Ewa

    2016-07-14

    Although it is well established that unique B-cell lineages develop through distinct regulatory mechanisms during embryonic development, much less is understood about the differences between embryonic and adult B-cell progenitor cells, likely to underpin the genetics and biology of infant and childhood PreB acute lymphoblastic leukemia (PreB-ALL), initiated by distinct leukemia-initiating translocations during embryonic development. Herein, we establish that a distinct subset of the earliest CD19(+) B-cell progenitors emerging in the E13.5 mouse fetal liver express the colony-stimulating factor-1 receptor (CSF1R), previously thought to be expressed, and play a lineage-restricted role in development of myeloid lineages, and macrophages in particular. These early embryonic CSF1R(+)CD19(+) ProB cells also express multiple other myeloid genes and, in line with this, possess residual myeloid as well as B-cell, but not T-cell lineage potential. Notably, these CSF1R(+) myeloid-primed ProB cells are uniquely present in a narrow window of embryonic fetal liver hematopoiesis and do not persist in adult bone marrow. Moreover, analysis of CSF1R-deficient mice establishes a distinct role of CSF1R in fetal B-lymphopoiesis. CSF1R(+) myeloid-primed embryonic ProB cells are relevant for infant and childhood PreB-ALLs, which frequently have a bi-phenotypic B-myeloid phenotype, and in which CSF1R-rearrangements have recently been reported. PMID:27207794

  11. Seroprevalence of Antibodies That React with Anaplasma phagocytophila, the Agent of Human Granulocytic Ehrlichiosis, in Different Populations in Westchester County, New York

    PubMed Central

    Aguero-Rosenfeld, Maria E.; Donnarumma, Lorraine; Zentmaier, Lois; Jacob, Jobby; Frey, Michael; Noto, Richard; Carbonaro, Carol A.; Wormser, Gary P.

    2002-01-01

    We determined the frequencies of antibodies to Anaplasma phagocytophila, the agent of human granulocytic ehrlichiosis (HGE), in different groups of adults and children from Westchester County, New York. The groups included 159 adult blood donors and 215 children who were seronegative for Borrelia burgdorferi antibodies, 118 adult patients and 57 children who were seropositive for B. burgdorferi antibodies, and 42 adult patients with culture-confirmed erythema migrans. Eighteen (11.3%) of the blood donors and 11 (5.1%) of the B. burgdorferi-seronegative children were found to have A. phagocytophila antibodies by indirect immunofluorescent-antibody assay (IFA). Nine of 42 (21.4%) patients with culture-confirmed erythema migrans tested at the baseline visit, 42 of 118 (35.6%) adults, and 3 of 57 (5.3%) children whose sera were reactive for B. burgdorferi antibodies also tested positive for A. phagocytophila antibodies. The geometric mean titer ranged from 219 to 315 for all groups, and the differences in titers among the groups were not statistically significant. Only one-third of the healthy blood donors reactive by IFA were confirmed to be positive by immunoblotting. We conclude that a significant proportion of adults and children without clinical evidence of HGE will test positive for A. phagocytophila antibodies when the conventional cutoff titer of 80 is used in the IFA. This information must be considered in interpretation of test results. PMID:12089287

  12. Direct evaluation of radiation damage in human hematopoietic progenitor cells in vivo.

    PubMed

    Kyoizumi, S; McCune, J M; Namikawa, R

    1994-01-01

    We have developed techniques by which normal functional elements of human bone marrow can be implanted into immunodeficient C.B-17 scid/scid (SCID) mice. Afterward, long-term multilineage human hematopoiesis is sustained in vivo. We evaluated the effect of irradiation on the function of human bone marrow with this in vivo model. After whole-body X irradiation of the engrafted animals, it was determined that the D0 value of human committed progenitor cells within the human marrow was 1.00 +/- 0.09 (SEM) Gy for granulocyte-macrophage colony-forming units (CFU-GM) and 0.74 +/- 0.12 Gy for erythroid burst-forming units (BFU-E). The effects of irradiation on the hematopoietic elements were reduced when the radioprotective agent WR-2721 was administered prior to irradiation. After low-dose irradiation, recovery of human myelopoiesis was accelerated by treatment with human granulocyte colony-stimulating factor (G-CSF). This small animal model may prove amenable for the analysis of the risk of the exposure of humans to radiation as well as for the development of new modalities for the prevention and treatment of radiation-induced hematopoietic damage. PMID:7505456

  13. Serologic evidence of a natural infection of white-tailed deer with the agent of human granulocytic ehrlichiosis in Wisconsin and Maryland.

    PubMed

    Walls, J J; Asanovich, K M; Bakken, J S; Dumler, J S

    1998-11-01

    White-tailed deer participate in the maintenance of the Ixodes tick life cycle and are reservoirs for some tick-borne infectious agents. Deer may be useful sentinels for tick-transmitted agents, such as ehrlichiae. In order to determine whether white-tailed deer are markers of natural transmission or are reservoirs for the human granulocytic ehrlichiosis (HGE) agent, we performed indirect immunofluorescent-antibody (IFA) tests and immunoblotting with the HGE agent and Ehrlichia chaffeensis on sera from 43 and 294 deer captured in northwest Wisconsin during 1994 and 1995, respectively, and 12 deer from southern Maryland. According to IFA testing, 47% of 1994 Wisconsin sera, 60% of 1995 Wisconsin sera, and 25% of Maryland sera contained HGE agent antibodies. All IFA-positive deer sera tested reacted with the 44-kDa band which is unique to the Ehrlichia phagocytophila group. Serologic reactions to E. chaffeensis were detected by IFA testing in 15 of 337 (4%) Wisconsin deer and in 10 of 12 (83%) Maryland deer, while 60 and 80% of E. chaffeensis IFA-positive Wisconsin and Maryland deer sera, respectively, reacted with the E. chaffeensis 28- to 29-kDa antigens by immunoblotting. A total of 4% of deer from Wisconsin and 25% of deer from Maryland were found by IFA testing to have antibodies to both the HGE agent and E. chaffeensis; 75% of these were confirmed to contain E. chaffeensis antibodies by immunoblotting. These results suggest that white-tailed deer in diverse geographical regions of the United States are naturally infected with the HGE agent, E. chaffeensis, or both and that these animals, and potentially humans, are exposed to infected ticks at a high frequency in nature. PMID:9801331

  14. Engraftment and long-term expression of human fetal hemopoietic stem cells in sheep following transplantation in utero.

    PubMed Central

    Zanjani, E D; Pallavicini, M G; Ascensao, J L; Flake, A W; Langlois, R G; Reitsma, M; MacKintosh, F R; Stutes, D; Harrison, M R; Tavassoli, M

    1992-01-01

    Hemopoietic stem cells from human fetal liver were transplanted in utero into preimmune fetal sheep (48-54 days of gestation). The fate of donor cells was followed using karyotype analysis, by immunofluorescence labeling with anti-CD antibodies, and by fluorescent in situ hybridization using human-specific DNA probes. Engraftment occurred in 13 of 33 recipients. Of five live born sheep that exhibited chimerism, all expressed human cells in the marrow, whereas three expressed them in blood as well. Engraftment was multilineage (erythroid, myeloid, and lymphoid) and human hemopoietic progenitors (multipotent colony-forming units, colony-forming units-granulocyte, macrophage, and erythroid burst-forming units) capable of forming colonies in vitro were detected in all five lambs for greater than 2 yr. These progenitors responded to human-specific growth factors both in vitro and in vivo. Thus the administration of recombinant human IL-3 and granulocyte macrophage-colony-stimulating factor to chimeric sheep resulted in a 2.1-3.4-fold increase in the relative expression of donor (human) cells. These results demonstrate that the permissive environment of the preimmune fetal sheep provides suitable conditions for the engraftment and long-term multilineage expression of human hemopoietic stem cells in a large animal model. In this model, donor human cells appear to retain certain phenotypic and functional characteristics that can be used to manipulate the size of donor cell pool. PMID:1348253

  15. Constitutively expressed Siglec-9 inhibits LPS-induced CCR7, but enhances IL-4-induced CD200R expression in human macrophages.

    PubMed

    Higuchi, Hiroshi; Shoji, Toru; Iijima, Shinji; Nishijima, Ken-Ichi

    2016-06-01

    Siglecs recognize the sialic acid moiety and regulate various immune responses. In the present study, we compared the expression levels of Siglecs in human monocytes and macrophages using a quantitative real-time reverse transcription-polymerase chain reaction analysis. The differentiation of monocytes into macrophages by macrophage colony-stimulating factor or granulocyte macrophage colony-stimulating factor enhanced the expression of Siglec-7 and Siglec-9. The differentiated macrophages were stimulated by lipopolysaccharide (LPS) plus interferon (IFN)-γ or interleukin (IL)-4. The expression of Siglec-10 was enhanced by IL-4, whereas that of Siglec-7 was reduced by LPS plus IFN-γ. The expression of Siglec-9 was not affected by these stimuli. The knockdown of Siglec-9 enhanced the expression of CCR7 induced by the LPS or the LPS plus IFN-γ stimulation, and decreased the IL-4-induced expression of CD200R. These results suggest that Siglec-9 is one of the main Siglecs in human blood monocytes/macrophages and modulates innate immunity. PMID:26923638

  16. Direct evaluation of radiation damage in human hematopoietic progenitor cells in vivo

    SciTech Connect

    Kyoizumi, Seishi; McCune, J.M.; Namikawa, Reiko

    1994-01-01

    We have developed techniques by which normal functional elements of human bone marrow can be implanted into immunodeficient C.B-17 scid/scid (SCID) mice. Afterward, long-term multilineage human hematopoiesis is sustained in vivo. We evaluated the effect of irradiation on the function of human bone marrow with this in vivo model. After whole-body X irradiation of the engrafted animals, it was determined that the D{sub 0} value of human committed progenitor cells within the human marrow was 1.00 {+-} 0.09 (SEM) Gy for granulocyte-macrophage colony-forming units (CFU-GM) and 0.74 {+-} 0.12 Gy for erythroidburst-forming units (BFU-E). The effects of irradiation on the hematopoietic elements were reduced when the radioprotective agent WR-2721 was administered prior to irradiation. After low-dose irradiation, recovery of human granulocyte colony-stimulating factor (G-CSF). This small animal model may prove amenable for the analysis of the risk of the exposure of humans to irradiation as well as for the development of new modalities for the prevention and treatment of radiation-induced hematopoietic damage. 41 refs., 5 figs., 1 tab.

  17. Cell surface antigens detected on mature and leukemic granulocytic populations by cytotoxicity testing.

    PubMed

    Drew, S I; Carter, B M; Terasaki, P I; Naiem, F; Nathanson, D S; Abromowitz, B; Gale, R P

    1978-08-01

    Using a microcytotoxicity assay, the serological reactivity of human granulocytes, namely neutrophils and eosinophils, and chronic myeloid leukemia (CML) cells and cultured CML cell lines (K562, NALM-1) were examined. Mature granulocyte forms and cord granulocytes are readily lysed by specific granulocyte cytotoxins that do not react with random T and B lymphocytes, monocytes, red blood cells, or platelets. Furthermore, certain antisera were preferentially cytotoxic for eosinophil-enriched populations. Granulocytotoxin detected antigens on one of three CML blast cell populations tested and K562, but failed to react with NALM-1. By cytotoxicity, mature granulocytes were poor targets for B2-microglobulin and the appropriate HLA antisera although both sera types are absorbed with granulocytes. Furthermore, granulocytes did not possess B-lymphocytes (Ia-like) or blood group A, B, and Rh (D) antigens. Except for K562, both HLA and heterologous B-lymphocyte antisera were cytotoxic for the CML blast cell populations tested.

  18. Targeting the colony stimulating factor 1 receptor alleviates two forms of Charcot-Marie-Tooth disease in mice.

    PubMed

    Klein, Dennis; Patzkó, Ágnes; Schreiber, David; van Hauwermeiren, Anemoon; Baier, Michaela; Groh, Janos; West, Brian L; Martini, Rudolf

    2015-11-01

    See Scherer (doi:10.1093/awv279) for a scientific commentary on this article.Charcot-Marie-Tooth type 1 neuropathies are inherited disorders of the peripheral nervous system caused by mutations in Schwann cell-related genes. Typically, no causative cure is presently available. Previous preclinical data of our group highlight the low grade, secondary inflammation common to distinct Charcot-Marie-Tooth type 1 neuropathies as a disease amplifier. In the current study, we have tested one of several available clinical agents targeting macrophages through its inhibition of the colony stimulating factor 1 receptor (CSF1R). We here show that in two distinct mouse models of Charcot-Marie-Tooth type 1 neuropathies, the systemic short- and long-term inhibition of CSF1R by oral administration leads to a robust decline in nerve macrophage numbers by ∼70% and substantial reduction of the typical histopathological and functional alterations. Interestingly, in a model for the dominant X-linked form of Charcot-Marie-Tooth type 1 neuropathy, the second most common form of the inherited neuropathies, macrophage ablation favours maintenance of axonal integrity and axonal resprouting, leading to preserved muscle innervation, increased muscle action potential amplitudes and muscle strengths in the range of wild-type mice. In another model mimicking a mild, demyelination-related Charcot-Marie-Tooth type 1 neuropathy caused by reduced P0 (MPZ) gene dosage, macrophage blockade causes an improved preservation of myelin, increased muscle action potential amplitudes, improved nerve conduction velocities and ameliorated muscle strength. These observations suggest that disease-amplifying macrophages can produce multiple adverse effects in the affected nerves which likely funnel down to common clinical features. Surprisingly, treatment of mouse models mimicking Charcot-Marie-Tooth type 1A neuropathy also caused macrophage blockade, but did not result in neuropathic or clinical improvements

  19. New Insight into Atherosclerosis in Hemodialysis Patients: Overexpression of Scavenger Receptor and Macrophage Colony-Stimulating Factor Genes

    PubMed Central

    Nishida, Miki; Ando, Minoru; Iwamoto, Yusuke; Tsuchiya, Ken; Nitta, Kosaku

    2016-01-01

    Background Scavenger receptors (SRs) play a pivotal role in atherogenesis. The mechanism of atherosclerosis, which is specific to hemodialysis (HD) patients, was studied on the basis of SR gene expressions. Methods The gene expressions of SR type A (SR-A) and CD36 were studied in peripheral monocytes by real-time reverse transcription polymerase chain reaction. Data were compared between HD (n = 30) and age-matched control subjects (n = 10). Serum levels of macrophage colony-stimulating factor (M-CSF) were measured with enzyme-linked immunosorbent assay to test its role in SR expression. The statistical differences and associations between two continuous variables were assessed using the Mann-Whitney U test and Pearson's correlation coefficient, respectively. Results The relative quantities of SR mRNAs were significantly greater in HD patients than in controls [median (interquartile range): SR-A, 1.67 (0.96-2.76) vs. 0.90 (0.60-1.04), p = 0.0060; CD36, 1.09 (0.88-1.74) vs. 0.74 (0.64-0.99), p = 0.0255]. The serum concentration of M-CSF was significantly higher in HD patients than in controls [1, 121 (999-1,342) vs. 176 (155-202) pg/ml, p < 0.0001]. In addition, the relative quantity of M-CSF mRNA was significantly greater in HD patients than in controls [0.79 (0.42-1.53) vs. 0.42 (0.28-0.66), p = 0.0392]. The serum M-CSF levels were positively correlated with both the relative quantity of SR-A mRNA (r2 = 0.1681, p = 0.0086) and that of CD36 mRNA (r2 = 0.1202, p = 0.0284) in all subjects (n = 40). Conclusion HD patients are predisposed to atherosclerosis as a consequence of their enhanced monocyte SR expressions. SRs and M-CSF are potential therapeutic targets for atherosclerosis in this high-risk population. PMID:27721822

  20. Phosphatidylcholine hydrolysis and c-myc expression are in collaborating mitogenic pathways activated by colony-stimulating factor 1.

    PubMed

    Xu, X X; Tessner, T G; Rock, C O; Jackowski, S

    1993-03-01

    Stimulation of diglyceride production via phospholipase C (PLC) hydrolysis of phosphatidylcholine was an early event in the mitogenic action of colony-stimulating factor 1 (CSF-1) in the murine macrophage cell line BAC1.2F5 and was followed by a second phase of diglyceride production that persisted throughout the G1 phase of the cell cycle. Addition of phosphatidylcholine-specific PLC (PC-PLC) from Bacillus cereus to the medium of quiescent cells raised the intracellular diglyceride concentration and stimulated [3H]thymidine incorporation, although PC-PLC did not support continuous proliferation. PC-PLC treatment did not induce tyrosine phosphorylation or turnover of the CSF-1 receptor. The major protein kinase C (PKC) isotype in BAC1.2F5 cells was PKC-delta. Diglyceride production from PC-PLC did not target PKC-delta, since unlike phorbol esters, PC-PLC treatment neither decreased the electrophoretic mobility of PKC-delta nor increased the amount of GTP bound to Ras, and PC-PLC was mitogenically active in BAC1.2F5 cells in which PKC-delta was downregulated by prolonged treatment with phorbol ester. PC-PLC mimicked CSF-1 action by elevating c-fos and junB mRNAs to 40% of the level induced by CSF-1; however, PC-PLC induced c-myc mRNA to only 5% of the level in CSF-1-stimulated cells. PC-PLC addition to CSF-1-dependent BAC1.2F5 clones that constitutively express c-myc increased [3H]thymidine incorporation to 86% of the level evoked by CSF-1 and supported slow growth in the absence of CSF-1. Therefore, PC-PLC is a component of a signal transduction pathway leading to transcription of c-fos and junB that collaborates with c-myc and is independent of PKC-delta and Ras activation.

  1. rhCSF3 accelerates the proliferation of human melanocytes in culture through binding CSF3R and the expression of CSF3R transcripts.

    PubMed

    Lu, Yan; Guo, Ze; Zhou, Mei-Hua; Li, Xue; Sun, Jie; Gong, Qing-Li; Zhu, Wen-Yuan

    2015-05-01

    Melanogenic paracrine and autocrine cytokine networks have recently been discovered in vitro between melanocytes and other types of skin cells. Granulocyte colony-stimulating factor receptor (CSF3R) controls the survival, proliferation and differentiation of many kinds of cells, including neutrophils. To understand the function of CSF3R and recombinant human granulocyte colony-stimulating factor (rhCSF3) on melanocyte proliferation, this study compared the expression of CSF3R and the effects of rhCSF3 in primary human melanocytes, neutrophils and HEL 92.1.7 cells. The results show that CSF3R is localized in the cytoplasm and on cell membranes of melanocytes and neutrophils. The percentage of CSF3R(+) melanocytes was higher than CSF3R(+) HEL 92.1.7 cells, but was lower than CSF3R(+) neutrophils. Both CSF3R mRNA and CSF3R protein levels in melanocytes were higher than in HEL 92.1.7 cells, but were lower than in neutrophils. Treatment with rhCSF3 increased the proliferation of human melanocytes, but not their tyrosinase activity. Transcripts of CSF3R in human melanocytes, M14, A375 melanoma and A431 squamous cell carcinoma cells were also detected. Expression of the CSF3R V3 transcript was lower in melanocytes than in M14, A375 melanoma and A431 squamous cell carcinoma cells. In conclusion, rhCSF3 can promote melanocyte proliferation through CSF3R without affecting tyrosinase activity.

  2. Constitutive production of multiple cytokines and a human chorionic gonadotrophin beta-subunit by a human bladder cancer cell line (KU-19-19): possible demonstration of totipotential differentiation.

    PubMed Central

    Tachibana, M.; Miyakawa, A.; Nakashima, J.; Murai, M.; Nakamura, K.; Kubo, A.; Hata, J. I.

    1997-01-01

    Bladder cancer cells have been shown to secrete a variety of factors that are not related to cells of urothelial origin. The histogenesis of these tumour developments is uncertain, and a variety of theories have been previously reported. In the present manuscript, we identify the factors constitutively produced by a human bladder cancer cell line (KU-19-19) that was found to produce beta human chorionic gonadotrophin (beta-hCG), granulocyte colony-stimulating factor (G-CSF), granulocyte-macrophage colony-stimulating factor (GM-CSF), interleukin 1alpha (IL-1alpha), interleukin 6 (IL-6) and interleukin 8 (IL-8). The cells were obtained from a case of metastatic carcinoma that was originally diagnosed to be a grade 3 (WHO classification), invasive transitional cell carcinoma of the bladder. On microscopic observation, the cultured cells exhibited an epithelial appearance with vacuole formation in their cytoplasm. Ultrastructural observations revealed relatively marked microvilli and a tight junction. Significant amounts of beta-hCG, G-CSF, GM-CSF, IL-1alpha, IL-6 and IL-8 concentrations in the supernatant from cultured cells were demonstrated by enzyme-linked immunosorbent assays, while the expression of mRNA of these marker proteins in cancer cells was also significantly exhibited by reverse transcription polymerase chain reaction (RT-PCR). In addition, the expression of G-CSF receptor and IL-6 receptor mRNA was also shown by RT-PCR. Xenograft transplantability using nude mice was observed in association with the presence of severe neutrophilia in the peripheral blood. These results indicate that this cell line appears to be an effective model for the study of transitional cell carcinoma of the bladder with multipotent differentiation potentials. Images Figure 1 Figure 4 Figure 5 Figure 6 Figure 7 Figure 8 PMID:9231915

  3. Identification of CD13+CD36+ cells as a common progenitor for erythroid and myeloid lineages in human bone marrow

    PubMed Central

    Chen, Ling; Gao, Zhigang; Zhu, Jianqiong; Rodgers, Griffin P.

    2008-01-01

    Objective To identify bi-potential precursor cells of erythroid and myeloid development in human bone marrow. Materials and Methods Cells co-expressing CD13 and CD36 (CD13+CD36+) were investigated by analyzing cell surface marker expression during erythroid development (induced with a combination of cytokines plus erythropoietin [EPO]), or myeloid development (induced with the same cocktail of cytokines plus granulocyte-colony stimulating factor [G-CSF]) of bone marrow derived CD133 cells in liquid cultures. CD13+CD36+ subsets were also isolated on the 14th day of cultures and further evaluated for their hematopoietic clonogenic capacity in methylcellulose. Results Colony-forming analysis of sorted CD13+CD36+ cells of committed erythroid and myeloid lineages demonstrated that these cells were able to generate erythroid, granulocyte, and mixed erythroid –granulocyte colonies. In contrast, CD13+CD36− or CD13−CD36+ cells exclusively committed to granulocyte/monocyte or erythroid colonies, respectively, but failed to form mixed erythroid –granulocyte colonies; no colonies were detected in CD13−CD36− cells with lineage-supporting cytokines. In addition, our data confirmed that EPO induced both erythroid and myeloid commitment, while G-CSF only supported the differentiation of the myeloid lineage. Conclusions The present data identify some CD13+CD36+ cells as bi-potential precursors of erythroid and myeloid commitment in normal hematopoiesis. They provide a physiological explanation for the cell identification of myeloid and erythroid lineages observed in hematopoietic diseases. This unique fraction of CD13+CD36+ cells may be useful for further studies on regulating erythroid and myeloid differentiation during normal and malignant hematopoiesis. PMID:17588473

  4. Characterization of a distinct binding site for the prokaryotic chaperone, GroEL, on a human granulocyte ribonuclease.

    PubMed

    Rosenberg, H F; Ackerman, S J; Tenen, D G

    1993-02-25

    Although ribonucleases fold into correct tertiary conformation in vitro guided solely by information contained in the primary amino acid sequence (Sela, M., White, F. H., and Anfinsen, C. B. (1957) Science 124, 691-693), it is not clear whether folding of these proteins proceeds unassisted in a complex intracellular environment. We describe here the specific and high affinity binding of groEL, the prokaryotic homolog of the heat shock protein 60 family of molecular chaperones, to recombinant eosinophil cationic protein and eosinophil-derived neurotoxin, two members of the human ribonuclease gene family. We have determined that groEL binds to a unique peptide sequence near the amino terminus of nascent eosinophil cationic protein that includes the first of eight cysteine residues. This binding site functions independently and can confer groEL binding activity on an unrelated carrier protein. GroEL dissociates from the binding site upon addition of ATP and Mg2+; no other cations or cofactors are necessary. These findings suggest the possibility that interaction with a groEL-like molecular chaperone may be a requirement for correct folding and/or translocation of eukaryotic ribonucleases in vivo.

  5. Megakaryocyte growth and development factor is a potent growth factor for primitive hematopoietic progenitors in the human fetus.

    PubMed

    Muench, Marcus O; Bárcena, Alicia

    2004-06-01

    Megakaryocyte growth and development factor (MGDF), or thrombopoietin, has received considerable attention as a therapeutic agent for treating thrombocytopenia or for its use in the ex vivo culture of hematopoietic stem cells. MGDF is known to support the growth of a broad spectrum of hematopoietic precursors obtained from adult or neonatal tissues, but its effects on the growth of fetal progenitors and stem cells has not been studied. Human CD38(+)CD34(2+) progenitors and CD38(-)CD34(2+) cells, a population that contains stem cells, were isolated from midgestation liver and grown under defined conditions with MGDF and various cytokines known to support the growth of primitive hematopoietic precursors. In clonal assays of colony-forming cells (CFCs), MGDF supported the growth of 15-25% of candidate stem cells when combined with granulocyte colony-stimulating factor, granulocyte-macrophage colony-stimulating factor (GM-CSF), flk-2/flt3 ligand, or stem cell factor. MGDF was observed to strongly support the early stages of hematopoiesis and expansion of high proliferative potential CFCs. More mature progenitors were expanded nearly 78-fold in 1 wk of culture with MGDF+SCF+GM-CSF. MGDF alone was also found to support the short-term (2 d) survival of CD38(-)CD34(2+) high proliferative potential CFCs. The effects of MGDF were more modest on CD38(+)CD34(2+) progenitors with only additive increases in colony formation being observed. These findings suggest that MGDF administration in fetuses and neonates may strongly affect the growth and mobilization of primitive hematopoietic progenitors and that MGDF may find use in the ex vivo growth and expansion of fetal stem cells.

  6. Colony-Stimulating Factor-1-Responsive Macrophage Precursors Reside in the Amphibian (Xenopus laevis) Bone Marrow Rather than the Hematopoietic Sub-Capsular Liver

    PubMed Central

    Grayfer, Leon; Robert, Jacques

    2013-01-01

    Macrophage precursors originate from, and undergo lineage commitment within designated sites of hematopoiesis, such as the mammalian bone marrow. These cells subsequently differentiate in response to stimulation with macrophage colony-stimulating factor (CSF-1). The amphibian bone marrow, unlike that of mammals, has been overlooked as a source of leukocyte precursors in favor of the liver sub-capsular region, where hematopoiesis occurs in anurans. Here we report that the bone marrow rather than the liver periphery provides macrophage progenitors to the amphibian Xenopus laevis. We identified the amphibian CSF-1, examined its gene expression in developing and virally infected X. laevis and produce it in recombinant form (rXlCSF-1). This rXlCSF-1 did not bind or elicit proliferation/differentiation of sub-cortical liver cells. Surprisingly, a sub-population of bone marrow cells engaged this growth factor and formed rXlCSF-1-concentration-dependant colonies in semi-solid medium. Furthermore, rXlCSF-1-treated bone marrow (but not liver) cultures comprised of cells with characteristic macrophage morphology and high gene expression of the macrophage marker, colony stimulating factor-1 receptor (CSF-1R). Together, our findings indicate that in contrast to all other vertebrates studied to date, Xenopus committed macrophage precursors populations are not present in the central site of hematopoiesis, but reside in the bone marrow. PMID:23485675

  7. Regulation of the colony-stimulating activity produced by a murine marrow-derived cell line (H-1)

    SciTech Connect

    Garnett, H.M.; Cronkite, E.P.; Harigaya, K.

    1982-03-01

    The production of molecular species that stimulate growth of granulocyte or macrophage colonies (GM-CSF) by the fibroblastoid H-1 cell line is unaffected by either native or iron-saturated lactoferrin, although some inhibition is detected with 10 ..mu..M prostaglandin E/sub 1/. The H-1 GM-CSF is able to support the formation of macrophage, neutrophil, and mixed colonies. This inhibitory effect is observed irrespective of the presence of an additional agar layer between the feeder cells and plated bone marrow cells, implying that diffusable substances are involved. Addition of indomethacin (10 ..mu..M) to feeder layers derived from 2.5 x 10/sup 3/ H-1 cells increases the number of GM-CFU/sub c/ detected to 50% of that seen with conditioned medium alone. This result suggests that released prostaglandin may be responsible for some, but not all, of the observed inhibition of colony formation. In the presence of the H-1 feeder layers, only macrophage colonies are detected and hence it appears that the H-1 cells produce, in addition to prostaglandin, a diffusible inhibitory substance that preferentially inhibits granulopoiesis.

  8. Giardia duodenalis Infection Reduces Granulocyte Infiltration in an In Vivo Model of Bacterial Toxin-Induced Colitis and Attenuates Inflammation in Human Intestinal Tissue

    PubMed Central

    Cotton, James A.; Motta, Jean-Paul; Schenck, L. Patrick; Hirota, Simon A.; Beck, Paul L.; Buret, Andre G.

    2014-01-01

    Giardia duodenalis (syn. G. intestinalis, G. lamblia) is a predominant cause of waterborne diarrheal disease that may lead to post-infectious functional gastrointestinal disorders. Although Giardia-infected individuals could carry as much as 106 trophozoites per centimetre of gut, their intestinal mucosa is devoid of overt signs of inflammation. Recent studies have shown that in endemic countries where bacterial infectious diseases are common, Giardia infections can protect against the development of diarrheal disease and fever. Conversely, separate observations have indicated Giardia infections may enhance the severity of diarrheal disease from a co-infecting pathogen. Polymorphonuclear leukocytes or neutrophils (PMNs) are granulocytic, innate immune cells characteristic of acute intestinal inflammatory responses against bacterial pathogens that contribute to the development of diarrheal disease following recruitment into intestinal tissues. Giardia cathepsin B cysteine proteases have been shown to attenuate PMN chemotaxis towards IL-8/CXCL8, suggesting Giardia targets PMN accumulation. However, the ability of Giardia infections to attenuate PMN accumulation in vivo and how in turn this effect may alter the host inflammatory response in the intestine has yet to be demonstrated. Herein, we report that Giardia infection attenuates granulocyte tissue infiltration induced by intra-rectal instillation of Clostridium difficile toxin A and B in an isolate-dependent manner. This attenuation of granulocyte infiltration into colonic tissues paralled decreased expression of several cytokines associated with the recruitment of PMNs. Giardia trophozoite isolates that attenuated granulocyte infiltration in vivo also decreased protein expression of cytokines released from inflamed mucosal biopsy tissues collected from patients with active Crohn’s disease, including several cytokines associated with PMN recruitment. These results demonstrate for the first time that certain

  9. Comparison of the nucleotide sequences of 16S rRNA, 444 Ep-ank, and groESL heat shock operon genes in naturally occurring Ehrlichia equi and human granulocytic ehrlichiosis agent isolates from Northern California.

    PubMed

    Chae, J S; Foley, J E; Dumler, J S; Madigan, J E

    2000-04-01

    We examined 11 naturally occurring isolates of Ehrlichia equi in horses and two human granulocytic ehrlichiosis agent isolates in California for sequence diversity in three genes. Ehrlichia equi isolates were from Sierra (n = 6), Mendocino (n = 3), Sonoma (n = 1), and Marin (n = 1) counties, and human granulocytic ehrlichiosis (HGE) agent isolates were obtained from Humboldt county. PCR with specific primers for 16S rRNA, 444 Ep-ank and groESL heat shock operon genes successfully produced amplicons for all 13 clinical samples. The 444 Ep-ank gene of the HGE agent and E. equi isolates from northern California is different from the eastern U.S. isolates BDS and USG3. The translated amino acid sequence of the groESL heat shock operon gene fragment is identical among E. equi, the HGE agent, and E. phagocytophila, with the exception of the northern Californian equine CASOLJ isolate. Microheterogeneity was observed in the 16S rRNA gene sequences of HGE agent and E. equi isolates from northern California. These results suggest that E. equi and the HGE agent found in California are similar or identical but may differ from the isolates of equine and human origin found in the eastern United States. PMID:10747108

  10. Adenosine potentiates stimulatory effects on granulocyte-macrophage hematopoietic progenitor cells in vitro of IL-3 and SCF, but not those of G-CSF, GM-CSF and IL-11.

    PubMed

    Hofer, M; Vacek, A; Pospísil, M; Weiterová, L; Holá, J; Streitová, D; Znojil, V

    2006-01-01

    The aim of the studies was to ascertain if adenosine is able to co-operate with selected hematopoietic growth factors and cytokines, namely with granulocyte colony-stimulating factor (G-CSF), granulocyte-macrophage colony-stimulating factor (GM-CSF), stem cell factor (SCF), interleukin-3 (IL-3), and interleukin-11 (IL-11), in inducing the growth of colonies from hematopoietic progenitor cells for granulocytes and macrophages (GM-CFC) from normal bone marrow cells in vitro. Adenosine was found not to produce any colonies when present in the cultures as the only potential stimulator. All the tested cytokines and growth factors were observed to induce the growth of distinct numbers of GM-CFC colonies, with the exception of IL-11. When suboptimal concentrations of the evaluated cytokines and growth factors were tested in the cultures in which various concentrations of adenosine were concomitantly present, mutually potentiating effects were found in the case of IL-3 and SCF. These results confirm the role of adenosine in regulation of granulopoiesis and predict IL-3 and SCF as candidates for further in vivo studies of their combined administration with adenosine.

  11. T(H)2 cytokines modulate the IL-9R expression on human neutrophils.

    PubMed

    Dragon, Stéphane; Takhar, Manrit Kaur; Shan, Lianyu; Hayglass, Kent T; Simons, F Estelle; Gounni, Abdelilah S

    2009-06-26

    Interleukin (IL)-9 is associated with key pathological features of asthma such as airway hyperresponsiveness, bronchoconstriction and mucus production. Inflammatory responses mediated by IL-9 rely on the expression of the IL-9R which has been reported on lung epithelial cells, T lymphocytes and recently on airway granulocyte infiltrates. In this study, we assessed the regulatory and constitutive cell surface expression of the IL-9Ralpha in unfractionated and purified human neutrophils from atopic asthmatics, atopic non-asthmatics and healthy normal controls. We demonstrate that T(H)2 cytokines (IL-4 or IL-13) and granulocyte macrophage-colony stimulating factor (GM-CSF) up-regulated mRNA and cell surface expression levels of the IL-9Ralpha in primary human and HL-60 differentiated neutrophils. Pharmacological inhibition of NF-kappaB did not affect T(H)2-mediated IL-9Ralpha expression in human neutrophils although IFN-gamma and IL-10 down-regulated IL-9Ralpha expression when co-incubated with IL-4, IL-13 or GM-CSF. Collectively, our results reveal a regulatory function for IFN-gamma and IL-10 on modulating the inducible IL-9Ralpha expression levels on peripheral blood neutrophils by T(H)2 cytokines. PMID:19401191

  12. T(H)2 cytokines modulate the IL-9R expression on human neutrophils.

    PubMed

    Dragon, Stéphane; Takhar, Manrit Kaur; Shan, Lianyu; Hayglass, Kent T; Simons, F Estelle; Gounni, Abdelilah S

    2009-06-26

    Interleukin (IL)-9 is associated with key pathological features of asthma such as airway hyperresponsiveness, bronchoconstriction and mucus production. Inflammatory responses mediated by IL-9 rely on the expression of the IL-9R which has been reported on lung epithelial cells, T lymphocytes and recently on airway granulocyte infiltrates. In this study, we assessed the regulatory and constitutive cell surface expression of the IL-9Ralpha in unfractionated and purified human neutrophils from atopic asthmatics, atopic non-asthmatics and healthy normal controls. We demonstrate that T(H)2 cytokines (IL-4 or IL-13) and granulocyte macrophage-colony stimulating factor (GM-CSF) up-regulated mRNA and cell surface expression levels of the IL-9Ralpha in primary human and HL-60 differentiated neutrophils. Pharmacological inhibition of NF-kappaB did not affect T(H)2-mediated IL-9Ralpha expression in human neutrophils although IFN-gamma and IL-10 down-regulated IL-9Ralpha expression when co-incubated with IL-4, IL-13 or GM-CSF. Collectively, our results reveal a regulatory function for IFN-gamma and IL-10 on modulating the inducible IL-9Ralpha expression levels on peripheral blood neutrophils by T(H)2 cytokines.

  13. Canine granulocytic anaplasmosis: a review.

    PubMed

    Carrade, D D; Foley, J E; Borjesson, D L; Sykes, J E

    2009-01-01

    Anaplasma phagocytophilum is an emerging pathogen of humans, horses, and dogs worldwide that is transmitted by Ixodid ticks and maintained in a variety of small wild mammal species. Recent studies suggest that multiple strains of A. phagocytophilum may be circulating in wild and domestic animal populations, and these strains may have differential host tropisms and pathogenicity. The organism infects and survives within neutrophils by disabling key neutrophil functions, including neutrophil motility, phagocytosis, the oxidative burst mechanism, and neutrophil-endothelial cell interactions, as well as interfering with neutrophil apoptosis. Coinfections with other tick-borne pathogens may occur, especially Borrelia burgdorferi. A. phagocytophilum causes an acute febrile illness in dogs with lethargy and inappetence. Less frequent signs include lameness, coughing, polydipsia, intermittent vomiting, and hemorrhages. Diagnosis is based on finding morulae within granulocytes in the peripheral blood, the combination of acute and convalescent serology using immunofluorescent antibody techniques, and detection of the DNA of A. phagocytophilum using specific polymerase chain reaction assays. Whether persistent infection or reinfection with A. phagocytophilum occurs after natural infection requires additional study, with most reports suggesting that anaplasmosis is a self-limiting disease in dogs that responds well to a 2-week course of doxycycline therapy.

  14. Impaired killing of Candida albicans by granulocytes mobilized for transfusion purposes: a role for granule components

    PubMed Central

    Gazendam, Roel P.; van de Geer, Annemarie; van Hamme, John L.; Tool, Anton T.J.; van Rees, Dieke J.; Aarts, Cathelijn E.M.; van den Biggelaar, Maartje; van Alphen, Floris; Verkuijlen, Paul; Meijer, Alexander B.; Janssen, Hans; Roos, Dirk; van den Berg, Timo K.; Kuijpers, Taco W.

    2016-01-01

    Granulocyte transfusions are used to treat neutropenic patients with life-threatening bacterial or fungal infections that do not respond to anti-microbial drugs. Donor neutrophils that have been mobilized with granulocyte-colony stimulating factor (G-CSF) and dexamethasone are functional in terms of antibacterial activity, but less is known about their fungal killing capacity. We investigated the neutrophil-mediated cytotoxic response against C. albicans and A. fumigatus in detail. Whereas G-CSF/dexamethasone-mobilized neutrophils appeared less mature as compared to neutrophils from untreated controls, these cells exhibited normal ROS production by the NADPH oxidase system and an unaltered granule mobilization capacity upon stimulation. G-CSF/dexamethasone-mobilized neutrophils efficiently inhibited A. fumigatus germination and killed Aspergillus and Candida hyphae, but the killing of C. albicans yeasts was distinctly impaired. Following normal Candida phagocytosis, analysis by mass spectrometry of purified phagosomes after fusion with granules demonstrated that major constituents of the antimicrobial granule components, including major basic protein (MBP), were reduced. Purified MBP showed candidacidal activity, and neutrophil-like Crisp-Cas9 NB4-KO-MBP differentiated into phagocytes were impaired in Candida killing. Together, these findings indicate that G-CSF/dexamethasone-mobilized neutrophils for transfusion purposes have a selectively impaired capacity to kill Candida yeasts, as a consequence of an altered neutrophil granular content. PMID:26802050

  15. Impaired killing of Candida albicans by granulocytes mobilized for transfusion purposes: a role for granule components.

    PubMed

    Gazendam, Roel P; van de Geer, Annemarie; van Hamme, John L; Tool, Anton T J; van Rees, Dieke J; Aarts, Cathelijn E M; van den Biggelaar, Maartje; van Alphen, Floris; Verkuijlen, Paul; Meijer, Alexander B; Janssen, Hans; Roos, Dirk; van den Berg, Timo K; Kuijpers, Taco W

    2016-05-01

    Granulocyte transfusions are used to treat neutropenic patients with life-threatening bacterial or fungal infections that do not respond to anti-microbial drugs. Donor neutrophils that have been mobilized with granulocyte-colony stimulating factor (G-CSF) and dexamethasone are functional in terms of antibacterial activity, but less is known about their fungal killing capacity. We investigated the neutrophil-mediated cytotoxic response against C. albicans and A. fumigatus in detail. Whereas G-CSF/dexamethasone-mobilized neutrophils appeared less mature as compared to neutrophils from untreated controls, these cells exhibited normal ROS production by the NADPH oxidase system and an unaltered granule mobilization capacity upon stimulation. G-CSF/dexamethasone-mobilized neutrophils efficiently inhibited A. fumigatus germination and killed Aspergillus and Candida hyphae, but the killing of C. albicans yeasts was distinctly impaired. Following normal Candida phagocytosis, analysis by mass spectrometry of purified phagosomes after fusion with granules demonstrated that major constituents of the antimicrobial granule components, including major basic protein (MBP), were reduced. Purified MBP showed candidacidal activity, and neutrophil-like Crisp-Cas9 NB4-KO-MBP differentiated into phagocytes were impaired in Candida killing. Together, these findings indicate that G-CSF/dexamethasone-mobilized neutrophils for transfusion purposes have a selectively impaired capacity to kill Candida yeasts, as a consequence of an altered neutrophil granular content. PMID:26802050

  16. 2-(trimethylammonium) ethyl (R)-3-methoxy-3-oxo-2-stearamidopropyl phosphate suppresses osteoclast maturation and bone resorption by targeting macrophage-colony stimulating factor signaling.

    PubMed

    Park, So Jeong; Park, Doo Ri; Bhattarai, Deepak; Lee, Kyeong; Kim, Jaesang; Bae, Yun Soo; Lee, Soo Young

    2014-08-01

    2-(Trimethylammonium) ethyl (R)-3-methoxy-3-oxo-2-stearamidopropyl phosphate [(R)-TEMOSPho], a derivative of an organic chemical identified from a natural product library, promotes highly efficient megakaryopoiesis. Here, we show that (R)-TEMOSPho blocks osteoclast maturation from progenitor cells of hematopoietic origin, as well as blocking the resorptive function of mature osteoclasts. The inhibitory effect of (R)-TEMOSPho on osteoclasts was due to a disruption of the actin cytoskeleton, resulting from impaired downstream signaling of c-Fms, a receptor for macrophage-colony stimulating factor linked to c-Cbl, phosphoinositol-3-kinase (PI3K), Vav3, and Rac1. In addition, (R)-TEMOSPho blocked inflammation-induced bone destruction by reducing the numbers of osteoclasts produced in mice. Thus, (R)-TEMOSPho may represent a promising new class of antiresorptive drugs for the treatment of bone loss associated with increased osteoclast maturation and activity.

  17. Pyrido[2,3-d]pyrimidin-5-ones: A Novel Class of Antiinflammatory Macrophage Colony-Stimulating Factor-1 Receptor Inhibitors

    SciTech Connect

    Huang, Hui; Hutta, Daniel A.; Rinker, James M.; Hu, Huaping; Parsons, William H.; Schubert, Carsten; DesJarlais, Renee L.; Crysler, Carl S.; Chaikin, Margery A.; Donatelli, Robert R.; Chen, Yanmin; Cheng, Deping; Zhou, Zhao; Yurkow, Edward; Manthey, Carl L.; Player, Mark R.

    2010-10-01

    A series of pyrido[2,3-d]pyrimidin-5-ones has been synthesized and evaluated as inhibitors of the kinase domain of macrophage colony-stimulating factor-1 receptor (FMS). FMS inhibitors may be useful in treating rheumatoid arthritis and other chronic inflammatory diseases. Structure-based optimization of the lead amide analogue 10 led to hydroxamate analogue 37, which possessed excellent potency and an improved pharmacokinetic profile. During the chronic phase of streptococcal cell wall-induced arthritis in rats, compound 37 (10, 3, and 1 mg/kg) was highly effective at reversing established joint swelling. In an adjuvant-induced arthritis model in rats, 37 prevented joint swelling partially at 10 mg/kg. In this model, osteoclastogenesis and bone erosion were prevented by low doses (1 or 0.33 mg/kg) that had minimal impact on inflammation. These data underscore the potential of FMS inhibitors to prevent erosions and reduce symptoms in rheumatoid arthritis.

  18. Pyrido[2,3-d]pyrimidin-5-ones: a novel class of antiinflammatory macrophage colony-stimulating factor-1 receptor inhibitors.

    PubMed

    Huang, Hui; Hutta, Daniel A; Rinker, James M; Hu, Huaping; Parsons, William H; Schubert, Carsten; DesJarlais, Renee L; Crysler, Carl S; Chaikin, Margery A; Donatelli, Robert R; Chen, Yanmin; Cheng, Deping; Zhou, Zhao; Yurkow, Edward; Manthey, Carl L; Player, Mark R

    2009-02-26

    A series of pyrido[2,3-d]pyrimidin-5-ones has been synthesized and evaluated as inhibitors of the kinase domain of macrophage colony-stimulating factor-1 receptor (FMS). FMS inhibitors may be useful in treating rheumatoid arthritis and other chronic inflammatory diseases. Structure-based optimization of the lead amide analogue 10 led to hydroxamate analogue 37, which possessed excellent potency and an improved pharmacokinetic profile. During the chronic phase of streptococcal cell wall-induced arthritis in rats, compound 37 (10, 3, and 1 mg/kg) was highly effective at reversing established joint swelling. In an adjuvant-induced arthritis model in rats, 37 prevented joint swelling partially at 10 mg/kg. In this model, osteoclastogenesis and bone erosion were prevented by low doses (1 or 0.33 mg/kg) that had minimal impact on inflammation. These data underscore the potential of FMS inhibitors to prevent erosions and reduce symptoms in rheumatoid arthritis.

  19. Use of a sentinel host system to study the questing behavior of Ixodes spinipalpis and its role in the transmission of Borrelia bissettii, human granulocytic ehrlichiosis, and Babesia microti.

    PubMed

    Burkot, T R; Maupin, G O; Schneider, B S; Denatale, C; Happ, C M; Rutherford, J S; Zeidner, N S

    2001-10-01

    Ixodes spinipalpis maintains Borrelia bissettii spirochetes in Colorado in a cycle involving wood rats and deer mice. This tick has been described as nidicolous, remaining either attached to its rodent hosts or in the rodent nest. Nidicolous ticks pose little risk of pathogen transmission to humans if they do not actively quest for hosts. To investigate the questing potential of I. spinipalpis, sentinel mice were placed in an area where I. spinipalpis had been commonly found on wood rats and deer mice. Concurrently, wild rodent populations were trapped and analyzed for Lyme disease spirochetes, the agent of human granulocytic ehrlichiosis (aoHGE), and Babesia microti. A total of 122 I. spinipalpis larvae and 10 nymphs were found on 19% of 244 sentinel mice. In addition, 4 sentinel mice became infested with Malaraeus telchinus or Orchopeas neotomae fleas. Questing I. spinipalpis were positively associated with woody shrubs and negatively associated with sunny and grassy areas. Four sentinel mice became infected with aoHGE after having been fed upon only by I. spinipalpis larvae. One sentinel mouse became infected with B. bissettii after having an I. spinipalpis nymph feed on it, and one sentinel mouse became coinfected with aoHGE and B. bissettii after it was fed upon by a single I. spinipalpis nymph. These sentinel mouse conversions suggest the possibility that the aoHGE is transovarially transmitted by I. spinipalpis, and that I. spinipalpis is capable of simultaneously transmitting B. bissettii and the aoHGE. The findings that I. spinipalpis quest away from rodent nests and will attach to and infect sentinel mice may be of public health importance. It suggests the potential transmission of the agents of human granulocytic ehrlichiosis and Lyme disease to other hosts by I. spinipalpis, in regions of the western United States where Ixodes pacificus is not found. PMID:11693872

  20. Efficient retroviral transduction of human B-lymphoid and myeloid progenitors: marked inhibition of their growth by the Pax5 transgene

    PubMed Central

    Sekine, Rieko; Kitamura, Toshio; Tsuji, Takashi

    2008-01-01

    We applied a coculture system for the genetic manipulation of human B-lymphoid and myeloid progenitor cells using murine bone marrow stromal cell support, and investigated the effects of forced Pax5 expression in both cell types. Cytokine-stimulated cord blood CD34+ cells could be transduced at 85% efficiency and 95% cell viability by a single 24-h infection with RD114-pseudotyped retroviral vectors, produced by the packaging cell line Plat-F and bicistronic vector plasmids pMXs-Ig, pMYs-Ig, or pMCs-Ig, encoding EGFP. Infected CD34+ cells were seeded onto HESS-5 cells in the presence of stem cell factor and granulocyte colony-stimulating factor, allowing the extensive production of B progenitors and granulocytic cells. We examined the cell number and CD34, CD33, CD19, and CD20 lambda and kappa expressions by flow cytometry. Ectopic expression of Pax5 in CD34+ cells resulted in small myeloid progenitors coexpressing CD33 and CD19 and inhibited myeloid differentiation. After 6 weeks, the number of Pax5-transduced CD19+ cells was 40-fold lower than that of control cells. However, the expression of CD20 and the κ/λ chain on Pax5-transduced CD19+ cells suggests that the Pax5 transgene may not interfere with their differentiation. This report is the first to describe the effects of forced Pax5 expression in human hematopoietic progenitors. PMID:18415655

  1. Nested PCR Assay for Detection of Granulocytic Ehrlichiae

    PubMed Central

    Massung, Robert F.; Slater, Kim; Owens, Jessica H.; Nicholson, William L.; Mather, Thomas N.; Solberg, Victoria B.; Olson, James G.

    1998-01-01

    A sensitive and specific nested PCR assay was developed for the detection of granulocytic ehrlichiae. The assay amplifies the 16S rRNA gene and was used to examine acute-phase EDTA-blood and serum samples obtained from seven humans with clinical presentations compatible with human granulocytic ehrlichiosis. Five of the seven suspected cases were positive by the PCR assay using DNA extracted from whole blood as the template, compared with a serologic assay that identified only one positive sample. The PCR assay using DNA extracted from the corresponding serum samples as the template identified three positive samples. The sensitivity of the assay on human samples was examined, and the limit of detection was shown to be fewer than 2 copies of the 16S rRNA gene. The application of the assay to nonhuman samples demonstrated products amplified from template DNA extracted from Ixodes scapularis ticks collected in Rhode Island and from EDTA-blood specimens obtained from white-tailed deer in Maryland. All PCR products were sequenced and identified as specific to granulocytic ehrlichiae. A putative variant granulocytic ehrlichia 16S rRNA gene sequence was detected among products amplified from both the ticks and the deer blood specimens. PMID:9542943

  2. Inhibition by nonsteroidal antiinflammatory drugs of luminol-dependent human-granulocyte chemiluminescence and /sup 3/H FMLP binding. Effect of sulindac sulfide, indomethacin metabolite, and optical enantiomers (+) and (-) MK830

    SciTech Connect

    Van Dyke, K.; Peden, D.; Van Dyke, C.; Jones, G.; Castranova, V.; Ma, J.

    1982-03-01

    A system is described to evaluate for nonsteroidal antiinflammatory drugs by means of luminol-dependent human-granulocyte chemiluminescence (CL) is described. The CL is produced using either opsonized zymosan (yeast cells) or the soluble chemotactic peptide f-Met-Leu-Phe as the perturbant of the granulocyte membrane. Using either system, the following drug effects 2 x 10(-5) M were noted: only sulindac sulfide, and not sulindac sulfone or sulindac, displayed marked inhibition of chemiluminescence, following the in vivo data regarding inflammatory effects. The 5-OH indomethacin metabolite was likewise inactive as an inhibitor of CL mirroring in vivo effects. MK(+)410, MK(-)830 and MK835 all showed approximately 50% inhibition of CL, displaying deviation from in vivo data. MK(+)830 markedly stimulated CL, 4-6 times the control (without drug), which is clearly different from its enantiomer, MK(-)830. The reasons for this behavior are unclear. However, receptor binding studies with /sup 3/H FMLP were accomplished in the presence and absence of the various drugs at 2 x 10(-5) M that were effective inhibitors of chemiluminescence (CL). Indomethacin, MK(-)830 and MK(+)410 had equivalent percent control binding and percent control CL. Sulindac sulfide and MK(+)835 both had higher percent control binding than percent control CL, with MK(+)835 displaying apparent increased numbers of available receptors relative to control. MK(+)830, which produces large increases in CL, produced a minor effect on percent control binding. A direct relationship between binding and CL does not exist with each drug. Chemiluminescence is dependent on ion movement and oxidative metabolism and is a secondary event to agonist-receptor occupation.

  3. Efficacy and toxicity of decitabine versus CHG regimen (low-dose cytarabine, homoharringtonine and granulocyte colony-stimulating factor) in patients with higher risk myelodysplastic syndrome: a retrospective study.

    PubMed

    Wu, Lingyun; Li, Xiao; Chang, Chunkang; Xu, Feng; He, Qi; Wu, Dong; Zhang, Zheng; Su, Jiying; Zhou, Liyu; Song, Luxi; Chao, Xiao; Zhao, Youshan

    2016-01-01

    Decitabine and CHG regimen (low-dose cytarabine and homoharringtonine with G-CSF) have been used for treating higher risk myelodysplastic syndrome (MDS). In this study, we retrospectively compared the efficacy and toxicity of the two regimens in 132 MDS patients. Complete remission (CR) was not significantly different between the groups (27.1% with decitabine vs. 30.6% with CHG, p = 0.657). The CR rate with decitabine (58.8%) was significantly higher than that with CHG (7.7%) (p = 0.007) among the patients with poor karyotypes. Five of 23 (21.7%) patients who failed to respond to decitabine achieved CR with CHG, while one of two patients achieved CR with decitabine after failure with CHG. Overall and relapse-free survival were not different between the groups. In conclusion, both decitabine and CHG regimen are effective for higher risk MDS; there is no cross resistance between the regimens. Decitabine might be a better choice for patients with poor karyotypes.

  4. Prophylactic administration of vector-encoded porcine granulocyte-colony stimulating factor reduces Salmonella shedding,tonsil colonization,& microbiota alterations of the gastrointestinal tract in Salmonella-challenged swine

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Salmonella colonization of food animals is a concern for animal health and public health as a food safety risk. Various obstacles impede the effort to reduce asymptomatic Salmonella carriage in food animals, including the existence of numerous serovars and the ubiquitous nature of Salmonella. To d...

  5. THE IMPACT OF CONCURRENT GRANULOCYTE MACROPHAGE-COLONY STIMULATING FACTOR ON QUALITY OF LIFE IN HEAD AND NECK CANCER PATIENTS: RESULTS OF THE RANDOMIZED, PLACEBO-CONTROLED RADIATION THERAPY ONCOLOGY GROUP 9901 TRIAL

    PubMed Central

    Hoffman, Karen E.; Pugh, Stephanie; James, Jennifer L.; Scarantino, Charles; Movsas, Benjamin; Valicenti, Richard K.; Fortin, Andre; Pollock, JonDavid; Kim, Harold; Brachman, David G.; Berk, Lawrence B.; Bruner, Deborah Watkins; Kachnic, Lisa A.

    2015-01-01

    Purpose The Radiation Therapy Oncology Group (RTOG) conducted a randomized, placebo-controlled, trial evaluating the efficacy of GM-CSF in reducing mucosal injury and symptom burden from curative radiotherapy for head-and-neck (H&N) cancer. Methods Eligible patients with H&N cancer receiving radiation encompassing ≥ 50% of the oral cavity or oropharynx received subcutaneous GM-CSF or placebo. Quality of life (QoL) was assessed using the RTOG modified University of Washington H&N symptom questionnaire at baseline, 4, 13, 26 and 48 weeks from radiation initiation. Results Of 125 eligible patients, 114 were evaluable for QoL (58 GM-CSF, 56 placebo). Patient demographics, clinical characteristics, and baseline symptom scores were well balanced between the treatment arms. At the end of the acute period (13 weeks) patients in both arms reported negative change in total symptom score indicating increase in symptom burden relative to baseline (mean −18.4 GM-CSF, −20.8 placebo). There was no difference in change in total symptom score (p>0.05) or change in mucous, pain, eating, or activity domain scores (p>0.01) between patients in the GM-CSF and placebo arms. Analysis limited to patients treated per protocol or with an acceptable protocol deviation also found no difference in change in total symptom score (p>0.05) or change in domain scores (p>0.01) between treatment arms. Provider assessment of acute mucositis during treatment did not correlate with patient-reported mucous domain and total symptom scores (p>0.05) Conclusion GM-CSF administered concurrently during head-and-neck radiation does not appear to significantly improve patient-reported QoL symptom burden. PMID:24492945

  6. Ca2+ ionophore A23187-dependent stabilization of granulocyte-macrophage colony-stimulating factor messenger RNA in murine thymoma EL-4 cells is mediated through two distinct regions in the 3'-untranslated region.

    PubMed

    Iwai, Y; Akahane, K; Pluznik, D H; Cohen, R B

    1993-05-15

    We analyze the role of the Ca2+ ionophore A23187 in the induction of GM-CSF mRNA expression in EL-4 thymoma cells. Northern analysis shows that A23187 increases the half-life of GM-CSF mRNA. To identify potential Ca2+ response elements in the GM-CSF mRNA, we produced stable transfectants containing pRSV-CAT (EL-4cat) or hybrid constructs in which most of the GM-CSF 3'-untranslated region (EL-4gm) or the adenosine-uridine boxes alone (EL-4au) were placed in a downstream position from the CAT coding region. A23187 induces a 4.4-fold increase in CAT activity in EL-4cat cells and a 210-fold and 48-fold increase in CAT activity in EL-4gm and EL-4au cells, respectively. Actinomycin D chase experiments in transfected cells demonstrate that A23187 increases the half-life of CAT mRNA from 15 min to 3 h in EL-4au cells and more than 3 h in EL-4gm cells, suggesting that the effect of Ca2+ is mediated predominantly by the adenosine-uridine boxes with a smaller contribution from upstream regions. To map these upstream regions, we transfected cells with constructs containing mutations of the 3'-untranslated region. With two of these mutations, corresponding to a region located about 160 bases upstream of the adenosine-uridine boxes, CAT activity was induced only 50-fold compared to 200-fold in EL-4gm cells. These data indicate that two regions within the GM-CSF 3'-untranslated region interact to modulate Ca2+ effects on GM-CSF mRNA half-life. PMID:8482841

  7. Race and ethnicity influences collection of granulocyte colony-stimulating factor-mobilized peripheral blood progenitor cells from unrelated donors, a Center for International Blood and Marrow Transplant Research analysis.

    PubMed

    Hsu, Jack W; Wingard, John R; Logan, Brent R; Chitphakdithai, Pintip; Akpek, Gorgun; Anderlini, Paolo; Artz, Andrew S; Bredeson, Chris; Goldstein, Steven; Hale, Gregory; Hematti, Peiman; Joshi, Sarita; Kamble, Rammurti T; Lazarus, Hillard M; O'Donnell, Paul V; Pulsipher, Michael A; Savani, Bipin N; Schears, Raquel M; Shaw, Bronwen E; Confer, Dennis L

    2015-01-01

    Little information exists on the effect of race and ethnicity on collection of peripheral blood stem cells (PBSC) for allogeneic transplantation. We studied 10,776 donors from the National Marrow Donor Program who underwent PBSC collection from 2006 to 2012. Self-reported donor race/ethnic information included Caucasian, Hispanic, Black/African American (AA), Asian/Pacific Islander (API), and Native American (NA). All donors were mobilized with subcutaneous filgrastim at an approximate dose of 10 μg/kg/day for 5 days. Overall, AA donors had the highest median yields of mononuclear cells per liter and CD34(+) cells per liter of blood processed (3.1 × 10(9) and 44 × 10(6), respectively), whereas Caucasians had the lowest median yields at 2.8 × 10(9) and 33.7 × 10(6), respectively. Multivariate analysis of CD34(+) per liter mobilization yields using Caucasians as the comparator and controlling for age, gender, body mass index, and year of apheresis revealed increased yields in overweight and obese AA and API donors. In Hispanic donors, only male obese donors had higher CD34(+) per liter mobilization yields compared with Caucasian donors. No differences in CD34(+) per liter yields were seen between Caucasian and NA donors. Characterization of these differences may allow optimization of mobilization regimens to allow enhancement of mobilization yields without compromising donor safety.

  8. Biologic Effects of Anti-Granulocyte-Macrophage Colony-Stimulating Factor (GM-CSF) Antibody Formation in Patients Treated with GM-CSF (Sargramostim) as Adjuvant Therapy of Melanoma

    PubMed Central

    Spitler, Lynn E.; Cao, Huynh; Piironen, Timo; Whiteside, Theresa L.; Weber, Robert W.; Cruickshank, Scott

    2014-01-01

    OBJECTIVES We investigated the development of binding and neutralizing antibodies to GM-CSF in patients receiving prolonged therapy with GM-CSF as adjuvant therapy of melanoma and the impact of these antibodies on biologic effects. METHODS Fifty-three patients with high-risk melanoma which had been surgically excised were treated with GM-CSF, 125 µg/m2 daily for 14 days every 28 days for 1 year following surgical resection of disease. Serum samples for antibodies to GM-CSF were measured before treatment and on Study Days 155 and 351. Blood draws for testing biologic effects were keyed to GM-CSF administration: Days 0 (before), 15 (after 14 days on GM-CSF), 29 (after 14 days off GM-CSF), 155, and 351 (after 14 days on GM-CSF in the 6th and 13th cycle of treatment). RESULTS Of 53 patients enrolled, 43 were evaluable for the development of anti-GM-CSF antibodies. Of these, 93% developed binding antibodies and 42% developed both binding and neutralizing antibodies. The increase in the white blood cell (WBC) count, percent eosinophils, or neopterin levels engendered by GM-CSF administration, was abrogated or markedly decreased in patients with neutralizing antibodies but not in patients who developed only binding antibodies. CONCLUSIONS Ninety-three percent of patients with melanoma treated with GM-CSF as adjuvant therapy develop antibodies to GM-CSF. In those with neutralizing antibodies, a diminution of the biologic effects of GM-CSF was observed. The development of neutralizing antibodies might also abrogate the potential clinical benefit of this treatment and should be considered in the design of future clinical trials. PMID:25286079

  9. Human Olfactory Mucosa Multipotent Mesenchymal Stromal Cells Promote Survival, Proliferation, and Differentiation of Human Hematopoietic Cells

    PubMed Central

    Diaz-Solano, Dylana; Wittig, Olga; Ayala-Grosso, Carlos; Pieruzzini, Rosalinda

    2012-01-01

    Multipotent mesenchymal stromal cells (MSCs) from the human olfactory mucosa (OM) are cells that have been proposed as a niche for neural progenitors. OM-MSCs share phenotypic and functional properties with bone marrow (BM) MSCs, which constitute fundamental components of the hematopoietic niche. In this work, we investigated whether human OM-MSCs may promote the survival, proliferation, and differentiation of human hematopoietic stem cells (HSCs). For this purpose, human bone marrow cells (BMCs) were co-cultured with OM-MSCs in the absence of exogenous cytokines. At different intervals, nonadherent cells (NACs) were harvested from BMC/OM-MSC co-cultures, and examined for the expression of blood cell markers by flow cytometry. OM-MSCs supported the survival (cell viability >90%) and proliferation of BMCs, after 54 days of co-culture. At 20 days of co-culture, flow cytometric and microscopic analyses showed a high percentage (73%) of cells expressing the pan-leukocyte marker CD45, and the presence of cells of myeloid origin, including polymorphonuclear leukocytes, monocytes, basophils, eosinophils, erythroid cells, and megakaryocytes. Likewise, T (CD3), B (CD19), and NK (CD56/CD16) cells were detected in the NAC fraction. Colony-forming unit–granulocyte/macrophage (CFU-GM) progenitors and CD34+ cells were found, at 43 days of co-culture. Reverse transcriptase–polymerase chain reaction (RT-PCR) studies showed that OM-MSCs constitutively express early and late-acting hematopoietic cytokines (i.e., stem cell factor [SCF] and granulocyte- macrophage colony-stimulating factor [GM-CSF]). These results constitute the first evidence that OM-MSCs may provide an in vitro microenvironment for HSCs. The capacity of OM-MSCs to support the survival and differentiation of HSCs may be related with the capacity of OM-MSCs to produce hematopoietic cytokines. PMID:22471939

  10. Human olfactory mucosa multipotent mesenchymal stromal cells promote survival, proliferation, and differentiation of human hematopoietic cells.

    PubMed

    Diaz-Solano, Dylana; Wittig, Olga; Ayala-Grosso, Carlos; Pieruzzini, Rosalinda; Cardier, Jose E

    2012-11-20

    Multipotent mesenchymal stromal cells (MSCs) from the human olfactory mucosa (OM) are cells that have been proposed as a niche for neural progenitors. OM-MSCs share phenotypic and functional properties with bone marrow (BM) MSCs, which constitute fundamental components of the hematopoietic niche. In this work, we investigated whether human OM-MSCs may promote the survival, proliferation, and differentiation of human hematopoietic stem cells (HSCs). For this purpose, human bone marrow cells (BMCs) were co-cultured with OM-MSCs in the absence of exogenous cytokines. At different intervals, nonadherent cells (NACs) were harvested from BMC/OM-MSC co-cultures, and examined for the expression of blood cell markers by flow cytometry. OM-MSCs supported the survival (cell viability >90%) and proliferation of BMCs, after 54 days of co-culture. At 20 days of co-culture, flow cytometric and microscopic analyses showed a high percentage (73%) of cells expressing the pan-leukocyte marker CD45, and the presence of cells of myeloid origin, including polymorphonuclear leukocytes, monocytes, basophils, eosinophils, erythroid cells, and megakaryocytes. Likewise, T (CD3), B (CD19), and NK (CD56/CD16) cells were detected in the NAC fraction. Colony-forming unit-granulocyte/macrophage (CFU-GM) progenitors and CD34(+) cells were found, at 43 days of co-culture. Reverse transcriptase-polymerase chain reaction (RT-PCR) studies showed that OM-MSCs constitutively express early and late-acting hematopoietic cytokines (i.e., stem cell factor [SCF] and granulocyte- macrophage colony-stimulating factor [GM-CSF]). These results constitute the first evidence that OM-MSCs may provide an in vitro microenvironment for HSCs. The capacity of OM-MSCs to support the survival and differentiation of HSCs may be related with the capacity of OM-MSCs to produce hematopoietic cytokines.

  11. Ectopic expression of a chimeric colony-stimulating factor-1/TrkB-receptor promotes CSF-1-dependent survival of cultured sympathetic neurons.

    PubMed

    Erdmann, K S; Kaiser, A D; Klinz, F J; Zhong, J; Krautwald, S; Heumann, R

    1998-08-28

    The regulation of the density of innervation and the promotion of survival of neurons are the original effects depending on neurotrophins. Here we analyse such effects evoked by trkB tyrosine kinase in transfected PC12 cells and transfected sympathetic neurons. In order to exclude the previously described modulation of trk kinase activity by the extracellular activation of the low-affinity p75 neurotrophin receptor, we applied a chimeric receptor approach: The extracellular domain of colony-stimulating factor-1 (CSF-1) receptor was fused to the transmembrane and cytoplasmic domain of the trkB tyrosine kinase receptor, allowing its selective activation by the heterologous ligand. Protein expression and CSF-1-induced tyrosine phosphorylation of the chimeric receptor protein was demonstrated in transfected COS cells. After stable transfection into nerve growth factor (NGF)-responsive PC12 cells, CSF-1 mediated the K252a-sensitive induction of fiber outgrowth. Furthermore, we were able to show by heterologous expression of the chimeric receptor, that activation of trkB tyrosine kinase activity is sufficient to promote survival of neurotrophin deprived sympathetic neurons. PMID:9731232

  12. Defective Self-Renewal and Differentiation of GBA-Deficient Neural Stem Cells Can Be Restored By Macrophage Colony-Stimulating Factor

    PubMed Central

    Lee, Hyun; Bae, Jae-sung; Jin, Hee Kyung

    2015-01-01

    Gaucher disease (GD) is an autosomal recessive lysosomal storage disorder caused by mutations in the glucocerebrosidase gene (GBA), which encodes the lysosomal enzyme glucosylceramidase (GCase). Deficiency in GCase leads to characteristic visceral pathology and lethal neurological manifestations in some patients. Investigations into neurogenesis have suggested that neurodegenerative disorders, such as GD, could be overcome or at least ameliorated by the generation of new neurons. Bone marrow-derived mesenchymal stem cells (BM-MSCs) are potential candidates for use in the treatment of neurodegenerative disorders because of their ability to promote neurogenesis. Our objective was to examine the mechanism of neurogenesis by BM-MSCs in GD. We found that neural stem cells (NSCs) derived from a neuronopathic GD model exhibited decreased ability for self-renewal and neuronal differentiation. Co-culture of GBA-deficient NSCs with BM-MSCs resulted in an enhanced capacity for self-renewal, and an increased ability for differentiation into neurons or oligodendrocytes. Enhanced proliferation and neuronal differentiation of GBA-deficient NSCs was associated with elevated release of macrophage colony-stimulating factor (M-CSF) from BM-MSCs. Our findings suggest that soluble M-CSF derived from BM-MSCs can modulate GBA-deficient NSCs, resulting in their improved proliferation and neuronal differentiation. PMID:26282862

  13. Colony-stimulating factor 1 receptor signaling is necessary for microglia viability, unmasking a microglia progenitor cell in the adult brain.

    PubMed

    Elmore, Monica R P; Najafi, Allison R; Koike, Maya A; Dagher, Nabil N; Spangenberg, Elizabeth E; Rice, Rachel A; Kitazawa, Masashi; Matusow, Bernice; Nguyen, Hoa; West, Brian L; Green, Kim N

    2014-04-16

    The colony-stimulating factor 1 receptor (CSF1R) is a key regulator of myeloid lineage cells. Genetic loss of the CSF1R blocks the normal population of resident microglia in the brain that originates from the yolk sac during early development. However, the role of CSF1R signaling in microglial homeostasis in the adult brain is largely unknown. To this end, we tested the effects of selective CSF1R inhibitors on microglia in adult mice. Surprisingly, extensive treatment results in elimination of ∼99% of all microglia brain-wide, showing that microglia in the adult brain are physiologically dependent upon CSF1R signaling. Mice depleted of microglia show no behavioral or cognitive abnormalities, revealing that microglia are not necessary for these tasks. Finally, we discovered that the microglia-depleted brain completely repopulates with new microglia within 1 week of inhibitor cessation. Microglial repopulation throughout the CNS occurs through proliferation of nestin-positive cells that then differentiate into microglia.

  14. Colony-stimulating factor-1 receptor protein expression is a specific marker for goldfish (Carassius auratus L.) macrophage progenitors and their differentiated cell types.

    PubMed

    Katzenback, Barbara A; Belosevic, Miodrag

    2012-03-01

    Signaling through the colony-stimulating factor-1 receptor (CSF-1R) mediates the proliferation, differentiation, and activation of macrophages and their progenitors. In this study we report on the use of an anti-goldfish CSF-1R antibody to specifically recognize a population of CSF-1R positive cells from goldfish tissues. Furthermore, using our previously characterized primary kidney macrophage culture system, we show that CSF-1R positive cells include monocytes, macrophages, and their progenitor cells. Freshly isolated progenitor cells had a higher median florescent intensity ratio than those progenitor cells cultured for up to four days. The decrease in CSF-1R expression on the progenitor cells coincides with the appearance and development of monocytes and macrophages. Monocytes were consistently CSF-1R+ and maintained the high level of CSF-1R expression as they developed into macrophages. Like that of mammalian systems, CSF-1R is expressed on all macrophage sub-populations (progenitors, monocytes, macrophages), and CSF-1R expression increases with macrophage development in teleosts.

  15. Accumulation of cells expressing macrophage colony-stimulating factor receptor gene in the ovary of a pregnant viviparous fish, Neoditrema ransonnetii (Perciformes, Embiotocidae).

    PubMed

    Ueda, Kazuki; Saito, Erina; Iwasaki, Kaoru; Tsutsui, Shigeyuki; Nozawa, Aoi; Kikuchi, Kiyoshi; Nakamura, Osamu

    2016-03-01

    Macrophage colony-stimulating factor receptor (M-CSFR), a member of the group of type III protein tyrosine kinase receptors, is expressed primarily by monocyte/macrophage lineage cells. In order to describe the distribution of macrophages at the maternal-fetal interface in Neoditrema ransonnetii, a viviparous fish species, M-CSFR cDNA was sequenced. Two sequences were obtained: NrM-CSFR1 (4381 bp, encoding 980 amino acids), and NrM-CSFR2 (3573 bp, encoding 1016 amino acids). Both the genes were expressed in the ovary of pregnant females. In situ hybridization revealed that a number of cells that were positive for NrM-CSFR1 and/or NrM-CSFR2 populated the ovigerous lamellae of the ovary during pregnancy. Following parturition, M-CSFR-positive cells disappeared from the subepithelial region of ovigerous lamellae, and were localized in perivascular tissues. These results suggest the role of M-CSFR-positive cells, which appear to be macrophages, in N. ransonnetii during pregnancy.

  16. Tyrosines 559 and 807 in the cytoplasmic tail of the macrophage colony-stimulating factor receptor play distinct roles in osteoclast differentiation and function.

    PubMed

    Feng, Xu; Takeshita, Sunao; Namba, Noriyuki; Wei, Shi; Teitelbaum, Steven L; Ross, F Patrick

    2002-12-01

    Osteoclast (OC) differentiation requires that precursors, such as macrophage colony-stimulating factor (M-CSF)-dependent bone marrow macrophages, receive signals transduced by receptor activator of nuclear factor kappaB (RANK) and c-Fms, receptors for RANK ligand (RANKL) and M-CSF, respectively. Activated c-Fms autophosphorylates cytoplasmic tail tyrosine residues, which, by recruiting adaptor molecules, initiate specific signaling pathways. To identify which tyrosine residues are involved in c-Fms signaling in primary cells, we retrovirally transduced M-CSF-dependent bone marrow macrophages with a chimera comprising the external domain of the erythropoietin (Epo) receptor linked to the transmembrane and cytoplasmic domains of c-Fms. Transduced cells differentiate into bone-resorbing osteoclasts when treated with RANKL and either M-CSF or Epo, confirming that both endogenous and chimeric receptors transmit osteoclastogenic signals. Cells expressing chimeric receptors with Y(697)F, Y(706)F, Y(721)F, and Y(921)F single point mutations generate normal numbers of bone-resorbing OCs, with normal bone-resorbing activity when treated with RANKL and Epo. In contrast, those expressing Y(559)F generate fewer OCs, whereas theY807F mutant is incapable of osteoclastogenesis. Finally, although mature OCs expressing Y(559)F exhibit impaired bone resorption, those bearing Y807F do not. Thus, we have identified specific tyrosine residues in the cytoplasmic tail of c-Fms that are critical for transmitting M-CSF-initiated signals individually required for OC formation or function, respectively.

  17. An antibody against the colony-stimulating factor 1 receptor depletes the resident subset of monocytes and tissue- and tumor-associated macrophages but does not inhibit inflammation.

    PubMed

    MacDonald, Kelli P A; Palmer, James S; Cronau, Stephen; Seppanen, Elke; Olver, Stuart; Raffelt, Neil C; Kuns, Rachel; Pettit, Allison R; Clouston, Andrew; Wainwright, Brandon; Branstetter, Dan; Smith, Jeffrey; Paxton, Raymond J; Cerretti, Douglas Pat; Bonham, Lynn; Hill, Geoffrey R; Hume, David A

    2010-11-11

    The development of the mononuclear phagocyte system requires macrophage colony-stimulating factor (CSF-1) signaling through the CSF-1 receptor (CSF1R, CD115). We examined the effect of an antibody against CSF1R on macrophage homeostasis and function using the MacGreen transgenic mouse (csf1r-enhanced green fluorescent protein) as a reporter. The administration of a novel CSF1R blocking antibody selectively reduced the CD115(+)Gr-1(neg) monocyte precursor of resident tissue macrophages. CD115(+)Gr-1(+) inflammatory monocytes were correspondingly increased, supporting the view that monocytes are a developmental series. Within tissue, the antibody almost completely depleted resident macrophage populations in the peritoneum, gastrointestinal tract, liver, kidney, and skin, but not in the lung or female reproductive organs. CSF1R blockade reduced the numbers of tumor-associated macrophages in syngeneic tumor models, suggesting that these cells are resident type macrophages. Conversely, it had no effect on inflammatory monocyte recruitment in models, including lipopolysaccharide-induced lung inflammation, wound healing, peritonitis, and severe acute graft-versus-host disease. Depletion of resident tissue macrophages from bone marrow transplantation recipients actually resulted in accelerated pathology and exaggerated donor T-cell activation. The data indicate that CSF1R signaling is required only for the maturation and replacement of resident-type monocytes and tissue macrophages, and is not required for monocyte production or inflammatory function.

  18. Use of mice tolerant to lipopolysaccharide to demonstrate requirement of cooperation between macrophages and lymphocytes to generate lipopolysaccharide-induced colony-stimulating factor in vivo.

    PubMed Central

    Williams, Z; Hertogs, C F; Pluznik, D H

    1983-01-01

    Injection of lipopolysaccharide (LPS) into mice was followed by a rapid elevation of colony-stimulating factor (CSF) in the serum. A second, challenging injection of LPS given 3 to 4 days later failed to induce elevated levels of CSF in the serum. Such mice tolerant to LPS were used as an experimental tool to identify the CSF-producing cells which respond to LPS. We observed that generation of LPS-induced CSF in mice tolerant to LPS could be restored by an intraperitoneal injection of spleen cells 24 h before the challenging injection of LPS. Depletion of the adherent cells from the spleen cells reduced the ability of the splenic lymphocytes to restore the capacity of the mice tolerant to LPS to generate serum CSF. Reconstitution of the splenic lymphocytes with 5% thioglycolate-elicited peritoneal macrophages, however, reestablished the restorative capacity of these cells, whereas almost no restoration was observed after direct injection of elicited peritoneal macrophages. These data suggest that the spleen cells are active in generating CSF, provided that macrophages are present and can interact with the splenic lymphocytes to generate LPS-induced CSF in the serum. PMID:6602767

  19. Diagnostic Power of Vascular Endothelial Growth Factor and Macrophage Colony-Stimulating Factor in Breast Cancer Patients Based on ROC Analysis

    PubMed Central

    Głażewska, Edyta Katarzyna; Będkowska, Grażyna Ewa; Chorąży, Przemysław; Szmitkowski, Maciej; Ławicki, Sławomir

    2016-01-01

    Breast cancer (BC) is the most common malignancy in women. Vascular endothelial growth factor (VEGF) has been described as an important regulator of angiogenesis which plays a vital role in the progression of tumor. Macrophage colony-stimulating factor (M-CSF) is a cytokine whose functions include regulation of hematopoietic lineages cells growth, proliferation, and differentiation. We investigated the diagnostic significance of these parameters in comparison to CA15-3 in BC patients and in relation to the control group (benign breast tumor and healthy women). Plasma levels of the tested parameters were determined by ELISA and CA15-3 was determined by CMIA. VEGF was shown to be comparable to CA15-3 values of sensitivity in BC group and, what is more important, higher values in early stages of BC. VEGF was also the only parameter which has statistically significant AUC in all stages of cancer. M-CSF has been shown to be comparable to CA15-3 and VEGF, specificity, and AUC values only in stages III and IV of BC. These results indicate the usefulness and high diagnostic power of VEGF in the detection of BC. Also, it occurred to be the best candidate for cancer diagnostics in stages I and II of BC and in the differentiation between BC and benign cases. PMID:27445439

  20. Therapeutic applications of macrophage colony-stimulating factor-1 (CSF-1) and antagonists of CSF-1 receptor (CSF-1R) signaling.

    PubMed

    Hume, David A; MacDonald, Kelli P A

    2012-02-23

    Macrophage-colony stimulating factor (CSF-1) signaling through its receptor (CSF-1R) promotes the differentiation of myeloid progenitors into heterogeneous populations of monocytes, macrophages, dendritic cells, and bone-resorbing osteoclasts. In the periphery, CSF-1 regulates the migration, proliferation, function, and survival of macrophages, which function at multiple levels within the innate and adaptive immune systems. Macrophage populations elicited by CSF-1 are associated with, and exacerbate, a broad spectrum of pathologies, including cancer, inflammation, and bone disease. Conversely, macrophages can also contribute to immunosuppression, disease resolution, and tissue repair. Recombinant CSF-1, antibodies against the ligand and the receptor, and specific inhibitors of CSF-1R kinase activity have been each been tested in a range of animal models and in some cases, in patients. This review examines the potential clinical uses of modulators of the CSF-1/CSF-1R system. We conclude that CSF-1 promotes a resident-type macrophage phenotype. As a treatment, CSF-1 has therapeutic potential in tissue repair. Conversely, inhibition of CSF-1R is unlikely to be effective in inflammatory disease but may have utility in cancer.

  1. Macrophage colony-stimulating factor (CSF1) controls monocyte production and maturation and the steady-state size of the liver in pigs.

    PubMed

    Sauter, Kristin A; Waddell, Lindsey A; Lisowski, Zofia M; Young, Rachel; Lefevre, Lucas; Davis, Gemma M; Clohisey, Sara M; McCulloch, Mary; Magowan, Elizabeth; Mabbott, Neil A; Summers, Kim M; Hume, David A

    2016-09-01

    Macrophage colony-stimulating factor (CSF1) is an essential growth and differentiation factor for cells of the macrophage lineage. To explore the role of CSF1 in steady-state control of monocyte production and differentiation and tissue repair, we previously developed a bioactive protein with a longer half-life in circulation by fusing pig CSF1 with the Fc region of pig IgG1a. CSF1-Fc administration to pigs expanded progenitor pools in the marrow and selectively increased monocyte numbers and their expression of the maturation marker CD163. There was a rapid increase in the size of the liver, and extensive proliferation of hepatocytes associated with increased macrophage infiltration. Despite the large influx of macrophages, there was no evidence of liver injury and no increase in circulating liver enzymes. Microarray expression profiling of livers identified increased expression of macrophage markers, i.e., cytokines such as TNF, IL1, and IL6 known to influence hepatocyte proliferation, alongside cell cycle genes. The analysis also revealed selective enrichment of genes associated with portal, as opposed to centrilobular regions, as seen in hepatic regeneration. Combined with earlier data from the mouse, this study supports the existence of a CSF1-dependent feedback loop, linking macrophages of the liver with bone marrow and blood monocytes, to mediate homeostatic control of the size of the liver. The results also provide evidence of safety and efficacy for possible clinical applications of CSF1-Fc.

  2. Macrophage colony-stimulating factor (CSF1) controls monocyte production and maturation and the steady-state size of the liver in pigs

    PubMed Central

    Sauter, Kristin A.; Waddell, Lindsey A.; Lisowski, Zofia M.; Young, Rachel; Lefevre, Lucas; Davis, Gemma M.; Clohisey, Sara M.; McCulloch, Mary; Magowan, Elizabeth; Mabbott, Neil A.; Summers, Kim M.

    2016-01-01

    Macrophage colony-stimulating factor (CSF1) is an essential growth and differentiation factor for cells of the macrophage lineage. To explore the role of CSF1 in steady-state control of monocyte production and differentiation and tissue repair, we previously developed a bioactive protein with a longer half-life in circulation by fusing pig CSF1 with the Fc region of pig IgG1a. CSF1-Fc administration to pigs expanded progenitor pools in the marrow and selectively increased monocyte numbers and their expression of the maturation marker CD163. There was a rapid increase in the size of the liver, and extensive proliferation of hepatocytes associated with increased macrophage infiltration. Despite the large influx of macrophages, there was no evidence of liver injury and no increase in circulating liver enzymes. Microarray expression profiling of livers identified increased expression of macrophage markers, i.e., cytokines such as TNF, IL1, and IL6 known to influence hepatocyte proliferation, alongside cell cycle genes. The analysis also revealed selective enrichment of genes associated with portal, as opposed to centrilobular regions, as seen in hepatic regeneration. Combined with earlier data from the mouse, this study supports the existence of a CSF1-dependent feedback loop, linking macrophages of the liver with bone marrow and blood monocytes, to mediate homeostatic control of the size of the liver. The results also provide evidence of safety and efficacy for possible clinical applications of CSF1-Fc. PMID:27445344

  3. Evaluation of radioimmunoassay and in vitro colony assay techniques for determination of colony-stimulating factor and inhibitory activity in murine serum and tissue

    SciTech Connect

    Boegel, F.; Waheed, A.; Shadduck, R.K.

    1981-12-01

    These studies have evaluated a newly developed radioimmunoassay (RIA) for the measurement of colony-stimulating factor (CSF) in murine serum and other biological fluids. The routine in vitro agar gel bioassay for CSF is influenced by high molecular weight serum lipoproteins and low molecular weight tissue-derived materials that are inhibitory to colony formation. Studies with normal serum revealed that in all cases, the levels of CSF detected by the RIA were equal to or greater than levels measured by the bioassay. Dose curves with varying quantities of serum had linear responses in the RIA but not the colony assay. Using Sephadex G-150 chromatography, the murine serum was separated into CSF active and CSF inhibitory peaks as determined by bioassay. Evaluation of these fractions by RIA indicated that the assay was unaffected by the serum inhibitors. Likewise, experiments with tissue lysates indicated that the RIA was not influenced by the low molecular weight tissue inhibitors. Instead, the radioimmunoassay revealed low levels of CSF that were not detectable by bioassay. These observations indicate that the RIA is superior to the bioassay for the estimation of CSF in murine serum and other biologic materials.

  4. Comparison of human fetal liver, umbilical cord blood, and adult blood hematopoietic stem cell engraftment in NOD-scid/gammac-/-, Balb/c-Rag1-/-gammac-/-, and C.B-17-scid/bg immunodeficient mice.

    PubMed

    Lepus, Christin M; Gibson, Thomas F; Gerber, Scott A; Kawikova, Ivana; Szczepanik, Marian; Hossain, Jaber; Ablamunits, Vitaly; Kirkiles-Smith, Nancy; Herold, Kevan C; Donis, Ruben O; Bothwell, Alfred L; Pober, Jordan S; Harding, Martha J

    2009-10-01

    Immunodeficient mice bearing components of a human immune system present a novel approach for studying human immune responses. We investigated the number, phenotype, developmental kinetics, and function of developing human immune cells following transfer of CD34(+) hematopoietic stem cell (HSC) preparations originating from second trimester human fetal liver (HFL), umbilical cord blood (UCB), or granulocyte colony-stimulating factor-mobilized adult blood (G-CSF-AB) delivered via intrahepatic injection into sublethally irradiated neonatal NOD-scid/gammac(-/-), Balb/c-Rag1(-/-)gammac(-/-), and C.B-17-scid/bg mice. HFL and UCB HSC provided the greatest number and breadth of developing cells. NOD-scid/gammac(-/-) and Balb/c-Rag1(-/-)gammac(-/-) harbored human B and dendritic cells as well as human platelets in peripheral blood, whereas NOD-scid/gammac(-/-) mice harbored higher levels of human T cells. NOD-scid/gammac(-/-) mice engrafted with HFL CD34(+) HSC demonstrated human immunological competence evidenced by white pulp expansion and increases in total human immunoglobulin following immunization with T-dependent antigens and delayed-type hypersensitivity-infiltrating leukocytes in response to antigenic challenge. In conclusion, we describe an encouraging base system for studying human hematopoietic lineage development and function utilizing human HFL or UCB HSC-engrafted NOD-scid/gammac(-/-) mice that is well suited for future studies toward the development of a fully competent humanized mouse model.

  5. Neurons and astroglia govern microglial endotoxin tolerance through macrophage colony-stimulating factor receptor-mediated ERK1/2 signals.

    PubMed

    Chu, Chun-Hsien; Wang, Shijun; Li, Chia-Ling; Chen, Shih-Heng; Hu, Chih-Fen; Chung, Yi-Lun; Chen, Shiou-Lan; Wang, Qingshan; Lu, Ru-Band; Gao, Hui-Ming; Hong, Jau-Shyong

    2016-07-01

    Endotoxin tolerance (ET) is a reduced responsiveness of innate immune cells like macrophages/monocytes to an endotoxin challenge following a previous encounter with the endotoxin. Although ET in peripheral systems has been well studied, little is known about ET in the brain. The present study showed that brain immune cells, microglia, being different from peripheral macrophages, displayed non-cell autonomous mechanisms in ET formation. Specifically, neurons and astroglia were indispensable for microglial ET. Macrophage colony-stimulating factor (M-CSF) secreted from these non-immune cells was essential for governing microglial ET. Neutralization of M-CSF deprived the neuron-glia conditioned medium of its ability to enable microglia to form ET when microglia encountered two lipopolysaccharide (LPS) treatments. Recombinant M-CSF protein rendered enriched microglia refractory to the second LPS challenge leading to microglial ET. Activation of microglial M-CSF receptor (M-CSFR; also known as CSF1R) and the downstream ERK1/2 signals was responsible for M-CSF-mediated microglial ET. Endotoxin-tolerant microglia in neuron-glia cultures displayed M2-like polarized phenotypes, as shown by upregulation of M2 marker Arg-1, elevated production of anti-inflammatory cytokine interleukin 10, and decreased secretion of pro-inflammatory mediators (tumor necrosis factor α, nitric oxide, prostaglandin E2 and interleukin 1β). Endotoxin-tolerant microglia protected neurons against LPS-elicited inflammatory insults, as shown by reduced neuronal damages in LPS pre-treatment group compared with the group without LPS pre-treatment. Moreover, while neurons and astroglia became injured during chronic neuroinflammation, microglia failed to form ET. Thus, this study identified a distinct non-cell autonomous mechanism of microglial ET. Interactions of M-CSF secreted by neurons and astroglia with microglial M-CSFR programed microglial ET. Loss of microglial ET could be an important

  6. Protein kinase A inhibition of macrophage maturation is accompanied by an increase in DNA methylation of the colony-stimulating factor 1 receptor gene.

    PubMed

    Zasłona, Zbigniew; Scruggs, Anne M; Peters-Golden, Marc; Huang, Steven K

    2016-10-01

    Macrophage colony-stimulating factor 1 (CSF-1) plays a critical role in the differentiation of mononuclear phagocytes from bone marrow precursors, and maturing monocytes and macrophages exhibit increased expression of the CSF-1 receptor, CSF1R. The expression of CSF1R is tightly regulated by transcription factors and epigenetic mechanisms. We previously showed that prostaglandin E2 and subsequent activation of protein kinase A (PKA) inhibited CSF1R expression and macrophage maturation. Here, we examine the DNA methylation changes that occur at the Csf1r locus during macrophage maturation in the presence or absence of activated PKA. Murine bone marrow cells were matured to macrophages by incubating cells with CSF-1-containing conditioned medium for up to 6 days in the presence or absence of the PKA agonist 6-bnz-cAMP. DNA methylation of Csf1r promoter and enhancer regions was assayed by bisulphite pyrosequencing. DNA methylation of Csf1r decreased during normal macrophage maturation in concert with an increase in Csf1r mRNA expression. Treatment with the PKA agonist inhibited Csf1r mRNA and protein expression, and increased DNA methylation at the Csf1r promoter. This was associated with decreased binding of the transcription factor PU.1 to the Csf1r promoter. Treatment with the PKA agonist inhibited the responsiveness of macrophages to CSF-1. Levels of endogenous PKA activity decreased during normal macrophage maturation, suggesting that attenuation of this signalling pathway contributes to the increase in CSF1R expression during macrophage maturation. Together, these results demonstrate that macrophage maturation is accompanied by Csf1r hypomethylation, and illustrates for the first time the ability of PKA to increase Csf1r DNA methylation.

  7. Regulation of pluripotency of inner cell mass and growth and differentiation of trophectoderm of the bovine embryo by colony stimulating factor 2.

    PubMed

    Dobbs, Kyle B; Khan, Firdous A; Sakatani, Miki; Moss, James I; Ozawa, Manabu; Ealy, Alan D; Hansen, Peter J

    2013-12-01

    Colony-stimulating factor 2 (CSF2) enhances competence of the bovine embryo to establish and maintain pregnancy after the embryo is transferred into a recipient. Mechanisms involved could include regulation of lineage commitment, growth, or differentiation of the inner cell mass (ICM) and trophectoderm (TE). Experiments were conducted to evaluate regulation by CSF2 of pluripotency of the ICM and differentiation and growth of the TE. Embryos were cultured with 10 ng/ml recombinant bovine CSF2 or a vehicle control from Days 5 to 7 or 6 to 8 postinsemination. CSF2 increased the number of putative zygotes that developed to blastocysts when the percent of embryos becoming blastocysts in the control group was low but decreased blastocyst yield when blastocyst development in controls was high. ICM isolated from blastocysts by lysing the trophectoderm using antibody and complement via immunosurgery were more likely to survive passage when cultured on mitomycin C-treated fetal fibroblasts if derived from blastocysts treated with CSF2 than if from control blastocysts. There was little effect of CSF2 on characteristics of TE outgrowths from blastocysts. The exception was a decrease in outgrowth size for embryos treated with CSF2 from Days 5 to 7 and an increase in expression of CDX2 when treatment was from Days 6 to 8. Expression of the receptor subunit gene CSF2RA increased from the zygote stage to the 9-16 cell stage before decreasing to the blastocyst stage. In contrast, CSF2RB was undetectable at all stages. In conclusion, CSF2 improves competence of the ICM to survive in a pluripotent state and alters TE outgrowths. Actions of CSF2 occur through a signaling pathway that is likely to be independent of CSF2RB.

  8. Neurons and astroglia govern microglial endotoxin tolerance through macrophage colony-stimulating factor receptor-mediated ERK1/2 signals

    PubMed Central

    Chu, Chun-Hsien; Wang, Shijun; Li, Chia-Ling; Chen, Shih-Heng; Hu, Chih-Fen; Chung, Yi-Lun; Chen, Shiou-Lan; Wang, Qingshan; Lu, Ru-Band; Gao, Hui-Ming; Hong, Jau-Shyong

    2016-01-01

    Endotoxin tolerance (ET) is a reduced responsiveness of innate immune cells like macrophages/monocytes to an endotoxin challenge following a previous encounter with the endotoxin. Although ET in peripheral systems has been well studied, little is known about ET in the brain. The present study showed that brain immune cells, microglia, being different from peripheral macrophages, displayed non-cell autonomous mechanisms in ET formation. Specifically, neurons and astroglia were indispensable for microglial ET. Macrophage colony-stimulating factor (M-CSF) secreted from these non-immune cells was essential for governing microglial ET. Neutralization of M-CSF deprived the neuron-glia conditioned medium of its ability to enable microglia to form ET when microglia encountered two lipopolysaccharide (LPS) treatments. Recombinant M-CSF protein rende