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Sample records for colony stimulating factors

  1. Granulocyte macrophage colony stimulating factor therapy for pulmonary alveolar proteinosis.

    PubMed

    Shende, Ruchira P; Sampat, Bhavin K; Prabhudesai, Pralhad; Kulkarni, Satish

    2013-03-01

    We report a case of 58 year old female diagnosed with Pulmonary Alveolar Proteinosis (PAP) with recurrence of PAP after 5 repeated whole lung lavage, responding to subcutaneous injections of Granulocyte Macrophage Colony Stimulating Factor therapy (GM-CSF). Thus indicating that GM-CSF therapy is a promising alternative in those requiring repeated whole lung lavage PMID:24475687

  2. Isolation of a murine osteoclast colony-stimulating factor.

    PubMed Central

    Lee, M Y; Eyre, D R; Osborne, W R

    1991-01-01

    Cultures of a cell line derived from a murine mammary carcinoma that induces hypercalcemia were examined for soluble products that could induce osteoclasts to differentiate from murine bone marrow cells. The serum-free culture supernatant of this cell line stimulated growth of colonies from bone marrow cells that exhibited tartrate-resistant acid phosphatase (TRAPase) activity. These TRAPase-positive cells demonstrated essential features of osteoclasts when cultured with mineralized bone or dentin. The culture period required for colony development and the frequency of colony-forming cells indicated that relatively primitive marrow progenitors were stimulated by a tumor-derived factor(s) to form immature osteoclasts. Other colony-stimulating factors (CSFs), including granulocyte CSF, macrophage CSF, granulocyte-macrophage CSF and interleukin 3, were ruled out as the source of the activity produced by the tumor cells. The biological activity was successfully purified by gel filtration chromatography and reverse-phase HPLC. By SDS/PAGE, the activity was traced to a protein of approximately 17 kDa. Functional and biochemical studies of the purified factor suggest that it is distinct from any known CSF of myeloid cells. This protein appears to be a CSF for the osteoclast lineage, osteoclast CSF (O-CSF). Images PMID:1924309

  3. 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

  4. 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

  5. Granulocyte colony-stimulating factor induces in vitro lymphangiogenesis

    SciTech Connect

    Lee, Ae Sin; Kim, Dal; Wagle, Susbin Raj; Lee, Jung Eun; Jung, Yu Jin; Kang, Kyung Pyo; Lee, Sik; Park, Sung Kwang; Kim, Won

    2013-07-12

    Highlights: •G-CSF induces tube formation, migration and proliferation of lymphatic cells. •G-CSF increases phosphorylation of MAPK and Akt in lymphatic endothelial cells. •MAPK and Akt pathways are linked to G-CSF-induced in vitro lymphangiogenesis. •G-CSF increases sprouting of a lymphatic ring. •G-CSF produces peritoneal lymphangiogenesis. -- Abstract: Granulocyte-colony stimulating factor (G-CSF) is reported to induce differentiation in cells of the monocyte lineage and angiogenesis in vascular endothelial cells, but its effects on lymphangiogenesis is uncertain. Here we examined the effects and the mechanisms of G-CSF-induced lymphangiogenesis using human lymphatic endothelial cells (hLECs). Our results showed that G-CSF induced capillary-like tube formation, migration and proliferation of hLECs in a dose- and time-dependent manner and enhanced sprouting of thoracic duct. G-CSF increased phosphorylation of Akt and ERK1/2 in hLECs. Supporting the observations, specific inhibitors of phosphatidylinositol 3′-kinase and MAPK suppressed the G-CSF-induced in vitro lymphangiogenesis and sprouting. Intraperitoneal administration of G-CSF to mice also stimulated peritoneal lymphangiogenesis. These findings suggest that G-CSF is a lymphangiogenic factor.

  6. Expression of granulocyte colony stimulating factor (GCSF) in Hansenula polymorpha

    PubMed Central

    Talebkhan, Yeganeh; Samadi, Tannaz; Samie, Armin; Barkhordari, Farzaneh; Azizi, Mohammad; Khalaj, Vahid; Mirabzadeh, Esmat

    2016-01-01

    Background and Objectives: During past decades Hansenula polymorpha has attracted global attention for the expression of recombinant proteins due to its high growth rate, minimal nutritional porequirements and use of methanol as a low cost inducer. Materials and Methods: The corresponding nucleotide sequences for the expression of heterologous genes in Hansenula poylmorpha were extracted and assembled in an E. coli vector. The constructed expression cassette included formate dehydrogenase promoter (pFMD), a secretory signal sequence, a multiple cloning site (MCS) and methanol oxidase (MOX) terminator. Zeocin resistance gene fragment and complete cDNA encoding granulocyte colony stimulating factor (GCSF) were cloned downstream of the expression cassette in-frame with signal sequence. Restriction mapping and sequence analysis confirmed the correct cloning procedures. Final vector was transformed into Hansenula and recombinant host was induced for the expression of GCSF protein by adding methanol. SDS-PAGE and immuno-blotting were performed to confirm the identity of r-GCSF. Results: The expression cassette containing gcsf gene (615bp) and zeocin resistance marker (sh-ble, 1200bp) was prepared and successfully transformed into competent Hansenula polymorpha cells via electroporation. Zeocin resistant colonies were selected and GCSF expression was induced in recombinant Hansenula transformants using 0.5% methanol and an approximately 19kDa protein was observed on SDS-PAGE. Western blot analysis using serum isolated from GCSF-treated rabbit confirmed the identity of the protein. Conclusions: Molecular studies confirmed the designed expression cassette containing gcsf gene along with pFMD and signal sequence. The expressed 19kDa protein also confirmed the ability of designed vector in expressing heterologous genes in Hansenula cells. PMID:27092221

  7. Colony-Stimulating Factor-1 Signaling Suppresses Renal Crystal Formation

    PubMed Central

    Taguchi, Kazumi; Kitamura, Hiroshi; Yasui, Takahiro; Naiki, Taku; Hamamoto, Shuzo; Ando, Ryosuke; Mizuno, Kentaro; Kawai, Noriyasu; Tozawa, Keiichi; Asano, Kenichi; Tanaka, Masato; Miyoshi, Ichiro; Kohri, Kenjiro

    2014-01-01

    We recently reported evidence suggesting that migrating macrophages (Mϕs) eliminate renal crystals in hyperoxaluric mice. Mϕs can be inflammatory (M1) or anti-inflammatory (M2), and colony-stimulating factor-1 (CSF-1) mediates polarization to the M2Mϕ phenotype. M2Mϕs promote renal tissue repair and regeneration, but it is not clear whether these cells are involved in suppressing renal crystal formation. We investigated the role of M2Mϕs in renal crystal formation during hyperoxaluria using CSF-1–deficient mice, which lack M2Mϕs. Compared with wild-type mice, CSF-1–deficient mice had significantly higher amounts of renal calcium oxalate crystal deposition. Treatment with recombinant human CSF-1 increased the expression of M2-related genes and markedly decreased the number of renal crystals in both CSF-1–deficient and wild-type mice. Flow cytometry of sorted renal Mϕs showed that CSF-1 deficiency resulted in a smaller population of CD11b+F4/80+CD163+CD206hi cells, which represent M2-like Mϕs. Additionally, transfusion of M2Mϕs into CSF-1–deficient mice suppressed renal crystal deposition. In vitro phagocytosis assays with calcium oxalate monohydrate crystals showed a higher rate of crystal phagocytosis by M2-polarized Mϕs than M1-polarized Mϕs or renal tubular cells. Gene array profiling showed that CSF-1 deficiency resulted in disordered M2- and stone-related gene expressions. Collectively, our results provide compelling evidence for a suppressive role of CSF-1 signaling in renal crystal formation. PMID:24578130

  8. 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

  9. 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.

  10. 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

  11. Vasculitis complicating granulocyte colony stimulating factor treatment of leukopenia and infection in Felty's syndrome.

    PubMed

    Farhey, Y D; Herman, J H

    1995-06-01

    Recombinant myeloid growth factors have been increasingly used in recent years to combat induced and disease associated neutropenia. Their application in the management of Felty's syndrome with intercurrent infection has raised concern that resultant neutrophilia and activation of a diverse array of polymorphonuclear cell functions may have an adverse effect on the rheumatoid disease process. We describe a patient with Felty's syndrome receiving short term treatment with recombinant human granulocyte colony stimulating factor (GCSF), who then developed acute renal failure in conjunction with leukocytoclastic vasculitis and presumptive gout. We address the issue of "adding fuel to the fire" and review reported implications of GCSF in induction of vasculitis. PMID:7545756

  12. Establishment of a cell line of gallbladder carcinoma (GBK-1) producing human colony stimulating factor.

    PubMed

    Egami, H; Sakamoto, K; Yoshimura, R; Kikuchi, H; Akagi, M

    1986-02-01

    A cell line designated GBK-1 was established from a patient with anaplastic carcinoma of the gallbladder, marked neutrophilia and fever, and has been propagated for the past 18 months. The cells grew as a monolayer sheet with a doubling time of 43 hr. The GBK-1 cells were of a pleomorphic polygonal epitheloid shape and they were transplantable into nude mice. Mice bearing the tumor developed marked granulocytosis. In the culture supernatant of GBK-1 cells, high colony stimulating factor (CSF) activity was evident with both human bone marrow cells and C57BL mouse bone marrow cells, and granulocytic colonies, macrophage colonies or granulocyte-macrophage mixed colonies were produced. The CSF activity was distributed in the molecular weight range of 25,000 to 60,000 with two distinct peaks at molecular weights of approximately 50,000 and 30,000. CSF activity was inactivated by heat treatment at 70 degrees for 30 min. GBK-1 is a new human cell line that produces heat-labile human GM-CSF. PMID:3082828

  13. Role of Stem Cell Factor and Granulocyte-Colony Stimulating Factor in Remodeling during Liver Regeneration

    PubMed Central

    Meng, Fanyin; Francis, Heather; Glaser, Shannon; Han, Yuyan; DeMorrow, Sharon; Stokes, Allison; Staloch, Dustin; Venter, Julie; White, Melanie; Ueno, Yoshiyuki; Reid, Lola M.; Alpini, Gianfranco

    2011-01-01

    Functional pluripotent characteristics have been observed in specific subpopulations of hepatic cells that express some of the known cholangiocyte markers. Although evidence indicates that specific cytokines, granulocyte-macrophage colony stimulating factors (GM-CSF) and stem cell factor (SCF) may be candidate treatments for liver injury, the role of these cytokines in intrahepatic biliary epithelium remodeling is unknown. Thus, our aim was to characterize the specific cytokines that regulate the remodeling potentials of cholangiocytes after 70% partial hepatectomy (PH). The expression of the cytokines and their downstream signaling molecules was studied in rats after 70% PH by immunoblots, and in small and large murine cholangiocyte cultures (SMCCs and LMCCs) by immunocytochemistry and real-time PCR. There was a significant and stable increase in SCF and GM-CSF levels until 7 days after PH. Real-time PCR analysis revealed significant increases of key remodeling molecules, such as S100A4 and miR-181b after SCF plus GM-CSF administration in SMCCs. SMCCs produced significant amounts of soluble and bound SCF and GM-CSF in response to TGF-β. When SMCCs were incubated with TGF-β plus anti–SCF and GM-CSF antibodies, there was a significant decrease in S100A4 expression. Furthermore, treatment of SMCCs with SCF + GM-CSF significantly increased matrix metalloproteinases (MMP-2 and MMP-9) mRNA as well as miR-181b expression along with a reduction of metalloproteinase inhibitor 3 (TIMP-3). The levels of MMP-2, MMP-9 and miR-181b were also up-regulated in rat liver and isolated cholangiocytes after PH. CONCLUSION Our data suggest that altered expression of SCF and GM-CSF following PH can contribute to biliary remodeling (for example post-transplantation) by functional deregulation of activity of key signaling intermediates involved in cell expansion and multipotent differentiation. PMID:21932404

  14. Neuroprotective Activities of Granulocyte-Macrophage Colony Stimulating Factor Following Controlled Cortical Impact

    PubMed Central

    Kelso, Matthew L.; Elliott, Bret R.; Haverland, Nicole A.; Mosley, R. Lee; Gendelman, Howard E.

    2014-01-01

    Neurodegeneration after traumatic brain injury (TBI) is facilitated by innate and adaptive immunity and can be harnessed to effect brain repair. In mice subjected to controlled cortical impact (CCI) we show that treatment with granulocyte macrophage colony stimulating factor (GM-CSF) affects regulatory T cell numbers coincident with decreased lesion volumes and increased cortical tissue sparing. This paralleled increases in neurofilament and diminished reactive microglial staining. Transcriptomic analysis showed that GM-CSF induces robust immune neuroprotective responses seven days following CCI. Together, these results support the therapeutic potential of GM-CSF for TBI. PMID:25468272

  15. Colony Stimulating Factor-1 exerts direct effects on the proliferation and invasiveness of endometrial epithelial cells

    PubMed Central

    Aligeti, Sabitha; Kirma, Nameer B.; Binkely, Peter A; Schenken, Robert S.; Tekmal, Rajeshwar Rao

    2011-01-01

    Although macrophage colony stimulating factor (CSF-1) has been suggested to play a role in part in maintaining the chronic inflammatory response in endometriosis, our previous data suggest that CSF-1 may also play a role in early endometriosis lesion formation. In the current studies, we have examined this possibility and have shown that CSF-1, in an autocrine fashion, has a direct effect on endometrial epithelial cell proliferation and attachment to peritoneal mesothelial cells, early steps in endometriosis lesion formation on the peritoneum. PMID:21481374

  16. Potentiation of photodynamic therapy by granulocyte-macrophage colony stimulating factor immunotherapy

    NASA Astrophysics Data System (ADS)

    Krosl, Gorazd; Korbelik, Mladen; Krosl, Jana; Dougherty, Graeme J.

    1995-03-01

    The murine squamous carcinoma cell line (SCCVII) was genetically engineered to produce high levels of granulocyte-macrophage colony stimulating factor (GM-CSF). Lethally irradiated GM-CSF producing cells were injected under the subcutaneously growing parental SCCVII tumor at various times before and/or after PDT. Even a single treatment with GM- CSF producing cells injected two days before PDT markedly enhanced the tumor cure rate when compared to the PDT treatment alone. Effective potentiation was observed with PDT mediated either by Photofrin or by benzoporphyrin derivative.

  17. Transcriptional mechanisms underlying sensitization of peripheral sensory neurons by Granulocyte-/Granulocyte-macrophage colony stimulating factors

    PubMed Central

    2013-01-01

    Background Cancer-associated pain is a major cause of poor quality of life in cancer patients and is frequently resistant to conventional therapy. Recent studies indicate that some hematopoietic growth factors, namely granulocyte macrophage colony stimulating factor (GMCSF) and granulocyte colony stimulating factor (GCSF), are abundantly released in the tumor microenvironment and play a key role in regulating tumor-nerve interactions and tumor-associated pain by activating receptors on dorsal root ganglion (DRG) neurons. Moreover, these hematopoietic factors have been highly implicated in postsurgical pain, inflammatory pain and osteoarthritic pain. However, the molecular mechanisms via which G-/GMCSF bring about nociceptive sensitization and elicit pain are not known. Results In order to elucidate G-/GMCSF mediated transcriptional changes in the sensory neurons, we performed a comprehensive, genome-wide analysis of changes in the transcriptome of DRG neurons brought about by exposure to GMCSF or GCSF. We present complete information on regulated genes and validated profiling analyses and report novel regulatory networks and interaction maps revealed by detailed bioinformatics analyses. Amongst these, we validate calpain 2, matrix metalloproteinase 9 (MMP9) and a RhoGTPase Rac1 as well as Tumor necrosis factor alpha (TNFα) as transcriptional targets of G-/GMCSF and demonstrate the importance of MMP9 and Rac1 in GMCSF-induced nociceptor sensitization. Conclusion With integrative approach of bioinformatics, in vivo pharmacology and behavioral analyses, our results not only indicate that transcriptional control by G-/GMCSF signaling regulates a variety of established pain modulators, but also uncover a large number of novel targets, paving the way for translational analyses in the context of pain disorders. PMID:24067145

  18. Development and characterization of antiserum to murine granulocyte-macrophage colony-stimulating factor

    SciTech Connect

    Mochizuki, D.Y.; Eisenman, J.R.; Conlon, P.J.; Park, L.S.; Urdal, D.L.

    1986-05-15

    The expression in yeast of a cDNA clone encoding murine granulocyte-macrophage colony-stimulating factor (GM-CSF) has made possible the purification of large quantities of this recombinant protein. Rabbits immunized with pure recombinant GM-CSF generated antibodies that were shown to be specific for both recombinant GM-CSF and GM-CSF isolated from natural sources. Other lymphokines, including IL 1..beta.., IL 2, IL 3, and recombinant human GM-CSF did not react with the antiserum. The antiserum together with recombinant GM-CSF that had been radiolabeled with /sup 125/I to high specific activity, formed the foundation for a rapid, sensitive, and quantitative radioimmunoassay specific for murine GM-CSF. Furthermore, the antiserum was found to inhibit the biologic activities of GM-CSF as measured in both a bone marrow proliferation assay and a colony assay, and thus should prove to be a useful reagent for dissecting the complex growth factor activities involved in murine hematopoiesis.

  19. Functional interaction between mutations in the granulocyte colony-stimulating factor receptor in severe congenital neutropenia.

    PubMed

    Ward, Alister C; Gits, Judith; Majeed, Fidel; Aprikyan, Andrew A; Lewis, Rowena S; O'Sullivan, Lynda A; Freedman, Melvin; Shigdar, Sarah; Touw, Ivo P; Dale, David C; Dror, Yigal

    2008-08-01

    Most severe congenital neutropenia (SCN) cases possess constitutive neutrophil elastase mutations; a smaller cohort has acquired mutations truncating the granulocyte colony-stimulating factor receptor (G-CSF-R). We have described a case with constitutive extracellular G-CSF-R mutation hyporesponsive to ligand. Here we report two independent acquired G-CSF-R truncation mutations and a novel constitutive neutrophil elastase mutation in this patient. Co-expression of a truncated receptor chain restored STAT5 signalling responses of the extracellular G-CSF-R mutant, while constitutively-active STAT5 enhanced its proliferative capacity. These data add to our knowledge of SCN and further highlight the importance of STAT5 in mediating proliferative responses to G-CSF. PMID:18513286

  20. 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

  1. 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.

  2. Applications of recombinant DNA technology in the production of glycosylated recombinant human granulocyte colony stimulating factor.

    PubMed

    Holloway, C J

    1994-01-01

    Lenograstim has been developed by recombinant DNA technology and is expressed in large-scale mammalian cell culture. It has been shown that lenograstim is indistinguishable in its physicochemical, structural and biological properties with respect to native granulocyte colony stimulating factor isolated from a human cell line. In particular, both the recombinant and natural proteins have identical amino acid sequences, contain the same intra-polypeptide chain disulphide bridges and exhibit the same posttranslational carbohydrate structures. Lenograstim is manufactured by expanding inoculum from vials of the Manufacturer's Working Cell Bank (from molecular cloning) followed by culture in a large bioreactor. Purification of lenograstim involves a four-step chromatographic process. The active ingredient is monitored by in-process controls at all stages of manufacture and routinely as purified bulk. The finished product is formulated into excipients reflecting conditions close to the natural environment of the protein with respect to pH, osmolarity and the presence of human serum albumin. PMID:7535067

  3. 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

  4. Expression and Control of Codon-Optimized Granulocyte Colony-Stimulating Factor in Pichia pastoris.

    PubMed

    Maity, Nitu; Thawani, Ankita; Sharma, Anshul; Gautam, Ashwani; Mishra, Saroj; Sahai, Vikram

    2016-01-01

    Granulocyte colony-stimulating factor (GCSF) has therapeutic applications due to its proven efficacy in different forms of neutropenia and chemotherapy-induced leucopenia. The original 564-bp nucleotide sequence from NCBI was codon optimized and assembled by overlapping PCR method comprising of 16 oligos of 50-nt length with 15 base overhang. The synthetic gene (CO-GCSF) was cloned under glucose utilizing glyceraldehyde 3-phosphate dehydrogenase (GAP) and methanol-utilizing alcohol oxidase (AOX1) promoters and expressed in Pichia pastoris SMD1168 strain. Constitutive expression under GAP resulted in cellular toxicity while AOX1 promoter controlled expression was stable. Variation in the levels of expression was observed among the transformant colonies with transformant #2 secreting up to ∼4 mg/L of GCSF. The molecular mass of the expressed GCSF in P. pastoris was ∼19.0 kDa. Quatitation of the expressed protein was carried out by a highly reproducible gel densitometric method. Effect of several operational and nutritional conditions was studied on GCSF production and the results suggest a general approach for increasing the yield of GCSF several folds (2- to 5-fold) over the standard conditions employed currently. Cultivation of the single-copy integrant in the chemically defined medium in a 5-L fermenter resulted in a volumetric productivity of ∼0.7 mg/L/h at the end of the induction phase, which was about 4-fold higher than attained in the shake flask. PMID:26410223

  5. 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.

  6. 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.

  7. 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.

  8. Improved granulocyte colony-stimulating factor mobilization of hemopoietic progenitors using cytokine combinations in primates.

    PubMed

    Larsen, Stephen R; Chng, Keefe; Battah, Fiona; Martiniello-Wilks, Rosetta; Rasko, John E J

    2008-11-01

    Peripheral blood stem cells (PBSCs), usually mobilized with granulocyte colony-stimulating factor (G-CSF) alone or in combination with chemotherapy, are the preferred source of cells for hemopoietic stem cell transplantation. Up to 25% of otherwise eligible transplant recipients fail to harvest adequate PBSCs. Therefore it is important to investigate existing and novel reagents to improve PBSC mobilization. Because of marked interindividual variation in humans, we developed a robust nonhuman primate model that allows the direct comparison of the efficacy of two PBSC mobilization regimens within the same animal. Using this model, we compared pegylated G-CSF (pegG-CSF) with standard G-CSF and compared the combination of G-CSF and pegylated megakaryocyte growth and development factor (pegMGDF) with G-CSF plus stem cell factor (SCF) by measuring the levels of CD34(+) cells, colony-forming cells (CFCs), and SCID repopulating cells (SRCs) before and after cytokine administration. Mobilization of CD34(+) cells, CFCs and SRCs using pegG-CSF achieved similar levels to those resulting from 5 days of standard G-CSF. The combination of G-CSF+pegMGDF mobilized progenitors to levels similar to G-CSF+SCF but greater than standard G-CSF for CD34(+) cells and CFC. This first direct comparison of PBSC mobilization in individual primates demonstrates that peg-G-CSF is equivalent to daily G-CSF and that the addition of pegMGDF to G-CSF improves mobilization. In light of the development of new thrombopoietin agonists, these data offer the potential for improved stem cell mobilization strategies. Disclosure of potential conflicts of interest is found at the end of this article. PMID:18719223

  9. Stimulation of monocytes, macrophages, and microglia by amphotericin B and macrophage colony-stimulating factor promotes remyelination.

    PubMed

    Döring, Axinia; Sloka, Scott; Lau, Lorraine; Mishra, Manoj; van Minnen, Jan; Zhang, Xu; Kinniburgh, David; Rivest, Serge; Yong, V Wee

    2015-01-21

    Approaches to stimulate remyelination may lead to recovery from demyelinating injuries and protect axons. One such strategy is the activation of immune cells with clinically used medications, since a properly directed inflammatory response can have healing properties through mechanisms such as the provision of growth factors and the removal of cellular debris. We previously reported that the antifungal medication amphotericin B is an activator of circulating monocytes, and their tissue-infiltrated counterparts and macrophages, and of microglia within the CNS. Here, we describe that amphotericin B activates these cells through engaging MyD88/TRIF signaling. When mice were subjected to lysolecithin-induced demyelination of the spinal cord, systemic injections of nontoxic doses of amphotericin B and another activator, macrophage colony-stimulating factor (MCSF), further elevated the representation of microglia/macrophages at the site of injury. Treatment with amphotericin B, particularly in combination with MCSF, increased the number of oligodendrocyte precursor cells and promoted remyelination within lesions; these pro-regenerative effects were mitigated in mice treated with clodronate liposomes to reduce circulating monocytes and tissue-infiltrated macrophages. Our results have identified candidates among currently used medications as potential therapies for the repair of myelin. PMID:25609628

  10. Granulocyte-macrophage colony-stimulating factor: pleiotropic cytokine with potential clinical usefulness.

    PubMed

    Ruef, C; Coleman, D L

    1990-01-01

    Granulocyte-macrophage colony-stimulating factor (GM-CSF) is a 23-kDa glycoprotein with remarkably diverse effects on immune and nonimmune cells. GM-CSF induces differentiation of granulocyte, macrophage, and eosinophil precursor cells. Proliferation of monocyte-macrophages, T lymphocytes, keratinocytes, and endothelial cells is also stimulated by GM-CSF. In addition, GM-CSF alters the functional properties of mature granulocytes, macrophages, eosinophils, and basophils. GM-CSF is produced by T lymphocytes, macrophages, and several cell types in extramedullary sites, where it may act in a paracrine manner to regulate the local response to antigenic challenge. Clinical trials of GM-CSF have been conducted in patients with AIDS, aplastic anemia, myelodysplastic syndromes, and sarcoma and following bone marrow transplantation and accidental radiation exposure. GM-CSF significantly increased circulating numbers of several myeloid cells and produced dose-dependent toxicity consisting primarily of myalgias, fever, fluid retention, and serosal effusions. Additional studies are needed to define the role of GM-CSF in treatment of patients with qualitative and quantitative dysfunction of immune cells. PMID:2405468

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

    PubMed Central

    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

  12. Granulocyte colony-stimulating factor improves alternative activation of microglia under microenvironment of spinal cord injury.

    PubMed

    Guo, Y; Zhang, H; Yang, J; Liu, S; Bing, L; Gao, J; Hao, A

    2013-05-15

    Granulocyte colony-stimulating factor (G-CSF) was investigated in the present study to examine whether it could affect the activation status of microglia under microenvironment of spinal cord injury and provide a potential therapeutic treatment for spinal cord injury. We established mouse spinal cord hemisection model and injected recombinant human G-CSF (rhG-CSF) subcutaneously. The results demonstrated that G-CSF could recruit microglia to the injury site in the first 72h after spinal cord injury. Moreover, G-CSF inhibits the expression of pro-inflammatory factors and promotes the expression of neurotrophic factors. Additionally, G-CSF also increases the expression of markers of M2 macrophage and inhibits the expression of markers of M1 macrophage in BV2 microglia in vitro model, favoring the M2 polarization of microglia under the microenvironment of spinal cord hemisection. NFκB signal pathway was involved in G-CSF-induced polarization of BV2 microglia. As a conclusion, we suggested that administration of G-CSF within the first 72h after spinal cord injury might reduce early inflammation-induced detrimental effect and promote an anti-inflammatory response that favors repair via improving alternative activation of microglia. Administration of G-CSF in the acute phase of spinal cord injury may be a promising strategy in restorative therapy after spinal cord injury. PMID:23419550

  13. Human Granulocyte Macrophage Colony-Stimulating Factor Enhances Antibiotic Susceptibility of Pseudomonas aeruginosa Persister Cells.

    PubMed

    Choudhary, Geetika S; Yao, Xiangyu; Wang, Jing; Peng, Bo; Bader, Rebecca A; Ren, Dacheng

    2015-01-01

    Bacterial persister cells are highly tolerant to antibiotics and cause chronic infections. However, little is known about the interaction between host immune systems with this subpopulation of metabolically inactive cells, and direct effects of host immune factors (in the absence of immune cells) on persister cells have not been studied. Here we report that human granulocyte macrophage-colony stimulating factor (GM-CSF) can sensitize the persister cells of Pseudomonas aeruginosa PAO1 and PDO300 to multiple antibiotics including ciprofloxacin, tobramycin, tetracycline, and gentamicin. GM-CSF also sensitized the biofilm cells of P. aeruginosa PAO1 and PDO300 to tobramycin in the presence of biofilm matrix degrading enzymes. The DNA microarray and qPCR results indicated that GM-CSF induced the genes for flagellar motility and pyocin production in the persister cells, but not the normal cells of P. aeruginosa PAO1. Consistently, the supernatants from GM-CSF treated P. aeruginosa PAO1 persister cell suspensions were found cidal to the pyocin sensitive strain P. aeruginosa PAK. Collectively, these findings suggest that host immune factors and bacterial persisters may directly interact, leading to enhanced susceptibility of persister cells to antibiotics. PMID:26616387

  14. Human Granulocyte Macrophage Colony-Stimulating Factor Enhances Antibiotic Susceptibility of Pseudomonas aeruginosa Persister Cells

    PubMed Central

    Choudhary, Geetika S.; Yao, Xiangyu; Wang, Jing; Peng, Bo; Bader, Rebecca A.; Ren, Dacheng

    2015-01-01

    Bacterial persister cells are highly tolerant to antibiotics and cause chronic infections. However, little is known about the interaction between host immune systems with this subpopulation of metabolically inactive cells, and direct effects of host immune factors (in the absence of immune cells) on persister cells have not been studied. Here we report that human granulocyte macrophage-colony stimulating factor (GM-CSF) can sensitize the persister cells of Pseudomonas aeruginosa PAO1 and PDO300 to multiple antibiotics including ciprofloxacin, tobramycin, tetracycline, and gentamicin. GM-CSF also sensitized the biofilm cells of P. aeruginosa PAO1 and PDO300 to tobramycin in the presence of biofilm matrix degrading enzymes. The DNA microarray and qPCR results indicated that GM-CSF induced the genes for flagellar motility and pyocin production in the persister cells, but not the normal cells of P. aeruginosa PAO1. Consistently, the supernatants from GM-CSF treated P. aeruginosa PAO1 persister cell suspensions were found cidal to the pyocin sensitive strain P. aeruginosa PAK. Collectively, these findings suggest that host immune factors and bacterial persisters may directly interact, leading to enhanced susceptibility of persister cells to antibiotics. PMID:26616387

  15. Granulocyte colony-stimulating factor promotes behavioral recovery in a mouse model of traumatic brain injury.

    PubMed

    Song, Shijie; Kong, Xiaoyuan; Acosta, Sandra; Sava, Vasyl; Borlongan, Cesar; Sanchez-Ramos, Juan

    2016-05-01

    Hematopoietic growth factors such as granulocyte colony-stimulating factor (G-CSF) represent a novel approach for treatment of traumatic brain injury (TBI). After mild controlled cortical impact (CCI), mice were treated with G-CSF (100 μg/kg) for 3 consecutive days. The primary behavioral endpoint was performance on the radial arm water maze (RAWM), assessed 7 and 14 days after CCI. Secondary endpoints included 1) motor performance on a rotating cylinder (rotarod), 2) measurement of microglial and astroglial response, 3) hippocampal neurogenesis, and 4) measures of neurotrophic factors (brain-derived neurotrophic factor [BDNF] and glial cell line-derived neurotrophic factor [GDNF]) and cytokines in brain homogenates. G-CSF-treated animals performed significantly better than vehicle-treated mice in the RAWM at 1 and 2 weeks but not on the rotarod. Cellular changes found in the G-CSF group included increased hippocampal neurogenesis as well as astrocytosis and microgliosis in both the striatum and the hippocampus. Neurotrophic factors GDNF and BDNF, elaborated by activated microglia and astrocytes, were increased in G-CSF-treated mice. These factors along with G-CSF itself are known to promote hippocampal neurogenesis and inhibit apoptosis and likely contributed to improvement in the hippocampal-dependent learning task. Six cytokines that were modulated by G-CSF treatment following CCI were elevated on day 3, but only one of them remained altered by day 7, and all of them were no different from vehicle controls by day 14. The pro- and anti-inflammatory cytokines modulated by G-CSF administration interact in a complex and incompletely understood network involving both damage and recovery processes, underscoring the dual role of inflammation after TBI. PMID:26822127

  16. Granulocyte-colony stimulating factor as a treatment for diabetic neuropathy in rat.

    PubMed

    Kim, Kyung-Soo; Song, Yi-Sun; Jin, Jiyong; Joe, Jun-Ho; So, Byung-Im; Park, Jun-Young; Fang, Cheng-Hu; Kim, Mi Jung; Cho, Youl-Hee; Hwang, Sejin; Ro, Young-Suck; Kim, Hyuck; Ahn, You-Hern; Sung, Hak-Joon; Sung, Jung-Joon; Park, Sung-Hye; Lipton, Stuart A

    2015-10-15

    Effective treatment of diabetic neuropathy (DN) remains unsolved. We serendipitously observed dramatic relief of pain in several patients with painful DN receiving granulocyte-colony stimulating factor (G-CSF). The aim of this study was to determine if G-CSF could treat DN in an animal model and to ascertain its mechanism of action. In a rodent model of DN, G-CSF dramatically recovered nerve function, retarded histological nerve changes and increased the expression of neurotrophic factors within nerve. A sex-mismatched bone marrow transplantation (BMT) study revealed that G-CSF treatment increased the abundance of bone marrow (BM)-derived cells in nerves damaged by DN. However, we did not observe evidence of transdifferentiation or cell fusion of BM-derived cells. The beneficial effects of G-CSF were dependent on the integrity of BM. In conclusion, G-CSF produced a therapeutic effect in a rodent model of DN, which was attributed, at least in part, to the actions of BM-derived cells. PMID:26190836

  17. Granulocyte macrophage colony-stimulating factor and the intestinal innate immune cell homeostasis in Crohn's disease.

    PubMed

    Däbritz, Jan

    2014-03-01

    Current literature consolidates the view of Crohn's disease (CD) as a form of immunodeficiency highlighting dysregulation of intestinal innate immunity in the pathogenesis of CD. Intestinal macrophages derived from blood monocytes play a key role in sustaining the innate immune homeostasis in the intestine, suggesting that the monocyte/macrophage compartment might be an attractive therapeutic target for the management of CD. Granulocyte macrophage colony-stimulating factor (GM-CSF) is a hematopoietic growth factor that also promotes myeloid cell activation, proliferation, and differentiation. GM-CSF has a protective effect in human CD and mouse models of colitis. However, the role of GM-CSF in immune and inflammatory reactions in the intestine is not well defined. Beneficial effects exerted by GM-CSF during intestinal inflammation could relate to modulation of the mucosal barrier function in the intestine, including epithelial cell proliferation, survival, restitution, and immunomodulatory actions. The aim of this review is to summarize potential mechanistic roles of GM-CSF in intestinal innate immune cell homeostasis and to highlight its central role in maintenance of the intestinal immune barrier in the context of immunodeficiency in CD. PMID:24503766

  18. Upregulation of glucose metabolism by granulocyte-monocyte colony-stimulating factor

    SciTech Connect

    Schuler, A.; Spolarics, Z.; Lang, C.H.; Bagby, G.J.; Nelson, S.; Spitzer, J.J. )

    1991-01-01

    Alterations of glucose metabolism were investigated for 6 hours following an intraarterial injection of murine recombinant granulocyte-monocyte colony-stimulating factor (GM-CSF). GM-CSF resulted in a transient elevation of plasma glucose. The rate of whole body glucose appearance, as measured by infusion of (6-{sup 3}H)glucose, was increased by about 10% between 0.5 and 3 hours following GM-CSF injection. In vivo glucose utilization of individual tissues was investigated by the tracer 2-deoxyglucose technique. At 30 min, GM-CSF increased glucose utilization by 80-90% in liver and lung, and 50-60% in skin and spleen. At 3 and 6 hours, glucose utilization by these tissues returned toward control levels except for lung. There was a 40-50% increase in glucose utilization by skeletal muscle 30 min after GM-CSF which was sustained for 6 hours. Glucose utilization of testis, ileum and kidney did not change significantly. Plasma concentrations of insulin, glucagon and tumor necrosis factor were not altered in response to GM-CSF. These findings indicate that some of the acute metabolic effects of a short-term administration of GM-CSF are observed in macrophage-rich tissues, and suggest that GM-CSF may be involved in the metabolic upregulation of immunologically active tissues.

  19. Sulfur mustard-induced neutropenia: treatment with granulocyte colony-stimulating factor.

    PubMed

    Anderson, Dana R; Holmes, Wesley W; Lee, Robyn B; Dalal, Stephen J; Hurst, Charles G; Maliner, Beverly I; Newmark, Jonathan; Smith, William J

    2006-05-01

    Although best known as a blistering agent, sulfur mustard (HD) can also induce neutropenia in exposed individuals, increasing their susceptibility to infection. Granulocyte colony-stimulating factor (G-CSF) and pegylated G-CSF (peg-G-CSF) have been approved by the U.S. Food and Drug Administration as hematopoietic growth factors to treat chemotherapy-induced neutropenia. The goal of this study was to determine the effectiveness of G-CSF and peg-G-CSF in ameliorating HD-induced neutropenia. African green monkeys (Chlorocebus aethiops) were challenged with HD and, at 1, 3, 5, or 7 days after exposure, G-CSF therapy (10 microg/kg per day for 21 days) was initiated. Peg-G-CSF (300 microg/kg, single treatment) was similarly tested, with treatment given at 3 days after exposure. Untreated HD-exposed animals recovered from neutropenia 28 days after exposure, whereas G-CSF- or peg-G-CSF-treated animals recovered 8 to 19 days after exposure (p < 0.05). These results indicate that G-CSF or peg-G-CSF may provide Food and Drug Administration-approved treatments that will reduce the duration of HD-induced neutropenia. PMID:16761898

  20. Characterisation of a Novel Fc Conjugate of Macrophage Colony-stimulating Factor

    PubMed Central

    Gow, Deborah J; Sauter, Kristin A; Pridans, Clare; Moffat, Lindsey; Sehgal, Anuj; Stutchfield, Ben M; Raza, Sobia; Beard, Philippa M; Tsai, Yi Ting; Bainbridge, Graeme; Boner, Pamela L; Fici, Greg; Garcia-Tapia, David; Martin, Roger A; Oliphant, Theodore; Shelly, John A; Tiwari, Raksha; Wilson, Thomas L; Smith, Lee B; Mabbott, Neil A; Hume, David A

    2014-01-01

    We have produced an Fc conjugate of colony-stimulating factor (CSF) 1 with an improved circulating half-life. CSF1-Fc retained its macrophage growth-promoting activity, and did not induce proinflammatory cytokines in vitro. Treatment with CSF1-Fc did not produce adverse effects in mice or pigs. The impact of CSF1-Fc was examined using the Csf1r-enhanced green fluorescent protein (EGFP) reporter gene in MacGreen mice. Administration of CSF1-Fc to mice drove extensive infiltration of all tissues by Csf1r-EGFP positive macrophages. The main consequence was hepatosplenomegaly, associated with proliferation of hepatocytes. Expression profiles of the liver indicated that infiltrating macrophages produced candidate mediators of hepatocyte proliferation including urokinase, tumor necrosis factor, and interleukin 6. CSF1-Fc also promoted osteoclastogenesis and produced pleiotropic effects on other organ systems, notably the testis, where CSF1-dependent macrophages have been implicated in homeostasis. However, it did not affect other putative CSF1 targets, notably intestine, where Paneth cell numbers and villus architecture were unchanged. CSF1 has therapeutic potential in regenerative medicine in multiple organs. We suggest that the CSF1-Fc conjugate retains this potential, and may permit daily delivery by injection rather than continuous infusion required for the core molecule. PMID:24962162

  1. 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.

  2. Enhanced and Secretory Expression of Human Granulocyte Colony Stimulating Factor by Bacillus subtilis SCK6

    PubMed Central

    Bashir, Shaista; Sadaf, Saima; Ahmad, Sajjad; Akhtar, Muhammad Waheed

    2015-01-01

    This study describes a simplified approach for enhanced expression and secretion of a pharmaceutically important human cytokine, that is, granulocyte colony stimulating factor (GCSF), in the culture supernatant of Bacillus subtilis SCK6 cells. Codon optimized GCSF and pNWPH vector containing SpymwC signal sequence were amplified by prolonged overlap extension PCR to generate multimeric plasmid DNA, which was used directly to transform B. subtilis SCK6 supercompetent cells. Expression of GCSF was monitored in the culture supernatant for 120 hours. The highest expression, which corresponded to 17% of the total secretory protein, was observed at 72 hours of growth. Following ammonium sulphate precipitation, GCSF was purified to near homogeneity by fast protein liquid chromatography on a QFF anion exchange column. Circular dichroism spectroscopic analysis showed that the secondary structure contents of the purified GCSF are similar to the commercially available GCSF. Biological activity, as revealed by the regeneration of neutrophils in mice treated with ifosfamine, was also similar to the commercial preparation of GCSF. This, to our knowledge, is the first study that reports secretory expression of human GCSF in B. subtilis SCK6 with final recovery of up to 96 mg/L of the culture supernatant, without involvement of any chemical inducer. PMID:26881203

  3. 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

  4. The Effects of Granulocyte-Colony Stimulating Factor on Regeneration in Nerve Crush Injuries in Rats.

    PubMed

    Song, Yi-Sun; Joe, Jun-Ho; Joo, Hyun-Woo; Park, In-Hwa; Shen, Guang-Yin; Kim, Ki-Jun; Lee, Yonggu; Shin, Jeong Hun; Kim, Hyuck; Kim, Kyung-Soo

    2016-07-01

    Granulocyte-colony stimulating factor (G-CSF) is widely known to have a neuroprotective effect, but its effects on function and morphology in mechanical nerve injury are not well understood. The aim of this study was to confirm the time course of the functional changes and morphological effects of G-CSF in a rat model of nerve crush injury. Twelve-eight rats were divided into three group: sham-operated control group, G-CSF-treated group, and saline treated group. 2 weeks after the nerve crush injury, G-CSF was injected for 5 days. After 4 weeks, functional tests such as motor nerve conduction velocity (MNCV), mechanical and cold allodynia tests, and morphological studies were performed. G-CSF-treated rats had significantly improved nerve function including MNCV and mechanical and cold allodynia. In addition, G-CSF-treated rats had significantly higher the density of myelinated fibers than saline-treated rats. In conclusion, we found that 100 μg/kg administration of G-CSF promoted long-term functional recovery in a rat model of nerve crush injury. PMID:26980007

  5. 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

  6. Granulocyte-Macrophage Colony-Stimulating Factor in Staphylococcus aureus-Induced Arthritis

    PubMed Central

    Verdrengh, Margareta; Tarkowski, Andrej

    1998-01-01

    Granulocyte-macrophage colony-stimulating factor (GM-CSF) is a cytokine that is able to increase not only the production of phagocytic cells but also their efficacy with respect to, e.g., bactericidal properties. In this study, we wanted to analyze the impact of GM-CSF on experimental Staphylococcus aureus-induced arthritis. For that purpose, mice were administered GM-CSF before and after bacterial inoculation. Although there was an increase in the total number of leukocytes as well as in the granulocyte fraction, there was no favorable effect on the severity of arthritis or on survival rates. There were no obvious differences between the GM-CSF-pretreated animals and controls with regard to growth of staphylococci in joints and kidneys 4 days after the bacterial inoculation. In contrast, mice that had been pretreated with GM-CSF prior to bacterial inoculation showed approximately four times lower numbers of bacteria in their blood 24 h later. These results, along with those of our previous studies, suggest that on the one hand the granulocyte is the main protective cell during the course of S. aureus infection but that on the other hand, upregulation of granulocyte-macrophage production will not exert any additional protective effects with respect to tissue injury. PMID:9453655

  7. [Emergency therapy with granulocyte-macrophage colony-stimulating factor (GM-CSF)].

    PubMed

    Gratwohl, A; Dazzi, H; Tichelli, A; Stebler, C; Wernli, M; Thomssen, C; Kim, I; Dieterle, A; Obrist, R; Stern, A

    1991-03-23

    Granulocyte-macrophage colony stimulating factor (GM-CSF) has been tested for tolerability and efficacy on a compassionate need case basis in 17 patients (5 females, 12 males aged 4-72 years, median 35 years). GM-CSF was given at the rate of 3.5-32 micrograms/kg for 2-64 days as a continuous infusion for the following indications: impending rejection following bone marrow transplantation (5 patients), severe neutropenia secondary to chemotherapy in tumor patients (5), severe aplastic anemia (3), immune granulocytopenia (2) and accidental overdose with cytostatic agents (2 patients). Tolerance of GM-CSF was good in regard to doses of up to 16 micrograms/kg. Fever, myalgia and eosinophilia were the most frequent side effects. The patient treated with 32 micrograms/kg developed thrombosis of the vena cava. Efficacy is more difficult to assess in this heterogenous population, but 11 of 17 patients showed increased granulocyte counts and 3 patients clearly recovered from severe neutropenia. The role of GM-CSF in this recovery, however, cannot be proven. The results further indicate that GM-CSF cannot reverse ongoing rejection following allogenic BMT and cannot correct immune neutropenia. The value of GM-CSF therapy in patients with severe aplastic anemia and in the context of chemotherapy still needs to be defined. It is certainly indicated in patients with an accidental overdose of chemotherapeutic agents. PMID:2028244

  8. Granulocyte-macrophage colony-stimulating factor and pulmonary surfactant homeostasis.

    PubMed

    Reed, J A; Whitsett, J A

    1998-01-01

    Pulmonary surfactant lining the alveolus of the lung is critical to postnatal adaptation to air breathing. Precise concentrations of surfactant proteins and lipids are maintained in the alveolar space by a careful balance among synthesis, recycling, and catabolism. Pulmonary alveolar proteinosis is a rare pulmonary disease associated with accumulation of surfactant lipids and proteins in the alveolar spaces. Recent work with transgenic mice demonstrated that disruption of the production of granulocyte-macrophage colony-stimulating factor (GM-CSF) or the common beta-subunit of the GM-CSF receptor caused alveolar proteinosis that was histologically similar to that seen in human patients. The defect in surfactant homeostasis is caused by decreased surfactant clearance, mediated (at least in part) by dysfunction of the alveolar macrophage. Local production of GM-CSF corrects the alveolar proteinosis in the GM-CSF knockout mouse. Likewise, transplantation of wild-type bone marrow cells expressing the common beta-chain of the GM-CSF receptor restores surfactant homeostasis in the GM-CSF receptor knockout mouse. These studies demonstrate the previously unanticipated role of GM-CSF signaling in surfactant homeostasis, mediated (at least in part) by its actions on the clearance of surfactant lipids and proteins by the alveolar macrophage. These findings may have important implications for the diagnosis and treatment of pulmonary alveolar proteinosis syndromes in humans. PMID:9686680

  9. Effect of granulocyte-colony stimulating factor on spinal cord tissue after experimental contusion injury.

    PubMed

    Sanli, A Metin; Serbes, Gökhan; Calişkan, Murat; Kaptanoğlu, Erkan; Sargon, Mustafa F; Kilinç, Kamer; Beşalti, Omer; Sekerci, Zeki

    2010-12-01

    The purpose of this study was to investigate the early effects of granulocyte-colony stimulating factor (G-CSF) on myeloperoxidase (MPO) activity, lipid peroxidation (LPO) and ultrastructural findings in rats after spinal cord injury (SCI). We also compared the effects of G-CSF and methylprednisolone sodium succinate (MPSS). Wistar rats were divided into four groups: control, SCI alone (50 g/cm weight drop trauma), SCI+MPSS (30 mg/kg), and SCI+G-CSF (50 μg/kg). Administration of G-CSF and MPSS significantly decreased LPO (p < 0.05) and MPO activity (p < 0.05) in the first 24 hours. MPSS was more effective than G-CSF in reducing LPO (p < 0.05) and in minimizing ultrastructure changes. The results of this study indicate that G-CSF exerts a beneficial effect by decreasing MPO activity and LPO and may reduce tissue damage in the first 24 hours after SCI. Our findings do not exclude the possibility that G-CSF has a protective effect on spinal cord ultrastructure after the first 24 hours following SCI. PMID:20801040

  10. Capillary electrophoretic separation of poly(ethylene glycol)-modified granulocyte-colony stimulating factor.

    PubMed

    Lee, Kyung Soo; Na, Dong Hee

    2010-03-01

    We evaluated the utility of capillary electrophoretic methods for analyzing poly(ethylene glycol) (PEG)-modified granulocyte-colony stimulating factor (G-CSF), a long-acting form of GCSF for the treatment of cancer therapy-induced neutropenia. Low- and high-molecularweight PEG-G-CSF conjugates prepared with aldehyde-activated PEG-5K and PEG-20K were separated by high-performance size-exclusion chromatography (HP-SEC), capillary zone electrophoresis (CZE), and sodium dodecyl sulfate-capillary gel electrophoresis (SDS-CGE). HPSEC showed low resolution for separating mono- and di-PEG-G-CSFs. SDS-CGE had higher resolution, but required a long analysis and had low peak efficiency. CZE could successfully separate both PEG-5K- and PEG-20K-conjugated G-CSFs with a running time of 20 min and high peak efficiency. In conclusion, CZE was better than SDS-CGE for separating PEG-G-CSF conjugates and will be useful for PEGylation studies, such as reaction monitoring for optimization of the PEGylation reaction, and purity and stability tests of PEG-G-CSF. PMID:20361316

  11. Granulocyte colony-stimulating factor delays neutrophil apoptosis by inhibition of calpains upstream of caspase-3

    PubMed Central

    Drewniak, Agata; Groenewold, Vincent; van den Berg, Timo K.; Kuijpers, Taco W.

    2008-01-01

    Neutrophils have a very short life span and undergo apoptosis within 24 hours after leaving the bone marrow. Granulocyte colony-stimulating factor (G-CSF) is essential for the recruitment of fresh neutrophils from the bone marrow but also delays apoptosis of mature neutrophils. To determine the mechanism by which G-CSF inhibits neutrophil apoptosis, the kinetics of neutrophil apoptosis during 24 hours in the absence or presence of G-CSF were analyzed in vitro. G-CSF delayed neutrophil apoptosis for approximately 12 hours and inhibited caspase-9 and -3 activation, but had virtually no effect on caspase-8 and little effect on the release of proapoptotic proteins from the mitochondria. However, G-CSF strongly inhibited the activation of calcium-dependent cysteine proteases calpains, upstream of caspase-3, via apparent control of Ca2+-influx. Calpain inhibition resulted in the stabilization of the X-linked inhibitor of apoptosis (XIAP) and hence inhibited caspase-9 and -3 in human neutrophils. Thus, neutrophil apoptosis is controlled by G-CSF after initial activation of caspase-8 and mitochondrial permeabilization by the control of postmitochondrial calpain activity. PMID:18524991

  12. Colony Stimulating Factors 1, 2, 3 and early pregnancy steps: from bench to bedside.

    PubMed

    Rahmati, Mona; Petitbarat, Marie; Dubanchet, Sylvie; Bensussan, Armand; Chaouat, Gerard; Ledee, Nathalie

    2015-06-01

    Reproductive immunology applies general immunology principles to specialised targets, reproduction and development. The involvement of colony-stimulating factors (CSFs) in reproduction illustrates this. The CSF family includes CSF-1 or macrophage CSF (M-CSF), CSF-2 or granulocyte macrophage CSF (GM-CSF), and CSF-3 or granulocyte CSF (G-CSF). Each member has a specific localisation and timed expression in the reproductive tract with specific functions involving them in ovulation, embryo implantation, placentation and further embryonic development. They are used in reproductive medicine, either as biomarkers of oocyte quality and competence (follicular G-CSF), or to supplement embryo culture media with human recombinant GM-CSF, or they are used as an innovative therapy by using human recombinant G-CSF for infertile patients. Given fundamental considerations on CSFs and their strong implication in reproduction, this review aimed to detail the current knowledge for each member of the family to improve our understanding of their implication in the maternal-foetal cytokinic dialogue and in possibly preventing reproductive disorders. PMID:25721620

  13. Granulocyte/Macrophage Colony-stimulating Factor-dependent Dendritic Cells Restrain Lean Adipose Tissue Expansion.

    PubMed

    Pamir, Nathalie; Liu, Ning-Chun; Irwin, Angela; Becker, Lev; Peng, YuFeng; Ronsein, Graziella E; Bornfeldt, Karin E; Duffield, Jeremy S; Heinecke, Jay W

    2015-06-01

    The physiological roles of macrophages and dendritic cells (DCs) in lean white adipose tissue homeostasis have received little attention. Because DCs are generated from bone marrow progenitors in the presence of granulocyte/macrophage colony-stimulating factor (GM-CSF), we used GM-CSF-deficient (Csf2(-/-)) mice fed a low fat diet to test the hypothesis that adipose tissue DCs regulate the development of adipose tissue. At 4 weeks of age, Csf2(-/-) mice had 75% fewer CD45(+)Cd11b(+)Cd11c(+)MHCII(+) F4/80(-) DCs in white adipose tissue than did wild-type controls. Furthermore, the Csf2(-/-) mice showed a 30% increase in whole body adiposity, which persisted to adulthood. Adipocytes from Csf2(-/-) mice were 50% larger by volume and contained higher levels of adipogenesis gene transcripts, indicating enhanced adipocyte differentiation. In contrast, adipogenesis/adipocyte lipid accumulation was inhibited when preadipocytes were co-cultured with CD45(+)Cd11b(+)Cd11c(+)MHCII(+)F4/80(-) DCs. Medium conditioned by DCs, but not by macrophages, also inhibited adipocyte lipid accumulation. Proteomic analysis revealed that matrix metalloproteinase 12 and fibronectin 1 were greatly enriched in the medium conditioned by DCs compared with that conditioned by macrophages. Silencing fibronectin or genetic deletion of matrix metalloproteinase 12 in DCs partially reversed the inhibition of adipocyte lipid accumulation. Our observations indicate that DCs residing in adipose tissue play a critical role in suppressing normal adipose tissue expansion. PMID:25931125

  14. Enhancement of human granulocyte-colony stimulating factor production in recombinant E. coli using batch cultivation.

    PubMed

    Babaeipour, Valiollah; Abbas, Mahdi Pesaran Haji; Sahebnazar, Zahra; Alizadeh, Reza

    2010-06-01

    Development of inexpensive and simple culture media is always favorable for recombinant protein over-expression in E. coli. The effects of medium composition on the production of recombinant human granulocyte-colony stimulating factor (rh-GCSF) were investigated in batch culture of E. coli BL21 (DE3) [pET23a-hgcsf]. First, the optimum medium for production of rh-GCSF was determined; and, then it was shown that mixture of amino acid addition at induction time, which was determined on the basis of amino acids frequency in the recombinant protein, increases recombinant protein expression level significantly. Furthermore, the effect of glucose concentration on productivity of rh-GCSF was investigated; 20 g/l of glucose will result in maximum attainable biomass and rh-GCSF in this process. At optimum conditions, a cell dry weight of 10.5 g/l, an expression level of about 35% of total cellular protein, rh-GCSF concentration of 1.75 +/- 0.1 g/l, and overall rh-GCSF yield of 165 +/- 5 mg/g were obtained. PMID:19859744

  15. Human Granulocyte Colony-Stimulating Factor (hG-CSF) Expression in Plastids of Lactuca sativa

    PubMed Central

    Sharifi Tabar, Mehdi; Habashi, Ali Akbar; Rajabi Memari, Hamid

    2013-01-01

    Background: Human granulocyte colony-stimulating factor (hG-CSF) can serve as valuable biopharmaceutical for research and treatment of the human blood cancer. Transplastomic plants have been emerged as a new and high potential candidate for production of recombinant biopharmaceutical proteins in comparison with transgenic plants due to extremely high level expression, biosafety and many other advantages. Methods: hG-CSF gene was cloned into pCL vector between prrn16S promoter and TpsbA terminator. The recombinant vector was coated on nanogold particles and transformed to lettuce chloroplasts through biolistic method. Callogenesis and regeneration of cotyledonary explants were obtained by Murashige and Skoog media containing 6-benzylaminopurine and 1-naphthaleneacetic acid hormones. The presence of hG-CSF gene in plastome was studied with four specific PCR primers and expression by Western immunoblotting. Results: hG-CSF gene cloning was confirmed by digestion and sequencing. Transplastomic lettuce lines were regenerated and subjected to molecular analysis. The presence of hG-CSF in plastome was confirmed by PCR using specific primers designed from the plastid genome. Western immunoblotting of extracted protein from transplastomic plants showed a 20-kDa band, which verified the expression of recombinant protein in lettuce chloroplasts. Conclusions: This study is the first report that successfully express hG-CSF gene in lettuce chloroplast. The lettuce plastome can provide a cheap and safe expression platform for producing valuable biopharmaceuticals for research and treatment. PMID:23748895

  16. Macrophage colony-stimulating factor is indispensable for both proliferation and differentiation of osteoclast progenitors.

    PubMed Central

    Tanaka, S; Takahashi, N; Udagawa, N; Tamura, T; Akatsu, T; Stanley, E R; Kurokawa, T; Suda, T

    1993-01-01

    The mechanism of action of macrophage colony-stimulating factor (M-CSF) in osteoclast development was examined in a co-culture system of mouse osteoblastic cells and spleen cells. In this co-culture, osteoclast-like multinucleated cells (MNCs) were formed within 6 d in response to 10 nM 1 alpha,25(OH)2D3 added only for the final 2 d of culture. Simultaneously adding hydroxyurea for the final 2 d completely inhibited proliferation of cultured cells without affecting 1 alpha,25(OH)2D3-stimulated MNC formation. Autoradiographic examination using [3H]-thymidine revealed that osteoclast progenitors primarily proliferated during the first 4 d, whereas their differentiation into MNCs occurred predominantly during the final 2 d of culture in response to 1 alpha,25(OH)2D3. When anti-M-CSF antibody or anti-M-CSF receptor antibody was added either for the first 4 d or for the final 2 d, the MNC formation was similarly inhibited. In co-cultures of normal spleen cells and osteoblastic cells obtained from op/op mice, which cannot produce functionally active M-CSF, the lack of M-CSF either for the first 4 d or for the final 2 d failed to form MNCs in response to 1 alpha,25(OH)2D3 added for the last 2 d. These results clearly indicate that M-CSF is indispensable for both proliferation of osteoclast progenitors and their differentiation into mature osteoclasts. Images PMID:8423223

  17. 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

  18. 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

  19. Cardiopulmonary effects of granulocyte colony-stimulating factor in a canine model of bacterial sepsis.

    PubMed

    Eichacker, P Q; Waisman, Y; Natanson, C; Farese, A; Hoffman, W D; Banks, S M; MacVittie, T J

    1994-11-01

    We investigated the effects of recombinant granulocyte colony-stimulating factor (G-CSF) in a canine model of septic shock. Awake 2-yr-old beagles were studied before and after intraperitoneal placement of an Escherichia coli-infected clot. Nine days before and until 3 days after clot placement, animals received daily high-dose (G-CSF (5 microgram/kg body wt; n = 17), low-dose G-CSF (0.1 microgram/kg body wt; n = 17), or a control protein (5 micrograms/kg body wt; n = 20). Survival rate was greater (P < 0.04, Wilcoxon test) in the high-dose G-CSF group (14/17) than in the low-dose G-CSF (10/17) and control (12/20) groups. High-dose G-CSF improved cardiovascular function, as evidenced by increased left ventricular ejection fraction (day 1 after clot; P < 0.001) and mean arterial pressure (day 2; P < 0.02) compared with low-dose G-CSF and control groups. High-dose G-CSF increased (P < 0.001) mean peripheral neutrophils before (-3 days) and after (2 h to 4 days) clot and produced a more rapid (P < 0.001) rise (day 2) and fall (day 4) in mean alveolar neutrophil numbers compared with the low-dose G-CSF and control groups. High-dose G-CSF decreased mean serum endotoxin (2-8 h; P < 0.002) and tumor necrosis factor (2 h; P < 0.02) levels and lowered blood bacteria counts (2-6 h; P < 0.04) compared with the low-dose G-CSF and control groups. Thus, in this canine model, G-CSF sufficient to increase peripheral neutrophils before and during peritonitis and septic shock enhances host defense, reduces cytokine (tumor necrosis factor) levels, and improves cardiovascular function and survival. PMID:7532649

  20. Administration of granulocyte colony-stimulating factor with radiotherapy promotes tumor growth by stimulating vascularization in tumor-bearing mice.

    PubMed

    Kim, Joong Sun; Son, Yeonghoon; Bae, Min Ji; Lee, Minyoung; Lee, Chang Geun; Jo, Wol Soon; Kim, Sung Dae; Yang, Kwangmo

    2015-07-01

    Although granulocyte-colony stimulating factor (G-CSF) is commonly used to support recovery from radiation-induced side-effects, the precise effects of G-CSF on colon cancer under radiotherapy remain poorly understood. In the present study, to investigate the effects of tumor growth following radiotherapy and G-CSF administration in a murine xenograft model of colon cancer, female BALB/c mice were injected with cells of a colon carcinoma cell line (CT26) with irradiation and G-CSF, alone or in combination. Mice received 2 Gy of focal radiation daily for 5 days and intraperitoneal injection of G-CSF (100 µg/kg/day) after irradiation for 7 days. Changes in the levels of myeloperoxidase (MPO), vascular endothelial growth factor (VEGF), matrix metalloproteinase type 9 (MMP-9) and CD31 were assessed in the mouse cancer induced by injection of colon cancer cells. We observed that G-CSF increased the number of circulating neutrophils, but facilitated tumor growth. However, G-CSF treatment did not affect radiation-induced cytotoxicity and cell viability in CT26 cells in vitro. Increased levels of myeloperoxidase, a neutrophil marker and those of vascular endothelial growth factor were observed in tumors with G-CSF supplementation. In addition, we found that increased levels of CD31 and matrix metalloproteinase-9 were correlated with the enhanced tumor growth after G-CSF treatment. Therefore, these data suggest that G-CSF may contribute to tumor growth and decrease the antitumor effect of radiotherapy, possibly by promoting vascularization in cancer lesions. PMID:25976379

  1. Safety of recombinant human granulocyte-macrophage colony-stimulating factor in healing pediatric severe burns.

    PubMed

    Chi, Y F; Chai, J K; Luo, H M; Zhang, Q X; Feng, R

    2015-01-01

    We explored the safety of recombinant human granulocyte-macrophage colony-stimulating factor (rhGM-CSF) for healing burns in children. Subjects were randomly assigned to two groups: the experimental group received external rhGM-CSF gel, and the control group received rhGM-CSF gel matrix components, applied to the burn surface. Neither group was given any other drugs that promote wound healing. Each day we recorded the pulse, body temperature, and respiration status in the two groups. We detected the blood routine, urine routine, and hepatic and renal function before the patients received drug treatment and after 72 h. The wound scab and healing states in the two groups were recorded every 4 days to evaluate wound healing rate and time taken for complete healing. Adverse reactions and their rate of occurrence were also recorded. The median time of healing was 15 days in the experimental group and 19 days in the control group (log-rank χ(2) = 5.139, P < 0.05). After 10 days, the experimental group healing rate was consistently higher than that of the control group (significantly different using intuitive analysis), suggesting the experimental group method was more effective. There were no obvious adverse reactions. There was no significant difference between the blood routine, urine routine, and liver and kidney function in the two groups before the treatment and after 3 days (P > 0.05). Compared with saline treatment of severe burns, rhGM-CSF can effectively shorten the healing time without significant adverse reactions, and is an effective and safe treatment for burns in children. PMID:25867422

  2. The effect of granulocyte-colony stimulating factor in global cerebral ischemia in rats

    PubMed Central

    Matchett, Gerald A.; Calinisan, Jason B.; Matchett, Genoveve C.; Martin, Robert D.; Zhang, John H.

    2007-01-01

    Granulocyte-colony stimulating factor (G-CSF) is an endogenous peptide hormone of the hematopoietic system that has entered Phase I/II clinical trials for treatment of ischemic stroke. Severe intraoperative hypotension can lead to global cerebral ischemia and apoptotic neuron loss within the hippocampus. We tested G-CSF in a rat model of global cerebral ischemia. Global cerebral ischemia was induced in male Sprague-Dawley rats (280–330g) with the 2-vessel occlusion model (hemorrhagic hypotension to a mean arterial pressure of 30–35 mmHg and bilateral common carotid artery occlusion for 8 minutes). Three groups of animals were used: global ischemia without treatment (GI, n=49), global ischemia with G-CSF treatment (GI+G-CSF, n=42), and sham surgery (Sham, n = 26). Rats in the treatment group received G-CSF (50 μg / kg, subcutaneously) 12 hours before surgery, on the day of surgery, and on post-operative Day 1, and were euthanized on Day 2, 3, and 14. Mild hyperglycemia was observed in all groups. T-maze testing for spontaneous alternation demonstrated initial improvement in the G-CSF treatment group but no long-term benefit. Measurement of daily body weight demonstrated an initial trend toward improvement in the G-CSF group. Quantitative Nissl histology of the hippocampus demonstrated equivalent outcomes on Day 3 and 14, which was supported by quantitative TUNEL stain. Immunohistochemistry and Western blot demonstrated an initial increase in phosphorylated-AKT in the GI+G-CSF group on Day 2. We conclude that G-CSF treatment is associated with transient early improvement in neurobehavioral outcomes after global ischemia complicated by mild hyperglycemia, but no long-term protection. PMID:17210148

  3. Soluble periplasmic production of human granulocyte colony-stimulating factor (G-CSF) in Pseudomonas fluorescens.

    PubMed

    Jin, Hongfan; Cantin, Greg T; Maki, Steven; Chew, Lawrence C; Resnick, Sol M; Ngai, Jerry; Retallack, Diane M

    2011-07-01

    Cost-effective production of soluble recombinant protein in a bacterial system remains problematic with respect to expression levels and quality of the expressed target protein. These constraints have particular meaning today as "biosimilar" versions of innovator protein drugs are entering the clinic and the marketplace. A high throughput, parallel processing approach to expression strain engineering was used to evaluate soluble expression of human granulocyte colony-stimulating factor (G-CSF) in Pseudomonas fluorescens. The human g-csf gene was optimized for expression in P. fluorescens and cloned into a set of periplasmic expression vectors. These plasmids were transformed into a variety of P. fluorescens host strains each having a unique phenotype, to evaluate soluble expression in a 96-well growth and protein expression format. To identify a strain producing high levels of intact, soluble Met-G-CSF product, more than 150 protease defective host strains from the Pfēnex Expression Technology™ toolbox were screened in parallel using biolayer interferometry (BLI) to quantify active G-CSF binding to its receptor. A subset of these strains was screened by LC-MS analysis to assess the quality of the expressed G-CSF protein. A single strain with an antibiotic resistance marker insertion in the pfaI gene was identified that produced>99% Met-GCSF. A host with a complete deletion of the autotransporter-coding gene pfaI from the genome was constructed, and expression of soluble, active Met-GSCF in this strain was observed to be 350mg/L at the 1 liter fermentation scale. PMID:21396452

  4. Effects of Granulocyte Colony-Stimulating Factor on Patients with Liver Failure: a Meta-Analysis

    PubMed Central

    Yang, Qiao; Yang, Ying; Shi, Yu; Lv, Fangfang; He, Jiliang; Chen, Zhi

    2016-01-01

    Abstract Background and Aims: It remains controversial whether granulocyte colony-stimulating factor (G-CSF) prolongs survival in liver failure (LF) patients. This meta-analysis was performed to evaluate the effect of G-CSF on patients with LF. Methods: PubMed, EMBASE, and Web of Science databases were searched to identify English language randomized controlled trials comparing G-CSF with control therapy published before14 February 2015. A meta-analysis was performed to examine changes in liver function and patient survival. The association was tested using odds ratio (OR) or risk ratio (RR) with 95% confidence intervals (CI). Results: Five randomized controlled trials were eligible for the meta-analysis. Significant amelioration of prothrombin time and total bilirubin in LF patients was attributed to G-CSF therapy (OR, −0.064; 95% CI,−0.481 to 0.353; p< 0.001; and OR, −0.803; 95% CI, −1.177 to −0.430; p = 0.000, respectively). Treatment with G-CSF resulted in improved Model for End-Stage Liver Disease and Child-Turcotte-Pugh scores (OR, −1.741; 95% CI, −2.234 to −1.250; p = 0.000; and OR, −0.830, 95% CI, −1.194 to −0.465; p = 0.000, respectively). A lower incidence of sepsis was found in patients treated with G-CSF (RR, 0.367; 95% CI, 0.158 to 0.854; p = 0.020). G-CSF therapy significantly increased survival rate in LF patients (RR, 2.25; 95% CI, 1.517 to 3.338; p = 0.000). Conclusions: The results of this meta-analysis indicate that G-CSF treatment in patients with LF significantly improved liver function, reduced the incidence of sepsis, and prolonged short-term survival. PMID:27350939

  5. Does granulocyte-colony stimulating factor administration induce damage or repair response in schistosomiasis?

    PubMed Central

    Ghanem, Lobna Y; Dahmen, Uta; Dirsch, Olaf; Nosseir, Mona MF; Mahmoud, Soheir S; Mansour, Wafaa AF

    2010-01-01

    AIM: To introduce Granulocyte-colony stimulating factor (G-CSF) as a new therapeutic modality for schistosomiasis through stem cell mobilization, immunomodulation or fibrosis remodeling. METHODS: In this study, a 5 d course of human recombinant G-CSF (100 μg/kg sc) was applied to Schistosoma mansoni-infected mice at different stages of disease (5 d before infection as well as 3, 5 and 7 wk post-infection). The animals were sacrificed at 10 d as well as 4, 6 and 8 wk post infection. Mice were examined for: (1) Total leukocyte count which is an accepted surrogate marker for the stem cell mobilization into the circulation; (2) Egg count in intestine and liver tissue to assess the parasitic load; and (3) Histopathological changes in Hx/E and Masson trichrome stained sections as well as collagen content in Sirius red-stained liver sections to determine the severity of liver fibrosis. RESULTS: Mice developed leukocytosis. The egg load and the number of granulomas were not affected by the G-CSF treatment but there was an obvious change in the composition of granulomas towards an increased cellularity. Moreover, fibrosis was significantly decreased in treated groups compared to untreated animals (collagen content either preinfection or at 3 and 5 wk post infection: 5.8 ± 0.5, 4.7 ± 0.5, 4.0 ± 0.7 vs 8.2 ± 0.9; P ≤ 0.01). CONCLUSION: Although G-CSF did not cause direct elimination of the parasite, it enhanced granulomatous reaction and reduced the fibrosis. Further investigation of the underlying mechanisms of these two actions is warranted. PMID:21191519

  6. Molecular Mechanism of Regulation of MTA1 Expression by Granulocyte Colony-stimulating Factor.

    PubMed

    Kumar, Arathy S; Jagadeeshan, Sankar; Subramanian, Anirudh; Chidambaram, Saravana Babu; Surabhi, Rohan Prasad; Singhal, Mahak; Bhoopalan, Hemadev; Sekar, Sathiya; Pitani, Ravi Shankar; Duvuru, Prathiba; Venkatraman, Ganesh; Rayala, Suresh K

    2016-06-01

    Parkinson disease (PD) is a neurodegenerative disorder with loss of dopaminergic neurons of the brain, which results in insufficient synthesis and action of dopamine. Metastasis-associated protein 1 (MTA1) is an upstream modulator of tyrosine hydroxylase (TH), the rate-limiting enzyme in dopamine synthesis, and hence MTA1 plays a significant role in PD pathogenesis. To impart functional and clinical significance to MTA1, we analyzed MTA1 and TH levels in the substantia nigra region of a large cohort of human brain tissue samples by Western blotting, quantitative PCR, and immunohistochemistry. Our results showed that MTA1 and TH levels were significantly down-regulated in PD samples as compared with normal brain tissue. Correspondingly, immunohistochemistry analysis for MTA1 in substantia nigra sections revealed that 74.1% of the samples had a staining intensity of <6 in the PD samples as compared with controls, 25.9%, with an odds ratio of 8.54. Because of the clinical importance of MTA1 established in PD, we looked at agents to modulate MTA1 expression in neuronal cells, and granulocyte colony-stimulating factor (G-CSF) was chosen, due to its clinically proven neurogenic effects. Treatment of the human neuronal cell line KELLY and acute 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine mouse model with G-CSF showed significant induction of MTA1 and TH with rescue of phenotype in the mouse model. Interestingly, the observed induction of TH was compromised on silencing of MTA1. The underlying molecular mechanism of MTA1 induction by G-CSF was proved to be through induction of c-Fos and its recruitment to the MTA1 promoter. PMID:27044752

  7. 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

  8. Granulocyte/Macrophage Colony-stimulating Factor-dependent Dendritic Cells Restrain Lean Adipose Tissue Expansion*

    PubMed Central

    Pamir, Nathalie; Liu, Ning-Chun; Irwin, Angela; Becker, Lev; Peng, YuFeng; Ronsein, Graziella E.; Bornfeldt, Karin E.; Duffield, Jeremy S.; Heinecke, Jay W.

    2015-01-01

    The physiological roles of macrophages and dendritic cells (DCs) in lean white adipose tissue homeostasis have received little attention. Because DCs are generated from bone marrow progenitors in the presence of granulocyte/macrophage colony-stimulating factor (GM-CSF), we used GM-CSF-deficient (Csf2−/−) mice fed a low fat diet to test the hypothesis that adipose tissue DCs regulate the development of adipose tissue. At 4 weeks of age, Csf2−/− mice had 75% fewer CD45+Cd11b+Cd11c+MHCII+ F4/80− DCs in white adipose tissue than did wild-type controls. Furthermore, the Csf2−/− mice showed a 30% increase in whole body adiposity, which persisted to adulthood. Adipocytes from Csf2−/− mice were 50% larger by volume and contained higher levels of adipogenesis gene transcripts, indicating enhanced adipocyte differentiation. In contrast, adipogenesis/adipocyte lipid accumulation was inhibited when preadipocytes were co-cultured with CD45+Cd11b+Cd11c+MHCII+F4/80− DCs. Medium conditioned by DCs, but not by macrophages, also inhibited adipocyte lipid accumulation. Proteomic analysis revealed that matrix metalloproteinase 12 and fibronectin 1 were greatly enriched in the medium conditioned by DCs compared with that conditioned by macrophages. Silencing fibronectin or genetic deletion of matrix metalloproteinase 12 in DCs partially reversed the inhibition of adipocyte lipid accumulation. Our observations indicate that DCs residing in adipose tissue play a critical role in suppressing normal adipose tissue expansion. PMID:25931125

  9. Granulocyte colony stimulating factor attenuates inflammation in a mouse model of amyotrophic lateral sclerosis

    PubMed Central

    2011-01-01

    Background Granulocyte colony stimulating factor (GCSF) is protective in animal models of various neurodegenerative diseases. We investigated whether pegfilgrastim, GCSF with sustained action, is protective in a mouse model of amyotrophic lateral sclerosis (ALS). ALS is a fatal neurodegenerative disease with manifestations of upper and lower motoneuron death and muscle atrophy accompanied by inflammation in the CNS and periphery. Methods Human mutant G93A superoxide dismutase (SOD1) ALS mice were treated with pegfilgrastim starting at the presymptomatic stage and continued until the end stage. After long-term pegfilgrastim treatment, the inflammation status was defined in the spinal cord and peripheral tissues including hematopoietic organs and muscle. The effect of GCSF on spinal cord neuron survival and microglia, bone marrow and spleen monocyte activation was assessed in vitro. Results Long-term pegfilgrastim treatment prolonged mutant SOD1 mice survival and attenuated both astro- and microgliosis in the spinal cord. Pegfilgrastim in SOD1 mice modulated the inflammatory cell populations in the bone marrow and spleen and reduced the production of pro-inflammatory cytokine in monocytes and microglia. The mobilization of hematopoietic stem cells into the circulation was restored back to basal level after long-term pegfilgrastim treatment in SOD1 mice while the storage of Ly6C expressing monocytes in the bone marrow and spleen remained elevated. After pegfilgrastim treatment, an increased proportion of these cells in the degenerative muscle was detected at the end stage of ALS. Conclusions GCSF attenuated inflammation in the CNS and the periphery in a mouse model of ALS and thereby delayed the progression of the disease. This mechanism of action targeting inflammation provides a new perspective of the usage of GCSF in the treatment of ALS. PMID:21711557

  10. 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

  11. Granulocyte-Colony Stimulating Factor Related Pathways Tested on an Endometrial Ex-Vivo Model

    PubMed Central

    Rahmati, Mona; Petitbarat, Marie; Dubanchet, Sylvie; Bensussan, Armand; Chaouat, Gerard; Ledee, Nathalie

    2014-01-01

    Introduction Recombinant human Granulocyte-Colony Stimulating Factor (rhG-CSF) supplementation seems to be a promising innovative therapy in reproductive medicine, used in case of recurrent miscarriage, embryo implantation failure or thin endometrium, although its mechanisms of action remain unknown. Our aim was to identify possible endometrial pathways influenced by rhG-CSF. Materials and Methods Hypothetical molecular interactions regulated by G-CSF were designed through a previous large scale endometrial microarray study. The variation of endometrial expression of selected target genes was confirmed in control and infertile patients. G-CSF supplementation influence on these targets was tested on an endometrial ex-vivo culture. Middle luteal phase endometrial biopsies were cultured on collagen sponge with or without rhG-CSF supplementation during 3 consecutive days. Variations of endometrial mRNA expression for the selected targets were studied by RT-PCR. Results At the highest dose of rhG-CSF stimulation, the mRNA expression of these selected target genes was significantly increased if compared with their expression without addition of rhG-CSF. The selected targets were G-CSF Receptor (G-CSFR), Integrin alpha-V/beta-3 (ITGB3) implicated in cell migration and embryo implantation, Plasminogen Activator Urokinase Receptor (PLAUR) described as interacting with integrins and implicated in cell migration, Thymidine Phosphorylase (TYMP) implicated in local angiogenesis, CD40 and its ligand CD40L involved in cell proliferation control. Conclusion RhG-CSF seems able to influence endometrial expressions crucial for implantation process involving endometrial vascular remodelling, local immune modulation and cellular adhesion pathways. These variations observed in an ex-vivo model should be tested in-vivo. The strict indications or counter indication of rhG-CSF supplementation in reproductive field are not yet established, while the safety of its administration in early

  12. Granulocyte-colony stimulating factor as treatment option in patients with recurrent miscarriage.

    PubMed

    Santjohanser, Claudia; Knieper, Catherine; Franz, Cordula; Hirv, Kaino; Meri, Osama; Schleyer, Manfred; Würfel, Wolfgang; Toth, Bettina

    2013-04-01

    In 1-5% of patients during childbearing years recurrent miscarriages (RM) occur. There are established risk factors like anatomical, endocrine and hemostatic disorders as well as immunological changes in the maternal immune system. Nevertheless, further elucidation of the pathogenesis remains a matter of debate. In addition, there are no standardized immunological treatment strategies. Recent studies indicate possible effects of tumor necrosis factor α blocker and granulocyte-colony stimulating factor (G-CSF) concerning live birth rate (LBR) in RM patients. Therefore, we performed a retrospective cohort study in patients undergoing assisted reproductive treatment (ART) with known RM analysing the possible benefits of G-CSF application. From January 2002 to December 2010, 127 patients (199 cylces) with RM (at least 2 early miscarriages) 49 (72 cycles) receiving G-CSF and 78 (127 cycles) controls receiving either no medication (subgroup 1) or Cortisone, intravenous immunoglobulins or low molecular weight heparin (subgroup 2) undergoing ART for in vitro fertilisation/intracytoplasmic sperm injection were analysed. G-CSF was administered weekly once (34 Mill) in 11 patients, 38 patients received 2 × 13 Mill G-CSF per week until the 12th week of gestation. Statistical analysis was performed with SPSS for Windows (19.0), p < 0.05 significant. The mean age of the study population was 37.3 ± 4.4 years (mean ± standard deviation) and differed not significantly between patients and subgroups. However, the number of early miscarriages was significantly higher in the G-CSF group as compared to the subgroups (G-CSF 2.67 ± 1.27, subgroup 1 0.85 ± 0.91, subgroup 2 0.64 ± 0.74) and RM patients receiving G-CSF had significantly more often a late embryo transfer (day 5) (G-CSF 36.7%, subgroup 1 12.1%, subgroup 2 8.9%). The LBR of patients and the subgroups differed significantly (G-CSF 32%, subgroup 1 13%, subgroup 2 14%). Side effects were present in less than 10% of

  13. Recombinant human granulocyte-macrophage colony-stimulating factor stimulates in vitro mature human neutrophil and eosinophil function, surface receptor expression, and survival.

    PubMed Central

    Lopez, A F; Williamson, D J; Gamble, J R; Begley, C G; Harlan, J M; Klebanoff, S J; Waltersdorph, A; Wong, G; Clark, S C; Vadas, M A

    1986-01-01

    A purified recombinant human granulocyte-macrophage colony stimulating factor (rH GM-CSF) was a powerful stimulator of mature human eosinophils and neutrophils. The purified rH GM-CSF enhanced the cytotoxic activity of neutrophils and eosinophils against antibody-coated targets, stimulated phagocytosis of serum-opsonized yeast by both cell types in a dose-dependent manner, and stimulated neutrophil-mediated iodination in the presence of zymosan. In addition, rH GM-CSF enhanced N-formylmethionylleucylphenylalanine(FMLP)-stimulated degranulation of Cytochalasin B pretreated neutrophils and FMLP-stimulated superoxide production. In contrast, rH GM-CSF did not promote adherence of granulocytes to endothelial cells or plastic surfaces. rH GM-CSF selectively enhanced the surface expression of granulocyte functional antigens 1 and 2, and the Mo1 antigen. rH GM-CSF induced morphological changes and enhanced the survival of both neutrophils and eosinophils by 6 and 9 h, respectively. These experiments show that granulocyte-macrophage colony stimulating factor can selectively stimulate mature granulocyte function. PMID:3021817

  14. 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

  15. Early Systemic Granulocyte-Colony Stimulating Factor Treatment Attenuates Neuropathic Pain after Peripheral Nerve Injury

    PubMed Central

    Lee, Yun-Lin; Chen, Jin-Chung; Wang, Hung-Li; Yang, Yi-Ling; Cheng, Mei-Yun; Liao, Ming-Feng; Ro, Long-Sun

    2012-01-01

    Recent studies have shown that opioid treatment can reduce pro-inflammatory cytokine production and counteract various neuropathic pain syndromes. Granulocyte colony-stimulating factor (G-CSF) can promote immune cell differentiation by increasing leukocytes (mainly opioid-containing polymorphonuclear (PMN) cells), suggesting a potential beneficial role in treating chronic pain. This study shows the effectiveness of exogenous G-CSF treatment (200 µg/kg) for alleviating thermal hyperalgesia and mechanical allodynia in rats with chronic constriction injury (CCI), during post-operative days 1–25, compared to that of vehicle treatment. G-CSF also increases the recruitment of opioid-containing PMN cells into the injured nerve. After CCI, single administration of G-CSF on days 0, 1, and 2, but not on day 3, relieved thermal hyperalgesia, which indicated that its effect on neuropathic pain had a therapeutic window of 0–48 h after nerve injury. CCI led to an increase in the levels of interleukin-6 (IL-6) mRNA and tumor necrosis factor-α (TNF-α) protein in the dorsal root ganglia (DRG). These high levels of IL-6 mRNA and TNF-α were suppressed by a single administration of G-CSF 48–144 h and 72–144 h after CCI, respectively. Furthermore, G-CSF administered 72–144 h after CCI suppressed the CCI-induced upregulation of microglial activation in the ipsilateral spinal dorsal horn, which is essential for sensing neuropathic pain. Moreover, the opioid receptor antagonist naloxone methiodide (NLXM) reversed G-CSF-induced antinociception 3 days after CCI, suggesting that G-CSF alleviates hyperalgesia via opioid/opioid receptor interactions. These results suggest that an early single systemic injection of G-CSF alleviates neuropathic pain via activation of PMN cell-derived endogenous opioid secretion to activate opioid receptors in the injured nerve, downregulate IL-6 and TNF-α inflammatory cytokines, and attenuate microglial activation in the spinal dorsal horn. This

  16. Effects of recombinant granulocyte colony-stimulating factor (rG-CSF) and recombinant granulocyte-macrophage colony-stimulating factor (rGM-CSF) on acute radiation hematopoietic injury in mice

    SciTech Connect

    Tanikawa, S.; Nakao, I.; Tsuneoka, K.; Nara, N. )

    1989-09-01

    We have attempted to evaluate in vivo effects of granulocyte colony-stimulating factor (G-CSF) and granulocyte-macrophage colony-stimulating factor (GM-CSF) on acute radiation hematopoietic injury in mice. BDF1 mice, irradiated with 7.5-Gy x-rays, were injected i.p. twice daily for 10 days with 10(5) U recombinant human G-CSF, 3.75 x 10(5) U recombinant murine GM-CSF, or a combination of both. G-CSF significantly enhanced the recovery of not only peripheral leukocytes but also platelets and hematocrit on days 14 and 21 after irradiation. GM-CSF significantly enhanced the recovery of platelets on day 14 and peripheral leukocytes on day 21. G-CSF markedly enhanced the recovery of spleen colony-forming units (CFU-S), colony-forming units in culture (CFU-C), erythroid burst-forming units (BFU-E), and megakaryocyte colony-forming units (CFU-Meg) both in bone marrow and in the spleen. GM-CSF significantly enhanced the recovery of CFU-Meg in bone marrow on day 14. We found synergistic effects between G-CSF and GM-CSF on CFU-S, CFU-C, and CFU-Meg in the spleen on day 14, although we found antagonistic effects between G-CSF and GM-CSF on CFU-S and CFU-C in bone marrow on day 7, and on platelet counts on day 7. These results indicate that the administration of recombinant G-CSF and GM-CSF may be useful in accelerating hematopoietic recovery in patients with acute radiation hematopoietic injuries.

  17. The synergistic therapeutic effect of hepatocyte growth factor and granulocyte colony-stimulating factor on pulmonary hypertension in rats.

    PubMed

    Guo, Yinghua; Su, Longxiang; Li, Yinghui; Guo, Na; Xie, Lixin; Zhang, Dong; Zhang, Xiaojun; Li, Hongxia; Zhang, Guizhi; Wang, Yajuan; Liu, Changting

    2014-07-01

    Pulmonary arterial hypertension (PAH) is characterized by a progressive increase in pulmonary arterial pressure and vascular resistance. Despite advances in therapy for PAH, its treatment and prognosis remain poor. We aimed to investigate whether the transplantation of bone marrow mesenchymal stem cells (MSCs) overexpressing hepatocyte growth factor (HGF), alone or in combination with granulocyte colony-stimulating factor (G-CSF), attenuates the development of experimental monocrotaline (MCT)-induced PAH. Three weeks after MCT administration, rats were divided into the following groups: (1) untreated (PAH); (2) HGF treated; (3) MSCs administered; (4) HGF-MSCs treated; and (5) HGF-MSCs plus G-CSF treated. After 3 weeks, hemodynamic changes, histomorphology, and angiogenesis were evaluated. To elucidate the molecular mechanisms of vascular remodeling and angiogenesis, serum levels of transforming growth factor (TGF)-β and endothelin-1 (ET-1) were measured, and the gene and protein expression levels of vascular cell adhesion molecule-1 (VCAM-1) and matrix metalloproteinase-9 (MMP-9) were determined. Compared with the PAH, MSC, and G-CSF groups, the HGF and HGF+G-CSF groups exhibited significantly reduced right ventricular hypertrophy and mean pulmonary arterial pressure (P < 0.05). Histologically, vessel muscularization or thickening and collagen deposition were also significantly decreased (P < 0.05). The number of vessels in the HGF+G-CSF group was higher than that in the other groups (P < 0.05). The TGF-β and ET-1 concentrations in the plasma of pulmonary hypertensive rats were markedly lower in the HGF and HGF+G-CSF groups (P < 0.05). Furthermore, HGF induced the expression of VCAM-1, and HGF treatment together with G-CSF synergistically stimulated MMP-9 expression. Transplanted HGF-MSCs combined with G-CSF potentially offer synergistic therapeutic benefit for the treatment of PAH. PMID:23933910

  18. 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.

  19. 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

  20. Absence of granulocyte colony-stimulating factor signaling and neutrophil development in CCAAT enhancer binding protein alpha-deficient mice.

    PubMed

    Zhang, D E; Zhang, P; Wang, N D; Hetherington, C J; Darlington, G J; Tenen, D G

    1997-01-21

    Transcription factors are master regulatory switches of differentiation, including the development of specific hematopoietic lineages from stem cells. Here we show that mice with targeted disruption of the CCAAT enhancer binding protein alpha gene (C/EBP alpha) demonstrate a selective block in differentiation of neutrophils. Mature neutrophils and eosinophils are not observed in the blood or fetal liver of mutant animals, while other hematopoietic lineages, including monocytes, are not affected. Instead, most of the white cells in the peripheral blood of mutant mice had the appearance of myeloid blasts. We also observed a selective loss of expression of a critical gene target of CCAAT enhancer binding protein alpha, the granulocyte colony-stimulating factor receptor. As a result, multipotential myeloid progenitors from the mutant fetal liver are unable to respond to granulocyte colony-stimulating factor signaling, although they are capable of forming granulocyte-macrophage and macrophage colonies in methylcellulose in response to other growth factors. Finally, we demonstrate that the lack of granulocyte development results from a defect intrinsic to the hematopoietic system; transplanted fetal liver from mutant mice can reconstitute lymphoid but not neutrophilic cells in irradiated recipients. These studies suggest a model by which transcription factors can direct the differentiation of multipotential precursors through activation of expression of a specific growth factor receptor, allowing proliferation and differentiation in response to a specific extracellular signal. In addition, the c/ebp alpha -/- mice may be useful in understanding the mechanisms involved in acute myelogenous leukemia, in which a block in differentiation of myeloid precursors is a key feature of the disease. PMID:9012825

  1. Successful Granulocyte Colony-stimulating Factor Treatment of Relapsing Candida albicans Meningoencephalitis Caused by CARD9 Deficiency.

    PubMed

    Celmeli, Fatih; Oztoprak, Nefise; Turkkahraman, Doga; Seyman, Derya; Mutlu, Esvet; Frede, Natalie; Köksoy, Sadi; Grimbacher, Bodo

    2016-04-01

    Caspase-associated recruitment domain-9 (CARD9) deficiency is an autosomal-recessive primary immunodeficiency with genetic defects in Th17 immunity marked by susceptibility to recurrent and invasive Candida infections. We present a case of relapsing Candida albicans meningoencephalitis over 1-year period despite appropriate antifungal therapy. We detected a homozygous p.Q295X mutation in CARD9 as well as a defective interleukin-17 and interferon gamma synthesis in Enzyme-Linked ImmunoSpot tests. We achieved complete clinical remission, and improvement of interleukin-17 secretion with subcutaneous granulocyte colony-stimulating factor) treatment. PMID:26658378

  2. Identification of critical regions in mouse granulocyte-macrophage colony-stimulating factor by scanning-deletion analysis.

    PubMed Central

    Shanafelt, A B; Kastelein, R A

    1989-01-01

    Structure-function relationships for mouse granulocyte-macrophage colony-stimulating factor were examined by generating a series of small deletions scanning the entire length of the molecule. Deletions of three amino acids were introduced at intervals of five amino acids by site-directed mutagenesis of the mature mouse granulocyte-macrophage colony-stimulating factor gene. The mutant proteins were expressed in Escherichia coli and assayed for biological activity. This procedure identified four regions critical to activity. These critical regions were further delineated by additional three-amino acid deletion mutants. Larger deletions at each terminus were also made, as well as changes of specific amino acid residues. The four critical regions span amino acid residues 18-22, 34-41, 52-61, and 94-115. The disulfide bridge between Cys-51 and Cys-93 was also shown to be essential for activity, whereas that between Cys-85 and Cys-118 could be removed without loss of activity. The possible structural and/or functional roles of the critical regions are discussed. Images PMID:2662186

  3. The expression and localization of mRNA for colony-stimulating factor (CSF)-1 in human term placenta.

    PubMed

    Kanzaki, H; Yui, J; Iwai, M; Imai, K; Kariya, M; Hatayama, H; Mori, T; Guilbert, L J; Wegmann, T G

    1992-04-01

    A 4-kb mRNA for colony-stimulating factor 1 (CSF-1) was detected in normal human placenta at term by Northern blot analysis. In-situ hybridization revealed that the mRNA for CSF-1 was localized in the mesenchymal cells of the chorionic villous stroma, but not in the trophoblasts or capillary epithelial cells. Because there are significant numbers of tissue macrophages (Hofbauer cells) in the placental stroma and because the receptor for CSF-1 (the c-fms proto-oncogene product) is known to be expressed by trophoblasts, our results suggest that CSF-1 produced by placental stromal cells may act as a growth and survival factor for human placental macrophages and trophoblasts. PMID:1522204

  4. 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

  5. Regulation of surface expression of the granulocyte/macrophage colony-stimulating factor receptor in normal human myeloid cells

    SciTech Connect

    Cannistra, S.A.; Groshek, P.; Griffin, J.D. ); Garlick, R.; Miller, J. )

    1990-01-01

    Recombinant human granulocyte/macrophage colony-stimulating factor (GM-CSF) exerts stimulatory effects on hematopoietic cells through binding to specific, high-affinity receptors. By using radiolabeled GM-CSF with high specific activity, the authors have investigated the factors and mechanisms that regulate GM-CSF receptor expression in normal human neutrophils, monocytes, and partially purified bone marrow myeloid progenitor cells. The neutrophil GM-CSF receptor was found to rapidly internalize in the presence of ligand through a mechanism that required endocytosis. Out of a large panel of naturally occurring humoral factors tested, only GM-CSF itself, tumor necrosis factor, and formyl-Met-Leu-Phe were found to down-regulate neutrophil GM-CSF receptor expression after a 2-hr exposure at biologically active concentrations. Since formyl-Met-Leu-Phe is known to stimulate neutrophil protein kinase C activity, they also tested the ability of protein kinase C agonists to modulate GM-CSF receptor expression. Phorbol 12-myristate 13-acetate, bryostatin-1, and 1,2-dioctanoylglycerol were found to induce rapid down-regulation of the GM-CSF receptor in neutrophils, monocytes, and partially purified myeloid progenitor cells, suggesting that this effect may be at least partially mediated by protein kinase C. These data suggest that certain activators of neutrophil function may negatively regulate their biological effects by inducing down-regulation of the GM-CSF receptor.

  6. 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.

  7. Localization of the murine macrophage colony-stimulating factor gene to chromosome 3 using interspecific backcross analysis.

    PubMed

    Buchberg, A M; Jenkins, N A; Copeland, N G

    1989-08-01

    The chromosomal location of the murine macrophage colony-stimulating factor (Csfm) gene was determined by interspecific backcross analysis. We mapped Csfm to mouse chromosome 3, 2.5 cM distal to Ngfb and Nras and 1.3 cM proximal to Amy-2. CSFM maps to human chromosome 5q, while AMY2, NGFB, and NRAS map to human chromosome 1p. The chromosomal location of Csfm thus disrupts a previously identified conserved linkage group between mouse chromosome 3 and human chromosome 1. The location of Csfm also identifies yet another mouse chromosome that shares synteny with human chromosome 5q, a region involved in several different types of myeloid disease. PMID:2676841

  8. Detection of transcripts for the receptor for macrophage colony-stimulating factor, c-fms, in murine osteoclasts.

    PubMed Central

    Hofstetter, W; Wetterwald, A; Cecchini, M C; Felix, R; Fleisch, H; Mueller, C

    1992-01-01

    Macrophage colony-stimulating factor (M-CSF), whose action is restricted to the cell populations of the mononuclear phagocyte system, has recently been found to be required for osteoclastogenesis and bone resorption. To investigate the cells involved in the action of M-CSF in these processes, expression of c-fms mRNA, encoding the M-CSF receptor, was studied by in situ hybridization. Paws from murine embryos and newborn mice, tibiae from 2-day-old animals, as well as isolated osteoclasts, were hybridized with a c-fms-specific RNA probe. In bone, c-fms mRNA was detected only in cells at the late stages of osteoclastogenesis and in mature osteoclasts. The findings strengthen the relation between osteoclasts and the mononuclear phagocyte system. Furthermore, they suggest that M-CSF acts directly on osteoclast precursors and on mature osteoclasts during osteoclastogenesis. Images PMID:1409676

  9. Critical analysis of an oncolytic herpesvirus encoding granulocyte-macrophage colony stimulating factor for the treatment of malignant melanoma

    PubMed Central

    Hughes, Tasha; Coffin, Robert S; Lilley, Caroline E; Ponce, Rafael; Kaufman, Howard L

    2014-01-01

    Oncolytic viruses that selectively lyse tumor cells with minimal damage to normal cells are a new area of therapeutic development in oncology. An attenuated herpesvirus encoding the granulocyte-macrophage colony stimulating factor (GM-CSF), known as talimogene laherparepvec (T-VEC), has been identified as an attractive oncolytic virus for cancer therapy based on preclinical tumor studies and results from early-phase clinical trials and a large randomized Phase III study in melanoma. In this review, we discuss the basic biology of T-VEC, describe the role of GM-CSF as an immune adjuvant, summarize the preclinical data, and report the outcomes of published clinical trials using T-VEC. The emerging data suggest that T-VEC is a safe and potentially effective antitumor therapy in malignant melanoma and represents the first oncolytic virus to demonstrate therapeutic activity against human cancer in a randomized, controlled Phase III study. PMID:27512660

  10. 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

  11. Application of microchip CGE for the analysis of PEG-modified recombinant human granulocyte-colony stimulating factors.

    PubMed

    Park, Eun Ji; Lee, Kyung Soo; Lee, Kang Choon; Na, Dong Hee

    2010-11-01

    The purpose of this study was to evaluate the microchip CGE (MCGE) for the analysis of PEG-modified granulocyte-colony stimulating factor (PEG-G-CSF) prepared with PEG-aldehydes. The unmodified and PEG-modified G-CSFs were analyzed by Protein 80 and 230 Labchips on the Agilent 2100 Bioanalyzer. The MCGE allowed size-based separation and quantitation of PEG-G-CSF. The Protein 80 Labchip was useful for PEG-5K-G-CSF, while the Protein 230 Labchip was more suitable for PEG-20K-G-CSF. The MCGE was also used to monitor a search for optimal PEG-modification (PEGylation) conditions to produce mono-PEG-G-CSF. This study demonstrates the usefulness of MCGE for monitoring and optimizing the PEGylation of G-CSF with the advantages of speed, minimal sample consumption, and automatic quantitation. PMID:20945411

  12. Colony stimulating factor-inducing activity of isoflavone C-glucosides from the bark of Dalbergia monetaria.

    PubMed

    Kawaguchi, K; Alves, S de M; Watanabe, T; Kikuchi, S; Satake, M; Kumazawa, Y

    1998-10-01

    To obtain immunomodulating substances from Amazonian medicinal plants, hot water extracts from 21 samples available commercially were tested in terms of mitogenic and colony-stimulating factor (CSF)-inducing activities. Among them, Dalbergia monetaria exhibited the highest CSF-inducing activity. Orobol 8-C-glucoside (OCG-8) and orobol 6-C-glucoside (OCG-6) were isolated from the bark of D. monetaria as major constituents. The CSF-inducing activity of OCG-8 was higher than that of OCG-6 and a dose-dependent manner at a range of 0.1-10 mg/mouse. Serum CSF production induced by an intraperitoneal (i.p.) injection with 1 mg OCG-8 reached a peak at 4-6 h later, suggesting that OCG-8 would act on hematopoietic system. PMID:9810271

  13. Expression of human granulocyte colony stimulating factor (hG-CSF) in colon adenocarcinoma cell line (Caco-2).

    PubMed

    Jana, Snehasis; Patel, Hitesh

    2012-10-01

    Growth and progression of many cancer cells are mediated by alterations in the microenvironment often caused by an aberrant expression of growth factors and receptors. There is no report on expression of growth factor granulocyte colony-stimulating factor (G-CSF) in the experimental model, colon adenocarcinoma cell line (Caco2), that is commonly used in drug permeability assays. We hypothesize that in vitro, the Caco2 model is associated with a constitutive neo-expression of the hematopoietic G-CSF thereby causing an autocrine stimulation of Caco2 growth and proliferation in vitro. To test our hypothesis, we analyzed mRNA and protein expression of G-CSF in Caco2 cells using reverse transcriptase-PCR and SDS-PAGE. G-CSF mRNA and protein were detected in Caco2 cells. Expression of G-CSF protein was similar at different passages of this cell line. The expression of G-CSF has a significant role in the autocrine regulation of Caco2 cell growth and proliferation. PMID:22714276

  14. Effects of recombinant human granulocyte colony-stimulating factor on the hematologic recovery and survival of irradiated mice

    SciTech Connect

    Tanikawa, S.; Nose, M.; Aoki, Y.; Tsuneoka, K.; Shikita, M.; Nara, N. )

    1990-08-01

    We studied the effects of intraperitoneal injections of recombinant human granulocyte colony-stimulating factor (rhG-CSF) according to various administration schedules on the recovery of spleen colony-forming units (CFU-S) and peripheral blood counts, and on the survival of irradiated mice. The sooner and more frequently the mice were injected with rhG-CSF after irradiation, the more enhanced the recovery of CFU-S in bone marrow was obtained on day 7. Twice-daily injections of rhG-CSF from day 0 to day 2 significantly enhanced the recovery of platelets and hematocrit, but two injections of rhG-CSF on only day 0 did not. Twice-daily injections of rhG-CSF from day 0 to day 6 enhanced the recovery of platelets more effectively than twice-daily injections of rhG-CSF from day 1 to day 7, and increased the survival of irradiated mice more effectively than any other examined administration schedules. Twice-daily injections of rhG-CSF from day 0 to day 6 were significantly effective in enhancing the survival of mice irradiated with 8.5-, 9.0-, and 9.5-Gy x-rays, although not effective after irradiation of 10.5-Gy x-rays.

  15. Granulocyte colony-stimulating factor enhances bone tumor growth in mice in an osteoclast-dependent manner

    PubMed Central

    Hirbe, Angela C.; Uluçkan, Özge; Morgan, Elizabeth A.; Eagleton, Mark C.; Prior, Julie L.; Piwnica-Worms, David; Trinkaus, Kathryn; Apicelli, Anthony

    2007-01-01

    Inhibition of osteoclast (OC) activity has been associated with decreased tumor growth in bone in animal models. Increased recognition of factors that promote osteoclastic bone resorption in cancer patients led us to investigate whether increased OC activation could enhance tumor growth in bone. Granulocyte colony-stimulating factor (G-CSF) is used to treat chemotherapy-induced neutropenia, but is also associated with increased markers of OC activity and decreased bone mineral density (BMD). We used G-CSF as a tool to investigate the impact of increased OC activity on tumor growth in 2 murine osteolytic tumor models. An 8-day course of G-CSF alone (without chemotherapy) significantly decreased BMD and increased OC perimeter along bone in mice. Mice administered G-CSF alone demonstrated significantly increased tumor growth in bone as quantitated by in vivo bioluminescence imaging and histologic bone marrow tumor analysis. Short-term administration of AMD3100, a CXCR4 inhibitor that mobilizes neutrophils with little effect on bone resorption, did not lead to increased tumor burden. However, OC-defective osteoprotegerin transgenic (OPGTg) mice and bisphosphonate-treated mice were resistant to the effects of G-CSF administration upon bone tumor growth. These data demonstrate a G-CSF–induced stimulation of tumor growth in bone that is OC dependent. PMID:17192391

  16. 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

  17. 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.

  18. Multipotent hematopoietic cell lines derived from C/EBPalpha(-/-) knockout mice display granulocyte macrophage-colony-stimulating factor, granulocyte- colony-stimulating factor, and retinoic acid-induced granulocytic differentiation.

    PubMed

    Collins, S J; Ulmer, J; Purton, L E; Darlington, G

    2001-10-15

    The transcription factor C/EBPalpha is an important mediator of granulocyte differentiation and regulates the expression of multiple granulocyte-specific genes including the granulocyte-colony-stimulating factor (G-CSF) receptor, neutrophil elastase, and myeloperoxidase. Indeed C/EBPalpha knockout mice display a profound block in granulocyte differentiation. To study this block in granulocytic differentiation in more detail, retroviral vector-mediated transduction of a dominant-negative retinoic acid receptor was used to establish hematopoietic growth factor-dependent, lympho-myeloid progenitor cell lines from the fetal livers of both the C/EBPalpha knockout animals (C/EBPalpha(-/-)) and their heterozygous littermates (C/EBPalpha(+/-)). Surprisingly, the C/EBPalpha(-/-) cell lines displayed significant spontaneous granulocytic differentiation, and this differentiation was markedly enhanced when the cells were stimulated with granulocyte macrophage (GM)-CSF. This GM-CSF-mediated differentiation was associated with the up-regulation of G-CSF receptor mRNA, and the combination of GM-CSF and G-CSF generated more than 95% mature neutrophils in the C/EBPalpha(-/-) cultures. The addition of all-trans retinoic acid also enhanced this granulocytic differentiation of the cultured C/EBPalpha(-/-) cells, indicating that the activated retinoic acid receptors can enhance granulocytic differentiation through a molecular pathway that is independent of C/EBPalpha. These studies clearly indicate that terminal granulocytic differentiation associated with the up-regulation of C/EBPalpha-responsive genes can occur in the absence of C/EBPalpha, and they indicate the existence of multiple independent molecular pathways potentially used by primitive hematopoietic precursors that can lead to the development of mature granulocytes. PMID:11588034

  19. Granulocyte colony-stimulating factor reprograms bone marrow stromal cells to actively suppress B lymphopoiesis in mice

    PubMed Central

    Day, Ryan B.; Bhattacharya, Deepta; Nagasawa, Takashi

    2015-01-01

    The mechanisms that mediate the shift from lymphopoiesis to myelopoiesis in response to infectious stress are largely unknown. We show that treatment with granulocyte colony-stimulating factor (G-CSF), which is often induced during infection, results in marked suppression of B lymphopoiesis at multiple stages of B-cell development. Mesenchymal-lineage stromal cells in the bone marrow, including CXCL12-abundant reticular (CAR) cells and osteoblasts, constitutively support B lymphopoiesis through the production of multiple B trophic factors. G-CSF acting through a monocytic cell intermediate reprograms these stromal cells, altering their capacity to support B lymphopoiesis. G-CSF treatment is associated with an expansion of CAR cells and a shift toward osteogenic lineage commitment. It markedly suppresses the production of multiple B-cell trophic factors by CAR cells and osteoblasts, including CXCL12, kit ligand, interleukin-6, interleukin-7, and insulin-like growth factor-1. Targeting bone marrow stromal cells is one mechanism by which inflammatory cytokines such as G-CSF actively suppress lymphopoiesis. PMID:25814527

  20. Granulocyte-colony stimulating factor promotes lung metastasis through mobilization of Ly6G+Ly6C+ granulocytes

    PubMed Central

    Kowanetz, Marcin; Wu, Xiumin; Lee, John; Tan, Martha; Hagenbeek, Thijs; Qu, Xueping; Yu, Lanlan; Ross, Jed; Korsisaari, Nina; Cao, Tim; Bou-Reslan, Hani; Kallop, Dara; Weimer, Robby; Ludlam, Mary J. C.; Kaminker, Joshua S.; Modrusan, Zora; van Bruggen, Nicholas; Peale, Franklin V.; Carano, Richard; Meng, Y. Gloria; Ferrara, Napoleone

    2010-01-01

    Priming of the organ-specific premetastatic sites is thought to be an important yet incompletely understood step during metastasis. In this study, we show that the metastatic tumors we examined overexpress granulocyte-colony stimulating factor (G-CSF), which expands and mobilizes Ly6G+Ly6C+ granulocytes and facilitates their subsequent homing at distant organs even before the arrival of tumor cells. Moreover, G-CSF–mobilized Ly6G+Ly6C+ cells produce the Bv8 protein, which has been implicated in angiogenesis and mobilization of myeloid cells. Anti–G-CSF or anti-Bv8 antibodies significantly reduced lung metastasis. Transplantation of Bv8 null fetal liver cells into lethally irradiated hosts also reduced metastasis. We identified an unexpected role for Bv8: the ability to stimulate tumor cell migration through activation of one of the Bv8 receptors, prokineticin receptor (PKR)-1. Finally, we show that administration of recombinant G-CSF is sufficient to increase the numbers of Ly6G+Ly6C+ cells in organ-specific metastatic sites and results in enhanced metastatic ability of several tumors. PMID:21081700

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

    PubMed

    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. PMID:25909052

  2. 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.

  3. 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

  4. Granulocyte colony-stimulating factor does not enhance phagocytosis or microbicidal activity of human mature polymorphonuclear neutrophils in vitro.

    PubMed Central

    Shimono, N; Okada, K; Takeda, D; Eguchi, K; Misumi, H; Sawae, Y; Niho, Y

    1994-01-01

    The direct effects of human granulocyte colony-stimulating factor (hG-CSF) on mature polymorphonuclear neutrophils (PMNs) in vitro were studied with regard to chemotaxis, superoxide production, and phagocytosis and microbicidal activity against the following viable microorganisms: Staphylococcus aureus, serum-resistant Pseudomonas aeruginosa, and Candida albicans. Recombinant hG-CSF (rhG-CSF) acted as a chemoattractant for human PMNs in a dose-dependent manner. The chemotactic response of PMNs to N-formyl-methionyl-leucyl-phenylalanine (FMLP) was not enhanced by rhG-CSF at any of the concentrations used. rhG-CSF did not induce the generation of superoxide by itself. However, rhG-CSF was able to prime human PMNs and to enhance O2- release stimulated by FMLP in a dose-dependent manner. rhg-CSF did not enhance phagocytosis or killing of the three species of microorganisms by normal PMNs. With PMNs obtained from patients who had hematological disorders or solid tumors, no enhancement of the microbicidal activity was observed in most cases. Microbial killing mediated by PMNs depended on the ratio of PMNs to target organisms. We concluded from these facts that the most important effect of rhG-CSF was to increase the number of the peripheral PMNs and not to enhance the functions of mature PMNs. PMID:8556501

  5. Effects of granulocyte-macrophage colony-stimulating factor and cyclic AMP interaction on human neutrophil apoptosis.

    PubMed Central

    Tortorella, C; Piazzolla, G; Spaccavento, F; Antonaci, S

    1998-01-01

    The current study was undertaken to evaluate the effects of granulocyte-macrophage colony-stimulating factor (GM-CSF) and cyclic AMP (cAMP) signaling interaction on human neutrophil apoptosis, either occurring spontaneously or induced by Fas antigen activation. Results show that GM-CSF, dibutyryl cAMP (a cAMP analog) and forskolin (an adenylate cyclase activator) are all able to suppress spontaneous neutrophil cell death. Of note however, when GM-CSF is used in combination with cAMP-elevating agents, an additive effect on neutrophil survival is observed with dibutyryl cAMP only, whereas supplementation of cell cultures with GM-CSF and forskolin results in a progressive reduction of antiapoptotic effects exerted by the single compounds. Moreover, although dibutyryl cAMP and forskolin do not affect Fas-triggered apoptotic events, they are still able to modulate the GM-CSF capacity to prolong neutrophil survival following anti-Fas IgM cell challenge, with effects similar to those respectively exerted on spontaneous neutrophil apoptosis. The data indicate that GM-CSF may negatively modulate the cAMP-mediated antiapoptotic pathway in human neutrophils, likely via the inhibition of adenylate cyclase activity. This would prevent an abnormal neutrophil survival as a result of cAMP signaling stimulation, which provides a novel insight into the role of GM-CSF as a physiological regulator of myeloid cell turnover. PMID:9927231

  6. Effect of recombinant human granulocyte colony-stimulating factor (rh G-CSF) on murine resistance against Listeria monocytogenes.

    PubMed Central

    Serushago, B A; Yoshikai, Y; Handa, T; Mitsuyama, M; Muramori, K; Nomoto, K

    1992-01-01

    Recombinant human granulocyte colony-stimulating factor (rh G-CSF) enhanced resistance of mice against Listeria monocytogenes (LM) as determined by survival and bacterial growth. Mice pretreated with rh G-CSF twice daily for 5 days survived better than untreated animals to the challenge with LM. Number of bacteria in peritoneal cavity (PC) and spleen was lower in treated mice than that in the control group. Rh G-CSF increased mainly polymorphonuclear cells (PMN) in blood and spleen. After LM inoculation, a larger number of PMN and monocyte-macrophages accumulated in PC and spleen of tested mice. In addition, PMN primed in vivo with rh G-CSF released more superoxide anions when stimulated with phorbol myristate acetate. The inhibition of bacterial growth in PC and spleen could be ascribed to the accumulation of phagocytic cells at the infection sites and the increased oxidative metabolism. The results provided further evidence of the important contribution of G-CSF and neutrophils, as target cells, to the host defence against the intracellular bacteria. PMID:1374055

  7. Porcine granulocyte-colony stimulating factor (G-CSF) delivered via replication-defective adenovirus induces a sustained increase in circulating peripheral blood neutrophils

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The use of immunomodulators is a promising area for biotherapeutic, prophylactic, and metaphylactic use to prevent and combat infectious disease, particularly during periods of peak disease incidence. Cytokines, including granulocyte colony-stimulating factor (G-CSF), are one class of compounds that...

  8. Prolactin, growth hormone, erythropoietin and granulocyte-macrophage colony stimulating factor induce MGF-Stat5 DNA binding activity.

    PubMed Central

    Gouilleux, F; Pallard, C; Dusanter-Fourt, I; Wakao, H; Haldosen, L A; Norstedt, G; Levy, D; Groner, B

    1995-01-01

    The molecular components which mediate cytokine signaling from the cell membrane to the nucleus were studied. Upon the interaction of cytokines with their receptors, members of the janus kinase (Jak) family of cytoplasmic protein tyrosine kinases and of the signal transducers and activators of transcription (Stat) family of transcription factors are activated through tyrosine phosphorylation. It has been suggested that the Stat proteins are substrates of the Jak protein tyrosine kinases. MGF-Stat5 is a member of the Stat family which has been found to confer the prolactin response. MGF-Stat5 can be phosphorylated and activated in its DNA binding activity by Jak2. The activation of MGF-Stat5 is not restricted to prolactin. Erythropoietin (EPO) and growth hormone (GH) stimulate the DNA binding activity of MGF-Stat5 in COS cells transfected with vectors encoding EPO receptor and MGF-Stat5 or vectors encoding GH receptor and MGF-Stat5. The activation of DNA binding by prolactin, EPO and GH requires the phosphorylation of tyrosine residue 694 of MGF-Stat5. The transcriptional induction of a beta-casein promoter luciferase construct in transiently transfected COS cells is specific for the prolactin activation of MGF-Stat5; it is not observed in EPO- and GH-treated cells. In the UT7 human hematopoietic cell line, EPO and granulocyte-macrophage colony stimulating factor activate the DNA binding activity of a factor closely related to MGF-Stat5 with respect to its immunological reactivity, DNA binding specificity and molecular weight. These results suggest that MGF-Stat5 regulates physiological processes in mammary epithelial cells, as well as in hematopoietic cells.(ABSTRACT TRUNCATED AT 250 WORDS) Images PMID:7744007

  9. Bone marrow cell mobilization by the systemic use of granulocyte colony-stimulating factor (GCSF) improves wound bed preparation.

    PubMed

    Iwamoto, Satori; Lin, Xiaofeng; Ramirez, Ron; Carson, Polly; Fiore, David; Goodrich, Jane; Yufit, Tatyana; Falanga, Vincent

    2013-12-01

    Innovative approaches are needed to accelerate the healing of human chronic wounds not responding to conventional therapies. An evolving and promising treatment is the use of stem cells. Our group has previously described the use of expanded (in vitro) autologous stem cells aspirated from human bone marrow and applied topically in a fibrin spray to human acute and chronic wounds. More recently, we have sought ways to mobilize stem cells directly from the bone marrow, without in vitro expansion. In this report, we show that systemic injections of granulocyte colony-stimulating factor (GCSF) can mobilize stem cells from bone marrow into the peripheral blood and then to the wound site. Our objectives were to optimize parameters for this method by using mouse models and proof of principle in a human chronic wound situation. Mice were injected for 5 days with 2 different formulations of GCSF and compared to control saline. To monitor stem cell mobilization, flow cytometric measurements of Sca-1 and c-Kit and colony-forming cell assays were performed. Full-thickness tail wounds in mice were created and monitored for healing, and polyvinyl alcohol sponges were implanted dorsally to assess collagen accumulation. To determine bone marrow stem cell homing to the wound site, chimeric mice transplanted with Green Fluorescent Protein bone marrow cells were scanned by live imaging. Additionally, as proof of principle, we tested the systemic GCSF approach in a patient with a nonhealing venous ulcer. Our findings lay the ground work and indicate that the systemic administration of GCSF is effective in mobilizing bone marrow stem cells into the peripheral blood and to the wound site. These findings are associated with an increased accumulation of collagen and promising results in terms of wound bed preparation and healing. PMID:24275756

  10. Granulocyte-macrophage colony-stimulating factor amplifies lipopolysaccharide-induced bronchoconstriction by a neutrophil- and cyclooxygenase 2-dependent mechanism.

    PubMed

    Wollin, L; Uhlig, S; Nüsing, R; Wendel, A

    2001-02-01

    Granulocyte-macrophage colony-stimulating factor (GM-CSF) is used to ameliorate neutropenia in patients after antineoplastic treatment. It has also been suggested as an adjunct treatment in septic patients; however, the recruitment and priming of leukocytes by GM-CSF bears the hazard of a hyperinflammatory response. In particular, the role of GM-CSF in pulmonary functions in septic lungs is still unclear. Therefore, we pretreated rats in vivo with GM-CSF (50 microg/kg, intravenous) and assessed the pulmonary functions of their subsequently prepared isolated perfused lungs when exposed to subtoxic concentrations of lipopolysaccharide (LPS, 2 microg/ml). These lungs showed enhanced expression of cyclooxygenase 2 (COX-2), a significant increase in thromboxane (TX) and tumor necrosis factor (TNF) release into the venous perfusate, and bronchoconstriction. COX-2 inhibition or blocking of the TX receptor abolished the GM-CSF/LPS-induced bronchoconstriction, but not the TNF release. Neutralizing antibodies against TNF did not prevent GM-CSF/LPS-induced bronchoconstriction. After GM-CSF pretreatment, massive neutrophil invasion into the lung occurred. Neutropenic rats were protected against GM-CSF/ LPS-induced lung injury. Similar results were obtained in rats pretreated with G-CSF instead of GM-CSF. We conclude that GM-CSF pretreatment exacerbates pulmonary injury by low-dose LPS via COX-2 expression, TX release, and bronchoconstriction by initiating neutrophil invasion and activation. PMID:11179120

  11. Radiation promotes invasiveness of non-small-cell lung cancer cells through granulocyte-colony-stimulating factor.

    PubMed

    Cui, Y-H; Suh, Y; Lee, H-J; Yoo, K-C; Uddin, N; Jeong, Y-J; Lee, J-S; Hwang, S-G; Nam, S-Y; Kim, M-J; Lee, S-J

    2015-10-16

    Despite ionizing radiation (IR) is being widely used as a standard treatment for lung cancer, many evidences suggest that IR paradoxically promotes cancer malignancy. However, its molecular mechanisms underlying radiation-induced cancer progression remain obscure. Here, we report that exposure to fractionated radiation (2 Gy per day for 3 days) induces the secretion of granulocyte-colony-stimulating factor (G-CSF) that has been commonly used in cancer therapies to ameliorate neutropenia. Intriguingly, radiation-induced G-CSF promoted the migratory and invasive properties by triggering the epithelial-mesenchymal cell transition (EMT) in non-small-cell lung cancer cells (NSCLCs). By irradiation, G-CSF was upregulated transcriptionally by β-catenin/TCF4 complex that binds to the promoter region of G-CSF as a transcription factor. Importantly, irradiation increased the stability of β-catenin through the activation of PI3K/AKT (phosphatidylinositol 3-kinase/AKT), thereby upregulating the expression of G-CSF. Radiation-induced G-CSF is recognized by G-CSFR and transduced its intracellular signaling JAK/STAT3 (Janus kinase/signal transducers and activators of transcription), thereby triggering EMT program in NSCLCs. Taken together, our findings suggest that the application of G-CSF in cancer therapies to ameliorate neutropenia should be reconsidered owing to its effect on cancer progression, and G-CSF could be a novel therapeutic target to mitigate the harmful effect of radiotherapy for the treatment of NSCLC. PMID:25639867

  12. Is primary prophylaxis with granulocyte colony stimulating factor (G-CSF) indicated in the treatment of lymphoma?

    PubMed

    Kansara, Roopesh; Kumar, Rajat; Seftel, Matthew

    2013-08-01

    Febrile neutropenia (FN) is a common complication of cancer therapy. It can contribute to delays in treatment, increased rates of hospitalization, and severe infections. FN may also hinder completion of intended chemotherapy. Granulocyte colony stimulating factors (G-CSF) lower the rates of FN, infections, and hospitalization. Multiple national and international guidelines advocate the use of G-CSF in primary prophylaxis if the overall risk of FN is >20% (accounting for both patient and treatment-related risks). Lymphoma specific guidelines recommend G-CSF use in similar fashion. However, based on our updated review of published literature, we note that primary prophylaxis (PP) with G-CSF fails to improve overall survival as well as infection-related mortality. Moreover, lymphoma specific cost-effectiveness analyses on the use of PP have shed further doubt on the optimal use of this myeloid growth factor. In this general review, we will discuss whether PP with GCSF has any role in the management of adults with non-Hodgkin lymphoma. PMID:23768687

  13. 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. PMID:25432887

  14. 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. PMID:18997822

  15. A Novel Combinatorial Therapy With Pulp Stem Cells and Granulocyte Colony-Stimulating Factor for Total Pulp Regeneration

    PubMed Central

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

    2013-01-01

    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. PMID:23761108

  16. Total absence of colony-stimulating factor 1 in the macrophage-deficient osteopetrotic (op/op) mouse.

    PubMed Central

    Wiktor-Jedrzejczak, W; Bartocci, A; Ferrante, A W; Ahmed-Ansari, A; Sell, K W; Pollard, J W; Stanley, E R

    1990-01-01

    Osteopetrotic (op/op) mutant mice suffer from congenital osteopetrosis due to a severe deficiency of osteoclasts. Furthermore, the total number of mononuclear phagocytes is extremely low in affected mice. Serum, 11 tissues, and different cell and organ conditioned media from op/op mice were shown to be devoid of biologically active colony-stimulating factor 1 (CSF-1), whereas all of these preparations from littermate control +/+ and +/op mice contained the growth factor. The deficiency was specific for CSF-1 in that serum or conditioned media from op/op mice possessed elevated levels of at least three other macrophage growth factors. Partial correction of the op/op defect was observed following intraperitoneal implantation of diffusion chambers containing L929 cells, which in culture produce CSF-1 as their sole macrophage growth factor. No rearrangement of the CSF-1 gene in op/op mice was detected by Southern analysis. However, in contrast to control lung fibroblasts, which contained 4.6- and 2.3-kilobase CSF-1 mRNAs, only the 4.6-kilobase species was detected in op/op cells. An alteration in the CSF-1 gene is strongly implicated as the primary defect in op/op mice because they do not contain detectable CSF-1, their defect is correctable by administration of CSF-1, the op locus and the CSF-1 gene map within the same region of mouse chromosome 3, their CSF-1 mRNA biosynthesis is altered, and the op/op phenotype is consistent with the phenotype expected in a CSF-1 deficient mouse. Images PMID:2191302

  17. Effect of granulocyte colony-stimulating factor administration on renal regeneration after experimentally induced acute kidney injury in dogs.

    PubMed

    Lim, Chae-Young; Han, Jae-Ik; Kim, Seung-Gon; Lee, Chang-Min; Park, Hee-Myung

    2016-02-01

    OBJECTIVE To evaluate the effects of granulocyte colony-stimulating factor (GCSF) administration in dogs with experimentally induced acute kidney injury. ANIMALS 6 healthy dogs. PROCEDURES After induction of kidney injury (day 0) with cisplatin (5 mg/kg, IV), the dogs were randomly assigned into 2 groups (n = 3 dogs/group). Then dogs immediately received GCSF (10 μg/kg) or 1 mL of saline (0.9% NaCl) solution (control group) SC; this treatment was repeated once daily for 4 additional days (days 1 through 4). A once-daily CBC (day 0 to 4), serum biochemical analysis (day 0 to 3), and urinalysis (day 0 to 3) were performed for each dog; samples were collected before administration of cisplatin (day 0) and before treatment with GCSF or saline solution (days 1 through 4). After sample collection and treatment on day 4, all dogs were euthanized; kidney tissue samples underwent histologic evaluation, immunohistochemical analyses, and cytokine profiling via reverse transcriptase PCR assay. RESULTS In the GCSF-treated group, the histologic evaluation and immunohistochemical analyses of kidney tissue revealed less fibrotic change and greater proliferation of renal tubular epithelial cells, compared with findings in the control group. The mRNA profiles of kidney tissue from the GCSF-treated group indicated lower expression of tumor necrosis factor-α and tumor growth factor-β, compared with findings in the control group; however, concentrations of factors related to renal regeneration were not greater in the GCSF-treated group. CONCLUSIONS AND CLINICAL RELEVANCE Results indicated that GCSF treatment can impede renal fibrosis and increase proliferation of renal tubules after experimentally induced acute kidney injury in dogs. (Am J Vet Res 2016;77:199-207). PMID:27027715

  18. Long-term evaluation of granulocyte-colony stimulating factor on hypoxic-ischemic brain damage in infant rats

    PubMed Central

    Fathali, Nancy; Lekic, Tim; Zhang, John H.

    2011-01-01

    Purpose Hypoxia-ischemia (HI), as a major cause of fetal brain damage, has long-lasting neurological implications. Therefore, therapeutic interventions that attenuate the neuropathological outcome of HI while also improving the neuro-functional outcome are of paramount clinical importance. The aim of this study was to investigate the long-term functional and protective actions of granulocyte-colony stimulating factor (G-CSF) treatment in an experimental model of cerebral HI. Methods Postnatal day-7 Sprague-Dawley rats were subjected to HI surgery, which entailed ligation of the right common carotid artery followed by 2 h of hypoxia (8% O2). Treatment consisted of subcutaneous injection of G-CSF at 1 h after hypoxia followed by an additional one injection per day for 5 days (6 total injections) or for 10 days (11 total injections). Animals were euthanized 5 weeks post-insult for extensive evaluation of neurological deficits and assessment of brain, spleen, heart, and liver damage. Results G-CSF treatment promoted somatic growth and prevented brain atrophy and under-development of the heart. Moreover, reflexes, limb placing, muscle strength, motor coordination, short-term memory, and exploratory behavior were all significantly improved by both G-CSF dosing regimens. Conclusions Long-term neuroprotection afforded by G-CSF in both morphological and functional parameters after a hypoxic-ischemic event in the neonate provides a rationale for exploring clinical translation. PMID:20461500

  19. 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

  20. An open-label trial of granulocyte macrophage colony stimulating factor therapy for moderate symptomatic pulmonary alveolar proteinosis.

    PubMed

    Venkateshiah, Saiprakash B; Yan, Tom D; Bonfield, Tracey L; Thomassen, Mary Jane; Meziane, Moulay; Czich, Carmen; Kavuru, Mani S

    2006-07-01

    Pulmonary alveolar proteinosis (PAP) is a rare idiopathic autoimmune lung disease in adults characterized by the accumulation of lipoproteinaceous material within the alveoli of the lung. The natural history of this disease is poorly defined. Current therapy of bilateral whole-lung lavage (WLL) under general anesthesia is invasive and has its limitations. Data suggest that relative granulocyte macrophage colony stimulating factor (GM-CSF) deficiency may be involved in the pathogenesis of this disease. There have been several case series that have described clinical improvement with exogenous GM-CSF therapy in a subset of patients with PAP. We describe the results of a prospective, open-label clinical trial of daily subcutaneous GM-CSF therapy in a group of adult patients with idiopathic PAP. In this series of 25 patients, the largest reported to date, administration of GM-CSF improved oxygenation as assessed by a 10 mm Hg decrease in alveolar-arterial oxygen gradient, as well as improvement in other clinical and quality of life parameters in 12 of 25 patients (48%) with moderate symptomatic disease who completed the trial. In addition, the serum anti-GM-CSF antibody titer correlated with lung disease activity and was a predictor for responsiveness to therapy. These data indicate that subcutaneous GM-CSF therapy is a promising alternative to WLL for symptomatic patients with PAP. PMID:16840407

  1. 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

  2. Analytical characterization of in vitro refolding in the quality by design paradigm: Refolding of recombinant human granulocyte colony stimulating factor.

    PubMed

    Pathak, Mili; Dixit, Shruti; Muthukumar, S; Rathore, Anurag S

    2016-07-15

    Protein based therapeutics dominate most pharmaceutical pipelines today. For a therapeutic product to be effective, it is important that it is in its native form as slight modifications have been known to result in significantly different performance in the clinic. When expressed in hosts such as Escherichia coli, formation of inactive insoluble aggregates of proteins popularly known as inclusion bodies occurs in most cases. This necessitates the need for in vitro refolding to generate the native (and active) form of the therapeutic protein. This paper aims to provide an approach to generate a deeper understanding of refolding of a therapeutic protein and then to use it for its optimal production commercially. Recombinant human granulocyte colony stimulating factor has been chosen as the model protein. Seven orthogonal analytical tools have been used to elucidate the refolding process. By strategically using these tools protein refolding has been segregated into a series of well-defined sequence of events, starting from the unfolded random coil and ending with the uniquely folded metastable state. The study also suggests the choice of tools that can be used to monitor each event. We believe that this paper successfully demonstrates an approach to generate deeper understanding of the protein refolding process as per the expectations laid out in the Quality by Design paradigm. PMID:27206104

  3. Kinetics of Neutrophils in Mice Exposed to Radiation and/or Granulocyte Colony-Stimulating Factor Treatment

    PubMed Central

    Romero-Weaver, A. L.; Wan, X. S.; Diffenderfer, E. S.; Lin, L.; Kennedy, A. R.

    2014-01-01

    Astronauts have the potential to develop the hematopoietic syndrome as a result of exposure to radiation from a solar particle event (SPE) during exploration class missions. This syndrome is characterized by a reduction in the number of circulating blood cells (cytopenias). In the present study the effects of SPE-like proton and γ radiation on the kinetics of circulating neutrophils were evaluated during a one-month time period using mice as a model system. The results revealed that exposure to a 2 Gy dose of either SPE-like proton or γ radiation significantly decreased the number of circulating neutrophils, with two nadirs observed on day 4 and day 16 postirradiation. Low circulating neutrophil count (neutropenia) is particularly important because it can increase the risk of astronauts developing infections, which can compromise the success of the mission. Thus, two granulocyte colony-stimulating factors (G-CSFs), filgrastim and pegfilgrastim were evaluated as countermeasures for this endpoint. Both forms of G-CSF significantly increased neutrophil counts in irradiated mice, however, the effect of pegfilgrastim was more potent and lasted longer than filgrastim. Using the expression of CD11b, CD18 and the production of reactive oxygen species (ROS) as markers of neutrophil activation, it was determined that the neutrophils in the irradiated mice treated with pegfilgrastim were physiologically active. Thus, these results suggest that pegfilgrastim could be a potential countermeasure for the reduced number of circulating neutrophils in irradiated animals. PMID:23829559

  4. 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

  5. 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

  6. Granulocyte macrophage colony-stimulating factor treatment results in recovery of motor function after white matter damage in mice.

    PubMed

    Theoret, Jennifer K; Jadavji, Nafisa M; Zhang, Min; Smith, Patrice D

    2016-01-01

    Clinical stroke usually results from a cerebral ischaemic event, and is frequently a debilitating condition with limited treatment options. A significant proportion of clinical strokes result from specific damage to the subcortical white matter (SWM), but currently there are few animal models available to investigate the pathogenesis and potential therapeutic strategies to promote recovery. Granulocyte macrophage colony-stimulating factor (GM-CSF) is a cytokine that has been previously shown to promote neuroprotective effects after brain damage; however, the mechanisms mediating this effect are not known. Here, it is reported that GM-CSF treatment results in dramatic functional improvement in a white matter model of stroke in mice. SWM stroke was induced in mice by unilateral injections of the vasoconstrictor, endothelin-1 (ET-1). The results reveal that ET-1-induced stroke impairs skilled motor function on the single pellet-reaching task and results in forelimb asymmetry, in adult mice. Treatment with GM-CSF, after stroke, restores motor function and abolishes forelimb asymmetry. The results also indicate that GM-CSF promotes its effects by activating mammalian target of rapamycin signalling mechanisms in the brain following stroke injury. Additionally, a significant increase in GM-CSF receptor expression was found in the ipsilateral hemisphere of the ET-1-injected brain. Taken together, the present study highlights the use of an under-utilized mouse model of stroke (using ET-1) and suggests that GM-CSF treatment can attenuate ET-1-induced functional deficits. PMID:26474338

  7. 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.

  8. Colony-Stimulating Factor 2 (CSF-2) Improves Development and Posttransfer Survival of Bovine Embryos Produced in Vitro

    PubMed Central

    Loureiro, Bárbara; Bonilla, Luciano; Block, Jeremy; Fear, Justin M.; Bonilla, Aline Q. S.; Hansen, Peter J.

    2009-01-01

    In this study, we tested the role of colony-stimulating factor 2 (CSF2) as one of the regulatory molecules that mediate maternal effects on embryonic development during the preimplantation period. Our objective was to verify effects of CSF2 on blastocyst yield, determine posttransfer survival, and evaluate properties of the blastocyst formed after CSF2 treatment. In vitro, CSF2 increased the percentage of oocytes that became morulae and blastocysts. Blastocysts that were treated with CSF2 tended to have a greater number of inner cell mass cells and had a higher ratio of inner cell mass to trophectoderm cells. There was no effect of CSF2 on the incidence of apoptosis. Treatment with CSF2 from d 5 to 7 after insemination increased embryonic survival as indicated by improved pregnancy rate at d 30–35 of gestation. Moreover, treatment with CSF2 from either d 1–7 or 5–7 after insemination reduced pregnancy loss after d 30–35. Results indicate that treatment with CSF2 can affect embryonic development and enhance embryo competence for posttransfer survival. The fact that treatment with CSF2 during such a narrow window of development altered embryonic function much later in pregnancy suggests that CSF2 may exert epigenetic effects on the developing embryo that result in persistent changes in function during the embryonic and fetal periods of development. PMID:19797121

  9. Nuclear proteins interacting with the promoter region of the human granulocyte/macrophage colony-stimulating factor gene

    SciTech Connect

    Shannon, M.F.; Gamble, J.R.; Vadas, M.A.

    1988-02-01

    The gene for human granulocyte/macrophage colony-stimulating factor (GM-CSF) is expressed in a tissue-specific as well as an activation-dependent manner. The interaction of nuclear proteins with the promoter region of the GM-CSF gene that is likely to be responsible for this pattern of GM-CSF expression was investigated. The authors show that nuclear proteins interact with DNA fragments from the GM-CSF promoter in a cell-specific manner. A region spanning two cytokine-specific sequences, cytokine 1 (CK-1, 5', GAGATTCCAC 3') and cytokine 2 (CK-2, 5' TCAGGTA 3') bound two nuclear proteins from GM-CSF-expressing cells in gel retardation assays. NF-GMb was inducible with phorbol 12-myristate 13-acetate and accompanied induction of GM-CSF message. NF-GMb was absent in cell lines not producing GM-CSF, some of which had other distinct binding proteins. NF-GMa and NF-GMb eluted from a heparin-Sepharose column at 0.3 and 0.6 M KCl, respectively. They hypothesize that the sequences CK-1 and CK-2 bind specific proteins and regulate GM-CSF transcription.

  10. A single injection of polyethylene-glycol granulocyte colony-stimulating factor strongly prolongs survival of mice with systemic candidiasis.

    PubMed

    van Spriel, A B; van den Herik-Oudijk, I E; van de Winkel, J G

    2000-06-01

    Systemic candidiasis is a life-threatening disease occurring in immunocompromized patients. Granulocyte colony-stimulating factor (G-CSF) reduces mortality in experimental invasive candidiasis. Covalent conjugation of polyethylene-glycol (peg) to proteins increases their stability and in vivo bioactivity. In this study, the effect of a single subcutaneous injection of peg-G-CSF on lethal candidiasis was assessed. This was performed in acute and chronic candidiasis models in non-neutropenic FVB/N mice. Peg-G-CSF rapidly increased circulating polymorphonuclear leukocyte (PMNL) numbers in mice, sustaining high for >4 days. Candida albicans outgrowth from kidneys of infected mice was strongly reduced after peg-G-CSF treatment (5.76 log cfu/g kidney vs 7.66 control), with absence of hyphal outgrowth and enhanced PMNL influx. Moreover, peg-G-CSF increased survival of C. albicans -infected mice, whereas efficacy of uncoupled G-CSF was obtained only after repeated treatment. These data document a potent in vivo biological effect of peg-G-CSF, resulting in strongly enhanced resistance against systemic candidiasis. PMID:10843742

  11. Using Hydrogen/Deuterium Exchange Mass Spectrometry to Study Conformational Changes in Granulocyte Colony Stimulating Factor upon PEGylation

    PubMed Central

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

    2012-01-01

    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 though 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. PMID:22227798

  12. Granulocyte colony-stimulating factor (G-CSF) administration in individuals with sickle cell disease: time for a moratorium?

    PubMed

    Fitzhugh, Courtney D; Hsieh, Matthew M; Bolan, Charles D; Saenz, Carla; Tisdale, John F

    2009-01-01

    Granulocyte colony-stimulating factor (G-CSF) is used commonly in an attempt to reduce the duration of neutropenia and hospitalization in patients undergoing chemotherapy and to obtain hematopoietic stem cells (HSC) for transplantation applications. Despite the relative safety of administration of G-CSF in most individuals, including subjects with sickle cell trait, severe and life-threatening complications have been reported when used in individuals with sickle cell disease (SCD), including those who were asymptomatic and undiagnosed prior to administration. The administration of G-CSF has now been reported in a total of 11 individuals with SCD. Seven developed severe adverse events, including vaso-occlusive episodes, acute chest syndrome, multi-organ system failure and death. Precautions, including minimizing the peak white blood cell count, dividing or reducing the G-CSF dose and red blood cell transfusions to reduce sickle hemoglobin (HbS) levels, have been employed with no consistent benefit. These reported data indicate that administration of G-CSF in individuals with SCD should be undertaken only in the absence of alternatives and after full disclosure of the risks involved. Unless further data demonstrate safety, routine usage of G-CSF in individuals with SCD should be avoided. PMID:19513902

  13. Fully Synthetic Granulocyte Colony-Stimulating Factor Enabled by Isonitrile-Mediated Coupling of Large, Side-Chain-Unprotected Peptides.

    PubMed

    Roberts, Andrew G; Johnston, Eric V; Shieh, Jae-Hung; Sondey, Joseph P; Hendrickson, Ronald C; Moore, Malcolm A S; Danishefsky, Samuel J

    2015-10-14

    Human granulocyte colony-stimulating factor (G-CSF) is an endogenous glycoprotein involved in hematopoiesis. Natively glycosylated and nonglycosylated recombinant forms, lenograstim and filgrastim, respectively, are used clinically to manage neutropenia in patients undergoing chemotherapeutic treatment. Despite their comparable therapeutic potential, the purpose of O-linked glycosylation at Thr133 remains a subject of controversy. In light of this, we have developed a synthetic platform to prepare G-CSF aglycone with the goal of enabling access to native and designed glycoforms with site-selectivity and glycan homogeneity. To address the synthesis of a relatively large, aggregation-prone sequence, we advanced an isonitrile-mediated ligation method. The chemoselective activation and coupling of C-terminal peptidyl Gly thioacids with the N-terminus of an unprotected peptide provide ligated peptides directly in a manner complementary to that with conventional native chemical ligation-desulfurization strategies. Herein, we describe the details and application of this method as it enabled the convergent total synthesis of G-CSF aglycone. PMID:26401918

  14. 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

  15. 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.

  16. Structure of macrophage colony stimulating factor bound to FMS: Diverse signaling assemblies of class III receptor tyrosine kinases

    SciTech Connect

    Chen, Xiaoyan; Liu, Heli; Focia, Pamela J.; Shim, Ann Hye-Ryong; He, Xiaolin

    2009-06-12

    Macrophage colony stimulating factor (M-CSF), through binding to its receptor FMS, a class III receptor tyrosine kinase (RTK), regulates the development and function of mononuclear phagocytes, and plays important roles in innate immunity, cancer and inflammation. We report a 2.4 {angstrom} crystal structure of M-CSF bound to the first 3 domains (D1-D3) of FMS. The ligand binding mode of FMS is surprisingly different from KIT, another class III RTK, in which the major ligand-binding domain of FMS, D2, uses the CD and EF loops, but not the {beta}-sheet on the opposite side of the Ig domain as in KIT, to bind ligand. Calorimetric data indicate that M-CSF cannot dimerize FMS without receptor-receptor interactions mediated by FMS domains D4 and D5. Consistently, the structure contains only 1 FMS-D1-D3 molecule bound to a M-CSF dimer, due to a weak, hydrophilic M-CSF:FMS interface, and probably a conformational change of the M-CSF dimer in which binding to the second site is rendered unfavorable by FMS binding at the first site. The partial, intermediate complex suggests that FMS may be activated in two steps, with the initial engagement step distinct from the subsequent dimerization/activation step. Hence, the formation of signaling class III RTK complexes can be diverse, engaging various modes of ligand recognition and various mechanistic steps for dimerizing and activating receptors.

  17. Functional Relationship between Tumor-Associated Macrophages and Macrophage Colony-Stimulating Factor as Contributors to Cancer Progression

    PubMed Central

    Laoui, Damya; Van Overmeire, Eva; De Baetselier, Patrick; Van Ginderachter, Jo A.; Raes, Geert

    2014-01-01

    The current review article describes the functional relationship between tumor-associated macrophages (TAM) as key cellular contributors to cancer malignancy on the one hand and macrophage-colony-stimulating factor (M-CSF or CSF-1) as an important molecular contributor on the other. We recapitulate the available data on expression of M-CSF and the M-CSF receptor (M-CSFR) in human tumor tissue as constituents of a stromal macrophage signature and on the limits of the predictive and prognostic value of plasma M-CSF levels. After providing an update on current insights into the nature of TAM heterogeneity at the level of M1/M2 phenotype and TAM subsets, we give an overview of experimental evidence, based on genetic, antibody-mediated, and pharmacological disruption of M-CSF/M-CSFR signaling, for the extent to which M-CSFR signaling can not only determine the TAM quantity, but can also contribute to shaping the phenotype and heterogeneity of TAM and other related tumor-infiltrating myeloid cells (TIM). Finally, we review the accumulating information on the – sometimes conflicting – effects blocking M-CSFR signaling may have on various aspects of cancer progression such as tumor growth, invasion, angiogenesis, metastasis, and resistance to therapy and we thereby discuss in how far these different effects actually reflect a contribution of TAM. PMID:25339957

  18. 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

  19. Granulocyte colony-stimulating factor as a potential inducer of ovulation in infertile women with luteinized unruptured follicle syndrome.

    PubMed

    Shibata, Takeo; Makinoda, Satoru; Waseda, Tomoo; Tomizawa, Hideki; Fujii, Ryota; Utsunomiya, Takafumi

    2016-05-01

    Luteinized unruptured follicle (LUF) syndrome is one of the intractable ovulation disorders that are commonly observed during cycles of treatment with ovulation inducers, for which no effective therapy other than assisted reproductive technology is available. Here, we investigated whether granulocyte colony-stimulating factor (G-CSF) could prevent the onset of LUF syndrome. We analyzed the effects of G-CSF in 68 infertile women with LUF syndrome who received ovulation induction (clomiphene + human chorionic gonadotropin [hCG] therapy or follicle-stimulating hormone + hCG therapy). G-CSF (lenograstim, 100 μg) was administered subcutaneously. Onsets of LUF syndrome were compared between the cycle during which G-CSF was given in combination with the ovulation inducer (ie, the G-CSF treatment cycle) and the subsequent cycle during which only the ovulation inducer was given (ie, the G-CSF nontreatment control cycle). The results showed that LUF syndrome recurred in only 3 cycles during the G-CSF treatment cycle (4.4% [3/68 cycles]), whereas LUF syndrome recurred in 13 cycles during the subsequent G-CSF nontreatment control cycle (19.1% [13/68 cycles]). The additional use of G-CSF significantly prevented the onset of LUF syndrome during ovulation induction (P = 0.013, McNemar test). No serious adverse reactions because of the administration of G-CSF were observed. In conclusion, our findings indicate that G-CSF may become a useful therapy for LUF syndrome. PMID:26518992

  20. 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

  1. Changes in the proteomic profile during differentiation and maturation of human monocyte-derived dendritic cells stimulated with granulocyte macrophage colony stimulating factor/interleukin-4 and lipopolysaccharide.

    PubMed

    Pereira, Sandra Rodrigues; Faça, Vitor Marcel; Gomes, Glauce Gaspar; Chammas, Roger; Fontes, Aparecida Maria; Covas, Dimas Tadeu; Greene, Lewis Joel

    2005-04-01

    Dendritic cells (DCs) are highly specialized antigen-presenting cells that play an essential role in the immune response. We used the proteomic approach based on two-dimensional gel electrophoresis and mass spectrometry to identify the protein changes that occur during differentiation of DCs from monocytes (Mo) stimulated with granulocyte macrophage colony stimulating factor/interleukin-4 (GM-CSF/IL-4) and during the maturation of immature DCs stimulated with lipopolysaccharide. Sixty-three differentially expressed proteins (+/- two-fold) were unambiguously identified with sequence coverage greater than 20%. They corresponded to only 36 different proteins, because 11 were present as 38 electrophoretic forms. Some proteins such as tropomyosin 4 and heat shock protein 71 presented differentially expressed electrophoretic forms, suggesting that many of the changes in protein expression that accompany differentiation and maturation of DCs occur in post-translationally modified proteins. The largest differences in expression were observed for actin (21-fold in Mo), Rho GDP-dissociation inhibitor 2 (20-fold in Mo), vimentin (eight-fold in immature DCs), lymphocyte-specific protein 1 (12-fold in mature DCs) and thioredoxin (14-fold in mature DCs). Several proteins are directly related to functional and morphological characteristics of DCs, such as cytoskeletal proteins (cytoskeleton rearrangement) and chaperones (antigen processing and presentation), but other proteins have not been assigned specific functions in DCs. Only a few proteins identified here were the same as those reported in proteomic studies of DCs, which used different stimuli to produce the cells (GM-CSF/IL-4 and tumor necrosis factor-alpha). These data suggest that the DC protein profile depends on the stimuli used for differentiation and especially for maturation. PMID:15800872

  2. 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

  3. Adjuvant granulocyte colony-stimulating factor therapy results in improved spatial learning and stimulates hippocampal neurogenesis in a mouse model of pneumococcal meningitis.

    PubMed

    Schmidt, Anna Kathrin; Reich, Arno; Falkenburger, Björn; Schulz, Jörg B; Brandenburg, Lars Ove; Ribes, Sandra; Tauber, Simone C

    2015-01-01

    Despite the development of new antibiotic agents, mortality of pneumococcal meningitis remains high. In addition, meningitis results in severe long-term morbidity, most prominently cognitive deficits. Granulocyte colony-stimulating factor (G-CSF) stimulates proliferation and differentiation of hematopoietic progenitor cells and increases the number of circulating neutrophil granulocytes. This study investigated the effect of adjuvant G-CSF treatment on cognitive function after pneumococcal meningitis. C57BL/6 mice were infected by subarachnoid injection of Streptococcus pneumoniae serotype 3 and treated with ceftriaxone and G-CSF subcutaneously or ceftriaxone alone for 5 days. Clinical scores, motor performance, and mortality during bacterial meningitis were unaffected by adjuvant G-CSF treatment. No effect of G-CSF treatment on production of proinflammatory cytokines or activation of microglia or astrocytes was observed. The G-CSF treatment did, however, result in hippocampal neurogenesis and improved spatial learning performance 6 weeks after meningitis. These results suggest that G-CSF might offer a new adjuvant therapeutic approach in bacterial meningitis to reduce long-term cognitive deficits. PMID:25470346

  4. Colony-stimulating factors and peripheral blood progenitor cell transplantation. Benefits and costs.

    PubMed

    Uyl-de Groot, C A; Huijgens, P C; Rutten, F F

    1996-07-01

    High dosage chemo- or radiotherapy followed by the administration of autologous bone marrow-derived stem cells [i.e. autologous bone marrow transplantation (ABMT)] is an established protocol for treating acute myeloid leukaemia and malignant lymphoma. The approach is also under investigation in the treatment of acute lymphocytic leukaemia, multiple myeloma and solid tumours. In all of these diseases, the optimisation of indications, conditioning schemes, stem cell harvest techniques and supportive care with growth factors is subject to continuous preclinical research and clinical phase II and III studies. Recently, the administration of peripheral blood stem cell preparations to cancer patients as rescue therapy after high dosage antitumour therapy has been received with much enthusiasm. At first glance, the technique looks rather easy to perform. The faster recovery of haemopoiesis, compared with ABMT, leads to shorter durations of hospitalisation. Moreover, in most studies, peripheral blood progenitor cell transplantation (PBPCT) resulted in fewer septic episodes, fewer intensive care admissions, fewer red blood cell and platelet transfusions, reduced use of anti-infectives and parenteral nutrition, and reduced hospital costs compared with ABMT. The overall conclusion is that the treatment costs of PBPCT are 15 to 30% lower than the treatment costs of ABMT. However, a formal comparison between PBPCT and ABMT, assessing the differences in toxicity, costs and quality of life, is still awaited. PMID:10160468

  5. 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

  6. Transcriptional regulation of mouse granulocyte-macrophage colony-stimulating factor/IL-3 locus

    SciTech Connect

    Osborne, C.S.; Vadas, M.A.; Cockerill, P.N.

    1995-07-01

    Granulocyte-macrophage (GM)-CSF and IL-3 are hemopoietic growth factors whose genes are closely linked in both humans and mice. In humans, the GM-CSF and IL-3 genes are regulated by a cyclosporin A-inhibitable enhancer located 3 kb upstream of the GM-CSF gene that is inducible by signals that mimic TCR activation. To search for a murine homologue of this enhancer we probed mouse genomic DNA and located a 400-bp element 2 kb upstream of the mouse GM-CSF gene that was 76% homologous with the human GM-CSF enhancer. Like the human GM-CSF enhancer, this element formed a cyclosporin A-inhibitable DNase I-hypersensitive site in the murine T cell line EL4 upon activation with phorbol ester and calcium ionophore. Transient transfection assays showed that this homologue of the human enhancer acted as an inducible enhancer of the thymidine kinase promoter, the mouse IL-3 promoter, and the human GM-CSF promoter. We observed, however, that the mouse GM-CSF promoter was significantly more active than the human GM-CSF promoter and found that it supported a level of activity equivalent to the combination of the human GM-CSF promoter and the human GM-CSF enhancer. Consequently, the activity of mouse GM-CSF promoter was not significantly elevated in the presence of the mouse GM-CSF enhancer. Because the mouse GM-CSF enhancer is considerably less active than its human homologue we suggest that the mouse GM-CSF gene has evolved with less dependence upon the upstream enhancer for its activation. 53 refs., 6 figs.

  7. Effect of granulocyte-macrophage colony-stimulating factor on hepatic regeneration after 70% hepatectomy in normal and cirrhotic rats

    PubMed Central

    Demirci, S; Akbulut, H; Sever, N; Demirer, S; Ünal, AE

    2002-01-01

    Background Post-hepatectomy liver insufficiency is one of the most serious postoperative problems and its prevention is important after major hepatic resection, especially in the cirrhotic liver. Some growth factors and cytokines appear to play important roles in liver regeneration. In the present study we have investigated the effects of granulocyte-macrophage colony-stimulating factor (GM-CSF) on hepatic regeneration after 70% partial hepatectomy (PH) in cirrhotic and non-cirrhotic rats. Methods A rat model of liver cirrhosis was prepared using thioacetamide (TAA) (a dose of 20 mg/100 g body w, intra-peritoneally) on three days a week for 12 weeks. Adult male rats were divided into four groups:Group 1 (n=10) no cirrhosis and no GM-CSF; Group 2 (n=10) no cirrhosis and GM-CSF; Group 3 (n=10) cirrhosis and no GM-CSF; and Group 4 (n=10) cirrhosis and GM-CSF. All the rats underwent a 70% hepatectomy, and GM-CSF was administrated immediately after operation in Groups 2 and 4. On postoperative days 2 and 7, fresh samples from the remnant liver were obtained to evaluate its regenerative capacity.The liver regenerative process was estimated by DNA synthesis, using flow cytometry. Results Proliferation index (PI) of hepatocytes at 48 h was higher in Group 4 rats than Group 3 rats (p<0.05). On postoperative day 7, PI was elevated in Group 3 rats compared with Group 4 rats, but this difference was not statistically significant. In non-cirrhotic rats given GM-CSF, PI was increased compared with Group 1 rats at day 2 (p<0.05), but not at day 7. Conclusions The findings suggest that the proliferative capacity of liver cells is impaired and delayed after 70% PH in cirrhotic rat liver. GM-CSF administration might enhance the liver PI in both normal and TAA-induced cirrhotic rats. PMID:18332927

  8. Treatment with recombinant granulocyte colony-stimulating factor (Filgrastin) stimulates neutrophils and tissue macrophages and induces an effective non-specific response against Mycobacterium avium in mice.

    PubMed Central

    Bermudez, L E; Petrofsky, M; Stevens, P

    1998-01-01

    A role of neutrophils in the host response against Mycobacterium avium (MAC) has recently been suggested. To investigate this matter further, we determined the effect of granulocyte colony-stimulating factor (G-CSF) on the outcome of MAC infection in mice. C57BL/6bg+/bg- black mice were intravenously infected with 1 x 10(7) MAC and then divided into four experimental groups to receive G-CSF as follows: (i) 10 micrograms/kg/day; (ii) 50 micrograms/kg/day; (iii) 100 micrograms/kg/day; (iv) placebo control. Mice were killed at 2 and 4 weeks of treatment to determine the bacterial load of liver and spleen. Treatment with G-CSF at both 10 and 50 micrograms/kg/day doses significantly decreased the number of viable bacteria in liver and spleen after 2 weeks (approximately 70.5% and 69.0%, respectively), and after 4 weeks (approximately 53% and 52%, respectively, P < 0.05 compared with placebo control). Treatment with 100 micrograms/kg/day did not result in decrease of bacterial colony-forming units in the liver and spleen after 4 weeks. Administration of G-CSF induced interleukin-10 (IL-10) and IL-12 production by splenocytes. To examine if the protective effect of G-CSF was accompanied by the activation of phagocytic cells, blood neutrophils and splenic macrophages were purified from mice receiving G-CSF and their ability to kill MAC was examined ex vivo. Neutrophils and macrophages from G-CSF-treated mice were able to inhibit the growth of or to kill MAC ex vivo, while phagocytic cells from untreated control mice had no anti-MAC effect. These results suggest that activation of neutrophils appears to induce an effective non-specific host defence against MAC, and further studies should aim for better understanding of the mechanisms of protection. Images Figure 3 Figure 4 Figure 5 PMID:9767410

  9. Pulmonary alveolar proteinosis: a bench-to-bedside story of granulocyte-macrophage colony-stimulating factor dysfunction.

    PubMed

    Greenhill, Sara R; Kotton, Darrell N

    2009-08-01

    Pulmonary alveolar proteinosis (PAP) is a rare disorder characterized by ineffective clearance of surfactant by alveolar macrophages. Through recent studies with genetically altered mice, the etiology of this idiopathic disease is becoming clearer. Functional deficiency of granulocyte-macrophage colony-stimulating factor (GM-CSF) appears to contribute to disease pathogenesis because mutant mice deficient in GM-CSF or its receptor spontaneously develop PAP. Recent human studies further suggest a connection between PAP and defective GM-CSF activity because inactivating anti-GM-CSF autoantibodies are observed in all patients with idiopathic PAP, and additional rare cases of PAP in children have been accompanied by genetic defects in the alpha chain of the GM-CSF receptor. In patients and mouse models of PAP, deficient GM-CSF activity appears to result in defective alveolar macrophages that are unable to maintain pulmonary surfactant homeostasis and display defective phagocytic and antigen-presenting capabilities. The most recent studies also suggest that neutrophil dysfunction additionally contributes to the increased susceptibility to lung infections seen in PAP. Because the phenotypic and immunologic abnormalities of PAP in mouse models can be corrected by GM-CSF reconstituting therapies, early clinical trials are underway utilizing administration of GM-CSF to potentially treat human PAP. The development of novel treatment approaches for PAP represents a dramatic illustration in pulmonary medicine of the "bench-to-bedside" process, in which basic scientists, translational researchers, and clinicians have joined together to rapidly take advantage of the unexpected observations frequently made in the modern molecular biology research laboratory. PMID:19666756

  10. Dermatitis during radiation for vulvar carcinoma: prevention and treatment with granulocyte-macrophage colony-stimulating factor impregnated gauze.

    PubMed

    Kouvaris, J R; Kouloulias, V E; Plataniotis, G A; Balafouta, E J; Vlahos, L J

    2001-01-01

    The aim of this study was to determine the effectiveness of granulocyte-macrophage colony-stimulating factor (GM-CSF) impregnated gauze in preventing or healing radiation-induced dermatitis. Sixty-one patients were irradiated for vulvar carcinoma. Thirty-seven applied steroid cream at irradiated areas throughout radiotherapy (Group A) and 24 patients applied additionally GM-CSF impregnated gauze (40 micrcog/cm2 of skin-irradiated area, twice per day) in addition to the steroid cream, after 20 Gy of irradiation (Group B). The score of skin reactions (P=0.008, chi2 test) and the time interval of radiotherapy interruption (P=0.037, Mann-Whitney U test) were statistically significantly reduced in Group B patients. Multivariate analysis of variance showed for this group not only a significant reduction in the Sum of Gross Dermatitis Scoring (P<0.001, adjusted for Duration of Dermatitis) but also a significant reduction of the healing time (P=0.02, adjusted for Sum of Gross Dermatitis Scoring). The pain grading was less (P=0.014, chi2 test) and pain reduction was noticed sooner after the application of GM-CSF impregnated gauze (P=0.0017, Mann-Whitney U test). Multivariate logistic regression analysis showed that the only significant effect on dermatitis score is due to Body Mass Index (P=0.034) and the application of GM-CSF (P=0.008). GM-CSF impregnated gauze can be effective in preventing and healing radiation-induced dermatitis and in reducing the interruption intervals of radiotherapy for vulvar carcinomas. PMID:11472614

  11. The combined effect of erythropoietin and granulocyte macrophage colony stimulating factor on liver regeneration after major hepatectomy in rats

    PubMed Central

    2010-01-01

    Background The liver presents a remarkable capacity for regeneration after hepatectomy but the exact mechanisms and mediators involved are not yet fully clarified. Erythropoietin (EPO) and Granulocyte-Macrophage Colony Stimulating Factor (GM-CSF) have been shown to promote liver regeneration after major hepatectomy. Aim of this experimental study is to compare the impact of exogenous administration of EPO, GM-CSF, as well as their combination on the promotion of liver regeneration after major hepatectomy. Methods Wistar rats were submitted to 70% major hepatectomy. The animals were assigned to 4 experimental groups: a control group (n = 21) that received normal saline, an EPO group (n = 21), that received EPO 500 IU/kg, a GM-CSF group (n = 21) that received 20 mcg/kg of GM-CSF and a EPO+GMCSF group (n = 21) which received a combination of the above. Seven animals of each group were killed on the 1st, 3rd and 7th postoperative day and their remnant liver was removed to evaluate liver regeneration by immunochemistry for PCNA and Ki 67. Results Our data suggest that EPO and GM-CSF increases liver regeneration following major hepatectomy when administered perioperatively. EPO has a more significant effect than GM-CSF (p < 0.01). When administering both, the effect of EPO seems to fade as EPO and GM-CSF treated rats have decreased regeneration compared to EPO administration alone (p < 0.01). Conclusion EPO, GM-CSF and their combination enhance liver regeneration after hepatectomy in rats when administered perioperatively. However their combination has a weaker effect on liver regeneration compared to EPO alone. Further investigation is needed to assess the exact mechanisms that mediate this finding. PMID:20604971

  12. Protein Tyrosine Phosphatase φ Regulates Paxillin Tyrosine Phosphorylation and Mediates Colony-Stimulating Factor 1-Induced Morphological Changes in Macrophages

    PubMed Central

    Pixley, Fiona J.; Lee, Pierre S. W.; Condeelis, John S.; Stanley, E. Richard

    2001-01-01

    Removal of colony-stimulating factor 1 (CSF-1) causes macrophages to round up and to increase their expression of protein tyrosine phosphatase φ (PTPφ). This is accompanied by the disruption of focal complexes and the formation of ruffles. Here we have overexpressed wild-type (WT) PTPφ and a phosphatase-inactive (C325S) mutant in a macrophage cell line in the presence and absence of CSF-1. In the presence of CSF-1, WT PTPφ induces cell rounding and ruffle formation, while C325S PTPφ has no effect. In contrast, in CSF-1-starved cells, C325S PTPφ behaves in a dominant negative fashion, preventing rounding and ruffling. Furthermore, C325S PTPφ increases adhesion in cycling cells, while WT PTPφ enhances motility. In WT PTPφ-overexpressing cells, the focal contact protein paxillin is selectively depleted from focal complexes and specifically dephosphorylated on tyrosine. In contrast, paxillin is hyperphosphorylated in C325S PTPφ-expressing cells. Moreover, a complex containing PTPφ, paxillin, and a paxillin-associated tyrosine kinase, Pyk2, can be immunoprecipitated from macrophage lysates, and the catalytic domain of PTPφ selectively binds paxillin and Pyk2 in vitro. Although PTPφ and Pyk2 do not colocalize with paxillin in focal complexes, all three proteins are colocalized in dorsal ruffles. The results suggest that paxillin is dephosphorylated by PTPφ in dorsal ruffles, using Pyk2 as a bridging molecule, resulting in a reduced pool of tyrosine-phosphorylated paxillin available for incorporation into focal complexes, thereby mediating CSF-1 regulation of macrophage morphology, adhesion, and motility. PMID:11238916

  13. A randomised trial of granulocyte-macrophage colony-stimulating factor for neonatal sepsis: outcomes at 2 years

    PubMed Central

    Marlow, Neil; Morris, Timothy; Brocklehurst, Peter; Carr, Robert; Cowan, Frances M; Patel, Nishma; Petrou, Stavros; Redshaw, Maggie E; Modi, Neena; Dore, Caroline

    2013-01-01

    Objective The authors performed a randomised trial in very preterm small-for-gestational age (SGA) babies to determine if prophylaxis with granulocyte-macrophage colony-stimulating factor (GM-CSF) improves outcomes (the PROGRAMS trial). Despite increased neutrophil counts following GM-CSF, the authors reported no significant difference in neonatal sepsis-free survival. Patients and methods 280 babies born <31 weeks of gestation and SGA were entered into the trial. Outcome was determined at 2 years to determine neurodevelopmental and general health outcomes, including economic costs. Results The authors found no significant differences in health outcomes or health and social care costs between the trial groups. In the GM-CSF arm, 87 of 134 (65%) babies survived to 2 years without severe disability compared with 87 of 131 (66%) controls (RR: 1·0, 95% CI 0·8 to 1·2). Marginally, more children receiving GM-CSF were reported to have cough (RR 1·7, 95% CI 1·1 to 2·6) and had signs of chronic respiratory disease (Harrison's sulcus; RR 2·0, 95% CI 1·0 to 3·9) though this was not reflected in bronchodilator use or need for hospitalisation for respiratory disease. Overall, the rate of neurologic abnormality (7%–9%) was similar but mean overall developmental scores were lower than expected for gestational age. Conclusions The administration of GM-CSF to very preterm SGA babies is not associated with improved or more adverse outcomes at 2 years of age. The apparent excess of developmental impairment in the entire PROGRAMS cohort, without corresponding increase in neurological abnormality, may represent diffuse brain injury attributable to intrauterine growth restriction. PMID:22542709

  14. Systematic review of the use of granulocyte-macrophage colony-stimulating factor in patients with advanced melanoma.

    PubMed

    Hoeller, Christoph; Michielin, Olivier; Ascierto, Paolo A; Szabo, Zsolt; Blank, Christian U

    2016-09-01

    Several immunomodulatory checkpoint inhibitors have been approved for the treatment of patients with advanced melanoma, including ipilimumab, nivolumab and pembrolizumab. Talimogene laherparepvec is the first oncolytic virus to gain regulatory approval in the USA; it is also approved in Europe. Talimogene laherparepvec expresses granulocyte-macrophage colony-stimulating factor (GM-CSF), and with other GM-CSF-expressing oncolytic viruses in development, understanding the clinical relevance of this cytokine in treating advanced melanoma is important. Results of trials of GM-CSF in melanoma have been mixed, and while GM-CSF has the potential to promote anti-tumor responses, some preclinical data suggest that GM-CSF may sometimes promote tumor growth. GM-CSF has not been approved as a melanoma treatment. We undertook a systematic literature review of studies of GM-CSF in patients with advanced melanoma (stage IIIB-IV). Of the 503 articles identified, 26 studies met the eligibility criteria. Most studies investigated the use of GM-CSF in combination with another treatment, such as peptide vaccines or chemotherapy, or as an adjuvant to surgery. Some clinical benefit was reported in patients who received GM-CSF as an adjuvant to surgery, or in combination with other treatments. In general, outcomes for patients receiving peptide vaccines were not improved with the addition of GM-CSF. GM-CSF may be a valuable therapeutic adjuvant; however, further studies are needed, particularly head-to-head comparisons, to confirm the optimal dosing regimen and clinical effectiveness in patients with advanced melanoma. PMID:27372293

  15. 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

  16. Granulocyte-colony stimulating factor-producing gallbladder carcinoma-include analysis all case reports: A case report

    PubMed Central

    Izumo, Wataru; Furukawa, Kenji; Katsuragawa, Hideo; Tezuka, Toru; Furukawa, Tatsuya; Hataji, Kenichirou; Komatsu, Akio; Shigematsu, Kyousuke; Yamamoto, Masakazu

    2016-01-01

    Introduction It is extremely rare for gallbladder carcinoma to produce granulocyte-colony stimulating factor (G-CSF) and such tumors have a poor prognosis. Presentation of case A 67-year-old man was admitted with continuous fever. Laboratory tests showed a leukocyte count of 27,980/μL, serum C-reactive protein (CRP) of 9.2 mg/dL and serum G-CSF of 225 pg/mL. Imaging revealed an irregular gallbladder mass about 90 mm in diameter with peripheral enhancement that also involved the liver and transverse colon. G-CSF producing gallbladder carcinoma was diagnosed. We performed cholecystectomy, partial resection of segments 4 and 5 of the liver, partial resection of the transverse colon, and gastrostomy. Histopathological examination showed gallbladder carcinoma (pT3, pN0, M0, G2, and pStage IIIA by the UICC classification, version 7). On immunohistochemical staining, tumor cells were positive for G-CSF. The leukocyte count was normalized postoperatively and fever subsided immediately after surgery. Two months later, the leukocyte count rose to 56,820/μL and metastases to the liver and lymph nodes were detected by CT. Chemotherapy (gemcitabine plus cisplatin) was started and the leukocyte count was normalized after the first course. The patient has continued chemotherapy and has survived for 16 months postoperatively. Discussion G-CSF producing gallbladder carcinoma has a poor prognosis and most patients die within 12 months of starting therapy. It is rare for patients with recurrence to survive for 16 months after surgery, as in the present case. Conclusion Multidisciplinary therapy (surgery and chemotherapy) may prolong the survival of patients with G-CSF producing gallbladder carcinoma, especially those with recurrence. PMID:26945490

  17. 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. PMID:25659501

  18. Granulocyte–Colony Stimulating Factor Promotes Liver Repair and Induces Oval Cell Migration and Proliferation in Rats

    PubMed Central

    PISCAGLIA, ANNA C.; SHUPE, THOMAS D.; OH, SEH–HOON; GASBARRINI, ANTONIO; PETERSEN, BRYON E.

    2011-01-01

    Background & Aims Hepatic regeneration is a heterogeneous phenomenon involving several cell populations. Oval cells are considered liver stem cells, a portion of which derive from bone marrow (BM). Recent studies have shown that granulocyte–colony stimulating factor (G-CSF) may be effective in facilitating liver repair. However, it remains unclear if G-CSF acts by mobilizing BM cells, or if it acts locally within the liver microenvironment to facilitate the endogenous restoration program. In the present study, we assessed the involvement of G-CSF during oval cell activation. Methods Dipeptidyl-peptidase-IV–deficient female rats received BM transplants from wild-type male donors. Four weeks later, rats were subjected to the 2-acetylaminofluorene/partial hepatectomy model of oval cell–mediated liver regeneration, followed by administration of either nonpegylated G-CSF or pegylated G-CSF. Control animals did not receive further treatments after surgery. The magnitude of oval cell reaction, the entity of BM contribution to liver repopulation, as well as the G-CSF/G-CSF–receptor expression levels were evaluated. In addition, in vitro proliferation and migration assays were performed on freshly isolated oval cells. Results Oval cells were found to express G-CSF receptor and G-CSF was produced within the regenerating liver. G-CSF administration significantly increased both the magnitude of the oval cell reaction, and the contribution of BM to liver repair. Finally, G-CSF acted as a chemoattractant and a mitogen for oval cells in vitro. Conclusions We have shown that G-CSF facilitates hepatic regeneration by increasing the migration of BM-derived progenitors to the liver, as well as enhancing the endogenous oval cell reaction. PMID:17681181

  19. Two protocols to treat thin endometrium with granulocyte colony-stimulating factor during frozen embryo transfer cycles.

    PubMed

    Xu, Bin; Zhang, Qiong; Hao, Jie; Xu, Dabao; Li, Yanping

    2015-04-01

    The efficacy of two granulocyte colony-stimulating factor (G-CSF) protocols for thin endometrium were investigated. Eighty-two patients were diagnosed with thin endometrium (<7 mm). Thirty patients with previously cancelled embryo transfers received intrauterine G-CSF in subsequent frozen embryo transfer (FET) cycles. Patients were divided into the G-CSF only and G-CSF with endometrial scratch subgroups. Compared with previous cycles, endometrial thickness increased from 5.7 ± 0.7 mm to 8.1 ± 2.1 mm after G-CSF treatment (P < 0.001). Endometrial thickness increases were not significantly different between the two subgroups. The G-CSF with endometrial scratch subgroup established nominally higher though non-significant clinical pregnancy and live birth rates than the G-CSF only subgroup (53.8 % versus 42.9% and 38.5% versus 28.6%, respectively). Fifty-two patients underwent FET despite edometrial thickness less than 7 mm, and were included as controls. Significantly higher embryo implantation and clinical pregnancy rates were observed in the G-CSF group compared with the control group (31.5% versus 13.9%; P < 0.01; 48.1% versus 25.0%; P = 0.038, respectively). Endometrial scracth did not impair G-CSF treatment for thin endometrium and favoured pregnancy and live birth rates. For patients with thin endometrium, embryo transfer cancellation and G-CSF treatment in subsequent FET cycles is beneficial. PMID:25682303

  20. 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.

  1. 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. PMID:26437001

  2. Plerixafor and abbreviated-course granulocyte colony-stimulating factor for mobilizing hematopoietic progenitor cells in light chain amyloidosis.

    PubMed

    Dhakal, Binod; Strouse, Christopher; D'Souza, Anita; Arce-Lara, Carlos; Esselman, Jeanie; Eastwood, Daniel; Pasquini, Marcelo; Saber, Wael; Drobyski, William; Rizzo, J Douglas; Hari, Parameswaran N; Hamadani, Mehdi

    2014-12-01

    Cytokine-based mobilization in light chain (AL) amyloidosis is frequently complicated by fluid overload, weight gain, cardiac arrhythmias, and peri-mobilization mortality. We analyzed hematopoietic progenitor cells (HPC) mobilization outcomes in 49 consecutive AL amyloidosis patients at our institution between 2004 and 2013 with granulocyte colony-stimulating factor (G) (10 μg/kg/day) (n = 25) versus an institutional protocol to limit G exposure using plerixafor (P) (.24 mg/kg s.c. starting day 3 of G 10 μg/kg) (n = 24). G+P strategy yielded higher total CD34(+) cells/kg (12.8 × 10(6) versus 6.3 × 10(6); P < .001) and CD34(+) cells/kg collected on day 1 (10.8 × 10(6) versus 4.9 × 10(6), P = .004) compared with the G cohort. More G+P patients collected ≥5 × 10(6) CD34(+) HPCs/kg (22 versus 16, P = .02) and ≥ 10 × 10(6) CD34(+) HPCs/kg (13 versus 5, P = .01). Four patients (16%) had mobilization failure with G; none with G+P. Peri-mobilization weight gain was lower with G+P strategy (median weight gain 1 versus 7 pounds, P = .009). Numbers of apheresis sessions (median, 1 versus 1, P = .52), number of hospitalization days (median, 1.1 versus 1.6, P = .52), transfusions, use of intravenous antibiotics, and cardiac arrhythmias were similar. In conclusion, our study demonstrates that upfront use of G+P as a mobilization strategy results in superior HPC collection, no mobilization failures, and less weight gain than G alone. PMID:25111581

  3. 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

  4. Timing of recombinant human granulocyte colony-stimulating factor administration on neutropenia induced by cyclophosphamide in normal mice.

    PubMed Central

    Misaki, M.; Ueyama, Y.; Tsukamoto, G.; Matsumura, T.

    1998-01-01

    The effects of altering the timing of recombinant human granulocyte colony-stimulating factor (rhG-CSF) administration on neutropenia induced by cyclophosphamide (CPA) were studied experimentally in a mouse model. Experimental mice were divided into three groups: (a) treatment with rhG-CSF after CPA administration (post-treatment group); (b) treatment with rhG-CSF both before and after CPA administration (pre- and post-treatment group); and (c) treatment with saline after CPA administration (control group). The results were as follows. Mice receiving rhG-CSF on the 2 days preceding CPA treatment, in which progenitor cell counts outside the S-phase when CPA was administered were the lowest of all the groups, showed accelerated neutrophil recovery but decreased neutrophil nadirs compared with the control group despite rhG-CSF treatment. The pre- and post-treatment group, consisting of mice who received rhG-CSF treatment on days -4 and -3 before CPA treatment, and in which progenitor cell counts when CPA was administered were increased to greater levels than in the other groups, showed remarkably accelerated neutrophil recovery and the greatest increase in the neutrophil nadirs of all the groups. These results suggested that the kinetics of progenitor cell populations when chemotherapeutic agents were administered seemed to play an important role in neutropenia after chemotherapy, and that not only peripheral neutrophil cell and total progenitor cell counts but also progenitor cell kinetics should be taken into consideration when administering rhG-CSF treatment against the effects of chemotherapy. PMID:9528829

  5. Granulocyte-Macrophage Colony Stimulating Factor Blockade Promotes CCR9+ Lymphocyte Expansion in Nod2 Deficient Mice

    PubMed Central

    Samson, Charles M.; Jurickova, Ingrid; Molden, Erin; Schreiner, William; Colliver, Joshua; Bonkowski, Erin; Han, Xiaonan; Trapnell, Bruce C.; Denson, Lee A.

    2011-01-01

    Background Ileal involvement in Crohn’s disease (CD) is associated with NOD2 mutations and Granulocyte-Macrophage Colony Stimulating Factor auto-antibodies (GM-CSF Ab), and GM-CSF blockade promotes ileitis in Nod2/Card15 deficient (C15KO) mice. RALDH2 expressing dendritic cells (DC) and IL-4 promote CCR9 imprinting and small bowel homing of T lymphocytes, in conjunction with CCL25 expression by ileal epithelial cells (IEC). We hypothesized that GM-CSF neutralization promotes ileal disease by modulating expression of CCL25 by IEC and CCR9 by T lymphocytes via Nod2 dependent and independent pathways. Methods CCL25 and CCR9 expression were determined in pediatric CD patients stratified by GM-CSF Ab. Ileitis was induced in C15KO mice via GM-CSF Ab administration followed by NSAID exposure, and expression of CCL25, CCR9, FOXP3, intracellular cytokines, and RALDH2 was determined in IEC and immune cell populations. Results The frequency of CCL25+ IEC and CCR9+ T lymphocytes was increased in CD patients with elevated GM-CSF Ab. In the murine model, GM-CSF blockade alone induced IEC CCL25 expression, and reduced the frequency of mesenteric lymph node (MLN) CD4+FOXP3+ cells, while Card15 deficiency alone enhanced MLN DC RALDH2 expression. Both GM-CSF neutralization and Card15 deficiency were required for down-regulation of MLN DC IL-10 expression; under these conditions NSAID exposure led to an expansion of IL-4+ and IL-17+ CCR9+ lymphocytes in the ileum. Conclusions GM-CSF prevents ileal expansion of CCR9+ lymphocytes via Nod2 dependent and independent pathways. CCR9 blockade may be beneficial in CD patients with elevated GM-CSF Ab. PMID:21381154

  6. Biotinylated granulocyte/macrophage colony-stimulating factor analogues: effect of linkage chemistry on activity and binding.

    PubMed

    Angelotti, T P; Clarke, M F; Longino, M A; Emerson, S G

    1991-01-01

    Biotinylated granulocyte/macrophage colony-stimulating factor (GM-CSF) analogues with different linkage chemistries and levels of conjugated biotin were synthesized by reacting recombinant human GM-CSF with sulfosuccinimidyl 6-biotinamidohexanoate or biotin hydrazide/1-[3-(dimethylamino)-propyl]-3-ethylcarbodiimide. These chemically reactive forms of biotin produced derivatives biotinylated at amine or carboxyl groups, respectively. Amine-derivatized analogues of 1.2 and 3.8 mol of biotin/mol of protein (N1-bGM-CSF and N4-bGM-CSF) and a carboxyl-modified analogue of 4.6 mol of biotin/mol of protein (C5-bGM-CSF) were synthesized. These analogues were compared to determine the effect of biotinylation on biological activity and GM-CSF receptor binding characteristics. The biotinylated proteins migrated with the same molecular weight as the native, unmodified protein as determined by SDS-PAGE and could be detected by Western blotting with alkaline phosphatase conjugated streptavidin, thus demonstrating the biotin linkage. All three analogues retained full agonist activity relative to the native protein (EC50 = 10-15 pM) when assayed for the stimulation of human bone marrow progenitor cell growth. Cell surface GM-CSF receptor binding was characterized by the binding of the analogues to human neutrophils, with detection by fluorescein-conjugated avidin and fluorescence-activated cell sorting. The N-bGM-CSFs demonstrated GM-CSF receptor specific binding that was displaceable by excess underivatized protein, with the detected fluorescence signal decreasing with increasing biotin to protein molar ratio. In contrast, C5-bGM-CSF binding above background fluorescence could not be detected using this system, suggesting that this derivative could bind to and activate the receptor, but not simultaneously bind fluorescein-conjugated avidin. The amine-derivatized biotinylated GM-CSF analogues retained biological activity, could specifically label cell surface receptors, and may be

  7. A sandwich enzyme-linked immunoabsorbent assay for measurement of picogram quantities of murine granulocyte colony-stimulating factor.

    PubMed

    Granger, J; Remick, D; Call, D; Ebong, S; Taur, A; Williams, B; Nauss, M; Millican, J; O'Reilly, M

    1999-05-27

    Granulocyte colony-stimulating factor (G-CSF) stimulates the proliferation and differentiation of hematopoietic progenitor cells of the neutrophil lineage. Measurement of murine G-CSF levels will allow examination of its role in host defense using murine models. Therefore, we developed a sensitive sandwich enzyme-linked immunoabsorbent assay (ELISA) for murine G-CSF. A polyclonal antibody to recombinant murine G-CSF was produced in rabbits and isolated using a protein A column. This purified native IgG served as the capture antibody and a portion of the IgG was biotinylated to serve as the developing antibody. Specificity was verified by lack of reactivity to GM-CSF, IL-6, IL-3, prolactin, and growth hormone. The lower limit of sensitivity routinely extended to 16 pg/ml in multiple ELISAs. Intra-assay coefficient of variation (CV) ranged from 3.4 to 21.5% across the detection limits of the assay, with the greatest variance occurring near the standard curve maximum. Interassay CV ranged from 11.5 to 23.3%. The ability of the ELISA to detect G-CSF in different sample preparations was examined in RPMI 1640 with 10% FCS, Hanks balanced salt solution, PBS/Tween-20/2% FCS, and the dilution media for ELISA (10% BLOTTO/PBS/0.05% Tween-20). Average recovery in these media ranged from 98 to 107%. Heparin anti-coagulated normal mouse plasma had a suppressive effect on the ELISA that varied between individual mice. Recovery was also determined from liver, spleen, and lung homogenate suspensions at dilutions of 1:5, 1:10, and 1:20 in dilution buffer. Recovery from liver was optimal at the 1:10 and 1:20 dilutions at 105%, with that of the 1:5 dilution at 135%. Recovery from spleen ranged from 94 to 96%. Lung homogenate displayed enhanced recovery (139% or greater) across all dilutions. The ability of the assay to detect G-CSF was explored by measurement of G-CSF levels in peritoneal lavage following polymicrobial intra-abdominal infection. Peak levels of G-CSF production

  8. 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

  9. Targeted overexpression of the two colony-stimulating factor-1 isoforms in osteoblasts differentially affects bone loss in ovariectomized mice

    PubMed Central

    Yao, Gang-Qing; Wu, Jian-Jun; Ovadia, Shira; Troiano, Nancy; Sun, Ben Hua; Insogna, Karl

    2009-01-01

    Colony-stimulating factor-1 (CSF1) is one of two cytokines required for normal osteoclastogenesis. There are two major isoforms of CSF1, the cell-surface or membrane-bound isoform (mCSF1) and soluble CSF1 (sCSF1). Whether these isoforms serve nonredundant functions in bone is unclear. To explore this question, we generated transgenic mice expressing human sCSF1, human mCSF1, or both (s/mCSF1) in osteoblasts using the 2.3-kb rat αI-collagen promoter. Bone density determined by peripheral quantitative computed tomography was significantly reduced in mCSF1, sCSF1, and s/mCSF1 transgenic mice compared with wild-type animals. When analyzed by sex, sCSF1, and s/mCSF1, female animals but not mCSF1 female mice were found to have greater bone loss than their male littermates (−20 vs. −9.2%; P < 0.05 for sCSF1 and −21.6 vs. −11.2% for s/mCSF1; P < 0.01). By breeding CSF1 isoform-selective transgenic mice to an op/op background, mice were generated in which a single CSF1 isoform was the only source of the cytokine (sCSF1op/op and mCSF1op/op). Unlike osteoblast-targeted overexpression of mCSF1, selective transgenic expression of sCSF1 did not completely correct the op/op phenotype in 5-mo-old animals. Interestingly, compared with sham-ovariectomized mice of the same genotype, ovariectomy in sCSF1op/op mice led to a greater loss of spinal bone mineral density (22.1%) than was seen in either mCSF1op/op mice (12.9%) or in wild-type animals (10.9%). Our findings support the conclusion that sCSF1 and mCSF1 serve nonredundant functions in bone and that sCSF1 may play a role in mediating estrogen-deficiency bone loss. PMID:19141689

  10. 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

  11. The Toll-like receptor 4-activated neuroprotective microglia subpopulation survives via granulocyte macrophage colony-stimulating factor and JAK2/STAT5 signaling.

    PubMed

    Kamigaki, Mayumi; Hide, Izumi; Yanase, Yuhki; Shiraki, Hiroko; Harada, Kana; Tanaka, Yoshiki; Seki, Takahiro; Shirafuji, Toshihiko; Tanaka, Shigeru; Hide, Michihiro; Sakai, Norio

    2016-02-01

    Toll-like receptor (TLR) 4 mediates inflammation and is also known to trigger apoptosis in microglia. Our time-lapse observations showed that lipopolysaccharide (LPS) stimulation induced rapid death in primary cultures of rat microglia, while a portion of the microglia escaped from death and survived for much longer than 2 days, in which time, all of the control cells had died. However, it remains unclear how the LPS-stimulated microglia subpopulation could continue to survive in the absence of any supplied growth factors. In the present study, to clarify the mechanism underlying the LPS-stimulated survival, we investigated whether microglia could produce their own survival factors in response to LPS, focusing on macrophage colony-stimulating factor (M-CSF), granulocyte macrophage colony-stimulating factor (GM-CSF) and interleukin (IL)-34, which are mainly supplied by astrocytes or neurons. The LPS-stimulated microglia drastically induced the expression of the GM-CSF mRNA and protein, while M-CSF and IL-34 levels were unchanged. The surviving microglia also significantly upregulated the expression of GM-CSF receptor (GM-CSFR) mRNA without affecting M-CSFR. As for the GM-CSFR downstream signal, LPS resulted in the phosphorylation of STAT5 and its translocation to the nucleus in the surviving microglia. Moreover, a specific JAK2 inhibitor, NVP-BSK805, suppressed STAT5 phosphorylation and microglia survival in response to LPS, indicating a critical role of the JAK2/STAT5 pathway in this survival mechanism. Together, these results suggest that a subpopulation of TLR4-activated microglia may survive by producing GM-CSF and up-regulating GM-CSFR. This autocrine GM-CSF pathway may activate the JAK2/STAT5 signaling pathway, which controls the transcription of survival-related genes. Finally, these surviving microglia may have neuroprotective functions because the neurons remained viable in co-cultures with these microglia. PMID:26802935

  12. Pulsed ultrasound promotes melanoblast migration through upregulation of macrophage colony-stimulating factor/focal adhesion kinase autocrine signaling and paracrine mechanisms.

    PubMed

    Liao, Yi-Hua; Huang, Yu-Ting; Deng, Jhu-Yun; Chen, Wen-Shiang; Jee, Shiou-Hwa

    2013-09-01

    Repigmentation of vitiliginous lesions relies on the proliferation and migration of melanoblasts from hair follicles to the epidermis. Pulsed ultrasound has been demonstrated to have stimulatory effects on cell proliferation and migration and has been applied clinically to enhance tissue repair. To clarify the biologic effects and signaling mechanisms of pulsed ultrasound on melanoblast proliferation and migration, two melanoblast cell lines, the undifferentiated NCCmelb4 cells and the differentiated NCCmelan5 cells, were examined. We demonstrated that pulsed ultrasound increased cell migration in a dose-dependent manner without altering cell proliferation. Pulsed ultrasound enhanced autocrine secretion of macrophage colony-stimulating factor (M-CSF), which subsequently activated the focal adhesion kinase (FAK) pathway to promote melanoblast migration. Furthermore, conditioned medium from mouse embryonic fibroblasts NIH 3T3 and primary human keratinocytes treated with pulsed ultrasound could stimulate melanoblast migration through a paracrine effect. Our results provide a novel mechanism to promote migration of melanoblasts by pulsed ultrasound stimulation. PMID:23725022

  13. 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

  14. Association of the T allele of an intronic single nucleotide polymorphism in the colony stimulating factor 1 receptor with Crohn's disease: a case-control study.

    PubMed

    Zapata-Velandia, Adriana; Ng, San-San; Brennan, Rebecca F; Simonsen, Neal R; Gastanaduy, Mariella; Zabaleta, Jovanny; Lentz, Jennifer J; Craver, Randall D; Correa, Hernan; Delgado, Alberto; Pitts, Angela L; Himel, Jane R; Udall, John N; Schmidt-Sommerfeld, Eberhard; Brown, Raynorda F; Athas, Grace B; Keats, Bronya B; Mannick, Elizabeth E

    2004-05-14

    BACKGROUND: Polymorphisms in several genes (NOD2, MDR1, SLC22A4) have been associated with susceptibility to Crohn's disease. Identification of the remaining Crohn's susceptibility genes is essential for the development of disease-specific targets for immunotherapy. Using gene expression analysis, we identified a differentially expressed gene on 5q33, the colony stimulating factor 1 receptor (CSF1R) gene, and hypothesized that it is a Crohn's susceptibility gene. The CSF1R gene is involved in monocyte to macrophage differentiation and in innate immunity. METHODS: Patients provided informed consent prior to entry into the study as approved by the Institutional Review Board at LSU Health Sciences Center. We performed forward and reverse sequencing of genomic DNA from 111 unrelated patients with Crohn's disease and 108 controls. We also stained paraffin-embedded, ileal and colonic tissue sections from patients with Crohn's disease and controls with a polyclonal antibody raised against the human CSF1R protein. RESULTS: A single nucleotide polymorphism (A2033T) near a Runx1 binding site in the eleventh intron of the colony stimulating factor 1 receptor was identified. The T allele of this single nucleotide polymorphism occurred in 27% of patients with Crohn's disease but in only 13% of controls (X2 = 6.74, p < 0.01, odds ratio (O.R.) = 2.49, 1.23 < O.R. < 5.01). Using immunohistochemistry, positive staining with a polyclonal antibody to CSF1R was observed in the superficial epithelium of ileal and colonic tissue sections. CONCLUSIONS: We conclude that the colony stimulating factor receptor 1 gene may be a susceptibility gene for Crohn's disease. PMID:15144560

  15. Integration of the BALB/c ecotropic provirus into the colony-stimulating factor-1 growth factor locus in a myc retrovirus-induced murine monocyte tumor.

    PubMed

    Baumbach, W R; Colston, E M; Cole, M D

    1988-09-01

    The development of tumors is thought to be a multistage process that requires an unknown number of genetic or epigenetic changes in a single cell. We previously described a murine monocyte tumor which was induced by a helper-free c-myc retrovirus and which also contained a DNA rearrangement at the colony-stimulating factor-1 (CSF-1) locus. The CSF-1 gene rearrangement gave rise to high levels of growth factor production and autocrine growth, implicating this secondary event in tumorigenesis. This CSF-1 gene rearrangement was found to be the result of integration of the BALB/c ecotropic retrovirus. Restriction enzyme mapping and DNA sequence analysis demonstrated that the novel provirus is identical to the BALB/c endogenous ecotropic provirus, indicating that infection was probably not due to the creation of a recombinant virus in vivo. The proviral integration site was mapped 3 kilobases 5' of the CSF-1 promoter and in an opposite transcriptional orientation, indicating that activation of CSF-1 expression was the result of the presence of the retroviral enhancer element. PMID:3261346

  16. 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

  17. Favorable outcome of granulocyte colony-stimulating factor use in neuromyelitis optica patients presenting with agranulocytosis in the setting of rituximab.

    PubMed

    Mealy, Maureen A; Levy, Michael

    2015-10-15

    Neuromyelitis optica is a severe autoimmune condition affecting the central nervous system characterized by a relapsing disease course. Rituximab is a chimeric monoclonal antibody against the protein CD20, and is one of the most utilized medications for management of this disease. A known complication of rituximab use is neutropenia. We report on two patients who developed symptomatic early-onset rituximab-induced agranulocytosis who safely received granulocyte colony-stimulating factor. Neutrophil counts recovered quickly and both patients continue to receive rituximab without further incident. PMID:26439958

  18. 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. PMID:26970370

  19. Macrophage production during murine listeriosis: colony-stimulating factor 1 (CSF-1) and CSF-1-binding cells in genetically resistant and susceptible mice.

    PubMed Central

    Cheers, C; Stanley, E R

    1988-01-01

    The concentration of the macrophage-specific colony-stimulating factor (CSF-1) and the numbers of bone marrow and spleen cells with specific receptors for that factor have been investigated in a number of mouse strains under normal conditions and after infection with the facultative intracellular bacterium Listeria monocytogenes. The CSF-1 concentration in serum and tissue was markedly elevated in infected mice, the degree of stimulation reflecting the dose of L. monocytogenes. The CSF-1 titer did not correlate with genetic resistance or susceptibility of the mice to L. monocytogenes. In contrast to the effect of lipopolysaccharide, Listeria infection was able to increase the level of CSF-1 in the lipopolysaccharide nonresponder strain C3H/HeJ. In line with earlier findings on colony-forming cells, cells bearing receptors for CSF-1 in uninfected susceptible BALB/cJ mice were only half those in resistant C57BL/6J mice. After infection the majority of these cells disappeared from the bone marrow and spleen cells of both resistant and susceptible mice. The number of CSF-1 receptor-bearing cells in the normal bone marrow may determine the degree of resistance to L. monocytogenes. PMID:3262588

  20. 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. PMID:26245897

  1. Differential utilization of Ras signaling pathways by macrophage colony-stimulating factor (CSF) and granulocyte-macrophage CSF receptors during macrophage differentiation.

    PubMed

    Guidez, F; Li, A C; Horvai, A; Welch, J S; Glass, C K

    1998-07-01

    Granulocyte-macrophage colony-stimulating factor (GM-CSF) and macrophage colony-stimulating factor (M-CSF) independently stimulate the proliferation and differentiation of macrophages from bone marrow progenitor cells. Although the GM-CSF and M-CSF receptors are unrelated, both couple to Ras-dependent signal transduction pathways, suggesting that these pathways might account for common actions of GM-CSF and M-CSF on the expression of macrophage-specific genes. To test this hypothesis, we have investigated the mechanisms by which GM-CSF and M-CSF regulate the expression of the macrophage scavenger receptor A (SR-A) gene. We demonstrate that induction of the SR-A gene by M-CSF is dependent on AP-1 and cooperating Ets domain transcription factors that bind to sites in an M-CSF-dependent enhancer located 4.1 to 4.5 kb upstream of the transcriptional start site. In contrast, regulation by GM-CSF requires a separate enhancer located 4.5 to 4.8 kb upstream of the transcriptional start site that confers both immediate-early and sustained transcriptional responses. Results of a combination of DNA binding experiments and functional assays suggest that immediate transcriptional responses are mediated by DNA binding proteins that are constitutively bound to the GM-CSF enhancer and are activated by Ras. At 12 to 24 h after GM-CSF treatment, the GM-CSF enhancer becomes further occupied by additional DNA binding proteins that may contribute to sustained transcriptional responses. In concert, these studies indicate that GM-CSF and M-CSF differentially utilize Ras-dependent signal transduction pathways to regulate scavenger receptor gene expression, consistent with the distinct functional properties of M-CSF- and GM-CSF-derived macrophages. PMID:9632769

  2. Antibodies binding granulocyte-macrophage colony stimulating factor produced by cord blood-derived B cell lines immortalized by Epstein-Barr virus in vitro.

    PubMed

    Revoltella, R P; Laricchia Robbio, L; Liberati, A M; Reato, G; Foa, R; Funaro, A; Vinante, F; Pizzolo, G

    2000-09-15

    We detected natural antibodies (auto-Abs) binding human granulocyte-macrophage colony stimulating factor (GM-CSF) in umbilical cord blood (CB) (23 of 94 samples screened) and peripheral blood of women at the end of pregnancy (6 of 42 samples tested). To demonstrate that Abs detected in CB were produced by the fetus, CB mononuclear cells were infected with Epstein-Barr virus in vitro. Ten cell lines producing constitutively anti-recombinant human GM-CSF (rhGM-CSF) Abs were isolated and characterized. These cells displayed a male karyotype, an early activated B cell phenotype, coexpressed surface IgM and IgD, and secreted only IgM with prevailing lambda clonal restriction. Specific cell surface binding of biotinylated rhGM-CSF and high-level anti-rhGM-CSF IgM Ab production were typical features of early cell cultures. In late cell passages the frequency of more undifferentiated B cells increased. Serum Abs of either maternal or fetal origin or Abs produced in culture did not affect the granulocyte and macrophage colony stimulating activity of rhGM-CSF from bone marrow progenitors in soft agar, suggesting that the Abs produced were nonneutralizing. PMID:11069719

  3. Characterization of a cell-type-restricted negative regulatory activity of the human granulocyte-macrophage colony-stimulating factor gene.

    PubMed Central

    Fraser, J K; Guerra, J J; Nguyen, C Y; Indes, J E; Gasson, J C; Nimer, S D

    1994-01-01

    Human granulocyte-macrophage colony-stimulating factor (GM-CSF) stimulates the proliferation and maturation of normal myeloid progenitor cells and can also stimulate the growth of acute myelogenous leukemia (AML) blasts. GM-CSF is not normally produced by resting cells but is expressed by a variety of activated cells including T lymphocytes, macrophages, and certain cytokine-stimulated fibroblasts and endothelial cells. Production of GM-CSF by cultured AML cells has been demonstrated, and GM-CSF expression by normal myeloid progenitors has been postulated to play a role in myelopoiesis. We have investigated the regulation of expression of GM-CSF in AML cell lines, and our results demonstrate the presence of a strong constitutive promoter element contained within 53 bp upstream of the cap site. We have also identified a negative regulatory element located immediately upstream of the positive regulatory element (within 69 bp of the cap site) that is active in AML cell lines but not T cells or K562 CML cells. Competition transfection and mobility shift studies demonstrate that this activity correlates with binding of a 45-kDa protein. Images PMID:8114751

  4. 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

  5. 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

  6. Neutrophil apoptosis: impact of granulocyte macrophage colony stimulating factor on cell survival and viability in chronic kidney disease and hemodialysis patients

    PubMed Central

    Zahran, Nariman; Sayed, Azza; William, Iman; Sabry, Omar; Rafaat, Manar

    2013-01-01

    Introduction Altered neutrophil apoptosis might be responsible for recurrent bacterial infections encountered in hemodialysis (HD) and chronic kidney disease (CKD) patients. This work was designed to assess the neutrophil apoptotic activity and the impact of implementation of granulocyte macrophage colony stimulating factor (GM-CSF), as a survival factor, on neutrophil apoptosis among these patients. Material and methods Twenty-five patients on regular HD along with 34 CKD patients on conservative treatment, as well as 15 healthy controls, were investigated for apoptotic rate via assessment of neutrophil expression of Annexin-V by flow cytometry, before and after 20 h culture in absence and presence of GM-CSF. Neutrophil viability was determined using light microscopy. The preservation of neutrophil activation in these patients was analyzed by flow cytometric CD18 neutrophil expression. Chronic inflammatory state was evaluated by estimating C-reactive protein (CRP) and soluble intercellular adhesion molecule-1 (sICAM-1). Obtained data were statistically analyzed. Results Compared to controls, both HD and CKD groups had a significant increase of Annexin-V and CD18 expression and significant decrease in neutrophil viability. Culture of their neutrophils with GM-CSF showed significant decrease of apoptosis accompanied by improvement of neutrophil viability compared to their cultured cells without GM-CSF. These patients also showed significant elevation of CRP and sICAM-1. Conclusions Granulocyte macrophage colony stimulating factor demonstrated an evident impact on improving in vitro neutrophil survival and viability in HD and CKD patients. Therefore, this may represent promising preventive and/or therapeutic strategies against infection frequently observed in these patients and causing morbidity and mortality. PMID:24482640

  7. Cytokine regulation of granulocyte-macrophage colony-stimulating factor (GM-CSF) production by human retinal pigment epithelial cells

    PubMed Central

    Crane, I J; Kuppner, M C; Mckillop-Smith, S; Wallace, C A; Forrester, J V

    1999-01-01

    GM-CSF is an important regulator of macrophage, granulocyte and dendritic cell behaviour and function. These cell types have been implicated in the retinal damage characteristic of endogenous posterior uveitis. Dendritic cells in the choroid have access to retinal antigens processed by the retinal pigment epithelial (RPE) cells of the blood–retinal barrier and are thought to be candidates for the presentation of antigen in uveoretinitis. We therefore investigated the production of GM-CSF and its regulation in human RPE cells. IL-1β, tumour necrosis factor-alpha (TNF-α) and transforming growth factor-beta (TGF-β) all stimulated GM-CSF production by RPE cells and a combination of these cytokines increased GM-CSF production over five-fold compared with that with the individual cytokines alone. Interferon-gamma (IFN-γ) rapidly down-regulated these responses. IFN-γ did not appear to be acting directly on IL-1β or via the synthesis of another protein. GM-CSF mRNA expression showed the same pattern of response to these cytokines, indicating transcriptional or pre-transcriptional regulation, and there was no evidence that IFN-γ was acting by destabilizing GM-CSF mRNA. These results are generally important in understanding the ways in which cytokine regulation differs between different cell types and also more specifically for determining ways in which a cytokine with a significant role in the development of autoimmune uveoretinitis may be manipulated. PMID:9933455

  8. 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.

  9. The costs of peripheral blood progenitor cell reinfusion mobilised by granulocyte colony-stimulating factor following high dose melphalan as compared with conventional therapy in multiple myeloma.

    PubMed

    Uyl-de Groot, C A; Ossenkoppele, G J; van Riet, A A; Rutten, F F

    1994-01-01

    In a retrospective study, we calculated the treatment costs of 26 patients, who received either high dose melphalan combined with granulocyte colony-stimulating factor (G-CSF; filgrastim)(n = 7) or without G-CSF (n = 11) or alternatively, peripheral blood progenitor cell reinfusion (PBPC) mobilised by G-CSF following high dose melphalan. In comparison with the control group, a shortening of the pancytopenic period and platelet recovery was noticed in the PBPC group. This resulted in a reduction in hospital costs, diagnostics, laboratory services, total parenteral nutrition and transfusions. The average costs per treatment in the PBPC group amounted to about US$ 17,908 as compared to US$ 32,223 in the control group, implying a cost reduction of 44% when changing to PBPC reinfusion. PMID:7517149

  10. Augmentation of granulocyte/macrophage colony-stimulating factor expression by ultraviolet irradiation is mediated by interleukin 1 in Pam 212 keratinocytes

    SciTech Connect

    Nozaki, S.; Abrams, J.S.; Pearce, M.K.; Sauder, D.N. )

    1991-07-01

    Keratinocytes are a potent source of a variety of cytokines including granulocyte-macrophage colony-stimulating factor (GM-CSF). In this study, we have shown that ultraviolet B (UVB) irradiation augments GM-CSF mRNA expression by murine keratinocytes. This is reflected in the increased production of GM-CSF protein by these cells. In the same cell population, exposure to UVB irradiation increases interleukin 1 alpha (IL-1 alpha) mRNA and IL-1 protein as detected by bioactivity. This increase in IL-1 alpha precedes the increase of GM-CSF mRNA. Addition of recombinant IL-1 alpha to the medium increases GM-CSF mRNA expression. Anti-IL-1 alpha antibodies can completely inhibit UV-augmented GM-CSF mRNA expression. These results demonstrate that UVB irradiation-induced augmentation of GM-CSF is mediated by UV-induced IL-1 alpha.

  11. Chemotherapy and recombinant human granulocyte colony-stimulating factor primed donor leukocyte infusion for treatment of relapse after allogeneic bone marrow transplantation.

    PubMed

    Sica, S; Di Mario, A; Salutari, P; Rutella, S; Chiusolo, P; Rumi, C; Menichella, G; D'Onofrio, G; Leone, G

    1995-09-01

    Two patients affected by acute leukemia relapsed 10 and 12 months respectively after allogeneic bone marrow transplantation. They were treated with aggressive chemotherapy and then infused with HLA-identical donor leukocytes (DLI) collected after recombinant human granulocyte colony-stimulating factor (rhG-CSF) administration. A total of 5.6 and 6.3 x 10(6)/kg CD34+ cells, 2.7 and 3.0 x 10(4)/kg CFU-GM, 4.7 and 4.4 x 10(8)/kg MNC, 4.6 and 3.9 x 10(9)/kg PMN respectively were infused. Both patients achieved complete remission (CR) and complete chimerism was re-established. One patient developed grade IV acute graft-versus-host disease of the liver requiring immunosuppression and he died in CR from disseminated aspergillosis, 7 months after chemotherapy; one patient is alive in relapse 12 months after treatment. PMID:8535325

  12. 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

  13. Anti-Granulocyte-Macrophage Colony-Stimulating Factor Autoantibodies Are a Risk Factor for Central Nervous System Infection by Cryptococcus gattii in Otherwise Immunocompetent Patients

    PubMed Central

    Saijo, Tomomi; Chen, Jianghan; Chen, Sharon C.-A.; Rosen, Lindsey B.; Yi, Jin; Sorrell, Tania C.; Bennett, John E.; Holland, Steven M.; Browne, Sarah K.; Kwon-Chung, Kyung J.

    2014-01-01

    ABSTRACT Cryptococcosis is caused by either Cryptococcus neoformans or C. gattii. While cryptococcal meningoencephalitis is caused mostly by C. neoformans in immunocompromised patients, the risk factors remain unclear for patients with no known immune defect. Recently, anti-granulocyte-macrophage colony-stimulating factor (GM-CSF) autoantibodies were detected in the plasma of seven “immunocompetent” cryptococcosis patients, and the cryptococcal strains from these patients were reported as C. neoformans (three strains), C. gattii (one strain), and Cryptococcus (three strains not identified to the species level). We identified all three strains that had not been identified to the species level as C. gattii. Notably, the three strains that were reported as C. neoformans but were unavailable for species confirmation originated from Sothern California and Thailand where C. gattii is endemic. Most clinical laboratories designate C. neoformans without distinguishing between the two species; hence, these three strains could have been C. gattii. Since C. gattii infects more immunocompetent patients than C. neoformans, we pursued the possibility that this antibody may be more prevalent in patients infected with C. gattii than in those infected with C. neoformans. We screened the plasma of 20 healthy controls and 30 “immunocompetent” patients with cryptococcal meningoencephalitis from China and Australia (multiple ethnicities). Anti-GM-CSF autoantibodies were detected only in the plasma of seven patients infected by C. gattii and one healthy volunteer and in none infected by C. neoformans. While plasma from these C. gattii patients completely prevented GM-CSF-induced p-STAT5 in normal human peripheral blood mononuclear cells (PBMCs), plasma from one healthy volunteer positive for anti-GM-CSF autoantibodies caused only partial blockage. Our results suggest that anti-GM-CSF autoantibodies may predispose otherwise immunocompetent individuals to

  14. Enhancement of the grafting efficiency of transplanted marrow cells by preincubation with interleukin-3 and granulocyte-macrophage colony-stimulating factor

    SciTech Connect

    Tavassoli, M.; Konno, M.; Shiota, Y.; Omoto, E.; Minguell, J.J.; Zanjani, E.D.

    1991-04-01

    To improve the grafting efficiency of transplanted murine hematopoietic progenitors, we briefly preincubated mouse bone marrow cells with interleukin-3 (IL-3) or granulocyte-macrophage colony-stimulating factor (GM-CSF) ex vivo before their transplantation into irradiated recipients. This treatment was translated into an increase in the seeding efficiency of colony-forming unit-spleen (CFU-S) and CFU-GM after transplantation. Not only was the concentration of CFU-S in the tibia increased 2 and 24 hours after transplantation, but the total cell number and CFU-S and CFU-GM concentrations were persistently higher in IL-3- and GM-CSF-treated groups 1 to 3 weeks after transplantation. In addition, the survival of animals as a function of transplanted cell number was persistently higher in IL-3- and GM-CSF-treated groups compared with controls. The data indicate that the pretreatment of marrow cells with IL-3 and GM-CSF before transplantation increases the seeding efficiency of hematopoietic stem cells and probably other progenitor cells after transplantation. This increased efficiency may be mediated by upward modulation of homing receptors. Therefore, ex vivo preincubation of donor marrow cells with IL-3 and GM-CSF may be a useful tactic in bone marrow transplantation.

  15. Granulocyte-macrophage colony-stimulating factor and interleukin-3 signaling pathways converge on the CREB-binding site in the human egr-1 promoter.

    PubMed Central

    Sakamoto, K M; Fraser, J K; Lee, H J; Lehman, E; Gasson, J C

    1994-01-01

    Granulocyte-macrophage colony-stimulating factor (GM-CSF) stimulates myeloid progenitor cell proliferation and enhances the function of terminally differentiated effector cells. Interleukin-3 (IL-3) stimulation results in the proliferation and maturation of early bone marrow progenitor cells. These activities are mediated by non-tyrosine kinase-containing receptors which consist of ligand-specific alpha subunits that complex with a common beta subunit required for signal transduction. Both GM-CSF and IL-3 rapidly and transiently induce expression of early growth response gene 1 (egr-1) in the human factor-dependent cell line TF-1. To define the mechanism of early response gene induction by GM-CSF and IL-3, growth factor- and serum-starved TF-1 cells transfected with recombinant constructs containing sequences of the human egr-1 promoter were stimulated with GM-CSF or IL-3. A 116-nucleotide (nt) region of the egr-1 promoter which contains sequences inducible by GM-CSF and IL-3 was defined. DNase I footprint analysis identified a 20-nt region, including nt -57 to -76, which contains a potential cyclic AMP (cAMP) response element (CRE). Electrophoretic mobility shift assays performed with CREB antibody confirmed the presence of CREB in the DNA-binding complex. Mutational analysis of the cytokine-responsive region of the egr-1 promoter revealed that both the cAMP response and serum response elements are required for induction by GM-CSF and IL-3. Nuclear extracts from GM-CSF- or IL-3-stimulated but not unstimulated TF-1 cells contain factors which specifically bind to the Egr-1-binding site in the nt -600 to -480 region of the promoter. Electrophoretic mobility shift assays were performed with antibodies against the Egr-1 protein to demonstrate the presence of the protein product in the shifted complex. Our studies suggest that the Egr-1 protein may further stimulate transcription of the egr-1 gene in response to GM-CSF as a secondary event. Images PMID:8065330

  16. 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

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

    PubMed

    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

  18. Myeloid cell kinetics in mice treated with recombinant interleukin-3, granulocyte colony-stimulating factor (CSF), or granulocyte-macrophage CSF in vivo

    SciTech Connect

    Lord, B.I.; Molineux, G.; Pojda, Z.; Souza, L.M.; Mermod, J.J.; Dexter, T.M. )

    1991-05-15

    Myeloid cell kinetics in mice treated with pure hematopoietic growth factors have been investigated using tritiated thymidine labeling and autoradiography. Mice were injected subcutaneously with 125 micrograms/kg granulocyte colony-stimulating factor (G-CSF) (in some cases 5 micrograms/kg), or 10 micrograms/kg of granulocyte-macrophage CSF (GM-CSF), or interleukin-3 (IL-3) every 12 hours for 84 hours. {sup 3}HTdR labeling was performed in vivo after 3 days of treatment. G-CSF increased the peripheral neutrophil count 14-fold and increased the proportion and proliferation rate of neutrophilic cells in the marrow, suppressing erythropoiesis at the same time. Newly produced mature cells were released into the circulation within 24 hours of labeling, compared with a normal appearance time of about 96 hours. By contrast, GM-CSF and IL-3 had little effect on either marrow cell kinetics or on the rate of release of mature cells, although GM-CSF did stimulate a 50% increase in peripheral neutrophils. Monocyte production was also increased about eightfold by G-CSF and 1.5-fold by GM-CSF, but their peak release was only slightly accelerated. While the peripheral half-lives of the neutrophilic granulocytes were normal, those of the monocytes were dramatically reduced, perhaps due to sequestration in the tissues for functional purposes. The stimulated monocyte production in the case of G-CSF required an additional five cell cycles, a level that might have repercussions on the progenitor compartments.

  19. 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

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

    PubMed

    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

  1. Constitutive receptor-independent low density lipoprotein uptake and cholesterol accumulation by macrophages differentiated from human monocytes with macrophage-colony-stimulating factor (M-CSF).

    PubMed

    Zhao, Bin; Li, Yifu; Buono, Chiara; Waldo, Stephen W; Jones, Nancy L; Mori, Masahiro; Kruth, Howard S

    2006-06-01

    Recently, we have shown that macrophage uptake of low density lipoprotein (LDL) and cholesterol accumulation can occur by nonreceptor mediated fluid-phase macropinocytosis when macrophages are differentiated from human monocytes in human serum and the macrophages are activated by stimulation of protein kinase C (Kruth, H. S., Jones, N. L., Huang, W., Zhao, B., Ishii, I., Chang, J., Combs, C. A., Malide, D., and Zhang, W. Y. (2005) J. Biol. Chem. 280, 2352-2360). Differentiation of human monocytes in human serum produces a distinct macrophage phenotype. In this study, we examined the effect on LDL uptake of an alternative macrophage differentiation phenotype. Differentiation of macrophages from human monocytes in fetal bovine serum with macrophage-colony-stimulating factor (M-CSF) produced a macrophage phenotype demonstrating constitutive fluid-phase uptake of native LDL leading to macrophage cholesterol accumulation. Fluid-phase endocytosis of LDL by M-CSF human macrophages showed non-saturable uptake of LDL that did not down-regulate over 48 h. LDL uptake was mediated by continuous actin-dependent macropinocytosis of LDL by these M-CSF-differentiated macrophages. M-CSF is a cytokine present within atherosclerotic lesions. Thus, macropinocytosis of LDL by macrophages differentiated from monocytes under the influence of M-CSF is a plausible mechanism to account for macrophage foam cell formation in atherosclerotic lesions. This mechanism of macrophage foam cell formation does not depend on LDL modification or macrophage receptors. PMID:16606620

  2. 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

  3. Colony-stimulating factor 1-mediated regulation of a chimeric c-fms/v-fms receptor containing the v-fms-encoded tyrosine kinase domain

    SciTech Connect

    Roussel, M.F.; Downing, J.R.; Ashmun, R.A.; Rettenmier, C.W.; Sherr, C.J. )

    1988-08-01

    A chimeric gene specifying the 308 N-terminal amino acids of the extracellular ligand binding domain of the human c-fms protooncogene joined to the remainder of the feline v-fms oncogene product encodes a functional colony-stimulating factor 1 (CSF-1) receptor. When expressed in mouse NIH 3T3 fibroblasts, the chimeric gene product was rapidly transported to the cell surface, was autophosphorylated on tyrosine only in response to human recombinant CSF-1, underwent ligand-induced but not phorbol ester-induced down-modulation, and stimulated CSF-1-dependent cell proliferation. By contrast, the C-terminally truncated glycoprotein encoded by the v-fms oncogene is partially inhibited in its transport to the plasma membrane, is constitutively phosphorylated on tyrosine, and is relatively refractory to both ligand-induced and phorbol ester-induced down-modulation. Although the v-fms oncogene can transform cells in the absence of CSF-1, its tyrosine kinase activity and turnover can be appropriately regulated by the human c-fms-encoded ligand binding domain. The results confirm that C-terminal truncation of the c-fms gene is insufficient to activate its transforming potential and suggest that an additional mutation in its distal extracellular domain is required for oncogenic activation.

  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. PU.1 (Spi-1) and C/EBP alpha regulate the granulocyte colony-stimulating factor receptor promoter in myeloid cells.

    PubMed

    Smith, L T; Hohaus, S; Gonzalez, D A; Dziennis, S E; Tenen, D G

    1996-08-15

    Cytokines, important for lineage commitment and differentiation during hematopoiesis, exert their influence by binding specific receptors. Receptor expression is tightly regulated and examining the factors that govern their expression will allow better understanding of the events that determine lineage commitment. The granulocyte colony-stimulating factor (G-CSF) receptor is expressed exclusively in myeloid cells and the placenta. We show here that the G-CSF receptor transcription start site is identical in each of these tissues. A 1,391-bp fragment of the G-CSF receptor promoter is both active in myeloid cell lines and tissue specific. We have also found two regions that are important for G-CSF receptor promoter activity. One region, located at bp -49, contains a GCAAT site that specifically binds the C/EBP alpha transcription factor in myeloid nuclear extracts. Mutation of this site prevents C/EBP alpha binding and reduces promoter activity by 60%. The other functionally important region of the G-CSF receptor promoter is in the 5' untranslated region, at bp +36 and +43, where there are two sites for the ets family member PU.1. Mutation of these sites prevents PU.1 binding and reduces promoter activity by 75%. These results reinforce the importance of both PU.1 and C/EBP alpha in the expression of myeloid-specific genes and neutrophil development. PMID:8695841

  6. Hypoxic tumor cell death and modulation of endothelial adhesion molecules in the regression of granulocyte colony-stimulating factor-transduced tumors.

    PubMed Central

    Colombo, M. P.; Lombardi, L.; Melani, C.; Parenza, M.; Baroni, C.; Ruco, L.; Stoppacciaro, A.

    1996-01-01

    C-26 colon adenocarcinoma cells transduced with the granulocyte colony-stimulating factor (G-CSF) gene form large tumors when injected into sublethally irradiated mice. These tumors regress when leukocyte function is reconstituted. Electron microscopy and immunocytochemical analysis of regressing C-26/G-CSF nodules indicates that tumor destruction is due mainly to hypoxia resulting from the functional loss of tumor vasculature and is only marginally due to direct cytolysis. Desegregation of basal lamina, cell swelling, and loss of junctions characterized the vessels within regressing tumors. Tumor cells were necrotic or filled with lipid vacuoles regardless of the distance from nearby vessels. Damage of tumor vasculature was dependent on the infiltrating leukocytes and the cytotoxic cytokines they produced. Locally produced interleukin-1 and tumor necrosis factor-alpha (TNF-alpha) induced vascular cellular adhesion molecule-1 (VCAM-1) and E-selectin on tumor vessels. Treatment with monoclonal antibodies to interferon-gamma (IFN-gamma) or TNF-alpha blocked tumor regression by inhibiting VCAM-1 and E-selectin expression on tumor-associated endothelial cells resulting in a reduced number of infiltrating leukocytes. Thus, C-26/G-CSF tumor regression presents features typical of hemorrhagic necrosis that occurs through the cytokines produced by infiltrating leukocytes in response to G-CSF. Images Figure 1 p477-a Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 PMID:8579110

  7. Continuous infusion or subcutaneous injection of granulocyte-macrophage colony-stimulating factor: increased efficacy and reduced toxicity when given subcutaneously.

    PubMed Central

    Honkoop, A. H.; Hoekman, K.; Wagstaff, J.; van Groeningen, C. J.; Vermorken, J. B.; Boven, E.; Pinedo, H. M.

    1996-01-01

    Granulocyte-macrophage colony-stimulating factor (GM-CSF) is a haematopoietic growth factor with a wide variety of applications in the clinic. In early phase I studies the continuous intravenous (c.i.) route of administration was often used. Later it was shown that subcutaneous (s.c.) administration was also effective. The optimal route of administration remains, however, poorly defined, and no studies have made a direct comparison between these two routes of administration. We treated patients with advanced breast cancer with moderately high-dose doxorubicin and cylophosphamide and GM-CSF. The first 14 patients received GM-CSF by c.i, while subsequently 47 patients received it s.c. Comparison between the two groups showed that c.i. GM-CSF was more toxic in several respects. There was a higher need for erythrocyte and platelet transfusions and a significant deterioration in the performance status. This study indicates that subcutaneous GM-CSF is the preferred route of administration. Randomised trials are, however, needed to confirm these conclusions. PMID:8855987

  8. 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

  9. Stromal derived factor‐1 and granulocyte‐colony stimulating factor treatment improves regeneration of Pax7−/− mice skeletal muscles

    PubMed Central

    Kowalski, Kamil; Archacki, Rafał; Archacka, Karolina; Stremińska, Władysława; Paciorek, Anna; Gołąbek, Magdalena; Ciemerych, Maria A.

    2015-01-01

    Abstract Background The skeletal muscle has the ability to regenerate after injury. This process is mediated mainly by the muscle specific stem cells, that is, satellite cells. In case of extensive damage or under pathological conditions, such as muscular dystrophy, the process of muscle reconstruction does not occur properly. The aim of our study was to test whether mobilized stem cells, other than satellite cells, could participate in skeletal muscle reconstruction. Methods Experiments were performed on wild‐type mice and mice lacking the functional Pax7 gene, that is, characterized by the very limited satellite cell population. Gastrocnemius mice muscles were injured by cardiotoxin injection, and then the animals were treated by stromal derived factor‐1 (Sdf‐1) with or without granulocyte‐colony stimulating factor (G‐CSF) for 4 days. The muscles were subjected to thorough assessment of the tissue regeneration process using histological and in vitro methods, as well as evaluation of myogenic factors' expression at the transcript and protein levels. Results Stromal derived factor‐1 alone and Sdf‐1 in combination with G‐CSF significantly improved the regeneration of Pax7−/− skeletal muscles. The Sdf‐1 and G‐CSF treatment caused an increase in the number of mononucleated cells associated with muscle fibres. Further analysis showed that Sdf‐1 and G‐CSF treatment led to the rise in the number of CD34+ and Cxcr4+ cells and expression of Cxcr7. Conclusions Stromal derived factor‐1 and G‐CSF stimulated regeneration of the skeletal muscles deficient in satellite cells. We suggest that mobilized CD34+, Cxcr4+, and Cxcr7+ cells can efficiently participate in the skeletal muscle reconstruction and compensate for the lack of satellite cells. PMID:27239402

  10. Sunlight Triggers Cutaneous Lupus through a Colony Stimulating Factor-1 (CSF-1) Dependent Mechanism in MRL-Faslpr mice

    PubMed Central

    Menke, Julia; Hsu, Mei-Yu; Byrne, Katelyn T.; Lucas, Julie A.; Rabacal, Whitney A.; Croker, Byron P.; Zong, Xiao-Hua; Stanley, E. Richard; Kelley, Vicki R.

    2008-01-01

    Sunlight (UVB) triggers cutaneous (CLE) and systemic lupus through an unknown mechanism. We tested the hypothesis that UVB triggers CLE through a CSF-1-dependent, macrophage (Mø) -mediated mechanism in MRL-Faslpr mice. By constructing mutant MRL-Faslpr strains expressing varying levels of CSF-1 (high, intermediate, none), and use of an ex-vivo gene transfer to deliver CSF-1 intra-dermally, we determined that CSF-1 induces CLE in lupus-susceptible, MRL-Faslpr mice, but not in lupus-resistant, BALB/c mice. Notably, UVB incites an increase in Mø, apoptosis in the skin and CLE in MRL-Faslpr, but not in CSF-1-deficient MRL-Faslpr mice. Furthermore, UVB did not induce CLE in BALB/c mice. Probing further, UVB stimulates CSF-1 expression by keratinocytes leading to recruitment and activation of Mø that, in turn, release mediators, which induce apoptosis in keratinocytes. Thus, sunlight triggers a CSF-1-dependent, Mø-mediated destructive inflammation in the skin leading to CLE in lupus-susceptible MRL-Faslpr, but not lupus-resistant BALB/c mice. Taken together, we envision CSF-1 as the “match” and lupus-susceptibility as the “tinder” leading to CLE. PMID:18981160

  11. 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

  12. 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

  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. The induction of prolonged myelopoietic effects in monkeys by GW003, a recombinant human granulocyte colony-stimulating factor genetically fused to recombinant human albumin.

    PubMed

    Xu, Xianxing; Yang, Jingwen; Liu, Yunlong; Shan, Chengqi; Wang, Qiushi; Chen, Zhihang; Cheng, Yuanguo

    2015-02-01

    GW003, a genetic fusion protein of human serum albumin and granulocyte colony-stimulating factor (G-CSF), was developed based on a novel strategy for producing long-acting proteins. The purpose of this study was to evaluate the hematologic, pharmacokinetic, and toxicokinetic effects of GW003 on cynomolgus monkeys. We show that following a single subcutaneous administration of GW003, the absolute neutrophil count increased significantly compared with monkeys that received only the vehicle, and the magnitude of the neutrophilic response to GW003 was dose dependent. After an injection at equal molar dose, the clearance of GW003 in the monkeys was approximately fourfold slower, and the terminal half-life (T1/2 ) was fivefold longer than the corresponding values for recombinant methionyl human G-CSF. Interestingly, both the clearance and T1/2 decreased with increasing doses of GW003, and much faster elimination was observed after multidose exposure. In toxicokinetic studies, the serum concentration of GW003 after the eighth injection was much lower than it was after the first injection, and a neutralizing antibody against G-CSF was found to have a dose-dependent effect upon the treatment groups. Overall, the favorable pharmacokinetic and pharmacodynamic properties supported the selection and development of GW003 as a promising candidate for neutropenia therapy. PMID:25174614

  15. A Chimeric HIV-1 Envelope Glycoprotein Trimer with an Embedded Granulocyte-Macrophage Colony-stimulating Factor (GM-CSF) Domain Induces Enhanced Antibody and T Cell Responses*

    PubMed Central

    van Montfort, Thijs; Melchers, Mark; Isik, Gözde; Menis, Sergey; Huang, Po-Ssu; Matthews, Katie; Michael, Elizabeth; Berkhout, Ben; Schief, William R.; Moore, John P.; Sanders, Rogier W.

    2011-01-01

    An effective HIV-1 vaccine should ideally induce strong humoral and cellular immune responses that provide sterilizing immunity over a prolonged period. Current HIV-1 vaccines have failed in inducing such immunity. The viral envelope glycoprotein complex (Env) can be targeted by neutralizing antibodies to block infection, but several Env properties limit the ability to induce an antibody response of sufficient quantity and quality. We hypothesized that Env immunogenicity could be improved by embedding an immunostimulatory protein domain within its sequence. A stabilized Env trimer was therefore engineered with the granulocyte-macrophage colony-stimulating factor (GM-CSF) inserted into the V1V2 domain of gp120. Probing with neutralizing antibodies showed that both the Env and GM-CSF components of the chimeric protein were folded correctly. Furthermore, the embedded GM-CSF domain was functional as a cytokine in vitro. Mouse immunization studies demonstrated that chimeric EnvGM-CSF enhanced Env-specific antibody and T cell responses compared with wild-type Env. Collectively, these results show that targeting and activation of immune cells using engineered cytokine domains within the protein can improve the immunogenicity of Env subunit vaccines. PMID:21515681

  16. 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

  17. 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

  18. Second-line treatment of non small cell lung cancer by biweekly gemcitabine and docetaxel +/- granulocyte-macrophage colony stimulating factor and low dose aldesleukine.

    PubMed

    Correale, Pierpaolo; Tindara Miano, Salvatora; Remondo, Cinzia; Migali, Cristina; Rotundo, Maria Saveria; Macrì, Paolo; Tagliaferri, Pierosandro; Caraglia, Michele; Gotti, Giuseppe; Francini, Guido

    2009-03-15

    Background. The antitumor activity of a novel biweekly gemcitabine (G) + docetaxel (D) regimen +/- granulocyte-macrophage colony stimulating factor (GM-CSF) and aldesleukine (IL-2) has been evaluated in a phase II trial in advanced pretreated non-small-cell lung cancer (NSCLC). Results. The treatment was well tolerated. The 42.3% response rate exceeded the predefined target activity, while time to progression (TTP) and overall survival (OS) were 7 and 11.2 months, respectively. A greater objective response rate (58.3% vs 28.6%) and an increased number of eosinophils, basophils and activated mononuclear blood cells were observed in those patients who also received cytokine administration. Methods. Twenty-six NSCLC patients received second line G (1000 mg/m2) and D (75 mg/m2) every 15 days. 12/26 patients also received s.c. GM-CSF (100µg, days 2-6) and s.c. IL-2 (0.5MIU/ twice daily, days 7-14 and 16-29) by random selection. Conclusion. The biweekly GD regimen is a safe and active second-line treatment in NSCLC. Addition of immune-adjuvant cytokines' may enhance the activity of this therapeutic combination. PMID:19242101

  19. 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

  20. Human recombinant granulocyte-macrophage colony-stimulating factor increases cell-to-cell adhesion and surface expression of adhesion-promoting surface glycoproteins on mature granulocytes.

    PubMed Central

    Arnaout, M A; Wang, E A; Clark, S C; Sieff, C A

    1986-01-01

    Human granulocyte-macrophage colony-stimulating factor (GM-CSF) has been shown to inhibit migration of mature granulocytes and to enhance their antibody-dependent cellular cytotoxicity. We found that human recombinant GM-CSF also enhanced granulocyte-granulocyte adhesion and increased by two- to threefold the surface expression of Mo1 and LeuM5 (P150, 95), two members of a family of leukocyte adhesion molecules (Leu-CAM). Increased Mo1 surface expression occurred within 15 min at 37 degrees C and was maximal at the migration inhibitory concentration of 500 pM. One-half maximal rise in the expression of Mo1 on the cell surface occurred at 5 pM. The chemotactic peptide f-Met-Leu-Phe produced a comparable rise in surface Mo1 with one-half maximal expression occurring at 7 nM. Both GM-CSF and f-Met-Leu-Phe produced optimal granulocyte-granulocyte adhesion at 500 pM and 100 nM, respectively. This adhesion-promoting effect induced by either stimulus was inhibited by a mouse monoclonal antibody directed against Mo1 antigen. These data indicate that GM-CSF promotes cell-to-cell adhesion, presumably through enhanced expression of leukocyte adhesion molecules. This mechanism may explain, in part, the known effects of GM-CSF on the function of mature granulocytes. Images PMID:3090106

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

    PubMed

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

    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

  2. Granulocyte colony-stimulating factor in secondary prophylaxis for advanced-stage Hodgkin lymphoma treated with ABVD chemotherapy: a cost-effectiveness analysis.

    PubMed

    Cheung, M C; Prica, A; Graczyk, J; Buckstein, R; Chan, K K W

    2016-08-01

    Granulocyte colony-stimulating factor (G-CSF) is commonly administered to patients with Hodgkin lymphoma (HL) with neutropenia. We constructed a decision-analytic model to compare the cost-effectiveness of secondary prophylaxis with G-CSF to a strategy of 'no G-CSF' in response to severe neutropenia for adults with advanced-stage HL treated with ABVD. A Canadian public health payer's perspective was considered and costs were presented in 2013 Canadian dollars. The quality-adjusted life years (QALYs) attained with the G-CSF and 'no G-CSF' strategies were 1.403 and 1.416, respectively. Costs for the strategies with and without G-CSF were $38,971 and $33,982, respectively. In the base case analysis, the 'no G-CSF' strategy was associated with cost savings and improved QALYs; therefore, 'no G-CSF' was the dominant approach. For patients with severe neutropenia during ABVD chemotherapy for advanced-stage HL, a strategy without G-CSF support is associated with improved quality-adjusted outcomes, cost savings, and is the preferred approach. PMID:26758765

  3. 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

  4. 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. PMID:26828262

  5. Efficacy of gene-therapy based on adenovirus encoding granulocyte-macrophage colony-stimulating factor in drug-sensitive and drug-resistant experimental pulmonary tuberculosis.

    PubMed

    Francisco-Cruz, Alejandro; Mata-Espinosa, Dulce; Ramos-Espinosa, Octavio; Marquina-Castillo, Brenda; Estrada-Parra, Sergio; Xing, Zhou; Hernández-Pando, Rogelio

    2016-09-01

    Tuberculosis (TB), although a curable disease, remains a major cause of morbidity and mortality worldwide. It is necessary to develop a short-term therapy with reduced drug toxicity in order to improve adherence rate and control disease burden. Granulocyte-macrophage colony-stimulating factor (GM-CSF) may be a key cytokine in the treatment of pulmonary TB since it primes the activation and differentiation of myeloid and non-myeloid precursor cells, inducing the release of protective Th1 cytokines. In this work, we administrated by intratracheal route recombinant adenoviruses encoding GM-CSF (AdGM-CSF). This treatment produced significant bacterial elimination when administered in a single dose at 60 days of infection with drug sensitive or drug resistant Mtb strains in a murine model of progressive disease. Moreover, AdGM-CSF combined with primary antibiotics produced more rapid elimination of pulmonary bacterial burdens than conventional chemotherapy suggesting that this form of treatment could shorten the conventional treatment. PMID:27553405

  6. 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

  7. Development and Characterization of a Novel Fusion Protein of a Mutated Granulocyte Colony-Stimulating Factor and Human Serum Albumin in Pichia pastoris

    PubMed Central

    Huang, Yan-Shan; Wen, Xiao-Fang; Yang, Zhi-Yu; Wu, Yi-Liang; Lu, You; Zhou, Lin-Fu

    2014-01-01

    The purpose of the present work was to develop a novel, long-acting and potent human serum albumin/granulocyte colony stimulating factor (HSA/G-CSF) therapeutic fusion protein. The novel fusion protein, called HMG, was constructed by genetically fusing mutated human derived G-CSF (mG-CSF) to the C-terminal of HSA and then prepared in Pichia pastoris. The molecular mass of HMG was about 85 kDa and the isoelectric point was 5.3. Circular dichroism spectroscopy suggested that mG-CSF retained nearly all of its native secondary structure, regardless of fusion. The binding capabilities of mG-CSF moiety to G-CSF receptor and HSA moiety to warfarin showed very little change after fusing. The bioactivity of HMG (11.0×106 IU/mg) was more than twice that of rHSA/G-CSF (4.6×106 IU/mg). A mutation was made at the 718th amino acid of HMG, substituting Ala for Thr, to investigate the glycosylation of HMG expressed in P. pastoris. Data indicated that HMG was modified at Thr718, speculatively with the addition of a mannose chain. In conclusion, a novel HSA/G-CSF fusion protein was successfully constructed based on a mutated G-CSF. This protein showed more potent bioactivity than rHSA/G-CSF and thus may be a suitable long-acting G-CSF. PMID:25535738

  8. 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

  9. 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

  10. 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

  11. 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

  12. A Soluble Granulocyte Colony Stimulating Factor Decoy Receptor as a Novel Tool to Increase Hematopoietic Cell Homing and Reconstitution in Mice

    PubMed Central

    Fortin, Audrey; Benabdallah, Basma; Palacio, Lina; Carbonneau, Cynthia L.; Le, Oanh N.; Haddad, Élie

    2013-01-01

    The relative ineffectiveness of hematopoietic stem cells in reaching the bone marrow upon transplantation combined with the limited number of these cells available is a major reason for graft failure and delayed hematopoietic recovery. Hence, the development of strategies that could enhance homing is of high interest. Here, we provide evidence that homing is severely impaired postexposure to ionizing radiation (IR) in mice, an effect we found was time dependent and could be partially rescued using mesenchymal stromal cell (MSC) therapy. In an attempt to further increase homing, we took advantage of our observation that the granulocyte colony stimulating factor (G-CSF), a cytokine known to induce cell mobilization, is increased in the marrow of mice shortly after their exposure to IR. As such, we developed a truncated, yet functional, soluble G-CSF receptor (solG-CSFR), which we hypothesized could act as a decoy and foster homing. Using MSCs or conditioned media as delivery vehicles, we show that an engineered solG-CSFR has the potential to increase homing and hematopoietic reconstitution in mice. Altogether, our results provide novel findings at the interplay of IR and stromal cell therapy and present the regulation of endogenous G-CSF as an innovative proof-of-concept strategy to manipulate hematopoietic cell homing. PMID:23205715

  13. Pretransplant mobilization with granulocyte colony-stimulating factor improves B-cell reconstitution by lentiviral vector gene therapy in SCID-X1 mice.

    PubMed

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

    2014-10-01

    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

  14. Role of Granulocyte Macrophage Colony-Stimulating Factor in Host Defense Against Pulmonary Cryptococcus neoformans Infection during Murine Allergic Bronchopulmonary Mycosis

    PubMed Central

    Chen, Gwo-Hsiao; Olszewski, Michal A.; McDonald, Roderick A.; Wells, Jason C.; Paine, Robert; Huffnagle, Gary B.; Toews, Galen B.

    2007-01-01

    We investigated the role of granulocyte macrophage colony-stimulating factor (GM-CSF) in host defense in a murine model of pulmonary cryptococcosis induced by intratracheal inoculation of Cryptococcus neoformans. Pulmonary C. neoformans infection of C57BL/6 mice is an established model of an allergic bronchopulmonary mycosis. Our objective was to determine whether GM-CSF regulates the pulmonary Th2 immune response in C. neoformans-infected C57BL/6 mice. Long-term pulmonary fungistasis was lost in GM-CSF knockout (GM−/−) mice, resulting in increased pulmonary burden of fungi between weeks 3 and 5. GM-CSF was required for the early influx of macrophages and CD4 and CD8 T cells into the lungs but was not required later in the infection. Lack of GM-CSF also resulted in reduced eosinophil recruitment and delayed recruitment of mononuclear cells into the airspace. Macrophages from GM+/+ mice showed numerous hallmarks of alternatively activated macrophages: higher numbers of intracellular cryptococci, YM1 crystals, and induction of CCL17. These hallmarks are absent in macrophages from GM−/− mice. Mucus-producing goblet cells were abundantly present within the bronchial epithelial layer in GM+/+ mice but not in GM−/− mice at week 5 after infection. Production of both Th1 and Th2 cytokines was impaired in the absence of GM-CSF, consistent with both reduced C. neoformans clearance and absence of allergic lung pathology. PMID:17322386

  15. 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

  16. Hematopoietic properties of granulocyte colony-stimulating factor/immunoglobulin (G-CSF/IgG-Fc) fusion proteins in normal and neutropenic rodents.

    PubMed

    Cox, George N; Chlipala, Elizabeth A; Smith, Darin J; Carlson, Sharon J; Bell, Stacie J; Doherty, Daniel H

    2014-01-01

    Previously we showed that granulocyte colony-stimulating factor (G-CSF) in vitro bioactivity is preserved when the protein is joined via a flexible 7 amino acid linker to an immunoglobulin-1 (IgG1)-Fc domain and that the G-CSF/IgG1-Fc fusion protein possessed a longer circulating half-life and improved hematopoietic properties compared to G-CSF in normal rats. We have extended this analysis by comparing the relative hematopoietic potencies of G-CSF/IgG1-Fc to G-CSF in normal mice and to G-CSF and polyethylene glycol (PEG) -modified G-CSF in neutropenic rats. Mice were treated for 5 days using different doses and dosing regimens of G-CSF/IgG1-Fc or G-CSF and circulating neutrophil levels in the animals measured on Day 6. G-CSF/IgG1-Fc stimulated greater increases in blood neutrophils than comparable doses of G-CSF when administered using daily, every other day or every third day dosing regimens. In rats made neutropenic with cyclophosphamide, G-CSF/IgG1-Fc accelerated recovery of blood neutrophils to normal levels (from Day 9 to Day 5) when administered as 5 daily injections or as a single injection on Day 1. By contrast, G-CSF accelerated neutrophil recovery when administered as 5 daily injections, but not when administered as a single injection. G-CSF/IgG1-Fc was as effective as PEG-G-CSF at accelerating neutrophil recovery following a single injection in neutropenic rats. G-CSF/IgG1-Fc and G-CSF/IgG4-Fc fusion proteins in which the 7 amino acid linker was deleted also were effective at accelerating neutrophil recovery following a single injection in neutropenic rats. These studies confirm the enhanced in vivo hematopoietic properties of G-CSF/IgG-Fc fusion proteins. PMID:24637521

  17. Effect of recombinant bovine granulocyte colony-stimulating factor covalently bound to polyethylene glycol injection on neutrophil number and function in periparturient dairy cows.

    PubMed

    Kimura, Kayoko; Goff, Jesse P; Canning, Peter; Wang, Chong; Roth, James A

    2014-01-01

    Dairy cows often experience decreased immune function around the time of calving, typified by impaired polymorphonuclear neutrophil (PMN) function and a transient neutropenia. This is associated with increased disease incidence, including mastitis, retained placenta, and metritis. In an attempt to improve PMN functional capacity during the periparturient period, we injected cows with recombinant bovine granulocyte colony-stimulating factor covalently bound to polyethylene glycol (PEG rbG-CSF) twice subcutaneously, about 6d before calving and within 24h after calving. Twenty-one cows in their second pregnancy were enrolled in this study and divided into 2 groups: PEG rbG-CSF treated (n=11) and saline-treated controls (n=10). The PMN numbers quickly and dramatically increased after PEG rbG-CSF administration and remained elevated through the end of the experiment (13d after calving). Exocytosis of myeloperoxidase by stimulated PMN, which is generally decreased in periparturient cows, was markedly increased by PEG rbG-CSF after injection. Higher myeloperoxidase exocytosis persisted for at least 10d after calving. The PMN superoxide anion release and phagocytosis activity did not differ between groups. Injection of PEG rbG-CSF was safe for cows, with no significant negative effects observed. The greatest single effect of PEG rbG-CSF administration was a dramatic increase in circulating numbers of PMN. The increased numbers of PMN ready to move to a site of infection early in the course of an infection may improve the ability of the cow to ward off clinical disease in the periparturient period. PMID:24881799

  18. miR-155 Is Associated with the Leukemogenic Potential of the Class IV Granulocyte Colony-Stimulating Factor Receptor in CD34+ Progenitor Cells

    PubMed Central

    Zhang, HaiJiao; Goudeva, Lilia; Immenschuh, Stephan; Schambach, Axel; Skokowa, Julia; Eiz-Vesper, Britta; Blasczyk, Rainer; Figueiredo, Constança

    2014-01-01

    Granulocyte colony-stimulating factor (G-CSF) is a major regulator of granulopoiesis on engagement with the G-CSF receptor (G-CSFR). The truncated, alternatively spliced, class IV G-CSFR (G-CSFRIV) has been associated with defective differentiation and relapse risk in pediatric acute myeloid leukemia (AML) patients. However, the detailed biological properties of G-CSFRIV in human CD34+ hematopoietic stem and progenitor cells (HSPCs) and the potential leukemogenic mechanism of this receptor remain poorly understood. In the present study, we observed that G-CSFRIV–overexpressing (G-CSFRIV+) HSPCs demonstrated an enhanced proliferative and survival capacity on G-CSF stimulation. Cell cycle analyses showed a higher frequency of G-CSFRIV+ cells in the S and G2/M phase. Also, apoptosis rates were significantly lower in G-CSFRIV+ HSPCs. These findings were shown to be associated with a sustained Stat5 activation and elevated miR-155 expression. In addition, G-CSF showed to further induce G-CSFRIV and miR-155 expression of peripheral blood mononuclear cells isolated from AML patients. A Stat5 pharmacological inhibitor or ribonucleic acid (RNA) interference–mediated silencing of the expression of miR-155 abrogated the aberrant proliferative capacity of the G-CSFRIV+ HSPCs. Hence, the dysregulation of Stat5/miR-155 pathway in the G-CSFRIV+ HSPCs supports their leukemogenic potential. Specific miRNA silencing or the inhibition of Stat5-associated pathways might contribute to preventing the risk of leukemogenesis in G-CSFRIV+ HSPCs. This study may promote the development of a personalized effective antileukemia therapy, in particular for the patients exhibiting higher expression levels of G-CSFRIV, and further highlights the necessity of pre-screening the patients for G-CSFR isoforms expression patterns before G-CSF administration. PMID:25730818

  19. 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.

  20. Hematopoietic Properties of Granulocyte Colony-Stimulating Factor/Immunoglobulin (G-CSF/IgG-Fc) Fusion Proteins in Normal and Neutropenic Rodents

    PubMed Central

    Cox, George N.; Chlipala, Elizabeth A.; Smith, Darin J.; Carlson, Sharon J.; Bell, Stacie J.; Doherty, Daniel H.

    2014-01-01

    Previously we showed that granulocyte colony-stimulating factor (G-CSF) in vitro bioactivity is preserved when the protein is joined via a flexible 7 amino acid linker to an immunoglobulin-1 (IgG1)-Fc domain and that the G-CSF/IgG1-Fc fusion protein possessed a longer circulating half-life and improved hematopoietic properties compared to G-CSF in normal rats. We have extended this analysis by comparing the relative hematopoietic potencies of G-CSF/IgG1-Fc to G-CSF in normal mice and to G-CSF and polyethylene glycol (PEG) - modified G-CSF in neutropenic rats. Mice were treated for 5 days using different doses and dosing regimens of G-CSF/IgG1-Fc or G-CSF and circulating neutrophil levels in the animals measured on Day 6. G-CSF/IgG1-Fc stimulated greater increases in blood neutrophils than comparable doses of G-CSF when administered using daily, every other day or every third day dosing regimens. In rats made neutropenic with cyclophosphamide, G-CSF/IgG1-Fc accelerated recovery of blood neutrophils to normal levels (from Day 9 to Day 5) when administered as 5 daily injections or as a single injection on Day 1. By contrast, G-CSF accelerated neutrophil recovery when administered as 5 daily injections, but not when administered as a single injection. G-CSF/IgG1-Fc was as effective as PEG-G-CSF at accelerating neutrophil recovery following a single injection in neutropenic rats. G-CSF/IgG1-Fc and G-CSF/IgG4-Fc fusion proteins in which the 7 amino acid linker was deleted also were effective at accelerating neutrophil recovery following a single injection in neutropenic rats. These studies confirm the enhanced in vivo hematopoietic properties of G-CSF/IgG-Fc fusion proteins. PMID:24637521

  1. 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

  2. Distribution of granulocyte-monocyte colony-stimulating factor and its receptor α-subunit in the adult human brain with specific reference to Alzheimer's disease.

    PubMed

    Ridwan, Sami; Bauer, Henrike; Frauenknecht, Katrin; von Pein, Harald; Sommer, Clemens J

    2012-11-01

    Granulocyte-monocyte colony-stimulating factor (GM-CSF) is a member of the hematopoietic growth factor family, promoting proliferation and differentiation of hematopoietic progenitor cells of the myeloid lineage. In recent years, GM-CSF has also proved to be an important neurotrophic factor in the central nervous system (CNS) via binding to the GM-CSF receptor (GM-CSF R). Furthermore, studies on rodent CNS revealed a wide distribution of both the major binding α-subunit of the GM-CSF R (GM-CSF Rα) and its ligand. Since respective data on the expression pattern of these two molecules are still lacking, the present study has been designed to systematically analyze the protein expression of GM-CSF and GM-CSF Rα in the human brain, with particular emphasis on their regulation in Alzheimer's disease (AD). One major finding is that both GM-CSF and GM-CSF Rα were ubiquitously but not uniformly expressed in neurons throughout the CNS. Protein expression of GM-CSF and GM-CSF Rα was not restricted to neurons but also detectable in astrocytes, ependymal cells and choroid plexus cells. Interestingly, distribution and intensity of immunohistochemical staining for GM-CSF did not differ among AD brains and age-matched controls. Concerning GM-CSF Rα, a marked reduction of protein expression was predominantly detected in the hippocampus although a slight reduction was also found in various cortical regions, thalamic nuclei and some brainstem nuclei. Since the hippocampus is one of the target regions of neurodegenerative changes in AD, reduction of GM-CSF Rα, with consecutive downregulation of GM-CSF signaling, may contribute to in the progressive course of neurodegeneration in AD. PMID:22430742

  3. Granulocyte colony stimulating factor (GCSF) improves memory and neurobehavior in an amyloid-β induced experimental model of Alzheimer's disease.

    PubMed

    Prakash, Ajay; Medhi, Bikash; Chopra, Kanwaljit

    2013-09-01

    GCSF is an endogenous neuronal hematopoietic factor that displays robust in vitro and in vivo neuroprotective activity. The present study aimed to evaluate the effect of GCSF on Aβ-induced memory loss in an Alzheimer's disease model of rats. A total of 42 male adult Wistar rats weighing 200-250 g were used in the study and were divided into 7 experimental groups. Animals were subjected to intracerebroventricular (ICV) injection stereotaxically at day 0 to instill amyloid-β(1-42) (Aβ(1-42)) or PBS (sham operated group) at 10 μl (5 μl bilaterally). GCSF treatment was given from day 7 to 12 of Aβ injection. On day 21, behavioral tests (short term memory, exploratory behavior and motor coordination) in all groups were evaluated. Biochemical parameters and RNA expression were measured to ensure the efficacy of GCSF. GCSF (35 and 70 μg/kg, s.c.) showed statistically significant improvement in memory as compared to control and sham operated groups (p<0.05). Mean time spent in the platform placed quadrant was found to be significantly increased in the GCSF (70 μg/kg, s.c.) as compared to GCSF (35 μg/kg, s.c.) and GCSF (10 μg/kg, s.c.) groups (p<0.001). GCSF (35 and 70 μg/kg, s.c.) also improved motor coordination and exploratory behavior significantly as compared to naïve sham operated and GCSF (10 μg/kg, s.c.) groups (p<0.05). Improvement in memory by GCSF (35 and 70 μg/kg, s.c.) was coupled with marked reduction of lipid peroxidation, acetylcholinesterase levels and a significant increase in antioxidant enzymes as well as total RNA expression in the brain. Additionally, GCSF (35 and 70 μg/kg, s.c.) significantly increased progenitor cells (iPSCs) and surface marker CD34+ in the brain and hence induced neurogenesis. The present findings demonstrate an improvement of memory and neurobehavioral function with GCSF in Aβ-induced Alzheimer's disease model in rats. PMID:23756182

  4. 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.

  5. 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 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. 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

  7. Substance P enhances tissue factor release from granulocyte-macrophage colony-stimulating factor-dependent macrophages via the p22phox/β-arrestin 2/Rho A signaling pathway.

    PubMed

    Yamaguchi, Rui; Yamamoto, Takatoshi; Sakamoto, Arisa; Ishimaru, Yasuji; Narahara, Shinji; Sugiuchi, Hiroyuki; Yamaguchi, Yasuo

    2016-03-01

    Granulocyte-macrophage colony stimulating factor (GM-CSF) induces procoagulant activity of macrophages. Tissue factor (TF) is a membrane-bound glycoprotein and substance P (SP) is a pro-inflammatory neuropeptide involved in the formation of membrane blebs. This study investigated the role of SP in TF release by GM-CSF-dependent macrophages. SP significantly decreased TF levels in whole-cell lysates of GM-CSF-dependent macrophages. TF was detected in the culture supernatant by enzyme-linked immunosorbent assay after stimulation of macrophages by SP. Aprepitant (an SP/neurokinin 1 receptor antagonist) reduced TF release from macrophages stimulated with SP. Pretreatment of macrophages with a radical scavenger(pyrrolidinedithiocarbamate) also limited the decrease of TF in whole-cell lysates after stimulation with SP. A protein kinase C inhibitor (rottlerin) partially blocked this macrophage response to SP, while it was significantly inhibited by a ROCK inhibitor (Y-27632) or a dynamin inhibitor (dinasore). An Akt inhibitor (perifosine) also partially blocked this response. Furthermore, siRNA targeting p22phox, β-arrestin 2, or Rho A, blunted the release of TF from macrophages stimulated with SP. In other experiments, visceral adipocytes derived from cryopreserved preadipocytes were found to produce SP. In conclusion, SP enhances the release of TF from macrophages via the p22phox/β-arrestin 2/Rho A signaling pathway. PMID:26852662

  8. Granulocyte colony-stimulating factor: A relation between serum and follicular fluid levels and in-vitro fertilization outcome in patients with polycystic ovary syndrome.

    PubMed

    Kahyaoglu, Inci; Yılmaz, Nafiye; Timur, Hakan; Inal, Hasan Ali; Erkaya, Salim

    2015-07-01

    Evidence is accumulating in the literature about the potential role of serum and follicular fluid (FF) granulocyte colony-stimulating factor (G-CSF) as a non-invasive biomarker of oocyte competence and embryo selection in in-vitro fertilization (IVF) cycles. In this study, we aimed to evaluate the effect of serum and FF G-CSF levels on IVF outcome in non-hyperandrogenic, non-obese patients with polycystic ovary syndrome (PCOS). Twenty-two patients with PCOS (Group I), and 22 patients with the etiology of male factor infertility (Group II) undergoing IVF treatment were included. Demographic features, controlled ovarian stimulation parameters, neutrophil count (NC), neutrophil/leukocyte (N/L) ratio, serum and FF G-CSF levels of the two groups were compared. Serum E2 level on the day of hCG (2982.5±171.4 vs. 2279.0±207.2 pg/mL), total number of retrieved oocytes (14.7±0.9 vs. 11.5±1.3) and mature oocytes (11.6±0.8 vs. 9.1±1.1) were significantly higher in group I when compared to group II (p<0.05). On the day of oocyte retrieval, both the mean serum (54.8±1.7 vs. 48.1±0.9 pg/mL) and FF G-CSF levels (48.8±1.4 vs. 44.1±0.5 pg/mL), NC (4.4±0.2×10(3) vs. 3.6±0.3×10(3)/μL) and N/L ratio (63.6±1.4 vs. 56.1±1.7) in group I were found to be significantly higher than group II ((p<0.05). Despite the increased levels of G-CSF both in the serum and follicular microenvironment in patients with PCOS, a relation between G-CSF and good ovarian response or clinical pregnancy rates could not be demonstrated in this study. PMID:25258001

  9. 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

  10. Tyrosine 763 of the murine granulocyte colony-stimulating factor receptor mediates Ras-dependent activation of the JNK/SAPK mitogen-activated protein kinase pathway.

    PubMed Central

    Rausch, O; Marshall, C J

    1997-01-01

    The receptor for granulocyte colony-stimulating factor (G-CSF) can mediate differentiation and proliferation of hemopoietic cells. A proliferative signal is associated with activation of the ERK mitogen-activated protein kinase (MAPK) pathway. To determine whether other MAPK pathways are activated by G-CSF signalling, we have investigated activation of JNK/SAPK in cells proliferating in response to G-CSF. Here we show that G-CSF and interleukin-3 activate JNK/SAPK in two hemopoietic cell lines. The region of the G-CSF receptor required for G-CSF-induced JNK/SAPK activation is located within the C-terminal 68 amino acids of the cytoplasmic domain, which contains Tyr 763. Mutation of Tyr 763 to Phe completely blocks JNK/SAPK activation. However, the C-terminal 68 amino acids are not required for ERK2 activation. We show that activation of JNK/SAPK, like that of ERK2, is dependent on Ras but that higher levels of Ras-GTP are associated with activation of JNK/SAPK than with activation of ERK2. Two separate functional regions of the G-CSF receptor contribute to activation of Ras. The Y763F mutation reduces G-CSF-induced Ras activation from 30 to 35% Ras-GTP to 10 to 13% Ras-GTP. Low levels of Ras activation (10 to 13% Ras-GTP), which are sufficient for ERK2 activation, require only the 100 membrane-proximal amino acids. High levels of Ras-GTP provided by expression of oncogenic Ras are not sufficient to activate JNK/SAPK. An additional signal, also mediated by Tyr 763, is required for activation of JNK/SAPK. PMID:9032244

  11. Extra-pulmonary aspects of acquired pulmonary alveolar proteinosis as predicted by granulocyte-macrophage colony-stimulating factor-deficient mice.

    PubMed

    Seymour, John F

    2006-01-01

    Granulocyte-macrophage colony-stimulating factor (GM-CSF)-/- mice are an invaluable model for exploring the effects of systemic GM-CSF deficiency. Their lung phenotype exactly reproduces the abnormalities seen in human pulmonary alveolar proteinosis (PAP). However, GM-CSF-/- mice also have significant systemic functional abnormalities. These include immune defects which result in a reduced susceptibility to a range of experimentally induced autoimmune disorders. These immunological defects are also functionally manifest as an impaired ability to resolve a range of infections under certain conditions, usually implicating cellular effectors, including Listeria, Group B streptococcus, adenovirus, Pneumocystis carinii, and malaria. These observations are consistent with the known propensity for patients with PAP to develop a range of opportunistic infections. Conversely, the diminished immunological response to inflammatory stimuli may be beneficial in some settings by limiting inflammatory cell recruitment and pro-inflammatory mediator-release. GM-CSF-/- mice also have distinct fertility defects, manifest as reduced litter size and an increased rate of early fetal loss. These observations may be clinically relevant for women affected by PAP and further support the evaluation of the role of GM-CSF in human reproduction. These observations reinforce the importance of clinicians viewing PAP as a state of systemic functional GM-CSF deficiency, albeit with prominent pulmonary manifestations, rather than purely a 'lung disease'. These systemic manifestations of GM-CSF deficiency should also be considered when deciding on the choice between pulmonary or systemic delivery of GM-CSF as therapy for PAP, as only systemic drug delivery has the potential capacity to correct the systemic manifestations of GM-CSF deficiency in these patients. PMID:16423263

  12. 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

  13. Induction of Monocyte Chemoattractant Proteins in Macrophages via the Production of Granulocyte/Macrophage Colony-Stimulating Factor by Breast Cancer Cells

    PubMed Central

    Yoshimura, Teizo; Imamichi, Tomozumi; Weiss, Jonathan M.; Sato, Miwa; Li, Liangzhu; Matsukawa, Akihiro; Wang, Ji Ming

    2016-01-01

    Monocyte chemoattractant protein-1 (MCP-1)/CCL2 plays an important role in the initiation and progression of cancer. We previously reported that in 4T1 murine breast cancer, non-tumor stromal cells, including macrophages, were the major source of MCP-1. In the present study, we analyzed the potential mechanisms by which MCP-1 is upregulated in macrophages infiltrating 4T1 tumors. We found that cell-free culture supernatants of 4T1 cells (4T1-sup) markedly upregulated MCP-1 production by peritoneal inflammatory macrophages. 4T1-sup also upregulated other MCPs, such as MCP-3/CCL7 and MCP-5/CCL12, but modestly upregulated neutrophil chemotactic chemokines, such as KC/CXCL1 or MIP-2/CXCL2. Physicochemical analysis indicated that an approximately 2–3 kDa 4T1 cell product was responsible for the capacity of 4T1-sup to upregulate MCP-1 expression by macrophages. A neutralizing antibody against granulocyte/macrophage colony-stimulating factor (GM-CSF), but not macrophage CSF, almost completely abrogated MCP-1-inducing activity of 4T1-sup, and recombinant GM-CSF potently upregulated MCP-1 production by macrophages. The expression levels of GM-CSF in 4T1 tumors in vivo were higher than other tumors, such as Lewis lung carcinoma. Treatment of mice with anti-GM-CSF antibody significantly reduced the growth of 4T1 tumors at the injection sites but did not reduce MCP-1 production or lung metastasis in tumor-bearing mice. These results indicate that 4T1 cells have the capacity to directly upregulate MCP-1 production by macrophages by releasing GM-CSF; however, other mechanisms are also involved in increased MCP-1 levels in the 4T1 tumor microenvironment. PMID:26834744

  14. 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

  15. Local transplantation of granulocyte colony stimulating factor-mobilized CD34+ cells for patients with femoral and tibial nonunion: pilot clinical trial.

    PubMed

    Kuroda, Ryosuke; Matsumoto, Tomoyuki; Niikura, Takahiro; Kawakami, Yohei; Fukui, Tomoaki; Lee, Sang Yang; Mifune, Yutaka; Kawamata, Shin; Fukushima, Masanori; Asahara, Takayuki; Kawamoto, Atsuhiko; Kurosaka, Masahiro

    2014-01-01

    Most bone fractures typically heal, although a significant proportion (5%-10%) of fractures fail to heal, resulting in delayed union or persistent nonunion. Some preclinical evidence shows the therapeutic potential of peripheral blood CD34(+) cells, a hematopoietic/endothelial progenitor cell-enriched population, for bone fracture healing; however, clinical outcome following transplantation of CD34(+) cells in patients with fracture has never been reported. We report a phase I/IIa clinical trial regarding transplantation of autologous, granulocyte colony stimulating factor-mobilized CD34(+) cells with atelocollagen scaffold for patients with femoral or tibial fracture nonunion (n = 7). The primary endpoint of this study is radiological fracture healing (union) by evaluating anteroposterior and lateral views at week 12 following cell therapy. For the safety evaluation, incidence, severity, and outcome of all adverse events were recorded. Radiological fracture healing at week 12 was achieved in five of seven cases (71.4%), which was greater than the threshold (18.1%) predefined by the historical outcome of the standard of care. The interval between cell transplantation and union, the secondary endpoint, was 12.6 ± 5.4 weeks (range, 8-24 weeks) for clinical healing and 16.1 ± 10.2 weeks (range, 8-36 weeks) for radiological healing. Neither deaths nor life-threatening adverse events were observed during the 1-year follow-up after the cell therapy. These results suggest feasibility, safety, and potential effectiveness of CD34(+) cell therapy in patients with nonunion. PMID:24307697

  16. Granulocyte-Macrophage Colony-Stimulating Factor Priming plus Papillomavirus E6 DNA Vaccination: Effects on Papilloma Formation and Regression in the Cottontail Rabbit Papillomavirus-Rabbit Model

    PubMed Central

    Leachman, Sancy A.; Tigelaar, Robert E.; Shlyankevich, Mark; Slade, Martin D.; Irwin, Michele; Chang, Ed; Wu, T. C.; Xiao, Wei; Pazhani, Sundaram; Zelterman, Daniel; Brandsma, Janet L.

    2000-01-01

    A cottontail rabbit papillomavirus (CRPV) E6 DNA vaccine that induces significant protection against CRPV challenge was used in a superior vaccination regimen in which the cutaneous sites of vaccination were primed with an expression vector encoding granulocyte-macrophage colony-stimulating factor (GM-CSF), a cytokine that induces differentiation and local recruitment of professional antigen-presenting cells. This treatment induced a massive influx of major histocompatibility complex class II-positive cells. In a vaccination-challenge experiment, rabbit groups were treated by E6 DNA vaccination, GM-CSF DNA inoculation, or a combination of both treatments. After two immunizations, rabbits were challenged with CRPV at low, moderate, and high stringencies and monitored for papilloma formation. As expected, all clinical outcomes were monotonically related to the stringency of the viral challenge. The results demonstrate that GM-CSF priming greatly augmented the effects of CRPV E6 vaccination. First, challenge sites in control rabbits (at the moderate challenge stringency) had a 0% probability of remaining disease free, versus a 50% probability in E6-vaccinated rabbits, and whereas GM-CSF alone had no effect, the interaction between GM-CSF priming and E6 vaccination increased disease-free survival to 67%. Second, the incubation period before papilloma onset was lengthened by E6 DNA vaccination alone or to some extent by GM-CSF DNA inoculation alone, and the combination of treatments induced additive effects. Third, the rate of papilloma growth was reduced by E6 vaccination and, to a lesser extent, by GM-CSF treatment. In addition, the interaction between the E6 and GM-CSF treatments was synergistic and yielded more than a 99% reduction in papilloma volume. Finally, regression occurred among the papillomas that formed in rabbits treated with the E6 vaccine and/or with GM-CSF, with the highest regression frequency occurring in rabbits that received the combination

  17. 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.

  18. A randomised trial of granulocyte-macrophage colony-stimulating factor for neonatal sepsis: childhood outcomes at 5 years

    PubMed Central

    Marlow, Neil; Morris, Timothy; Brocklehurst, Peter; Carr, Robert; Cowan, Frances; Patel, Nishma; Petrou, Stavros; Redshaw, Margaret; Modi, Neena; Doré, Caroline J

    2015-01-01

    Objective We performed a randomised trial in very preterm, small for gestational age (SGA) babies to determine if prophylaxis with granulocyte macrophage colony stimulating factor (GM-CSF) improves outcomes (the PROGRAMS trial). GM-CSF was associated with improved neonatal neutrophil counts, but no change in other neonatal or 2-year outcomes. As subtle benefits in outcome may not be ascertainable until school age we performed an outcome study at 5 years. Patients and methods 280 babies born at 31 weeks of gestation or less and SGA were entered into the trial. Outcomes were assessed at 5 years to determine neurodevelopmental and general health status and educational attainment. Results We found no significant differences in cognitive, general health or educational outcomes between 83 of 106 (78%) surviving children in the GM-CSF arm compared with 81 of 110 (74%) in the control arm. Mean mental processing composite (equivalent to IQ) at 5 years were 94 (SD 16) compared with 95 (SD 15), respectively (difference in means −1 (95%CI −6 to 4), and similar proportions were in receipt of special educational needs support (41% vs 35%; risk ratio 1.2 (95% CI 0.8 to 1.9)). Performance on Kaufmann-ABC subscales and components of NEPSY were similar. The suggestion of worse respiratory outcomes in the GM-CSF group at 2 years was replicated at 5 years. Conclusions The administration of GM-CSF to very preterm SGA babies is not associated with improved or more adverse neurodevelopmental, general health or educational outcomes at 5 years. Trial registration number ISRCTN42553489. PMID:25922190

  19. 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. PMID:26376304

  20. The High Effect of Chemomobilization with High-Dose Etopside + Granulocyte-Colony Stimulating Factor in Autologous Hematopoietic Peripheral Blood Stem Cell Transplantation: A Single Center Experience

    PubMed Central

    Yanmaz, Mustafa Teoman; Selvi, Ahmet; Usul, Cigdem

    2016-01-01

    Autologous hematopoietic stem cell transplantation (auto-HSCT) provides hematopoietic support after high-dose chemotherapy and is the standard of care for patients with multiple myeloma (MM), chemo sensitive relapsed high or intermediate grade non-Hodgkin’s lymphoma (NHL) and Hodgkin’s lymphoma (HL). However, yields of hematopoietic stem cells vary greatly between patients, and the optimal strategy to mobilize hematopoietic stem cells into peripheral blood for collection has not been defined yet. We investigated the efficacy and safety of chemo mobilization with an intermediate dose etoposide (VP-16; 200 mg/m2 on days 1-3) and granulocyte-colony stimulating factor (G-CSF)(5 µg/kg twice daily from day 4 through the final day of collection). We reviewed our institutional experience with 91 patients (71 MM, 12 HL, 8 NHL) mobilized with this regimen. VP-16 + G-CSF resulted in successful mobilization in 95.55% of the patients (on one patient stem cell collection with plerixafor was applied), including 76 patients (83.52%) whose stem cells were collected successfully in a single day. Collection was managed between min. D8 and max. D17. Patient age, gender, exposure to previous irradiation and chemotherapy, previous mobilization attempts, and disease characteristics were not considered during selection. Adverse effects of the regimen included supportive transfusions and fevers requiring hospitalization or intravenous antibiotics. VP-16 and G-CSF appears to be a safe and effective mobilization regimen for patients with multiple myeloma, non-Hodgkin’s lymphoma and Hodgkin’s lymphoma undergoing autologous stem cell transplantation, producing excellent stem cell yield with the majority of patients requiring 1 day of apheresis. PMID:27103979

  1. Granulocyte Colony-Stimulating Factor Use in a Large Iranian Hospital: Comparison with American Society of Clinical Oncology (ASCO) Clinical Practice Guideline

    PubMed Central

    Mousavi, Sarah; Dadpoor, Mina; Ashrafi, Farzaneh

    2016-01-01

    Background: Granulocyte Colony Stimulating Factors (GCSF) is high-cost agents commonly recommended for primary and secondary prophylaxis of chemotherapy-induced neutropenia and febrile neutropenia. GCSFs have been shown to be beneficial in some patient subgroups, although they are probably overused in clinical settings. The American Society of Clinical Oncology (ASCO) guidelines summarize current data on the appropriate use of CSFs. The aim of this study was to assess and audit the use of GCSF in a tertiary care center according to the recommendation of ASCO guideline. Subjects and Methods: A prospective observational study from November 2014 to June 2015 was performed on all patients prescribed with filgrastim in the large teaching hospital (Isfahan, Iran). Data was collected on demographics, indication, dosing regimen and duration of treatment, the Absolute Neutrophil Count (ANC) and patient outcome. Results: 91 patients were recorded over the period of the study. 63.7% of prescription complied with the ASCO guideline. Febrile neutropenia post chemotherapy/radiotherapy was the most common appropriate indication (29.3%) followed by primary prophylaxis (25.8%). Fourteen (32%) patients showed ANC recovery in 1-3 days and 16 (37%) within 4-7 days. Ten patients (23%) showed no recovery. The overall mortality was 8 (8.8%) patients. Conclusion: This study revealed that at least one-third of prescribed GCSF was not in accordance with ASCO guideline. Considering the high cost of GCSF in our country and limitation of our resources, we proposed cost-effectiveness studies on GCSF treatment and also the development of a national guideline for optimizing GCSF use. PMID:27252808

  2. Autocrine protective mechanisms of human granulocyte colony-stimulating factor (G-CSF) on retinal ganglion cells after optic nerve crush.

    PubMed

    Huang, Shun-Ping; Fang, Kan-Tang; Chang, Chung-Hsing; Huang, Tzu-Lun; Wen, Yao-Tseng; Tsai, Rong-Kung

    2016-02-01

    This study investigated the role of autocrine mechanisms in the anti-apoptotic effects of human granulocyte colony-stimulating factor (G-CSF) on retinal ganglion cells (RGCs) after optic nerve (ON) crush. We observed that both G-CSF and G-CSF receptor (G-CSFR) are expressed in normal rat retina. Further dual immunofluorescence staining showed G-CSFR immunoreactive cells were colocalized with RGCs, Müller cells, horizontal and amacrine cells. These results confirm that G-CSF is an endogenous ligand in the retina. The semi-quantitative RT-PCR finding demonstrated the transcription levels of G-CSF and G-CSFR were up-regulated after ON crush injury. G-CSF treatment further increased and prolonged the expression level of G-CSFR in the retina. G-CSF has been shown to enhance transdifferentiation of the mobilized hematopoietic stem cells into tissue to repair central nervous system injury. We test the hypothesis that the hematopoietic stem cells recruited by G-CSF treatment can transdifferentiate into RGCs after ON crush by performing sublethal irradiation of the rats 5 days before ON crush. The flow cytometric analysis showed the number of CD34 positive cells in the peripheral blood is significantly lower in the irradiated, crushed and G-CSF-treated group than the sham control group or crush and G-CSF treated group. Nevertheless, the G-CSF treatment enhances the RGC survival after sublethal irradiation and ON crush injury. These data indicate that G-CSF seems unlikely to induce hematopoietic stem cell transdifferentiation into RGCs after ON crush injury. In conclusion, G-CSF may serve an endogenous protective signaling in the retina through direct activation of intrinsic G-CSF receptors and downstream signaling pathways to rescue RGCs after ON crush injury, exogenous G-CSF administration can enhance the anti-apoptotic effects on RGCs. PMID:26518178

  3. The Development of Murine Plasmacytoid Dendritic Cell Precursors Is Differentially Regulated by FLT3-ligand and Granulocyte/Macrophage Colony-Stimulating Factor

    PubMed Central

    Gilliet, Michel; Boonstra, Andre; Paturel, Carine; Antonenko, Svetlana; Xu, Xiu-Ling; Trinchieri, Giorgio; O'Garra, Anne; Liu, Yong-Jun

    2002-01-01

    Plasmacytoid predendritic cells or type 1 interferon (IFN)-producing cells (IPCs) have recently been identified in mice. Although culture systems giving rise to different murine dendritic cell subsets have been established, the developmental regulation of murine plasmacytoid IPCs and the culture conditions leading to their generation remain unknown. Here we show that large numbers of over 40% pure CD11c+CD11b−B220+Gr-1+ IPCs can be generated from mouse bone marrow cultures with FLT3-ligand. By contrast GM-CSF or TNF-α, which promote the generation of CD11c+CD11b+B220− myeloid DCs, block completely the development of IPCs. IPCs generated display similar features to human IPCs, such as the plasmacytoid morphology, the ability to produce large amounts of IFN-α in responses to herpes simplex virus, and the capacity to respond to ligands for Toll-like receptor 9 (TLR-9; CpG ODN 1668), but not to ligands for TLR-4 (lipopolysaccharide [LPS]). Unlike human IPCs which produce little IL-12p70, mouse IPCs produce IL-12p70 in response to CpG ODN 1668 and herpes simplex virus. This study demonstrates that the development of murine CD11c+CD11b−B220+Gr-1+ IPCs and CD11c+CD11b+B220− myeloid DCs is differentially regulated by FLT3-ligand and granulocyte/macrophage colony-stimulating factor. Human IPCs and mouse IPCs display different ability to produce IL-12p70. Large numbers of mouse IPCs can now be obtained from total bone marrow culture. PMID:11927638

  4. Double-Blinded, Randomized Phase II Study Using Embolization with or without Granulocyte–Macrophage Colony-Stimulating Factor in Uveal Melanoma with Hepatic Metastases

    PubMed Central

    Valsecchi, Matias E.; Terai, Mizue; Eschelman, David J.; Gonsalves, Carin F.; Chervoneva, Inna; Shields, Jerry A.; Shields, Carol L.; Yamamoto, Akira; Sullivan, Kevin L.; Laudadio, MaryAnn; Berd, David; Mastrangelo, Michael J.

    2015-01-01

    Purpose To investigate the effects of immunoembolization with granulocyte–macrophage colony-stimulating factor (GM-CSF) in patients with uveal melanoma (UM) with liver-only metastasis. Materials and Methods In this double-blind phase II clinical trial, patients were randomized to undergo immunoembolization or bland embolization (BE). Lobar treatment was performed with GM-CSF or normal saline solution mixed with ethiodized oil followed by embolization with gelatin sponge emulsified with iodinated contrast medium. Fifty-two patients (immunoembolization, n = 25; BE, n = 27) were enrolled. Response was assessed after every two treatments. The primary endpoint was overall response rate (ORR) of liver metastases. Progression-free survival (PFS), overall survival (OS), and immunologic responses were secondary endpoints. Results There were five partial responses in the immunoembolization group (ORR, 21.2%; 90% confidence interval [CI], 10.3%–30.5%) and three in the BE group (ORR, 16.7%; 90% CI, 6.3%–26.9%). Stable disease was seen in 12 patients in the immunoembolization group and 19 in the BE group. OS times were 21.5 months (95% CI, 18.5–24.8 mo) with immunoembolization and 17.2 months (95% CI, 11.9–22.4 mo) with BE. The degree of proinflammatory cytokine production was more robust after immunoembolization and correlated with time to “systemic” extrahepatic progression. In the immunoembolization group, interleukin (IL)-6 levels at 1 hour (P = .001) and IL-8 levels at 18 hours after the procedure (P < .001) were significant predictors of longer systemic PFS. Moreover, a dose–response pattern was evident between posttreatment serum cytokine concentrations and systemic PFS. Conclusions Immunoembolization induced more robust inflammatory responses, which correlated with the delayed progression of extrahepatic systemic metastases. PMID:25678394

  5. Four-Week Repeated Intravenous Dose Toxicity and Toxicokinetic Study of TS-DP2, a Novel Human Granulocyte Colony Stimulating Factor in Rats

    PubMed Central

    Lee, JooBuom; Lee, Kyungsun; Choe, Keunbum; Jung, Hyunseob; Cho, Hyunseok; Choi, Kiseok; Kim, Taegon; Kim, Seojin; Lee, Hyeong-Seok; Cha, Mi-Jin; Song, Si-Whan; Lee, Chul Kyu; Chun, Gie-Taek

    2015-01-01

    TS-DP2 is a recombinant human granulocyte colony stimulating factor (rhG-CSF) manufactured by TS Corporation. We conducted a four-week study of TS-DP2 (test article) in repeated intravenous doses in male and female Sprague-Dawley (SD) rats. Lenograstim was used as a reference article and was administered intravenously at a dose of 1000 μg/kg/day. Rats received TS-DP2 intravenously at doses of 250, 500, and 1000 μg/kg/day once daily for 4 weeks, and evaluated following a 2-week recovery period. Edema in the hind limbs and loss of mean body weight and body weight gain were observed in both the highest dose group of TS-DP2 and the lenograstim group in male rats. Fibro-osseous lesions were observed in the lenograstim group in both sexes, and at all groups of TS-DP2 in males, and at doses of TS-DP2 500 μg/kg/day and higher in females. The lesion was considered a toxicological change. Therefore, bone is the primary toxicological target of TS-DP2. The lowest observed adverse effect level (LOAEL) in males was 250 μg/kg/day, and no observed adverse effect level (NOAEL) in females was 250 μg/kg/day in this study. In the toxicokinetic study, the serum concentrations of G-CSF were maintained until 8 hr after administration. The systemic exposures (AUC0-24h and C0) were not markedly different between male and female rats, between the administration periods, or between TS-DP2 and lenograstim. In conclusion, TS-DP2 shows toxicological similarity to lenograstim over 4-weeks of repeated doses in rats. PMID:26877840

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

    PubMed Central

    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. PMID:26376304

  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. 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

  9. 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.

  10. Granulocyte colony-stimulating factor inhibits CXCR4/SDF-1α signaling and overcomes stromal-mediated drug resistance in the HL-60 cell line

    PubMed Central

    SHENG, XIANFU; ZHONG, HUA; WAN, HAIXIA; ZHONG, JIHUA; CHEN, FANGYUAN

    2016-01-01

    Combining cytarabine, aclarubicin and granulocyte colony-stimulating factor (G-CSF) has demonstrated marked efficacy in the treatment of elderly and relapsed/refractory patients with acute myeloid leukemia (AML); however, the role of G-CSF remains poorly understood. The present study aimed to investigate the ability of G-CSF to overcome stromal-mediated drug resistance and the underlying molecular mechanism. Two types of co-culture models were established in the HS-5 human bone marrow/stromal and HL-60 human promyelocytic leukemia cell lines, in order to imitate the interactions between stromal and leukemia cells in vitro, which is mediated by the stromal cell-derived factor (SDF)-1α signaling axis. In the present study, HL-60 cells were attracted and adhered to HS-5 cells using migration assay and flow cytometry, respectively; however, these interactions were inhibited by treatment with G-CSF and/or the C-X-C chemokine receptor type 4 (CXCR4) antagonist, AMD3100. Co-culture with HS-5 cells, including direct and indirect contact, protected HL-60 cells against spontaneous apoptosis or drug-induced apoptosis; however, these protective effects were disrupted by treatment with G-CSF and/or AMD3100. Notably, G-CSF and/or AMD3100 did not alter cell viability or apoptosis when HL-60 cells were cultured with medium alone. In addition, G-CSF significantly reduced the expression levels of surface CXCR4 protein, total CXCR4 protein and CXCR4 mRNA, and significantly upregulated the expression of microRNA (miR)-146a. Conversely, AMD3100 significantly reduced surface CXCR4 expression levels, but not the total CXCR4, CXCR4 mRNA or miR-146a expression levels. The results of the present study suggested that interfering with the CXCR4/SDF-1α signaling axis via G-CSF inhibited the migration and adhesion of HL-60 cells to HS-5 cells and eliminated HS5 cell-mediated protective effects. Furthermore, G-CSF administration reduced CXCR4 expression levels by upregulating the expression of

  11. PU.1 (Spi-1) and C/EBP alpha regulate expression of the granulocyte-macrophage colony-stimulating factor receptor alpha gene.

    PubMed Central

    Hohaus, S; Petrovick, M S; Voso, M T; Sun, Z; Zhang, D E; Tenen, D G

    1995-01-01

    Growth factor receptors play an important role in hematopoiesis. In order to further understand the mechanisms directing the expression of these key regulators of hematopoiesis, we initiated a study investigating the transcription factors activating the expression of the granulocyte-macrophage colony-stimulating factor (GM-CSF) receptor alpha gene. Here, we demonstrate that the human GM-CSF receptor alpha promoter directs reporter gene activity in a tissue-specific fashion in myelomonocytic cells, which correlates with its expression pattern as analyzed by reverse transcription PCR. The GM-CSF receptor alpha promoter contains an important functional site between positions -53 and -41 as identified by deletion analysis of reporter constructs. We show that the myeloid and B cell transcription factor PU.1 binds specifically to this site. Furthermore, we demonstrate that a CCAAT site located upstream of the PU.1 site between positions -70 and -54 is involved in positive-negative regulation of the GM-CSF receptor alpha promoter activity. C/EBP alpha is the major CCAAT/enhancer-binding protein (C/EBP) form binding to this site in nuclear extracts of U937 cells. Point mutations of either the PU.1 site or the C/EBP site that abolish the binding of the respective factors result in a significant decrease of GM-CSF receptor alpha promoter activity in myelomonocytic cells only. Furthermore, we demonstrate that in myeloid and B cell extracts, PU.1 forms a novel, specific, more slowly migrating complex (PU-SF) when binding the GM-CSF receptor alpha promoter PU.1 site. This is the first demonstration of a specific interaction with PU.1 on a myeloid PU.1 binding site. The novel complex is distinct from that described previously as binding to B cell enhancer sites and can be formed by addition of PU.1 to extracts from certain nonmyeloid cell types which do not express PU.1, including T cells and epithelial cells, but not from erythroid cells. Furthermore, we demonstrate that the PU

  12. Role of the pituitary–adrenal axis in granulocyte-colony stimulating factor-induced neuroprotection against hypoxia–ischemia in neonatal rats

    PubMed Central

    Charles, Mélissa S.; Ostrowski, Robert P.; Manaenko, Anatol; Duris, Kamil; Zhang, John H.; Tang, Jiping

    2013-01-01

    Several reports indicate that the activity of the hypothalamic–pituitary–adrenal axis (HPA) is increased after a brain insult and that its down-regulation can improve detrimental outcomes associated with ischemic brain injuries. Granulocyte-colony stimulating factor (G-CSF) is a neuroprotective drug shown in the naïve rat to regulate hormones of the HPA axis. In this study we investigate whether G-CSF confers its neuroprotective properties by influencing the HPA response after neonatal hypoxia–ischemia (HI). Following the Rice–Vannucci model, seven day old rats (P7) were subjected to unilateral carotid ligation followed by 2.5 h of hypoxia. To test our hypothesis, metyrapone was administered to inhibit the release of rodent specific glucocorticoid, corticosterone, at the adrenal level. Dexamethasone, a synthetic glucocorticoid, was administered to agonize the effects of corticosterone. Our results show that both G-CSF and metyrapone significantly reduced infarct volume while dexamethasone treatment did not reduce infarct size even when combined with G-CSF. The protective effects of G-CSF do not include blood brain barrier preservation as suggested by the brain edema results. G-CSF did not affect the pituitary released adrenocorticotropic hormone (ACTH) levels in the blood plasma at 4 h, but suppressed the increase of corticosterone in the blood. The administration of G-CSF and metyrapone increased weight gain, and significantly reduced the Bax/Bcl-2 ratio in the brain while dexamethasone reversed the effects of G-CSF. The combination of G-CSF and metyrapone significantly decreased caspase-3 protein levels in the brain, and the effect was antagonized by dexamethasone. We report that G-CSF is neuroprotective in neonatal HI by reducing infarct volume, by suppressing the HI-induced increase of the Bax/Bcl-2 ratio, and by decreasing corticosterone in the blood. Metyrapone was able to confer similar neuroprotection as G-CSF while dexamethasone reversed the

  13. 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

  14. Multimodal Approaches for Regenerative Stroke Therapies: Combination of Granulocyte Colony-Stimulating Factor with Bone Marrow Mesenchymal Stem Cells is Not Superior to G-CSF Alone

    PubMed Central

    Balseanu, Adrian Tudor; Buga, Ana-Maria; Catalin, Bogdan; Wagner, Daniel-Christoph; Boltze, Johannes; Zagrean, Ana-Maria; Reymann, Klaus; Schaebitz, Wolf; Popa-Wagner, Aurel

    2014-01-01

    Attractive therapeutic strategies to enhance post-stroke recovery of aged brains include methods of cellular therapy that can enhance the endogenous restorative mechanisms of the injured brain. Since stroke afflicts mostly the elderly, it is highly desirable to test the efficacy of cell therapy in the microenvironment of aged brains that is generally refractory to regeneration. In particular, stem cells from the bone marrow allow an autologous transplantation approach that can be translated in the near future to the clinical practice. Such a bone marrow-derived therapy includes the grafting of stem cells as well as the delayed induction of endogenous stem cell mobilization and homing by the stem cell mobilizer granulocyte colony-stimulating factor (G-CSF). We tested the hypothesis that grafting of bone marrow-derived pre-differentiated mesenchymal cells (BM-MSCs) in G-CSF-treated animals improves the long-term functional outcome in aged rodents. To this end, G-CSF alone (50 μg/kg) or in combination with a single dose (106 cells) of rat BM MSCs was administered intravenously to Sprague-Dawley rats at 6 h after transient occlusion (90 min) of the middle cerebral artery. Infarct volume was measured by magnetic resonance imaging at 3 and 48 days post-stroke and additionally by immunhistochemistry at day 56. Functional recovery was tested during the entire post-stroke survival period of 56 days. Daily treatment for post-stroke aged rats with G-CSF led to a robust and consistent improvement of neurological function after 28 days. The combination therapy also led to robust angiogenesis in the formerly infarct core and beyond in the “islet of regeneration.” However, G-CSF + BM MSCs may not impact at all on the spatial reference-memory task or infarct volume and therefore did not further improve the post-stroke recovery. We suggest that in a real clinical practice involving older post-stroke patients, successful regenerative therapies would have to be

  15. Recombinant human granulocyte colony-stimulating factor (rh-G-CSF) may accelerate hematopoietic recovery after HLA-identical sibling allogeneic peripheral blood stem cell transplantation.

    PubMed

    Ozcan, M; Ustün, C; Akçağlayan, E; Akan, H; Arslan, O; Ilhan, O; Beksaç, M; Gürman, G; Demirer, T; Arat, M; Celebi, H; Konuk, N; Uysal, A; Koç, H

    2001-03-01

    We studied the effects of recombinant human granulocyte colony-stimulating factor (G-CSF) on hematopoietic recovery and clinical outcome in patients undergoing allogeneic peripheral blood stem cell (PBSC) transplantation. Fifty-six patients with hematological malignancies who underwent allogeneic PBSC transplantation between 1995 and 1998 were entered into this study. Twenty-eight patients who received daily G-CSF from day +1 after allogeneic PBSC transplantation until the absolute neutrophil count (ANC) reached >0.5 x 10(9)/l for 3 consecutive days were compared with 28 patients (control group) who did not receive G-CSF in a non-randomized manner. The study group and the control group were comparable with respect to baseline patient and transplantation characteristics. Median times to ANC of >0.5 x 10(9)/l and 1 x 10(9)/l with or without G-CSF were 12 days (range 8-21), 13 days (10-32) (P = 0.04) and 13 days (9-21), 15 days (11-44) (P = 0.02), respectively. Median times to reach a platelet count of >20 x 10(9)/l with and without G-CSF were 11 days (0-20) and 13 days (9-26), respectively (P = 0.03). The incidence of febrile episodes was significantly lower with G-CSF, 75% vs 100% (P = 0.008). Patients receiving G-CSF had less grade III-IV mucositis than those who did not receive G-CSF (P = 0.01). There was also no increase in the incidence and severity of acute GVHD in patients using G-CSF (P = 0.22). Although the number of relapsing patients was greater in the G-CSF group (seven vs three patients), this was not statistically significant (P = 0.24). Disease-free and overall survival rates did not differ between the two groups (P = 0.58 and 0.53, respectively). The administration of G-CSF after allogeneic PBSC transplantation provided faster neutrophil and platelet engraftment associated with less severe mucositis and less febrile episodes. PMID:11313683

  16. 9- and 13-Hydroxy-octadecadienoic acids (9+13 HODE) are inversely related to granulocyte colony stimulating factor and IL-6 in runners after 2h running.

    PubMed

    Nieman, David C; Meaney, Mary Pat; John, Casey S; Knagge, Kevin J; Chen, Huiyuan

    2016-08-01

    This study utilized a pro-inflammatory exercise mode to explore potential linkages between increases in 9- and 13-hydroxy-octadecadienoic acid (9+13 HODE) and biomarkers for inflammation, oxidative stress, and muscle damage. Male (N=10) and female (N=10) runners ran at ∼70% VO2max for 1.5h followed by 30min of downhill running (-10%). Blood samples were taken pre-run and immediately-, 1-h-, and 24-h post-run, and analyzed for 9+13 HODE, F2-isoprostanes, six cytokines, C-reactive protein (CRP), creatine kinase (CK), and myoglobin (MYO). Gender groups performed at comparable relative heart rate and oxygen consumption levels during the 2-h run. All outcome measures increased post-run (time effects, P⩽0.001), with levels near pre-run levels by 24h except for CRP, CK, MYO, and delayed onset of muscle soreness (DOMS). Plasma 9+13 HODE increased 314±38.4% post-run (P<0.001), 77.3±15.8% 1-h post-run (P<0.001), and 40.6±16.4% 24-h post-exercise (P=0.024), and F2-isoprostanes increased 50.8±8.9% post-run (P<0.001) and 19.0±5.3% 1-h post-run (P=0.006). Post-run increases were comparable between genders for all outcomes except for 9+13 HODE (interaction effect, P=0.024, post-run tending higher in females), IL-10 (P=0.006, females lower), and DOMS (P=0.029, females lower). The pre-to-post-run increase in 9+13 HODEs was not related to other outcomes except for plasma granulocyte colony stimulating factor (GCSF) (r=-0.710, P<0.001) and IL-6 (r=-0.457, P=0.043). Within the context of this study, exercise-induced increases in 9+13 HODEs tended higher in females, and were not related to increases in F2-isoprostanes, muscle damage, or soreness. The negative relationships to GCSF and IL-6 suggest a linkage between 9+13 HODES and exercise-induced neutrophil chemotaxis, degranulation, and inflammation. PMID:27018002

  17. 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

  18. 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. PMID:16818292

  19. Generation of dendritic cells in vitro from peripheral blood mononuclear cells with granulocyte-macrophage-colony-stimulating factor, interleukin-4, and tumor necrosis factor-alpha for use in cancer immunotherapy.

    PubMed Central

    Morse, M A; Zhou, L J; Tedder, T F; Lyerly, H K; Smith, C

    1997-01-01

    OBJECTIVE: The purpose of the study was to characterize the requirements in terms of precursors, developmental pathways, and media for the generation of large numbers of mature dendritic cells (DC) under conditions acceptable for use in adjuvant, active immunotherapy strategies for surgically treated malignancies. SUMMARY BACKGROUND DATA: Although limited previously by the small numbers accessible, DC-based immunotherapies for malignancy have become more realistic with the development of methods for efficiently generating larger numbers of DC from peripheral blood mononuclear cells (PBMC) in vitro, but these methods rely on clinically unacceptable culture conditions (such as inclusion of fetal bovine serum), necessitating the development of methods for generating functionally equivalent DC in serum-free conditions. METHODS: Plastic-adherent PBMC (from healthy donors and patients with cancer) were incubated for 7 days with granulocyte-macrophage-colony-stimulating factor (GM-CSF) and interleukin-4 (IL-4) with and without tumor necrosis factor-alpha (TNF-alpha) in fetal bovine serum-containing and serum-free media and were analyzed by Wright's stain for morphology, flow cytometry for phenotype, and mixed lymphocyte reaction for allostimulatory function. RESULTS: Growth in either serum-containing or serum-free media supplemented with GM-CSF and IL-4 yielded a similarly heterogeneous population of cells, 6% to 10% of which had the morphology (large cells with thin projections), immunophenotype (including CD83+), and function of mature DC. Tumor necrosis factor-alpha significantly augmented the number of these mature DC, whereas preculture depletion of CD14+ PBMC virtually eliminated them. CONCLUSIONS: Generation of mature DC in the authors' serum-free clinically applicable conditions is similar to serum-containing conditions and requires CD14+ precursors, differentiation through a CD14-CD83- immature stage under the influence of GM-CSF and IL-4, and maturation into a

  20. Synthesis of membrane-bound colony-stimulating factor 1 (CSF-1) and downmodulation of CSF-1 receptors in NIH 3T3 cells transformed by cotransfection of the human CSF-1 and c-fms (CSF-1 receptor) genes.

    PubMed Central

    Rettenmier, C W; Roussel, M F; Ashmun, R A; Ralph, P; Price, K; Sherr, C J

    1987-01-01

    NIH 3T3 cells cotransfected with the human c-fms proto-oncogene together with a 1.6-kilobase cDNA clone encoding a 256-amino-acid precursor of the human mononuclear phagocyte colony-stimulating factor CSF-1 (M-CSF) undergo transformation by an autocrine mechanism. The number of CSF-1 receptors on the surface of transformed cells was regulated by ligand-induced receptor degradation and was inversely proportional to the quantity of CSF-1 produced. A tyrosine-to-phenylalanine mutation at position 969 near the receptor carboxyl terminus potentiated its transforming efficiency in cells cotransfected by the CSF-1 gene but did not affect receptor downmodulation. CSF-1 was synthesized as an integral transmembrane glycoprotein that was rapidly dimerized through disulfide bonds. The homodimer was externalized at the cell surface, where it underwent proteolysis to yield the soluble growth factor. Trypsin treatment of viable cells cleaved the plasma membrane form of CSF-1 to molecules of a size indistinguishable from that of the extracellular growth factor, suggesting that trypsinlike proteases regulate the rate of CSF-1 release from transformed cells. The data raise the possibility that this form of membrane-bound CSF-1 might stimulate receptors on adjacent cells through direct cell-cell interactions. Images PMID:3039346

  1. 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.

  2. The toothless osteopetrotic rat has a normal vitamin D-binding protein-macrophage activating factor (DBP-MAF) cascade and chondrodysplasia resistant to treatments with colony stimulating factor-1 (CSF-1) and/or DBP-MAF.

    PubMed

    Odgren, P R; Popoff, S N; Safadi, F F; MacKay, C A; Mason-Savas, A; Seifert, M F; Marks, S C

    1999-08-01

    The osteopetrotic rat mutation toothless (tl) is characterized by little or no bone resorption, few osteoclasts and macrophages, and chondrodysplasia at the growth plates. Short-term treatment of tl rats with colony-stimulating factor-1 (CSF-1) has been shown to increase the number of osteoclasts and macrophages, producing dramatic resolution of skeletal sclerosis at some, but not all, sites. Defects in production of vitamin D-binding protein-macrophage activating factor (DBP-MAF) have been identified in two other independent osteopetrotic mutations of the rat (op and ia), and two in the mouse (op and mi), in which macrophages and osteoclasts can be activated by the administration of exogenous DBP-MAF. The present studies were undertaken to examine the histology and residual growth defects in tl rats following longer CSF-1 treatments, to investigate the possibility that exogenous DBP-MAF might act synergistically with CSF-1 to improve the tl phenotype, and to assess the integrity of the endogenous DBP-MAF pathway in this mutation. CSF-1 treatment-with or without DBP-MAF-induced resorption of metaphyseal bone to the growth plate on the marrow side, improved slightly but did not normalize long bone growth, and caused no improvement in the abnormal histology of the growth plate. Injections of lysophosphatidylcholine (lyso-Pc) to prime macrophage activation via the DBP-MAF pathway raised superoxide production to similar levels in peritoneal macrophages from both normal and mutant animals, indicating no defect in the DBP-MAF pathway in tl rats. Interestingly, pretreatments with CSF-1 alone also increased superoxide production, although the mechanism for this remains unknown. In summary, we find that, unlike other osteopetrotic mutations investigated to date, the DBP-MAF pathway does not appear to be defective in the tl rat; that additional DBP-MAF does not augment the beneficial skeletal effects seen with CSF-1 alone; and that the growth plate chondrodystrophy seen in

  3. Protein phosphatase 2A is expressed in response to colony-stimulating factor 1 in macrophages and is required for cell cycle progression independently of extracellular signal-regulated protein kinase activity.

    PubMed Central

    Wilson, N J; Moss, S T; Csar, X F; Ward, A C; Hamilton, J A

    1999-01-01

    Colony-stimulating factor 1 (CSF-1) is required for the development of monocytes/macrophages from progenitor cells and for the survival and activation of mature macrophages. The receptor for CSF-1 is the product of the c-fms proto-oncogene, which, on binding ligand, can stimulate a mitogenic response in the appropriate cells. To investigate which genes are regulated in response to CSF-1-stimulation in murine bone-marrow-derived macrophages (BMM), we employed mRNA differential display reverse transcriptase-mediated PCR to identify cDNA species induced by CSF-1. Both Northern and Western blot analyses confirmed the increased expression of one of the cDNA species identified as coding for the catalytic subunit of protein phosphatase 2A (PP2A), an observation not previously reported during the response to a growth factor. To determine the significance of the increased expression of PP2A in response to CSF-1, the PP2A inhibitor okadaic acid (OA) was added to CSF-1-treated BMM and found to inhibit DNA synthesis in a dose-dependent manner. Further analysis with flow cytometry in the presence of OA led to the novel conclusion that PP2A activity is critical for CSF-1-driven BMM cell cycle progression in both early G1 and S phases. Surprisingly, in the light of previous studies with other cells, the PP2A-dependent proliferation could be dissociated from activation by extracellular signal-regulated protein kinase (ERK) in macrophages because OA did not affect either the basal or CSF-1-induced ERK activity in BMM. Two-dimensional SDS/PAGE analysis of lysates of 32P-labelled BMM, which had been treated with CSF-1 in the presence or absence of OA, identified candidate substrates for PP2A. PMID:10215588

  4. 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

  5. A low-dose cytarabine, aclarubicin and granulocyte colony-stimulating factor priming regimen versus a daunorubicin plus cytarabine regimen as induction therapy for older patients with acute myeloid leukemia: A propensity score analysis.

    PubMed

    Minakata, Daisuke; Fujiwara, Shin-Ichiro; Ito, Shoko; Mashima, Kiyomi; Umino, Kento; Nakano, Hirofumi; Kawasaki, Yasufumi; Sugimoto, Miyuki; Yamasaki, Ryoko; Yamamoto, Chihiro; Ashizawa, Masahiro; Hatano, Kaoru; Okazuka, Kiyoshi; Sato, Kazuya; Oh, Iekuni; Ohmine, Ken; Suzuki, Takahiro; Muroi, Kazuo; Kanda, Yoshinobu

    2016-03-01

    This retrospective analysis compared the efficacy of intensive induction therapy consisting of daunorubicin and cytarabine (DNR-AraC) to that of less-intensive therapy including low-dose cytarabine, aclarubicin and granulocyte colony-stimulating factor priming (CAG). Patients aged 60 years or older who were newly diagnosed as acute myeloid leukemia (AML) were analyzed. Sixty-four and 48 patients were treated with DNR-AraC and CAG, respectively. The complete remission rates, 3-year overall survival and event-free survival in the DNR-AraC group were significantly superior to those in the CAG group (65.6% vs. 29.2%, p<0.001, 38.4% vs. 12.3%, p=0.0033, and 20.3% vs. 7.8%, p=0.0030, respectively), although these differences were not statistically significant in multivariate analyses. Next, we calculated a propensity score for selecting the CAG regimen from six factors. The DNR-AraC regimen was associated with better survival than the CAG regimen in a low propensity score group, but there was no difference in survival between regimens in a high propensity score group. Intensive therapy should be performed for patients with sufficient general and comorbid conditions, but less-intensive therapy may be sufficient for patients with higher age, myelodysplasia-related changes, and lower white blood cell counts, which were relevant factors in the propensity score calculation. PMID:26790727

  6. Effects of molecular liposomal hybrid compositions with oxidized dextrans and isonicotinic acid hydrazide on production of granulocytic macrophage colony-stimulating factor by macrophages.

    PubMed

    Shkurupy, V A; Arkhipov, S A; Troitsky, A V; Luzgina, N G; Zaikovskaja, M V; Ufimceva, E G; Iljine, D A; Akhramenko, E S; Gulyaeva, E P; Bistrova, T N

    2009-10-01

    The effects of molecular liposomal hybrid compositions consisting of liposomes (200-450 nm) containing oxidized dextrans (dextranals; 35-60 kDa) conjugated with isonicotinic acid hydrazide (dextrazides), their components, and native dextrans on the production of granulocytic macrophage CSF by peritoneal macrophages were studied in vitro. Dextranals proved to be more potent inductors of granulocytic macrophage CSF than native dextrans. Conjugation of nicotinic acid hydrazide with dextranals did not modify their capacity to stimulate the production of granulocytic macrophage CSF. Liposomes in the molecular liposomal hybrid compositions did not attenuate the dextrazide capacity to stimulate the production of granulocytic macrophage CSF. Molecular liposomal compositions containing 60 kDa dextrazide exhibited the most potent stimulatory effect on macrophage production of granulocytic macrophage CSF. PMID:20396775

  7. 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

  8. CBL Linker Region and RING Finger Mutations Lead to Enhanced Granulocyte-Macrophage Colony-stimulating Factor (GM-CSF) Signaling via Elevated Levels of JAK2 and LYN*

    PubMed Central

    Javadi, Mojib; Richmond, Terri D.; Huang, Kai; Barber, Dwayne L.

    2013-01-01

    Juvenile myelomonocytic leukemia (JMML) is characterized by hypersensitivity to granulocyte-macrophage colony-stimulating factor (GM-CSF). SHP2, NF-1, KRAS, and NRAS are mutated in JMML patients, leading to aberrant regulation of RAS signaling. A subset of JMML patients harbor CBL mutations associated with 11q acquired uniparental disomy. Many of these mutations are in the linker region and the RING finger of CBL, leading to a loss of E3 ligase activity. We investigated the mechanism by which CBL-Y371H, a linker region mutant, and CBL-C384R, a RING finger mutant, lead to enhanced GM-CSF signaling. Expression of CBL mutants in the TF-1 cell line resulted in enhanced survival in the absence of GM-CSF. Cells expressing CBL mutations displayed increased phosphorylation of GM-CSF receptor βc subunit in response to stimulation, although expression of total GM-CSFR βc was lower. This suggested enhanced kinase activity downstream of GM-CSFR. JAK2 and LYN kinase expression is elevated in CBL-Y371H and CBL-C384R mutant cells, resulting in enhanced phosphorylation of CBL and S6 in response to GM-CSF stimulation. Incubation with the JAK2 inhibitor, TG101348, abolished the increased phosphorylation of GM-CSFR βc in cells expressing CBL mutants, whereas treatment with the SRC kinase inhibitor dasatinib resulted in equalization of GM-CSFR βc phosphorylation signal between wild type CBL and CBL mutant samples. Dasatinib treatment inhibited the elevated phosphorylation of CBL-Y371H and CBL-C384R mutants. Our study indicates that CBL linker and RING finger mutants lead to enhanced GM-CSF signaling due to elevated kinase expression, which can be blocked using small molecule inhibitors targeting specific downstream pathways. PMID:23696637

  9. Co-administration of plasmid-encoded granulocyte-macrophage colony-stimulating factor increases human immunodeficiency virus-1 DNA vaccine-induced polyfunctional CD4+ T-cell responses

    PubMed Central

    Santana, Vinicius Canato; Almeida, Rafael Ribeiro; Ribeiro, Susan Pereira; Ferreira, Luís Carlos de Souza; Kalil, Jorge; Rosa, Daniela Santoro; Cunha-Neto, Edecio

    2015-01-01

    T-cell based vaccines against human immunodeficiency virus (HIV) generate specific responses that may limit both transmission and disease progression by controlling viral load. Broad, polyfunctional, and cytotoxic CD4+T-cell responses have been associated with control of simian immunodeficiency virus/HIV-1 replication, supporting the inclusion of CD4+ T-cell epitopes in vaccine formulations. Plasmid-encoded granulocyte-macrophage colony-stimulating factor (pGM-CSF) co-administration has been shown to induce potent CD4+ T-cell responses and to promote accelerated priming and increased migration of antigen-specific CD4+ T-cells. However, no study has shown whether co-immunisation with pGM-CSF enhances the number of vaccine-induced polyfunctional CD4+ T-cells. Our group has previously developed a DNA vaccine encoding conserved, multiple human leukocyte antigen (HLA)-DR binding HIV-1 subtype B peptides, which elicited broad, polyfunctional and long-lived CD4+ T-cell responses. Here, we show that pGM-CSF co-immunisation improved both magnitude and quality of vaccine-induced T-cell responses, particularly by increasing proliferating CD4+ T-cells that produce simultaneously interferon-γ, tumour necrosis factor-α and interleukin-2. Thus, we believe that the use of pGM-CSF may be helpful for vaccine strategies focused on the activation of anti-HIV CD4+ T-cell immunity. PMID:26602876

  10. Co-administration of plasmid-encoded granulocyte-macrophage colony-stimulating factor increases human immunodeficiency virus-1 DNA vaccine-induced polyfunctional CD4+ T-cell responses.

    PubMed

    Santana, Vinicius Canato; Almeida, Rafael Ribeiro; Ribeiro, Susan Pereira; Ferreira, Luís Carlos de Souza; Kalil, Jorge; Rosa, Daniela Santoro; Cunha-Neto, Edecio

    2015-12-01

    T-cell based vaccines against human immunodeficiency virus (HIV) generate specific responses that may limit both transmission and disease progression by controlling viral load. Broad, polyfunctional, and cytotoxic CD4+T-cell responses have been associated with control of simian immunodeficiency virus/HIV-1 replication, supporting the inclusion of CD4+ T-cell epitopes in vaccine formulations. Plasmid-encoded granulocyte-macrophage colony-stimulating factor (pGM-CSF) co-administration has been shown to induce potent CD4+ T-cell responses and to promote accelerated priming and increased migration of antigen-specific CD4+ T-cells. However, no study has shown whether co-immunisation with pGM-CSF enhances the number of vaccine-induced polyfunctional CD4+ T-cells. Our group has previously developed a DNA vaccine encoding conserved, multiple human leukocyte antigen (HLA)-DR binding HIV-1 subtype B peptides, which elicited broad, polyfunctional and long-lived CD4+ T-cell responses. Here, we show that pGM-CSF co-immunisation improved both magnitude and quality of vaccine-induced T-cell responses, particularly by increasing proliferating CD4+ T-cells that produce simultaneously interferon-γ, tumour necrosis factor-α and interleukin-2. Thus, we believe that the use of pGM-CSF may be helpful for vaccine strategies focused on the activation of anti-HIV CD4+ T-cell immunity. PMID:26602876

  11. 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

  12. 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

  13. 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.

  14. Characterization of the human granulocyte-macrophage colony-stimulating factor gene promoter: an AP1 complex and an Sp1-related complex transactivate the promoter activity that is suppressed by a YY1 complex.

    PubMed Central

    Ye, J; Zhang, X; Dong, Z

    1996-01-01

    It is well documented that a repeated CATT element in the human granulocyte-macrophage colony-stimulating factor (GM-CSF) gene promoter is required for promoter activity. However, the transcription factors that are able to transactivate this enhancer element remain unidentified. Recently, we have found that nuclear factor YY1 can interact with the enhancer element. Here, we report that in addition to YY1, two other nuclear factors have been identified in the DNA-protein complexes formed by the CATT oligonucleotide and the Jurkat T-cell nuclear protein. One of these factors is AP1, and the other one is an Sp1-related protein. Results from transient transfection of Jurkat T cells have revealed that formation of both AP1 and the Sp1-related complex is required for the full enhancer activity of the CATT element. This result is supported by cotransfection of a c-jun expression vector and mutational analysis of the AP1 site or the Sp1-related protein binding site. In contrast, formation of the YY1 complex suppresses enhancer activity, since deletion of the YY1 complex induces an augmentation of the enhancer activity and overexpression of YY1 results in an attenuation of the enhancer activity. Results from the mechanism study have revealed that YY1 is able to inhibit transactivation mediated by either AP1 or the Sp1-related protein, and YY1 suppressive activity is DNA binding dependent. Taken together, these data support the ideas that AP1 and the Sp1-related nuclear protein are required for transactivation of the human GM-CSF gene promoter and that YY1 can suppress transactivation of the promoter even under inducible conditions. PMID:8524292

  15. 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

  16. 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. PMID:26694130

  17. Colony-stimulating factor-1 (CSF-1) receptor-mediated macrophage differentiation in myeloid cells: a role for tyrosine 559-dependent protein phosphatase 2A (PP2A) activity.

    PubMed Central

    McMahon, K A; Wilson, N J; Marks, D C; Beecroft, T L; Whitty, G A; Hamilton, J A; Csar, X F

    2001-01-01

    M1 myeloid cells transfected with the wild-type (WT) colony-stimulating factor-1 (CSF-1) receptor (CSF-1R; M1/WT cells) undergo CSF-1-dependent macrophage differentiation. By mutation studies, we have provided prior evidence that tyrosine 559 in the CSF-1R cytoplasmic domain governs the Src-dependent differentiation pathway. Further components of this pathway were then sought. We report that the extent of CSF-1-mediated tyrosine phosphorylation of protein phosphatase 2A (PP2A), and the associated loss of its activity were reduced in M1 cells transfected with the CSF-1R with a tyrosine-to-phenylalanine mutation at position 559 (M1/559 cells), compared with the corresponding responses in CSF-1-treated M1/WT cells. This evidence for an involvement of a reduction in PP2A activity in the differentiation process was supported by the restoration of the defect in the CSF-1-mediated differentiation of M1/559 cells by the addition of the PP2A inhibitor, okadaic acid. It was also found that the degree of activation of extracellular-signal-regulated kinase (ERK) activities by CSF-1 was reduced in M1/559 cells, suggesting their involvement in the differentiation process. These data suggest that PP2A and ERK form part of the Src-dependent signal-transduction cascade governing CSF-1-mediated macrophage differentiation in M1 cells. PMID:11513742

  18. A point mutation at tyrosine-809 in the human colony-stimulating factor 1 receptor impairs mitogenesis without abrogating tyrosine kinase activity, association with phosphatidylinositol 3-kinase, or induction of c-fos and junB genes

    SciTech Connect

    Roussel, M.F. ); Shurtleff, S.A.; Downing, J.R. ); Sherr, C.J. Saint Jude Children's Research Hospital, Memphis, TN )

    1990-09-01

    Substitution of phenylalanine for tyrosine-809 in the human colony-stimulating factor 1 receptor (CSF-1R) inhibited its ability to transduce ligand-dependent mitogenic signals in mouse NIH 3T3 cells. When combined with an activating mutation at codon 301 that induces constitutive CSF-1R tyrosine kinase activity, the codon 809 mutation suppressed ligand-independent cell transformation. Comparative mapping tryptic phosphopeptides from mutant and wild-type CSF-1R indicated that tyrosine-809 is a site of ligand-dependent receptor phosphorylation in vivo. The mutant receptor was active as a tyrosine kinase in vitro and in vivo, underwent CSF-1-dependent association with a phosphatidylinositol 3-kinase, and induced expression of the protooncogenes c-fos and junB, underscoring its ability to trigger some of the known cellular responses to CSF-1. The mutant receptor is likely to be impaired in its ability to interact with critical cellular effectors whose activity is required for mitogenesis.

  19. Combined plerixafor and granulocyte colony-stimulating factor for harvesting high-dose hematopoietic stem cells: Possible niche for plerixafor use in pediatric patients.

    PubMed

    Bitan, Menachem; Eshel, Rinat; Sadot, Efraim; Friedman, Shirley; Pinhasov, Aviva; Levin, Dror; Dvir, Rina; Manisterski, Michal; Berger-Achituv, Sivan; Rosenfeld-Keidar, Hila; Elhasid, Ronit

    2016-06-01

    PB is a source of HSC, especially for autologous HCT in solid tumors. However, there is a risk of failing to achieve the target number of SC after mobilization with growth factors alone in patients who were heavily pretreated with chemotherapy or those in need for tandem transplants. SC were harvested from seven pediatric patients with solid tumors who were in need of autologous HCT following combination GCSF and plerixafor. Six of them received plerixafor after failing to achieve enough SC with GCSF only, while the seventh patient received the combined protocol upfront. All seven patients achieved the target number of SC according to their treatment protocol. There were no adverse events. All patients underwent autologous HCT using the harvested HSC and achieved full engraftment. A protocol for harvesting autologous HCT using GCSF and plerixafor is feasible and safe in children with solid tumors who had been heavily pretreated with chemotherapy or needed tandem transplants. PMID:26991903

  20. Granulocyte-macrophage colony stimulating factor (GM-CSF) enhances the clinical responses to Interferon-α (IFN) in newly diagnosed chronic myeloid leukemia (CML)

    PubMed Central

    Zeidner, Joshua F; Gladstone, Douglas E; Zahurak, Marianna; Matsui, William H; Gocke, Christopher; Jones, Richard J; Smith, B Douglas

    2014-01-01

    The majority of chronic myeloid leukemia (CML) patients treated with tyrosine kinase inhibitors (TKIs) remain with residual disease. In contrast to TKIs, interferon (IFN) is directly toxic to CML progenitor cells, and myeloid growth factors such as GM-CSF may enhance IFN’s cytotoxicity. We performed a phase 2 study of IFN+GM-CSF in 58 newly diagnosed CML patients before imatinib approval. Short-term clinical responses included: 60% major cytogenetic response, 28% complete cytogenetic response and 19% complete molecular response. Six patients remain off all therapy for CML (range: 15 months–12 years) after IFN+GM-CSF treatment. IFN+GM-CSF shows promise as an adjunctive therapy for CML. PMID:25012565

  1. Routine Primary Prophylaxis for Febrile Neutropenia with Biosimilar Granulocyte Colony-Stimulating Factor (Nivestim) or Pegfilgrastim Is Cost Effective in Non-Hodgkin Lymphoma Patients undergoing Curative-Intent R-CHOP Chemotherapy

    PubMed Central

    Wang, Xiao Jun; Tang, Tiffany; Farid, Mohamad; Quek, Richard; Tao, Miriam; Lim, Soon Thye; Wee, Hwee Lin; Chan, Alexandre

    2016-01-01

    Objective This study aims to compare the cost-effectiveness of various strategies of myeloid growth factor prophylaxis for reducing the risk of febrile neutropenia (FN) in patients with non-Hodgkin lymphoma in Singapore who are undergoing R-CHOP chemotherapy with curative intent. Methods A Markov model was created to compare seven prophylaxis strategies: 1) primary prophylaxis (PP) with nivestim (biosimilar filgrastim) throughout all cycles of chemotherapy; 2) PP with nivestim during the first two cycles of chemotherapy; 3) secondary prophylaxis (SP) with nivestim; 4) PP with pegfilgrastim throughout all cycles of chemotherapy; 5) PP with pegfilgrastim during the first two cycles of chemotherapy; 6) SP with pegfilgrastim; and 7) no prophylaxis (NP). The perspective of a hospital was taken and cost-effectiveness was expressed as the cost per episode of FN avoided over six cycles of chemotherapy. A probabilistic sensitivity analysis was conducted. Results Strategies 3, 6, and 7 were dominated in the base case analysis by strategy 5. The costs associated with strategies 2, 5, 1, and 4 were US$3,813, US$4,056, US$4,545, and US$5,331, respectively. The incremental cost-effectiveness ratios for strategy 5 vs. strategy 2, strategy 1 vs. strategy 5, and strategy 4 vs. strategy 1 were US$13,532, US$22,565, and US$30,452, respectively, per episode of FN avoided. Strategy 2 has the highest probability to be cost-effective (ranged from 48% to 60%) when the willingness to pay (WTP) threshold is lower than US$10,000 per FN episode prevented. Conclusion In Singapore, routine PP with granulocyte colony-stimulating factor (nivestim or pegfilgrastim) is cost-effective for reducing the risk of FN in patients receiving R-CHOP. PMID:26871584

  2. Safety and efficacy of pegfilgrastim compared to granulocyte colony stimulating factor (G-CSF) supporting a dose-intensive, rapidly cycling anti-metabolite containing chemotherapy regimen (Hyper-CVAD) for lymphoid malignancy.

    PubMed

    Lane, Steven W; Crawford, Julie; Kenealy, Melita; Cull, Gavin; Seymour, John F; Prince, H Miles; Marlton, Paula; Gill, Devinder; Mollee, Peter N

    2006-09-01

    Pegfilgrastim (Neulasta) has proven efficacy as supportive therapy in a variety of 21-day chemotherapy regimens, but has not been studied in dose intensive, rapidly cycling regimens utilising cell-cycle active drugs (e.g. anti-metabolites) such as hyper-CVAD. This study examined whether pegfilgrastim was safe and lead to similar kinetics of neutrophil recovery as daily granulocyte colony stimulating factor (G-CSF). Using retrospective analysis, patients receiving pegfilgrastim (6 mg) were matched with controls (G-CSF 5 microg kg-1 per day) for a cycle of chemotherapy, prior chemotherapy, dose of cytarabine received, age (<60 or >60 years), diagnosis and bone marrow involvement. The primary endpoint was duration of grade IV neutropenia (absolute neutrophil count, ANC < 500 microl-1). Secondary endpoints included time to neutrophil recovery, incidence of febrile neutropenia, positive blood cultures and delay in subsequent chemotherapy. This study identified 124 pegfilgrastim supported cycles in 43 patients and successfully matched them to 124 G-CSF supported cycles from 38 patients treated between January 1999 and July 2005. There were no significant differences between pegfilgrastim and G-CSF groups in baseline or treatment-related variables. The median duration of grade IV neutropenia was 4 days in both groups (P = 0.55). Time to neutrophil recovery, incidence of febrile neutropenia, positive blood cultures and delay in subsequent chemotherapy were similar in both groups. Once per cycle dosing of pegfilgrastim appears safe and as effective as daily G-CSF for supporting the hyper-CVAD chemotherapy regimen. PMID:17064993

  3. 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

  4. Septoclast Deficiency Accompanies Postnatal Growth Plate Chondrodysplasia in the Toothless (tl) Osteopetrotic, Colony-Stimulating Factor-1 (CSF-1)-Deficient Rat and Is Partially Responsive to CSF-1 Injections

    PubMed Central

    Gartland, Alison; Mason-Savas, April; Yang, Meiheng; MacKay, Carole A.; Birnbaum, Mark J.; Odgren, Paul R.

    2009-01-01

    The septoclast is a specialized, cathepsin B-rich, perivascular cell type that accompanies invading capillaries on the metaphyseal side of the growth plate during endochondral bone growth. The putative role of septoclasts is to break down the terminal transverse septum of growth plate cartilage and permit capillaries to bud into the lower hypertrophic zone. This process fails in osteoclast-deficient, osteopetrotic animal models, resulting in a progressive growth plate dysplasia. The toothless rat is severely osteopetrotic because of a frameshift mutation in the colony-stimulating factor-1 (CSF-1) gene (Csf1tl). Whereas CSF-1 injections quickly restore endosteal osteoclast populations, they do not improve the chondrodysplasia. We therefore investigated septoclast populations in Csf1tl/Csf1tl rats and wild-type littermates, with and without CSF-1 treatment, at 2 weeks, before the dysplasia is pronounced, and at 4 weeks, by which time it is severe. Tibial sections were immunolabeled for cathepsin B and septoclasts were counted. Csf1tl/Csf1tl mutants had significant reductions in septoclasts at both times, although they were more pronounced at 4 weeks. CSF-1 injections increased counts in wild-type and mutant animals at both times, restoring mutants to normal levels at 2 weeks. In all of the mutants, septoclasts seemed misoriented and had abnormal ultrastructure. We conclude that CSF-1 promotes angiogenesis at the chondroosseous junction, but that, in Csf1tl/Csf1tl rats, septoclasts are unable to direct their degradative activity appropriately, implying a capillary guidance role for locally supplied CSF-1. PMID:19893052

  5. Granulocyte colony-stimulating factor and interleukin-1β are important cytokines in repair of the cirrhotic liver after bone marrow cell infusion: comparison of humans and model mice.

    PubMed

    Mizunaga, Yuko; Terai, Shuji; Yamamoto, Naoki; Uchida, Koichi; Yamasaki, Takahiro; Nishina, Hiroshi; Fujita, Yusuke; Shinoda, Koh; Hamamoto, Yoshihiko; Sakaida, Isao

    2012-01-01

    We previously described the effectiveness of autologous bone marrow cell infusion (ABMi) therapy for patients with liver cirrhosis (LC). We analyzed chronological changes in 19 serum cytokines as well as levels of specific cytokines in patients after ABMi therapy and in a mouse model of cirrhosis generated using green fluorescent protein (GFP)/carbon tetrachloride (CCl4). We measured expression profiles of cytokines in serum samples collected from 13 patients before and at 1 day and 1 week after ABMi. Child-Pugh scores significantly improved in all of these patients. To analyze the meaning of early cytokine change, we infused GFP-positive bone marrow cells (BMCs) into mice with CCl4-induced LC and obtained serum and tissue samples at 1 day and as well as at 1, 2, 3, and 4 weeks later. We compared chronological changes in serum cytokine expression in humans and in the model mice at 1 day and 1 week after BMC infusion. Among 19 cytokine, both granulocyte colony-stimulating factor (G-CSF) and interleukin-1β(IL-1β) in serum was found to show the same chronological change pattern between human and mice model. Next, we examined changes in cytokine expression in cirrhosis liver before and at 1, 2, 3, and 4 weeks after BMC infusion. Both G-CSF and IL-1β were undetectable in the liver tissues before and at 1 week after BMC infusion but increased at 2 weeks and continued until 4 weeks after infusion. The infused BMCs induced an early decrease of both G-CSF and IL-1β in serum and an increase in the model mice with LC. These dynamic cytokine changes might be important to repair liver cirrhosis after BMC infusion. PMID:22507241

  6. Molecular cloning of a second subunit of the receptor for human granulocyte - macrophage colony-stimulating factor (GM-CSF): Reconstitution of a high-affinity GM-CSF receptor

    SciTech Connect

    Hayashida, Kazuhiro; Kitamura, Toshio; Gorman, D.M.; Miyajima, Atsushi ); Arai, Kenichi; Yokota, Takashi )

    1990-12-01

    Using the mouse interleukin 3 (IL-3) receptor cDNA as a probe, the authors obtained a monologous cDNA (KH97) from a cDNA library of a human hemopoietic cell line, TF-1. The protein encoded by the KH97 cDNA has 56% amino acid sequence identity with the mouse IL-3 receptor and retains features common to the family of cytokine receptors. Fibroblasts transfected with the KH97 cDNA expressed a protein of 120 kDa but did not bind any human cytokines, including IL-3 and granulocyte - macrophage colony-stimulating factor (GM-CSF). Interestingly, cotransfection of cDNAs for KH97 and the low-affinity human GM-CSF receptor in fibroblasts resulted in formation of a high-affinity receptor for GM-CSF. The dissociation rate of GM-CSF from the reconstituted high-affinity receptor was slower than that from the low-affinity site, whereas the association rate was unchanged. Cross-linking of {sup 125}I-labeled GM-CSF to fibroblasts cotransfected with both cDNAs revealed the same cross-linking patterns as in TF-1 cells - i.e., two major proteins of 80 and 120 kDa which correspond to the low-affinity GM-CSF receptor and the KH97 protein, respectively. These results indicate that the high-affinity GM-CSF receptor is composed of at least two components in a manner analogous to the IL-2 receptor. They therefore propose to designate the low-affinity GM-CSF receptor and the KH97 protein as the {alpha} and {beta} subunits of the GM-CSF receptor, respectively.

  7. Living T9 glioma cells expressing membrane macrophage colony-stimulating factor produce immediate tumor destruction by polymorphonuclear leukocytes and macrophages via a "paraptosis"-induced pathway that promotes systemic immunity against intracranial T9 gliomas.

    PubMed

    Chen, Yijun; Douglass, Thomas; Jeffes, Edward W B; Xu, Qingcheng; Williams, Christopher C; Arpajirakul, Neary; Delgado, Christina; Kleinman, Michael; Sanchez, Ramon; Dan, Qinghong; Kim, Ronald C; Wepsic, H Terry; Jadus, Martin R

    2002-08-15

    Cloned T9-C2 glioma cells transfected with membrane macrophage colony-stimulating factor (mM-CSF) never formed subcutaneous tumors when implanted into Fischer rats, whereas control T9 cells did. The T9-C2 cells were completely killed within 1 day through a mechanism that resembled paraptosis. Vacuolization of the T9-C2 cell's mitochondria and endoplasmic reticulum started within 4 hours after implantation. By 24 hours, the dead tumor cells were swollen and terminal deoxynucleotide transferase-mediated dUTP nick-end labeling (TUNEL)-positive. Bcl2-transduced T9-C2 cells failed to form tumors in rats. Both T9 and T9-C2 cells produced cytokine-induced neutrophil chemoattractant that recruited the granulocytes into the tumor injection sites, where they interacted with the tumor cells. Freshly isolated macrophages killed the T9-C2 cells in vitro by a mechanism independent of phagocytosis. Nude athymic rats treated with antiasialo GM1 antibody formed T9-C2 tumors, whereas rats treated with a natural killer cell (NK)-specific antibody failed to form tumors. When treated with antipolymorphonuclear leukocyte (anti-PMN) and antimacrophage antibodies, 80% of nude rats formed tumors, whereas only 40% of the rats developed a tumor when a single antibody was used. This suggests that both PMNs and macrophages are involved in the killing of T9-C2 tumor cells. Immunocompetent rats that rejected the living T9-C2 cells were immune to the intracranial rechallenge with T9 cells. No vaccinating effect occurred if the T9-C2 cells were freeze-thawed, x-irradiated, or treated with mitomycin-C prior to injection. Optimal tumor immunization using mM-CSF-transduced T9 cells requires viable tumor cells. In this study optimal tumor immunization occurred when a strong inflammatory response at the injection of the tumor cells was induced. PMID:12149220

  8. 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.

  9. Monoclonal antibody against macrophage colony-stimulating factor suppresses circulating monocytes and tissue macrophage function but does not alter cell infiltration/activation in cutaneous lesions or clinical outcomes in patients with cutaneous lupus erythematosus.

    PubMed

    Masek-Hammerman, K; Peeva, E; Ahmad, A; Menon, S; Afsharvand, M; Peng Qu, R; Cheng, J B; Syed, J; Zhan, Y; O'Neil, S P; Pleasic-Williams, S; Cox, L A; Beidler, D

    2016-02-01

    This study's objective was to assess the effects of PD-0360324, a fully human immunoglobulin G2 monoclonal antibody against macrophage colony-stimulating factor in cutaneous lupus erythematosus (CLE). Patients with active subacute CLE or discoid lupus erythematosus were randomized to receive 100 or 150 mg PD-0360324 or placebo via intravenous infusion every 2 weeks for 3 months. Blood and urine samples were obtained pre- and post-treatment to analyse pharmacokinetics and pharmacodynamic changes in CD14(+) CD16(+) monocytes, urinary N-terminal telopeptide (uNTX), alanine/aspartate aminotransferases (ALT/AST) and creatine kinase (CK); tissue biopsy samples were taken to evaluate macrophage populations and T cells using immunohistochemistry. Clinical efficacy assessments included the Cutaneous Lupus Erythematosus Disease Area and Severity Index (CLASI). Among 28 randomized/analysed patients, peak/trough plasma concentrations increased in a greater-than-dose-proportional manner with dose increases from 100 to 150 mg. Statistically significant differences were observed between active treatment and placebo groups in changes from baseline in CD14(+) CD16(+) cells, uNTX, ALT, AST and CK levels at most time-points. The numbers, density and activation states of tissue macrophages and T cells did not change from baseline to treatment end. No between-group differences were seen in CLASI. Patients receiving PD-0360324 reported significantly more adverse events than those receiving placebo, but no serious adverse events. In patients with CLE, 100 and 150 mg PD-0360324 every 2 weeks for 3 months suppressed a subset of circulating monocytes and altered activity of some tissue macrophages without affecting cell populations in CLE skin lesions or improving clinical end-points. PMID:26376111

  10. Interleukin-3, but not granulocyte-macrophage colony-stimulating factor and interleukin-5, inhibits apoptosis of human basophils through phosphatidylinositol 3-kinase: requirement of NF-kappaB-dependent and -independent pathways.

    PubMed

    Zheng, Xueyan; Karsan, Aly; Duronio, Vincent; Chu, Fanny; Walker, David C; Bai, Tony R; Schellenberg, R Robert

    2002-11-01

    Basophils are key effector cells of allergic reactions. Although proinflammatory cytokines, such as interleukin (IL)-3, granulocyte-macrophage colony-stimulating factor (GM-CSF) and IL-5, inhibit eosinophil apoptosis in vitro, little is known about basophil apoptosis, and the signalling mechanisms required for basophil survival remain undefined. To address this issue, we used a novel negative-selection system to isolate human basophils to a purity of > 95%, and evaluated apoptosis by morphology using light and transmission electron microscopy, and by annexin-V binding and propidium iodide incorporation using flow cytometry. In this study, we demonstrated that the spontaneous rate of apoptotic basophils was higher than that of eosinophils as, at 24 hr, 57.6 +/- 4.7% of basophils underwent apoptosis compared with 39.5 +/- 3.8% of eosinophils. In addition, basophil cell death was significantly inhibited when cultured with IL-3 for 48 hr (84.6 +/- 4.9% vehicle-treated cells versus 40.9 +/- 3.9% IL-3-treated cells). IL-3 also up-regulated basophil CD69 surface expression. The effects of IL-3 on apoptosis and CD69 surface expression of human basophils were completely blocked by LY294002 (LY), a potent inhibitor of phosphatidylinositol 3-kinase (PI3-K), but only partially inhibited by lactacystin, a proteasome inhibitor that prevents degradation of IkappaB and NF-kappaB translocation. These observations reveal the novel finding that IL-3 prevents basophil apoptosis through the activation of PI3-K, which is only partially NF-kappaB dependent. As basophils are active participants in allergic reactions and IL-3 is one of the abundant proinflammatory cytokines in secretions from allergic tissue, we suggest that IL-3-mediated inhibition of basophil apoptosis may exacerbate the inflammation associated with allergic disorders. PMID:12423306

  11. 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.

  12. DNA-Protein Immunization Using Leishmania Peroxidoxin-1 Induces a Strong CD4+ T Cell Response and Partially Protects Mice from Cutaneous Leishmaniasis: Role of Fusion Murine Granulocyte-Macrophage Colony-Stimulating Factor DNA Adjuvant

    PubMed Central

    Bayih, Abebe Genetu; Daifalla, Nada S.; Gedamu, Lashitew

    2014-01-01

    Background To date, no universally effective and safe vaccine has been developed for general human use. Leishmania donovani Peroxidoxin-1 (LdPxn-1) is a member of the antioxidant family of proteins and is predominantly expressed in the amastigote stage of the parasite. The aim of this study was to evaluate the immunogenicity and protective efficacy of LdPxn-1 in BALB/c mice in heterologous DNA-Protein immunization regimen in the presence of fusion murine granulocyte-macrophage colony-stimulating factor (mGMCSF) DNA adjuvant. Methodology and Principal Findings A fusion DNA of LdPxn1 and mGMCSF was cloned into a modified pcDNA vector. To confirm the expression in mammalian system, Chinese hamster ovary cells were transfected with the plasmid vector containing LdPxn1 gene. BALB/c mice were immunized twice with pcDNA-mGMCSF-LdPxn-1 or pcDNA-LdPxn1 DNA and boosted once with recombinant LdPxn-1 protein. Three weeks after the last immunization, mice were infected with Leishmania major promastigotes. The result showed that immunization with pcDNA-mGMCSF-LdPxn1 elicited a mixed Th-1/Th-2 immune response with significantly higher production of IFN-γ than controls. Intracellular cytokine staining of antigen-stimulated spleen cells showed that immunization with this antigen elicited significantly higher proportion of CD4+ T cells that express IFN-γ, TNF-α, or IL-2. The antigen also induced significantly higher proportion of multipotent CD4+ cells that simultaneously express the three Th-1 cytokines. Moreover, a significant reduction in the footpad swelling was seen in mice immunized with pcDNA-mGMCSF-LdPxn1 antigen. Expression study in CHO cells demonstrated that pcDNA-mGMCSF-LdPxn-1 was expressed in mammalian system. Conclusion The result demonstrates that immunization of BALB/c mice with a plasmid expressing LdPxn1 in the presence of mGMCSF adjuvant elicits a strong specific immune response with high level induction of multipotent CD4+ cells that mediate protection of the

  13. 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

  14. 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. PMID:24644014

  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. Phase I study of simultaneous dose escalation and schedule acceleration of cyclophosphamide-doxorubicin-etoposide using granulocyte colony-stimulating factor with or without antimicrobial prophylaxis in patients with small-cell lung cancer.

    PubMed Central

    Ardizzoni, A.; Pennucci, M. C.; Danova, M.; Viscoli, C.; Mariani, G. L.; Giorgi, G.; Venturini, M.; Mereu, C.; Scolaro, T.; Rosso, R.

    1996-01-01

    A phase I study was designed to assess whether dose intensity of an 'accelerated' cyclophosphamide-doxorubicin-etoposide (CDE) regimen plus granulocyte colony-stimulating factor (G-CSF) could be increased further, in an outpatient setting, by escalating the dose of each single drug of the regimen. Patients with previously untreated small-cell lung cancer (SCLC) received escalating doses of cyclophosphamide (C) 1100-1300 mg m-2 intravenously (i.v.) on day 1, doxorubicin (D) 50-60 mg m-2 i.v. on day 1, etoposide (E) 110-130 mg m-2 i.v. on days 1, 2, 3 and every 14 days for at least three courses. Along with chemotherapy, G-CSF (filgastrim) 5 micrograms kg-1 from day 5 to day 11 was administered subcutaneously (s.c.) to all patients. Twenty-five patients were enrolled into the study. All patients at the first dose level (C 1100, D 50, E 110 x 3) completed three or more cycles at the dose and schedule planned by the protocol and no 'dose-limiting toxicity' (DLT) was seen. At the second dose level (C 1200, D 55, E 120 x 3) three out of five patients had a DLT consisting of 'granulocytopenic fever' (GCPF). Another six patients were treated at this dose level with the addition of ciprofloxacin 500 mg twice a day and only two patients had a DLT [one episode of documented oral candidiasis and one of 'fever of unknown origin' (FUO) with generalised mucositis]. Accrual of patients proceeded to the third dose level (C 1300, D 60, E 130 x 3) with the prophylactic use of ciprofloxacin. Four out of six patients experienced a DLT consisting of GCPF or documented non-bacterial infection. Accrual of patients at the third dose level was then resumed adding to ciprofloxacin anti-fungal prophylaxis (fluconazole 100 mg daily) and anti-viral prophylaxis (acyclovir 800 mg twice a day) from day 5 to 11. Out of five patients treated three experienced a DLT consisting of severe leucopenia and fever or infection. With a simultaneous dose escalation and schedule acceleration it is indeed

  17. 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

  18. 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

  19. Monoclonal Lym-1 antibody-dependent cytolysis by neutrophils exposed to granulocyte-macrophage colony-stimulating factor: intervention of FcgammaRII (CD32), CD11b-CD18 integrins, and CD66b glycoproteins.

    PubMed

    Ottonello, L; Epstein, A L; Dapino, P; Barbera, P; Morone, P; Dallegri, F

    1999-05-15

    Murine monoclonal antibody (MoAb) Lym-1 is an IgG2a able to bind HLA-DR variants on malignant B cells and suitable for serotherapeutic approaches in B-lymphoma patients. We have previously shown that Lym-1 can synergize with granulocyte-macrophage colony-stimulating factor (GM-CSF) to trigger neutrophil cytolysis towards Raji cells used as a model of B-lymphoma targets. Here we provide evidence for the intervention of certain neutrophil receptors or surface molecules in this model of cell-mediated lysis. The lysis was completely inhibited by the anti-FcgammaRII MoAb IV.3 and unaffected by the anti-FcgammaRIII MoAb 3G8. This suggests that neutrophil cytolysis involves FcgammaRII without cooperation of this receptor with FcgammaRIII. Moreover, the lysis was inhibited by an anti-CD18 MoAb (MEM48) and by a MoAb specific for carcinoembryonic antigen (CEA)-like and glycophosphatidyl inositol (GPI)-linked glycoproteins (CD66b). Using an immunofluorescence staining procedure, cross-linking of CD66b induced the redistribution of CD11b on neutrophils with distinct areas of CD11b clustering via a process susceptible of inhibition by D-mannose. This is consistent with the ability of CD11b-CD18 and CD66b to undergo lectin-like physical interactions on the neutrophil surface. Such a type of interaction is presumably instrumental for neutrophil cytolytic activity in that the lysis was inhibited by D-mannose and enhanced by the MoAb VIM-12, which mimics the cooperation between CD11b and GPI-anchored molecules by specifically interacting with CD11b lectin-like sites. Therefore, the present results prove the absolute requirement for FcgammaRII in neutrophil GM-CSF/Lym-1-mediated cytolysis and, on the other hand, define the crucial role of CD66b and CD11b/CD18 in the expression of the cell lytic potential. PMID:10233903

  20. Laevofolinic acid, 5-fluorouracil, cyclophosphamide and escalating doses of epirubicin with granulocyte colony-stimulating factor support in locally advanced and/or metastatic breast carcinoma: a phase I-II study of the Southern Italy Oncology Group (GOIM).

    PubMed Central

    Colucci, G.; Romito, S.; Gebbia, V.; Pacilio, G.; Giotta, F.; Testa, A.; Pezzella, G.; Durini, E.; Agostara, B.; Cariello, S.

    1995-01-01

    Sixty-four consecutive patients with locally advanced (n = 7) or metastatic breast cancer (n = 57), were treated with a combination of laevofolinic acid 100 mg m-2 plus 5-fluorouracil 340 mg m-2 i.v. on days 1-3, cyclophosphamide 600 mg m-2 i.v. on day 1 and epirubicin 90 mg m-2 i.v. on day 1. Epirubicin dose was progressively escalated by 10 mg m-2 per cycle up to 120 mg m-2 in the absence of dose-limiting toxicities. Granulocyte colony-stimulating factor (G-CSF) was given subcutaneously in order to prevent neutropenia. Epirubicin dosage could be increased to 100 mg m-2 in 53 patients (87%), to 110 mg m-2 in 31 patients (51%) and to 120 mg m-2 in 18 cases (30%). In most patients the dose-limiting toxicity was represented by myelosuppression. A statistically significant correlation was found between median white blood count (WBC) or absolute neutrophil count (ANC) nadir and epirubicin dose level (P = 0.009; P = 0.008). Moreover, a statistically significant correlation was observed between the number of chemotherapeutic cycles, nadir ANC and WBC and the occurrence of anaemia and thrombocytopenia of increasing severity. These data suggest the occurrence of progressive cumulative bone marrow toxicity. Although patients who reached different epirubicin levels showed differences in mean dose intensity, such differences were not statistically significant. No correlation was found between the increase in dose intensity and type, rate or duration of objective responses. In patients with metastatic breast cancer the overall response rate was 72% (95% CL 66-78%) with a 25% complete response rate. Median duration of response was 10 and 13 months respectively for complete and partial responses. All patients with locally advanced breast cancer had an objective response and underwent radical mastectomy. Projected median survival of the whole series of patients with metastatic breast cancer was 20 + months. These data demonstrate that the combination of 5-fluorouracil with

  1. Mobilization of peripheral blood stem cells with chemotherapy and recombinant human granulocyte colony-stimulating factor (rhG-CSF): a randomized evaluation of different doses of rhG-CSF.

    PubMed

    Demirer, T; Ayli, M; Ozcan, M; Gunel, N; Haznedar, R; Dagli, M; Fen, T; Genc, Y; Dincer, S; Arslan, O; Gürman, G; Demirer, S; Ozet, G; Uysal, A; Konuk, N; Ilhan, O; Koc, H; Akan, H

    2002-02-01

    To date, no randomized study has compared different doses of recombinant human granulocyte colony-stimulating factor (rhG-CSF) following submyeloablative mobilization chemotherapy. Therefore, we evaluated the effect of different doses of rhG-CSF following mobilization chemotherapy on yields of CD34+ peripheral blood stem cells (PBSC). Fifty patients were randomized to receive 8 (n = 25) versus 16 microg/kg/d (n = 25) of rhG-CSF following mobilization chemotherapy. The median number of CD34+ cells collected after 8 microg/kg/d of rhG-CSF was 2.36 x 10(6)/kg (range, 0.21-7.80), compared with 7.99 (2.76-14.89) after 16 microg/kg/d (P < 0.001). Twenty out of 25 (80%) patients in the low-dose and 23 out of 25 (92%) in the high-dose rhG-CSF arm underwent high-dose chemotherapy (HDC) and autologous stem cell transplantation (ASCT). Median days to white blood cell engraftment in patients mobilized with 8 microg/kg and 16 microg/kg of rhG-CSF were 12 (10-20) and 9 (8-11) respectively (P < 0.001). There was no difference between the two groups regarding the other parameters of peritransplant morbidity: days to platelet engraftment (P = 0.10), number of red blood cell (P = 0.56) and platelet transfusions (P = 0.22), days of total parenteral nutrition requirement (P = 0.84), fever (P = 0.93) and antibiotics (P = 0.77), and number of different antibiotics used (P = 0.58). These data showed that higher doses of rhG-CSF following submyeloablative mobilization chemotherapy were associated with a clear dose-response effect based on the collected cell yields. Based on the parameters of peritransplant morbidity, 8 microg/kg/d was as effective as 16 microg/kg/d except for a rapid neutrophil engraftment in the high-dose arm. Therefore, in routine clinical practice, despite some advantage in the use of higher doses of rhG-CSF, lower doses may be used for PBSC collections following chemotherapy-based mobilization regimens in this cost-conscious era. PMID:11841454

  2. 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

  3. Involvement of the high-affinity receptor for IgG (Fc gamma RI; CD64) in enhanced tumor cell cytotoxicity of neutrophils during granulocyte colony-stimulating factor therapy.

    PubMed

    Valerius, T; Repp, R; de Wit, T P; Berthold, S; Platzer, E; Kalden, J R; Gramatzki, M; van de Winkel, J G

    1993-08-01

    Three different classes of Fc receptors for IgG (Fc gamma R) are currently distinguished in humans, of which polymorphonuclear phagocytes (PMN) normally express both low-affinity receptor classes--Fc gamma RII (CD32) and Fc gamma RIII (CD16). During therapy with granulocyte colony-stimulating factor (G-CSF), neutrophils from patients with various malignancies and different hematologic disorders were found to additionally express high levels of the receptor with high affinity for IgG (Fc gamma RI; CD64). For these patients, the relative fluorescence intensity (rFI) for Fc gamma RI was 5.3 (range, 1.7 to 10.3; n = 19), compared with 1.0 (range, 1.0 to 1.1; n = 8) for healthy donors. The expression of Fc gamma RI during G-CSF therapy could be confirmed by using a panel of six CD64-specific antibodies, and by showing mRNA for Fc gamma RI. So far, three genes for Fc gamma RI have been identified, encoding four distinct transcription products. By reverse transcriptase-polymerase chain reaction technology, transcripts for both membrane-associated isoforms (hFc gamma RIa and hFc gamma RIb2) could be detected. The functional activity of Fc gamma RI on PMN during G-CSF therapy was shown by measuring binding of monomeric human IgG and antibody-dependent cellular cytotoxicity (ADCC). Thus, Fc gamma RI-positive neutrophils displayed enhanced ADCC activity to glioma (A1207), squamous cell (A431), and ovarian (SK-ov3) carcinoma cell lines. The involvement of Fc gamma RI in this increased cytotoxic activity was shown by blocking Fc gamma receptors with monoclonal antibodies, and by using F(ab')2 x F(ab')2-bispecific antibodies with specificities against tumor-related antigens and Fc gamma RI, resulting in solely Fc gamma RI-mediated cytotoxicity. Therapeutically, this additional Fc receptor on PMN may increase the efficacy of experimental antibody therapy. PMID:7687898

  4. Scatter factor influences the formation of prostate epithelial cell colonies on bone marrow stroma in vitro.

    PubMed

    Lang, S H; Clarke, N W; George, N J; Testa, N G

    1999-06-01

    Prostate cancer metastases form selectively in the bone marrow. Previously we demonstrated motility was important for the formation of primary prostatic epithelial cell colonies in bone marrow stroma (BMS) co-culture. In this study we looked at the influence of motility factors on the colony formation of epithelial cells derived from benign (bPEC) or malignant (mPEC) prostate tissue. After 7 days co-culture we found that anti-scatter factor consistently inhibited prostate epithelial cell colony formation on BMS (7/7 mPEC and 4/7 bPEC samples showed significant inhibition). Antibodies against bFGF and 5T4 did not significantly affect colony formation. Addition of fibroblast conditioned media (derived from benign prostates) to co-cultures stimulated the colony formation of bPEC (170%) and mPEC (252%). This stimulation was eliminated by depletion of SF from the conditioned media. Immunohistochemical staining found c-Met expression in 5/6 bPEC cultures and 7/9 mPEC cultures. When grown in BMS co-culture expression of c-Met was positive in 3/6 bPEC and 2/7 mPEC samples. In conclusion, scatter factor influences the in vitro formation of prostate epithelial cell colonies on BMS co-culture. PMID:10545020

  5. Production of granulocyte/macrophage-colony-stimulating factor by human natural killer cells. Modulation by the p75 subunit of the interleukin 2 receptor and by the CD2 receptor.

    PubMed

    Levitt, L J; Nagler, A; Lee, F; Abrams, J; Shatsky, M; Thompson, D

    1991-07-01

    Resting natural killer (NK) cells express the p75 chain of the IL-2 receptor (IL-2R beta) and most NK cells express the CD2 (erythrocyte rosette) receptor. The cell adhesion molecule, LFA-3, is a natural co-ligand for CD2. Tac antigen (IL-2R alpha), a p55 IL-2R subunit, can be expressed after NK activation and may play a role in IL-2-induced NK proliferation. Little is known of the molecular mechanisms underlying cytokine production in NK cells. We investigated the roles of IL-2R alpha, IL-2R beta, and CD2/LFA-3 in the molecular regulation of NK cell granulocyte/macrophage-colony-stimulating factor (GM-CSF) production. Enriched populations of peripheral blood NK cells were separated into CD16-positive and CD16-negative fractions by flow cytometry; positively selected cells were greater than 97% positive for CD16 (the FcIII receptor for IgG which is present on almost all NK cells), less than 1% positive for the T cell antigen CD3, and did not demonstrate rearrangement of the T cell receptor beta chain gene by Southern blot. NK cell supernatants were harvested after 3-4 d of incubation with 0-100 U/ml IL-2, or after incubation with anti-CD2 (T11(3] MAb and sheep red blood cells (SRBC are a homologue for LFA-3). Parallel cell aliquots were harvested at 3-16 h for transcriptional run-on assays, S1 nuclease assays, and actinomycin D mRNA t1/2 determinations. IL-2-activated NK supernatants contained large amounts of GM-CSF (178 +/- 35 pg/ml) by ELISA as did supernatants from CD2-activated NK cells (T11(3) MAb + SRBC: 212 +/- 42) vs. less than 20 pg/ml for NK cells incubated alone or with either SRBC or T11(3) MAb alone. Sepharose-linked anti-CD3 MAb did not induce GM-CSF release from NK cells. By S1 analysis, both IL-2 and CD2 stimulation markedly augmented GM-CSF mRNA expression but with very different latencies of onset. IL-2R beta MAb inhibited greater than 85% of GM-CSF release from IL-2-activated NK cells and markedly suppressed IL-2-induced GM-CSF mRNA expression

  6. Tumor necrosis factor induced stimulation of granulopoiesis and radioprotection.

    PubMed

    Urbaschek, R; Männel, D N; Urbaschek, B

    1987-01-01

    Human recombinant tumor necrosis factor, TNF, was used to assess its ability to stimulate granulopoiesis and to protect mice against lethal irradiation, effects known to be inducable with TNF-rich postendotoxin serum from BCG infected mice (BCG/ET serum). Although the endotoxin contamination of this TNF preparation is extremely low its effects were compared in endotoxin low responder C3H/HeJ mice and susceptible NMRI mice. TNF is a potent inducer of serum colony stimulating activity, CSA, in both mouse strains. In peripheral blood a marked granulocytosis with a concomitant decrease in lymphocytes and monocytopenia occurs at 2 hours after injection of TNF. Moreover, TNF induces an increase in the number of splenic myelopoietic committed stem cells (GM-CFC, granulocyte-macrophage colony forming cells) determined five days after injection. The lethality rate, registered over 30 days after exposure to 660 cGy whole body X-irradiation is reduced to 40% in C3H/HeJ mice as compared to 75% in control animals. The reduction in lethality is observed both, when TNF was injected 24 hours before or after irradiation. In vitro, TNF significantly increases the number of colonies in the presence of CSA in bone marrow cultures. TNF per se does not effect colony growth. The studies reported here demonstrate that TNF is a myelopoiesis stimulating factor in mice which may be related to the reduction in lethality following whole body irradiation. PMID:3306175

  7. Prognostic factors for re-mobilization using plerixafor and granulocyte colony-stimulating factor (G-CSF) in patients with malignant lymphoma or multiple myeloma previously failing mobilization with G-CSF with or without chemotherapy: the Korean multicenter retrospective study.

    PubMed

    Kim, Jin Seok; Yoon, Dok Hyun; Park, Seonyang; Yoon, Sung-Soo; Cho, Seok-Goo; Min, Chang-Ki; Lee, Je-Jung; Yang, Deok-Hwan; Kwak, Jae-Yong; Eom, Hyeon-Seok; Kim, Won Seog; Kim, Hawk; Do, Young Rok; Moon, Joon Ho; Lee, Jihye; Suh, Cheolwon

    2016-03-01

    Plerixafor in combination with granulocyte colony-stimulating factor (G-CSF) has been shown to improve the rates of successful peripheral blood stem cell (PBSC) mobilization in patients with malignant lymphoma or multiple myeloma (MM) who experienced prior failure of PBSC mobilization. We evaluated the mobilization results of re-mobilization using plerixafor and G-CSF in insufficiently mobilizing patients. Forty-four patients with lymphoma (n = 29) or MM (n = 15) were included in the study. The median age was 50 (range, 24-64) years. Previous mobilization regimens were chemotherapy with G-CSF (n = 28), including cyclophosphamide with G-CSF (n = 15), and G-CSF only (n = 16). All patients with lymphoma achieved at least partial response (PR) before the mobilization, including 13 complete responses (CRs). Eleven patients with MM achieved at least PR and four patients with MM were in stable disease before mobilization. The median number of apheresis was 3 (range, 1-6). The median yield of PBSC collections was 3.41 (0.13-38.11) × 10(6) CD34(+) cells/kg. Thirty-four (77.3 %) patients had successful collections defined as at least 2 × 10(6) CD34(+) cells/kg. The rate of successful collections was not different between the two underlying diseases (79.3 % in lymphoma and 73.3 % in MM). Of the entire cohort, 38 (86.4 %) of patients went on to receive an autologous transplant. Previous long-term use of high-risk drugs (>4 cycles use of alkylating agents, platinum-containing agents, or thalidomide) (HR 10.8, 95 % CI 1.1-110.0, P = 0.043) and low platelet count (<100 × 10(9)/L) 1 day before the first apheresis (HR 27.9, 95 % CI 2.9-273.7, P = 0.004) were independent prognostic factors for predicting failure of PBSC re-mobilization using plerixafor and G-CSF. In conclusion, re-mobilization using plerixafor and G-CSF showed a success rate of 77.3 % in patients with lymphoma or MM who experienced prior failure of PBSC

  8. Human U251MG glioma cells expressing the membrane form of macrophage colony-stimulating factor (mM-CSF) are killed by human monocytes in vitro and are rejected within immunodeficient mice via paraptosis that is associated with increased expression of three different heat shock proteins.

    PubMed

    Jadus, Martin R; Chen, Yijun; Boldaji, Mehrdokht Tarbiyat; Delgado, Christina; Sanchez, Ramon; Douglass, Thomas; Al-Atar, Usama; Schulz, William; Lloyd, Cheri; Wepsic, H Terry

    2003-05-01

    Human U251MG glioma cells retrovirally transduced with the human gene for the membrane form of macrophage colony-stimulating factor (mM-CSF) were investigated. The clones, MG-2F11 and MG-2C4, that expressed the most mM-CSF, but not the viral vector or the parental U251MG cells, were killed by both murine and human monocyte/macrophages in cytotoxicity assays. MG-2F11 cells failed to form subcutaneous tumors in either nude or NIH-bg-nu-xidBR mice, while mice inoculated with the U251MG viral vector (MG-VV) cells developed tumors. Electron microscopy studies showed that 4 hours after subcutaneous injection, the mM-CSF-transduced cells began dying of a process that resembled paraptosis. The dying tumor cells were swollen and had extensive vacuolization of their mitochondria and endoplasm reticulum. This killing process was complete within 24 hours. Macrophage-like cells were immediately adjacent to the killed MG-2F11 cells. Immunohistological staining for the heat shock proteins HSP60, HSP70 and GRP94 (gp96) showed that 18 hours after inoculation into nude mice, the MG-2F11 injection site was two to four times more intensely stained than the MG-VV cells. This study shows that human gliomas transduced with mM-CSF have the potential to be used as a safe live tumor cell vaccine. PMID:12719711

  9. Antilymphocyte globulin, cyclosporine, prednisolone, and granulocyte colony-stimulating factor for severe aplastic anemia: an update of the GITMO/EBMT study on 100 patients. European Group for Blood and Marrow Transplantation (EBMT) Working Party on Severe Aplastic Anemia and the Gruppo Italiano Trapianti di Midolio Osseo (GITMO).

    PubMed

    Bacigalupo, A; Bruno, B; Saracco, P; Di Bona, E; Locasciulli, A; Locatelli, F; Gabbas, A; Dufour, C; Arcese, W; Testi, G; Broccia, G; Carotenuto, M; Coser, P; Barbui, T; Leoni, P; Ferster, A

    2000-03-15

    One hundred consecutive patients with severe aplastic anemia (SAA) received horse antilymphocyte globulin (ALG), cyclosporin A (CyA), 6-methylprednisolone (6Mpred), and granulocyte colony-stimulating factor (G-CSF) as first-line therapy. The median age was 16 years (range, 1-72 years) and median neutrophil count was 0.2 x 10(9)/L (range, 0-0.5 x 10(9)/L). Trilineage hematologic recovery (at a median interval of 96 days from treatment) was seen in 77 patients (48 complete, 29 partial) after 1 (n = 50) or more courses of ALG (n = 27). Of the 23 nonresponders, 11 patients died at a median interval of 83 days (range, 16-1132 days), 6 were considered treatment failures and underwent transplantation, and 6 were pancytopenic. Cytogenetic abnormalities were seen in 11% of patients, clonal hematologic disease in 8%, and relapse of marrow aplasia in 9%. The actuarial survival at 5 years was 87% (median follow-up 1424 days): 76% versus 98% for patients with neutrophil counts less than versus greater than 0.2 x 10(9)/L (P =.001) and 88% versus 87% for patients aged less than versus more than 16 years (P =.8). The actuarial probability of discontinuing CyA was 38%. Patients who did not achieve a white blood cell (WBC) count of 5 x 10(9)/L during G-CSF treatment have a low probability of responding (37%) and a high mortality rate (42%). This update confirms a high probability for SAA patients of becoming transfusion independent and of surviving after treatment with ALG, CyA, 6Mpred, and G-CSF, with a significant effect of neutrophil counts on outcome. Problems still remain, such as absent or incomplete responses, clonal evolution, relapse of the original disease, and cyclosporine dependence. Early transplantation, also from alternative donors, may be warranted in patients with poor WBC response to G-CSF. (Blood. 2000;95:1931-1934) PMID:10706857

  10. Inadvertent Propagation of Factor VII Deficiency in a Canine Mucopolysaccharidosis Type I Research Breeding Colony

    PubMed Central

    Carlstrom, Lucas P; Jens, Jackie K; Dobyns, Marley E; Passage, Merry; Dickson, Patricia I; Ellinwood, N Matthew

    2009-01-01

    Issues of cost and genetics can result in inbreeding of canine genetic disease colonies. Beagles often are used to maintain such colonies, providing stock for outcrosses. Factor VII (FVII) deficiency is a hemostatic disorder found at increased frequency in beagles and has been characterized at the DNA level. Deficiency of FVII presents obstacles in colonies founded with beagles. An initial finding of a FVII-deficient pup from a longstanding colony prompted us to evaluate FVII deficiency fully in this colony. Current and archival records and tissues were used to reconstruct the colony pedigree, assess the contribution from beagles, and test samples to document the source and frequency of the mutant FVII allele. As part of this study we developed a PCR-based diagnostic assay that was simpler than what was previously available. Pedigree analysis revealed a founder effect implicating beagles that led to high frequency (55%) of the mutant allele. In addition, affected animals were identified. The complete picture of the clinical effect within the colony remains unclear, but unusual neonatal presentations, including hemoabdomen, have occurred in pups affected with FVII deficiency. Use of a PCR-based diagnostic assay to screen all potential beagle breeding stock will prevent similar occurrences of FVII deficiency in future canine research colonies. PMID:19712579

  11. Effects of granulocyte-macrophage colony-stimulating factor (GM-CSF), IL-2, interferon-gamma (IFN-gamma), tumour necrosis factor-alpha (TNF-alpha) and IL-6 on the production of immunoreactive IL-1 and TNF-alpha by human monocytes.

    PubMed Central

    Danis, V A; Franic, G M; Rathjen, D A; Brooks, P M

    1991-01-01

    The effects of GM-CSF, IL-2, IFN-gamma, TNF-alpha and IL-6 on the production of IL-1 (both secreted and cell associated) and TNF-alpha by peripheral blood monocytes were studied. Monocytes were cultured for 20 h in suspension and in serum-free conditions which minimized background stimulation of monokine production. GM-CSF, IL-2 and TNF-alpha directly induced the production of cell-associated IL-1 but little or no IL-1 or TNF-alpha secretion. Combination of GM-CSF with IFN-gamma, IL-2 or TNF-alpha synergistically enhanced IL-1 secretion and had an additive effect on cell-associated IL-1 production. Combination of IL-2 with IFN-gamma or TNF-alpha also synergistically enhanced IL-1 secretion but the effect on cell-associated IL-1 production was less than additive. GM-CSF synergistically enhanced TNF-alpha secretion induced by IFN-gamma but not by lipopolysaccharide. GM-CSF did not enhance TNF-alpha secretion induced by IL-2 or TNF-alpha. In contrast, IL-2 synergistically enhanced TNF-alpha secretion induced by IFN-gamma. These results are discussed in relation to cytokine involvement in rheumatoid arthritis. PMID:1906383

  12. Inhibition of CD26 enzyme activity with pro-boropro stimulates rat granulocyte/macrophage colony formation and thymocyte proliferation in vitro.

    PubMed

    Bristol, L A; Bachovchin, W; Takács, L

    1995-06-15

    CD26 dipeptidyl peptidase (DPPIV) is involved in the regulation of proliferation of some hematopoietic and T-lineage cells. Here, we show that Pro-boropro a potent inhibitor of DPP activity has a costimulating effect in hematopoietic colony assays for macrophage and, to a lesser extent, for granulocyte colonies and has a stimulating effect in organ cultures of immature thymocytes. Based on these and other evidences, we propose that the mechanism by which CD26 regulates proliferation is associated with its DPP activity. PMID:7780144

  13. Multiple-factor influences upon feeding flight rates at wading bird colonies (Alias: Are flight-line counts useful?)

    USGS Publications Warehouse

    Erwin, R.M.; Ogden, J.C.

    1979-01-01

    The temporal patterns of feeding, resting, and reproductive behavior in colonial wading birds have been studied by a number of investigators, both on a short-term (daily) and long-term (annual) basis. In coastal marine environments, activities at colonies are influenced by tides, time of day and phase of the nesting cycle. The purpose of this paper is twofold: (1) to examine the effects of tide, time of day (physical factors), nesting phase, colony site, and species identity (biological factors) on feeding flight rates at breeding colonies and, as a result of this, (2) to evaluate the usefulness of feeding flight counts as an index of the number of nests in the colony. Earlier work suggests that the relationship between the number of individuals flying to and from the nesting colony may be quite consistent with nest numbers. Thus, by monitoring flights from remote locations, observers might obtain relatively accurate census data while minimizing time and disturbance at colonies. Recent concern for the deleterious impact of humans at waterbird colonies underscores the need to investigate alternative census methods.

  14. Pre-B Cell Colony Enhancing Factor (PBEF), a Cytokine with Multiple Physiological Functions

    PubMed Central

    Sun, Zhongjie; Lei, Han; Zhang, Zhonge

    2013-01-01

    Pre-B cell colony enhancing factor (PBEF) is regarded as a proinflammatory cytokine. Named for its first discovered function as a pre-B cell colony enhancing factor, it has since been found to have many other functions relating to cell metabolism, inflammation, and immune modulation. It has also been found to have intracellular and extracellular forms, with the two overlapping in function. Most of the intracellular functions of PBEF are due to its role as a nicotinamide phosphoribosyltransferase (Nampt). It has been found in human endothelial cells, where it is able to induce angiogenesis through upregulation of VEGF and VEGFR and secretion of MCP-1. In human umbilical endothelial cells, PBEF increases levels of the protease MMP 2/9. PBEF has also been found in a variety of immune cells other than B cells and has been shown to inhibit apoptosis of macrophages. Extracellular PBEF has been shown to increase inflammatory cytokines, such as TNF-α, IL-1β, IL-16, and TGF-β1, and the chemokine receptor CCR3. PBEF also increases the production of IL-6, TNF-α, and IL-1β in CD14+ monocyctes, macrophages, and dendritic cells, enhances the effectiveness of T cells, and is vital to the development of both B and T lymphocytes. The purpose of this review is to summarize the recent advances in PBEF research. PMID:23787158

  15. The Importance of Microbes in Nutrition and Health of Honey Bee Colonies Part-2: Factors Affecting the Microbial Community in Honey Bee Colonies

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Honey bee colonies have innumerable symbiotic bacteria and fungi that are essential to the health of the colony. In the first part of this series, we discussed the importance of microbes in maintaining the health of honey bee colonies. The bacteria, yeasts and molds that live in a healthy colony a...

  16. 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

  17. Colony Failure Linked to Low Sperm Viability in Honey Bee (Apis mellifera) Queens and an Exploration of Potential Causative Factors.

    PubMed

    Pettis, Jeffery S; Rice, Nathan; Joselow, Katie; vanEngelsdorp, Dennis; Chaimanee, Veeranan

    2016-01-01

    to colony performance and laboratory and field data provide evidence that temperature extremes are a potential causative factor. PMID:26863438

  18. Colony Failure Linked to Low Sperm Viability in Honey Bee (Apis mellifera) Queens and an Exploration of Potential Causative Factors

    PubMed Central

    Pettis, Jeffery S.; Rice, Nathan; Joselow, Katie; vanEngelsdorp, Dennis; Chaimanee, Veeranan

    2016-01-01

    linked to colony performance and laboratory and field data provide evidence that temperature extremes are a potential causative factor. PMID:26863438

  19. First analysis of risk factors associated with bee colony collapse disorder by classification and regression trees

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Sudden losses of managed honey bee (Apis mellifera L.) colonies are considered an important problem worldwide but the underlying cause or causes of these losses are currently unknown. In the United States, this syndrome was termed Colony Collapse Disorder (CCD), since the defining trait was a rapid ...

  20. Rac regulates vascular endothelial growth factor stimulated motility.

    PubMed

    Soga, N; Connolly, J O; Chellaiah, M; Kawamura, J; Hruska, K A

    2001-01-01

    During angiogenesis endothelial cells migrate towards a chemotactic stimulus. Understanding the mechanism of endothelial cell migration is critical to the therapeutic manipulation of angiogenesis and ultimately cancer prevention. Vascular endothelial growth factor (VEGF) is a potent chemotactic stimulus of endothelial cells during angiogenesis. The endothelial cell signal transduction pathway of VEGF represents a potential target for cancer therapy, but the mechanisms of post-receptor signal transduction including the roles of rho family GTPases in regulating the cytoskeletal effects of VEGF in endothelial cells are not understood. Here we analyze the mechanisms of cell migration in the mouse brain endothelial cell line (bEND3). Stable transfectants containing a tetracycline repressible expression vector were used to induce expression of Rac mutants. Endothelial cell haptotaxis was stimulated by constitutively active V12Rac on collagen and vitronectin coated supports, and chemotaxis was further stimulated by VEGF. Osteopontin coated supports were the most stimulatory to bEND3 haptotaxis, but VEGF was not effective in further increasing migration on osteopontin coated supports. Haptotaxis on support coated with collagen, vitronectin, and to a lesser degree osteopontin was inhibited by N17 Rac. N17 Rac expression blocked stimulation of endothelial cell chemotaxis by VEGF. As part of the chemotactic stimulation, VEGF caused a loss of actin organization at areas of cell-cell contact and increased stress fiber expression in endothelial cells which were directed towards pores in the transwell membrane. N17 Rac prevented the stimulation of cell-cell contact disruption and the stress fiber stimulation by VEGF. These data demonstrate two pathways of regulating endothelial cell motility, one in which Rac is activated by matrix/integrin stimulation and is a crucial modulator of endothelial cell haptotaxis. The other pathway, in the presence of osteopontin, is Rac independent

  1. Abiotic factors in colony formation: effects of nutrition and light on extracellular polysaccharide production and cell aggregates of Microcystis aeruginosa

    NASA Astrophysics Data System (ADS)

    Yang, Zhen; Kong, Fanxiang

    2013-07-01

    Colony morphology is important for Microcystis to sustain a competitive advantage in eutrophic lakes. The mechanism of colony formation in Microcystis is currently unclear. Extracellular polysaccharide (EPS) has been reported to play an important role in cell aggregate formation of some phytoplankton. Microcystis aeruginosa was cultivated under varied abiotic conditions, including different nutrient, light, and temperature conditions, to investigate their effects on EPS production and morphological change. The results show that nutrient concentration and light intensity have great effects on EPS productionin M. aeruginosa. There was a considerable increase in EPS production after M. aeruginosa was cultivated in adjusted culture conditions similar to those present in the field (28.9 mg C/L, 1.98 mg N/L, 0.65 mg P/L, light intensity: 100 μmol/(m2 · s)). These results indicate that abiotic factors might be one of the triggers for colony formation in Microcystis.

  2. Physical Factors Influencing Pleasant Touch during Passive Fingertip Stimulation

    PubMed Central

    Klöcker, Anne; Oddo, Calogero Maria; Camboni, Domenico; Penta, Massimo; Thonnard, Jean-Louis

    2014-01-01

    Objective Tactile explorations with the fingertips provide information regarding the physical properties of surfaces and their relative pleasantness. Previously, we performed an investigation in the active touch domain and linked several surface properties (i.e. frictional force fluctuations and net friction) with their pleasantness levels. The aim of the present study was to investigate physical factors being important for pleasantness perception during passive fingertip stimulation. Specifically we were interested to see whether factors, such as surfaces' topographies or their frictional characteristics could influence pleasantness. Furthermore, we ascertained how the stimulus pleasantness level was impacted by (i) the normal force of stimulus application (FN) and (ii) the stimulus temperature (TS). Methods and Results The right index fingertips of 22 blindfolded participants were stimulated using 27 different stimuli, which varied in average roughness (Ra) and TS. A 4-axis robot moved the stimuli horizontally under participants' fingertips with three levels of FN. The robot was equipped with force sensors, which recorded the FN and friction force (FT) during stimulation. Participants rated each stimulus according to a three-level pleasantness scale, as very pleasant (scored 0), pleasant (scored 1), or unpleasant (scored 2). These ordinal pleasantness ratings were logarithmically transformed into linear and unidimensional pleasantness measures with the Rasch model. Statistical analyses were conducted to investigate a possible link between the stimulus properties (i.e. Ra, FN, FT, and TS) and their respective pleasantness levels. Only the mean Ra and FT values were negatively correlated with pleasantness. No significant correlation was detected between FN or TS and pleasantness. Conclusion Pleasantness perception, resulting from passive fingertip stimulation, seems to be influenced by the surfaces' average roughness levels and average FT occurring during fingertip

  3. Risk factors associated with the presence of Varroa destructor in honey bee colonies from east-central Argentina.

    PubMed

    Giacobino, A; Bulacio Cagnolo, N; Merke, J; Orellano, E; Bertozzi, E; Masciangelo, G; Pietronave, H; Salto, C; Signorini, M

    2014-08-01

    Varroa destructor is considered one of the major threats for worldwide apiculture. Damage caused by varroa mite includes body weight loss, malformation and weakening of the bees. It was also suggested as the main cause associated with colony winter mortality and as an important vector for several honey bee viruses. Little is known about multiple factors and their interaction affecting V. destructor prevalence in apiaries from South America. The aim of this study was to identify risk factors associated with V. destructor prevalence in east-central Argentina. Parasitic mite infestation level and colony strength measures were evaluated in 63 apiaries distributed in 4 different regions in east-central Argentina in a cross sectional study. Data regarding management practices in each apiary were collected by means of a questionnaire. A mixed-effects logistic regression model was constructed to associate management variables with the risk of achieving mite infestation higher than 3%. Colonies owned by beekeepers who indicated that they did not monitor colonies after mite treatment (OR=2.305; 95% CI: 0.944-5.629) nor disinfect hives woodenware material (OR=2.722; 95% CI: 1.380-5.565) were associated with an increased risk of presenting high intensity infestation with V. destructor (>3%). On the other hand, beekeepers who reported replacing more than 50% of the queens in their operation (OR=0.305; 95% CI: 0.107-0.872), feeding colonies protein substitute containing natural pollen (OR=0.348; 95% CI: 0.129-0.941) and feeding colonies High Fructose Corn Syrup (HFCS) (OR=0.108; 95% CI: 0.032-0.364), had colonies that were less likely to have V. destructor infestations above 3%, than beekeepers who did not report using these management practices. Further research should be conducted considering that certain management practices were associated to mite infestation level in order to improve the sanitary condition in the colonies. Epidemiological studies provide key information to

  4. Granulocyte-Colony Stimulating Factor Improves MDX Mouse Response to Peripheral Nerve Injury

    PubMed Central

    Simões, Gustavo Ferreira; de Oliveira, Alexandre Leite Rodrigues

    2012-01-01

    Background G-CSF has been shown to increase neuronal survival, which may positively influence the spinal cord microenvironment during the course of muscular dystrophies. Methodology/Principal Findings Male MDX mice that were six weeks of age received a left sciatic nerve transection and were treated with intraperitoneal injections of 200 µg/kg/day of G-CSF 7 days before and 7 days after the transection. The axotomy was performed after the cycles of muscular degeneration/regeneration, consistent with previous descriptions of this model of muscular dystrophy. C57BL/10 mice were used as control subjects. Seven days after the surgery, the animals were sacrificed and their lumbar spinal cords were processed for immunohistochemistry (anti-MHC I, anti-Synaptophysin, anti-GFAP and anti-IBA-1) and transmission electron microscopy. MHC I expression increased in both strains of mice after the axotomy. Nevertheless, the MDX mice displayed a significantly smaller MHC I upregulation than the control mice. Regarding GFAP expression, the MDX mice showed a stronger astrogliosis compared with the C57BL/10 mice across all groups. Both groups that were treated with G-CSF demonstrated preservation of synaptophysin expression compared with the untreated and placebo groups. The quantitative analysis of the ultrastructural level showed a preservation of the synaptic covering for the both groups that were treated with G-CSF and the axotomized groups showed a smaller loss of synaptic contact in relation to the treated groups after the lesion. Conclusions/Significance The reduction of active inputs to the alpha-motoneurons and increased astrogliosis in the axotomized and control groups may be associated with the cycles of muscle degeneration/regeneration that occur postnatally. The G-CSF treated group showed a preservation of the spinal cord microenvironment after the lesion. Moreover, the increase of MHC I expression in the MDX mice that were treated with G-CSF may indicate that this drug performs an active role in regenerative potential after lesions. PMID:22912741

  5. Potentiation by granulocyte macrophage colony-stimulating factor of lipopolysaccharide toxicity in mice.

    PubMed Central

    Tiegs, G; Barsig, J; Matiba, B; Uhlig, S; Wendel, A

    1994-01-01

    GM-CSF is known to prime leukocytes for inflammatory stimuli in vitro. The objective of this study was to investigate the role of GM-CSF in vivo in a systemic inflammatory reaction syndrome. The results demonstrate a potentiation of LPS toxicity by GM-CSF in a mortality model as well as in a septic liver failure model in mice. Pretreatment of animals with 50 micrograms/kg GM-CSF induced lethality within 24 h in mice challenged with a subtoxic dose of LPS while controls survived > 72 h. A monoclonal anti-GM-CSF antibody significantly protected against a lethal LPS dose. Serum GM-CSF was inducible by LPS and peaked at 2 h. GM-CSF pretreatment dramatically potentiated systemic TNF release and hepatotoxicity induced by a subtoxic dose of LPS in galactosamine-sensitized mice. Potentiation of LPS hepatotoxicity was possible until 30 min after LPS challenge. Polyclonal anti-GM-CSF IgG protected against septic liver failure in this model and attenuated serum TNF concentrations. In vitro an ex vivo experiments revealed that after GM-CSF pretreatment LPS-induced IL-1 release from bone marrow or spleen cells was also enhanced. These findings suggest that GM-CSF represents an endogenous enhancer of LPS-induced organ injury, possibly by potentiating the release of proinflammatory cytokines such as TNF and IL-1. Images PMID:8201000

  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. Granulocyte colony-stimulating factor-primed leukocyte transfusions in candida tropicalis fungemia in neutropenic patients.

    PubMed

    Di Mario, A; Sica, S; Salutari, P; Ortu La Barbera, E; Marra, R; Leone, G

    1997-01-01

    Optimal management of fungemia in neutropenic patients is still controversial. Several reports have already stressed the poor prognosis in invasive candidiasis (80% mortality in several reports). Therefore granulocyte transfusions would appear to be useful in the management of these infections. We report the use of rhG-CSF-primed granulocyte transfusions plus amphotericin B in two neutropenic patients who developed life-threatening systemic fungal infections. This approach was successful and both patients fully recovered from the infection. PMID:9234594

  8. 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 ...

  9. An Observational Study of Honey Bee Colony Winter Losses and Their Association with Varroa destructor, Neonicotinoids and Other Risk Factors

    PubMed Central

    van der Zee, Romée; Gray, Alison; Pisa, Lennard; de Rijk, Theo

    2015-01-01

    This article presents results of an analysis of honey bee losses over the winter of 2011-2012 in the Netherlands, from a sample of 86 colonies, located at 43 apiaries. The apiaries were selected using spatially stratified random sampling. Colony winter loss data were collected and related to various measures of colony strength recorded in summer, as well as data from laboratory analysis of sample material taken from two selected colonies in each of the 43 apiaries. The logistic regression model which best explained the risk of winter loss included, in order of statistical importance, the variables (1) Varroa destructor mite infestation rate in October 2011, (2) presence of the cyano-substituted neonicotinoids acetamiprid or thiacloprid in the first 2 weeks of August 2011 in at least one of the honey bee matrices honey, bees or bee bread (pollen), (3) presence of Brassica napus (oilseed rape) or Sinapis arvensis (wild mustard) pollen in bee bread in early August 2011, and (4) a measure of the unexplained winter losses for the postal code area where the colonies were located, obtained from a different dataset. We consider in the discussion that reduced opportunities for foraging in July and August because of bad weather may have added substantially to the adverse effects of acetamiprid and thiacloprid. A novel feature of this work is its use of postal code random effects from two other independent datasets collected in the annual national monitoring by questionnaires of winter losses of honey bees in the Netherlands. These were used to plan the sample selection and also in the model fitting of the data in this study. It should however be noted that the results of the present pilot study are based on limited data, which may consequently reveal strong factors but fail to demonstrate possible interaction effects. PMID:26154346

  10. An Observational Study of Honey Bee Colony Winter Losses and Their Association with Varroa destructor, Neonicotinoids and Other Risk Factors.

    PubMed

    van der Zee, Romée; Gray, Alison; Pisa, Lennard; de Rijk, Theo

    2015-01-01

    This article presents results of an analysis of honey bee losses over the winter of 2011-2012 in the Netherlands, from a sample of 86 colonies, located at 43 apiaries. The apiaries were selected using spatially stratified random sampling. Colony winter loss data were collected and related to various measures of colony strength recorded in summer, as well as data from laboratory analysis of sample material taken from two selected colonies in each of the 43 apiaries. The logistic regression model which best explained the risk of winter loss included, in order of statistical importance, the variables (1) Varroa destructor mite infestation rate in October 2011, (2) presence of the cyano-substituted neonicotinoids acetamiprid or thiacloprid in the first 2 weeks of August 2011 in at least one of the honey bee matrices honey, bees or bee bread (pollen), (3) presence of Brassica napus (oilseed rape) or Sinapis arvensis (wild mustard) pollen in bee bread in early August 2011, and (4) a measure of the unexplained winter losses for the postal code area where the colonies were located, obtained from a different dataset. We consider in the discussion that reduced opportunities for foraging in July and August because of bad weather may have added substantially to the adverse effects of acetamiprid and thiacloprid. A novel feature of this work is its use of postal code random effects from two other independent datasets collected in the annual national monitoring by questionnaires of winter losses of honey bees in the Netherlands. These were used to plan the sample selection and also in the model fitting of the data in this study. It should however be noted that the results of the present pilot study are based on limited data, which may consequently reveal strong factors but fail to demonstrate possible interaction effects. PMID:26154346

  11. Identification of osteoblast stimulating factor 5 as a negative regulator in the B-lymphopoietic niche.

    PubMed

    Fujita, Natsuko; Ichii, Michiko; Maeda, Tetsuo; Saitoh, Norimitsu; Yokota, Takafumi; Yamawaki, Kengo; Kakitani, Makoto; Tomizuka, Kazuma; Oritani, Kenji; Kanakura, Yuzuru

    2015-11-01

    Recent studies have revealed the crucial role of the niche which supports B-lymphocyte differentiation from hematopoietic stem cells. In this study, we aimed to identify a novel regulator of B lymphopoiesis secreted in the specific niche using the signal sequence trap method. Among the identified proteins from MS5 stromal cells, expression of pleiotrophin, placental proliferin 2, and osteoblast stimulating factor 5 (OSF-5) was dominantly high in several stromal cell lines. We found that OSF-5 suppressed early B lymphopoiesis in transgenic mice producing the target protein. The number of pre-B and immature B cells was reduced by more than half compared with control in the transgenic mice. In vitro studies showed that a secreted variant of OSF-5 inhibited the proliferation and colony formation of pre-B cells, whereas cell-intrinsic form had no influence on B lymphopoiesis. The main components of the B-lymphopoietic niche, osteoblasts in mice and mesenchymal cells in humans, are primary producers of OSF-5. These results define a novel mechanism of B lymphopoiesis in bone marrow. In the specific niche, B-lymphocyte differentiation is fine-tuned by negative regulators as well as supportive factors. PMID:26213229

  12. Factors stimulating propagation of legionellae in cooling tower water

    SciTech Connect

    Yamamoto, Hiroyuki; Sugiura, Minoru; Kusunoki, Shinji; Ezaki, Takayuki; Ikedo, Masanari; Yabuuchi, Eiko )

    1992-04-01

    The authors survey of cooling tower water demonstrated that the highest density of legionellae, {ge}10{sup 4} CFU/100 ml, appeared in water containing protozoa, {ge}10{sup 2} MPN/100 ml, and heterotrophic bacteria, {ge}10{sup 6} CFU/100 ml, at water temperatures between 25 and 35C. Viable counts of legionellae were detected even in the winter samples, and propagation, up to 10{sup 5} CFU/100 ml, occurs in summer. The counts of legionellae correlated positively with increases in water temperature, pH, and protozoan counts, but not with heterotrophic bacterial counts. The water temperature of cooling towers may promote increases in the viable counts of legionellae, and certain microbes, e.g., protozoa or some heterotrophic bacteria, may be a factor stimulating the propagation of legionellae.

  13. Expression and function of hematopoiesis-stimulating factor receptors on the GPI− and GPI+ hematopoietic stem cells of patients with paroxysmal nocturnal hemoglobinuria/aplastic anemia syndrome

    PubMed Central

    FU, RONG; DING, SHAO-XUE; LIU, YI; LI, LI-JUAN; LIU, HUI; WANG, HONG-LEI; ZHANG, TIAN; SHAO, ZONG-HONG

    2016-01-01

    Paroxysmal nocturnal hemoglobinuria/aplastic anemia (PNH/AA) syndrome presents a markedly increased population of cells deficient in glycophosphatidylinositol (GPI− cells) and signs of bone marrow failure, which requires treatment with hematopoiesis-stimulating factors, such as granulocyte colony-stimulating factor (G-CSF) and stem cell factor (SCF). However, little is known about the effects of these stimulating factors on GPI− cells. In order to explore the effects of stimulating factors in PNH/AA, G-CSF receptor (CD114) and SCF receptor (CD117) expression levels on GPI+ and GPI− hematopoietic stem cells (HSCs) were measured by flow cytometry (FCM). The mean fluorescence intensity (MFI) values of signal transducer and activator of transcription 5 (STAT5) and phosphorylated (P)-STAT5 were measured in GPI+ and GPI− HSCs by FCM following stimulation with G-CSF or SCF in vitro. The expression levels of CD114 and CD117 on GPI− HSCs were significantly lower (P<0.01) than those on GPI+ HSCs in PNH/AA patients and normal controls. The MFI values of STAT5 in the GPI− and GPI+ HSCs of PNH/AA patients and normal controls were not significantly different. However, the MFI values of P-STAT5 in the GPI− HSCs of PNH/AA patients were significantly lower than those in the GPI+ HSCs of PNH/AA patients and normal controls prior to and following stimulation with G-CSF or SCF (P<0.01). The GPI− HSCs of PNH/AA patients responded poorly to stimulation by hematopoiesis-stimulating factors, which indicates that these factors can be used safely in patients with PNH/AA. PMID:27168787

  14. In vitro expansion of CD34(+)CD38(-) cells under stimulation with hematopoietic growth factors on AGM-S3 cells in juvenile myelomonocytic leukemia.

    PubMed

    Sakashita, K; Kato, I; Daifu, T; Saida, S; Hiramatsu, H; Nishinaka, Y; Ebihara, Y; Ma, F; Matsuda, K; Saito, S; Hirabayashi, K; Kurata, T; Uyen, L T N; Nakazawa, Y; Tsuji, K; Heike, T; Nakahata, T; Koike, K

    2015-03-01

    Using serum-containing culture, we examined whether AGM-S3 stromal cells, alone or in combination with hematopoietic growth factor(s), stimulated the proliferation of CD34(+) cells from patients with juvenile myelomonocytic leukemia (JMML). AGM-S3 cells in concert with stem cell factor plus thrombopoietin increased the numbers of peripheral blood CD34(+) cells to approximately 20-fold of the input value after 2 weeks in nine JMML patients with either PTPN11 mutations or RAS mutations, who received allogeneic hematopoietic transplantation. Granulocyte-macrophage colony-stimulating factor (GM-CSF) also augmented the proliferation of JMML CD34(+) cells on AGM-S3 cells. The expansion potential of CD34(+) cells was markedly low in four patients who achieved spontaneous hematological improvement. A large proportion of day-14-cultured CD34(+) cells were negative for CD38 and cryopreservable. Cultured JMML CD34(+)CD38(-) cells expressed CD117, CD116, c-mpl, CD123, CD90, but not CXCR4, and formed GM and erythroid colonies. Day-7-cultured CD34(+) cells from two of three JMML patients injected intrafemorally into immunodeficient mice stimulated with human GM-CSF after transplantation displayed significant hematopoietic reconstitution. The abilities of OP9 cells and MS-5 cells were one-third and one-tenth, respectively, of the value obtained with AGM-S3 cells. Our culture system may provide a useful tool for elucidating leukemogenesis and for therapeutic approaches in JMML. PMID:25102944

  15. A discrete artificial bee colony algorithm for detecting transcription factor binding sites in DNA sequences.

    PubMed

    Karaboga, D; Aslan, S

    2016-01-01

    The great majority of biological sequences share significant similarity with other sequences as a result of evolutionary processes, and identifying these sequence similarities is one of the most challenging problems in bioinformatics. In this paper, we present a discrete artificial bee colony (ABC) algorithm, which is inspired by the intelligent foraging behavior of real honey bees, for the detection of highly conserved residue patterns or motifs within sequences. Experimental studies on three different data sets showed that the proposed discrete model, by adhering to the fundamental scheme of the ABC algorithm, produced competitive or better results than other metaheuristic motif discovery techniques. PMID:27173272

  16. Colony failure linked to low sperm viability in honey bee (Apis mellifera) queens and an exploration of potential causative factors

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Queen health is closely linked to colony performance in honey bees as a single queen is normally responsible for all egg laying and brood production within the colony. In the U. S. in recent years, queens have been failing at a high rate; with 50% or greater of queens replaced in colonies within 6 m...

  17. Epidermal growth factor receptor-dependent stimulation of amphiregulin expression in androgen-stimulated human prostate cancer cells.

    PubMed Central

    Sehgal, I; Bailey, J; Hitzemann, K; Pittelkow, M R; Maihle, N J

    1994-01-01

    Amphiregulin is a heparin-binding epidermal growth factor (EGF)-related peptide that binds to the EGF receptor (EGF-R) with high affinity. In this study, we report a role for amphiregulin in androgen-stimulated regulation of prostate cancer cell growth. Androgen is known to enhance EGF-R expression in the androgen-sensitive LNCaP human prostate carcinoma cell line, and it has been suggested that androgenic stimuli may regulate proliferation, in part, through autocrine mechanisms involving the EGF-R. In this study, we demonstrate that LNCaP cells express amphiregulin mRNA and peptide and that this expression is elevated by androgenic stimulation. We also show that ligand-dependent EGF-R stimulation induces amphiregulin expression and that androgenic effects on amphiregulin synthesis are mediated through this EGF-R pathway. Parallel studies using the estrogen-responsive breast carcinoma cell line, MCF-7, suggest that regulation of amphiregulin by estrogen may also be mediated via an EGF-R pathway. In addition, heparin treatment of LNCaP cells inhibits androgen-stimulated cell growth further suggesting that amphiregulin can mediate androgen-stimulated LNCaP proliferation. Together, these results implicate an androgen-regulated autocrine loop composed of amphiregulin and its receptor in prostate cancer cell growth and suggest that the mechanism of steroid hormone regulation of amphiregulin synthesis may occur through androgen upregulation of the EGF-R and subsequent receptor-dependent pathways. Images PMID:8049525

  18. Stimulation of hormone-responsive adenylate cyclase activity by a factor present in the cell cytosol.

    PubMed Central

    MacNeil, S; Crawford, A; Amirrasooli, H; Johnson, S; Pollock, A; Ollis, C; Tomlinson, S

    1980-01-01

    1. Homogenates of whole tissues were shown to contain both intracellular and extracellular factors that affected particulate adenylate cyclase activity in vitro. Factors present in the extracellular fluids produced an inhibition of basal, hormone- and fluoride-stimulated enzyme activity but factors present in the cell cytosol increased hormone-stimulated activity with relatively little effect on basal or fluoride-stimulated enzyme activity. 2. The existence of this cytosol factor or factors was investigated using freshly isolated human platelets, freshly isolated rat hepatocytes, and cultured cells derived from rat osteogenic sarcoma, rat calvaria, mouse melanoma, pig aortic endothelium, human articular cartilage chondrocytes and human bronchial carcinoma (BEN) cells. 3. The stimulation of the hormone response by the cytosol factor ranged from 60 to 890% depending on the tissue of origin of the adenylate cyclase. 4. In each case the behaviour of the factor was similar to the action of GTP on that particular adenylate cyclase preparation. 5. No evidence of tissue or species specificity was found, as cytosols stimulated adenylate cyclase from their own and unrelated tissues to the same degree. 6. In the human platelet, the inclusion of the cytosol in the assay of adenylate cyclase increased the rate of enzyme activity in response to stimulation by prostaglandin E1 without affecting the amount of prostaglandin E1 required for half-maximal stimulation or the characteristics of enzyme activation by prostaglandin E. PMID:7396869

  19. Colonial America.

    ERIC Educational Resources Information Center

    Web Feet K-8, 2001

    2001-01-01

    Presents resources for grades K-8, on the subject of Colonial America. Describes Web sites; CD-ROMs and software; videos; books; audios; magazines; and professional resources. Includes two articles, "Native Americans in the Colonies," and "The Golden Age of Pirates," which also highlight resources. Presents a Web activity focusing on daily life in…

  20. Impact of West Nile virus and other mortality factors on American white pelicans at breeding colonies in the northern plains of North America

    USGS Publications Warehouse

    Sovada, M.A.; Pietz, P.J.; Converse, K.A.; Tommy, King D.; Hofmeister, E.K.; Scherr, P.; Ip, H.S.

    2008-01-01

    American white pelicans (Pelecanus erythrorhynchos) are colonial-nesting birds and their breeding sites are concentrated in a few small areas, making this species especially vulnerable to factors that can influence productivity, such as disease, disturbance, predation, weather events and loss of nesting habitat. Nearly half of the American white pelican population breeds at four colonies in the northern plains: Chase Lake National Wildlife Refuge (NWR) in North Dakota, Bitter Lake (Waubay NWR) in South Dakota, Medicine Lake NWR in Montana, and Marsh Lake in Minnesota. Thus, sustained productivity at these colonies is crucial to the health of the entire species. During the latter half of the 2002 and 2003 breeding seasons, unusually high mortality of pelican chicks was observed at these colonies. West Nile virus (WNv) was identified as one source of these losses. In 2004-2007 we monitored three major colonies in the northern plains to assess mortality of chicks during the late breeding season. We documented severe weather events, disturbance, and WNv as factors contributing to chick mortality. Before WNv arrived in the region in 2002, chick mortality after mid-July was ???4%, and then jumped to as high as 44% in the years since WNv arrived. WNv kills older chicks that are no longer vulnerable to other common mortality factors (e.g., severe weather, gull predation) and typically would have survived to fledge; thus WNv appears to be an additive mortality factor. Persistence of lower productivity at American white pelican colonies in the northern plains might reduce the adult breeding population of this species in the region.

  1. Pre-B-cell colony-enhancing factor protects against apoptotic neuronal death and mitochondrial damage in ischemia.

    PubMed

    Wang, Xiaowan; Li, Hailong; Ding, Shinghua

    2016-01-01

    We previously demonstrated that Pre-B-cell colony-enhancing factor (PBEF), also known as nicotinamide phosphoribosyltransferase (NAMPT), the rate-limiting enzyme in mammalian NAD(+) biosynthesis pathway, plays a brain and neuronal protective role in ischemic stroke. In this study, we further investigated the mechanism of its neuroprotective effect after ischemia in the primary cultured mouse cortical neurons. Using apoptotic cell death assay, fluorescent imaging, molecular biology, mitochondrial biogenesis measurements and Western blotting analysis, our results show that the overexpression of PBEF in neurons can significantly promote neuronal survival, reduce the translocation of apoptosis inducing factor (AIF) from mitochondria to nuclei and inhibit the activation of capase-3 after glutamate-induced excitotoxicity. We further found that the overexpression of PBEF can suppress glutamate-induced mitochondrial fragmentation, the loss of mitochondrial DNA (mtDNA) content and the reduction of PGC-1 and NRF-1 expressions. Furthermore, these beneficial effects by PBEF are dependent on its enzymatic activity of NAD(+) synthesis. In summary, our study demonstrated that PBEF ameliorates ischemia-induced neuronal death through inhibiting caspase-dependent and independent apoptotic signaling pathways and suppressing mitochondrial damage and dysfunction. Our study provides novel insights into the mechanisms underlying the neuroprotective effect of PBEF, and helps to identify potential targets for ischemic stroke therapy. PMID:27576732

  2. Pre-B-cell colony-enhancing factor protects against apoptotic neuronal death and mitochondrial damage in ischemia

    PubMed Central

    Wang, Xiaowan; Li, Hailong; Ding, Shinghua

    2016-01-01

    We previously demonstrated that Pre-B-cell colony-enhancing factor (PBEF), also known as nicotinamide phosphoribosyltransferase (NAMPT), the rate-limiting enzyme in mammalian NAD+ biosynthesis pathway, plays a brain and neuronal protective role in ischemic stroke. In this study, we further investigated the mechanism of its neuroprotective effect after ischemia in the primary cultured mouse cortical neurons. Using apoptotic cell death assay, fluorescent imaging, molecular biology, mitochondrial biogenesis measurements and Western blotting analysis, our results show that the overexpression of PBEF in neurons can significantly promote neuronal survival, reduce the translocation of apoptosis inducing factor (AIF) from mitochondria to nuclei and inhibit the activation of capase-3 after glutamate-induced excitotoxicity. We further found that the overexpression of PBEF can suppress glutamate-induced mitochondrial fragmentation, the loss of mitochondrial DNA (mtDNA) content and the reduction of PGC-1 and NRF-1 expressions. Furthermore, these beneficial effects by PBEF are dependent on its enzymatic activity of NAD+ synthesis. In summary, our study demonstrated that PBEF ameliorates ischemia-induced neuronal death through inhibiting caspase-dependent and independent apoptotic signaling pathways and suppressing mitochondrial damage and dysfunction. Our study provides novel insights into the mechanisms underlying the neuroprotective effect of PBEF, and helps to identify potential targets for ischemic stroke therapy. PMID:27576732

  3. Mechanically stimulated bone cells secrete paracrine factors that regulate osteoprogenitor recruitment, proliferation, and differentiation

    SciTech Connect

    Brady, Robert T.; O'Brien, Fergal J.; Hoey, David A.

    2015-03-27

    Bone formation requires the recruitment, proliferation and osteogenic differentiation of mesenchymal progenitors. A potent stimulus driving this process is mechanical loading, yet the signalling mechanisms underpinning this are incompletely understood. The objective of this study was to investigate the role of the mechanically-stimulated osteocyte and osteoblast secretome in coordinating progenitor contributions to bone formation. Initially osteocytes (MLO-Y4) and osteoblasts (MC3T3) were mechanically stimulated for 24hrs and secreted factors within the conditioned media were collected and used to evaluate mesenchymal stem cell (MSC) and osteoblast recruitment, proliferation and osteogenesis. Paracrine factors secreted by mechanically stimulated osteocytes significantly enhanced MSC migration, proliferation and osteogenesis and furthermore significantly increased osteoblast migration and proliferation when compared to factors secreted by statically cultured osteocytes. Secondly, paracrine factors secreted by mechanically stimulated osteoblasts significantly enhanced MSC migration but surprisingly, in contrast to the osteocyte secretome, inhibited MSC proliferation when compared to factors secreted by statically cultured osteoblasts. A similar trend was observed in osteoblasts. This study provides new information on mechanically driven signalling mechanisms in bone and highlights a contrasting secretome between cells at different stages in the bone lineage, furthering our understanding of loading-induced bone formation and indirect biophysical regulation of osteoprogenitors. - Highlights: • Physically stimulated osteocytes secrete factors that regulate osteoprogenitors. • These factors enhance recruitment, proliferation and osteogenic differentiation. • Physically stimulated osteoblasts secrete factors that also regulate progenitors. • These factors enhance recruitment but inhibit proliferation of osteoprogenitors. • This study highlights a contrasting

  4. STIMULATION OF DEFENSE FACTORS FOR OYSTERS DEPLOYED TO CONTAMINATED SITES IN PENSACOLA BAY, FLORIDA

    EPA Science Inventory

    A positive association between chemical contaminants and defense factors has been established for eastern oysters (Crassostrea virginica) from Florida, but it is unknown whether such factors can be stimulated through short-term exposure to contaminants in the field. Hatchery oyst...

  5. Human tumor clonogenic assay for carcinoma of the lung. II. Factors that influence colony formation in soft agar.

    PubMed

    Kanzawa, F; Matsushima, Y; Hamburger, A W; Ishihara, J; Sasaki, Y; Shimizu, E; Eguchi, K; Shinkai, T; Saijo, N; Miyazawa, N

    1987-01-01

    The human tumor clonogenic assay (HTCA) has potential value for studies of both the chemosensitivity and biology of human tumors. However, many technical problems including low plating efficiencies and the preparation of sufficient numbers of viable cells remain. In this study, an improved method for disaggregation of solid tumors increased the yield of single cells. Consequently, more than 10 anticancer drugs could be tested in 94 of 168 specimens (56%). Removal of peripheral blood lymphocytes from cell suspensions derived from effusions also improved colony formation. Adequate growth for sensitivity testing (greater than 30 colonies/plate) was obtained in 122 cases (73%), inadequate growth for drug evaluation (5-29 colonies/plate) in 29 cases (17%), and no colony formation (less than 5 colonies/plate) in 17 cases (10%) of the 168 viable samples. The cloning efficiencies of cells derived from primary tumors (median 0.015%) were higher than those of cells derived from metastatic tumors (0.012%), and they varied with the location of the metastatic site. Cloning efficiencies varied markedly from specimen to specimen, and were unaffected by tumor histology, grade of differentiation, patient age, stage of disease, or prior chemotherapy. The HTCA is promising as a potential tool for studying the biology of tumors. PMID:3601316

  6. Epidermal growth factor receptor is required for estradiol-stimulated bovine satellite cell proliferation.

    PubMed

    Reiter, B C; Kamanga-Sollo, E; Pampusch, M S; White, M E; Dayton, W R

    2014-07-01

    The objective of this study was to assess the role of the epidermal growth factor receptor (EGFR) in estradiol-17β (E2)-stimulated proliferation of cultured bovine satellite cells (BSCs). Treatment of BSC cultures with AG1478 (a specific inhibitor of EGFR tyrosine kinase activity) suppresses E2-stimulated BSC proliferation (P < 0.05). In addition, E2-stimulated proliferation is completely suppressed (P < 0.05) in BSCs in which EGFR expression is silenced by treatment with EGFR small interfering RNA (siRNA). These results indicate that EGFR is required for E2 to stimulate proliferation in BSC cultures. Both AG1478 treatment and EGFR silencing also suppress proliferation stimulated by LR3-IGF-1 (an IGF1 analogue that binds normally to the insulin-like growth factor receptor (IGFR)-1 but has little or no affinity for IGF binding proteins) in cultured BSCs (P < 0.05). Even though EGFR siRNA treatment has no effect on IGFR-1β mRNA expression in cultured BSCs, IGFR-1β protein level is substantially reduced in BSCs treated with EGFR siRNA. These data suggest that EGFR silencing results in post-transcriptional modifications that result in decreased IGFR-1β protein levels. Although it is clear that functional EGFR is necessary for E2-stimulated proliferation of BSCs, the role of EGFR is not clear. Transactivation of EGFR may directly stimulate proliferation, or EGFR may function to maintain the level of IGFR-1β which is necessary for E2-stimulated proliferation. It also is possible that the role of EGFR in E2-stimulated BSC proliferation may involve both of these mechanisms. PMID:24906928

  7. Platelet-derived growth factor stimulated mechanisms of glucosamine incorporation

    SciTech Connect

    Harrington, M.A.; Pledger, W.J. )

    1987-10-01

    Platelet-derived growth factor (PDGF) treatment of density-arrested BALB/c-3T3 cells results in increased ({sup 3}H)glucosamine (GlcN) incorporation into cellular material. The enhanced GlcN incorporation is not due to a preferential increase in proteoglycan synthesis as measured by ({sup 35}S)H{sub 2}SO{sub 4} incorporation. Approximately 50% of the GlcN incorporated in PDGF or platelet-poor plasma (PPP)-treated cultures enters N-linked glycoproteins. Addition of dolichol-phosphate (dolichol-P), a required intermediate in N-linked glycosylation, did not alter ({sup 3}H)GlcN incorporation in PDGF-treated cells but did increase incorporation in PPP-treated cultures to a level comparable to that observed for PDGF-treated cultures. PDGF-treated cultures contained twofold greater quantities of ({sup 3}H)GlcN dolichol intermediates and lipid-free glycoprotein. Over a 12-h time course 3-hydroxy-3-methylglutaryl coenzyme A reductase (HMG CoA reductase) activity was similar in cultures treated with PDGF or PPP. Results of these studies reveal that enhanced protein glycosylation in response to PDGF treatment is not the result of a direct effect on HMG CoA reductase.

  8. Carbachol stimulates a different phospholipid metabolism than nerve growth factor and basic fibroblast growth factor in PC12 cells.

    PubMed Central

    Pessin, M S; Altin, J G; Jarpe, M; Tansley, F; Bradshaw, R A; Raben, D M

    1991-01-01

    We have examined 1,2-diglycerides (DGs) generated in PC12 cells in response to the muscarinic agonist carbachol and compared them with those generated in response to the differentiation factors nerve growth factor and basic fibroblast growth factor. Whereas carbachol stimulates a greater release of inositol phosphates, all three agonists generate similar levels of DGs. In this report, we have analyzed the molecular species of PC12 DGs generated in response to these three agonists. Additionally, we have analyzed the molecular species of PC12 phospholipids. The data indicate that 1) after 1 min of either nerve growth factor or basic fibroblast growth factor stimulation, DGs arise primarily from phosphoinositide hydrolysis; 2) in contrast, after 1 min of carbachol stimulation, DG are generated equally by both phosphoinositide and phosphatidylcholine hydrolysis; and 3) after 15 min of stimulation by any of these agonists, DGs are generated largely by phosphatidylcholine hydrolysis, with a smaller component arising from the phosphoinositides. These results suggest that at least part of the mechanism by which PC12 cells distinguish between different agonists is via alterations in phospholipid sources and kinetics of DG generation. PMID:1892912

  9. Factors influencing the prevalence and infestation levels of Varroa destructor in honeybee colonies in two highland agro-ecological zones of Uganda.

    PubMed

    Chemurot, Moses; Akol, Anne M; Masembe, Charles; de Smet, Lina; Descamps, Tine; de Graaf, Dirk C

    2016-04-01

    Varroa mites are ecto-parasites of honeybees and are a threat to the beekeeping industry. We identified the haplotype of Varroa mites and evaluated potential factors that influence their prevalence and infestation levels in the eastern and western highland agro-ecological zones of Uganda. This was done by collecting samples of adult worker bees between December 2014 and September 2015 in two sampling moments. Samples of bees were screened for Varroa using the ethanol wash method and the mites were identified by molecular techniques. All DNA sequences obtained from sampled mite populations in the two zones were 100 % identical to the Korean Haplotype (AF106899). Mean mite prevalence in the apiaries was 40 and 53 % for the western and eastern zones, respectively, during the first sampling. Over the second sampling, mean mite prevalence increased considerably in the western (59 %) but not in the eastern (51 %) zone. Factors that were associated with Varroa mite infestation levels include altitude, nature of apiary slope and apiary management practices during the first sampling. Our results further showed that Varroa mites were spreading from lower to higher elevations. Feral colonies were also infested with Varroa mites at infestation levels not significantly different from those in managed colonies. Colony productivity and strength were not correlated to mite infestation levels. We recommend a long-term Varroa mite monitoring strategy in areas of varying landscape and land use factors for a clear understanding of possible changes in mite infestation levels among African honeybees for informed decision making. PMID:26801158

  10. Epidermal growth factor (EGF) inhibits stimulated thyroid hormone secretion in the mouse

    SciTech Connect

    Ahren, B.

    1987-07-01

    It is known that epidermal growth factor (EGF) inhibits iodide uptake in the thyroid follicular cells and lowers plasma levels of thyroid hormones upon infusion into sheep and ewes. In this study, the effects of EGF on basal and stimulated thyroid hormone secretion were investigated in the mouse. Mice were pretreated with /sup 125/I and thyroxine; the subsequent release of /sup 125/I is an estimation of thyroid hormone secretion. It was found that basal radioiodine secretion was not altered by intravenous injection of EGF (5 micrograms/animal). However, the radioiodine secretion stimulated by both TSH (120 microU/animal) and vasoactive intestinal peptide (VIP; 5 micrograms/animal) were inhibited by EGF (5 micrograms/animal). At a lower dose level (0.5 microgram/animal), EGF had no influence on stimulated radioiodine secretion. In conclusion, EGF inhibits stimulated thyroid hormone secretion in the mouse.

  11. Mechanically stimulated bone cells secrete paracrine factors that regulate osteoprogenitor recruitment, proliferation, and differentiation.

    PubMed

    Brady, Robert T; O'Brien, Fergal J; Hoey, David A

    2015-03-27

    Bone formation requires the recruitment, proliferation and osteogenic differentiation of mesenchymal progenitors. A potent stimulus driving this process is mechanical loading, yet the signalling mechanisms underpinning this are incompletely understood. The objective of this study was to investigate the role of the mechanically-stimulated osteocyte and osteoblast secretome in coordinating progenitor contributions to bone formation. Initially osteocytes (MLO-Y4) and osteoblasts (MC3T3) were mechanically stimulated for 24 hrs and secreted factors within the conditioned media were collected and used to evaluate mesenchymal stem cell (MSC) and osteoblast recruitment, proliferation and osteogenesis. Paracrine factors secreted by mechanically stimulated osteocytes significantly enhanced MSC migration, proliferation and osteogenesis and furthermore significantly increased osteoblast migration and proliferation when compared to factors secreted by statically cultured osteocytes. Secondly, paracrine factors secreted by mechanically stimulated osteoblasts significantly enhanced MSC migration but surprisingly, in contrast to the osteocyte secretome, inhibited MSC proliferation when compared to factors secreted by statically cultured osteoblasts. A similar trend was observed in osteoblasts. This study provides new information on mechanically driven signalling mechanisms in bone and highlights a contrasting secretome between cells at different stages in the bone lineage, furthering our understanding of loading-induced bone formation and indirect biophysical regulation of osteoprogenitors. PMID:25721667

  12. Pre-B cell colony enhancing factor (PBEF/NAMPT/Visfatin) and vascular endothelial growth factor (VEGF) cooperate to increase the permeability of the human placental amnion

    PubMed Central

    Astern, J.M.; Collier, A.C.; Kendal-Wright, C.E.

    2012-01-01

    Fluid efflux across the region of the amnion overlying the placenta is an essential component of the intramembranous absorption pathway that maintains amniotic fluid volume homeostasis. Dysregulation of this pathway may result in adverse pregnancy outcomes, however the factors controlling amnion permeability are unknown. Here, we report a novel mechanism that increases placental amnion permeability. Pre-B Cell Colony Enhancing Factor (PBEF) is a stress-responsive cytokine expressed by the human amnion, and is known to induce Vascular Endothelial Growth Factor (VEGF) production by other cell types. Interestingly, VEGF is up-regulated in the ovine amnion when intramembranous absorption is augmented. In this study, we show that PBEF induced VEGF secretion by primary human amniotic epithelial cells (AEC) derived from the placental amnion, as well as from the reflected amnion that lines the remainder of the gestational sac. Further, PBEF treatment led to the increased expression of VEGFR2 in placental AEC, but not reflected AEC. To test the hypothesis that PBEF and VEGF increase placental amnion permeability, we monitored the transfer of 2′,7′-dichlorofluorescein (DCF) from the fetal to the maternal side of human amnion explants. A treatment regimen including both PBEF and VEGF increased the rate of DCF transfer across the placental amnion, but not the reflected amnion. In summary, our results suggest that by augmenting VEGFR2 expression in the placental amnion, PBEF primes the tissue for a VEGF-mediated increase in permeability. This mechanism may have important implications in amniotic fluid volume control throughout gestation. PMID:23151382

  13. Granulocyte-macrophage colony stimulatory factor enhances the pro-inflammatory response of interferon-γ-treated macrophages to Pseudomonas aeruginosa infection.

    PubMed

    Singh, Sonali; Barr, Helen; Liu, Yi-Chia; Robins, Adrian; Heeb, Stephan; Williams, Paul; Fogarty, Andrew; Cámara, Miguel; Martínez-Pomares, Luisa

    2015-01-01

    Pseudomonas aeruginosa is an opportunistic pathogen that can cause severe infections at compromised epithelial surfaces, such those found in burns, wounds, and in lungs damaged by mechanical ventilation or recurrent infections, particularly in cystic fibrosis (CF) patients. CF patients have been proposed to have a Th2 and Th17-biased immune response suggesting that the lack of Th1 and/or over exuberant Th17 responses could contribute to the establishment of chronic P. aeruginosa infection and deterioration of lung function. Accordingly, we have observed that interferon (IFN)-γ production by peripheral blood mononuclear cells from CF patients positively correlated with lung function, particularly in patients chronically infected with P. aeruginosa. In contrast, IL-17A levels tended to correlate negatively with lung function with this trend becoming significant in patients chronically infected with P. aeruginosa. These results are in agreement with IFN-γ and IL-17A playing protective and detrimental roles, respectively, in CF. In order to explore the protective effect of IFN-γ in CF, the effect of IFN-γ alone or in combination with granulocyte-macrophage colony-stimulating factor (GM-CSF), on the ability of human macrophages to control P. aeruginosa growth, resist the cytotoxicity induced by this bacterium or promote inflammation was investigated. Treatment of macrophages with IFN-γ, in the presence and absence of GM-CSF, failed to alter bacterial growth or macrophage survival upon P. aeruginosa infection, but changed the inflammatory potential of macrophages. IFN-γ caused up-regulation of monocyte chemoattractant protein-1 (MCP-1) and TNF-α and down-regulation of IL-10 expression by infected macrophages. GM-CSF in combination with IFN-γ promoted IL-6 production and further reduction of IL-10 synthesis. Comparison of TNF-α vs. IL-10 and IL-6 vs. IL-10 ratios revealed the following hierarchy in regard to the pro-inflammatory potential of human macrophages

  14. Which Factors Obstruct or Stimulate Teacher Educators to Use ICT Innovatively?

    ERIC Educational Resources Information Center

    Drent, Marjolein; Meelissen, Martina

    2008-01-01

    This article discusses the factors which stimulate or limit the innovative use of ICT by teacher educators in the Netherlands. Innovative use of ICT is defined as the use of ICT applications that support the educational objectives based on the needs of the current knowledge society. Explorative path analysis and case studies were used to study the…

  15. Childhood Conduct Problems and Other Early Risk Factors in Rural Adult Stimulant Users

    ERIC Educational Resources Information Center

    Kramer, Teresa L.; Han, Xiaotong; Leukefeld, Carl; Booth, Brenda M.; Edlund, Carrie

    2009-01-01

    Context: Understanding childhood risk factors associated with adult substance use and legal problems is important for treatment and prevention. Purpose: To examine the relationship of early substance use, conduct problems before age 15, and family history of substance abuse on adult outcomes in rural, stimulant users. Methods: Adult cocaine and…

  16. Evaluation of transduction efficiency in macrophage colony-stimulating factor differentiated human macrophages using HIV-1 based lentiviral vectors

    PubMed Central

    2011-01-01

    Background Monocyte-derived macrophages contribute to atherosclerotic plaque formation. Therefore, manipulating macrophage function could have significant therapeutic value. The objective of this study was to determine transduction efficiency of two HIV-based lentiviral vector configurations as delivery systems for the transduction of primary human blood monocyte-derived macrophages. Results Human blood monocytes were transduced using two VSV-G pseudotyped HIV-1 based lentiviral vectors containing EGFP expression driven by either native HIV-LTR (VRX494) or EF1α promoters (VRX1090). Lentiviral vectors were added to cultured macrophages at different times and multiplicities of infection (MOI). Transduction efficiency was assessed using fluorescence microscopy and flow cytometry. Macrophages transduced between 2 and 120 hours after culturing showed the highest transduction efficiency at 2-hours transduction time. Subsequently, cells were transduced 2 hours after culturing at various vector concentrations (MOIs of 5, 10, 25 and 50) to determine the amount of lentiviral vector particles required to maximally transduce human monocyte-derived macrophages. On day 7, all transduced cultures showed EGFP-positive cells by microscopy. Flow cytometric analysis showed with all MOIs a peak shift corresponding to the presence of EGFP-positive cells. For VRX494, transduction efficiency was maximal at an MOI of 25 to 50 and ranged between 58 and 67%. For VRX1090, transduction efficiency was maximal at an MOI of 10 and ranged between 80 and 90%. Thus, transductions performed with VRX1090 showed a higher number of EGFP-positive cells than VRX494. Conclusions This report shows that VSV-G pseudotyped HIV-based lentiviral vectors can efficiently transduce human blood monocyte-derived macrophages early during differentiation using low particle numbers that do not interfere with differentiation of monocytes into macrophages. PMID:21281514

  17. Efficacy of deferred dosing of granulocyte colony-stimulating factor in autologous hematopoietic transplantation for multiple myeloma.

    PubMed

    Cox, J E; Campos, S; Wu, J; May, R; Liu, H; Ramos, C A; Carrum, G; Heslop, H E; Brenner, M K; Kamble, R T

    2014-02-01

    Routine administration of G-CSF following autologous hematopoietic SCT (ASCT) expedites ANC recovery and reduces hospitalization by 1-2 days; it has no impact on febrile neutropenia, infections, morbidity, mortality, event-free survival or OS. To determine whether delayed G-CSF dosage could result in equivalent ANC recovery and thereby improve cost effectiveness, we deferred the administration of G-CSF until WBC recovery had begun. A total of 117 patients with multiple myeloma received ASCT from January 2005 to September 2012. Of these, 52 were in the conventional dosing group (CGD) and received G-CSF from Day +7 for a median of five doses. In the deferred dosing group (DGD), 65 patients received G-CSF from median day 14 post transplant for a median of zero doses. There was no difference between groups in the incidence or duration of febrile neutropenia, duration of grade III mucositis, weight gain, rash, engraftment syndrome or early death (100 days). The DGD group had a significantly longer time to neutrophil engraftment than the CGD group (15 days vs 12 days; P<0.0001), a longer period of severe neutropenia (<100/μL; 8 days vs 6 days; P<0.0001), longer treatment with intravenous antibiotics (7 days vs 5 days; P=0.016) and longer hospital stay (19 days vs 17 days; P=<0.0001). Although the cost of G-CSF was lower in the DGD group (mean $308 vs $2467), the additional hospitalization raised the median total cost of ASCT in this group by 17%. There was, however, no adverse effect of deferred dosing on the rate of febrile neuropenic episodes or Day 100 survival, so that deferred dosing of G-CSF may be suitable for patients receiving ASCT as outpatients, for whom longer hospital stay would not be an offsetting cost. PMID:24096822

  18. Polymorphism at codon 117 of the granulocyte-macrophage colony-stimulating factor gene (GM-CSF)

    SciTech Connect

    Tagiev, A.F.; Surin, V.L.; Osokina, A.V.

    1995-10-01

    A T-to-C substitution, replacing a hydrophobic isoleucine residue with a hydrophilic threonine residue in position 100 of a mature protein molecule, was found at codon 117 of the GM-CSF gene. The mutation frequencies were estimated in 51 DNA samples from healthy adult donors and also in 20 samples from patients with different neoplastic myeloid disorders. Almost equal substitution frequencies in patients and normal individuals were observed, suggesting that the defect was not associated with leukemia. Additionally, the GM-CSF gene intron 1 sequence was refined. 44 refs., 2 tabs.

  19. Production of tumor necrosis factor alpha in human leukocytes stimulated by Cryptococcus neoformans.

    PubMed Central

    Levitz, S M; Tabuni, A; Kornfeld, H; Reardon, C C; Golenbock, D T

    1994-01-01

    Tumor necrosis factor alpha (TNF-alpha) is a key mediator of inflammation and may promote human immunodeficiency virus replication in latently infected cells. Since cryptococcosis often is associated with aberrations in the host inflammatory response and occurs preferentially in persons with AIDS, we defined the conditions under which human leukocytes produce TNF-alpha when stimulated by Cryptococcus neoformans. Peripheral blood mononuclear cells (PBMC) produced comparable amounts of TNF-alpha following stimulation with C. neoformans and lipopolysaccharide. Detectable TNF-alpha release in response to C. neoformans occurred only when fungi with small-sized capsules were used and complement-sufficient serum was added. Fractionation of PBMC established that monocytes were the predominant source of TNF-alpha. TNF-alpha gene expression and release occurred significantly later in PBMC stimulated with C. neoformans than in PBMC stimulated with LPS. C. neoformans was also a potent inducer of TNF-alpha from freshly isolated bronchoalveolar macrophages (BAM). Upon in vitro culture, BAM and monocytes bound greater numbers of fungal cells, yet their capacity to produce TNF-alpha following cryptococcal stimulation declined by 74 to 100%. However, this decline was reversed if the BAM and monocytes were cultured with gamma interferon. These data establish that C. neoformans can potently stimulate TNF-alpha release from human leukocytes. However, several variables profoundly affected the amount of TNF-alpha released, including the type of leukocyte and its state of activation, the size of the cryptococcal capsule, and the availability of opsonins. PMID:8168965

  20. Chronic sublethal stress causes bee colony failure

    PubMed Central

    Bryden, John; Gill, Richard J; Mitton, Robert A A; Raine, Nigel E; Jansen, Vincent A A; Hodgson, David

    2013-01-01

    Current bee population declines and colony failures are well documented yet poorly understood and no single factor has been identified as a leading cause. The evidence is equivocal and puzzling: for instance, many pathogens and parasites can be found in both failing and surviving colonies and field pesticide exposure is typically sublethal. Here, we investigate how these results can be due to sublethal stress impairing colony function. We mathematically modelled stress on individual bees which impairs colony function and found how positive density dependence can cause multiple dynamic outcomes: some colonies fail while others thrive. We then exposed bumblebee colonies to sublethal levels of a neonicotinoid pesticide. The dynamics of colony failure, which we observed, were most accurately described by our model. We argue that our model can explain the enigmatic aspects of bee colony failures, highlighting an important role for sublethal stress in colony declines. PMID:24112478

  1. Factor Xa stimulates fibroblast procollagen production, proliferation, and calcium signaling via PAR{sub 1} activation

    SciTech Connect

    Blanc-Brude, Olivier P. . E-mail: olivier.blanc-brude@larib.inserm.fr; Archer, Fabienne; Leoni, Patricia; Derian, Claudia; Bolsover, Steven; Laurent, Geoffrey J.; Chambers, Rachel C.

    2005-03-10

    Fibroblast proliferation and procollagen production are central features of tissue repair and fibrosis. In addition to its role in blood clotting, the coagulation cascade proteinase thrombin can contribute to tissue repair by stimulating fibroblasts via proteolytic activation of proteinase-activated receptor-1 (PAR{sub 1}). During hemostasis, the coagulation cascade proteinase factor X is converted into factor Xa. We have previously shown that factor Xa upregulates fibroblast proliferation via production of autocrine PDGF. In this study, we further examined the effects of factor Xa on fibroblast function and aimed to identify its signaling receptor. We showed that factor Xa stimulates procollagen promoter activity and protein production by human and mouse fibroblasts. This effect was independent of PDGF and thrombin production, but dependent on factor Xa proteolytic activity. We also showed that PAR{sub 1}-deficient mouse fibroblasts did not upregulate procollagen production, mobilize cytosolic calcium, or proliferate in response to factor Xa. Desensitization techniques and PAR{sub 1}-specific agonists and inhibitors were used to demonstrate that PAR{sub 1} mediates factor Xa signaling in human fibroblasts. This is the first report that factor Xa stimulates extracellular matrix production. In contrast with endothelial cells and vascular smooth muscle cells, fibroblasts appear to be the only cell type in which the effects of factor Xa are mediated mainly via PAR{sub 1} and not PAR{sub 2}. These findings are critical for our understanding of tissue repair and fibrotic mechanisms, and for the design of novel approaches to inhibit the profibrotic effects of the coagulation cascade without compromising blood hemostasis.

  2. Glioma-secreted soluble factors stimulate microglial activation: The role of interleukin-1β and tumor necrosis factor-α.

    PubMed

    Hwang, Ji-Sun; Jung, Eun-Hye; Kwon, Mi-Youn; Han, Inn-Oc

    2016-09-15

    We aimed to elucidate the effect of soluble factors secreted by glioma on microglial activation. Conditioned medium (CM) from glioma cells, CRT-MG and C6, significantly induced nitric oxide (NO) production and stimulated the mRNA expression of inducible NO synthase (iNOS), interleukin (IL)-1beta, IL-6, tumor necrosis factor-alpha (TNF-α) and cyclooxygenase 2 (COX-2) in BV2 cells. Glioma CM stimulated p38 mitogen-activated protein kinase (MAPK) phosphorylation, and a p38 MAPK inhibitor, SB203580, suppressed CM-induced NO production in BV2 cells. In addition, CM stimulated nuclear factor-kappaB (NF-κB) DNA binding and transcriptional activity, which was repressed by SB203580. Gliomas displayed higher mRNA expression and release of TNF-α and IL-1β than primary astrocyte cells. Neutralization of TNF-α and IL-1β in C6-CM using a neutralizing antibody inhibited NO/iNOS expression in BV-2 cells. These results indicate potential contribution of diffusible tumor-derived factors to regulate microglial activation and subsequent tumor microenvironment. PMID:27609291

  3. Imatinib mesylate inhibits platelet derived growth factor stimulated proliferation of rheumatoid synovial fibroblasts

    SciTech Connect

    Sandler, Charlotta; Joutsiniemi, Saima; Lindstedt, Ken A.; Juutilainen, Timo; Kovanen, Petri T.; Eklund, Kari K. . E-mail: kari.eklund@hus.fi

    2006-08-18

    Synovial fibroblast is the key cell type in the growth of the pathological synovial tissue in arthritis. Here, we show that platelet-derived growth factor (PDGF) is a potent mitogen for synovial fibroblasts isolated from patients with rheumatoid arthritis. Inhibition of PDGF-receptor signalling by imatinib mesylate (1 {mu}M) completely abrogated the PDGF-stimulated proliferation and inhibited approximately 70% of serum-stimulated proliferation of synovial fibroblasts. Similar extent of inhibition was observed when PDGF was neutralized with anti-PDGF antibodies, suggesting that imatinib mesylate does not inhibit pathways other than those mediated by PDGF-receptors. No signs of apoptosis were detected in synovial fibroblasts cultured in the presence of imatinib. These results suggest that imatinib mesylate specifically inhibits PDGF-stimulated proliferation of synovial fibroblasts, and that inhibition of PDGF-receptors could represent a feasible target for novel antirheumatic therapies.

  4. Secretion of corticotrophin releasing factor from the adrenal during splanchnic nerve stimulation in conscious calves.

    PubMed Central

    Edwards, A V; Jones, C T

    1988-01-01

    1. The output of corticotrophin releasing factor-like immunoreactivity (CRF) from the adrenal gland has been investigated using the 'adrenal clamp' technique in conscious calves. 2. Stimulation of the peripheral end of the splanchnic nerve for 10 min increased the mean output of CRF progressively, so that it had risen by about twentyfold, to a peak incremental value of 24 +/- 4 pmol min-1 kg-1 at 10 min. This response was significantly increased by stimulating in bursts at 40 Hz for 1 s at 10 s intervals, which raised the mean CRF output by 44 +/- 7 pmol min-1 kg-1 at 10 min (P less than 0.05). 3. The mean output of adrenaline and noradrenaline rose more abruptly in response to splanchnic nerve stimulation with peak incremental values realized within 2.5 min. However, the ratios of adrenal CRF to catecholamine output were closely similar during the later stages of stimulation (7.5-10 min). There was a similarly abrupt rise in adrenal cortisol output in response to splanchnic nerve stimulation which was, nevertheless, linearly related to arterial plasma ACTH concentration throughout. 4. In hypophysectomized calves, administration of adrenocorticotrophic hormone (ACTH1-24) at a dose of 5 ng min-1 kg-1 reduced the output of adrenal CRF in response to splanchnic nerve stimulation by about 50% (P less than 0.05). 5. CRF isolated from adrenal venous effluent plasma, collected both at rest and during splanchnic nerve stimulation, was separated by reverse-phase high-pressure liquid chromatography and found to elute in a position identical to that of human 41CRF. This suggests that adrenal CRF is structurally closely similar to its pituitary counterpart. PMID:2843642

  5. The role of glucocorticoids in naturally fasting grey seal (Halichoerus grypus) pups: dexamethasone stimulates mass loss and protein utilisation, but not departure from the colony.

    PubMed

    Bennett, K A; Fedak, M A; Moss, S E W; Pomeroy, P P; Speakman, J R; Hall, A J

    2013-03-15

    Seals must manage their energy reserves carefully while they fast on land to ensure that they go to sea with sufficient fuel to sustain them until they find food. Glucocorticoids (GCs) have been implicated in the control of fuel metabolism and termination of fasting in pinnipeds. Here we tested the hypothesis that dexamethasone, an artificial GC, increases fat and protein catabolism, and induces departure from the breeding colony in wild, fasting grey seal pups. A single intramuscular dose of dexamethasone completely suppressed cortisol production for 24-72 h, demonstrating activation of GC receptors. In experiment 1, we compared the effects of a single dose of dexamethasone or saline administered 10 days after weaning on fasting mass and body composition changes, cortisol, blood urea nitrogen (BUN) and glucose levels, and timing of departure from the colony. In experiment 2, we investigated the effects of dexamethasone on short-term (5 days) changes in mass loss, body composition and BUN levels. In experiment 1, dexamethasone induced a short-lived increase in mass loss, but there was no difference in timing of departure between dexamethasone- and saline-treated pups (N=10). In experiment 2, dexamethasone increased protein and water loss and prevented a decrease in BUN levels (N=11). Our data suggest changes in cortisol contribute to regulation of protein catabolism in fasting seal pups, irrespective of the sex of the animal, but do not terminate fasting. By affecting the rate of protein depletion, lasting changes in cortisol levels could influence the amount of time seal pups have to find food, and thus may have important consequences for their survival. PMID:23197086

  6. Childhood Conduct Problems and Other Early Risk Factors in Rural Adult Stimulant Users

    PubMed Central

    Kramer, Teresa L.; Han, Xiaotong; Leukefeld, Carl; Booth, Brenda M.; Edlund, Carrie

    2009-01-01

    Context Understanding childhood risk factors associated with adult substance use and legal problems is important for treatment and prevention. Purpose To examine the relationship of early substance use, conduct problems before age 15, and family history of substance abuse on adult outcomes in rural, stimulant users. Methods Adult cocaine and methamphetamine users (N=544) in rural Arkansas and Kentucky were interviewed. Data were analyzed using both bivariate analyses and multiple logistic and log-linear regression models, with dependent variables being any substance abuse/dependence, stimulant abuse/dependence, total number of arrests since age 18 and days incarcerated since age 18. Findings One-third reported three or more conduct disorder problems prior to age 15; half reported initiation of substances (excluding alcohol) before age 15; and 60% reported family history of substance problems. All three variables were associated with adult substance abuse/dependence but only the latter two were associated with stimulant abuse/dependence. Conclusions This study highlights early risk factors for adult substance abuse/dependence among rural stimulant users. PMID:19166561

  7. Cohesion Establishment Factors Stimulate Endonuclease Activity of hFen1 Independently and Cooperatively.

    PubMed

    Kim, Do-Hyung; Kim, Jeong-Hoon; Park, Byoung Chul; Cho, Sayeon; Park, Sung Goo

    2015-10-01

    Human Fen1 protein (hFen1) plays an important role in Okazaki fragment processing by cleaving the flap structure at the junction between single-stranded (ss) DNA and doublestranded (ds) DNA, an intermediate formed during Okazaki fragment processing, resulting in ligatable nicked dsDNA. It was reported that hChlR1, a member of the cohesion establishment factor family, stimulates hFen1 nuclease activity regardless of its ATPase activity. In this study, we found that cohesion establishment factors cooperatively stimulate endonuclease activity of hFen1 in in vivo mimic condition, including replication protein-A-coated DNA and high salt. Our findings are helpful to explain how a DNA replication machinery larger than the cohesion complex goes through the cohesin ring structure on DNA during S phase in the cell cycle. PMID:26032365

  8. PXR stimulates growth factor-mediated hepatocyte proliferation by cross-talk with the FOXO transcription factor.

    PubMed

    Shizu, Ryota; Abe, Taiki; Benoki, Satoshi; Takahashi, Miki; Kodama, Susumu; Miayata, Masaaki; Matsuzawa, Atsushi; Yoshinari, Kouichi

    2016-02-01

    Growth factor-mediated hepatocyte proliferation is crucial in liver regeneration and the recovery of liver function after injury. The nuclear receptor, pregnane X receptor (PXR), is a key transcription factor for the xenobiotic-induced expression of genes associated with various liver functions. Recently, we reported that PXR activation stimulates xenobiotic-induced hepatocyte proliferation. In the present study, we investigated whether PXR activation also stimulates growth factor-mediated hepatocyte proliferation. In G0 phase-synchronized, immortalized mouse hepatocytes, serum or epidermal growth factor treatment increased cell growth and this growth was augmented by the expression of mouse PXR and co-treatment with pregnenolone 16α-carbonitrile (PCN), a PXR ligand. In a liver regeneration model using carbon tetrachloride, PCN treatment enhanced the injury-induced increase in the number of Ki-67-positive nuclei as well as Ccna2 and Ccnb1 mRNA levels in wild-type (WT) but not Pxr-null mice. Chronological analysis of this model demonstrated that PCN treatment shifted the maximum cell proliferation to an earlier time point and increased the number of M-phase cells at those time points. In WT but not Pxr-null mice, PCN treatment reduced hepatic mRNA levels of genes involved in the suppression of G0/G1- and G1/S-phase transition, e.g. Rbl2, Cdkn1a and Cdkn1b. Analysis of the Rbl2 promoter revealed that PXR activation inhibited its Forkhead box O3 (FOXO3)-mediated transcription. Finally, the PXR-mediated enhancement of hepatocyte proliferation was inhibited by the expression of dominant active FOXO3 in vitro. The results of the present study suggest that PXR activation stimulates growth factor-mediated hepatocyte proliferation in mice, at least in part, through inhibiting FOXO3 from accelerating cell-cycle progression. PMID:26574435

  9. Differential and synergistic effects of mechanical stimulation and growth factor presentation on vascular wall function

    PubMed Central

    Liang, Mao-Shih; Koobatian, Maxwell T.; Lei, Pedro; Swartz, Daniel D.; Andreadis, Stelios T.

    2013-01-01

    We investigated the hypothesis that immobilizing TGF-β1 within fibrin hydrogels may act in synergy with cyclic mechanical stimulation to enhance the properties of vascular grafts. To this end, we engineered a fusion TGF-β1 protein that can covalently anchor to fibrin during polymerization upon the action of factor XIII. We also developed a 24-well based bioreactor in which vascular constructs can be mechanically stimulated by distending the silastic mandrel in the middle of each well. TGF-β1 was either conjugated to fibrin or supplied in the culture medium and the fibrin based constructs were cultured statically for a week followed by cyclic distention for another week. The tissues were examined for myogenic differentiation, vascular reactivity, mechanical properties and ECM content. Our results showed that some aspects of vascular function were differentially affected by growth factor presentation vs. pulsatile force application, while others were synergistically enhanced by both. Overall, this two-prong biomimetic approach improved ECM secretion, vascular reactivity and mechanical properties of vascular constructs. These findings may be applied in other tissue engineering applications such as cartilage, tendon or cardiac regeneration where growth factors TGF-β1 and mechano-stimulation play critical roles. PMID:23810080

  10. Some growth factors stimulate cultured adult rabbit ventricular myocyte hypertrophy in the absence of mechanical loading

    NASA Technical Reports Server (NTRS)

    Decker, R. S.; Cook, M. G.; Behnke-Barclay, M.; Decker, M. L.

    1995-01-01

    Cultured adult rabbit cardiac myocytes treated with recombinant growth factors display enhanced rates of protein accumulation (ie, growth) in response to insulin and insulin-like growth factors (IGFs), but epidermal growth factor, acidic or basic fibroblast growth factor, and platelet-derived growth factor failed to increase contractile protein synthesis or growth of the heart cells. Insulin and IGF-1 increased growth rates by stimulating anabolic while simultaneously inhibiting catabolic pathways, whereas IGF-2 elevated growth modestly by apparently inhibiting lysosomal proteolysis. Neutralizing antibodies directed against either IGF-1 or IGF-2 or IGF binding protein 3 blocked protein accumulation. A monoclonal antibody directed against the IGF-1 receptor also inhibited changes in protein turnover provoked by recombinant human IGF-1 but not IGF-2. Of the other growth factors tested, only transforming growth factor-beta 1 increased the fractional rate of myosin heavy chain (MHC) synthesis, with beta-MHC synthesis being elevated and alpha-MHC synthesis being suppressed. However, the other growth factors were able to modestly stimulate the rate of DNA synthesis in this preparation. Bromodeoxyuridine labeling revealed that these growth factors increased DNA synthesis in myocytes and nonmyocytes alike, but the heart cells displayed neither karyokinesis or cytokinesis. In contrast, cocultures of cardiac myocytes and nonmyocytes and nonmyocyte-conditioned culture medium failed to enhance the rate of cardiac MHC synthesis or its accumulation, implying that quiescent heart cells do not respond to "conditioning" by cardiac nonmyocytes. These findings demonstrated that insulin and the IGFs promote passively loaded cultured adult rabbit heart cells to hypertrophy but suggest that other growth factors tested may be limited in this regard.

  11. ORF3 of Hepatitis E Virus Inhibits the Expression of Proinflammatory Cytokines and Chemotactic Factors in LPS-Stimulated Human PMA-THP1 Cells by Inhibiting NF-κB Pathway.

    PubMed

    Lei, Qingsong; Li, Lin; Cai, Jia; Huang, Wenxiang; Qin, Bo; Zhang, Shujun

    2016-03-01

    Hepatitis E virus (HEV) is one of the primary causative agents of acute hepatitis. It is noteworthy that HEV can develop chronic infection and even lead to liver cirrhosis; however, the mechanism has not been revealed. In this study, the ELISA assay was used to detect protein levels, and we found that HEV open reading frame 3 (ORF3) protein inhibited the expression of proinflammatory cytokines (tumor necrosis factor-alpha [TNF-α], interleukin [IL]-1β, IL-6, IL-8, IL-12p40, and IL-18) and chemotactic factors (nitric oxide [NO], interferon-inducible protein-10 (IP-10), macrophage inflammatory protein (MIP)-1α, monocyte chemoattractant protein-1 (MCP-1), granulocyte colony-stimulating factor (G-CSF), granulocyte macrophage colony-stimulating factor (GM-CSF)] in lipopolysaccharide (LPS)-stimulated human PMA-THP1 cells. Further study showed that mRNA and protein levels of pattern recognition receptors (PRRs), such as Toll-like receptor 4 (TLR4), TNF receptor-associated factor 6 (TRAF6), and nucleotide-binding oligomerization domain containing 2 (NOD2), decreased after infection of pLL3.7-ORF3 (pORF3); moreover, the inhibition produced corresponding upregulation of IκBα and downregulation of phosphorylated IκB kinase IKKɛ (p-IKKɛ) and phosphorylated nuclear factor (NF)-κB (p-NF-κB), but little variation was found in the concentration of IKKɛ and NF-κB. Taken together, our results demonstrated that HEV ORF3 attenuated LPS-induced cytokine production and chemotactic factors, predominantly by inhibiting various PRRs-mediated NF-κB signaling pathways. The anti-inflammatory properties might be of great importance to clarify the role and mechanism of macrophages in chronic HEV infection and cirrhosis. PMID:26771290

  12. Cancer drug troglitazone stimulates the growth and response of renal cells to hypoxia inducible factors.

    PubMed

    Taub, Mary

    2016-03-11

    Troglitazone has been used to suppress the growth of a number of tumors through apoptosis and autophagy. However, previous in vitro studies have employed very high concentrations of troglitazone (≥10(-5) M) in order to elicit growth inhibitory effects. In this report, when employing lower concentrations of troglitazone in defined medium, troglitazone was observed to stimulate the growth of primary renal proximal tubule (RPT) cells. Rosiglitazone, like troglitazone, is a thiazolidinedione (TZD) that is known to activate Peroxisome Proliferator Activated Receptor Υ (PPARΥ). Notably, rosiglitazone also stimulates RPT cell growth, as does Υ-linolenic acids, another PPARΥ agonist. The PPARΥ antagonist GW9662 inhibited the growth stimulatory effect of troglitazone. In addition, troglitazone stimulated transcription by a PPAR Response Element/Luciferase construct. These results are consistent with the involvement of PPARΥ as a mediator of the growth stimulatory effect of troglitazone. In a number of tumor cells, the expression of hypoxia inducible factor (HIF) is increased, promoting the expression of HIF inducible genes, and vascularization. Troglitazone was observed to stimulate transcription by a HIF/luciferase construct. These observations indicate that troglitazone not only promotes growth, also the survival of RPT cells under conditions of hypoxia. PMID:26869517

  13. Overlapping sites for constitutive and induced DNA binding factors involved in interferon-stimulated transcription.

    PubMed Central

    Dale, T C; Rosen, J M; Guille, M J; Lewin, A R; Porter, A G; Kerr, I M; Stark, G R

    1989-01-01

    A 14 bp interferon (IFN)-stimulated response element (ISRE) from 6-16, a human gene regulated by alpha-IFN, confers IFN inducibility on a heterologous thymidine kinase promoter. A 39 bp double-stranded oligonucleotide corresponding to a 5' region of 6-16 which includes the ISRE competes for factors required for gene expression by alpha-IFN in transfected cells and a single base change (A-11 to C) within the ISRE (GGGAAAATGAAACT) abolishes this competition. Band-shift assays performed with whole-cell extracts and the 39 bp oligonucleotide reveal specific complexes formed by rapidly induced and constitutive factors, both of which fail to bind to the A-11 to C oligonucleotide. A detailed footprinting analysis reveals that these two types of factors bind to overlapping sites within the ISRE, but in very different ways. These data were used to design oligonucleotides which decreased the formation of the inducible complex without affecting the constitutive one. Changes at the 5' margin of the ISRE and upstream of it markedly decrease formation of the induced but not the constitutive complex and also abolish the ability of the 39 bp sequence to function as an inducible enhancer with the thymidine kinase promoter. Thus, induction of 6-16 transcription in IFN-treated cells is likely to be stimulated by binding of the induced factor to the ISRE and upstream sequences, while the subsequent suppression of transcription may involve competition for the ISRE by the other class of factors. Images PMID:2721502

  14. Ex Vivo Assay of Electrical Stimulation to Rat Sciatic Nerves: Cell Behaviors and Growth Factor Expression.

    PubMed

    Du, Zhiyong; Bondarenko, Olexandr; Wang, Dingkun; Rouabhia, Mahmoud; Zhang, Ze

    2016-06-01

    Neurite outgrowth and axon regeneration are known to benefit from electrical stimulation. However, how neuritis and their surroundings react to electrical field is difficult to replicate by monolayer cell culture. In this work freshly harvested rat sciatic nerves were cultured and exposed to two types of electrical field, after which time the nerve tissues were immunohistologically stained and the expression of neurotrophic factors and cytokines were evaluated. ELISA assay was used to confirm the production of specific proteins. All cell populations survived the 48 h culture with little necrosis. Electrical stimulation was found to accelerate Wallerian degeneration and help Schwann cells to switch into migratory phenotype. Inductive electrical stimulation was shown to upregulate the secretion of multiple neurotrophic factors. Cellular distribution in nerve tissue was altered upon the application of an electrical field. This work thus presents an ex vivo model to study denervated axon in well controlled electrical field, bridging monolayer cell culture and animal experiment. It also demonstrated the critical role of electrical field distribution in regulating cellular activities. PMID:26516696

  15. GROWTH REGULATING FACTOR5 Stimulates Arabidopsis Chloroplast Division, Photosynthesis, and Leaf Longevity1[OPEN

    PubMed Central

    Vercruyssen, Liesbeth; Tognetti, Vanesa B.; Gonzalez, Nathalie; Van Dingenen, Judith; De Milde, Liesbeth; Bielach, Agnieszka; De Rycke, Riet; Van Breusegem, Frank; Inzé, Dirk

    2015-01-01

    Arabidopsis (Arabidopsis thaliana) leaf development relies on subsequent phases of cell proliferation and cell expansion. During the proliferation phase, chloroplasts need to divide extensively, and during the transition from cell proliferation to expansion, they differentiate into photosynthetically active chloroplasts, providing the plant with energy. The transcription factor GROWTH REGULATING FACTOR5 (GRF5) promotes the duration of the cell proliferation period during leaf development. Here, it is shown that GRF5 also stimulates chloroplast division, resulting in a higher chloroplast number per cell with a concomitant increase in chlorophyll levels in 35S:GRF5 leaves, which can sustain higher rates of photosynthesis. Moreover, 35S:GRF5 plants show delayed leaf senescence and are more tolerant for growth on nitrogen-depleted medium. Cytokinins also stimulate leaf growth in part by extending the cell proliferation phase, simultaneously delaying the onset of the cell expansion phase. In addition, cytokinins are known to be involved in chloroplast development, nitrogen signaling, and senescence. Evidence is provided that GRF5 and cytokinins synergistically enhance cell division and chlorophyll retention after dark-induced senescence, which suggests that they also cooperate to stimulate chloroplast division and nitrogen assimilation. Taken together with the increased leaf size, ectopic expression of GRF5 has great potential to improve plant productivity. PMID:25604530

  16. Hepatic nuclear factor 3 is an accessory factor required for the stimulation of phosphoenolpyruvate carboxykinase gene transcription by glucocorticoids.

    PubMed

    Wang, J C; Strömstedt, P E; O'Brien, R M; Granner, D K

    1996-07-01

    Transcription of the hepatic phosphoenolpyruvate carboxykinase gene is stimulated by glucocorticoids and inhibited by insulin. The glucocorticoid response is mediated by a complex glucocorticoid response unit that consists of two glucocorticoid receptor (GR)-binding sites (GR1 and GR2) and two accessory factor-binding sites (AF1 and AF2). The complete unit is required for the full glucocorticoid response. The dominant insulin effect is mediated in part through an insulin response sequence that is coincident with the AF2 element. Members of the hepatic nuclear factor 3 (HNF3) and CCAAT enhancer binding protein (C/EBP) families bind to the AF2 element; however, there is no correlation between binding of these factors and the ability of the AF2 element to mediate an insulin response. We show here that binding of HNF3 does correlate with the stimulation of the glucocorticoid response by the AF2 element and that C/EBP is apparently not involved in this effect. This requirement for HNF3 is quite specific since the substitution of elements known to enhance the action of the GR in other promoters fails to recapitulate AF2 accessory factor activity. By contrast, an HNF3-binding site from the transthyretin gene is able to substitute for the wild type AF2 sequence and elicit a maximal glucocorticoid response. Based on current and previous observations, the glucocorticoid response unit consists of four DNA elements that bind four different proteins. These are: AF1 (hepatic nuclear factor 4/chicken ovalbumin upstream promoter transcription factor), AF2 (HNF3), GR1 (GR), and GR2 (GR). PMID:8813720

  17. Characteristics of rat megakaryocyte colonies and their progenitors in agar culture

    SciTech Connect

    Kellar, K.L.; Rolovic, Z.; Evatt, B.L.; Sewell, E.T.

    1985-11-01

    The characteristics of megakaryocyte colonies that develop from megakaryocyte progenitors of rat bone marrow stimulated by rat spleen-conditioned medium (SCM) in agar culture were investigated. Colony frequency was optimal on day 7 and increased relative to both the number of cells plated and the concentration of SCM used. Colonies were categorized as small cell and big cell. Small-cell colonies had a greater proliferative potential, with a mean of 25 cells/colony. Big-cell colonies averaged 15 cells/colony. The ratio of big-cell to small-cell colonies was 0.69 +/- 0.29. Granulocyte-macrophage colonies, which were also stimulated by SCM, accounted for 70% +/- 15% of the total colonies in the cultures. Cytocidal experiments with tritiated thymidine reduced megakaryocyte colony formation by 45% and granulocyte-macrophage colony formation by 21%. The properties of rat, mouse, and human megakaryocyte progenitors as assayed in vitro are compared.

  18. Attractant and stimulant factors for oviposition of Culex pipiens fatigans in natural breeding-sites*

    PubMed Central

    Ikeshoji, Toshiaki

    1966-01-01

    The breeding of mosquito larvae in the field is determined by the ovipositing behaviour of the gravid females. Investigation of the chemical factors that induce oviposition is therefore important for understanding mosquito ecology. These substances may also prove to be useful in assessing and controlling mosquito populations. The author has demonstrated two chemical factors, an ovipositing attractant and an ovipositing stimulant, in surface-water. The ovipositing attractant, extracted from surface-water by distillation and extraction with diethyl ether, was found to be quite effective when used to recapture known numbers of gravid mosquitos released in a large calf-shed. The presence of the stimulant factor was established by forcing gravid females to touch the testing water with tarsi and proboscis. After such contact, they began oviposition three times more rapidly on surface-water than on tap-water. The importance of these factors was demonstrated in the larval populations of Culex pipiens fatigans in pit-latrines in Rangoon, Burma. PMID:5298039

  19. Stimulation of LDL receptor activity in Hep-G2 cells by a serum factor(s)

    SciTech Connect

    Ellsworth, J.L.; Brown, C.; Cooper, A.D.

    1988-05-01

    The regulation of low-density lipoprotein (LDL) receptor activity in the human hepatoma cell line Hep-G2 by serum components was examined. Incubation of dense monolayers of Hep-G2 cells with fresh medium containing 10% fetal calf serum (FM) produced a time-dependent increase in LDL receptor activity. Uptake and degradation of 125I-LDL was stimulated two- to four-fold, as compared with that of Hep-G2 cells cultured in the same media in which they had been grown to confluence (CM); the maximal 125I-LDL uptake plus degradation increased from 0.2 microgram/mg cell protein/4 h to 0.8 microgram/mg cell protein/4 h. In addition, a two-fold increase in cell surface binding of 125I-LDL to Hep-G2 cells was observed when binding was measured at 4 degrees C. There was no change in the apparent Kd. The stimulation of LDL receptor activity was suppressed in a concentration-dependent manner by the addition of cholesterol, as LDL, to the cell medium. In contrast to the stimulation of LDL receptor activity, FM did not affect the uptake or degradation of 125I-asialoorosomucoid. Addition of FM increased the protein content per dish, and DNA synthesis was stimulated approximately five-fold, as measured by (3H)thymidine incorporation into DNA; however, the cell number did not change. Cellular cholesterol biosynthesis was also stimulated by FM; (14C)acetate incorporation into unesterified and esterified cholesterol was increased approximately five-fold. Incubation of Hep-G2 cells with high-density lipoproteins (200 micrograms protein/ml) or albumin (8.0 mg/ml) in the absence of the serum factor did not significantly increase the total processed 125I-LDL. Stimulation of LDL receptor activity was dependent on a heat-stable, nondialyzable serum component that eluted in the inclusion volume of a Sephadex G-75 column.

  20. Priming effect of platelet activating factor on leukotriene C4 from stimulated eosinophils of asthmatic patients.

    PubMed Central

    Shindo, K.; Koide, K.; Hirai, Y.; Sumitomo, M.; Fukumura, M.

    1996-01-01

    BACKGROUND: Eosinophils from asthmatic patients are known to release greater amounts of leukotrienes than normal eosinophils when stimulated by the calcium ionophore A23187. The effect of platelet activating factor (PAF) in priming eosinophils was investigated. METHODS: Eosinophils were obtained from 18 asthmatic patients and 18 healthy donors. Cells separated by the Percoll gradients were incubated with PAF (C-18) for 30 minutes and then stimulated with the calcium ionophore A23187 (2.5 microM) for 15 minutes. The amount of leukotriene C4 (LTC4) in supernatants was measured using a combination of high pressure liquid chromatography and radioimmunoassay. RESULTS: The mean (SD) amount of LTC4 released by eosinophils from asthmatic patients upon stimulation with the calcium ionophore A23187 alone was 27.9 (9.9) ng/10(6) cells (n = 6). The amount of LTC4 released following stimulation with the calcium ionophore A23187 after pretreatment with PAF (1, 5, and 10 microM) was 57.2 (8.9), 75.1 (14.3), and 52.6 (10.7) ng/10(6) cells (n = 6), respectively. Trace amounts of LTC4 (0.9 (0.02) ng/10(6) cells, n = 6) were detected in the supernatant of the cells after stimulation by PAF alone (5 microM). The amount of LTC4 released upon stimulation by calcium ionophore A23187 alone in eosinophils from healthy donors was 10.3 (3.7) ng/10(6) cells (n = 4). The amounts of LTC4 released upon stimulation with calcium ionophore A23187 after pretreatment with PAF at concentrations of 1, 5, and 10 microM were 11.9 (3.5), 17.8 (5.6), and 12.7 (5.1) ng/10(6) cells (n = 4), respectively. Trace amounts of LTC4 (0.6 (0.02) ng/10(6) cells, n = 4) were detected in the supernatant of the cells upon stimulation with PAF alone (5 microM). The amounts of LTC4 released upon stimulation with calcium ionophore A23187 after pretreatment with lyso-PAF at concentrations of 1, 5, and 10 microM (n = 4 or 6) were 30.8 (5.2), 22.9 (5.1), and 27.3 (4.3) ng/10(6) cells (n = 6) from the eosinophils of asthmatic

  1. Enhanced jun gene expression is an early genomic response to transforming growth factor beta stimulation.

    PubMed Central

    Pertovaara, L; Sistonen, L; Bos, T J; Vogt, P K; Keski-Oja, J; Alitalo, K

    1989-01-01

    Transforming growth factor beta (TGF beta) is a multifunctional polypeptide that regulates proliferation, differentiation, and other functions of many cell types. The pathway of TGF beta signal transduction in cells is unknown. We report here that an early effect of TGF beta is an enhancement of the expression of two genes encoding serum- and phorbol ester tumor promoter-regulated transcription factors: the junB gene and the c-jun proto-oncogene, respectively. This stimulation was observed in human lung adenocarcinoma A549 cells which were growth inhibited by TGF beta, AKR-2B mouse embryo fibroblasts which were growth stimulated by TGF beta, and K562 human erythroleukemia cells, which were not appreciably affected in their growth by TGF beta. The increase in jun mRNA occurred with picomolar TGF beta concentrations within 1 h of TGF beta stimulation, reached a peak between 1 and 5 h in different cells, and declined gradually to base-line levels. This mRNA response was followed by a large increase in the biosynthesis of the c-jun protein (AP-1), as shown by metabolic labeling and immunoprecipitation analysis. However, differential and cell type-specific regulation appeared to determine the timing and magnitude of the response of each jun gene in a given cell. In AKR-2B and NIH 3T3 cells, only junB was induced by TGF beta, evidently in a protein synthesis-independent fashion. The junB response to TGF beta was maintained in c-Ha-ras and neu oncogene-transformed cells. Thus, one of the earliest genomic responses to TGF beta may involve nuclear signal transduction and amplification by the junB and c-jun transcription factors in concert with c-fos, which is also induced. The differential activation of the jun genes may explain some of the pleiotropic effects of TGF beta. Images PMID:2725496

  2. Domains of ERRgamma that mediate homodimerization and interaction with factors stimulating DNA binding.

    PubMed

    Hentschke, Moritz; Süsens, Ute; Borgmeyer, Uwe

    2002-08-01

    The estrogen receptor-related receptor gamma (ERRgamma/ERR3/NR3B3) is an orphan member of the nuclear receptor superfamily closely related to the estrogen receptors. To explore the DNA binding characteristics, the protein-DNA interaction was studied in electrophoretic mobility shift assays (EMSAs). In vitro translated ERRgamma binds as a homodimer to direct repeats (DR) without spacing of the nuclear receptor half-site 5'-AGGTCA-3' (DR-0), to extended half-sites, and to the inverted estrogen response element. Using ERRgamma deletion constructs, binding was found to be dependent on the presence of sequences in the ligand binding domain (LBD). A far-Western analysis revealed that ERRgamma forms dimers even in the absence of DNA. Two elements, located in the hinge region and in the LBD, respectively, are necessary for DNA-independent dimerization. DNA binding of bacterial expressed ERRgamma requires additional factors present in the serum and in cellular extracts. Fusion proteins of the germ cell nuclear factor (GCNF/NR6A1) with ERRgamma showed that the characteristic feature to be stimulated by additional factors can be transferred to a heterologous protein. The stimulating activity was further characterized and its target sequence narrowed down to a small element in the hinge region. PMID:12180985

  3. Mast Cell Proteases 6 and 7 Stimulate Angiogenesis by Inducing Endothelial Cells to Release Angiogenic Factors

    PubMed Central

    de Souza, Devandir Antonio; Borges, Antonio Carlos; Santana, Ana Carolina; Oliver, Constance; Jamur, Maria Célia

    2015-01-01

    Mast cell proteases are thought to be involved with tumor progression and neo-vascularization. However, their exact role is still unclear. The present study was undertaken to further elucidate the function of specific subtypes of recombinant mouse mast cell proteases (rmMCP-6 and 7) in neo-vascularization. SVEC4-10 cells were cultured on Geltrex® with either rmMCP-6 or 7 and tube formation was analyzed by fluorescence microscopy and scanning electron microscopy. Additionally, the capacity of these proteases to induce the release of angiogenic factors and pro and anti-angiogenic proteins was analyzed. Both rmMCP-6 and 7 were able to stimulate tube formation. Scanning electron microscopy showed that incubation with the proteases induced SVEC4-10 cells to invade the gel matrix. However, the expression and activity of metalloproteases were not altered by incubation with the mast cell proteases. Furthermore, rmMCP-6 and rmMCP-7 were able to induce the differential release of angiogenic factors from the SVEC4-10 cells. rmMCP-7 was more efficient in stimulating tube formation and release of angiogenic factors than rmMCP-6. These results suggest that the subtypes of proteases released by mast cells may influence endothelial cells during in vivo neo-vascularization. PMID:26633538

  4. Platelet-derived growth factor (PDGF) stimulates glycogen synthase activity in 3T3 cells

    SciTech Connect

    Chan, C.P.; Bowen-Pope, D.F.; Ross, R.; Krebs, E.G.

    1986-05-01

    Hormonal regulation of glycogen synthase, an enzyme that can be phosphorylated on multiple sites, is often associated with changes in its phosphorylation state. Enzyme activation is conventionally monitored by determining the synthase activity ratio ((activity in the absence of glucose 6-P)/(activity in the presence of glucose 6-P)). Insulin causes an activation of glycogen synthase with a concomitant decrease in its phosphate content. In a previous report, the authors showed that epidermal growth factor (EGF) increases the glycogen synthase activity ratio in Swiss 3T3 cells. The time and dose-dependency of this response was similar to that of insulin. Their recent results indicate that PDGF also stimulates glycogen synthase activity. Enzyme activation was maximal after 30 min. of incubation with PDGF; the time course observed was very similar to that with insulin and EGF. At 1 ng/ml (0.03nM), PDGF caused a maximal stimulation of 4-fold in synthase activity ratio. Half-maximal stimulation was observed at 0.2 ng/ml (6 pM). The time course of changes in enzyme activity ratio closely followed that of /sup 125/I-PDGF binding. The authors data suggest that PDGF, as well as EFG and insulin, may be important in regulating glycogen synthesis through phosphorylation/dephosphorylation mechanisms.

  5. Inositol lipid metabolism in vasopressin stimulated hepatocytes from rats infused with tumor necrosis factor

    SciTech Connect

    Spitzer, J.A.; Rodriguez de Turco, E.B. )

    1989-05-30

    We studied the effect of i.v. infusion of human recombinant tumor necrosis factor alpha (rHuTNF alpha, Cetus, 15 micrograms/100 g bw over 3 h) on vasopressin (VP)-stimulated {sup 32}P-inositol lipid turnover and the release of {sup 3}H-inositol phosphates in isolated rat hepatocytes. The early VP-induced decrease (within 30 s) in {sup 32}P-phosphatidylinositol 4-phosphate and {sup 32}P-phosphatidylinositol 4,5-bisphosphate labeling was significantly reduced (-40%) and at the same time the uptake of {sup 32}P into phosphatidic acid was 50% lower than in saline-infused (matched control) rats. Within 5 min of VP-stimulation, lower {sup 32}P phosphatidylinositol (-40%) and higher {sup 32}P-phosphatidic acid (+30%) labeling were observed in rHuTNF alpha-infused rats. Infusion of rHuTNF alpha also affected the VP-induced release of {sup 3}H-inositol phosphates. The accumulation of {sup 3}H-inositol-labeled water soluble products was decreased by 25% and 17% at 30 s and 10 min, respectively. These data show that rHuTNF alpha mimics early perturbations induced by Escherichia coli endotoxin infusion in VP-stimulated inositol lipid metabolism in rat hepatocytes.

  6. Transforming growth factor-beta stimulates epithelial-mesenchymal transformation in the proepicardium.

    PubMed

    Olivey, Harold E; Mundell, Nathan A; Austin, Anita F; Barnett, Joey V

    2006-01-01

    The proepicardium (PE) migrates over the heart and forms the epicardium. A subset of these PE-derived cells undergoes epithelial-mesenchymal transformation (EMT) and gives rise to cardiac fibroblasts and components of the coronary vasculature. We report that transforming growth factor-beta (TGFbeta) 1 and TGFbeta2 increase EMT in PE explants as measured by invasion into a collagen gel, loss of cytokeratin expression, and redistribution of ZO1. The type I TGFbeta receptors ALK2 and ALK5 are both expressed in the PE. However, only constitutively active (ca) ALK2 stimulates PE-derived epithelial cell activation, the first step in transformation, whereas caALK5 stimulates neither activation nor transformation in PE explants. Overexpression of Smad6, an inhibitor of ALK2 signaling, inhibits epithelial cell activation, whereas BMP7, a known ligand for ALK2, has no effect. These data demonstrate that TGFbeta stimulates transformation in the PE and suggest that ALK2 partially mediates this effect. PMID:16245329

  7. Activation of Sterol Regulatory Element Binding Factors by Fenofibrate and Gemfibrozil Stimulates Myelination in Zebrafish

    PubMed Central

    Ashikawa, Yoshifumi; Nishimura, Yuhei; Okabe, Shiko; Sasagawa, Shota; Murakami, Soichiro; Yuge, Mizuki; Kawaguchi, Koki; Kawase, Reiko; Tanaka, Toshio

    2016-01-01

    Oligodendrocytes are major myelin-producing cells and play essential roles in the function of a healthy nervous system. However, they are also one of the most vulnerable neural cell types in the central nervous system (CNS), and myelin abnormalities in the CNS are found in a wide variety of neurological disorders, including multiple sclerosis, adrenoleukodystrophy, and schizophrenia. There is an urgent need to identify small molecular weight compounds that can stimulate myelination. In this study, we performed comparative transcriptome analysis to identify pharmacodynamic effects common to miconazole and clobetasol, which have been shown to stimulate myelination by mouse oligodendrocyte progenitor cells (OPCs). Of the genes differentially expressed in both miconazole- and clobetasol-treated mouse OPCs compared with untreated cells, we identified differentially expressed genes (DEGs) common to both drug treatments. Gene ontology analysis revealed that these DEGs are significantly associated with the sterol biosynthetic pathway, and further bioinformatics analysis suggested that sterol regulatory element binding factors (SREBFs) might be key upstream regulators of the DEGs. In silico screening of a public database for chemicals associated with SREBF activation identified fenofibrate, a peroxisome proliferator-activated receptor α (PPARα) agonist, as a drug that increases the expression of known SREBF targets, raising the possibility that fenofibrate may also stimulate myelination. To test this, we performed in vivo imaging of zebrafish expressing a fluorescent reporter protein under the control of the myelin basic protein (mbp) promoter. Treatment of zebrafish with fenofibrate significantly increased expression of the fluorescent reporter compared with untreated zebrafish. This increase was attenuated by co-treatment with fatostatin, a specific inhibitor of SREBFs, confirming that the fenofibrate effect was mediated via SREBFs. Furthermore, incubation of zebrafish

  8. Exploring bikeability in a metropolitan setting: stimulating and hindering factors in commuting route environments

    PubMed Central

    2012-01-01

    Background Route environments may influence people's active commuting positively and thereby contribute to public health. Assessments of route environments are, however, needed in order to better understand the possible relationship between active commuting and the route environment. The aim of this study was, therefore, to assess the potential associations between perceptions of whether the route environment on the whole hinders or stimulates bicycle commuting and perceptions of environmental factors. Methods The Active Commuting Route Environment Scale (ACRES) was used for the assessment of bicycle commuters' perceptions of their route environments in the inner urban parts of Greater Stockholm, Sweden. Bicycle commuters (n = 827) were recruited by advertisements in newspapers. Simultaneous multiple regression analyses were used to assess the relation between predictor variables (such as levels of exhaust fumes, noise, traffic speed, traffic congestion and greenery) and the outcome variable (hindering - stimulating route environments). Two models were run, (Model 1) without and (Model 2) with the item traffic: unsafe or safe included as a predictor. Results Overall, about 40% of the variance of hindering - stimulating route environments was explained by the environmental predictors in our models (Model 1, R2 = 0.415, and Model 2, R 2= 0.435). The regression equation for Model 1 was: y = 8.53 + 0.33 ugly or beautiful + 0.14 greenery + (-0.14) course of the route + (-0.13) exhaust fumes + (-0.09) congestion: all types of vehicles (p ≤ 0.019). The regression equation for Model 2 was y = 6.55 + 0.31 ugly or beautiful + 0.16 traffic: unsafe or safe + (-0.13) exhaust fumes + 0.12 greenery + (-0.12) course of the route (p ≤ 0.001). Conclusions The main results indicate that beautiful, green and safe route environments seem to be, independently of each other, stimulating factors for bicycle commuting in inner urban areas. On the other hand, exhaust fumes, traffic

  9. Hepatoma-derived growth factor stimulates smooth muscle cell growth and is expressed in vascular development

    PubMed Central

    Everett, Allen D.; Lobe, David R.; Matsumura, Martin E.; Nakamura, Hideji; McNamara, Coleen A.

    2000-01-01

    Hepatoma-derived growth factor (HDGF) is the first member identified of a new family of secreted heparin-binding growth factors highly expressed in the fetal aorta. The biologic role of HDGF in vascular growth is unknown. Here, we demonstrate that HDGF mRNA is expressed in smooth muscle cells (SMCs), most prominently in proliferating SMCs, 8–24 hours after serum stimulation. Exogenous HDGF and endogenous overexpression of HDGF stimulated a significant increase in SMC number and DNA synthesis. Rat aortic SMCs transfected with a hemagglutinin-epitope–tagged rat HDGF cDNA contain HA-HDGF in their nuclei during S-phase. We also detected native HDGF in nuclei of cultured SMCs, of SMCs and endothelial cells from 19-day fetal (but not in the adult) rat aorta, of SMCs proximal to abdominal aortic constriction in adult rats, and of SMCs in the neointima formed after endothelial denudation of the rat common carotid artery. Moreover, HDGF colocalizes with the proliferating cell nuclear antigen (PCNA) in SMCs in human atherosclerotic carotid arteries, suggesting that HDGF helps regulate SMC growth during development and in response to vascular injury. PMID:10712428

  10. Stimulation of body weight increase and epiphyseal cartilage growth by insulin like growth factor

    NASA Technical Reports Server (NTRS)

    Ellis, S.

    1981-01-01

    The ability of insulin-like growth factor (IGF) to induce growth in hypophysectomized immature rats was tested by continuous infusion of the partially purified factor at daily doses of 6, 21, and 46 mU for an 8-day period. A dose-dependent growth of the proximal epiphyseal cartilage of the tibia and an associated stimulation of the primary spongiosa were produced by these amounts of IGF. The two highest doses of IGF also resulted in dose-dependent increases of body weight. Gel permeation of the sera at neutrality showed that the large-molecular-weight IGF binding protein was not induced by the infusion of IGF, whereas it ws generated in the sera of hypophysectomized rats that were infused with daily doses of 86 mU of human growth hormone.

  11. Immunosuppressive drugs prevent a rapid dephosphorylation of transcription factor NFAT1 in stimulated immune cells.

    PubMed Central

    Shaw, K T; Ho, A M; Raghavan, A; Kim, J; Jain, J; Park, J; Sharma, S; Rao, A; Hogan, P G

    1995-01-01

    The immunosuppressive drugs cyclosporin A and FK506 interfere with the inducible transcription of cytokine genes in T cells and in other immune cells, in part by preventing the activation of NF-AT (nuclear factor of activated T cells). We show that transcription factor NFAT1 in T cells is rapidly dephosphorylated on stimulation, that dephosphorylation occurs before translocation of NFAT1 into the cell nucleus, and that dephosphorylation increases the affinity of NFAT1 for its specific sites in DNA. Cyclosporin A prevents the dephosphorylation and the nuclear translocation of NFAT1 in T cells, B cells, macrophages, and mast cells, delineating at least one mechanism that contributes to the profound immunosuppressive effects of this compound. Images Fig. 1 Fig. 2 Fig. 3 Fig. 4 PMID:7479966

  12. Multiple Growth Factors, But Not VEGF, Stimulate Glycosaminoglycan Hyperelongation in Retinal Choroidal Endothelial Cells

    PubMed Central

    Al Gwairi, Othman; Osman, Narin; Getachew, Robel; Zheng, Wenhua; Liang, X-L.; Kamato, Danielle; Thach, Lyna; Little, Peter J.

    2016-01-01

    A major feature of early age-related macular degeneration (AMD) is the thickening of Bruch's membrane in the retina and an alteration in its composition with increased lipid deposition. In certain pathological conditions proteoglycans are responsible for lipid retention in tissues. Growth factors are known to increase the length of glycosaminoglycan chains and this can lead to a large increase in the interaction between proteoglycans and lipids. Using choroidal endothelial cells, we investigated the effects of a number of AMD relevant growth factors TGFβ, thrombin, PDGF, IGF and VEGF on proteoglycan synthesis. Cells were characterized as of endothelial origin using the specific cell markers endothelial nitric oxide synthesis and von Willebrand factor and imaged using confocal microscopy. Cells were treated with growth factors in the presence and absence of the appropriate inhibitors and were radiolabeled with [35S]-SO4. Proteoglycans were isolated by ion exchange chromatography and sized using SDS-PAGE. Radiosulfate incorporation was determined by the cetylpyridinium chloride (CPC) precipitation technique. To measure cellular glycosaminoglycan synthesizing capacity we added xyloside and assessed the xyloside-GAGs by SDS-PAGE. TGFβ, thrombin, PDGF & IGF dose-dependently stimulated radiosulfate incorporation and GAG elongation as well as xyloside-GAG synthesis, however VEGF treatment did not stimulate any changes in proteoglycan synthesis. VEGF did not increase pAKT but caused a large increase in pERK relative to the response to PDGF. Thus, AMD relevant agonists cause glycosaminoglycan hyperelongation of proteoglycans synthesised and secreted by retinal choroidal endothelial cells. The absence of a response to VEGF is intriguing and identifies proteoglycans as a novel potential target in AMD. Future studies will examine the relevance of these changes to enhanced lipid binding and the development of AMD. PMID:27570478

  13. Palmitoleate is a mitogen, formed upon stimulation with growth factors, and converted to palmitoleoyl-phosphatidylinositol.

    PubMed

    Koeberle, Andreas; Shindou, Hideo; Harayama, Takeshi; Shimizu, Takao

    2012-08-01

    Controversial correlations between biological activity and concentration of the novel lipokine palmitoleate (9Z-hexadecenoate, 16:1) might depend on the formation of an active 16:1 metabolite. For its identification, we analyzed the glycerophospholipid composition of mouse Swiss 3T3 fibroblasts in response to 16:1 using LC-MS/MS. 16:1 was either supplemented to the cell culture medium or endogenously formed when cells were stimulated with insulin or growth factors as suggested by the enhanced mRNA expression of 16:1-biosynthetic enzymes. The proportion of 1-acyl-2-16:1-sn-phosphatidylinositol (16:1-PI) was time-dependently and specifically increased relative to other glycerophospholipids under both conditions and correlated with the proliferation of fatty acid (16:1, palmitate, oleate, or arachidonate)-supplemented cells. Accordingly, cell proliferation was impaired by blocking 16:1 biosynthesis using the selective stearoyl-CoA desaturase-1 inhibitor CAY10566 and restored by supplementation of 16:1. The accumulation of 16:1-PI occurred throughout cellular compartments and within diverse mouse cell lines (Swiss 3T3, NIH-3T3, and 3T3-L1 cells). To elucidate further whether 16:1-PI is formed through the de novo or remodeling pathway of PI biosynthesis, phosphatidate levels and lyso-PI-acyltransferase activities were analyzed as respective markers. The proportion of 16:1-phosphatidate was significantly increased by insulin and growth factors, whereas lyso-PI-acyltransferases showed negligible activity for 16:1-coenzyme A. The relevance of the de novo pathway for 16:1-PI biosynthesis is supported further by the comparable incorporation rate of deuterium-labeled 16:1 and tritium-labeled inositol into PI for growth factor-stimulated cells. In conclusion, we identified 16:1 or 16:1-PI as mitogen whose biosynthesis is induced by growth factors. PMID:22700983

  14. Multiple Growth Factors, But Not VEGF, Stimulate Glycosaminoglycan Hyperelongation in Retinal Choroidal Endothelial Cells.

    PubMed

    Al Gwairi, Othman; Osman, Narin; Getachew, Robel; Zheng, Wenhua; Liang, X-L; Kamato, Danielle; Thach, Lyna; Little, Peter J

    2016-01-01

    A major feature of early age-related macular degeneration (AMD) is the thickening of Bruch's membrane in the retina and an alteration in its composition with increased lipid deposition. In certain pathological conditions proteoglycans are responsible for lipid retention in tissues. Growth factors are known to increase the length of glycosaminoglycan chains and this can lead to a large increase in the interaction between proteoglycans and lipids. Using choroidal endothelial cells, we investigated the effects of a number of AMD relevant growth factors TGFβ, thrombin, PDGF, IGF and VEGF on proteoglycan synthesis. Cells were characterized as of endothelial origin using the specific cell markers endothelial nitric oxide synthesis and von Willebrand factor and imaged using confocal microscopy. Cells were treated with growth factors in the presence and absence of the appropriate inhibitors and were radiolabeled with [35S]-SO4. Proteoglycans were isolated by ion exchange chromatography and sized using SDS-PAGE. Radiosulfate incorporation was determined by the cetylpyridinium chloride (CPC) precipitation technique. To measure cellular glycosaminoglycan synthesizing capacity we added xyloside and assessed the xyloside-GAGs by SDS-PAGE. TGFβ, thrombin, PDGF & IGF dose-dependently stimulated radiosulfate incorporation and GAG elongation as well as xyloside-GAG synthesis, however VEGF treatment did not stimulate any changes in proteoglycan synthesis. VEGF did not increase pAKT but caused a large increase in pERK relative to the response to PDGF. Thus, AMD relevant agonists cause glycosaminoglycan hyperelongation of proteoglycans synthesised and secreted by retinal choroidal endothelial cells. The absence of a response to VEGF is intriguing and identifies proteoglycans as a novel potential target in AMD. Future studies will examine the relevance of these changes to enhanced lipid binding and the development of AMD. PMID:27570478

  15. Angiotensin II stimulated expression of transforming growth factor-beta1 in cardiac fibroblasts and myofibroblasts.

    PubMed

    Campbell, S E; Katwa, L C

    1997-07-01

    Angiotensin II (Ang II) stimulates pathologic myocardial fibrosis. Cardiac fibroblasts (CFb) and myofibroblasts mediate this response, perhaps in part by indirect production of specific cytokines. We sought to determine if Ang II could stimulate transforming growth factor-beta1 (TGF-beta1) gene expression and protein production in adult rat CFb and two cardiac myofibroblast cell types, scar myofibroblasts (MyoFb) and valvular interstitial cells (VIC). Confluent CFb, MyoFb, and VIC in serum-deprived (0.4% FCS) media were treated with Ang II (10(-7) m for CFb; 10(-9) m for MyoFb, VIC) for 24 h. Untreated cells served as controls. Culture media was collected and TGF-beta1 levels determined in triplicate using a sandwich ELISA. Reverse transcriptase-polymerase chain reaction (RT-PCR) analysis was performed to determine TGF-beta1 mRNA expression. Ang II increased CFb (P<0.02) and VIC (P<0.04) TGF-beta1 mRNA expression, while the increase in MyoFb was not statistically significant. MyoFb produced the highest TGF-beta1 levels under control conditions compared to VIC and CFb. Ang II stimulated further TGF-beta1 secretion in VIC and CFb, but not MyoFb. The AT1 receptor antagonist Losartan (10(-7) m) greatly attenuated Ang II-stimulated TGF-B1 secretion and decreased TGF-beta1 immunostaining in VIC. The AT2 receptor antagonist PD123177 (10(-7) m) also decreased secretion and immunostaining of TGF-beta1 in VIC, but to a lesser extent than Losartan. TGF-beta1 secretion by MyoFb was unaffected by Losartan and PD123177, although TGF-B1 immunostaining was absent or greatly decreased, respectively, compared to Ang II-treated MyoFb. Ang II stimulates TGF-beta1 gene expression and/or protein production in cardiac fibroblast-like cells which may act as an autocrine/paracrine stimulus to collagen formation. Furthermore, TGF-beta1 production and secretion in these cells can be modulated by specific Ang II receptor antagonists, suggesting a potential benefit in preventing

  16. Long-Term Expansion in Platelet Lysate Increases Growth of Peripheral Blood-Derived Endothelial-Colony Forming Cells and Their Growth Factor-Induced Sprouting Capacity

    PubMed Central

    Tasev, Dimitar; van Wijhe, Michiel H.; Weijers, Ester M.; van Hinsbergh, Victor W. M.; Koolwijk, Pieter

    2015-01-01

    Introduction Efficient implementation of peripheral blood-derived endothelial-colony cells (PB-ECFCs) as a therapeutical tool requires isolation and generation of a sufficient number of cells in ex vivo conditions devoid of animal-derived products. At present, little is known how the isolation and expansion procedure in xenogeneic-free conditions affects the therapeutical capacity of PB-ECFCs. Results The findings presented in this study indicate that human platelet lysate (PL) as a serum substitute yields twice more colonies per mL blood compared to the conventional isolation with fetal bovine serum (FBS). Isolated ECFCs displayed a higher proliferative ability in PL supplemented medium than cells in FBS medium during 30 days expansion. The cells at 18 cumulative population doubling levels (CPDL) retained their proliferative capacity, showed higher sprouting ability in fibrin matrices upon stimulation with FGF-2 and VEGF-A than the cells at 6 CPDL, and displayed low β-galactosidase activity. The increased sprouting of PB-ECFCs at 18 CPDL was accompanied by an intrinsic activation of the uPA/uPAR fibrinolytic system. Induced deficiency of uPA (urokinase-type plasminogen activator) or uPAR (uPA receptor) by siRNA technology completely abolished the angiogenic ability of PB-ECFCs in fibrin matrices. During the serial expansion, the gene induction of the markers associated with inflammatory activation such as VCAM-1 and ICAM-1 did not occur or only to limited extent. While further propagation up to 31 CPDL proceeded at a comparable rate, a marked upregulation of inflammatory markers occurred in all donors accompanied by a further increase of uPA/uPAR gene induction. The observed induction of inflammatory genes at later stages of long-term propagation of PB-ECFCs underpins the necessity to determine the right time-point for harvesting of sufficient number of cells with preserved therapeutical potential. Conclusion The presented isolation method and subsequent cell

  17. Ability of Post-endotoxin serum from BCG-infected mice to induce nonspecific resistance and stimulation of granulopoiesis.

    PubMed Central

    Urbaschek, R; Urbaschek, B

    1983-01-01

    Serum from BCG-infected mice obtained 2 h after injection with endotoxin induced elevated levels of colony-stimulating factor and an increase in splenic granulocyte-macrophage progenitor cells in C3H/HeJ mice. The capacity of such serum to stimulate granulopoiesis may be related to its ability to increase nonspecific resistance to lethal irradiation. PMID:6341239

  18. BEEHAVE: a systems model of honeybee colony dynamics and foraging to explore multifactorial causes of colony failure

    PubMed Central

    Becher, Matthias A; Grimm, Volker; Thorbek, Pernille; Horn, Juliane; Kennedy, Peter J; Osborne, Juliet L

    2014-01-01

    A notable increase in failure of managed European honeybee Apis mellifera L. colonies has been reported in various regions in recent years. Although the underlying causes remain unclear, it is likely that a combination of stressors act together, particularly varroa mites and other pathogens, forage availability and potentially pesticides. It is experimentally challenging to address causality at the colony scale when multiple factors interact. In silico experiments offer a fast and cost-effective way to begin to address these challenges and inform experiments. However, none of the published bee models combine colony dynamics with foraging patterns and varroa dynamics. We have developed a honeybee model, BEEHAVE, which integrates colony dynamics, population dynamics of the varroa mite, epidemiology of varroa-transmitted viruses and allows foragers in an agent-based foraging model to collect food from a representation of a spatially explicit landscape. We describe the model, which is freely available online (www.beehave-model.net). Extensive sensitivity analyses and tests illustrate the model's robustness and realism. Simulation experiments with various combinations of stressors demonstrate, in simplified landscape settings, the model's potential: predicting colony dynamics and potential losses with and without varroa mites under different foraging conditions and under pesticide application. We also show how mitigation measures can be tested. Synthesis and applications. BEEHAVE offers a valuable tool for researchers to design and focus field experiments, for regulators to explore the relative importance of stressors to devise management and policy advice and for beekeepers to understand and predict varroa dynamics and effects of management interventions. We expect that scientists and stakeholders will find a variety of applications for BEEHAVE, stimulating further model development and the possible inclusion of other stressors of potential importance to honeybee

  19. Modulation of cultured porcine granulosa cell responsiveness to follicle stimulating hormone and epidermal growth factor

    SciTech Connect

    Buck, P.A.

    1986-01-01

    Ovarian follicular development is dependent upon the coordinated growth and differentiation of the granulosa cells which line the follicle. Follicle stimulating hormone (FSH) induces granulosa cell differentiation both in vivo and in vitro. Epidermal growth factor (EGF) stimulates granulosa cell proliferation in vitro. The interaction of these two effectors upon selected parameters of growth and differentiation was examined in monolayer cultures of porcine granulose cells. Analysis of the EGF receptor by /sup 125/I-EGF binding revealed that the receptor was of high affinity with an apparent dissociation constant of 4-6 x 10/sup -10/ M. The average number of receptors per cell varied with the state of differentiation both in vivo and in vitro; highly differentiated cells bound two-fold less /sup 125/I-EGF and this effect was at least partially induced by FSH in vitro. EGF receptor function was examined by assessing EGF effects on cell number and /sup 3/H-thymidine incorporation. EGF stimulated thymidine incorporation in both serum-free and serum-supplemented culture, but only in serum-supplemented conditions was cell number increased. EGF receptor function was inversely related to the state of differentiation and was attenuated by FSH. The FSH receptor was examined by /sup 125/I-FSH binding. EGF increased FSH receptor number, and lowered the affinity of the receptor. The function of these receptors was assessed by /sup 125/I-hCG binding and progesterone radioimmunoassay. If EGF was present continuously in the cultures. FSH receptor function was attenuated regardless of FSH receptor number. A preliminary effort to examine the mechanism of this interaction was performed by analyzing hormonally controlled protein synthesis with /sup 35/S-methionine labeling, SDS polyacrylamide gel electrophoresis and fluorography. FSH promoted the expression of a 27,000 dalton protein. This effect was attenuated by EGF.

  20. The effect of diet on tumor necrosis factor stimulation of hepatic lipogenesis

    SciTech Connect

    Feingold, K.R.; Soued, M.; Serio, M.K.; Adi, S.; Moser, A.H.; Grunfeld, C. )

    1990-06-01

    In this study, we determined the effects of tumor necrosis factor (TNF) on serum lipid levels and hepatic lipid synthesis in animals whose diets and feeding conditions were varied to induce changes in baseline serum lipid levels and/or rates of hepatic lipid synthesis. In animals studied at both the nadir and peak of the diurnal cycle of hepatic lipid synthesis, TNF acutely increases serum triglyceride levels, stimulates hepatic fatty acid synthesis, and increases the quantity of newly synthesized fatty acids found in the serum. Similarly, in animals ingesting either high-sucrose or cholesterol-enriched diets, TNF induces the characteristic rapid increase in serum triglyceride levels, hepatic fatty acid synthesis, and quantity of labeled fatty acids in the serum. In animals fed a diet high in triglycerides, using either corn oil or lard, TNF stimulates hepatic fatty acid synthesis and increases the quantity of newly synthesized fatty acids in the serum, but serum triglyceride levels do not change. However, TNF inhibits gastric emptying, which results in a marked decrease in fat absorption in TNF-treated animals. It is likely that a decrease in the dietary contribution to serum triglyceride levels during high-triglyceride feeding counterbalances the increased hepatic contribution induced by TNF treatment. In animals fasted before TNF administration there was no acute change in either serum lipid levels, hepatic fatty acid synthesis, or the quantity of labeled fatty acids in the serum. Thus, TNF stimulates hepatic fatty acid synthesis and increases serum triglyceride levels under many diverse dietary conditions, suggesting that there is a strong linkage between the immune system and lipid metabolism that is independent of most dietary manipulations and may be of fundamental importance in the body's response to infection.

  1. Phosphorylation and Activation of RhoA by ERK in Response to Epidermal Growth Factor Stimulation.

    PubMed

    Tong, Junfeng; Li, Laiji; Ballermann, Barbara; Wang, Zhixiang

    2016-01-01

    The small GTPase RhoA has been implicated in various cellular activities, including the formation of stress fibers, cell motility, and cytokinesis. In addition to the canonical GTPase cycle, recent findings have suggested that phosphorylation further contributes to the tight regulation of Rho GTPases. Indeed, RhoA is phosphorylated on serine 188 (188S) by a number of protein kinases. We have recently reported that Rac1 is phosphorylated on threonine 108 (108T) by extracellular signal-regulated kinases (ERK) in response to epidermal growth factor (EGF) stimulation. Here, we provide evidence that RhoA is phosphorylated by ERK on 88S and 100T in response to EGF stimulation. We show that ERK interacts with RhoA and that this interaction is dependent on the ERK docking site (D-site) at the C-terminus of RhoA. EGF stimulation enhanced the activation of the endogenous RhoA. The phosphomimetic mutant, GFP-RhoA S88E/T100E, when transiently expressed in COS-7 cells, displayed higher GTP-binding than wild type RhoA. Moreover, the expression of GFP-RhoA S88E/T100E increased actin stress fiber formation in COS-7 cells, which is consistent with its higher activity. In contrast to Rac1, phosphorylation of RhoA by ERK does not target RhoA to the nucleus. Finally, we show that regardless of the phosphorylation status of RhoA and Rac1, substitution of the RhoA PBR with the Rac1 PBR targets RhoA to the nucleus and substitution of Rac1 PBR with RhoA PBR significantly reduces the nuclear localization of Rac1. In conclusion, ERK phosphorylates RhoA on 88S and 100T in response to EGF, which upregulates RhoA activity. PMID:26816343

  2. Phosphorylation and Activation of RhoA by ERK in Response to Epidermal Growth Factor Stimulation

    PubMed Central

    Tong, Junfeng; Li, Laiji; Ballermann, Barbara; Wang, Zhixiang

    2016-01-01

    The small GTPase RhoA has been implicated in various cellular activities, including the formation of stress fibers, cell motility, and cytokinesis. In addition to the canonical GTPase cycle, recent findings have suggested that phosphorylation further contributes to the tight regulation of Rho GTPases. Indeed, RhoA is phosphorylated on serine 188 (188S) by a number of protein kinases. We have recently reported that Rac1 is phosphorylated on threonine 108 (108T) by extracellular signal-regulated kinases (ERK) in response to epidermal growth factor (EGF) stimulation. Here, we provide evidence that RhoA is phosphorylated by ERK on 88S and 100T in response to EGF stimulation. We show that ERK interacts with RhoA and that this interaction is dependent on the ERK docking site (D-site) at the C-terminus of RhoA. EGF stimulation enhanced the activation of the endogenous RhoA. The phosphomimetic mutant, GFP-RhoA S88E/T100E, when transiently expressed in COS-7 cells, displayed higher GTP-binding than wild type RhoA. Moreover, the expression of GFP-RhoA S88E/T100E increased actin stress fiber formation in COS-7 cells, which is consistent with its higher activity. In contrast to Rac1, phosphorylation of RhoA by ERK does not target RhoA to the nucleus. Finally, we show that regardless of the phosphorylation status of RhoA and Rac1, substitution of the RhoA PBR with the Rac1 PBR targets RhoA to the nucleus and substitution of Rac1 PBR with RhoA PBR significantly reduces the nuclear localization of Rac1. In conclusion, ERK phosphorylates RhoA on 88S and 100T in response to EGF, which upregulates RhoA activity. PMID:26816343

  3. Growth differentiation factor 15 stimulates rapamycin-sensitive ovarian cancer cell growth and invasion.

    PubMed

    Griner, Samantha E; Joshi, Jayashree P; Nahta, Rita

    2013-01-01

    Identification of novel molecular markers and therapeutic targets may improve survival rates for patients with ovarian cancer. In the current study, immunohistochemical (IHC) analysis of two human ovarian tumor tissue arrays showed high staining for GDF15 in a majority of tissues. Exogenous stimulation of ovarian cancer cell lines with recombinant human GDF15 (rhGDF15) or stable over-expression of a GDF15 expression plasmid promoted anchorage-independent growth, increased invasion, and up-regulation of matrix metalloproteinases (MMPs) and vascular endothelial growth factor (VEGF). MMP inhibition suppressed GDF15-mediated invasion. In addition, IHC analysis of human ovarian tumor tissue arrays indicated that GDF15 expression correlated significantly with high MMP2 and MMP9 expression. Exogenous and endogenous GDF15 over-expression stimulated phosphorylation of p38, Erk1/2, and Akt. Pharmacologic inhibition of p38, MEK, or PI3K suppressed GDF15-stimulated growth. Further, proliferation, growth, and invasion of GDF15 stable clones were blocked by rapamycin. IHC analysis demonstrated significant correlation between GDF15 expression and phosphorylation of mTOR. Finally, knockdown of endogenous GDF15 or neutralization of secreted GDF15 suppressed invasion and growth of a GDF15-over-expressing ovarian cancer cell line. These data indicate that GDF15 over-expression, which occurred in a majority of human ovarian cancers, promoted rapamycin-sensitive invasion and growth of ovarian cancer cells. Inhibition of mTOR may be an effective therapeutic strategy for ovarian cancers that over-express GDF15. Future studies should examine GDF15 as a novel molecular target for blocking ovarian cancer progression. PMID:23085437

  4. Hepatic transforming growth factor-β 1 stimulated clone-22 D1 controls systemic cholesterol metabolism☆

    PubMed Central

    Jäger, Julia; Greiner, Vera; Strzoda, Daniela; Seibert, Oksana; Niopek, Katharina; Sijmonsma, Tjeerd P.; Schäfer, Michaela; Jones, Allan; De Guia, Roldan; Martignoni, Marc; Dallinga-Thie, Geesje M.; Diaz, Mauricio B.; Hofmann, Thomas G.; Herzig, Stephan

    2014-01-01

    Disturbances in lipid homeostasis are hallmarks of severe metabolic disorders and their long-term complications, including obesity, diabetes, and atherosclerosis. Whereas elevation of triglyceride (TG)-rich very-low-density lipoproteins (VLDL) has been identified as a risk factor for cardiovascular complications, high-density lipoprotein (HDL)-associated cholesterol confers atheroprotection under obese and/or diabetic conditions. Here we show that hepatocyte-specific deficiency of transcription factor transforming growth factor β 1-stimulated clone (TSC) 22 D1 led to a substantial reduction in HDL levels in both wild-type and obese mice, mediated through the transcriptional down-regulation of the HDL formation pathway in liver. Indeed, overexpression of TSC22D1 promoted high levels of HDL cholesterol in healthy animals, and hepatic expression of TSC22D1 was found to be aberrantly regulated in disease models of opposing energy availability. The hepatic TSC22D1 transcription factor complex may thus represent an attractive target in HDL raising strategies in obesity/diabetes-related dyslipidemia and atheroprotection. PMID:24634828

  5. Sox9 is a β-catenin-regulated transcription factor that enhances the colony-forming activity of squamous cell carcinoma cells.

    PubMed

    Li, Xue Mei; Piao, Yong Jun; Sohn, Kyung-Cheol; Ha, Jeong-Min; Im, Myung; Seo, Young-Joon; Whang, Kyu Uang; Lee, Jeung-Hoon; Lee, Young; Kim, Chang Deok

    2016-07-01

    Squamous cell carcinoma (SCC) is a common skin cancer, of which the incidence is relatively high, ranking second among the non‑melanoma skin cancers. It is known that numerous intracellular signal regulators are involved in the pathogenesis of SCC. The Wnt/β-catenin signaling pathway serves an important role in cancer development. However, the downstream effectors of β‑catenin remain to be clearly elucidated yet. The present study investigated the functional importance of Wnt/β‑catenin signaling in cutaneous SCC. β‑catenin expression was reduced using recombinant adenovirus expressing specific microRNA (miR). Knockdown of β‑catenin resulted in a marked reduction of the colony-forming activity of the SCC cells, SCC12. In an attempt to identify the β‑catenin downstream genes, it was found that Sox9 was regulated by β‑catenin in SCC12 cells. Overexpression of a constitutively active form of β‑catenin led to the induction of Sox9, while knockdown of β‑catenin resulted in downregulation of Sox9. When the expression of Sox9 was reduced using specific miR, colony-forming activity of the SCC12 cells was significantly reduced. When Sox9 was overexpressed in cells where β‑catenin was knocked down, it partially restored the colony‑forming potential. Taken together, the present results suggested that Sox9 is a β-catenin downstream transcription factor and is positively involved in SCC development. PMID:27151141

  6. Increase in Bacterial Colony Formation from a Permafrost Ice Wedge Dosed with a Tomitella biformata Recombinant Resuscitation-Promoting Factor Protein.

    PubMed

    Puspita, Indun Dewi; Kitagawa, Wataru; Kamagata, Yoichi; Tanaka, Michiko; Nakatsu, Cindy H

    2015-01-01

    Resuscitation-promoting factor (Rpf) is a protein that has been found in a number of different Actinobacteria species and has been shown to promote the growth of active cells and resuscitate dormant (non-dividing) cells. We previously reported the biological activity of an Rpf protein in Tomitella biformata AHU 1821(T), an Actinobacteria isolated from a permafrost ice wedge. This protein is excreted outside the cell; however, few studies have investigated its contribution in environmental samples to the growth or resuscitation of bacteria other than the original host. Therefore, the aim of the present study was to determine whether Rpf from T. biformata impacted the cultivation of other bacteria from the permafrost ice wedge from which it was originally isolated. All experiments used recombinant Rpf proteins produced using a Rhodococcus erythropolis expression system. Dilutions of melted surface sterilized ice wedge samples mixed with different doses of the purified recombinant Rpf (rRpf) protein indicated that the highest concentration tested, 1250 pM, had a significantly (p <0.05) higher number of CFUs on agar plates after 8 d, approximately 14-fold higher than that on control plates without rRpf. 16S rRNA gene sequences revealed that all the colonies on plates were mainly related to Brevibacterium antiquum strain VKM Ac-2118 (AY243344), with 98-99% sequence identity. This species is also a member of the phylum Actinobacteria and was originally isolated from Siberian permafrost sediments. The results of the present study demonstrated that rRpf not only promoted the growth of T. biformata from which it was isolated, but also enhanced colony formation by another Actinobacteria in an environmental sample. PMID:25843055

  7. Enhanced jun gene expression is an early genomic response to transforming growth factor. beta. stimulation

    SciTech Connect

    Pertovaara, L.; Sistonen, L.; Keski-Oja, J.; Alitalo, K. ); Bos, T.J.; Vogt, P.K. . Dept. of Microbiology)

    1989-03-01

    Transforming growth factor {beta} (TGF{beta}) is a multifunctional polypeptide4 that regulates proliferation, differentiation, and other functions of many cell types. The pathway of TGF{beta} signal transduction in cells is unknown. The authors report here that an early effect of TGF{beta} is an enhancement of the expression of two genes encoding serum- and phorbol ester tumor promoter-regulated transcription factors: the junB gene and the c-jun proto-oncogene, respectively. This stimulation was observed in human lung adenocarcinoma A549 cells which were growth inhibited by TGF{beta}, AKR-2B mouse embryo fibroblasts which were growth stimulated by TGF{beta}, and K562 human erythroleukemia cells, which were not appreciably affected in their growth by TFG{beta}. The increase in jun mRNA occurred with picomolar TGF{beta} concentrations within 1 h of TGF{beta} stimulation, reached a peak between 1 and 5 h in different cells, and declined gradually to base-fine levels. This mRNA response was followed by a large increase in the biosynthesis of the c-jun protein (AP-1), as shown by metabolic labeling and immunoprecipitation analysis. However, differential and cell type-specific regulation appeared to determine the timing and magnitude of the response of each jun gene in a given cell. In AKR-2B and NIH 3T3 cells, only junB was induced by TGF{beta}, evidently in a protein synthesis-independent fashion. The junB response to TGF{beta} was maintained in c-Ha-ras and neu oncogene-transformed cells. Thus, one of the earliest genomic responses to TGF{beta} may involve nuclear signal transduction and amplification by the junB and c-jun transcription factors in concert with c-fos, which is also induced. The differential activation of the jun genes may explain some of the pleiotropic effects of TGF{beta}.

  8. Pancreatic adenocarcinoma upregulated factor serves as adjuvant by activating dendritic cells through stimulation of TLR4.

    PubMed

    Kang, Tae Heung; Kim, Young Seob; Kim, Seokho; Yang, Benjamin; Lee, Je-Jung; Lee, Hyun-Ju; Lee, Jaemin; Jung, In Duk; Han, Hee Dong; Lee, Seung-Hyun; Koh, Sang Seok; Wu, T-C; Park, Yeong-Min

    2015-09-29

    Dendritic cell (DC) based cancer vaccines represent a promising immunotherapeutic strategy against cancer. To enhance the modest immunogenicity of DC vaccines, various adjuvants are often incorporated. Particularly, most of the common adjuvants are derived from bacteria. In the current study, we evaluate the use of a human pancreatic cancer derived protein, pancreatic adenocarcinoma upregulated factor (PAUF), as a novel DC vaccine adjuvant. We show that PAUF can induce activation and maturation of DCs and activate NFkB by stimulating the Toll-like receptor signaling pathway. Furthermore, vaccination with PAUF treated DCs pulsed with E7 or OVA peptides leads to generation of E7 or OVA-specific CD8+ T cells and memory T cells, which correlate with long term tumor protection and antitumor effects against TC-1 and EG.7 tumors in mice. Finally, we demonstrated that PAUF mediated DC activation and immune stimulation are dependent on TLR4. Our data provides evidence supporting PAUF as a promising adjuvant for DC based therapies, which can be applied in conjunction with other cancer therapies. Most importantly, our results serve as a reference for future investigation of human based adjuvants. PMID:26336989

  9. The contribution of interindividual factors to variability of response in transcranial direct current stimulation studies

    PubMed Central

    Li, Lucia M.; Uehara, Kazumasa; Hanakawa, Takashi

    2015-01-01

    There has been an explosion of research using transcranial direct current stimulation (tDCS) for investigating and modulating human cognitive and motor function in healthy populations. It has also been used in many studies seeking to improve deficits in disease populations. With the slew of studies reporting “promising results” for everything from motor recovery after stroke to boosting memory function, one could be easily seduced by the idea of tDCS being the next panacea for all neurological ills. However, huge variability exists in the reported effects of tDCS, with great variability in the effect sizes and even contradictory results reported. In this review, we consider the interindividual factors that may contribute to this variability. In particular, we discuss the importance of baseline neuronal state and features, anatomy, age and the inherent variability in the injured brain. We additionally consider how interindividual variability affects the results of motor-evoked potential (MEP) testing with transcranial magnetic stimulation (TMS), which, in turn, can lead to apparent variability in response to tDCS in motor studies. PMID:26029052

  10. Insulin-like growth factor-1 stimulates regulatory T cells and suppresses autoimmune disease

    PubMed Central

    Bilbao, Daniel; Luciani, Luisa; Johannesson, Bjarki; Piszczek, Agnieszka; Rosenthal, Nadia

    2014-01-01

    The recent precipitous rise in autoimmune diseases is placing an increasing clinical and economic burden on health systems worldwide. Current therapies are only moderately efficacious, often coupled with adverse side effects. Here, we show that recombinant human insulin-like growth factor-1 (rhIGF-1) stimulates proliferation of both human and mouse regulatory T (Treg) cells in vitro and when delivered systemically via continuous minipump, it halts autoimmune disease progression in mouse models of type 1 diabetes (STZ and NOD) and multiple sclerosis (EAE) in vivo. rhIGF-1 administration increased Treg cells in affected tissues, maintaining their suppressive properties. Genetically, ablation of the IGF-1 receptor specifically on Treg cell populations abrogated the beneficial effects of rhIGF-1 administration on the progression of multiple sclerotic symptoms in the EAE model, establishing a direct effect of IGF-1 on Treg cell proliferation. These results establish systemically delivered rhIGF-1 as a specific, effective stimulator of Treg cell action, underscoring the clinical feasibility of manipulating natural tolerance mechanisms to suppress autoimmune disease. PMID:25339185

  11. Pancreatic adenocarcinoma upregulated factor serves as adjuvant by activating dendritic cells through stimulation of TLR4

    PubMed Central

    Yang, Benjamin; Lee, Je-Jung; Lee, Hyun-Ju; Lee, Jaemin; Jung, In Duk; Han, Hee Dong; Lee, Seung-Hyun; Koh, Sang Seok; Wu, T.-C.; Park, Yeong-Min

    2015-01-01

    Dendritic cell (DC) based cancer vaccines represent a promising immunotherapeutic strategy against cancer. To enhance the modest immunogenicity of DC vaccines, various adjuvants are often incorporated. Particularly, most of the common adjuvants are derived from bacteria. In the current study, we evaluate the use of a human pancreatic cancer derived protein, pancreatic adenocarcinoma upregulated factor (PAUF), as a novel DC vaccine adjuvant. We show that PAUF can induce activation and maturation of DCs and activate NFkB by stimulating the Toll-like receptor signaling pathway. Furthermore, vaccination with PAUF treated DCs pulsed with E7 or OVA peptides leads to generation of E7 or OVA-specific CD8+ T cells and memory T cells, which correlate with long term tumor protection and antitumor effects against TC-1 and EG.7 tumors in mice. Finally, we demonstrated that PAUF mediated DC activation and immune stimulation are dependent on TLR4. Our data provides evidence supporting PAUF as a promising adjuvant for DC based therapies, which can be applied in conjunction with other cancer therapies. Most importantly, our results serve as a reference for future investigation of human based adjuvants. PMID:26336989

  12. Nicotine Stimulates Nerve Growth Factor in Lung Fibroblasts through an NFκB-Dependent Mechanism

    PubMed Central

    Wongtrakool, Cherry; Grooms, Kora; Bijli, Kaiser M.; Crothers, Kristina; Fitzpatrick, Anne M.; Hart, C. Michael

    2014-01-01

    Rationale Airway hyperresponsiveness (AHR) is classically found in asthma, and persistent AHR is associated with poor asthma control. Although airway smooth muscle (ASM) cells play a critical pathophysiologic role in AHR, the paracrine contributions of surrounding cells such as fibroblasts to the contractile phenotype of ASM cells have not been examined fully. This study addresses the hypothesis that nicotine promotes a contractile ASM cell phenotype by stimulating fibroblasts to increase nerve growth factor (NGF) secretion into the environment. Methods Primary lung fibroblasts isolated from wild type and α7 nicotinic acetylcholine receptor (α7 nAChR) deficient mice were treated with nicotine (50 µg/ml) in vitro for 72 hours. NGF levels were measured in culture media and in bronchoalveolar lavage (BAL) fluid from asthmatic, smoking and non-smoking subjects by ELISA. The role of the NFκB pathway in nicotine-induced NGF expression was investigated by measuring NFκB nuclear translocation, transcriptional activity, chromatin immunoprecipitation assays, and si-p65 NFκB knockdown. The ability of nicotine to stimulate a fibroblast-mediated, contractile ASM cell phenotype was confirmed by examining expression of contractile proteins in ASM cells cultured with fibroblast-conditioned media or BAL fluid. Results NGF levels were elevated in the bronchoalveolar lavage fluid of nicotine-exposed mice, current smokers, and asthmatic children. Nicotine increased NGF secretion in lung fibroblasts in vitro in a dose-dependent manner and stimulated NFκB nuclear translocation, p65 binding to the NGF promoter, and NFκB transcriptional activity. These responses were attenuated in α7 nAChR deficient fibroblasts and in wild type fibroblasts following NFκB inhibition. Nicotine-treated, fibroblast-conditioned media increased expression of contractile proteins in ASM cells. Conclusion Nicotine stimulates NGF release by lung fibroblasts through α7 nAChR and NFκB dependent pathways

  13. Role of granulocyte‐macrophage colony‐stimulating factor in pulmonary fibrosis following pulmonary alveolar proteinosis

    PubMed Central

    Sha, Joy

    2016-01-01

    Abstract Pulmonary alveolar proteinosis (PAP) is a rare diffuse lung disease characterized by accumulation of lipoproteinacious material in alveoli, with distinct features on high resolution computed tomography and biopsy. Its association with pulmonary fibrosis is infrequently encountered, and a clear understanding of the underlying pathogenesis is yet to be established. We report the case of a 48‐year‐old woman with known autoimmune PAP (aPAP) first diagnosed 20 years ago, who presented with worsening hypoxemia and radiological features consistent with pulmonary fibrosis, after many years of stable disease. We present a review of previously considered mechanisms of causation behind such changes, and in particular, postulate the role of granulocyte‐macrophage colony‐stimulating factor deficiency in pulmonary fibrosis seen in aPAP. PMID:27512562

  14. Using Student-Centred Learning Environments to Stimulate Deep Approaches to Learning: Factors Encouraging or Discouraging Their Effectiveness

    ERIC Educational Resources Information Center

    Baeten, Marlies; Kyndt, Eva; Struyven, Katrien; Dochy, Filip

    2010-01-01

    This review outlines encouraging and discouraging factors in stimulating the adoption of deep approaches to learning in student-centred learning environments. Both encouraging and discouraging factors can be situated in the context of the learning environment, in students' perceptions of that context and in characteristics of the students…

  15. Stimulation of prostaglandin E/sub 2/ production by phorbol esters and epidermal growth factor in porcine thyroid cells

    SciTech Connect

    Kasai, K.; Hiraiwa, M.; Emoto, T.; Akimoto, K.; Takaoka, T.; Shimoda, S.I.

    1987-07-13

    Effects of phorbol esters and epidermal growth factor (EGF) on prostaglandin E/sub 2/ production by cultured porcine thyroid cells were examined. Both phorbol 12-myristate 13-acetate (PMA) and EGF stimulated prostaglandin E/sub 2/ production by the cells in dose related fashion. PMA stimulated prostaglandin E/sub 2/ production over fifty-fold with the dose of 10/sup -7/ M compared with control. EGF (10/sup -7/ M) also stimulated it about ten-fold. The ED/sub 50/ values of PMA and EGF were respectively around 1 x 10/sup -9/ M and 5 x 10/sup -10/ M. Thyroid stimulating hormone (TSH), however, did not stimulate prostaglandin E/sub 2/ production from 1 to 24-h incubation. The release of radioactivity from (/sup 3/H)-arachidonic acid prelabeled cells was also stimulated by PMA and EGF, but not by TSH. These results indicate that both PMA and EGF are potent stimulators of prostaglandin E/sub 2/ production, associated with the activity to stimulate arachidonic acid release in porcine thyroid cells. 36 references, 2 figures, 1 table.

  16. Mechanism of arachidonic acid liberation in platelet-activating factor-stimulated human polymorphonuclear neutrophils

    SciTech Connect

    Nakashima, S.; Suganuma, A.; Sato, M.; Tohmatsu, T.; Nozawa, Y. )

    1989-08-15

    Upon stimulation of human polymorphonuclear neutrophils with platelet-activating factor (PAF), arachidonic acid (AA) is released from membrane phospholipids. The mechanism for AA liberation, a key step in the synthesis of biologically active eicosanoids, was investigated. PAF was found to elicit an increase in the cytoplasmic level of free Ca2+ as monitored by fluorescent indicator fura 2. When (3H) AA-labeled neutrophils were exposed to PAF, the enhanced release of AA was observed with a concomitant decrease of radioactivity in phosphatidylinositol and phosphatidylcholine fractions. The inhibitors of phospholipase A2, mepacrine and 2-(p-amylcinnamoyl)-amino-4-chlorobenzoic acid, effectively suppressed the liberation of (3H)AA from phospholipids, indicating that liberation of AA is mainly catalyzed by the action of phospholipase A2. The extracellular Ca2+ is not required for AA release. However, intracellular Ca2+ antagonists, TMB-8 and high dose of quin 2/AM drastically reduced the liberation of AA induced by PAF, indicating that Ca2+ is an essential factor for phospholipase A2 activation. PAF raised the fluorescence of fura 2 at concentrations as low as 8 pM which reached a maximal level about 8 nM, whereas more than nM order concentrations of PAF was required for the detectable release of (3H)AA. Pretreatment of neutrophils with pertussis toxin resulted in complete abolition of AA liberation in response to PAF. However, the fura 2 response to PAF was not effectively inhibited by toxin treatment. In human neutrophil homogenate and membrane preparations, guanosine 5'-O-(thiotriphosphate) stimulated AA release and potentiated the action of PAF. Guanosine 5'-O-(thiodiphosphate) inhibited the effects of guanosine 5'-O-(thiotriphosphate).

  17. Subthalamic Nucleus Stimulation Increases Brain Derived Neurotrophic Factor in the Nigrostriatal System and Primary Motor Cortex

    PubMed Central

    Spieles-Engemann, Anne L.; Steece-Collier, Kathy; Behbehani, Michael M.; Collier, Timothy J.; Wohlgenant, Susan L.; Kemp, Christopher J.; Cole-Strauss, Allyson; Levine, Nathan D.; Gombash, Sara E.; Thompson, Valerie B.; Lipton, Jack W.; Sortwell, Caryl E.

    2011-01-01

    The mechanisms underlying the effects of long-term deep brain stimulation of the subthalamic nucleus (STN DBS) as a therapy for Parkinson’s disease (PD) remain poorly understood. The present study examined whether functionally effective, long-term STN DBS modulates glial cell line-derived neurotrophic factor (GDNF) and/or brain-derived neurotrophic factor (BDNF) in both unlesioned and unilateral 6-hydroxydopamine lesioned rats. Lesioned rats that received two weeks of continuous unilateral STN DBS exhibited significant improvements in parkinsonian motor behaviors in tests of forelimb akinesia and rearing activity. Unilateral STN DBS did not increase GDNF in the nigrostriatal system, primary motor cortex (M1), or hippocampus of unlesioned rats. In contrast, unilateral STN DBS increased BDNF protein 2–3 fold bilaterally in the nigrostriatal system with the location (substantia nigra vs. striatum) dependent upon lesion status. Further, BDNF protein was bilaterally increased in M1 cortex by as much as 2 fold regardless of lesion status. STN DBS did not impact cortical regions that receive less input from the STN. STN DBS also was associated with bilateral increases in BDNF mRNA in the substantia nigra (SN) and internal globus pallidus (GPi). The increase observed in GPi was completely blocked by pretreatment with 5-Methyl-10,11-dihydro-5 H-dibenzo[a,d]cyclohepten-5,10-imine (MK-801), suggesting that the activation of N-methyl-D-aspartate (NMDA) receptors was involved in this phenomenon. The upregulation of BDNF associated with long term STN DBS suggest that this therapy may exert pronounced and underappreciated effects on plasticity in the basal ganglia circuitry that may play a role in the symptomatic effects of this therapy as well as support the neuroprotective effect of stimulation documented in this rat model. PMID:22328911

  18. Immobilized epidermal growth factor stimulates persistent, directed keratinocyte migration via activation of PLCγ1.

    PubMed

    Kim, Chloe S; Mitchell, Isaiah P; Desotell, Anthony W; Kreeger, Pamela K; Masters, Kristyn S

    2016-07-01

    Epidermal growth factor (EGF) is a critical element in dermal repair, but EGF-containing wound dressings have not been successful clinically. However, these dressings have delivered only soluble EGF, and the native environment provides both soluble and matrix-bound EGF. To address our hypothesis that tethered EGF can stimulate cell behaviors not achievable with soluble EGF, we examined single-cell movement and signaling in human immortalized HaCaT keratinocytes treated with soluble or immobilized EGF. Although both EGF treatments increased collective sheet displacement and individual cell speed, only cells treated with immobilized EGF exhibited directed migration, as well as 2-fold greater persistence compared with soluble EGF. Immunofluorescence showed altered EGF receptor (EGFR) trafficking, where EGFR remained membrane-localized in the immobilized EGF condition. Cells treated with soluble EGF demonstrated higher phosphorylated ERK1/2, and cells on immobilized EGF exhibited higher pPLCγ1, which was localized at the leading edge. Treatment with U0126 inhibited migration in both conditions, demonstrating that ERK1/2 activity was necessary but not responsible for the observed differences. In contrast, PLCγ1 inhibition with U73122 significantly decreased persistence on immobilized EGF. Combined, these results suggest that immobilized EGF increases collective keratinocyte displacement via an increase in single-cell migration persistence resulting from altered EGFR trafficking and PLCγ1 activation.-Kim, C. S., Mitchell, I. P., Desotell, A. W., Kreeger, P. K., Masters, K. S. Immobilized epidermal growth factor stimulates persistent, directed keratinocyte migration via activation of PLCγ1. PMID:27025961

  19. FGF2 stimulates SDF-1 expression through the Erm transcription factor in Sertoli cells.

    PubMed

    Yoon, Kyung-Ae; Chae, Young-Mi; Cho, Je-Yoel

    2009-07-01

    Ets-related molecule (Erm) is a member of the Ets transcription factor family. Erm is known to be an important factor for the self-renewal of Spermatogonial stem cells (SSCs) and the maintenance of spermatogenesis. We investigated the molecular mechanism of Erm regulation on SDF-1 in TM4 Sertoli cells. Erm and Sdf-1 levels were up-regulated after FGF2 treatment in TM4 cells, whereas these levels were significantly decreased by FGF2 in ST2 bone marrow stromal cells. Knockdown of Erm by siRNA in the presence of FGF2 decreased the Sdf-1 levels in TM4 cells. The expression levels of Erm were similar and Erm overexpression increased the Sdf-1 in both TM4 and ST2 cells. FGFR subtype analysis revealed that FGFR4 was expressed in TM4 cells but not in ST2 cells. A blocking experiment also confirmed that FGFR4 is partly responsible for the up-regulation of Erm and SDF-1 induced by FGF2 stimulation in TM4 cells. FGF2 and ERM increased the activity of Sdf-1 gene promoter region in a dose-dependent manner. EMSA revealed that ERM strongly binds to the -846 to -851 nucleotide region of the potential Ets binding site (EBS) in the Sdf-1 promoter. In addition, CXCR4, the SDF-1 receptor, was expressed in spermatogonia and Sertoli cells in the seminiferous tubules of the mouse testis. Our results indicate that ERM directly regulates Sdf-1 gene expression by interacting with its cis-acting element in response to FGF2 stimulation in TM4 cells. PMID:19301256

  20. Prostacyclin synthesis stimulating plasma factor in patients with peripheral vascular disease.

    PubMed

    Strobl-Jäger, E; Fitscha, P; Kaliman, J; Sinzinger, H; Peskar, B A

    1987-08-01

    Human plasma contains a factor capable of stimulating vascular prostacyclin generation even in atherosclerotic vessels with minimal in-vitro capacity for PGI2-synthesis. The activity of this prostacyclin stimulating plasma factor (PSPF) has been reported to be elevated in renal failure and hepatic coma. We are not aware of any data as to whether this PSPF plays a role in maintaining hemostatic balance in patients with peripheral vascular lesions. Therefore, we examined 62 patients with peripheral vascular disease (PVD). This study group was subdivided into normo- and hyperlipemic subjects, patients with and without maturity onset diabetes, and plasma beta-thromboglobulin levels higher and lower than 50 ng/ml. 10 healthy sex and age matched persons served as controls. Vascular prostacyclin formation was studied in vitro after incubation of the patients' plasma and a buffer control with various tissue samples (human femoral artery, rat abdominal and thoracic aorta of healthy and of streptozotocin induced diabetic animals, swine endothelial layer and remaining tissue (media and adventitia) and cultured endothelial (EC) and smooth muscle cells (SMC) of minipig arota. In addition, 6-oxo-PFG1 alpha formation by cultured EC and SMC (minipig aorta source) after incubation with tris HCl-buffer or plasma were estimated by means of specific radioimmunoassays. In general, tissue samples and cells incubated in plasma exhibit a marked increase of in-vitro PGI2-formation as compared to buffer. No difference could be found between PSPF of CHD-patients and healthy controls. Similar findings were obtained using incubated vascular tissue and cultured cells by means of the bioassay and specific RIA, respectively. These findings indicate that the PSPF does not seem to be of any clinical relevance in hemostatic regulation in patients with advanced atherosclerosis. PMID:2958884

  1. Free bone graft reconstruction of irradiated facial tissue: Experimental effects of basic fibroblast growth factor stimulation

    SciTech Connect

    Eppley, B.L.; Connolly, D.T.; Winkelmann, T.; Sadove, A.M.; Heuvelman, D.; Feder, J. )

    1991-07-01

    A study was undertaken to evaluate the potential utility of basic fibroblast growth factor in the induction of angiogenesis and osseous healing in bone previously exposed to high doses of irradiation. Thirty New Zealand rabbits were evaluated by introducing basic fibroblast growth factor into irradiated mandibular resection sites either prior to or simultaneous with reconstruction by corticocancellous autografts harvested from the ilium. The fate of the free bone grafts was then evaluated at 90 days postoperatively by microangiographic, histologic, and fluorochrome bone-labeling techniques. Sequestration, necrosis, and failure to heal to recipient osseous margins was observed both clinically and histologically in all nontreated irradiated graft sites as well as those receiving simultaneous angiogenic stimulation at the time of graft placement. No fluorescent activity was seen in these graft groups. In the recipient sites pretreated with basic fibroblast growth factor prior to placement of the graft, healing and reestablishment of mandibular contour occurred in nearly 50 percent of the animals. Active bone formation was evident at cortical margins adjacent to the recipient sites but was absent in the more central cancellous regions of the grafts.

  2. Osteoblast-derived paracrine factors regulate angiogenesis in response to mechanical stimulation.

    PubMed

    Liu, Chao; Cui, Xin; Ackermann, Thomas M; Flamini, Vittoria; Chen, Weiqiang; Castillo, Alesha B

    2016-07-11

    Angiogenesis is a process by which new blood vessels emerge from existing vessels through endothelial cell sprouting, migration, proliferation, and tubule formation. Angiogenesis during skeletal growth, homeostasis and repair is a complex and incompletely understood process. As the skeleton adapts to mechanical loading, we hypothesized that mechanical stimulation regulates "osteo-angio" crosstalk in the context of angiogenesis. We showed that conditioned media (CM) from osteoblasts exposed to fluid shear stress enhanced endothelial cell proliferation and migration, but not tubule formation, relative to CM from static cultures. Endothelial cell sprouting was studied using a dual-channel collagen gel-based microfluidic device that mimics vessel geometry. Static CM enhanced endothelial cell sprouting frequency, whereas loaded CM significantly enhanced both frequency and length. Both sprouting frequency and length were significantly enhanced in response to factors released from osteoblasts exposed to fluid shear stress in an adjacent channel. Osteoblasts released angiogenic factors, of which osteopontin, PDGF-AA, IGBP-2, MCP-1, and Pentraxin-3 were upregulated in response to mechanical loading. These data suggest that in vivo mechanical forces regulate angiogenesis in bone by modulating "osteo-angio" crosstalk through release of paracrine factors, which we term "osteokines". PMID:27332785

  3. 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.

  4. Mononuclear cells from the cord blood and granulocytecolony stimulating factor-mobilized peripheral blood: is there a potential for treatment of cerebral palsy?

    PubMed Central

    Koh, Hani; Hwang, Kyoujung; Lim, Hae-Young; Kim, Yong-Joo; Lee, Young-Ho

    2015-01-01

    To investigate a possible therapeutic mechanism of cell therapy in the field of cerebral palsy us