scholarly journals 125 CHANGES OF PLASMA MACROPHAGE COLONY-STIMULATING FACTOR LEVELS AND ITS GENE EXPRESSION IN PERIPHERAL WHITE BLOOD CELLS DURING PREGNANCY IN JAPANESE BLACK CATTLE

2005 ◽  
Vol 17 (2) ◽  
pp. 213
Author(s):  
K. Oshima ◽  
K. Yoshihara ◽  
H. Watanabe ◽  
T. Kojima ◽  
M. Komatsu ◽  
...  
Keyword(s):  
Gene Expression ◽  
White Blood Cells ◽  
Colony Stimulating Factor ◽  
Rt Pcr ◽  
Relative Level ◽  
Csf Levels ◽  
Macrophage Colony Stimulating Factor ◽  
Macrophage Colony ◽  
Peripheral White Blood Cells ◽  
Stimulating Factor

Macrophage colony-stimulating factor (M-CSF) is a hemopoietic cytokine that plays a primary role in placental physiology. Gene expression of M-CSF in bovine intercaruncular endometrium shows an upward trend in mid-pregnancy. The objective of this study was to determine the plasma M-CSF levels and the M-CSF gene expression levels in maternal peripheral white blood cells (PWBCs) during pregnancy using ELISA and quantitative RT-PCR. In Experiment 1, the plasma M-CSF levels in 112 Japanese Black heifers or cows were determined. Animals were divided into four groups according to pregnancy stage: first- (n = 29), second- (n = 33), third- (n = 26) trimester, and non-pregnant (n = 24). ELISA for bovine M-CSF established by Yoshihara et al. (2003 Vet. Immunol. Immunopathol. 95, 103–111) was used according to their instructions. The absorbance was measured at 405 nm in the Biomek Plate Reader (Beckman Coulter, Fullerton, CA, USA). In Experiment 2, the plasma M-CSF levels and M-CSF gene expression levels in PWBCs during pregnancy were determined. The plasma samples for ELISA were obtained from 8 heifers and 3 cows every 1 and/or 2 weeks. The PWBCs samples for quantitative RT-PCR were obtained from 4 heifers every 1 and/or 4 weeks. All quantitative RT-PCR protocols were carried out according to the previous report (Oshima et al. 2003 Theriogenology 60, 1217–1226). The quantitative PCR assay used an ABI Prism 7700 Sequence Detector (Applied Biosystems, Foster City, CA, USA). Signals were detected according to the manufacturer's instructions. The relative level of M-CSF expression was calculated on the basis of glyceraldehyde-phosphate-dehydrogenase (GAPDH) quantity (in the method of calculation, the relative level = M-CSF quantity/GAPDH quantity). Data were analyzed by Kruskal-Wallis test. In Experiment 1, the plasma M-CSF level in second-trimester cows was significantly higher than those in other stages (P < 0.05). In Experiment 2, the plasma M-CSF levels were significantly higher in gestational age from −4 to 1 weeks compared with the last stage of pregnancy (P < 0.05). The levels decreased until 6 weeks, appeared to temporarily increase, and were relatively constant until 35 weeks. Macrophage colony-stimulating factor genes were expressed in all samples examined; the levels were relatively constant in early pregnancy, and then were widely varied until parturition. These results suggest that plasma M-CSF levels may be related to the maternal condition of pregnancy and to a slight extent to M-CSF gene expression in PWBCs. This work was supported in part by a Grant-in-aid from the Recombinant Cytokine Project (RCP2002-2110), provided by the Ministry of Agriculture, Forestry, and Fisheries, Japan.

scholarly journals cAMP attenuates interleukin-1-stimulated macrophage colony-stimulating factor (M-CSF) expression

2000 ◽  
Vol 350 (1) ◽  
pp. 115-122 ◽  
Author(s):  
Pisate J. KAMTHONG ◽  
Fu-mei WU ◽  
Ming-chi WU
Keyword(s):  
Pancreatic Cancer Cell Line ◽  
Human Pancreatic Cancer ◽  
Luciferase Reporter ◽  
Transcriptional Level ◽  
Colony Stimulating Factor ◽  
Rt Pcr ◽  
Macrophage Colony Stimulating Factor ◽  
Macrophage Colony ◽  
Stimulating Factor ◽  
Camp Elevation

Macrophage colony-stimulating factor (M-CSF) is a multifunctional cytokine attributed with key biological functions beyond the first discovered role in promoting proliferation of myeloid cell lineage. The human pancreatic cancer cell line MIA PaCa-2, from which the M-CSF gene was originally cloned, was used to study regulation of M-CSF expression. Expression of M-CSF was inducible by interleukin-1α (IL-1α), lipopolysaccharide (LPS) and PMA as demonstrated by a biological activity assay, Northern-blot analysis and reverse transcriptase (RT) PCR. Treatment of the cells with forskolin or dibutyryl-cAMP attenuated the expression of M-CSF induced by IL-1α or LPS, but not by PMA. Electromobility shift assays showed that IL-1α predominantly activated nuclear factor κB (NF-κB), while PMA preferentially activated activator protein-1 (AP-1). The activation of NF-κB, but not AP-1, could be attenuated by cAMP elevation. Relative RT-PCR demonstrated that the expression of a 1.6-kb M-CSF mRNA transcript was more effectively induced by IL-1α than a 4.0-kb transcript. By and large the induced expression of both mRNA transcripts could be attenuated by cAMP. M-CSF promoter-driven luciferase reporter-gene assays revealed that cAMP elevation attenuated the IL-1-induced transcription activation of the M-CSF promoter, but it had no effect on PMA-induced transcription. Our findings suggest that cAMP regulates M-CSF gene expression at the transcriptional level and that its inhibitory effect involves NF-κB signalling pathway.

scholarly journals Role of Granulocyte-Macrophage Colony-Stimulating Factor in Acute Myeloid Leukemia/Myelodysplastic Syndromes

2018 ◽  
pp. 1-6
Author(s):  
Neemat M. Kassem ◽  
Alya M. Ayad ◽  
Noha M. El Husseiny ◽  
Doaa M. El-Demerdash ◽  
Hebatallah A. Kassem ◽  
...  
Keyword(s):  
Gene Expression ◽  
Myeloid Leukemia ◽  
Serum Levels ◽  
Colony Stimulating Factor ◽  
Gm Csf ◽  
Macrophage Colony Stimulating Factor ◽  
Macrophage Colony ◽  
Stimulating Factor ◽  
Acute Myeloid

Purpose Granulocyte-macrophage colony-stimulating factor (GM-CSF) cytokine stimulates growth, differentiation, and function of myeloid progenitors. We aimed to study the role of GM-CSF gene expression, its protein, and antibodies in patients with acute myeloid leukemia/myelodysplastic syndromes (AML/MDS) and their correlation to disease behavior and treatment outcome. The study included 50 Egyptian patients with AML/MDS in addition to 20 healthy volunteers as control subjects. Patients and Methods Assessment of GM-CSF gene expression was performed by quantitative real-time polymerase chain reaction. GM-CSF proteins and antibodies were assessed by enzyme-linked immunosorbent assay. Results There was significant decrease in GM-CSF gene expression ( P = .008), increase in serum level of GM-CSF protein ( P = .0001), and increase in anti–GM-CSF antibodies ( P = .001) in patients with AML/MDS compared with healthy control subjects. In addition, there was a significant negative correlation between serum levels of GM-CSF protein and initial peripheral blood blasts, percentage as well as response to therapy. Conclusion Any alteration in GM-CSF gene expression could have implications in leukemogenesis. In addition, GM-CSF protein serum levels could be used to predict outcome of therapy. GM-CSF antibodies may also play a role in the pathogenesis of AML/MDS. The use of these GM-CSF parameters for disease monitoring and as markers of disease activity needs further research.

scholarly journals STAT5A-Deficient Mice Demonstrate a Defect in Granulocyte-Macrophage Colony-Stimulating Factor–Induced Proliferation and Gene Expression

Blood ◽  
1997 ◽  
Vol 90 (5) ◽  
pp. 1768-1776 ◽  
Author(s):  
Gerald M. Feldman ◽  
Louis A. Rosenthal ◽  
Xiuwen Liu ◽  
Mark P. Hayes ◽  
Anthony Wynshaw-Boris ◽  
...  
Keyword(s):  
Gene Expression ◽  
Dna Binding Proteins ◽  
Colony Stimulating Factor ◽  
Wild Type ◽  
Stat Proteins ◽  
Gm Csf ◽  
Macrophage Colony Stimulating Factor ◽  
Macrophage Colony ◽  
Stimulating Factor ◽  
In Cells

Abstract Responses of cells to cytokines typically involve the activation of a family of latent DNA binding proteins, referred to as signal transducers and activators of transcription (STAT) proteins, which are critical for the expression of early response genes. Of the seven known STAT proteins, STAT5 (originally called mammary gland factor) has been shown to be activated by several cytokines, such as granulocyte-macrophage colony-stimulating factor (GM-CSF), interleukin-3 (IL-3), and IL-5, which are known to play important roles in growth and differentiation of hematopoietic precursors. In this report we have used mice that are deficient in STAT5A (one of two homologues of STAT5) to study the role of STAT5A in GM-CSF stimulation of cells. When bone marrow–derived macrophages were generated by differentiation with macrophage-CSF (M-CSF), exposure of cells from wild-type mice to GM-CSF resulted in a typical pattern of assembly of DNA binding proteins specific for the gamma activation sequence (GAS) element within the β-casein promoter. However, in cells from the STAT5A null mouse one of the shifted bands was absent. Immunoblotting analysis in the null mice showed that lack of STAT5A protein resulted in no alteration in activation of STAT5B by tyrosine phosphorylation. Proliferation experiments revealed that, when exposed to increasing concentrations of GM-CSF, cells derived from the null mice grew considerably more slowly than cells derived from the wild-type mice. Moreover, expression of GM-CSF–dependent genes, CIS and A1, was markedly inhibited in cells derived from null mice as compared with those of wild-type mice. The decreased expression observed with A1, a bcl-2 like gene, may account in part for the suppression of growth in cells from the null mice. These data suggest that the presence of STAT5A during the GM-CSF–induced assembly of STAT5 dimers is critical for the formation of competent transcription factors that are required for both gene expression and cell proliferation.

scholarly journals Secretion of cytokines (interleukins-1 alpha, -3, and -6 and granulocyte-macrophage colony-stimulating factor) by normal human bone marrow megakaryocytes

Blood ◽  
1995 ◽  
Vol 85 (3) ◽  
pp. 685-691 ◽  
Author(s):  
C Wickenhauser ◽  
J Lorenzen ◽  
J Thiele ◽  
A Hillienhof ◽  
K Jungheim ◽  
...  
Keyword(s):  
Plaque Assay ◽  
Secretory Activity ◽  
Colony Stimulating Factor ◽  
Rt Pcr ◽  
Gm Csf ◽  
Clear Cut ◽  
Normal Human ◽  
Macrophage Colony Stimulating Factor ◽  
Macrophage Colony ◽  
Stimulating Factor

The effects of cytokine stimulation [recombinant human interleukin (rhIL)-1 alpha, rhIL-3, rhIL-6, rhIL-11, and rh granulocyte-macrophage colony-stimulating factor (GM-CSF)] on the secretory activity of normal human megakaryocytes were studied by means of the reverse hemolytic plaque assay (RHPA) in enriched cell preparations. This test facilitates an extremely sensitive determination of cytokine secretion at the single-cell level, together with the clear-cut identification of each immunostained (CD61) secretory active megakaryocyte. Moreover, the reverse transcriptase-polymerase chain reaction (RT-PCR) was used to investigate the expression of IL-6, IL-6 receptor (IL-6R), IL-9, IL-10, IL-12, and IL-13 mRNA in highly concentrated megakaryocyte preparations. In comparison with the spontaneous secretion rate, stimulation with rhIL-1 alpha, rhIL-6, and rhGM-CSF failed to induce a significant increase in the release of cytokines by CD61+ cells. On the other hand, both rhIL-3 and, in a less pronounced way, rhIL-11 exerted a marked effect on IL-6 secretion. Additionally, after stimulation with rhIL-3, a significant enhancement of the secretion of IL-3 and GM-CSF, but not of IL-1 alpha, could be observed. Using the RT-PCR, a significant induction of IL-6 expression could be appreciated in the enriched megakaryocyte population (60% to 80%) stimulated with rhIL-3. The results of this study provide persuasive evidence that a number of cytokines are synthesized and secreted by human megakaryocytes and not only by hematopoietic stroma cells. These data suggest the existence of autocrine and paracrine mechanisms that may influence maturation and differentiation of megakaryocytes as well as act on various stroma cells to sustain an appropriate hematopoietic micro-environment.

Generation of Functional Human Dendritic Cells From Adherent Peripheral Blood Monocytes by CD40 Ligation in the Absence of Granulocyte-Macrophage Colony-Stimulating Factor

Blood ◽  
1998 ◽  
Vol 92 (11) ◽  
pp. 4238-4247 ◽  
Author(s):  
Peter Brossart ◽  
Frank Grünebach ◽  
Gernot Stuhler ◽  
Volker L. Reichardt ◽  
Robert Möhle ◽  
...  
Keyword(s):  
Peripheral Blood ◽  
Colony Stimulating Factor ◽  
Blood Monocytes ◽  
Rt Pcr ◽  
Cd40 Ligation ◽  
Peripheral Blood Monocytes ◽  
Gm Csf ◽  
Macrophage Colony Stimulating Factor ◽  
Macrophage Colony ◽  
Stimulating Factor

Abstract Recently it has been shown that dendritic cells (DC) can develop from peripheral blood monocytes when grown in the presence of granulocyte-macrophage colony-stimulating factor (GM-CSF) and interleukin-4 (IL-4). However, it is unclear whether DC can also develop from monocytes in absence of these cytokines. We therefore analyzed the effect of Flt-3 ligand (Flt3L) and of CD40 ligand on the development of human DC from blood monocytes in the absence of GM-CSF. Adherent peripheral blood mononuclear cells (PBMNC) were cultured in the presence of different cytokine combinations and analyzed for the expression of surface molecules and antigen presenting capacity. For functional analyses, cells were tested for their ability to stimulate allogeneic T lymphocytes in a mixed lymphocyte reaction (MLR), to present soluble antigens, and to induce primary HIV-peptide–specific cytotoxic T-cell (CTL) responses in vitro. Furthermore, expression of DC-CK1, a recently identified chemokine with specific expression in DC, and of IL-18 (IGIF), a growth and differentiation factor for Th 1 lymphocytes, was analyzed by reverse-transcription polymerase chain reaction (RT-PCR). In our study, Flt3L alone was not sufficient to generate DC and required addition of IL-4. DC generated with Flt3L and IL-4 underwent maturation after stimulation with tumor necrosis factor- (TNF-) or CD40L, characterized by CD83 expression, upregulation of MHC, adhesion, and costimulatory molecules as well as increased allogeneic proliferative response. In contrast, CD40 ligation alone promoted differentiation of adherent blood monocytes into functional DC in the absence of GM-CSF and IL-4. These cells displayed all phenotypic and functional characteristics of mature DC and were potent stimulatory cells in priming of major histocompatibility complex (MHC) class I–restricted CTL responses against an HIV-peptide, whereas their ability to present soluble protein antigens was reduced. Using a semiquantitative RT-PCR, DC-CK1 and IL-18 transcripts were detected in all generated DC populations, independent of growth factors used. Our findings provide further evidence for the importance of CD40-CD40L interaction for initiation and maintenance of T-cell responses and confirm the emerging concept that blood monocytes provide an additional source of DC depending on external stimuli.

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