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

Blood ◽  
1990 ◽  
Vol 76 (9) ◽  
pp. 1783-1787 ◽  
Author(s):  
CS Rosenfeld ◽  
C Evans ◽  
RK Shadduck
Keyword(s):  
Bone Marrow ◽  
Conditioned Media ◽  
Human Bone ◽  
Human Bone Marrow ◽  
Human Macrophage ◽  
Colony Stimulating Factor ◽  
Colony Stimulating Activity ◽  
Macrophage Colony Stimulating Factor ◽  
Macrophage Colony ◽  
Stimulating Factor

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

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

Blood ◽  
1990 ◽  
Vol 76 (9) ◽  
pp. 1783-1787
Author(s):  
CS Rosenfeld ◽  
C Evans ◽  
RK Shadduck
Keyword(s):  
Bone Marrow ◽  
Conditioned Media ◽  
Human Bone ◽  
Human Bone Marrow ◽  
Human Macrophage ◽  
Colony Stimulating Factor ◽  
Colony Stimulating Activity ◽  
Macrophage Colony Stimulating Factor ◽  
Macrophage Colony ◽  
Stimulating Factor

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

scholarly journals Fractionation of subsets of BFU-E from normal human bone marrow: responsiveness to erythropoietin, human placental-conditioned medium, or granulocyte-macrophage colony-stimulating factor

Blood ◽  
1988 ◽  
Vol 71 (3) ◽  
pp. 758-765 ◽  
Author(s):  
G Kannourakis ◽  
GR Johnson
Keyword(s):  
Bone Marrow ◽  
Conditioned Medium ◽  
Human Bone ◽  
Human Bone Marrow ◽  
Colony Stimulating Factor ◽  
Normal Human ◽  
Macrophage Colony Stimulating Factor ◽  
Macrophage Colony ◽  
Stimulating Factor ◽  
Normal Human Bone Marrow

Abstract Normal human bone marrow mononuclear cells were fractionated by differential adherence, immunomagnetic separation, and fluorescence- activated cell sorting (FACS). The resultant fractionated cells were cultured in semisolid medium to monitor the presence of BFU-E, Mix-CFC, and nonerythroid CFC. Two populations of cells were recovered on the basis of binding by the monoclonal antibody (MoAb) RFB-1. One of these populations contained BFU-E that were stimulated only by erythropoietin (Epo), whereas the second population contained BFU-E responsive to Epo, Epo and recombinant human granulocyte-macrophage colony-stimulating factor (rHGM-CSF), or Epo and human placental-conditioned medium (HPCM). Prior enrichment of clonogenic cells by removal of adherent and Leu-M3+ve, Leu-4+ve, Leu-7+ve, B1+ve, WEMG1+ve, and Glycophorin A+ve cells, followed by FACS fractionation on the basis of RFB-1 binding, consistently resulted in recoveries of BFU-E, Mix-CFC, and nonerythroid CFC of greater than 100% (up to 800%). These procedures also resulted in enrichment of up to 200-fold and frequencies of 1:6 for BFU-E, 1:5 for CFC, and 1:130 for Mix-CFC.

scholarly journals Fractionation of subsets of BFU-E from normal human bone marrow: responsiveness to erythropoietin, human placental-conditioned medium, or granulocyte-macrophage colony-stimulating factor

Blood ◽  
1988 ◽  
Vol 71 (3) ◽  
pp. 758-765 ◽  
Author(s):  
G Kannourakis ◽  
GR Johnson
Keyword(s):  
Bone Marrow ◽  
Conditioned Medium ◽  
Human Bone ◽  
Human Bone Marrow ◽  
Colony Stimulating Factor ◽  
Normal Human ◽  
Macrophage Colony Stimulating Factor ◽  
Macrophage Colony ◽  
Stimulating Factor ◽  
Normal Human Bone Marrow

Normal human bone marrow mononuclear cells were fractionated by differential adherence, immunomagnetic separation, and fluorescence- activated cell sorting (FACS). The resultant fractionated cells were cultured in semisolid medium to monitor the presence of BFU-E, Mix-CFC, and nonerythroid CFC. Two populations of cells were recovered on the basis of binding by the monoclonal antibody (MoAb) RFB-1. One of these populations contained BFU-E that were stimulated only by erythropoietin (Epo), whereas the second population contained BFU-E responsive to Epo, Epo and recombinant human granulocyte-macrophage colony-stimulating factor (rHGM-CSF), or Epo and human placental-conditioned medium (HPCM). Prior enrichment of clonogenic cells by removal of adherent and Leu-M3+ve, Leu-4+ve, Leu-7+ve, B1+ve, WEMG1+ve, and Glycophorin A+ve cells, followed by FACS fractionation on the basis of RFB-1 binding, consistently resulted in recoveries of BFU-E, Mix-CFC, and nonerythroid CFC of greater than 100% (up to 800%). These procedures also resulted in enrichment of up to 200-fold and frequencies of 1:6 for BFU-E, 1:5 for CFC, and 1:130 for Mix-CFC.

scholarly journals Purification and properties of bacterially synthesized human granulocyte-macrophage colony stimulating factor

Blood ◽  
1987 ◽  
Vol 69 (1) ◽  
pp. 43-51 ◽  
Author(s):  
AW Burgess ◽  
CG Begley ◽  
GR Johnson ◽  
AF Lopez ◽  
DJ Williamson ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Conditioned Medium ◽  
Human Bone ◽  
Human Bone Marrow ◽  
Colony Stimulating Factor ◽  
Gm Csf ◽  
Adult Human ◽  
Macrophage Colony Stimulating Factor ◽  
Macrophage Colony ◽  
Stimulating Factor

Abstract Human granulocyte-macrophage colony stimulating factor (GM-CSF) has been synthesized in high yield using a temperature inducible plasmid in Escherichia coli. The human GM-CSF is readily isolated from the bacterial proteins because of its differential solubility and chromatographic properties. The bacterially synthesized form of the human GM-CSF contains an extra methionine residue at position 1, but otherwise it is identical to the polypeptide predicted from the cDNA sequence. The specific activity of 2.9 X 10(7) units/mg of protein for purified bacterially synthesized human GM-CSF indicates that despite the lack of glycosylation, the molecule is substantially in its native conformation. This molecule stimulated the same number and type of both seven- and 14-day human bone marrow colonies as the CSF alpha preparation from human placental conditioned medium. Human GM-CSF had no activity on murine bone marrow or murine leukemic cells. There was no detectable, direct stimulation of adult human erythroid burst forming units (BFU-E) by the bacterially synthesized human GM-CSF. Although impure preparations containing native human GM-CSF (eg, human placental conditioned medium) stimulated the formation of mixed colonies, even in the presence of erythropoietin, the bacterially synthesized human GM-CSF failed to stimulate the formation of mixed colonies from adult human bone marrow cells. The bacterially synthesized human GM-CSF increased N-formyl-methionyl-leucyl- phenylalanine (FMLP)-induced superoxide production and lysozyme secretion. Antibody-dependent cytotoxicity and phagocytosis by human neutrophils was stimulated by the bacterially synthesized human GM-CSF and eosinophils were also activated in the antibody-dependent cytotoxicity assay.

scholarly journals Purification and properties of bacterially synthesized human granulocyte-macrophage colony stimulating factor

Blood ◽  
1987 ◽  
Vol 69 (1) ◽  
pp. 43-51 ◽  
Author(s):  
AW Burgess ◽  
CG Begley ◽  
GR Johnson ◽  
AF Lopez ◽  
DJ Williamson ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Conditioned Medium ◽  
Human Bone ◽  
Human Bone Marrow ◽  
Colony Stimulating Factor ◽  
Gm Csf ◽  
Adult Human ◽  
Macrophage Colony Stimulating Factor ◽  
Macrophage Colony ◽  
Stimulating Factor

Human granulocyte-macrophage colony stimulating factor (GM-CSF) has been synthesized in high yield using a temperature inducible plasmid in Escherichia coli. The human GM-CSF is readily isolated from the bacterial proteins because of its differential solubility and chromatographic properties. The bacterially synthesized form of the human GM-CSF contains an extra methionine residue at position 1, but otherwise it is identical to the polypeptide predicted from the cDNA sequence. The specific activity of 2.9 X 10(7) units/mg of protein for purified bacterially synthesized human GM-CSF indicates that despite the lack of glycosylation, the molecule is substantially in its native conformation. This molecule stimulated the same number and type of both seven- and 14-day human bone marrow colonies as the CSF alpha preparation from human placental conditioned medium. Human GM-CSF had no activity on murine bone marrow or murine leukemic cells. There was no detectable, direct stimulation of adult human erythroid burst forming units (BFU-E) by the bacterially synthesized human GM-CSF. Although impure preparations containing native human GM-CSF (eg, human placental conditioned medium) stimulated the formation of mixed colonies, even in the presence of erythropoietin, the bacterially synthesized human GM-CSF failed to stimulate the formation of mixed colonies from adult human bone marrow cells. The bacterially synthesized human GM-CSF increased N-formyl-methionyl-leucyl- phenylalanine (FMLP)-induced superoxide production and lysozyme secretion. Antibody-dependent cytotoxicity and phagocytosis by human neutrophils was stimulated by the bacterially synthesized human GM-CSF and eosinophils were also activated in the antibody-dependent cytotoxicity assay.

scholarly journals Recombinant human macrophage colony-stimulating factor (M-CSF) requires subliminal concentrations of granulocyte/macrophage (GM)-CSF for optimal stimulation of human macrophage colony formation in vitro.

1987 ◽  
Vol 166 (6) ◽  
pp. 1851-1860 ◽  
Author(s):  
D Caracciolo ◽  
N Shirsat ◽  
G G Wong ◽  
B Lange ◽  
S Clark ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Colony Formation ◽  
Human Macrophage ◽  
Colony Stimulating Factor ◽  
Colony Assay ◽  
Gm Csf ◽  
Macrophage Colony Stimulating Factor ◽  
Macrophage Colony ◽  
Stimulating Factor

Human macrophage colony-stimulating factor (M-CSF or CSF-1), either in purified or in recombinant form, is able to generate macrophagic colonies in a murine bone marrow colony assay, but only stimulates small macrophagic colonies of 40-50 cells in a human bone marrow colony assay. We report here that recombinant human granulocytic/macrophage colony stimulating factor (rhGM-CSF) at concentrations in the range of picograms enhances the responsiveness of bone marrow progenitors to M-CSF activity, resulting in an increased number of macrophagic colonies of up to 300 cells. Polyclonal antiserum against M-CSF did not alter colony formation of bone marrow progenitors incubated with GM-CSF at optimal concentration (1-10 ng/ml) for these in vitro assays. Thus, GM-CSF at higher concentrations (nanogram range) can by itself, elicit macrophagic colonies, and at lower concentrations (picogram range) acts to enhance the responsiveness of these progenitors to M-CSF.

scholarly journals T3 Regulates a Human Macrophage-Derived TSH-β Splice Variant: Implications for Human Bone Biology

Endocrinology ◽  
2016 ◽  
Vol 157 (9) ◽  
pp. 3658-3667 ◽  
Author(s):  
R. Baliram ◽  
R. Latif ◽  
S. A. Morshed ◽  
M. Zaidi ◽  
T. F. Davies
Keyword(s):  
Bone Marrow ◽  
Real Time ◽  
Real Time Pcr ◽  
Splice Variant ◽  
Human Bone ◽  
Colony Stimulating Factor ◽  
Bone Biology ◽  
Macrophage Colony Stimulating Factor ◽  
Macrophage Colony ◽  
Stimulating Factor

TSH and thyroid hormones (T3 and T4) are intimately involved in bone biology. We have previously reported the presence of a murine TSH-β splice variant (TSH-βv) expressed specifically in bone marrow-derived macrophages and that exerted an osteoprotective effect by inducing osteoblastogenesis. To extend this observation and its relevance to human bone biology, we set out to identify and characterize a TSH-β variant in human macrophages. Real-time PCR analyses using human TSH-β-specific primers identified a 364-bp product in macrophages, bone marrow, and peripheral blood mononuclear cells that was sequence verified and was homologous to a human TSH-βv previously reported. We then examined TSH-βv regulation using the THP-1 human monocyte cell line matured into macrophages. After 4 days, 46.1% of the THP-1 cells expressed the macrophage markers CD-14 and macrophage colony-stimulating factor and exhibited typical morphological characteristics of macrophages. Real-time PCR analyses of these cells treated in a dose-dependent manner with T3 showed a 14-fold induction of human TSH-βv mRNA and variant protein. Furthermore, these human TSH-βv-positive cells, induced by T3 exposure, had categorized into both M1 and M2 macrophage phenotypes as evidenced by the expression of macrophage colony-stimulating factor for M1 and CCL-22 for M2. These data indicate that in hyperthyroidism, bone marrow resident macrophages have the potential to exert enhanced osteoprotective effects by oversecreting human TSH-βv, which may exert its local osteoprotective role via osteoblast and osteoclast TSH receptors.

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