Paradoxical drop in circulating neutrophil count following granulocyte-colony stimulating factor and stem cell factor administration in rhesus macaques

Background Stem cell factor (SCF) and granulocyte-colony stimulating factor (G-CSF) are well-characterized vital hematopoietic growth factors that regulate hematopoiesis. G-CSF and SCF synergistically exhibit a stimulatory effect on hematopoietic progenitors. The combination of G-CSF and SCF has been used for mobilization of peripheral blood progenitor cells in cancer and non-cancerous conditions. To overcome challenges connected with the administration of two cytokines, we developed two fusion proteins composed of human SCF and human G-CSF interspaced by an alpha-helix-forming peptide linker. Methods The recombinant proteins SCF-Lα-GCSF and GCSF-Lα-SCF were purified in three steps using an ion-exchange and mixed-mode chromatography. The purity and quantity of the proteins after each stage of purification was assessed using RP-HPLC, SDS-PAGE, and the Bradford assays. Purified proteins were identified using high-performance liquid chromatography/electrospray ionization mass spectrometry (HPLC/ESI-MS) and the Western blot analyses. The molecular weight was determined by size exclusion HPLC (SE-HPLC). The activity of heterodimers was assessed using cell proliferation assays in vitro. The capacity of recombinant fusion proteins to stimulate the increase of the absolute neutrophil count in rats was determined in vivo. The binding kinetics of the proteins to immobilized G-CSF and SCF receptors was measured using total internal reflection ellipsometry and evaluated by a standard Langmuir kinetics model. Results The novel SCF-Lα-GCSF and GCSF-Lα-SCF proteins were synthesized in Escherichia coli. The purity of the heterodimers reached >90% as determined by RP-HPLC. The identity of the proteins was confirmed using the Western blot and HPLC/ESI-MS assays. An array of multimeric forms, non-covalently associated dimers or trimers were detected in the protein preparations by SE-HPLC. Each protein induced a dose-dependent proliferative response on the cell lines. At equimolar concentration, the heterodimers retain 70–140% of the SCF monomer activity (p ≤ 0.01) in promoting the M-07e cells proliferation. The G-CSF moiety in GCSF-Lα-SCF retained 15% (p ≤ 0.0001) and in SCF-Lα-GCSF retained 34% (p ≤ 0.01) of the monomeric G-CSF activity in stimulating the growth of G-NFS-60 cells. The obtained results were in good agreement with the binding data of each moiety in the fusion proteins to their respective receptors. The increase in the absolute neutrophil count in rats caused by the SCF-Lα-GCSF protein corresponded to the increase induced by a mixture of SCF and G-CSF.

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Reparative Effects of Stem Cell Factor and Granulocyte Colony-Stimulating Factor in Aged APP/PS1 Mice

Aging and Disease ◽

10.14336/ad.2020.0201 ◽

2020 ◽

Vol 11 (6) ◽

pp. 1423

Author(s):

Xingzhi Guo ◽

Yanying Liu ◽

David Morgan ◽

Li-Ru Zhao

Keyword(s):

Stem Cell ◽

Stem Cell Factor ◽

Granulocyte Colony Stimulating Factor ◽

Colony Stimulating Factor ◽

Stimulating Factor

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Granulocyte-colony Stimulating Factor in Combination with Stem Cell Factor Confers Greater Neuroprotection after Hypoxic–Ischemic Brain Damage in the Neonatal Rats than a Solitary Treatment

Translational Stroke Research ◽

10.1007/s12975-012-0225-2 ◽

2012 ◽

Vol 4 (2) ◽

pp. 171-178 ◽

Cited By ~ 34

Author(s):

Desislava Doycheva ◽

Gary Shih ◽

Hank Chen ◽

Richard Applegate ◽

John H. Zhang ◽

...

Keyword(s):

Stem Cell ◽

Brain Damage ◽

Stem Cell Factor ◽

Granulocyte Colony Stimulating Factor ◽

Neonatal Rats ◽

Colony Stimulating Factor ◽

Ischemic Brain Damage ◽

Ischemic Brain ◽

Stimulating Factor

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Stem cell factor in combination with granulocyte colony-stimulating factor (CSF) or granulocyte-macrophage CSF synergistically increases granulopoiesis in vivo

Blood ◽

10.1182/blood.v78.8.1954.1954 ◽

1991 ◽

Vol 78 (8) ◽

pp. 1954-1962 ◽

Cited By ~ 41

Author(s):

TR Ulich ◽

J del Castillo ◽

IK McNiece ◽

ES Yi ◽

CP Alzona ◽

...

Keyword(s):

Stem Cell ◽

Peripheral Blood ◽

Stem Cell Factor ◽

Granulocyte Colony Stimulating Factor ◽

Colony Stimulating Factor ◽

Gm Csf ◽

Colony Forming Unit ◽

Stimulating Factor ◽

Synergistic Increase

Abstract Recombinant rat stem cell factor (rrSCF) and recombinant human granulocyte colony-stimulating factor (G-CSF) coinjected for 1 week in rats cause a synergistic increase in mature marrow neutrophils accompanied by a striking decrease in erythroid and lymphoid marrow elements. The spleens of the same rats show increased granulopoiesis as well as increased erythropoiesis as compared with the spleens of rats treated with either growth factor alone. Splenic extramedullary erythropoiesis may act to compensate for the decrease in marrow erythropoiesis. The coinjection of rrSCF and G-CSF causes an increase in marrow mast cells at the end of 1 week, but the increase is much less than in rrSCF-alone-treated rats. The combination of rrSCF and G- CSF increases the rate of release of marrow neutrophils into the circulation and causes a dramatic synergistic peripheral neutrophilia, beginning especially after 4 days of treatment. Colony-forming assays of all experimental groups showed a synergistic increase in colony- forming unit granulocyte-macrophage (CFU-GM) in the marrow, but not in peripheral blood, after coincubation with SCF plus granulocyte- macrophage CSF (GM-CSF) as opposed to GM-CSF alone, showing anatomic compartmentalization between a more primitive marrow CFU-GM subset and a more mature peripheral blood CFU-GM subset. In vivo daily administration of SCF plus GM-CSF results in a synergistic increase in marrow neutrophils, but not the striking synergistic increase in circulating neutrophils that is observed with SCF plus G-CSF.

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Stem Cell Factor in Combination with Granulocyte Colony-Stimulating Factor reduces Cerebral Capillary Thrombosis in a Mouse Model of CADASIL

Cell Transplantation ◽

10.1177/0963689718766460 ◽

2018 ◽

Vol 27 (4) ◽

pp. 637-647 ◽

Cited By ~ 6

Author(s):

Suning Ping ◽

Xuecheng Qiu ◽

Maria E Gonzalez-Toledo ◽

Xiaoyun Liu ◽

Li-Ru Zhao

Keyword(s):

Bone Marrow ◽

Stem Cell ◽

Mouse Model ◽

Stem Cell Factor ◽

Granulocyte Colony Stimulating Factor ◽

Cerebral Thrombosis ◽

Colony Stimulating Factor ◽

Cerebral Capillaries ◽

Cerebral Capillary ◽

Stimulating Factor

Cerebral autosomal dominant arteriopathy with subcortical infarcts and leucoencephalopathy (CADASIL) is a cerebral small vascular disease caused by NOTCH3 mutation-induced vascular smooth muscle cell (VSMC) degeneration, leading to ischemic stroke and vascular dementia. Our previous study has demonstrated that repeated treatment with a combination of stem cell factor (SCF) and granulocyte colony-stimulating factor (G-CSF) reduces VSMC degeneration and cerebral endothelial cell (EC) damage and improves cognitive function in a mouse model of CADASIL (TgNotch3R90C). This study aimed to determine whether cerebral thrombosis occurs in TgNotch3R90C mice and whether repeated SCF+G-CSF treatment reduces cerebral thrombosis in TgNotch3R90C mice. Using the approaches of bone marrow transplantation to track bone marrow-derived cells and confocal imaging, we observed bone marrow-derived blood cell occlusion in cerebral small vessels and capillaries (thrombosis). Most thrombosis occurred in the cerebral capillaries (93% of total occluded vessels), and the thrombosis showed an increased frequency in the regions of capillary bifurcation. Degenerated capillary ECs were seen inside and surrounding the thrombosis, and the bone marrow-derived ECs were also found next to the thrombosis. IgG extravasation was seen in and next to the areas of thrombosis. SCF+G-CSF treatment significantly reduced cerebral capillary thrombosis and IgG extravasation. These data suggest that the EC damage is associated with thrombosis and blood–brain barrier leakage in the cerebral capillaries under the CADASIL-like condition, whereas SCF+G-CSF treatment diminishes these pathological alterations. This study provides new insight into the involvement of cerebral capillary thrombosis in the development of CADASIL and potential approaches to reduce the thrombosis, which may restrict the pathological progression of CADASIL.

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Efficient retrovirus transduction of mouse pluripotent hematopoietic stem cells mobilized into the peripheral blood by treatment with granulocyte colony-stimulating factor and stem cell factor

Blood ◽

10.1182/blood.v84.5.1482.1482 ◽

1994 ◽

Vol 84 (5) ◽

pp. 1482-1491 ◽

Cited By ~ 81

Author(s):

DM Bodine ◽

NE Seidel ◽

MS Gale ◽

AW Nienhuis ◽

D Orlic

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Gene Transfer ◽

Hematopoietic Stem Cells ◽

Peripheral Blood ◽

Stem Cell Factor ◽

Granulocyte Colony Stimulating Factor ◽

Colony Stimulating Factor ◽

Hematopoietic Stem ◽

Stimulating Factor

Abstract Cytokine-mobilized peripheral blood cells have been shown to participate in hematopoietic recovery after bone marrow (BM) transplantation, and are proposed to be useful targets for retrovirus- mediated gene transfer protocols. We treated mice with granulocyte colony-stimulating factor (G-CSF) and stem cell factor (SCF) to mobilize hematopoietic progenitor cells into the peripheral blood. These cells were analyzed for the number and frequency of pluripotent hematopoietic stem cells (PHSC). We found that splenectomized animals treated for 5 days with G-CSF and SCF showed a threefold increase in the absolute number of PHSC over normal mice. The number of peripheral- blood PHSC increased 250-fold from 29 per untreated mouse to 7,200 in peripheral-blood PHSC in splenectomized animals treated for 5 days with G-CSF and SCF. Peripheral blood PHSC mobilized by treatment with G-CSF and SCF were analyzed for their ability to be transduced by retroviral vectors. Peripheral-blood PHSC from splenectomized animals G-CSF and SCF were transduced with a recombinant retrovirus containing the human MDR-1 gene. The frequency of gene transfer into peripheral blood PHSC from animals treated for 5 and 7 days was two-fold and threefold higher than gene transfer into PHSC from the BM of 5-fluorouracil-treated mice (P < .01). We conclude that peripheral blood stem cells mobilized by treatment with G-CSF and SCF are excellent targets for retrovirus- mediated gene transfer.

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Recombinant rat stem cell factor synergizes with recombinant human granulocyte colony-stimulating factor in vivo in mice to mobilize peripheral blood progenitor cells that have enhanced repopulating potential

Blood ◽

10.1182/blood.v82.6.1720.1720 ◽

1993 ◽

Vol 82 (6) ◽

pp. 1720-1723 ◽

Cited By ~ 91

Author(s):

RA Briddell ◽

CA Hartley ◽

KA Smith ◽

IK McNiece

Keyword(s):

Stem Cell ◽

Progenitor Cells ◽

Peripheral Blood ◽

Stem Cell Factor ◽

Granulocyte Colony Stimulating Factor ◽

Colony Stimulating Factor ◽

Peripheral Blood Progenitor Cells ◽

Stimulating Factor ◽

Synergistic Increase

Abstract Splenectomized mice treated for 7 days with pegylated recombinant rat stem cell factor (rrSCF-PEG) showed a dose-dependent increase in peripheral blood progenitor cells (PBPC) that have enhanced in vivo repopulating potential. A dose of rrSCF-PEG at 25 micrograms/kg/d for 7 days produced no significant increase in PBPC. However, when this dose of rrSCF-PEG was combined with an optimal dose of recombinant human granulocyte colony-stimulating factor (rhG-CSF; 200 micrograms/kg/d), a synergistic increase in PBPC was observed. Compared with treatment with rhG-CSF alone, the combination of rrSCF-PEG plus rhG-CSF resulted in a synergistic increase in peripheral white blood cells, in the incidence and absolute numbers of PBPC, and in the incidence and absolute numbers of circulating cells with in vivo repopulating potential. These data suggest that low doses of SCF, which would have minimal, if any, effects in vivo, can synergize with optimal doses of rhG-CSF to enhance the mobilization of PBPC stimulated by rhG-CSF alone.

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