Journal: International Journal of Molecular Sciences

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

doi: 10.3390/ijms17071158

Figure Lengend Snippet: The effect of granulocyte colony-stimulating factor (G-CSF) treatment, apheresis procedures and allogeneic stem cell transplantation on plasma osteopontin (OPN; Upper part ) and G-CSF ( Lower part ) concentration. ( Upper part ) From the top, the plasma OPN levels are presented for the four study groups: (i) prior to and after G-CSF treatment of allogeneic stem cell donors; (ii) immediately before and after apheresis and in the apheresis product for each study group undergoing apheresis; and (iii) in allotransplanted patients 8–12 h prior to start of stem cell infusion and 12–16 h after infusion; ( Lower part ) Plasma G-CSF concentrations are given for allogeneic stem cell donors prior to and after G-CSF treatment and for autologous stem cell donors only after the G-CSF therapy. All concentrations are given as medians with variation ranges in parentheses.

Article Snippet: The medium was supplemented with recombinant human osteopontin 50 ng/mL (R&D Systems, Minneapolis, MN, USA) and eventually recombinant human G-CSF 10 pg/mL (PeproTech EC Ltd., Rocky Hill, NJ, USA).

Techniques: Transplantation Assay, Clinical Proteomics, Concentration Assay

Journal: International Journal of Molecular Sciences

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

doi: 10.3390/ijms17071158

Figure Lengend Snippet: Plasma osteopontin levels in healthy allogeneic stem cell donors during stem cell mobilization and harvesting. Peripheral blood plasma osteopontin concentrations were determined prior to stimulation with granulocyte colony-stimulating factor (G-CSF) (A), after stem cell mobilization and immediately prior to apheresis (B), immediately after apheresis (C) and approximately 24 h after start of apheresis (D).

Article Snippet: The medium was supplemented with recombinant human osteopontin 50 ng/mL (R&D Systems, Minneapolis, MN, USA) and eventually recombinant human G-CSF 10 pg/mL (PeproTech EC Ltd., Rocky Hill, NJ, USA).

Techniques: Clinical Proteomics

Journal: International Journal of Molecular Sciences

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

doi: 10.3390/ijms17071158

Figure Lengend Snippet: Plasma osteopontin levels in autologous stem cell donors (myeloma patients) after stem cell mobilization and immediately prior to apheresis (B), immediately after apheresis (C) and approximately 24 h after start of apheresis (D).

Article Snippet: The medium was supplemented with recombinant human osteopontin 50 ng/mL (R&D Systems, Minneapolis, MN, USA) and eventually recombinant human G-CSF 10 pg/mL (PeproTech EC Ltd., Rocky Hill, NJ, USA).

Techniques: Clinical Proteomics

Journal: International Journal of Molecular Sciences

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

doi: 10.3390/ijms17071158

Figure Lengend Snippet: Osteopontin levels in apheresis products, i.e., peripheral blood stem cell grafts and platelet concentrates. The osteopontin levels were determined in allogeneic stem cell products from G-CSF-mobilized healthy stem cell donors ( n = 22), autologous stem cell products derived from myeloma patients mobilized by chemotherapy plus G-CSF ( n = 15), and platelet concentrates from unstimulated healthy platelet donors ( n = 15). The osteopontin levels measured in platelet concentrate supernatants were adjusted for dilution of the products with platelet additive solution (37% plasma, 63% solution).

Article Snippet: The medium was supplemented with recombinant human osteopontin 50 ng/mL (R&D Systems, Minneapolis, MN, USA) and eventually recombinant human G-CSF 10 pg/mL (PeproTech EC Ltd., Rocky Hill, NJ, USA).

Techniques: Derivative Assay, Clinical Proteomics

Journal: International Journal of Molecular Sciences

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

doi: 10.3390/ijms17071158

Figure Lengend Snippet: Peripheral blood mononuclear cells (PBMC) from eight healthy unstimulated donors were cultured in serum-free medium and stimulated with anti-CD3 and anti-CD28. The effect of osteopontin 50 ng/mL without G-CSF ( left ) and with G-CSF 10 pg/mL ( right ) on in vitro T cell proliferation was assayed as 3 H-thymidine incorporation expressed as median counts per minute (cpm). The proliferation of normal PBMC in control cultures containing isotypic control antibodies instead of anti-CD3/anti-CD28 antibodies corresponded to <1000 cpm.

Article Snippet: The medium was supplemented with recombinant human osteopontin 50 ng/mL (R&D Systems, Minneapolis, MN, USA) and eventually recombinant human G-CSF 10 pg/mL (PeproTech EC Ltd., Rocky Hill, NJ, USA).

Techniques: Cell Culture, In Vitro, Control

Journal: Laboratory investigation; a journal of technical methods and pathology

Article Title: Osteopontin isoforms differentially promote arteriogenesis in response to ischemia via macrophage accumulation and survival.

doi: 10.1038/s41374-018-0094-8

Figure Lengend Snippet: Fig. 3 OPN isoforms differentially influence macrophage accumula- tion in vivo. a To determine if OPN isoforms differentially promote macrophage accumulation, histology sections from WT, OPN−/−, and OPN isoform a-, b-, or c-treated animals were stained for CD68 (macrophage marker; pink stain) 5 days post HLI. The 20× repre- sentative images show detail. The 2.5× images stitched together across the hindlimb demonstrate tissue-wide differences. Scale bars for 2.5× stitched = 200 μm. Scale bars for 20× = 50 μm. b To quantitatively assess macrophage accumulation in response to OPN isoforms, qRT- PCR on mRNA isolated from the muscles of the IL was used to quantify CD68 expression. CD68 expression was compared across all treatment groups at day 7; #p < 0.05 vs. OPN−/−, LV-GFP; *p < 0.05 vs. OPN−/−, LV-GFP, and LV-OPNb, †p < 0.001 vs. OPN−/−, LV- GFP, LV-OPNa, and LV-OPNb; n = 4

Article Snippet: For macrophage polarization studies, 3 h after plating cells were stimulated with 10% FBS–RPMI with 100 ng/mL purified recombinant human OPNa, OPNb, or OPNc (cat# TP304803, TP305118, and TP31059; Origene, Rockville, MD), or were stimulated with 20 ng/mL of either interferon gamma (INFγ) or interleukin (IL)-4 for 48 h (cat# 554587 and 550067; BD Biosciences) as positive controls for macrophage polarization.

Techniques: In Vivo, Staining, Marker, Quantitative RT-PCR, Isolation, Muscles, Expressing

Journal: Laboratory investigation; a journal of technical methods and pathology

Article Title: Osteopontin isoforms differentially promote arteriogenesis in response to ischemia via macrophage accumulation and survival.

doi: 10.1038/s41374-018-0094-8

Figure Lengend Snippet: Fig. 6 Effects of MCP-1 and OPN isoforms on macrophage migration in vitro. Peritoneal macrophages were used as an in vitro cell model to measure isoform effects on macrophage migration using a modified Boyden chamber assay. a Monocyte chemoattractant protein-1 (MCP-1), a known stimulus for macrophage migration, was used to first determine if OPN−/−and WT macrophages exhibit differences in MCP-1-mediated migration after 2 h (n = 4, *p = 0.0027 vs. WT−no trx, †p < 0.001 vs. WT+ MCP-1). b To assess if OPN isoforms differentially promote macrophage migration, OPN−/−macrophages were stimulated with 50 ng/mL of recombinant OPN a, b, or c and macrophage migration was quantified after 2 h (n = 4, *p < 0.05 vs. no trx and OPNb, †p < 0.001 vs. no trx, OPNa, and OPNb). c Proposed mechanism by which OPN isoforms a and c increase arteriogenesis in vivo by increasing macrophage migration and survival

Article Snippet: For macrophage polarization studies, 3 h after plating cells were stimulated with 10% FBS–RPMI with 100 ng/mL purified recombinant human OPNa, OPNb, or OPNc (cat# TP304803, TP305118, and TP31059; Origene, Rockville, MD), or were stimulated with 20 ng/mL of either interferon gamma (INFγ) or interleukin (IL)-4 for 48 h (cat# 554587 and 550067; BD Biosciences) as positive controls for macrophage polarization.

Techniques: Migration, In Vitro, Boyden Chamber Assay, Recombinant, In Vivo