Sdf 1α

Manufactured by Thermo Fisher Scientific

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SDF-1α is a recombinant protein produced in E. coli. It is a chemokine that belongs to the CXC chemokine family. SDF-1α is involved in the regulation of various cellular processes.

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Recombinant human SDF-1α (13 citations) SDF-1α/CXCL12 (9 citations) Human SDF-1α (8 citations) Recombinant SDF-1α (4 citations) Recombinant murine SDF-1α (CXCL12) (4 citations) Recombinant human SDF-1α/CXCL12 (4 citations) Stromal Cell-derived Factor-1α (SDF-1α) (2 citations) Recombinant (CXCL12) SDF-1α (2 citations) Recombinant murine SDF-1α (2 citations) Recombinant mouse stromal cell-derived factor-1α (SDF-1α) (2 citations)

85 protocols using sdf 1α

Heparin-Modulated SDF-1α Release from Hydrogels

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To determine the effect of increasing heparin content on the release of soluble SDF-1α from our hydrogels, we first formed gels as described above (same 4 ratios), with the following addition: prior to crosslinking, we added 75 μL of a 2000 ng/mL solution of SDF-1α (Peprotech) in ultrapure (Type 1) water per gram of polysaccharide solution. Additionally, we sterile filtered (0.22 μm, Pall Corporation) the heparin, SDF-1α, and photoinitiator solutions, and autoclaved the MAC solution prior to crosslinking. We carried out the entire release study under sterile conditions.
We prepared 100 μL hydrogels for each condition (n = 4) and placed them in 1.5 mL centrifuge tubes (sterilized by autoclave). To each tube, we added 500 μL sterile phosphate buffered saline (PBS, pH = 7.4). We placed the gels in an incubator set to 37 °C. At specified timepoints (1.5h, 3h, 6h, 12h, 24h. then daily for 2 weeks), we removed the entire volume of PBS and replaced it with fresh PBS.^{27 (link)} We stored the collected release samples at −80 °C for the duration of the study. To determine the amount of released SDF-1α at each timepoint, we performed an enzyme-linked immunosorbent assay (ELISA, Peprotech) according to the manufacturer’s instructions. We calculated the cumulative amount of SDF-1α released at each timepoint, and propagated the error through each successive calculation.

Ham T.R., Cox D.G, & Leipzig N.D. (2019). Concurrent delivery of soluble and immobilized proteins to recruit and differentiate neural stem cells. Biomacromolecules, 20(9), 3445-3452.

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SDF1α Stimulation Kinetics

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Cells were plated at 1 × 10⁵ - 1×10⁶ in 6 well plates and either stimulated with 100 ng/mL SDF1α (PeproTech) for 0, 2, 10 and 30 minutes, or with 1 μg/mL SDF1α for 48 hours in serum-free media.

Buss M.C., Remke M., Lee J., Gandhi K., Schniederjan M.J., Kool M., Northcott P.A., Pfister S.M., Taylor M.D, & Castellino R.C. (2014). The WIP1 Oncogene Promotes Progression and Invasion of Aggressive Medulloblastoma Variants. Oncogene, 34(9), 1126-1140.

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SDF1α Stimulation Kinetics

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Hydrogel Crosslinking with BMP-7 and SDF-1α

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BMP-7 (Miltenyi Biotec B.V. Co. KG, Bergisch Gladbach, Deutschland) and SDF-1α (Peprotech Inc. Rocky Hill, SC, USA) were diluted in RB to get the desired concentration (BMP-7 100/500 ng, SDF-1α 500 ng). Since this leads to a smaller RB concentration, a higher RB concentration was used to get a final concentration of 0.01% on the sheets. After 2 h of swelling, crosslinking occurred.

Braun J., Eckes S., Kilb M.F., Fischer D., Eßbach C., Rommens P.M., Drees P., Schmitz K., Nickel D, & Ritz U. (2021). Mechanical characterization of rose bengal and green light crosslinked collagen scaffolds for regenerative medicine. Regenerative Biomaterials, 8(6), rbab059.

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SDF-1α Loading on OPF/BP Hydrogels

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The recombinant murine SDF-1α (PeproTech, Rocky Hill, NJ) was loaded onto the microporous OPF/BP by adsorption. Firstly, the SDF-1α was diluted in phosphate-buffered saline (PBS) (Gibco, Carlsbad, CA) in concentrations of 0 ng/ml, 250 ng/ml, 500 ng/ml, and 1000 ng/ml. Then, 50 μl SDF-1α solutions at different concentrations were pipetted evenly on one side of hydrogels. After 1 hour, other 50 μl SDF-1α solutions were pipetted evenly on the other side of the hydrogels. SDF-1α was loaded onto the OPF/BP hydrogels to generate four groups of composites with final SDF-1α concentrations of 0 ng/ml (control), 50 ng/ml (SDF50), 100 ng/ml (SDF100), or 200 ng/ml (SDF200), according to the culture system of 500 μl. Finally, the SDF-1α/OPF/BP composites were dried in the hood for 4 hours prior to further use.

Li L., Liu X., Gaihre B., Park S., Li Y., Terzic A, & Lu L. (2021). SDF-1α/OPF/BP Composites Enhance the Migrating and Osteogenic Abilities of Mesenchymal Stem Cells. Stem Cells International, 2021, 1938819.

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Transwell Assay for Fused RPMI8226 Cells

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Fused RPMI8226 cells and RPMI8226 cells (1 × 10⁴/100 μ L/well) were seeded into 96-well culture plates (Corning, USA) at 37° C for 24, 48, 72 and 96 h. The cells were incubated with CCK-8 (10 μL/well) (Dojindo, Japan) for 3 h. Then, optical density (OD) values were determined by an enzyme-labeled instrument.
We performed a transwell migration assay (Costar, Cambridge, MA, USA) using the fusion cell in the presence of BM-MSCs, which were cultured in the lower chambers. Three groups were included in the assay: (1) control, upper chamber: fused RPMI8226 cells, and lower chamber: DMEM-LG supplemented with 0.5% FBS; (2) MM cell, upper chamber: RPMI8226 cells, and lower chamber: DMEM-LG supplemented with 0.5% FBS and 50 ng/ml SDF-1α (Peprotech, Rocky Hill, NJ, USA); (3) fusion cell, upper chamber: fused RPMI8226, and lower chamber: DMEM-LG supplemented with 0.5% FBS and 50 ng/ml SDF-1α. In brief, the fused cells and RPMI8226 cells were suspended in 0.5% FBS medium, and 5 × 10⁵ cells were placed in the upper chambers of the transwell plates with or without SDF-1αin the lower chambers. After 4 h at 37° C, cells that migrated to the lower chambers were counted. Triplicate experiments were performed in each group, and the means and standard deviations were calculated.

Wang Z., Yuan Y., Zhang L., Min Z., Zhou D., Yu S., Wang P., Ju S., Jun L, & Fu J. (2018). Impact of cell fusion in myeloma marrow microenvironment on tumor progression. Oncotarget, 9(57), 30997-31006.

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Fabrication of Biodegradable Vascular Grafts

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Biodegradable TEVGs (4 mm diameter, ≈400 μm thickness and 4 cm length) were fabricated using emulsion electrospinning (Nanon-01A, MECC, Tokyo, Japan) from PHBV (403105, Sigma-Aldrich, Saint Louis, MO, USA): PCL (440744, Sigma-Aldrich, Saint Louis, MO, USA) (5:10%)/chloroform (366927, Sigma-Aldrich, Saint Louis, MO, USA) solution using the following parameters: 23 kV voltage, 0.5 mL/h feed rate, 2 mm rotating drum diameter, 22G needle, and 150 mm tip-to-collector distance. Abovementioned polymer ratio was determined in our previous studies [25 (link),31 (link),32 (link)]. In all these investigations, PHBV/PCL vascular grafts did not show any signs of dissolution as long as 1 year after implantation into rat abdominal aorta. Either VEGF (V7259, Sigma-Aldrich, St. Louis, MO, USA), bFGF (SRP4037, Sigma-Aldrich, St. Louis, MO, USA), or SDF-1α (SRP3276, Sigma-Aldrich, St. Louis, MO, USA) were dissolved in phosphate buffered saline (10010023, Thermo Fisher Scientific, Waltman, MA, USA) to 10 µg/mL concentration and then added to PHBV/PCL/chloroform solution (1:20), with the final concentration of 500 ng/mL. Grafts with the combination of VEGF, bFGF, and SDF-1α were two-layered, with the inner layer fabricated using 27G needle and containing VEGF (500 ng/mL) and the outer layer prepared utilizing 22G needle and containing bFGF and SDF-1α (500 ng/mL each).

Antonova L.V., Krivkina E.O., Sevostianova V.V., Mironov A.V., Rezvova M.A., Shabaev A.R., Tkachenko V.O., Krutitskiy S.S., Khanova M.Y., Sergeeva T.Y., Matveeva V.G., Glushkova T.V., Kutikhin A.G., Mukhamadiyarov R.A., Deeva N.S., Akentieva T.N., Sinitsky M.Y., Velikanova E.A, & Barbarash L.S. (2021). Tissue-Engineered Carotid Artery Interposition Grafts Demonstrate High Primary Patency and Promote Vascular Tissue Regeneration in the Ovine Model. Polymers, 13(16), 2637.

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Dual-layer Vascular Graft Fabrication

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Small-diameter vascular grafts were fabricated using emulsion electrospinning (Nanon-01A, MECC, Tokyo, Japan) from PHBV (403105, Sigma-Aldrich, Saint Louis, MO, USA):PCL (440744, Sigma-Aldrich, Saint Louis, MO, USA) (5%:10%)/chloroform (366927, Sigma-Aldrich, Saint Louis, MO, USA) solution using the following parameters: 23 kV voltage, 0.5 mL/h feed rate, 2 mm rotating drum diameter, 22G needle, and 150 mm tip-to-collector distance. Either VEGF (SRP4365, Sigma-Aldrich, St. Louis, MO, USA), bFGF (SRP4039, Sigma-Aldrich, St. Louis, MO, USA), or SDF-1α (SRP3252, Sigma-Aldrich, St. Louis, MO, USA) were dissolved in phosphate buffered saline (10010023, Thermo Fisher Scientific, Waltman, MA, USA) to 10 µg/mL concentration and then added to PHBV/PCL/chloroform solution (1:20), with the final concentration of 500 ng/mL. Grafts with the combination of VEGF, bFGF, and SDF-1α were two-layered, with the inner layer fabricated using 27G needle and containing VEGF (500 ng/mL) and the outer layer prepared utilizing 22G needle and containing bFGF and SDF-1α (500 ng/mL each).

Antonova L., Kutikhin A., Sevostianova V., Velikanova E., Matveeva V., Glushkova T., Mironov A., Krivkina E., Shabaev A., Senokosova E, & Barbarash L. (2021). bFGF and SDF-1α Improve In Vivo Performance of VEGF-Incorporating Small-Diameter Vascular Grafts. Pharmaceuticals, 14(4), 302.

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Hydrogel-Based Cell Migration Promotion

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The HA/gelatin hydrogel (HyStem-HP, ESI-BIO, Alameda, CA) was prepared as described elsewhere.^{[17 (link),43 ,50 (link)]} Briefly, sterile water was mixed with 0.05% w/v 2-hydroxy-4′-(2-hydroxyethoxy)-2-methylpropiophenone photoinitiator (Sigma Aldrich, St. Louis, MO) and then used to produce solutions of thiol-modified hyaluronan (Heparasil), thiol-modified gelatin (Gelin-S), and thiol-reactive PEGDA crosslinker (Extralink) at concentrations of 1% w/v each. The Heparasil, Gelin-S, and Extralink solutions were then mixed at a ratio of 2:2:1 v/v, respectively, to form the HA hydrogel precursor. For experiments requiring downstream dissolving of hydrogel, the PEGDA crosslinker was replaced with PEGSSDA crosslinker. Remaining components were prepared as described above. To accelerate cell migration in co-culture, 200 ng mL⁻¹ of human stromal cell-derived factor 1-alpha (SDF-1α or CXCL12, Peprotech, Inc., Rocky Hill, NJ) were added to the PEGSSDA mixture.

Rajan S.A., Skardal A, & Hall A.R. (2020). Multi-Domain Photopatterned 3D Tumor Constructs in a Micro-Physiological System for Analysis, Quantification, and Isolation of Infiltrating Cells. Advanced biosystems, 4(4), e1900273.

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Trans-well migration of megakaryocytes

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Mk migration and invasion assays were performed as described previously (36 (link)). Briefly, 8-μm trans-well migration inserts (Millipore) were coated with 25 μg/ml type III collagen overnight at 4 °C. Mks (25 × 10³) were seeded on the upper well in 100 μl of StemSpan and incubated at 37 °C and 5% CO₂. After 16 h, Mks that had passed through the trans-well to the other side of the filters and in the outer wells, which contained StemSpan medium with 100 ng/ml SDF1-α (PeproTech, London, UK), were recovered and counted under an inverted microscope. Thereafter, the upper side of the filters was carefully washed with cold PBS, and cells remaining on the upper face of the filters were removed with a cotton wool swab. Trans-well filters were fixed in 4% paraformaldehyde for 20 min at room temperature, stained using a monoclonal antibody against CD61 (Santa Cruz Biotechnology) and with Hoechst 33258, cut out with a scalpel, and mounted onto glass slides, putting the lower face on the top. Each experiment was performed in at least triplicates. Data are expressed as numbers of total migrated cells per insert or as percentages of cells related to that of the control. Images were acquired using an Olympus BX51 using ×20/0.5 UPlanF1 objective.

An B., Abbonante V., Xu H., Gavriilidou D., Yoshizumi A., Bihan D., Farndale R.W., Kaplan D.L., Balduini A., Leitinger B, & Brodsky B. (2015). Recombinant Collagen Engineered to Bind to Discoidin Domain Receptor Functions as a Receptor Inhibitor. The Journal of Biological Chemistry, 291(9), 4343-4355.

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