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In situ modulation of cell behavior via smart dual-ligand surfaces.

Pulsipher A, Park S, Dutta D, Luo W, Yousaf MN - Langmuir (2014)

Bottom Line: Such tools provide strategies for identifying specific ligand-receptor interactions that induce vital biological consequences.A redox-responsive trigger was incorporated into this surface strategy to spontaneously release ligands in the presence of adhered cells, and cell spreading, growth, and migration responses were measured and compared.The identity and nature of the dual-ligand combination directly influenced cell behavior.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemistry, University of North Carolina at Chapel Hill , Chapel Hill, North Carolina 27599-3290, United States.

ABSTRACT
Due to the highly complex nature of the extracellular matrix (ECM), the design and implementation of dynamic, stimuli-responsive surfaces that present well-defined ligands and serve as model ECM substrates have been of tremendous interest to biomaterials, biosensor, and cell biology communities. Such tools provide strategies for identifying specific ligand-receptor interactions that induce vital biological consequences. Herein, we report a novel dual-ligand-presenting surface methodology that modulates dynamic ECM properties to investigate various cell behaviors. Peptides PHSRN, cRGD, and KKKTTK, which mimic the cell- and heparan sulfate-binding domains of fibronectin, and carbohydrates Gal and Man were combined with cell adhesive RGD to survey possible synergistic or antagonist ligand effects on cell adhesion, spreading, growth, and migration. Soluble molecule and enzymatic inhibition assays were also performed, and the levels of focal adhesion kinase in cells subjected to different ligand combinations were quantified. A redox-responsive trigger was incorporated into this surface strategy to spontaneously release ligands in the presence of adhered cells, and cell spreading, growth, and migration responses were measured and compared. The identity and nature of the dual-ligand combination directly influenced cell behavior.

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Related in: MedlinePlus

Synthetic Scheme of HQ-RGD: (A) SolutionSynthesis of Fmoc-glycine-HQ and (B) Solid-PhasePeptide Synthesis of HQ-RGDUsing MBHA ResinReagents and conditions: (i)DHP (4.5 equiv), HCl (cat.), THF, 16 h, 67.4%; (ii) tert-butyllithium (2 equiv), 1,6-dibromohexane (2 equiv), dry THF, 0–25°C, 20 h, 50.6%; (iii) NaN3 (1.5 equiv), DMF, 2 h,80 °C; and (iv) Fmoc-propargylglycine (0.5 equiv), CuSO4·5H2O (1.5 equiv), NaAsc (1.5 equiv), DMF/H2O/EtOH (2:1:1), 12 h, 61.1%.Reagents and conditions: (i) piperidine (20%); (ii) repetitionof DIEA (3 equiv), HBTU (3 equiv), Fmoc-amino acid (SSDGRG, 3 equiv),DMF, then piperidine (20%); (iii) DIEA (3 equiv), HBTU (3 equiv),Fmoc-Gly-HQ (3 equiv), DMF, then piperidine (20%); (iv) DIEA (3 equiv),HBTU (3 equiv), Fmoc-C6-linker (3 equiv), Fmoc-Lys-N3 (3 equiv), piperidine (20%), acetic anhydride (10%), thenTFA/H2O (9.75/0.25, 2 h), ether suspension, centrifugation(2 × 2000 rpm, 10 min), dissolve and freeze in H2O(10 mL), lyophilize. Overall yield of 54 mg, 65%.
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sch1: Synthetic Scheme of HQ-RGD: (A) SolutionSynthesis of Fmoc-glycine-HQ and (B) Solid-PhasePeptide Synthesis of HQ-RGDUsing MBHA ResinReagents and conditions: (i)DHP (4.5 equiv), HCl (cat.), THF, 16 h, 67.4%; (ii) tert-butyllithium (2 equiv), 1,6-dibromohexane (2 equiv), dry THF, 0–25°C, 20 h, 50.6%; (iii) NaN3 (1.5 equiv), DMF, 2 h,80 °C; and (iv) Fmoc-propargylglycine (0.5 equiv), CuSO4·5H2O (1.5 equiv), NaAsc (1.5 equiv), DMF/H2O/EtOH (2:1:1), 12 h, 61.1%.Reagents and conditions: (i) piperidine (20%); (ii) repetitionof DIEA (3 equiv), HBTU (3 equiv), Fmoc-amino acid (SSDGRG, 3 equiv),DMF, then piperidine (20%); (iii) DIEA (3 equiv), HBTU (3 equiv),Fmoc-Gly-HQ (3 equiv), DMF, then piperidine (20%); (iv) DIEA (3 equiv),HBTU (3 equiv), Fmoc-C6-linker (3 equiv), Fmoc-Lys-N3 (3 equiv), piperidine (20%), acetic anhydride (10%), thenTFA/H2O (9.75/0.25, 2 h), ether suspension, centrifugation(2 × 2000 rpm, 10 min), dissolve and freeze in H2O(10 mL), lyophilize. Overall yield of 54 mg, 65%.


In situ modulation of cell behavior via smart dual-ligand surfaces.

Pulsipher A, Park S, Dutta D, Luo W, Yousaf MN - Langmuir (2014)

Synthetic Scheme of HQ-RGD: (A) SolutionSynthesis of Fmoc-glycine-HQ and (B) Solid-PhasePeptide Synthesis of HQ-RGDUsing MBHA ResinReagents and conditions: (i)DHP (4.5 equiv), HCl (cat.), THF, 16 h, 67.4%; (ii) tert-butyllithium (2 equiv), 1,6-dibromohexane (2 equiv), dry THF, 0–25°C, 20 h, 50.6%; (iii) NaN3 (1.5 equiv), DMF, 2 h,80 °C; and (iv) Fmoc-propargylglycine (0.5 equiv), CuSO4·5H2O (1.5 equiv), NaAsc (1.5 equiv), DMF/H2O/EtOH (2:1:1), 12 h, 61.1%.Reagents and conditions: (i) piperidine (20%); (ii) repetitionof DIEA (3 equiv), HBTU (3 equiv), Fmoc-amino acid (SSDGRG, 3 equiv),DMF, then piperidine (20%); (iii) DIEA (3 equiv), HBTU (3 equiv),Fmoc-Gly-HQ (3 equiv), DMF, then piperidine (20%); (iv) DIEA (3 equiv),HBTU (3 equiv), Fmoc-C6-linker (3 equiv), Fmoc-Lys-N3 (3 equiv), piperidine (20%), acetic anhydride (10%), thenTFA/H2O (9.75/0.25, 2 h), ether suspension, centrifugation(2 × 2000 rpm, 10 min), dissolve and freeze in H2O(10 mL), lyophilize. Overall yield of 54 mg, 65%.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4334223&req=5

sch1: Synthetic Scheme of HQ-RGD: (A) SolutionSynthesis of Fmoc-glycine-HQ and (B) Solid-PhasePeptide Synthesis of HQ-RGDUsing MBHA ResinReagents and conditions: (i)DHP (4.5 equiv), HCl (cat.), THF, 16 h, 67.4%; (ii) tert-butyllithium (2 equiv), 1,6-dibromohexane (2 equiv), dry THF, 0–25°C, 20 h, 50.6%; (iii) NaN3 (1.5 equiv), DMF, 2 h,80 °C; and (iv) Fmoc-propargylglycine (0.5 equiv), CuSO4·5H2O (1.5 equiv), NaAsc (1.5 equiv), DMF/H2O/EtOH (2:1:1), 12 h, 61.1%.Reagents and conditions: (i) piperidine (20%); (ii) repetitionof DIEA (3 equiv), HBTU (3 equiv), Fmoc-amino acid (SSDGRG, 3 equiv),DMF, then piperidine (20%); (iii) DIEA (3 equiv), HBTU (3 equiv),Fmoc-Gly-HQ (3 equiv), DMF, then piperidine (20%); (iv) DIEA (3 equiv),HBTU (3 equiv), Fmoc-C6-linker (3 equiv), Fmoc-Lys-N3 (3 equiv), piperidine (20%), acetic anhydride (10%), thenTFA/H2O (9.75/0.25, 2 h), ether suspension, centrifugation(2 × 2000 rpm, 10 min), dissolve and freeze in H2O(10 mL), lyophilize. Overall yield of 54 mg, 65%.
Bottom Line: Such tools provide strategies for identifying specific ligand-receptor interactions that induce vital biological consequences.A redox-responsive trigger was incorporated into this surface strategy to spontaneously release ligands in the presence of adhered cells, and cell spreading, growth, and migration responses were measured and compared.The identity and nature of the dual-ligand combination directly influenced cell behavior.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemistry, University of North Carolina at Chapel Hill , Chapel Hill, North Carolina 27599-3290, United States.

ABSTRACT
Due to the highly complex nature of the extracellular matrix (ECM), the design and implementation of dynamic, stimuli-responsive surfaces that present well-defined ligands and serve as model ECM substrates have been of tremendous interest to biomaterials, biosensor, and cell biology communities. Such tools provide strategies for identifying specific ligand-receptor interactions that induce vital biological consequences. Herein, we report a novel dual-ligand-presenting surface methodology that modulates dynamic ECM properties to investigate various cell behaviors. Peptides PHSRN, cRGD, and KKKTTK, which mimic the cell- and heparan sulfate-binding domains of fibronectin, and carbohydrates Gal and Man were combined with cell adhesive RGD to survey possible synergistic or antagonist ligand effects on cell adhesion, spreading, growth, and migration. Soluble molecule and enzymatic inhibition assays were also performed, and the levels of focal adhesion kinase in cells subjected to different ligand combinations were quantified. A redox-responsive trigger was incorporated into this surface strategy to spontaneously release ligands in the presence of adhered cells, and cell spreading, growth, and migration responses were measured and compared. The identity and nature of the dual-ligand combination directly influenced cell behavior.

Show MeSH
Related in: MedlinePlus