title: Fibroblast growth factor 2 dimer with superagonist in vitro activity improves granulation tissue formation during wound healing.
abstract: Site-specific chemical dimerization of fibroblast growth factor 2 (FGF2) with the optimal linker length resulted in a FGF2 homodimer with improved granulation tissue formation and blood vessel formation at exceptionally low concentrations. Homodimers of FGF2 were synthesized through site-specific linkages to both ends of different molecular weight poly(ethylene glycols) (PEGs). The optimal linker length was determined by screening dimer-induced metabolic activity of human dermal fibroblasts and found to be that closest to the inter-cysteine distance, 70 Å, corresponding to 2 kDa PEG. A straightforward analysis of the kinetics of second ligand binding as a function of tether length showed that, as the polymerization index (the number of monomer repeat units in the polymer, N) of the tether decreases, the mean time for second ligand capture decreases as ∼N(3/2), leading to an enhancement of the number of doubly bound ligands in steady-state for a given (tethered) ligand concentration. FGF2-PEG2k-FGF2 induced greater fibroblast metabolic activity than FGF2 alone, all other dimers, and all monoconjugates, at each concentration tested, with the greatest difference observed at low (0.1 ng/mL) concentration. FGF2-PEG2k-FGF2 further exhibited superior activity compared to FGF2 for both metabolic activity and migration in human umbilical vein endothelial cells, as well as improved angiogenesis in a coculture model in vitro. Efficacy in an in vivo wound healing model was assessed in diabetic mice. FGF2-PEG2k-FGF2 increased granulation tissue and blood vessel density in the wound bed compared to FGF2. The results suggest that this rationally designed construct may be useful for improving the fibroblast matrix formation and angiogenesis in chronic wound healing.
authors.affiliation: Department of Chemistry and Biochemistry and California NanoSystems Institute, University of California, Los Angeles, 607 Charles E. Young Drive South, Los Angeles, CA 90095-1569, United States.
authors.forename: Caitlin G
authors.initials: CG
authors.lastname: Decker
authors.affiliation: Department of Dermatology, Johns Hopkins University School of Medicine, 1550 Orleans St., Baltimore, MD 21231, United States.
authors.forename: Yu
authors.initials: Y
authors.lastname: Wang
authors.affiliation: Department of Chemistry and Biochemistry and California NanoSystems Institute, University of California, Los Angeles, 607 Charles E. Young Drive South, Los Angeles, CA 90095-1569, United States.
authors.forename: Samantha J
authors.initials: SJ
authors.lastname: Paluck
authors.affiliation: Department of Chemistry and Biochemistry and California NanoSystems Institute, University of California, Los Angeles, 607 Charles E. Young Drive South, Los Angeles, CA 90095-1569, United States.
authors.forename: Lu
authors.initials: L
authors.lastname: Shen
authors.affiliation: Department of Chemistry and Biochemistry and California NanoSystems Institute, University of California, Los Angeles, 607 Charles E. Young Drive South, Los Angeles, CA 90095-1569, United States; Department of Biological Chemistry, David Geffen School of Medicine, and UCLA/DOE Institute of Genomics and Proteomics, University of California, Los Angeles, CA 90095, United States.
authors.forename: Joseph A
authors.initials: JA
authors.lastname: Loo
authors.affiliation: Department of Chemistry and Biochemistry and California NanoSystems Institute, University of California, Los Angeles, 607 Charles E. Young Drive South, Los Angeles, CA 90095-1569, United States; Department of Physics and Astronomy, University of California, Los Angeles, CA 90095-1569, United States; Department of Biomathematics, University of California, Los Angeles, CA 90095-1569, United States.
authors.forename: Alex J
authors.initials: AJ
authors.lastname: Levine
authors.affiliation: Department of Dermatology, Johns Hopkins University School of Medicine, 1550 Orleans St., Baltimore, MD 21231, United States.
authors.forename: Lloyd S
authors.initials: LS
authors.lastname: Miller
authors.affiliation: Department of Chemistry and Biochemistry and California NanoSystems Institute, University of California, Los Angeles, 607 Charles E. Young Drive South, Los Angeles, CA 90095-1569, United States. Electronic address: maynard@chem.ucla.edu.
authors.forename: Heather D
authors.initials: HD
authors.lastname: Maynard
mesh_terms: D000818:Animals; D001808:Blood Vessels / Q000187:drug effects / Q000473:pathology; D015153:Blotting, Western; D002465:Cell Movement / Q000187:drug effects; D003432:Cross-Linking Reagents / Q000737:chemistry; D020405:Dermis / Q000166:cytology; D003921:Diabetes Mellitus, Experimental / Q000473:pathology; D016222:Fibroblast Growth Factor 2 / Q000819:agonists* / Q000737:chemistry / Q000494:pharmacology*; D005347:Fibroblasts / Q000166:cytology / Q000187:drug effects; D006097:Granulation Tissue / Q000187:drug effects / Q000473:pathology*; D061307:Human Umbilical Vein Endothelial Cells / Q000187:drug effects / Q000378:metabolism; D008297:Male; D051379:Mice; D018919:Neovascularization, Physiologic / Q000187:drug effects; D011092:Polyethylene Glycols / Q000737:chemistry; D055503:Protein Multimerization; D014945:Wound Healing / Q000187:drug effects*
chemical_list: D003432:Cross-Linking Reagents; D016222:Fibroblast Growth Factor 2; D011092:Polyethylene Glycols
keywords: Angiogenesis; Diabetic wounds; Dimerization; Growth factor; PEGylation; Wound healing