title: Enzymatically cross-linked gelatin-phenol hydrogels with a broader stiffness range for osteogenic differentiation of human mesenchymal stem cells.
abstract: An injectable hydrogel system, composed of gelatin-hydroxyphenylpropionic acid (Gtn-HPA) conjugates chemically cross-linked by an enzyme-mediated oxidation reaction, has been designed as a biodegradable scaffold for tissue engineering. In light of the role of substrate stiffness on cell differentiation, we herein report a newly improved Gtn hydrogel system with a broader range of stiffness control that uses Gtn-HPA-tyramine (Gtn-HPA-Tyr) conjugates to stimulate the osteogenic differentiation of human mesenchymal stem cells (hMSCs). The Gtn-HPA-Tyr conjugate was successfully synthesized through a further conjugation of Tyr to Gtn-HPA conjugate by means of a general carbodiimide/active ester-mediated coupling reaction. Proton nuclear magnetic resonance and UV-visible measurements showed a higher total phenol content in the Gtn-HPA-Tyr conjugate than that content in the Gtn-HPA conjugate. The Gtn-HPA-Tyr hydrogels were formed by the oxidative coupling of phenol moieties catalyzed by hydrogen peroxide (H(2)O(2)) and horseradish peroxidase (HRP). Rheological studies revealed that a broader range of storage modulus (G') of Gtn-HPA-Tyr hydrogel (600-26,800 Pa) was achieved using different concentrations of H(2)O(2), while the G' of the predecessor Gtn-HPA hydrogels was limited to the range of 1000 to 13,500 Pa. The hMSCs on Gtn-HPA-Tyr hydrogel with G' greater than 20,000 showed significantly up-regulated expressions of osteocalcin and runt-related transcription factor 2 (RUNX2) on both the gene and protein level, with the presence of alkaline phosphatase, and the evidence of calcium accumulation. These studies with the Gtn-HPA-Tyr hydrogel with G' greater than 20,000 collectively suggest the stimulation of the hMSCs into osteogenic differentiation, while these same observations were not found with the Gtn-HPA hydrogel with a G' of 13,500.
authors.affiliation: Institute of Bioengineering and Nanotechnology, The Nanos, Singapore, Singapore.
authors.forename: Li-Shan
authors.initials: LS
authors.lastname: Wang
authors.forename: Chan
authors.initials: C
authors.lastname: Du
authors.forename: Joo Eun
authors.initials: JE
authors.lastname: Chung
authors.forename: Motoichi
authors.initials: M
authors.lastname: Kurisawa
mesh_terms: D001672:Biocompatible Materials / Q000138:chemical synthesis; D002454:Cell Differentiation; D002478:Cells, Cultured; D003432:Cross-Linking Reagents / Q000737:chemistry; D055119:Elastic Modulus; D005780:Gelatin / Q000737:chemistry; D006801:Humans; D020100:Hydrogels / Q000737:chemistry*; D008422:Materials Testing; D059630:Mesenchymal Stem Cells / Q000166:cytology* / Q000502:physiology; D010006:Osteoblasts / Q000166:cytology* / Q000502:physiology; D010012:Osteogenesis / Q000502:physiology*; D019800:Phenol / Q000737:chemistry*; D023822:Tissue Engineering / Q000379:methods*
chemical_list: D001672:Biocompatible Materials; D003432:Cross-Linking Reagents; D020100:Hydrogels; D019800:Phenol; D005780:Gelatin