Healing and Angiogenic Properties of Collagen/Chitosan Scaffolds Enriched with Hyperstable FGF2-STAB® Protein: In Vitro, Ex Ovo and In Vivo Comprehensive Evaluation

Abstract

Wound healing is a process regulated by a complex interaction of multiple growth factors including fibroblast growth factor 2 (FGF2). Although FGF2 appears in several tissue engineered studies, its applications are limited due to its low stability both in vitro and in vivo. Here, this shortcoming is overcome by a unique nine-point mutant of the low molecular weight isoform FGF2 retaining full biological activity even after twenty days at 37 °C. Crosslinked freeze-dried 3D porous collagen/chitosan scaffolds enriched with this hyper stable recombinant human protein named FGF2-STAB® were tested for in vitro biocompatibility and cytotoxicity using murine 3T3-A31 fibroblasts, for angiogenic potential using an ex ovo chick chorioallantoic membrane assay and for wound healing in vivo with 3-month old white New Zealand rabbits. Metabolic activity assays indicated the positive effect of FGF2-STAB® already at very low concentrations (0.01 µg/mL). The angiogenic properties examined ex ovo showed enhanced vascularization of the tested scaffolds. Histological evaluation and gene expression analysis by RT-qPCR proved newly formed granulation tissue at the place of a previous skin defect without significant inflammation infiltration in vivo. This work highlights the safety and biocompatibility of newly developed crosslinked collagen/chitosan scaffolds involving FGF2-STAB® protein. Moreover, these sponges could be used as scaffolds for growing cells for dermis replacement, where neovascularization is a crucial parameter for successful skin regeneration.

Lumbar Interbody Fusion Conducted on a Porcine Model with a Bioresorbable Ceramic/Biopolymer Hybrid Implant Enriched with Hyperstable Fibroblast Growth Factor 2

“…The bioresorption is enabled via chemical depolarization and enzymatic hydrolysis mediated by glycosidases, which leads to nontoxic final products of glucuronic acid and glucose [28]. Newly, we verified the safety, healing, and angiogenic potential of collagenbased scaffolds enriched with FGF2-STAB ® as full skin replacement in vivo on 3-month-old White New Zealand rabbits [29]. Biodegradable scaffolds revealed promising results in cell culture experiments and have displayed high suitability and biocompatibility to be used as a transferable scaffold for tissue reconstruction [30].…”

“…Recently, we examined the FGF2-STAB ® release from collagen/chitosan scaffolds cross-linked with EDC/NHS system as the growth factor biological activity both in vitro on murine 3T3-A31 mouse fibroblast cells and in vivo on New Zealand white rabbits when supporting full-skin thickness regeneration [29]. Broad range of FGF2-STAB ® concentrations from 0 to 100 µg/mL was evaluated for cell metabolic activity, where lower concentrations from 0.01 up to 0.5 µg/mL showed enhanced cell metabolic activity and collagen expression compared to proteins with higher concentrations that may be beneficial for wound healing and tissue regeneration [47].…”

“…Therefore, in the present study, we used the same BCP 3D matrix as a control group for 21 days cytotoxicity evaluation of the BCP matrix impregnated with collagen, oxidized cellulose, and FGF2-STAB ® (BHI). The amount of FGF2-STAB ® protein has been widely evaluated in our previous studies [29,30].…”

A Biological Study of Composites Based on the Blends of Nanohydroxyapatite, Silk Fibroin and Chitosan

“…The mixed solution was poured into 24-well polystyrene culture plates, and frozen overnight in a −20 °C freezer, followed by lyophilization for 48 h. Dry samples were removed from the molds and etched with methanol (for 30 min) to induce crystallization and water stability. Following methanol evaporation at room temperature, the scaffolds were then crosslinked in 1-(3-dimetylaminopropyl)-3-etylcarbodiimide hydrochloride (EDC) in N-hydroxysuccinimide (NHS) [ 20 ]. Following this, the sponges were further immersed in sodium phosphate dibasic 0.1M Na 2 HPO 4 , neutralized 0.5M NaOH (eliminate the acid environment, which remained from the chitosan solvent) and rinsed several times in deionized water (in preparation for the biological tests).…”

Effect of Polymeric Nanoparticles with Entrapped Fish Oil or Mupirocin on Skin Wound Healing Using a Porcine Model

“…Rodents are often used as an animal model for testing drugs active in wound healing in humans [ 9 , 10 ]. However, the driving force behind wound closure in rodents is contraction [ 11 ], whereas the closure of human wounds is primarily accomplished through the proliferation and migration of cells at the wound edge.…”