Novel osteoinductive photo-cross-linkable chitosan-lactide-fibrinogen hydrogels enhance bone regeneration in critical size segmental bone defects

Abstract

The purpose of this study was to develop and characterize a novel photo-cross-linkable chitosan-lactide-fibrinogen (CLF) hydrogel and evaluate the efficacy of bone morphogenetic protein-2 (BMP-2) containing CLF hydrogel for osteogenesis in vitro and in vivo. We synthesized the CLF hydrogels and characterized their chemical structure, degradation rate, compressive modulus, and in vitro BMP-2 release kinetics. We evaluated bioactivities of the BMP-2 containing CLF hydrogels (0, 50, 100, and 500 ng/ml) in vitro using W-20-17 preosteoblast mouse bone marrow stromal cells and C2C12 mouse myoblast cells. The effect of BMP-2 containing CLF gels (0, 0.5, 1, 2, and 5μg) on bone formation was evaluated using rat critical size segmental bone defects for 4 weeks. FTIR spectra and SEM images showed chemical and structural changes by addition of fibrinogen into chitosan-lactide copolymer. Incorporation of fibrinogen molecules significantly increased compressive modulus of the hydrogels. In vitro BMP-2 release study showed initial burst releases from the CLF hydrogels followed by sustained releases, regardless of the concentration of the BMP-2 over 4 weeks. Cells in all groups were viable in the presence of the hydrogels regardless of BMP-2 doses, indicating non-cytotoxicity of hydrogels. Alkaline phosphate activity and mineralization of cells exhibited dose dependence on BMP-2 containing CLF hydrogels. Radiographs, microcomputed tomography, and histology confirmed that the BMP-2 containing CLF hydrogels prompted neo-osteogenesis and accelerated healing of the defects in a dose-dependent manner. Thus the CLF hydrogel is a promising delivery system of growth factors for bone regeneration.

History and Applications of Hydrogels

“…Cytotoxicity and cell viability suggest that GDF-5loaded HA hydrogel has proper biocompatibility for use as a scaffold which can induce osteogenesis. Moreover, the HA hydrogel showed improved osteogenesis in both in-vitro tests, the results from these tests show that the HA hydrogel can be used as a scaffold for bone tissue regeneration [149]. Gelatin hydrogels were also used for bone regeneration at both the critical-sized bone defect.…”

Chitosan Hydrogels for Regenerative Engineering

“…In vitro and in vivo studies demonstrated the ability of the hydrogel to deliver nucleic acid to the surrounding tissues and cells leading to effective blocking of TNF-α [99]. Recently, Kim et al developed a photocrosslinked hydrogel made of chitosan-lactide-fibrinogen (CLF) as a delivery system for BMP-2 [100]. Chitosan and lactide in the hydrogel controlled the physiochemical properties of the hydrogel [101].…”

“…The results indicated increased cell viability, neo-osteogenesis, and bone healing. The responses were dependent on the dose of the growth factor [100]. …”

Morphological and mechanical characterization of chitosan-calcium phosphate composites for potential application as bone-graft substitutes

“…The use of β-tri-calcium phosphate that is more soluble than hydroxyapatite (Yamada et al, 1997) and chitosan, a lysozyme-biodegradable biopolymer (New et al, 2009), makes the CS-CP composite susceptible for bioresorption and biodegradation. Moreover, CS composites have been demonstrated osteoinductive effects (Jebahi et al, 2013;Kim et al, 2014;Kung et al, 2011) in vitro and in vivo. Thus, the CS-CP composites herein prepared are potentially biodegradable and osteoinductive, and demonstrated adequate porosity and mechanical properties for treating bone defects.…”