Cross-linking methods of electrospun fibrinogen scaffolds for tissue engineering applications

Abstract

The purpose of this study was to enhance the mechanical properties and slow the degradation of an electrospun fibrinogen scaffold, while maintaining the scaffold's high level of bioactivity. Three different cross-linkers were used to achieve this goal: glutaraldehyde vapour, 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC) in ethanol and genipin in ethanol. Scaffolds with a fibrinogen concentration of 120 mg ml(-1) were electrospun and cross-linked with one of the aforementioned cross-linkers. Mechanical properties were determined through uniaxial tensile testing performed on scaffolds incubated under standard culture conditions for 1 day, 7 days and 14 days. Cross-linked scaffolds were seeded with human foreskin fibroblasts (BJ-GFP-hTERT) and cultured for 7, 14 and 21 days, with histology and scanning electron microscopy performed upon completion of the time course. Mechanical testing revealed significantly increased peak stress and modulus values for the EDC and genipin cross-linked scaffolds, with significantly slowed degradation. However, cross-linking with EDC and genipin was shown to have some negative effect on the bioactivity of the scaffolds as cell migration throughout the thickness of the scaffold was slowed.

Characterization of Leukocyte-platelet Rich Fibrin, A Novel Biomaterial

“…To prepare genipin cross-linked L-PRF, rinse membranes with PBS and soak in 4 ml of 1% genipin solution (in 70% ethanol) for 48 hr. Rinse with PBS prior to experiments to remove excess genipin 15,16 . 2.…”

Modeling early stage bone regeneration with biomimetic electrospun fibrinogen nanofibers and adipose-derived mesenchymal stem cells

“…However, prior publications with electrospun Fg matrix lacked mechanical stability to serve as tissue engineering scaffolds without crosslinking, as the Fg scaffolds completely dissociated in 1-2 weeks of culture, a problem consistently observed with electrospun collagen scaffolds [38]. While the mechanical properties of electrospun Fg may be augmented by chemical crosslinking, we have previously reported that chemical crosslinking inhibits cell growth and differentiation [37]. Blending Fg with a synthetic polymer, such as PDO, improves scaffold stability in media [33,37].…”

“…Solutions of lyophilized bovine Fg (Fraction 1, Type 1-S from bovine plasma, Sigma Aldrich) were made with a 10% (by volume) 10X minimal essential medium (MEM, Sigma Aldrich) and 90% 1,1,1,3,3,3 hexafluoro-2-propanol (HFP; TCI America, Portland, OR) at concentrations of 100 mg/mL (ratio determined to make biomimetic fiber diameters as described in our prior work [37]), and allowed to dissolve overnight with shaking. Solutions of PDO (Ethicon, Somerville, NJ) were made at 100 mg/ml concentrations, dissolved in HFP overnight with shaking at room temperature (RT).…”

“…We previously reported that crosslinking Fg by various methods leaves the scaffold impermeable to cells [37]; therefore, scaffolds were left in their native, uncrosslinked state after electrospinning. As shown in Figure 1, viable ASCs were abundantly present on pure Fg scaffold (panel B) with a modest reduction in cellularity on blended Fg:PDO scaffolds (panel A) after 2 weeks in culture.…”

The Use of Natural Polymers in Tissue Engineering: A Focus on Electrospun Extracellular Matrix Analogues

“…Cross-linking of fibrinogen scaffolds has also been used to increase their mechanical strength and slow their rate of degradation. The authors of this review [106] have used both EDC and genipin to successfully alter the properties of electrospun fibrinogen, with cross-linked scaffolds remaining mechanically viable after 14 days in culture compared to complete loss of mechanical integrity after 7 days for non-cross-linked scaffolds. The degree to which these scaffolds were cross-linked decreased the rate of scaffold remodeling, with little new collagen matrix visible in Masson's trichrome histological stains after 21 days in culture with FBs.…”