In vitro and in vivo Evaluation of the Bioactive Nanofibers-Encapsulated Benzalkonium Bromide for Accelerating Wound Repair with MRSA Skin Infection

Abstract

Purpose: Developing the ideal drug or dressing is a serious challenge to controlling the occurrence of antibacterial infection during wound healing. Thus, it is important to prepare novel nanofibers for a wound dressing that can control bacterial infections. In our study, the novel self-assembled nanofibers of benzalkonium bromide with bioactive peptide materials of IKVAV and RGD were designed and fabricated.

Methods: Different drug concentration effects of encapsulation efficacy, swelling ratio and strength were determined. Its release profile in simulated wound fluid and its cytotoxicity were studied in vitro. Importantly, the antibacterial efficacy, inhibition of biofilm formation effect and wound healing against MRSA infections in vitro and in vivo were performed after observing the tissue toxicity in vivo.

Results: It was found that the optimized drug load (0.8%) was affected by the encapsulation efficacy, swelling ratio, and strength. In addition, the novel nanofibers with average diameter (222.0 nm) and stabile zeta potential (-11.2 mV) have good morphology and characteristics. It has a delayed released profile in the simulated wound fluid and good biocompatibility with L929 cells and most tissues. Importantly, the nanofibers were shown to improve antibacterial efficacy, inhibit biofilm formation, and lead to accelerated wound healing following infection with methicillin-resistant Staphylococcus aureus.

Conclusion: These data suggest that novel nanofibers could effectively shorten the wound-healing time by inhibiting biofilm formation, which make it promising candidates for treatment of MRSA-induced wound infections.

Keywords: antibacterial effect; biofilm; methicillin-resistant Staphylococcus aureus; nanofibers; wound enclosure.

Figure 1

Research schematic diagram of the novel bioactive antimicrobial nanofibers with benzalkonium bromide.

Figure 2

Effects of six different drug loading on the encapsulation efficiency (A), swelling ratio (B) and tensile strength (C).

Figure 3

Morphological and physicochemical characteristics of the nanofibers. (A) Transmission electron microscopy morphological image (45,000× magnification), (B) transmission electron microscopy morphological image (65,000× magnification), (C) scanning electron microscopy morphological image (15,000× magnification), (D) scanning electron microscopy morphological image (30,000× magnification), (E) the diameter distribution, (F) zeta potential distribution.

Figure 4

Release profile, cytotoxicity, and antimicrobial activity of the novel nanofibers. (A) In vitro release profiles. (B) In vitro L929 cytotoxicity. (C) Minimum inhibitory concentration. (D) Time-kill analysis.

Figure 5

Surface structure of inhibited biofilm formation observed by SEM. (A) 1 μg/mL novel nanofibers, BBF, treatment group (2,000× magnification). (B) 1 μg/mL benzalkonium bromide treatment group (2,000× magnification). (C) Blank nanofibers, BNF, control (2,000× magnification). (D) 1 μg/mL novel nanofibers, BBF, treatment group (8,000× magnification). (E) 1 μg/mL BZK treatment group (8,000× magnification). (F) Blank nanofibers, BNF, control (8,000× magnification).

Figure 6

Histotoxicity images of the novel nanofibers (BBFs).

Figure 7

Antibacterial activity and wound closure against MRSA infection in vivo. (A) Bacterial burden on days 7 after wounding; (B) relative healing area ratio of wounds. (C) Histological microscopy images of untreated wounds and infections treated with BZK aqueous solution, BBF, vancomycin control, and blank nanofibers (BNF).