Development and In Vivo Characterization of Probiotic Lysate-Treated Chitosan Nanogel as a Novel Biocompatible Formulation for Wound Healing

Abstract

Wound healing is a physiological reaction to tissue injuries which plays a crucial role in replacing the destroyed tissues. Probiotics produce valuable compounds that possess antibacterial and anti-inflammatory activities, immunomodulatory effects, and angiogenesis traits leading to the promotion of wound healing. Chitosan nanostructures have versatile properties making them quickly produced into gels, scaffolds, nanoparticles, beads, and sponge structures that can be incorporated into wound healing processes. In the current study, three formulations from nanogel consisting of probiotic supernatant (Lactobacillus reuteri, Lactobacillus fermentum, and Bacillus subtilis sp. natto)-loaded chitosan nanogels were prepared from the culture of corresponding cultures. The chitosan nanogels were previously characterized by Zetasizer, FTIR, and TEM. The prepared formulations' effectiveness and dressing activity were assessed by evaluating wound closure and histological trials in Sprague-Dawley rats. The results indicated that all probiotic lysate formulations have advantages over the wound healing process. However, Bacillus subtilis natto has a better wound healing quality, which is well known in pathology examination. The favorable effects of probiotic lysate nanogels, including the reasonable wound closing rate, good wound appearance, and satisfactory histological observation via in vivo examination, suggest it as a promising nominee for wound healing purposes.

Figure 1

Characterization of chitosan nanogel: (a) FT-IR spectra, (b) appearance before formulation, (c) TEM image, and (d) mean size distribution.

Figure 2

Photographs of the wound healing process in different groups: Lactobacillus fermentum supernatant-loaded chitosan nanogel as group 1, Lactobacillus reuteri supernatant-loaded chitosan nanogel as group 2, Bacillus subtilis sp. natto supernatant-loaded chitosan nanogel as group 3, chitosan nanogel without probiotic lysate as group 4, and the control group which receives no treatment as group 5. The photographs were taken after day 2, day 6, and day 14.

Figure 3

Effects of different groups on wound healing: Lactobacillus fermentum supernatant-loaded chitosan nanogel as group 1, Lactobacillus reuteri supernatant-loaded chitosan nanogel as group 2, Bacillus subtilis sp. natto supernatant-loaded chitosan nanogel as group 3, chitosan nanogel without probiotic lysate as group 4, and the control group which receives no treatment as group 5. Data are demonstrated as mean ± SEM (n = 6). ∗ indicates a significant difference from the fructose control group (P < 0.05).

Figure 4

The histopathological examination in the different groups: Lactobacillus fermentum supernatant-loaded chitosan nanogel as group 1, Lactobacillus reuteri supernatant-loaded chitosan nanogel as group 2, Bacillus subtilis sp. natto supernatant-loaded chitosan nanogel as group 3, chitosan nanogel without probiotic lysate as group 4, and the control group which receives no treatment as group 5.