Synthesis of a Two-Dimensional Molybdenum Disulfide Nanosheet and Ultrasensitive Trapping of Staphylococcus Aureus for Enhanced Photothermal and Antibacterial Wound-Healing Therapy

Abstract

Photothermal therapy has been widely used in the treatment of bacterial infections. However, the short photothermal effective radius of conventional nano-photothermal agents makes it difficult to achieve effective photothermal antibacterial activity. Therefore, improving composite targeting can significantly inhibit bacterial growth. We inhibited the growth of Staphylococcus aureus (S. aureus) by using an extremely low concentration of vancomycin (Van) and applied photothermal therapy with molybdenum disulfide (MoS₂). This simple method used chitosan (CS) to synthesize fluorescein 5(6)-isothiocyanate (FITC)-labeled and Van-loaded MoS₂-nanosheet hydrogels (MoS₂-Van-FITC@CS). After modifying the surface, an extremely low concentration of Van could inhibit bacterial growth by trapping bacteria synergistically with the photothermal effects of MoS₂, while FITC labeled bacteria and chitosan hydrogels promoted wound healing. The results showed that MoS₂-Van-FITC@CS nanosheets had a thickness of approximately 30 nm, indicating the successful synthesis of the nanosheets. The vitro antibacterial results showed that MoS₂-Van-FITC with near-infrared irradiation significantly inhibited S. aureus growth, reaching an inhibition rate of 94.5% at nanoparticle concentrations of up to 100 µg/mL. Furthermore, MoS₂-Van-FITC@CS could exert a healing effect on wounds in mice. Our results demonstrate that MoS₂-Van-FITC@CS is biocompatible and can be used as a wound-healing agent.

Keywords: MoS2 nanosheet; antibacterial activity; photothermal therapy; wound-healing.

Figure 1

Schematic illustration of the synergism between the two components of MoS₂ and Van in MoS₂-Van-FITC@CS, which was constructed and applied to wound healing in vivo with their highly efficiently antibacterial properties.

Figure 2

Synthesis and characterization of MoS₂-Van-FITC@CS. (A) Synthesis illustration of MoS₂-Van-FITC@CS. (B) SEM images of MoS₂ NPs. (C) Zeta potential of MoS₂, MoS₂-Van, MoS₂-Van-FITC, and MoS₂-Van-FITC@CS. (D) UV-vis absorption spectra of MoS₂, Van, MoS₂-Van, MoS₂-Van-FITC, and MoS₂-Van-FITC@CS. (E) FT-IR spectrometry of MoS₂, Van, MoS₂-Van, MoS₂-Van-FITC, and MoS₂-Van-FITC@CS. (F) Digital images of I (Milli-water), II (FITC), III (MoS₂), IV (MoS₂-Van), V (MoS₂-Van-FITC), and VI (MoS₂-Van-FITC@CS) under bright and UV light. (G) Fluorescence emission spectra of MoS₂-Van-FITC@CS.

Figure 3

In vitro photothermal efficiency. (A) The photothermal responses of MoS₂-Van-FITC@CS. Different concentrations (50, 100, 200 and 400 µg/mL) were exposed to NIR irradiation (808 nm; 2 W/cm²). PBS served as a control. (B) The photothermal responses of MoS₂-Van-FITC@CS (400 µg/mL), which was exposed to different power density of NIR irradiation (808 nm; 0.5, 1, 1.5 and 2 W/cm²). (C) The heating and cooling curves of MoS₂-Van-FITC@CS (400 μg/mL) and PBS (laser irradiation at 808 nm). (D) The linear regression between the cooling period and −ln(θ) of the driving force temperature. Results shown are mean ± SD, n = 3.

Figure 4

In vitro antibacterial activity. (A) The results of the CFU assay for the blank group. PBS as a blank group. (B) The results of the CFU assay for MoS₂ (100 μg/mL) for different times (0, 150 and 300 s) at different power densities (0.5, 1 and 1.5 W/cm²). (C) The results of the CFU assay for MoS₂-Van-FITC (100 μg/mL) for different times (0, 150 and 300 s) at different power densities (0.5, 1 and 1.5 W/cm²). (D) Quantitative statistical results of (A–C) (PD = Power Density). graphed using the Origin software. Results shown are mean ± SD, n = 3.

Figure 5

Fluorescence microscopy images of S. aureus cultures treated with MoS₂-Van-FITC at various concentrations (15, 30, 45 µg/mL) for 12 h. PBS served as a control for the blank group. The cells stained with DAPI for 30 min were fluorescent blue, and those stained with MoS₂-Van-FITC were fluorescent green. Scale bar = 15 μm. (DAPI, Ex = 358 nm and Em = 461 nm; FITC, Ex = 490 nm and Em = 525 nm).

Figure 6

Confocal fluorescence microscopy assay (A). S. aureus cultures after treatment with MoS₂-Van NPs + NIR (100 µg/mL). Van solution (1 µg/mL), MoS₂ NPs + NIR (100 µg/mL) and MoS₂-Van (100 µg/mL) served as control groups. PBS served as a blank group. The cells were stained with SYTO 9 (green fluorescence) and PI (red fluorescence) for 30 min. The cells underwent NIR irradiation at 808 nm (1.5 W/cm², 6 min). The results of the apoptotic assay by flow cytometry analysis were statistically analyzed by CytExpert software (version 2.4.0.28) (B). Scale bar = 15 μm. The data are expressed as mean ± SD (n = 3).

Figure 7

SEM images of S. aureus cells. The bacterial cultures were treated with MoS₂-Van-FITC NPs at various concentrations (25, 50 and 100 µg/mL). The bacterial cultures treated with Van (1 µg/mL) and MoS₂ NPs + NIR (100 µg/mL) served as control groups. PBS + NIR served as a blank group. The red squares indicate the enlarged regions. The blue area is the elemental analysis chart. The cells underwent NIR irradiation at 808 nm (1.5 W/cm², 6 min).

Figure 8

In vivo wound healing analysis. (A) The relative wound area curve shows the MoS₂-Van-FITC@CS hydrogel had a significant positive effect on wound healing. (B) The body weights of different groups after treatment. (C) The thermal imaging of mice after treatment. The cells underwent NIR irradiation at 808 nm (1.5 W/cm², 6 min). (D) The photographs of wounds were taken every 2 days after treatment. MoS₂-Van-FITC@CS hydrogel + NIR (100 µg/mL), MoS₂ NPs (100 µg/mL), MoS₂ NPs + NIR (100 µg/mL) and MoS₂-Van-FITC@CS hydrogel served as control groups. PBS + NIR served as a blank group. The cells underwent NIR irradiation at 808 nm (1.5 W/cm², 6 min). Scale bar = 1 cm. Results shown are mean ± SD, n = 5–7.

Figure 9

In vivo toxicity evaluation. The hematoxylin–eosin-stained images of major organs following different treatments of normal mice. Group 1 was the no treatment group, group 2 was the NIR irradiation (1.5 W/cm², 6 min) group, group 3 was the MoS₂-Van-FITC@CS group and group 4 was the MoS₂-Van-FITC@CS + NIR (1.5 W/cm², 6 min) group.