Bioheterojunctions Prevent Tooth Caries via Cascade Antibacterial Strategy

Abstract

Tooth caries is a prevalent chronic oral disease with microorganisms as the initiating factor and carbohydrates as the key environmental factor. Clinical antimicrobial therapies rely mainly on broad-spectrum antibiotics, which usually increase the risk of bacterial resistance. Recently, phototherapy has shown powerful antibacterial effects, although it cannot effectively eliminate cariogenic microenvironments and the antibacterial effect is not sustained after the light is removed. Here, we developed novel bioheterojunctions (bio-HJs) comprising MXene/Ag₃PO₄ (MX/AgP) and glucose oxidase (GOx), denoted MX/AgP-GOx, aiming at both the chemical and biological components of dental plaque biofilm. The bio-HJs decomposed the glucose rich in the cariogenic environment through GOx while providing abundant H₂O₂ for subsequent Fenton reaction. Under near-infrared (NIR) light, the bio-HJs produced hyperthermia and generated large amounts of reactive oxygen species based on the above H₂O₂, exerting powerful phototherapy properties (log reduction: 1.45 log 10 CFU/mL). It is worth noting that MX/AgP-GOx still exerted antibacterial effects in the dark via Ag⁺ bactericidal effects, Ag⁰ NPs catalytic activity, and GOx-mediated glucose depletion (log reduction: 0.39 log 10 CFU/mL), ensuring a sustained anticaries effect after the removal of NIR light. In addition, the rat caries model revealed that MX/AgP-GOx significantly reduced enamel mineral loss and had good biocompatibility. This study constructed efficacy-cascade bio-HJs targeting the sugar-rich cariogenic microenvironment, which promotes subsequent photodynamic therapy and combines photothermal and metal ion synergistic antibacterial means to continuously and effectively eliminate biofilm and prevent the occurrence and development of tooth caries.

Keywords

MXene glucose oxidase glucose biofilms phototherapy reactive oxygen species

Introduction

Tooth caries is a multifactorial, sugar-dependent disease driven by microorganisms, representing a global public health challenge (Peres et al. 2019; Wen et al. 2022). Specifically, Streptococcus mutans metabolizes dietary sucrose into glucose and forms dextran polymer, known as exopolysaccharides (EPSs), which protect and nourish microorganisms while contributing to antimicrobial resistance (Bertolini et al. 2022). Therefore, targeting the microbial etiology in sugar-rich cariogenic microenvironments is the key to preventing caries initiation and progression. However, conventional broad-spectrum antibiotics in clinical practice show limited efficacy due to poor penetration of low-permeability plaque biofilms and disrupt the natural diversity of the oral microbiome (Chatzigiannidou et al. 2020).

Photothermal therapy (PTT) has emerged as a promising alternative to antibiotics by destroying pathogens’ integrity through photothermal conversion processes (Qi et al. 2023). However, the temperatures required to eliminate biofilm would exceed the thermal tolerance of healthy cells (Huo et al. 2021). To enhance antibacterial efficiency with less light energy, researchers have explored synergistic strategies that combine PTT with other technologies, such as photodynamic therapy (PDT) (Hu et al. 2022). Yet, the PTT/PDT strongly depends on the external energy supply, whose effect disappears with the withdrawal of light, allowing biofilm formation to occur. Continuous biofilm inhibition after light withdrawal remains a challenge. In addition, blocking EPS formation is also important given its central role in plaque formation and bacterial defense, which is beyond the scope of phototherapy. Accordingly, a comprehensive strategy incorporating chemical and biological perspectives is essential for effective biofilm elimination.

Biologically compatible heterojunctions (bio-HJs) comprise 2 functional semiconductors with different bandgaps and energy levels, which is a superior platform for biological systems by integrating PTT, PDT, chemodynamic therapy (CDT), and metal ion therapy (MIT) through structural and compositional modulation (Low et al. 2017; Geng et al. 2024). Here, we proposed cascade anti-biofilm bio-HJs constructed of functionalizing MXene-Ag₃PO₄ HJs with glucose oxidase (GOx) through polydopamine (pDA) linkage, designated as MXene/Ag₃PO₄-GOx (MX/AgP-GOx). MXene is a category of 2-dimensional layered materials with broad specific surfaces, atomically thin thickness, tunable bandgaps, and facile surface functionalization (Naguib et al. 2014; Hao et al. 2022). Ag₃PO₄ serves as a photocatalyst with a narrow bandgap and high visible light responsiveness, releasing Ag⁺ to destroy bacterial membrane proteins and forming catalytically active Ag⁰ nanoparticles (NPs) reduced by pDA (Ma et al. 2022; Yang et al. 2022). GOx catalyzes glucose in cariogenic biofilms to generate hydrogen peroxide (H₂O₂) (Fu et al. 2018), providing substrates for chemodynamic and photodynamic processes. Under near-infrared (NIR) radiation, the bio-HJs demonstrate photothermal efficiency and generate reactive oxygen species (ROS) via Fenton-like reactions, chemical reactions that generate ROS through catalytic H₂O₂ decomposition (Tang et al. 2021). Furthermore, bio-HJs are sustainedly anticaries in darkness through the Ag⁺ bactericidal effects, Ag⁰ NPs catalytic activity, and GOx-mediated glucose depletion, overcoming the light-dependency limitation of conventional phototherapy. The MX/AgP-GOx bio-HJs effectively eliminate sugar-rich cariogenic environments while enhancing ROS generation, achieving cascading anticaries effects in dark and NIR environments.

Materials and Methods

Synthesis of Bio-HJ

MXene was synthesized through liquid-phase exfoliation of Ti₃AlC₂ using a 1:1 mixture of LiF and HCl at 45 °C for 12 h (Wang et al. 2022). The product was washed, centrifuged at 4,000 rpm, and lyophilized. MX/AgP was constructed via electrostatic self-assembly by adding silver nitrate (AgNO₃, 0.1 M) to dispersed MXene, dialyzed overnight, and tested with sodium phosphate. Aqueous sodium phosphate solution (Na₂HPO₄, 0.05 M) was then applied to the dialysate and vigorously agitated. The MX/AgP composite was modified with pDA (2 mg mL⁻¹) and GOx (2 mg mL⁻¹) through sequential addition and incubation to obtain MX/AgP-GOx.

Sample morphology and microstructure were characterized using scanning electron microscopy (SEM) equipped with energy dispersive spectrometer (EDS) mapping (JSM-7500-F, JEOL), transmission electron microscopy (TEM; Tecnai F20), and X-ray photoelectron spectroscopy (XPS; ESCALAB 250 instrument, Thermo).

Photothermal and Chemophotodynamic Performance

The photothermal performance of MX/AgP-GOx was analyzed under different powers of 808 nm NIR (0.5, 1.0, and 1.5 W cm⁻²). The temperature was monitored using an infrared radiant thermal camera (E6, FLIR) at 30-s intervals. The photothermal stability of MX/AgP-GOx was assessed under 1.5 W cm⁻² NIR illumination through 3 heating–cooling cycles (10-min heating, 15-min natural cooling) (He et al. 2022). Photodynamic performance was evaluated by detecting hydroxyl radicals (·OH), superoxide radicals (·O₂⁻), and singlet oxygen (¹O₂) using methylene blue (MB) spectrophotometry (Huang et al. 2023) and 1,3-diphenylisobenzofuran (DPBF) assays (Shu et al. 2023). Glutathione (GSH) depletion was assessed through the Ellman assay (Deng et al. 2022).

Antibacterial Activity

S. mutans (UA159) was cultured anaerobically in brain heart infusion broth (Oxoid). Bacterial suspensions of S. mutans were treated with different samples under NIR irradiation or in darkness. Antibacterial activity was evaluated using the plate spread method, and bacterial morphology was observed by SEM. In addition, antibiofilm performance was assessed using crystal violet (CV) staining.

The log reduction was calculated by

Log reduction = {lgA}_{C} - {lgA}_{E}

where A_E is the average CFU of the experimental groups and A_C is the average CFU of the control group.

Cytocompatibility

Human oral keratinocytes (HOK) cells were cultured in high-glucose DMEM with samples under standard conditions. Cell morphology was observed using SEM and confocal laser scanning microscope (CLSM, Nikon) after fluorescent staining. Cell proliferation was evaluated using a cell counting kit (CCK-8) assay.

Anticaries Evaluation In Vivo

Twelve male Sprague–Dawley rats (4 wk old) were used to establish a caries model following ARRIVE guidelines and permitted by the Animal Care and Experiment Committee of West China Stomatology Hospital of Sichuan University. After streptomycin pretreatment for 4 d, oral swabs were taken from the rats’ mouths for the plate spread method to confirm no S. mutans infection (Tao et al. 2022). Then, the rats’ mouths were infected with S. mutans suspension (10⁸ CFU/mL) for 3 d and fed with a cariogenic diet (Diet 2000, Trophic Animal Feed) and sucrose aqueous solution (10% w/v). Sterile oral swabs and plate spread method were used again to confirm the successful infection of S. mutans. The rats were randomly divided into 4 groups (n = 3): phosphate-buffered saline (PBS) (NIR−), MX/AgP-GOx (NIR−), PBS (NIR+), and MX/AgP-GOx (NIR+). Treatments were applied twice daily with or without NIR irradiation (808 nm, 1.5 W cm⁻²) for 4 wk.

Caries progression was evaluated using micro–computed tomography (micro-CT; Quantum GX system, PerkinElmer) and Keyes scoring after ammonium purpurate staining (Wu et al. 2018). Biocompatibility was assessed through hematoxylin and eosin (H&E) staining of oral mucosa and major organs.

Statistical Analysis

The results are presented as the mean ± standard deviation from at least 3 independent experiments. Statistical significance was determined using one-way analysis of variance or t test (SPSS 24.0), with P < 0.05 considered significant.

Further details are included in the Appendix.

Results

Characterization of MX/AgP-GOx Hjs

SEM and TEM were employed to observe the micromorphology and structure of the samples. As the zoomed-out image SEM in the upper right corner of Figure 1B shows, MXene presented a typical accordion-like multilayer nanostructure (Venkateshalu and Grace 2020). After stratification, the SEM (Fig. 1B) and TEM (Fig. 1C) images of MXene exhibited single-layer nanostructures with larger relative surface areas. In Figure 1D, MX/AgP were evenly coated with particles (ø = 10~200 nm). Corresponding EDS results (Fig. 1F) reflect that Ag, C, O, and Ti elements were uniformly distributed in MX/AgP.

Figure 1.

Bioheterojunctions (bio-HJs) assessment characterization. (A) Preparation diagram of MX/AgP-GOx. (B) Scanning electron microscopy (SEM) of multilayer MXene and MXene nanosheets. (C) Transmission electron microscopy (TEM) of MXene nanosheets. (D, E) SEM images of MX/AgP and MX/AgP-GOx. (F) Energy dispersive spectrometry (EDS) of MX/AgP. (G) X-ray photoelectron spectroscopy (XPS) survey spectra and (H) fine spectra of C 1s of MX/AgP and MX/AgP-GOx. Fine spectra of (I) N 1s and (J) Ag 3d of MX/AgP-GOx.

The surface of the MX/AgP-GOx (Fig. 1E) shows an incorporation of spherical particles. The chemical valence and elemental composition of MX/AgP and MX/AgP-GOx were detected by XPS spectra (Fig. 1G). The MX/AgP-GOx shows a distinct O-C = O bond (Fig. 1H) and N 1s peak (Fig. 1I), confirming the incorporation of GOx (C₆H₁₂O₆) and pDA (C₈H₁₁NO₂) (Guiomar et al. 1999). In addition, it exhibits peaks of Ag 3d_5/2 and Ag 3d_3/2 (Fig. 1J), which further split into signals for Ag⁺ in Ag₃PO₄ and Ag⁰ NPs reduced by pDA (Duan et al. 2023).

Photothermal Performance of MX/AgP-GOx Hjs

The photothermal effect was studied by monitoring temperature changes under NIR light. As shown in Figure 2A, the temperature of MX/AgP-GOx increased to 36.7, 45.4, and 54.2 °C after 10-min 808 nm NIR illumination at powers of 0.5, 1.0, and 1.5 W cm⁻². The heating images and curves (Fig. 2B and C) show the temperature of MX/AgP-GOx significantly increased to 54.2 °C, higher than those of Ag₃PO₄ and MX/AgP. The photothermal conversion efficiency of MX/AgP-GOx attained 20.5% under 1.5 W cm⁻² NIR irradiation (Fig. 2D), and it had no significant damping in the heating curves during 3 heating–natural cooling cycles (Fig. 2E).

Figure 2.

Photothermal and chemophotodynamic performance. (A) Photothermal curves of MX/AgP-GOx under different powers. (B) Instantaneous infrared thermal photos and (C) photothermal curves of Ag₃PO₄, MX/AgP, and MX/AgP-GOx. (D) Photothermal conversion efficiency of MX/AgP-GOx. (E) Photothermal curves of MX/AgP-GOx with 3 heating/cooling cycles. (F) The principle and related MB consumption spectra of (G) MXene and (H) MX/AgP-GOx. (I) The principle and related 1,3-diphenylisobenzofuran (DPBF) consumption spectra of (J) MXene and (K) MX/AgP-GOx. (L) The mechanism of Ellman’s method and (M) consumption of Glutathione (GSH) (n = 3). ****P < 0.0001.

Chemophotodynamic Performance of MX/AgP-GOx Hjs

The production of OH was detected by MB, which caused a peak decline at about 664 nm, whose principle is illustrated in Figure 2F. After a 15-min NIR irradiation, the MB absorbance of MXene decreased very slightly (Fig. 2G). However, the MB absorption intensity of MX/AgP-GOx decreased obviously after 5-min NIR irradiation and continued to decline with the increase in irradiation time (Fig. 2H). The ·O₂⁻ and ¹O₂ was captured by DPBF, which caused a peak decline at about 410 nm, whose principle is illustrated in Figure 2I. Under NIR irradiation, the DPBF absorbance of MXene decreased slightly (Fig. 2J), while that of MX/AgP-GOx decreased more distinctly (Fig. 2K).

The GSH consumption capacity (Fig. 2L) was determined by the sulfhydryl reaction of 5,5′-dithiolide (2-nitrobenzoic acid) (DTNB) with glutathione to form yellow 5-thiole-2-nitrobenzoic acid (TNB) (Rahman et al. 2006). Compared with the blank group, the absorbance of MX/AgP (NIR−) and MX/AgP (NIR+) slightly reduced. Notably, MX/AgP-GOx consumed a significant amount of GSH in the dark, and the GSH consumption under NIR radiation was close to the H₂O₂ group (Fig. 2M).

Antibacterial Properties In Vitro

The spread plate method (Fig. 3A and B) shows that in the absence of NIR light, the log reduction of the MX/AgP and MX/AgP-GOx groups was 0.43 and 0.39 log 10 CFU/mL, respectively. Under NIR irradiation, the log reduction of MX/AgP-GOx reached 1.45 log 10 CFU/mL. Under 2.0 W cm⁻² and 3.0 W cm⁻² NIR irradiation, the log reduction of MX/AgP-GOx further reached 2.53 and 3.02 log 10 CFU/mL, respectively, in just 5 min (Appendix Fig. 1A and B).

Figure 3.

Antibacterial properties in vitro. (A) Bacterial colonies and (B) counts (CFU/mL, log 10) of S. mutans treated with different materials (n = 3). (C) Scanning electron microscopy morphologies of S. mutans (the yellow arrows indicate shrinking membranes). (D) Quantitative analysis and (E) typical pictures of crystal violet (CV) staining biofilms (n = 3). (F) Schematic diagram of the antibiofilm mechanism of MX/AgP-GOx bioheterojunctions (bio-HJs). *P < 0.05, ***P < 0.001, ****P < 0.0001.

Furthermore, the morphology and the cell membrane integrity of S. mutans were observed by SEM. As Figure 3C shows, S. mutans in the PBS group appeared to have a typical oval shape with a smooth surface. In contrast, bacterial cytoplasm contraction was observed in the MX/AgP (NIR−) group, while the contraction was more pronounced in the MX/AgP (NIR+) and MX/AgP-GOx (NIR−) groups. Notably, S. mutans in the MX/AgP-GOx (NIR+) group exhibited more severe morphological shrinkage and cell membrane rupture.

The influence of different materials on S. mutans biofilm was investigated by CV staining (Fig. 3E). Mature S. mutans biofilm in PBS groups was stained purple. In the darkness, the MX/AgP-GOx group (≈75%) exhibited much higher biofilm clearance than the MX/AgP group (≈20%) (Fig. 3D). Under NIR irradiation, the residual amount of biofilm in the MX/AgP-GOx group was less, and the biofilm clearance rate reached greater than 80%.

Cytocompatibility In Vitro

The HOK cells in the MX/AgP and MX/AgP-GOx groups presented a round and smooth morphology similar to that in the control group (Fig. 4A). All the HOK cells exhibited clear, complete nuclei and an extended cytoskeleton (Fig. 4B). The CCK-8 results (Fig. 4C) show that the cell number in all groups was close to that of the control group after 1 d of culture. The cell proliferation rate in the MX/AgP-GOx groups under dark and NIR light was 88% and 79% of the control group.

Figure 4.

Cytocompatibility evaluation. (A) Scanning electron microscopy (SEM) images of cell morphologies cultured in different materials. (B) Confocal laser scanning microscope (CLSM) images of cell structure (green, cytoskeleton, stained with FITC phalloidin; blue nuclei, stained with DAPI). (C) The optical density (OD) value at 450 nm of human oral keratinocyte (HOK) proliferation measured by cell counting kit–8 (CCK-8) without or with near-infrared (NIR) light (n = 3). *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001.

Anticaries Property In Vivo

Given the excellent antibacterial activity and good biocompatibility of MX/AgP-GOx in vitro, we established a rat caries model to verify its anticaries performance in vivo (Fig. 5A).

Figure 5.

Anticaries performance in vivo. (A) Schematic diagram of rat caries model establishment and following treatments. (B) Three-dimensional (3D) reconstructed images and (C) micro–computed tomography (CT) images of rat molars (the purple arrows indicate the demineralization area). Quantitative results of the (D) surface area and (E) volume of enamel (n = 3). (F) Caries staining images of rat molars (the blue arrows indicate caries lesions) and (G) modified Keyes scores (n = 3). (H) Hematoxylin and eosin (H&E) staining images of the dental pulp. *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001.

From the top view of micro-CT (Fig. 5B), the PBS (NIR−) group involves a large area of enamel loss in the tooth cusps and fossa, while the MX/AgP-GOx (NIR+) group involves only a small amount of enamel wear in the cusps. The tomography images (Fig. 5C) show a density reduction shadow at the bottom of the fossa and groove of the teeth in the PBS (NIR−) group, which reflects tooth demineralization, whereas there was no significant density reduction shadow in the MX/AgP-GOx (NIR+) group. The enamel surface area (Fig. 5D) and volume (Fig. 5E) of the MX/AgP-GOx (NIR+) group and MX/AgP-GOx (NIR−) group were higher than that of the PBS (NIR−) group.

After the noninvasive CT scan, we adopted the modified Keyes scoring, one of the most commonly used methods to assess the biological course of caries, which evaluates the lesions from 3 planes of depth and width and provides more morphological details. The images of stained teeth sections (Fig. 5F) show the PBS (NIR−) group had extensive caries (stained red) that invaded the dentin, while barely red-stained lesions were observed in the MX/AgP-GOx (NIR+) group. The score (Fig. 5G) of the MX/AgP-GOx (NIR−) group decreased significantly compared with the PBS (NIR−) group. The score of the MX/AgP-GOx (NIR+) group (0.3 ± 0.6) further decreased with only slight plane.

Histopathological Analysis

Hematoxylin and eosin (H&E) staining was performed on sections of oral tissues and vital organs to assess biosafety in vivo. The H&E staining images of the dental pulp (Fig. 5H) and oral mucosa (Appendix Fig. 2) of all groups showed no obvious inflammatory infiltration. In addition, there was no obvious damage or inflammation in the H&E staining images of the vital organ sections (Appendix Fig. 3).

Discussion

Dental caries is a biofilm-mediated disease in which S. mutans exploits sugar-rich microenvironments, metabolizing carbohydrates into glucose. This process leads to EPSs formation, nourishing microorganisms, and inducing antibiotic resistance (Du et al. 2020; Bertolini et al. 2022; Ciofu et al. 2022). While phototherapy is a promising alternative antibiotic strategy due to its deep penetration and high bactericidal efficiency, its reliance on NIR allows for potential bacterial growth following the withdrawal of light. To address this limitation, we developed MX/AgP-GOx bio-HJs that target the cariogenic microenvironment to consume glucose while generating H₂O₂ to enhance subsequent CDT and phototherapeutic efficacy. After removing NIR, the system maintains antibacterial activity through the MIT of Ag⁺, the chemodynamic property of Ag⁰ NPs, and the high-sugar environment elimination of GOx, continuously blocking the caries process.

The construction of MX/AgP-GOx (Fig. 1A) starts with extracting single-layer MXene (Ti₃C₂Tx), which provides abundant nucleation sites (Fig. 1C) for Ag₃PO₄. Via an electrostatically driven self-assembly strategy, Ag₃PO₄ was uniformly bound to MXene (Fig. 1F) and formed HJs due to different bandgaps (Yang et al. 2022). Eventually, by the excellent adhesion property of pDA, GOx was attached to the MX/AgP HJs surface and obtained MX/AgP-GOx bio-HJs. With redox-active catechol groups, pDA can reduce some Ag⁺ to Ag⁰ NPs, which has catalytic activity due to the large specific surface area and quantum effects (Yang et al. 2023).

The photothermal and photodynamic properties are critical determinants of bactericidal efficiency. The photothermal curves (Fig. 2A) show that the photothermal performance of MX/AgP-GOx increases proportionally with NIR power intensity. Under 1.5 W cm⁻² irradiation, the final temperature exceeded 50 °C, surpassing the bacterial thermal ablation threshold (Huo et al. 2021), so it was chosen for subsequent experiments. MX/AgP-GOx exhibited superior heating efficiency compared with AgP and MX/AgP under NIR irradiation (Fig. 2B and C), indicating its enhanced capacity to convert NIR light energy into thermal energy for PTT. Besides, it possesses good photothermal stability (Fig. 2E).

The photodynamic efficiency correlates with ROS production. MB and DPBF assays revealed that MX/AgP-GOx HJs function as photosensitizers to produce various ROS under NIR irradiation, including ·OH, ·O₂⁻, and ¹O₂. The outstanding ROS generation efficiency of MX/AgP-GOx is due to the establishment of HJs between MXene and Ag₃PO₄, and the energy generated by hot electrons during the surface plasmon resonance decay of MXene promotes the isolation of electron-hole pairs (Chan et al. 2021). Under NIR light excitation, the holes will remain and extend to the surface of MXene, reacting with the surrounding O₂ to form ¹O₂, while the hot electrons will be reverse transferred to Ag₃PO₄, reacting with H₂O to form ·OH.

GSH is a nonenzymatic antioxidant that can oxidize organic thiols (R-SH) to disulfides (R-S-S-R) and mediate ROS neutralization (Zhou et al. 2022). Extracellular GSH uptake is essential for the antioxidant stress responses and protein function regulation in S. mutans (Cheng et al. 2024). MX/AgP-GOx consumed large amounts of GSH in both darkness and NIR irradiation, reflecting its sustaining chemodynamic and photodynamic ROS generation functionality. The chemodynamic process is attributed to the ROS generation from H₂O₂ catalyzed by the reduced Ag⁰ NPs, which makes up for the dependence of the photodynamic ROS generation on NIR.

The above photothermal, photodynamic, and chemodynamic efficiency of MX/AgP-GOx indicates its sustained antimicrobial potential under NIR and dark conditions. Considering S. mutans is recognized as the predominant cariogenic bacteria in caries initiation and progression (Zhang et al. 2021), we chose it for antibacterial evaluation. In darkness, MX/AgP and MX/AgP-GOx exhibited a certain antibacterial ability, attributed to the direct bactericidal action of Ag⁺ and ROS produced by the chemodynamic process. Under NIR irradiation, the antibacterial ability of MX/AgP-GOx was significantly enhanced, demonstrating its potent photothermal and photodynamic bactericidal effect. Bacterial morphological analysis revealed that when MX/AgP-GOx was exposed to NIR light, it destroyed the bacterial cell membrane integrity under the synergistic effects of Ag⁺, heating, and ROS, thereby significantly inhibiting bacterial growth and proliferation.

Furthermore, dental plaque biofilm is the initiating factor of caries (Jakubovics et al. 2021), making biofilm inhibition efficacy critical in evaluating potential anticaries materials. The biofilm biomass of S. mutans in the MX/AgP-GOx (NIR−) group was significantly reduced compared with the MX/AgP (NIR−) group, demonstrating GOx’s capacity for EPSs degradation. Under NIR irradiation, MX/AgP-GOx suppressed biofilm formation more obviously, owing to the EPSs degradation producing H₂O₂, which enhanced ROS generation. Eventually, MX/AgP-GOx effectively inhibited biofilm formation through synergistic therapy of MIT, PTT, CDT, and PDT, as depicted in the proposed antibiofilm mechanism (Fig. 3F).

This study successfully developed MX/AgP-GOx bio-HJs that catalyze glucose and produce H₂O₂ in cariogenic conditions, triggering cascade antibacterial actions. Under NIR irradiation, MX/AgP-GOx bio-HJs effectively clear biofilms through the combination of MIT, PTT, and enhanced CDT/PDT. In addition, due to the Ag⁺ bactericidal effects, Ag⁰ NPs’ catalytic activity, and GOx-mediated glucose depletion, MX/AgP-GOx continues to work in the dark, making up for the dependence on external irradiation. In the rat caries model, MX/AgP-GOx bio-HJs reduced mineral loss of the teeth in the dark and further reduced mineral loss under NIR light. In addition, MX/AgP-GOx showed good biocompatibility both in vitro and in vivo. The MX/AgP-GOx bio-HJs provide a multidirectional template for blocking biofilm initiation diseases not only limited to tooth caries.

The current study has some limitations. Although MX/AgP-GOx showed effective antibacterial activity at 1.5 W cm⁻² NIR irradiation for 10 min, its antibacterial efficiency can be further improved by enhancing NIR intensity and shortening the duration of irradiation in the future. While our study focused on S. mutans to evaluate the antibacterial efficacy of MX/AgP-GOx bio-HJs and the ability to target primary cariogenic factors, we recognize that the oral cavity hosts a diverse microbial community. This includes beneficial symbiotic bacteria that promote oral health by competing with cariogenic bacteria and maintaining ecological balance. To minimize the risk of microbial dysbiosis, we designed the MX/AgP-GOx bio-HJs to amplify their antibacterial effect specifically in cariogenic high-sugar environments. However, more assessments of the impact of MX/AgP-GOx bio-HJs on oral microbial diversity and community composition are needed in further research.

Author Contributions

J. Zhu, contributed to conception, design, data acquisition and interpretation, drafted and critically revised the manuscript; L. He, contributed to conception, design, data analysis and interpretation, critically revised the manuscript; X. Xu, H. Wu, and J. Li, contributed to data analysis and interpretation, critically revised the manuscript; B. Yan, Y. Deng, contributed to data acquisition, critically revised the manuscript; Y. Gao, K. Liang, contributed to conception and design, critically revised the manuscript. All authors gave final approval and agree to be accountable for all aspects of the work.

Supplemental Material

sj-docx-1-jdr-10.1177_00220345251329334 – Supplemental material for Bioheterojunctions Prevent Tooth Caries via Cascade Antibacterial Strategy

Supplemental material, sj-docx-1-jdr-10.1177_00220345251329334 for Bioheterojunctions Prevent Tooth Caries via Cascade Antibacterial Strategy by J. Zhu, L. He, X. Xu, H. Wu, J. Li, B. Yan, Y. Deng, Y. Gao and K. Liang in Journal of Dental Research

Footnotes

A supplemental appendix to this article is available online.

Declaration of Conflicting Interests

The authors declared no potential conflicts of interest with respect to the research, authorship, and/or publication of this article.

Funding

The authors disclosed receipt of the following financial support for the research, authorship, and/or publication of this article: This work was supported by the National Natural Science Foundation of China (82270970, 82470967), Natural Science Foundation of Sichuan Province (2023NSFSC1504), and Sichuan Science and Technology Program (2023YFQ0014).

ORCID iDs

J. Li

K. Liang

References

Bertolini

Costa

Barao

VAR

Cunha Villar

Retamal-Valdes

Feres

Silva Souza

. 2022. Oral microorganisms and biofilms: new insights to defeat the main etiologic factor of oral diseases. Microorganisms. 10(12):2413. doi: 10.3390/microorganisms10122413

Chan

Chen

Gao

Xie

Zhang

Zhao

Chen

2021. Coordination-driven enhancement of radiosensitization by black phosphorus via regulating tumor metabolism. ACS Nano. 15(2):3047–3060.

Chatzigiannidou

Teughels

Van de Wiele

Boon

2020. Oral biofilms exposure to chlorhexidine results in altered microbial composition and metabolic profile. NPJ Biofilms Microbiomes. 6(1):13. doi: 10.1038/s41522-020-0124-3

Cheng

Zhou

Ning

. 2024. Oxidative stress response: A critical factor affecting the ecological competitiveness of streptococcus mutans. J Oral Microbiol. 16(1):2292539. https://doi.org/10.1080/20002297.2023.2292539

Ciofu

Moser

Jensen

Hoiby

2022. Tolerance and resistance of microbial biofilms. Nat Rev Microbiol. 20(10):621–635.

Deng

Ouyang

Sun

Shi

Chan

Yang

Peng

2022. Rapid sterilisation and diabetic cutaneous regeneration using cascade bio-heterojunctions through glucose oxidase-primed therapy. Bioact Mater. 25:748–765.

Zhou

Cao

Guo

Zhou

Peng

Zheng

, et al. 2020. Sucrose promotes caries progression by disrupting the microecological balance in oral biofilms: an in vitro study. Sci Rep. 10(1):2961. doi: 10.1038/s41598-020-59733-6

Duan

Fang

Guo

Zou

Xiang

Wang

2023. Efficient and stable monolithic microreactor with Ag/AgCl photocatalysts coated on polydopamine modified melamine sponge for photocatalytic water purification. Colloids Surf A. 659:130759. doi: 10.1016/j.colsurfa.2022.130759

Lin

Huang

2018. Catalytic chemistry of glucose oxidase in cancer diagnosis and treatment. Chem Soc Rev. 47(17):6454–6472.

10.

Geng

Johnson

Shi

Chen

Chan

Qin

, et al. 2024. Achieving clearance of drug-resistant bacterial infection and rapid cutaneous wound regeneration using an ROS-balancing-engineered heterojunction. Adv Mater. 36(16):e2310599. doi: 10.1002/adma.202310599

11.

Guiomar

Guthrie

Evans

. 1999. Use of mixed self-assembled monolayers in a study of the effect of the microenvironment on immobilized glucose oxidase. Langmuir. 15(4):1198–1207.

12.

Hao

Han

Yang

Chen

Jiang

Dong

Wen

Liu

, et al. 2022. Recent advancements on photothermal conversion and antibacterial applications over MXenes-based materials. Nanomicro Lett. 14(1):178. doi: 10.1007/s40820-022-00901-w

13.

Duan

Wang

Kei Chan

Shi

Huang

Wang

Peng

Deng

2022. Fusion peptide-engineered polyetheretherketone implants with photo-assisted anti-pathogen and enhanced angiogenesis for in vivo osseointegrative fixation. Chem Eng J. 446:137453. doi: 10.1016/j.cej.2022.137453

14.

Zhang

Wang

Shiu

B-C

Lin

J-H

Zhang

Lou

C-W

T-T.

2022. Synergistic antibacterial strategy based on photodynamic therapy: progress and perspectives. Chem Eng J. 450:138129. doi: 10.1016/j.cej.2022.138129

15.

Huang

Wang

Johnson

Yang

Deng

Liang

2023. Engineered bio-heterojunction with infection-primed H₂S liberation for boosted angiogenesis and infectious cutaneous regeneration. Small. 19(45):e2304324. doi: 10.1002/smll.202304324

16.

Huo

Jia

Huang

Zhang

Dong

Huang

2021. Emerging photothermal-derived multimodal synergistic therapy in combating bacterial infections. Chem Soc Rev. 50(15):8762–8789.

17.

Jakubovics

Goodman

Mashburn-Warren

Stafford

Cieplik

2021. The dental plaque biofilm matrix. Periodontol 2000. 86(1):32–56.

18.

Low

Jaroniec

Wageh

Al-Ghamdi

. 2017. Heterojunction photocatalysts. Adv Mater. 29(20):1601694. doi: 10.1002/adma.201601694

19.

Yang

Mei

Zhu

Liu

Yao

Yang

2022. Z-scheme Fe₂(MoO₄)₃/Ag/Ag₃PO₄ heterojunction with enhanced degradation rate by in-situ generated H₂O₂: turning waste (H₂O₂) into wealth (•OH). J Colloid Interface Sci. 606(pt 2):1800–1810.

20.

Naguib

Mochalin

Barsoum

Gogotsi

2014. 25th anniversary article: MXenes: a new family of two-dimensional materials. Adv Mater. 26(7):992–1005.

21.

Peres

Macpherson

LMD

Weyant

Daly

Venturelli

Mathur

Listl

Celeste

Guarnizo-Herreno

Kearns

, et al. 2019. Oral diseases: a global public health challenge. Lancet. 394(10194):249–260.

22.

Xiang

Cai

Chen

Zhang

Shen

2023. Inorganic-organic hybrid nanomaterials for photothermal antibacterial therapy. Coord Chem Rev. 496:215426. doi: 10.1016/j.ccr.2023.215426

23.

Rahman

Kode

Biswas

. 2006. Assay for quantitative determination of glutathione and glutathione disulfide levels using enzymatic recycling method. Nat Protoc. 1(6):3159–3165.

24.

Shu

Sun

Gao

Chan

Shi

Bai

Yang

Deng

2023. Self-tandem bio-heterojunctions empower orthopedic implants with amplified chemo-photodynamic anti-pathogenic therapy and boosted diabetic osseointegration. Adv Funct Mater. 33(27):2214873. doi: 10.1002/adfm.202214873

25.

Tang

Zhao

Wang

Liu

2021. Biomedicine meets Fenton chemistry. Chem Rev. 121(4):1981–2019.

26.

Tao

Yang

Zhou

Wang

Yang

Sun

Deng

Liang

2022. A dentin biomimetic remineralization material with an ability to stabilize collagen. Small. 18(38):e2203644. doi: 10.1002/smll.202203644

27.

Venkateshalu

Grace

. 2020. MXenes—a new class of 2D layered materials: synthesis, properties, applications as supercapacitor electrode and beyond. Appl Mater Today. 18:100509. doi: 10.1016/j.apmt.2019.100509

28.

Wang

Gao

Huang

Shi

Deng

Liang

2022. Endogenous oxygen-evolving bio-catalytic fabrics with fortified photonic disinfection for invasive bacteria-caused refractory cutaneous regeneration. Nano Today. 46:101595. doi: 10.1016/j.nantod.2022.101595

29.

Wen

Chen

Zhong

Dong

Wong

2022. Global burden and inequality of dental caries, 1990 to 2019. J Dent Res. 101(4):392–399.

30.

Zhou

Zhang

Huang

HHK

Guo

Feng

, et al. 2018. Evaluation of novel anticaries adhesive in a secondary caries animal model. Caries Res. 52(1–2):14–21.

31.

Yang

Mao

Y-Y

Liu

W-W.

2023. Metal nanozymes with multiple catalytic activities: regulating strategies and biological applications. Rare Met. 42(9):2928–2948.

32.

Yang

Zhou

Chan

Wang

Liang

Deng

2022. Photo-activated nanofibrous membrane with self-rechargeable antibacterial function for stubborn infected cutaneous regeneration. Small. 18(12):e2105988. doi: 10.1002/smll.202105988

33.

Zhang

Wang

Zou

2021. Molecular mechanisms of inhibiting glucosyltransferases for biofilm formation in Streptococcus mutans. Int J Oral Sci. 13(1):30. doi: 10.1038/s41368-021-00137-1

34.

Zhou

Wang

Chan

Yang

Jiao

Liang

Deng

. 2022. Infection micromilieu-activated nanocatalytic membrane for orchestrating rapid sterilization and stalled chronic wound regeneration. Adv Funct Mater. 32(7):2109469. https://doi.org/10.1002/adfm.202109469

Supplementary Material

Please find the following supplemental material available below.

For Open Access articles published under a Creative Commons License, all supplemental material carries the same license as the article it is associated with.

For non-Open Access articles published, all supplemental material carries a non-exclusive license, and permission requests for re-use of supplemental material or any part of supplemental material shall be sent directly to the copyright owner as specified in the copyright notice associated with the article.

0.00 MB

2.55 MB