Department of Orthodontics, School of Dentistry, Shahed University, Tehran, Iran.
Dental Research Center, Dentistry Research Institute, Tehran University of Medical Sciences, Tehran, Iran.
Photodiagnosis Photodyn Ther. 2023 Dec;44:103866. doi: 10.1016/j.pdpdt.2023.103866. Epub 2023 Oct 27.
White spot lesions (WSLs) are a common side effect of fixed orthodontic treatment. Streptococcus mutans is the primary causative agent of WSLs and dental caries on the teeth during treatment. According to the unique features of reduced graphene oxide-nano curcumin (rGO-nCur), this study aimed to investigate the mechanical properties and antimicrobial potency of rGO-nCur coated orthodontic elastomeric ligatures as a novel coating composite following dual-modal photodynamic inactivation (PDI) and photothermal inactivation (PTI) against S. mutans biofilms.
After confirmation of rGO-nCur synthesis and coating elastomeric ligatures with different concentration levels of 1.25, 2.5, 5, 7.5, and 10 % of rGO-nCur, tensile strength, force decay, extension to tensile strength, and contact angle of the coated elastomeric ligatures were measured using universal testing machine and sessile drop method, respectively. To investigate the mechanism through which irradiated rGO-nCur can inhibit the formation of S. mutans biofilms, intracellular reactive oxygen species (ROS) generation, and increase in temperature of rGO-nCur solutions under the 450 and 980 nm laser irradiation, respectively, were measured. The anti-biofilm activity and inhibition of water-insoluble extracellular polysaccharide (EPS) production ability of irradiated rGO-nCur coated elastomeric ligatures using a 450 nm diode laser (195 J/cm), a 980 nm diode laser (195 J/cm), and a combination of both (78 J/cm of irradiation from each one) (i.e., PDI, PTI, and dual-modal PDI/PTI, respectively) were determined. Also, the expression of virulence genes involved in biofilm formation (comDE, gtfD, and smuT) was assessed by quantitative real-time polymerase chain reaction (RT-qPCR) following the mentioned treatment. One-way ANOVA test and Tukey post-hoc test at a p-value equal to/or less than 0.05 were used to analyze the obtained data.
The synthesis of GO nano-sheets in a layered structure with a thickness of 0.76 nm was confirmed by AFM analysis. FESEM showed that the exfoliated sheet of synthesized GO had several micrometers in lateral size. DLS revealed that the mean particle size and density index of synthesized nCur were 57.47 ± 2.14 nm and 10 % respectively. In DLS analysis, rGO-nCur showed more positive surface charge (24 mV) than the nano-sheets of GO. FESEM confirmed the coating of rGO-nCur on elastomeric ligatures. ANOVA revealed that tensile strength of 1.25, 2.5, and 5 % rGO-nCur coated elastomeric ligatures were not decreased statistically significantly (P > 0.05). Mean tensile strength and recorded force of 7.5 and 10 % rGO-nCur coated elastomeric ligatures decreased significantly after 14 days' immersion in the artificial saliva (P < 0.05). On the 28th day of the study, the mean of the tensile strength of elastomeric ligatures coated with 10 % rGO-nCur (13.03 ± 0.10 N) was recorded as 55.90 % of the initial tensile strength (23.31 ± 0.41 N in uncoated elastomeric ligatures), while the mean tensile strength of elastomeric ligatures coated with 7.5 % rGO-nCur (16.01 ± 0.10 N) was measured as 68.94 % of the initial tensile strength (23.22 ± 0.09 N in uncoated elastomeric ligatures). When comparing the coated elastomeric ligatures at 7.5 % and 10 % to the original uncoated elastomeric ligatures at similar time intervals, statistically significant decreases in extension to tensile strength (0.42 to 0.71 mm or 3.02 to 5.05 %; all P < 0.05) were observed. The largest contact angle was measured in elastomeric ligatures coated by 10 % rGO-nCur followed by 7.5 and 5 % rGO-nCur (128 ± 2.19°, 117 ± 2.23°, and 99 ± 1.83°; respectively). The results revealed a rise of 6.4-fold in intracellular ROS and an 11.2 °C increase in the temperature of rGO-nCur solutions following the 450 nm and 980 nm laser irradiation, respectively. The 5 % rGO-nCur coated elastomeric ligature mediated dual-modal PDI/PTI showed the most inhibition of the biofilm formation of S. mutans by 83.62 % (P = 0.00). Significant reductions in water-insoluble EPS were detected in biofilm cultures of S. mutans on 1.25 % rGO-nCur coated elastomeric ligatures following irradiation with dual waves of the 450 nm and 980 nm diode lasers (i.e., dual-modal PDI/PTI; 96.17 %; P = 0.00). The expression levels of comDE, gtfD, and smuT virulence genes were significantly downregulated (7.52-, 13.92-, and 8.23-fold, respectively) in the biofilm cultures of S. mutans on 1.25 % rGO-nCur coated elastomeric ligatures following dual-modal PDI/PTI in comparison with biofilm cultures on non-coated elastomeric ligatures.
5 % rGO-nCur coated elastomeric ligatures following irradiation with dual waves of the 450 and 980 nm diode lasers (dual-modal PDI/PTI), without adverse effects on the physico-mechanical properties of elastomeric ligatures, can be used to inhibit the formation of S. mutans biofilms on the coated elastomeric ligatures around orthodontic brackets.
固定正畸治疗的一个常见副作用是白色斑点病变（WSLs）。变形链球菌是 WSLs 和治疗过程中牙齿龋齿的主要致病因子。根据还原氧化石墨烯-纳米姜黄素（rGO-nCur）的独特特性，本研究旨在研究 rGO-nCur 涂层正畸弹性结扎线的机械性能和抗菌效力，作为一种新型涂层复合材料，用于对变形链球菌生物膜进行双模式光动力失活（PDI）和光热失活（PTI）。
在确认 rGO-nCur 的合成和不同浓度水平（1.25%、2.5%、5%、7.5%和 10%）的 rGO-nCur 涂层弹性结扎线的拉伸强度、力衰减、拉伸强度至延伸率和接触角后，使用万能试验机和静态滴液法分别进行测量。为了研究辐照 rGO-nCur 抑制变形链球菌生物膜形成的机制，分别测量了 450nm 和 980nm 激光照射下 rGO-nCur 溶液中细胞内活性氧（ROS）的产生和温度的升高。使用 450nm 二极管激光（195J/cm）、980nm 二极管激光（195J/cm）和两者的组合（各 78J/cm 照射）（即 PDI、PTI 和双模式 PDI/PTI），分别测定了辐照 rGO-nCur 涂层弹性结扎线的抗生物膜活性和抑制水不溶性胞外多糖（EPS）产生能力。此外，通过定量实时聚合酶链反应（RT-qPCR）评估了参与生物膜形成的毒力基因（comDE、gtfD 和 smuT）的表达。使用单因素方差分析和 Tukey 事后检验，p 值等于或小于 0.05，用于分析获得的数据。
原子力显微镜分析证实了 GO 纳米片层结构的合成，厚度为 0.76nm。FESEM 显示，合成的 GO 层状薄片具有数微米的横向尺寸。动态光散射显示，合成的 nCur 的平均粒径和密度指数分别为 57.47±2.14nm 和 10%。在 DLS 分析中，rGO-nCur 比 GO 纳米片具有更大的正表面电荷（24mV）。FESEM 证实了 rGO-nCur 对弹性结扎线的涂层。方差分析表明，1.25%、2.5%和 5%rGO-nCur 涂层弹性结扎线的拉伸强度没有统计学显著降低（P>0.05）。7.5%和 10%rGO-nCur 涂层弹性结扎线在人工唾液中浸泡 14 天后，平均拉伸强度和记录力显著降低（P<0.05）。在研究的第 28 天，记录了 10%rGO-nCur 涂层弹性结扎线的拉伸强度（13.03±0.10N）为未涂层弹性结扎线初始拉伸强度（23.31±0.41N）的 55.90%，而 7.5%rGO-nCur 涂层弹性结扎线的平均拉伸强度（16.01±0.10N）为未涂层弹性结扎线初始拉伸强度（23.22±0.09N）的 68.94%。在相似的时间间隔内，与原始未涂层弹性结扎线相比，涂层弹性结扎线的延伸至拉伸强度（0.42 至 0.71mm 或 3.02 至 5.05%；均 P<0.05）显著降低。接触角最大的是用 10%rGO-nCur 涂层的弹性结扎线，其次是 7.5%和 5%rGO-nCur 涂层的弹性结扎线（128±2.19°、117±2.23°和 99±1.83°；分别）。结果表明，450nm 和 980nm 激光照射后，rGO-nCur 溶液中的细胞内 ROS 增加了 6.4 倍，温度升高了 11.2°C。5%rGO-nCur 涂层弹性结扎线介导的双模式 PDI/PTI 对变形链球菌生物膜形成的抑制作用最强，抑制率为 83.62%（P=0.00）。在 450nm 和 980nm 二极管激光双波照射下，变形链球菌生物膜培养物中 rGO-nCur 涂层弹性结扎线的水不溶性 EPS 显著减少（双模式 PDI/PTI；96.17%；P=0.00）。在 1.25%rGO-nCur 涂层弹性结扎线双模式 PDI/PTI 处理后，变形链球菌生物膜培养物中 comDE、gtfD 和 smuT 毒力基因的表达水平显著下调（分别为 7.52 倍、13.92 倍和 8.23 倍）与非涂层弹性结扎线的生物膜培养物相比。
在双波 450nm 和 980nm 二极管激光照射下（双模式 PDI/PTI），无弹性结扎线物理机械性能不良的 5%rGO-nCur 涂层可用于抑制正畸支架周围弹性结扎线变形链球菌生物膜的形成。
