Pourhajibagher Maryam, Bahador Abbas
Dental Research Center, Dentistry Research Institute, Tehran University of Medical Sciences, Tehran, Iran.
Dental Research Center, Dentistry Research Institute, Tehran University of Medical Sciences, Tehran, Iran; Fellowship in Clinical Laboratory Sciences, BioHealth Lab, Tehran, Iran.
Photodiagnosis Photodyn Ther. 2022 Sep;39:103020. doi: 10.1016/j.pdpdt.2022.103020. Epub 2022 Jul 16.
Despite the high success rate of root canal treatment, failures are observed in a broad range of cases. Therefore, the need for novel approaches with the development of new generations of antimicrobial agents and intracellular drug delivery systems as adjunctive therapy is undeniable. In this study, we investigated the antimicrobial effects of antimicrobial photodynamic therapy (aPDT) using dermcidin‑derived peptide DCD‑1L loaded onto aptamer-functionalized emodin nanoparticles (Apt@EmoNp-DCD-1L) against Enterococcus faecalis as one of the most common bacteria involved in recurrent root canal treatment failures.
Following preparation of EmoNp-DCD-1L, the binding of selected labeled Apt to EmoNp-DCD-1L was performed, followed by the specificity of Apt@EmoNp-DCD-1L to E. faecalis was determined. The antimicrobial potential of aPDT was then assessed after the determination of the minimum inhibitory concentration (MIC) of Apt@EmoNp-DCD-1L. The molecular docking analysis was conducted to evaluate the potential binding modes of EmoNp to the proteins involved in E. faecalis pathogenesis. Eventually, the anti-virulence capacity of Apt@EmoNp-DCD-1L-mediated aPDT was investigated via quantitative real-time PCR (qRT-PCR) assay following measurement of intracellular reactive oxygen species (ROS) generation.
The binding specificity of Apt@EmoNp-DCD-1L to E. faecalis was confirmed by flow cytometry. The results showed that the cell viability of E. faecalis exposed to aPDT groups employing the sub-MIC doses of Apt@EmoNp-DCD-1L (7.8 and 15.6 µM) was significantly reduced compared to the control group (P < 0.05). Also, Apt@EmoNp-DCD-1L in combination with a blue laser light was capable of enhancing the anti-biofilm activity of aPDT against E. faecalis biofilm. Data obtained from the qRT-PCR analysis showed significant downregulation in the expression level of genes involved in bacterial biofilm formation after exposure to aPDT (P < 0.05).
This in vitro study highlights that aPDT with the minimum concentration of Apt@EmoNp-DCD-1L can be considered as a targeted bio-theragnostic agent for the detection and elimination of E. faecalis in the dispersed and biofilm states.
尽管根管治疗成功率很高，但在很多病例中仍会出现治疗失败的情况。因此，随着新一代抗菌剂和细胞内药物递送系统的发展，需要采用新方法作为辅助治疗，这一点不可否认。在本研究中，我们研究了载有防御素衍生肽DCD-1L的适配体功能化大黄素纳米颗粒（Apt@EmoNp-DCD-1L）介导的抗菌光动力疗法（aPDT）对粪肠球菌的抗菌作用，粪肠球菌是导致根管再治疗失败的最常见细菌之一。
制备EmoNp-DCD-1L后，将选定的标记适配体与EmoNp-DCD-1L进行结合，随后测定Apt@EmoNp-DCD-1L对粪肠球菌的特异性。在确定Apt@EmoNp-DCD-1L的最低抑菌浓度（MIC）后，评估aPDT的抗菌潜力。进行分子对接分析以评估EmoNp与粪肠球菌致病相关蛋白的潜在结合模式。最终，在测量细胞内活性氧（ROS）生成后，通过定量实时PCR（qRT-PCR）测定法研究Apt@EmoNp-DCD-1L介导的aPDT的抗毒力能力。
通过流式细胞术证实了Apt@EmoNp-DCD-1L对粪肠球菌的结合特异性。结果表明，与对照组相比，暴露于采用亚MIC剂量的Apt@EmoNp-DCD-1L（7.8和15.6 μM）的aPDT组中的粪肠球菌细胞活力显著降低（P < 0.05）。此外，Apt@EmoNp-DCD-1L与蓝光激光联合能够增强aPDT对粪肠球菌生物膜的抗生物膜活性。从qRT-PCR分析获得的数据显示，暴露于aPDT后，参与细菌生物膜形成的基因表达水平显著下调（P < 0.05）。
这项体外研究强调，使用最低浓度的Apt@EmoNp-DCD-1L的aPDT可被视为一种靶向生物诊断治疗剂，用于检测和消除分散状态和生物膜状态下的粪肠球菌。
