Key Laboratory of Dental Maxillofacial Reconstruction and Biological Intelligence Manufacturing, Gansu Province, School of Stomatology, Lanzhou University, Lanzhou, China.
Front Cell Infect Microbiol. 2021 Nov 19;11:784153. doi: 10.3389/fcimb.2021.784153. eCollection 2021.
Secondary caries caused by oral microbiome dysbiosis and hybrid layer degradation are two important contributors to the poor resin-dentin bond durability. Cavity cleansers with long-term antimicrobial and anti-proteolytic activities are in demand for eliminating bacteria-induced secondary caries and preventing hybrid layers from degradation. The objectives of the present study were to examine the long-term antimicrobial effect and anti-proteolytic potential of poly(amidoamine) dendrimers with amino terminal groups (PAMAM-NH) cavity cleanser.
Adsorption tests by attenuated total reflectance-infrared (ATR-IR) spectroscopy and confocal laser scanning microscopy (CLSM) were first performed to evaluate whether the PAMAM-NH cavity cleanser had binding capacity to dentin surface to fulfill its relatively long-term antimicrobial and anti-proteolytic effects. For antibacterial testing, , , and were grown on dentin surfaces, prior to the application of cavity cleanser. Colony-forming unit (CFU) counts and live/dead bacterial staining were performed to assess antibacterial effects. Gelatinolytic activity within the hybrid layers was directly detected by zymography. Adhesive permeability of bonded interface and microtensile bond strength were employed to assess whether the PAMAM-NH cavity cleanser adversely affected resin-dentin bonding. Finally, the cytotoxicity of PAMAM-NH was evaluated by the Cell Counting Kit-8 (CCK-8) assay.
Adsorption tests demonstrated that the binding capacity of PAMAM-NH on dentin surface was much stronger than that of 2% chlorhexidine (CHX) because its binding was strong enough to resist phosphate-buffered saline (PBS) washing. Antibacterial testing indicated that PAMAM-NH significantly inhibited bacteria grown on the dentin discs as compared with the control group (p < 0.05), which was comparable with the antibacterial activity of 2% CHX (p > 0.05). Hybrid layers conditioned with PAMAM-NH showed significant decrease in gelatin activity as compared with the control group. Furthermore, PAMAM-NH pretreatment did not adversely affect resin-dentin bonding because it did not decrease adhesive permeability and microtensile strength. CCK-8 assay showed that PAMAM-NH had low cytotoxicity on human dental pulp cells (HDPCs) and L929.
PAMAM-NH cavity cleanser developed in this study could provide simultaneous long-term antimicrobial and anti-proteolytic activities for eliminating secondary caries that result from a dysbiosis in the oral microbiome and for preventing hybrid layers from degradation due to its good binding capacity to dentin collagen matrix, which are crucial for the maintenance of resin-dentin bond durability.
口腔微生物群落失调和混合层降解引起的继发龋是导致树脂-牙本质粘结耐久性差的两个重要原因。具有长期抗菌和抗蛋白水解活性的窝洞清洁剂可用于消除细菌引起的继发龋，并防止混合层降解。本研究的目的是研究具有氨基末端的聚（酰胺-胺）树枝状大分子（PAMAM-NH）窝洞清洁剂的长期抗菌效果和抗蛋白水解潜力。
通过衰减全反射-红外（ATR-IR）光谱和共焦激光扫描显微镜（CLSM）进行吸附试验，以评估 PAMAM-NH 窝洞清洁剂是否具有与牙本质表面结合的能力，以实现其相对长期的抗菌和抗蛋白水解作用。在应用窝洞清洁剂之前，将 、 、 和 种植在牙本质表面上进行抗菌试验。通过平板计数法和活菌/死菌染色评估抗菌效果。通过 zymography 直接检测混合层内的明胶酶活性。采用粘结界面的胶渗透和微拉伸粘结强度评估 PAMAM-NH 是否对树脂-牙本质粘结产生不利影响。最后，通过细胞计数试剂盒-8（CCK-8）测定法评估 PAMAM-NH 的细胞毒性。
吸附试验表明，PAMAM-NH 与牙本质表面的结合能力远强于 2%洗必泰（CHX），因为其结合能力足以抵抗磷酸盐缓冲盐水（PBS）的冲洗。抗菌试验表明，与对照组相比，PAMAM-NH 显著抑制了种植在牙本质片上的细菌（p<0.05），与 2%CHX 的抗菌活性相当（p>0.05）。用 PAMAM-NH 处理的混合层的明胶活性明显低于对照组。此外，PAMAM-NH 预处理不会降低树脂-牙本质粘结的粘结强度，因为它不会增加胶渗透。CCK-8 测定表明 PAMAM-NH 对人牙髓细胞（HDPCs）和 L929 的细胞毒性较低。
本研究开发的 PAMAM-NH 窝洞清洁剂具有良好的牙本质胶原基质结合能力，可同时提供长期的抗菌和抗蛋白水解活性，以消除由口腔微生物群落失调引起的继发龋，并防止混合层降解，这对维持树脂-牙本质粘结耐久性至关重要。
