Department of Biomaterials Science, Osaka University, Graduate School of Dentistry, Osaka, Japan
Department of Biomaterials Science, Osaka University, Graduate School of Dentistry, Osaka, Japan.
J Dent Res. 2015 Aug;94(8):1085-91. doi: 10.1177/0022034515589282. Epub 2015 Jun 3.
Nanoparticles (NPs) are currently the focus of considerable attention for dental applications; however, their biological effects have not been fully elucidated. The long-term, slow release of matrix metalloproteases (MMPs) digests collagen fibrils within resin-dentin bonds. Therefore, MMP inhibitors can prolong the durability of resin-dentin bonds. However, there have been few reports evaluating the combined effect of MMP inhibition and the cytotoxic effects of NPs for dentin bonding. The aim of this study was to evaluate MMP inhibition and cytotoxic responses to gold (AuNPs) and platinum nanoparticles (PtNPs) stabilized by polyvinylpyrrolidone (PVP) in cultured murine macrophages (RAW264) by using MMP inhibition assays, measuring cell viability and inflammatory responses (quantitative reverse transcription polymerase chain reaction [RT-qPCR]), and conducting a micromorphological analysis by fluorescence and transmission electron microscopy. Cultured RAW264 cells were exposed to metal NPs at various concentrations (1, 10, 100, and 400 µg/mL). AuNPs and PtNPs markedly inhibited MMP-8 and MMP-9 activity. Although PtNPs were cytotoxic at high concentrations (100 and 400 µg/mL), no cytotoxic effects were observed for AuNPs at any concentration. Transmission electron microscopy images showed a significant nonrandom intercellular distribution for AuNPs and PtNPs, which were mostly observed to be localized in lysosomes but not in the nucleus. RT-qPCR analysis demonstrated inflammatory responses were not induced in RAW264 cells by AuNPs or PtNPs. The cytotoxicity of nanoparticles might depend on the core metal composition and arise from a "Trojan horse" effect; thus, MMP inhibition could be attributed to the surface charge of PVP, which forms the outer coating of NPs. The negative charge of the surface coating of PVP binds to Zn(2+) from the active center of MMPs by chelate binding and results in MMP inhibition. In summary, AuNPs are attractive NPs that effectively inhibit MMP activity without cytotoxicity or inflammatory responses.
纳米粒子（NPs）目前是牙科应用的研究热点，但其生物学效应尚未完全阐明。基质金属蛋白酶（MMPs）的长期缓慢释放会消化树脂-牙本质结合处的胶原纤维。因此，MMP 抑制剂可以延长树脂-牙本质结合的耐久性。然而，很少有研究评估 MMP 抑制和纳米粒子的细胞毒性对牙本质结合的联合作用。本研究旨在通过 MMP 抑制测定、细胞活力和炎症反应（定量逆转录聚合酶链反应[RT-qPCR]）的测量以及荧光和透射电子显微镜的微观形态分析，评估由聚乙烯吡咯烷酮（PVP）稳定的金（AuNPs）和铂纳米粒子（PtNPs）对培养的小鼠巨噬细胞（RAW264）的 MMP 抑制和细胞毒性反应。培养的 RAW264 细胞暴露于不同浓度（1、10、100 和 400 µg/mL）的金属 NPs 下。AuNPs 和 PtNPs 显著抑制 MMP-8 和 MMP-9 活性。尽管 PtNPs 在高浓度（100 和 400 µg/mL）时具有细胞毒性，但在任何浓度下均未观察到 AuNPs 的细胞毒性。透射电子显微镜图像显示 AuNPs 和 PtNPs 呈明显的非随机细胞内分布，主要观察到它们定位于溶酶体中，而不是核内。RT-qPCR 分析表明，AuNPs 或 PtNPs 未诱导 RAW264 细胞产生炎症反应。纳米粒子的细胞毒性可能取决于核心金属成分，并源于“木马”效应；因此，MMP 抑制可能归因于 PVP 的表面电荷，PVP 形成 NPs 的外壳。PVP 表面涂层的负电荷通过螯合结合与 MMPs 的活性中心的 Zn(2+)结合，从而导致 MMP 抑制。总之，AuNPs 是一种有吸引力的 NPs，它可以有效抑制 MMP 活性而无细胞毒性或炎症反应。
