Graduate Institute of Clinical Dentistry, School of Dentistry, College of Medicine, National Taiwan University, Taipei, Taiwan.
Department of Dentistry, National Taiwan University Hospital, Taipei, Taiwan.
J Formos Med Assoc. 2021 Jun;120(6):1332-1339. doi: 10.1016/j.jfma.2020.11.022. Epub 2020 Dec 17.
BACKGROUND/PURPOSE: 2-hydroxyethyl methacrylate (HEMA) is one of the most major components in dentin bonding systems. Uncured HEMA is eluted through the dentin and harmful to pulp cells. The study aimed to investigate the death pattern, morphological change and factors of human dental pulp cells (HDPCs) cultured with low-dose HEMA.
HDPCs were cultured with low-dose concentration of HEMA at 0 mM (control), 0.125 mM, 0.25 mM, 1 mM, 2 mM and 4 mM on Day 3 and 5. The cell morphology was observed with F-actin immunocytochemical staining. The flow cytometry was used to analyze the death pattern. NF-κB and Trx-1 were measured using ELISA kits.
The major death pattern was early apoptosis and late apoptosis. The morphological characteristics of apoptosis were observed clearly at 4 mM on Day 3 and Day 5. The phosphorylated NF-κB normalized to total NF-κB protein was significantly higher at 2 mM and 4 mM on Day 5. There was no difference of Trx-1 on Day 3, but significantly higher at 0.25 mM and 1 mM on Day 5. The trend line of phosphorylated NF-κB and Trx-1 showed highly positive correlations with HEMA concentration.
The significant cellular morphology characteristics of apoptosis can be observed at higher dose and longer period after exposed to uncured HEMA. The expression of NF-κB was following the ratio of late apoptosis at longer exposure period. Clinically, the remaining dentin thickness should be enough to decrease HEMA concentration and thus to protect pulp cells free from harm.
背景/目的：2-羟乙基甲基丙烯酸酯（HEMA）是牙本质粘结系统中最主要的成分之一。未固化的 HEMA 通过牙本质洗脱，对牙髓细胞有害。本研究旨在探讨低浓度 HEMA 培养人牙髓细胞（HDPCs）的死亡模式、形态变化及相关因素。
HDPCs 在 0 mM（对照）、0.125 mM、0.25 mM、1 mM、2 mM 和 4 mM HEMA 低浓度下培养 3 天和 5 天。用 F-肌动蛋白免疫细胞化学染色观察细胞形态。采用流式细胞术分析死亡模式。采用 ELISA 试剂盒测定 NF-κB 和 Trx-1。
主要死亡模式为早期凋亡和晚期凋亡。在 3 天和 5 天的 4 mM 时，观察到凋亡的形态特征清晰。第 5 天，2 mM 和 4 mM 时磷酸化 NF-κB 与总 NF-κB 蛋白的比值显著升高。第 3 天 Trx-1 无差异，但第 5 天 0.25 mM 和 1 mM 时明显升高。磷酸化 NF-κB 和 Trx-1 的趋势线与 HEMA 浓度呈高度正相关。
在暴露于未固化 HEMA 后更高剂量和更长时间，可观察到明显的细胞形态凋亡特征。NF-κB 的表达在更长的暴露时间后与晚期凋亡的比例一致。临床上，剩余的牙本质厚度应足以降低 HEMA 浓度，从而保护牙髓细胞免受伤害。
