Jaiklaew Sukrit, Tansriratanawong Kallapat
Department of Oral Medicine and Periodontology, Faculty of Dentistry, Mahidol University, Bangkok, Thailand.
Eur J Dent. 2025 Feb;19(1):70-79. doi: 10.1055/s-0044-1786844. Epub 2024 May 17.
This study aimed to investigate and compare the influence of hypoxic conditions on cytotoxicity, cellular migration, and osteogenic differentiation of aged periodontal ligament (PDL) cells.
Isolated human PDL cells from aged and young subjects were cultured under hypoxic conditions, which were treated with hydrogen peroxide (HO) (0, 25, 50, 100, 200, and 500 µM). To assess cytotoxicity, lactate dehydrogenase release was determined by the optical density at 490 nm, and the percentage of cell death was calculated. An wound healing assay was performed over 24 to 48 hours for cellular migration determination. Osteogenic differentiation was determined by alizarin red staining and osteogenic gene expression, including the expression of runt-related transcription factor 2 (RUNX2), alkaline phosphatase (ALP), and osteopontin (OPN).
There was a significant difference in the percentage of cell death with high hypoxic condition (200 and 500 µM) compared to low hypoxic conditions on both day 1 and 2. The highest cellular migration was depicted at 50 µM in both young and aged groups of the wound healing assay. Osteogenic gene expression of RUNX2 in the aged group was increased at 25 and 50 µM hypoxic condition at day 7, but the expression was gradually decreased after 14 days. On the contrary, the expression of ALP and OPN in the aged group was increased at day 14. Only OPN had been found to be statistically significantly different when compared with gene expression at day 7 and 14 ( < 0.05). The results showed no statistically significant differences when compared with the young and aged groups in all genes and all concentrations.
The concentration of low hypoxic condition (25-50 µM) was proposed to promote cell viability, cellular migration, and osteogenic differentiation in aged PDL cells. We suggested that the potential of aged PDL cells for use in cell therapy for periodontal regeneration might possibly be similar to that of young PDL cells.
本研究旨在调查和比较低氧条件对老年牙周膜（PDL）细胞的细胞毒性、细胞迁移和成骨分化的影响。
从老年和年轻受试者中分离出的人PDL细胞在低氧条件下培养，并用过氧化氢（HO）（0、25、50、100、200和500μM）处理。为评估细胞毒性，通过490nm处的光密度测定乳酸脱氢酶释放量，并计算细胞死亡百分比。在24至48小时内进行伤口愈合试验以测定细胞迁移。通过茜素红染色和成骨基因表达，包括 runt相关转录因子2（RUNX2）、碱性磷酸酶（ALP）和骨桥蛋白（OPN）的表达来确定成骨分化。
与低氧条件相比，高氧条件（200和500μM）下第1天和第2天的细胞死亡百分比存在显著差异。在伤口愈合试验的年轻和老年组中，50μM时细胞迁移率最高。老年组中RUNX2的成骨基因表达在第7天的低氧条件为25和50μM时增加，但在14天后表达逐渐降低。相反，老年组中ALP和OPN的表达在第14天增加。与第7天和第14天的基因表达相比，仅发现OPN有统计学显著差异（P<0.05）。在所有基因和所有浓度下，与年轻和老年组相比，结果均无统计学显著差异。
建议低氧条件浓度（25 - 50μM）可促进老年PDL细胞的细胞活力、细胞迁移和成骨分化。我们认为，老年PDL细胞用于牙周再生细胞治疗的潜力可能与年轻PDL细胞相似。
