Taira Masayuki, Nezu Takashi, Sasaki Minoru, Kimura Shigenobu, Kagiya Tadayoshi, Harada Hidemitsu, Narushima Takayuki, Araki Yoshima
Department of Dental Materials Science and Technology, Iwate Medical University School of Dentistry, Morioka, Japan.
Biomed Mater Eng. 2009;19(1):63-70. doi: 10.3233/BME-2009-0564.
The purpose of this study was to examine gene expressions of macrophage phagocytizing sub-micro Ti particles by a DNA chip. Human monocytic cell line THP-1 was differentiated into macrophages by culturing for two days in medium supplemented with 200 nM phorbol ester (PMA). The macrophages were then cultured in four media: medium without PMA (control); medium with suspended sub-micro Ti particles (0.5 wt%); medium with 1.0 microg/ml lipopolysaccharide (LPS); and medium with LPS and Ti particles. After 6 hours' culture, total RNA were extracted and gene expressions were evaluated by DNA allergy chip with 205 allergy and inflammation related gene spots. We found that phagocytosis of sub-micro Ti particles and LPS independently and synergistically up-regulated 17 inflammation-related genes more than two-fold. The extensive expressions of four genes (CCL1, IL1B, IL6 and IL8) were further confirmed by real-time quantitative PCR. It turned out that dual stimulation of LPS and Ti particles most up-regulated three genes (IL1B, IL6 and IL8), followed by LPS while Ti particles moderately but least increased, suggesting that phagocytosis of sub-micro Ti particles induces moderate inflammation with its degree less than LPS, but phagocytosis of sub-micro Ti particles has the potential to worsen inflammation caused by LPS-stimulated macrophages.
本研究的目的是通过DNA芯片检测巨噬细胞吞噬亚微米级钛颗粒的基因表达。人单核细胞系THP-1在补充有200 nM佛波酯（PMA）的培养基中培养两天后分化为巨噬细胞。然后将巨噬细胞在四种培养基中培养：无PMA的培养基（对照）；含有悬浮亚微米级钛颗粒（0.5 wt%）的培养基；含有1.0 μg/ml脂多糖（LPS）的培养基；以及含有LPS和钛颗粒的培养基。培养6小时后，提取总RNA，并通过具有205个过敏和炎症相关基因点的DNA过敏芯片评估基因表达。我们发现，亚微米级钛颗粒和LPS的吞噬作用独立且协同地上调了17个炎症相关基因两倍以上。通过实时定量PCR进一步证实了四个基因（CCL1、IL1B、IL6和IL8）的广泛表达。结果表明，LPS和钛颗粒的双重刺激对三个基因（IL1B、IL6和IL8）的上调作用最强，其次是LPS，而钛颗粒的上调作用适中但最弱，这表明亚微米级钛颗粒的吞噬作用诱导了中度炎症，其程度低于LPS，但亚微米级钛颗粒的吞噬作用有可能加重LPS刺激的巨噬细胞引起的炎症。
