Jarrar Hala, Çetİn Altindal Damla, GÜmÜŞderelİoĞlu Menemşe
Bioengineering Division, Department of Chemical Engineering, Faculty of Engineering, Hacettepe University, Ankara Turkey.
Turk J Biol. 2020 Dec 14;44(6):427-436. doi: 10.3906/biy-2007-85. eCollection 2020.
RAW 264.7 cells are one of the most recommended cell lines for investigating the activity and differentiation of osteoclasts. These cells differentiate into osteoclasts in the presence of two critical components: receptor activator of nuclear factor kappa B ligand (RANKL) and macrophage colony stimulating factor (MCSF). Melatonin (MEL) hormone has recently become one of the small molecules used in the field of bone regeneration and bone disease treatment, as it has the ability to inhibit the differentiation of osteoclasts directly by suppression of the NF-κB signaling pathway. The main aim of the current study is to determine sufficient RANKL/MCSF concentrations for differentiation of the cells to osteoclasts and to describe the repressive effect of MEL on the osteoclastogenesis of these cells. In this regard, it was found that 10 ng/mL of RANKL- and MCSF-containing medium is suitable for inducing osteoclastogenesis of the cells. In addition, melatonin at doses in the range of 100-1000 µM does not have a cytotoxic effect. Subsequently, results of tartrate resistant acid phosphatase (TRAP) activity, TRAP staining, and relative expressions of cathepsin K, nuclear factor of activated T cells one (NFATC1), and TRAP genes showed a suppressive effect of MEL -especially 800 µM- on RANKL-induced osteoclastogenesis of these cells.
RAW 264.7细胞是研究破骨细胞活性和分化时最常被推荐的细胞系之一。这些细胞在两种关键成分存在的情况下会分化为破骨细胞：核因子κB受体激活剂配体（RANKL）和巨噬细胞集落刺激因子（MCSF）。褪黑素（MEL）激素最近已成为骨再生和骨疾病治疗领域中使用的小分子之一，因为它能够通过抑制NF-κB信号通路直接抑制破骨细胞的分化。本研究的主要目的是确定使细胞分化为破骨细胞所需的足够的RANKL/MCSF浓度，并描述MEL对这些细胞破骨细胞生成的抑制作用。在这方面，发现含有10 ng/mL RANKL和MCSF的培养基适合诱导细胞的破骨细胞生成。此外，100-1000 µM剂量范围内的褪黑素没有细胞毒性作用。随后，抗酒石酸酸性磷酸酶（TRAP）活性、TRAP染色以及组织蛋白酶K、活化T细胞核因子1（NFATC1）和TRAP基因的相对表达结果显示，MEL（尤其是800 µM）对RANKL诱导的这些细胞的破骨细胞生成具有抑制作用。
