Departamento de Biología, Bioquímica y Farmacia, Universidad Nacional del Sur (UNS), 8000, Bahía Blanca, Buenos Aires, Argentina; Instituto de Ciencias Biológicas y Biomédicas del Sur (INBIOSUR), Argentina, Consejo Nacional de Investigaciones Científicas y Técnicas (CONICET), 8000, Bahía Blanca, Buenos Aires, Argentina.
Departamento de Biología, Bioquímica y Farmacia, Universidad Nacional del Sur (UNS), 8000, Bahía Blanca, Buenos Aires, Argentina; Instituto de Ciencias Biológicas y Biomédicas del Sur (INBIOSUR), Argentina, Consejo Nacional de Investigaciones Científicas y Técnicas (CONICET), 8000, Bahía Blanca, Buenos Aires, Argentina.
Biochimie. 2022 Aug;199:46-59. doi: 10.1016/j.biochi.2022.04.003. Epub 2022 Apr 18.
There is a global trend in the use of natural bioactive compounds to complement conventional therapies in bone diseases. In this work, we studied the effects of the phytoestrogen quercetin (QUE) in healthy and tumor osteoblasts. We found that QUE (1 μM, 48 h) significantly increased the cell number and the viability of healthy human osteoblasts (hFOB cells) determined by a trypan blue and a MTS assay, respectively, among other concentrations tested. In addition, wound healing and cellular adhesion assays also demonstrated that 1 μM of QUE significantly stimulated both parameters in osteoblasts. Moreover, osteoblast differentiation was also triggered by QUE in an osteogenic medium by measuring alkaline phosphatase activity, calcium deposition, and collagen levels. Herein, a concentration of 0.01 μM of QUE showed an increment in these differentiation markers and an activation of AKT/GSK3β/β-catenin pathway, determined by a Western blot analysis. In addition, immunocytochemistry and subcellular fraction studies indicated an increase of β-catenin localization in the plasma membrane after QUE treatment. Otherwise, QUE (20-100 μM) decreased the cell number and the viability in tumor osteoblasts (ROS 17/2.8 cells) after 48 h. Furthermore, QUE (100 μM) decreased AKT(Ser473) and the pro-apoptotic protein BAD(Ser136) phosphorylation. In addition, the ERK1/2 phosphorylation increased leading to osteosarcoma cell death since pre-treatment with the MEK inhibitor PD98059 had reverted QUE effect. Altogether, these results indicate that low concentrations of QUE stimulate osteoblastogenesis but have no effect on the growth of tumor osteoblast cells, for which only high concentrations are efficient.
在骨骼疾病的常规治疗中，利用天然生物活性化合物来补充治疗的方法已成为一种全球趋势。在这项工作中，我们研究了植物雌激素槲皮素（QUE）对健康和肿瘤成骨细胞的影响。我们发现，QUE（1 μM，48 h）在其他测试浓度中，能显著增加健康人成骨细胞（hFOB 细胞）的细胞数量和活力，分别通过台盼蓝和 MTS 测定。此外，划痕愈合和细胞黏附试验也表明，1 μM 的 QUE 能显著刺激成骨细胞的这两个参数。此外，在成骨培养基中，QUE 还能通过测量碱性磷酸酶活性、钙沉积和胶原蛋白水平来触发成骨细胞的分化。在此，QUE 的浓度为 0.01 μM 时，能增加这些分化标志物，并激活 AKT/GSK3β/β-catenin 通路，通过 Western blot 分析来确定。此外，免疫细胞化学和亚细胞部分研究表明，在 QUE 处理后，β-catenin 在质膜中的定位增加。另一方面，QUE（20-100 μM）在 48 h 后降低了肿瘤成骨细胞（ROS 17/2.8 细胞）的细胞数量和活力。此外，QUE（100 μM）降低了 AKT（Ser473）和促凋亡蛋白 BAD（Ser136）的磷酸化。此外，ERK1/2 磷酸化增加，导致骨肉瘤细胞死亡，因为用 MEK 抑制剂 PD98059 预处理已经逆转了 QUE 的作用。总之，这些结果表明，低浓度的 QUE 能刺激成骨细胞生成，但对肿瘤成骨细胞的生长没有影响，只有高浓度的 QUE 才有效。
