Regulation of DMT1 on autophagy and apoptosis in osteoblast.
Regulation of DMT1 on autophagy and apoptosis in osteoblast.
复制标题
DMT1对成骨细胞自噬和凋亡的调节
DOI：
10.7150/ijms.17860
复制
发表时间：
2017
影响因子：
3.6
通讯作者：
Yang MW
中科院分区：
文献类型：
--
作者：
Liu F;Zhang WL;Meng HZ;Cai ZY;Yang MW
最近，铁超载与骨质疏松症中发生的骨微结构的变化有关。然而，铁超载对成骨细胞的影响尚不清楚。本研究的目的是探讨二价金属转运蛋白1（DMT1）在骨质疏松病理过程中的功能。成骨细胞 hFOB1.19 细胞在添加不同浓度（0、50、100、200、300、400、500 μmol/L）柠檬酸铁铵（FAC）作为铁离子供体的培养基中培养。我们使用蛋白质印迹和免疫荧光来测定 FAC 处理后 DMT1 的水平。通过使用蛋白质印迹法检测裂解的 caspase 3、BCL2 和 BAX 的水平来评估细胞凋亡。通过蛋白质印迹和免疫荧光检测 LC3 的水平来评估自噬。还通过蛋白质印迹法评估了 Beclin-1 表达。使用自噬抑制剂3-甲基腺嘌呤来确定自噬是否影响FAC诱导的细胞凋亡。我们的结果表明，FAC 增加了 DMT1 的水平，上调了 BCL2 的表达，并下调了凋亡相关蛋白 cleaved caspase 3 和 BAX。 LC3I/LC3II 水平和 beclin-1 也增加，表明 FAC 增加了 hFOB1.19 细胞中自噬体的积累。凋亡抑制剂 3-MA 增加了 FAC 诱导的自噬，但在 DMT1 shRNA hFOB1.19 细胞中却减少了自噬。这些结果提示DMT1表达增加诱导铁超载，铁超载又诱导成骨细胞自噬和凋亡，从而影响骨质疏松的病理过程。阐明 DMT1 作用的机制将有助于确定预防和治疗骨质疏松症的新靶点。
Iron overload has recently been associated with the changes in the bone microstructure that occur in osteoporosis. However, the effect of iron overload on osteoblasts is unclear. The purpose of this study was to explore the function of divalent metal transporter 1 (DMT1) in the pathological processes of osteoporosis. Osteoblast hFOB1.19 cells were cultured in medium supplemented with different concentrations (0, 50, 100, 200, 300, 400, 500 μmol/L) of ferric ammonium citrate (FAC) as a donor of ferric ions. We used western blotting and immunofluorescence to determine the levels of DMT1 after treatment with FAC. Apoptosis was evaluated by detecting the levels of cleaved caspase 3, BCL2, and BAX with western blotting. Autophagy was evaluated by detecting the levels of LC3 with western blotting and immunofluorescence. Beclin-1 expression was also assessed with western blotting. The autophagy inhibitor 3-methyladenine was used to determine whether autophagy affects the apoptosis induced by FAC. Our results show that FAC increased the levels of DMT1, upregulated the expression of BCL2, and downregulated the apoptosis-related proteins cleaved caspase 3 and BAX. Both LC3I/LC3II levels and beclin-1 were also increased, indicating that FAC increases the accumulation of autophagosomes in hFOB1.19 cells. FAC-induced autophagy was increased by the apoptosis inhibitor 3-MA but was reduced in DMT1 shRNA hFOB1.19 cells. These results suggest that the increased expression of DMT1 induces iron overload and iron overload induces osteoblast autophagy and apoptosis, thus affecting the pathological processes of osteoporosis. Clarifying the mechanisms underlying the effects of DMT1 will allow the identification of novel targets for the prevention and treatment of osteoporosis.