摘要
该研究旨在探究高糖抑制骨髓基质细胞(bone marrow stroma cells,BMSCs)成骨分化的作用及其可能的机制。采用25.5 mmol/L的葡萄糖(HG)模拟高糖微环境,通过碱性磷酸酶(ALP)染色及生化定量检测成骨分化水平,qRT-PCR检测成骨分化相关基因mRNA表达水平,CCK-8检测细胞增殖活性,荧光探针H2DCFDA检测细胞活性氧(reactive oxygen species,ROS)水平,脂质过氧化氢荧光探针Liperfluo检测脂质过氧化产物(lipid peroxidation products,LPO)水平,流式细胞仪定量检测平均荧光强度,Western blot检测谷胱甘肽过氧化物酶4(glutathione peroxidase 4,GPX4)的表达水平,透射电子显微镜(transmission electron microscopy,TEM)观察线粒体超微结构。结果显示,高糖(HG)显著抑制BMSCs成骨分化及细胞增殖活性,导致细胞内ROS及LPO水平显著升高,而铁死亡抑制剂可以恢复BMSCs成骨分化及细胞增殖活性,降低ROS及LPO水平,除此以外高糖(HG)导致BMSCs线粒体皱缩、体积变小、膜密度增高、嵴减少、细胞核形态变化不明显等铁死亡样改变。综上所述,高糖通过诱导铁死亡从而抑制BMSCs成骨分化功能。
This study investigated the effect of high glucose on osteoblast differentiation of BMSCs (bone marrow stroma cells) and its possible mechanism.Using 25.5 mmol/L glucose (HG) to simulate high glucose microenvironment,the level of osteoblast differentiation was detected by ALP (alkaline phosphatase) staining and biochemical quantification.Cell proliferation activity was detected by CCK-8.ROS (reactive oxygen species) was detected by fluorescence probe H2DCFDA and Liperfluo probe was used to detect LPO (lipid peroxidation products),and the mean fluorescence intensity was quantitatively detected by flow cytometry.The expression level of GPX4 (glutathione peroxidase 4) was detected by Western blot.The ultrastructure of mitochondria was observed by TEM (transmission electron microscopy).These results showed that high glucose significantly inhibited osteoblast differentiation and cell proliferation activity of BMSCs,leading to a significant increase in intracellular ROS and LPO levels,while ferroptosis inhibitors could restore osteoblast differentiation and cell proliferation activity of BMSCs,and reduce ROS and LPO levels.In addition,high glucose resulted in mitochondrial shrinkage,smaller volume,increased membrane density,reduced cristae and nucleus.In conclusion,high glucose inhibits osteoblast differentiation of BMSCs by inducing ferroptosis.
作者
张希
邵高海
ZHANG Xi;SHAO Gaohai(Yongchuan Hospital,Chongqing Medical University,Chongqing 402177,China)
出处
《中国细胞生物学学报》
CAS
CSCD
2021年第9期1810-1817,共8页
Chinese Journal of Cell Biology
基金
重庆市科委技术创新与应用示范项目(批准号:cstc2018jscx-msyb X0132)
重庆医科大学附属永川医院院级项目(批准号:YJLC201719)资助的课题。
关键词
高糖
铁死亡
骨髓基质细胞
成骨分化
high glucose
ferroptosis
bone marrow stroma cells
osteoblast differentiation
