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| 低分子量褐藻糖胶对破骨细胞凋亡的影响 |
| Effect of low-molecular-weight fucoidan on osteoclast apoptosis |
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| DOI：10.3969/j.issn.1006-7108.2016.06.001 |
| 中文关键词: 低分子量褐藻糖胶 破骨细胞 骨质疏松 凋亡 caspase-3 BCL-2 |
| 英文关键词:Low-molecular-weight fucoidan Osteoclasts Osteoporosis Apoptosis Caspase-3 BCL-2 |
| 基金项目:天津市卫生局科技基金（2011KZ57）；天津市中医药管理局（13123） |
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| 中文摘要: |
| 目的 探讨低分子量褐藻糖胶（LMWF）对小鼠单核细胞RAW264.7诱导成熟破骨细胞凋亡的影响。方法 通过100ng/mL RANKL诱导RAW264.7细胞株分化为破骨细胞，经TRAP特异性染色和骨吸收陷窝对诱导后的细胞进行鉴定。鉴定成功后，用100ng/mL RANKL诱导RAW264.7细胞株5d后，使用含有LMWF的培养基继续培养3d，通过对TRAP阳性细胞计数和分析骨吸收面积来观察低分子量褐藻糖胶对破骨细胞的抑制和骨吸收功能情况；采用流式细胞术检测LMWF对破骨细胞凋亡的影响，capsase-3活性测试试剂盒检测LMWF对capsase-3活性进行测定；RT-PCR检测LMWF对成熟破骨细胞BAX与BCL-2基因表达的影响。结果 单纯采用100ng/mL的RANKL可成功诱导成熟的、有功能的破骨细胞。LMWF可以明显抑制RANKL诱导成熟破骨细胞的形成以及成熟破骨细胞的骨吸收功能；流式细胞术显示LMWF可增加成熟破骨细胞的早期凋亡率；并且能升高capsase-3的活性；PCR显示LMWF可明显下调破骨细胞凋亡相关的BCL-2和上调BAX基因mRNA表达，降低BCL-2/BAX的比值。结论 低分子量褐藻糖胶可抑制破骨细胞的活性与骨吸收能力，促进破骨细胞凋亡，其主要机制是通过下调BCL-2和上调BAX mRNA基因表达实现的。 |
| 英文摘要: |
| Abstract: Objective To investigate the effect of low-molecular-weight fucoidan (LMWF) on the apoptosis of RAW264.7 murine monocyte-induced mature osteoclasts. Methods RAW264.7 cells were induced to differentiate into osteoclasts by 100ng/ml RANKL. The induced cells were identified using TRAP staining and specific bone resorption pits. After successful authentication, RAW264.7 cells were cultured with 100ng/ml RANKL for 5 days, and then with LMWF (5ng/ml) for 3 days. TRAP-positive cells and bone resorption area were observed to analyze the inhibition effect of LMWF on osteoclasts and bone resorption. The effect of LMWF on apoptosis of osteoclasts was detected using flow cytometry. Caspase-3 activity was measured using the caspase-3 activity assay kit. The effect of LMWF on BAX and BCL-2 gene expression was detected using RT-PCR. Results Mature and functional osteoclasts were successfully induced by 100ng/ml RANKL. LMWF significantly inhibited the activity of mature osteoclasts and osteoclast resorption. LMWF promoted osteoclast apoptosis and caspases-3 activity. LMWF down-regulated apoptosis-associated BCL-2 and up-regulated BAX mRNA expression, thus decreasing the BCL-2/BAX ratio. Conclusion LMWF inhibits osteoclast activity and promotes osteoclast apoptosis by down-regulation of BCL-2 and up-regulation of BAX mRNA gene expression. |
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