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| 阿司匹林体外抑制破骨细胞生成的分子机制研究 |
| The molecular mechanism of aspirin in the inhibition of osteoclastogenesis in vitro |
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| DOI：10.3969/j.issn.1006.7108.2015.03.006 |
| 中文关键词: 阿司匹林 破骨细胞 NF-κB MAPKs 骨质疏松 |
| 英文关键词:Aspirin Osteoclast NF-κB MAPKs Osteoporosis |
| 基金项目:国家自然科学基金面上项目（81270959) |
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| 中文摘要: |
| 目的 探讨阿司匹林对破骨细胞生成的影响及其分子机制。方法 体外培养RAW264.7细胞，以100 ng/ml核激活因子κB( NF-κB)受体配体(RANKL)诱导培养，并同时添加不同溶度的阿司匹林(0,0. 25,0. 5,1. 0,1. 5 mmol/L)培养5天。在不同时间点，通过抗酒石酸酸性磷酸酶(TRAP)染色的方法来观察破骨细胞诱导生成能力，用实时荧光PCR方法检测其破骨细胞系标志基因，包括组织蛋白酶K( CTSK)、TRAP、基质金属蛋白酶9( MMP-9)和降钙素受体(CTR) mRNA的表达。裂解不同培养条件的细胞并提取蛋白上样，行western印迹检测NF-κB通道蛋白的表达以及有丝分裂原激活蛋白激酶(MAPKS)通道蛋白的表达，通过免疫荧光的方法分析确定NF-κB的P65的核易位。结果 阿司匹林抑制RANKL诱导的破骨细胞生成，随着阿司匹林浓度增加，破骨细胞形成数量明显减少;其标志性基因TRAP、CTSK、MMP9及CTR的mRNA表达均有所下调;磷酸化的P65、P50、IKB-a、P38以及氨基端激酶(C-JNK)和胞外信号调节蛋白激酶(ERK)蛋白表达均有所减少，而其非磷酸化的蛋 白表达水平无明显变化。阿司匹林同时对NF-κB P65的核易位也表现出抑制效果。结论 在RAW264. 7细胞系中，阿司匹林通过抑制NF-κB系统(P65、P50、IKB-a)和MAPKS系统(P38、C-JNK和ERK)通道的激活来抑制破骨细胞的生成，且在一定范围内和阿司匹林浓度呈正相关。阿司匹林可能具有临床预防及治疗骨质疏松的潜能。 |
| 英文摘要: |
| Objective To explore the influence of aspirin in the generation and differentiation of osteoclasts and its molecular mechanism. Methods RAW264. 7 cells were cultured with 100 ng/ml RANKL treatment and additioned with aspirin solution (0， 0.25，0. 5，1.0，1.5 mmol/L) of 5 days. TRAP staining was used to observe the generation of osteoclasts at different time points. Expressions of osteoclast marker mRNAs，including CTSK，MMP-9，and CTR，were detected with real-time PCR. NF-κB and MAPKS protein expressions were detected using Western blotting. Nuclear translocation of NF-κB p65 was analyzed using immunofluorescence technique. Results Aspirin suppressed RANKL-induced osteoclast formation. With the increase of aspirin concentration，the expressions of marker genes including TRAP，CTSK，MMP9，and CTR mRNA decreased，and the expressions of phosphorylated P65,P50,IKB- a,P38,C-JNK,and ERK protein reduced. However,aspirin showed no effect on non- phosphorylated proteins and inhibited P65 dislocation. Conclusion Aspirin inhibits the osteoclastogenesis of RAW264. 7 cells by inhibiting the activation of NF-κB and MAPK signaling pathways，indicating that aspirin has the potential of preventing and treating osteoporosis. |
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