摘要
目的研究天山雪莲细胞培养物对破骨细胞形成以及成熟破骨细胞活性的影响。方法小鼠巨噬细胞RAW264.7在核因子κB受体活化因子配体(RANKL)诱导下形成破骨细胞,诱导过程中加入天山雪莲细胞培养物(以天山雪莲总黄酮计算),通过抗酒石酸酸性磷酸酶(TRAP)和Hoechst 33342染色观察破骨细胞形成,以对硝基苯磷酸酯作底物检测TRAP活性;对诱导形成的破骨细胞进行乳酸脱氢酶(LDH)活性检测和Annexin V-PI双染流式细胞术进行细胞凋亡检测,RT-PCR检测Trap和基质金属蛋白酶(Mmp9)m RNA的表达。结果天山雪莲细胞培养物能够显著抑制破骨细胞的形成以及TRAP酶活;天山雪莲总黄酮小于125μg/m L时,对破骨前体细胞RAW264.7无细胞毒性,但会破坏成熟破骨细胞细胞膜,显著提高胞外LDH酶活性,并诱导破骨细胞凋亡;62.5μg/m L天山雪莲总黄酮会显著下调破骨细胞的标志物Trap和Mmp9 mRNA表达。结论天山雪莲细胞培养物对破骨细胞形成有抑制作用,并能破坏已形成的破骨细胞,降低骨吸收活性。
AIM To investigate the effects of Saussurea involucrata cell cultures on the formation of osteoclast as well as the activity of mature osteoporosis. METHODS Except in the control group,murine macrophage RAW264. 7 cells were given the inducing medium receptor activator of NF-κB ligand（ RANKL）. The effect of S.involucrata cell cultures（ total flovenoids from S. involucrata as calculated） on osteoclastogenesis was observed by tartrate-resistant acid phosphatase（ TRAP） and Hoechst 33342 staining and measured by TRAP activity. The viability of osteoclast was detected by LDH assay and flow cytometry assay. RT-PCR was employed to determine the expression levels of Trap and matrix metalloproteinase-9（ Mmp9） m RNA in osteoclast. RESULTS S. involucrata cell cultures significantly inhibited RANKL-induced formation of multinucleated osteoclast and TRAP activity in a dose-dependent manner. MTT activity assay showed that S. involucrata cell cultures（ ≤125 μg / m L） were nontoxic to RAW264. 7 cells. But it could destroy the cell membrane of osteoclast,significantly improving the extracellular LDH activity and induced apoptosis. S. involucrata cell culture markedly down-regulated Trap and Mmp9 m RNA expression at the concentration of 62. 5 μg / m L. CONCLUSION S. involucrata cell cultures have the potential to inhibit the osteoclast differentiation,induce mature osteoclast apoptosis and decrease the activity of bone resorption.
出处
《中成药》
CAS
CSCD
北大核心
2016年第1期1-6,共6页
Chinese Traditional Patent Medicine
