CAJ | 학술논문

目的：研究小白菊内酯对NF-κB受体活化因子配体（RANKL）诱导的RAW264.7细胞向破骨细胞分化的影响。方法以 RANKL 单独诱导小鼠单核巨噬细胞 RAW264.7细胞为对照组，分别采用不同浓度（0.5、1、2μmol/L）小白菊内酯联合RANKL诱导RAW264.7细胞分化培养。第3、5、7天采用抗酒石酸酸性磷酸酶（TRAP）染色法检测破骨细胞样细胞并计数； ELISA法检测第7天各组培养液上清中抗酒石酸酸性磷酸酶5b（TRAP5b）含量；实时（RT）-PCR检测第7天各组破骨细胞标志基因降钙素受体（CTR）和MMP-9的表达。使用SPSS 17.0统计学软件进行方差分析和t检验以比较组间差异。结果不同的培养条件下RANKL均能诱导RAW264.7细胞分化为成熟的破骨细胞。与同期对照组比较，诱导第3、5、7天，小白菊内酯各组破骨细胞数均明显减少；随小白菊内酯浓度的增大而破骨细胞数减少幅度更显著，0.5、1、2μmol/L各组第7天破骨细胞数较对照组分别下降为36.3%、40.8%、49.3%，细胞数差异有统计学意义（t=7.758，8.742，10.56；P<0.05）。各组第7天培养液中TRAP5b含量变化与细胞计数结果相符（P<0.05）。 TRAP阳性破骨细胞的CTR、MMP-9表达量随着小白菊内酯浓度增加而减少，2μmol/L小白菊内酯组最低，与对照组比较小白菊内酯0.5、1、2μmol/L各组差异均有统计学意义（P<0.05）。结论小白菊内酯对RANKL诱导下RAW264.7细胞向破骨细胞的分化具有抑制作用，且抑制作用具有剂量依赖性。
목적：연구소백국내지대NF-κB수체활화인자배체（RANKL）유도적RAW264.7세포향파골세포분화적영향。방법이 RANKL 단독유도소서단핵거서세포 RAW264.7세포위대조조，분별채용불동농도（0.5、1、2μmol/L）소백국내지연합RANKL유도RAW264.7세포분화배양。제3、5、7천채용항주석산산성린산매（TRAP）염색법검측파골세포양세포병계수； ELISA법검측제7천각조배양액상청중항주석산산성린산매5b（TRAP5b）함량；실시（RT）-PCR검측제7천각조파골세포표지기인강개소수체（CTR）화MMP-9적표체。사용SPSS 17.0통계학연건진행방차분석화t검험이비교조간차이。결과불동적배양조건하RANKL균능유도RAW264.7세포분화위성숙적파골세포。여동기대조조비교，유도제3、5、7천，소백국내지각조파골세포수균명현감소；수소백국내지농도적증대이파골세포수감소폭도경현저，0.5、1、2μmol/L각조제7천파골세포수교대조조분별하강위36.3%、40.8%、49.3%，세포수차이유통계학의의（t=7.758，8.742，10.56；P<0.05）。각조제7천배양액중TRAP5b함량변화여세포계수결과상부（P<0.05）。 TRAP양성파골세포적CTR、MMP-9표체량수착소백국내지농도증가이감소，2μmol/L소백국내지조최저，여대조조비교소백국내지0.5、1、2μmol/L각조차이균유통계학의의（P<0.05）。결론소백국내지대RANKL유도하RAW264.7세포향파골세포적분화구유억제작용，차억제작용구유제량의뢰성。
Objective To study the effects of parthenolide on osteoclast differentiation of RAW264. 7 cell induced by receptor activator of nuclear factor κB ligand (RANKL). Methods The mouse macrophage RAW264.7 cells induced by RANKL was used alone as the control group, different concentrations of par-thenolide (0.5, 1, 2 μmol/L) were added to culture the RAW264.7 cells. On the third, fifth and seventh day, the tartrate resistant acid phosphatase (TRAP) staining method was used to detect osteodast-like cells and the cell number was count;the contents of tartrate resistant acid phosphatase (TRAP5b) in the Culture supernatant of each groups were detected by enzyme linked immunosorbent assay (ELISA) and the expression of osteodast marker gene alcitonin receptor (CTR) and matrix metalloproteinase (MMP)-9 in each groups were detected by realtime-polymerase chain reaction (PCR) on the seventh day. We use Chi-square test and t test to test the differences between groups by SPSS 17.0. Results In different culture conditions, RANKL could always induce the RAW264.7 cell differentiate into mature osteoclasts. Compared with the control group at the same time control group, on the third, fifth and seventh day, he number of mature osteoclasts induced were obviously decreased in groups adding different concentration of PAR; the number of induced osteoclasts decreased along with the increase of parthenolide concentration, on the seventh day in 0.5, 1, 2 μmol/L concentration PAR groups, the number of mature osteoclasts compared with the control group were descended 36.3%, 40.8%, 49.3%(t=7.758, 8.742, 10.56;P<0.05);the contents of TRAP5b in the culture supernatant were consistent with the cell counting results on the seventh day (P<0.05). The expression of CTR and MMP-9 by TRAP positive osteoclasts decreased along with the increase of parthenolide concentration, and the 2 μmol/L group was the lowest. Compared with the control group, there were statistically significant differences with the different PAR concentration groups 0.5, 1, 2 μmol/L (P<0.05). Conclusion Parthenolide can inhibit RANKL induced RAW264.7 differentiation into osteoclast cells, and the inhibition is dose dependent.