S100A4和S100A6在成骨细胞和破骨细胞分化中表达及意义

Expression and Signficant of S100A4 and S100A6 Proteins in the Differentiation of Osteoblast and Osteoclast

目的探索S100A4和S100A6 mRNA在成骨细胞和破骨细胞分化中表达及意义。方法将MC3T3-E1细胞分成成骨诱导组和对照组,在有或无成骨诱导液的培养基中培养3、7、14和21天时,分别检测各时间段的茜素红染色和碱性磷酸酶染色情况,使用TRIZOL法提取细胞总RNA,通过real time RT-PCR的方法检测S100A4、S100A6、碱性磷酸酶(ALP)、骨保护素(OPG)及核因子-κβ受体活化因子配体(RANKL)mRNA表达情况。RAW264.7细胞分成破骨细胞诱导组和对照组,在有破骨细胞诱导液的培养基中培养1、3和5天时,收集各时间段细胞,通过real time RT-PCR的方法检测S100A4、S100A6及酒石酸抗酸性磷酸酶(TRAP)mRNA等表达情况。结果与未加诱导液组作为对照,MC3T3-E1细胞诱导3、7和14天时,S100A4 mRNA表达逐渐减弱,而诱导21天时S100A4 mRNA表达比14天时要增加;S100A6 mRNA表达在诱导7天和14天时较低,在3天和21天时表达逐渐较高,两者在各时间段比较差异有统计学意义(P<0.05)。与未加诱导液对照,随着RAW264.7细胞逐渐分化,TRAP mRNA表达逐渐增加,S100A4 mRNA表达逐渐增加,S100A6 mRNA表达逐渐下降,差异有统计学意义(P<0.05)。结论S100A4 mRNA参与调控成骨细胞分化过程,而S100A6 mRNA可能参与调控成骨细胞分化过程某个阶段。S100A4和S100A6 mRNA参与骨吸收过程。

Objective To investigate the expression and signficant of S100A4 and S100A6 proteins in the differentiation of osteoblast and osteoclast.Methods MC3T3-E1 cell were divided into osteoblast induced group and control group,cultured in a-MEM with or without osteogenesis induced liquid for 3,7,14,21days.The stain of ALP and alizarin red were detected.Total mRNA were extracted by using improved TRIzol method.We performed real-time RT-PCR on tibia to examine S100A4,S100A6,OPG,RANKL,ALP and RUNX2 mRNA expression.RAW264.7 cells were divided into osteoclast induced group and control group,cultured in a-MEM with osteoclast induced liquid for 1,3,5days.Total mRNA were extracted by using improved TRIzol method.We performed real-time RT-PCR on tibia to examine S100A4,S100A6 and TRAP mRNA expression.Results S100A4 mRNA expressed the lowest on 14 days.Their expression were less and less,while the MC3T3-E1 cells were cultured in normal medium containing osteogenesis induced liquid on 3,7 and 14 days compared with conteol,and the expression of S100A6 were lower on 7 and 14 days.The expression of S100A4 and S100A6 mRNA were statistically significant at each timepoints between osteogenesis induced groups and control groups in the MC3T3 cells(P<0.05).While the RAW264.7 cells were cultured in normal medium containing osteoclast induced liquid on 1,3and 5 days compared with conteol,the expression of S100A4 and TRAP were gradually increased,but S100A6 were gradually decreased.The expression of S100A4 and TRAP were statistically significant on 3 and 5 day(P<0.05),and the expression of S100A6 were statistically significant on 1 and 3 days(P<0.05).Conclusion S100A4 mRNA may participate in regulating osteoblast cell differentiation,but S100A6 mRNA may participate in regulating osteoblast cell differentiation for a certain stage.S100A4 and S100A6 mRNA were possible associated with bone resorption process.