运动员骨关节损伤体外细胞成骨分化机制探讨  

Study on the relationship between osteogenic differentiation mechanism of bone and joint injury of athletes in vitro

作　　者：唐雪娇 刘晶莹 闫英梅 TANG Xue-jiao;LIU Jing-ying;YAN Ying-mei(Department of prosthodontics,Tianjin stomatological hospital,School of Medicine,Nankai University,Tianjin,300041,China)

机构地区：[1]天津市口腔医院修复二科/南开大学医学院,天津300041

出　　处：《工业卫生与职业病》2023年第2期151-154,共4页Industrial Health and Occupational Diseases

基　　金：天津市自然科学基金(21JCYBJC01320);天津市口腔医院口腔修复学重点学科建设培育项目(22XFZD7);天津市卫生健康科技青年项目(TJWJ2021QN069)。

摘　　要：目的通过体外实验探讨P物质受体NK1(NK1-R)在骨髓间充质干细胞(大鼠骨髓基质干细胞,ST2)中的表达及其与成骨分化标记物的关系。方法将骨髓间充质干细胞(ST2)细胞株接种于培养皿传代培养,培养基为高糖DMEM培养基(含10%FBS),放置于5%CO_(2)细胞培养箱中37℃孵育72 h传代,至第三四代时使用。实验组加入含10-6 mol/L浓度的P物质的成骨诱导培养液,对照组为不含P物质的成骨诱导培养液,培养7天,分别在第1,3,5,7天收获细胞进行NK1受体的免疫细胞化学染色,分析其阳性表达率。同一批细胞在1,3,5,7天收获细胞培养液,用ELISA检测ALP、OCN(骨钙素),分析其表达量的变化。结果细胞免疫化学显示第1天时NK1受体即开始表达,第3,5天表达上升,1周时NK1受体明显表达,其存在于细胞膜和细胞质,主要存在于细胞质中,成骨分化标记物中成骨分化早期标记物ALP第1,3,5,7天表达量为1.695 ng/ml,5.013 ng/ml,7.94 ng/ml和9.932 ng/ml,随着时间推移表达量逐渐增加,成骨分化晚期标记物OCN在1,3,5天表达量很少,分别为1.497 ng/ml,2.663 ng/ml,5.633 ng/ml,第7天开始有明显增加表达,为18.53 ng/ml。结论NK1受体在骨髓基质干细胞成骨分化的早期即开始出现,并且与促进ST2成骨分化成熟密切相关。Objective To investigate the expression of substance P receptor NK1 in ST2 cells and its relationship with osteogenic differentiation markers.Methods ST2 cells were cultured in high glucose DMEM medium(10%FBS),and placed in 5%CO2 incubator at 37℃for 72 h,passaged to 3 or 4 generations when used.In the experimental group,osteogenic induction medium containing substance P was added,and osteoblast inducing medium without substance P was used in the control group for 7 days.Cells were harvested on day 1,3,5 and 7.Expression of NK1 receptor was detected by using immunocytochemical staining.Simultaneously,the expression of ALP,OCN(osteocalcin)in cell culture medium was detected by using ELISA on 1,3,5,7 days.Results Cell immunochemistry showed that NK1 receptor began to express on day 1,and expression was increased at day 3 and day 5,greatly at day 7.The NK1 receptor was expressed in the cell membrane and cytoplasm,mainly in the cytoplasm.The expression of ALP which is an early osteogenic differentiation marker was 1.695 ng/ml,5.013 ng/ml,7.94 ng/ml and 9.932 ng/ml at day 1,3,5,7,and the expression increased gradually with time.The expression of OCN was less expressed at 1,3 and 5 days,which was 1.497 ng/ml,2.663 ng/ml,5.633 ng/ml,but began to significantly increase at the 7th day,which was 18.53 ng/ml.Conclusions NK1 receptors begin to appear early in osteogenesis of ST2 cells and are closely related to the promotion of osteogenesis and differentiation of ST2.

关 键 词：P物质 NK1受体 成骨分化 

分 类 号：R135[医药卫生—劳动卫生]