生长停滞特异性蛋白6在人牙周膜细胞迁移及成骨分化中的作用Role of growth arrest-specific protein 6 in migration and osteogenic differentiation of human periodontal ligament cells
张胜男;安娜;欧阳翔英;刘颖君;王雪奎;
摘要(Abstract)：
目的:探讨AXL受体酪氨酸激酶配体——生长停滞特异性蛋白6(growth arrest-specific protein 6,Gas6)在人牙周膜细胞(human periodontal ligament cells,h PDLCs)迁移及成骨诱导液培养下成骨分化中发挥的作用。方法:在对hPDLCs进行体外培养的培养液中加入不同浓度的外源性人重组Gas6(recombinant human Gas6,rhGas6),通过细胞增殖实验(CCK-8)检测rhGas6对h PDLCs细胞增殖的影响,通过细胞划痕实验和细胞迁移实验(Transwell)检测rhGas6对hPDLCs迁移的影响。用小干扰RNA(siRNA)下调hPDLCs中Gas6基因表达,然后进行成骨诱导,利用实时荧光定量聚合酶链式反应(real-time PCR)检测runt相关转录因子2(runt-related transcription factor 2,Runx2)和碱性磷酸酶(alkaline phosphatase,ALP)基因表达变化,用ALP染色检测其对矿化结节形成的影响。结果:不同浓度rhGas6对24、48、72 h的h PDLCs增殖的影响与对照组差异均无统计学意义(P> 0.05)。划痕后24 h,800μg/L rhGas6组愈合面积百分比(31.06%±13.70%)大于对照组(21.79%±9.51%),但差异无统计学意义(P> 0.05);迁移实验中,24 h后800μg/L rhGas6组迁移细胞数显著多于对照组(P <0.01)。加入rhGas6并成骨诱导后,800μg/L组Runx2、ALP基因表达量显著高于对照组(1.60±0.30 vs.0.91±0.10,2.81±0.61 vs.0.86±0.12,P <0.01)。敲低Gas6后,ALP表达显著低于对照组(0.39±0.07 vs.0.92±0.14,P <0.01),Runx2表达无明显变化(P>0.05)。成骨诱导7 d后Gas6敲低组矿化结节形成显著少于对照组(0.25±0.04 vs.1.00±0.11,P <0.001),14 d后Gas6敲低组矿化结节形成少于对照组,但两组间差异无统计学意义(0.86±0.04 vs.1.00±0.16,P> 0.05)。结论:下调Gas6基因后成骨诱导早期的矿化结节形成减少,ALP表达减少,加入rhGas6后Runx2、ALP表达增多,细胞迁移数量增多,提示Gas6在牙周膜细胞迁移及成骨分化中可能存在促进作用。
关键词(KeyWords)： 牙周韧带;生长停滞特异性蛋白6;细胞分化;骨生成;碱性磷酸酶
基金项目(Foundation): 国家自然科学基金(81500859、82071118、81870772、81570986)~~
作者(Author): 张胜男;安娜;欧阳翔英;刘颖君;王雪奎;
Email:
DOI: 10.19723/j.issn.1671-167X.2021.01.003
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