北京大学学报(医学版) ›› 2025, Vol. 57 ›› Issue (1): 7-12. doi: 10.19723/j.issn.1671-167X.2025.01.002
帅婷1, 郭艳艳1, 林春平2, 侯晓玫1,*(), 金婵媛1,*(
)
Ting SHUAI1, Yanyan GUO1, Chunping LIN2, Xiaomei HOU1,*(), Chanyuan JIN1,*(
)
摘要:
目的: 初步探究神经元正五聚体1(neuronal pentraxin 1,NPTX1)基因对人骨髓间充质干细胞(human bone marrow mesenchymal stem cells,hBMSCs)成骨分化调控的作用机制。方法: 将hBMSCs进行成骨诱导培养,在不同的时间点(0、3、7、10、14 d)收集RNA,通过实时荧光定量聚合酶链式反应(quantitative real-time polymerase chain reaction,qPCR)技术检测成骨分化调控相关的Runt相关转录因子2(runt-related transcription factor 2,RUNX2)、碱性磷酸酶(alkaline phosphatase,ALP)、骨钙素(osteocalcin,OCN)以及NPTX1的mRNA表达水平。通过构建NPTX1 shRNA慢病毒并感染hBMSCs,建立NPTX1稳定敲减的hBMSCs细胞系;采用ALP染色、茜素红(alizarin red,AR)染色和qPCR等方法检测敲减NPTX1对hBMSCs成骨分化能力的影响。结果: 体外诱导hBMSCs成骨分化时,随着成骨诱导时间的延长,与第0天相比,成骨基因RUNX2、ALP和OCN表达量均显著升高,而NPTX1表达量明显降低(P < 0.01)。慢病毒感染hBMSCs 72 h后,qPCR结果显示NPTX1敲减效率高于60%。hBMSCs敲减NPTX1后,将敲减NPTX1组(sh NPTX1组)及其对照组(shNC组)在普通增殖培养基下进行培养后提取RNA,qPCR结果显示和shNC组相比,sh NPTX1组的成骨相关基因RUNX2和成骨细胞特异性转录因子(osterix,OSX)表达量显著增高(P < 0.01)。ALP染色试验显示成骨诱导7 d时,sh NPTX1组较shNC组显色明显加深;AR染色试验显示成骨诱导14 d时,sh NPTX1组较shNC组矿化结节明显增多。结论: NPTX1对hBMSCs成骨分化具有调控作用,且其敲减可促进hBMSCs的成骨分化,该结果提示NPTX1可能成为治疗骨质疏松症等成骨异常疾病的潜在靶点。
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