载氨基胍的铜介孔二氧化硅对MGO刺激下大鼠BMSCs成骨分化的影响

doi:10.12439/kqhm.1005-4979.2026.04.004

摘要/Abstract

摘要：

目的：

制备载氨基胍（aminoguanidine，AG）的铜复合介孔二氧化硅纳米颗粒（copper-composite mesoporous silica nanoparticles，Cu-MSNs），探究其对甲基乙二醛（methylglyoxal，MGO）刺激下大鼠颌骨骨髓间充质基质细胞（maxillary bone marrow stromal cells，M-BMSCs）成骨分化的影响。

方法：

通过溶胶-凝胶法制备Cu-MSNs，并将Cu-MSNs分散至含AG的去离子水中制备载氨基胍的铜复合介孔二氧化硅纳米颗粒（AG@Cu-MSNs），通过扫描电子显微镜、透射电子显微镜等技术对复合纳米颗粒进行表征；培养大鼠M-BMSCs，使用CCK-8试剂盒确定MGO对M-BMSCs的刺激浓度和复合纳米颗粒与M-BMSCs的共培养浓度；使用活/死细胞染色实验评估MGO刺激下复合纳米颗粒对细胞活力的影响；矿化诱导7 d后使用碱性磷酸酶（alkaline phosphatase，ALP）染色法评估ALP强度，应用实时定量聚合酶链反应（real-time quantitative polymerase chain reaction，RT-qPCR）检测部分成骨相关基因的表达情况，使用蛋白质印迹法检测部分成骨相关蛋白的表达情况。

结果：

应用CCK-8实验确定MGO的应用浓度为0.5 mmol/L。CCK-8实验及活/死细胞染色结果显示，复合纳米颗粒具有良好的生物相容性，其中，AG@Cu-MSNs组细胞受MGO影响最低，细胞增殖最活跃。矿化诱导7 d后，与MGO组相比，AG@Cu-MSNs组细胞ALP染色最明显，成骨相关基因（Alpl、Runx2、Osx）和相关蛋白（RUNX2、OSX）的表达均上调。

结论：

在MGO刺激环境中，AG@Cu-MSNs能够促进M-BMSCs的增殖及成骨分化。

关键词: 甲基乙二醛, 纳米颗粒, 氨基胍, 铜离子, 成骨分化

Abstract:

Objective:

To synthesize copper-composite mesoporous silica nanoparticles loaded with aminoguanidine (AG@Cu-MSNs) and investigate their effects on the osteogenic differentiation of methylglyoxal (MGO)-stimulated rat maxillary bone marrow stromal cells (M-BMSCs).

Methods:

Copper-composite mesoporous silica nanoparticles (Cu-MSNs) were prepared by the sol-gel method, and AG@Cu-MSNs were prepared by dispersing Cu-MSNs in deionized water containing aminoguanidine. The nanoparticles were characterized by scanning electron microscopy and transmission electron microscopy, and other techniques. M-BMSCs were cultured. The CCK-8 assay kit was used to determine the optimal stimulatory concentration of MGO on M-BMSCs and the co-culture concentration of composite nanoparticles with M-BMSCs. The live/dead staining assay was employed to assess the impact of composite nanoparticles on cell viability in MGO-stimulated conditions. After 7 days of mineralization induction, alkaline phosphatase (ALP) activity was assessed using an ALP assay kit. Real-time quantitative polymerase chain reaction (RT-qPCR) was used to detect the expression of some osteogenic related gene. Western blotting was used to detect the expression of osteogenesis-related proteins.

Results:

The application concentration of MGO was determined to be 0.5 mmol/L by CCK-8 assay. The results of CCK-8 assay and live/dead staining assay showed that the composite nanoparticles had good biocompatibility. Among them, the AG@Cu-MSNs group exhibited the highest cell viability and proliferation in MGO-stimulated conditions. After 7 days of mineralization induction, compared with the MGO stimulation group, the AG@Cu-MSNs group exhibited the most pronounced ALP staining intensity. Additionally, the expression levels of osteogenesis-related genes (Alpl, Runx2, Osx) and corresponding proteins (RUNX2, OSX) were significantly upregulated.

Conclusion:

In MGO-stimulated conditions, AG@Cu-MSNs significantly promotes the proliferation and osteogenic differentiation of M-BMSCs.

Key words: methylglyoxal, nanoparticles, aminoguanidine, copper ions, osteogenic differentiation

保静, 胡丝桐, 苏俭生. 载氨基胍的铜介孔二氧化硅对MGO刺激下大鼠BMSCs成骨分化的影响[J]. 《口腔颌面外科杂志》, 2026, 36(4): 290-298.

BAO Jing, HU Sitong, SU Jiansheng. Effect of copper-composite mesoporous silica nanoparticles loaded with aminoguanidine on osteogenic differentiation of MGO-stimulated rat BMSCs[J]. Journal of Oral and Maxillofacial Surgery(CN), 2026, 36(4): 290-298.

https://journal06.magtech.org.cn/Jweb_joms/CN/Y2026/V36/I4/290

图/表 8

表1 引物序列表

Table 1 Primer sequences

基因	正向引物（5'-3'）	反向引物（5'-3'）
GAPDH	TCTCTGCTCCTCCCTGTTCT	TACGGCCAAATCCGTTCACA
Alpl	CCAGAGAAAGAGAGAGACCCC	GAGACGCCCATACCATCTCC
Runx2	ATGATGACACTGCCACCTCTGAC	AACTGCCTGGGGTCTGAAAAAGG
Osx	GCCTACTTACCCGTCTGACTTTG	TGCCCACTATTGCCAACTGC

图1 Cu-MSNs和AG@Cu-MSNs的表征 A. Cu-MSNs的扫描电镜图（左图为放大100 000倍，右图为放大200 000倍）；B. Cu-MSNs的透射电镜图（左图为放大60 000倍，右图为放大70 000倍）；C. Cu-MSNs的能谱仪结果；D. AG@Cu-MSNs、AG、Cu-MSNs的傅里叶变换红外光谱仪结果；E. AG@Cu-MSNs中AG的累积释放曲线。

Figure 1 Characterization of Cu-MSNs and AG@Cu-MSNs

图2 不同浓度MGO对M-BMSCs细胞活力的影响（n=3） A、B.分别为培养1、3 d时的CCK-8结果。***表示P<0.001，与0 mmol/L组比较。

Figure 2 Effects of different concentrations of MGO on the viability of M-BMSCs（n=3）

图3 不同浓度纳米颗粒对M-BMSCs细胞活力的影响（n=3） A. MSNs CCK-8实验结果；B. Cu-MSNs CCK-8实验结果；C. AG@MSNs CCK-8实验结果；D. AG@Cu-MSNs CCK-8实验结果。*表示P<0.05，**表示P<0.01，***表示P<0.001，与对照组比较。

Figure 3 Effects of different concentrations of nanoparticles on the viability of M-BMSCs （n=3）

图4 AG@Cu-MSNs对MGO刺激下M-BMSCs细胞活力的影响 A.各组M-BMSCs共培养的活/死细胞染色照片（×10）；B.活/死细胞染色半定量数据。n=3，***表示P<0.001，与对照组比较；###表示P<0.001，与MGO组比较。

Figure 4 Effects of AG@Cu-MSNs on the viability of MGO-stimulated M-BMSCs

图5 ALP染色结果 A.各组M-BMSCs ALP染色结果（×20）；B. ALP染色面积半定量数据。n=3，***表示P<0.001，与对照组比较；###表示P<0.001，与MGO组比较。

Figure 5 Results of ALP staining

图6 各组成骨相关基因mRNA的表达（n=3） A—C.分别为各组Alpl、Runx2、Osx mRNA相对表达量比较。*表示P<0.05，**表示P<0.01，与对照组比较；##表示P<0.01，###表示P<0.001，与MGO组比较。

Figure 6 Relative mRNA expression levels of osteogenesis-related genes in each group （n=3）

图7 M-BMSCs成骨相关蛋白的表达

Figure 7 Expression of osteogenesis-related proteins of M-BMSCs

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