| 丁海,田波,李想,等.负载丹酚酸B的破骨前体细胞膜纳米颗粒对成骨细胞和破骨细胞分化的影响.骨科,2023,14(5): 453-458. |
| 负载丹酚酸B的破骨前体细胞膜纳米颗粒对成骨细胞和破骨细胞分化的影响 |
| Effect of Osteoclast Precursor Membrane Nanoparticles Loaded with Salvianolic Acid B on Osteoblast and Osteoclast Differentiation |
| 投稿时间：2023-04-22 |
| DOI：10.3969/j.issn.1674-8573.2023.05.011 |
| 中文关键词: 骨质疏松 骨稳态 纳米颗粒 细胞膜 成骨细胞 破骨细胞 |
| 英文关键词: Osteoporosis Bone homeostasis Nanoparticles Cell membrane Osteoblasts Osteoclasts |
| 基金项目:国家自然科学基金（82072416）；安徽省教育厅重点项目（KJ2020A0579、2023AH051987）；蚌埠医学院自然科学重点项目（2021byzd106） |
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| 中文摘要: |
| 目的 通过制备破骨前体细胞膜纳米颗粒（nanoparticles，NPs）负载丹酚酸B（salvianolic acid，SalB），构建载药纳米颗粒SalB-NPs，观察其对破骨细胞及成骨细胞分化的影响。方法 采用超声裂解、挤膜的方法制备NPs，并将NPs与SalB共孵育后，使用200 nm聚碳酸酯膜挤出，获得SalB-NPs。在诱导小鼠原代破骨细胞分化和成骨前体细胞（MC3T3-E1）成骨分化的过程中，按照处理方式不同，分为对照组、SalB组、SalB-NPs组。采用透射电镜、纳米粒度及ZETA电位仪和Western Blot对材料进行表征，采用噻唑蓝检测试剂盒检测材料对RAW 264.7和MC3T3-E1细胞活力的影响，采用高效液相色谱法检测SalB的释放率和装载率。采用抗酒石酸酸性磷酸酶（TRAP）染色评价破骨细胞分化能力，通过碱性磷酸酶（ALP）染色和茜素红染色评估成骨分化能力，采用Real time-PCR检测破骨细胞分化及成骨分化相关基因表达水平。结果 制备的纳米颗粒直径在200 nm左右，同时表达RANK蛋白。TRAP染色显示SalB-NPs显著抑制破骨细胞的形成，下调破骨分化相关基因水平，与对照组和SalB组相比，差异有统计学意义（P＜0.05）。使用成骨诱导培养基诱导MC3T3-E1细胞成骨分化，14 d ALP染色和21 d茜素红染色均显示SalB-NPs组ALP活性和钙盐沉积量较SalB组明显增加，差异有统计学意义（P＜0.05）。结论 SalB-NPs体外发挥促进成骨分化、抑制破骨细胞形成双重功效，作为骨质疏松的治疗药物开发，有很好的应用前景，未来仍需在骨质疏松模型动物进一步验证其治疗效果。 |
| 英文摘要: |
| Objective To evaluate the effect of osteoclast precursor cell membrane nanoparticles (NPs) loaded with salvianolic acid B (SalB) on osteoclast differentiation and osteoblast differentiation. Methods The NPs were prepared by sonication and membrane extrusion. After co-incubation with SalB, SalB-NPs were obtained by extrusion through 200 nm polycarbonate membrane. In the process of inducing the differentiation of primary osteoclasts and osteogenic precursor cells (MC3T3-E1), three groups were set up according to different treatment methods: control group, SalB group and SalB-NPs group. The materials were characterized by transmission electron microscopy, nano-particle size and ZETA potentiometer and Western blotting. The viability of RAW 264.7 and MC3T3-E1 cells was detected by thiazole blue detection assay. The SalB release rate and loading rate were detected by high performance liquid chromatography. Tartrate-resistant acid phosphatase (TRAP) staining was used to evaluate osteoclast differentiation. The ability of SalB-NPs to promote osteogenic differentiation was confirmed by alkaline phosphatase (ALP) and alizarin red staining (ARS). In addition, osteoclast and osteoblast differentiation related genes were also detected by Real time-PCR. Results SalB-NPs were about 200 nm in diameter with negative charge. TRAP staining confirmed that SalB-NPs significantly inhibited osteoclast differentiation and down-regulated osteoclast differentiation-related genes with statistically significant differences as compared with SalB (P＜0.05). ALP staining at 14 days and ARS staining at 21 days after osteogenic induction showed that SalB-NPs significantly increased ALP activity and calcium salt deposition in comparison with SalB (P＜0.05). Conclusion SalB-NPs exhibited amazing bilateral effects on promoting bone formation and inhibiting bone resorption both in vitro. SalB-NPs are promising for clinical transformation as anti-osteoporosis drugs in the future and further validation of its therapeutic effect is needed in osteoporosis model animals in the future. |
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