Page 36 - 南京医科大学学报自然科学版

南京医科大学学报（自然科学版） 第43卷第3期
·326 · Journal of Nanjing Medical University（Natural Sciences） 2023年3月

·基础研究·

负载γ⁃Fe2O3的壳聚糖多孔海绵对大鼠骨髓间质干细胞增殖和

成骨分化的影响

孙上雯 ，陈汉帮 ，胡姝颖 ，章非敏 1，2*
1，2
1，2
1，2
南京医科大学口腔疾病研究江苏省重点实验室，南京医科大学附属口腔医院修复科，江苏 南京 210029
1 2

［摘 要］ 目的：研究负载γ⁃Fe2O3的壳聚糖多孔海绵对大鼠骨髓间充质干细胞（rat bone marrow mesenchymal stem cell，rBMSC）增
殖和成骨分化的影响。方法：使用冻干⁃交联法制备负载γ⁃Fe2O3浓度分别为1%、5%、10%和20%的壳聚糖海绵，并制备空白对
照组。将rBMSC培养于海绵上，通过扫描电镜和共聚焦显微镜观察细胞黏附及增殖情况，通过CCK⁃8法检测细胞第1、3、5、7天的
增殖情况，通过碱性磷酸酶（alkaline phosphatases，ALP）染色及活性检测和荧光定量PCR检测第7、14天ALP活性和成骨指标

ALP、骨形态发生蛋白（bone morphogenetic protein 2，Bmp2）、胶原蛋白（collagen Ⅰ，Col1）和 Runt 相关转录因子 2（core binding
factor alphal 1，Runx2）的表达；使用茜素红定量法评估第21、28天细胞外基质矿化情况。结果：CCK⁃8结果显示rBMSC均能在
材料上持续增殖，添加γ⁃Fe2O3对rBMSC增殖有促进作用；ALP染色及活性检测结果和PCR结果显示添加γ⁃Fe2O3能提高ALP活
性并促进成骨指标表达；茜素红定量结果显示添加浓度为 5%和 10%时，矿化物形成量高于对照组（P < 0.05）。结论：负载γ⁃
Fe2O3的壳聚糖海绵能够促进 rBMSC 的增殖和早期成骨分化，浓度为 5%和 10%时对 rBMSC 成骨分化晚期矿化物形成有促
进作用。
［关键词］ γ⁃Fe2O3磁性纳米颗粒；壳聚糖多孔海绵；骨组织工程；成骨分化
［中图分类号］ R336 ［文献标志码］ A ［文章编号］ 1007⁃4368（2023）03⁃326⁃08
doi：10.7655/NYDXBNS20230305

Effects of γ ⁃ Fe2O3 ⁃ loaded chitosan porous sponge on proliferation and osteogenic

differentiation of rat bone marrow mesenchymal stem cells
1，2
1，2
1，2
SUN Shangwen ，CHEN Hanbang ，HU Shuying ，ZHANG Feimin 1，2*
1 Jiangsu Province Key Laboratory of Oral Diseases，Nanjing Medical University；Department of Prosthodontics，the
2
Affiliated Stomatological Hospital of Nanjing Medical University，Nanjing 210029，china
［Abstract］ Objective： This study aims to observe the effects of γ⁃Fe2O3⁃loaded chitosan porous sponge on the proliferation and early
osteogenic differentiation of rat bone marrow mesenchymal stem cells（rBMSC）. Methods：Chitosan sponges loaded with γ⁃Fe2O3 at
concentrations of 1%，5%，10% and 20% were prepared by freeze⁃drying and cross⁃linking，and blank control group was prepared.
Scanning electron microscopy and confocal microscopy were used to evaluate the adhesion and proliferation of rBMSC on the sponges.
Cell proliferation at 1，3，5 and 7 days was detected by CCK⁃8. The early osteogenic differentiation of rBMSC at 7 and 14 days was
evaluated by alkaline phosphatases（ALP）staining and activity detection. Real ⁃ time fluorescence quantitative reverse transcription
PCR was used to detect the expression of ALP，bone morphogenetic protein 2（Bmp2），collagen I（Col1）and Runt⁃related transcription
factor 2（Runx2）after 7 and 14 days of osteogenic induction. The mineralization of extracellular matrix at 21 and 28 days was assessed
by alizarin red staining quantitative method. Results：CCK⁃8 results showed each group added with γ⁃Fe2O3 promoted the proliferation
of rBMSC. ALP staining and activity detection results showed the addition of γ⁃Fe2O3 can improve the activity of ALP. The results of
real ⁃ time fluorescence quantitative reverse transcription PCR showed that the addition of γ ⁃ Fe2O3 can promote the expression of
osteogenic indexes（ALP，Bmp2，Col1 and Runx2）. The quantitative detection results of alizarin red staining showed that the amount of
mineralization in the groups with γ⁃Fe2O3 added concentration of 5% and 10% was higher than that in the control group（P < 0.05）.

［基金项目］ 国家自然科学基金（81870807）；江苏省高校优势学科建设工程资助项目（2018⁃87）
∗
通信作者（Corresponding author），E⁃mail： fmzhang@ njmu.edu.cn