Wu Jianfei, Wang Binglong, Liu Yu, Wei Daixu
Zigong Affiliated Hospital of Southwest Medical University, Zigong Psychiatric Research Center, Zigong Institute of Brain Science, Zigong Sichuan, 643020, P. R. China.
Department of Life Sciences and Medicine, Northwest University, Xi'an Shaanxi, 710069, P. R. China.
Zhongguo Xiu Fu Chong Jian Wai Ke Za Zhi. 2023 Aug 15;37(8):929-936. doi: 10.7507/1002-1892.202303136.
To construct polyhydroxyalkanoate (PHA) microspheres loaded with bone morphogenetic protein 2 (BMP-2) and human β-defensin 3 (HBD3), and evaluate the antibacterial activity of microspheres and the effect of promoting osteogenic differentiation, aiming to provide a new option of material for bone tissue engineering.
The soybean lecithin (SL)-BMP-2 and SL-HBD3 were prepared by SL-mediated introduction of growth factors into polyesters technology, and the functional microsphere (f-PMS) containing BMP-2 and HBD3 were prepared by microfluidic technology, while pure microsphere (p-PMS) was prepared by the same method as the control. The morphology of microspheres was observed by scanning electron microscopy and the water absorption was detected; the release curves of BMP-2 and HBD3 in f-PMS were detected by ELISA kit. The antibacterial effect of microspheres in and was tested with the LIVE/DEAD BacLight bacterial staining kit; the biocompatibility of microspheres was tested using Transwell and cell counting kit 8 (CCK-8). The effect of microspheres on osteogenic differentiation was determined by collagen type Ⅰ (COL-1) immunofluorescence staining and alkaline phosphatase (ALP) concentration.
In this experiment, the f-PMS and p-PMS were successfully constructed. Morphological characteristics showed that p-PMS surface was rough and distributed with micropores of 1-3 μm, while f-PMS surface was smooth and existed white granular material. There was no significant difference in water absorption between the two groups (>0.05). The release curves of BMP-2 and HBD3 in the f-PMS and p-PMS were basically the same, showing both early sudden release and late slow release. The antibacterial activity of f-PMS was significantly higher than that of p-PMS in the test that against and (<0.05), but there was no significant difference in biocompatibility between the two groups (>0.05). The results of osteogenic differentiation of human BMSCs showed that the fluorescence intensity of osteogenic specific protein COL-1 of f-PMS was significantly higher than that in p-PMS, and the activity of ALP in f-PMS was also significantly higher than that in p-PMS (<0.05).
The p-PHA have good antibacterial activity and biocompatibility, and can effectively promote the osteogenic differentiation of human BMSCs, which is expected to be applied to bone tissue engineering in the future.
构建负载骨形态发生蛋白2（BMP-2）和人β-防御素3（HBD3）的聚羟基脂肪酸酯（PHA）微球，评价微球的抗菌活性及促进成骨分化的效果，旨在为骨组织工程提供新的材料选择。
采用大豆卵磷脂（SL）介导生长因子导入聚酯技术制备SL-BMP-2和SL-HBD3，通过微流控技术制备含BMP-2和HBD3的功能微球（f-PMS），同时采用相同方法制备纯微球（p-PMS）作为对照。通过扫描电子显微镜观察微球形态并检测吸水率；采用ELISA试剂盒检测f-PMS中BMP-2和HBD3的释放曲线。使用LIVE/DEAD BacLight细菌染色试剂盒检测微球对金黄色葡萄球菌和大肠杆菌的抗菌效果；采用Transwell和细胞计数试剂盒8（CCK-8）检测微球的生物相容性。通过Ⅰ型胶原（COL-1）免疫荧光染色和碱性磷酸酶（ALP）浓度测定微球对成骨分化的影响。
本实验成功构建了f-PMS和p-PMS。形态学特征显示，p-PMS表面粗糙，分布有1-3μm的微孔，而f-PMS表面光滑，存在白色颗粒状物质。两组吸水率无显著差异（>0.05）。f-PMS和p-PMS中BMP-2和HBD3的释放曲线基本相同，均表现为早期突释和后期缓释。在对金黄色葡萄球菌和大肠杆菌的测试中，f-PMS的抗菌活性显著高于p-PMS（<0.05），但两组生物相容性无显著差异（>0.05）。人骨髓间充质干细胞成骨分化结果显示，f-PMS中成骨特异性蛋白COL-1的荧光强度显著高于p-PMS，f-PMS中ALP活性也显著高于p-PMS（<0.05）。
p-PHA具有良好的抗菌活性和生物相容性，能有效促进人骨髓间充质干细胞的成骨分化，有望在未来应用于骨组织工程。
