搭载依达拉奉的活性氧响应型水凝胶的构建及其对成骨细胞的影响和抑菌性的研究

doi:10.12439/kqhm.1005-4979.2026.03.004

摘要/Abstract

摘要：

目的：

构建搭载依达拉奉（Edaravone）的活性氧（reactive oxygen species，ROS）响应型水凝胶，并评价其理化性能、促成骨作用及抑菌性能。

方法：

使用冻融循环法制备复合水凝胶，通过CCK-8法筛选Edaravone的工作浓度，通过测量孔隙率、力学性能、溶胀率与降解率优化各组分配比，筛选出最佳配比的复合水凝胶，并评价药物释放行为及ROS清除能力。将水凝胶与金黄色葡萄球菌（Staphylococcus aureus）共培养，通过菌落计数法检测其抑菌性能；将水凝胶与小鼠胚胎成骨细胞前体细胞（mouse embryo osteoblast precursor cells，MC3T3-E1 cells）共培养后，采用碱性磷酸酶（alkaline phosphatase，ALP）染色、茜素红S（alizarin red S，ARS）染色、实时定量聚合酶链反应（real-time quantitative polymerase chain reaction，RT-qPCR）检测其对MC3T3-E1细胞的生长、矿化及成骨相关基因表达的影响。

结果：

成功制备了具有三维多孔网状结构的复合水凝胶(P3ATE)，其力学性能、溶胀性能及降解率良好，可持续释放Edaravone，并具有较强的ROS清除能力（P<0.05）及良好的抑菌活性，可显著抑制金黄色葡萄球菌生长（P<0.05）；与对照组比较，P3ATE水凝胶可显著促进MC3T3-E1细胞增殖与矿化，提高ALP活性，上调Bmp2、Spp1、Runx2及Col1a1的表达（P<0.05）。

结论：

所构建的Edaravone搭载ROS响应型复合水凝胶具有良好的促成骨、抗氧化及抑菌性能，在颌骨缺损修复中具有潜在应用价值。

关键词: 颌骨缺损, 水凝胶, 活性氧, 聚乙烯醇, 依达拉奉

Abstract:

Objective:

To construct an Edaravone-loaded reactive oxygen species (ROS)-responsive hydrogel and to evaluate its physicochemical properties, osteogenic effects and antibacterial properties.

Methods:

The composite hydrogel was prepared using the freeze-thaw cycle method. The working concentration of Edaravone was screened using the CCK-8 assay. The composition ratios were then optimized through porosity, mechanical property, swelling rate and degradation rate tests to identify the optimal composite hydrogel, and the drug release behavior and ROS scavenging ability were evaluated. The hydrogel was co-cultured with Staphylococcus aureus, and its antibacterial activity was assessed by the colony counting method. After co-culturing the hydrogel with mouse embryo osteoblast precursor cells (MC3T3-E1 cells), alkaline phosphatase (ALP) staining, alizarin red S (ARS) staining and real-time quantitative polymerase chain reaction (RT-qPCR) were used to evaluate the effects of the hydrogel on the proliferation, mineralization and osteogenesis-related gene expression of MC3T3-E1 cells.

Results:

A composite hydrogel (P3ATE) with a three-dimensional porous network structure was successfully prepared. It exhibited favorable mechanical, swelling and degradation properties, and provided sustained release of Edaravone. It demonstrated strong ROS scavenging ability (P<0.05) and good antibacterial activity, significantly inhibiting the growth of Staphylococcus aureus (P<0.05). Compared with the control group, the P3ATE hydrogel significantly promoted the proliferation and mineralization of MC3T3-E1 cells, enhanced ALP activity, and upregulated the expression of Bmp2, Spp1, Runx2 and Col1a1 (P<0.05).

Conclusion:

The constructed Edaravone-loaded ROS-responsive composite hydrogel exhibits favorable osteogenic, antioxidant and antibacterial properties, suggesting potential application value in the repair of jaw defects.

Key words: jaw defects, hydrogel, reactive oxygen species, polyvinyl alcohol, Edaravone

王思丹, 刘雨璐, 沈程奕, 关键. 搭载依达拉奉的活性氧响应型水凝胶的构建及其对成骨细胞的影响和抑菌性的研究[J]. 《口腔颌面外科杂志》, 2026, 36(3): 196-205.

WANG Sidan, LIU Yulu, SHEN Chengyi, GUAN Jian. Construction of an Edaravone-loaded reactive oxygen species-responsive hydrogel and its effects on osteoblasts and antibacterial activity[J]. Journal of Oral and Maxillofacial Surgery(CN), 2026, 36(3): 196-205.

https://journal06.magtech.org.cn/Jweb_joms/CN/Y2026/V36/I3/196

图/表 11

表1 RT-qPCR引物序列

Table 1 Sequences of the primers used in RT-qPCR

基因	正向引物（5'-3'）	反向引物（5'-3'）
Col1a1	ACAGGCGAACAAGGTGACAGAG	AGGAGAACCAGGAGAACCAGGAG
Runx2	CCAGGCAGGCACAGTCTTCC	CGTGGTGGAGTGGATGGATGG
Spp1	GATGACGATGATGATGACGATGGAG	TTGTGTACTAGCAGTGACGGTCTC
Bmp2	AGCCAAACACAAACAGCGGAAG	AGGTGCCACGATCCAGTCATTC

图1 Edaravone对MC3T3-E1细胞活力的影响 *表示P<0.05，**表示 P<0.01，***表示P<0.001，与0 μmol/L组比较。

Figure 1 Effect of Edaravone on the viability of MC3T3-E1 cells

图2 各组复合水凝胶的形貌及孔隙率表征 A—C.从左至右依次为P组、PAT组、P3ATE组水凝胶外观（×1）；D—F.从左至右依次为P组、PAT组、P3ATE组水凝胶的FE-SEM图像（×1000）；G.各组水凝胶孔隙率。***表示P<0.001，与P组比较。

Figure 2 Morphological characterization and porosity analysis of the composite hydrogels in each group

图3 各组复合水凝胶的力学性能 A—C.从上至下分别为P3ATE组水凝胶在初始、压缩、复原时的效果图；D.各组水凝胶应力-应变曲线图。

Figure 3 Mechanical properties of the composite hydrogels in each group

图4 各组复合水凝胶的理化性能 A.初始含水率；B.溶胀率；C.剩余质量。***表示P<0.001，与P组比较。

Figure 4 Physicochemical properties of the composite hydrogels in each group

图5 P3ATE复合水凝胶中Edaravone的体外释放曲线

Figure 5 In vitro release profile of Edaravone from the P3ATE composite hydrogel

图6 P3ATE复合水凝胶的ROS清除能力 A. 各组DPPH自由基清除率；B. P3ATE水凝胶抗氧化效果（×50）；C.各组抗氧化染色平均荧光强度。*表示P<0.05，***表示P<0.001，与对照组比较。

Figure 6 ROS scavenging ability of the P3ATE composite hydrogel

图7 P3ATE复合水凝胶的抗菌性能 A.对照组与各组水凝胶与菌液共培养后的吸光度值比较；B.各组水凝胶抑菌率；C.对照组与各组水凝胶与菌液共培养后的菌落图。*表示P<0.05，**表示P<0.01，与对照组比较。

Figure 7 Antibacterial activity of the P3ATE composite hydrogel

图8 P3ATE复合水凝胶的生物相容性评价 A. P组、P3ATE组、阴性对照组和阳性对照组的溶血率；B.活/死细胞双染法测定PAT组、Edaravone组及P3ATE组细胞活性（×50）；C.接受不同处理的MC3T3-E1细胞染色后的荧光强度定量分析。*表示P<0.05，***表示P<0.001，与P3ATE组比较。

Figure 8 Biocompatibility evaluation of the P3ATE composite hydrogel

图9 P3ATE复合水凝胶促进MC3T3-E1细胞成骨分化的体外评价 A—B.分别为诱导培养7 d和14 d后各组MC3T3-E1细胞ALP活性定量结果；C—F.分别为对照组、Edaravone组、PAT组及P3ATE组处理14 d后，MC3T3-E1细胞的ARS染色图像（×20）；G.各组ARS染色定量结果。***表示P<0.001，与对照组比较。

Figure 9 In vitro evaluation of osteogenic differentiation of MC3T3-E1 cells induced by the P3ATE composite hydrogel

图10 各组成骨相关基因mRNA表达分析 A—D.各组细胞Bmp2、Col1a1、Runx2、Spp1的mRNA相对表达水平。*表示P<0.05，**表示P<0.01，***表示P<0.001，与P3ATE组比较。

Figure 10 mRNA expression analysis of osteogenesis-related genes in each group

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