4-辛基衣康酸靶向脂质体的微流控制备及修复牙周炎骨缺损的研究

doi:10.12439/kqhm.1005-4979.2026.05.004

摘要/Abstract

摘要：

目的：

采用微流控技术构建叶酸（folic acid，FA）修饰的4-辛基衣康酸（4-octyl itaconate，4-OI）负载脂质体（liposomes，LPs），评价其理化特性及其对牙周炎性骨缺损的抗炎和促修复作用。

方法：

应用微流控技术构建FA修饰、负载4-OI的靶向LPs（4-OI/FA-LPs），利用透射电子显微镜（transmission electron microscope，TEM）、颗粒粒度和Zeta电位分析对其进行表征；通过超滤离心法和平衡透析法分析其载药量（drug loading，DL）、包封率（encapsulation efficiency，EE）及体外释药行为；采用CCK-8法检测4-OI/FA-LPs的生物相容性。以脂多糖（lipopolysaccharide，LPS）诱导RAW264.7巨噬细胞炎症模型，应用实时定量聚合酶链反应（real-time quantitative polymerase chain reaction，RT-qPCR）、免疫荧光法检测4-OI/FA-LPs的炎症调控作用。建立丝线结扎诱导的小鼠牙周炎模型，通过Micro-CT和苏木精-伊红（hematoxylin and eosin，HE）染色分析4-OI/FA-LPs对牙槽骨吸收和牙周组织损伤的影响。

结果：

4-OI/FA-LPs呈规则球形结构，分散均匀，粒径为（138.80±0.60） nm，带负电荷。其EE为81.94%，DL为4.49%，并可持续释放4-OI达2周。CCK-8结果显示，4-OI/FA-LPs具有良好的生物相容性。RT-qPCR结果显示，与LPS组相比，4-OI/FA-LPs可下调IL-6、IL-1β和MMP13 mRNA表达，并上调Arg-1 mRNA表达；免疫荧光结果显示，iNOS表达降低，Arg-1表达升高。Micro-CT及HE染色结果显示，4-OI/FA-LPs可减轻牙周炎小鼠牙槽骨吸收及牙周组织损伤，提高骨密度（bone mineral density，BMD）和骨体积分数（bone volume fraction，BV/TV）。

结论：

本研究成功构建4-OI/FA-LPs，其具有良好的理化性能和生物相容性，能够减轻炎症反应并改善牙周炎相关牙槽骨吸收，对牙周炎性骨缺损具有一定的修复促进作用。

关键词: 微流控, 靶向脂质体, 4-辛基衣康酸, 牙周炎

Abstract:

Objective:

To construct folic acid (FA)-modified liposomes (LPs) loaded with 4-octyl itaconate (4-OI) using microfluidic technology and to evaluate their physicochemical properties, anti-inflammatory effects, and reparative potential in periodontal inflammatory bone defects.

Methods:

FA-modified, 4-OI-loaded targeted liposomes (4-OI/FA-LPs) were prepared using a microfluidic approach and characterized by transmission electron microscopy (TEM), particle size analysis, and zeta potential measurement. Encapsulation efficiency (EE), drug loading (DL), and in vitro drug-release behavior were determined by ultrafiltration centrifugation and equilibrium dialysis. The biocompatibility of 4-OI/FA-LPs was evaluated using the CCK-8 assay. Lipopolysaccharide (LPS)-stimulated RAW264.7 macrophages were used as an in vitro inflammatory model, and the inflammation-regulating effects of 4-OI/FA-LPs were assessed by real-time quantitative polymerase chain reaction (RT-qPCR) and immunofluorescence staining. A ligature-induced mouse model of periodontitis was established, and the effects of 4-OI/FA-LPs on alveolar bone resorption and periodontal tissue damage were evaluated by Micro-CT and hematoxylin and eosin (HE) staining.

Results:

The 4-OI/FA-LPs exhibited a regular spherical morphology with uniform distribution, an average particle size of 138.80±0.60 nm, and a negative surface charge. The EE and DL capacity were 81.94% and 4.49%, respectively, and sustained release of 4-OI was observed for up to 2 weeks. The CCK-8 assay demonstrated good biocompatibility of 4-OI/FA-LPs. RT-qPCR showed that, compared with the LPS group, 4-OI/FA-LPs significantly downregulated the mRNA expression of IL-6, IL-1β, and MMP13 while upregulating Arg-1 mRNA expression. Immunofluorescence analysis showed decreased iNOS expression and increased Arg-1 expression. Micro-CT and HE staining revealed that 4-OI/FA-LPs alleviated alveolar bone resorption and periodontal tissue damage in mice with periodontitis and improved bone mineral density (BMD) and bone volume fraction (BV/TV).

Conclusion:

The 4-OI/FA-LPs were successfully constructed and exhibited favorable physicochemical properties and biocompatibility. The LPs attenuated inflammatory responses and alleviated periodontitis-related alveolar bone resorption, suggesting potential for promoting the repair of periodontal inflammatory bone defects.

Key words: microfluidics, targeted liposomes, 4-octyl itaconate, periodontitis

中图分类号:

R781.4

罗茜, 解健, 苏俭生. 4-辛基衣康酸靶向脂质体的微流控制备及修复牙周炎骨缺损的研究[J]. 《口腔颌面外科杂志》, 2026, 36(5): 384-392.

LUO Xi, XIE Jian, SU Jiansheng. Microfluidic preparation of 4-octyl itaconate-loaded targeted liposomes and repair of bone defects in periodontitis[J]. Journal of Oral and Maxillofacial Surgery(CN), 2026, 36(5): 384-392.

https://journal06.magtech.org.cn/Jweb_joms/CN/Y2026/V36/I5/384

图/表 7

表1 基因引物序列

Table 1 The gene primer sequences

基因	上游引物序列（5'-3'）	下游引物序列（5'-3'）
β-actin	GTCCCTCACCCTCCCAAAAG	GCTGCCTCAACACCTCAACCC
Arg-1	CTCCAAGCCAAAGTCCTTAGAG	AGGAGCTGTCATTAGGGACATC
Mmp13	CTTCTTCTTGTTGAGCTGGACTC	CTGTGGAGGTCACTGTAGACT
IL-6	TGCCTTCTTGGGACTGAT	CTGGCTTTGTCTTTCTTGTT
IL-1β	AGCTTCAGGCAGGCAGTATCAC	CCAGCAGGTTATCATCATCATCC

图1 FA-LPs及4-OI/FA-LPs的纳米表征 A. FA-LPs TEM图（×120 000）；B. 4-OI/FA-LPs TEM图（×120 000）；C. FA-LPs的粒径分布图；D. 4-OI/FA-LPs的粒径分布图；E. LPs Zeta电位图。

Figure 1 Nano-characterization of FA-LPs and 4-OI/FA-LPs

图2 药物体外释放曲线 A. 4-OI/FA-LPs体外释放曲线；B. 4-OI体外释放曲线。

Figure 2 Drug release profile in vitro

表2 释药模型拟合结果

Table 2 Fitting release model and coefficient

模型	方程	R²
零级动力学	Q=4.795 t+15.603	0.906
一级动力学	Q=77.571×（1-e^-0.455t）	0.990
Higuchi	Q=19.61 t^1/2 +7.12	0.968

图3 不同LPs处理下RAW264.7细胞活性 **表示P<0.01，与Control组比较。

Figure 3 RAW264.7 cell activity under different LPs treatments

图4 LPs对炎症的调控能力 A. RT-qPCR检测结果；B.免疫荧光检测结果（×20）；C.免疫荧光相对强度定量分析，*表示P<0.05，***表示P<0.001，与LPS组比较。

Figure 4 Effects of LPs on inflammatory regulation

图5 4-OI/FA-LPs对牙周炎模型小鼠牙槽骨吸收的治疗效果 A.各组Micro-CT三维重建影像（×10），橙色线条示意CEJ-ABC；B.各组HE染色结果（×10）；C—E.依次为CEJ-ABC、BMD和BV/TV的定量分析结果；**表示P<0.01，***表示P<0.001，与Control组比较；##表示P<0.01，###表示P<0.001，与PBS组比较；△△表示P<0.01，与FA-LPs组比较。

Figure 5 Effect of 4-OI/FA-LPs on alveolar bone resorption in periodontitis model of mice

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