负载淫羊藿苷的壳聚糖基仿生支架的促软骨形成和炎症缓解作用

Chondrogenic and ameliorated inflammatory effects of chitosan-based biomimetic scaffold loaded with icariin

摘要:

淫羊藿苷（icariin，ICA）是具有促软骨修复和抗炎作用的小分子药物，将ICA负载到软骨组织工程支架将能增强支架的生物功能性。本研究将聚l-丙交酯-己内酯（poly （L-lactide-co-caprolactone，PLCL）电纺纤维经聚多巴胺（polydopamine，PDA）涂覆处理后制备成的短纤维与柠檬酸/壳聚糖溶液（citric acid/chitosan solution，CC）混合，通过冻-融和冷冻干燥方法制备成短纤维增强的壳聚糖水凝胶（PDA@PLCL/CC）；然后通过物理吸附方法负载ICA构建新型工程软骨仿生支架（ICA-PDA@PLCL/CC）；最后通过体外兔软骨细胞培养试验，研究ICA介导的软骨仿生支架对软骨细胞功能及炎症反应的影响。结果表明，PDA@PLCL/CC具有适当的孔径和孔隙率（>80%），具有较高的吸水率和力学性能，能够促进软骨细胞增殖和黏附。ICA-PDA@PLCL/CC支架利于维持软骨细胞形态，且能促进软骨生成性基因（sox-9）、糖胺聚糖（gag）和Ⅱ型胶原（colⅡ）基因的表达和软骨基质的合成。在模拟炎症存在的情形下，ICA-PDA@PLCL/CC仿生支架可降低软骨细胞纤维化，有效下调软骨细胞促炎症因子白介素-6（il-6）、白介素-1β（il-1β）和诱导型一氧化氮合成酶（inos）的表达，减少基质金属蛋白酶-3（mmp-3）的表达，提高软骨胞外基质糖胺聚糖（glycosaminoglycan，GAG）和Ⅱ型胶原（type Ⅱ collagen，Col Ⅱ）的合成。该研究发展的ICA-PDA@PLCL/CC工程软骨仿生支架在软骨损伤的再生修复中具有应用潜力。

Abstract:

Icariin (ICA) is a small molecule drug capable of promoting cartilage repair and ameliorating inflammation. Loading ICA into a biomaterial scaffold for cartilage tissue engineering will thus potentially enhance the biological functionality of the engineered scaffold. In this study, short fibers processed from electrospun poly(l-lactide-co-caprolactone) (PLCL) fibers which were prior coated with polydopamine (PDA), were mixed with citric acid doped chitosan solution (CC) for preparing short fibers reinforced chitosan hydrogel (PDA@PLCL/CC) by a freeze-thawing combined freeze-drying method. Thereafter, ICA was loaded into the PDA@PLCL/CC scaffold through physical adsorption to generate a newly engineered biomimetic cartilage scaffold (ICA-PDA@PLCL/CC). Finally, ICA-mediated chondrogenic and ameliorated inflammatory effects of the ICA-PDA@PLCL/CC scaffold were examined in vitro using rabbit chondrocytes. The results showed that the ICA-free PDA@PLCL/CC scaffold possessed appropriate pore size and porosity (>80%), high water absorbance capacity and improved mechanical performance, and also promoted chondrocyte proliferation and adhesion. The ICA-laden ICA-PDA@PLCL/CC scaffold was evidenced to maintain cytomorphology, upregulate the expression of chondrogenic gene (sox-9), glycosaminoglycan gene (gag), and type Ⅱ collagen gene (col Ⅱ) as well as the synthesis of the cartilage matrix. In the presence of a simulated inflammation, the ICA-PDA@PLCL/CC scaffold was found to reduce chondrocyte fibrosis, effectively downregulate the expression of proinflammatory factors interleukin-6 (il-6), interleukin-1 (il-1), and inducible nitric oxide synthase (inos) in chondrocytes. It can also reduce matrix metalloproteinase-3 (mmp-3) expression and promote the synthesis of the extracellular matrix glycosaminoglycan (GAG) and type II collagen (Col II). The newly developed ICA-PDA@PLCL/CC scaffold may find applications in the regeneration and repair of cartilage defects.