【摘要】 目的：开发E7肽功能化聚酰亚胺（polyimide,PI）纤维支架，用于巨大不可修复性肩袖撕裂（massive irreparable rotator cuff tears,MIRCT）的组织工程修复，评估其理化性能及生物活性。方法：通过表面预处理、羧基化改性、聚多巴胺（polydopamine,PDA）涂层构建及E7肽偶联制备PI-PDA-E7支架。同时制备PI支架和PI-PDA支架作为对照。采用扫描电子显微镜、原子力显微镜、傅里叶变换红外光谱、X射线光电子能谱、热重分析、接触角测试及拉伸测试，表征各支架的表面形貌、化学结构、热稳定性、亲水性及力学性能；采用蠕变实验和动态弯折实验评价PI-PDA-E7支架的长期力学稳定性和抗疲劳性能。骨髓间充质干细胞（bone marrow mesenchymal stem cells,BMSCs）取自大鼠骨髓，经体外分离、扩增培养后，以一定密度接种于三组支架表面进行共培养。通过活/死细胞染色、细胞计数试剂盒-8(cell counting kit-8,CCK-8)、细胞骨架染色、Transwell试验、实时荧光定量PCR(real-time quantitative polymerase chain reaction,RT-qPCR)、Western Blot、碱性磷酸酶（alkaline phosphatase,ALP）活性检测及茜素红染色实验评估支架对BMSCs的生物相容性、增殖、招募及成骨分化能力。结果：PI-PDA-E7支架表面均匀致密，亲水性显著增强（接触角降至42°±2°），热稳定性优异，拉伸强度达271±6 MPa，弹性模量3854±154 MPa，均优于PI及PI-PDA组；PI-PDA-E7支架高温蠕变及15000次动态弯折实验显示良好耐疲劳性。三组支架BMSCs存活率均>90%,PI-PDA-E7支架对BMSCs的增殖及招募能力显著高于PI及PI-PDA组（P<0.01）。与PI组和PI-PDA组相比，PI-PDA-E7支架上BMSCs成骨相关基因[Runt相关转录因子2(runt-related transcription factor 2,Runx2)、ALP、Ⅰ型胶原α1链（collagen type Ⅰ alpha1 chain,Col1α1）、骨钙素（Osteocalcin,OCN）、骨形态发生蛋白-2(bone morphogenetic protein 2,BMP-2)]及蛋白[Runx2、骨桥蛋白（Osteopontin ,OPN）、Ⅰ型胶原（Collagen Ⅰ）]表达显著上调（P<0.05）,ALP活性显著增强（P<0.01），成骨诱导培养14天后矿化结节形成量（OD405值）显著高于PI组和PI-PDA组（P<0.01）。结论：PI-PDA-E7支架兼具优异力学性能和生物活性，能有效招募BMSCs并促进成骨分化，为MIRCT修复提供新型组织工程材料，具备临床应用潜力。更多还原

【Abstract】 Objective To develop E7 peptide-functionalized polyimide(PI) fiber scaffolds for tissue engineering repair of massive irreparable rotator cuff tears(MIRCT),and to evaluate their physicochemical properties and biological activity. Methods PI-PDA-E7 scaffolds were prepared through surface pretreatment, carboxylation modification, polydopamine(PDA) coating construction, and subsequent E7 peptide conjugation. Meanwhile,PI scaffolds and PI-PDA scaffolds were fabricated as controls. The surface morphology, chemical structure, thermal stability, hydrophilicity, and mechanical properties of all scaffolds were characterized using the scanning electron microscopy,atomic force microscopy,Fourier transform infrared spectroscopy,X-ray photoelectron spectroscopy,thermogravimetric analysis,contact angle measurement,and tensile testing. Additionally,creep tests and dynamic folding tests were performed to evaluate the long-term mechanical stability and fatigue resistance of the PIPDA-E7 scaffolds. Bone marrow mesenchymal stem cells(BMSCs) were isolated from rat bone marrow,expanded in vitro,and seeded onto the three groups of scaffolds at a defined density for co-culture. The biocompatibility,proliferation,recruitment,and osteogenic differentiation capacity of the scaffolds toward BMSCs were evaluated through live/dead cell staining,cell counting kit-8(CCK-8) assay,cytoskeletal staining,Transwell migration assay,real-time quantitative polymerase chain reaction(RT-qPCR),Western Blot,alkaline phosphatase(ALP) activity assay,and Alizarin Red staining. Results The PI-PDA-E7 scaffold exhibited a uniform and dense surface morphology, significantly enhanced hydrophilicity(water contact angle reduced to 42° ± 2°),excellent thermal stability,and superior mechanical properties,with a tensile strength of 271 ± 6 MPa and an elastic modulus of 3854 ±154 MPa,both significantly higher than those of the PI and PI-PDA groups. Moreover,the PI-PDAE7 scaffold demonstrated good fatigue resistance in high-temperature creep and 15,000-cycle dynamic bending tests. Moreover,the BMSC survival rate of all three scaffold groups exceeded 90%. Compared with the PI and PI-PDA groups,the PI-PDA-E7 scaffold significantly promoted BMSC proliferation and recruitment(P<0.01). Furthermore,a significant upregulation was observed in the expression of osteogenic-related genes [Runt-related transcription factor 2(Runx2),ALP,Collagen type Ⅰ alpha 1 chain(Col1α1), Osteocalcin(OCN), and bone morphogenetic protein 2(BMP-2)] and proteins[Runx2,Osteopontin(OPN),and Collagen Ⅰ] in BMSCs cultured on the PI-PDA-E7 scaffold(P<0.05). What’s more,its ALP activity was markedly enhanced(P<0.01),and the mineralized nodules formed after 14 days of osteogenic induction(OD405 value) were significantly higher than the PI and PI-PDA groups(P<0.01). Conclusion The PI-PDA-E7 scaffold possesses both excellent mechanical properties and favorable bioactivity,effectively recruiting BMSCs and promoting their osteogenic differentiation. Therefore,it provides a novel tissue engineering material for MIRCT repair and holds promising potentials for clinical application.更多还原