负载巨噬细胞-骨髓间充质干细胞的3D打印预构建血管化定压复合体对股骨头缺血性坏死修复的研究

Osteonecrosis of femoral head is a major challenge for orthopaedic surgeons because of its high morbidity. How to acquire the vascularization meanwhile the osteogenesis remains difficult.Will be on the basis of previous studies,this study utilizes BMSCs both inducing phenotypic transformation of macrophagocyte leading to vasifaction and osteogenic differentiation when influenced by a certain pressure to be a great seeding cell. We apply parallel-plate flow chamber to obtain the marker of stress answer using RNA-Seq and get the optimal stress for BMSCs osteogenic differentiation, and analysis the stress by Bernoulli's equation and Hagen-Poiseuille law in fluid dynamics to get appropriate data of diameter and length of main channel when in vivo. Then we apply 3 d printing technology using the data calculating by fluid dynamics and acquiring from the necrosis area to prepare beta - CS/ESHU composite scaffold, which contain a main and several minor channels to provide certain pressur. In this way SPIO marked BMSCs loaded by alginate adhered to the prostheses matching with main channel can achieve osteogenic differentiation beyond the stress of flowing blood, and transfer to the minor channels to induce the phenotypic transformation of macrophages there to achieve vascularization. Ultimately we verify the new composite bracket with constant pressure for the repair of the necrosis of the femoral head by in vivo animal experiments.

股骨头无菌性坏死因其极高的致残率一直是骨科医生面临的一个重大挑战。如何在早期同时实现成骨与成血管是治疗难点。本研究将在前期研究的基础上，利用骨髓间充质干细胞可诱导巨噬细胞表型转化促进成血管，其本身又可在一定应力刺激下成骨分化的特点，设计体外实验，通过RNA-Seq及生物信息学检索获得应力相关指示蛋白，利用流体动力学粘性流体的伯努利方程与泊肃叶定律分析骨髓间充质干细胞最优成骨分化时的应力，获得体内血流状态下满足最佳应力时的管径、管长等数据作为支架主管道数据，结合股骨头坏死区影像学扫描数据，利用3D打印技术精确制备具有主副管道的 β- CS/ESHU定压复合支架，将负载有SPIO标记的骨髓间充质干细胞藻酸盐凝胶附着在人工血管内壁，使其在一定的血流应力下成骨分化，并迁移至副管道与附着在副管道的巨噬细胞相互作用实现成血管，最终通过体内动物实验验证这种新型的支架对股骨头坏死的修复作用。

股骨头无菌性坏死因其极高的致残率一直是骨科医生面临的一个重大挑战。如何在早期诊断同时实现成骨与成血管是治疗的难点。本研究首先使用生物信息学方法，分析确定了14个关键的microRNAs(miRNAs）作为股骨头坏死潜在的生物标志物，获得了302个miRNA靶基因，构建了miRNA-mRNA相互作用网络。分析发现，这14个关键miRNAs与股骨头坏死密切相关。通过定量实时聚合酶链反应的结果证实了股骨头坏死患者血清中miR-93-5p和miR-320a的表达显著增高。这些循环中的miRNAs可作为股骨头坏死的早期诊断指标和潜在的治疗靶点。然后我们通过建立小鼠骨缺损模型、分离培养小鼠的脂肪来源的间充质干细胞和骨髓来源的巨噬细胞发现脂肪组织来源的间充质干细胞(ADSCs)具有促进骨损伤的修复的能力;脂肪组织来源的间充质干细胞具有促进骨髓巨噬细胞向M2型极化;M2型巨噬细胞具有促进脂肪组织来源的间充质干细胞成骨分化的能力，两种细胞能够协同促进骨修复。本研究筛选出了股骨头坏死的早期诊断指标，探索了组织工程学中种子细胞在成骨方面的相互作用，为股骨头无菌性坏死的早期诊断和治疗提供了新的思路。