3D打印静电纺定向沉积细胞与纳米纤维仿生构筑三维空间内“细胞和细胞外基质”

Biomimetical construction of cells and extracellular matrix within three-dimensional space is a critical scientific problem of design and preparation of tissue regenerative materials, and cells deposition during the process of building multilayered scaffolds based biomaterials, accompanying with the regulation and control of cell biological characteristics, is significant to the reparation and reconstruction of multilayer cells based tissues, and up to now rare related researches have been reported. Previous research results indicated that cells electrospinning has little effect on the cytoactive and cellular morphology of cells, and nanofibrous scaffolds could simulate the microstructure of extracellular matrix deeply. In this study, 3D printing electrospinning technique will be employed to fabricate 3D printing electrospun chitin and silk-fibrion composite nanofibrous mats, after that, 3D printing electrospun targeted cells on their surface orientedly. Repeat the above fibers electrospinning and cells electrospinning process alternately till the desired number obtained to get the sandwich-like composite active mats with the property of structural controllability for targeted tissue defect replacement. This project aims to establish an efficient method of 3D printing electrospinning for cells deposition, select the key parameters for keeping the best cell biological characteristics, and investigate the influence of changing chemical composition and topological structure of the prepared nanofibrous mats on the cell behavior such as adhesion, proliferation, and differentiation. Besides, we also will elucidate the structure-activity relationship and mechanism of action between the microstructure control of the scaffolds and the effects in the targeted tissue reconstruction, ultimately set up a referenced model of 3D printing electrospinning for depositing various kinds of cells or multilayered cells. The implementation will employ 3D printing electrospinning technique for cells immobilization and the cell biological characteristics regulation and control, which may also help us to gain a new insight into intelligent biomimetic construction of scaffolds for rebuilding the multicellular tissues.

在三维空间内实现细胞和细胞外基质的仿生构筑是组织再生材料设计与制备的关键科学问题，其中支架构建过程中伴随细胞沉积及调控细胞生物学性状对多层细胞源组织仿生重构至关重要。研究证实：静电纺细胞对细胞活性和形态无明显影响，纳米纤维能有效模拟细胞外基质结构。本项目拟采用3D打印静电纺技术构筑甲壳素/丝素蛋白纳米纤维支架仿生细胞外基质，随后通过3D打印静电纺定向沉积不同目标细胞并加入纳米纤维隔离层，根据目标组织重建需求沉积细胞与纳米纤维。项目拟建立3D打印静电纺沉积细胞有效方法并筛选有利于保持细胞生物学性状的关键参数；弄清支架拓扑结构调控对细胞粘附、增殖及分化等行为的影响规律；研究支架结构调控与目标组织重建效果间构效关系及作用机制；建立3D打印静电纺沉积多种/多层细胞的可供参考模型。本项目将3D打印静电纺用于活细胞固定化及其生物学性状调控，有望为多层细胞源三维组织支架的智能仿生构建提供新思路。

多层生物支架材料构建伴随细胞沉积及细胞生物学性状调控对多层细胞源组织修复重建具有重要意义。交替静电纺-电喷可实现纳米纤维支架上活性微生物细胞均匀沉积，且微生物层厚度可控。本项目采用交替电纺-电喷打印技术，电纺醋酸纤维素/累托石复合纳米纤维膜，电喷定向沉积酵母细胞，构建了夹心状纤维/酵母复合纳米纤维支架。以壳聚糖/丝素蛋白修饰的多层静电纺丝纤维素纳米纤维为材料，制备了负载间充质干细胞的壳聚糖/丝素纤维支架，大幅提升了干细胞的存活效率，所得的细胞-纤维支架层级材料被成功用于心肌修复。在丝素-甲壳素纳米纤维膜上负载生长因子（TGF-β1）获得3D复合纤维支架，该支架能在六周内极大促进成骨细胞的粘附、增殖、分化，提高骨修复效果。基于上述研究，本项目建立了电纺/喷沉积细胞有效方法并筛选有利于保持细胞生物学性状的关键参数；弄清了纳米纤维组分及拓扑结构对细胞粘附、增殖及分化等行为影响规律；最终建立了电纺/喷沉积多种/多层细胞的参考模型，累计发表SCI论文15篇，授权发明专利2项，有望为新型三维支架的构建提供新思路。