P（LLA-CL）复合纳米纤维的制备、表征及在生物医学领域的应用
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摘要
天然细胞外基质是由邻近细胞分泌的三种主要成分即蛋白纤维、蛋白聚糖和非纤维蛋白成分通过化学和物理的交联组成的,其中直径分布在几十到几百纳米的蛋白纤维,主要是胶原蛋白和弹性蛋白纤维通过相互的交织形成具有三维结构的网状结构为组织的生长提供支撑和弹性。聚合物纳米纤维在形态上类似于天然细胞外基质,可以为细胞的生长提供一个三维的空间和更多的黏附位点。本文利用静电纺丝制备聚合物纳米纤维以仿生天然细胞外基质作为生物医用材料用于组织工程支架、药物缓释等相关领域。本文内容涉及材料加工、机械设计、医疗器械领域,主要分为三个部分:
(1)首次制备由天然材料壳聚糖和人工聚合材料P(LLA-CL)复合的纳米纤维仿生细胞外基质,并通过SEM、FTIR、XRD、接触角、机械性能测定分析该纳米纤维的物理、化学性能,并通过成纤维细胞和PIECs细胞在纤维膜上的黏附和增殖行为对其安全性进行了评估。实验发现纤维的直径随纺丝溶液的浓度和P(LLA-CL)所占比例的增加而递增;纤维膜的亲水性随着壳聚糖组分的逐渐增加依次增强;该纳米纤维膜能为成纤维细胞和PIECs细胞的生长提供一个适宜的三维的空间环境,有助于细胞的粘附和增殖。此后,为得到具三维结构的纳米纤维管状组织工程支架,本文设计了一套静电纺接收装置,可将电纺纤维成型为长度、厚度和管腔直径可控的管状支架。另外,通过对该装置的改进我们得到了由排列纤维组成的膜状织物,并发现通过调节织物中纤维的排列度可以调节断裂拉伸力学性能。
(2)通过同轴共纺技术首次成功制备出了芯层载有肝素或肝素与明胶复合物的超细纤维及织物。通过TEM、SEM及Image J软件,我们对其形态及壳-芯结构进行了分析。肝素及其复合物可被完全包裹于P(LLA-CL)中,其含量及纤维形态与芯层供给速率相关。纤维内部细胞实验中发现对成纤维细胞的生长具有明显的抑制作用。
(3)成功制备出以静电纺P(LLA-CL)纤维或同轴共纺载肝素P(LLA-CL)壳-芯纤维为覆膜的新型血管内支架。P(LLA-CL)纤维覆膜支架易被固定在传输器的球囊上,最大扩张压低于8个大气压,扩张后的回弹率分布在3.9%-6.1%,在液压P=16.0+0.3kPa时水的渗透量为3.23±1.62ml/cm2/min,动物实验发现对动脉瘤具有较好封堵效果,能够初步满足临床要求。
Native extracellular matrix (ECM) is a chemically and physically cross-linked complex network of three important classes of biomacromolecules secreted locally by cells. These are fibrous proteins, proteoglycans, and non-fibrous proteins. In a typical connective tissue, structural fibers such as collagen fibers and elastin fibers have diameters ranging from several ten to several hundred nanometers. The nanoscaled fibers entangle with each other to form a nonwoven mesh that provides tensile strength and elasticity for the tissues. The morphology of polymer nanofibers are similar to ECM, and can supply a 3-D environment and more acceptor sites for cell adherence and growth. In this study, electrospinning has been used to fabricate nanofibers as biomemitic ECM for excellent biomaterials. There are many research fields such as materials fabrication, mechanical design and medical device were involved as following in this paper.
(1) Chitosan-P(LLA-CL) complex was electrospun into nanofibers as biomemitic ECM first time. In this study, SEM, FTIR, XRD, contact angle, and materials test machine were used to characterize the nanofiber. The cells of fibroblast and PIECs were seeded onto nanofiber mats for adherence and proliferation studies. A new collector with a rotary mandrel has been design and used to fabricate tubular scaffold or aligned nanofiber fibric. The results showed that chitosan-P(LLA-CL) complex nanofiber and its fibric have shown valuable potential application as biomaterials for tissue engineering scaffold, wound dressing, and so on.
(2) Heparin contained core-shell fibers and fibric were fabricated using coaxial electrospinning technique. Through TEM, SEM test and Image J software, the morphology and diameter of core-shell fiber were investigated. In cell culturing test, heparin contained fibers have shown obvious inhibit affect on proliferation of fibroblast. Due to the special structure, this kind of fibers are very useful in application as heparin release and tissue engineering scaffold.
(3) A new type of covered stent was successfully developed with electropinning technique and the rotary mandrel collector in our study. Electrospun fibers covered stent could be well fixed on the balloon of delivery device, and showed good mechanical properties and morphology in dilation process. After expanding, the fiber fabric on stent could maintain their morphology well and be fixed in artery. As a new type of covered stent with low recoil and infiltration, electropun fibers covered stent showed potential application in clinical treatment of aneurysm..
(1)首次制备由天然材料壳聚糖和人工聚合材料P(LLA-CL)复合的纳米纤维仿生细胞外基质,并通过SEM、FTIR、XRD、接触角、机械性能测定分析该纳米纤维的物理、化学性能,并通过成纤维细胞和PIECs细胞在纤维膜上的黏附和增殖行为对其安全性进行了评估。实验发现纤维的直径随纺丝溶液的浓度和P(LLA-CL)所占比例的增加而递增;纤维膜的亲水性随着壳聚糖组分的逐渐增加依次增强;该纳米纤维膜能为成纤维细胞和PIECs细胞的生长提供一个适宜的三维的空间环境,有助于细胞的粘附和增殖。此后,为得到具三维结构的纳米纤维管状组织工程支架,本文设计了一套静电纺接收装置,可将电纺纤维成型为长度、厚度和管腔直径可控的管状支架。另外,通过对该装置的改进我们得到了由排列纤维组成的膜状织物,并发现通过调节织物中纤维的排列度可以调节断裂拉伸力学性能。
(2)通过同轴共纺技术首次成功制备出了芯层载有肝素或肝素与明胶复合物的超细纤维及织物。通过TEM、SEM及Image J软件,我们对其形态及壳-芯结构进行了分析。肝素及其复合物可被完全包裹于P(LLA-CL)中,其含量及纤维形态与芯层供给速率相关。纤维内部细胞实验中发现对成纤维细胞的生长具有明显的抑制作用。
(3)成功制备出以静电纺P(LLA-CL)纤维或同轴共纺载肝素P(LLA-CL)壳-芯纤维为覆膜的新型血管内支架。P(LLA-CL)纤维覆膜支架易被固定在传输器的球囊上,最大扩张压低于8个大气压,扩张后的回弹率分布在3.9%-6.1%,在液压P=16.0+0.3kPa时水的渗透量为3.23±1.62ml/cm2/min,动物实验发现对动脉瘤具有较好封堵效果,能够初步满足临床要求。
Native extracellular matrix (ECM) is a chemically and physically cross-linked complex network of three important classes of biomacromolecules secreted locally by cells. These are fibrous proteins, proteoglycans, and non-fibrous proteins. In a typical connective tissue, structural fibers such as collagen fibers and elastin fibers have diameters ranging from several ten to several hundred nanometers. The nanoscaled fibers entangle with each other to form a nonwoven mesh that provides tensile strength and elasticity for the tissues. The morphology of polymer nanofibers are similar to ECM, and can supply a 3-D environment and more acceptor sites for cell adherence and growth. In this study, electrospinning has been used to fabricate nanofibers as biomemitic ECM for excellent biomaterials. There are many research fields such as materials fabrication, mechanical design and medical device were involved as following in this paper.
(1) Chitosan-P(LLA-CL) complex was electrospun into nanofibers as biomemitic ECM first time. In this study, SEM, FTIR, XRD, contact angle, and materials test machine were used to characterize the nanofiber. The cells of fibroblast and PIECs were seeded onto nanofiber mats for adherence and proliferation studies. A new collector with a rotary mandrel has been design and used to fabricate tubular scaffold or aligned nanofiber fibric. The results showed that chitosan-P(LLA-CL) complex nanofiber and its fibric have shown valuable potential application as biomaterials for tissue engineering scaffold, wound dressing, and so on.
(2) Heparin contained core-shell fibers and fibric were fabricated using coaxial electrospinning technique. Through TEM, SEM test and Image J software, the morphology and diameter of core-shell fiber were investigated. In cell culturing test, heparin contained fibers have shown obvious inhibit affect on proliferation of fibroblast. Due to the special structure, this kind of fibers are very useful in application as heparin release and tissue engineering scaffold.
(3) A new type of covered stent was successfully developed with electropinning technique and the rotary mandrel collector in our study. Electrospun fibers covered stent could be well fixed on the balloon of delivery device, and showed good mechanical properties and morphology in dilation process. After expanding, the fiber fabric on stent could maintain their morphology well and be fixed in artery. As a new type of covered stent with low recoil and infiltration, electropun fibers covered stent showed potential application in clinical treatment of aneurysm..
引文
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