组织工程用PLA纤维基支架的制备、成型及细胞相容性研究
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摘要
论文基于静电纺丝技术，制备出PLA(polylactic acid)基支架材料：溶液静电纺PLA纤维支架，熔融静电纺PLA纤维支架及溶液静电纺PLA纱线支架。
探讨了溶液静电纺成纤过程中纺丝液射流运动过程：超过临界电压后，纺丝液射流所受电相斥力与表面张力的相互作用，决定着射流从直线区域进入到鞭动区域时刻。而正是射流在鞭动区域的运动引起直径剧减，形成超细纤维。基于此，介入不同成分与组分的溶剂，以干涉纺丝液射流在运动过程所受的电相斥力对射流在运动过程，特别是鞭动区域的运动构成影响，成功调控制备出直径分布差异的溶液静电纺PLA纤维支架材料及溶液静电纺PLA纱线支架材料。分别借助Fridrikh S V模型与Rutledge G C模型，进一步探讨了不同溶剂成分与组分对溶液静电纺PLA纤维直径差异的调控机制。
借助高速相机在纺丝过程中拍摄的射流照片，探讨了熔融静电纺成纤过程中聚合物射流的运动过程：超过临界电压后，喷丝口处聚合物熔滴从球形被拉伸成锥形，形成泰勒锥，射流向收集端作加速运动；射流运动过程中，表面张力与电相斥力之间的相互竞争作用决定了射流的鞭笞稳定性，进而决定了射流成纤时的细化程度。并基于熔融静电纺丝技术，制备出高度定向排列熔融静电纺PLA纤维支架材料。
采用表面沉积的方法，分别对溶液静电纺PLA纤维支架和熔融静电纺PLA纤维支架进行表面Ti/Cu沉积处理。形貌观察发现：对于溶液静电纺PLA纤维支架，表面Ti/Cu沉积造成了纤维表面凹凸不平的类似多孔结构；而对于熔融静电纺PLA纤维支架，Ti/Cu以纳米级的颗粒沉积在纤维表面的部分区域，并在纤维表面呈岛状生长。接触角测试表明：溶液静电纺制备的PLA纤维支架具有一定的疏水性，而熔融静电纺PLA纤维支架则具有一定的亲水性；表面Ti/Cu沉积使溶液静电纺和熔融静电纺PLA纤维均具有较好的亲水性，特别是表面Ti沉积的熔融静电纺PLA纤维，亲水性大大增加，1s时间内，完全润湿。结合形貌观察结果，探讨Ti/Cu金属颗粒在PLA纤维基表面生长机理发现：在表面Ti/Cu沉积初始阶段，金属颗粒主要以核生长型在静电纺PLA纤维基表面进行生长。
分别针对溶液静电纺PLA纤维支架材料，表面Ti/Cu沉积熔融静电纺PLA纤维支架材料以及静电纺PLA纱线支架材料的细胞相容性进行了研究。MTT (4-甲偶氮唑蓝)检测结果发现：NIH3T3细胞在表面Ti沉积的浸提液中，细胞活力随着培养时间的推移而增大，在72h时甚至超过未处理支架材料；体外毒性评定发现，表面Ti沉积样品，在24h时，具有轻微毒性，而到72h时，毒性逐渐消失至0级。而NIH3T3细胞在表面Cu沉积熔融静电纺PLA纤维支架材料浸提液中，细胞活力随培养时间推移稍有降低。体外毒性评定发现，24h，48h及72h时，Cu沉积熔融静电纺PLA纤维支架材料均表现出轻微毒性。
对不同直径范围的溶液静电纺PLA纤维/纱线支架材料的增殖与粘附结果表明：纤维基支架材料均比浇铸膜支架材料更有利于NIH3T3细胞的粘附和增殖；培养72h后，NIH3T3细胞在所有PLA纤维基支架材料上以不同的增长率进行了增殖；纱线的加捻结构，对细胞在PLA纤维支架材料上的增殖有着较大影响；基于纤维之间孔隙结构差异，孔隙率等因素，无论是在不同直径分布的PLA纤维支架材料，还是纱线支架材料上，难以单独从细胞粘附和增殖，判断纤维直径对细胞生长情况的影响。
In our work, PLA fibrous scaffolds such as solution electrospinning(S-ES) PLA fibrousscaffolds melt-electrospinning(M-ES) PLA fibrous scaffolds as well as solutionelectrospun(S-ES) yarns were fabricated based on electrospinning.
Jet motion in the fiber formation process of was investigated. As a voltage exceeds thecritical voltage, stability of the jet may be viewed as competition between surface tension andsurface charge repulsion. While it is jet motion that gives rise to a sharp decrease in diameter,forming ultra-fine fibers. With this in mind, solvents with different composition andproportion were introduced to affect the charge repulsion in jet motion, especially in thewhipping section. Hence, S-ES PLA fibrous scaffolds and S-ES PLA yarns with differentfiber diameter distribution were successfully prepared. Fridrikh S V model and Rutledge G Cmodel were applied for further investigation on the regulating mechanism.
Motion of polymer jet in the fiber formation process during melt-elctrospinning wasinvestigated based on pictures recorded by a high-speed CCD camera. When a voltage abovethe critical voltage applied, the sphere polymer melt at the nozzle tip will be stretched to acone shaped droplet, forming the Taylor cone and jet will then be accelerated downfield. Andthe competition between surface tension and surface charge repulsion results in a jetinstability to whipping, as well as the jet thinning in fiber formation. In addition, M-ES PLAfibrous scaffolds with well aligned fiber structure were prepared by melt-electrospinning.
Surface deposition was adopted for surface functionalization of both S-ES and M-ESPLA fibrous scaffolds. Morphology observation shows that surface Ti/Cu deposition on S-ESPLA fibrous scaffold created an uneven and porous structure on the fiber surface. And themetal Ti/Cu show a deposition of certain regions on the fiber surface of M-ES PLA fibrousscaffold in the form of nano-scale particles with an island growth model. Results of contactangle measurement indicate that S-ES fibrous scaffolds are hydrophobic to some extent, whileM-ES fibrous scaffolds are hydrophilic. Both Ti/Cu sputtered S-ES PLA and M-ES fibrousscaffolds exhibit a significant increasing of hydrophilicity as a result of the surface deposition,espesically for the Ti sputtered M-ES PLA fibrous scaffolds. In less than1s, fiber surface ofthe M-ES PLA fibrous scaffolds was entirely wetted. Considering with morphologyobservation results, it can be concluded that the metal particles exhibit a nucleation-growthmodel on the electrospun PLA fiber surface.
Investigations on the cytocompatibility of S-ES PLA fibrous scaffold, Ti/Cu sputteredM-ES PLA fibrous scaffolds as well as S-ES PLA fibrous scaffolds were conducted. MTTresults indicate that NIH3T3fibroblasts show an increasing viability over time in theextraction of Ti sputtered scaffolds, even with a higher OD value when cultured for72h.Results of cytotoxicity in vitro show that the Ti sputtered PLA fibrous scaffolds exhibit aslight toxic effects when cultured for24h. However, the toxicity disappear to0degrade. Andviability of cells in the extract of Cu sputtered scaffolds show a slight decrease over time.Evaluation of cytotoxicity in vitro indicates that Cu sputtered scaffolds show a slight toxiceffects on NIH3T3fibroblasts when cultured for as long as72h.
Adhesion and proliferation results of S-ES PLA fibrous/yarn scaffolds with differentdiameter distribution indicate that:1) compared with PLA casting film, all fibrous scaffoldsshow a better support both for adhesion and proliferation of NIH3T3fibroblasts;2) NIH3T3fibroblasts proliferated on all fibrous scaffolds at different levels;3) twists of yarnsignificantly affect the proliferation of NIH3T3fibroblasts;4) it is difficult to give aconclusion on the effects of fiber diameter on cell growth due to the complex structure offibrous scaffold.
探讨了溶液静电纺成纤过程中纺丝液射流运动过程：超过临界电压后，纺丝液射流所受电相斥力与表面张力的相互作用，决定着射流从直线区域进入到鞭动区域时刻。而正是射流在鞭动区域的运动引起直径剧减，形成超细纤维。基于此，介入不同成分与组分的溶剂，以干涉纺丝液射流在运动过程所受的电相斥力对射流在运动过程，特别是鞭动区域的运动构成影响，成功调控制备出直径分布差异的溶液静电纺PLA纤维支架材料及溶液静电纺PLA纱线支架材料。分别借助Fridrikh S V模型与Rutledge G C模型，进一步探讨了不同溶剂成分与组分对溶液静电纺PLA纤维直径差异的调控机制。
借助高速相机在纺丝过程中拍摄的射流照片，探讨了熔融静电纺成纤过程中聚合物射流的运动过程：超过临界电压后，喷丝口处聚合物熔滴从球形被拉伸成锥形，形成泰勒锥，射流向收集端作加速运动；射流运动过程中，表面张力与电相斥力之间的相互竞争作用决定了射流的鞭笞稳定性，进而决定了射流成纤时的细化程度。并基于熔融静电纺丝技术，制备出高度定向排列熔融静电纺PLA纤维支架材料。
采用表面沉积的方法，分别对溶液静电纺PLA纤维支架和熔融静电纺PLA纤维支架进行表面Ti/Cu沉积处理。形貌观察发现：对于溶液静电纺PLA纤维支架，表面Ti/Cu沉积造成了纤维表面凹凸不平的类似多孔结构；而对于熔融静电纺PLA纤维支架，Ti/Cu以纳米级的颗粒沉积在纤维表面的部分区域，并在纤维表面呈岛状生长。接触角测试表明：溶液静电纺制备的PLA纤维支架具有一定的疏水性，而熔融静电纺PLA纤维支架则具有一定的亲水性；表面Ti/Cu沉积使溶液静电纺和熔融静电纺PLA纤维均具有较好的亲水性，特别是表面Ti沉积的熔融静电纺PLA纤维，亲水性大大增加，1s时间内，完全润湿。结合形貌观察结果，探讨Ti/Cu金属颗粒在PLA纤维基表面生长机理发现：在表面Ti/Cu沉积初始阶段，金属颗粒主要以核生长型在静电纺PLA纤维基表面进行生长。
分别针对溶液静电纺PLA纤维支架材料，表面Ti/Cu沉积熔融静电纺PLA纤维支架材料以及静电纺PLA纱线支架材料的细胞相容性进行了研究。MTT (4-甲偶氮唑蓝)检测结果发现：NIH3T3细胞在表面Ti沉积的浸提液中，细胞活力随着培养时间的推移而增大，在72h时甚至超过未处理支架材料；体外毒性评定发现，表面Ti沉积样品，在24h时，具有轻微毒性，而到72h时，毒性逐渐消失至0级。而NIH3T3细胞在表面Cu沉积熔融静电纺PLA纤维支架材料浸提液中，细胞活力随培养时间推移稍有降低。体外毒性评定发现，24h，48h及72h时，Cu沉积熔融静电纺PLA纤维支架材料均表现出轻微毒性。
对不同直径范围的溶液静电纺PLA纤维/纱线支架材料的增殖与粘附结果表明：纤维基支架材料均比浇铸膜支架材料更有利于NIH3T3细胞的粘附和增殖；培养72h后，NIH3T3细胞在所有PLA纤维基支架材料上以不同的增长率进行了增殖；纱线的加捻结构，对细胞在PLA纤维支架材料上的增殖有着较大影响；基于纤维之间孔隙结构差异，孔隙率等因素，无论是在不同直径分布的PLA纤维支架材料，还是纱线支架材料上，难以单独从细胞粘附和增殖，判断纤维直径对细胞生长情况的影响。
In our work, PLA fibrous scaffolds such as solution electrospinning(S-ES) PLA fibrousscaffolds melt-electrospinning(M-ES) PLA fibrous scaffolds as well as solutionelectrospun(S-ES) yarns were fabricated based on electrospinning.
Jet motion in the fiber formation process of was investigated. As a voltage exceeds thecritical voltage, stability of the jet may be viewed as competition between surface tension andsurface charge repulsion. While it is jet motion that gives rise to a sharp decrease in diameter,forming ultra-fine fibers. With this in mind, solvents with different composition andproportion were introduced to affect the charge repulsion in jet motion, especially in thewhipping section. Hence, S-ES PLA fibrous scaffolds and S-ES PLA yarns with differentfiber diameter distribution were successfully prepared. Fridrikh S V model and Rutledge G Cmodel were applied for further investigation on the regulating mechanism.
Motion of polymer jet in the fiber formation process during melt-elctrospinning wasinvestigated based on pictures recorded by a high-speed CCD camera. When a voltage abovethe critical voltage applied, the sphere polymer melt at the nozzle tip will be stretched to acone shaped droplet, forming the Taylor cone and jet will then be accelerated downfield. Andthe competition between surface tension and surface charge repulsion results in a jetinstability to whipping, as well as the jet thinning in fiber formation. In addition, M-ES PLAfibrous scaffolds with well aligned fiber structure were prepared by melt-electrospinning.
Surface deposition was adopted for surface functionalization of both S-ES and M-ESPLA fibrous scaffolds. Morphology observation shows that surface Ti/Cu deposition on S-ESPLA fibrous scaffold created an uneven and porous structure on the fiber surface. And themetal Ti/Cu show a deposition of certain regions on the fiber surface of M-ES PLA fibrousscaffold in the form of nano-scale particles with an island growth model. Results of contactangle measurement indicate that S-ES fibrous scaffolds are hydrophobic to some extent, whileM-ES fibrous scaffolds are hydrophilic. Both Ti/Cu sputtered S-ES PLA and M-ES fibrousscaffolds exhibit a significant increasing of hydrophilicity as a result of the surface deposition,espesically for the Ti sputtered M-ES PLA fibrous scaffolds. In less than1s, fiber surface ofthe M-ES PLA fibrous scaffolds was entirely wetted. Considering with morphologyobservation results, it can be concluded that the metal particles exhibit a nucleation-growthmodel on the electrospun PLA fiber surface.
Investigations on the cytocompatibility of S-ES PLA fibrous scaffold, Ti/Cu sputteredM-ES PLA fibrous scaffolds as well as S-ES PLA fibrous scaffolds were conducted. MTTresults indicate that NIH3T3fibroblasts show an increasing viability over time in theextraction of Ti sputtered scaffolds, even with a higher OD value when cultured for72h.Results of cytotoxicity in vitro show that the Ti sputtered PLA fibrous scaffolds exhibit aslight toxic effects when cultured for24h. However, the toxicity disappear to0degrade. Andviability of cells in the extract of Cu sputtered scaffolds show a slight decrease over time.Evaluation of cytotoxicity in vitro indicates that Cu sputtered scaffolds show a slight toxiceffects on NIH3T3fibroblasts when cultured for as long as72h.
Adhesion and proliferation results of S-ES PLA fibrous/yarn scaffolds with differentdiameter distribution indicate that:1) compared with PLA casting film, all fibrous scaffoldsshow a better support both for adhesion and proliferation of NIH3T3fibroblasts;2) NIH3T3fibroblasts proliferated on all fibrous scaffolds at different levels;3) twists of yarnsignificantly affect the proliferation of NIH3T3fibroblasts;4) it is difficult to give aconclusion on the effects of fiber diameter on cell growth due to the complex structure offibrous scaffold.
引文
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