生物電紡技術製備含PC12細胞之順向中空纖維薄膜及管內細胞生長行為評估

In Vitro Study of Bio-Electrospun Microtube Array Membranes Containing PC12 Cells

陳建華(Chien-Hwa Chen)

指導教授 ： 陳建中

臺北醫學大學/口腔醫學院/生醫材料暨工程研究所/碩士(2012年)

https://doi.org/10.6831/TMU.2012.00062

摘要

神經再生醫學中，理想神經導管設計概念是以具生物相容性與可降解性高分子為基材(scaffold)，且須具備適當的微孔及順向結構提供神經細胞貼附生長與利於養分與代謝物置換，亦有接觸導引(contact guidance)之作用，使神經細胞有序列地向斷端處生長，幫助神經系統修復與再生。本研究目的在於以生物電紡技術(bio-electrospinning)技術整合電氣紡絲(electrospinning)製程與細胞，製備含有神經細胞之聚乳酸順向中空纖維薄膜，並評估薄膜內細胞生長形態以及細胞活性與分化能力。生物電紡技術可直接以一製程結合組織工程要素，可將神經細胞導入支架中，相較於傳統單一管道神經修復導管有較多比表面積，可增加細胞貼附面積同時也能導引神經細胞生長方向性。研究結果顯示，PC12類神經細胞經電場強度0~2.2 k V/cm刺激時，仍具有細胞活性與增生能力。 4% w/v (PEO:PEG = 1:1)內管溶液對於PC12類神經細胞具有良好細胞相容性，BCA assay 數據顯示，PC12類神經細胞於順向中空纖維薄膜內亦有生物活性與增生能力，將細胞排列角度計算為細胞順向度，結果顯示控制組順向度較低，約為14.2 ± 1%；而Pure PLA順向中空纖維薄膜組別順向度為94.9 ± 9%； PLA/PEG40 %組別順向度為90 ± 7%，顯示順向中空結構可導引PC12類神經細胞軸突生長方向性。本研究以生物電紡技術設計概念，利用靜電紡絲技術將PC12類神經細胞導入聚乳酸順向中空纖維薄膜內，其薄膜具導引細胞生長方向之特性，管內細胞具有活性與增生能力，經過一系列細胞體外評估，期望未來能應用神經修復與再生。

關鍵字

PC12類神經細胞；生物電紡；順向中空纖維薄膜；神經再生

並列摘要

Injuries to the nervous system lead to permanent loss of neurologic function. The common way to restore the directed axonal growth across the lesion site is to provide a physical support and bridging devices, such as synthetic or natural conduits, which have several advantages over auto graft counterpart, especially, no source restraint. It was hence considered a promising candidate for such surgical needs. The objective of this study is to demonstrate the possibility of using bio-electrospinning to prepare cell-containing, biodegradable microtube array scaffold as an improved nerve guide conduit (NGC). Poly-l-lactic acid (PLLA) dope solution was delivered as outer shell, while the PC12 cell in the 4wt% PEO/PEG (1:1) solution as the core components, for the co-axial electrospinning to the fabricate novel PC12 cells containing PLLA microtube array (MTA) membrane. With certain operation condition, smooth PC12 cell in PLLA (CiPLLA) MTA membranes were obtained and the cells were found to evenly distribute within the MTA membrane. The viability of PC12 cells after electrospinning was found to be around 75% at electric field strengths
of 1、1.5、2.2 kV/cm and dropped to less than 60% at 3kV/cm. BCA assay data showed PC12 cell proliferated normally in PLLA MTA membrane. The PC12 cell differentiation induced by the added NGF was found to be successful with the neurite extended more than 90 microns after 5 days culture, however, still shorter than 130 microns of control group. On the other hand, the alignment of the neurite extension of PC12 cell within MTA (94.9±9%) was significantly better than those of control group (14.2±1%). These results demonstrated the highly aligned PLLA MTA scaffold can served as an effective guiding scaffold for PC12 cell and bio-electrospinning process provide an avenue to integrate cell, growth factor and scaffold in situ for tissue engineering regeneration. The PC12 cells seeded PLLA microtube array scaffold, based upon the advanced nerve guide conduit (NGC) model, has the potential application as an effective NGC for nerve regeneration.

並列關鍵字

PC12 cells；Bio-electrospinning；PLA microtube array；Nerve repair

參考文獻

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