應用射出成形／粒子析出法於三維多孔性鼻支架之製造及細胞培養研究

Research on Fabrication and Cell Culture of Three-Dimensional Porous Nasal Scaffolds by Injection Molding/Particulate Leaching Technique

江裕群(Yuh-Chyun Chiang) ； 薛如茵(Ju-Yin Hsueh) ； 許信德(Hsin-Te Hsu) ； 李飛鵬(Fei-Peng Lee) ； 沈永康(Yung-Kang Shen) ； 洪嘉祥(Jia-Hsiang Hung)

《台灣耳鼻喉頭頸外科雜誌》 47卷3期 (2012/06) Pp. 173-180

https://doi.org/10.6286/2012.47.3.173

摘要

背景：我們延續先前『應用溶劑澆鑄/粒子過濾法結合模流分析於三維多孔性生物鼻支架之研究』成果，以射出成形/粒子析出法製作出實體的L型鼻支架，並研究該支架相關降解測試及細胞培養的結果。方法：實驗分成三部份：首先利用模具和射出成形/粒子析出法製造以聚甘醇酸（PLGA；polylactideco-glycolide）為主材料的L型鼻用多孔性生物支架，並引入田口法，找出射出成形的最佳化製程。實驗的第二部份是為期20週的降解測試，測試本研究製作出的L型鼻用多孔性生物支架重量損失的百分率，和磷酸緩衝液（PBS; phosphate buffer solution）的酸鹼值變化。實驗的第三部份為細胞培養，以類骨母細胞（MG63；osteoblast-like cells）培養在本研究製作出的L型鼻用多孔性生物支架，透過MTT試驗（MTT；Microculture Tetrazolium Test）量測光密度值（OD；Optical Density）值，並以顯微鏡觀察細胞貼附型態。結果：實驗的第一部份：我們成功地以射出成形/粒子析出法製作出實體的L型鼻支架，並列出過程中使用的射出參數。透過電子掃描顯微鏡所拍攝的孔洞圖，孔洞平均大小為250μm，孔隙率高達88％。實驗的第二部份：降解材料重量損失的百分率在第7週後開始急遽下降，至第15週相對重量為0.4。磷酸緩衝液的酸鹼值在第8週開始下降，到第12週酸鹼值緩慢上升。實驗的第三部份：經過4天培養，本研究製作出的支架量測的光密度值明顯高於控制組；在第7天，光密度值則達控制組的5倍。由細胞在支架上的電子掃描顯微鏡圖觀察，在培養4小時後，可以看出細胞在支架上附著；到第7天，細胞已經在支架上明顯的延展開。結論：以射出成形/粒子析出法製作出實體的L型鼻支架，在孔徑分布、大小與孔隙率部份已可依指定要求完全掌控。在降解實驗中，以聚甘醇酸為主材料的L型鼻用多孔性生物支架在第7週開始降解。細胞培養的結果，明顯看出用射出成形/粒子析出法所製作的聚甘醇酸支架適合類骨母細胞的生長。

關鍵字

組織工程 ； L型鼻支架 ； 聚甘醇酸 ； 射出成形/粒子析出法 ； 類骨母細胞

並列摘要

BACKGROUND: The aim of this research combines the injection molding /particulate leaching method to fabricate the L-type nasal scaffold with PLGA to do the degradation test and cell culture.METHODS: This research firstly to fabricate the L-type nasal scaffold by injection molding/particulate leaching method with PLGA (Polylactide-co-glycolicle)/NaCl. This research also to find the optimal processing by Taguchi method. Then this research does the degradation test for 20 weeks. The relative weight loss, pH variation, nanoindentor (ASMEC UNAT-M) test are determined after incubation. In order to investigate whether cells can grow in the resultant scaffold or not, cell culture is conducted in the scaffolds. MG-63 osteoblastlike cells are chosen to culture in the scaffolds. This research measures the OD (Optical density) value by MTT (Microculture tetrazolium test) assay and observes the cell adhesion by SEM (Scanner electron microscope).RESULTS: The L-shaped nasal scaffold is success fabricated by injection molding/particulate leaching method. The product is then immersed in D.I. water to remove NaCl. As a result, the scaffold has a well interconnected structure of average pore size ranging from 250 μm and 88% porosity according to SEM images. The degradation test of the PLGA scaffold reveals that the structure degrades gradually after 7 weeks. The relative weight just is 0.4 on 15 weeks. The pH value decreases as the eighth week and increases as the twelfth week. Cell culture is undertaken to cell proliferation in the scaffold. In terms of the results of cell culture, it turns out that high cell proliferation and viability is found in the PLGA scaffold. The OD value of nasal scaffold is higher than it of control group as the fourth day and its value is five times than control group on the seventh day. The cell acts as the adhesion behavior at the scaffold on the fourth day and it expands at the scaffold on the seventh day.CONCLUSIONS: This study can be concluded that the porous nasal scaffold made by the IM/PL method shows the acceptable pore distribution, pore size, porosity, biocompatibility and degradation. The nasal scaffold begins to degradate on the seventh week. The nasal scaffold may degrade well after surgery and the cartilage tissue may grow well in it. The nasal scaffold is suitable for osteoblast cell growth by injection molding/particulate leaching method.

並列關鍵字

Tissue engineering ； L-shaped nasal scaffold ； Poly(lactic-co-glycolic acid) (PLGA) ； Injection molding/particulate leaching technique ； MG63 osteoblast-like cells