聚乙烯醇／柞蚕丝素蛋白材料作为肌腱组织工程支架材料的初步研究  

PRELIMINARY STUDY ON POLYVINYL ALCOHOL/WILD ANTHERAEA PERNYI SILK FIBROIN AS NANOFIBER SCAFFOLDS FOR TISSUE ENGINEERED TENDON

作　　者：吴琳琳[1] 李敏[1] 赵晋[1] 陈登龙[2] 周志华[1] 

机构地区：[1]福建师范大学生命科学学院福州,350108 [2]福建师范大学化学与材料学院

出　　处：《中国修复重建外科杂志》2011年第2期181-186,共6页Chinese Journal of Reparative and Reconstructive Surgery

基　　金：国家自然科学基金资助项目(30570956);福建省科技厅重点资助项目(2010Y0020)~~

摘　　要：目的检测聚乙烯醇(polyvinyl alcohol,PVA)/柞蚕丝素蛋白(wild antheraea pernyi silk fibroin,WSF)电纺膜与肌腱细胞的细胞相容性,探讨其作为肌腱组织工程支架材料的可行性。方法以质量分数为98%的甲酸为溶剂,配制浓度均为11%的WSF溶液、PVA溶液及PVA/WSF(质量配比为9∶1)溶液,采用电纺丝技术制备WSF、PVA及PVA/WSF电纺膜,扫描电镜观察电纺成丝情况。取5只3日龄SD大鼠尾腱,体外分离培养肌腱细胞并传至第3代进行实验。取浓度为1×106个/mL肌腱细胞分别与PVA及PVA/WSF电纺膜复合培养,4、12 h采用MTT法测量细胞黏附率。取浓度为4.5×103个/mL肌腱细胞,分别于PVA、1/2 PVA、1/4 PVA、PVA/WSF、1/2 PVA/WSF、1/4 PVA/WSF电纺膜浸提液中培养,MTT法测定培养1、3、5、7 d的吸光度(A)值,并进行毒性等级评价。取PVA、PVA/WSF电纺膜分别与细胞密度为1×106个/mL的肌腱细胞悬液100μL复合培养制备细胞-材料复合物,7 d后行HE染色观察,1、3、5、7 d行扫描电镜观察。结果 WSF甲酸溶液无法电纺成丝,PVA和PVA/WSF甲醛溶液均可成功电纺成丝。肌腱细胞与PVA、PVA/WSF电纺膜复合培养4 h后,其细胞黏附率分别为26.9%±0.4%、87.0%±1.0%,差异有统计学意义(t=100.400,P=0.000);复合培养12 h后,其细胞黏附率分别为35.2%±0.6%、110.0%±1.7%,差异有统计学意义(t=42.500,P=0.000)。体外浸提液毒性评价结果显示,培养后1、3、5、7 d各种电纺膜间细胞A值比较,差异均有统计学意义(P<0.05);1/2 PVA和1/4 PVA浸提液细胞毒性等级差异有统计学意义(P<0.05)。细胞-材料复合物的组织学及扫描电镜观察显示肌腱细胞均可黏附于PVA、PVA/WSF电纺膜表面,但PVA/WSF电纺膜上细胞较多。结论 PVA/WSF电纺膜支架材料具有良好的细胞相容性,有望成为肌腱组织工程的理想支架。Objective To investigate the cellular compatibility of polyvinyl alcohol（PVA）/wild antheraea pernyi silk fibroin（WSF）,and to explore the feasibility for tendon tissue engineering scaffold in vitro.Methods The solutions of WSF（11%）,PVA（11%）,and PVA/WSF（11%） were prepared with 98% formic acid（mass fraction） at a mass ratio of 9 ： 1.The electrospinning membranes of WSF,PVA,and PVA/WSF were prepared by electrostatic spinning apparatus.The morphologies of scaffolds were evaluated using scanning electronic microscope（SEM）.The tendon cells were isolated from tail tendon of 3-day-old Sprague Dawley rats in vitro.The experiment was performed using the 3rd generation cells.The tendon cells（1 × 106/mL） were cocultured with PVA and PVA/WSF electrospinning film,respectively,and MTT test was used to assess the cell adhesion rate 4,12 hours after coculture.The tendon cells were cultured in PVA and PVA/WSF extraction medium of different concentration（1,1/2,and 1/4）,respectively;and the absorbance（A） values were detected at 1,3,5,and 7 days to evaluate the cytotoxicity.The composite of tendon cells and the PVA or PVA/WSF scaffold were observed by HE staining at 7 days and characterized by SEM at 1,3,5,and 7 days.Results The solution of WSF could not be used to electrospin;and the solution of PVA and PVA/WSF could be electrospun.After coculture of tendon and PVA or PVA/WSF electrospinning membranes,the cell adhesion rates were 26.9% ± 0.4% and 87.0% ± 1.0%,respectively for 4 hours,showing significant difference（t=100.400,P=0.000）;the cell adhesion rates were 35.2% ± 0.6% and 110.0% ± 1.7%,respectively for 12 hours,showing significant difference（t=42.500,P=0.000）.The cytotoxicity of PVA/WSF was less significantly than that of PVA（P 〈 0.05） and significant difference was observed between 1/2 PVA and 1/4 PVA（P 〈 0.05）.HE staining and SEM images showed that the tendon cells could adhere to PVA and PVA/WSF scaffolds,but that the cells grew better in PVA/WSF scaffold tha

关 键 词：肌腱组织工程 聚乙烯醇 柞蚕丝素蛋白 电纺丝技术 支架材料 肌腱细胞 大鼠 

分 类 号：R318.08[医药卫生—生物医学工程]