CAJ | 학술논문

生物材料%材料力学及表面改性%肝细胞特异性%大孔微载体%丝素蛋白%细胞培养%壳聚糖%半乳糖基%冷冻干燥法%生物人工肝%863项目生物材料%材料力學及錶麵改性%肝細胞特異性%大孔微載體%絲素蛋白%細胞培養%殼聚糖%半乳糖基%冷凍榦燥法%生物人工肝%863項目
생물재료%재료역학급표면개성%간세포특이성%대공미재체%사소단백%세포배양%각취당%반유당기%냉동간조법%생물인공간%863항목
biomaterials%material mechanics and surface modification%hepatocyte-specific characteristics%macroporous microcarrier%silk fibroin%cell culture%chitosan%galactosyl%freeze-drying%bioartificial liver%National“863”Project of China

背景:在肝细胞移植及生物人工肝的研究中,肝细胞培养是关键环节,肝细胞表面存在着去唾液酸糖蛋白受体和半乳糖基之间的分子识别功能,如何方便地获取足够数量、活性较好、功能良好的肝细胞已经成为其首要问题. 目的:制备一种新的体外三维环境下具有肝细胞特异性的高性能的大孔微载体. 方法:以丝素蛋白、壳聚糖及乳糖酸为基本材料,通过乳化-化学交联、冷冻干燥、极性溶液处理制备一种新型丝素蛋白/半乳糖基化壳聚糖大孔微载体.采用光学显微镜、扫描电子显微镜和通过超导核磁共振波谱、傅里叶变换红外光谱及细胞相容性等对其形态结构及理化性质进行表征. 结果与结论:实验所制备的丝素蛋白/半乳糖基化壳聚糖大孔微载体,内外多孔且相互连通,具有肝细胞特异性.扫描电子显微镜下观察显示,该大孔微载体表面呈多孔结构,孔径为40-80μm,开口向外,呈喇叭状,孔的分布均匀,内部亦为多孔结构,适合于肝细胞高密度培养.超导核磁共振波谱、傅里叶变换红外光谱检测结果显示,半乳糖基已经成功复合到支架材料之中.细胞相容性实验结果显示,肝细胞能与材料特异性黏附、细胞围绕微载体聚集生长,并表现出很好的活力.结果证实,应用乳化-化学交联、极性溶液处理、冷冻干燥等方法可制备出具有肝细胞特异性亲和力的丝素蛋白/半乳糖基化壳聚糖大孔微载体.
배경:재간세포이식급생물인공간적연구중,간세포배양시관건배절,간세포표면존재착거타액산당단백수체화반유당기지간적분자식별공능,여하방편지획취족구수량、활성교호、공능량호적간세포이경성위기수요문제. 목적:제비일충신적체외삼유배경하구유간세포특이성적고성능적대공미재체. 방법:이사소단백、각취당급유당산위기본재료,통과유화-화학교련、냉동간조、겁성용액처리제비일충신형사소단백/반유당기화각취당대공미재체.채용광학현미경、소묘전자현미경화통과초도핵자공진파보、부리협변환홍외광보급세포상용성등대기형태결구급이화성질진행표정. 결과여결론:실험소제비적사소단백/반유당기화각취당대공미재체,내외다공차상호련통,구유간세포특이성.소묘전자현미경하관찰현시,해대공미재체표면정다공결구,공경위40-80μm,개구향외,정나팔상,공적분포균균,내부역위다공결구,괄합우간세포고밀도배양.초도핵자공진파보、부리협변환홍외광보검측결과현시,반유당기이경성공복합도지가재료지중.세포상용성실험결과현시,간세포능여재료특이성점부、세포위요미재체취집생장,병표현출흔호적활력.결과증실,응용유화-화학교련、겁성용액처리、냉동간조등방법가제비출구유간세포특이성친화력적사소단백/반유당기화각취당대공미재체.
@@@@BACKGROUND:Hepatocyte culture is the key to the hepatocyte transplantation and bioartificial liver study. It has turned into the first problem how to get adequate number of better activity and wel-functioning hepatocytes. @@@@OBJECTIVE:To prepare a high-performance silk fibroin/galactosylated chitosan macroporous microcarrier with hepatocyte-specific characteristics, for hepatocyte adhesion culture in the three-dimensional environment in vitro. @@@@METHODS:Silk fibroin, chitosan, and lactose acid served as raw materials to prepare a hepatocyte-specific silk fibroin/galactosylated chitosan macroporous microcarrier using comprehensive application of emulsified-chemical crosslinking, polarity solution and freeze-drying. Optical microscope, scanning electron microscope, proton nuclear magnetic resonance spectroscopy, Fourier transform infrared spectroscopy, and cel biocompatibility test were used to characterize its morphology, physical and chemical properties. @@@@RESULTS AND CONCLUSION:The microcarrier was successful y prepared in the project. Furthermore, the pores were occupied inside and outside of the microcarrier and interconnected, which presented hepatocyte-specific characteristics. Under the scanning electron microscope, the macroporous microcarrier exhibited a trumpet-shaped porous structure with the pore size of 40-80 μm and even pore distribution, providing an appropriate environment for high-density culture of hepatocytes. The galactosyl was successful y compounded into the scaffold material which was confirmed by the proton nuclear magnetic resonance spectroscopy and Fourier transform infrared spectroscopy. Cel biocompatibility test showed a hepatocyte-specific affinity to the silk fibroin/galactosylated chitosan macroporous microcarrier, and the cel s grew and aggregated around the microcarrier exhibiting a good activity. These findings indicate that combination of emulsified-chemical crosslinking, polarity solution and freeze-drying can prepare a silk fibroin/galactosylated chitosan macroporous microcarrier with hepatocyte-specific characteristics.