成年猪胰岛细胞的分离纯化及其微囊包裹  

Isolation,Purification and Microencapsulation of the Adult Porcine Pancreatic Islets

作　　者：杨学伟[1] 章乐虹[1] 胡以则[1] 刘应生[1] 

机构地区：[1]广州医学院第二附属医院普通外科,广东广州510260

出　　处：《武汉大学学报（医学版）》2007年第4期496-499,I0004,共5页Medical Journal of Wuhan University

摘　　要：目的:探索成年猪胰岛细胞分离纯化的有效方法,获取具有免疫隔离作用的小微囊化胰岛细胞。方法:①用胶原酶灌注,人工、静态的方法分离成年猪胰腺;用Ficoll400不连续密度梯度离心纯化猪胰岛,从形态和功能学(体外培养)鉴别分离细胞的活力。②用自制的静电微囊发生装置、采用纯化的海藻酸钠和多聚赖氨酸材料制备小微囊化胰岛细胞,并鉴别微囊胰岛的活力和微囊膜的通透性。结果:①每只猪胰腺可获得(292647±79909)个胰岛,纯化前胰岛产量为(5360±1779)IEQ/g胰腺组织,纯化后产量(3239±1116)IEQ/g胰腺组织,胰岛纯度为90%。胰岛组织学结构完整,体外培养胰岛素分泌良好,葡萄糖刺激释放反应明显,显示了良好的细胞活力。②微囊化胰岛细胞圆形,直径315-350μm,表面光滑,大小相对均一,其内可见1-2个胰岛团。微囊胰岛也具良好活力,微囊膜通透性良好。结论:①本方法分离成年猪胰岛能获得较高数量和具有良好活力的猪胰岛细胞。②自制的静电微囊发生装置能制成具有生物活性的大小相对均一,圆形,表面光滑的小APA微囊化胰岛细胞(直径315-350μm),为进行肝内微囊化胰岛移植提供了可能。Objective： To study the effective methods for the isolation and purification of adult porcine pancreatic islets and to obtain the smaller microencapsulated porcine islets with immunoisolation. Methods： Porcine islets were isolated by using manual and static digestion by ductal injection of collagenase, and purified by centrifugation on Ficoll 400 discontinuons density gradients. The viability of the islets was assessed by histological examination and the glucose stimulated insulin secretion test. Small alginate-poly-L-lysine-alginate microencapsulated islets were produced with a self-made high voltage electrostatic pulse system. The viability of the microencapsulated islets and the penetrability of the capsules were assessed too. Results： The number of isolated porcine islets was （292 647＋79 909） islets/porcine pancreas （n=12）, corresponding to （5 360＋1 779） IEQ/g pancreas tissue; after purification, the number of islets was （3 239＋1 116） IEQ/g pancreas. The purity of recovered porcine islets was 90 %. The isolated islets showed the almost normal cell structure by histology. The concentration of insulin released in the culture medium and insulin secretion in response to the different glucose concentrations of the islets after purification confirmed good viability in vitro. Microencapsulated islets had a diameter of 315-350 μm. Capsules were more uniform and its surface were smooth, most capsules contained one or two islets. The culture of the microencapsulated islets in vitro also demonstrated the viable cells and the capsules had good penetrability. Conclusion： A considerable amounts of porcine islets with high quality can be obtained from an adult porcine pancreas using our methods. Microencapsulated porcine islets can be gained which are smaller （315-350 μm diameter） with smooth surface and viable cells by this self-made high voltage electrostatic pulse system, which provide a basis for intrahepatic xenotransplantation of microencapsulated adult porcine islets.

关 键 词：猪 胰岛 分离 纯化 微囊化 

分 类 号：R587.1[医药卫生—内分泌]